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Hameed A, Nawaz I, Alrokayan S, Hussain T, Iqbal J. Synthesis, structure activity relationship and biological evaluation of indole sulfonohydrazide derivatives as antagonists of P2Y1 and P2Y6 receptors. Bioorg Chem 2025; 160:108499. [PMID: 40280013 DOI: 10.1016/j.bioorg.2025.108499] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/11/2025] [Revised: 04/15/2025] [Accepted: 04/19/2025] [Indexed: 04/29/2025]
Abstract
The P2Y receptors play a significant role in regulating various physiological functions, such as neurotransmission and inflammatory response. They are also considered promising therapeutic targets for the treatment and prevention of conditions like neurological disorders, pain, cardiovascular diseases, thrombosis, and cancer. Active research is ongoing to identify antagonists of P2Y receptor. Although extensive research has been conducted on P2Y receptors inhibitors, only a limited number of P2Y receptors antagonists have been identified and approved by regulatory authority. In the current research, new indole sulfonohydrazide derivatives (3a-3 k) were synthesized in good yield. Based on toxicity assays performed on h-1321 N1 astrocytoma cell line, these low molecular weight compound showed a safe toxicity profile. The synthesized derivatives were also screened against tP2Y1 and rP2Y6 receptors using a calcium mobilization assay. The results showed that compounds 3a, 3b, 3 h and 3 k were potent against tP2Y1 with IC50 values of 9.91 ± 1.01 μM, 3.49 ± 0.31 μM, 9.72 ± 0.82 μM, and 6.14 ± 0.17 μM, respectively. Additionally, three compounds, i.e., 3d, 3f, and 3 h, exhibited potency against rP2Y6 with IC50 value of 9.22 ± 1.10 μM, 16.25 ± 0.27 μM, and 1.89 ± 0.11 μM, respectively. Molecular docking study was conducted to support the in vitro analysis, which revealed that the tested compounds showed favorable interaction with the amino acids of the target P2Y1 receptor, including Phe62, Phe66, Leu102, Thr103, Pro105, Ala106, Phe119 and Met123. An in silico pharmacokinetic study was also performed, which revealed that the synthesized compounds met all the criteria for favorable gastrointestinal absorption, indicating potential for oral bioavailability. The stability and reactivity of compounds were determined by using the Guassian09 programme in which the density functional theory (DFT) calculations were performed by using the B3LYP/SVP level.
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Affiliation(s)
- Abdul Hameed
- Centre for Advanced Drug Research, COMSATS University Islamabad, Abbottabad Campus, Abbottabad 22060, Pakistan; Department of Chemistry, COMSATS University Islamabad, Abbottabad Campus, Abbottabad, 22060, Pakistan
| | - Ismat Nawaz
- Centre for Advanced Drug Research, COMSATS University Islamabad, Abbottabad Campus, Abbottabad 22060, Pakistan
| | - Salman Alrokayan
- Research Chair for Biomedical Application of Nanomaterials, Biochemistry Department, College of Science, King Saud University, Riyadh 11451, Saudi Arabia
| | - Tajamul Hussain
- Research Chair for Biomedical Application of Nanomaterials, Biochemistry Department, College of Science, King Saud University, Riyadh 11451, Saudi Arabia; Center of Excellence in Biotechnology Research, King Saud University, Riyadh 11451, Saudi Arabia.
| | - Jamshed Iqbal
- Centre for Advanced Drug Research, COMSATS University Islamabad, Abbottabad Campus, Abbottabad 22060, Pakistan; Department of Chemistry, COMSATS University Islamabad, Abbottabad Campus, Abbottabad, 22060, Pakistan.
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Kristof Z, Szabo D, Sperlagh B, Torok D, Gonda X. From Childhood Woes to Adult Blues: Unmasking the Role of Early Traumas, P2X7 Receptor, and Neuroinflammation in Anxiety and Depression. Int J Mol Sci 2025; 26:4687. [PMID: 40429831 PMCID: PMC12111330 DOI: 10.3390/ijms26104687] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/13/2025] [Revised: 05/07/2025] [Accepted: 05/10/2025] [Indexed: 05/29/2025] Open
Abstract
Early-life stress may increase the risk of neuropsychiatric disorders via immune activation. While the purinergic signaling pathway is implicated in psychiatric disorders, the specific role of the P2X7 receptor (P2X7R) in anxiety, depression, and childhood trauma still requires further clarification. Upon chronic stress, excessive ATP release activates purinergic P2X7R signalling in the brain contributing to long-lasting neuroinflammation, which potentially promotes the development of psychiatric disorders. There is also a putative link between the P2X7 receptor gene, located on chromosome 12q24, and the development of anxiety and depression. This review aims to systematically examine how P2X7R contributes to the pathophysiology of anxiety and depressive disorders, with a particular focus on early-life stress (ELS). It offers a comprehensive synthesis of the current findings, emphasizing the previously unexplored intersections between P2X7R signaling, early-life stress, and psychiatric disorders. These interactions may shape long-term neuroinflammation, contributing to the development of anxiety and depression, and offer new insights into potential therapeutic targets. The review integrates the role of P2X7R regarding both indirect mechanisms-such as the modulation and long-term transmission of neuroinflammation following environmental stressors and vulnerability-and direct genetic associations with psychiatric conditions, including the influence of single-nucleotide polymorphisms (SNPs), haplotypes, and other variants within the P2X7 gene. Special emphasis is placed on the impact of early-life stress, drawing primarily on preclinical findings to elucidate underlying mechanisms.
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Affiliation(s)
- Zsuliet Kristof
- Discipline of Psychiatry, Adelaide Medical School, The University of Adelaide, Adelaide, SA 5000, Australia;
| | - Dorottya Szabo
- Laboratory of Molecular Pharmacology, HUN-REN Institute of Experimental Medicine, 1083 Budapest, Hungary; (D.S.); (B.S.)
- Janos Szentagothai Doctoral School, Semmelweis University, 1085 Budapest, Hungary
| | - Beata Sperlagh
- Laboratory of Molecular Pharmacology, HUN-REN Institute of Experimental Medicine, 1083 Budapest, Hungary; (D.S.); (B.S.)
| | - Dora Torok
- Department of Pharmacodynamics, Semmelweis University, 1089 Budapest, Hungary;
- Center of Pharmacology and Drug Research & Development, Semmelweis University, 1089 Budapest, Hungary
- Hungarian Brain Research Program, NAP3.0-SE Neuropsychopharmacology Research Group, 1089 Budapest, Hungary
| | - Xenia Gonda
- Department of Pharmacodynamics, Semmelweis University, 1089 Budapest, Hungary;
- Center of Pharmacology and Drug Research & Development, Semmelweis University, 1089 Budapest, Hungary
- Hungarian Brain Research Program, NAP3.0-SE Neuropsychopharmacology Research Group, 1089 Budapest, Hungary
- Department of Psychiatry and Psychotherapy, Semmelweis University, 1082 Budapest, Hungary
- Department of Clinical Psychology, Semmelweis University, 1092 Budapest, Hungary
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Wei S, Song X, Mou Y, Yang T, Wang Y, Wang H, Ren C, Song X. New insights into pathogenisis and therapies of P2X7R in Parkinson's disease. NPJ Parkinsons Dis 2025; 11:108. [PMID: 40325043 PMCID: PMC12053563 DOI: 10.1038/s41531-025-00980-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2024] [Accepted: 04/27/2025] [Indexed: 05/07/2025] Open
Abstract
Parkinson's disease (PD), a prevalent neurodegenerative disorder, is linked to genetics and environment, but its mechanisms remain unclear. Emerging evidence connects purinergic signaling-particularly ATP-sensitive P2X7 receptor (P2X7R)-to PD. P2X7R expression is elevated in PD patients, and its antagonist BBG mitigates 6-OHDA-induced dopaminergic neuron death. This review discusses P2X7R's structure, neural functions, PD-related mechanisms, and therapeutic potential as a targert.
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Affiliation(s)
- Shizhuang Wei
- Department of Otorhinolaryngology, Head and Neck Surgery, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
- Shandong Provincial Key Laboratory of Neuroimmune Interaction and Regulation, Yantai Yuhuangding Hospital, Yantai, China
- Shandong Provincial Clinical Research Center for Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Yantai, China
- Yantai Key Laboratory of Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
| | - Xiaoyu Song
- Department of Otorhinolaryngology, Head and Neck Surgery, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
- Shandong Provincial Key Laboratory of Neuroimmune Interaction and Regulation, Yantai Yuhuangding Hospital, Yantai, China
- Shandong Provincial Clinical Research Center for Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Yantai, China
- Yantai Key Laboratory of Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
| | - Yakui Mou
- Department of Otorhinolaryngology, Head and Neck Surgery, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
- Shandong Provincial Key Laboratory of Neuroimmune Interaction and Regulation, Yantai Yuhuangding Hospital, Yantai, China
- Shandong Provincial Clinical Research Center for Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Yantai, China
- Yantai Key Laboratory of Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
| | - Ting Yang
- Department of Otorhinolaryngology, Head and Neck Surgery, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
- Shandong Provincial Key Laboratory of Neuroimmune Interaction and Regulation, Yantai Yuhuangding Hospital, Yantai, China
- Shandong Provincial Clinical Research Center for Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Yantai, China
- Yantai Key Laboratory of Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
| | - Yao Wang
- Department of Otorhinolaryngology, Head and Neck Surgery, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
- Shandong Provincial Key Laboratory of Neuroimmune Interaction and Regulation, Yantai Yuhuangding Hospital, Yantai, China
- Shandong Provincial Clinical Research Center for Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Yantai, China
- Yantai Key Laboratory of Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
| | - Hanrui Wang
- Department of Otorhinolaryngology, Head and Neck Surgery, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
- Shandong Provincial Key Laboratory of Neuroimmune Interaction and Regulation, Yantai Yuhuangding Hospital, Yantai, China
- Shandong Provincial Clinical Research Center for Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Yantai, China
- Yantai Key Laboratory of Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China
| | - Chao Ren
- Department of Otorhinolaryngology, Head and Neck Surgery, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China.
- Shandong Provincial Key Laboratory of Neuroimmune Interaction and Regulation, Yantai Yuhuangding Hospital, Yantai, China.
- Shandong Provincial Clinical Research Center for Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Yantai, China.
- Yantai Key Laboratory of Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China.
- Department of Neurology, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China.
| | - Xicheng Song
- Department of Otorhinolaryngology, Head and Neck Surgery, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China.
- Shandong Provincial Key Laboratory of Neuroimmune Interaction and Regulation, Yantai Yuhuangding Hospital, Yantai, China.
- Shandong Provincial Clinical Research Center for Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Yantai, China.
- Yantai Key Laboratory of Otorhinolaryngologic Diseases, Yantai Yuhuangding Hospital, Qingdao University, Yantai, China.
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Zhang J, Ju T, Qin Y, Hou M, Gao L, Wu LA. Therapeutic targeting of P2X receptors for orofacial pain. Brain Res Bull 2025; 224:111301. [PMID: 40086763 DOI: 10.1016/j.brainresbull.2025.111301] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/08/2025] [Revised: 02/16/2025] [Accepted: 03/10/2025] [Indexed: 03/16/2025]
Abstract
P2X receptors (P2XRs), membrane ion channels activated by extracellular adenosine 5' -triphosphate (ATP), play a pivotal role in nociception by directly promoting pain signaling. Currently, antagonists targeting P2XRs have taken effect in alleviating various pain. The therapeutic potential of the P2X receptor family has become a central focus. Consequently, numerous research groups and pharmaceutical companies are actively engaged in developing novel P2XR antagonists. Furthermore, an increasing number of clinical trials on P2XR antagonists have obtained encouraging results. This review provides an overview of the structural characteristics and cellular localization of P2XRs, their molecular mechanisms and signaling pathways implicated in orofacial pain. Additionally, it explores the development of P2XR antagonists and their therapeutic application for managing orofacial pain. In conclusion, this review highlights the promising role of P2XRs as therapeutic targets for orofacial pain treatment.
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Affiliation(s)
- Jing Zhang
- State Key Laboratory of Oral & Maxillofacial Reconstruction and Regeneration, National Clinical Research Center for Oral Diseases, Shaanxi Clinical Research Center for Oral Diseases, Department of Pediatric Den-tistry, School of Stomatology, The Fourth Military Medical University, Xi'an 710032, China.
| | - Tianjuan Ju
- State Key Laboratory of Oral & Maxillofacial Reconstruction and Regeneration, National Clinical Research Center for Oral Diseases, Shaanxi Clinical Research Center for Oral Diseases, Department of Pediatric Den-tistry, School of Stomatology, The Fourth Military Medical University, Xi'an 710032, China.
| | - Yaru Qin
- State Key Laboratory of Oral & Maxillofacial Reconstruction and Regeneration, National Clinical Research Center for Oral Diseases, Shaanxi Clinical Research Center for Oral Diseases, Department of Pediatric Den-tistry, School of Stomatology, The Fourth Military Medical University, Xi'an 710032, China.
| | - Ming Hou
- State Key Laboratory of Oral & Maxillofacial Reconstruction and Regeneration, National Clinical Research Center for Oral Diseases, Shaanxi Clinical Research Center for Oral Diseases, Department of Pediatric Den-tistry, School of Stomatology, The Fourth Military Medical University, Xi'an 710032, China.
| | - Lei Gao
- State Key Laboratory of Oral & Maxillofacial Reconstruction and Regeneration, National Clinical Research Center for Oral Diseases, Shaanxi Clinical Research Center for Oral Diseases, Department of Pediatric Den-tistry, School of Stomatology, The Fourth Military Medical University, Xi'an 710032, China.
| | - Li-An Wu
- State Key Laboratory of Oral & Maxillofacial Reconstruction and Regeneration, National Clinical Research Center for Oral Diseases, Shaanxi Clinical Research Center for Oral Diseases, Department of Pediatric Den-tistry, School of Stomatology, The Fourth Military Medical University, Xi'an 710032, China.
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Cen YY, Gao XL, Feng YH, Zhou C, Li CJ, Liu F, Shen JF, Zhang YY. The Double-Edged Effect of Connexins and Pannexins of Glial Cells in Central and Peripheral Nervous System After Nerve Injury. Mol Neurobiol 2025:10.1007/s12035-025-04991-6. [PMID: 40310549 DOI: 10.1007/s12035-025-04991-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2025] [Accepted: 04/21/2025] [Indexed: 05/02/2025]
Abstract
Glial cells play pivotal roles in homeostatic regulation and driving reactive pathologic changes after nerve injury. Connexins (Cxs) and pannexins (Panxs) have emerged as seminal proteins implicated in cell-cell communication, exerting a profound impact on the response processes of glial cell activation, proliferation, protein synthesis and secretion, as well as apoptosis following nerve injury. These influences are mediated through various forms, including protein monomers, hemichannel (HC), and gap junction (GJ), mainly by regulating intercellular or intracellular signaling pathways. Multiple Cx and Panx isoforms have been detected in central nervous system (CNS) or peripheral nervous system (PNS). Each isoform exhibits distinct cellular and subcellular localization, and the differential regulation and functional roles of various protein isoforms are observed post-injury. The quantitative and functional alterations of the same protein isoform in different studies remain inconsistent, attributable to factors such as the predominant mode of protein polymerization, the specific injury model, and the injury site. Similarly, the same protein isoforms have different roles in regulating the response processes after nerve injury, thus exerting a double-edged sword effect. This review describes the regulatory mechanisms and bidirectional effects of Cxs and Panxs. Additionally, it surveys the current status of research and application of drugs as therapeutic targets for neuropathic injuries. We summarize comprehensive and up-to-date information on these proteins in the glial cell response to nerve injury, providing new perspectives for future mechanistic exploration and development of targeted therapeutic approaches.
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Affiliation(s)
- Yue-Yan Cen
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, National Center for Stomatology, West China School of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China
- Department of Prosthodontics, West China Hospital of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China
| | - Xin-Lin Gao
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, National Center for Stomatology, West China School of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China
- Department of Prosthodontics, West China Hospital of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China
| | - Yu-Heng Feng
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, National Center for Stomatology, West China School of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China
- Department of Prosthodontics, West China Hospital of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China
| | - Cheng Zhou
- Laboratory of Anesthesia and Critical Care Medicine, Translational Neuroscience Center, West China Hospital of Sichuan University, Chengdu, China
| | - Chun-Jie Li
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, National Center for Stomatology, West China School of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China
- Department of Head and Neck Oncology, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Fei Liu
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, National Center for Stomatology, West China School of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China
- Department of Prosthodontics, West China Hospital of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China
| | - Jie-Fei Shen
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, National Center for Stomatology, West China School of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China.
- Department of Prosthodontics, West China Hospital of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China.
| | - Yan-Yan Zhang
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, National Center for Stomatology, West China School of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China.
- Department of Prosthodontics, West China Hospital of Stomatology, Sichuan University, No. 14, Section 3, Renminnan Road, Chengdu, 610041, China.
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Chaves MM. Neutrophils and purinergic signaling: Partners in the crime against Leishmania parasites? Biochimie 2025; 232:43-53. [PMID: 39855456 DOI: 10.1016/j.biochi.2025.01.004] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2024] [Revised: 12/18/2024] [Accepted: 01/17/2025] [Indexed: 01/27/2025]
Abstract
The parasite of the genus Leishmania is the causative agent of diseases that affect humans called leishmaniasis. These diseases affect millions of people worldwide and the currently existing drugs are either very toxic or the parasites acquire resistance. Therefore, new elimination mechanisms need to be elucidated so that new therapeutic strategies can be developed. Much has already been discussed about the role of neutrophils in Leishmania infection, and their participation is still controversial. A recent study showed that receptors present in the neutrophil membrane, the purinergic receptors, can control the infection when activated, but the triggering mechanism has not been elucidated. In this review, we will address the possible participation of purinergic receptors expressed in the neutrophil extracellular membrane that may be participating in the detection of Leishmania infection and their possible effects during parasitism.
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Affiliation(s)
- Mariana M Chaves
- Laboratory on Thymus Research, Oswaldo Cruz Institute, Oswaldo Cruz Foundation, Rio de Janeiro, Brazil; Bio-Manguinhos, Oswaldo Cruz Foundation, Brazilian Ministry of Health, Rio de Janeiro, Brazil.
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Li C, Wang B. Role of P2X7R in Retinal Diseases: A Review. Immun Inflamm Dis 2025; 13:e70203. [PMID: 40396593 DOI: 10.1002/iid3.70203] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/04/2025] [Revised: 03/16/2025] [Accepted: 04/25/2025] [Indexed: 05/22/2025] Open
Abstract
BACKGROUND P2X purinoceptor 7 receptor (P2X7R) is an ATP-gated ion channel that, upon activation by ATP, triggers the release of inflammatory mediators and induces apoptosis in cells. This channel plays a crucial role in the onset and progression of various diseases. Recently, there has been a growing body of research focused on the function of P2X7R receptors in ophthalmic conditions, particularly concerning retinal diseases such as age-related macular degeneration, diabetic retinopathy, and retinitis pigmentosa. OBJECTIVE This article is to provide a comprehensive review of the advancements in the study of P2X7R and its association with retinal diseases, elucidating its role in these conditions and identifying potential avenues for future research. METHODS Electronic databases, including PubMed, Web of Science, and Wan fang Data were searched for relevant literature. The following keywords were used: "P2X7R", Age-related macular degeneration", "Diabetic retinopathy", "Retinitis pigmentosa". Both preclinical and clinical studies were included to provide a holistic understanding of P2X7R's role in retinal pathology. RESULTS P2X7R activation exacerbates retinal diseases by promoting inflammation and apoptosis. However, its role in disease progression and homeostasis complicates therapeutic targeting, highlighting the need for selective inhibitors and further research into its context-dependent functions. CONCLUSION P2X7R plays a critical role in the pathogenesis of retinal diseases. At the same time, preclinical studies suggest that P2X7R inhibition holds promise as a therapeutic strategy. Future research should focus on developing selective P2X7R inhibitors, elucidating the receptor's role in different disease stages, and identifying biomarkers to guide personalized treatment. Addressing these challenges will be essential for translating P2X7R-targeted therapies into clinical practice and improving outcomes for patients with retinal diseases.
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Affiliation(s)
- Chunli Li
- Department of Ophthalmology, The Second Hospital of Lanzhou University, Lanzhou, Gansu, China
| | - Binsheng Wang
- Department of General Surgery, The First Hospital of Lanzhou University, Lanzhou, Gansu, China
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Hoffrogge R, Karachunskaya A, Heitmann N, Pedreiturria X, Klöster K, Bader V, Winklhofer KF, Hamacher M, Klebl B, Gold R, Dinkel K, Kleiter I, Faissner S. The P2X7R-antagonist AFC-5128 ameliorates chronic experimental autoimmune encephalomyelitis in a preventive and therapeutic paradigm. Front Immunol 2025; 16:1554999. [PMID: 40308584 PMCID: PMC12040686 DOI: 10.3389/fimmu.2025.1554999] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2025] [Accepted: 03/31/2025] [Indexed: 05/02/2025] Open
Abstract
Background Multiple sclerosis (MS) is characterized by chronic inflammation driven by central nervous system (CNS)-resident immune cells such as microglia, especially during the progressive phase of the disease. The P2X7 receptor (P2X7R), a risk protein for MS, is ubiquitously expressed on immune cells. AFC-5128, a CNS-penetrating small molecule inhibitor of P2X7R, is a promising agent for the treatment of autoimmune diseases such as MS. Methods In vitro, the effects on the calcium influx of primary murine microglia were assessed via Fluo-4 calcium imaging. In vivo, MOG35-55 immunized C57BL/6 mice were treated with AFC-5128, fingolimod (FTY) or vehicle in different treatment paradigms. The mice were scored daily. Microglial marker expression, immune cell phenotyping and serum cytokine analyses were performed via flow cytometry. Immune cell infiltration, demyelination and Iba1+/CD3+ cells were detected in spinal cord cross-sections. The effects of MOG35-55 T-cell restimulation were assessed in vitro. Results In vitro, treatment of primary microglia with 10 µM AFC-5128 reduced the influx of calcium following ATP stimulation (p<0.0001). In vivo, treatment of mice with AFC-5128 led to a reduction in overall EAE scores in acute and chronic EAE, with the best effects using 200 mg/kg body weight AFC-5128 (p<0.0001). Peripheral immune cell subsets (B cells, T cells and macrophages) and serum cytokine levels of chronic EAE mice treated in a therapeutic paradigm were not affected. While the expression of homeostasis markers of microglia in AFC-5128-treated mice was not affected, there was a trend toward lower expression of phagocytosis-associated markers. Late therapeutic treatment with AFC-5128 had only mild effects on chronic EAE. Conclusion The treatment of EAE mice with AFC-5128 improved acute and chronic EAE in different treatment paradigms, with positive effects on histological markers and slight modulation of microglial marker expression. Mechanistically, calcium influx of microglia was reduced following AFC-5128 treatment, which implies the ability of AFC-5128 to stabilize calcium homeostasis. Therefore, therapeutic inhibition of P2X7R via AFC-5128 has the potential for translation into a treatment of both relapsing and progressive forms of multiple sclerosis.
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MESH Headings
- Animals
- Encephalomyelitis, Autoimmune, Experimental/drug therapy
- Encephalomyelitis, Autoimmune, Experimental/immunology
- Encephalomyelitis, Autoimmune, Experimental/prevention & control
- Encephalomyelitis, Autoimmune, Experimental/metabolism
- Mice
- Microglia/drug effects
- Microglia/immunology
- Microglia/metabolism
- Purinergic P2X Receptor Antagonists/pharmacology
- Purinergic P2X Receptor Antagonists/therapeutic use
- Receptors, Purinergic P2X7/metabolism
- Mice, Inbred C57BL
- Female
- Disease Models, Animal
- Multiple Sclerosis/drug therapy
- Myelin-Oligodendrocyte Glycoprotein/immunology
- Cytokines/blood
- Calcium/metabolism
- Cells, Cultured
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Affiliation(s)
| | | | - Neele Heitmann
- Department of Neurology, St. Josef-Hospital, Bochum, Germany
| | | | | | - Verian Bader
- Molecular Cell Biology, Ruhr-University, Bochum, Germany
| | | | | | - Bert Klebl
- KHAN Technology Transfer Fund I GmbH & Co KG, Dortmund, Germany
| | - Ralf Gold
- Department of Neurology, St. Josef-Hospital, Bochum, Germany
| | | | - Ingo Kleiter
- Department of Neurology, St. Josef-Hospital, Bochum, Germany
- Behandlungszentrum Kempfenhausen für Multiple Sklerose Kranke gemeinnützige GmbH, Berg, Germany
| | - Simon Faissner
- Department of Neurology, St. Josef-Hospital, Bochum, Germany
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Erlitz KS, Prinz AK, Wagner S, Massa J, Dunker C, Höhl M, Griep A, McManus RM, Schelhaas S, Koch O, Junker A. Naphtho[1,2- b][1,4]diazepinedione-Based P2X4 Receptor Antagonists from Structure-Activity Relationship Studies toward PET Tracer Development. J Med Chem 2025; 68:6965-7002. [PMID: 39805099 DOI: 10.1021/acs.jmedchem.4c02171] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/16/2025]
Abstract
The P2X4 receptor is implicated in various pathological conditions, including neuropathic pain and cancer. This study reports the development of 1,4-naphthodiazepinedione-based P2X4 receptor antagonists aimed at both therapeutic applications and potential use as PET tracers for imaging P2X4 receptor expression in cancer. Structure-activity relationship studies aided by docking studies and molecular dynamics simulations led to a series of compounds with potent P2X4 receptor antagonism, promising in vitro inhibition of interleukin-1β release in THP-1 cells and suitability for radiolabeling with fluorine-18. The most potent compounds were further evaluated for their physicochemical properties, metabolic stability, and in vivo biodistribution using PET imaging in mice. While these antagonists exhibited strong receptor binding and serum stability, rapid in vivo metabolism limited their potential as PET tracers, highlighting the need for further structural optimization. This study advances the understanding of P2X4 receptor antagonism and underscores the challenges in developing effective PET tracers for this target.
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Affiliation(s)
- Katharina Sophie Erlitz
- European Institute for Molecular Imaging (EIMI), University of Muenster, Roentgenstr. 16, 48149 Muenster, Germany
- Department of Preclinical Imaging and Radiopharmacy, University of Tuebingen, Roentgenweg 1372076Tuebingen, Germany
| | - Ann-Kathrin Prinz
- Institut für Pharmazeutische und Medizinische Chemie der Universität Münster, Corrensstr. 48, 48149 Münster, Germany
| | - Stefan Wagner
- Department of Nuclear Medicine, University Hospital Münster, Albert-Schweitzer-Campus 1, Building A148149 Münster, Germany
| | - Joana Massa
- Institut für Pharmazeutische und Medizinische Chemie der Universität Münster, Corrensstr. 48, 48149 Münster, Germany
| | - Calvin Dunker
- European Institute for Molecular Imaging (EIMI), University of Muenster, Roentgenstr. 16, 48149 Muenster, Germany
- Department of Preclinical Imaging and Radiopharmacy, University of Tuebingen, Roentgenweg 1372076Tuebingen, Germany
| | - Meike Höhl
- European Institute for Molecular Imaging (EIMI), University of Muenster, Roentgenstr. 16, 48149 Muenster, Germany
- Department of Preclinical Imaging and Radiopharmacy, University of Tuebingen, Roentgenweg 1372076Tuebingen, Germany
| | - Angelika Griep
- German Center for Neurodegenerative Diseases (DZNE) within the Helmholtz Association, Venusberg-Campus 1/99, 53127 Bonn, Germany
| | - Róisín M McManus
- German Center for Neurodegenerative Diseases (DZNE) within the Helmholtz Association, Venusberg-Campus 1/99, 53127 Bonn, Germany
| | - Sonja Schelhaas
- European Institute for Molecular Imaging (EIMI), University of Muenster, Roentgenstr. 16, 48149 Muenster, Germany
| | - Oliver Koch
- Institut für Pharmazeutische und Medizinische Chemie der Universität Münster, Corrensstr. 48, 48149 Münster, Germany
| | - Anna Junker
- European Institute for Molecular Imaging (EIMI), University of Muenster, Roentgenstr. 16, 48149 Muenster, Germany
- Department of Preclinical Imaging and Radiopharmacy, University of Tuebingen, Roentgenweg 1372076Tuebingen, Germany
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10
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Kodirov SA, Plakhova VB, Hamill OP, Krylov BV. Long-term spontaneous membrane currents in DRG neurons. J Recept Signal Transduct Res 2025:1-8. [PMID: 40186880 DOI: 10.1080/10799893.2025.2477925] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 04/07/2025]
Abstract
We have experimented with freshly isolated single DRG neurons from neonatal (P0-5) rats to study currents mediated by voltage dependent Na+ (Nav) channels. All experiments were performed using the whole-cell mode of patch-clamp electrophysiology and following the standard steps of this technique. However, in a subgroup of neurons, spontaneous events resembling neurotransmitter release were observed under conditions optimized for whole-cell patch-clamp recordings of INa. All events have a fast rise phase (similar to responses of receptor channels), but decay in a heterogeneous manner. The waveform of the event closely matches that of the response of the purinergic receptor P2X type to ATP. This new activity in neurons was observed at -60 mV and was facilitated during relatively strong hyperpolarization. Although spontaneous fluctuations, termed membrane potential instabilities, are described in DRG neurons, the observed inward currents at more hyperpolarized states are distinct and novel. The spontaneous heterogeneous activities could be relevant to the elucidation of pain mechanisms by distinct pharmacological tools.
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Affiliation(s)
- Sodikdjon A Kodirov
- Pavlov Institute of Physiology, Russian Academy of Sciences, Saint Petersburg, Russia
- Instituto de Medicina Molecular, Universidade de Lisboa, Lisbon, Portugal
- I. P. Pavlov Department of Physiology, Institute of Experimental Medicine, Russian Academy of Medical Sciences, Saint Petersburg, Russia
- Institute of Biophysics, Johannes Kepler University, Linz, Austria
| | - Vera B Plakhova
- Pavlov Institute of Physiology, Russian Academy of Sciences, Saint Petersburg, Russia
| | - Owen P Hamill
- Department of Neuroscience and Cell Biology, UTMB, Galveston, TX, USA
| | - Boris V Krylov
- Pavlov Institute of Physiology, Russian Academy of Sciences, Saint Petersburg, Russia
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11
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Ježek P. Physiological Fatty Acid-Stimulated Insulin Secretion and Redox Signaling Versus Lipotoxicity. Antioxid Redox Signal 2025; 42:566-622. [PMID: 39834189 DOI: 10.1089/ars.2024.0799] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/22/2025]
Abstract
Significance: Type 2 diabetes as a world-wide epidemic is characterized by the insulin resistance concomitant to a gradual impairment of β-cell mass and function (prominently declining insulin secretion) with dysregulated fatty acids (FAs) and lipids, all involved in multiple pathological development. Recent Advances: Recently, redox signaling was recognized to be essential for insulin secretion stimulated with glucose (GSIS), branched-chain keto-acids, and FAs. FA-stimulated insulin secretion (FASIS) is a normal physiological event upon postprandial incoming chylomicrons. This contrasts with the frequent lipotoxicity observed in rodents. Critical Issues: Overfeeding causes FASIS to overlap with GSIS providing repeating hyperinsulinemia, initiates prediabetic states by lipotoxic effects and low-grade inflammation. In contrast the protective effects of lipid droplets in human β-cells counteract excessive lipids. Insulin by FASIS allows FATP1 recruitment into adipocyte plasma membranes when postprandial chylomicrons come late at already low glycemia. Future Directions: Impaired states of pancreatic β-cells and peripheral organs at prediabetes and type 2 diabetes should be revealed, including the inter-organ crosstalk by extracellular vesicles. Details of FA/lipid molecular physiology are yet to be uncovered, such as complex phenomena of FA uptake into cells, postabsorptive inactivity of G-protein-coupled receptor 40, carnitine carrier substrate specificity, the role of carnitine-O-acetyltransferase in β-cells, and lipid droplet interactions with mitochondria. Antioxid. Redox Signal. 42, 566-622.
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Affiliation(s)
- Petr Ježek
- Department of Mitochondrial Physiology, No.75, Institute of Physiology of the Czech Academy of Sciences, Prague, Czech Republic
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12
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Wand PL, Brünings X, Tewari D, Reuter S, Mrowka R, Benndorf K, Zimmer T, Sattler C. Differential Effects of Hearing Loss Mutations in Homomeric P2X2 and Heteromeric P2X2/3 Receptors. Cells 2025; 14:510. [PMID: 40214464 PMCID: PMC11987926 DOI: 10.3390/cells14070510] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/03/2025] [Revised: 03/24/2025] [Accepted: 03/27/2025] [Indexed: 04/14/2025] Open
Abstract
P2X receptors are unspecific cation channels activated by ATP. They are expressed in various tissues and found in neuronal and immune cells. In mammals, seven subunits are described, which can assemble into homomeric and heteromeric trimers. P2X2 receptors play important roles in cochlear adaptation to elevated sound levels. Three mutations causing inherited progressive hearing loss have been identified. These mutations localize to the transmembrane domain 1 (V60L), the transmembrane domain 2 (G353R) and a β-sheet linking the ATP binding site to the pore (D273Y). Herein, mutations were studied in human homomeric P2X2 as well as in heteromeric P2X2/3 receptors. We measured their binding of a fluorescently labeled ATP derivative (fATP) and characterized the constructs using the patch-clamp technique. The conclusions from our results are as follows: 1. The mutations V60L and G353R show robust localization on the plasma membrane and binding of fATP, whereas the mutant D273Y has no binding to fATP. 2. The mutation V60L has an increased affinity to fATP compared with the wildtype. 3. The expression of hP2X2 V60L channels reduces cell viability, which may support its role in the pathogenesis of hearing loss. 4. All mutant P2X2 subunits can assemble into P2X2/3 heteromeric channels with distinct phenotypes.
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Affiliation(s)
- Paula-Luise Wand
- Institute of Physiology II, Jena University Hospital, Friedrich Schiller University Jena, 07743 Jena, Thüringen, Germany; (P.-L.W.); (X.B.); (D.T.); (K.B.); (T.Z.)
| | - Xenia Brünings
- Institute of Physiology II, Jena University Hospital, Friedrich Schiller University Jena, 07743 Jena, Thüringen, Germany; (P.-L.W.); (X.B.); (D.T.); (K.B.); (T.Z.)
| | - Debanjan Tewari
- Institute of Physiology II, Jena University Hospital, Friedrich Schiller University Jena, 07743 Jena, Thüringen, Germany; (P.-L.W.); (X.B.); (D.T.); (K.B.); (T.Z.)
| | - Stefanie Reuter
- Experimentelle Nephrologie, KIM III, Universitätsklinikum Jena, Nonnenplan 4, 07743 Jena, Thüringen, Germany; (S.R.); (R.M.)
- Thimedop, ThIMEDOP—Thüringer Innovationszentrum für Medizintechnik-Lösungen, Nonnenplan 4, UKJ, 07743 Jena, Thüringen, Germany
| | - Ralf Mrowka
- Experimentelle Nephrologie, KIM III, Universitätsklinikum Jena, Nonnenplan 4, 07743 Jena, Thüringen, Germany; (S.R.); (R.M.)
- Thimedop, ThIMEDOP—Thüringer Innovationszentrum für Medizintechnik-Lösungen, Nonnenplan 4, UKJ, 07743 Jena, Thüringen, Germany
| | - Klaus Benndorf
- Institute of Physiology II, Jena University Hospital, Friedrich Schiller University Jena, 07743 Jena, Thüringen, Germany; (P.-L.W.); (X.B.); (D.T.); (K.B.); (T.Z.)
| | - Thomas Zimmer
- Institute of Physiology II, Jena University Hospital, Friedrich Schiller University Jena, 07743 Jena, Thüringen, Germany; (P.-L.W.); (X.B.); (D.T.); (K.B.); (T.Z.)
| | - Christian Sattler
- Institute of Physiology II, Jena University Hospital, Friedrich Schiller University Jena, 07743 Jena, Thüringen, Germany; (P.-L.W.); (X.B.); (D.T.); (K.B.); (T.Z.)
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13
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Barraco M, Kudova E, Bucolo C, Ciranna L, Sortino MA, Chisari M. Cholesterol metabolites modulate ionotropic P2X4 and P2X7 receptor current in microglia cells. Neuropharmacology 2025; 266:110294. [PMID: 39755203 DOI: 10.1016/j.neuropharm.2024.110294] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2024] [Revised: 12/20/2024] [Accepted: 12/30/2024] [Indexed: 01/06/2025]
Abstract
The central nervous system is a well-known steroidogenic tissue producing, among others, cholesterol metabolites such as neuroactive steroids, oxysterols and steroid hormones. It is well known that these endogenous molecules affect several receptor classes, including ionotropic GABAergic and NMDA glutamatergic receptors in neurons. It has been shown that also ionotropic purinergic (P2X) receptors are cholesterol metabolites' targets. Among P2X receptors, purinergic P2X4 and P2X7 receptors are expressed in microglia, the innate immune cells involved in the brain inflammatory response. In this study, we explore the ionotropic purinergic receptors modulation by cholesterol metabolites in microglia. Patch-clamp experiments were performed in BV2 cells, a murine microglia cell line, to evaluate effects of cholesterol metabolites using micro- and nanomolar concentrations. About P2X4 receptor, we found that testosterone butyrate (20 μM and 200 nM) and allopregnanolone (10 μM and 100 nM) both potentiated its current, while neither 25-hydroxycholesterol (10 μM and 100 nM) nor 17β-estradiol (1 μM) showed any effects. On the other hand, P2X7 receptor current was potentiated by allopregnanolone (10 μM) and 25-hydroxycholesterol (10 μM and 100 nM). Taken together, our data show that modulation of either P2X4 and P2X7 current is affected differently by cholesterol metabolites, suggesting a structure-activity relationship among these players. Identifying the possible link between purinergic transmission, microglia and cholesterol metabolites will allow to define new targets for drug development to treat neuroinflammation.
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Affiliation(s)
- Michele Barraco
- Dept. of Biomedical and Biotechnological Sciences, Section of Pharmacology, University of Catania, Catania, Italy
| | - Eva Kudova
- Institute of Organic Chemistry and Biochemistry, Czech Academy of Sciences, Prague, Czech Republic
| | - Claudio Bucolo
- Dept. of Biomedical and Biotechnological Sciences, Section of Pharmacology, University of Catania, Catania, Italy
| | - Lucia Ciranna
- Dept. of Biomedical and Biotechnological Sciences, Physiology, University of Catania, Catania, Italy
| | - Maria Angela Sortino
- Dept. of Biomedical and Biotechnological Sciences, Section of Pharmacology, University of Catania, Catania, Italy
| | - Mariangela Chisari
- Dept. of Biomedical and Biotechnological Sciences, Section of Pharmacology, University of Catania, Catania, Italy.
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14
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Brooke AK, Ojha S, Murrow DP, Ross AE. Purinergic Receptor P2Y1 Modulates Catecholamine Signaling in Murine Mesenteric Lymph Nodes. ACS Chem Neurosci 2025; 16:772-780. [PMID: 39988830 DOI: 10.1021/acschemneuro.4c00435] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/25/2025] Open
Abstract
Neuroimmune communication is crucial for the body's response to physiological challenges, homeostasis, and immune stress response. Adrenergic and purinergic neurotransmission in the sympathetic nervous system is vital for this communication. This study achieves the first co-detection of adenine-based purines and catecholamines in mesenteric lymph nodes via fast-scan cyclic voltammetry. Additionally, we reveal that manipulating an ATP receptor can impact catecholamine signaling in the lymph node for the first time. The G-protein-coupled receptor P2Y1, which controls intracellular Ca2+ levels, was targeted with the antagonist MRS2179. MRS2179 decreased catecholamine concentrations, increased inter-event times, and prolonged event durations. These results suggest that events became smaller, less frequent, and longer-lasting, possibly attributable to decreased intracellular Ca2+ levels. These findings indicate that ATP release in the lymph node can partially regulate norepinephrine signaling, providing mechanistic insight into sympathetic neuronal neurotransmitter control. A deeper understanding of more complicated neuroimmune mechanisms could potentially influence the development of therapeutic strategies in immunology and neurobiology.
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Affiliation(s)
- Alexandra K Brooke
- Department of Chemistry, University of Cincinnati, Cincinnati, Ohio 45221. United States
| | - Sarbeshwar Ojha
- Department of Chemistry, University of Cincinnati, Cincinnati, Ohio 45221. United States
| | - Daniel P Murrow
- Department of Chemistry, University of Cincinnati, Cincinnati, Ohio 45221. United States
| | - Ashley E Ross
- Department of Chemistry, University of Cincinnati, Cincinnati, Ohio 45221. United States
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15
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Mousavikia SN, Matin MM, Bahreyni Tossi MT, Azimian H. Unraveling the role of the P2X7 receptor in cancer radioresistance: Molecular insights and therapeutic implications. BIOCHIMICA ET BIOPHYSICA ACTA. MOLECULAR CELL RESEARCH 2025; 1872:119910. [PMID: 39889832 DOI: 10.1016/j.bbamcr.2025.119910] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/10/2024] [Revised: 01/19/2025] [Accepted: 01/22/2025] [Indexed: 02/03/2025]
Abstract
The P2X7 receptor, a key player in purinergic signaling, is a crucial factor in modulating the response of cancer cells to radiotherapy. The aim of this study was to elucidate the molecular mechanisms by which P2X7 receptor activation contributes to radioresistance in different cancer types. P2X7 receptor signaling influences cellular processes such as DNA damage repair and inflammatory responses, thereby improving tumor survival after radiation exposure. Activation of the P2X7 receptor leads to changes in the tumor microenvironment and promotes an adaptive response that enables cancer cells to resist therapeutic interventions. Therefore, targeting the P2X7 receptor could represent a new therapeutic strategy against cancer. By linking molecular insights with therapeutic implications, this research highlights the P2X7 receptor as a promising target for overcoming radioresistance in cancer therapy and paves the way for novel combination approaches that could significantly improve patient outcomes.
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Affiliation(s)
- Seyedeh Nasibeh Mousavikia
- Department of Medical Physics, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran; Medical Physics Research Center, Basic Sciences Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Maryam M Matin
- Department of Biology, Faculty of Science, Ferdowsi University of Mashhad, Mashhad, Iran; Novel Diagnostics and Therapeutics Research Group, Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran
| | - Mohammad Taghi Bahreyni Tossi
- Medical Physics Research Center, Basic Sciences Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Hosein Azimian
- Department of Medical Physics, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran; Medical Physics Research Center, Basic Sciences Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran.
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16
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Vinhais da Silva AV, Chesseron S, Benouna O, Rollin J, Roger S, Bourguignon T, Chadet S, Ivanes F. P2 purinergic receptors at the heart of pathological left ventricular remodeling following acute myocardial infarction. Am J Physiol Heart Circ Physiol 2025; 328:H550-H564. [PMID: 39884315 DOI: 10.1152/ajpheart.00599.2024] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/30/2024] [Revised: 09/17/2024] [Accepted: 01/15/2025] [Indexed: 02/01/2025]
Abstract
Pathological left ventricular remodeling is a complex process following an acute myocardial infarction, leading to architectural disorganization of the cardiac tissue. This phenomenon is characterized by sterile inflammation and the exaggerated development of fibrotic tissue, which is noncontractile and poorly conductive, responsible for organ dysfunction and heart failure. At present, specific therapies are lacking for both prevention and treatment of this condition, and no biomarkers are currently validated to identify at-risk patients. Physiopathological understanding of this process is limited, probably due to the combination of the multicellular responses involved that are initially necessary for tissue healing but may be detrimental in the longer term. Current research focuses on understanding and modulating the inflammatory response, a key aspect of the tissue healing process. Inflammation is triggered by the release of inflammatory mediators from cardiomyocytes undergoing cell death in the context of ischemia-reperfusion injury. Among them, extracellular ATP is a strong mediator of inflammation through the activation of P2 purinergic receptors, regulating the behavior of all the cellular actors of the postmyocardial infarction response and impacting organ function and recovery. Rather than considering each cellular protagonist independently, this review provides an integrated overview of the inflammatory and tissue response to myocardial infarction by members of the P2 receptor family. Finally, it explores the possibility of reducing pathological left ventricular remodeling through the modulation of these receptors and their associated signaling pathways.
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Affiliation(s)
- Ana Valéria Vinhais da Silva
- Inserm UMR1327 ISCHEMIA Membrane Signalling and Inflammation in reperfusion injuries, Université de Tours, Tours, France
| | - Simon Chesseron
- Inserm UMR1327 ISCHEMIA Membrane Signalling and Inflammation in reperfusion injuries, Université de Tours, Tours, France
| | - Oumnia Benouna
- Inserm UMR1327 ISCHEMIA Membrane Signalling and Inflammation in reperfusion injuries, Université de Tours, Tours, France
| | - Jérôme Rollin
- Inserm UMR1327 ISCHEMIA Membrane Signalling and Inflammation in reperfusion injuries, Université de Tours, Tours, France
- Service d'Hématologie-Hémostase, CHU de Tours, Tours, France
| | - Sébastien Roger
- Inserm UMR1327 ISCHEMIA Membrane Signalling and Inflammation in reperfusion injuries, Université de Tours, Tours, France
| | - Thierry Bourguignon
- Inserm UMR1327 ISCHEMIA Membrane Signalling and Inflammation in reperfusion injuries, Université de Tours, Tours, France
- Service de Chirurgie Cardiaque, CHU de Tours, Tours, France
| | - Stéphanie Chadet
- Inserm UMR1327 ISCHEMIA Membrane Signalling and Inflammation in reperfusion injuries, Université de Tours, Tours, France
| | - Fabrice Ivanes
- Inserm UMR1327 ISCHEMIA Membrane Signalling and Inflammation in reperfusion injuries, Université de Tours, Tours, France
- Service de Cardiologie, CHU de Tours, Tours, France
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17
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Singh U, Bernstein JA. Investigation of the Pathomechanism of Chronic Cough Using an In Vitro Approach. Clin Exp Allergy 2025; 55:247-249. [PMID: 39838624 PMCID: PMC11908833 DOI: 10.1111/cea.14628] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2024] [Revised: 11/29/2024] [Accepted: 01/12/2025] [Indexed: 01/23/2025]
Affiliation(s)
- Umesh Singh
- Department of Internal Medicine, Division of Rheumatology, Allergy and Immunology, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA
| | - Jonathan A Bernstein
- Department of Internal Medicine, Division of Rheumatology, Allergy and Immunology, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA
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18
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da Silva Portilho R, Brito IL, Santos AN, Moreschi BP, de Lucena MN, Otsubo Jaques JA. First evidence of Tityus confluens Borelli, 1899 (Buthidae) venom altering purine metabolism in rat blood cells. Purinergic Signal 2025:10.1007/s11302-025-10076-9. [PMID: 39992594 DOI: 10.1007/s11302-025-10076-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2024] [Accepted: 02/14/2025] [Indexed: 02/26/2025] Open
Abstract
Purinergic signaling pathways play crucial roles in regulating hemostatic and inflammatory responses, both of which are impacted by scorpion envenomation. Scorpion venoms are complex mixtures of various toxins, such as peptides, enzymes, and nucleotides. Previous research showed that the action of scorpion toxins on the purinergic system stems from their effects on purinergic receptors. Additionally, a study identified a putative ectonucleotidase in scorpion venom. This study aimed to investigate the ability of Tityus confluens venom (10, 50, and 100 µg/mL) to metabolize adenine nucleotides and its potential effects on purinergic enzyme activity in rat platelets and lymphocytes. The effects of T. confluens venom on E-NTPDase (ATP and ADP hydrolysis), E-5'-NT (AMP hydrolysis), and E-ADA (ADO hydrolysis) activities were analyzed. The results revealed that crude venom from T. confluens exhibited ATP hydrolysis activity at all tested concentrations. In lymphocytes, ADP hydrolysis was inhibited by 100 µg/mL crude venom, whereas ADO hydrolysis was increased by all venom concentrations. In platelets, ATP hydrolysis was inhibited by 50 and 100 µg/mL crude venom, whereas AMP and ADO hydrolysis were inhibited by all concentrations. When considered collectively, the data suggested an elevation in extracellular ATP levels and a reduction in extracellular ADO. These findings are in alignment with clinical manifestations of scorpion envenomation characterized by a pro-inflammatory milieu. Furthermore, this study demonstrated the intrinsic ATPase activity of T. confluens venom and its ability to modulate E-NTPDase, E-5'-NT, and E-ADA activities in rat blood cells.
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Affiliation(s)
- Romário da Silva Portilho
- Biochemistry Sector, Institute of Biosciences, Federal University of Mato Grosso Do Sul (UFMS), Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil
- Graduate Program in Pharmaceutical Sciences, UFMS, Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil
| | - Igor Leal Brito
- Biochemistry Sector, Institute of Biosciences, Federal University of Mato Grosso Do Sul (UFMS), Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil
- Multicenter Graduate Program in Biochemistry and Molecular Biology, UFMS, Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil
| | - Andreza Negreli Santos
- Biochemistry Sector, Institute of Biosciences, Federal University of Mato Grosso Do Sul (UFMS), Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil
- Multicenter Graduate Program in Biochemistry and Molecular Biology, UFMS, Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil
| | - Bruna Pache Moreschi
- Biochemistry Sector, Institute of Biosciences, Federal University of Mato Grosso Do Sul (UFMS), Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil
- Graduate Program in Pharmaceutical Sciences, UFMS, Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil
| | - Malson Neilson de Lucena
- Biochemistry Sector, Institute of Biosciences, Federal University of Mato Grosso Do Sul (UFMS), Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil
- Multicenter Graduate Program in Biochemistry and Molecular Biology, UFMS, Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil
| | - Jeandre Augusto Otsubo Jaques
- Biochemistry Sector, Institute of Biosciences, Federal University of Mato Grosso Do Sul (UFMS), Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil.
- Graduate Program in Pharmaceutical Sciences, UFMS, Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil.
- Multicenter Graduate Program in Biochemistry and Molecular Biology, UFMS, Campo Grande, Mato Grosso Do Sul, 79070-900, Brazil.
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19
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Bockstiegel J, Engelhardt J, Schuchardt M, Tölle M, Weindl G. The vasoconstrictor adenosine 5'-tetraphosphate is a danger signal that induces IL-1β. Mol Med 2025; 31:72. [PMID: 39984847 PMCID: PMC11844157 DOI: 10.1186/s10020-025-01116-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/24/2024] [Accepted: 02/05/2025] [Indexed: 02/23/2025] Open
Abstract
BACKGROUND The endogenous nucleotide adenosine 5'-tetraphosphate (Ap4) is a potent vasoconstrictor. Despite its structural similarity to the danger signal adenosine 5'-triphosphate (ATP), the immunomodulatory effects of Ap4 remain unclear. METHODS Modulation of interleukin (IL)-1β secretion by Ap4 was studied in both immune cells lines (THP-1, U937) and primary immune cells. Genetic and pharmacological approaches were used to characterize signaling. Cytokine production was measured using ELISA and multiplex assays, while cell viability was determined by MTT and LDH assays. Calcium influx and YO-PRO-1 uptake were assessed via microplate assays and flow cytometry, respectively. RNA sequencing and Western blotting were performed to analyze global gene expression and protein levels. RESULTS We demonstrate that Ap4 stimulates IL-1β release in primed immune cells without affecting the levels of other cytokines, suggesting specificity in its immunomodulatory actions. Mechanistically, Ap4-induced IL-1β release was partially modulated by the P2X7 receptor, a key mediator of inflammation. However, unlike canonical inflammasome activators, this process was independent of potassium efflux, the NLRP3 inflammasome, and caspase-1. Ap4 specifically increased LDH release in macrophages irrespective of priming. Furthermore, Ap4-mediated calcium influx, crucial for immune cell activation, predominantly occurred through P2Y receptors rather than P2X7 receptors. Transcriptomic analysis highlighted Ap4-induced upregulation of metallothioneins, implicating metal ion homeostasis in Ap4-mediated responses. CONCLUSIONS Collectively, our findings suggest Ap4 as a novel pro-inflammatory mediator capable of inducing IL-1β release in innate immune cells through distinct mechanisms from classical NLRP3 inflammasome activators, shedding light on its potential role in inflammatory diseases and vascular disorders.
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Affiliation(s)
- Judith Bockstiegel
- Pharmaceutical Institute, Pharmacology and Toxicology Section, University of Bonn, Gerhard- Domagk-Str. 3, 53121, Bonn, Germany
| | - Jonas Engelhardt
- Pharmaceutical Institute, Pharmacology and Toxicology Section, University of Bonn, Gerhard- Domagk-Str. 3, 53121, Bonn, Germany
| | - Mirjam Schuchardt
- Department of Nephrology and Medical Intensive Care, Charité-Universitätsmedizin Berlin, Freie Universität Berlin, Humboldt Universität zu Berlin, Hindenburgdamm 30, 12203, Berlin, Germany
- Medical School Berlin, Faculty of Medicine, Rüdesheimer Straße 50, 14513, Berlin, Germany
| | - Markus Tölle
- Department of Nephrology and Medical Intensive Care, Charité-Universitätsmedizin Berlin, Freie Universität Berlin, Humboldt Universität zu Berlin, Hindenburgdamm 30, 12203, Berlin, Germany
| | - Günther Weindl
- Pharmaceutical Institute, Pharmacology and Toxicology Section, University of Bonn, Gerhard- Domagk-Str. 3, 53121, Bonn, Germany.
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20
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Yu H, Ren K, Jin Y, Zhang L, Liu H, Huang Z, Zhang Z, Chen X, Yang Y, Wei Z. Mitochondrial DAMPs: Key mediators in neuroinflammation and neurodegenerative disease pathogenesis. Neuropharmacology 2025; 264:110217. [PMID: 39557152 DOI: 10.1016/j.neuropharm.2024.110217] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/17/2024] [Revised: 11/02/2024] [Accepted: 11/13/2024] [Indexed: 11/20/2024]
Abstract
Neurodegenerative diseases such as Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD), and amyotrophic lateral sclerosis (ALS) are increasingly linked to mitochondrial dysfunction and neuroinflammation. Central to this link are mitochondrial damage-associated molecular patterns (mtDAMPs), including mitochondrial DNA, ATP, and reactive oxygen species, released during mitochondrial stress or damage. These mtDAMPs activate inflammatory pathways, such as the NLRP3 inflammasome and cGAS-STING, contributing to the progression of neurodegenerative diseases. This review delves into the mechanisms by which mtDAMPs drive neuroinflammation and discusses potential therapeutic strategies targeting these pathways to mitigate neurodegeneration. Additionally, it explores the cross-talk between mitochondria and the immune system, highlighting the complex interplay that exacerbates neuronal damage. Understanding the role of mtDAMPs could pave the way for novel treatments aimed at modulating neuroinflammation and slowing disease progression, ultimately improving patient outcome.
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Affiliation(s)
- Haihan Yu
- Department of Neurology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, 450052, PR China
| | - Kaidi Ren
- Department of Pharmacy, the First Affiliated Hospital of Zhengzhou University, Zhengzhou, 450052, PR China
| | - Yage Jin
- Department of Cardiology, the First Affiliated Hospital of Zhengzhou University, Zhengzhou, 450052, PR China
| | - Li Zhang
- Key Clinical Laboratory of Henan Province, Department of Clinical Laboratory, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, 450052, PR China
| | - Hui Liu
- Henan Key Laboratory of Immunology and Targeted Drug, Henan Collaborative Innovation Center of Molecular Diagnosis and Laboratory Medicine, School of Medical Technology, Xinxiang Medical University, Xinxiang, 453003, PR China
| | - Zhen Huang
- Henan Key Laboratory of Immunology and Targeted Drug, Henan Collaborative Innovation Center of Molecular Diagnosis and Laboratory Medicine, School of Medical Technology, Xinxiang Medical University, Xinxiang, 453003, PR China
| | - Ziheng Zhang
- College of Life Sciences, Xinjiang University, Urumqi, Xinjiang, 830046, PR China
| | - Xing Chen
- Clinical Systems Biology Laboratories, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, 450052, PR China.
| | - Yang Yang
- Clinical Systems Biology Laboratories, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, 450052, PR China.
| | - Ziqing Wei
- Department of Neurology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, 450052, PR China.
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21
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Oparin P, Khokhlova O, Cherkashin A, Nadezhdin K, Palikov V, Palikova Y, Korolkova Y, Mosharova I, Rogachevskaja O, Baranov M, Shaidullova K, Ermakova E, Lushpa V, Bruter A, Deykin A, Ivanova E, Silaeva Y, Dyachenko I, Bocharov E, Sitdikova G, Andreev-Andrievskiy A, Poteryaev D, Shuster A, Murashev A, Kolesnikov S, Stepanenko V, Grishin E, Vassilevski A. Potent painkiller from spider venom antagonizes P2X3 receptors without dysgeusia. Mol Ther 2025; 33:771-785. [PMID: 39960544 PMCID: PMC11852983 DOI: 10.1016/j.ymthe.2024.12.036] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/24/2023] [Revised: 06/15/2024] [Accepted: 12/27/2024] [Indexed: 02/28/2025] Open
Abstract
P2X3 receptors are a validated molecular target in pain syndromes and chronic cough. Known P2X3 inhibitors generally suffer from poor selectivity and efficacy. Taking advantage of peptide combinatorial libraries found in venoms, we describe a P2X3 antagonist from the crab spider Thomisus onustus. This peptide potently inhibits P2X3 in the dorsal root and trigeminal ganglia neurons of rodents, as well as recombinant human P2X3, showing no effect on P2X2 or P2X2/3 receptors. PT6 presents a compact and rigid structure and produces pronounced antinociception in animal models of inflammatory and neuropathic pain at low doses (0.01-0.1 mg/kg subcutaneously). It does not show antinociceptive activity in P2rx3-knockout mice, providing further evidence in favor of its specificity. Importantly, PT6 shows no dysgeusia or ageusia effects, notoriously characteristic of small-molecule P2X3 ligands, and therefore stands out as an attractive hit for analgesic drug discovery.
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Affiliation(s)
- Peter Oparin
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia; Future Analgesics Ltd, Moscow 123060, Russia
| | - Oksana Khokhlova
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia
| | - Aleksandr Cherkashin
- Institute of Cell Biophysics, Russian Academy of Sciences, Federal Research Center "Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences", Pushchino 142290, Moscow Region, Russia
| | - Kirill Nadezhdin
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia
| | - Victor Palikov
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia
| | - Yulia Palikova
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia
| | - Yuliya Korolkova
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia
| | - Irina Mosharova
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia
| | - Olga Rogachevskaja
- Institute of Cell Biophysics, Russian Academy of Sciences, Federal Research Center "Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences", Pushchino 142290, Moscow Region, Russia
| | - Mikhail Baranov
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia
| | - Ksenia Shaidullova
- Institute of Fundamental Medicine and Biology, Kazan Federal University, Kazan 420012, Russia
| | - Elizaveta Ermakova
- Institute of Fundamental Medicine and Biology, Kazan Federal University, Kazan 420012, Russia
| | - Vladislav Lushpa
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia
| | - Alexandra Bruter
- Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Institute of Gene Biology, Russian Academy of Sciences, Moscow 119334, Russia
| | - Alexey Deykin
- Joint Center for Genetic Technology, Belgorod State University, Belgorod 308015, Russia
| | - Elena Ivanova
- Zakusov Institute of Pharmacology, Moscow 125315, Russia
| | - Yulia Silaeva
- Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Institute of Gene Biology, Russian Academy of Sciences, Moscow 119334, Russia
| | - Igor Dyachenko
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia
| | - Eduard Bocharov
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia; Moscow Institute of Physics and Technology (National Research University), Dolgoprudny 141701, Moscow Region, Russia
| | - Guzel Sitdikova
- Institute of Fundamental Medicine and Biology, Kazan Federal University, Kazan 420012, Russia
| | - Alexander Andreev-Andrievskiy
- M.V. Lomonosov Moscow State University, Moscow 119991, Russia; Institute for Biomedical Problems, Russian Academy of Sciences, Moscow 123007, Russia
| | | | | | - Arkady Murashev
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia
| | - Stanislav Kolesnikov
- Institute of Cell Biophysics, Russian Academy of Sciences, Federal Research Center "Pushchino Scientific Center for Biological Research of the Russian Academy of Sciences", Pushchino 142290, Moscow Region, Russia
| | - Vasiliy Stepanenko
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia
| | - Eugene Grishin
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia; Future Analgesics Ltd, Moscow 123060, Russia
| | - Alexander Vassilevski
- Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia; Future Analgesics Ltd, Moscow 123060, Russia; Moscow Institute of Physics and Technology (National Research University), Dolgoprudny 141701, Moscow Region, Russia.
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22
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Carvalho-Barbosa N, Zeidler JD, Savio LEB, Coutinho-Silva R. Purinergic signaling in the battlefield of viral infections. Purinergic Signal 2025; 21:83-98. [PMID: 38038801 PMCID: PMC11958901 DOI: 10.1007/s11302-023-09981-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/28/2023] [Accepted: 11/19/2023] [Indexed: 12/02/2023] Open
Abstract
Purinergic signaling has been associated with immune defenses against pathogens such as bacteria, protozoa, fungi, and viruses, acting as a sentinel system that signals to the cells when a threat is present. This review focuses on the roles of purinergic signaling and its therapeutic potential for viral infections. In this context, the purinergic system may play potent antiviral roles by boosting interferon signaling. In other cases, though, it can contribute to a hyperinflammatory response and disease severity, resulting in poor outcomes, such as during flu and potentially COVID-19. Lastly, a third situation may occur since viruses are obligatory intracellular parasites that hijack the host cell machinery for their infection and replication. Viruses such as HIV-1 use the purinergic system to favor their infection and persistence within the host cell. Therefore, understanding the particular nuances of purinergic signaling in each viral infection may contribute to designing proper therapeutic strategies to treat viral diseases.
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Affiliation(s)
- Nayara Carvalho-Barbosa
- Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Edifício do Centro de Ciências da Saúde, Bloco G. Av. Carlos Chagas Filho, 373. Cidade Universitária, Ilha do Fundão, Rio de Janeiro, RJ, 21941-902, Brazil
| | - Julianna Dias Zeidler
- Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Edifício do Centro de Ciências da Saúde, Bloco G. Av. Carlos Chagas Filho, 373. Cidade Universitária, Ilha do Fundão, Rio de Janeiro, RJ, 21941-902, Brazil
| | - Luiz Eduardo Baggio Savio
- Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Edifício do Centro de Ciências da Saúde, Bloco G. Av. Carlos Chagas Filho, 373. Cidade Universitária, Ilha do Fundão, Rio de Janeiro, RJ, 21941-902, Brazil
| | - Robson Coutinho-Silva
- Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Edifício do Centro de Ciências da Saúde, Bloco G. Av. Carlos Chagas Filho, 373. Cidade Universitária, Ilha do Fundão, Rio de Janeiro, RJ, 21941-902, Brazil.
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23
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Ho VR, Goss GG, Leys SP. ATP and glutamate coordinate contractions in the freshwater sponge Ephydatia muelleri. J Exp Biol 2025; 228:JEB248010. [PMID: 39936310 PMCID: PMC11883242 DOI: 10.1242/jeb.248010] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/30/2024] [Accepted: 12/20/2024] [Indexed: 02/13/2025]
Abstract
Sponges (phylum Porifera) are an early diverging animal lineage without nervous and muscular systems, and yet they are able to produce coordinated whole-body contractions in response to disturbances. Little is known about the underlying signalling mechanisms in coordinating such responses. Previous studies demonstrated that sponges respond specifically to chemicals such as l-glutamate and γ-amino-butyric acid (GABA), which trigger and prevent contractions, respectively. Genes for purinergic P2X-like receptors are present in several sponge genomes, leading us to ask whether ATP works with glutamate to coordinate contractions in sponges as it does in other animal nervous systems. Using pharmacological approaches on the freshwater sponge Ephydatia muelleri, we show that ATP is involved in coordinating contractions. Bath application of ATP caused a rapid, sustained expansion of the excurrent canals in a dose-dependent manner. Complete contractions occurred when ATP was added in the presence of apyrase, an enzyme that hydrolyses ATP. Application of ADP, the first metabolic product of ATP hydrolysis, triggered complete contractions, whereas AMP, the subsequent metabolite, did not trigger a response. Blocking ATP from binding and activating P2X receptors with pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) prevented both glutamate- and ATP-triggered contractions, suggesting that ATP works downstream of glutamate. Bioinformatic analysis revealed two P2X receptor sequences, one of which groups with other vertebrate P2X receptors. Altogether, our results confirm that purinergic signalling by ATP is involved in coordinating contractions in the freshwater sponge.
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Affiliation(s)
- Vanessa R. Ho
- Department of Biological Sciences, University of Alberta, Edmonton, AB, Canada, T6G 2R3
| | - Greg G. Goss
- Department of Biological Sciences, University of Alberta, Edmonton, AB, Canada, T6G 2R3
| | - Sally P. Leys
- Department of Biological Sciences, University of Alberta, Edmonton, AB, Canada, T6G 2R3
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24
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Nikolenko VN, Proshkina AS, Zharikov YO, Rizaeva NA, Lapina TA, Sharifullina PA, Aniskin DA, Zharikova TS, Pontes-Silva A, Oganesyan MV. Vestibular Otolithic Receptors: Association Between Molecular Mechanism of Degeneration and Benign Paroxysmal Positional Vertigo. Curr Behav Neurosci Rep 2025; 12:6. [DOI: 10.1007/s40473-025-00299-w] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 01/20/2025] [Indexed: 05/03/2025]
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25
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Shi H, Ditter IA, Oken AC, Mansoor SE. Human P2X4 receptor gating is modulated by a stable cytoplasmic cap and a unique allosteric pocket. SCIENCE ADVANCES 2025; 11:eadr3315. [PMID: 39823330 PMCID: PMC11740937 DOI: 10.1126/sciadv.adr3315] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/26/2024] [Accepted: 12/17/2024] [Indexed: 01/19/2025]
Abstract
P2X receptors (P2XRs) are adenosine 5'-triphosphate (ATP)-gated ion channels comprising homomeric and heteromeric trimers of seven subtypes (P2X1-P2X7) that confer different rates of desensitization. The helical recoil model of P2XR desensitization proposes stability of the cytoplasmic cap sets the rate of desensitization, but timing of its formation is unclear for slow-desensitizing P2XRs. We report cryo-electron microscopy structures of full-length wild-type human P2X4 receptor in apo closed, antagonist-bound inhibited, and ATP-bound desensitized states. Because the apo closed and antagonist-bound inhibited state structures of this slow-desensitizing P2XR include an intact cytoplasmic cap while the ATP-bound desensitized state structure does not, the cytoplasmic cap is formed before agonist binding. Furthermore, structural and functional data suggest the cytoplasmic cap is stabilized by lipids to modulate desensitization, and P2X4 is modified by glycosylation and palmitoylation. Last, our antagonist-bound inhibited state structure reveals features specific to the allosteric ligand-binding pocket in human receptors that facilitates development of small-molecule modulators.
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Affiliation(s)
- Haoyuan Shi
- Department of Chemical Physiology & Biochemistry, Oregon Health & Science University, Portland, OR 97239, USA
| | - Ismayn A. Ditter
- Department of Chemical Physiology & Biochemistry, Oregon Health & Science University, Portland, OR 97239, USA
| | - Adam C. Oken
- Department of Chemical Physiology & Biochemistry, Oregon Health & Science University, Portland, OR 97239, USA
| | - Steven E. Mansoor
- Department of Chemical Physiology & Biochemistry, Oregon Health & Science University, Portland, OR 97239, USA
- Division of Cardiovascular Medicine, Knight Cardiovascular Institute, Oregon Health & Science University, Portland, OR 97239, USA
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26
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Li D, Dai Y, Li Z, Bi H, Li H, Wang Y, Liu Y, Tian X, Chen L. Resveratrol Upregulates miR-124-3p Expression to Target DAPK1, Regulating the NLRP3/Caspase-1/GSDMD Pathway to Inhibit Pyroptosis and Alleviate Spinal Cord Injury. J Cell Mol Med 2025; 29:e70338. [PMID: 39833100 PMCID: PMC11745821 DOI: 10.1111/jcmm.70338] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/28/2024] [Revised: 11/29/2024] [Accepted: 12/23/2024] [Indexed: 01/30/2025] Open
Abstract
Currently, an effective treatment for spinal cord injury (SCI) is not available. Due to the irreversible primary injury associated with SCI, the prevention and treatment of secondary injury are very important. In the secondary injury stage, pyroptosis exacerbates the deterioration of the spinal cord injury, and inhibiting pyroptosis is beneficial for recovery from SCI. The aim of this study was to clarify the role of resveratrol (RES) and the antipyroptotic mechanisms of RES and miR-124-3p in SCI to lay a theoretical foundation for the clinical treatment of SCI and provide new therapeutic approaches. Using cell staining and related molecular protein detection techniques to assess DAPK1, the effects of miR-124-3p and RES on pyroptosis were investigated, and the effects of RES on injured spinal cord repair in rats were evaluated using tissue staining and related functional recovery experiments. In vitro, DAPK1 interacts with NLRP3, exerting a pyroptotic effect through the NLRP3/Caspase-1/GSDMD pathway and DAPK1 knockdown inhibits pyroptosis. miR-124-3P negatively regulates the level of DAPK1 and reduced cell pyroptosis. RES increased miR-124-3p expression and reduces DAPK1 expression, affecting the NLRP3/Caspase-1/GSDMD pathway and inhibiting pyroptosis. In vivo, RES reduces GSDMD-N levels in rats with SCI, promotes functional recovery, and thus promotes recovery from SCI. Therefore, we concluded that RES increases the level of miR-124-3p, which targets DAPK1, regulates the NLRP3/Caspase-1/GSDMD pathway, inhibits pyroptosis and alleviates SCI.
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Affiliation(s)
- Daohui Li
- Department of OrthopedicsThe First Affiliated Hospital of Kunming Medical UniversityYunnanChina
| | - Yongwen Dai
- Department of OrthopedicsThe First Affiliated Hospital of Kunming Medical UniversityYunnanChina
| | - Zhengtao Li
- Department of OrthopedicsThe First Affiliated Hospital of Kunming Medical UniversityYunnanChina
| | - Hangchuan Bi
- Department of OrthopedicsThe First Affiliated Hospital of Kunming Medical UniversityYunnanChina
| | - Haotian Li
- Department of OrthopedicsThe First Affiliated Hospital of Kunming Medical UniversityYunnanChina
| | - Yongquan Wang
- Department of OrthopedicsThe First Affiliated Hospital of Kunming Medical UniversityYunnanChina
| | - Yuan Liu
- Department of OrthopedicsThe First Affiliated Hospital of Kunming Medical UniversityYunnanChina
| | - Xinpeng Tian
- Department of Critical Care MedicineXi Chang People's HospitalSichuanChina
| | - Lingqiang Chen
- Department of OrthopedicsThe First Affiliated Hospital of Kunming Medical UniversityYunnanChina
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27
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Verkhratsky A, Semyanov A. Physiology of neuroglia of the central nervous system. HANDBOOK OF CLINICAL NEUROLOGY 2025; 209:69-91. [PMID: 40122632 DOI: 10.1016/b978-0-443-19104-6.00005-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 03/25/2025]
Abstract
Neuroglia of the central nervous system (CNS) are a diverse and highly heterogeneous population of cells of ectodermal, neuroepithelial origin (macroglia, that includes astroglia and oligodendroglia) and mesodermal, myeloid origin (microglia). Neuroglia are primary homeostatic cells of the CNS, responsible for the support, defense, and protection of the nervous tissue. The extended class of astroglia (which includes numerous parenchymal astrocytes, such as protoplasmic, fibrous, velate, marginal, etc., radial astrocytes such as Bergmann glia, Muller glia, etc., and ependymoglia lining the walls of brain ventricles and central canal of the spinal cord) is primarily responsible for overall homeostasis of the nervous tissue. Astroglial cells control homeostasis of ions, neurotransmitters, hormones, metabolites, and are responsible for neuroprotection and defense of the CNS. Oligodendroglia provide for myelination of axons, hence supporting and sustaining CNS connectome. Microglia are tissue macrophages adapted to the CNS environment which contribute to the host of physiologic functions including regulation of synaptic connectivity through synaptic pruning, regulation of neurogenesis, and even modifying neuronal excitability. Neuroglial cells express numerous receptors, transporters, and channels that allow neuroglia to perceive and follow neuronal activity. Activation of these receptors triggers intracellular ionic signals that govern various homeostatic cascades underlying glial supportive and defensive capabilities. Ionic signaling therefore represents the substrate of glial excitability.
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Affiliation(s)
- Alexei Verkhratsky
- Faculty of Biology, Medicine and Health, University of Manchester, Manchester, United Kingdom; Department of Neurosciences, University of the Basque Country UPV/EHU and CIBERNED, Leioa, Bizkaia, Spain; IKERBASQUE, Basque Foundation for Science, Bilbao, Spain
| | - Alexey Semyanov
- Department of Physiology, Jiaxing University College of Medicine, Jiaxing, Zhejiang, China
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28
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Chen YH, Lin S, Jin SY, Gao TM. Extracellular ATP Is a Homeostatic Messenger That Mediates Cell-Cell Communication in Physiological Processes and Psychiatric Diseases. Biol Psychiatry 2025; 97:41-53. [PMID: 38679359 DOI: 10.1016/j.biopsych.2024.04.013] [Citation(s) in RCA: 3] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/03/2023] [Revised: 03/14/2024] [Accepted: 04/15/2024] [Indexed: 05/01/2024]
Abstract
Neuronal activity is the basis of information encoding and processing in the brain. During neuronal activation, intracellular ATP (adenosine triphosphate) is generated to meet the high-energy demands. Simultaneously, ATP is secreted, increasing the extracellular ATP concentration and acting as a homeostatic messenger that mediates cell-cell communication to prevent aberrant hyperexcitability of the nervous system. In addition to the confined release and fast synaptic signaling of classic neurotransmitters within synaptic clefts, ATP can be released by all brain cells, diffuses widely, and targets different types of purinergic receptors on neurons and glial cells, making it possible to orchestrate brain neuronal activity and participate in various physiological processes, such as sleep and wakefulness, learning and memory, and feeding. Dysregulation of extracellular ATP leads to a destabilizing effect on the neural network, as found in the etiopathology of many psychiatric diseases, including depression, anxiety, schizophrenia, and autism spectrum disorder. In this review, we summarize advances in the understanding of the mechanisms by which extracellular ATP serves as an intercellular signaling molecule to regulate neural activity, with a focus on how it maintains the homeostasis of neural networks. In particular, we also focus on neural activity issues that result from dysregulation of extracellular ATP and propose that aberrant levels of extracellular ATP may play a role in the etiopathology of some psychiatric diseases, highlighting the potential therapeutic targets of ATP signaling in the treatment of these psychiatric diseases. Finally, we suggest potential avenues to further elucidate the role of extracellular ATP in intercellular communication and psychiatric diseases.
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Affiliation(s)
- Yi-Hua Chen
- State Key Laboratory of Organ Failure Research, Key Laboratory of Mental Health of the Ministry of Education, Guangdong-Hong Kong-Macao Greater Bay Area Center for Brain Science and Brain-Inspired Intelligence, Guangdong-Hong Kong Joint Laboratory for Psychiatric Disorders, Guangdong Province Key Laboratory of Psychiatric Disorders, Guangdong Basic Research Center of Excellence for Integrated Traditional and Western Medicine for Qingzhi Diseases, Department of Neurobiology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, China
| | - Song Lin
- Department of Physiology, School of Medicine, Jinan University, Guangzhou, China
| | - Shi-Yang Jin
- State Key Laboratory of Organ Failure Research, Key Laboratory of Mental Health of the Ministry of Education, Guangdong-Hong Kong-Macao Greater Bay Area Center for Brain Science and Brain-Inspired Intelligence, Guangdong-Hong Kong Joint Laboratory for Psychiatric Disorders, Guangdong Province Key Laboratory of Psychiatric Disorders, Guangdong Basic Research Center of Excellence for Integrated Traditional and Western Medicine for Qingzhi Diseases, Department of Neurobiology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, China
| | - Tian-Ming Gao
- State Key Laboratory of Organ Failure Research, Key Laboratory of Mental Health of the Ministry of Education, Guangdong-Hong Kong-Macao Greater Bay Area Center for Brain Science and Brain-Inspired Intelligence, Guangdong-Hong Kong Joint Laboratory for Psychiatric Disorders, Guangdong Province Key Laboratory of Psychiatric Disorders, Guangdong Basic Research Center of Excellence for Integrated Traditional and Western Medicine for Qingzhi Diseases, Department of Neurobiology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, China.
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Magham S, Kumar ML, Krishnamurthy PT, Shaji N, Ramakkamma AR. Purinergic Receptor (P2X7R): A Promising Anti-Parkinson's Drug Target. Adv Pharm Bull 2024; 14:807-818. [PMID: 40190682 PMCID: PMC11970502 DOI: 10.34172/apb.43206] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/28/2024] [Revised: 11/16/2024] [Accepted: 12/03/2024] [Indexed: 04/09/2025] Open
Abstract
Purpose Parkinson's disease (PD) is the fourth most common neurodegenerative disorder, characterized by degeneration of basal ganglia and a decrease in dopamine levels in the brain. Purinergic 2X7 receptors (P2X7Rs) serve as inflammation gatekeepers. They are found in both central and peripheral nervous systems (CNS & PNS), and are activated in glial cells during inflammation. Purinergic 2X receptors (P2XRs) have been extensively studied in recent decades, particularly P2X7R, because of their important role in neuroinflammation caused by selective overexpression in glial cells. As P2X7R and its selective antagonists may provide neuroprotection by preventing the release of inflammatory mediators such as IL-1, they have become a research focus in PD. The review covers structure, signalling, molecular mechanisms, neuroprotective role, and current developments of P2X7R antagonists in PD. Methods A systematic analysis and review of the potential prospects of P2X7R antagonists in the treatment of PD were conducted by analyzing existing research data and reports published between 1996 and present. Results There is a substantial body of evidence linking P2X7R to pathology of PD. As a result, P2X7R antagonists may have therapeutic potential in treatment of PD. Conclusion P2X7R has been demonstrated as an efficacious target in PD. Recent advances in rational drug design have paved the way for development of therapeutically valuable P2X7R antagonists such as adamantyl cyanoguanides, small molecular weight compounds, and PET ligands for the treatment of PD. However, the exact molecular mechanism and therapeutic potential of P2X7R antagonists in treatment of PD are yet to be fully explored.
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Affiliation(s)
- Saivarshini Magham
- Department of Pharmacology, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Ooty-643001, The Nilgiris, Tamil Nadu, India
| | - M. Lalith Kumar
- Department of Pharmacology, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Ooty-643001, The Nilgiris, Tamil Nadu, India
| | - Praveen Thaggikuppe Krishnamurthy
- Department of Pharmacology, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Ooty-643001, The Nilgiris, Tamil Nadu, India
| | - Neenu Shaji
- Department of Pharmacology, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Ooty-643001, The Nilgiris, Tamil Nadu, India
| | - Aishwarya Reddy Ramakkamma
- Department of Pharmaceutics, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Ooty-643001, The Nilgiris, Tamil Nadu, India
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Domingos LB, Silva Júnior AFD, Diniz CRAF, Rosa J, Terzian ALB, Resstel LBM. P2X7 receptors modulate acquisition of cue fear extinction and contextual background memory generalization in male mice. Neuropharmacology 2024; 261:110177. [PMID: 39366651 DOI: 10.1016/j.neuropharm.2024.110177] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/17/2024] [Revised: 09/22/2024] [Accepted: 10/01/2024] [Indexed: 10/06/2024]
Abstract
The purinergic P2X7 receptors (P2X7R) are activated by adenosine triphosphate (ATP) in several brain regions, particularly those involved with emotional control and the regulation of fear-related memories. Here, we investigate the role of P2X7R in fear learning memory, specifically in the acquisition and consolidation phases of the cued fear conditioning paradigm. C57Bl/6 wildtype (WT) male mice that received a single i.p. injection of the selective P2X7R antagonist A438079 prior the conditioning session showed generalization of cued fear memory and impaired fear extinction recall in the test session, while those treated prior the extinction session exhibited a similar behavior profile accompanied by resistance in the extinction learning. However, no effects were observed when this drug was administered immediately after the conditioning, extinction, or before the test session. Our results with P2X7R knockout (P2X7 KO) mice showed a behavioral profile that mirrored the collective effects observed across all pharmacological treatment conditions. This suggests that the P2X7R KO model effectively replicates the behavioral changes induced by the pharmacological interventions, demonstrating that we have successfully isolated the role of P2X7R in the fear and extinction phases of memory. These findings highlight the role of P2X7R in the acquisition and recall of extinction memory and supports P2X7R as a promising candidate for controlling abnormal fear processing, with potential applications for stress exposure-related disorders such as post-traumatic stress disorder (PTSD).
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Affiliation(s)
- Luana Barreto Domingos
- Department of Pharmacology, School of Medicine of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil; Department of Biomedicine, Aarhus University, Aarhus, Denmark; Translational Neuropsychiatry Unit, Aarhus University, Denmark
| | | | - Cassiano Ricardo Alves Faria Diniz
- Department of Pharmacology, School of Medicine of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil; Center for Neuroscience, University of California, Davis, CA, USA
| | | | - Ana Luisa B Terzian
- Department of Pharmacology, School of Medicine of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil
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Santoro A, De Santis S, Palmieri F, Vozza A, Agrimi G, Andolfo I, Russo R, Palazzo A, Storlazzi CT, Ferrucci A, Jun YW, Kool ET, Fiermonte G, Iolascon A, Paradies E, Marobbio CMT, Palmieri L. P2 Receptor Antagonists Rescue Defective Heme Content in an In Vitro SLC25A38-Associated Congenital Sideroblastic Anemia Cell Model. Int J Mol Sci 2024; 25:13314. [PMID: 39769087 PMCID: PMC11679772 DOI: 10.3390/ijms252413314] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2024] [Revised: 12/02/2024] [Accepted: 12/04/2024] [Indexed: 01/30/2025] Open
Abstract
Mutations in the SLC25A38 gene are responsible for the second most common form of congenital sideroblastic anemia (CSA), a severe condition for which no effective treatment exists. We developed and characterized a K562 erythroleukemia cell line with markedly reduced expression of the SLC25A38 protein (A38-low cells). This model successfully recapitulated the main features of CSA, including reduced heme content and mitochondrial respiration, increase in mitochondrial iron, ROS levels and sensitivity to oxidative stress. Notably, our study uncovered a new role for extracellular pyridoxal 5'-phosphate (PLP) and other P2 receptor antagonists in rescuing the altered parameters of A38-low cells (for example, the heme content of the A38-low cells was increased from about 50% to about 80% by the P2 receptor antagonists treatment compared with the value of the controls). These findings suggest that targeting P2 receptors could represent a promising therapeutic approach for SLC25A38-associated CSA.
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Affiliation(s)
- Antonella Santoro
- Department of Biosciences, Biotechnology and Environment, University of Bari Aldo Moro, 70125 Bari, Italy; (A.S.); (S.D.S.); (F.P.); (A.V.); (G.A.); (A.P.); (C.T.S.); (G.F.); (L.P.)
| | - Silvia De Santis
- Department of Biosciences, Biotechnology and Environment, University of Bari Aldo Moro, 70125 Bari, Italy; (A.S.); (S.D.S.); (F.P.); (A.V.); (G.A.); (A.P.); (C.T.S.); (G.F.); (L.P.)
| | - Ferdinando Palmieri
- Department of Biosciences, Biotechnology and Environment, University of Bari Aldo Moro, 70125 Bari, Italy; (A.S.); (S.D.S.); (F.P.); (A.V.); (G.A.); (A.P.); (C.T.S.); (G.F.); (L.P.)
- CNR Institute of Biomembranes, Bioenergetics and Molecular Biotechnologies (IBIOM), 70126 Bari, Italy;
- Center of Excellence in Comparative Genomics, University of Bari Aldo Moro, 70125 Bari, Italy
| | - Angelo Vozza
- Department of Biosciences, Biotechnology and Environment, University of Bari Aldo Moro, 70125 Bari, Italy; (A.S.); (S.D.S.); (F.P.); (A.V.); (G.A.); (A.P.); (C.T.S.); (G.F.); (L.P.)
| | - Gennaro Agrimi
- Department of Biosciences, Biotechnology and Environment, University of Bari Aldo Moro, 70125 Bari, Italy; (A.S.); (S.D.S.); (F.P.); (A.V.); (G.A.); (A.P.); (C.T.S.); (G.F.); (L.P.)
| | - Immacolata Andolfo
- Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, 80131 Naples, Italy; (I.A.); (R.R.); (A.I.)
- CEINGE-Biotecnologie Avanzate Franco Salvatore, 80145 Naples, Italy
| | - Roberta Russo
- Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, 80131 Naples, Italy; (I.A.); (R.R.); (A.I.)
- CEINGE-Biotecnologie Avanzate Franco Salvatore, 80145 Naples, Italy
| | - Antonio Palazzo
- Department of Biosciences, Biotechnology and Environment, University of Bari Aldo Moro, 70125 Bari, Italy; (A.S.); (S.D.S.); (F.P.); (A.V.); (G.A.); (A.P.); (C.T.S.); (G.F.); (L.P.)
| | - Clelia Tiziana Storlazzi
- Department of Biosciences, Biotechnology and Environment, University of Bari Aldo Moro, 70125 Bari, Italy; (A.S.); (S.D.S.); (F.P.); (A.V.); (G.A.); (A.P.); (C.T.S.); (G.F.); (L.P.)
| | - Arianna Ferrucci
- CNR Institute of Biomembranes, Bioenergetics and Molecular Biotechnologies (IBIOM), 70126 Bari, Italy;
| | - Yong Woong Jun
- Department of Chemistry, Korea Advanced Institute Science and Technology (KAIST), Daejeon 34141, Republic of Korea;
| | - Eric T. Kool
- Department of Chemistry, Stanford University, Stanford, CA 94305, USA;
- Sarafan ChEM-H Institute, Stanford University, Stanford, CA 94305, USA
| | - Giuseppe Fiermonte
- Department of Biosciences, Biotechnology and Environment, University of Bari Aldo Moro, 70125 Bari, Italy; (A.S.); (S.D.S.); (F.P.); (A.V.); (G.A.); (A.P.); (C.T.S.); (G.F.); (L.P.)
| | - Achille Iolascon
- Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, 80131 Naples, Italy; (I.A.); (R.R.); (A.I.)
- CEINGE-Biotecnologie Avanzate Franco Salvatore, 80145 Naples, Italy
| | - Eleonora Paradies
- CNR Institute of Biomembranes, Bioenergetics and Molecular Biotechnologies (IBIOM), 70126 Bari, Italy;
| | - Carlo Marya Thomas Marobbio
- Department of Biosciences, Biotechnology and Environment, University of Bari Aldo Moro, 70125 Bari, Italy; (A.S.); (S.D.S.); (F.P.); (A.V.); (G.A.); (A.P.); (C.T.S.); (G.F.); (L.P.)
| | - Luigi Palmieri
- Department of Biosciences, Biotechnology and Environment, University of Bari Aldo Moro, 70125 Bari, Italy; (A.S.); (S.D.S.); (F.P.); (A.V.); (G.A.); (A.P.); (C.T.S.); (G.F.); (L.P.)
- CNR Institute of Biomembranes, Bioenergetics and Molecular Biotechnologies (IBIOM), 70126 Bari, Italy;
- Center of Excellence in Comparative Genomics, University of Bari Aldo Moro, 70125 Bari, Italy
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Mou YJ, Li FM, Zhang R, Sheng R, Han R, Zhang ZL, Hu LF, Zhao YZ, Wu JC, Qin ZH. The P2X7 receptor mediates NADPH transport across the plasma membrane. Biochem Biophys Res Commun 2024; 737:150500. [PMID: 39142135 DOI: 10.1016/j.bbrc.2024.150500] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/24/2024] [Accepted: 08/02/2024] [Indexed: 08/16/2024]
Abstract
Nicotinamide Adenine Dinucleotide Phosphate (NADPH) plays a vital role in regulating redox homeostasis and reductive biosynthesis. However, if exogenous NADPH can be transported across the plasma membrane has remained elusive. In this study, we present evidence supporting that NADPH can traverse the plasma membranes of cells through a mechanism mediated by the P2X7 receptor (P2X7R). Notably, we observed an augmentation of intracellular NADPH levels in cultured microglia upon exogenous NADPH supplementation in the presence of ATP. The P2X7R-mediated transmembrane transportation of NADPH was validated with P2X7R antagonists, including OX-ATP, BBG, and A-438079, or through P2X7 knockdown, which impeded NADPH transportation into cells. Conversely, overexpression of P2X7 resulted in an enhanced capacity for NADPH transport. Furthermore, transfection of hP2X7 demonstrated the ability to complement NADPH uptake in native HEK293 cells. Our findings provide evidence for the first time that NADPH is transported across the plasma membrane via a P2X7R-mediated pathway. Additionally, we propose an innovative avenue for modulating intracellular NADPH levels. This discovery holds promise for advancing our understanding of the role of NADPH in redox homeostasis and neuroinflammation.
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Affiliation(s)
- Yu-Jie Mou
- Department of Pharmacology and Laboratory of Aging and Nervous Diseases, Jiangsu Key Laboratory of Neuropsychiatric Diseases, College of Pharmaceutical Sciences, Soochow University, Suzhou, Jiangsu 215123, China.
| | - Feng-Min Li
- Department of Pharmacology and Laboratory of Aging and Nervous Diseases, Jiangsu Key Laboratory of Neuropsychiatric Diseases, College of Pharmaceutical Sciences, Soochow University, Suzhou, Jiangsu 215123, China.
| | - Rong Zhang
- Department of Pharmacology and Laboratory of Aging and Nervous Diseases, Jiangsu Key Laboratory of Neuropsychiatric Diseases, College of Pharmaceutical Sciences, Soochow University, Suzhou, Jiangsu 215123, China.
| | - Rui Sheng
- Department of Pharmacology and Laboratory of Aging and Nervous Diseases, Jiangsu Key Laboratory of Neuropsychiatric Diseases, College of Pharmaceutical Sciences, Soochow University, Suzhou, Jiangsu 215123, China.
| | - Rong Han
- Department of Pharmacology and Laboratory of Aging and Nervous Diseases, Jiangsu Key Laboratory of Neuropsychiatric Diseases, College of Pharmaceutical Sciences, Soochow University, Suzhou, Jiangsu 215123, China.
| | - Zhong-Ling Zhang
- Department of Neurology, The First Affiliated Hospital of Harbin Medical University, 23 Post Street, Nangang District, Harbin, HeiLongjiang 150081, China.
| | - Li-Fang Hu
- Department of Neurology and Clinical Research Center of Neurological Disease, The Second Affiliated Hospital of Soochow University, Suzhou, Jiangsu 215004, China.
| | - Yu-Zheng Zhao
- Optogenetics & Synthetic Biology Interdisciplinary Research Center, State Key Laboratory of Bioreactor Engineering, Shanghai Frontiers Science Center of Optogenetic Techniques for Cell Metabolism, School of Pharmacy, East China University of Science and Technology, 130 Mei Long Road, Shanghai 200237, China.
| | - Jun-Chao Wu
- Department of Pharmacology and Laboratory of Aging and Nervous Diseases, Jiangsu Key Laboratory of Neuropsychiatric Diseases, College of Pharmaceutical Sciences, Soochow University, Suzhou, Jiangsu 215123, China.
| | - Zheng-Hong Qin
- Department of Pharmacology and Laboratory of Aging and Nervous Diseases, Jiangsu Key Laboratory of Neuropsychiatric Diseases, College of Pharmaceutical Sciences, Soochow University, Suzhou, Jiangsu 215123, China; Institute of Health Science and Technology, Suzhou Gaobo Vocational College, Qingshan Road, Suzhou Science and Technology Tower, Hi-Tech Area, Suzhou, Jiangsu 215163, China.
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Qin J, Yang T, Li K, Liu T, Zhang W. Pharyngeal mechanosensory neurons control food swallow in Drosophila melanogaster. eLife 2024; 12:RP88614. [PMID: 39630079 PMCID: PMC11616994 DOI: 10.7554/elife.88614] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/07/2024] Open
Abstract
As the early step of food ingestion, the swallow is under rigorous sensorimotor control. Nevertheless, the mechanisms underlying swallow control at a molecular and circuitry level remain largely unknown. Here, we find that mutation of the mechanotransduction channel genes nompC, Tmc, or piezo impairs the regular pumping rhythm of the cibarium during feeding of the fruit fly Drosophila melanogaster. A group of multi-dendritic mechanosensory neurons, which co-express the three channels, wrap the cibarium and are crucial for coordinating the filling and emptying of the cibarium. Inhibition of them causes difficulty in food emptying in the cibarium, while their activation leads to difficulty in cibarium filling. Synaptic and functional connections are detected between the pharyngeal mechanosensory neurons and the motor circuit that controls swallow. This study elucidates the role of mechanosensation in swallow, and provides insights for a better understanding of the neural basis of food swallow.
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Affiliation(s)
- Jierui Qin
- School of Life Sciences, IDG/McGovern Institute for Brain Research, Tsinghua UniversityBeijingChina
- Tsinghua-Peking Center for Life ScienceBeijingChina
| | - Tingting Yang
- School of Life Sciences, IDG/McGovern Institute for Brain Research, Tsinghua UniversityBeijingChina
- Tsinghua-Peking Center for Life ScienceBeijingChina
| | - Kexin Li
- School of Life Sciences, IDG/McGovern Institute for Brain Research, Tsinghua UniversityBeijingChina
- Tsinghua-Peking Center for Life ScienceBeijingChina
| | - Ting Liu
- School of Life Sciences, IDG/McGovern Institute for Brain Research, Tsinghua UniversityBeijingChina
| | - Wei Zhang
- Tsinghua-Peking Center for Life ScienceBeijingChina
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Ma XB, Yue CX, Liu Y, Yang Y, Wang J, Yang XN, Huang LD, Zhu MX, Hattori M, Li CZ, Yu Y, Guo CR. A shared mechanism for TNP-ATP recognition by members of the P2X receptor family. Comput Struct Biotechnol J 2024; 23:295-308. [PMID: 38173879 PMCID: PMC10762375 DOI: 10.1016/j.csbj.2023.12.005] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2023] [Revised: 11/30/2023] [Accepted: 12/03/2023] [Indexed: 01/05/2024] Open
Abstract
P2X receptors (P2X1-7) are non-selective cation channels involved in many physiological activities such as synaptic transmission, immunological modulation, and cardiovascular function. These receptors share a conserved mechanism to sense extracellular ATP. TNP-ATP is an ATP derivative acting as a nonselective competitive P2X antagonist. Understanding how it occupies the orthosteric site in the absence of agonism may help reveal the key allostery during P2X gating. However, TNP-ATP/P2X complexes (TNP-ATP/human P2X3 (hP2X3) and TNP-ATP/chicken P2X7 (ckP2X7)) with distinct conformations and different mechanisms of action have been proposed. Whether these represent species and subtype variations or experimental differences remains unclear. Here, we show that a common mechanism of TNP-ATP recognition exists for the P2X family members by combining enhanced conformation sampling, engineered disulfide bond analysis, and covalent occupancy. In this model, the polar triphosphate moiety of TNP-ATP interacts with the orthosteric site, while its TNP-moiety is deeply embedded in the head and dorsal fin (DF) interface, creating a restrictive allostery in these two domains that results in a partly enlarged yet ion-impermeable pore. Similar results were obtained from multiple P2X subtypes of different species, including ckP2X7, hP2X3, rat P2X2 (rP2X2), and human P2X1 (hP2X1). Thus, TNP-ATP uses a common mechanism for P2X recognition and modulation by restricting the movements of the head and DF domains which are essential for P2X activation. This knowledge is applicable to the development of new P2X inhibitors.
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Affiliation(s)
- Xiao-Bo Ma
- Department of Pharmacology and Chemical Biology, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Chen-Xi Yue
- School of Basic Medicine and Clinical Pharmacy, and State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009, China
| | - Yan Liu
- Department of Pharmacology and Chemical Biology, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Yang Yang
- Department of Pharmacology and Chemical Biology, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Jin Wang
- Department of Pharmacology and Chemical Biology, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
- School of Basic Medicine and Clinical Pharmacy, and State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009, China
| | - Xiao-Na Yang
- Department of Pharmacology and Chemical Biology, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
- School of Basic Medicine and Clinical Pharmacy, and State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009, China
| | - Li-Dong Huang
- Department of Pharmacology and Chemical Biology, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Michael X. Zhu
- Department of Integrative Biology and Pharmacology, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, TX 77030, USA
| | - Motoyuki Hattori
- State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Shanghai Key Laboratory of Bioactive Small Molecules, Department of Physiology and Neurobiology, School of Life Sciences, Fudan University, Shanghai 200438, China
| | - Chang-Zhu Li
- State Key Laboratory of Utilization of Woody Oil Resource, Hunan Academy of Forestry, Changsha 410004, China
| | - Ye Yu
- Department of Pharmacology and Chemical Biology, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
- School of Basic Medicine and Clinical Pharmacy, and State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009, China
| | - Chang-Run Guo
- School of Traditional Chinese Pharmacy, and State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009, China
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Montalbetti N, Manrique-Maldonado G, Ikeda Y, Dalghi M, Kanai A, Apodaca G, Carattino MD. Expression of Acid-Sensing Ion Channel 3 in Afferents Averts Long-Term Sensitization and the Development of Visceral Pain. Int J Mol Sci 2024; 25:12503. [PMID: 39684215 DOI: 10.3390/ijms252312503] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2024] [Revised: 11/07/2024] [Accepted: 11/18/2024] [Indexed: 12/18/2024] Open
Abstract
Sensitization of primary afferents is essential for the development of pain, but the molecular events involved in this process and its reversal are poorly defined. Recent studies revealed that acid-sensing ion channels (ASICs) control the excitability of nociceptors in the urinary bladder. Using genetic and pharmacological tools we show that ASICs are functionally coupled with voltage-gated Ca2+ channels to mediate Ca2+ transients evoked by acidification in sensory neurons. Genetic deletion of Asic3 of these sensory neurons does not alter the mechanical response of bladder afferents to distension in naïve mice. Both control and sensory neuron conditional Asic3 knockout (Asic3-KO) mice with chemical cystitis induced by cyclophosphamide (CYP) administration exhibit frequent low volume voiding events. However, these changes are transient and revert over time. Of major significance, in Asic3-KO mice, CYP treatment results in the sensitization of a subset of bladder afferents and pelvic allodynia that persist beyond the resolution of the inflammatory process. Thus, ASICs function is necessary to prevent long-term sensitization of visceral nociceptors.
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Affiliation(s)
- Nicolas Montalbetti
- Renal-Electrolyte Division, Department of Medicine, University of Pittsburgh, Pittsburg, PA 15261, USA
| | | | - Youko Ikeda
- Renal-Electrolyte Division, Department of Medicine, University of Pittsburgh, Pittsburg, PA 15261, USA
- Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburg, PA 15261, USA
| | - Marianela Dalghi
- Renal-Electrolyte Division, Department of Medicine, University of Pittsburgh, Pittsburg, PA 15261, USA
| | - Anthony Kanai
- Renal-Electrolyte Division, Department of Medicine, University of Pittsburgh, Pittsburg, PA 15261, USA
- Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburg, PA 15261, USA
| | - Gerard Apodaca
- Renal-Electrolyte Division, Department of Medicine, University of Pittsburgh, Pittsburg, PA 15261, USA
- Department of Cell Biology, University of Pittsburgh, Pittsburg, PA 15261, USA
| | - Marcelo D Carattino
- Renal-Electrolyte Division, Department of Medicine, University of Pittsburgh, Pittsburg, PA 15261, USA
- Department of Cell Biology, University of Pittsburgh, Pittsburg, PA 15261, USA
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Duret T, Elmallah M, Rollin J, Gatault P, Jiang LH, Roger S. Role of purinoreceptors in the release of extracellular vesicles and consequences on immune response and cancer progression. Biomed J 2024; 48:100805. [PMID: 39510381 DOI: 10.1016/j.bj.2024.100805] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/04/2024] [Revised: 10/24/2024] [Accepted: 11/02/2024] [Indexed: 11/15/2024] Open
Abstract
Cell-to-cell communication is a major process for accommodating cell functioning to changes in the environments and to preserve tissue and organism homeostasis. It is achieved by different mechanisms characterized by the origin of the message, the molecular nature of the messenger, its speed of action and its reach. Purinergic signalling is a powerful mechanism initiated by extracellular nucleotides, such as ATP, acting on plasma membrane purinoreceptors. Purinergic signalling is tightly controlled in time and space by the action of ectonucleotidases. Recent studies have highlighted the critical role of purinergic signalling in controlling the generation, release and fate of extracellular vesicles and, in this way, mediating long-distance responses. Most of these discoveries have been made in immune and cancer cells. This review is aimed at establishing the current knowledge on the way which purinoreceptors control extracellular vesicle-mediated communications and consequences for recipient cells.
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Affiliation(s)
- Thomas Duret
- Université de Tours, Inserm UMR1327 ISCHEMIA « Membrane Signalling and Inflammation in Reperfusion Injuries », Tours, France; Fédération Hospitalo-Universitaire Survival Optimization in Organ Transplantation (FHU SUPORT), Tours, France
| | - Mohammed Elmallah
- Université de Tours, Inserm UMR1327 ISCHEMIA « Membrane Signalling and Inflammation in Reperfusion Injuries », Tours, France
| | - Jérôme Rollin
- Université de Tours, Inserm UMR1327 ISCHEMIA « Membrane Signalling and Inflammation in Reperfusion Injuries », Tours, France; Service d'Hématologie-Hémostase, CHRU de Tours, Tours, France
| | - Philippe Gatault
- Université de Tours, Inserm UMR1327 ISCHEMIA « Membrane Signalling and Inflammation in Reperfusion Injuries », Tours, France; Service de Néphrologie, Hypertension, Dialyse et Transplantation Rénale, Tours, France; Fédération Hospitalo-Universitaire Survival Optimization in Organ Transplantation (FHU SUPORT), Tours, France
| | - Lin-Hua Jiang
- Université de Tours, Inserm UMR1327 ISCHEMIA « Membrane Signalling and Inflammation in Reperfusion Injuries », Tours, France; School of Basic Medical Sciences, Xinxiang Medical University, Henan, China; School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom
| | - Sébastien Roger
- Université de Tours, Inserm UMR1327 ISCHEMIA « Membrane Signalling and Inflammation in Reperfusion Injuries », Tours, France; Fédération Hospitalo-Universitaire Survival Optimization in Organ Transplantation (FHU SUPORT), Tours, France.
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Dunker C, Vinnenberg L, Isaak A, Karabatak E, Hundehege P, Budde T, Murakami K, Junker A. Exploring P2X receptor activity: A journey from cellular impact to electrophysiological profiling. Biochem Pharmacol 2024; 229:116543. [PMID: 39304104 DOI: 10.1016/j.bcp.2024.116543] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/25/2024] [Revised: 08/12/2024] [Accepted: 09/13/2024] [Indexed: 09/22/2024]
Abstract
The development of in vitro pharmacological assays relies on creating genetically modified cell lines that overexpress the target protein of interest. However, the choice of the host cell line can significantly impact the experimental outcomes. This study explores the functional characterization of P2X7 and P2X4 receptor modulators through cellular assays and advanced electrophysiological techniques. The influence of different host cell lines (HEK-293, HEK-293FT, and 1321N1) on the activity of reference agonists and antagonists targeting human and murine P2X4 and P2X7 receptors was systematically investigated, highlighting the significant impact of the host cell on experimental results. The 1321N1 cell line was identified as the preferred host cell line when investigating the human P2X4 receptor due to more consistent agonist activities, antagonist potencies, and a more stable assay signal window. Furthermore, a patch-clamp protocol that allows for the repetitive recording of ATP-mediated inward currents from isolated human CD4+ T-cells was established, revealing that both P2X7 and P2X4 receptors are crucial for immune cell regulation, positioning them as promising therapeutic targets for managing inflammatory disorders.
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Affiliation(s)
- Calvin Dunker
- European Institute for Molecular Imaging (EIMI), Roentgenstr 16, University of Muenster, 48149 Muenster, Germany; Werner Siemens Imaging Center, Department of Preclinical Imaging and Radiopharmacy, University of Tübingen, Roentgenweg 13, 72076, Tuebingen, Germany
| | - Laura Vinnenberg
- University Hospital Muenster, Department of Neurology with Institute of Translational Neurology, Albert-Schweitzer-Campus 1, 48149 Muenster, Germany
| | - Andreas Isaak
- European Institute for Molecular Imaging (EIMI), Roentgenstr 16, University of Muenster, 48149 Muenster, Germany
| | - Elif Karabatak
- Institute of Physiology I, University of Muenster, Robert-Koch-Str. 27a, 48149 Muenster, Germany
| | - Petra Hundehege
- University Hospital Muenster, Department of Neurology with Institute of Translational Neurology, Albert-Schweitzer-Campus 1, 48149 Muenster, Germany
| | - Thomas Budde
- Institute of Physiology I, University of Muenster, Robert-Koch-Str. 27a, 48149 Muenster, Germany
| | - Kazuhiro Murakami
- Division of Epithelial Stem Cell Biology, Cancer Research Institute, Kanazawa University, Kanazawa, Ishikawa 920-1192, Japan
| | - Anna Junker
- European Institute for Molecular Imaging (EIMI), Roentgenstr 16, University of Muenster, 48149 Muenster, Germany; Werner Siemens Imaging Center, Department of Preclinical Imaging and Radiopharmacy, University of Tübingen, Roentgenweg 13, 72076, Tuebingen, Germany.
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38
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Hao JW, Liu TT, Qiu CY, Li XM, Qiao WL, Li Q, Qin QR, Hu WP. Lipid mediator resolvin D2 inhibits ATP currents in rat primary sensory neurons. J Neurochem 2024; 168:3715-3726. [PMID: 37987505 DOI: 10.1111/jnc.16009] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2023] [Revised: 10/19/2023] [Accepted: 10/19/2023] [Indexed: 11/22/2023]
Abstract
Resolvin D2 (RvD2), an endogenous lipid mediator derived from docosahexaenoic acid, has been demonstrated to have analgesic effects. However, little is known about the mechanism underlying RvD2 in pain relief. Herein, we demonstrate that RvD2 targeted the P2X3 receptor as an analgesic. The electrophysiological activity of P2X3 receptors was suppressed by RvD2 in rat dorsal root ganglia (DRG) neurons. RvD2 pre-application dose-dependently decreased α,β-methylene-ATP (α,β-meATP)-induced inward currents. RvD2 remarkably decreased the maximum response to α,β-meATP, without influencing the affinity of P2X3 receptors. RvD2 also voltage-independently suppressed ATP currents. An antagonist of the G protein receptor 18 (GPR18), O-1918, prevented the RvD2-induced suppression of ATP currents. Additionally, intracellular dialysis of the Gαi/o-protein antagonist pertussis toxin (PTX), the PKA antagonist H89, or the cAMP analog 8-Br-cAMP also blocked the RvD2-induced suppression. Furthermore, α,β-meATP-triggered depolarization of membrane potential along with the action potential bursts in DRG neurons were inhibited by RvD2. Lastly, RvD2 attenuated spontaneous nociceptive behaviors as well as mechanical allodynia produced by α,β-meATP in rats via the activation of the peripheral GPR18. These findings indicated that RvD2 inhibited P2X3 receptors in rat primary sensory neurons through GPR18, PTX-sensitive Gαi/o-proteins, and intracellular cAMP/PKA signaling, revealing a novel mechanism that underlies its analgesic effects by targeting P2X3 receptors.
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Affiliation(s)
- Jia-Wei Hao
- School of Pharmacy, School of Basic Medical Sciences, Xianning Medical College, Hubei University of Science and Technology, Xianning, Hubei, PR China
| | - Ting-Ting Liu
- School of Pharmacy, School of Basic Medical Sciences, Xianning Medical College, Hubei University of Science and Technology, Xianning, Hubei, PR China
| | - Chun-Yu Qiu
- School of Pharmacy, School of Basic Medical Sciences, Xianning Medical College, Hubei University of Science and Technology, Xianning, Hubei, PR China
| | - Xue-Mei Li
- School of Pharmacy, School of Basic Medical Sciences, Xianning Medical College, Hubei University of Science and Technology, Xianning, Hubei, PR China
| | - Wen-Long Qiao
- School of Pharmacy, School of Basic Medical Sciences, Xianning Medical College, Hubei University of Science and Technology, Xianning, Hubei, PR China
| | - Qing Li
- School of Pharmacy, School of Basic Medical Sciences, Xianning Medical College, Hubei University of Science and Technology, Xianning, Hubei, PR China
| | - Qing-Rui Qin
- School of Pharmacy, School of Basic Medical Sciences, Xianning Medical College, Hubei University of Science and Technology, Xianning, Hubei, PR China
| | - Wang-Ping Hu
- School of Pharmacy, School of Basic Medical Sciences, Xianning Medical College, Hubei University of Science and Technology, Xianning, Hubei, PR China
- Department of Physiology, Hubei College of Chinese Medicine, Jingzhou, Hubei, PR China
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39
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Bai Y, Zhou Z, Han B, Xiang X, Huang W, Yao H. Revisiting astrocytic calcium signaling in the brain. FUNDAMENTAL RESEARCH 2024; 4:1365-1374. [PMID: 39734522 PMCID: PMC11670731 DOI: 10.1016/j.fmre.2023.11.021] [Citation(s) in RCA: 6] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2023] [Revised: 11/28/2023] [Accepted: 11/30/2023] [Indexed: 12/31/2024] Open
Abstract
Astrocytes, characterized by complex spongiform morphology, participate in various physiological processes, and abnormal changes in their calcium (Ca2+) signaling are implicated in central nervous system disorders. However, medications targeting the control of Ca2+ have fallen short of the anticipated therapeutic outcomes in clinical applications. This underscores the fact that our comprehension of this intricate regulation of calcium ions remains considerably incomplete. In recent years, with the advancement of Ca2+ labeling, imaging, and analysis techniques, Ca2+ signals have been found to exhibit high specificity at different spatial locations within the intricate structure of astrocytes. This has ushered the study of Ca2+ signaling in astrocytes into a new phase, leading to several groundbreaking research achievements. Despite this, the comprehensive understanding of astrocytic Ca2+ signaling and their implications remains challenging area for future research.
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Affiliation(s)
- Ying Bai
- Department of Pharmacology, Jiangsu Provincial Key Laboratory of Critical Care Medicine, School of Medicine, Southeast University, Nanjing 210009, China
| | - Zhongqiu Zhou
- Department of Pharmacology, Jiangsu Provincial Key Laboratory of Critical Care Medicine, School of Medicine, Southeast University, Nanjing 210009, China
| | - Bing Han
- Department of Pharmacology, Jiangsu Provincial Key Laboratory of Critical Care Medicine, School of Medicine, Southeast University, Nanjing 210009, China
| | - Xianyuan Xiang
- Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen 518055, China
| | - Wenhui Huang
- Molecular Physiology, CIPMM, University of Saarland, Homburg 66421, Germany
| | - Honghong Yao
- Department of Pharmacology, Jiangsu Provincial Key Laboratory of Critical Care Medicine, School of Medicine, Southeast University, Nanjing 210009, China
- Co-innovation Center of Neuroregeneration, Nantong University, Nantong 226001, China
- Institute of Life Sciences, Key Laboratory of Developmental Genes and Human Disease, Southeast University, Nanjing 210096, China
- Center for Global Health, School of Public Health, Nanjig Medical University, Nanjing 211166, China
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40
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Delafenêtre A, Chapotte-Baldacci CA, Dorémus L, Massouridès E, Bernard M, Régnacq M, Piquereau J, Chatelier A, Cognard C, Pinset C, Sebille S. Duchenne muscular dystrophy skeletal muscle cells derived from human induced pluripotent stem cells recapitulate various calcium dysregulation pathways. Cell Calcium 2024; 123:102943. [PMID: 39154623 DOI: 10.1016/j.ceca.2024.102943] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2024] [Revised: 07/23/2024] [Accepted: 08/11/2024] [Indexed: 08/20/2024]
Abstract
Duchenne muscular dystrophy (DMD) is an X-linked progressive muscle degenerative disease, caused by mutations in the dystrophin gene and resulting in premature death. As a major secondary event, an abnormal elevation of the intracellular calcium concentration in the dystrophin-deficient muscle contributes to disease progression in DMD. In this study, we investigated the specific functional features of induced pluripotent stem cell-derived muscle cells (hiPSC-skMCs) generated from DMD patients to regulate intracellular calcium concentration. As compared to healthy hiPSC-skMCs, DMD hiPSC-skMCs displayed specific spontaneous calcium signatures with high levels of intracellular calcium concentration. Furthermore, stimulations with electrical field or with acetylcholine perfusion induced higher calcium response in DMD hiPSC-skMCs as compared to healthy cells. Finally, Mn2+ quenching experiments demonstrated high levels of constitutive calcium entries in DMD hiPSC-skMCs as compared to healthy cells. Our findings converge on the fact that DMD hiPSC-skMCs display intracellular calcium dysregulation as demonstrated in several other models. Observed calcium disorders associated with RNAseq analysis on these DMD cells highlighted some mechanisms, such as spontaneous and activated sarcoplasmic reticulum (SR) releases or constitutive calcium entries, known to be disturbed in other dystrophin-deficient models. However, store operated calcium entries (SOCEs) were not found to be dysregulated in our DMD hiPSC-skMCs model. These results suggest that all the mechanisms of calcium impairment observed in other animal models may not be as pronounced in humans and could point to a preference for certain mechanisms that could correspond to major molecular targets for DMD therapies.
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Affiliation(s)
| | | | - Léa Dorémus
- PRETI laboratory, University of Poitiers, France
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41
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Lim XR, Mercer M, Harraz OF, Hollywood MA, Sergeant GP, Thornbury KD. Evidence of an excitatory purinergic innervation in mouse corpus cavernosum smooth muscle. J Sex Med 2024; 21:977-985. [PMID: 39234981 DOI: 10.1093/jsxmed/qdae107] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2023] [Revised: 06/14/2024] [Accepted: 08/14/2024] [Indexed: 09/06/2024]
Abstract
BACKGROUND Evidence suggests that the corpus cavernosum smooth muscle (CCSM) cells of several species, including humans, express purinergic P2X receptors, but it is not known if the corpus cavernosum has an excitatory purinergic innervation. AIM In this study we aimed to determine if the mouse CCSM has a functional purinergic innervation. METHODS Mouse CCSM myocytes were enzymatically isolated and studied using the perforated patch configuration of the patch clamp technique. Isometric tension was measured in whole cavernosum tissue subjected to electrical field stimulation (EFS) to evoke nerve-mediated responses. OUTCOMES The mouse CCSM myocytes expressed P2X1 receptors, and adenosine triphosphate (ATP) evoked inward currents in these cells. In addition, P2X1-mediated contractions were recorded in whole tissue in response to EFS. RESULTS In cells held under a voltage clamp at -60 mV, ATP (1 μm) evoked large inward currents (mean approximately 900 pA). This current rapidly declined but was repeatable at 8-minute intervals. α,β-methylene ATP (10 μM), an agonist of P2X1 and P2X3 receptors, caused a similar current that also rapidly declined. Desensitization to α,β-methylene ATP negated the effect of ATP, but the ATP effect was restored 8 minutes after washout of α,β-methylene ATP. The effect of ATP was reversibly blocked by NF449 (1 μm), a selective antagonist of P2X1 receptors. In isometric tension experiments electrical field stimulation (EFS) at 0.5-8 Hz evoked frequency-dependent contractions in the presence of l-nitro arginine (l-NO-Arg) (100 μm). When phentolamine (3 μm) and atropine (1 μm) were applied, there remained a nonadrenergic, noncholinergic component of the response to EFS, consisting mainly of a transient contraction. This was significantly reduced by NF449 (1 μm). Finally, in immunocytochemistry experiments, isolated CCSM myocytes stained positively when exposed to an antibody raised against P2X1 receptors. CLINICAL IMPLICATIONS Previous studies have shown that P2X1 receptors in CCSM are upregulated in diabetes. These findings, taken together with the functional evidence presented here, indicate that P2X1 receptors may provide an alternative therapeutic target for treatment of erectile dysfunction in patients with diabetes, which is known to be relatively resistant to treatment with phosphodiesterase 5 inhibitors. STRENGTHS AND LIMITATIONS Strengths of this study are the use of a combination of functional experiments (patch clamp) and immunocytochemical analyses to show expression of P2X1 receptors on CCSM myocytes while also performing functional experiments to show that stimulation these receptors results in contraction of CCSM. A limitation of this study was the use of animal rather than human tissue. CONCLUSION This investigation provides evidence that mouse corpus cavernosum smooth muscle cells express P2X1 receptors and that these receptors are involved in mediating part of the contractile response to nerve stimulation evoked by EFS.
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Affiliation(s)
- Xin Rui Lim
- Department of Pharmacology, Larner College of Medicine, University of Vermont, Burlington VT 05405, United States
| | - Mitchell Mercer
- Smooth Muscle Research Centre, Dundalk Institute of Technology, Dundalk, County Louth A91 K584, Ireland
| | - Osama F Harraz
- Department of Pharmacology, Larner College of Medicine, University of Vermont, Burlington VT 05405, United States
| | - Mark A Hollywood
- Smooth Muscle Research Centre, Dundalk Institute of Technology, Dundalk, County Louth A91 K584, Ireland
| | - Gerard P Sergeant
- Smooth Muscle Research Centre, Dundalk Institute of Technology, Dundalk, County Louth A91 K584, Ireland
| | - Keith D Thornbury
- Smooth Muscle Research Centre, Dundalk Institute of Technology, Dundalk, County Louth A91 K584, Ireland
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42
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Oken AC, Ditter IA, Lisi NE, Krishnamurthy I, Godsey MH, Mansoor SE. P2X 7 receptors exhibit at least three modes of allosteric antagonism. SCIENCE ADVANCES 2024; 10:eado5084. [PMID: 39365862 PMCID: PMC11451537 DOI: 10.1126/sciadv.ado5084] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/07/2024] [Accepted: 08/28/2024] [Indexed: 10/06/2024]
Abstract
P2X receptors are trimeric ion channels activated by adenosine triphosphate (ATP) that contribute to pathophysiological processes ranging from asthma to neuropathic pain and neurodegeneration. A number of small-molecule antagonists have been identified for these important pharmaceutical targets. However, the molecular pharmacology of P2X receptors is poorly understood because of the chemically disparate nature of antagonists and their differential actions on the seven constituent subtypes. Here, we report high-resolution cryo-electron microscopy structures of the homomeric rat P2X7 receptor bound to five previously known small-molecule allosteric antagonists and a sixth antagonist that we identify. Our structural, biophysical, and electrophysiological data define the molecular determinants of allosteric antagonism in this pharmacologically relevant receptor, revealing three distinct classes of antagonists that we call shallow, deep, and starfish. Starfish binders, exemplified by the previously unidentified antagonist methyl blue, represent a unique class of inhibitors with distinct functional properties that could be exploited to develop potent P2X7 ligands with substantial clinical impact.
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Affiliation(s)
- Adam C. Oken
- Department of Chemical Physiology and Biochemistry, Oregon Health & Science University, Portland, OR 97239, USA
| | - Ismayn A. Ditter
- Department of Chemical Physiology and Biochemistry, Oregon Health & Science University, Portland, OR 97239, USA
| | - Nicolas E. Lisi
- Department of Chemical Physiology and Biochemistry, Oregon Health & Science University, Portland, OR 97239, USA
| | - Ipsita Krishnamurthy
- Department of Chemical Physiology and Biochemistry, Oregon Health & Science University, Portland, OR 97239, USA
| | - Michael H. Godsey
- Department of Chemical Physiology and Biochemistry, Oregon Health & Science University, Portland, OR 97239, USA
| | - Steven E. Mansoor
- Department of Chemical Physiology and Biochemistry, Oregon Health & Science University, Portland, OR 97239, USA
- Division of Cardiovascular Medicine, Knight Cardiovascular Institute, Oregon Health & Science University, Portland, OR 97239, USA
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Segars KL, Azzari N, Cole M, Kushimi L, Rapaka S, Rich CB, Trinkaus-Randall V. Diverse calcium signaling profiles regulate migratory behavior in avascular wound healing and aberrant signal hierarchy occurs early in diabetes. Am J Physiol Cell Physiol 2024; 327:C1051-C1072. [PMID: 39129489 PMCID: PMC11482046 DOI: 10.1152/ajpcell.00249.2024] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/16/2024] [Revised: 08/06/2024] [Accepted: 08/06/2024] [Indexed: 08/13/2024]
Abstract
In avascular wound repair, calcium signaling events are the predominant mechanism cells use to transduce information about stressors in the environment into an effective and coordinated migratory response. Live cell imaging and computational analysis of corneal epithelial wound healing revealed that signal initiation and propagation at the wound edge are highly ordered, with groups of cells engaging in cyclical patterns of initiation and propagation. The cells in these groups exhibit a diverse range of signaling behavior, and dominant "conductor cells" drive activity in groups of lower-signaling neighbors. Ex vivo model systems reveal that conductor cells are present in wing cell layers of the corneal epithelium and that signaling propagates both within and between wing and basal layers. There are significant aberrations in conductor phenotype and interlayer propagation in type II diabetic murine models, indicating that signal hierarchy breakdown is an early indicator of disease. In vitro models reveal that signaling profile diversity and conductor cell phenotype is eliminated with P2X7 inhibition and is altered in Pannexin-1 or P2Y2 but not Connexin-43 inhibition. Conductor cells express significantly less P2X7 than their lower-signaling neighbors and exhibit significantly less migratory behavior after injury. Together, our results show that the postinjury calcium signaling cascade exhibits significantly more ordered and hierarchical behavior than previously thought, that proteins previously shown to be essential for regulating motility are also essential for determining signaling phenotype, and that loss of signal hierarchy integrity is an early indicator of disease state. NEW & NOTEWORTHY Calcium signaling in corneal epithelial cells after injury is highly ordered, with groups of cells engaged in cyclical patterns of event initiation and propagation driven by high-signaling cells. Signaling behavior is determined by P2X7, Pannexin-1, and P2Y2 and influences migratory behavior. Signal hierarchy is observed in healthy ex vivo models after injury and becomes aberrant in diabetes. This represents a paradigm shift, as signaling was thought to be random and determined by factors in the environment.
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Affiliation(s)
- Kristen L Segars
- Department of Pharmacology, Physiology, and Biophysics, Boston University Chobanian and Avedisian School of Medicine, Boston, Massachusetts, United States
| | - Nicholas Azzari
- Department of Biochemistry and Cell Biology, Boston University Chobanian and Avedisian School of Medicine, Boston, Massachusetts, United States
| | - Malia Cole
- STaRS Program, Boston University Chobanian and Avedisian School of Medicine, Boston, Massachusetts, United States
| | - Landon Kushimi
- Department of Computer Science, Boston University Center for Computing and Data Sciences, Boston, Massachusetts, United States
| | - Srikar Rapaka
- Department of Medicine, Boston University Chobanian and Avedisian School of Medicine, Boston, Massachusetts, United States
| | - Celeste B Rich
- Department of Biochemistry and Cell Biology, Boston University Chobanian and Avedisian School of Medicine, Boston, Massachusetts, United States
| | - Vickery Trinkaus-Randall
- Department of Biochemistry and Cell Biology, Boston University Chobanian and Avedisian School of Medicine, Boston, Massachusetts, United States
- Department of Ophthalmology, Boston University Chobanian and Avedisian School of Medicine, Boston, Massachusetts, United States
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Oken AC, Lisi NE, Ditter IA, Shi H, Nechiporuk NA, Mansoor SE. Cryo-EM structures of the human P2X1 receptor reveal subtype-specific architecture and antagonism by supramolecular ligand-binding. Nat Commun 2024; 15:8490. [PMID: 39353889 PMCID: PMC11448502 DOI: 10.1038/s41467-024-52636-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/03/2024] [Accepted: 09/16/2024] [Indexed: 10/03/2024] Open
Abstract
P2X receptors are a family of seven trimeric non-selective cation channels that are activated by extracellular ATP to play roles in the cardiovascular, neuronal, and immune systems. Although it is known that the P2X1 receptor subtype has increased sensitivity to ATP and fast desensitization kinetics, an underlying molecular explanation for these subtype-selective features is lacking. Here we report high-resolution cryo-EM structures of the human P2X1 receptor in the apo closed, ATP-bound desensitized, and the high-affinity antagonist NF449-bound inhibited states. The apo closed and ATP-bound desensitized state structures of human P2X1 define subtype-specific properties such as distinct pore architecture and ATP-interacting residues. The NF449-bound inhibited state structure of human P2X1 reveals that NF449 has a unique dual-ligand supramolecular binding mode at the interface of neighboring protomers, inhibiting channel activation by overlapping with the canonical P2X receptor ATP-binding site. Altogether, these data define the molecular pharmacology of the human P2X1 receptor laying the foundation for structure-based drug design.
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Affiliation(s)
- Adam C Oken
- Department of Chemical Physiology & Biochemistry, Oregon Health & Science University, Portland, OR, 97239, USA
| | - Nicolas E Lisi
- Department of Chemical Physiology & Biochemistry, Oregon Health & Science University, Portland, OR, 97239, USA
| | - Ismayn A Ditter
- Department of Chemical Physiology & Biochemistry, Oregon Health & Science University, Portland, OR, 97239, USA
| | - Haoyuan Shi
- Department of Chemical Physiology & Biochemistry, Oregon Health & Science University, Portland, OR, 97239, USA
| | - Nadia A Nechiporuk
- Department of Chemical Physiology & Biochemistry, Oregon Health & Science University, Portland, OR, 97239, USA
| | - Steven E Mansoor
- Department of Chemical Physiology & Biochemistry, Oregon Health & Science University, Portland, OR, 97239, USA.
- Division of Cardiovascular Medicine, Knight Cardiovascular Institute, Oregon Health & Science University, Portland, OR, 97239, USA.
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Di Virgilio F, Vultaggio-Poma V, Tarantini M, Giuliani AL. Overview of the role of purinergic signaling and insights into its role in cancer therapy. Pharmacol Ther 2024; 262:108700. [PMID: 39111410 DOI: 10.1016/j.pharmthera.2024.108700] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/07/2024] [Revised: 07/05/2024] [Accepted: 07/31/2024] [Indexed: 08/30/2024]
Abstract
Innovation of cancer therapy has received a dramatic acceleration over the last fifteen years thanks to the introduction of the novel immune checkpoint inhibitors (ICI). On the other hand, the conspicuous scientific knowledge accumulated in purinergic signaling since the early seventies is finally being transferred to the clinic. Several Phase I/II clinical trials are currently underway to investigate the effect of drugs interfering with purinergic signaling as stand-alone or combination therapy in cancer. This is supporting the novel concept of "purinergic immune checkpoint" (PIC) in cancer therapy. In the present review we will address a) the basic pharmacology and cell biology of the purinergic system; b) principles of its pathophysiology in human diseases; c) implications for cell death, cell proliferation and cancer; d) novel molecular tools to investigate nucleotide homeostasis in the extracellular environment; e) recent developments in the pharmacology of P1, P2 receptors and related ecto-enzymes; f) P1 and P2 ligands as novel diagnostic tools; g) current issues in PIC-based anti-cancer therapy. This review will provide an appraisal of the current status of purinergic signaling in cancer and will help identify future avenues of development.
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Affiliation(s)
| | | | - Mario Tarantini
- Department of Medical Sciences, University of Ferrara, Italy
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Bennetts FM, Venugopal H, Glukhova A, Mobbs JI, Ventura S, Thal DM. Structural insights into the human P2X1 receptor and ligand interactions. Nat Commun 2024; 15:8418. [PMID: 39341830 PMCID: PMC11439047 DOI: 10.1038/s41467-024-52776-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2024] [Accepted: 09/20/2024] [Indexed: 10/01/2024] Open
Abstract
The P2X1 receptor is a trimeric ligand-gated ion channel that plays an important role in urogenital and immune functions, offering the potential for new drug treatments. However, progress in this area has been hindered by limited structural information and a lack of well-characterised tool compounds. In this study, we employ cryogenic electron microscopy (cryo-EM) to elucidate the structures of the P2X1 receptor in an ATP-bound desensitised state and an NF449-bound closed state. NF449, a potent P2X1 receptor antagonist, engages the receptor distinctively, while ATP, the endogenous ligand, binds in a manner consistent with other P2X receptors. To explore the molecular basis of receptor inhibition, activation, and ligand interactions, key residues involved in ligand and metal ion binding were mutated. Radioligand binding assays with [3H]-α,β-methylene ATP and intracellular calcium ion influx assays were used to evaluate the effects of these mutations. These experiments validate key ligand-receptor interactions and identify conserved and non-conserved residues critical for ligand binding or receptor modulation. This research expands our understanding of the P2X1 receptor structure at a molecular level and opens new avenues for in silico drug design targeting the P2X1 receptor.
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Affiliation(s)
- Felix M Bennetts
- Drug Discovery Biology, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, VIC, Australia
- Australian Research Council Centre for Cryo-Electron Microscopy of Membrane Proteins, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, VIC, Australia
| | - Hariprasad Venugopal
- Ramaciotti Centre for Cryo-Electron Microscopy, Monash University, Clayton, VIC, Australia
| | - Alisa Glukhova
- Australian Research Council Centre for Cryo-Electron Microscopy of Membrane Proteins, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, VIC, Australia
- Structural Biology Division, Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia
| | - Jesse I Mobbs
- Drug Discovery Biology, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, VIC, Australia.
- Australian Research Council Centre for Cryo-Electron Microscopy of Membrane Proteins, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, VIC, Australia.
| | - Sabatino Ventura
- Drug Discovery Biology, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, VIC, Australia.
| | - David M Thal
- Drug Discovery Biology, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, VIC, Australia.
- Australian Research Council Centre for Cryo-Electron Microscopy of Membrane Proteins, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, VIC, Australia.
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Cheng XY, Ren WJ, Li X, Deussing JM, Illes P, Tang Y, Rubini P. Astrocytic P2X7 receptor regulates depressive-like behavioral reactions of mice in response to acute stressful stimulation. Purinergic Signal 2024:10.1007/s11302-024-10047-6. [PMID: 39325357 DOI: 10.1007/s11302-024-10047-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/11/2024] [Accepted: 08/25/2024] [Indexed: 09/27/2024] Open
Abstract
Acute stress causes depressive-like reactions in the tail suspension (TST) and forced swim tests (FST) of mice. Similarly, inescapable foot shock is able to promote the development of anhedonia as indicated by decreased sucrose consumption of treated mice in the sucrose preference test (SPT). The astrocyte-specific deletion of the P2X7R by a conditional knockout strategy or its knockdown by the intracerebroventricular (i.c.v.) delivery of an adeno-associated virus (AAV) expressing P2X7R-specific shRNA in astrocytes significantly prolonged the immobility time in TST and FST. In contrast, the shRNA-induced downregulation of the P2X7R in neurons, oligodendrocytes, or microglia had no detectable effect on the behavior of treated mice in these tests. Moreover, sucrose consumption in the SPT was not altered following inescapable foot shock treatment in any of these cell type-specific approaches. Immunohistochemistry indicated that the administered astrocyte-specific AAV efficiently conveyed expression of shRNA by hippocampal CA1 astrocytes, but not by neurons. In conclusion, P2X7R in astrocytes of this area of the brain appears to be involved in depressive-like reactions to acute stressors.
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Affiliation(s)
- Xin-Yi Cheng
- International Research Centre on Purinergic Signalling, School of Acupuncture and Tuina, Chengdu University of Traditional Chinese Medicine, Chengdu, China
| | - Wen-Jing Ren
- International Research Centre on Purinergic Signalling, School of Acupuncture and Tuina, Chengdu University of Traditional Chinese Medicine, Chengdu, China
| | - Xuan Li
- International Research Centre on Purinergic Signalling, School of Acupuncture and Tuina, Chengdu University of Traditional Chinese Medicine, Chengdu, China
| | - Jan M Deussing
- Molecular Neurogenetics, Max Planck Institute of Psychiatry, Munich, Germany
| | - Peter Illes
- International Research Centre on Purinergic Signalling, School of Acupuncture and Tuina, Chengdu University of Traditional Chinese Medicine, Chengdu, China.
- Acupuncture and Chronobiology Key Laboratory of Sichuan Province, Chengdu, China.
- Rudolf Boehm Institute for Pharmacology and Toxicology, University of Leipzig, Leipzig, Germany.
| | - Yong Tang
- International Research Centre on Purinergic Signalling, School of Acupuncture and Tuina, Chengdu University of Traditional Chinese Medicine, Chengdu, China.
- Acupuncture and Chronobiology Key Laboratory of Sichuan Province, Chengdu, China.
- School of Health and Rehabilitation, Chengdu University of Traditional Chinese Medicine, Chengdu, China.
| | - Patrizia Rubini
- International Research Centre on Purinergic Signalling, School of Acupuncture and Tuina, Chengdu University of Traditional Chinese Medicine, Chengdu, China.
- Acupuncture and Chronobiology Key Laboratory of Sichuan Province, Chengdu, China.
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Choi S, Dalloul RSD, Vemulapalli PB, Yousef S, Goswami N, Schmidt F. Comprehensive Proteomic Profiling of Converted Adipocyte-like Cells from Normal Human Dermal Fibroblasts Using Data-Independent Acquisition Mass Spectrometry. ACS OMEGA 2024; 9:40034-40050. [PMID: 39346858 PMCID: PMC11425921 DOI: 10.1021/acsomega.4c05852] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 06/24/2024] [Revised: 08/28/2024] [Accepted: 09/01/2024] [Indexed: 10/01/2024]
Abstract
Adipocytes play an important role in the regulation of systemic energy homeostasis and are closely related to metabolic disorders, such as type-2 diabetes and inflammatory bowel diseases. Particularly, there is an increasing need for a human adipocyte model for studying metabolic diseases and obesity. However, utilizing human primary adipocyte culture and stem-cell-based models presents several practical limitations due to their time-consuming nature, requirement for relatively intensive labor, and high cost. Here, we applied direct conversion of normal human dermal fibroblasts (NHDFs) into adipocyte-like cells using an adipogenic cocktail containing 3-isobutyl-1-methylxanthine (IBMX), dexamethasone, insulin, and rosiglitazone and confirmed prominent lipid droplet accumulation in the converted cells. For profiling the proteome changes in the converted cells, we conducted a comprehensive quantitative proteome analysis of both the intracellular and extracellular proteome fractions using data-independent acquisition mass spectrometry. We observed that several proteins, which are known to be highly expressed in adipocytes specifically, were dominantly increased in the converted cells. In this study, we suggest that NHDFs can be converted into adipocyte-like cells by an adipogenic cocktail and can serve as a useful tool for studying human adipocytes and their metabolism.
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Affiliation(s)
- Sunkyu Choi
- Proteomics Core, Research, Weill Cornell Medicine-Qatar, Education City, Qatar Foundation, P.O. 24144 Doha, Qatar
| | - Rajaa S D Dalloul
- Proteomics Core, Research, Weill Cornell Medicine-Qatar, Education City, Qatar Foundation, P.O. 24144 Doha, Qatar
| | - Praveen Babu Vemulapalli
- Proteomics Core, Research, Weill Cornell Medicine-Qatar, Education City, Qatar Foundation, P.O. 24144 Doha, Qatar
| | - Sondos Yousef
- Proteomics Core, Research, Weill Cornell Medicine-Qatar, Education City, Qatar Foundation, P.O. 24144 Doha, Qatar
| | - Neha Goswami
- Proteomics Core, Research, Weill Cornell Medicine-Qatar, Education City, Qatar Foundation, P.O. 24144 Doha, Qatar
| | - Frank Schmidt
- Proteomics Core, Research, Weill Cornell Medicine-Qatar, Education City, Qatar Foundation, P.O. 24144 Doha, Qatar
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Leite-Aguiar R, Bello-Santos VG, Castro NG, Coutinho-Silva R, Savio LEB. Techniques for evaluating the ATP-gated ion channel P2X7 receptor function in macrophages and microglial cells. J Immunol Methods 2024; 532:113727. [PMID: 38997100 DOI: 10.1016/j.jim.2024.113727] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/19/2024] [Revised: 05/31/2024] [Accepted: 07/08/2024] [Indexed: 07/14/2024]
Abstract
Resident macrophages are tissue-specific innate immune cells acting as sentinels, constantly patrolling their assigned tissue to maintain homeostasis, and quickly responding to pathogenic invaders or molecular danger signals molecules when necessary. Adenosine triphosphate (ATP), when released to the extracellular medium, acts as a danger signal through specific purinergic receptors. Interaction of ATP with the purinergic receptor P2X7 activates macrophages and microglial cells in different pathological conditions, triggering inflammation. The highly expressed P2X7 receptor in these cells induces cell membrane permeabilization, inflammasome activation, cell death, and the production of inflammatory mediators, including cytokines and nitrogen and oxygen-reactive species. This review explores the techniques to evaluate the functional and molecular aspects of the P2X7 receptor, particularly in macrophages and microglial cells. Polymerase chain reaction (PCR), Western blotting, and immunocytochemistry or immunohistochemistry are essential for assessing gene and protein expression in these cell types. Evaluation of P2X7 receptor function involves the use of ATP and selective agonists and antagonists and diverse techniques, including electrophysiology, intracellular calcium measurements, ethidium bromide uptake, and propidium iodide cell viability assays. These techniques are crucial for studying the role of P2X7 receptors in immune responses, neuroinflammation, and various pathological conditions. Therefore, a comprehensive understanding of the functional and molecular aspects of the P2X7 receptor in macrophages and microglia is vital for unraveling its involvement in immune modulation and its potential as a therapeutic target. The methodologies presented and discussed herein offer valuable tools for researchers investigating the complexities of P2X7 receptor signaling in innate immune cells in health and disease.
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Affiliation(s)
- Raíssa Leite-Aguiar
- Laboratório de Imunofisiologia, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil
| | | | | | - Robson Coutinho-Silva
- Laboratório de Imunofisiologia, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil
| | - Luiz Eduardo Baggio Savio
- Laboratório de Imunofisiologia, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil..
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50
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Shukla S, Dalai P, Agrawal-Rajput R. Metabolic crosstalk: Extracellular ATP and the tumor microenvironment in cancer progression and therapy. Cell Signal 2024; 121:111281. [PMID: 38945420 DOI: 10.1016/j.cellsig.2024.111281] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2024] [Revised: 06/25/2024] [Accepted: 06/27/2024] [Indexed: 07/02/2024]
Abstract
Adenosine 5'-triphosphate (ATP) is a vital element in energy information. It plays a critical role in transmitting signals inside the body, which is necessary for controlling the life activities of all cells, including tumor cells [1]. Its significance extends from intracellular signaling pathways to tumor regression. Purinergic signaling, a form of extracellular paracrine signaling, relies on purine nucleotides. Extracellular ectonucleotidases convert these purine nucleotides to their respective di and mono-phosphate nucleoside forms, contributing significantly to immune biology, cancer biology, and inflammation studies. ATP functions as a mighty damage-linked molecular pattern when released outside the cell, accumulating in inflammatory areas. In the tumor microenvironment (TME), purinergic receptors such as ATP-gated ion channels P2X1-5 and G protein-coupled receptors (GPCR) (P2Y) interact with ATP and other nucleotides, influencing diverse immune cell activities. CD39 and CD73-mediated extracellular ATP degradation contributes to immunosuppression by diminishing ATP-dependent activation and generating adenosine (ADO), potentially hindering antitumor immunity and promoting tumor development. Unraveling the complexities of extracellular ATP (e-ATP) and ADO effects on the TME poses challenges in identifying optimal treatment targets, yet ongoing investigations aim to devise strategies combating e-ATP/ADO-induced immunosuppression, ultimately enhancing anti-tumor immunity. This review explores e-ATP metabolism, its purinergic signaling, and therapeutic strategies targeting associated receptors and enzymes.
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Affiliation(s)
- Sourav Shukla
- Immunology Lab, Biotechnology and Bioengineering, Indian Institute of Advanced Research, Gandhinagar 382426, Gujarat, India
| | - Parameswar Dalai
- Immunology Lab, Biotechnology and Bioengineering, Indian Institute of Advanced Research, Gandhinagar 382426, Gujarat, India
| | - Reena Agrawal-Rajput
- Immunology Lab, Biotechnology and Bioengineering, Indian Institute of Advanced Research, Gandhinagar 382426, Gujarat, India.
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