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Zheng W, Lai W, Zhang Y, Wang X, Yang K, Zhou X, Chen Z, Yang R, Deng L. Type 2 diabetes as a risk factor for promoting hypertrophic scar formation after wound healing: a two-sample mendelian randomization study. Arch Dermatol Res 2025; 317:656. [PMID: 40167795 PMCID: PMC11961473 DOI: 10.1007/s00403-025-04101-5] [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: 12/22/2024] [Revised: 02/21/2025] [Accepted: 02/25/2025] [Indexed: 04/02/2025]
Abstract
Individuals with type 2 diabetes mellitus (T2DM) not only face the risk of difficult wound healing after trauma or surgery but also struggle with scar formation after wound healing. However, the relationship between T2DM and hypertrophic scars (HS) remains unclear. First, the data sets related to T2DM and HS were downloaded from the OpenGWAS database. Second, the inverse variance-weighted (IVW) technique was employed to assess the causal impact of T2DM on HS utilizing mendelian randomization (MR). Finally, sensitivity analyses and reverse MR analyses were performed to determine the robustness of the findings and to examine the potential for reverse causation. A positive causal relationship between T2DM and HS was observed. Three suitable datasets for T2DM analysis were obtained using the ids ebi-a-GCST006867, ebi-a-GCST007515, and ebi-a-GCST007516 (A total of 206,572 cases and 753,191 controls were included). The HS (finn-b-L12_HYPETROPHICSCAR) dataset comprised 766 cases and 207,482 controls. The IVW model results showed odds ratio (OR) of 1.213 (95% confidence interval (CI): 1.04-1.41, p = 0.0137), 1.300 (95% CI: 1.05-1.61, p = 0.017), and 1.284 (95% CI: 1.03-1.60, p = 0.0276), respectively. The analysis results were stable without any significant bias or reverse causality. This study systematically explored the causal relationship between T2DM and the risk of HS for the first time, and provided a quantitative effect assessment. The results showed consistency, which indicated the potential biological mechanism and clinical intervention direction.
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Affiliation(s)
- Wenlian Zheng
- Department of Burn and Wound Repair, Shaoguan First People's Hospital, No. 3 Dongdi South Road, Zhenjiang, Shaoguan, Guangdong, 512000, China
| | - Wenxiu Lai
- Department of Urology, Yuebei People's Hospital, No. 133 Huimin South Road, Wujiang, Shaoguan, Guangdong, 512026, China
| | - Yixun Zhang
- Department of Burn and Plastic Surgery, Guangzhou First People's Hospital, Guangzhou Medical University, No. 1 Panfu Road, Yuexiu, Guangzhou, Guangdong, 510180, China
| | - Xiaoxiang Wang
- Department of Burn Surgery, The First Affiliated Hospital of Sun Yat-Sen University, No. 58, Zhongshan 2nd Road, Yuexiu, Guangzhou, Guangdong, 510080, China
| | - Kaibo Yang
- Department of Burn and Wound Repair, Shaoguan First People's Hospital, No. 3 Dongdi South Road, Zhenjiang, Shaoguan, Guangdong, 512000, China
| | - Xin Zhou
- Department of Burn and Plastic Surgery, The Second Affiliated Hospital, School of Medicine, South China University of Technology, No. 1 Panfu Road, Yuexiu, Guangzhou, Guangdong, 510180, China
| | - Zheng Chen
- Department of Burn and Wound Repair, Shaoguan First People's Hospital, No. 3 Dongdi South Road, Zhenjiang, Shaoguan, Guangdong, 512000, China.
| | - Ronghua Yang
- Department of Burn and Plastic Surgery, Guangzhou First People's Hospital, Guangzhou Medical University, No. 1 Panfu Road, Yuexiu, Guangzhou, Guangdong, 510180, China.
| | - Liaoyuan Deng
- Department of Burn and Wound Repair, Shaoguan First People's Hospital, No. 3 Dongdi South Road, Zhenjiang, Shaoguan, Guangdong, 512000, China.
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Rzhanova LA, Alpeeva EV, Aleksandrova MA. Using Small Molecules to Reprogram RPE Cells in Regenerative Medicine for Degenerative Eye Disease. Cells 2024; 13:1931. [PMID: 39682681 PMCID: PMC11640686 DOI: 10.3390/cells13231931] [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: 09/26/2024] [Revised: 11/13/2024] [Accepted: 11/18/2024] [Indexed: 12/18/2024] Open
Abstract
The main purpose of regenerative medicine for degenerative eye diseases is to create cells to replace lost or damaged ones. Due to their anatomical, genetic, and epigenetic features, characteristics of origin, evolutionary inheritance, capacity for dedifferentiation, proliferation, and plasticity, mammalian and human RPE cells are of great interest as endogenous sources of new photoreceptors and other neurons for the degrading retina. Promising methods for the reprogramming of RPE cells into retinal cells include genetic methods and chemical methods under the influence of certain low-molecular-weight compounds, so-called small molecules. Depending on the goal, which can be the preservation or the replacement of lost RPE cells and cellular structures, various small molecules are used to influence certain biological processes at different levels of cellular regulation. This review discusses the potential of the chemical reprogramming of RPE cells in comparison with other somatic cells and induced pluripotent stem cells (iPSCs) into neural cells of the brain and retina. Possible mechanisms of the chemically induced reprogramming of somatic cells under the influence of small molecules are explored and compared. This review also considers other possibilities in using them in the treatment of retinal degenerative diseases based on the protection, preservation, and support of survived RPE and retinal cells.
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Affiliation(s)
- Lyubov A. Rzhanova
- Koltzov Institute of Developmental Biology of the Russian Academy of Sciences, 26 Vavilov Street, 119334 Moscow, Russia;
| | - Elena V. Alpeeva
- Koltzov Institute of Developmental Biology of the Russian Academy of Sciences, 26 Vavilov Street, 119334 Moscow, Russia;
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Perez-Estrada JR, Tangeman JA, Proto-Newton M, Sanaka H, Smucker B, Del Rio-Tsonis K. Metabolic states influence chicken retinal pigment epithelium cell fate decisions. Development 2024; 151:dev202462. [PMID: 39120084 PMCID: PMC11708821 DOI: 10.1242/dev.202462] [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: 10/25/2023] [Accepted: 07/08/2024] [Indexed: 08/10/2024]
Abstract
During tissue regeneration, proliferation, dedifferentiation and reprogramming are necessary to restore lost structures. However, it is not fully understood how metabolism intersects with these processes. Chicken embryos can regenerate their retina through retinal pigment epithelium (RPE) reprogramming when treated with fibroblast factor 2 (FGF2). Using transcriptome profiling, we uncovered extensive regulation of gene sets pertaining to proliferation, neurogenesis and glycolysis throughout RPE-to-neural retina reprogramming. By manipulating cell media composition, we determined that glucose, glutamine or pyruvate are individually sufficient to support RPE reprogramming, identifying glycolysis as a requisite. Conversely, the activation of pyruvate dehydrogenase by inhibition of pyruvate dehydrogenase kinases, induces epithelial-to-mesenchymal transition, while simultaneously blocking the activation of neural retina fate. We also identified that epithelial-to-mesenchymal transition fate is partially driven by an oxidative environment. Our findings provide evidence that metabolism controls RPE cell fate decisions and provide insights into the metabolic state of RPE cells, which are prone to fate changes in regeneration and pathologies, such as proliferative vitreoretinopathy.
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Affiliation(s)
- J. Raúl Perez-Estrada
- Department of Biology, Miami University, Oxford, OH 45056, USA
- Center for Visual Sciences, Miami University, Oxford, OH 45056, USA
| | - Jared A. Tangeman
- Department of Biology, Miami University, Oxford, OH 45056, USA
- Center for Visual Sciences, Miami University, Oxford, OH 45056, USA
| | | | | | - Byran Smucker
- Center for Visual Sciences, Miami University, Oxford, OH 45056, USA
- Department of Statistics, Miami University, Oxford, OH 45056, USA
| | - Katia Del Rio-Tsonis
- Department of Biology, Miami University, Oxford, OH 45056, USA
- Center for Visual Sciences, Miami University, Oxford, OH 45056, USA
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Mysore Y, Hytti M, Deen AJ, Ranta-Aho S, Piippo N, Toppila M, Loukovaara S, Harju N, Kauppinen A. Epithelial-mesenchymal Transition (EMT) and the Effect of Atorvastatin on it in ARPE-19 cells. Cell Biochem Biophys 2024; 82:1523-1536. [PMID: 38777991 PMCID: PMC11344705 DOI: 10.1007/s12013-024-01305-w] [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] [Accepted: 05/07/2024] [Indexed: 05/25/2024]
Abstract
Proliferative vitreoretinopathy (PVR) develops after an unsuccessful or complicated recovery from rhegmatogenous retinal detachment (RRD) surgery. Intraocular scar formation with the contribution of epithelial-mesenchymal transition (EMT) in RPE cells is prominent in the pathology of PVR. In the present study, the EMT process was experimentally induced in human retinal pigment epithelium (RPE; ARPE-19) cells, and the effect of atorvastatin on the process was studied. The mRNA and protein levels of mesenchymal markers actin alpha 2 (ACTA2) / alpha-smooth muscle actin (α-SMA) and fibronectin (FN), and epithelial markers occludin (OCLN) and zonula occludens-1 (ZO-1) were measured using quantitative real-time PCR (qRT-PCR) and western blot methods, respectively. In addition, α-SMA and FN were visualized using immunofluorescence staining. Cells were photographed under a phase contrast light microscope. Changes in the functionality of cells following the EMT process were studied using the IncuCyte scratch wound cell migration assay and the collagen cell invasion assay with confocal microscopy. The induction of EMT in ARPE-19 cells increased the expression of mesenchymal markers ACTA2/α-SMA and fibronectin and reduced the expression of epithelial marker OCLN both at mRNA and protein levels. The mRNA levels of ZO-1 were lower after EMT, as well. Increased levels of α-SMA and FN were confirmed by immunofluorescence staining. Atorvastatin further increased the mRNA levels of mesenchymal markers ACTA2 and FN as well as the protein levels of α-SMA and reduced the mRNA levels of epithelial markers OCLN and ZO-1 under the EMT process. EMT promoted wound closure and cell invasion into the 3D collagen matrix when compared to untreated control cells. These data present cellular changes upon the induction of the EMT process in ARPE-19 cells and the propensity of atorvastatin to complement the effect. More studies are needed to confirm the exact influence of the EMT process and atorvastatin treatment on the PVR development after RRD surgery.
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Affiliation(s)
- Yashavanthi Mysore
- School of Pharmacy, Faculty of Health Sciences, University of Eastern Finland, Kuopio, Finland
| | - Maria Hytti
- School of Pharmacy, Faculty of Health Sciences, University of Eastern Finland, Kuopio, Finland
- Department of Ophthalmology, Kuopio University Hospital and School of Medicine, Faculty of Health Sciences, University of Eastern Finland, Kuopio, Finland
| | - Ashik Jawahar Deen
- A.I. Virtanen Institute for Molecular Sciences, University of Eastern Finland, Kuopio, Finland
| | - Sofia Ranta-Aho
- School of Pharmacy, Faculty of Health Sciences, University of Eastern Finland, Kuopio, Finland
| | - Niina Piippo
- School of Pharmacy, Faculty of Health Sciences, University of Eastern Finland, Kuopio, Finland
| | - Maija Toppila
- School of Pharmacy, Faculty of Health Sciences, University of Eastern Finland, Kuopio, Finland
| | - Sirpa Loukovaara
- Department of Ophthalmology, Unit of Vitreoretinal Surgery, Helsinki University Central Hospital, and Individualized Drug Therapy Research Program, University of Helsinki, Helsinki, Finland
| | - Niina Harju
- School of Pharmacy, Faculty of Health Sciences, University of Eastern Finland, Kuopio, Finland.
- Head and Neck Center, Ophthalmology Research Unit, Helsinki University Central Hospital, Helsinki, Finland.
| | - Anu Kauppinen
- School of Pharmacy, Faculty of Health Sciences, University of Eastern Finland, Kuopio, Finland.
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Zou G, Que L, Liu Y, Lu Q. Interplay of endothelial-mesenchymal transition, inflammation, and autophagy in proliferative diabetic retinopathy pathogenesis. Heliyon 2024; 10:e25166. [PMID: 38327444 PMCID: PMC10847601 DOI: 10.1016/j.heliyon.2024.e25166] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2023] [Revised: 01/13/2024] [Accepted: 01/22/2024] [Indexed: 02/09/2024] Open
Abstract
Background Assessment and validation of endothelial-mesenchymal transition (EndoMT) in the retinal endothelium of patients with proliferative diabetic retinopathy (PDR) at the level of retinal and vitreous specimens, and preliminary discussion of its regulatory mechanisms. Methods Transcriptome sequencing profiles of CD31+ cells from 9 retinal fibrovascular mem-branes (FVMs) and 4 postmortem retinas were downloaded from GEO databases to analyze EndoMT-related differentially expressed genes (DEGs). Then, 42 PDR patients and 34 idiopathic macular holes (IMH) patients were enrolled as the PDR and control groups, respectively. Vitreous humor (VH) samples were collected, and the expression of EndoMT-related proteins was quantified by enzyme-linked immunosorbent assay. Results A total of 5845 DEGs were identified, and we subsequently focused on the analysis of 24 EndoMT-related marker genes, including the trigger of EndoMT, endothelial genes, mesenchymal genes, transcription factors, inflammatory factors, and autophagy markers. Six of these genes were selected for protein assay validation in VH, showing increased mesenchymal marker (type I collagen α 2 chain, COL1A2) and decreased endothelial marker (VE-cadherin, CDH5) accompanied by increased TGFβ, IL-1β, LC3B and P62 in PDR patients. In addition, anti-VEGF therapy could enhance EndoMT-related phenotypes. Conclusions EndoMT may underlie the pathogenesis of PDR, and the induction and regulation correlate with autophagy defects and the inflammatory response.
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Affiliation(s)
- Gaocheng Zou
- Department of Ophthalmology, The First Affiliated Hospital of Anhui University of Traditional Chinese Medicine, Hefei, China
| | - Lijuan Que
- Department of Ophthalmology, The First Affiliated Hospital of Soochow University, Suzhou, China
| | - Yaping Liu
- State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guang-dong Provincial Key Laboratory of Ophthalmology and Visual Science, Guangzhou, China
| | - Qianyi Lu
- Department of Ophthalmology, The First Affiliated Hospital of Soochow University, Suzhou, China
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Shakour N, Karami S, Iranshahi M, Butler AE, Sahebkar A. Antifibrotic effects of sodium-glucose cotransporter-2 inhibitors: A comprehensive review. Diabetes Metab Syndr 2024; 18:102934. [PMID: 38154403 DOI: 10.1016/j.dsx.2023.102934] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/14/2023] [Revised: 11/25/2023] [Accepted: 12/20/2023] [Indexed: 12/30/2023]
Abstract
BACKGROUND AND AIMS Scar tissue accumulation in organs is the underlying cause of many fibrotic diseases. Due to the extensive array of organs affected, the long-term nature of fibrotic processes and the large number of people who suffer from the negative impact of these diseases, they constitute a serious health problem for modern medicine and a huge economic burden on society. Sodium-glucose cotransporter-2 inhibitors (SGLT2is) are a relatively new class of anti-diabetic pharmaceuticals that offer additional benefits over and above their glucose-lowering properties; these medications modulate a variety of diseases, including fibrosis. Herein, we have collated and analyzed all available research on SGLT2is and their effects on organ fibrosis, together with providing a proposed explanation as to the underlying mechanisms. METHODS PubMed, ScienceDirect, Google Scholar and Scopus were searched spanning the period from 2012 until April 2023 to find relevant articles describing the antifibrotic effects of SGLT2is. RESULTS The majority of reports have shown that SGLT2is are protective against lung, liver, heart and kidney fibrosis as well as arterial stiffness. According to the results of clinical trials and animal studies, many SGLT2 inhibitors are promising candidates for the treatment of fibrosis. Recent studies have demonstrated that SGLT2is affect an array of cellular processes, including hypoxia, inflammation, oxidative stress, the renin-angiotensin system and metabolic activities, all of which have been linked to fibrosis. CONCLUSION Extensive evidence indicates that SGLT2is are promising treatments for fibrosis, demonstrating protective effects in various organs and influencing key cellular processes linked to fibrosis.
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Affiliation(s)
- Neda Shakour
- Department of Medicinal Chemistry, School of Pharmacy, Mashhad University of Medical Sciences, Mashhad, Iran; Student Research Committee, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Shima Karami
- Department of Clinical Biochemistry, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Mehrdad Iranshahi
- Biotechnology Research Center, Pharmaceutical Technology Institute, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Alexandra E Butler
- Research Department, Royal College of Surgeons in Ireland, Adliya, Bahrain
| | - Amirhossein Sahebkar
- Biotechnology Research Center, Pharmaceutical Technology Institute, Mashhad University of Medical Sciences, Mashhad, Iran; Applied Biomedical Research Center, Mashhad University of Medical Sciences, Mashhad, Iran.
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Cao Z, Gao X, Meng J, Guo X, Xu J, Cui J, Zhou X. ACSL1: A preliminary study that provides a new target for the treatment of renal fibrosis could bring new insights in diabetic kidney disease. Nefrologia 2023; 43 Suppl 2:38-46. [PMID: 38245444 DOI: 10.1016/j.nefroe.2023.05.008] [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: 10/07/2022] [Accepted: 02/20/2023] [Indexed: 01/22/2024] Open
Abstract
BACKGROUND Renal fibrosis is the main cause of the development of diabetic kidney disease (DKD). ACSL1 plays an important role in colon cancer and liver fibrosis. METHODS We screened ACSL1 by proteomics analysis and then verified the expression of ACSL1 in the urine of diabetic nephropathy patients by WB and ELISA. Then, a total of 12db/m and db/db mice were used to verify the association between renal fibrosis and ACSL1. Periodic acid-Schiff (PAS) staining, Masson staining, and immunostaining were performed for histological studies. The relationship between ACSL1 and renal fibrosis was studied by knocking down ACSL1 in cell experiments. RESULTS The expression of ACSL1 was significantly increased in the exfoliated urine cells and urine supernatant of diabetic nephropathy patients and was closely related to renal function. In addition, the expression of ACSL1 was significantly increased in the renal tissues of db/db mice with fibrosis. Knocking down ACSL1 in HK-2 cells was shown to reverse renal fibrosis induced by high glucose. CONCLUSIONS We found a potential therapeutic target for preventing or ameliorating the progression of DKD fibrosis. Reducing ACSL1 expression may be a new strategy for the treatment of renal fibrosis caused by DKD, which provides an experimental theoretical basis for new drug research.
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Affiliation(s)
- Zhonghui Cao
- Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, College of Pharmacy, Xuzhou Medical University, Xuzhou 221004, China; Department of Pharmacy, Affiliated Hospital of Xuzhou Medical University, Xuzhou 221004, China
| | - Xiao Gao
- Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, College of Pharmacy, Xuzhou Medical University, Xuzhou 221004, China
| | - Jing Meng
- Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, College of Pharmacy, Xuzhou Medical University, Xuzhou 221004, China
| | - Xiaoli Guo
- Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, College of Pharmacy, Xuzhou Medical University, Xuzhou 221004, China
| | - Jiahao Xu
- Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, College of Pharmacy, Xuzhou Medical University, Xuzhou 221004, China
| | - Junchao Cui
- Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, College of Pharmacy, Xuzhou Medical University, Xuzhou 221004, China
| | - Xueyan Zhou
- Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, College of Pharmacy, Xuzhou Medical University, Xuzhou 221004, China.
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Perez-Estrada JR, Tangeman JA, Proto-Newton M, Sanaka H, Smucker B, Del Rio-Tsonis K. DISTINCT METABOLIC STATES DIRECT RETINAL PIGMENT EPITHELIUM CELL FATE DECISIONS. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.09.26.559631. [PMID: 37808829 PMCID: PMC10557760 DOI: 10.1101/2023.09.26.559631] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 10/10/2023]
Abstract
During tissue regeneration, proliferation, dedifferentiation, and reprogramming are necessary to restore lost structures. However, it is not fully understood how metabolism intersects with these processes. Chicken embryos can regenerate their retina through retinal pigment epithelium (RPE) reprogramming when treated with fibroblast factor 2 (FGF2). Using transcriptome profiling, we uncovered extensive regulation of gene sets pertaining to proliferation, neurogenesis, and glycolysis throughout RPE-to-neural retina reprogramming. By manipulating cell media composition, we determined that glucose, glutamine, or pyruvate are sufficient to support RPE reprogramming identifying glycolysis as a requisite. Conversely, the induction of oxidative metabolism by activation of pyruvate dehydrogenase induces Epithelial-to-mesenchymal transition (EMT), while simultaneously blocking the activation of neural retina fate. We also identify that EMT is partially driven by an oxidative environment. Our findings provide evidence that metabolism controls RPE cell fate decisions and provide insights into the metabolic state of RPE cells, which are prone to fate changes in regeneration and pathologies, such as proliferative vitreoretinopathy.
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9
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Shen G, Li Y, Zeng Y, Hong F, Zhang J, Wang Y, Zhang C, Xiang W, Wang J, Fang Z, Qi W, Yang X, Gao G, Zhou T. Kallistatin Deficiency Induces the Oxidative Stress-Related Epithelial-Mesenchymal Transition of Retinal Pigment Epithelial Cells: A Novel Protagonist in Age-Related Macular Degeneration. Invest Ophthalmol Vis Sci 2023; 64:15. [PMID: 37682567 PMCID: PMC10500364 DOI: 10.1167/iovs.64.12.15] [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: 04/03/2023] [Accepted: 08/15/2023] [Indexed: 09/09/2023] Open
Abstract
Purpose Retinal pigment epithelium (RPE) dysfunction induced by oxidative stress-related epithelial-mesenchymal transition (EMT) of RPE is the primary underlying mechanism of age-related macular degeneration (AMD). Kallistatin (KAL) is a secreted protein with an antioxidative stress effect. However, the relationship between KAL and EMT in RPE has not been determined. Therefore we aimed to explore the impact and mechanism of KAL in oxidative stress-induced EMT of RPE. Methods Sodium iodate (SI) was injected intraperitoneally to construct the AMD rat model and investigate the changes in RPE morphology and KAL expression. KAL knockout rats and KAL transgenic mice were used to explain the effects of KAL on EMT and oxidative stress. In addition, Snail overexpressed adenovirus and si-RNA transfected ARPE19 cells to verify the involvement of Snail in mediating KAL-suppressed EMT of RPE. Results AMD rats induced by SI expressed less KAL in the retina, and KAL knockout rats showed RPE dysfunction spontaneously where EMT and reactive oxygen species (ROS) production increased in RPE. In contrast, KAL overexpression attenuated EMT and ROS levels in RPE, even in TGF-β treatment. Mechanistically, Snail reversed the beneficial effect of KAL on EMT and ROS reduction. Moreover, KAL ameliorated SI-induced AMD-like pathological changes. Conclusions Our findings demonstrated that KAL inhibits oxidative stress-induced EMT by downregulating the transcription factor Snail. Herein, KAL knockout rats may be an appropriate animal model for observing spontaneous RPE dysfunction for AMD-like retinopathy, and KAL may represent a novel therapeutic target for treating dry AMD.
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Affiliation(s)
- Gang Shen
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
- Department of Laboratory Medicine, Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, China
| | - Yanmei Li
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Yongcheng Zeng
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Fuyan Hong
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Jing Zhang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Yan Wang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Chengwei Zhang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Wei Xiang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Jinhong Wang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Zhenzhen Fang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Weiwei Qi
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Xia Yang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
- Guangdong Province Key Laboratory of Brain Function and Disease, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Guoquan Gao
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
- Guangdong Engineering & Technology Research Center for Gene Manipulation and Biomacromolecular Products, Sun Yat-Sen University, Guangzhou, China
| | - Ti Zhou
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
- Advanced Medical Technology Center, The First Affiliated Hospital, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
- China Key Laboratory of Tropical Disease Control (Sun Yat-Sen University), Ministry of Education, Guangzhou, China
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Puddu A, Maggi DC. Klotho: A new therapeutic target in diabetic retinopathy? World J Diabetes 2023; 14:1027-1036. [PMID: 37547589 PMCID: PMC10401458 DOI: 10.4239/wjd.v14.i7.1027] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/16/2023] [Revised: 05/12/2023] [Accepted: 05/22/2023] [Indexed: 07/12/2023] Open
Abstract
Klotho (Kl) is considered an antiaging gene, mainly for the inhibition of the insulin-like growth factor-1 signaling. Kl exists as full-length transmembrane, which acts as co-receptor for fibroblast growth factor receptor, and in soluble forms (sKl). The sKl may exert pleiotropic effects on organs and tissues by regulating several pathways involved in the pathogenesis of diseases associated with oxidative and inflammatory state. In diabetic Patients, serum levels of Kl are significantly decreased compared to healthy subjects, and are related to duration of diabetes. In diabetic retinopathy (DR), one of the most common microvascular complications of type 2 diabetes, serum Kl levels are negatively correlated with progression of the disease. A lot of evidences showed that Kl regulates several mechanisms involved in maintaining homeostasis and functions of retinal cells, including phagocytosis, calcium signaling, secretion of vascular endothelial growth factor A (VEGF-A), maintenance of redox status, and melanin biosynthesis. Experimental data have been shown that Kl exerts positive effects on several mechanisms involved in onset and progression of DR. In particular, treatment with Kl: (1) Prevents apoptosis induced by oxidative stress in human retinal endothelial cells and in retinal pigment epithelium (RPE) cells; (2) reduces secretion of VEGF-A by RPE cells; and (3) decreases subretinal fibrosis and preserves autophagic activity. Therefore, Kl may become a novel biomarker and a good candidate for the treatment of DR.
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Affiliation(s)
- Alessandra Puddu
- Department of Internal Medicine and Medical Specialties, University of Genova, Genova 16132, Italy
| | - Davide Carlo Maggi
- Department of Internal Medicine and Medical Specialties, University of Genova, Genova 16132, Italy
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11
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Daley R, Maddipatla V, Ghosh S, Chowdhury O, Hose S, Zigler JS, Sinha D, Liu H. Aberrant Akt2 signaling in the RPE may contribute to retinal fibrosis process in diabetic retinopathy. Cell Death Discov 2023; 9:243. [PMID: 37443129 DOI: 10.1038/s41420-023-01545-4] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2023] [Revised: 06/12/2023] [Accepted: 07/04/2023] [Indexed: 07/15/2023] Open
Abstract
Diabetic Retinopathy (DR) is a complication of diabetes that causes blindness in adults. Retinal fibrosis is closely associated with developing proliferative diabetic retinopathy (PDR). Clinical studies have shown that fibrotic membranes exhibit uncontrolled growth in PDR and contribute to retinal detachment from RPE cells, ultimately leading to vision loss. While anti-VEGF agents and invasive laser treatments are the primary treatments for PDR, retinal fibrosis has received minimal attention as a potential target for therapeutic intervention. Therefore, to investigate the potential role of Akt2 in the diabetes-induced retinal fibrosis process, we generated RPE-specific Akt2 conditional knockout (cKO) mice and induced diabetes in these mice and Akt2fl/fl control mice by intraperitoneal injection of streptozotocin. After an 8-month duration of diabetes (10 months of age), the mice were euthanized and expression of tight junction proteins, epithelial-mesenchymal transition (EMT), and fibrosis markers were examined in the RPE. Diabetes induction in the floxed control mice decreased levels of the RPE tight junction protein ZO-1 and adherens junction proteins occludin and E-cadherin; these decreases were rescued in Akt2 cKO diabetic mice. Loss of Akt2 also inhibited diabetes-induced elevation of RNA and protein levels of the EMT markers Snail/Slug and Twist1 in the RPE as compared to Akt2fl/fl diabetic mice. We also found that in Akt2 cKO mice diabetes-induced increase of fibrosis markers, including collagen IV, Connective tissue growth factor (CTGF), fibronectin, and alpha-SMA was attenuated. Furthermore, we observed that high glucose-induced alterations in EMT and fibrosis markers in wild-type (WT) RPE explants were rescued in the presence of PI3K and ERK inhibitors, indicating diabetes-induced retinal fibrosis may be mediated via the PI3K/Akt2/ERK signaling, which could provide a novel target for DR therapy.
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Affiliation(s)
- Rachel Daley
- Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - Vishnu Maddipatla
- Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - Sayan Ghosh
- Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - Olivia Chowdhury
- Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - Stacey Hose
- Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
| | - J Samuel Zigler
- Wilmer Eye Institute, The Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Debasish Sinha
- Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA.
- Wilmer Eye Institute, The Johns Hopkins University School of Medicine, Baltimore, MD, USA.
| | - Haitao Liu
- Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA.
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12
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Yang J, Hua Z, Zheng Z, Ma X, Zhu L, Li Y. Acteoside inhibits high glucose-induced oxidative stress injury in RPE cells and the outer retina through the Keap1/Nrf2/ARE pathway. Exp Eye Res 2023; 232:109496. [PMID: 37268044 DOI: 10.1016/j.exer.2023.109496] [Citation(s) in RCA: 11] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/19/2022] [Revised: 04/20/2023] [Accepted: 05/03/2023] [Indexed: 06/04/2023]
Abstract
Diabetes retinopathy (DR) is one of the most common microvascular complications of diabetes. Retinal pigment epithelial (RPE) cells exposed to a high glucose environment experience a series of functional damages, which is an important factor in promoting the progression of DR. Acteoside (ACT) has strong antioxidant and anti-apoptotic properties, but the mechanism of ACT in DR is not completely clear. Therefore, the purpose of the present study was to explore whether ACT inhibits the damage to RPE cells in a high glucose environment through antioxidative effects to alleviate the DR process. The DR in vitro cell model was constructed by treating RPE cells with high glucose, and the DR in vivo animal model was constructed by injecting streptozotocin (STZ) into the peritoneal cavity of mice to induce diabetes. The proliferation and apoptosis of RPE cells were detected by CCK-8 and flow cytometry assays, respectively. The expression changes in Nrf2, Keap1, NQO1 and HO-1 were evaluated by qRT‒PCR, Western blot and immunohistochemistry analyses. The MDA, SOD, GSH-Px and T-AOC contents were detected by kits. The changes in ROS and nuclear translocation of Nrf2 were observed by immunofluorescence assays. HE staining was used to measure the thickness of the outer nuclear layer (ONL) of the retina, and TUNEL staining was used to detect the number of apoptotic cells in the retinas of mice. In the present study, ACT effectively ameliorated outer retina damage in diabetic mice. In high glucose (HG)-induced RPE cells, ACT treatment had the following effects: improved proliferation, decreased apoptosis, inhibited Keap1 expression, promoted the nuclear translocation and expression of Nrf2, upregulated NQO1 and HO-1 (the target genes of Nrf2) expression, decreased ROS concentration, and increased the levels of the SOD, GSH-Px and T-AOC antioxidant indicators. However, knockdown of Nrf2 reversed the above phenomena, which indicated that the protective function of ACT in HG-induced RPE cells are closely related to Nrf2. In summary, the present study demonstrated that HG-induced oxidative stress injury is inhibited by ACT in RPE cells and the outer retina through the Keap1/Nrf2/ARE pathway.
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Affiliation(s)
- Jingfei Yang
- Department of Ophthalmology, The First Affiliated Hospital of Kunming Medical University, Kunming, 650032, China; Department of Ophthalmology, The Affiliated Hospital of Yunnan University, Kunming, 650021, China
| | - Zhijuan Hua
- Department of Pediatric Ophthalmology, The Affiliated Hospital of Yunnan University, Kunming, 650021, China
| | - Zhikun Zheng
- Department of Ophthalmology, The Affiliated Hospital of Yunnan University, Kunming, 650021, China
| | - Xuan Ma
- Department of Ophthalmology, The Affiliated Hospital of Yunnan University, Kunming, 650021, China
| | - Liang Zhu
- Department of Ophthalmology, The First Affiliated Hospital of Kunming Medical University, Kunming, 650032, China
| | - Yan Li
- Department of Ophthalmology, The First Affiliated Hospital of Kunming Medical University, Kunming, 650032, China.
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13
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Xiang W, Li L, Hong F, Zeng Y, Zhang J, Xie J, Shen G, Wang J, Fang Z, Qi W, Yang X, Gao G, Zhou T. N-cadherin cleavage: A critical function that induces diabetic retinopathy fibrosis via regulation of β-catenin translocation. FASEB J 2023; 37:e22878. [PMID: 36939278 DOI: 10.1096/fj.202201664rr] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2022] [Revised: 02/15/2023] [Accepted: 03/02/2023] [Indexed: 03/21/2023]
Abstract
Retinal fibrosis is a severe pathological change in the late stage of diabetic retinopathy and is also the leading cause of blindness. We have previously revealed that N-cadherin was significantly increased in type 1 and type 2 diabetic mice retinas and the fibrovascular membranes from proliferative diabetic retinopathy (PDR) patients. However, whether N-cadherin directly induces retinal fibrosis in DR and the related mechanism is unknown. Here, we investigated the pathogenic role of N-cadherin in mediating retinal fibrosis and further explored the relevant therapeutic targets. We found that the level of N-cadherin was significantly increased in PDR patients and STZ-induced diabetic mice and positively correlated with the fibrotic molecules Connective Tissue Growth Factor (CTGF) and fibronectin (FN). Moreover, intravitreal injection of N-cadherin adenovirus significantly increased the expression of FN and CTGF in normal mice retinas. Mechanistically, overexpression of N-cadherin promotes N-cadherin cleavage, and N-cadherin cleavage can further induce translocation of non-p-β-catenin in the nucleus and upregulation of fibrotic molecules. Furthermore, we found a novel N-cadherin cleavage inhibitor, pigment epithelial-derived factor (PEDF), which ameliorated the N-cadherin cleavage and subsequent retinal fibrosis in diabetic mice. Thus, our findings provide novel evidence that elevated N-cadherin level not only acts as a classic EMT maker but also plays a causative role in diabetic retinal fibrosis, and targeting N-cadherin cleavage may provide a strategy to inhibit retinal fibrosis in DR patients.
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Affiliation(s)
- Wei Xiang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
- Department of Clinical Laboratory Medicine, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Southern Medical University, Guangzhou, China
| | - Longhui Li
- State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, China
| | - Fuyan Hong
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
| | - Yongcheng Zeng
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
| | - Jin Zhang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
| | - Jinye Xie
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
| | - Gang Shen
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
| | - Jinhong Wang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
| | - Zhenzhen Fang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
| | - Weiwei Qi
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
- Guangdong Province Key Laboratory of Brain Function and Disease, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Xia Yang
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
- Guangdong Province Key Laboratory of Brain Function and Disease, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, China
| | - Guoquan Gao
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
- Guangdong Engineering & Technology Research Center for Gene Manipulation and Biomacromolecular Products, Sun Yat-Sen University, Guangzhou, China
| | - Ti Zhou
- Department of Biochemistry and Molecular Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
- Advanced Medical Technology Center, The First Affiliated Hospital, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
- China Key Laboratory of Tropical Disease Control (Sun Yat-sen University), Ministry of Education, Guangzhou, China
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14
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Syed MA, Bhat B, Wali A, Saleem A, Ahmad Dar L, Gugjoo MB, Bhat S, Saleem Bhat S. Epithelial to mesenchymal transition in mammary gland tissue fibrosis and insights into drug therapeutics. PeerJ 2023; 11:e15207. [PMID: 37187521 PMCID: PMC10178283 DOI: 10.7717/peerj.15207] [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: 11/14/2022] [Accepted: 03/19/2023] [Indexed: 05/17/2023] Open
Abstract
Background The epithelial-mesenchymal transition (EMT) is a multi-step morphogenetic process in which epithelial cells lose their epithelial properties and gain mesenchymal characteristics. The process of EMT has been shown to mediate mammary gland fibrosis. Understanding how mesenchymal cells emerge from an epithelial default state will aid in unravelling the mechanisms that control fibrosis and, ultimately, in identifying therapeutic targets to alleviate fibrosis. Methods The effects of EGF and high glucose (HG) on EMT in mammary epithelial cells, MCF10A and GMECs, as well as their pathogenic role, were studied. In-silico analysis was used to find interacting partners and protein-chemical/drug molecule interactions. Results On treatment with EGF and/or HG, qPCR analysis showed a significant increase in the gene expression of EMT markers and downstream signalling genes. The expression of these genes was reduced on treatment with EGF+HG combination in both cell lines. The protein expression of COL1A1 increased as compared to the control in cells treated with EGF or HG alone, but when the cells were treated with EGF and HG together, the protein expression of COL1A1 decreased. ROS levels and cell death increased in cells treated with EGF and HG alone, whereas cells treated with EGF and HG together showed a decrease in ROS production and apoptosis. In-silico analysis of protein-protein interactions suggest the possible role of MAPK1, actin alpha 2 (ACTA2), COL1A1, and NFκB1 in regulating TGFβ1, ubiquitin C (UBC), specificity protein 1 (SP1) and E1A binding protein P300 (EP300). Kyoto Encyclopaedia of Genes and Genomes (KEGG) enrichment suggests advanced glycation end products-receptor for advanced glycation end products (AGE-RAGE) signalling pathway, relaxin signalling pathway and extra cellular matrix (ECM) receptor interactions underlying fibrosis mechanism. Conclusion This study demonstrates that EGF and HG induce EMT in mammary epithelial cells and may also have a role in fibrosis.
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Affiliation(s)
- Mudasir Ahmad Syed
- Division of Animal Biotechnology, Faculty of Veterinary Sciences, Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir, India, Srinagar, India
| | - Basharat Bhat
- Division of Animal Biotechnology, Faculty of Veterinary Sciences, Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir, India, Srinagar, India
| | - Abiza Wali
- Department of Clinical Biochemistry, University of Kashmir, Srinagar, Jammu and Kashmir, India
| | - Afnan Saleem
- Division of Animal Biotechnology, Faculty of Veterinary Sciences, Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir, India, Srinagar, India
| | - Lateef Ahmad Dar
- Division of Animal Biotechnology, Faculty of Veterinary Sciences, Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir, India, Srinagar, India
| | - Mudasir Bashir Gugjoo
- Division of Veterinary Surgery, Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir, Faculty of Veterinary Sciences and Animal Husbandry, Shuhama, SKUAST-K, India, Srinagar, Jammu and Kashmir, India
| | - Shakil Bhat
- Division of Animal Biotechnology, Faculty of Veterinary Sciences, Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir, India, Srinagar, India
| | - Sahar Saleem Bhat
- Division of Animal Biotechnology, Faculty of Veterinary Sciences, Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir, India, Srinagar, India
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15
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Exosomal miRNA Profiling in Vitreous Humor in Proliferative Diabetic Retinopathy. Cells 2022; 12:cells12010123. [PMID: 36611916 PMCID: PMC9818905 DOI: 10.3390/cells12010123] [Citation(s) in RCA: 9] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2022] [Revised: 12/14/2022] [Accepted: 12/22/2022] [Indexed: 12/31/2022] Open
Abstract
MicroRNAs (miRNAs) are small noncoding RNAs which mediate some of the pathological mechanisms of diabetic retinopathy. The aim of this study was to identify differentially expressed miRNAs in the vitreal exosomes of proliferative diabetic retinopathy (PDR) patients and non-diabetic controls. Exosomes were extracted from the vitreous samples of 10 PDR patients and 10 controls. The expression of 372 miRNAs was determined using a quantitative polymerase chain reaction (qPCR) panel. We have demonstrated a significant dysregulation in 26 miRNAs. The most remarkable findings include a profound attenuation of the miR-125 family, as well as enhanced miR-21-5p expression in the diabetic samples. We also showed the downregulation of miR-204-5p and the upregulation of let-7g in PDR compared to the controls. This study identified miR-125 and miR-21 as potential targets for further functional analysis regarding their putative role in the pathogenesis of PDR.
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16
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CircRNA SCMH1 regulates the miR-200a-3p/ZEB1 signaling axis to promote diabetes-induced retinal epithelial-mesenchymal transition. Exp Eye Res 2022; 224:109264. [PMID: 36162459 DOI: 10.1016/j.exer.2022.109264] [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: 01/11/2022] [Revised: 08/23/2022] [Accepted: 09/19/2022] [Indexed: 11/22/2022]
Abstract
Diabetic retinopathy (DR) is one of the common systemic complications of diabetes. Epithelial-mesenchymal transition (EMT) is required for DR progression. Previous studies have explored that circular RNAs (circRNAs) are crucial for DR development. Herein, we focused on the biological functions of circSCMH1 in DR. RT-qPCR determined the expression of circSCMH1, miR-200a-3p and ZEB1. EMT-related proteins were measured by Western blot. Gene combinations were validated by RIP and dual luciferase reporter assays. CCK-8, EdU, TUNEL staining and Transwell analysis were used to assess the cellular function. FISH analysis assessed the localization of circSCMH1 and miR-200a-3p. HE staining was used to detect retinal structures in a mouse DR model. High-glucose (HG) significantly increased circSCMH1 expression in ARPE-19 cells. Additionally, circSCMH1 silencing repressed proliferation, migration, and EMT in HG cells. Mechanistically, circSCMH1 positively regulated ZEB1 expression via targeting miR-200a-3p. Furthermore, circSCMH1 was observed to induce HG cell growth and EMT by regulating the miR-200a-3p/ZEB1 axis. Finally, we verified that downregulation of circSCMH1 or ZEB1 alleviated EMT in the retina of diabetic mice. These findings have implications for new therapeutic targets for DR.
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17
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Trotta MC, Petrillo F, Gesualdo C, Rossi S, Corte AD, Váradi J, Fenyvesi F, D’Amico M, Hermenean A. Effects of the Calix[4]arene Derivative Compound OTX008 on High Glucose-Stimulated ARPE-19 Cells: Focus on Galectin-1/TGF-β/EMT Pathway. MOLECULES (BASEL, SWITZERLAND) 2022; 27:molecules27154785. [PMID: 35897964 PMCID: PMC9332238 DOI: 10.3390/molecules27154785] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 06/27/2022] [Revised: 07/18/2022] [Accepted: 07/22/2022] [Indexed: 11/16/2022]
Abstract
Diabetic retinopathy (DR) is a neurovascular disease characterized by the reduction of retina integrity and functionality, as a consequence of retinal pigment epithelial cell fibrosis. Although galectin-1 (a glycan-binding protein) has been associated with dysregulated retinal angiogenesis, no evidence has been reported about galectin-1 roles in DR-induced fibrosis. ARPE-19 cells were cultured in normal (5 mM) or high glucose (35 mM) for 3 days, then exposed to the selective galectin-1 inhibitor OTX008 (2.5–5–10 μM) for 6 days. The determination of cell viability and ROS content along with the analysis of specific proteins (by immunocytochemistry, Western blotting, and ELISA) or mRNAs (by real time-PCR) were performed. OTX008 5 μM and 10 μM improved cell viability and markedly reduced galectin-1 protein expression in cells exposed to high glucose. This was paralleled by a down-regulation of the TGF-β/, NF-kB p65 levels, and ROS content. Moreover, epithelial–mesenchymal transition markers were reduced by OTX008 5 μM and 10 μM. The inhibition of galectin-1 by OTX008 in DR may preserve retinal pigment epithelial cell integrity and functionality by reducing their pro-fibrotic phenotype and epithelial–mesenchymal transition phenomenon induced by diabetes.
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Affiliation(s)
- Maria Consiglia Trotta
- Department of Experimental Medicine, University of Campania “Luigi Vanvitelli”, 80138 Naples, Italy; (M.C.T.); (M.D.)
| | - Francesco Petrillo
- PhD Course in Translational Medicine, Department of Experimental Medicine, University of Campania “Luigi Vanvitelli”, 80138 Naples, Italy;
| | - Carlo Gesualdo
- Multidisciplinary Department of Medical, Surgical and Dental Sciences, Eye Clinic, University of Campania “Luigi Vanvitelli”, 80138 Naples, Italy; (C.G.); (S.R.); (A.D.C.)
| | - Settimio Rossi
- Multidisciplinary Department of Medical, Surgical and Dental Sciences, Eye Clinic, University of Campania “Luigi Vanvitelli”, 80138 Naples, Italy; (C.G.); (S.R.); (A.D.C.)
| | - Alberto Della Corte
- Multidisciplinary Department of Medical, Surgical and Dental Sciences, Eye Clinic, University of Campania “Luigi Vanvitelli”, 80138 Naples, Italy; (C.G.); (S.R.); (A.D.C.)
| | - Judit Váradi
- Department of Pharmaceutical Technology, Faculty of Pharmacy, University of Debrecen, Nagyerdei St. 98, H-4032 Debrecen, Hungary; (J.V.); (F.F.)
| | - Ferenc Fenyvesi
- Department of Pharmaceutical Technology, Faculty of Pharmacy, University of Debrecen, Nagyerdei St. 98, H-4032 Debrecen, Hungary; (J.V.); (F.F.)
| | - Michele D’Amico
- Department of Experimental Medicine, University of Campania “Luigi Vanvitelli”, 80138 Naples, Italy; (M.C.T.); (M.D.)
| | - Anca Hermenean
- Faculty of Medicine, Vasile Goldis Western University of Arad, 310414 Arad, Romania
- Correspondence:
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18
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Liu YY, Liu HY, Yu TJ, Lu Q, Zhang FL, Liu GY, Shao ZM, Li DQ. O-GlcNAcylation of MORC2 at threonine 556 by OGT couples TGF-β signaling to breast cancer progression. Cell Death Differ 2022; 29:861-873. [PMID: 34974534 PMCID: PMC8991186 DOI: 10.1038/s41418-021-00901-0] [Citation(s) in RCA: 51] [Impact Index Per Article: 17.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2021] [Revised: 11/03/2021] [Accepted: 11/10/2021] [Indexed: 12/11/2022] Open
Abstract
MORC family CW-type zinc finger 2 (MORC2) is a newly identified chromatin-remodeling enzyme involved in DNA damage response and gene transcription, and its dysregulation has been linked with Charcot-Marie-Tooth disease, neurodevelopmental disorder, and cancer. Despite its functional importance, how MORC2 is regulated remains enigmatic. Here, we report that MORC2 is O-GlcNAcylated by O-GlcNAc transferase (OGT) at threonine 556. Mutation of this site or pharmacological inhibition of OGT impairs MORC2-mediated breast cancer cell migration and invasion in vitro and lung colonization in vivo. Moreover, transforming growth factor-β1 (TGF-β1) induces MORC2 O-GlcNAcylation through enhancing the stability of glutamine-fructose-6-phosphate aminotransferase (GFAT), the rate-limiting enzyme for producing the sugar donor for OGT. O-GlcNAcylated MORC2 is required for transcriptional activation of TGF-β1 target genes connective tissue growth factor (CTGF) and snail family transcriptional repressor 1 (SNAIL). In support of these observations, knockdown of GFAT, SNAIL or CTGF compromises TGF-β1-induced, MORC2 O-GlcNAcylation-mediated breast cancer cell migration and invasion. Clinically, high expression of OGT, MORC2, SNAIL, and CTGF in breast tumors is associated with poor patient prognosis. Collectively, these findings uncover a previously unrecognized mechanistic role for MORC2 O-GlcNAcylation in breast cancer progression and provide evidence for targeting MORC2-dependent breast cancer through blocking its O-GlcNAcylation.
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Affiliation(s)
- Ying-Ying Liu
- grid.8547.e0000 0001 0125 2443Fudan University Shanghai Cancer Center and Shanghai Key Laboratory of Medical Epigenetics, International Co-laboratory of Medical Epigenetics and Metabolism, Ministry of Science and Technology, Institutes of Biomedical Sciences, Fudan University, Shanghai, 200032 China ,grid.8547.e0000 0001 0125 2443Cancer Institute, Shanghai Medical College, Fudan University, Shanghai, 200032 China ,grid.8547.e0000 0001 0125 2443Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, 200032 China ,grid.8547.e0000 0001 0125 2443Department of Breast Surgery, Shanghai Medical College, Fudan University, Shanghai, 200032 China
| | - Hong-Yi Liu
- grid.8547.e0000 0001 0125 2443Fudan University Shanghai Cancer Center and Shanghai Key Laboratory of Medical Epigenetics, International Co-laboratory of Medical Epigenetics and Metabolism, Ministry of Science and Technology, Institutes of Biomedical Sciences, Fudan University, Shanghai, 200032 China
| | - Tian-Jian Yu
- grid.8547.e0000 0001 0125 2443Fudan University Shanghai Cancer Center and Shanghai Key Laboratory of Medical Epigenetics, International Co-laboratory of Medical Epigenetics and Metabolism, Ministry of Science and Technology, Institutes of Biomedical Sciences, Fudan University, Shanghai, 200032 China ,grid.8547.e0000 0001 0125 2443Cancer Institute, Shanghai Medical College, Fudan University, Shanghai, 200032 China ,grid.8547.e0000 0001 0125 2443Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, 200032 China ,grid.8547.e0000 0001 0125 2443Department of Breast Surgery, Shanghai Medical College, Fudan University, Shanghai, 200032 China
| | - Qin Lu
- grid.8547.e0000 0001 0125 2443Fudan University Shanghai Cancer Center and Shanghai Key Laboratory of Medical Epigenetics, International Co-laboratory of Medical Epigenetics and Metabolism, Ministry of Science and Technology, Institutes of Biomedical Sciences, Fudan University, Shanghai, 200032 China
| | - Fang-Lin Zhang
- grid.8547.e0000 0001 0125 2443Fudan University Shanghai Cancer Center and Shanghai Key Laboratory of Medical Epigenetics, International Co-laboratory of Medical Epigenetics and Metabolism, Ministry of Science and Technology, Institutes of Biomedical Sciences, Fudan University, Shanghai, 200032 China ,grid.8547.e0000 0001 0125 2443Cancer Institute, Shanghai Medical College, Fudan University, Shanghai, 200032 China ,grid.8547.e0000 0001 0125 2443Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, 200032 China
| | - Guang-Yu Liu
- Department of Breast Surgery, Shanghai Medical College, Fudan University, Shanghai, 200032, China.
| | - Zhi-Ming Shao
- Fudan University Shanghai Cancer Center and Shanghai Key Laboratory of Medical Epigenetics, International Co-laboratory of Medical Epigenetics and Metabolism, Ministry of Science and Technology, Institutes of Biomedical Sciences, Fudan University, Shanghai, 200032, China. .,Cancer Institute, Shanghai Medical College, Fudan University, Shanghai, 200032, China. .,Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, 200032, China. .,Department of Breast Surgery, Shanghai Medical College, Fudan University, Shanghai, 200032, China. .,Shanghai Key Laboratory of Breast Cancer, Shanghai Medical College, Fudan University, Shanghai, 200032, China.
| | - Da-Qiang Li
- Fudan University Shanghai Cancer Center and Shanghai Key Laboratory of Medical Epigenetics, International Co-laboratory of Medical Epigenetics and Metabolism, Ministry of Science and Technology, Institutes of Biomedical Sciences, Fudan University, Shanghai, 200032, China. .,Cancer Institute, Shanghai Medical College, Fudan University, Shanghai, 200032, China. .,Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, 200032, China. .,Department of Breast Surgery, Shanghai Medical College, Fudan University, Shanghai, 200032, China. .,Shanghai Key Laboratory of Breast Cancer, Shanghai Medical College, Fudan University, Shanghai, 200032, China. .,Shanghai Key Laboratory of Radiation Oncology, Shanghai Medical College, Fudan University, Shanghai, 200032, China.
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Cliff CL, Williams BM, Chadjichristos CE, Mouritzen U, Squires PE, Hills CE. Connexin 43: A Target for the Treatment of Inflammation in Secondary Complications of the Kidney and Eye in Diabetes. Int J Mol Sci 2022; 23:600. [PMID: 35054783 PMCID: PMC8776095 DOI: 10.3390/ijms23020600] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/26/2021] [Revised: 01/03/2022] [Accepted: 01/04/2022] [Indexed: 02/06/2023] Open
Abstract
Of increasing prevalence, diabetes is characterised by elevated blood glucose and chronic inflammation that precedes the onset of multiple secondary complications, including those of the kidney and the eye. As the leading cause of end stage renal disease and blindness in the working population, more than ever is there a demand to develop clinical interventions which can both delay and prevent disease progression. Connexins are membrane bound proteins that can form pores (hemichannels) in the cell membrane. Gated by cellular stress and injury, they open under pathophysiological conditions and in doing so release 'danger signals' including adenosine triphosphate into the extracellular environment. Linked to sterile inflammation via activation of the nod-like receptor protein 3 inflammasome, targeting aberrant hemichannel activity and the release of these danger signals has met with favourable outcomes in multiple models of disease, including secondary complications of diabetes. In this review, we provide a comprehensive update on those studies which document a role for aberrant connexin hemichannel activity in the pathogenesis of both diabetic eye and kidney disease, ahead of evaluating the efficacy of blocking connexin-43 specific hemichannels in these target tissues on tissue health and function.
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Affiliation(s)
- Chelsy L. Cliff
- Joseph Banks Laboratories, School of Life, Sciences University of Lincoln, Lincoln LN6 7DL, UK; (C.L.C.); (B.M.W.); (P.E.S.)
| | - Bethany M. Williams
- Joseph Banks Laboratories, School of Life, Sciences University of Lincoln, Lincoln LN6 7DL, UK; (C.L.C.); (B.M.W.); (P.E.S.)
| | - Christos E. Chadjichristos
- National Institutes for Health and Medical Research, UMR-S1155, Batiment Recherche, Tenon Hospital, 4 Rue de la Chine, 75020 Paris, France;
| | - Ulrik Mouritzen
- Ciana Therapeutics, Ole Maaloes Vej 3, 2200 Copenhagen N, Denmark;
| | - Paul E. Squires
- Joseph Banks Laboratories, School of Life, Sciences University of Lincoln, Lincoln LN6 7DL, UK; (C.L.C.); (B.M.W.); (P.E.S.)
| | - Claire E. Hills
- Joseph Banks Laboratories, School of Life, Sciences University of Lincoln, Lincoln LN6 7DL, UK; (C.L.C.); (B.M.W.); (P.E.S.)
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20
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Recent Advances in Diabetic Kidney Diseases: From Kidney Injury to Kidney Fibrosis. Int J Mol Sci 2021; 22:ijms222111857. [PMID: 34769288 PMCID: PMC8584225 DOI: 10.3390/ijms222111857] [Citation(s) in RCA: 74] [Impact Index Per Article: 18.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2021] [Revised: 10/08/2021] [Accepted: 10/30/2021] [Indexed: 12/14/2022] Open
Abstract
Diabetic kidney disease (DKD) is the leading cause of chronic kidney disease and end-stage renal disease. The natural history of DKD includes glomerular hyperfiltration, progressive albuminuria, declining estimated glomerular filtration rate, and, ultimately, kidney failure. It is known that DKD is associated with metabolic changes caused by hyperglycemia, resulting in glomerular hypertrophy, glomerulosclerosis, and tubulointerstitial inflammation and fibrosis. Hyperglycemia is also known to cause programmed epigenetic modification. However, the detailed mechanisms involved in the onset and progression of DKD remain elusive. In this review, we discuss recent advances regarding the pathogenic mechanisms involved in DKD.
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21
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Lyon H, Yin N, Rupenthal ID, Green CR, Mugisho OO. Blocking connexin43 hemichannels prevents TGF-β2 upregulation and epithelial-mesenchymal transition in retinal pigment epithelial cells. Cell Biol Int 2021; 46:323-330. [PMID: 34719065 DOI: 10.1002/cbin.11718] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/30/2021] [Revised: 10/10/2021] [Accepted: 10/23/2021] [Indexed: 12/14/2022]
Abstract
Epithelial-mesenchymal transition (EMT) occurs when polarised epithelial cells change to a mesenchymal phenotype. EMT plays a role in several chronic conditions, including ocular diseases with retinal pigment epithelium (RPE) EMT associated with retinal diseases such as diabetic retinopathy (DR). Here, EMT results in breakdown of the blood-retinal barrier (BRB) leading to sub-retinal fluid deposition and retinal detachment. Previous studies have shown that blocking connexin43 (Cx43) hemichannels can protect against RPE BRB breakdown, but the underlying mechanism is unknown. To determine whether open Cx43 hemichannels may enable EMT of RPE cells and thus result in BRB breakdown, ARPE-19 cells were either challenged with high glucose plus the inflammatory cytokines IL-1β and TNF-α (HG + Cyt) to simulate DR or treated with the Cx43 hemichannel blocker tonabersat alongside the HG + Cyt challenge. HG + Cyt induced a morphological change in RPE cells to a fibroblastic phenotype with a corresponding decrease in epithelial zonular occludens-1 and an increase in the fibroblastic marker α-SMA. The HG + Cyt challenge also induced loss of transepithelial electrical resistance while increasing dye passage between RPE cells. All of these changes were significantly reduced with tonabersat treatment, which also prevented HG + Cyt-induced transforming growth factor-β2 (TGF-β2) release. In conclusion, Cx43 hemichannel block with tonabersat attenuated both TGF-β2 release and RPE EMT under disease-mimicking conditions, offering the potential to ameliorate the progression of EMT-associated retinal diseases.
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Affiliation(s)
- Heather Lyon
- Buchanan Ocular Therapeutics Unit, Department of Ophthalmology, New Zealand National Eye Centre, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand
| | - Naibo Yin
- Buchanan Ocular Therapeutics Unit, Department of Ophthalmology, New Zealand National Eye Centre, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand
| | - Ilva D Rupenthal
- Buchanan Ocular Therapeutics Unit, Department of Ophthalmology, New Zealand National Eye Centre, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand
| | - Colin R Green
- Department of Ophthalmology, New Zealand National Eye Centre, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand
| | - Odunayo O Mugisho
- Buchanan Ocular Therapeutics Unit, Department of Ophthalmology, New Zealand National Eye Centre, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand
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22
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Yang Y, Zhou J, Li WH, Zhou ZX, Xia XB. LncRNA NEAT1 regulated diabetic retinal epithelial-mesenchymal transition through regulating miR-204/SOX4 axis. PeerJ 2021; 9:e11817. [PMID: 34386303 PMCID: PMC8312494 DOI: 10.7717/peerj.11817] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/09/2020] [Accepted: 06/29/2021] [Indexed: 12/18/2022] Open
Abstract
AIM Epithelial-mesenchymal transition (EMT) of retinal pigment epithelium (RPE) cells is the key of the development of diabetic retinopathy (DR), and lncRNA NEAT1 could accelerate EMT in diabetic nephropathy. Meanwhile, as a diabetes susceptibility gene, whether sex-determining region Y-related (SRY) high-mobility group box 4 (SOX4) has relationship with lncRNA NEAT1 in DR remains unclear. METHODS Firstly, NEAT1, SOX4 and miR-204 were evaluated by qRT-PCR (quantitative reverse-transcriptase PCR) under high glucose condition. Then, cell viability, proliferation, migration and invasion were respectively detected by MTT, BrdU staining, wound healing and transwell assay after NEAT1 knockdown or miR-204 overexpression. Also, the EMT-related proteins were examined by western blot and cell immunofluorescence assay. In order to confirm the relationship between miR-204 and NEAT1 or SOX4, dual luciferase reporter gene assay was conducted. At the same time, the protein levels of SOX4 and EMT-related proteins were investigated by immunohistochemistry in vivo. RESULTS High glucose upregulated NEAT1 and SOX4 and downregulated miR-204 in ARPE19 cells. NEAT1 knockdown or miR-204 overexpression inhibited the proliferation and EMT progression of ARPE19 cells induced by high glucose. NEAT1 was identified as a molecular sponge of miR-204 to increase the level of SOX4. The effect of NEAT1 knockdown on the progression of EMT under high glucose condition in ARPE19 cells could be reversed by miR-204 inhibitor. Also, NEAT1 knockdown inhibited retinal EMT in diabetic mice. CONCLUSION NEAT1 regulated the development of EMT in DR through miR-204/SOX4 pathway, which could provide reference for clinical prevention and treatment.
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Affiliation(s)
- Yang Yang
- Eye center of Xiangya Hospital, Central South University, Changsha, Hunan Province, China
- Department of Ophthalmology, the First People’s Hospital of Yueyang, Yueyang, Hunan, China
| | - Jing Zhou
- Department of Ophthalmology, the First People’s Hospital of Yueyang, Yueyang, Hunan, China
| | - Wei hong Li
- Department of Ophthalmology, the First People’s Hospital of Yueyang, Yueyang, Hunan, China
| | - Zhi xiong Zhou
- Department of Ophthalmology, the First People’s Hospital of Yueyang, Yueyang, Hunan, China
| | - Xiao bo Xia
- Eye center of Xiangya Hospital, Central South University, Changsha, Hunan Province, China
- Hunan Key Laboratory of Ophthalmology, Central South University, Chang sha, Hunan, China
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Yang J, Yang K, Meng X, Liu P, Fu Y, Wang Y. Silenced SNHG1 Inhibited Epithelial-Mesenchymal Transition and Inflammatory Response of ARPE-19 Cells Induced by High Glucose. J Inflamm Res 2021; 14:1563-1573. [PMID: 33907437 PMCID: PMC8068511 DOI: 10.2147/jir.s299010] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/24/2020] [Accepted: 03/09/2021] [Indexed: 12/27/2022] Open
Abstract
PURPOSE The lncRNA small nucleolar RNA host gene 1 (SNHG1) is a cerebral infarction-associated gene, its biological role and mechanism in diabetic retinopathy remain to be illuminated. The present study was designed to investigate the role of SNHG1 in high glucose induced human retinal pigment epithelial cells (ARPE-19). METHODS ARPE-19 cells were cultured and exposed to 60 mM high glucose for 48h, and 5.5mM glucose-exposed ARPE-19 cells were used as the control. The levels of the epithelial-mesenchymal transition (EMT) markers E-cadherin, ZO-1, vimentin and α-SMA were measured, and the Cell inflammatory response was evaluated by detecting IL-6 and IL-1β levels. Then, cell migration, proliferation and apoptosis were detected. The expression of the lncRNA SNHG1 in ARPE-19 cells was detected by quantitative real-time PCR. SNHG1 was knocked down by small interfering RNA (siRNA) transfection. The effects of SNHG1 inhibition on inflammation, EMT, migration, proliferation and apoptosis were observed. RESULTS The results showed that the expression of SNHG1 was significantly increased in ARPE-19 cells exposed to high glucose. Silencing SNHG1 reduced the expression of vimentin, α-SMA, and the expression of inflammatory chemokines IL-6 and IL-1β, inhibited migration and proliferation, elevated the expression of E-cadherin and ZO-1, and promoted apoptosis in ARPE-19 cells. CONCLUSION The lncRNA SNHG1 is involved in hyperglycemia-induced EMT and the inflammatory response of ARPE-19 cells and provides a new understanding of the pathogenesis of DR.
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Affiliation(s)
- Jing Yang
- Ophthalmology, Affiliated Hospital of Qingdao University, Qingdao, 266500, Shandong Province, People’s Republic of China
| | - Kun Yang
- Central Laboratory, Affiliated Hospital of Qingdao University, Qingdao, 266500, Shandong Province, People’s Republic of China
| | - Xuxia Meng
- Ophthalmology, Affiliated Hospital of Qingdao University, Qingdao, 266500, Shandong Province, People’s Republic of China
| | - Penghui Liu
- Ophthalmology, Affiliated Hospital of Qingdao University, Qingdao, 266500, Shandong Province, People’s Republic of China
| | - Yudong Fu
- Ophthalmology, Affiliated Hospital of Qingdao University, Qingdao, 266500, Shandong Province, People’s Republic of China
| | - Yibo Wang
- Ophthalmology, Affiliated Hospital of Qingdao University, Qingdao, 266500, Shandong Province, People’s Republic of China
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Astaxanthin mediated regulation of VEGF through HIF1α and XBP1 signaling pathway: An insight from ARPE-19 cell and streptozotocin mediated diabetic rat model. Exp Eye Res 2021; 206:108555. [PMID: 33789142 DOI: 10.1016/j.exer.2021.108555] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2020] [Revised: 03/08/2021] [Accepted: 03/23/2021] [Indexed: 11/21/2022]
Abstract
Breakdown of outer blood-retina barrier (BRB) has been associated with the pathogenesis of diabetic retinopathy (DR) and diabetic macular edema (DME). Vascular endothelial growth factor (VEGF) might play a detrimental role in the pathogenesis of DME, a major clinical manifestation of DR. In the present study, we investigated the inhibitory mechanism of astaxanthin on VEGF and its upstream signaling pathways under in vitro and in vivo conditions. Astaxanthin has been observed to downregulate VEGF expression under hyperglycemic (HG) and CoCl2 induced hypoxic conditions in ARPE-19 cells. There were compelling pieces of evidence for the involvement of transcription factors like HIF1α and XBP1 in the upregulation of VEGF under HG and hypoxic conditions. Thus, we investigated the role of astaxanthin in the expression and nuclear translocation of HIF1α and XBP1. The activation and translocation of HIF1α and XBP1 induced by HG or CoCl2 conditions were hindered by astaxanthin. Additionally, treatment with HIF1α siRNA and IRE1 inhibitor STF-083010 also inhibited the expression of VEGF induced by HG and CoCl2 conditions. These results indicated that the anti-VEGF property of astaxanthin might be associated with the downregulation of HIF1α and XBP1. Furthermore, astaxanthin mitigated the enhanced migration of retinal pigment epithelial (RPE) cells under DR conditions. As well, astaxanthin protected disorganization of zona occludin-1 (ZO-1) tight junction protein in RPE and reduced HG or hypoxic induced permeability of RPE cells. In streptozotocin-induced diabetic rat model, astaxanthin reduced the expression of HIF1α, XBP1, and VEGF as well as protected the abnormalities in the retinal layers induced by diabetes condition. Thus, astaxanthin may be used as a potential nutraceutical to prevent or treat retinal dysfunction in diabetic patients.
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Astragaloside IV attenuates high glucose-induced EMT by inhibiting the TGF-β/Smad pathway in renal proximal tubular epithelial cells. Biosci Rep 2021; 40:225214. [PMID: 32515466 PMCID: PMC7313447 DOI: 10.1042/bsr20190987] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/24/2019] [Revised: 05/28/2020] [Accepted: 06/03/2020] [Indexed: 12/13/2022] Open
Abstract
In the present study, we examined the molecular mechanism of astragaloside IV (AS-IV) in high glucose (HG)-induced epithelial-to-mesenchymal transition (EMT) in renal proximal tubular epithelial cells (PTCs). NRK-52E cell viability and apoptosis were determined by the cell counting kit-8 (CCK-8) assay and flow cytometric analysis, respectively. Expressions of E-cadherin, N-cadherin, vimentin, and occludin were measured by Western blot, and those of E-cadherin and N-cadherin were additionally measured by immunofluorescence analysis. Transforming growth factor-β1 (TGF-β1) and α-smooth muscle actin (α-SMA) expressions were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. The expressions of Smad2, Smad3, phosphorylated-Smad2 (p-Smad2), and p-Smad3 were measured using Western blot. We found that AS-IV could recover NRK-52E cell viability and inhibit HG-induced cell apoptosis. TGF-β1, α-SMA, Smad2, Smad3, p-Smad2, and p-Smad3 expressions were decreased in the AS-IV-treated groups compared with the HG group. Moreover, the expressions of E-cadherin and occludin were remarkably up-regulated and those of N-cadherin and vimentin were down-regulated in the AS-IV-treated groups compared with the HG group. Interestingly, the TGF-β1 activator SRI-011381 hydrochloride had an antagonistic effect to AS-IV on HG-induced EMT behavior. In conclusion, AS-IV attenuates HG-induced EMT by inhibiting the TGF-β/Smad pathway in renal PTCs.
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26
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Wang JX, Yang Y, Li WY. SIRT3 deficiency increases mitochondrial oxidative stress and promotes migration of retinal pigment epithelial cells. Exp Biol Med (Maywood) 2021; 246:877-887. [PMID: 33423553 DOI: 10.1177/1535370220976073] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
Retinal pigment epithelial cells are closely associated with the pathogenesis of diabetic retinopathy. The mechanism by which diabetes impacts retinal pigment epithelial cell function is of significant interest. Sirtuins are an important class of proteins that primarily possess nicotinamide adenine dinucleotide-dependent deacetylases activity and involved in various cellular physiological and pathological processes. Here, we aimed to examine the role of sirtuins in the induction of diabetes-associated retinal pigment epithelial cell dysfunction. High glucose and platelet-derived growth factor (PDGF) treatment induced epithelial-mesenchymal transition and the migration of retinal pigment epithelial cells, and decreased sirtuin-3 expression. Sirtuin-3 knockdown using siRNA increased epithelial-mesenchymal transition and migration of retinal pigment epithelial cells. In contrast, sirtuin-3 overexpression attenuated the effects caused by high glucose and PDGF on epithelial-mesenchymal transition and migration of retinal pigment epithelial cells, suggesting that sirtuin-3 deficiency contributed to retinal pigment epithelial cell dysfunction induced by high glucose and PDGF. Mechanistically, sirtuin-3 deficiency induced retinal pigment epithelial cell dysfunction by the overproduction of mitochondrial reactive oxygen species. These results suggest that sirtuin-3 deficiency mediates the migration of retinal pigment epithelial cells, at least partially by increasing mitochondrial oxidative stress, and shed light on the importance of sirtuin-3 and mitochondrial reactive oxygen species as potential targets in diabetic retinopathy therapy.
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Affiliation(s)
- Jing-Xian Wang
- Department of Medical Plastic and Cosmetic, Cangzhou Central Hospital, Cangzhou 061001, China
| | - Yuan Yang
- Department of Ophthalmology, Cangzhou Central Hospital, Cangzhou 061001, China
| | - Wen-Ying Li
- Department of Ophthalmology, Cangzhou Central Hospital, Cangzhou 061001, China
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27
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Positive Feedback Loop of SNAIL-IL-6 Mediates Myofibroblastic Differentiation Activity in Precancerous Oral Submucous Fibrosis. Cancers (Basel) 2020; 12:cancers12061611. [PMID: 32570756 PMCID: PMC7352888 DOI: 10.3390/cancers12061611] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/17/2020] [Revised: 06/11/2020] [Accepted: 06/11/2020] [Indexed: 12/17/2022] Open
Abstract
Oral submucosal fibrosis (OSF) is a premalignant disorder of the oral cavity, and areca nut chewing is known to be a major etiological factor that could induce epithelial to mesenchymal transition (EMT) and activate buccal mucosal fibroblasts (BMFs). However, this detailed mechanism is not fully understood. In this study, we showed that the upregulation of Snail in OSF samples and fibrotic BMFs (fBMFs) may result from constant irritation by arecoline, a major alkaloid of the areca nut. The elevation of Snail triggered myofibroblast transdifferentiation and was crucial to the persistent activation of fBMFs. Meanwhile, Snail increased the expression of numerous fibrosis factors (e.g., α-SMA and collagen I) as well as IL-6. Results from bioinformatics software and a luciferase-based reporter assay revealed that IL-6 was a direct target of Snail. Moreover, IL-6 in BMFs was found to further increase the expression of Snail and mediate Snail-induced myofibroblast activation. These findings suggested that there was a positive loop between Snail and IL-6 to regulate the areca nut-associated myofibroblast transdifferentiation, which implied that the blockage of Snail may serve as a favorable therapeutic strategy for OSF treatment.
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28
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EMT and EndMT: Emerging Roles in Age-Related Macular Degeneration. Int J Mol Sci 2020; 21:ijms21124271. [PMID: 32560057 PMCID: PMC7349630 DOI: 10.3390/ijms21124271] [Citation(s) in RCA: 162] [Impact Index Per Article: 32.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2020] [Revised: 06/12/2020] [Accepted: 06/14/2020] [Indexed: 02/06/2023] Open
Abstract
Epithelial–mesenchymal transition (EMT) and endothelial–mesenchymal transition (EndMT) are physiological processes required for normal embryogenesis. However, these processes can be hijacked in pathological conditions to facilitate tissue fibrosis and cancer metastasis. In the eye, EMT and EndMT play key roles in the pathogenesis of subretinal fibrosis, the end-stage of age-related macular degeneration (AMD) that leads to profound and permanent vision loss. Predominant in subretinal fibrotic lesions are matrix-producing mesenchymal cells believed to originate from the retinal pigment epithelium (RPE) and/or choroidal endothelial cells (CECs) through EMT and EndMT, respectively. Recent evidence suggests that EMT of RPE may also be implicated during the early stages of AMD. Transforming growth factor-beta (TGFβ) is a key cytokine orchestrating both EMT and EndMT. Investigations in the molecular mechanisms underpinning EMT and EndMT in AMD have implicated a myriad of contributing factors including signaling pathways, extracellular matrix remodelling, oxidative stress, inflammation, autophagy, metabolism and mitochondrial dysfunction. Questions arise as to differences in the mesenchymal cells derived from these two processes and their distinct mechanistic contributions to the pathogenesis of AMD. Detailed discussion on the AMD microenvironment highlights the synergistic interactions between RPE and CECs that may augment the EMT and EndMT processes in vivo. Understanding the differential regulatory networks of EMT and EndMT and their contributions to both the dry and wet forms of AMD can aid the development of therapeutic strategies targeting both RPE and CECs to potentially reverse the aberrant cellular transdifferentiation processes, regenerate the retina and thus restore vision.
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29
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Zhang NN, Kang JS, Liu SS, Gu SM, Song ZP, Li FX, Wang LF, Yao L, Li T, Li LL, Wang Y, Li XJ, Mao XM. Flavanomarein inhibits high glucose-stimulated epithelial-mesenchymal transition in HK-2 cells via targeting spleen tyrosine kinase. Sci Rep 2020; 10:439. [PMID: 31949205 PMCID: PMC6965095 DOI: 10.1038/s41598-019-57360-4] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2019] [Accepted: 12/27/2019] [Indexed: 01/14/2023] Open
Abstract
Flavanomarein (FM) is a major natural compound of Coreopsis tinctoria Nutt with protective effects against diabetic nephropathy (DN). In this study, we investigated the effects of FM on epithelial-mesenchymal transition (EMT) in high glucose (HG)-stimulated human proximal tubular epithelial cells (HK-2) and the underlying mechanisms, including both direct targets and downstream signal-related proteins. The influence of FM on EMT marker proteins was evaluated via western blot. Potential target proteins of FM were searched using Discovery Studio 2017 R2. Gene Ontology (GO) analysis was conducted to enrich the proteins within the protein-protein interaction (PPI) network for biological processes. Specific binding of FM to target proteins was examined via molecular dynamics and surface plasmon resonance analyses (SPR). FM promoted the proliferation of HK-2 cells stimulated with HG and inhibited EMT through the Syk/TGF-β1/Smad signaling pathway. Spleen tyrosine kinase (Syk) was predicted to be the most likely directly interacting protein with FM. Combined therapy with a Syk inhibitor and FM presents significant potential as an effective novel therapeutic strategy for DN.
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Affiliation(s)
- Nan-Nan Zhang
- State Key Laboratory of Pathogenesis, Prevention, Treatment of Central Asian High Incidence Diseases, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China.,Department of Pharmacology, Pharmacy College, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China
| | - Jin-Sen Kang
- Department of Pharmacology, Pharmacy College, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China
| | - Shuai-Shuai Liu
- Department of Pharmacology, School of Basic Medical Sciences, Peking University, Beijing, 100191, China
| | - Si-Meng Gu
- Department of Pharmacology, School of Basic Medical Sciences, Peking University, Beijing, 100191, China
| | - Zhi-Peng Song
- State Key Laboratory of Pathogenesis, Prevention, Treatment of Central Asian High Incidence Diseases, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China.,Department of Pharmacology, Pharmacy College, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China
| | - Feng-Xiang Li
- Department of Pharmacology, Pharmacy College, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China
| | - Li-Feng Wang
- Department of Physiology, Preclinical School, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China
| | - Lan Yao
- College of Traditional Chinese Medicine, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China
| | - Tian Li
- Department of Histology and Embryology, Preclinical College, XinJiang Medical University, Urumqi, Xinjiang, 830011, China
| | - Lin-Lin Li
- Department of Pharmacology, Pharmacy College, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China
| | - Ye Wang
- Department of Pharmacology, Pharmacy College, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China
| | - Xue-Jun Li
- Department of Pharmacology, School of Basic Medical Sciences, Peking University, Beijing, 100191, China.
| | - Xin-Min Mao
- State Key Laboratory of Pathogenesis, Prevention, Treatment of Central Asian High Incidence Diseases, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China. .,College of Traditional Chinese Medicine, Xinjiang Medical University, Urumqi, Xinjiang, 830011, China.
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Tenconi PE, Bermúdez V, Oresti GM, Giusto NM, Salvador GA, Mateos MV. High glucose-induced phospholipase D activity in retinal pigment epithelium cells: New insights into the molecular mechanisms of diabetic retinopathy. Exp Eye Res 2019; 184:243-257. [PMID: 31059692 DOI: 10.1016/j.exer.2019.04.028] [Citation(s) in RCA: 20] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2018] [Revised: 04/05/2019] [Accepted: 04/30/2019] [Indexed: 02/02/2023]
Abstract
Chronic hyperglycemia, oxidative stress and inflammation are key players in the pathogenesis of diabetic retinopathy (DR). In this work we study the role of phospholipase D (PLD) pathway in an in vitro model of high glucose (HG)-induced damage. To this end, we exposed human retinal pigment epithelium (RPE) cell lines (ARPE-19 and D407) to HG concentrations (16.5 or 33 mM) or to normal glucose concentration (NG, 5.5 mM) for 4, 24 or 72 h. Exposure to HG increased reactive oxygen species levels and caspase-3 cleavage and reduced cell viability after 72 h of incubation. In addition, short term HG exposure (4 h) induced the activation of early events, that involve PLD and ERK1/2 signaling, nuclear factor kappa B (NFκB) nuclear translocation and IκB phosphorylation. The increment in pro-inflammatory interleukins (IL-6 and IL-8) and cyclooxygenase-2 (COX-2) mRNA levels was observed after 24 h of HG exposure. The effect of selective pharmacological PLD1 (VU0359595) and PLD2 (VU0285655-1) inhibitors demonstrated that ERK1/2 and NFκB activation were downstream events of both PLD isoforms. The increment in IL-6 and COX-2 mRNA levels induced by HG was reduced to control levels in cells pre-incubated with both PLD inhibitors. Furthermore, the inhibition of PLD1, PLD2 and MEK/ERK pathway prevented the loss of cell viability and the activation of caspase-3 induced by HG. In conclusion, our findings demonstrate that PLD1 and PLD2 mediate the inflammatory response triggered by HG in RPE cells, pointing to their potential use as a therapeutic target for DR treatment.
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Affiliation(s)
- Paula E Tenconi
- Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), 8000, Bahía, Blanca, Argentina; Departamento de Biología, Bioquímica y Farmacia (DBByF), Universidad Nacional del Sur (UNS), 8000, Bahía, Blanca, Argentina
| | - Vicente Bermúdez
- Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), 8000, Bahía, Blanca, Argentina
| | - Gerardo M Oresti
- Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), 8000, Bahía, Blanca, Argentina; Departamento de Biología, Bioquímica y Farmacia (DBByF), Universidad Nacional del Sur (UNS), 8000, Bahía, Blanca, Argentina
| | - Norma M Giusto
- Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), 8000, Bahía, Blanca, Argentina; Departamento de Biología, Bioquímica y Farmacia (DBByF), Universidad Nacional del Sur (UNS), 8000, Bahía, Blanca, Argentina
| | - Gabriela A Salvador
- Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), 8000, Bahía, Blanca, Argentina; Departamento de Biología, Bioquímica y Farmacia (DBByF), Universidad Nacional del Sur (UNS), 8000, Bahía, Blanca, Argentina
| | - Melina V Mateos
- Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), 8000, Bahía, Blanca, Argentina; Departamento de Biología, Bioquímica y Farmacia (DBByF), Universidad Nacional del Sur (UNS), 8000, Bahía, Blanca, Argentina.
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Ji T, Su SL, Zhu Y, Guo JM, Qian DW, Tang YP, Duan JA. The mechanism of mulberry leaves against renal tubular interstitial fibrosis through ERK1/2 signaling pathway was predicted by network pharmacology and validated in human tubular epithelial cells. Phytother Res 2019; 33:2044-2055. [PMID: 31209937 DOI: 10.1002/ptr.6390] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2018] [Revised: 02/18/2019] [Accepted: 04/23/2019] [Indexed: 12/24/2022]
Abstract
Mulberry leaf was reported that it has antidiabetic activity, although the mechanisms underlying the function have not been fully elucidated. In the present study, the results of network pharmacology suggested that mulberry leaves could regulate key biological process in development of diabetes, and the process implicates multiple signaling pathways, such as JAK-STAT, MAPK, VEGF, PPAR, and Wnt. Then, the research in vitro indicated that mulberry leaves remarkably ameliorated high glucose-induced epithelial to mesenchymal transition, which was characterized with significant reduction of intracellular reactive oxygen species (ROS) levels as well as downregulation of NADPH oxidase subunits NOX1, NOX2, and NOX4, and it was found to be connected with the ERK1/2 signaling pathway in human tubular epithelial cells (HK-2). Moreover, the results of bioinformatics and the dual luciferase report showed that ZEB1 might be a target gene of miR-302a; decreased miR-302a and increased ZEB1 expressions could significantly promote epithelial to mesenchymal transition. However, mulberry leaves could reverse these modulations. Our results demonstrated that network pharmacology could provide a guidance role for traditional Chinese medicine research, and mulberry leaves could be of benefit in preventing high glucose-induced EMT in HK-2 cells, which proved that it was related to the upregulation of miR-302a by targeting ZEB1 and the inhibition of NADPH oxidase/ROS/ERK1/2 pathway.
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Affiliation(s)
- Tao Ji
- Jiangsu Collaborative Innovation Center of Chinese Medicinal Resources Industrialization, National and Local Collaborative Engineering Center of Chinese Medicinal Resources Industrialization and Formulae Innovative Medicine, and Jiangsu Key Laboratory for High Technology Research of TCM Formulae, Nanjing University of Chinese Medicine, Nanjing, China.,Institute of traditional Chinese medicine, Zhejiang pharmaceutical college, Ningbo, 310053, China
| | - Shu-Lan Su
- Jiangsu Collaborative Innovation Center of Chinese Medicinal Resources Industrialization, National and Local Collaborative Engineering Center of Chinese Medicinal Resources Industrialization and Formulae Innovative Medicine, and Jiangsu Key Laboratory for High Technology Research of TCM Formulae, Nanjing University of Chinese Medicine, Nanjing, China
| | - Yue Zhu
- Jiangsu Collaborative Innovation Center of Chinese Medicinal Resources Industrialization, National and Local Collaborative Engineering Center of Chinese Medicinal Resources Industrialization and Formulae Innovative Medicine, and Jiangsu Key Laboratory for High Technology Research of TCM Formulae, Nanjing University of Chinese Medicine, Nanjing, China
| | - Jian-Ming Guo
- Jiangsu Collaborative Innovation Center of Chinese Medicinal Resources Industrialization, National and Local Collaborative Engineering Center of Chinese Medicinal Resources Industrialization and Formulae Innovative Medicine, and Jiangsu Key Laboratory for High Technology Research of TCM Formulae, Nanjing University of Chinese Medicine, Nanjing, China
| | - Da-Wei Qian
- Jiangsu Collaborative Innovation Center of Chinese Medicinal Resources Industrialization, National and Local Collaborative Engineering Center of Chinese Medicinal Resources Industrialization and Formulae Innovative Medicine, and Jiangsu Key Laboratory for High Technology Research of TCM Formulae, Nanjing University of Chinese Medicine, Nanjing, China
| | - Yu-Ping Tang
- Jiangsu Collaborative Innovation Center of Chinese Medicinal Resources Industrialization, National and Local Collaborative Engineering Center of Chinese Medicinal Resources Industrialization and Formulae Innovative Medicine, and Jiangsu Key Laboratory for High Technology Research of TCM Formulae, Nanjing University of Chinese Medicine, Nanjing, China
| | - Jin-Ao Duan
- Jiangsu Collaborative Innovation Center of Chinese Medicinal Resources Industrialization, National and Local Collaborative Engineering Center of Chinese Medicinal Resources Industrialization and Formulae Innovative Medicine, and Jiangsu Key Laboratory for High Technology Research of TCM Formulae, Nanjing University of Chinese Medicine, Nanjing, China
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Chen X, Jiang C, Yang D, Sun R, Wang M, Sun H, Xu M, Zhou L, Chen M, Xie P, Yan B, Liu Q, Zhao C. CRB2 mutation causes autosomal recessive retinitis pigmentosa. Exp Eye Res 2018; 180:164-173. [PMID: 30593785 DOI: 10.1016/j.exer.2018.12.018] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2018] [Revised: 12/21/2018] [Accepted: 12/21/2018] [Indexed: 01/29/2023]
Abstract
Retinitis pigmentosa (RP), the most common form of inherited retinal dystrophies, exhibits significant genetic heterogeneity. The crumbs homolog 2 (CRB2) protein, together with CRB1 and CRB3, belongs to the Crumbs family. Given that CRB1 mutations account for 4% of RP cases, the role of CRB2 mutations in RP etiology has long been hypothesized but never confirmed. Herein, we report the identification of CRB2 as a novel RP causative gene in a Chinese consanguineous family and have analyzed its pathogenic effects. Comprehensive ophthalmic and systemic evaluations confirmed the clinical diagnosis of the two patients in this family as RP. WES revealed a homozygous missense mutation, CRB2 p.R1249G, to segregate the RP phenotype, which was highly conserved among multiple species. In vitro cellular study revealed that this mutation not only interrupted the stability of the transcribed CRB2 mRNA and the encoded CRB2 protein, but also interfered with the wild type CRB2 mRNA/protein and decreased their expression. This mutation was also shown to trigger epithelial-mesenchymal transition (EMT) in retinal pigment epithelium (RPE) cells, thus impairing regular RPE phagocytosis and induce RPE degeneration and apoptosis. Thus, we conclude that CRB2 p.R1249G mutation causes RP via accelerating EMT, dysfunction and loss of RPE cells, and establish CRB2 as a novel Crumbs family member associated with non-syndromic RP. We provide important hints for understanding of CRB2 defects and retinopathy, and for the involvement of EMT of RPE cells in RP pathogenesis.
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Affiliation(s)
- Xue Chen
- Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China; Department of Ophthalmology and Vision Science, Eye & ENT Hospital, Shanghai Medical College, Fudan University, Shanghai, 200023, China; Key Laboratory of Myopia of State Health Ministry (Fudan University) and Shanghai Key Laboratory of Visual Impairment and Restoration, Shanghai, 200023, China
| | - Chao Jiang
- Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China
| | - Daidi Yang
- Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China
| | - Ruxu Sun
- Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China
| | - Min Wang
- Department of Ophthalmology and Vision Science, Eye & ENT Hospital, Shanghai Medical College, Fudan University, Shanghai, 200023, China; Key Laboratory of Myopia of State Health Ministry (Fudan University) and Shanghai Key Laboratory of Visual Impairment and Restoration, Shanghai, 200023, China
| | - Hong Sun
- Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China
| | - Min Xu
- Department of Ophthalmology, Northern Jiangsu People's Hospital, Yangzhou, 211406, China
| | - Luyin Zhou
- Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China
| | - Mingkang Chen
- Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China
| | - Ping Xie
- Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China
| | - Biao Yan
- Department of Ophthalmology and Vision Science, Eye & ENT Hospital, Shanghai Medical College, Fudan University, Shanghai, 200023, China; Key Laboratory of Myopia of State Health Ministry (Fudan University) and Shanghai Key Laboratory of Visual Impairment and Restoration, Shanghai, 200023, China
| | - Qinghuai Liu
- Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China.
| | - Chen Zhao
- Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China; Department of Ophthalmology and Vision Science, Eye & ENT Hospital, Shanghai Medical College, Fudan University, Shanghai, 200023, China; Key Laboratory of Myopia of State Health Ministry (Fudan University) and Shanghai Key Laboratory of Visual Impairment and Restoration, Shanghai, 200023, China.
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You ZP, Chen SS, Yang ZY, Li SR, Xiong F, Liu T, Fu SH. GEP100/ARF6 regulates VEGFR2 signaling to facilitate high-glucose-induced epithelial-mesenchymal transition and cell permeability in retinal pigment epithelial cells. Am J Physiol Cell Physiol 2018; 316:C782-C791. [PMID: 30540496 DOI: 10.1152/ajpcell.00312.2018] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Cell permeability and epithelial-mesenchymal transition (EMT) were found to be enhanced in diabetic retinopathy, and the aim of this study was to investigate the underlying mechanism. ARPE-19 cell line or primary retinal pigment epithelial (RPE) cells were cultured under high or normal glucose conditions. Specific shRNAs were employed to knock down ADP-ribosylation factor 6 (ARF6), GEP100, or VEGF receptor 2 (VEGFR2) in ARPE-19 or primary RPE cells. Cell migration ability was measured using Transwell assay. Western blotting was used to measure indicated protein levels. RPE cells treated with high glucose showed increased cell migration, paracellular permeability, EMT, and expression of VEGF. Knockdown of VEGFR2 inhibited the high-glucose-induced effects on RPE cells via inactivation of ARF6 and MAPK pathways. Knockdown ARF6 or GEP100 led to inhibition of high-glucose-induced effects via inactivation of VEGFR2 pathway. Knockdown of ARF6, but not GEP100, decreased high-glucose-induced internalization of VEGFR2. High-glucose enhances EMT and cell permeability of RPE cells through activation of VEGFR2 and ARF6/GEP100 pathways, which form a positive feedback loop to maximize the activation of VEGF/VEGFR2 signaling.
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Affiliation(s)
- Zhi-Peng You
- Department of Ophthalmology, The Second Affiliated Hospital of Nanchang University , Nanchang , People's Republic of China
| | - Shan-Shan Chen
- Department of Ophthalmology, The Second Affiliated Hospital of Nanchang University , Nanchang , People's Republic of China
| | - Zhong-Yi Yang
- Department of Ophthalmology, The Second Affiliated Hospital of Nanchang University , Nanchang , People's Republic of China
| | - Shu-Rong Li
- Department of Ophthalmology, The Second Affiliated Hospital of Nanchang University , Nanchang , People's Republic of China
| | - Fan Xiong
- Department of Ophthalmology, The Second Affiliated Hospital of Nanchang University , Nanchang , People's Republic of China
| | - Ting Liu
- Department of Ophthalmology, The Second Affiliated Hospital of Nanchang University , Nanchang , People's Republic of China
| | - Shu-Hua Fu
- Department of Ophthalmology, The Second Affiliated Hospital of Nanchang University , Nanchang , People's Republic of China
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