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Lyu G, Dai L, Deng X, Liu X, Guo Y, Zhang Y, Wang X, Huang Y, Wu S, Guo J, Liu Y. Integrative Analysis of Cuproptosis-Related Mitochondrial Depolarisation Genes for Prognostic Prediction in Non-Small Cell Lung Cancer. J Cell Mol Med 2025; 29:e70438. [PMID: 40008552 PMCID: PMC11862892 DOI: 10.1111/jcmm.70438] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2024] [Revised: 01/27/2025] [Accepted: 02/12/2025] [Indexed: 02/27/2025] Open
Abstract
In this research, we conducted an in-depth analysis of differentially expressed genes associated with mitochondrial depolarisation in non-small cell lung cancer (NSCLC) using single-cell sequencing. By combining our findings with cuproptosis-related genes, we identified 10 significant risk genes: DCN, PTHLH, CRYAB, HMGCS1, DSG3, ZFP36L2, SCAND1, NUDT4, NDUFA4L2 and RPL36A, using univariate Cox regression analysis and machine learning methods. These genes form the core of our prognosis risk prediction model, which demonstrated high specificity and accuracy in predicting patient outcomes, as evidenced by ROC curve analysis. Kaplan-Meier curves further confirmed that patients in the low-risk group had significantly better survival rates compared to those in the high-risk group. Our models also provided valuable insights into the tumour microenvironment, immunotherapy sensitivity and chemotherapy response. To facilitate the quantification of the probability of patient survival, we incorporated clinical data into a nomogram. We comprehensively analysed the mutation status and expression patterns of the 10 risk genes using bulk transcriptomic, single-cell and spatial transcriptomic datasets. Drug target predictions highlighted DSG3, PTHLH, ZFP36L2, DCN and NDUFA4L2 as promising therapeutic targets. Notably, RPL36A emerged as a potential tumour marker for NSCLC, with its expression validated in lung cancer cell lines through qPCR. This study has established a predictive models based on mitochondrial depolarisation genes associated with cuproptosis, aiding clinicians in forecasting overall survival and guiding personalised treatment strategies. The identification of novel tumour markers has paved the way for targeted therapies, and therapeutic targets are critical for advancing the treatment of NSCLC.
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Affiliation(s)
- Guoqing Lyu
- Department of HematologyThe First Affiliated Hospital of Xinxiang Medical UniversityWeihuiHenan ProvinceChina
- Life Science CenterThe First Affiliated Hospital of Xinxiang Medical UniversityWeihuiHenan ProvinceChina
- Key Laboratory for Leukemia Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
- Key Laboratory for Lymphoma Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
| | - Lihua Dai
- Prenatal Diagnosis CenterShanghai General Hospital Affiliated to Shanghai JiaoTong University School of MedicineShanghaiChina
| | - Xin Deng
- Beijing University of Chinese MedicineBeijingChina
| | - Xiankai Liu
- Department of HematologyThe First Affiliated Hospital of Xinxiang Medical UniversityWeihuiHenan ProvinceChina
- Key Laboratory for Leukemia Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
- Key Laboratory for Lymphoma Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
| | - Yan Guo
- Department of HematologyThe First Affiliated Hospital of Xinxiang Medical UniversityWeihuiHenan ProvinceChina
- Key Laboratory for Leukemia Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
- Key Laboratory for Lymphoma Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
| | - Yuan Zhang
- Department of HematologyThe First Affiliated Hospital of Xinxiang Medical UniversityWeihuiHenan ProvinceChina
- Key Laboratory for Leukemia Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
- Key Laboratory for Lymphoma Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
| | - Xiufeng Wang
- Department of HematologyThe First Affiliated Hospital of Xinxiang Medical UniversityWeihuiHenan ProvinceChina
- Key Laboratory for Leukemia Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
- Key Laboratory for Lymphoma Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
| | - Yan Huang
- Department of HematologyThe First Affiliated Hospital of Xinxiang Medical UniversityWeihuiHenan ProvinceChina
- Key Laboratory for Leukemia Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
- Key Laboratory for Lymphoma Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
| | - Sun Wu
- Department of HematologyThe First Affiliated Hospital of Xinxiang Medical UniversityWeihuiHenan ProvinceChina
- Key Laboratory for Leukemia Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
- Key Laboratory for Lymphoma Molecular Diagnosis and Treatment in Xinxiang CityWeihuiHenan ProvinceChina
| | | | - Yanting Liu
- Department of HematologyThe First Affiliated Hospital of Xinxiang Medical UniversityWeihuiHenan ProvinceChina
- Department of OncologyThe First Affiliated Hospital of Xinxiang Medical UniversityWeihuiHenan ProvinceChina
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Sun B, Jiang J, Zhu X, Yang D, Cui Z, Zhang Y, Zhang M, Qian Y, Liu R, Yang W. Protective effects of gastrodin pretreatment on mouse hepatic ischemia-reperfusion occurring through antioxidant and anti-apoptotic mechanisms. Exp Ther Med 2021; 21:471. [PMID: 33767766 DOI: 10.3892/etm.2021.9902] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/26/2020] [Accepted: 02/08/2021] [Indexed: 12/21/2022] Open
Abstract
Hepatic ischemia-reperfusion injury (HIRI) often occurs following surgical procedures such as liver resection and transplantation. However, despite its clinical prominence, to the best of our knowledge, there remain no effective strategies to treat HIRI. Therefore, the aim of present study was to identify therapeutic agents that can exert beneficial effects against HIRI. The present study found that following hepatic IR modeling in mice, gastrodin (Gas) pretreatment improved the IR outcomes in terms of the serum biochemical indexes (alanine transaminase and aspartate transaminase), tissue biochemical indexes (superoxide dismutase, malondialdehyde and reduced glutathione content) and tissue pathology (H&E staining). In addition, compared with those in the IR + vehicle group, the IR + Gas group showed upregulated expression levels of nuclear erythroid 2-related factor 2, heme oxygenase 1 and Bcl-2 as detected by western blotting and reverse transcription-quantitative PCR. The mRNA and protein expression levels of Bax and caspase-3 were downregulated in the IR + Gas group compared with the IR + vehicle group. Concurrently, no significant differences were observed in the parameters between the Sham + vehicle and the Sham + Gas groups, indicating that Gas pretreatment may not cause liver damage. In conclusion, the findings of the present study revealed that Gas pretreatment exerted a protective effect in HIRI through both antioxidant and anti-apoptotic mechanisms.
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Affiliation(s)
- Bo Sun
- Department of Gastroenterology and Hepatology, Institute of Digestive Disease, Tongji Hospital, Tongji University School of Medicine, Shanghai 200065, P.R. China
| | - Jie Jiang
- Department of Gastroenterology and Hepatology, Institute of Digestive Disease, Tongji Hospital, Tongji University School of Medicine, Shanghai 200065, P.R. China
| | - Xinyan Zhu
- Department of Gastroenterology, Shanghai East Hospital, Tongji University School of Medicine, Shanghai 200120, P.R. China
| | - Dan Yang
- Department of Gastroenterology and Hepatology, Institute of Digestive Disease, Tongji Hospital, Tongji University School of Medicine, Shanghai 200065, P.R. China
| | - Zhenyu Cui
- Department of Gastroenterology and Hepatology, Institute of Digestive Disease, Tongji Hospital, Tongji University School of Medicine, Shanghai 200065, P.R. China
| | - Yu Zhang
- Department of Gastroenterology and Hepatology, Institute of Digestive Disease, Tongji Hospital, Tongji University School of Medicine, Shanghai 200065, P.R. China
| | - Minbo Zhang
- Department of Gastroenterology and Hepatology, Institute of Digestive Disease, Tongji Hospital, Tongji University School of Medicine, Shanghai 200065, P.R. China
| | - Yiting Qian
- Department of Gastroenterology and Hepatology, Institute of Digestive Disease, Tongji Hospital, Tongji University School of Medicine, Shanghai 200065, P.R. China
| | - Ruilin Liu
- Department of Pulmonary and Critical Care Medicine, Tongji Hospital, Tongji University School of Medicine, Shanghai 200065, P.R. China
| | - Wenzhuo Yang
- Department of Gastroenterology and Hepatology, Institute of Digestive Disease, Tongji Hospital, Tongji University School of Medicine, Shanghai 200065, P.R. China
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Grynberg K, Ozols E, Mulley WR, Davis RJ, Flavell RA, Nikolic-Paterson DJ, Ma FY. JUN Amino-Terminal Kinase 1 Signaling in the Proximal Tubule Causes Cell Death and Acute Renal Failure in Rat and Mouse Models of Renal Ischemia/Reperfusion Injury. THE AMERICAN JOURNAL OF PATHOLOGY 2021; 191:817-828. [PMID: 33607044 DOI: 10.1016/j.ajpath.2021.02.004] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/10/2020] [Revised: 02/02/2021] [Accepted: 02/04/2021] [Indexed: 10/22/2022]
Abstract
Activation of the JUN amino-terminal kinase (JNK) pathway is prominent in most forms of acute and progressive tubulointerstitial damage, including acute renal ischemia/reperfusion injury (IRI). Two forms of JNK, JNK1 and JNK2, are expressed in the kidney. Systemic administration of pan-JNK inhibitors suppresses renal IRI; however, the contribution of JNK1 versus JNK2, and the specific role of JNK activation in the proximal tubule in IRI, remains unknown. These questions were addressed in rat and mouse models of acute bilateral renal IRI. Administration of the JNK inhibitor, CC-930, substantially reduced the severity of renal failure, tubular damage, and inflammation at 24 hours in a rat IRI model. Additionally, Jnk1-/- mice, but not Jnk2-/- mice, were shown to be significantly protected against acute renal failure, tubular damage, and inflammation in the IRI model. Furthermore, mice with conditional Jnk1 deletion in the proximal tubule also showed considerable protection from IRI-induced renal failure, tubular damage, and inflammation. Finally, primary cultures of Jnk1-/-, but not Jnk2-/-, tubular epithelial cells were protected from oxidant-induced cell death, in association with preventing phosphorylation of proteins (receptor interacting serine/threonine kinase 3 and mixed lineage kinase domain-like pseudokinase) in the necroptosis pathway. In conclusion, JNK1, but not JNK2, plays a specific role in IRI-induced cell death in the proximal tubule, leading to acute renal failure.
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Affiliation(s)
- Keren Grynberg
- Department of Nephrology, Monash Medical Centre, Clayton, Victoria, Australia; Centre for Inflammatory Diseases, Monash University, Clayton, Victoria, Australia
| | - Elyce Ozols
- Department of Nephrology, Monash Medical Centre, Clayton, Victoria, Australia; Centre for Inflammatory Diseases, Monash University, Clayton, Victoria, Australia
| | - William R Mulley
- Department of Nephrology, Monash Medical Centre, Clayton, Victoria, Australia; Centre for Inflammatory Diseases, Monash University, Clayton, Victoria, Australia
| | - Roger J Davis
- Howard Hughes Medical Institute and Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts
| | - Richard A Flavell
- Department of Immunobiology, Yale University School of Medicine, New Haven, Connecticut
| | - David J Nikolic-Paterson
- Department of Nephrology, Monash Medical Centre, Clayton, Victoria, Australia; Centre for Inflammatory Diseases, Monash University, Clayton, Victoria, Australia.
| | - Frank Y Ma
- Department of Nephrology, Monash Medical Centre, Clayton, Victoria, Australia; Centre for Inflammatory Diseases, Monash University, Clayton, Victoria, Australia
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Zhang Y, Pan Q, Cheng Y, Liu Y. Effects of SP600125 and hypothermic machine perfusion on livers donated after cardiac death in a pig allograft transplantation model. Eur J Med Res 2021; 26:15. [PMID: 33546770 PMCID: PMC7863371 DOI: 10.1186/s40001-020-00472-9] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2019] [Accepted: 12/14/2020] [Indexed: 12/12/2022] Open
Abstract
BACKGROUND Hypothermic machine perfusion (HMP) improves the quality of donor livers for transplantation, both in animal models and in clinical practice. Treatment with SP600125, an inhibitor of c-Jun N-terminal kinase (JNK), can suppress the JNK signaling pathway to alleviate donor liver ischemia-reperfusion injury (IRI). We performed the present study with the objective of exploring the protective effects exerted by a combination of HMP and SP600125 on liver xenograft viability for donation after cardiac death (DCD) in a porcine model. METHODS 54 adult BAMA mini-pigs were randomly assigned to 5 groups, including sham, cold storage for 4 h (CS 4 h), CS 4 h + SP600125, CS 2 h + HMP 2 h, and CS 2 h + HMP 2 h + SP600125 groups. Donor livers in the CS 4 h and CS 4 h + SP600125 groups were conventionally cold preserved for 4 h, whereas donor livers in the CS 2 h + HMP 2 h and CS 2 h + HMP 2 h + SP600125 groups were cold preserved for 2 h and then treated with HMP for 2 h. The preservation and perfusion solutions contained SP600125 (20 µM). Follow-up was conducted for 5 days after liver transplantation to compare the surgical outcomes by means of serological examination, pathological results, and survival rate. RESULTS The most satisfactory outcome after liver transplantation was observed in the CS 2 h + HMP 2 h + SP600125 group, which presented with minimal damage of donor livers during 5 days' follow-up. Additionally, serological examination, pathological results, and survival rate concurred in showing better results in the CS 2 h + HMP 2 h ± SP600125 group than in the CS 4 h ± SP600125 group. CONCLUSION HMP in combination with SP600125 has hepatoprotective properties and improves the quality and viability of porcine livers collected after DCD, thus improving prognosis after liver transplantation.
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Affiliation(s)
- Yijie Zhang
- Department of Organ Transplantation and Hepatobiliary, The First Affiliated Hospital of China Medical University, No. 155, Nanjing Street, Shenyang, 110001, Liaoning, People's Republic of China.,The Key Laboratory of Organ Transplantation of Liaoning Province, The First Affiliated Hospital of China Medical University, No. 155, Nanjing Street, Shenyang, 110001, Liaoning, People's Republic of China
| | - Qi Pan
- Department of Organ Transplantation and Hepatobiliary, The First Affiliated Hospital of China Medical University, No. 155, Nanjing Street, Shenyang, 110001, Liaoning, People's Republic of China.,The Key Laboratory of Organ Transplantation of Liaoning Province, The First Affiliated Hospital of China Medical University, No. 155, Nanjing Street, Shenyang, 110001, Liaoning, People's Republic of China
| | - Ying Cheng
- Department of Organ Transplantation and Hepatobiliary, The First Affiliated Hospital of China Medical University, No. 155, Nanjing Street, Shenyang, 110001, Liaoning, People's Republic of China.,The Key Laboratory of Organ Transplantation of Liaoning Province, The First Affiliated Hospital of China Medical University, No. 155, Nanjing Street, Shenyang, 110001, Liaoning, People's Republic of China
| | - Yongfeng Liu
- Department of Organ Transplantation and Hepatobiliary, The First Affiliated Hospital of China Medical University, No. 155, Nanjing Street, Shenyang, 110001, Liaoning, People's Republic of China. .,The Key Laboratory of Organ Transplantation of Liaoning Province, The First Affiliated Hospital of China Medical University, No. 155, Nanjing Street, Shenyang, 110001, Liaoning, People's Republic of China.
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Thomson AW, Vionnet J, Sanchez-Fueyo A. Understanding, predicting and achieving liver transplant tolerance: from bench to bedside. Nat Rev Gastroenterol Hepatol 2020; 17:719-739. [PMID: 32759983 DOI: 10.1038/s41575-020-0334-4] [Citation(s) in RCA: 67] [Impact Index Per Article: 13.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 06/11/2020] [Indexed: 02/07/2023]
Abstract
In the past 40 years, liver transplantation has evolved from a high-risk procedure to one that offers high success rates for reversal of liver dysfunction and excellent patient and graft survival. The liver is the most tolerogenic of transplanted organs; indeed, immunosuppressive therapy can be completely withdrawn without rejection of the graft in carefully selected, stable long-term liver recipients. However, in other recipients, chronic allograft injury, late graft failure and the adverse effects of anti-rejection therapy remain important obstacles to improved success. The liver has a unique composition of parenchymal and immune cells that regulate innate and adaptive immunity and that can promote antigen-specific tolerance. Although the mechanisms underlying liver transplant tolerance are not well understood, important insights have been gained into how the local microenvironment, hepatic immune cells and specific molecular pathways can promote donor-specific tolerance. These insights provide a basis for the identification of potential clinical biomarkers that might correlate with tolerance or rejection and for the development of novel therapeutic targets. Innovative approaches aimed at promoting immunosuppressive drug minimization or withdrawal include the adoptive transfer of donor-derived or recipient-derived regulatory immune cells to promote liver transplant tolerance. In this Review, we summarize and discuss these developments and their implications for liver transplantation.
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Affiliation(s)
- Angus W Thomson
- Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA. .,Department of Immunology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA.
| | - Julien Vionnet
- Institute of Liver Studies, Medical Research Council (MRC) Centre for Transplantation, School of Immunology and Infectious Diseases, King's College London University, King's College Hospital, London, UK.,Transplantation Center, University Hospital of Lausanne, Lausanne, Switzerland.,Service of Gastroenterology and Hepatology, University Hospital of Lausanne, Lausanne, Switzerland
| | - Alberto Sanchez-Fueyo
- Institute of Liver Studies, Medical Research Council (MRC) Centre for Transplantation, School of Immunology and Infectious Diseases, King's College London University, King's College Hospital, London, UK
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Yu Y, Li J, Bi Z, Wu C, Yang S, Jiang Q, Deng R, Fu Q, Liu L, Wang C. Cuff Anastomosis of Both Renal Artery and Vein to Minimize Thrombosis: A Novel Method of Kidney Transplantation in Mice. J INVEST SURG 2020; 35:56-60. [PMID: 32965132 DOI: 10.1080/08941939.2020.1821264] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
Abstract
OBJECTIVES Anastomosis of renal artery and renal vein in mouse models of kidney transplantation is technically challenging. Conventional technique using suture may result in vascular thrombosis. We developed a simple cuff method to anastomose both renal artery and vein. MATERIALS AND METHODS Briefly, the left renal artery was occluded at the junction with abdominal aorta using a small vessel clip, transected at the renal hilum, irrigated with heparinized saline, and passed through the lumen of a seamless tubing made of polyimide. The loose end of the artery was everted over the cuff and secured using an 8-0 silk suture. The cuffed artery was inserted into the donor renal artery and secured with an 8-0 suture. Anastomosis of the renal vein was performed similarly. Isograft transplantation was conducted using BALB/c mice as donor and recipient mice (n = 20). The total operative time was 77 ± 3 min, and the cold ischemic time of the graft kidney was minimized to 20 min. One animal was excluded due to anatomic variant vessels and another one died at three day after surgery without thrombosis. RESULTS Serum creatinine increased insignificantly after transplantation and remained stable over 12 weeks posttransplant. Five recipient mice were sacrificed for histologic examination at 12 weeks after transplantation. No vascular thrombosis was observed at the site of anastomosis. The isografts showed no evidence of acute and chronic lesions such as extinctive ischemic sclerosis and interstitial fibrosis. CONCLUSION In summary, cuff anastomosis can be used to eliminate thrombosis formation in the mouse model of kidney transplantation.
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Affiliation(s)
- Yi Yu
- Organ Transplant Center, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.,Organ Transplant Center, Zhongshan People's Hospital, Zhongshan, China
| | - Jun Li
- Organ Transplant Center, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China
| | - Zirong Bi
- Organ Transplant Center, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China
| | - Chenglin Wu
- Organ Transplant Center, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China
| | - Shicong Yang
- Department of Pathology, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China
| | - Qifeng Jiang
- Department of Pathology, Guangzhou Huayin Medical Laboratory Center, Guangzhou, China
| | - Ronghai Deng
- Organ Transplant Center, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China
| | - Qian Fu
- Organ Transplant Center, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China
| | - Longshan Liu
- Organ Transplant Center, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China
| | - Changxi Wang
- Organ Transplant Center, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.,Guangdong Provincial Key Laboratory of Organ Donation and Transplant Immunology, Guangzhou, China.,Guangdong Provincial International Cooperation Base of Science and Technology (Organ Transplantation), Guangzhou, China
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Tan J, Gao W, Yang W, Zeng X, Wang L, Cui X. Isoform-specific functions of c-Jun N-terminal kinase 1 and 2 in lung ischemia-reperfusion injury through the c-Jun/activator protein-1 pathway. J Thorac Cardiovasc Surg 2020; 162:e143-e156. [PMID: 32414595 DOI: 10.1016/j.jtcvs.2020.03.083] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/30/2019] [Revised: 03/23/2020] [Accepted: 03/25/2020] [Indexed: 02/07/2023]
Abstract
BACKGROUND c-Jun N-terminal kinase 1 (JNK1) and JNK2 regulate distinct pathological processes in lung diseases. Here we discriminated the respective roles of these kinases in lung transplantation-induced ischemia-reperfusion injury (IRI). METHODS Rat pulmonary microvascular endothelial cells were transfected with JNK1 small-interfering RNA (siRNA) and JNK2 siRNA and then subjected to in vitro IRI. For the isoform confirmed to aggravate IRI, the delivery of short-hairpin RNA (shRNA) plasmid was performed by intratracheal administration 48 hours before transplantation into donor rats. After a 3-hour reperfusion, the samples were collected. RESULTS JNK1 siRNA decreased but JNK2 siRNA increased JNK phosphorylation and activity, phosphorylated and total c-Jun, and activator protein-1 activity. Although JNK1 siRNA decreased apoptosis and the levels of malondialdehyde, interleukin (IL)-1, IL-6, and tumor necrosis factor (TNF-α), it increased the levels of superoxide dismutase, S-phase percentage, and cyclin D1; JNK2 siRNA had a converse effect. JNK1 siRNA decreased the level of lactate dehydrogenase and increased the levels of VE-cadherin, nitric oxide, phosphorylated nitric oxide synthase, and cell viability; JNK2 si RNA had a converse effect. Compared with the control group, the JNK1 shRNA group exhibited a higher lung oxygenation index and lower lung apoptosis index, injury score, wet weight:dry weight ratio, and levels of IL-1, IL-6, and TNF-α. CONCLUSIONS JNK1 aggravated, but JNK2 alleviated, IRI through differential regulation of the JNK1 pathway in in vitro ischemia-reperfusion. JNK1 silence attenuated lung graft dysfunction by inhibiting inflammation and apoptosis. These findings provide a theoretical basis for devising therapeutic strategies against IRI after lung transplantation.
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Affiliation(s)
- Jing Tan
- Department of Anesthesiology, Hei Long Jiang Province Key Lab of Research on Anesthesiology and Critical Care Medicine, Second Affiliated Hospital, Harbin Medical University, Harbin, China
| | - Wei Gao
- Department of Anesthesiology, Hei Long Jiang Province Key Lab of Research on Anesthesiology and Critical Care Medicine, Second Affiliated Hospital, Harbin Medical University, Harbin, China
| | - Wanchao Yang
- Department of Anesthesiology, Hei Long Jiang Province Key Lab of Research on Anesthesiology and Critical Care Medicine, Second Affiliated Hospital, Harbin Medical University, Harbin, China
| | - Xianzhang Zeng
- Department of Anesthesiology, Hei Long Jiang Province Key Lab of Research on Anesthesiology and Critical Care Medicine, Second Affiliated Hospital, Harbin Medical University, Harbin, China
| | - Linlin Wang
- Department of Anesthesiology, Hei Long Jiang Province Key Lab of Research on Anesthesiology and Critical Care Medicine, Second Affiliated Hospital, Harbin Medical University, Harbin, China
| | - Xiaoguang Cui
- Department of Anesthesiology, Hei Long Jiang Province Key Lab of Research on Anesthesiology and Critical Care Medicine, Second Affiliated Hospital, Harbin Medical University, Harbin, China; Department of Anesthesiology, First Affiliated Hospital, Hainan Medical University, Hainan, China.
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Ying D, Zhou X, Ruan Y, Wang L, Wu X. LncRNA Gm4419 induces cell apoptosis in hepatic ischemia-reperfusion injury via regulating the miR-455-SOX6 axis. Biochem Cell Biol 2020; 98:474-483. [PMID: 32114773 DOI: 10.1139/bcb-2019-0331] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Long non-coding RNA (lncRNA) is known to be involved in a variety of diseases. However, the role of Gm4419 in hepatic ischemia-reperfusion (I/R) injury remains unknown. To study this, we first established a rat model of hepatic I/R, and a BRL-3A cell model of hypoxia-reoxygenation (H/R) for in vivo and in vitro studies. Staining with hematoxylin and eosin and hepatic injury scores were used to evaluate the degree of hepatic I/R injury. Cell apoptosis was assessed via staining with Edu, and with annexin V-FITC-propidium iodide assays. The interactions between Gm4419 and miR-455, as well as miR-455 and SOX6 were evaluated via luciferase reporter activity assays and RNA immunoprecipitation assays. In vivo, we found that Gm4419 was up-regulated in the rats subjected to I/R. Moreover, knockdown of Gm4419 alleviated the I/R-induced liver damage in the rats. In vitro, knockdown of Gm4419 alleviated H/R-induced apoptosis in BRL-3A cells. Interestingly, we found that miR-455 is a target of Gm4419, and Gm4419 regulates the expression of miR-455 via sponging. Furthermore, SOX6 was proven to be the target of miR-455. Finally, rescue experiments confirmed that knockdown of Gm4419 inhibits apoptosis by regulating miR-455 and SOX6 in H/R-treated BRL-3A cells. Therefore, our findings show that the lncRNA Gm4419 accelerates hepatic I/R injury by targeting the miR-455-SOX6 axis, which suggests a novel therapeutic target for hepatic I/R injury.
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Affiliation(s)
- Dongjian Ying
- Department of Minimal Invasive Surgery, Ningbo Medical Center Lihuili Hospital, Ningbo City, Zhejiang Province, 315040, China.,Department of Minimal Invasive Surgery, Ningbo Medical Center Lihuili Hospital, Ningbo City, Zhejiang Province, 315040, China
| | - Xinhua Zhou
- Department of Minimal Invasive Surgery, Ningbo Medical Center Lihuili Hospital, Ningbo City, Zhejiang Province, 315040, China.,Department of Minimal Invasive Surgery, Ningbo Medical Center Lihuili Hospital, Ningbo City, Zhejiang Province, 315040, China
| | - Yi Ruan
- Department of Minimal Invasive Surgery, Ningbo Medical Center Lihuili Hospital, Ningbo City, Zhejiang Province, 315040, China.,Department of Minimal Invasive Surgery, Ningbo Medical Center Lihuili Hospital, Ningbo City, Zhejiang Province, 315040, China
| | - Luoluo Wang
- Department of Minimal Invasive Surgery, Ningbo Medical Center Lihuili Hospital, Ningbo City, Zhejiang Province, 315040, China.,Department of Minimal Invasive Surgery, Ningbo Medical Center Lihuili Hospital, Ningbo City, Zhejiang Province, 315040, China
| | - Xiang Wu
- Department of Minimal Invasive Surgery, Ningbo Medical Center Lihuili Hospital, Ningbo City, Zhejiang Province, 315040, China.,Department of Minimal Invasive Surgery, Ningbo Medical Center Lihuili Hospital, Ningbo City, Zhejiang Province, 315040, China
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9
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Hu C, Li L. Pre-conditions for eliminating mitochondrial dysfunction and maintaining liver function after hepatic ischaemia reperfusion. J Cell Mol Med 2017; 21:1719-1731. [PMID: 28301072 PMCID: PMC5571537 DOI: 10.1111/jcmm.13129] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2016] [Accepted: 01/13/2017] [Indexed: 12/16/2022] Open
Abstract
The liver, the largest organ with multiple synthesis and secretion functions in mammals, consists of hepatocytes and Kupffer, stem, endothelial, stellate and other parenchymal cells. Because of early and extensive contact with the external environment, hepatic ischaemia reperfusion (IR) may result in mitochondrial dysfunction, autophagy and apoptosis of cells and tissues under various pathological conditions. Because the liver requires a high oxygen supply to maintain normal detoxification and synthesis functions, it is extremely susceptible to ischaemia and subsequent reperfusion with blood. Consequently, hepatic IR leads to acute or chronic liver failure and significantly increases the total rate of morbidity and mortality through multiple regulatory mechanisms. An increasing number of studies indicate that mitochondrial structure and function are impaired after hepatic IR, but that the health of liver tissues or liver grafts can be effectively rescued by attenuation of mitochondrial dysfunction. In this review, we mainly focus on the subsequent therapeutic interventions related to the conservation of mitochondrial function involved in mitigating hepatic IR injury and the potential mechanisms of protection. Because mitochondria are abundant in liver tissue, clarification of the regulatory mechanisms between mitochondrial dysfunction and hepatic IR should shed light on clinical therapies for alleviating hepatic IR‐induced injury.
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Affiliation(s)
- Chenxia Hu
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, China
| | - Lanjuan Li
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, China
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10
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Bettermann K. NF-κB and Its Implication in Liver Health and Cancer Development. MECHANISMS OF MOLECULAR CARCINOGENESIS – VOLUME 1 2017:87-114. [DOI: 10.1007/978-3-319-53659-0_6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/04/2025]
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11
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Li DY, Shi XJ, Li W, Du XH, Wang GY. Key Points in Establishing a Model of Mouse Liver Transplantation. Transplant Proc 2016; 47:2683-9. [PMID: 26680072 DOI: 10.1016/j.transproceed.2015.07.031] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2015] [Accepted: 07/14/2015] [Indexed: 12/14/2022]
Abstract
The explosion of interest in research into the mouse genome and immune system has meant that the mouse orthotopic liver transplantation (MOLT) model has become a popular means of studying transplantation immunity, organ preservation, ischemia-reperfusion injury, and surgical techniques, among others. Although numerous modifications and refinements of surgical techniques have simplified the operation, the relatively short duration of postoperative survival after MOLT remains an obstacle to longer-term follow-up studies. Here, we summarize the scientific basis of MOLT and our experience improving and refining the model in six key areas: anesthesia, operative technique, perfusion and preservation of the liver, cuff technique, anhepatic time, and the value of rearterialization for the liver graft. We also compare the characteristics of different surgical techniques, and give recommendations for the best means of tailoring technique to the objectives of a study. In doing so, we aim to assist other investigators in establishing and perfecting the MOLT model in their routine research practice.
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Affiliation(s)
- D-Y Li
- Department of Hepatobiliary & Pancreatic Surgery, the First Norman Bethune Hospital Affiliated to Jilin University, Jilin Province, China
| | - X-J Shi
- Department of Hepatobiliary & Pancreatic Surgery, the First Norman Bethune Hospital Affiliated to Jilin University, Jilin Province, China
| | - W Li
- Department of Hepatobiliary & Pancreatic Surgery, Third Hospital (China-Japan Union Hospital) of Jilin University, Jilin Province, China
| | - X-H Du
- Department of Hepatobiliary & Pancreatic Surgery, the First Norman Bethune Hospital Affiliated to Jilin University, Jilin Province, China
| | - G-Y Wang
- Department of Hepatobiliary & Pancreatic Surgery, the First Norman Bethune Hospital Affiliated to Jilin University, Jilin Province, China.
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Yokota S, Yoshida O, Ono Y, Geller DA, Thomson AW. Liver transplantation in the mouse: Insights into liver immunobiology, tissue injury, and allograft tolerance. Liver Transpl 2016; 22:536-46. [PMID: 26709949 PMCID: PMC4811737 DOI: 10.1002/lt.24394] [Citation(s) in RCA: 40] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/04/2015] [Revised: 12/18/2015] [Accepted: 12/21/2015] [Indexed: 12/13/2022]
Abstract
The surgically demanding mouse orthotopic liver transplant model was first described in 1991. It has proved to be a powerful research tool for the investigation of liver biology, tissue injury, the regulation of alloimmunity and tolerance induction, and the pathogenesis of specific liver diseases. Liver transplantation in mice has unique advantages over transplantation of the liver in larger species, such as the rat or pig, because the mouse genome is well characterized and there is much greater availability of both genetically modified animals and research reagents. Liver transplant experiments using various transgenic or gene knockout mice have provided valuable mechanistic insights into the immunobiology and pathobiology of the liver and the regulation of graft rejection and tolerance over the past 25 years. The molecular pathways identified in the regulation of tissue injury and promotion of liver transplant tolerance provide new potential targets for therapeutic intervention to control adverse inflammatory responses/immune-mediated events in the hepatic environment and systemically. In conclusion, orthotopic liver transplantation in the mouse is a valuable model for gaining improved insights into liver biology, immunopathology, and allograft tolerance that may result in therapeutic innovation in the liver and in the treatment of other diseases.
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Affiliation(s)
- Shinichiro Yokota
- Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, Address correspondence to: Angus W. Thomson, PhD DSc, FRCPath, FAST, University of Pittsburgh, 200 Lothrop Street, BST W1540, Pittsburgh, PA 15235; ; (412) 624-6392
| | - Osamu Yoshida
- Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261
| | | | - David A. Geller
- Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261,Liver Cancer Center, University of Pittsburgh Medical Center, Pittsburgh, PA 15261
| | - Angus W. Thomson
- Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261,Department of Immunology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261
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13
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Zhu P, Esckilsen S, Atkinson C, Chen XP, Nadig SN. A simplified cuff technique for abdominal aortic transplantation in mice. J Surg Res 2015; 200:707-13. [PMID: 26375503 DOI: 10.1016/j.jss.2015.08.039] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2015] [Revised: 08/06/2015] [Accepted: 08/21/2015] [Indexed: 12/18/2022]
Abstract
BACKGROUND Allograft arteriopathy is still a leading cause of late organ failure. The aortic allograft model in mice has been used to study chronic rejection and has given useful information in the development of graft arteriosclerosis. However, the technical difficulties of small vessel anastomoses still continue to limit its widespread use. We introduce a new simple method for aortic transplantation in mice. METHODS The descending aorta or infrarenal aorta from the donor mouse was anastomosed to the infrarenal aorta using a cuff technique. Aortic transplantation was performed in 30 mice, 10 isografts and 20 allografts. No immunosuppression was administered, and the recipients were sacrificed at day 28. The grafts were histologically analyzed. RESULTS Implantation of grafts could be completed in an average of 23 min. There was no technical failure in all 60 anastomoses. The overall survival rate was 93.3%. Histology of aortas revealed typical aspects of chronic rejection in the allografts at day 28. No significant lesion was observed in isografts. CONCLUSIONS We have developed an innovative, stable, and simple aortic transplantation model in mice, which is useful for vascular research in transplantation and beyond.
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MESH Headings
- Allografts/pathology
- Allografts/transplantation
- Anastomosis, Surgical
- Animals
- Aorta, Abdominal/pathology
- Aorta, Abdominal/transplantation
- Aorta, Thoracic/pathology
- Aorta, Thoracic/transplantation
- Graft Rejection/pathology
- Isografts/pathology
- Isografts/transplantation
- Male
- Mice
- Mice, Inbred BALB C/surgery
- Mice, Inbred C57BL/surgery
- Models, Animal
- Transplantation, Homologous/methods
- Transplantation, Isogeneic/methods
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Affiliation(s)
- Peng Zhu
- Division of Transplantation, Department of Surgery, Hepatic Surgery Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China; Department of Microbiology and Immunology, Medical University of South Carolina, Charleston, South Carolina
| | - Scott Esckilsen
- Department of Microbiology and Immunology, Medical University of South Carolina, Charleston, South Carolina
| | - Carl Atkinson
- Department of Microbiology and Immunology, Medical University of South Carolina, Charleston, South Carolina; Division of Transplantation, Department of Surgery, Members of the South Carolina Investigators in Transplantation (SCIT), Charleston, SC
| | - Xiao-Ping Chen
- Division of Transplantation, Department of Surgery, Hepatic Surgery Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Satish N Nadig
- Department of Microbiology and Immunology, Medical University of South Carolina, Charleston, South Carolina; Division of Transplantation, Department of Surgery, Members of the South Carolina Investigators in Transplantation (SCIT), Charleston, SC.
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14
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Liu Q, Rehman H, Krishnasamy Y, Schnellmann RG, Lemasters JJ, Zhong Z. Improvement of liver injury and survival by JNK2 and iNOS deficiency in liver transplants from cardiac death mice. J Hepatol 2015; 63:68-74. [PMID: 25703084 PMCID: PMC4475508 DOI: 10.1016/j.jhep.2015.02.017] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/23/2014] [Revised: 02/09/2015] [Accepted: 02/11/2015] [Indexed: 01/09/2023]
Abstract
BACKGROUND & AIMS Inclusion of liver grafts from cardiac death donors (CDD) would increase the availability of donor livers but is hampered by a higher risk of primary non-function. Here, we seek to determine mechanisms that contribute to primary non-function of liver grafts from CDD with the goal to develop strategies for improved function and outcome, focusing on c-Jun-N-terminal kinase (JNK) activation and mitochondrial depolarization, two known mediators of graft failure. METHODS Livers explanted from wild-type, inducible nitric oxide synthase knockout (iNOS(-/-)), JNK1(-/-) or JNK2(-/-) mice after 45-min aorta clamping were implanted into wild-type recipients. Mitochondrial depolarization was detected by intravital confocal microscopy in living recipients. RESULTS After transplantation of wild-type CDD livers, graft iNOS expression and 3-nitrotyrosine adducts increased, but hepatic endothelial NOS expression was unchanged. Graft injury and dysfunction were substantially higher in CDD grafts than in non-CDD grafts. iNOS deficiency and inhibition attenuated injury and improved function and survival of CDD grafts. JNK1/2 and apoptosis signal-regulating kinase-1 activation increased markedly in wild-type CDD grafts, which was blunted by iNOS deficiency. JNK inhibition and JNK2 deficiency, but not JNK1 deficiency, decreased injury and improved function and survival of CDD grafts. Mitochondrial depolarization and binding of phospho-JNK2 to Sab, a mitochondrial protein linked to the mitochondrial permeability transition, were higher in CDD than in non-CDD grafts. iNOS deficiency, JNK inhibition and JNK2 deficiency all decreased mitochondrial depolarization and blunted ATP depletion in CDD grafts. JNK inhibition and deficiency did not decrease 3-nitrotyrosine adducts in CDD grafts. CONCLUSION The iNOS-JNK2-Sab pathway promotes CDD graft failure via increased mitochondrial depolarization, and is an attractive target to improve liver function and survival in CDD liver transplantation recipients.
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Affiliation(s)
- Qinlong Liu
- Department of Drug Discovery & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, United States; The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
| | - Hasibur Rehman
- Department of Drug Discovery & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, United States
| | - Yasodha Krishnasamy
- Department of Drug Discovery & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, United States
| | - Rick G Schnellmann
- Department of Drug Discovery & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, United States; Ralph H. Johnson Veterans Affairs Medical Center, Charleston, SC 29403, United States
| | - John J Lemasters
- Department of Drug Discovery & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, United States; Department of Biochemistry & Molecular Biology, Medical University of South Carolina, Charleston, SC 29425, United States
| | - Zhi Zhong
- Department of Drug Discovery & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, United States.
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Mobasher MA, Valverde ÁM. Signalling pathways involved in paracetamol-induced hepatotoxicity: new insights on the role of protein tyrosine phosphatase 1B. Arch Physiol Biochem 2014; 120:51-63. [PMID: 24738658 DOI: 10.3109/13813455.2014.893365] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Acute hepatic failure secondary to paracetamol poisoning is associated with high mortality. Paracetamol-induced hepatotoxicity causes oxidative stress that triggers signalling pathways and ultimately leads to lethal hepatocyte injury. We will review the signalling pathways activated by paracetamol in the liver emphasizing the role of protein tyrosine phosphatase 1B (PTP1B) in the balance between cell death and survival in hepatocytes. PTP1B has emerged as a key modulator of the antioxidant system mediated by the nuclear factor erythroid-2-related factor 2 (Nrf2) in hepatic cells in response to paracetamol overdose. Also, this phosphatase modulates the classical survival pathways triggered by the activation of the insulin-like growth factor-I (IGF-I) signalling cascade. Therefore, PTP1B is a novel therapeutic target against paracetamol-induced liver failure.
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Affiliation(s)
- Maysa Ahmed Mobasher
- Instituto de Investigaciones Biomédicas Alberto Sols (CSIC-UAM), 28029 Madrid, Spain, and Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM) , ISCIII , Spain
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Chen J, Gong W, Ge F, Huang T, Wu D, Liang T. A review of various techniques of mouse liver transplantation. Transplant Proc 2014; 45:2517-21. [PMID: 23953573 DOI: 10.1016/j.transproceed.2013.03.035] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2012] [Revised: 12/10/2012] [Accepted: 03/06/2013] [Indexed: 12/15/2022]
Abstract
OBJECTIVES Liver transplantation in a mouse model is a valuable tool for studying transplantation immunobiology and clinic-relevant issues. However, the successful establishment is highly technical and demanding, impeding its widespread use. Herein, the aims of this study were to review and analyze the various techniques of liver transplantation in mice to circumvent pitfalls and minimize the incidence of complications. MATERIALS AND METHODS A search of PubMed was made by using the key words "mouse liver transplantation" for articles published between January 1973 and July 2012. Of the 473 publications identified, 14 were shown to be closely associated with mouse liver transplantation and 4 articles discussed specific microsurgical techniques. Through reviewing these articles, a series of potential factors were collected and analyzed in combination with other murine transplantation models, which might influence successfully establishing a mouse model for liver transplantation. RESULTS A mouse liver transplantation model is feasible and practical for experimental studies. Mouse strain, type of anesthesia, type of perfusion and storage solution, and reconstruction of bile duct are relevant factors but not determinants for a successful transplantation. Cold and warm ischemia time should be less than 4.0 hours and 20 minutes, respectively. CONCLUSIONS The cuff preparation, reconstruction of the hepatic artery, and length of the anhepatic phase play critical roles in successfully establishing a liver transplantation model in mice.
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Affiliation(s)
- J Chen
- Department of Surgery, Transplant International Research Centre, Second Affiliated Hospital of School of Medicine, Zhejiang University, Hangzhou City, People's Republic of China
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Pan N, Lv X, Liang R, Wang L, Liu Q. Suppression of graft regeneration, not ischemia/reperfusion injury, is the primary cause of small-for-size syndrome after partial liver transplantation in mice. PLoS One 2014; 9:e93636. [PMID: 24709852 PMCID: PMC3977924 DOI: 10.1371/journal.pone.0093636] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2013] [Accepted: 03/06/2014] [Indexed: 01/10/2023] Open
Abstract
Background Ischemia/reperfusion injury (IRI) is commonly considered to play a crucial role in the pathogenesis of small-for-size syndrome (SFSS) after liver transplantation. Rapid regeneration is also considered essential for the survival of SFS grafts. Methods Mouse models of full-size orthotopic liver transplantation, 50% partial liver transplantation and 30% partial liver transplantation were established. Survival rate and serum alanine aminotransferase were observed. IRI was assessed by hepatic pathologic alterations, apoptosis and necrosis. Regeneration response was detected by mitotic index, BrdU incorporation and PCNA, Cyclin D1 and Cyclin E expression. The expression of mTOR, AKT, ERK, JNK2 and p70S6K, also involved in regeneration signaling pathways, were analyzed as well. Results 30% partial liver graft resulted in a significantly low 7-day survival rate (P = 0.002) with no marked difference in tissue injury compared with the 50% partial graft group. Serum alanine aminotransferase levels were not significantly different between partial transplantation and full-size transplantation. Western blot analysis of caspase-3 and TUNEL staining also indicated no significant difference in apoptosis response between 30% partial transplantation and half-size or full-size transplantation (P = 0.436, P = 0.113, respectively). However, liver regeneration response indicators, mitotic index (P<0.0001) and BrdU (P = 0.0022), were markedly lower in 30% LTx compared with 50% LTx. Suppressed expression of PCNA, cyclin D1, cyclin E, mTOR, JNK2, AKT, ERK and p70S6K was also detected by western blot. Conclusions Liver regeneration is markedly suppressed in SFSS, and is more likely the primary cause of SFSS, rather than ischemia/reperfusion injury. Therapy for recovering graft regeneration could be a potentially important strategy to reduce the incidence of SFSS.
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Affiliation(s)
- Ning Pan
- Department of Anesthesiology, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
| | - Xiangwei Lv
- Department of General Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
| | - Rui Liang
- Department of General Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
| | - Liming Wang
- Department of General Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
| | - Qinlong Liu
- Department of General Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian, Liaoning Province, China
- * E-mail:
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18
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Peralta C, Jiménez-Castro MB, Gracia-Sancho J. Hepatic ischemia and reperfusion injury: effects on the liver sinusoidal milieu. J Hepatol 2013; 59:1094-106. [PMID: 23811302 DOI: 10.1016/j.jhep.2013.06.017] [Citation(s) in RCA: 456] [Impact Index Per Article: 38.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/30/2013] [Revised: 06/17/2013] [Accepted: 06/18/2013] [Indexed: 12/16/2022]
Abstract
Ischemia-reperfusion injury is an important cause of liver damage occurring during surgical procedures including hepatic resection and liver transplantation, and represents the main underlying cause of graft dysfunction post-transplantation. Cellular and biochemical processes occurring during hepatic ischemia-reperfusion are diverse and complex, and include the deregulation of the healthy phenotype of all liver cellular components. Nevertheless, a significant part of these processes are still unknown or unclear. The present review aims at summarizing the current knowledge in liver ischemia-reperfusion, but specifically focusing on liver cell phenotype and paracrine interaction deregulations. Moreover, the most updated therapeutic strategies including pharmacological, genetic and surgical interventions, as well as some of the scientific controversies in the field will be described. Finally, the importance of considering the subclinical situation of liver grafts when translating basic knowledge to the bedside is discussed.
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Affiliation(s)
- Carmen Peralta
- Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Centro de Investigación Biomédica en Red en Enfermedades Hepáticas y Digestivas (CIBEREHD), Barcelona, Spain
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Datta G, Fuller BJ, Davidson BR. Molecular mechanisms of liver ischemia reperfusion injury: Insights from transgenic knockout models. World J Gastroenterol 2013; 19:1683-98. [PMID: 23555157 PMCID: PMC3607745 DOI: 10.3748/wjg.v19.i11.1683] [Citation(s) in RCA: 152] [Impact Index Per Article: 12.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/13/2012] [Revised: 06/29/2012] [Accepted: 07/09/2012] [Indexed: 02/06/2023] Open
Abstract
Ischemia reperfusion injury is a major obstacle in liver resection and liver transplantation surgery. Understanding the mechanisms of liver ischemia reperfusion injury (IRI) and developing strategies to counteract this injury will therefore reduce acute complications in hepatic resection and transplantation, as well as expanding the potential pool of usable donor grafts. The initial liver injury is initiated by reactive oxygen species which cause direct cellular injury and also activate a cascade of molecular mediators leading to microvascular changes, increased apoptosis and acute inflammatory changes with increased hepatocyte necrosis. Some adaptive pathways are activated during reperfusion that reduce the reperfusion injury. IRI involves a complex interplay between neutrophils, natural killer T-cells cells, CD4+ T cell subtypes, cytokines, nitric oxide synthases, haem oxygenase-1, survival kinases such as the signal transducer and activator of transcription, Phosphatidylinositol 3-kinases/Akt and nuclear factor κβ pathways. Transgenic animals, particularly genetic knockout models, have become a powerful tool at elucidating mechanisms of liver ischaemia reperfusion injury and are complementary to pharmacological studies. Targeted disruption of the protein at the genetic level is more specific and maintained than pharmacological inhibitors or stimulants of the same protein. This article reviews the evidence from knockout models of liver IRI about the cellular and molecular mechanisms underlying liver IRI.
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20
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Ocuin LM, Zeng S, Cavnar MJ, Sorenson EC, Bamboat ZM, Greer JB, Kim TS, Popow R, DeMatteo RP. Nilotinib protects the murine liver from ischemia/reperfusion injury. J Hepatol 2012; 57:766-73. [PMID: 22641092 PMCID: PMC3437237 DOI: 10.1016/j.jhep.2012.05.012] [Citation(s) in RCA: 45] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/30/2012] [Revised: 05/15/2012] [Accepted: 05/19/2012] [Indexed: 12/13/2022]
Abstract
BACKGROUND & AIMS The mitogen-activated protein kinases (MAPKs), c-Jun N-terminal kinase (JNK), and p38, mediate liver ischemia/reperfusion (I/R) injury via cell death and inflammatory cytokine expression, respectively. Nilotinib is an orally available receptor tyrosine kinase inhibitor used for chronic myelogenous leukemia that also has in vitro activity against JNK and p38. In this study, we examine its therapeutic potential against hepatic I/R injury. METHODS The effects of nilotinib on liver I/R injury were tested using a murine model of warm, segmental liver I/R. Serum ALT was measured and livers were analyzed by histology, RT-PCR, Western blot, and flow cytometry. The in vitro effects of nilotinib on hepatocyte and non-parenchymal cell (NPC) MAPK activation and cytokine production were also tested. RESULTS Mice receiving nilotinib had markedly lower serum ALT levels and less histologic injury and apoptosis following liver I/R. Nilotinib did not inhibit its known receptor tyrosine kinases. Nilotinib lowered intrahepatic expression of IL-1β, IL-6, MCP-1, and MIP-2 and systemic levels of IL-6, MCP-1, and TNF. Nilotinib reduced NPC activation of p38 MAPK signaling and decreased the recruitment of inflammatory monocytes and their production of TNF. Nilotinib attenuated JNK phosphorylation and hepatocellular apoptosis. In vitro, nilotinib demonstrated direct inhibition of JNK activation in isolated hepatocytes cultured under hypoxic conditions, and blocked activation of p38 MAPK and cytokine production by stimulated NPCs. CONCLUSIONS Nilotinib lowers both liver JNK activation and NPC p38 MAPK activation and may be useful for ameliorating liver I/R injury in humans.
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Affiliation(s)
- Lee M Ocuin
- Hepatopancreatobiliary Service, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA
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21
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Liu Q, Rehman H, Krishnasamy Y, Haque K, Schnellmann R, Lemasters J, Zhong Z. Amphiregulin stimulates liver regeneration after small-for-size mouse liver transplantation. Am J Transplant 2012; 12:2052-61. [PMID: 22694592 PMCID: PMC3409348 DOI: 10.1111/j.1600-6143.2012.04069.x] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/25/2023]
Abstract
This study investigated whether amphiregulin (AR), a ligand of the epidermal growth factor receptor (EGFR), improves liver regeneration after small-for-size liver transplantation. Livers of male C57BL/6 mice were reduced to ~50% and ~30% of original sizes and transplanted. After transplantation, AR and AR mRNA increased in 50% but not in 30% grafts. 5-Bromodeoxyuridine (BrdU) labeling, proliferating cell nuclear antigen (PCNA) expression and mitotic index increased substantially in 50% but not 30% grafts. Hyperbilirubinemia and hypoalbuminemia occurred and survival decreased after transplantation of 30% but not 50% grafts. AR neutralizing antibody blunted regeneration in 50% grafts whereas AR injection (5 μg/mouse, iv) stimulated liver regeneration, improved liver function and increased survival after transplantation of 30% grafts. Phosphorylation of EGFR and its downstream signaling molecules Akt, mTOR, p70S6K, ERK and JNK increased markedly in 50% but not 30% grafts. AR stimulated EGFR phosphorylation and its downstream signaling pathways. EGFR inhibitor PD153035 suppressed regeneration of 50% grafts and largely abrogated stimulation of regeneration of 30% grafts by AR. AR also increased cyclin D1 and cyclin E expression in 30% grafts. Together, liver regeneration is suppressed in small-for-size grafts, as least in part, due to decreased AR formation. AR supplementation could be a promising therapy to stimulate regeneration of partial liver grafts.
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Affiliation(s)
- Q. Liu
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425,Department of General Surgery, the Second Affiliated Hospital of Dalian Medical University, Dalian, China
| | - H. Rehman
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425
| | - Y. Krishnasamy
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425
| | - K. Haque
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425
| | - R.G. Schnellmann
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425,Ralph H. Johnson VA Medical Center, Charleston, SC 29403
| | - J.J. Lemasters
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425,Department of Biochemistry & Molecular Biology, Medical University of South Carolina, Charleston, SC 29425,Hollings Cancer Center, Medical University of South Carolina, Charleston, SC 29425
| | - Z. Zhong
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425,Hollings Cancer Center, Medical University of South Carolina, Charleston, SC 29425
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Seki E, Brenner DA, Karin M. A liver full of JNK: signaling in regulation of cell function and disease pathogenesis, and clinical approaches. Gastroenterology 2012; 143:307-20. [PMID: 22705006 PMCID: PMC3523093 DOI: 10.1053/j.gastro.2012.06.004] [Citation(s) in RCA: 389] [Impact Index Per Article: 29.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/06/2012] [Revised: 06/05/2012] [Accepted: 06/11/2012] [Indexed: 02/08/2023]
Abstract
c-Jun-N-terminal kinase (JNK) is a mitogen-activated protein kinase family member that is activated by diverse stimuli, including cytokines (such as tumor necrosis factor and interleukin-1), reactive oxygen species (ROS), pathogens, toxins, drugs, endoplasmic reticulum stress, free fatty acids, and metabolic changes. Upon activation, JNK induces multiple biologic events through the transcription factor activator protein-1 and transcription-independent control of effector molecules. JNK isozymes regulate cell death and survival, differentiation, proliferation, ROS accumulation, metabolism, insulin signaling, and carcinogenesis in the liver. The biologic functions of JNK are isoform, cell type, and context dependent. Recent studies using genetically engineered mice showed that loss or hyperactivation of the JNK pathway contributes to the development of inflammation, fibrosis, cancer growth, and metabolic diseases that include obesity, hepatic steatosis, and insulin resistance. We review the functions and pathways of JNK in liver physiology and pathology and discuss findings from preclinical studies with JNK inhibitors.
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Affiliation(s)
- Ekihiro Seki
- Department of Medicine, University of California, La Jolla, CA, USA.
| | - David A. Brenner
- Department of Medicine, Division of Gastroenterology, University of California, San Diego, School of Medicine, La Jolla, CA, USA
| | - Michael Karin
- Laboratory of Gene Regulation and Signal Transduction, Department of Pharmacology, University of California, San Diego, School of Medicine, La Jolla, CA, USA
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Liu Q, Rehman H, Shi Y, Krishnasamy Y, Lemasters JJ, Smith CD, Zhong Z. Inhibition of sphingosine kinase-2 suppresses inflammation and attenuates graft injury after liver transplantation in rats. PLoS One 2012; 7:e41834. [PMID: 22848628 PMCID: PMC3405047 DOI: 10.1371/journal.pone.0041834] [Citation(s) in RCA: 33] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/18/2012] [Accepted: 06/25/2012] [Indexed: 11/28/2022] Open
Abstract
Inflammation mediates/promotes graft injury after liver transplantation (LT). This study investigated the roles of sphingosine kinase-2 (SK2) in inflammation after LT. Liver grafts were stored in UW solution with and without ABC294640 (100 µM), a selective inhibitor of SK2, before implantation. Hepatic sphingosine-1-phosphate (S1P) levels increased ∼4-fold after LT, which was blunted by 40% by ABC294640. Hepatic toll-like receptor-4 (TLR4) expression and nuclear factor-κB (NF-κB) p65 subunit phosphorylation elevated substantially after transplantation. The pro-inflammatory cytokines/chemokines tumor necrosis factor-α, interleukin-1β and C-X-C motif chemokine 10 mRNAs increased 5.9-fold, 6.1-fold and 16-fold, respectively following transplantation, while intrahepatic adhesion molecule-1 increased 5.7-fold and monocytes/macrophage and neutrophil infiltration and expansion of residential macrophage population increased 7.8-13.4 fold, indicating enhanced inflammation. CD4+ T cell infiltration and interferon-γ production also increased. ABC294640 blunted TLR4 expression by 60%, NF-κB activation by 84%, proinflammatory cytokine/chemokine production by 45-72%, adhesion molecule expression by 54% and infiltration of monocytes/macrophages and neutrophils by 62-67%. ABC294640 also largely blocked CD4+ T cell infiltration and interferon-γ production. Focal necrosis and apoptosis occurred after transplantation with serum alanine aminotransferase (ALT) reaching ∼6000 U/L and serum total bilirubin elevating to ∼1.5 mg/dL. Inhibition of SK2 by ABC294640 blunted necrosis by 57%, apoptosis by 74%, ALT release by ∼68%, and hyperbilirubinemia by 74%. Most importantly, ABC294640 also increased survival from ∼25% to ∼85%. In conclusion, SK2 plays an important role in hepatic inflammation responses and graft injury after cold storage/transplantation and represents a new therapeutic target for liver graft failure.
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Affiliation(s)
- Qinlong Liu
- Departments of Pharmaceutical & Biomedical Sciences and Medical University of South Carolina, Charleston, South Carolina, United States of America
| | - Hasibur Rehman
- Departments of Pharmaceutical & Biomedical Sciences and Medical University of South Carolina, Charleston, South Carolina, United States of America
| | - Yanjun Shi
- Departments of Pharmaceutical & Biomedical Sciences and Medical University of South Carolina, Charleston, South Carolina, United States of America
| | - Yasodha Krishnasamy
- Departments of Pharmaceutical & Biomedical Sciences and Medical University of South Carolina, Charleston, South Carolina, United States of America
| | - John J. Lemasters
- Departments of Pharmaceutical & Biomedical Sciences and Medical University of South Carolina, Charleston, South Carolina, United States of America
- Biochemistry & Molecular Biology, and Medical University of South Carolina, Charleston, South Carolina, United States of America
- Hollings Cancer Center, Medical University of South Carolina, Charleston, South Carolina, United States of America
| | - Charles D. Smith
- Departments of Pharmaceutical & Biomedical Sciences and Medical University of South Carolina, Charleston, South Carolina, United States of America
- Apogee Biotechnology Corporation, Hummelstown, Pennsylvania, United States of America
| | - Zhi Zhong
- Departments of Pharmaceutical & Biomedical Sciences and Medical University of South Carolina, Charleston, South Carolina, United States of America
- Hollings Cancer Center, Medical University of South Carolina, Charleston, South Carolina, United States of America
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Liu Q, Rehman H, Krishnasamy Y, Ramshesh VK, Theruvath TP, Chavin KD, Schnellmann RG, Lemasters JJ, Zhong Z. Role of inducible nitric oxide synthase in mitochondrial depolarization and graft injury after transplantation of fatty livers. Free Radic Biol Med 2012; 53:250-9. [PMID: 22609250 PMCID: PMC3392495 DOI: 10.1016/j.freeradbiomed.2012.05.012] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/08/2011] [Revised: 04/12/2012] [Accepted: 05/07/2012] [Indexed: 12/23/2022]
Abstract
This study investigated the role of inducible nitric oxide synthase (iNOS) in failure of ethanol-induced fatty liver grafts. Rat livers were explanted 20 h after gavaging with ethanol (5 g/kg) and storing in UW solution for 24h before implantation. Hepatic oil red O staining-positive areas increased from ∼2 to ∼33% after ethanol treatment, indicating steatosis. iNOS expression increased ∼8-fold after transplantation of lean grafts (LG) and 25-fold in fatty grafts (FG). Alanine aminotransferase release, total bilirubin, hepatic necrosis, TUNEL-positive cells, and cleaved caspase-3 were higher in FG than LG. A specific iNOS inhibitor 1400W (5 μM in the cold-storage solution) blunted these alterations by >42% and increased survival of fatty grafts from 25 to 88%. Serum nitrite/nitrate and hepatic nitrotyrosine adducts increased to a greater extent after transplantation of FG than LG, indicating reactive nitrogen species (RNS) overproduction. Phospho-c-Jun and phospho-c-Jun N-terminal kinase-1/2 (JNK1/2) were higher in FG than in LG, indicating more JNK activation in fatty grafts. RNS formation and JNK activation were blunted by 1400W. Mitochondrial polarization and cell death were visualized by intravital multiphoton microscopy of rhodamine 123 and propidium iodide, respectively. After implantation, viable cells with depolarized mitochondria were 3-fold higher in FG than in LG and 1400W decreased mitochondrial depolarization in FG to the levels of LG. Taken together, iNOS is upregulated after transplantation of FG, leading to excessive RNS formation, JNK activation, mitochondrial dysfunction, and severe graft injury. The iNOS inhibitor 1400W could be an effective therapy for primary nonfunction of fatty liver grafts.
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Affiliation(s)
- Qinlong Liu
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA
- Department of General Surgery, the Second Affiliated Hospital of Dalian Medical University, Dalian, China
| | - Hasibur Rehman
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA
| | - Yasodha Krishnasamy
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA
| | - Venkat K. Ramshesh
- Hollings Cancer Center, Medical University of South Carolina, Charleston, SC 29425, USA
| | - Tom P. Theruvath
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA
| | - Kenneth D. Chavin
- Department of Surgery, Medical University of South Carolina, Charleston, SC 29425, USA
| | - Rick G. Schnellmann
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA
- Hollings Cancer Center, Medical University of South Carolina, Charleston, SC 29425, USA
- Ralph H. Johnson VA Medical Center, Charleston, SC 29403, USA
| | - John J. Lemasters
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA
- Department of Biochemistry & Molecular Biology, Medical University of South Carolina, Charleston, SC 29425, USA
- Hollings Cancer Center, Medical University of South Carolina, Charleston, SC 29425, USA
| | - Zhi Zhong
- Department of Pharmaceutical & Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA
- Hollings Cancer Center, Medical University of South Carolina, Charleston, SC 29425, USA
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C-Jun N-Terminal Kinase 2 Promotes Liver Injury via the Mitochondrial Permeability Transition after Hemorrhage and Resuscitation. HPB SURGERY : A WORLD JOURNAL OF HEPATIC, PANCREATIC AND BILIARY SURGERY 2012; 2012:641982. [PMID: 22791932 PMCID: PMC3390051 DOI: 10.1155/2012/641982] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 02/16/2012] [Accepted: 03/24/2012] [Indexed: 12/11/2022]
Abstract
Hemorrhagic shock leads to hepatic hypoperfusion and activation of mitogen-activated stress kinases (MAPK) like c-Jun N-terminal kinase (JNK) 1 and 2. Our aim was to determine whether mitochondrial dysfunction leading to hepatic necrosis and apoptosis after hemorrhage/resuscitation (H/R) was dependent on JNK2. Under pentobarbital anesthesia, wildtype (WT) and JNK2 deficient (KO) mice were hemorrhaged to 30 mm Hg for 3 h and then resuscitated with shed blood plus half the volume of lactated Ringer's solution. Serum alanine aminotransferase (ALT), necrosis, apoptosis and oxidative stress were assessed 6 h after resuscitation. Mitochondrial polarization was assessed by intravital microscopy. After H/R, ALT in WT-mice increased from 130 U/L to 4800 U/L. In KO-mice, ALT after H/R was blunted to 1800 U/l (P < 0.05). Necrosis, caspase-3 activity and ROS were all substantially decreased in KO compared to WT mice after H/R. After sham operation, intravital microscopy revealed punctate mitochondrial staining by rhodamine 123 (Rh123), indicating normal mitochondrial polarization. At 4 h after H/R, Rh123 staining became dim and diffuse in 58% of hepatocytes, indicating depolarization and onset of the mitochondrial permeability transition (MPT). By contrast, KO mice displayed less depolarization after H/R (23%, P < 0.05). In conclusion, JNK2 contributes to MPT-mediated liver injury after H/R.
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Johnston A, Ponzetti K, Anwer MS, Webster CRL. cAMP-guanine exchange factor protection from bile acid-induced hepatocyte apoptosis involves glycogen synthase kinase regulation of c-Jun NH2-terminal kinase. Am J Physiol Gastrointest Liver Physiol 2011; 301:G385-400. [PMID: 21546580 PMCID: PMC3280825 DOI: 10.1152/ajpgi.00430.2010] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/23/2010] [Accepted: 05/01/2011] [Indexed: 01/31/2023]
Abstract
Cholestatic liver disorders are accompanied by the hepatic accumulation of cytotoxic bile acids that induce cell death. Increases in cAMP protect hepatocytes from bile acid-induced apoptosis by a cAMP-guanine exchange factor (cAMP-GEF)/phosphoinositide-3-kinase (PI3K)/Akt pathway. The aim of these studies was to identify the downstream substrate in this pathway and to determine at what level in the apoptotic cascade cytoprotection occurs. Since inhibitory phosphorylation of glycogen synthase kinase-3 (GSK) occurs downstream of PI3K/Akt and this phosphorylation has been implicated in cell survival, we conducted studies to determine whether GSK was downstream in cAMP-GEF/PI3K/Akt-mediated cytoprotection. Our results show that treatment of hepatocytes with the cAMP-GEF-specific analog, 4-(4-chlorophenylthio)-2'-O-methyladenosine-3',5'-cAMP, results in PI3K-dependent phosphorylation of GSK. Direct chemical inhibition of GSK in rat hepatocytes or human HUH7-NTCP cells with several structurally and functionally distinct inhibitors including bromoindirubin-3'-oxime (BIO), maleimides (SB216763, SB415286), thiadiazolidine derivatives, and LiCl attenuates apoptosis induced by glycochenodeoxycholate (GCDC). In addition, genetic silencing of the GSK β isoform with small interfering RNA attenuates GCDC apoptosis in HUH7-NTCP cells. Adenoviral inhibition of the Rap1 blocks both cAMP-GEF-mediated cytoprotection against GCDC-induced apoptosis and Akt/GSK3β phosphorylation. GCDC-induced phosphorylation of the proapoptotic kinase, c-Jun NH(2)-terminal kinase (JNK) is inhibited by GSK inhibition or cAMP-GEF activation. GCDC-induced apoptosis is accompanied by phosphorylation of the endoplasmic reticulum stress markers pIEF2α and IRE-1, and pretreatment with the cAMP-GEF analog or GSK inhibitors prevents this phosphorylation. Collectively, our results support the presence of a cAMP/cAMP-GEF/Rap1/PI3K/Akt/GSKβ survival pathway in hepatocytes that inhibits bile acid-induced JNK phosphorylation.
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Affiliation(s)
| | | | - M. S. Anwer
- Biomedical Science, Tufts Cummings School of Veterinary Medicine, Grafton, Massachusetts
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Shi Y, Rehman H, Wright GL, Zhong Z. Inhibition of inducible nitric oxide synthase prevents graft injury after transplantation of livers from rats after cardiac death. Liver Transpl 2010; 16:1267-77. [PMID: 21031542 PMCID: PMC2967449 DOI: 10.1002/lt.22148] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
This study investigated the roles of inducible nitric oxide synthase (iNOS) in the failure of rat liver grafts from cardiac death donors (GCDD). Livers were explanted after 30-minute aorta clamping and implanted after 4-hour storage in University of Wisconsin solution. The iNOS expression increased slightly in grafts from non-cardiac death donors (GNCDD) but markedly in GCDD. Serum nitrite and nitrate and hepatic 3-nitrotyrosine adducts, indicators of NO and peroxynitrite production, respectively, were substantially higher after transplantation of GCDD than GNCDD. Production of reactive nitrogen species (RNS) was largely blocked by 1400W (N-[1-naphthyl]ethylenediamine dihydrochloride; 5 μM), a specific iNOS inhibitor. Alanine aminotransferase release, bilirubin, necrosis, and apoptosis were 6.4-fold, 6.5-fold, 2.3-fold, and 2.7-fold higher, respectively, after transplantation of GCDD than GNCDD. The inhibitor 1400W effectively blocked these alterations and also increased survival of GCDD to 80% from 33%. Increased RNS production and failure of GCDD were associated with activation of c-Jun-N-terminal kinase (JNK), an effect that was blocked by inhibition of iNOS. Inhibition of JNK also improved the outcome after transplantation of GCDD. Together, the data indicate that iNOS increases substantially in GCDD, leading to RNS overproduction, JNK activation, and more severe graft injury. Inhibitors of iNOS are suggested as effective therapies to improve the outcome after transplantation of GCDD.
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Affiliation(s)
- Yanjun Shi
- Department of Pharmaceutical and Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA
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Pallet N, Bouvier N, Beaune P, Legendre C, Anglicheau D, Thervet E. [Involvement of endoplasmic reticulum stress in solid organ transplantation]. Med Sci (Paris) 2010; 26:397-403. [PMID: 20412745 DOI: 10.1051/medsci/2010264397] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
Endoplasmic reticulum (ER) stress is a situation caused by the accumulation of unfolded proteins in the endoplasmic reticulum, triggering an evolutionary conserved adaptive response termed the unfolded protein response. When adaptation fails, excessive and prolonged ER stress triggers cell suicide. Important roles for ER-initiated cell death pathways have been recognized for several diseases, including diabetes, hypoxia, ischemia/reperfusion injury, neurodegenerative and heart diseases. The implication of the ER stress is not well recognized in solid organ transplantation, but increasing evidence suggests its implication in mediating allograft injury. The purpose of this review is to summarize the mechanisms of ER stress and to discuss its implication during tissue injury in solid organ transplantation. The possible implications of the ER stress in the modifications of cell functional properties and phenotypic changes are also discussed beyond the scope of adaptation and cell death. Increasing the understanding of the cellular and molecular mechanisms of acute and chronic allograft damages could lead to the development of new biomarkers and to the discovery of new therapeutic strategies to prevent the initiation of graft dysfunction or to promote the tissue regeneration after injury.
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Affiliation(s)
- Nicolas Pallet
- Inserm U775, Centre universitaire des Saints-Pères, Paris, France.
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Han D, Shinohara M, Ybanez MD, Saberi B, Kaplowitz N. Signal transduction pathways involved in drug-induced liver injury. Handb Exp Pharmacol 2010:267-310. [PMID: 20020266 DOI: 10.1007/978-3-642-00663-0_10] [Citation(s) in RCA: 84] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
Hepatocyte death following drug intake is the critical event in the clinical manifestation of drug-induced liver injury (DILI). Traditionally, hepatocyte death caused by drugs had been attributed to overwhelming oxidative stress and mitochondria dysfunction caused by reactive metabolites formed during drug metabolism. However, recent studies have also shown that signal transduction pathways activated/inhibited during oxidative stress play a key role in DILI. In acetaminophen (APAP)-induced liver injury, hepatocyte death requires the sustained activation of c-Jun kinase (JNK), a kinase important in mediating apoptotic and necrotic death. Inhibition of JNK using chemical inhibitors or knocking down JNK can prevent hepatocyte death even in the presence of extensive glutathione (GSH) depletion, covalent binding, and oxidative stress. Once activated, JNK translocates to mitochondria, to induce mitochondria permeability transition and trigger hepatocyte death. Mitochondria are central targets where prodeath kinases such as JNK, prosurvival death proteins such as bcl-xl, and oxidative damage converge to determine hepatocyte survival. The importance of mitochondria in DILI is also observed in the Mn-SOD heterozygous (+/-) model, where mice with less mitochondrial Mn-SOD are sensitized to liver injury caused by certain drugs. An extensive body of research is accumulating suggesting a central role of mitochondria in DILI. Drugs can also cause redox changes that inhibit important prosurvival pathways such as NF-kappaB. The inhibition of NF-kappaB by subtoxic doses of APAP sensitizes hepatocyte to the cytotoxic actions of tumor necrosis factor (TNF). Many drugs will induce liver injury if simultaneously treated with LPS, which promotes inflammation and cytokine release. Drugs may be sensitizing hepatocytes to the cytotoxic effects of cytokines such as TNF, or vice versa. Overall many signaling pathways are important in regulating DILI, and represent potential therapeutic targets to reduce liver injury caused by drugs.
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Affiliation(s)
- Derick Han
- Research Center for Liver Diseases, Keck School of Medicine, University of Southern California, 2011 Zonal Ave, HMR 101, Los Angeles, CA 90089-9121, USA.
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Endoplasmic reticulum stress: an unrecognized actor in solid organ transplantation. Transplantation 2009; 88:605-13. [PMID: 19741454 DOI: 10.1097/tp.0b013e3181b22cec] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
Endoplasmic reticulum (ER) stress is an adaptive response to the accumulation of misfolded proteins within the ER, which can trigger cell dedifferentiation and cell suicide. Increasing evidences suggest its implication in mediating allograft injury. Herein, we summarize the mechanisms of ER stress and discuss its implication in allograft injury. Increasing our understanding of the cellular and molecular mechanisms of acute and chronic allograft damages could lead to the development of new biomarkers and to the discovery of new therapeutic strategies to prevent the initiation of graft dysfunction or to promote the tissue regeneration after injury.
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31
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He S, Atkinson C, Evans Z, Ellett JD, Southwood M, Elvington A, Chavin KD, Tomlinson S. A role for complement in the enhanced susceptibility of steatotic livers to ischemia and reperfusion injury. THE JOURNAL OF IMMUNOLOGY 2009; 183:4764-72. [PMID: 19752222 DOI: 10.4049/jimmunol.0900550] [Citation(s) in RCA: 37] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Hepatic steatosis typically renders the donor organ unusable, as donor organs with >30% steatosis are more likely to develop graft failure. The mechanisms leading to failure are not well defined, but steatosis enhances hepatic susceptibility to ischemia reperfusion injury (IRI). We investigated the role of complement in hepatic IRI in lean and steatotic (diet-induced) mice. Steatotic mice were significantly more susceptible to total warm hepatic IRI than lean mice as determined by serum alanine aminotransferase, histopathologically assessed damage, and 24-h survival. C3 deficiency protected both lean and steatotic mice from IRI, as determined by all measured outcomes. Furthermore, treatment of wild-type mice with the complement inhibitor CR2-Crry provided protection equivalent to that seen in C3-deficient mice. Importantly, although steatotic livers were much more susceptible to IRI than lean livers, by most measures there was no statistical difference between the level of IRI to steatotic or lean livers when complement was inhibited. To investigate the clinical relevance of these findings in the context of transplantation, we treated recipients of lean or steatotic liver grafts with saline or CR2-Crry. There was a marked reduction in graft inflammation and injury and significantly improved 7-day survival in CR2-Crry-treated recipients of either lean or steatotic grafts. These data indicate that complement plays a key role in the enhanced susceptibility of steatotic livers to IRI and suggest that complement inhibition represents a potential strategy to reduce the donor shortage by allowing the more routine use of marginal steatotic donor livers.
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Affiliation(s)
- Songqing He
- Department of Microbiology and Immunology, Children's Research Institute, Medical University of South Carolina, Charleston, South Carolina 29245, USA
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Cardinal J, Pan P, Dhupar R, Ross M, Nakao A, Lotze M, Billiar T, Geller D, Tsung A. Cisplatin prevents high mobility group box 1 release and is protective in a murine model of hepatic ischemia/reperfusion injury. Hepatology 2009; 50:565-74. [PMID: 19492424 DOI: 10.1002/hep.23021] [Citation(s) in RCA: 65] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
UNLABELLED The nuclear protein high mobility group box 1 (HMGB1) is an important inflammatory mediator involved in the pathogenesis of liver ischemia/reperfusion (I/R) injury. Strategies aimed at preventing its release from stressed or damaged cells may be beneficial in preventing inflammation after I/R. Cisplatin is a member of the platinating chemotherapeutic agents and can induce DNA lesions that are capable of retaining high mobility group proteins inside the nucleus of cells. In vitro studies in primary cultured rat hepatocytes show that nontoxic concentrations of cisplatin can sequester HMGB1 inside the nucleus of hypoxic cells. Similarly, the in vivo administration of nontoxic doses of cisplatin prevents liver damage associated with a well-established murine model of hepatic I/R as measured by lower circulating serum aminotransferase levels, lower hepatic inflammatory cytokine levels including tumor necrosis factor alpha and interleukin-6, lower inducible NO synthase expression, and fewer I/R-associated histopathologic changes. The mechanism of action in vivo appears to involve the capacity of cisplatin to prevent the I/R-induced release of HMGB1 as well as to alter cell survival and stress signaling in the form of autophagy and mitogen-activated protein kinase activation, respectively. CONCLUSION Low, nontoxic doses of cisplatin can sequester HMGB1 inside the nucleus of redox-stressed hepatocytes in vitro and prevent its release in vivo in a murine model of hepatic I/R. Furthermore, cell survival and stress signaling pathways are altered by low-dose cisplatin. Therefore, platinating agents may provide a novel approach to mitigating the deleterious effects of I/R-mediated disease processes.
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Affiliation(s)
- Jon Cardinal
- Department of Surgery, University of Pittsburgh, Pittsburgh, PA 15213, USA
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33
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Kaplowitz N, Shinohara M, Liu ZX, Han D. How to protect against acetaminophen: don't ask for JUNK. Gastroenterology 2008; 135:1047-51. [PMID: 18782575 DOI: 10.1053/j.gastro.2008.08.031] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
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