亚低温通过磷脂酰肌醇3-激酶/蛋白激酶B信号通路对肝缺血再灌注损伤大鼠模型的保护作用

刘剑; 李立; 王晓川; 张升宁; 李来邦; 莽源祎; 高扬; 陈永林; 任刚; 李望

doi:10.3969/j.issn.1001-5256.2019.04.027

亚低温通过磷脂酰肌醇3-激酶/蛋白激酶B信号通路对肝缺血再灌注损伤大鼠模型的保护作用

DOI: 10.3969/j.issn.1001-5256.2019.04.027

1. 昆明市第一人民医院附属甘美医院肝胆胰血管外科云南省器官移植中心
2. 云南省第一人民医院皮肤科

基金项目:

云南省应用基础研究联合专项[2017FE467(-101)]；中国肝炎防治基金会天晴肝病研究基金(TQGB20180114)；云南省卫生科技计划项目(2016NS327)；

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中图分类号: R-332;R575
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出版历程
- 收稿日期: 2018-10-17
- 出版日期: 2019-04-20

Mild hypothermia exerts a protective effect against hepatic ischemia-reperfusion injury in rats by activating the PI3K/Akt signaling pathway

Department of Hepatopancreatobiliary Vascular Surgery, Ganmei Hospital Affiliated to The First People’s Hospital of Kunming & Yunnan Province Organ Transplant Center

Research funding:

摘要

摘要:
目的探讨亚低温预处理肝缺血再灌注损伤（HIRI）大鼠模型的保护作用机理。方法将40只SD大鼠分为4组:假手术（Sham）组、HIRI组、亚低温处理（MH）组和MH+LY294002处理（MH+LY）组,每组10只,分别于肝缺血再灌注3、6、12、24 h后收集大鼠的血清及肝组织标本。采用Western Blot检测大鼠肝组织中磷脂酰肌醇3-激酶（PI3K）、磷酸化PI3K（p-PI3K）、蛋白激酶B（Akt）和磷酸化Akt（p-Akt,Ser308）蛋白的表达水平。采用TUNEL法和Western Blot检测分析肝组织中细胞凋亡和凋亡相关蛋白的表达水平。采用ELISA试剂盒检测肝组织中超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH-Px）和丙二醛（MDA）水平。采用全自动生化分析仪检测大鼠血清ALT、AST活性。计量资料多组间比较采用单因素方差分析,进一步两两比较采用SNK-q检验。结果 HIRI组p-PI3K、p-Akt的相对表达水平明显低于Sham组,差异均有统计学意义（q值分别为5. 217、5. 456,P值均<0. 01）;经亚低温处理后大鼠肝组织中p-PI3K...
- 肝疾病 /
- 再灌注损伤 /
- 1-磷脂酰肌醇3-激酶 /
- 蛋白激酶类 /
- 大鼠,Sprague-Dawley
Abstract:
Objective To investigate the protective mechanism of mild hypothermia pretreatment against hepatic ischemia-reperfusion injury in rats. Methods A total of 40 male Sprague-Dawley rats were randomly divided into sham-operation group ( Sham group) , hepatic ischemia-reperfusion injury group ( HIRI group) , mild hypothermia group ( MH group) , and mild hypothermia + LY294002 group ( MH +LY group) , with 10 rats in each group. Serum and liver tissue samples were collected at 3, 6, 12, and 24 hours of hepatic ischemia-reperfusion. Western blotting was used to measure the protein expression of phosphoinositide 3-kinase ( PI3 K) , phosphorylated PI3 K ( p-PI3 K) , protein kinase-B ( Akt) , and phosphorylated Akt ( p-Akt, Ser308) in liver tissue. TUNEL and Western blotting were used to measure cell apoptosis and expression of apoptosis-related proteins in liver tissue. ELISA was used to measure the levels of superoxide dismutase ( SOD) , glutathione peroxidase ( GSH-Px) , and malondialdehyde ( MDA) in liver tissue. An automatic biochemical analyzer was used to measure the activities of alanine aminotransferase ( ALT) and aspartate aminotransferase ( AST) in serum. A one-way analysis of variance was used for comparison between multiple groups, and the SNK-q test was used for further comparison between two groups.Results The HIRI group had significantly lower relative expression levels of p-PI3 K and p-Akt than the Sham group ( q = 5. 217 and5. 456, both P < 0. 01) , and the MH group had significantly higher relative expression levels of p-PI3 K and p-Akt in liver tissue than the HIRI group ( q = 10. 434 and 14. 116, both P < 0. 01) . At 3, 6, 12, and 24 hours of hepatic ischemia-reperfusion, the HIRI group, the MH group, and the MH + LY group had significantly higher activities of AST and ALT in serum than the Sham group ( all P < 0. 001) .TUNEL staining showed that the HIRI group had a significantly higher proportion of apoptotic cells in liver tissue than the Sham group ( 42. 25% ± 3. 50% vs 3. 21% ± 0. 5%, q = 10. 187, P < 0. 01) . Compared with the HIRI group, the MH group had a significant reduction in the proportion of apoptotic cells ( q = 7. 784, P < 0. 01) , while there was no significant difference in the proportion of apoptotic cells between the HIRI group and the MH + LY group ( 42. 25% ± 3. 50% vs 38. 19% ± 2. 8%, q = 1. 059, P > 0. 05) . Western blotting showed that compared with the Sham group, the HIRI group had a significant reduction in the expression of the anti-apoptotic protein Bcl-2 ( q =2. 101, P < 0. 05) and significant increases in the expression of the pro-apoptotic proteins Bax, Fas, and Fasl ( q = 11. 016、4. 735, and10. 201, all P < 0. 01) , and the regulatory effect of HIRI on apoptotic-related proteins was downregulated by mild hypothermia treatment.According to the results of oxidative indices, the HIRI group, the MH group, and the MH + LY group had significantly lower expression of SOD and GSH-Px and significantly higher expression of MDA in liver tissue than the Sham group ( all P < 0. 05) ; compared with the HIRI group, the MH group had significantly higher expression of SOD and GSH-Px ( q = 2. 894 and 2. 731, both P < 0. 05) and significantly lower expression of MDA ( q = 5. 888, P < 0. 05) in liver tissue. In addition, mild hypothermia + LY294002 treatment significantly reduced the regulatory effect on the expression of SOD, MDA, and GSH-Px in liver tissue in the MH group ( q = 2. 999, 2. 944, and 2. 620, all P < 0. 05) . Conclusion MH exerts a protective effect against HIRI in rats, possibly by activating the PI3 K/Akt pathway to downregulate hepatocyte apoptosis and enhance antioxidant capacity in vivo.
- liver diseases /
- reperfusion injury /
- 1-phosphatidylinositol 3-kinase /
- protein kinases /
- rats, Sprague-Dawley

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参考文献(22)

[1]SUN P, LU YX, CHENG D, et al.Monocyte Chemoattractant protein-induced protein 1 targets hypoxia-inducible factor1αto protect against hepatic ischemia/reperfusion injury[J].Hepatology, 2018, 68 (6) :2359-2375.

[2]FRATTE S, GENDRAULT JL, STEFFAN AM, et al.Comparative ultrastructural study of rat livers preserved in Euro-Collins or University of Wisconsin solution[J].Hepatology, 1991, 13 (6) :1173-1180.

[3]DAI HB, XU MM, LV J, et al.Mild hypothermia combined with hydrogen sulfide treatment during resuscitation reduces hippocampal neuron apoptosis via NR2A, NR2B, and PI3K-Akt signaling in a rat model of cerebral ischemia-reperfusion injury[J].Mol Neurobiol, 2016, 53 (7) :4865-4873.

[4]CHENG BC, HUANG HS, CHAO CM, et al.Hypothermia may attenuate ischemia/reperfusion-induced cardiomyocyte death by reducing autophagy[J].Int J Cardiol, 2013, 168 (3) :2064-2069.

[5]TSUCHIYA D, HONG S, SUH SW, et al.Mild hypothermia reduces zinc translocation, neuronal cell death, and mortality after transient global ischemia in mice[J].J Cereb Blood Flow Metab, 2002, 22 (10) :1231-1238.

[6]SHRIVASTAVA S, JEENGAR MK, REDDY VS, et al.Anticancer effect of celastrol on human triple negative breast cancer:Possible involvement of oxidative stress, mitochondrial dysfunction, apoptosis and PI3K/Akt pathways[J].Exp Mol Pathol, 2015, 98 (3) :313-327.

[7]WANG XM, YAO M, LIU SX, et al.Interplay between the Notch and PI3K/Akt pathways in high glucose-induced podocyte apoptosis[J].Am J Physiol Renal Physiol, 2014, 306 (2) :F205-F213.

[8]CHEN Y, LIU G, HE F, et al.MicroRNA 375 modulates hyperglycemia-induced enteric glial cell apoptosis and diabetes-induced gastrointestinal dysfunction by targeting Pdk1 and repressing PI3K/Akt pathway[J].Sci Rep, 2018, 8 (1) :12681.

[9]KOHLI V, SELZNER M, MADDEN JF, et al.Endothelial cell and hepatocyte deaths occur by apoptosis after ischemiareperfusion injury in the rat liver[J].Transplantation, 1999, 67 (8) :1099-1105.

[10]GRANGER DN, KVIETYS PR.Reperfusion injury and reactive oxygen species:The evolution of a concept[J].Redox Biol, 2015, 6:524-551.

[11]PELL VR, SPIROSKI AM, MULVEY J, et al.Ischemic preconditioning protects against cardiac ischemia reperfusion injury without affecting succinate accumulation or oxidation[J].JMol Cell Cardiol, 2018, 123:88-91.

[12]YANG F, WANG S, LIU Y, et al.IRE1alpha aggravates ischemia reperfusion injury of fatty liver by regulating phenotypic transformation of kupffer cells[J].Free Radic Biol Med, 2018, 124:395-407.

[13]BEHRENDS M, HIROSE R, SERKOVA NJ, et al.Mild hypothermia reduces the inflammatory response and hepatic ischemia/reperfusion injury in rats[J].Liver Int, 2006, 26 (6) :734-741.

[14]ZHU P, ZHAO MY, LI XH, et al.Effect of low temperatures on BAX and BCL2 proteins in rats with spinal cord ischemia reperfusion injury[J].Genet Mol Res, 2015, 14 (3) :10490-[[[[[[[[10499.

[15]SANTOS EB, KOFF WJ, GREZZANA FILHO TDE J, et al.Oxidative stress evaluation of ischemia and reperfusion in kidneys under various degrees of hypothermia in rats[J].Acta Cir Bras, 2013, 28 (8) :568-573.

[16]ZHOU T, LIANG L, LIANG Y, et al.Mild hypothermia protects hippocampal neurons against oxygen-glucose deprivation/reperfusion-induced injury by improving lysosomal function and autophagic flux[J].Exp Cell Res, 2017, 358 (2) :147-160.

[17]YANG YF, ZHANG MF, TIAN QH, et al.SPAG5 interacts with CEP55 and exerts oncogenic activities via PI3K/AKT pathway in hepatocellular carcinoma[J].Mol Cancer, 2018, 17 (1) :117.

[18]WEI L, CHEN Q, GUO A, et al.Asiatic acid attenuates CCl4-induced liver fibrosis in rats by regulating the PI3K/AKT/m TOR and Bcl-2/Bax signaling pathways[J].Int Immunopharmacol, 2018, 60:1-8.

[19]YIN W, HAN J, ZHANG Z, et al.Aloperine protects mice against bleomycin-induced pulmonary fibrosis by attenuating fibroblast proliferation and differentiation[J].Sci Rep, 2018, 8 (1) :6265.

[20]LI Z, ZHAO F, CAO Y, et al.DHA attenuates hepatic ischemia reperfusion injury by inhibiting pyroptosis and activating PI3K/Akt pathway[J].Eur J Pharmacol, 2018, 835:1-10.

[21]OUYANG ZH, WANG WJ, YAN YG, et al.The PI3K/Akt pathway:A critical player in intervertebral disc degeneration[J].Oncotarget, 2017, 8 (34) :57870-57881.

[22]XIAO Q, YE QF, WANG W, et al.Mild hypothermia pretreatment protects hepatocytes against ischemia reperfusion injury via down-regulating miR-122 and IGF-1R/AKT pathway[J].Cryobiology, 2017, 75:100-105.

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