β-arrestin-2 inhibits autophagy by up-regulating PI3K/Akt signal to alleviate liver ischemia-reperfusion injury in mice
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摘要:
目的 验证β-arrestin-2是否通过上调PI3K/Akt信号抑制自噬对小鼠肝脏缺血-再灌注损伤（IRI）发挥保护作用。 方法 将C57BL/6背景的β-arrestin-2基因敲除（KO）及野生型（WT）小鼠各12只，随机分为KO小鼠假手术组（KO+sham组），KO小鼠IRI组（KO+IRI组），WT小鼠假手术组（WT+sham组）和WT小鼠IRI组（WT+IRI组），每组各6只。分别建立70%肝脏IRI模型或进行假手术处理，于肝脏再灌注或手术后6 h进行相关研究。采用免疫组织化学（免疫组化）染色检测凋亡信号蛋白裂解半胱氨酸天冬氨酸蛋白酶3（cleaved Caspase-3）、增殖信号蛋白Ki-67及PI3K/Akt通路信号蛋白p-Akt的表达。 结果 免疫组化染色结果显示，与相应的sham组比较，KO+IRI组和WT+IRI组小鼠肝组织中的cleaved Caspase-3、Ki-67和p-Akt阳性细胞计数均增加（均为P < 0.01）；与WT+IRI组比较，KO+IRI组小鼠肝组织中cleaved Caspase-3阳性细胞计数增加，Ki-67和p-Akt阳性细胞计数均减少（均为P < 0.05）。 结论 β-arrestin-2可减轻小鼠IRI后肝细胞凋亡、促进其损伤修复，其通过上调PI3K/Akt信号抑制自噬来减轻小鼠肝脏IRI。 -
关键词:
- 肝脏 /
- 缺血-再灌注损伤 /
- 自噬 /
- β-arrestin-2 /
- 基因敲除 /
- PI3K/Akt信号 /
- 免疫组织化学
Abstract:Objective To verify whether β-arrestin-2 inhibits autophagy by up-regulating PI3K/Akt signal to protect the liver from ischemia-reperfusion injury (IRI) in mice. Methods Twelve β-arrestin-2 knockout (KO) and twelve wild-type (WT) C57BL/6 mice were randomly divided into the KO+sham group, KO+IRI group, WT+sham group and WT+IRI group, six mice in each group. The mouse models with 70% liver IRI were established or sham operation was performed. Relevant experiments were carried out at 6 h after liver reperfusion or operation. The expression levels of apoptosis signal protein cleaved Caspase-3, proliferation signal protein Ki-67 and the PI3K/Akt signal protein p-Akt were detected by immunohistochemical staining. Results Immunohistochemical staining demonstrated that compared with the corresponding sham group, the positive cell count for cleaved Caspase-3, Ki-67 and p-Akt in liver tissues of mice was significantly increased in the KO+IRI and WT+IRI groups (all P < 0.01). Compared with the WT+IRI group, the positive cell count for cleaved Caspase-3 in liver tissues of mice was significantly increased, whereas the positive cell count forKi-67 and p-Akt was significantly decreased in the KO+IRI group (both P < 0.05). Conclusions β-arrestin-2 can mitigate the liver cell apoptosis and promote the repair of injury after IRI in mice. Moreover, β-arrestin-2 inhibits autophagy by up-regulating the PI3K/Akt signal to alleviate liver IRI in mice. -
Key words:
- Liver /
- Ischemia-reperfusion injury /
- Autophagy /
- β-arrestin-2 /
- Gene knock out /
- PI3K/Akt signal /
- Immunohistochemistry
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图 1 各组小鼠肝组织中cleaved Caspase-3表达水平的比较（免疫组化, ×200）
注：与相应的sham组比较，aP < 0.01；与WT+IRI组比较，bP < 0.05。
Figure 1. Comparison of the expression of cleaved Caspase-3 in liver tissues of mice among each group
图 2 各组小鼠肝组织中Ki-67表达水平的比较（免疫组化, ×200）
注：与相应的sham组比较，aP < 0.01；与WT+IRI组比较，bP < 0.05。
Figure 2. Comparison of the expression of Ki-67 in liver tissues of mice among each group
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[1] SUN P, ZHANG P, WANG PX, et al. Mindin deficiency protects the liver against ischemia/reperfusion injury[J]. J Hepatol, 2015, 63(5):1198-1211. DOI:10.1016/j.jhep. 2015.06.033. [2] LEVINE B, KROEMER G. Biological functions of autophagy genes: a disease perspective[J]. Cell, 2019, 176(1/2):11-42. DOI: 10.1016/j.cell.2018.09.048. [3] LIU A, FANG H, WEI W, et al. Ischemic preconditioning protects against liver ischemia/reperfusion injury via heme oxygenase-1-mediated autophagy[J]. CritCare Med, 2014, 42(12):e762-e771. DOI:10.1097/CCM. 0000000000000659. [4] CZAJA MJ, DING WX, DONOHUE TM, et al. Functions of autophagy in normal and diseased liver[J]. Autophagy, 2013, 9(8):1131-1158. DOI: 10.4161/auto.25063. [5] WANG C, CHEN K, XIA Y, et al. N-acetylcysteine attenuates ischemia-reperfusion-induced apoptosis and autophagy in mouse liver via regulation of the ROS/JNK/Bcl-2 pathway[J]. PLoS One, 2014, 9(9):e108855. DOI: 10.1371/journal.pone.0108855. [6] JEAN-CHARLES PY, KAUR S, SHENOY SK. G protein-coupled receptor signaling through β-arrestin-dependent mechanisms[J]. J Cardiovasc Pharmacol, 2017, 70(3):142-158. DOI:10.1097/FJC. 0000000000000482. [7] WEN Y, HE J, XUE X, et al. β-arrestin2 inhibits apoptosis and liver inflamation induced by ischemia-reperfusion in mice via Akt and TLR4 pathway[J]. Arch Med Res, 2019, 50(7):413-422. DOI: 10.1016/j.arcmed.2019.10.012. [8] CARNEIRO AP, FONSECA-ALANIZ MH, DALLAN LAO, et al. β-arrestin is critical for early shear stress-induced Akt/eNOS activation in human vascular endothelial cells[J]. Biochem Biophys Res Commun, 2017, 483(1):75-81. DOI: 10.1016/j.bbrc.2017.01.003. [9] HERAS-SANDOVAL D, PÉREZ-ROJAS JM, HERNÁNDEZ-DAMIÁN J, et al. The role of PI3K/Akt/mTOR pathway in the modulation of autophagy and the clearance of protein aggregates in neurodegeneration[J]. Cell Signal, 2014, 26(12):2694-2701. DOI: 10.1016/j.cellsig.2014.08.019. [10] DECUYPERE JP, PARYS JB, BULTYNCK G. Regulation of the autophagic bcl-2/beclin 1 interaction[J]. Cells, 2012, 1(3):284-312. DOI: 10.3390/cells1030284. [11] 王励, 汪根树. β-arrestin-2通过抑制自噬减轻小鼠肝脏缺血-再灌注损伤[J].器官移植, 2015, 6(3): 139-145, 156. DOI: 10.3969/j.issn.1674-7445.2015.03.003.WANG L, WANG GS. β-arrestin-2 alleviates mouse hepatic ischemia-reperfusion injury by inhibiting autophagy[J]. Organ Transplant, 2015, 6(3): 139-145, 156.DOI: 10.3969/j.issn.1674-7445.2015.03.003. [12] RAO J, YUE S, FU Y, et al. ATF6 mediates apro-inflammatory synergy between ER stress and TLR activation in the pathogenesis of liver ischemia-reperfusion injury[J]. Am J Transplant, 2014, 14(7):1552-1561. DOI: 10.1111/ajt.12711. [13] LUEDDE T, KAPLOWITZ N, SCHWABE RF. Cell death and cell death responses in liver disease: mechanisms and clinical relevance[J]. Gastroenterology, 2014, 147(4):765-783. DOI: 10.1053/j.gastro.2014.07.018. [14] YANG M, ANTOINE DJ, WEEMHOFF JL, et al. Biomarkers distinguish apoptotic and necrotic cell death during hepatic ischemia/reperfusion injury in mice[J]. Liver Transpl, 2014, 20(11):1372-1382. DOI: 10.1002/lt.23958. [15] DREFS M, THOMAS MN, GUBA M, et al. Modulation of glutathione hemostasis by inhibition of 12/15-lipoxygenase prevents ROS-mediated cell death after hepatic ischemia and reperfusion[J]. Oxid Med Cell Longev, 2017:8325754. DOI: 10.1155/2017/8325754. [16] JADHAV PV, KOTHAKOTA SR, SASIDHARAN M, et al. Effect of donor hepatic steatosis on ischemia reperfusion injury in liver transplant recipient[J]. J Clin Exp Hepatol, 2020, 10(3):236-244. DOI: 10.1016/j.jceh.2019.07.004. [17] WANG Y, SHEN J, XIONG X, et al. Remote ischemic preconditioning protects against liver ischemia-reperfusion injury via heme oxygenase-1-induced autophagy[J]. PLoS One, 2014, 9(6):e98834. DOI: 10.1371/journal.pone.0098834. [18] WANG JH, BEHRNS KE, LEEUWENBURGH C, et al. Critical role of autophage in ischemia/reperfusion injury to aged livers[J]. Autophagy, 2012, 8(1):140-141. DOI: 10.4161/auto.8.1.18391. [19] WANG W, WU L, LI J, et al. Alleviation of hepatic ischemia reperfusion injury by oleanolic acid pretreating via reducing HMGB1 release and inhibiting apoptosis and autophagy[J]. Mediators Inflamm, 2019:3240713. DOI: 10.1155/2019/3240713. [20] SHEN M, LU J, DAI W, et al. Ethyl pyruvate ameliorates hepatic ischemia-reperfusion injury by inhibiting intrinsic pathway of apoptosis and autophagy[J]. Mediators Inflamm, 2013:461536. DOI: 10.1155/2013/461536. [21] CHEN Z, LI T, KAREEM K, et al. The role of PI3K/Akt signaling pathway in non-physiological shear stress-induced platelet activation[J]. Artif Organs, 2019, 43(9):897-908. DOI: 10.1111/aor.13465. [22] ALYAHYA AM, AL-MASRI A, HERSI A, et al. The effects of progranulin in a rat model of acute myocardial ischemia/reperfusion are mediated by activation of the P13K/Akt signaling pathway[J]. Med Sci Monit Basic Res, 2019, 25:229-237. DOI: 10.12659/MSMBR.916258. [23] LI H, XU W, MA Y, et al. Milk fat globule membrane protein promotes C2C12 cell proliferation through the PI3K/Akt signaling pathway[J]. Int J Biol Macromol, 2018, 114:1305-1314. DOI: 10.1016/j.ijbiomac.2018.04.026. [24] ZHU JF, HUANG W, YI HM, et al. Annexin A1-suppressed autophagy promotes nasopharyngeal carcinoma cell invasion and metastasis by PI3K/Akt signaling activation[J]. Cell Death Dis, 2018, 9(12):1154. DOI: 10.1038/s41419-018-1204-7. [25] LIN Q, WANG Y, CHEN D, et al. Cisplatin regulates cell autophagy in endometrial cancer cells via the PI3K/Akt/mTOR signalling pathway[J]. Oncol Lett, 2017, 13(5):3567-3571. DOI: 10.3892/ol.2017.5894. -
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