促肝细胞生长素对大鼠肝缺血再灌注损伤的保护作用及其机制探讨
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摘要
目的探讨促肝细胞生长素(PHGF)对大鼠肝缺血再灌注损伤的保护作用及机制。方法将80只健康雄性SD大鼠随机分为实验组(PHGF组)和对照组(NS组),每组各40只。通过阻断肝中叶及肝左叶血流造成70%肝脏缺血建立大鼠肝缺血再灌注损伤模型,缺血时间为21 min。PHGF组术前予PHGF腹腔注射干预,NS组术前给予等量生理盐水。术前及术后1、3、5、7 d取大鼠血清及肝组织,检测ALT、AST、TBil水平; HE染色观察病理改变; real-time PCR检测肝组织线粒体转录因子A(TFAM) mRNA水平。计量资料两组间比较采用独立样本t检验。结果 HE染色结果显示,光镜下PHGF组肝细胞水肿、炎细胞浸润程度及肝细胞坏死程度较NS组明显减轻。肝功能生化检测结果显示,PHGF组大鼠术后1、3、5、7 d血清ALT、AST、TBil水平低于同时相NS组,差异均有统计学意义(t值分别为11. 879、16. 019、22. 168、10. 235、9. 041、12. 936、18. 759、8. 142、10. 108、11. 014、13. 245、9. 968,P值均<0. 001)。real-time PCR结果显示,PHGF组大鼠术后1、3、5 d肝组织TFAM mRNA水平较NS组明显升高(t值分别为7. 998、14. 764、13. 861,P值均<0. 001)。结论 PHGF预处理对大鼠肝缺血再灌注损伤具有保护作用,其作用机制可能是通过上调TFAM表达减轻肝缺血再灌注损伤。
Objective To investigate the protective effect of hepatocyte growth-promoting factor( PHGF) against liver ischemia-reperfusion injury in rats and its mechanism of its action. Methods A total of 80 healthy male Sprague-Dawley rats were randomly divided into experimental group( PHGF group) and control group( NS group),with 40 rats in each group. A rat model of liver ischemia-reperfusion injury was established by 70% liver ischemia caused by the occlusion of blood flow in the middle and left lobes of the liver,with an ischemia time of 21 minutes. The rats in the PHGF group were given intraperitoneal injection of PHGF for intervention before surgery,and those in the NS group were given an equal volume of normal saline. Serum and liver tissue samples were collected before surgery and on days 1,3,5,and 7 after surgery,and the levels of alanine aminotransferase( ALT),aspartate aminotransferase( AST),and total bilirubin( TBil) were measured; HE staining was used to observe pathological changes; real-time PCR was used to measure the mRNA expression of mitochondrial transcription factor A( TFAM) in the liver. The independent samples t-test was used for comparison of continuous data between two groups. Results HE staining showed that compared with the NS group,the PHGF group had significantly lower degrees of hepatocyte swelling,inflammatory cell infiltration,and hepatocyte necrosis under a light microscope. Liver biochemistry showed that on days 1,3,5,and 7 after surgery,the PHGF group had significantly lower serum levels of ALT,AST,and TBil than the NS group( t = 11. 879,16. 019,22. 168,10. 235,9. 041,12. 936,18. 759,8. 142,10. 108,11. 014,13. 245,and 9. 968,all P < 0. 001). Real-time PCR showed that on days 1,3,and 5 after surgery,the PHGF group had a significantly higher mRNA level of TFAM in the liver than the NS group( t =7. 998,14. 764,and 13. 861,all P < 0. 001). Conclusion PHGF preconditioning exerts a protective effect against liver ischemia-reperfusion injury in rats,possibly by upregulating the expression of TFAM to alleviate liver ischemia-reperfusion injury.
Objective To investigate the protective effect of hepatocyte growth-promoting factor( PHGF) against liver ischemia-reperfusion injury in rats and its mechanism of its action. Methods A total of 80 healthy male Sprague-Dawley rats were randomly divided into experimental group( PHGF group) and control group( NS group),with 40 rats in each group. A rat model of liver ischemia-reperfusion injury was established by 70% liver ischemia caused by the occlusion of blood flow in the middle and left lobes of the liver,with an ischemia time of 21 minutes. The rats in the PHGF group were given intraperitoneal injection of PHGF for intervention before surgery,and those in the NS group were given an equal volume of normal saline. Serum and liver tissue samples were collected before surgery and on days 1,3,5,and 7 after surgery,and the levels of alanine aminotransferase( ALT),aspartate aminotransferase( AST),and total bilirubin( TBil) were measured; HE staining was used to observe pathological changes; real-time PCR was used to measure the mRNA expression of mitochondrial transcription factor A( TFAM) in the liver. The independent samples t-test was used for comparison of continuous data between two groups. Results HE staining showed that compared with the NS group,the PHGF group had significantly lower degrees of hepatocyte swelling,inflammatory cell infiltration,and hepatocyte necrosis under a light microscope. Liver biochemistry showed that on days 1,3,5,and 7 after surgery,the PHGF group had significantly lower serum levels of ALT,AST,and TBil than the NS group( t = 11. 879,16. 019,22. 168,10. 235,9. 041,12. 936,18. 759,8. 142,10. 108,11. 014,13. 245,and 9. 968,all P < 0. 001). Real-time PCR showed that on days 1,3,and 5 after surgery,the PHGF group had a significantly higher mRNA level of TFAM in the liver than the NS group( t =7. 998,14. 764,and 13. 861,all P < 0. 001). Conclusion PHGF preconditioning exerts a protective effect against liver ischemia-reperfusion injury in rats,possibly by upregulating the expression of TFAM to alleviate liver ischemia-reperfusion injury.
引文
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[13]HEBA SE DEEN,HEMMAT EE HADDAD.Significance of hepatocyte growth factor concentrations in serum of patients with liver cirrhosis and patients with hepatocellular carcinoma[J].Egypt J Intern Med,2015,3(27):92-102.
[14]CAO XF,JIN SZ,SUN L,et al.Therapeutic effects of hepatocyte growth factor-overexpressing dental pulp stem cells on liver cirrhosis in a rat model[J].Sci Rep,2017,7(1):15812.
[15]CHEN BH,LI WM.Effect of ulinastatin combined with ischemic preconditioning on hepatic ischemia-reperfusion injury in rats[J].J Clin Hepatol,2018,34(2):368-372.(in Chinese)陈宝鹤,李文美.乌司他丁联合缺血预处理对大鼠肝缺血再灌注损伤的影响[J].临床肝胆病杂志,2018,34(2):368-372.
[16]FALKENBERG M,LARSSON NG,GUSTAFSSON CM.DNAreplication and transcription in mammalian mitochondria[J].Annu Rev Biochem,2007,76:679-699.
[17]NGO HB,LOVELY GA,PHILLIPS R,et al.Distinct structural features of TFAM drive mitochondrial DNA packaging versus transcriptional activation[J].Nat Commun,2014,5:3077.
[18]CAMPBELL CT,KOLESAR JE,KAUFMAN BA.Mitochondrial transcription factor A regulates mitochondrial transcription initiation,DNA packaging,and genome copy number[J].Biochim Biophys Acta,2012,1819(9-10):921-929.
[2]SAAT TC,van den ENGEL S,BIJMAN-LACHGER W,et al.Fate and effect of intravenously infused mesenchymal stem cells in a mouse model of hepatic ischemia reperfusion injury and resection[J].Stem Cells Int,2016,2016:5761487.
[3]TSUBOUCHI H.Discovery and clinical application of hepatocyte growth factor[J].Nihon Shokakibyo Gakkai Zasshi,2013,110(12):2033-2041.
[4]OU HL,MA L.Hepatocyte growth promoting factor combined with antiviral drug in the treatment of chronic hepatitis B and its effect on liver function,viral replication and fibrosis index[J].Biomed Res,2017,28(13):5764-5767.
[5]KANG I,CHU CT,KAUFMAN BA.The mitochondrial transcription factor TFAM in neurodegeneration:Emerging evidence and mechanisms[J].FEBS Lett,2018,592(5):793-811.
[6]FRANCO DG,MORETTI IF,MARIE SKN.Mitochondria transcription factor A:A putative target for the effect of melatonin on U87MG malignant glioma cell line[J].Molecules,2018,23(5):1129.
[7]JULIAN MW,SHAO G,BAO S,et al.Mitochondrial transcription factor A serves as a danger signal by augmenting plasmacytoid dendritic cell responses to DNA[J].J Immunol,2012,189(1):433-443.
[8]CALATAYUD D,SNCHEZ CABU'S S,SAMPSON J,et al.Hepatic resection:A safe and effective surgery[J].Cir Esp,2017,95(8):437-446.
[9]SCATTON O,ZALINSKI S,JEGOU D,et al.Randomized clinical trial of ischaemic preconditioning in major liver resection with intermittent Pringle manoeuvre[J].Br J Surg,2011,98(9):1236-1243.
[10]FU SY,LAU WY,LI GG,et al.A prospective randomized controlled trial to compare Pringle maneuver,hemihepatic vascular inflow occlusion,and main portal vein inflow occlusion in partial hepatectomy[J].Am J Surg,2011,201(1):62-69.
[11]RICHARDSON AJ,LAURENCE JM,LAM VW.Portal triad clamping versus other methods of vascular control in liver resection:Asystematic review and meta-analysis[J].HPB(Oxford),2012,14(6):355-364.
[12]YAN LP,LING S,LI H,et al.Effect of hepatocyte growthpromoting factor on acute lung injury induced by paraquat poisoning in rats[J].China Med Herald,2017,14(12):29-32.(in Chinese)颜丽萍,凌顺,李弘,等.促肝细胞生长素对百草枯中毒引起大鼠急性肺损伤的影响[J].中国医药导报,2017,14(12):29-32.
[13]HEBA SE DEEN,HEMMAT EE HADDAD.Significance of hepatocyte growth factor concentrations in serum of patients with liver cirrhosis and patients with hepatocellular carcinoma[J].Egypt J Intern Med,2015,3(27):92-102.
[14]CAO XF,JIN SZ,SUN L,et al.Therapeutic effects of hepatocyte growth factor-overexpressing dental pulp stem cells on liver cirrhosis in a rat model[J].Sci Rep,2017,7(1):15812.
[15]CHEN BH,LI WM.Effect of ulinastatin combined with ischemic preconditioning on hepatic ischemia-reperfusion injury in rats[J].J Clin Hepatol,2018,34(2):368-372.(in Chinese)陈宝鹤,李文美.乌司他丁联合缺血预处理对大鼠肝缺血再灌注损伤的影响[J].临床肝胆病杂志,2018,34(2):368-372.
[16]FALKENBERG M,LARSSON NG,GUSTAFSSON CM.DNAreplication and transcription in mammalian mitochondria[J].Annu Rev Biochem,2007,76:679-699.
[17]NGO HB,LOVELY GA,PHILLIPS R,et al.Distinct structural features of TFAM drive mitochondrial DNA packaging versus transcriptional activation[J].Nat Commun,2014,5:3077.
[18]CAMPBELL CT,KOLESAR JE,KAUFMAN BA.Mitochondrial transcription factor A regulates mitochondrial transcription initiation,DNA packaging,and genome copy number[J].Biochim Biophys Acta,2012,1819(9-10):921-929.