苦参碱对大鼠供肝冷保存再灌注损伤中肝细胞凋亡及调控基因表达的影响

Effects of matrine on apoptosis of hepatocytes and expression of regulating gene during cold preservation and reperfusion injury in rat donor liver

目的:探讨苦参碱对大鼠原位肝移植供肝冷保存再灌注损伤中肝细胞凋亡及调控基因表达的影响.方法:应用延长保存的大鼠原位肝移植模型,大鼠84只随机分为对照组、低剂量苦参碱治疗组(40mg/kg)、高剂量苦参碱治疗组(80mg/kg)和假手术组,将供肝在4℃林格液中保存5h后植入受体,各组大鼠于再灌注后4和24h后取样.采用TUNEL法分别检测移植术后肝细胞凋亡,流式细胞仪检测凋亡相关基因Bcl-2,FasL蛋白的表达,光镜下观察移植肝脏病理形态学的改变.结果:与对照组比较,苦参碱低、高剂量治疗组术后肝脏细胞凋亡指数显著降低(6.07±1.68,6.17±0.83vs14.87±2.10,P<0.01),肝组织Bcl-2表达增加(59.32±14.09,58.90±16.70vs17.00±8.01,P<0.01),但FasL表达量无显著差异(P>0.05),肝细胞凋亡指数、Bcl-2和FasL表达在苦参碱低、高剂量组间也无显著差异.对照组肝细胞损伤的病理表现相当严重,而治疗组的病理表现显著改善.结论:苦参碱通过促进抑制凋亡基因Bcl-2的表达来抑制冷保存再灌注导致的肝细胞凋亡.

AIM： To investigate the effect of Matrine on the apoptosis of hepatocytes and expression of regulating gene during cold preservation and reperfusion injury in rat orthotopic liver transplantation （OLT）. METHODS： Eighty-four syngeneic SD rats were randomly divided into control, Matrine, and pseudo-treatment group. The rats in Matrine group were treated with low （40 mg/kg） and high dose （80 mg/kg） of Matrine, respectively. After the donor liver was preserved in Ringer＇s （LR） solution for 5 h, the orthotopic implantation was performed. Four and 24 h after reperfusion of the portal vein, the serum and tissue samples were collected for analysis. The cell apoptosis was detected by TUNEL, and the expression of Fas-L and Bcl-2 protein were determined by flow cytometry. The histopathological changes were observed under light and electron microscope. RESULTS： In comparison with that in the control group, the apoptosis index was significantly decreased in low and high dose Matrine treatment group （6.07 ± 1.68, 6.17 ± 0.83 vs 14.87 ± 2.10, P 〈 0.01）, while the level of Bcl-2 expression was markedly elevated （59.32 ± 14.09, 58.90 ± 16.70 vs 17.00 ±8.01, P 〈 0.01）. The expression of FasL was not notably different between the control and Matrine group. The apoptosis index, Bcl-2 and FasL expression had no distinct difference between low and high dose treatment group either. The pathological changes of liver cells were severe in the control group, but they were obviously ameliorated in Matrine group. CONCLUSION： Matrine can protect hepatocyts against apoptosis by inhibiting the Bcl-2 expression during cold preservation and reperfusion injury.