三七三醇皂苷通过PI3K/Akt通路减轻大鼠离体心脏缺血/再灌注损伤的研究

The study of panax notoginseng saponins through PI3K/Akt pathway to reduce ischemia/reperfusion injury in isolated rat hearts

目的探讨三七三醇皂苷(PTS)对心肌缺血/再灌注(I/R)大鼠心肌功能及磷酸肌醇—蛋白激酶B(PI3K/Akt)信号通路的影响。方法将40只雄性Wistar大鼠随机数字表法分为5组,每组8只。对照组、I/R组、PTS组、LY294002组、LY294002+PTS组,通过Langendorff离体心脏灌流装置建立心脏缺血/再灌注模型,收集不同时刻的灌流液测量心肌酶学指标(CK-MB、LDH),并记录不同时刻左心室血流动力学参数(HR、LVESD、LVEDD、LVDP、±dp/dtmax);Western-blot检测心肌组织中Akt、p-Akt、GSK-3β、p-GSK-3β的表达,免疫组化检测Bax、Bcl-2、Caspase-3的表达。结果(1)血流动力学参数:对照组HR不同时点无明显变化(P>0.05),其余各组均不同程度上升,且除对照组外其余4组在再灌注75 min时无明显差异(P>0.05)。对照组不同时点LVESP、LVEDP、LVDP、±dp/dt max无明显变化,其余各组随时间延长出现不同程度下降。再灌注75 min时,PTS组明显低于模型组(P<0.05),而LY294002+PTS组与模型组无明显差异,与PTS组差异显著(P<0.05)。(2)CK-MB、LDH:对照组不同时点无显著变化,其余各组再灌注后心肌酶水平均较缺血30 min有显著提升(P<0.05)。各组平衡20 min时无明显差异(P>0.05);局部缺血30 min时,各组CK-MB无统计学差异(P>0.05),而各组LDH较对照组有所升高(P<0.05);再灌注15min、75 min时,各组均较对照组明显升高,PTS组明显低于模型组,LY294002+PTS组高于PTS组(P均<0.05)。(3)Akt、pAkt、GSK-3β、pGSK-3β的表达:缺血再灌注损伤后,各组心肌组织Akt、GSK-3β表达无统计学差异(P>0.05),pAkt、pGSK在PTS组中表达较模型组有显著提高(P<0.05),在应用LY294002的2组中表达较模型组显著下降(P<0.05),且LY294002+PTS与PTS组差异存在统计学意义(P<0.05)。(4)Bax、Bcl-2、Caspase-3的表达:模型组Bax、Caspase-3显著高于对照组(P<0.05),PTS与LY+PTS 2组Bax、Caspase-3表达低于模型组(P<0.05)。对照组Bcl-2强阳性,模型组显著低于对照组(P<0.05),PTS组较模型组显著提高(P<0.05),LY294002+PTS组显著低于PTS组(P<0.05)。结论PTS可改善心肌I/R大鼠离体心脏功能,可能与其调节PI3K/Akt信号通路有关。

Objective To explore the effect of notoginseng triol saponins(PTS)on myocardial function and PI3K/Akt signaling pathway in myocardial ischemia/reperfusion(I/R)rats.Methods Forty male Wistar rats were randomly divided into 5 groups with 8 rats in each group.In the control group,I/R group,PTS group,LY294002 group,LY294002+PTS group,the Langendorff isolated cardiac perfusion device was used to establish a cardiac ischemia/reperfusion model,and the perfusion fluid at different times was collected to measure myocardial enzyme indexes(CK-MB,LDH),and record left ventricular hemodynamic parameters(HR,LVESD,LVEDD,LVDP,±dp/dtmax)at different moments;Western blot detection of Akt,pAkt,GSK-3β,pGSK-3βexpression in myocardial tissue,immune Histochemical detection of Bax,Bcl-2,Caspase-3 expression.Results(1)Hemodynamic parameters:the HR of the control group did not change significantly at different time points(P>0.05),the other groups increased to varying degrees,and the remaining four groups except the control group had no significant difference at 75 minutes of reperfusion(P>0.05).There was no significant change in LVESP,LVEDP,LVDP,±dp/dt max in the control group at different time points,and the other groups decreased to varying degrees over time.At 75 minutes of reperfusion,the PTS group was significantly lower than the model group(P<0.05),while the LY294002+PTS group was not significantly different from the model group,but was significantly different from the PTS group(P<0.05).(2)CK-MB,LDH:The control group did not change significantly at different time points,and the myocardial enzyme levels in the other groups were significantly increased after reperfusion compared with 30 min ischemia(P<0.05).There was no significant difference between the groups at 20 min of equilibrium(P>0.05);at 30 min of ischemia,there was no statistical difference between the CK-MB groups(P>0.05),while the LDH groups were higher than the control group(P<0.05);at 15min and 75min of reperfusion,all groups were significantly higher than the control group,the PTS group was significantly lower than the model group,and the LY294002+PTS group was higher than the PTS group(all P<0.05).(3)Expression of Akt,pAkt,GSK-3β,and pGSK-3β:After ischemia-reperfusion injury,there was no statistical difference in the expression of Akt and GSK-3βin myocardial tissue of each group(P>0.05).pAkt and pGSK were expressed in the PTS group Compared with the model group,there was a significant increase(P<0.05).The expression in the two groups using LY294002 was significantly lower than that in the model group(P<0.05),and the difference between LY294002+PTS and PTS group was statistically significant(P<0.05).(4)The expression of Bax,Bcl-2 and Caspase-3:Bax and Caspase-3 in the model group were significantly higher than those in the control group(P<0.05),and the expression of Bax and Caspase-3 in the PTS and LY+PTS 2 groups was lower than that in the model group(P<0.05).The control group was Bcl-2 strongly positive,the model group was significantly lower than the control group(P<0.05),the PTS group was significantly higher than the model group(P<0.05),and the LY294002+PTS group was significantly lower than the PTS group(P<0.05).Conclusion Panax notoginseng saponins can improve isolated heart function damage in myocardial ischemia-reperfusion rats,which may be related to the regulation of PI3K/Akt signaling pathway.