目的 观察降钙素基因相关肽(CGRP)预处理对高糖孵育H9c2细胞缺氧/复氧(A/R)损伤的影响.方法 采用大鼠的成肌细胞H9c2细胞株作为研究对象,细胞在含10%胎牛血清DMEM/F12完全培养基中培养(条件为37 ℃,5% CO2),2~3 d传代1次.细胞进行缺氧3 h复氧3 h作为损伤模型.实验细胞随机分为五组,每组6孔细胞:①正常糖对照组(NC组);②高糖对照组(HC组);③高糖缺氧复氧组(HAR组);④高糖缺氧/复氧+CGRP组(HCP组);⑤高糖缺氧/复氧+CGRP+CGRP8-37组(HCCP组).缺氧/复氧处理后测定细胞凋亡率、培养液中乳酸脱氢酶(LDH)含量以及细胞匀浆中SOD含量.结果 与NC组比较,高糖环境HC组的细胞凋亡率、培养液中LDH显著升高,细胞匀浆SOD含量显著降低(P<0.01);缺氧复氧后HAR组的细胞凋亡率、培养液中LDH含量均显著高于HC组,细胞匀浆SOD含量显著低于HC组(P<0.01);与HAR组相比,HCP组细胞凋亡率、培养液上清中LDH含量较低,SOD含量较高(P<0.01);给予CGRP1受体阻断剂后HCCP组细胞凋亡率、培养液中LDH含量均显著高于HCP组,细胞匀浆SOD含量显著低于HCP组(P<0.01).结论 高糖环境及缺氧/复氧过程均可造成H9c2细胞损伤和凋亡,CGRP预处理可部分减轻H9c2细胞在缺氧/复氧过程中的损伤,该作用可能由CGRP受体介导.
Objective To investigate the effect of calcitonin gene-related peptide (CGRP)pretreatment on anoxia/rexogenation-induced injury of H9c2 cell cultured in high-glucose culture medium.Methods H9c2 cells was cultured in DMEM/F12 culture with low-glucose and 10% fetal bovine serum.For establishing anoxia-rexogenation(AR) injury model of cardiac muscle cells,rat cardiac myoblasts (H9c2 cells) were exposed to 3 hours of anoxia followed by 3 hours of reoxygenation.Cultured H9c2 cells were divided randomly into five groups(n=6): Normal glucose-cultured control group(group NC),which was cultured in low-glucose culture for 48 h;High glucose-cultured control group(group HC),which was cultured in high-glucose culture for 48 h;High glucose-cultured AR group(group HAR),which was exposed to 3 hours of anoxia followed by 3 hours of reoxygenation(AR) after cultured in high-glucose culture for 48 h;High glucose-cultured pretreatment group(group HCP),which was pretreatmented by CGRP(final concentration for 10-8 mol/L) for 30 min followed by AR;High glucose-cultured CGRP and CGRP8-37 pretreatment group(Group HCCP),which was pretreatmented by CGRP 8-37 by 10 min followed by CGRP (final concentration for 10-7 mol/L) pretreatment for 30 min and then suffer AR progress.Results Compared with group NC,all cells of other groups had significant lower SOD activity and higher levels of LDH and apoptosis ratio of H9c2 cell(P<0.01).Compared to group HC,cells of group HAR and HNAR had significant decreased SOD activity of the cells and higher released LDH in cultured supernatant and apoptosis ratio of H9c2 cell(P<0.01).Compared with group HAR,cells of group HCP had significantly high SOD activity and decreased levels of the LDH and apoptosis ratio of H9c2 cell(P<0.01);Compared with group HCP,cells of group HCCP had significant decreased SOD activity of the cells and higher released LDH in cultured supernatant and apoptosis ratio of H9c2 cell(P<0.01).Conclusions It is suggested that CGRP pretreatment has direct role for decreaseing the AR-induced cell injury cultured in higher glucose medium,which is mostly associated with the activation of CGRP receptor.