基于PERK信号通路介导的细胞凋亡及杯状细胞破坏探讨芍药汤对溃疡性结肠炎大鼠的作用机制

The mechanism of Shaoyao decoction on UC rats based on cell apoptosis and goblet cell destruction mediated by PERK signaling pathway

目的:通过检测溃疡性结肠炎(UC)大鼠p-PERK通路蛋白、C/EBP同源蛋白的表达及结肠杯状细胞数量,探讨芍药汤对UC大鼠的作用机制。方法:84只SD大鼠中随机抽取14只作为空白组,余下70只大鼠通过三硝基苯磺酸建立UC大鼠模型,除去死亡大鼠,随机分为模型组12只、柳氮磺吡啶组13只、芍药汤低剂量组12只、芍药汤中剂量组13只、芍药汤高剂量组13只。空白组、模型组用等体积的0.9%氯化钠溶液灌胃,柳氮磺吡啶组采用柳氮磺吡啶灌胃,芍药汤高、中、低剂量组按照芍药汤不同剂量灌胃。第14天进行疾病活动指数(DAI)评分。随后将大鼠处死并解剖,进行结肠黏膜损伤指数(CMDI)评分,并检测大鼠结肠p-PERK通路蛋白、C/EBP同源蛋白的表达及结肠杯状细胞数量。结果:与空白组比较,柳氮磺吡啶组及芍药汤不同剂量组大鼠DAI、CMDI评分均增高(P<0.05),与模型组比较,柳氮磺吡啶组和芍药汤高、中、低剂量组DAI、CMDA评分均降低(P<0.05),柳氮磺吡啶组与芍药汤高、中、低剂量组DAI、CMDA评分比较无统计学差异(P>0.05);与空白组比较,模型组、柳氮磺吡啶组和芍药汤高、中、低剂量组p-PERK、CHOP蛋白表达均上升(P<0.05);与模型组比较,柳氮磺吡啶组和芍药汤高、中、低剂量组p-PERK、CHOP蛋白表达均下降(P<0.05);与芍药汤低剂量组比较,柳氮磺吡啶组和芍药汤高、中剂量组p-PERK、CHOP蛋白表达均下降(P<0.05);柳氮磺吡啶组和芍药汤高、中剂量组p-PERK、CHOP蛋白表达无统计学差异(P>0.05);与空白组比较,其他各组杯状细胞数量均下降(P<0.05);与模型组比较,柳氮磺吡啶组和芍药汤高、中、低剂量组杯状细胞数量均明显增加(P<0.05);与芍药汤低剂量组比较,柳氮磺吡啶组和芍药汤高、中剂量组杯状细胞数量均增加(P<0.05);柳氮磺吡啶组和芍药汤高、中剂量组杯状细胞数量比较无统计学差异(P>0.05)。结论:芍药汤能抑制内质网应激PERK信号通路被激活,降低CHOP表达活性,减少细胞凋亡,抑制杯状细胞被破坏,保护肠黏膜屏障,改善UC大鼠的症状和体征,这可能是芍药汤治疗UC的作用机制。

Objective:Through the detection the expression of p-PERK pathway protein,C/EBP homologous protein and the number of colon goblet cells in ulcerative colitis(UC)rats,investigate the mechanism of Shaoyao decoction in UC rats.Methods:The 84 SD rats were randomly selected as blank group,and the remaining 70 rats were model as UC rats by trinitrophenylsulfonic acid.Excluding the dead rats,it was randomly divided into 12 rats in the model group,13 rats in the sulfasalazine group,12 rats in the low-dose group of Shaoyao decoction,13 rats in the medium-dose group of Shaoyao decoction,and 13 rats in the high-dose group of Shaoyao decoction.Blank and model groups were infused with equal volume of saline,SASP with sulfasalazine,and high,medium and low doses with different doses of Shaoyao decoction.The Disease Activity Index(DAI)score was performed on day 14.Rats were subsequently sacrificed and dissected for a Colonic Mucal Injury Index(CMDI)score.The expression of rat colonic p-PERK pathway protein,C/EBP homologs and the number of colonic goblet cells were examined.Results:Compared with the model group,the general situation of rats in the SASP group and different doses of Shaoyao decoction group has improved significantly.In contrast to the blank group,DAI and CMDI scores increased in model,SASP,high,medium and low-dose Shaoyao decoction groups(P<0.05).Compared with the model group,the DAI and CMDI scores were lower in the SASP and Shaoyao decoction high,medium and low-dose groups(P<0.05).There was no significant difference in DAI and CMDI scores between the SASP and Shaoyao decoction high,medium and low-dose groups(P>0.05).Relative expression of rat p-PERK and CHOP proteins:in contrast to the blank group.The expression of p-PERK and CHOP protein increased in model,SASP,high,medium and low-dose groups of Shaoyao decoction(P<0.05).Compared to the model group.the expression of p-PERK and CHOP protein all decreased in SASP group,high,medium and low dose of Shaoyao decoction group(P<0.05).Compared with the Shaoyao decoction low-dose group,the expression of p-PERK and CHOP protein all decreased in SASP,high-dose Shaoyao decoction and medium-dose groups(P<0.05).No differences of p-PERK and CHOP protein expression between SASP,Shaoyao decoction high and medium-dose groups(P>0.05).Goblet cells number:compared with the blank group,the number of goblet cells decreased in the other groups(P<0.05).Compared with the model group,the number of goblet cells increased significantly in the SASP group and Shaoyao decoction group with high,medium and low-dose(P<0.05).Compared with the low-dose group of Shaoyao decoction,the number of goblet cells increased in SASP,Shaoyao decoction high and medium-dose groups(P<0.05).There was no significant difference in the number of goblet cells decreased in SASP,Shaoyao decoction high and medium-dose(P>0.05).Conclusion:Shaoyao decoction can improve the symptoms and signs of UC rats by inhibiting the activation of PERK signaling in ER stress,reducing CHOP expression activity and cell apoptosis,inhibiting goblet cell destruction,then protect intestinal mucosal barrier,which may be the mechanism of Shaoyao decoction in treating UC.