乳源六肽预防和降低小鼠急性酒精性肝损伤及其机制

Experimental study on prevention and reduction of alcoholic liver injury in mice by priming hexapeptide

目的研究乳源六肽(PGPIPN)在降低小鼠酒精性肝损伤方面的作用及其机制。方法建立小鼠急性酒精肝损伤的动物模型,健康雄性昆明种小鼠60只,体质量18~22 g,动物预饲养1周后分成6组:对照组、模型组、PGPIPN L组、PGPIPN M组、PGPIPN H组和谷胱甘肽(GSH)组,每组10只小鼠。为了制作小鼠急性酒精性肝损伤的动物模型,实验最后3 d,PGPIPN低、中、高剂量组、GSH组和模型对照组以56°红星二锅头酒灌胃,对照组使用等量蒸馏水灌胃,建立小鼠的急性酒精性肝损伤的对照模型。12 h后,将小鼠颈椎脱臼处死,取材检测相关指标。在实验结束时,将小鼠称重并麻醉,收集血清和肝脏样品,取肝脏称重后保存,计算出肝脏指数。同时,测定小鼠血清中天冬氨酸氨基转移酶(AST)和丙氨酸氨基转移酶(ALT)含量,血清和肝匀浆中三酰甘油(TG)和总胆固醇(TC)的含量。取肝脏组织制作成病理切片,观察肝脏细胞的病理学变化。原代细胞培养,流式细胞术检测肝脏组织的凋亡状况。结果生化分析显示模型组小鼠血清TG、TC和肝脏TG(肝匀浆中TG的浓度)高于对照组,PGPIPN H剂量组、PGPIPN M剂量组和PGPIPN L剂量组血清中ALT、AST含量与模型组相比降低,PGPIPN高剂量组中TNF-α含量较模型组也有减少。对照组中,肝细胞结构完整,凋亡指数为3.61%。而模型组中肝索结构紊乱,液泡数目增加,肝脏细胞的边界不清晰,细胞核消失,细胞皱缩,凋亡指数为36.87%,出现了明显的细胞凋亡状态。PGPIPN L组细胞出现凋亡状态,而PGPIPN H组细胞状态良好,无凋亡现象发生。流式细胞术检测细胞凋亡,对照组为4.558%,模型组为31.50%。结论PGPIPN能够预防和降低小鼠急性酒精性肝损伤。

Objective To study the role and mechanism of immunomodulating peptide(PGPIPN)in reducing alcoholic liver injury in mice.Methods Sixty healthy male Kunming mice weighing 18~22 g.Animals were pre-fed for one week and then divided into 6 groups(control,model,PGPIPN low,PGPIPN mediun,PGPIPN high and GSH)with 10 mice per group.In order to make an animal model of acute alcoholic liver in mice,in the last 3 days of the experiment,PGPIPN low,medium and high dose groups,GSH group and model control group were intragastrically administered with 56 degree red star Erguotou wine,and the control group was given the same amount of distilled water.A mice model of acute alcoholic liver injury was established.After 12 h,the mice were sacrificed by cervical dislocation and the relevant indicators were taken.At the end of the experiment,the mice were weighed and anesthetized,serum and liver samples were collected,weighed(liver)and stored,calculate the size of the liver index at the same time.At the same time,serum aspartate aminotransferase(AST)and alanine aminotransferase(ALT)levels,serum and liver homogenate triglyceride(TG)and total cholesterol(TC)content were determined.Liver tissue was taken to make pathological sections,and changes in cytopathology were observed.In primary cell culture experiment,flow cytometry was used to detect the apoptosis of liver tissue.Results Biochemical analysis showed that serum TG,TC and liver TG(concentration of TG in liver homogenate)were higher in the model group than that in control group,and the contents of ALT and AST in serum of PGPIPN high dose group,PGPIPN middle dose group and PGPIPN low dose group were lower than those in the model group,and the content of tumor necrosis factorα(TNF-α)in PGPIPN high dose group was also lower than that in the model group.In the control group,the hepatocyte structure was intact and the apoptosis index was 3.61%.In the model group,the liver cable structure was disordered,the number of vacuoles was increased,the boundary of liver cells was not clear,the nucleus disappeared,the cells collapsed,and the apoptosis index was 36.87%,showing obvious apoptosis.The cells in the PGPIPN low dose group showed apoptosis,while PGPIPN high dose improved apoptosis significantly.Cell apoptosis was detected by flow cytometry,4.558%in the control group and 31.50%in the model group.Conclusion The PGPIPN can prevent and reduce acute alcoholic liver injury in mice.