当归多糖通过抑制GRP78、p-PERK、p-Eif2α表达改善糖尿病KK-Ay小鼠肝内质网应激

Angelica polysaccharides improve hepatic endoplasmic reticulum stress by inhibiting the expression of GRP78,p-PERK and p-Eif2αin diabetic KK-Ay mice

目的:探讨当归多糖对糖尿病KK-Ay小鼠肝内质网应激的调节机制。方法:将40只糖尿病KK-Ay小鼠随机分为模型组、二甲双胍组、当归多糖高、中、低剂量组,每组8只。C57BL/6J小鼠8只作为空白对照组。当归多糖高、中、低剂量组分别灌胃当归多糖400、200、100 mg/kg,二甲双胍组灌胃盐酸二甲双胍200 mg/kg,正常组和模型组灌胃等体积生理盐水,每周测小鼠空腹血糖和体质量。灌胃4周后,测量小鼠血清中甘油三酯(TG)、胆固醇(TC)、低密度脂蛋白胆固醇(LDL-C)、高密度脂蛋白胆固醇(HDL-C)含量;RT-PCR观察肝组织中葡萄糖调节蛋白78(GRP78)、磷酸化胰腺内质网激酶(p-PERK)、磷酸化α亚基的真核起始因子2(p-Eif2α)的mRNA表达;Westernblot、免疫组化检测小鼠肝组织中GRP78、p-PERK、p-Eif2α的蛋白表达。HE染色观察肝脏组织病理学变化。结果:与空白组相比,模型组TC、TG、LDL-C的水平显著升高(P<0.01),HDL-C的水平明显降低(P<0.01);与模型组相比,二甲双胍组,当归多糖高、中剂量组TC、TG、LDL-C的水平显著降低(P<0.05,P<0.01),HDL-C的水平明显升高(P<0.05,P<0.01);与空白组相比,模型组小鼠GRP78、p-PERK、p-Eif2α的表达显著上调(P<0.01),与模型组相比,当归多糖高、中、低剂量组GRP78、p-PERK、p-Eif2α的表达显著下调(P<0.05,P<0.01),且高剂量组效果最佳;与模型组相比,当归多糖组小鼠肝组织致密,空泡样变性减少,肝脏脂肪化降低,肝细胞逐渐排列整齐,肝窦结构趋于清晰,且效果呈剂量依赖性。结论:当归多糖可显著改善糖尿病KK-Ay小鼠肝损伤,其作用机制可能与当归多糖抑制内质网应激相关蛋白和因子GRP78、p-PERK、p-Eif2α的表达,改善内质网应激有关。

AIM:To investigate the regulatory mechanism of Angelica polysaccharide on hepatic endoplasmic reticulum stress in diabetic KK-Ay mice.MEHTODS:Forty diabetic KK-Ay mice were randomly divided into model group,metformin group,and angelica polysaccharide high,medium,and low dose groups,with 8 mice in each group.8 C57BL/6J mice were used as blank control group.The mice were gavaged with 400 mg/kg,200 mg/kg and 100 mg/kg of angelica polysaccharide in the high,medium and low dose groups,respectively,and 200 mg/kg of metformin hydrochloride in the metformin group,while the normal and model groups were gavaged with equal volume of saline,and fasting blood glucose and body weight were measured weekly.After 4 weeks of gavage,triglyceride(TG),cholesterol(TC),low-density lipoprotein cholesterol(LDL-C)and high-density lipoprotein cholesterol(HDL-C)levels were measured in mice serum;RT-PCR was performed to observe the expression of glucose-regulated protein 78(GRP78),phosphorylated pancreatic endoplasmic reticulum kinase(p-PERK)and phosphorylatedα-subunit eukaryotic initiation factor 2(p-Eif2α)in liver tissues.mRNA expression;Western blot,immunohistochemistry to detect the protein expression of GRP78,p-PERK,p-Eif2αin mouse liver tissues.HE staining:to observe the histopathological changes in the liver.RESULT:Compared with the blank group,the levels of TC,TG and LDL-C were significantly increased(P<0.01)and the levels of HDL-C were significantly decreased(P<0.01)in the model group;compared with the model group,the levels of TC,TG and LDL-C were significantly decreased(P<0.05,P<0.01)and the levels of HDL-C were significantly increased in the metformin group,angelica polysaccharide high and medium dose groups.Compared with the blank group,the expression of GRP78,p-PERK and p-Eif2αin the model group was significantly upregulated(P<0.01),and the expression of GRP78,p-PERK and p-Eif2αin the angelica polysaccharide high,medium and low dose groups was significantly downregulated(P<0.05,P<0.01),and the high dose group had the best effect compared with the model group.Compared with the model group,the mice in the angelica polysaccharide group showed dense liver tissue,reduced vacuole-like degeneration,reduced liver steatosis,gradually aligned hepatocytes,and clear hepatic sinusoidal structure,and the effect was dose-dependent.CONCLUSION:Angelica polysaccharide significantly improved liver injury in diabetic KK-Ay mice,and its mechanism of action may be related to the inhibition of endoplasmic reticulum stress-related proteins and factors GRP78,p-PERK and p-Eif2αexpression by Angelica polysaccharide and improvement of endoplasmic reticulum stress.