基于视网膜MEG3表达变化探讨活血通络方对db/db小鼠视网膜病变的保护作用  

Protective Effect of Huoxue Tongluo Decoction(活血通络方)on Retinopathy in db/db Mice Based on Changes of MEG3 Expression in Retina Tissue

作　　者：刘晓丹 詹晓峰 李能[2] LIU Xiaodan;ZHAN Xiaofeng;LI Neng(Traditional Chinese Osteopathy&Traumatology Hospital of Fuyang,Hangzhou,Zhejiang 311400;Hangzhou Hospital of Chinese Medicine,Hangzhou,Zhejiang 310007)

机构地区：[1]杭州市富阳中医骨伤医院,浙江杭州311400 [2]杭州市中医院,浙江杭州310007

出　　处：《中国中医药科技》2025年第3期404-409,共6页Chinese Journal of Traditional Medical Science and Technology

基　　金：浙江省自然科学基金(LGF21H120001)。

摘　　要：目的:基于视网膜母系表达基因3(MEG3)表达的变化探究活血通络方对db/db小鼠视网膜损伤的保护作用及机制。方法:db/db小鼠随机分为模型组、活血通络方组、空载组、MEG3敲减组、MEG3敲减+活血通络方组,应用重组腺病毒载体shRNA-LncRNA MEG3敲减小鼠视网膜MEG3表达,并予活血通络方(15.4 g/kg)干预8周。qRT-PCR检测小鼠视网膜MEG3 mRNA表达;EB染色观察血-视网膜屏障(blood retina barrier,BRB)损伤,HE染色观察视网膜病理改变;ELISA测定视网膜TNF-α、IL-1β、MDA水平和SOD活性;WB测定视网膜还原型烟酰胺腺嘌呤二核苷酸磷酸氧化酶4(NOX4)、鸟苷酸交换因子3(VAV3)、磷酸化-磷脂酰肌醇-3-激酶(p-PI3K)、磷脂酰肌醇-3-激酶(PI3K)、磷酸化-蛋白激酶B(p-AKT)、蛋白激酶B(Akt)、磷酸化哺乳动物雷帕霉素(p-mTOR)、哺乳动物雷帕霉素(mTOR)蛋白表达。结果:敲减MEG3表达后,db/db小鼠BRB损伤加重,视网膜上皮细胞数量减少,TNF-α、IL-1β、MDA水平上升(P<0.01)、SOD活性降低(P<0.01),NOX4蛋白、VAV2蛋白、PI3K/AKT/mTOR磷酸化蛋白表达增加(P<0.01)。活血通络方干预组小鼠视网膜MEG3表达上调(P<0.01),BRB损伤改善,TNF-α、IL-1β、MDA水平下降(P<0.01),SOD活性增加(P<0.01),NOX4、VAV2、p-PI3K、PI3K/AKT/mTOR蛋白表达降低(P<0.01)。结论:敲减MEG3表达加重小鼠视网膜炎症和BRB损伤,活血通络方可通过上调MEG3表达减轻db/db小鼠视网膜病变。Objective:To investigate the protective effect and mechanism of Huoxue Tongluo Decoction(HTD)on retina injury of db/db mice mediated by MEG3.Methods:db/db mice were randomly divided into model group,HTD group,empty group,MEG3 knockdown group,and MEG3 knockdown+HTD group,shRNA-LncRNA MEG3 was used to knockdown MEG3 expression of retina.After 8 weeks of HTD intervention,qRT-PCR was used to detect the expression of MEG3 mRNA expression in retina tissue;EB staining was use detect injury of blood retina barrier(BRB),HE staining was used to observe retinal epithelial pathological changes;ELISA was used to detect TNF-α,IL-1β,MDA levels and SOD activity in retina tissue;Western blot was used to detect NOX4,VAV2,p-PI3K,PI3K/AKT/mTOR protein expressions in retina tissue.Results:Knockdown of MEG3 increased the BRB injury,decreased number of epithelial cells,activated the levels of TNF-α,IL-1β,and MDA(P<0.01),decreased SOD activity(P<0.01),and increased NOX4,VAV2,p-PI3K,PI3K/AKT/mTOR protein expressions(P<0.01)in retina tissues of db/db mice.Intervention of HTD increased MEG3 mRNA expression(P<0.01),alleviated BRB injury,decreased TNF-α,IL-1βand MDA levels(P<0.01),increased SOD activity(P<0.01),decreased NOX4,VAV2,p-PI3K,PI3K/AKT/mTOR protein expressions(P<0.01)in retina tissues.Conclusion:Knockdown of MEG3 expression promots db/db mice retina inflammation and BRB injury,HTD protect db/db mice from retinal injury by up-regulating the expression of MEG3.

关 键 词：活血通络方 糖尿病视网膜病变 MEG3 PI3K/AKT/MTOR 炎性因子 氧化应激 DB/DB小鼠 

分 类 号：R285.5[医药卫生—中药学]