GnT-V通过影响自噬调控糖尿病小鼠的心肌损伤  

GnT-V modulates myocardial injury in diabetic mice by affecting autophagy

作　　者：赵然 王婷婷 赵杰琼 李妍 ZHAO Ran;WANG Tingting;ZHAO Jieqiong;LI Yan(Department of Cardiology,Second Affiliated Hospital,Air Force Medical University,Xi′an 710068,China)

机构地区：[1]空军军医大学第二附属医院心血管内科,西安710068

出　　处：《山西医科大学学报》2024年第7期842-848,共7页Journal of Shanxi Medical University

基　　金：国家自然科学基金资助项目(82200404);唐都医院社会人才基金项目(2021SHRC017)。

摘　　要：目的探究N-乙酰氨基葡萄糖转移酶V(N-acetylglucosaminetransferase-V,GnT-V)对糖尿病心肌病小鼠心肌损伤的影响及潜在分子机制。方法30只6~8周龄雄性C57BL/6小鼠,随机分为对照组(Con组)、糖尿病心肌病组(DCM组)和糖尿病心肌病+干扰GnT-V基因的腺相关病毒组(DCM+AAV-shGnT-V组),每组10只。DCM组小鼠腹腔注射50 mg/kg链脲佐菌素,连续5 d,诱导糖尿病模型(随机血糖≥16.7 mmol/L),继续普通饮食喂养12周诱导糖尿病心肌病模型;对照组小鼠腹腔注射相应剂量的的柠檬酸盐缓冲液;DCM+AAV-shGnT-V组在糖尿病模型诱导成功后4周,通过心肌注射40μL滴度为1×10^(10)PFU/mL的腺相关病毒(AAV)-shGnT-V以敲低GnT-V基因。超声心动图检测各组小鼠心脏功能,Masson染色观察各组小鼠心肌纤维化情况,TUNEL染色检测各组小鼠心肌细胞凋亡程度,免疫组化染色观察各组心肌组织中GnT-V和Beclin1的表达情况,Western blot法检测自噬蛋白LC3、P62及PI3K/AKT通路的蛋白表达情况。结果与Con组相比,DCM组小鼠心脏收缩功能显著降低(P<0.05),心肌组织纤维化程度明显增加(P<0.05),心肌细胞的凋亡率明显上升(P<0.05),心肌组织中GnT-V、自噬蛋白Beclin1、LC3Ⅱ/LC3Ⅰ表达显著增加,P62的表达明显减少(P<0.05),且PI3K和AKT磷酸化程度明显降低(P<0.05)。与DCM组相比,DCM+AAV-shGnT-V组小鼠心脏收缩功能明显改善(P<0.05),心肌组织纤维化程度显著下降(P<0.05),心肌细胞的凋亡率明显降低(P<0.05),GnT-V、Beclin1及LC3Ⅱ/LC3Ⅰ表达显著下调,P62的表达明显增加(P<0.05),且PI3K和AKT磷酸化程度明显增强(P<0.05)。结论敲低GnT-V基因可以显著改善糖尿病心肌病小鼠的心肌组织纤维化进展和心肌细胞凋亡程度,其作用可能与激活PI3K/AKT信号通路,进而抑制心肌自噬有关。Objective To investigate the effect of N-acetylglucosaminetransferase-V on myocardial injury in diabetic cardiomyopathy(DCM)mice and its potential molecular mechanism.Methods Thirty 6-8-week-old male C57BL/6 mice were randomly divided into control group(Con group),DCM group,and DCM+adeno-associated virus carrying GnT-V-specific small hairpin RNA(DCM+AAV-shGnT-V group),with 10 mice in each group.The mice in DCM group were injected intraperitoneally with 50 mg/kg streptozotocin for 5 d to induce a diabetes model(randomized blood glucose≥16.7 mmol/L),and then continued to be fed with universal diet for 12 weeks to induce a diabetic cardiomyopathy model.The mice in Con group were intraperitoneally injected with the corresponding dose of citrate buffer.The mice in DCM+AAV-shGnT-V group were given 40μL of AAV-shGnT-V with a titer of 1×10^(10) PFU/mL by myocardial injection 4 weeks after the successful induction in diabetes model to knock down the GnT-V gene.Echocardiography was used to detect the cardiac function,Masson staining was used to observe the myocardial fibrosis,TUNEL staining was used to detect the apoptosis,immunohistochemical staining was used to observe the expressions of GnT-V and Beclin1 in myocardial tissues,and Western blot was used to detect the expressions of autophagy-related proteins LC3,P62,and PI3K/AKT pathway-related proteins.Results Compared to Con group,the cardiac contractile function was significantly reduced in DCM group(P<0.05),the fibrosis of myocardial tissue was significantly increased(P<0.05),the apoptosis rate of cardiomyocytes was significantly increased(P<0.05),the expressions of GnT-V,Beclin1,and LC3Ⅱ/LC3Ⅰwere significantly increased in myocardial tissue,the expression of P62 was significantly reduced(P<0.05),and PI3K and AKT phosphorylation levels were significantly reduced(P<0.05).Compared to DCM group,the cardiac contractile function was significantly improved in DCM+AAV-shGnT-V group(P<0.05),the myocardial tissue fibrosis was significantly decreased(P<0.05),the apoptos

关 键 词：糖尿病心肌病 自噬 凋亡 心肌纤维化 GNT-V PI3K/AKT 

分 类 号：R587.2[医药卫生—内分泌]