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目的 探讨二氢杨梅素对高糖诱导血管内皮细胞凋亡的影响及其与磷酸腺苷活化蛋白激酶(AMP-activated protein kinase,AMPK)的关系。方法体外培养原代人脐带静脉内皮细胞(humanumbilicalvein endothelial cells,HUVECs),分为正常对照组(葡萄糖浓度5.56 mmol/L)、高糖处理组(葡萄糖浓度33.36 mmol/L)、甘露醇高渗对照组(5.56 mmol/L葡萄糖+27.8 mmol/L甘露醇)、AMPK激动剂5-氨基咪唑-4-甲酰胺-1-β-D-呋喃核糖苷(5-Aminoimidazole-4-carboxamide-1-β-D-ribofuranoside,AICAR)(10nmol/L)+高糖处理组、二氢杨梅素(1μmol/L)+高糖处理组、AMPK抑制剂compound C(5mmol/L)+AICAR+高糖处理组和AMPK抑制剂compound C(5mmol/L)+二氢杨梅素+高糖处理组。处理36 h后,CCK-8法检测细胞增殖活力,Annexin V-FITC/PI双标记流式细胞术检测细胞凋亡,Western blot法检测AMPK、p-AMPK、乙酰辅酶A羧化酶(Acetyl-Co A carboxylase,ACC)和p-ACC蛋白水平。结果与正常对照组比较,高糖处理组细胞活力明显降低(P<0.05),细胞凋亡率显著增加(P<0.05),p-AMPK、p-ACC水平显著降低(P<0.05);二氢杨梅素或AICAR预处理均明显抑制高糖诱导的HUVECs细胞活力下降、细胞凋亡增加和p-AMPK、p-ACC水平降低(P<0.05);Compound C预处理明显抑制AICAR和二氢杨梅素对高糖诱导HUVECs细胞凋亡的保护作用(P<0.05)。结论 二氢杨梅素通过促进AMPK活化抑制高糖诱导的血管内皮细胞凋亡。
Objective To investigate the effect of dihydromyricetin on apoptosis of vascular endothelial cells induced by high glucose and its relationship with AMP-activated protein kinase (AMPK). Methods Primary human umbilical vein endothelial cells (HUVECs) were cultured in vitro and divided into normal control group (glucose concentration 5.56 mmol / L), high glucose treatment group (glucose concentration 33.36 mmol / L), mannitol hypertonic control Group (5.56 mmol / L glucose + 27.8 mmol / L mannitol), AMPK agonist 5-Aminoimidazole-4-carboxamide-1- (10 mmol / L) + high glucose treatment group, 1 μmol / L glycerophosphate treatment group and 5 mmol / L AMPA inhibitor + AICAR + high glucose treatment group AMPK inhibitor compound C (5mmol / L) + dihydromyricetin + high glucose treatment group. Cell viability was measured by CCK-8 assay after 36 h of treatment. Apoptosis was detected by Annexin V-FITC / PI double-labeled flow cytometry. AMPK, p-AMPK and Acetyl- Co A carboxylase, ACC) and p-ACC protein levels. Results Compared with the normal control group, the viability of cells in high glucose treatment group was significantly decreased (P <0.05), the apoptosis rate was significantly increased (P <0.05), the levels of p-AMPK and p-ACC were significantly decreased Hydromobendin or AICAR pretreatment significantly inhibited cell viability, apoptosis and P-AMPK and p-ACC levels in HUVECs induced by high glucose (P <0.05). Compound C pretreatment significantly inhibited AICAR and dihydromyricetin (P <0.05) on the apoptosis of HUVECs induced by high glucose. Conclusion Dihydromyricetin inhibits high glucose-induced apoptosis of vascular endothelial cells by promoting AMPK activation.