Hu Yuxin, Xu Qian, Li Hongxue, Meng Ziyu, Hao Ming, Ma Xuefei, Lin Wenjian, Kuang Hongyu
The Department of Endocrinology, The First Affiliated Hospital of Harbin Medical University, Harbin, China.
Front Pharmacol. 2022 Feb 24;13:827896. doi: 10.3389/fphar.2022.827896. eCollection 2022.
It is known that the metabolic disorder caused by high glucose is one of pathogenesis in diabetic retinopathy (DR), the leading cause of blindness, due to the main pathological change of apoptosis of endothelial cells (ECs). In previous studies, the potential impact of sodium glucose cotransporter-2 (SGLT-2), whose inhibitors slow the progression of DR, has not been elucidated. The purpose of the presented study was to explore the effect of SGLT-2 inhibitors dapagliflozin (DAPA) on apoptosis of diabetic mice retina and human retinal microvascular endothelial cells (HRMECs), examine the effects of dapagliflozin on HRMECs metabolism, and explore the molecular processes that affect DR. The eyeballs of male streptozotocin (STZ)-induced diabetic C57BL/6N mice were evaluated. C57BL/6N mice were divided into control group (CON), diabetic untreated group (DM), diabetic dapagliflozin treatment group (DM + DAPA) and diabetic insulin treatment group (DM + INS). Hematoxylin-Eosin (HE) staining was performed to observe the pathological structure of the mice retina, and TUNEL staining to detect apoptosis of mice retinal cells. , DCFH-DA and western blot (WB) were used to evaluate ROS, Bcl-2, BAX, cleaved-caspase 3 in HRMECs and metabolomics detected the effect of dapagliflozin on the metabolism of HRMECs. And then, we performed correlation analysis and verification functions for significantly different metabolites. , dapagliflozin reduced the apoptosis of diabetic mice retina independently of hypoglycemic. , SGLT-2 protein was expressed on HRMECs. Dapagliflozin reduced the level of ROS caused by high glucose, decreased the expression of cleaved-caspase3 and the ratio of BAX/Bcl-2. Metabolomics results showed that dapagliflozin did not affect the intracellular glucose level. Compared with the high glucose group, dapagliflozin reduced the production of arachidonic acid (AA) and inhibited the phosphorylation of ERK1/2, therefore, reducing the phosphorylation of cPLA2, which is a key enzyme for arachidonic acid release. Collectively, results unearthed for the first time that dapagliflozin reduced apoptosis of retina induced by DM whether or . Dapagliflozin did not affect the glucose uptake while mitigated intracellular arachidonic acid in HRMECs. Dapagliflozin alleviated HRMECs apoptosis induced by high glucose through ERK/1/2/cPLA2/AA/ROS pathway.
已知高糖引起的代谢紊乱是糖尿病视网膜病变（DR）的发病机制之一，DR是导致失明的主要原因，其主要病理变化是内皮细胞（ECs）凋亡。在先前的研究中，尚未阐明钠-葡萄糖协同转运蛋白2（SGLT-2）的潜在影响，其抑制剂可减缓DR的进展。本研究的目的是探讨SGLT-2抑制剂达格列净（DAPA）对糖尿病小鼠视网膜和人视网膜微血管内皮细胞（HRMECs）凋亡的影响，研究达格列净对HRMECs代谢的作用，并探索影响DR的分子机制。对雄性链脲佐菌素（STZ）诱导的糖尿病C57BL/6N小鼠的眼球进行评估。将C57BL/6N小鼠分为对照组（CON）、糖尿病未治疗组（DM）、糖尿病达格列净治疗组（DM + DAPA）和糖尿病胰岛素治疗组（DM + INS）。进行苏木精-伊红（HE）染色以观察小鼠视网膜的病理结构，采用TUNEL染色检测小鼠视网膜细胞凋亡。采用2',7'-二氯二氢荧光素二乙酸酯（DCFH-DA）和蛋白质免疫印迹法（WB）评估HRMECs中的活性氧（ROS）、Bcl-2、BAX、裂解的半胱天冬酶3，并通过代谢组学检测达格列净对HRMECs代谢的影响。然后，对显著差异的代谢物进行相关性分析和验证功能。达格列净可独立于降糖作用减少糖尿病小鼠视网膜的凋亡。SGLT-2蛋白在HRMECs上表达。达格列净降低高糖引起的ROS水平，降低裂解的半胱天冬酶3的表达以及BAX/Bcl-2的比值。代谢组学结果表明，达格列净不影响细胞内葡萄糖水平。与高糖组相比，达格列净减少花生四烯酸（AA）的产生并抑制细胞外信号调节激酶1/2（ERK1/2）的磷酸化，从而减少花生四烯酸释放的关键酶——胞浆型磷脂酶A2（cPLA2）的磷酸化。总的来说，本研究首次发现达格列净可减少糖尿病诱导的视网膜凋亡。达格列净不影响葡萄糖摄取，但可减轻HRMECs中的细胞内花生四烯酸水平。达格列净通过ERK1/2/cPLA2/AA/ROS途径减轻高糖诱导的HRMECs凋亡。
