Department of Cardiology, The Second Affiliated Hospital of Xi'an Medical University, Xi'an, P.R. China.
Experimental Center, The Second Affiliated Hospital of Xi'an Medical University, Xi'an, P.R. China.
Bioengineered. 2022 Jan;13(1):1476-1490. doi: 10.1080/21655979.2021.2019874.
To investigate the role of hypoxia-inducible factor 1-alpha (HIF1A) in hypoxia/reoxygenation (H/R) injury of cardiomyocytes induced by high glucose (HG). The in vitro model of coronary heart disease with diabetes was that H9c2 cells were stimulated by H/R and HG. Quantitative reverse transcription PCR (RT-qPCR) and Western blot analysis were used to detect the expression of HIF1A and angiopoietin-like protein 2 (ANGPTL2) in H9c2 cells. Cell viability and apoptosis were, respectively, estimated by Cell Counting Kit 8 (CCK-8) and TUNEL assays. Lactate dehydrogenase (LDH) activity, inflammation and oxidative stress were in turn detected by their commercial assay kits. Luciferase reporter assay and chromatin immunoprecipitation (ChIP) assay were used to confirm the association between HIF1A and ANGPTL2 promoter. The expression of nuclear factor E2-related factor 2 (Nrf2)/heme oxygenase 1 (HO-1) pathway-related proteins and apoptosis-related proteins were also detected by Western blot analysis. As a result, ANGPTL2 expression was upregulated in H9c2 cells induced by HG or/and H/R. ANGPTL2 positively modulated HIF1A expression in H9c2 cells. HG or/and H/R suppressed the cell viability and promoted apoptosis, inflammatory response and oxidative stress levels in H9c2 cells. However, the knockdown of ANGPTL2 could reverse the above phenomena in H/R-stimulated-H9c2 cells through activation of Nrf2/HO-1 pathway. HIF1A transcriptionally activated ANGPTL2 expression. The effect of knockdown of ANGPTL2 on H/R triggered-H9c2 cells was weakened by HIF1A overexpression. In conclusion, knockdown of HIF1A downregulated ANGPTL2 to alleviate H/R injury in HG-induced H9c2 cells by activating the Nrf2/HO-1 pathway.
为了研究低氧诱导因子 1 ɑ（HIF1A）在高糖（HG）诱导的心肌细胞缺氧/复氧（H/R）损伤中的作用。采用体外糖尿病冠心病模型，用 H/R 和 HG 刺激 H9c2 细胞。采用定量逆转录 PCR（RT-qPCR）和 Western blot 分析检测 H9c2 细胞中 HIF1A 和血管生成素样蛋白 2（ANGPTL2）的表达。分别用细胞计数试剂盒（CCK-8）和 TUNEL 测定法检测细胞活力和细胞凋亡。用其商业测定试剂盒依次检测乳酸脱氢酶（LDH）活性、炎症和氧化应激。用荧光素酶报告基因测定和染色质免疫沉淀（ChIP）测定证实 HIF1A 与 ANGPTL2 启动子之间的关联。用 Western blot 分析检测核因子 E2 相关因子 2（Nrf2）/血红素加氧酶 1（HO-1）通路相关蛋白和凋亡相关蛋白的表达。结果显示，HG 或/和 H/R 诱导的 H9c2 细胞中 ANGPTL2 表达上调。ANGPTL2 正向调节 H9c2 细胞中的 HIF1A 表达。HG 或/和 H/R 抑制 H9c2 细胞活力，促进细胞凋亡、炎症反应和氧化应激水平。然而，ANGPTL2 的敲低可通过激活 Nrf2/HO-1 通路逆转 H/R 刺激的 H9c2 细胞中的上述现象。HIF1A 转录激活 ANGPTL2 的表达。ANGPTL2 敲低对 H/R 触发 H9c2 细胞的影响因 HIF1A 过表达而减弱。结论：敲低 HIF1A 下调 ANGPTL2 通过激活 Nrf2/HO-1 通路减轻 HG 诱导的 H9c2 细胞 H/R 损伤。
