| ghrelin-GHSR-1a对高糖环境下视网膜血管内皮细胞的保护作用 |
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| 引用本文： | 李蓉,姚国敏,周凌霄,张敏,闫瑾.ghrelin-GHSR-1a对高糖环境下视网膜血管内皮细胞的保护作用[J].眼科新进展,2022,0(3):205-209. |
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| 作者姓名： | 李蓉 姚国敏 周凌霄 张敏 闫瑾 |
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| 作者单位： | 710077 陕西省西安市，西安医学院第一附属医院眼科（李蓉，姚国敏，周凌霄）；710077 西安医学院第一附属医院内分泌科（张敏）；710021 陕西省西安市，西安医学院医学技术学院眼视光教研室（闫瑾） |
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| 摘 要： | 目的 观察ghrelin及其受体生长激素分泌素受体1a（GHSR-1a）在高糖环境下对视网膜血管内皮细胞的保护作用。方法 将体外培养的人视网膜血管内皮细胞（HRMECs）分为对照组和高浓度葡萄糖组。对照组细胞在含5.5 mmol·L-1葡萄糖的M199培养基中培养48 h，高浓度葡萄糖组细胞在含30.0 mmol·L-1葡萄糖的M199培养基中培养48 h，采用免疫荧光染色法检测两组细胞中GHSR-1a的表达。然后另取HRMECs随机分为5组:正常对照（NC）组、高糖（HG）组、HG+ghrelin组、HG+ghrelin+siGHSR-1a组和HG+ghrelin+NC-siGHSR-1a组，孵育48 h。NC组处理同对照组，HG组处理同高浓度葡萄糖组，HG+ghrelin组细胞在含30.0 mmol·L-1葡萄糖、10.0 nmol·L-1 ghrelin的M199培养基中培养，HG+ghrelin+siGHSR-1a组细胞先转染GHSR-1a-特异性siRNA，24 h后在含30.0 mmol·L-1葡萄糖和10.0 nmol·L-1 ghrelin的M199培养基中培养，HG+ghrelin+NC-siGHSR-1a组细胞先转染非特异性序列，24 h后在含30.0 mmol·L-1葡萄糖和10.0 nmol·L-1 ghrelin的M199培养基中培养。分别采用CCK-8和Annexin-V/FITC-PI凋亡检测试剂盒检测各组HRMECs细胞活力和细胞凋亡情况。Western blot检测凋亡相关蛋白Bax和Bcl-2的表达。结果 高浓度葡萄糖组的GHSR-1a表达低于对照组（P<0.05）。NC组、HG组、HG+ghrelin组、HG+ghrelin+siGHSR-1a组和HG+ghrelin+NC-siGHSR-1a组的细胞活力分别为1.00、69.87%±0.68%、92.31%±3.62%、75.98%±4.67%和90.87%±1.95%，总体比较差异有统计学意义（P<0.001）；两两比较显示，除HG+ghrelin组与HG+ghrelin+NC-siGHSR-1a组的细胞活力差异无统计学意义（P>0.05）外，其余各组细胞活力比较差异均有统计学意义（均为P<0.05）。NC组、HG组、HG+ghrelin组、HG+ghrelin+siGHSR-1a组和HG+ghrelin+NC-siGHSR-1a组的细胞凋亡率分别为5.76%±0.36%、20.53%±0.38%、10.84%±0.52%、14.93%±0.18%和11.03%±0.62%，总体比较差异有统计学意义（F=468.88，P<0.001）；两两比较显示，除HG+ghrelin组与HG+ghrelin+NC-siGHSR-1a组细胞凋亡率差异无统计学意义（P>0.05）外，其余各组细胞凋亡率比较差异均有统计学意义（均为P<0.05）。NC组、HG组、HG+ghrelin组、HG+ghrelin+siGHSR-1a组和HG+ghrelin+NC-siGHSR-1a组细胞的Bax蛋白和Bcl-2蛋白相对表达量分别为0.17±0.03和0.64±0.05，0.75±0.04和0.14±0.02，0.38±0.04和0.49±0.08，0.56±0.04和0.33±0.07，0.40±0.06和0.46±0.09，总体比较差异均有统计学意义（均为P<0.001）；两两比较显示，除HG+ghrelin组与HG+ghrelin+NC-siGHSR-1a组Bax蛋白和Bcl-2蛋白表达差异均无统计学意义（均为P>0.05）外，其余各组Bax蛋白和Bcl-2蛋白表达比较差异均有统计学意义（均为P<0.05）。结论 ghrelin-GHSR-1a系统对高糖环境下的HRMECs功能具有保护作用，并抑制高糖诱导的HRMECs凋亡。
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| 关 键 词： | ghrelin 生长激素分泌素受体1a 糖尿病视网膜病变 高糖 视网膜血管内皮细胞 |
Protective effects of ghrelin and growth hormone secretin receptor 1a on retinal microvascular endothelial cells under high glucose conditions |
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| LI Rong,YAO Guomin,ZHOU Lingxiao,ZHANG Min,YAN Jin.Protective effects of ghrelin and growth hormone secretin receptor 1a on retinal microvascular endothelial cells under high glucose conditions[J].Recent Advances in Ophthalmology,2022,0(3):205-209. |
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| Authors: | LI Rong YAO Guomin ZHOU Lingxiao ZHANG Min YAN Jin |
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| Affiliation: | 1.Department of Ophthalmology,the First Affiliated Hospital of Xi’an Medical University,Xi’an 710077,Shaanxi Province,China2.Department of Endocrinology,the First Affiliated Hospital of Xi’an Medical University,Xi’an 710077,Shaanxi Province,China3.Department of Optometry Teaching and Research Section,College of Medical Technology,Xi’an Medical University,Xi’an 710021,Shaanxi Province,China |
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| Abstract: | Objective To investigate the protective effects of ghrelin and its receptor, the growth hormone secretin receptor 1a (GHSR-1a), on retinal microvascular endothelial cells under high glucose conditions.Methods Human retinal microvascular endothelial cells (HRMECs) cultured in vitro were divided into the control and high glucose groups. Cells in the control group were cultured in M199 medium containing 5.5 mmol·L-1 glucose for 48 h, while cells in the high glucose group were cultured in M199 medium containing 30.0 mmol·L-1 glucose for 48 h. The expression of GHSR-1a in each group was detected by immunofluorescence staining. Additional HRMECs were randomly divided into five groups and incubated for 48 h: normal control (NC) group, high glucose (HG) group, HG+ghrelin group, HG+ghrelin+siGHSR-1a group, and HG+ghrelin+NC-siGHSR-1a group. Cells in the NC group were treated the same as the control group, and cells in the HG group were treated the same as the high glucose group. Cells in the HG+ghrelin group were cultured in M199 medium containing 30.0 mmol·L-1 glucose and 10.0 nmol·L-1 ghrelin. Cells in the HG+ghrelin+siGHSR-1a group were first transfected with GHSR-1a-specific siRNA, and 24 h later, cultured in M199 medium containing 30.0 mmol·L-1 glucose and 10.0 nmol·L-1 ghrelin. Cells in the HG+ghrelin+NC-siGHSR-1a group were transfected with non-specific sequence, and 24 h later, cultured in M199 medium containing 30.0 mmol·L-1 glucose and 10.0 nmol·L-1 ghrelin. The viability and apoptosis of HRMECs were detected by CCK-8 and Annexin-V/FITC-PI kit, respectively. Western blot was used to detect the expression of apoptosis-related proteins Bax and Bcl-2.Results The expression of GHSR-1a in the high glucose group was lower than that in the control group (P<0.05). The cell viability in the NC, HG, HG+ghrelin, HG+ghrelin+siGHSR-1a, and HG+ghrelin+NC-siGHSR-1a groups were 1.00, (69.87±0.68) %, (92.31±3.62) %, (75.98±4.67) %, and (90.87±1.95) %, respectively, and the overall difference was statistically significant (P<0.001). The cell viability showed no statistical difference between the HG+ghrelin group and HG+ghrelin+NC-siGHSR-1a group (P>0.05), but in the remaining groups, the cell viability showed significant statistical difference (all P<0.05). The cell apoptosis in the NC, HG, HG+ghrelin, HG+ghrelin+siGHSR-1a, and HG+ghrelin+NC-siGHSR-1a groups were (5.76±0.36)%, (20.53±0.38) %, (10.84±0.52) %, (14.93±0.18) %, and (11.03±0.62) %, respectively, and the overall difference was statistically significant (F=468.88, P<0.001). The cell apoptosis showed no statistical difference between the HG+ghrelin group and HG+ghrelin+NC-siGHSR-1a group (P>0.05), but in the remaining groups, the cell apoptosis showed significant statistical difference (all P<0.05). The relative expression levels of Bax in the NC, HG, HG+ghrelin, HG+ghrelin+siGHSR-1a, and HG+ghrelin+NC-siGHSR-1a groups were 0.17±0.03, 0.75±0.04, 0.38±0.04, 0.56±0.04, and 0.40±0.06, respectively, while the relative expression levels of Bcl-2 in the NC, HG, HG+ghrelin, HG+ghrelin+siGHSR-1a, and HG+ghrelin+NC-siGHSR-1a groups were 0.64±0.05, 0.14±0.02, 0.49±0.08, 0.33±0.07, and 0.46±0.09, respectively, and the overall difference was statistically significant (both P<0.001). The expression levels of Bax protein and Bcl-2 protein showed no statistical difference between the HG+ghrelin group and HG+ghrelin+NC-siGHSR-1a group (both P>0.05), but in the remaining groups, the expression levels of Bax protein and Bcl-2 protein showed significant statistical difference (all P<0.05).Conclusion Ghrelin-GHSR-1a system can protect the function of HRMECs under high glucose conditions and inhibit their apoptosis induced by high glucose. |
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| Keywords: | ghrelin growth hormone secretin receptor 1a diabetic retinopathy high glucose retinal microvascular endothelial cells |
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