石斛碱通过miR-499-5p调控糖尿病肾病系膜细胞增殖、上皮间质转化和纤维化  

Dendrobiine regulates mesangial cell proliferation,epithelial mesangial transformation and fibrosis in diabetic nephropathy through miR-499-5p

作　　者：陈玉凤 孔亚坤 李江雁 毛小芳 周艳红 CHEN Yu-feng;KONG Ya-kun;LI Jiang-yan;MAO Xiao-fang;ZHOU Yan-hong(Department of Endocrinology,Xinxiang Central Hospital(Fourth Clinical College,Xinxiang Medical College),Xinxiang 453000,Henan Province,China)

机构地区：[1]新乡市中心医院(新乡医学院第四临床学院)内分泌科,河南新乡453000

出　　处：《中国临床药理学杂志》2023年第2期211-215,共5页The Chinese Journal of Clinical Pharmacology

摘　　要：目的 探讨石斛碱(Den)对高糖致人肾小球系膜细胞(HGMCs)增殖、上皮间质转化(EMT)和纤维化的影响及其机制。方法 将体外培养的HGMCs分为正常组、模型组和低、中、高剂量实验组。正常组以DMEM培养基培养,模型组以高糖DMEM培养基培养,低、中、高剂量实验组分别以含0.05、0.50、5.00μg·mL^(-1)Den的高糖DMEM培养基培养;将anti-miR-NC、anti-miR-499-5p分别转染至高剂量实验组细胞,标记为高剂量实验组+anti-miR-NC组、高剂量实验组+anti-miR-499-5p组。以细胞计数试剂盒-8(CCK-8)法检测细胞增殖率;以实时荧光定量聚合酶链反应(qRT-PCR)法检测细胞中miR-499-5p表达情况;以蛋白质印迹(Western blot)法检测细胞中EMT、纤维化和核因子-κB(NF-κB)通路相关蛋白的表达情况。结果 正常组、模型组、低、中、高剂量实验组细胞中miR-499-5p表达水平分别为1.00±0.08,0.32±0.06,0.47±0.04,0.59±0.08和0.81±0.05,正常组与模型组比较,低、中、高剂量实验组与模型组比较,差异均有统计学意义(均P<0.05)。正常组、模型组、高剂量实验组、高剂量实验组+anti-miR-NC组、高剂量实验组+anti-miR-499-5p组细胞增殖率分别为(100.00±6.85)%,(159.02±10.60)%,(118.41±10.63)%、(119.30±6.88)%和(149.46±14.20)%;细胞中上皮性钙黏附素(E-cadherin)蛋白表达水分别为0.96±0.06,0.35±0.03,0.86±0.05,0.84±0.04和0.43±0.06;I型胶原酶(Col-1)蛋白表达水平分别为0.28±0.03,0.94±0.11,0.33±0.02,0.38±0.04和0.78±0.10;NF-κB蛋白表达水平分别为0.37±0.03,1.18±0.10,0.53±0.04、0.50±0.05和0.91±0.08。上述指标,模型组与正常组比较,高剂量实验组与模型组比较,高剂量实验组+anti-miR-499-5p组与高剂量实验组+anti-miR-NC组比较,差异均有统计学意义(均P<0.05)。结论 Den可通过上调miR-499-5p表达改善糖尿病肾病系膜细胞增殖、EMT和纤维化,其作用机制可能与抑制NF-κB通路活化有关。Objective To investigate the effects of dendrobiine(Den) on proliferation, epithelial mesangial transformation(EMT) and fibrosis of human glomerular mesangial cells(HGMCs) induced by high glucose and its mechanism. Methods The HGMCs cultured in vitro were divided into normal group, model group, experimental-L group, experimental-M group and experimental-H group. The normal group was cultured with DMEM medium;the model group was cultured with high glucose DMEM medium;the experimental-L,-M,-H groups were cultured in high-glucose DMEM medium containing 0. 05,0. 50 and 5. 00 μg · m L-1Den,respectively;anti-miR-NC and anti-miR-499-5p were transfected into experimental-H group cells,which were marked as experimental-H group + anti-miR-NC group and experimental-H group + anti-miR-499-5p group. Cell counting kit( CCK-8) method was used to detect cell proliferation rate;real-time fluorescence quantitative polymerase chain reaction( qRT-PCR) method was used to the expression of miR-499-5p in cells was detected;Western blot was used to detect the expression of EMT,fibrosis and nuclear factor-κB( NF-κB) pathway-related proteins in cells. Results The expression levels of miR-499-5p in normal group,model group and experimental-L,-M,-H groups were 1. 00 ± 0. 08,0. 32 ± 0. 06,0. 47 ± 0. 04,0. 59 ± 0. 08 and 0. 81 ± 0. 05,respectively. There were statistically significant differences in the above indexes between model group and control group or between experimental-L,-M,-H groups and model group( all P < 0. 05). The cell proliferation rates of normal group,model group,experimental-H group,experimental-H group+ anti-miR-NC group,experimental-H group + anti-miR-499-5p group were( 100. 00 ± 6. 85) %,( 159. 02 ± 10. 60) %,( 118. 41 ± 10. 63) %,( 119. 30 ± 6. 88) % and( 149. 46 ± 14. 20) %,respectively;E-cadherin protein expressions in cells were 0. 96 ± 0. 06,0. 35 ± 0. 03,0. 86 ± 0. 05,0. 84 ± 0. 04 and 0. 43 ± 0. 06,respectively;collagenase I( Col-1) protein expression levels were 0. 28 ± 0. 03,0. 94 ± 0. 11,0.

关 键 词：石斛碱 微小RNA-499-5p 糖尿病肾病 肾小球系膜细胞 细胞增殖 上皮间质转化 纤维化 

分 类 号：R97[医药卫生—药品]