摘要
目的:分析中性粒细胞明胶酶相关脂钙蛋白受体(NGALR)干扰对高糖诱导HK-2细胞的活性和上皮间质细胞转分化(EMT)的影响及其机制。方法:将HK-2细胞分成对照组(正常培养的HK-2细胞)、模型组(用30mM高浓度的葡萄糖诱导HK-2细胞24h)、si-NGALR NC组(先用30mM高浓度的葡萄糖诱导HK-2细胞24h,然后再转染si-NGALR NC质粒4h)、si-NGALR组(先用30mM高浓度的葡萄糖诱导HK-2细胞24h,然后再转染NGALR干扰质粒4h)、PDTC组(先用30mM高浓度的葡萄糖诱导HK-2细胞24h,然后加入100μM NF-κB抑制剂PDTC)、PDTC+si-NGALR组(先用30mM高浓度的葡萄糖诱导HK-2细胞24h,然后加入100μM NF-κB抑制剂PDTC作用1h,再转染NGALR干扰质粒4h)。各组细胞均在37℃、5%CO2的环境中培养。CCK8法检测细胞增殖,qRT-PCR检测NGALR mRNA,流式细胞术检测细胞凋亡,Western Blot检测NGALR、NGAL蛋白和NF-κB通路蛋白(NF-κB p65和p-NF-κB p65)及EMT相关蛋白(Snail和Vimentin)表达。结果:与对照组相比,模型组细胞增殖能力显著下降(P<0.01),凋亡率上升(P<0.01),NGALR、NGAL、Snail、Vimentin及NF-κB p65磷酸化的蛋白表达均上升(P<0.01)。与模型组相比,si-NGALR组和PDTC组细胞增殖能力显著上升(P<0.01),凋亡率下降(P<0.01),NGALR、NGAL、Snail、Vimentin及NF-κB p65磷酸化的蛋白表达均下降(P<0.01)。与PDTC组相比,PDTC+si-NGALR组变化趋势更加显著。结论:NGALR干扰通过调控NF-κB通路阻断高糖诱导肾小管EMT,抑制细胞的异常增殖和凋亡,减轻肾小管损伤。
Objective:To investigate the effect of neutrophil gelatinase-associated lipocalin receptor(NGALR)interference on the activity and epithelial mesenchymal transdifferentiation(EMT)of HK-2 cells induced by high glucose and its mechanism.Method:The HK-2 cells were divided into control group,model group,si-NGALR NC group,si-NGALR group,PDTC(NF-κB inhibitor)group and PDTC+si-NGALR group.CCK8 assay was used to detect cell proliferation,qRT-PCR was performed to detect the mRNA expression of NGALR,flow cytometry was used to detect apoptosis,Western Blot was used to detect protein expression of NGALR,NGAL,NF-κB pathway proteins(NF-κB p65 and p-NF-κB p65)and EMT-related proteins(Snail and Vimentin).Results:Compared with control group,the proliferation capacity of model cells was significantly reduced(P<0.01),the apoptosis rate was increased(P<0.01),and the expressions of NGALR,NGAL,Snail,Vimentin and p-p65/p65 proteins were increased(P<0.01).Compared with model group,the proliferation ability of cells in si-NGALR groups and PDTC groups were increased significantly(P<0.01),the apoptosis rate were decreased(P<0.01),and the expressions of NGALR,NGAL,Snail,Vimentin and p-p65/p65 proteins wre decreased(P<0.01).The trend was more significant in PDTC+si-NGALR group compared to the PDTC group.Conclusion:NGALR interference can block high glucose-induced epithelial mesenchymal transdifferentiation in renal tubular cells by regulating the NF-κB pathway,thereby inhibiting abnormal cell proliferation and apoptosis to relieve renal tubule injury.
作者
方珣
曾星若
陈隆敏
申江曼
董艳
吕会娟
高思
陈文莉
FANG Xun;ZENG Xing-ruo;CHEN Long-min;SHEN Jiang-man;DONG Yan;LU Hui-juan;GAO Si;CHEN Wen-li(Department of Rheumatology and Immunology,The Central Hospital of Wuhan,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430014,China)
出处
《微循环学杂志》
2023年第4期6-12,共7页
Chinese Journal of Microcirculation
基金
武汉市卫健委医学科研项目(WX19Y17)
