四环素对纳米细菌感染的人肾小管上皮细胞HK-2中CaSR、Claudin-14表达的影响

Effects of Tetracycline on the Expression of CaSR and Claudin-14 in Nanobacteria-infected Human Renal Tubular Epithelial Cells HK-2

目的:研究四环素对纳米细菌(NB)感染的人肾小管上皮细胞HK-2中钙敏感受体(CaSR)、紧密连接蛋白14(Claudin-14)表达的影响,为抑制肾结石的形成提供参考。方法:将HK-2细胞分为空白对照组、NB组和四环素组(NB+四环素),每组3个复孔,各组分别加入相应NB和药物后共培养24 h。采用透射电子显微镜观察各组细胞的形态和超微结构变化,运用实时荧光定量聚合酶链式反应(qRT-PCR)和Western blot法分别检测细胞中CaSR、Claudin-14 mRNA和蛋白的表达水平。结果:与空白对照组比较,NB组细胞膜结构破损,胞核裸露在外,微绒毛基本消失,线粒体数量较少,结构肿胀,细胞中CaSR、Claudin-14 mRNA和蛋白表达水平明显升高(P<0.05);四环素组细胞膜结构完整,微绒毛结构稍紊乱,局部绒毛消失,部分线粒体结构肿胀不明显,细胞中CaSR、Claudin-14 mRNA和蛋白表达水平明显升高(P<0.05)。与NB组比较,四环素组细胞中CaSR、Claudin-14 mRNA和蛋白表达水平明显降低(P<0.05)。结论:NB可能通过上调HK-2细胞中CaSR、Claudin-14 mRNA和蛋白的表达促进肾结石的形成,而四环素可能通过作用于NB降低CaSR、Claudin-14 mRNA和蛋白的表达抑制肾结石的形成。

OBJECTIVE:To study the effects of tetracycline on the expression of calcium-sensing receptor(CaSR)and Claudin-14 in nanobacteria-infected(NB)human renal tubular epithelial cells HK-2,and to provide reference for inhibiting the formation of kidney stone.METHODS:HK-2 cells were divided into blank control group,NB group and tetracycline group(NB+tetracycline),with 3 complex holes in each group.They were given relevant nanobacteria and drug for 24 h.Morphology and ultrastructural changes of cells were observed by TEM.mRNA and protein expression of CaSR and Claudin-14 were measured by real-time quantitative PCR(qRT-PCR)and Western blot assay.RESULTS:Compared with blank control group,the damaged structure of cell membrane,exposed nucleus,disappeared microvilli,small number of mitochondria and swollen structure were found in NB group;mRNA and protein expression of CaSR and Claudin-14 were increased significantly(P<0.05).The complete structure of cell membrane,disordered structure of microvilli,disappeared local villi and not obvious swollen of some mitochondrial structure were found in tetracycline group;mRNA and protein expression of CaSR and Claudin-14 were increased significantly(P<0.05).Compared with NB group,mRNA and protein expression of CaSR and Claudin-14 were decreased significantly in tetracycline group(P<0.05).CONCLUSIONS:NB may improve the formation of kidney stones by up-regulating the mRNA and protein expression of CaSR and Claudin-14 in HK-2 cells,while tetracycline may reduce the mRNA and protein expression of CaSR and Claudin-14 by anti-NB effect so as to inhibit kidney stone formation.