LncRNA NEAT1参与钙离子诱导HK-2细胞成骨样分化调控Randall钙斑形成的机制研究

LncRNA NEAT1 in the regulation of randall's plaque formation by calcium ion-induced osteo-genic differentiation of the HK-2 cell

目的研究lncRNA NEAT1在Randall钙斑中的表达水平以及lncRNA NEAT1在钙离子诱导的人肾近端小管上皮细胞(HK-2细胞)中成骨样分化的分子机制。方法通过应用Real-time PCR和Western blot来检测肾结石患者Randall钙斑组织和正常肾乳头组织成骨蛋白(BMP-2、OCN)以及lncRNA NEAT1的mRNA表达水平。体外培养HK-2细胞,将其分为对照组、10 mmol钙离子组、20 mmol钙离子组,各组细胞培养至第15天时,采用细胞茜素红钙染色液对细胞进行染色,观察各组细胞钙结节形成的表达情况。采用Real-time PCR和Western blot检测各组细胞lncRNA NEAT1及成骨蛋白的mRNA表达水平。结果 lncRNA NEAT1和成骨蛋白在Randall钙斑组织中显著高表达。细胞染色结果发现不同浓度的钙离子组均可以见到明显被染成红色的钙结节,且lncRNA NEAT1和成骨蛋白的mRNA表达呈钙离子浓度依赖性升高(P<0.05)。结论在体外培养的环境下,HK-2细胞可以在高钙离子的刺激作用下发生成骨分化。lncRNA NEAT1可能通过参与钙离子诱导HK-2成骨样分化调控Randall钙斑形成。

Objective To investigate the expression Ievel of lncRNA NEAT1 in Randall plaques and the molecular mechanism of osteoid differentiation of lncRNA NEAT1 in calcium induced human proximal tubular epithelial cells(HK-2 cell),.Methods Firstly,real-tine PCR was used to test the mRNA expression levels of bone morpbogenetic protein 2(BMP-2),ostecalein(0CN)and lncRNA NEAT1 in of Randalls plaque tssue and normal renal papillary issue.Then human renal proximal tuo-bular epithelial cells were cultured in vitro(control group,10 mmol calcium group,20 mmol calcium group).The cells in each group were cultured to the 15th day respectively,and the cells in each group were stained with the cell Alizarin red S calcium staining to observe the expression of calcium nodule formation.In addition,real-time PCR was used to detect the mRNA expression levels of IncRNA NEAT1 and osteogenic protein(BMP-2 and OCN)in each group.Results LncRNA NEATI and 0s-teogenic proteins(BMP-2 and 0CN)were highly expressed in Randall plaque.The cell staining re-sults confirmed that the typical red caleium nodules could be seen in the calcium ion-induced group,and the mRNA exresions of lncRNA NEATI and oatogenie proteins(BMP-2 and OCN)were elevated in a calcium ioo conentration dependent monner when compured with coctrol group(P<0.05).Conclu-sions Human kidney procimal tubular epithelial cells can undergo cstogenie diferentiation under the stimulaion of the high cncentration of ealciun ion in vitro.In:RNA NEATI may regulate Randall's plaque frmtion by participating in calcum ion-induced etetgenie diferentiaio of the HK-2 ell.