缺氧环境下的肝星状细胞中缺氧诱导因子1α与缺氧诱导因子2α对CD248表达水平影响

Effects of hypoxia inducible factor-1αand hypoxia inducible factor-2α on CD248 expression in hepatic stellate cells under hypoxicconditions

目的探究缺氧状态下的肝星状细胞(HSC)中缺氧诱导因子1α(HIF-1α)、缺氧诱导因子2α(HIF-2α)对内皮唾液酸蛋白(endosialin/CD248/TEM1)表达水平的影响。方法首先,通过对布-加综合征(BCS)患者肝组织样本进行HIF-1α、HIF-2α和CD248免疫组化染色,并通过计算HIF-1α、HIF-2α和CD248的免疫组化累积光密度(IOD)值来分析CD248与HIF-1α、HIF-2α之间表达的相关性。其次,将成功包装的HIF-1α、HIF-2α过表达慢病毒和空载慢病毒分别感染体外培养的LX-2人肝星状细胞(HSC-LX2),并使用CoCl2进行低氧诱导处理。使用β-actin作为内参基因,通过定量聚合酶链反应技术观察缺氧环境下过表达慢病毒和空载慢病毒感染的HSC-LX2细胞中CD248基因表达水平,并对数据进行统计学分析。结果通过对BCS患者肝组织样本HIF-1α、HIF-2α和CD248的IOD值进行检测,显示CD248与HIF-1α(Pearson相关系数:r=0.285,P=0.010)和HIF-2α(Pearson相关系数:r=0.382,P<0.001)的表达水平呈现显著的正相关。定量聚合酶链反应技术检测显示,与空载慢病毒感染的HSC-LX2细胞比较,HIF-1α与HIF-2α过表达慢病毒感染的HSC-LX2细胞中CD248的表达丰度均明显升高(P均<0.05)。结论在缺氧状态下的HSC中,HIF-1α、HIF-2α均可能参与调控CD248基因表达增加。

Objective To investigate the effect of hypoxia inducible factor 1α(HIF-1α)and hypoxia inducible factor 2α(HIF-2α)on the expression of endosialin(CD248/TEM1)in hepatic stellate cells(HSC)under hypoxia.Methods Immunohistochemical analyses of HIF-1α,HIF-2α and CD248 expression in liver tissue samples of Budd-Chiari syndrome(BCS)patients were performed,immunohistochemical cumulative optical density(IOD)values of HIF-1α,HIF-2α and CD248were calculated,and then their correlation was evaluated.Lentivirus packaged with HIF-1α,HIF-2α overexpression and empty lentivirus were separately introduced into LX-2 human hepatic stellate cell(HSC-LX2)cultured in vitro and treated with CoCl_2 for hypoxia induction.Subsequently,quantitative polymerase chain reaction technology was used to observe the CD248 gene expression level in HSC-LX2 cells infected with overexpressed lentivirus and empty lentivirus under hypoxic environment by using β-actin as the internal reference gene,and the data were statistically analyzed.Results IOD values of HIF-1α,HIF-2α and CD248 in liver tissue samples from BCS patients were detected and analyzed.The CD248 expression level was correlated with HIF-1α(pearson correlation coefficient:r=0.285,P=0.010)and HIF-2α(pearson correlation coefficient:r=0.382,P<0.001).Compared with HSC-LX2 cells infected with empty lentivirus,the CD248 expression abundance in HSC-LX2 cells infected with HIF-1α and HIF-2α overexpression lentivirus were significantly increased(all P<0.05).Conclusion Both HIF-1α and HIF-2α may be involved in upregulating CD248 gene expression in HSC under hypoxia.