miR-143在肝脏组织中的表达及调控机制

Study on the loss of miR-143, targeting to SHP2, aggravates liver cancer cells malignancy through promoting cancer stem cells population

目的探讨miR-143在肝癌组织中的表达及其调控机制。方法采用qRT-PCR实验用来检测肝癌组织与肝癌细胞系中的miR-143表达;慢病毒感染实验用来敲低或者过表达miR-143。流式细胞术检测CD133阳性的肿瘤干细胞比例。miRNA靶基因预测(Targetscan)与荧光素酶报告基因实验用验证miR-143的靶基因。Westernblot实验用来检测SHP2-ERK1/2信号通路在肝癌干细胞干性维持中的作用。结果miR-143在肝癌组织与细胞系中皆呈低表达趋势(P<0.01)。敲低miR-143能够进一步促进肝癌细胞的恶性生物学行为,相反,过表达miR-143可以逆转此效应。miR-143的敲低可以显著提高肝癌细胞中干细胞的比例。这一效应是通过调控SHP2-ERK1/2信号通路来实现的。过表达miR-143可以显著降低肿瘤干细胞比例。结论miR-143通过调控SHP2-ERK1/2信号通路来调节肿瘤干细胞的比例进而调控肿瘤的恶性程度。

Objective To understand the mechanism dysfunction of miR-143 in liver cancer patients. Methods qRT-PCR was used to examine the expression of miR-143 in the liver cancer tissues and cell lines. Lentiviral infection assay was adopted to establish the liver cells with knock down or overexpression of miR-143. Clony formation, MTT assays were used to examine the proliferation of liver cancer cells with miR-143 inhibition or overexpression. Transwell assay was applicable to determine the capacity of liver cancer cells migration and invasion. Flow cytometry was used to examine the population of CD133 positive cancer stem cells. Targetscan was used to predict the target gene of miR-143 and luciferase reporter assay confirmed the interaction between miR-143 and SHP2. Western blot (WB) assay was used to confirm the function of SHP2-ERK1/2 signaling in cancer stem cells maintenance. Results The expression of miR-143 was significantly lower ( P <0.01) in the liver cancer tissues and cell lines than normal tissues and cell lines. Down-regulation of miR-143 further aggravated the malignancy of liver cancer cells and promoted the population of cancer stem cells. Interestingly, miR-143 regulated the phosphorylation of ERK through targeting to SHP2. Conclusion Identification of the miR-143 function in liver cancer cells provided a new insight into liver carcinogenesis and was aslo critical to next-step liver cancer diagnosis and treatment.