Study on the molecular mechanism of Musashi RNA binding protein 2 in regulating the progression of hepatocellular carcinoma
10.3760/cma.j.cn113884-20231203-00152
- VernacularTitle:RNA结合蛋白Musashi-2调控肝细胞癌进展的分子机制研究
- Author:
Rui LI
1
;
Haichao ZHAO
;
Yanjun LI
Author Information
1. 山西医科大学第三医院 山西白求恩医院 山西医学科学院 同济山西医院肝胆外科，太原 030032
- Keywords:
Carcinoma, hepatocellular;
Musashi RNA binding protein 2;
Proliferation;
Wnt/β-catenin signaling pathway
- From:
Chinese Journal of Hepatobiliary Surgery
2024;30(8):606-612
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the role and related mechanisms of Musashi RNA binding protein 2 (MSI2) in hepatocellular carcinoma (HCC).Methods:The mRNA expression data of HCC patients from the cancer genome atlas (TCGA) and the genotype-tissue expression (GTEX) as well as the corresponding clinical information were obtained, and the expression level of MSI2 was evaluated in 50 pairs of matched tumor/normal HCC samples. Furthermore, the protein-protein interaction (PPI) network, the Kyoto encyclopedia of genes and genomes (KEGG) and the encyclopedia of RNA interactions (ENCORI) databases were used to explore how MSI2 affects the progression of HCC. Three pairs of short hairpin RNA (shRNA) sequences targeting MSI2 and β-catenin were designed and cloned into lentiviral vectors and transfected into HepG2, MHCC97H and MHCC97L cells as knockdown plasmids for the construction of MSI2 knockdown group cells (HepG2 sh-MSI2 group, MHCC97H sh-MSI2 group) and β-catenin knockdown group cells (MHCC97L sh-β-catenin group) and their respective negative control cells (HepG2 sh-Ctrl group, MHCC97H sh-Ctrl group, and MHCC97L sh-Ctrl group). The overexpression level of MSI2 was detected by Western blotting. CCK-8 and colony formation assay were used to detect the proliferation of hepatocellular carcinoma cells. Cell suspensions of HepG2 sh-Ctrl group, HepG2 sh-MSI2 group, MHCC97L sh-Ctrl group and MHCC97L sh-β-catenin group were injected subcutaneously into the front thighs of the nude mice in each group. Nude mice were sacrificed after 4 weeks, and tumors were removed for volume measurement. Results:The analysis based on TCGA and GTEX databases showed that the expression of MSI2 in 50 pairs of matched tumors was higher than that in normal samples (2.073±0.767 vs. 1.256±0.260), and the difference was statistically different ( P<0.001). The results showed that MSI2 was positively correlated with β-catenin ( r=0.455, P<0.001), transcription factor 7 (TCF7) ( r=0.142, P=0.006) and lymphatic enhancement factor 1 (LEF1) ( r=0.246, P<0.001) in HCC. Compared with the HepG2 sh-Ctrl group, the expressions of MSI2, β-catenin, TCF7 and LEF1 in the HepG2 sh-MSI2 group were down-regulated, and the expressions of MSI2, β-catenin, TCF7 and LEF1 in the MHCC97H sh-MSI2 group were down-regulated compared with the MHCC97H sh-Ctrl group, and the differences were statistically significant (all P<0.001). Compared with the MHCC97L sh-Ctrl group, the expression of β-catenin in the MHCC97L sh-β-catenin group decreased statistically significant ( P<0.001). The results of CCK-8 proliferation assay showed that the activity of HCC cells in the HepG2 sh-MSI2 group was reduced compared with the HepG2 sh-Ctrl group, in the MHCC97H sh-MSI2 group was decreased compared with the MHCC97H sh-Ctrl group, and compared with the MHCC97L sh-Ctrl group, the activity of HCC cells in the MHCC97L sh-β-catenin group was decreased (all P<0.001). The results of colony formation experiments showed that the ability of HCC cells to form colonies in the HepG2 sh-MSI2 group was reduced compared with the HepG2 sh-Ctrl group, and in the MHCC97H sh-MSI2 group was reduced compared with the MHCC97H sh-Ctrl group, furthermore, compared with the MHCC97L sh-Ctrl group, the ability in the MHCC97L sh-β-catenin group was significantly lower (all P<0.001). Compared with the HepG2 sh-Ctrl group, the tumor volume of nude mice in the HepG2 sh-MSI2 group decreased [(160.19±60.22) mm 3 vs. (480.46±65.28) mm 3], compared with the MHCC97L sh-Ctrl group, the tumor volume in the MHCC97L sh-β-catenin group also decreased [(456.48±38.23) mm 3 vs. (845.67±33.19) mm 3], and the differences were statistically significant (both P<0.05). Conclusion:Up-regulation of MSI2 expression promotes the proliferation of HCC cells through the Wnt/β-catenin signaling pathway.
