ABSTRACT
Objective: To study the expression and effect of small nuclear ribonucleoprotein-associated protein B (SNRPB) on proliferation and metastasis of liver cancer tissues and cells. Methods: The bioinformatics database starBase v3.0 and GEPIA were used to analyze the expression of SNRPB in liver cancer tissue and normal liver tissue, as well as the survival and prognosis of liver cancer patients. The expression of SNRPB mRNA and protein in liver cancer cell lines were analyzed by qRT-PCR and Western blot. RNA interference technique (siRNA) was used to determine SNRPB protein expression down-regulation. The proliferation effect on hepatocellular carcinoma cells was observed by MTT assay. Transwell invasion and migration assay was used to detect the changes in the metastatic ability of liver cancer cells after SNRPB down-regulation. Western blot was used to detect the changes of epithelial mesenchymal transition (EMT) markers in liver cancer cells after down-regulation of SNRPB expression. Data were compared between two groups and multiple groups using t-test and analysis of variance. Results: The expression of SNRPB was significantly higher in liver cancer tissue than normal liver tissue, and its expression level was correlated with the prognosis of liver cancer patients. Compared with the immortalized hepatocyte LO(2), the expression of SNRPB was significantly increased in the liver cancer cells (P < 0.01). siRNA-SNRPB had significantly inhibited the expression of SNRPB mRNA and protein in liver cancer cells. MTT results showed that the absorbance value was lower in SNRPB knockdown group than negative control group, and the difference at 96 h after transfection was most significant (P < 0.01). Transwell assay results showed that compared with the negative control group, the SNRPB knockdown group (MHCC-97H: 121.27 ± 8.12 vs. 46.38 ± 7.54; Huh7: 126.50 ± 6.98 vs. 41.10 ± 8.01) invasion and migration (MHCC-97H: 125.20 ± 4.77 vs. 43.18 ± 7.32; Huh7: 132.22 ± 8.21 vs. 38.00 ± 6.78) ability was significantly reduced (P < 0.01) in liver cancer cells. Western blot showed that the expression level of epithelial phenotype marker E-cadherin was decreased after down-regulation of SNRPB, while the expression levels of mesenchymal phenotype markers N-cadherin and vimentin was increased, suggesting that down-regulation of SNRPB inhibited EMT in liver cancer cells. Conclusion: SNRPB expression is significantly increased in liver cancer tissues and cells, and it is involved in regulating the proliferation, metastasis and EMT of liver cancer cells.
Subject(s)
Humans , Carcinoma, Hepatocellular/genetics , Cell Line, Tumor , Cell Movement , Cell Proliferation , Epithelial-Mesenchymal Transition , Gene Expression Regulation, Neoplastic , Liver Neoplasms/genetics , snRNP Core ProteinsABSTRACT
Thrombospondin 4 (THBS4), a member of the THBS family, is a protein secreted by the extracellular matrix and is involved in regulating various physiological processes, such as cell proliferation, adhesion and angiogenesis. Recent studies have shown that the inflammation stimulates THBS4 production and induces the adhesion and accumulation of macrophages. Our previous study confirmed that THBS4 acts as an oncogene in hepatocellular carcinoma (HCC), the effect of THBS4 on the immune microenvironment of HCC remains unclear. This study aims to analyze the role of THBS4 in promoting the metastasis of HCC cells by inducing M2-type polarization of tumor-associated macrophages. We simulate the tumor microenvironment through HCC conditioned medium (HCM) and found that the expression of THBS4 in macrophages increased in a time-dependent manner under the action of HCM (P<0.05); THBS4 knockdown promotes the expression of M1 macrophages markers IL-1β and CD86 (P<0.01), while the expression of M2-type markers IL-10 and CD206 were decreased (P<0.01). Transwell co-culture assay was used to further detect the effect of THBS4-induced M2-type macrophages on HCC metastasis. Results from co-culture of THBS4-downregulated M2 macrophages with HepG2 cells showed that THBS4-downregulated M2-TAMs significantly inhibited the invasion and migration ability of HepG2 cells (all P < 0.01). In conclusion, the tumor microenvironment promotes the expression of THBS4 in macrophages, and THBS4 may promote the invasion and metastasis of HCC cells by inducing M2-type polarization of macrophages. This study provides some new experimental basis for exploring the establishment of THBS4-induced HCC immune microenvironment.
ABSTRACT
Objective: To investigate the expression of small nucleolar RNA host gene 5 (SNHG5) in hepatocellular carcinoma (HCC) and its effect on epithelial-mesenchymal transition (EMT) and tumor stem cell proliferation. Methods: The expression of SNHG5 gene was detected by qRT-PCR in 48 cases of HCC tissues and corresponding adjacent tissues. The results and clinical pathological parameters were analyzed. The SNHG5 knockdown lentivirus was constructed and transfected into HCC cell lines, and the efficiency was verified by qRT-PCR. The cell invasion and migration abilities were detected by Transwell test; the formation and proliferation of HCC stem cells were detected by stem cell glomeration test. The expressions of EMT and stem cell-related genes mRNA and protein were detected by qRT-PCR and Western blot. Results: In 48 HCC patients, the expression of SNHG5 was significantly higher in HCC than in adjacent normal tissue. The high expression of SNHG5 was associated with tumor size, HBV infection, pathological differentiation type and clinical stage of HCC (P<0.05). At the cellular level, knockdown of SNHG5 inhibited the invasion and migration of HCC lines HepG2 and Huh7 and the number and diameter of cancer stem cells (CSCs). The expression of E-cadherin was significantly increased in the SNHG5 knockdown group while the expressions of N-cadherin and stem cell-associated proteins OCT4 and SOX2 were significantly reduced (P<0.05). Conclusion: SNHG5 is highly expressed in HCC. Knockdown of SNHG5 inhibits the invasion and migration of HCC cells and the proliferation of CSCs. The mechanism may be related to the knockdown of SNHG5 to promote the reversal of HCC cell EMT and reduce the characteristics of HCC cancer stem cells.