Circular RNA CircGSK3B Promotes Cell Proliferation,Migration,and Invasion In Hepatocellular Carcinoma

ObjectiveHepatocellular carcinoma(HCC)is the third leading cause of cancer-related deaths worldwide.It is considered an intractable disease due to its concealed early clinical symptoms and high rate of metastasis.At present,standard clinical practice of HCC is surgery supplemented with chemotherapy and treatment with multi-target tyrosine kinase inhibitors such as sorafenib and regorafenib.However,the 5-year survival rate of HCC patients is still very low.The dysregulation of circular RNAs(circ RNAs)has been shown to play an important regulatory role in the development of various cancers.However,the biological functions and potential molecular mechanisms of circ RNAs in HCC are still unclear.Methods1.Identification of circ GSK3 B,verifying the expression and function of circ GSK3 B in HCC:First,circ GSK3 B was screened from the GEO database,its ring structure was identified,and its intracellular localization was detected.The high expression of circ GSK3 B was verified in HCC tissues and cell lines,and the functional experiment verified the promoting effect of circ GSK3 B on HCC.Analyzed the relationship between circ GSK3 B expression and clinicopathological characteristics.2.Exploring the molecular mechanism of circ GSK3 B : We used Bioinformatics analysis to find out the downstream candidate miRNAs of circ GSK3 B and their target genes,and verified their mutual combination through luciferase experiments to preliminarily determine the downstream pathways of circ GSK3 B.On this basis,FISH experiments confirmed that circ GSK3 B co-localizes with miR-1265 in the cytoplasm,which provides a basis for circ GSK3 B to have miRNA sponge effect.To verify the low expression of miR-1265 in HCC and its negative regulation of target genes,functional experiments were used to verify the inhibitory effect of miR-1265 in HCC.Reverse experiments ware used to confirm that circ GSK3 B can promote the development of HCC through the miRNA sponge mechanism.3.Exploring the relationship between high expression of circ GSK3 B and glutamine metabolism in HCC: First,it was confirmed that circ GSK3 B can up-regulate GLS expression at the cellular level,and the next step was to verify that circ GSK3 B was positively correlated with GLS expression in HCC tissues.On this basis,we then explored the mechanism that circ GSK3 B promotes the production of glutamine metabolites and regulates the role of intracellular reactive oxygen species.ResultsWe found that circ GSK3 B was highly expressed in HCC tissues and HCC cell lines.Additionally,the expression level of circ GSK3 B significantly correlated with HCC tumor size and vascular invasion.Functionally,we confirmed that circ GSK3 B can promote the proliferation,migration,and invasion of HCC cells in vivo and in vitro.In terms of mechanism,we demonstrated that circ GSK3 B acts as a miR-1265 sponge,positively regulates the target gene CAB39,and promotes the reprogramming of glutamine metabolism,thereby promoting the progression of HCC.Finally,the classic RNA binding protein QKI was observed to participate in the biogenesis of circ GSK3 B.ConclusionsIn summary,we proved that the circ GSK3B-miR-1265-CAB39 axis can promote the proliferation,migration,invasion of HCC cells,indicating that circ GSKB may serve as a promising diagnostic and prognostic marker in HCC.