The Role Of PTBP1 And Other Genes In Malignant Proliferation Of Hepatocellular Carcinoma Cells And Their Relation With Related TCM Syndromes

ObjectiveOn the basis of the previous research on the prevention and treatment of hepatocellular carcinoma with traditional Chinese medicine,we selected several genes that were up-regulated in the malignant proliferation of hepatocellular carcinoma cells.We used RNA interference and other experimental techniques to explore the role of these genes in the malignant proliferation of hepatocellular carcinoma cells,in order to enrich the pathogenesis of hepatocellular carcinoma and malignant proliferation mechanism,as well as some mechanisms of hepatocellular carcinoma heat-toxicity syndrome.MethodsIt mainly includes: culture of hepatocellular carcinoma cells,preparation of target gene interfering plasmids,transient transfection of interfering plasmids by liposome,MTT assay for cell proliferation ability after silencing target genes,change of gene expression by RT-q PCR after silencing target genes,change of protein expression by Western blot after silencing target genes,fine flow cytometry after silencing target genes and detection of cell cycle and apoptosis,exploration and improvement of related methodologies and literature review.Results(1)The interfering plasmids of 10 stress-related genes were successfully constructed.(2)Fluorescence control showed that plasmid transfection was successful.(3)72 hours after transfection of si RNA,the number of cells in PTBP1,RPS18,RPL13 A and RPL37 A gene silencing groups decreased significantly compared with the normal group.(4)72 hours after transfection of si RNA,the relative expression of PTBP1,RPS18,RPL13 A and RPL37 A genes were down-regulated in varying degrees compared with negative ones,and the extent of down-regulation was obvious.(5)72 hours after transfection of si RNA,the corresponding protein expression of PTBP1,RPS18,RPL13 A and RPL37 A gene silencing groups decreased in varying degrees compared with the control group.(6)72 hours after si RNA transfection,flow cytometry showed that compared with the negative plasmid transfection group,the peak area of G2 phase in PTBP1,RPS18,RPL13 A and RPL37 A gene knockout groups increased in varying degrees,but the change was not obvious.(7)72 hours after si RNA transfection,flow cytometry showed that compared with negative plasmid transfection group,the number of apoptotic cells in PTBP1,RPS18,RPL13 A and RPL37 A gene knockout group increased to some extent,but the change was not obvious.ConclusionPTBP1,RPS18,RPL13 A,RPL37A and other stress-related genes can maintain and promote the malignant proliferation of SMMC-7721 hepatocellular carcinoma cells,and their effects should not be achieved directly or indirectly by regulating cell cycle and apoptosis.Among the common complex syndromes in patients with hepatocellular carcinoma,heat toxin/stasis toxin syndrome is one of the basic syndromes,which is mainly manifested in the malignant proliferation and refractory treatment of hepatocellular carcinoma.The corresponding clinical treatment will be heat-clearing and detoxification herbs.Therefore,the results of this study suggest that the overexpression of PTBP1,RPS18,RPL13 A,RPL37A and other genes is related to the heat/blood stasis syndrome of hepatocellular carcinoma,which is the material basis for its occurrence.