The Relationship Between The Expression Of RSPO3 And Tumor Progression In Hepatocellular Carcinoma

Objectives:1 To study the different expression of RSP03 in hepatocellular carcinoma tissues and peritumoral tissues,and to further study the relationship between RSP03 expression with clinicopathological grades of HCC in hepatocellular carcinoma tissues.2 To explore the relationship between the expression of RSP03 and the invasion and migration of HCC cell lines,and analyze the relationship between RSP03 expression and HCC epithelial-mesenchymal transition by protein and gene level detection.Methods:1 Immunohistochemical staining method was used to detect the expression of RSP03 in HCC and peritumoral tissues and study the relationship with clinicopathological features.The expression of RSP03 mRNA in HCC and adjacent tissues was detected by real-time fluorescent quantitative PCR.Western blot was used to detect the expression of RSP03 in HCC and adjacent tissues.2 Immunohistochemical staining was used to detect the protein expression of RSP03,E-cadherin,Vimentin,Snail,and explore the relationship between RSP03 with EMT in the protein level of tissues.3 Real-time fluorescent quantitative PCR and Western blot were used to detect the expression of RSP03 in HCC cell lines at the gene level and translation level,and to screen out the cell lines with high and low expression of RSP03.4 Inhibition of RSP03 expressions in HCCLM3,SMMC-7721 cell lines and overexpression of RSP03 in Hep3B cell line:(1)Real-time fluorescent quantitative PCR was used to explore transcription changes of RSP03 and E-cadherin,Vimentin,Snail in transfected cells;(2)Western blot and immunofluorescence staining were used to detect the expression of RSP03 and E-cadherin,Vimentin and Snail in transfected cells.5 Transfected cell lines were used to explore the effects of RSP03 expression on cell migration by using chamber experiments and wound healing experiments.Results:1 Real-time quantitative PCR showed that the expression of RSP03 mRNA:cancer tissues were higher than that of adjacent tissues.2 Western blot showed that RSP03 expressed higher in cancer tissue than in adjacent tissues.3 Immunohistochemical staining,the expression of RSP03 in grade ⅣEdmondson-Steiner pathological tissues with HCC was significantly higher than that in grade Ⅱ tissues;the expression of Vimentin and Snail was higher than that in gradeⅡ tissues,while E-cadherin on the contrary,the same trend was observed in the RSP03 low expression organization.4 The expression of Vimentin and Snail was consistent with RSP03 decreased,while E-cadherin expression was up-regulated after inhibition of RSP03 expression in cell lines HCCLM3 and SMMC-7721.Vimentin and Snail expression were up-regulated and E-cadherin expression was down-regulated after overexpression of RSP03 in Hep3B cell line;5 The number of cells in HCCLM3 and SMMC-7721 cells passing through the chamber to the serum side was reduced in the chamber experiment,and the scratch healing area was reduced in the scratch healing experiment after interfering with RSP03 expression;The number of cells that passed through increased in the chamber experiment,and the area of scratch healing increased in the scratch healing experiment after overexpression of RSP03 in Hep3B cells.Conclusions:1 In HCC tissues,RSP03 expression is closely related to clinical pathology and positively correlated with pathological grade;2 The expression of RSPO3 is related to EMT-related molecules in HCC,RSPO3 may promote tumor invasion and migration by regulating the process of EMT and lead to tumor progression.