犚犐犆犜犗犚在食管鳞状细胞癌增殖中的功能研究

Objective To explore the effect of RICTOR on the proliferation ability of esophageal squamous cell carcinoma cells and analyze its biological function in esophageal squamous cell carcinoma. Methods The expression of RICTOR and its impact on the prognosis of tumor patients were analyzed by using the cBioPortal database. SiRNA interference technology was used to construct siRNA-NC, siRNAl-RICTOR, and siRNA2-RICTOR cells of esophageal squamous cell carcinoma cells KYSE30 and KYSE150, respectively. The cell counting kit 8(CCK8) and clone formation experiments were used to detect the differences in cell proliferation and clone formation abilities among the groups. The slow virus transfec-tion method was used to construct KYSE30 stable knockdown RICTOR shRNA-RICTOR and shRNA-NC cells, and the effect of RICTOR knockdown on the growth of transplanted tumors in nude mice was tested through subcutaneous tumor-igenesis experiments. Protein blotting was used to detect the expression level of downstream target proteins of RICTOR, and the effect of RICTOR inhibitor JR-AB2-011 on the proliferation of esophageal squamous cell carcinoma cells was preliminarily explored through in vitro experiments. Results The database analysis results showed that high expression of RICTOR was associated with poor prognosis in tumor patients (P:0. 013). The CCK8 detection results showed that at 96 hours after knocking down the RICTOR gene, in the KYSE30 cell line, compared with siRNA-NC (1. 46 + 0. 02), siR-NA1-RICTOR (0. 85 + 0. 03) and siRNA2-RICTOR (0. 70 + 0. 03) showed a decrease in cell proliferation ability, r-values were 33.51 and 47. 13, respectively, both P<0. 001; In the KYSE150 cell line, compared with siRNA-NC (2.37 + 0. 10), the proliferation ability of siRNAl-RICTOR (1. 63 + 0. 02) and siRNA2-RICTOR (1. 39 + 0.03) cells was also reduced, ^-values were 16. 14 and 20. 48, respectively, both P<0. 001. The results of clone formation experiments showed that in the KYSE30 cell line, compared with siRNA-NC (74. 00 + 3. 26), siRNAl-RICTOR (50. 67+1. 24) and siRNA2-RICTOR (46. 33+1. 24) cells showed a decrease in clone formation ability, ^-values were 9. 44 and 11. 19, respectively, both P<0. 001; In the KYSE150 cell line, compared with siRNA-NC (86. 00 + 2. 44), siRNAl-RICTOR (35. 33 + 2. 05) and siRNA2-RICTOR (40. 33+ 1. 69) also showed a decrease in cell clone formation ability, ^-values were 22. 41 and 21. 66, respectively, both P<0. 001. The nude mouse tumor formation experiment showed that at the 5th week after subcutaneous inoculation, the tumor volumes of shRNA-RICTOR and shRNA-NC cells in nude mice were (104.42 + 50. 84) and (235. 32 + 85. 94) mm3, respectively, 1: 2. 62, P:0. 031. The results of Western blot analysis showed that RICTOR was involved in regulating the AKT signaling pathway, and the phosphorylation level of AKT Ser473 site in e-sophageal squamous cell carcinoma cells with RICTOR knockdown was significantly down-regulated. In addition, in vitro experiments have shown that the inhibitor JR-AB2-011 of RICTOR which was cultivating KYSE30 and KYSE150 cells at a concentration of 0. 25 (xmol/L for 72 hours inhibited cell proliferation, with statistically significant differences, t-values were 39. 64 and 28. 22, respectively, both P<0. 001. Conclusion RICTOR may affect the proliferation and cloning ability of esophageal squamous cell carcinoma cells by regulating the AKT signaling pathway, and inhibiting the RICTOR signaling pathway has potential anti-tumor clinical value.