The Research Of The Effect Of IL-17 On The Biological Behavior And PD-L1 Expression Level Of Human Intrahepatic Cholangiocarcinoma Cells

Objective: To investigate the role of interleukin-17（IL-17）in the proliferation,invasion and migration of human intrahepatic cholangiocarcinoma（iCCA）cells and its effect on the expression level of programmed cell death ligand 1（PD-L1）,and to further explore the possible molecular mechanisms.Methods: The expression of interleukin-17 receptor（IL-17R）of two human iCCA cell lines,RBE and HCCC9810 cells,was detected separately using Westen blot technique.The effect of IL-17 on the proliferative capacity of RBE and HCCC9810 cells was examined using the Cell Counting Kit-8（CCK-8）.Western blot was used to detect the activation of signaling pathways in both cell lines after IL-17 action,and according to the results of Western blot,the activation of signaling pathways was blocked using specific inhibitors of the corresponding signaling pathways.After blocking the signaling pathway,the effect of IL-17 on the proliferation,migration and invasion ability of RBE and HCCC9810 cells was detected again.Western blot was used to detect the effect of IL-17 on the expression level of PD-L1 in two human iCCA cell lines,and real-time quantitative PCR was used to detect the expression of PD-L1 at the m RNA level.And the effect of IL-17 on PD-L1 expression levels in RBE and HCCC9810 cells after the pathway was blocked was detected using Western blot.Results:（1）Western blot assay revealed that IL-17 R was expressed on the surface of both human iCCA cell lines RBE and HCCC9810 cells.（2）CCK-8 assay suggested that different concentrations of IL-17 significantly promoted the proliferation ability of both human iCCA cells（P < 0.05）.（3）Scratch assay and Transwell assay showed that IL-17 significantly promoted the migration and invasion ability of RBE and HCCC9810 cells（P < 0.05）.（4）Western blot results showed that IL-17 action on RBE and HCCC9810 cells promoted the phosphorylation levels of STAT3 and JAK2 proteins（P < 0.01）,and after blocking the activation of the signaling pathway using the JAK2/STAT3 signaling pathway-specific blocker AG490,the ability of IL-17 stimulated RBE and HCCC9810 cell proliferation,migration and invasion were significantly attenuated（P < 0.05）.（5）IL-17 at different concentrations（0,5,10,20,50 and 100 ng/ml）acting on human intrahepatic cholangiocarcinoma cell lines RBE and HCCC9810 induced the expression of PD-L1 in RBE and HCCC9810 cells,and the upregulation of PD-L1 was correlated with the concentration of IL-17 when the concentration was 50 ng/ml and 100 ng/ml,the induction of PD-L1 was most significant.Meanwhile,IL-17 likewise significantly stimulated PD-L1 expression at the m RNA level（P < 0.01）.（6）The ability of IL-17 to induce PD-L1 protein expression was significantly diminished after blocking the JAK2/STAT3 signaling pathway using AG490（P < 0.05）.Conclusions:（1）IL-17 promoted the proliferation,migration and invasion of intrahepatic cholangiocarcinoma cells.（2）IL-17 induced the expression of PD-L1 in intrahepatic cholangiocarcinoma cells.（3）IL-17 could induce the proliferation,migration,invasion and PD-L1 expression of intrahepatic cholangiocarcinoma cells by promoting the phosphorylation level of JAK2/ STAT3 signal pathway proteins.