β₂-AR Study On The Effect Of Blockers On Liver Metastasis Of Colorectal Cancer By Inhibiting The M2 Polarization Of Kupffer Cell

Purpose:We found that the norepinephrine can promote liver metastasis of colorectal cancer in our previous study.In the current study,we will investigate the effects ofβ₂-adrenergic receptor（β₂-AR）blockers on the Kupffer cell（KC）polarization and the migration of colon cancer,and explore its mechanism.Methods:The experiment was divided into two groups,the control group and the experimental group.The control group was treated with PBS,and the experimental group pretreated KC with gradient concentrationβ₂-AR blocker ICI118551(10^-9M,10^-8M,10^-7M,10^-6M,10^-5M),then added the same concentration of NE(10^-9M)to culture 24 hours.The polarization of KC as a marker of M2 type using CD206,was observed by flow cytometry method.And we measure the cytokine secretion of KC by ELISA.We also used Transwell cell migration assay to detect the migration of CT26 cells.Western blot method was used to detect the expression of PI3K/Akt signal pathway related proteins of KC.RT-PCR method was used to detect the expression of CXCL12.Results:（1）Compared with the control group,it showed a decreasing trend of CD206 on KC after addingβ₂-AR blocker,and it has a statistical difference when the concentration reached to 10^-6M,10^-5M（P<0.05）.（2）After using gradient concentration ofβ₂-AR blocker to treat KC,the secretion of IL-6 has an incresing trend,and it has a statistical difference when the concentration reached to 10^-5M（P<0.01）.Although it doesn’t reach out to statistical difference,the amount of secreting cytokines of VEGF and TGF-βshowed a decresing trend by using treated KC withβ₂-AR blocker.（3）KC treated by high concentration(10^-6M,10^-5M)β₂-AR blocker significantly reduced the migration effect of CT26.（P<0.01）.（4）β₂-AR blocker inhibits the PI3K,p-Akt,AKT’s protein expression in KC,and reduce the expression of CXCL12.Conclusion:β₂-AR blocker may block the polarization of KC to M2 type by inhibit the PI3K/Akt signal pathway,and reduce the expression of CXCL12 in KC.Finally,it inhibit the intrahepatic migration of colon cancer cells.