The Mechanisim Of IL-33/ST2 Axis On Regulation Of Melanoma Migration, Invasion And Vasculogenic Mimicry

Objective: Melanoma is the most deadly skin cancer.Although melanoma is a preventable malignant tumor,the incidence of melanoma is increasing year by year.Vasculogenic mimicry(VM),blood vessels formed by tumor cells instead of endothelial cells,is the cause of resistance to conventional anti-vascular treatment.Chronic inflammation and tumors are closely related.Inflammatory factors are an important feature of the tumor microenvironment.Interleukin-33(IL-33),as a common inflammatory factor in the tumor microenvironment,plays an important role in the progression of various tumors.IL-33 binding with ST2 can activate immune cells to produce a series of immune responses against tumors.IL-33 also affects metastasis by participating in immune responses.IL-33 can also act as ligands and directly bind to tumor cells.However,the impact of IL-33 on melanoma is not clear.This study aims to explore the effect of IL-33 on melanoma and its mechanism of action.Providing ideas for finding new targets for clinical melanoma immunotherapy or targeted therapy.Methods: 1.Case data and paraffin-embedded melanoma tissue from 66 cases were collected from Tianjin Cancer Hospital of China.Each case was diagnosed by two pathologists.Immunohistochemical staining and CD34/PAS staining were used to detect the expression level of ST2,MMP-2 and the presence of VM in the melanoma tissues and analyze the results.2.Two melanoma cell lines(A375,MUM-2B)from different tissue sources were used to build ST2 silenced stable cell lines.Transfection efficiency was detected by Western-blot.Colony formation assay,MTT assay,wound healing assay,transwell migration,invasion assay and three-dimensional culture were performed to detect proliferation,migration and invasion of melanoma cells in ST2 down-regulated and control cells under different concentrations of IL-33.And the impact of VM formation capabilities.3.Western-blot and RT-PCR were used to detect the expression of MMP-2 and MMP-9,VM-related markers,in A375 and MUM-2B cells under different concentrations of IL-33.Introducing MKK pathway blocker U0126 to block ERK1/2 phosphorylation of A375 and MUM-2B cells,then measure the expression of MMP-2/9 and the abilities of migration and invasion under IL-33.Results: 1.In the collected melanoma tissue from 66 patients,32 cases were found to have VM phenomenon,32 cases were MMP-2 positive,and 38 cases were ST2 positive.The results showed that there was a significant correlation between VM and the expression of MMP-2 in human melanoma tissues,and the expression of ST2 was significantly correlated with MMP-2 expression and VM(P<0.05).2.The results of plate cloning and MTT assay showed that the proliferation of A375 and MUM-2B cells were up-regulated under the action of a certain concentration of IL-33.Transwell experiments under different concentrations of IL-33 showed that the migration and invasion ability of A375 and MUM-2B cells increased with the increase of IL-33 concentration.Under IL-33,A375 and MUM-2B cells have enhanced the ability of forming tubes.3.Western-blot and PCR experiments showed that the expression of MMP-2/9 and p-ERK1/2 in A375 and MUM-2B cells was up-regulated by IL-33.The expression level of MMP-2/9 increased as the concentration of IL-33 increases.Under IL-33,the expression of MMP-2/9,the abilities of migration,invasion and tube formation in cells with U0126 were decreased.Conclusions: 1.VM phenomenon exists in human melanoma,and there is a correlation between VM and ST2 expression.ST2 expression is associated with MMP-2 expression.2.In vitro,IL-33/ST2 promotes the proliferation and VM formation of melanoma cells and promotes melanoma cells migration and invasion in a dose-dependent manner.3.IL-33/ST2 promotes melanoma cells migration and invasion and VM formation through the ERK1/2-MMP-2/9 pathway.