Research On The Mechanism Of SLPI Promoting Melanoma A375 Cell Migration

Experimental background: Melanoma,as one of the most rapidly developing malignant tumors with the worst prognosis,has attracted more and more attention.Invasion and metastasis are the most prominent features of melanoma and the primary factors affecting the survival of patients,and an important reason for its treatment difficulties.The local proliferation and metastasis of tumors are the main causes of many cancer deaths.Infiltration and metastasis of tumors is a multi-step,multi-factor complex and involves a variety of adhesion molecules,stromal proteases,cytokines,and corresponding signal transduction.Mechanisms and related genetic changes,the study of its transfer mechanism are currently difficult and hot spots.Recent studies have shown that secretory leukocyte protease inhibitor(SLPI)is both an inflammatory inhibitor and a protease inhibitor participates in the occurrence and metabolism of tumors by degrading peripheral tissues.Studies have shown that the expression level of SLPI is positively correlated with the ability of tumor migration and invasion.So far,the malignant phenotype of how SLPI enhances tumor migration is not clear.In this study,melanoma A375 cells were used as the research object,and the molecular mechanism of SLPI promoting A375 cell migration was verified by RT-PCR,Western Blot,gelatin zymography and Transwell cell migration assay.Experimental results: 1.RT-PCR results showed that MMP-2 m RNA levels did not change significantly in A375 SLPI(+)cells,but MMP-9 m RNA levels increased with treatment of A375 SLPI(+)cells with 5 μmol/L U0126 for 9h.The transcription levels of SLPI and MMP-2 were not affected,but the level of MMP-9 m RNA was significantly lower than that of untreated A375 SLPI(+)cells.2.Western Blot results showed that the expression of MMP-2/MMP-9 in A375 SLPI(+)cells increased,while the phosphorylation of extracellular signal-regulated kinase ERK1/2 increased in A375 SLPI(+)cells.Based on the interference of SLPI expression,the expression of MMP-2/9 in the cells decreased,and the phosphorylation of ERK1/2 decreased.After treating A375 SLPI(+)cells with 5 μmol/L U0126 for 9h,A375 SLPI(+)Compared with cells,the expression of MMP-2/9 was decreased,and the level of ERK1/2 phosphorylation was significantly decreased.3.Gelatin zymography results showed that the activity of MMP-9 in A375 SLPI(+)cells was stronger than that of normal A375 cells and SLPI si RNA based on A375 SLPI(+),with 5 μmol/L.After 9h treatment of A375 SLPI(+)cells with U0126,the activity of MMP-9 was significantly decreased,but there was no significant change in the activity of secreted MMP-2 in each experimental group.4.Transwell cell migration and invasion assay results showed that compared with other experimental cells,the migration and invasion of A375 SLPI(+)cells were faster,and the number of cells passing through the Transwell chamber membrane was significantly increased.The results were statistically significant;compared with A375 SLPI(+)cells,the number of transmembrane cells was significantly reduced after 5 μmol/L U0126 treatment of A375 SLPI(+)cells for 9h,and the transmembrane number of A375 cells without any treatment in the blank control group was similar.All of the above results indicate that SLPI,as a key signal molecule that promotes cell migration and invasion in melanoma A375 cells,activates MEK signaling pathway,increases ERK1/2 phosphorylation,induces expression of downstream MMP-2/9,and promotes melanin production.Migration and invasion of tumor cell A375 cells.