| Objective:(1) To investigate the the methylation status of uPA and MMP-9 gene is relationship with its mRNA expression of these genes in human cutaneous melanoma.(2) Explore the mechanism of DNA methylation of uPA and MMP-9 gene in the pathogenesis of human cutaneous malignant melanoma.Method:(1) Human cutaneous melanoma A375 cell line was cultured in vitro, the impact of S-Adenosylmethionine (SAM) on the proliferation and growth of A375 cell line was observed with MTT assay.(2) According to unaccepted or accepted treatment of A375 cell line with different concentration of SAM (100,250,400μmol/L) for different period (2,3,5days) as control or treated group, using reverse-transcription polymerase chain reaction (RT-PCR) technique, detect the difference of expression of uPA and MMP-9 mRNA expression between two groups.(3) Methylation specific PCR (MS-PCR) was used to determine the alteration of the methylation status of uPA and MMP-9 promoter between two groups.Result:(1) SAM was demonstrated to inhibit the proliferation and growth of A375 cell line.(2) Both uPA and MMP-9 mRNA are significantly increased expression in control group, whereas the gene promoter status is completely or partially demethylated.(3) After treatment of A375 cell lines with SAM, resulted in decrease tumoral uPA and MMP-9 mRNA expression , at the same time showed that the ability of SAM to a dose- and time-dependent inhibition of uPA and MMP-9 expression compared with control one , whereas the decreased of demethylation level as well as hypermethylation is gradully occurred in gene promoter. Conculusin:after treatment A375 cell line with SAM showed that the inhibition of both uPA and MMP-9 mRNA expression was due to hypermethylation of their promoter region. These studies support the hypothesis that DNA hypomethylation might play a causal role in tumorgenesis and progression is activation of multiple tumor-promoting genes such as uPA and MMP-9 in cutaneous malignant melanoma.
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