The Effects Of DJ-1 Knockdown For The Proliferation, Migration And Invasion In MGC803 Cells Inhibited By Diallyl Disulfide

Objective: The aim of this study was to investigate the effects of DJ-1knockdown and diallyl disulfide(DADS) on proliferation, migration and invasion in human gastric cancer MGC803 cells and to explore its molecular mechanism.Methods: DJ-1 knockdown MGC803 cell lines was established by transfecting DJ-1-specific shRNAs into the human gastric cancer MGC803 cells. Then, qRT-PCR and Western Blot assay were used to detect the interference efficiency. After treated with 30mg/L DADS, using MTT assay,clone formation assay, scratch woud healing assay,Transwell migration assay and Xenograft tumor assay to evaluate the effects of DJ-1 knockdown and DADS on proliferation, migration and invasion of MGC803 cells. In addition,Immunoflurescence assay, Western Blot and qRT-PCR determined the expression of DJ-1 in MGC803 cells. The expression of protein of PTEN and p-Akt in MGC803 cells was examined by Western Blot analysis.Results: 1. Stable DJ-1 knockdown MGC803 cell lines was established successfully.DJ-1 interference vector and negative control vector were transfected intoMGC803 cells. Then, using qRT-PCR and Western Blot assays detected the expressions of DJ-1 to identify the interference efficiency. Finally, after screening with Puromycin, stable DJ-1 knockdown MGC803 cell lines was achieved. Western Blot and qRT-PCR assays shows that the expressions of DJ-1 in transfected MGC803 cells was significantly decreased.2. The effects of DJ-1 knockdown and DADS on proliferation, migration and invasion in MGC803 cells.MTT and clone formation assays showed that DJ-1 knockdown and DADS could significantly inhibite the ability of proliferation in MGC803 cells,and silencing DJ-1 could strengthen the DADS inhibition on the proliferation in MGC803 cells. The scratch woud healing assay and Transwell migration assay showed, DADS and DJ-1 knockdown resulted in decreased migration and invasion of MGC803 cells(P<0.05). In addition, the ability of DADS inhibiting MGC803 cells migration and invasion could be enhanced after silencing DJ-1.3. The expression of DJ-1 in MGC803 cells after treated with DADS.The results of Immunoflurescence assay shows that DJ-1 are mainly distributed in the cytoplasm. Knockdown DJ-1 and DADS could significantly decrease the expression of DJ-1 protein in MGC803 cells. And that DADS could further decrease DJ-1 protein expression in DJ-1 Knockdown MGC803 cells. And the same results have been confirmed by qRT-PCR and Western Blot. In addition, Western Blot also showed that DADS and DJ-1 silencedcould up-regulated the expression levels of PTEN protein, as well as down-regulated phosphorylated Akt levels in MGC803 cells.4. The effect of DJ-1 silenced and DADS on growth of MGC803-induced tumor in nude mice.The result of Xenograft tumor growth assay showed that silencing DJ-1and DADS could inhibit the growth of MGC803-induced tumor in vivo.Immunohistochemical results show that, the expression of Ki-67, CD34 and Vimentin was obviously decreased in DJ-1 silenced group and DADS treated group comparing with control group, while the expression of PTEN and E-cadherin increased.Conclusion: 1. DADS can down-regulated DJ-1 inhibit the proliferation,migration and invasion through PTEN/Akt pathway in MGC803 cells.2. DJ-1 knockdown can inhibit MGC803 cells proliferation, migration and invasion and can increase the capacity of DADS inhibiting the proliferation, migration and invasion of MGC803 cells.3. DADS can inhibit the growth of MGC803-induced tumor in nude mice cooperating with DJ-1 silenced.