The Research On The Effect Of PARG On The Invasion Of Human Colon Carcinoma LOVO Cell Lines And Its Mechanism In Vitro

Objective: To study the effect of poly(ADP-ribose) glycohydrolase(PARG) on the invasion of human colon carcinoma Lovo cell lines in vitro,and its mechanism.Methods: PARG was suppressed stably on colon carcinoma Lovo cells by Lentivirus Vector-mediated PARG-shRNA transfection. LY294002(LY) was served as Akt phosphorylation inhibitor. The effects of PARG gene silencing on the Lovo cells involved in matrix adhesion, invasion and migration were observed by cell matrix adhesion, invasion and migration assay, separately. The expression of PARP mRNA was detected by RT-PCR. While the proteins expressions of PARP, NF-κB, Akt phosphorylation, MMP-2 and MMP-9 were investigated by western blot.Results: 1. Lovo cell line with PARG gene silenced was perpetually . The result of cell matrix adhesion assay displayed that Absorbance values of PARG-shRNA groups were clearly decreased, compared with blank control groups (P<0.05). Meanwhile, comparing with blank control groups, the invasive and migratory potencies of Lovo cells were both decreased in PARG-shRNA groups. The inhibitory rates of Lovo cells matrix adhesion, invasion and migration were 25.22%, 35.29%, 38.71% respectively. However, there was no significant difference on the adhesive ,invasive and migratory potencies of Lovo cells between PARG-shRNA+LY groups and blank control groups (P>0.05).2. The result of RT-PCR analysis showed that the expression of PARP mRNA in PARG-shRNA group was significantly weaker than that in the blank control group (p<0.05).3. The expressions of PARP, NF-κB, MMP-2 and MMP-9 in PARG-shRNA group were all conspicuously lower than that in the blank control group in western blot analysis (P<0.05,P<0.05,P<0.05,P<0.05 respectively); but however the expression of PI-Akt473 in PARG-shRNA group was stronger than that in the blank control group(P<0.05); Nevertheless, in PARG-shRNA+LY groups, the changes of expression of NF-κB, MMP-2 and MMP-9 were not conspicuous, comparing with in the blank control group(P>0.05).Conclusion: 1.The data suggested that PARG deficiency could decrease cells matrix adhesive, invasive and migratory potencies of Lovo cells. PARG probably plays an important role in the infiltration of human colon carcinoma.2. The expression of PARP, NF-κB, MMP-2 and MMP-9 in Lovo cells were decreased by PARG-deficiency synchronously, but PARG-deficiency was shown to upregulate Akt phosphorylation. Meanwhile, the inhibition of expression of NF-κB, MMP-2 and MMP-9 because of PARG-deficiency was prevented by the increase of Akt phosphorylation. The data suggest that PARG deficiency could reduce the cell-matrix adhesion, migration and invasion in Lovo cells. It probably because of PARG deficiency that weakens PARP activity; downregulating expression of NF-KB and its dependent genes such as MMP-2 and MMP-9 through PI3K /Akt signal pathway.