The Mechanism Of PRDM2Gene Promoter Methylation Involved In The Lung Squamous Cell Carcinoma

【Objective】PRDM2gene is an important member of the tumor suppressor PRDM gene family, ina variety of tumors gene expression appears to reduce or missing because of promotermethylation.The research about PRDM2in lung cancer has not been reported.5-aza-2’-deoxycytidine (5-aza-2dC) is methyl transferase inhibitor, can reduce the level ofmethylated genes.【Methods】1.Cultured lung squamous cell carcinoma SK-MES-1cells,A gradient concentration of5-aza-2dC were processed to the cells,using the MTT assay cell growth curve; MSP,RT-PCR and Western-blot method were determined PRDM2gene promoter methylationstatus, mRNA and protein expression levels;2.22BAL B/c nude mice were randomly divided into a control group and5-aza-2dCtreatment group (treatment group), using subcutaneous SK-MES-1cells to establishxenograft model; when the xenograft’s short diameter reaches0.5cm, the treatmentgroup in accordance with1μg/g body weight three times a week5-aza-2-dC peritonealcavity injection4weeks, the control group received the same amount of5-aza-2-dCvehicle (PBS) three times weekly peritoneal cavity injection control for4weeks,observed the xenograft’s growth;after four weeks the xenograft was removed, usingMSP assay detect the PRDM2methylation status;RT-PCR and Western-Blot methodfor detecting PRDM2mRNA and protein expression levels.【Results】5-aza-2-dC for SK-MES-1cells with growth inhibitory effect in a concentration-dependent effect; SK-MES-1cells in a highly PRDM2gene promoter methylationstatus,5-aza-2-dC treatment can reduce the degree of PRDM2gene promotermethylation with concentration-dependent manner, and concentration-dependent up-regulate PRDM2gene mRNA, protein expression levels (0μmol/L5-aza-2-dCgroup,1μmol/L5-aza-2-dC group,5μmol/L5-aza-2-dC group mRNA levels were0.18±0.04,0.32±0.07,0.41±0.09, comparison between groups p <0.05, havesignificant difference; protein expression levels were0.21±0.05,0.38±0.09,0.62±0.11, comparison between groups p <0.05, have a significant difference). The processof tumor-bearing nude mice, the treatment group tumor growth is slower than thecontrol group. At the end, the xenografts’volume of treatment and control groups,respectively are (570.±196) mm3and (1499±350) mm3, have a significant difference(t=7.303, p <0.01) between groups. MSP test results showed that: the control groupxenograft PRDM2gene promoter methylation frequency of81.8, the treatment groupxenograft PRDM2gene promoter methylation frequency of27.3%. The PRDM2mRNA in the control group and the treatment group were0.1703±0.1201and0.5322±0.090, protein expression levels were0.1451±0.1012and0.5125±0.4902. Thedifference was significant (p <0.05).【Conclusions】1.PRDM2in the SK-MES-1cells with a highly promoter methylation status,5-aza-2-dCable to concentration-dependently reduced PRDM2gene promoter methylation levelsand correspondingly increase PRDM2mRNA and protein expression levels;2.5-aza-2-dC intraperitoneal injection therapy can reduce nude’s PRDM2genepromoter methylation status, up-regulate the mRNA and protein expression levels, andinhibit the tumor growth.