Increased cell proliferation and R.Msp1 fragmentation induced by 5-aza-2 '-deoxycytidine in rat testes related to the gene imprinting mechanism

DNA methylation is one of the crucial mechanisms for cellular and tissue di fferentiation during developmental stages in mammals. 5-aza-2'-deoxycytidin e, a specific cytosine DNA Methyltransferase inhibitor, is known to cause D NA hypomethylation in CpG, CpNpG and CCGG sequences. Therefore the present study was designed to determine the effects of 5-aza-2'-deoxycytidine on th e germinal cells of the adult ra I: testicular tissue. Rat testicular tissues from the 5-aza-2'-deoxycytidine treated experimental and non-treated control groups were processed for light microscopy and als o for genomic DNA isolation assays. The isolated genomic DNAs were digest e d with R.Msp1 in order to determine the methyl pattern differences in the e nzyme cognate CCGG sequence. Testicular tissues from treated rats showed increased cell proliferation wh en investigated at the light microscopical level. On the other hand, genomi c DNA of these proliferative tissue showed high fragmentation sizes of R.Ms p1 digestion when compared to controls. While the R.Msp1 digested control g roup DNA fragmentation condensed at approximately 4700-5100 bps size, the e xperimental group DNA fragmentation was condensed at 700-900 bps size. In a ddition, 5-aza-2'-deoxycytidine has effects on increased ce:ll proliferatio n via the loss of somatic de novo gene imprinting. These results imply that abnormally imprinted normal somatic cells in mammals are susceptible to ep igenetic modification. These results also suggest that the genomic DNA of t esticular tissues from control rats is resistant to R.Msp1 while DNA from t he experimental group testicular cells demonstrating high proliferation rat e could not resist to R.Msp1 digestion due to DNA hypomethylation in CCGG s equence. In conclusion, it could be suggested that the reversal of gene imprinting i n germinal cells may cause an increased cellular proliferation and R.Msp1 f ragmentation when induced by 5-aza-2'-deoxycytidine.