Expression Of Dnmt3b And Pdcd4 In Hepatocellular Carcinoma And Its Mechanism Study

DNA methylation is a natural DNA modification method, and is one of the main content of epigenetics. This regulation does not change DNA sequence and is reversible, which play an important role in many aspects such as regulating gene expression, regulating embryonic development, genomic imprinting, X chromosome inactivation and apoptosis.DNA methylation is catalyzed by various methyltransferase enzymes. DNMT3b is one of the essential kind, which catalyze de-novo-methylation in the region of CpG island. The over-expression of DNMT3b has been found in in a variety of tumor, it lead to a variety of tumor suppressor gene hypermethylation and play an important role at the occurrence and development of tumor.Programmed cell death factor 4(PDCD4) is a suppressor gene related to apoptosis, it inhibits cell growth at the level of protein translation by impact the translation initiation factor eIF4A, eIF4G The current study found that its expression decreased in lung and digestive carcinomas, and induced apoptosis by caspase pathway, but its expression in the mechanism is unclear. Study shows that, there is many CpG islands exist in its promoter region, so we speculate that its expression and regulation may be related to DNA methyltransferase enzymes.Purpose Our study aims to detect the expression of DNA methylation transferase 3b, programmed cell death 4 in hepatocellular carcinoma and liver cancer cell line SMMC7721 and their relationship. At the same time, we also analysis PDCD4 in hepatocellular carcinoma occurrence and development, its clinical significance.Methods1. Immunohistochemical method was used to test the exoression of DNMT3b and PDCD4 in 31 normal liver tissue, 72 liver cancer tissue and its adjacent tissues for the purpose of penetrating their relevance, and at the same time the pathological parameters of PDCD4 in hepatocellular carcinoma.2. Cultivate liver cancer cell line SMMC7721, total protein and genomic DNA were extrcated to use.3. With different concentrations of methylase inhibitor 5-aza-CdR cultivate SMMC7721 cells, and two groups of cells extracted total protein and genomic DNA to use.4. Western-blotting was used to test the changes of DNMT3b and PDCD4 proteins, and the methylation level of PDCD4's promoter was tested by methylation specific polymerase chain reaction(MSP) method.5. To deal with the data by SPSS13.0, and consequently come to a conclusion.Results1. The positive rate of PDCD4 in normal tissues, cancer tissues and adjacent tissues is 70.97%, 45.83%, 34.72%, there are differences between normal tissues and cancer tissues, adjacent tissues and normal tissues, but there are no differences between cancer tissues and its adjacent tissues; The positive rate of DNMT3b in the three tissues is 35.48%,59.72%,70.83%, the differences between them are same to the PDCD4; The expression of PDCD4 and DNMT3b in cancer tissue was a negative correlation, r =- 0.059.2. Western-blotting showed that with 5-aza-CdR treatment DNMT3b protein expression in a lower level, while the expression level of PDCD4 increased, there is difference between the concentration.3. The methylation level of PDCD4's promoter was high in SMMC7721,but after the treatment of 5-aza-CdR, the promoter of PDCD4 was demethylation.Conclusions1. The expression of DNMT3b protein was decreased, while the PDCD4 was increased in hepatocellular carcinoma, their relationship was a negative correlation.2. Methylase inhibitor 5-aza-CdR can inhibit the DNMT3b expression in hepatocellular carcinoma, and the inhibition enhanced as the concentration increased; The expression level of PDCD4 increased as 5-aza-CdR inhibited the expression of DNMT3b.3. DNMT3b may control the expression of PDCD4 by influencing the methylation status of its promoter.