RESUMO
Objective: Epigenetic alterations, including any change in DNA methylation pattern, could be the missing link of understanding radiation-induced genomic instability. Dapper, Dishevelled-associated antagonist of ß-catenin homolog 2 (DACT2) is a tumor suppressor gene regulating Wnt/ß-catenin. In hepatocellular carcinoma (HCC), DACT2 is hypermethylated, while methylation status of its promoter regulates the corresponding expression. Radionuclides have been used to reduce proliferation and induce apoptosis in cancerous cells. Epigenetic impact of radionuclides as therapeutic agents for treatment of HCC is still unknown. The aim of this study was to evaluate epigenetic impact of 188Rhenium perrhenate (188ReO4) on HCC cells. Materials and Methods: In this in vitro experimental study, HepG2 and Huh7 cells were treated with 188ReO4, receiving 55 and 73 Mega Becquerel (MBq) exposures, respectively. For cell viability measurement, live/dead staining was carried out 18, 24, and 48 hours post-exposure. mRNA expression level of ß-Catenin, Wnt1, DNMT1, DACT2 and WIF- 1 genes were quantified by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Then, possible regulatory impact of DACT2 upregulation was investigated through evaluating methylation-specific PCR (MS-PCR). Results: Results showed that viability of both cells was reduced after treatment with 188ReO4 at three time points postexposure compared to the control groups. The qRT-PCR results showed that DACT2 mRNA level was significantly increased at 24, and 48 hours post-exposure in HepG2 cells compared to the control group, while, no significant change was observed in Huh7 cells. Methylation pattern of DACT2 promoter remained unchanged in HepG2 and Huh7 cells. Conclusion: Treatment with 188ReO4 reduced viability of HepG2 and Huh7 cells. Although DACT2 expression was increased after 188ReO4 exposure in HepG2 cells, methylation pattern of its promoter remained unchanged. This study assessed impacts of the 188ReO4 ß-irradiation on expression and induction of DACT2 epigenetic aberrations as well as the correlation of this agent with viability of cells.
RESUMO
BACKGROUND: Betatrophin is a member of the angiopoietin-like (ANGPTL) family that has been implicated in both triglyceride and glucose metabolism. The physiological functions and molecular targets of this protein remain largely unknown; hence, a purified available protein would aid study of the exact role of betatrophin in lipid or glucose metabolism. METHODS: In this study, we cloned the full-length cDNA of betatrophin from a human liver cDNA library. Betatrophin was expressed in the pET-21b-E. coli Bl21 (DE3) system and purified by immobilized metal-affinity chromatography and ion-exchange chromatography. RESULTS: Circular dichroism spectroscopy revealed α-helix as the major regular secondary structure in recombinant betatrophin. CONCLUSION: The production method is based on commonly available resources; therefore, it can be readily implemented.
