RESUMEN
OBJECTIVE: To explore functions of the histone H3 lysine 79 (K79) methyltransferase Dot1L in the development of pancreatic cancer and evaluate the possibility of targeting Dot1L to inhibit pancreatic cancer progression. METHODS: Patient samples were used to detect differences in Dot1L expression between tumor and adjacent tissues and to determine correlations between Dot1L expression in patients with different stages of pancreatic cancer. Lentiviral-mediated knockdown of Dot1L expression and flow cytometry were used to detect apoptosis in pancreatic cancer lacking Dot1L expression; chromatin immunoprecipitation and quantitative PCR were used to detect downstream target genes of Dot1L. RESULTS: We show that Dot1L is highly expressed in pancreatic cancer, and that its expression is related to pancreatic cancer stage. Knocking down Dot1L significantly promoted apoptosis in pancreatic cancer cells, while overexpressing Dot1L inhibited apoptosis. Mechanistically, Dot1L regulated apoptosis in pancreatic cancer cells by promoting NUPR1 expression. The enriched H3K79 trimethylation in the transcription initiation region of NUPR1 promoted its expression. Overexpressing NUPR1 inhibited the pancreatic cancer cell apoptosis caused by Dot1L knockdown. CONCLUSIONS: Dot1L inhibits pancreatic cancer cell apoptosis by targeting NUPR1; thus, Dot1L is a promising target for pancreatic cancer treatment.
Asunto(s)
Histonas , Neoplasias Pancreáticas , Apoptosis/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , N-Metiltransferasa de Histona-Lisina/genética , Histonas/genética , Histonas/metabolismo , Humanos , Proteínas de Neoplasias , Neoplasias Pancreáticas/genéticaRESUMEN
In this study, the mechanism by which Ad-p27mt inhibits the growth, invasion and metastasis of transplanted liver tumor was studied by examining the effects of Ad-27mt gene transfer on the expression of Bax, Bcl-2, VEGF and MMP-9 in the transplanted liver tumors in nude mice. The model of transplanted hepatic tumor was established in nude mice. The mice were then divided into three groups, which were injected with PBS, Ad-LacZ and Ad-p27mt and the growth of the transplanted liver tumor was observed. The expressions of P27, Bax and Bcl-2 proteins were detected by Western blotting and the expressions of VEGF and MMP-9 were immunohistochemically determined. Our result showed that the tumor size, expressions of Bax, Bcl-2 proteins, VEGF and MMP-9 were all lower than those in PBS and Ad-LacZ groups and the differences were statistically significant (P<0.05). Our study suggested that Ad-p27mt could inhibit the growth, invasion and metastasis of hepatic cancer by lowering the expressions of VEGF and MMP-9.
RESUMEN
Objective To assess the effects of p27mt gene transfection on the proliferation and apoptosis of human hepatocellular carcinoma cell (HCC) lines SMMC-7721. Methods A replication deficient adenovirus vector encoding p27mt (Ad-p27mt) was used and p27mt cDNA was transfected into human SMMC-7721 cell lines in vitro. The synthesis of DNA in SMMC-7721 cells was determined by using 3H-thymidine incorporation; the cell apoptosis was determined by flow cytometry, TUNEL method and DNA fragmentation analysis. Results The virus titer was 7.95?1012 cfu/ml, the transduction efficiency was 100 % when multiplicity of infection ≥50, FCM analysis revealed a sub-G1 cell peak in Ad-p27mt transduced hepatocellular carcinoma cell lines. Agarose electrophoresis showed marked ladder .The difference of apoptotic index between the Ad-p27mt group and the control group was statistically significant (58.6?4.3, vs 4.5?1.6, P