Wang Hr

Targeting Smad4 links microRNA-146a to the TGF-β pathway during retinoid acid induction in acute promyelocytic leukemia cell line

The expression pattern of microRNAs (miRNAs) and their potential target genes were investigated in acute promyelocytic leukemia (APL) cell line NB4 cells during all-trans-retinoid acid (ATRA) treatment by using a miRNA microarrays platform and real-time quantitative PCR (RTQ-PCR). MiR-146a as one of the miRNAs down-regulated by ATRA during APL differentiation was identified. Direct interaction between miR146a and its predictive target gene Smad4 were confirmed by Luciferase assay. Down-regulation of miR-146a and upregulation of Smad4 at protein levels were demonstrated. These data suggested that miR-146a might influence proliferation of APL cells through TGF-β1/Smad signal transduction pathway during ATRA induction.

Modification of TGF-β1 signaling pathway during NB4 cells differentiation by all-trans retinoid acid induction

The aim of the study was to present the possible mechanisms of transforming growth factor beta 1(TGF-β1) signal pathway during cell differentiation by studying the expression levels of six components of TGF-β1 pathway (TGF-β1, two TGF-β1 receptors and three Smad proteins). The morphology change, the CD11 expression levels, and the mRNA and protein expression levels of TGF-β1, TGF-β ReceptorI (TβRI), TGF-β ReceptorII (TβRII), Smad2, Smad4 and Smad7 were assessed by exposing NB4 cells to all-trans retinoid acid (ATRA) using Wright’s stain, flow cytometry, real-time PCR assay and Western blot analysis. The mRNA and protein expression levels of all six components increased during NB4 cells differentiation induced by ATRA. They were most significantly increased after 24–72 h individually when cells were induced by ATRA (the mRNA and protein expression levels of TGF-β1, TβRI, TβRII and Smad2 reached their peaks at 48 and 48 h individually after the treatment, Smad4 at 48 and 72 h, and Smad7 at 72 and 72 h). The change in mRNA expression levels was earlier than the change in the same gene controlling protein. These results indicate that the upregulation of TGF-β1 pathway plays an important role in NB4 cells differentiation induced by ATRA.

PNAS-2: a novel gene probably participating in leukemogenesis.

Objective:As4S4 is an effective drug for the treatment of acute promyelocytic leukemia but its mechanism of action remains largely unknown. In a previous study, we identified PNAS-2, a human apoptosis-related protein gene, using gene expression profiling. In this study, we tried to clarify the role of PNAS-2 in apoptosis and leukemogenesis. Methods: NB4 and U937 leukemia cell lines and serial clinical samples were studied. RNA interference (RNAi) and RNA overexpression were used to address the potential role of PNAS-2 in apoptosis. PNAS-2 expression was examined using Northern blot in multiple tissues, and real-time PCR was applied to analyze PNAS-2 expression in various patient samples. Results: Functional analyses of PNAS-2 by RNAi and RNA overexpression indicate PNAS-2 is an anti-apoptosis gene. PNAS-2 expression is significantly increased in de novo or relapsed acute leukemia, but in patients in complete remission PNAS-2 levels decrease to levels comparable to those found in normal controls. In carcinomas, PNAS-2 expression was not upregulated, indicating that PNAS-2 overexpression was specific for leukemia. Conclusion: Based on the preliminary data, we suggest that the PNAS-2 gene functions as an anti-apoptotic gene and probably participates in leukemogenesis.