No abstract

Bcl-2 is an anti-apoptotic protein. If the level of Bcl-2 protein can be reduced sufficiently in tumors using RNA interference (RNAi) to target the gene message, the apoptosis of tumor cells may be promoted. In this study, we synthesized 19 nucleotides (nts) small interference RNA (siRNA) constructs suppressing bcl-2 gene expression in human tumor cells (HeLaB2 and BGC-823 cell lines) in vitro. The bcl-2 gene expression levels were significantly reduced when these siRNA were transfected into experimental two tumor cells for 72 hours. The apoptosis process was also examined in the tumor cells. Here we synthesized siRNA from a DNA template under the control of the RNA polymerase III promoter in transfected tumor cells. Using this DNA vector-based approach, we found that the siRNA efficiently and specifically inhibited the synthesis of protein encoded by the bcl-2 gene in HeLaB2 and BGC-823 tumor cells. Tumor growth was inhibited by 66.5% with 2mg/kg pSilencer 3.1H1-bcl-2 in mouse liver tumor-bearing BALB/c mice. This approach may prove to be a valuable clinical technique for the analysis of specific gene functions and gene therapy of malignant tumors that utilize the bcl-2 gene via RNA interference.

The Wnt/beta-catenin pathway is a conserved signalling pathway that regulates gene expression and controls diverse developmental processes such as cell fate specification, cell proliferation and cell differentiation. To our knowledge, the potential role of this pathway in the adult ovary has been poorly addressed. To this end, we investigated the expression pattern of Frizzled 2 in the mouse ovary during oestrous cycle by real-time Q-PCR, in situ hybridization, Western blotting and immunohistochemistry. In this study, we used uterine wet weight and H3.2 mRNA level as two markers for classification of mouse oestrous cycle. We found that both uterine wet weight and H3.2 mRNA are the highest in the proestrus and decrease from oestrus to diestrus. During oestrous cycle, Frizzled 2 mRNA and protein exhibited the highest level in the proestrus stage and rapidly reduced from oestrus to diestrus. In situ hybridization results showed that the positive signals for Frizzled 2 are highly detected in the oocyte and stroma in proestrus stage, while moderate or weak Frizzled 2 mRNAs are localized in the oocyte, granulosa cells, stroma and corpus luteum from oestrus to diestrus. The localization pattern of Frizzled 2 protein is mostly consistent with its mRNA, but stronger Frizzled 2 proteins are present in the granulosa cells and membrane of oocyte in oestrous cycle. Our data suggest that Frizzled 2 may be involved in regulating follicle growth, oocyte maturation and luteinization during oestrous cycle.