Induced apoptosis in human prostate cancer cells by blocking of vascular endothelial growth factor by siRNA
A Deezagi1, S Ansari-Majd, N Vaseli-Hagh
1Department of Biochemistry, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran. deezagi@nigeb.ac.ir
Introduction:
Vascular endothelial growth factor (VEGF) regulates several cell functions including; proliferation, differentiation, permeability, vascular tone, and the production of vasoactive molecules. The purpose of this study was to evaluate the potency of specific short-interfering RNA (siRNA) to suppress human VEGF expression by siRNA and investigate the effects of VEGF down-regulation on the cell proliferation and apoptosis of the human prostate cancer cell lines DU-145.
Methods:
Transfection was performed using X-tremeGENE siRNA transfection reagent. At different time intervals, transfected cells were harvested and total RNA was extracted for RT-PCR. The VEGF content in supernatants were measured by ELISA. Inhibition of cell growth by hVEGF-siRNA was measured by using cell proliferation ELISA BrdU assay. Apoptotic cells were evaluated by using annexin-V-FITC apoptotic detection method.
Results:
Transfection of hVEGF-siRNA resulted in statistically significant inhibition of hVEGF-mRNA that in turn caused a marked reduction in the expression of hVEGF. The cell growth was assessed every 24 h for 4 days after siRNA treatment resulted in a marked inhibition of cell proliferation as compared to scramble siRNA. The results of apoptosis showed that approximately 15 % of the cells treated with control-siRNA manifested evident apoptotic changes after 24 hpt, whereas DU-145 cells treated with hVEGF-siRNA significantly were positive, that is to say, 53 % at 72 hpt 23.9 ± 2.78 % (P < 0.001) and 13 ± 1.57 % at 96 hpt.
Conclusion:
Our findings indicate that siRNA are effective in eliciting the RNAi pathway in cancerous cells and that specific siRNA efficiently down-regulate VEGF expression. They could decrease VEGF production and induce apoptosis, which may also be linked to the inhibition of cancerous cell proliferation. Therefore, it can be concluded that siRNA-mediated suppression of VEGF represents a powerful tool against prostate cancer cell proliferation. VEGF down-regulation exerts a direct anti-apoptotic function in the DU-145 cell lines and promises the development of drugs for cancer therapy.
Insights
Short-interfering RNA (siRNA) effectively suppresses vascular endothelial growth factor (VEGF) in prostate cancer cells. This VEGF down-regulation inhibits cell proliferation and significantly increases apoptosis, offering a promising therapeutic strategy.
Area of Science:
- Molecular Biology
- Cancer Research
- Gene Silencing
Background:
- Vascular Endothelial Growth Factor (VEGF) is crucial for cell proliferation, differentiation, and vascular permeability.
- Dysregulation of VEGF is implicated in various cancers, including prostate cancer.
- Targeting VEGF offers a potential therapeutic avenue for cancer treatment.
Purpose of the Study:
- To evaluate the efficacy of specific short-interfering RNA (siRNA) in suppressing human VEGF expression.
- To investigate the impact of VEGF down-regulation on the proliferation and apoptosis of DU-145 prostate cancer cells.
Main Methods:
- Transfection of DU-145 cells with human VEGF siRNA (hVEGF-siRNA) using X-tremeGENE reagent.
- Quantification of VEGF mRNA and protein levels via RT-PCR and ELISA.
- Assessment of cell proliferation using BrdU assay and apoptosis using Annexin-V-FITC staining.
Main Results:
- hVEGF-siRNA significantly inhibited VEGF mRNA and protein expression in DU-145 cells.
- A marked reduction in prostate cancer cell proliferation was observed following siRNA treatment.
- siRNA-mediated VEGF suppression led to a significant increase in cellular apoptosis (53% at 72h post-transfection).
Conclusions:
- siRNA effectively induces the RNA interference pathway to down-regulate VEGF expression in cancer cells.
- VEGF down-regulation by siRNA inhibits prostate cancer cell proliferation and promotes apoptosis.
- siRNA-mediated VEGF suppression is a potent strategy for targeting prostate cancer and developing novel therapies.
