In vitro bioactivity and gene silencing effect of shRNA-VEGF loaded chitosan nanoparticles

Abstract

Purpose: In this study, it is aimed to prepare chitosan nanoparticles containing shRNA-VEGF and evaluate their bioactivity by in vitro cell culture studies and to perform mechanical characterization of nanoparticles. Material and Methods: Ionic chelation method was used to prepare nanoparticles. The XTT assay was used to assess the cytotoxic activity of shRNA-VEGF and shRNA-VEGF loaded NP on the HeLa and NIH 3T3 cells. Results: According to the results IC50 values of shRNA-VEGF and NP including shRNA-VEGF were calculated. IC50 values of shRNA-VEGF and NP including shRNA-VEGF were 0.89±0.010 µg/mL and 0.52±0.004 µg/mL on HeLa cell line. Bax quantities of control, shRNA-VEGF, and shRNA-VEGF loaded NP was measured as 23.70±0.27 ng/mg protein, 34.64±0.36 ng/mg protein, and 39.46±0.54 ng/mg protein, respectively. According to the results, cleaved caspase 3 quantities of control, shRNA-VEGF, and shRNA-VEGF loaded NP was measured as 711.70±4.40 pg/mg protein, 767.23±3.82 pg/mg protein, and 825.32±5.06 pg/mg protein, respectively. Conclusion: shRNA-VEGF and shRNA-VEGF loaded NP significantly reduced HeLa cell reproduction in a concentration-dependent manner while generating no cytotoxicity in NIH 3T3 cells. The expression of pro-apoptotic Bax and cleaved caspase 3 proteins was significantly increased by shRNA-VEGF and shRNA-VEGF loaded NP.

Keywords

• shRNA-VEGF
• Nanoparticle
• RNA interference
• Cell viability
• ELISA kits

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