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STAT3-siRNA induced B16.F10 melanoma cell death: more association with VEGF downregulation than p-STAT3 knockdown.
Saudi Pharmaceutical Journal : SPJ : the Official Publication of the Saudi Pharmaceutical Society 2018 December
STAT3 knockdown by small interfering RNA (siRNA) has been described to inhibit carcinogenic growth in various types of tumors. Earlier we have reported delivery of siRNA by oleic acid- and stearic acid-modified-polyethylenimine and enhancement of silencing of STAT3 by small interfering RNA (siRNA) in B16.F10 melanoma cell lines and consequent tumor suppression. Present investigation mainly focused on the downstream events involved in B16.F10 melanoma cell death and consequent tumor suppression following knockdown of p-STAT3 by siRNA. Lipid-substituted polyethylenimine (PEI)-p-STAT3-siRNA were prepared and characterized by measuring its N/P ratio, zeta potential, size, association and dissociation with siRNA. B16.F10 melanoma cells were treated with six different concentrations of PEI-p-STAT3-siRNA (200, 100, 50, 25, 12.5 and 6.25 nM). Downregulation of p-STAT3 and VEGF were studied using western blot and ELISA in association with the melanoma cell death. PEI-p-STAT3-siRNA hydrodynamic diameter ranged from 110 to 270 nm. PEI assisted p-STAT3-siRNA delivery exhibited increased uptake by B16.F10, when analyzed by fluorescent and confocal microscopy along with flowcytometry. It induced concentration-dependent knockdown of the p-STAT3 that also downregulated VEGF expression in similar fashion and induced B16.F10 cell death. Higher concentrations of p-STAT3-siRNA appear to significantly downregulate the VEGF expression via p-STAT3 knockdown. Decreasing survival of B16.F10 cells with the increasing concentration of p-STAT3-siRNA significantly correlated with VEGF downregulation, not with p-STAT3 expression. Data suggest that VEGF downregulation following knockdown of p-STAT3 may be a key event in survival reduction in B16.F10 melanoma cells and.
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