Functional TNFalpha gene silencing mediated by polyethyleneimine/TNFalpha siRNA nanocomplexes in inflamed colon.

摘要

During inflammatory bowel disease, TNFalpha is the major pro-inflammatory cytokine mainly secreted from macrophages and dendritic cells. Here, we have demonstrated that TNFalpha siRNA/polyethyleneimine loaded into polylactide at an optimal concentration of 20 g/L nanoparticles covered with polyvinyl alcohol are efficiently taken up by inflamed macrophages and inhibit TNFalpha secretion by the macrophages. Those nanoparticles have a diameter of approximately 380 nm and zeta potential of -8 mV at pH 7.2, and are non-cytotoxic. Complexation, interactions and protection from RNAse between TNFalpha siRNA and polyethyleneimine were higher than those using chitosan. Importantly, complexation between TNFalpha siRNA and polyethyleneimine facilitated higher rates of siRNA loading into nanoparticles, compared to Chi or free siRNA mixed with Lipofectamine. Oral administration of encapsulated TNFalpha siRNA-loaded nanoparticles specifically reduced the TNFalpha expression/secretion in colonic tissue in LPS-treated mice. In conclusion, we have shown: (1) that proposed TNFalpha siRNA-loaded NPs are prepared via a non-denaturing synthetic process; (2) a high encapsulation rate of TNFalpha siRNA complexed to polyethyleneimine into NPs; (3) effective enzymatic protection of TNFalpha siRNA by polyethyleneimine; (4) non-cytotoxicity and biodegradability of nanoparticles loaded with polyethyleneimine/TNFalpha siRNA; and (5) in vitro and in vivo significant anti-inflammatory effects at low TNFalpha siRNA dose that is specific and restricted to the colonic cells. Our results collectively indicate that polyethyleneimine/TNFalpha siRNA nanocomplexes represent an efficient therapeutic option for diseases such as IBD.