title: Polyethyleneimine-based core-shell nanogels: a promising siRNA carrier for argininosuccinate synthetase mRNA knockdown in HeLa cells.
abstract: RNA interference using small interfering RNA (siRNA) is a promising biological strategy for treatment of diverse diseases; however, application of siRNA is severely hindered by its poor stability and low cellular uptake efficiency. We have recently demonstrated that polyethyleneimine (PEI)-based amphiphilic core-shell particles have several distinguishing advantages over native PEI and its derivatives. This paper presents a novel type of PEI-based nanogels with a biodegradable gelatin core. The core-shell nanogels were synthesized via a two-stage reaction: (1) preparation of highly uniform gelatin nanoparticles through appropriate treatment of gelatin solution; and (2) conjugation of branched PEI to the preformed gelatin nanoparticles, followed by repeated cycles of desolvation and drying of the gelatin-PEI nanogels in ethanol/water mixture. The resulting nanogels have a well-defined nanostructure that contains a gelatin core and a PEI shell. They have an average diameter of 200 ± 40nm with high uniformity. The nanogel particles possess positive zeta-potential values of up to +40mV at neutral pH, indicating that they are highly positive and very stable in aqueous medium. The gelatin-PEI nanogels were able to completely condense siRNA at N/P ratios of as low as 5:1, and effectively protected siRNA against enzymatic degradation. Furthermore, the nanogels were four times less toxic than native PEI. Besides low toxicity, the nanogels were able to effectively deliver siRNA into HeLa cells. It was found that increasing the N/P ratio from 10 to 30 significantly increased the intracellular uptake efficiency of siRNA from 41 to 84%. Confocal laser scanning microscopic images confirmed that the nanogels were able to effectively deliver siRNA in the cytoplasm of HeLa cells. The delivered siRNA could inhibit 70% of human argininosuccinate synthetase 1 (ASS1) gene expression. This gene silencing percentage is much higher than that of the commercial Lipofectamine(TM) 2000. Our studies demonstrate that gelatin-PEI core-shell nanogels have promising potential to act as an effective siRNA carrier.
authors.affiliation: Department of Applied Biology and Chemical Technology, The Hong Kong Polytechnic University, Hung Hom, Kowloon, Hong Kong, P.R. China.
authors.forename: Hetti
authors.initials: H
authors.lastname: Mimi
authors.forename: Kin Man
authors.initials: KM
authors.lastname: Ho
authors.forename: Yuen Shan
authors.initials: YS
authors.lastname: Siu
authors.forename: Aihua
authors.initials: A
authors.lastname: Wu
authors.forename: Pei
authors.initials: P
authors.lastname: Li
mesh_terms: D001124:Argininosuccinate Synthase / Q000235:genetics*; D002470:Cell Survival; D005780:Gelatin / Q000008:administration & dosage / Q000737:chemistry; D055785:Gene Knockdown Techniques; D020868:Gene Silencing; D018014:Gene Transfer Techniques; D006367:HeLa Cells; D006801:Humans; D000080385:Nanogels; D011092:Polyethylene Glycols / Q000008:administration & dosage* / Q000737:chemistry; D011094:Polyethyleneimine / Q000008:administration & dosage* / Q000737:chemistry; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000737:chemistry
chemical_list: D000080385:Nanogels; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C413692:polyethylene glycol polyethyleneimine nanogel; D011092:Polyethylene Glycols; D005780:Gelatin; D011094:Polyethyleneimine; D001124:Argininosuccinate Synthase