Preliminary Study on the Feasibility of p53 Gene Delivery Mediated by Novel Vectors Constructed with CTS and PEI

Huang Yu; Luo Rui; Fang Yiping; Li Sha

Preliminary Study on the Feasibility of p53 Gene Delivery Mediated by Novel Vectors Constructed with CTS and PEI

Huang Yu, Luo Rui, Fang Yiping, Li Sha

Abstract

Cancer is one of the serious diseases threatening human health. Surgery, radiotherapy and chemotherapy are major conventional therapies for cancer treatment. The pain, toxic and side effects limit their efficacy. Gene therapy, which has promising potential in cancer therapy, attracts much interest. p53 gene is one of the genes found to have the highest correlation to human tumor. Chitosan (CTS) and polyethyleneimine (PEI) were reported to be potent carrier materials for preparing effective gene vectors delivering DNA and siRNA. In this work, novel vectors were developed using CTS and PEI together to explore the feasibility in delivery of p53 gene in Hela cells. The plasmid pEGFP-N1-p53 was incubated with CTS and PEI in different sequences to obtain two kinds of sequential ternary nanocomplexes, PEI/CTS/pEGFP-N1-p53 and CTS/PEI/pEGFP-N1-p53, which has a size of (109.2 ± 12.50) nm and (228.4 ± 15.78) nm, respectively. The cytotoxicity of the nanocomplex was measured by methy1 thiazoly1 tetrazolium (MTT) method and the transfection efficiency was investigated by fluorescence microscope and flow cytometry in Hela cells. Both PEI/CTS/pEGFP-N1-p53 and CTS/PEI/pEGFP-N1-p53 showed lower toxicity than PEI/pEGFP-N1-p53. Compared to the naked plasmid, positive control Lip2000/pEGFP-N1-p53 and binary nanocomplex PEI/pEGFP-N1-p53 and CTS/pEGFP-N1-p53, PEI/CTS/pEGFP-N1-p53 achieved the highest transfection rate of 65% and that of CTS/PEI/pEGFP-N1-p53 was second only to PEI/CTS/pEGFP-N1-P53 with a rate of 46%. The results suggested that combined use of CTS and PET as carrier materials constructed novel gene vectors of high transfection capacity and low cytotoxicity.

Keywords

pEGFP-N1-p53 plasmid, polyethyleneimine, chitosan, gene vector, toxicity, transfection, Hela cells

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