Study On The Antimicrobial Activities Of The Culture Supernatant Of 293 Cells Transfected With Human Beta-defensin-2 Gene

Background: As the important mediators of natural immunity and molecular basis of protection from inflammation reaction, endogenous peptides are small molecule cationic polypeptides with broad-spectrum antimicrobial activities. Human defensins can be divided into two types as alpha-defensins and beta-defensins according to the location of cysteine residues and the connection of disulfide-bonding. Human beta-defensin (HBD) is cationic antimicrobial peptides consists of 34-42 amino acid residues with small molecular weights (3.5-4.5 kDa). The molecular is composed of 6 conservative cysteine residues which are proposed to form 3 disulphide bonds. Human beta-defensins are the important mediators of the mucosal protection system including respiratory, gastrointestinal, and urogenital tracts. 4 kinds of HBD have been found, and HBD-2 was found from psoriasis skin in 1997 and it is made up of 41 amino acid with abundant cationic amino acid and cysteine. Generally, the HBD-2 does not or seldom express in normal epithelia, but when it is infected, HBD-2 can be induced to express by Proinflammatory mediators such as TNF-α,IL-1β,LPS and bacteria. Patients with Cysitc fibrosis are likely to have pulmonary infection, and its mechanism is concerned with the restrain of sterilization of HBD-2 resulted from the gene defects of CFTR in respiratory tract. In the post-antibiotic age, the bacteria with drug resistance bring the new threats to our health. The unique mechanism of human defensins can prevent bacteria from drug resistance and so have bright future in pharmacy. With the current development of gene project, a certain scholars have researched the express of human defensin with prokayotic microorganism , eucaryotic microorganisms plant, COS-7 cells and etc.Objective: The objective of this paper is to assess the antimicrobial activities of the protein secreted from 293 cells transfected with human beta-defensin-2 gene and to provide the foundation of the further researches in bronchus epithelial HBD-2 gene transferr.Methods: HBD-2 eukaryotic expression recombinant plasmid pcDNA3.1 -zeo-HBD-2 was constructed by the molecular cloning technologies, and transfected 293 cells by liposome. The cell culture supernatants were collected and examined antimicrobial activities against several bacteria.Results: RT-PCR detection showed that HBD-2 gene expressed in the transfected 293 cells. The antimicrobial activities of the culture supernatant of the HBD-2 gene transfected- 239 cells were stronger than that of the control group.Conclusion: This study comfirmed that HBD-2 gene could transferred into and expressed in 239 cells. The culture supernatant of HBD-2 gene-transfected 293 cells had a increased antimicrobial activities.