title: Macrophage-specific overexpression of antimicrobial peptide PR-39 inhibits intracellular growth of Salmonella enterica serovar Typhimurium in macrophage cells.
abstract: Although purified and synthesised PR-39 shows potent antibacterial effects in vitro, its ability to kill intracellular bacteria in macrophages, which are a major cause of refractory intracellular infection, has not yet been demonstrated. Both to enhance its antimicrobial potential and to reduce systemic side effects, it would be desirable to deliver PR-39 into macrophage cells and to limit its activation to the site of infection. To address this issue, PR-39 DNA was inserted into the eukaryotic expression plasmid pIRES2-EGFP and the adenoviral vector Ad-MSP, from which PR-39 can be specifically expressed in macrophage cells from the macrophage-specific promoter. pIRES2-EGFP/PR39 and Ad-MSP/PR-39 were either transduced or infected into macrophage RAW264.7 cells for stable or transient expression of PR-39. PR-39 expression in macrophage cells was subsequently confirmed by reverse transcription polymerase chain reaction (RT-PCR) and immunocytochemistry. Furthermore, its antimicrobial activity in macrophage cells was evaluated by colony enumeration assay. Results showed that the macrophage-specific promoter could initiate targeted expression of PR-39 only in macrophage RAW264.7 cells. Moreover, either stable or transient expression of PR-39 in macrophage cells conferred enhanced antimicrobial activity against Salmonella enterica serovar Typhimurium. Our results have demonstrated that macrophage-specific expression of antimicrobial peptide PR-39 in macrophages could inhibit the growth of intracellular S. Typhimurium and indicated it to be a novel and promising approach for the control of refractory intracellular infection.
authors.affiliation: Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Faculty of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
authors.forename: Yang-An
authors.initials: YA
authors.lastname: Wen
authors.forename: Xiao-Lin
authors.initials: XL
authors.lastname: Yu
authors.forename: Qian-Feng
authors.initials: QF
authors.lastname: Xia
authors.forename: Dong
authors.initials: D
authors.lastname: Cen
authors.forename: Jin-Bo
authors.initials: JB
authors.lastname: Liu
authors.forename: Zhi-Guang
authors.initials: ZG
authors.lastname: Tu
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D000890:Anti-Infective Agents / Q000378:metabolism* / Q000494:pharmacology; D023181:Antimicrobial Cationic Peptides / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D002460:Cell Line; D006801:Humans; D008264:Macrophages / Q000378:metabolism* / Q000382:microbiology* / Q000821:virology; D051379:Mice; D010957:Plasmids; D012486:Salmonella typhimurium / Q000187:drug effects* / Q000254:growth & development* / Q000472:pathogenicity; D014162:Transfection; D015854:Up-Regulation
chemical_list: D000890:Anti-Infective Agents; D023181:Antimicrobial Cationic Peptides; C072212:PR 39