title: [Zinc-dependent metalloprotease 1 promotes apoptosis of RAW264.7 macrophages].
abstract: OBJECTIVE
  To construct the eukaryotic expression vector of zinc-dependent metalloprotease 1 (zmp1) gene from Bacillus Calmette-Guerin (BCG) and investigate its impact on the apoptosis of RAW264.7 macrophages.
  
  
  METHODS
  Zmp1 gene was amplified from the genome of BCG by PCR. The zmp1 gene fragment was inserted into multiple cloning sites of pEGFP-N1 to construct the eukaryotic expression vector pEGFP-N1-zmp1. The constructed pEGFP-N1-zmp1 was transfected into RAW264.7 cells by Lipofectamine(TM) 2000. The expression of green fluorescent protein (GFP) was observed by fluorescence microscopy. The zmp1 mRNA was detected by quantitative real-time PCR (qR-PCR). The effect of Zmp1 protein on the apoptosis of RAW264.7 macrophages was detected by flow cytometry (FCM).
  
  
  RESULTS
  With zmp1 gene amplified by PCR, we successfully constructed the recombinant vector pEGFP-N1-zmp1 as demonstrated by restriction enzyme analysis and sequencing. GFP was seen in RAW264.7 cells 24 hours after transfected with the recombinant plasmid. As qRT-PCR showed, the expression level of zmp1 mRNA was up-regulated. The early apoptotic rate increased 48 hours after transfection.
  
  
  CONCLUSIONS
  The increased expression of Zmp1 in RAW264.7 cells promotes the apoptosis of RAW264.7 cells.
authors.affiliation: Center for Molecular Medicine and Tumor Research, Department of Pathogen Biology, School of Basic Medicine, Chongqing Medical University, Chongqing 400016, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Li
authors.affiliation: Center for Molecular Medicine and Tumor Research, Department of Pathogen Biology, School of Basic Medicine, Chongqing Medical University, Chongqing 400016, China. *Corresponding author.
authors.forename: Yonglin
authors.initials: Y
authors.lastname: He
authors.affiliation: Center for Molecular Medicine and Tumor Research, Department of Pathogen Biology, School of Basic Medicine, Chongqing Medical University, Chongqing 400016, China.
authors.forename: Jiming
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Center for Molecular Medicine and Tumor Research, Department of Pathogen Biology, School of Basic Medicine, Chongqing Medical University, Chongqing 400016, China.
authors.forename: Chencheng
authors.initials: C
authors.lastname: Fang
mesh_terms: D000818:Animals; D017209:Apoptosis; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D008264:Macrophages / Q000166:cytology*; D020781:Matrix Metalloproteinase 1 / Q000235:genetics / Q000378:metabolism*; D045726:Metalloproteases / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics; D000067996:RAW 264.7 Cells; D014376:Tuberculosis / Q000503:physiopathology*; D015032:Zinc / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D045726:Metalloproteases; C568073:Zmp1 protein, Mycobacterium tuberculosis; D020781:Matrix Metalloproteinase 1; D015032:Zinc