title: [Cell death of THP-1 induced by puried Rv3671c protein of tuberculosis and the detection of TNF-α and IL-1β in Mycobacterium tuberculosis].
abstract: OBJECTIVE
  To assess the response in THP-1 treated with Rv3671c protein in Mycobacterium tuberculosis (M.tuberculosis).
  
  
  METHODS
  The gene encoding Rv3671c protein of M.tuberculosis was cloned into pET-28a vector and then expressed in Escherichia coli. The Rv3671c was purified with Ni-NTA affinity and ion exchange chromatography. The detection of protein concentration was by Lowry method.THP-1 cell was stimulated with Rv3671c protein and cells were analyzed by Hochest staining under fluorescence microscopy to assay cell death (apoptosis and necrosis). TNF-α and IL-1β were detected by ELISA at each stimulating time.
  
  
  RESULTS
  The Rv3671c protein of M.tuberculosis was successfully expressed in Escherichia coli. The purity of recombinant Rv3671c protein was 95%, and the protein concentration was up to 0.4 mg/ml. The nucleus of THP-1 was isolated and necrosis-like under fluorescence when cells were stimulated by Rv3671c protein. The levels of TNF-α and IL-1β in supernatant were 19 000 and 16 500 pg/ml respectively, and were significantly higher than control cells with the levels of 2100 and 3800 pg/ml separately.
  
  
  CONCLUSIONS
  The necrosis of THP-1 cells could be stimulated by Rv3671c protein of M.tuberculosis and it was probably associated with high cytokines TNF-α and IL-1β levels.
authors: The Fifth People's Hospital of Wuxi, Wuxi, China.
authors: Shou-gang
authors: SG
authors: Kuai
authors: Hao
authors: H
authors: Pei
authors: Li-hua
authors: LH
authors: Huang
authors: Zhong-hua
authors: ZH
authors: Liu
authors: Guang-liang
authors: GL
authors: Mai
authors: Jun
authors: J
authors: Liu
authors: Zhen-ling
authors: ZL
authors: Cui
mesh_terms: D001426:Bacterial Proteins / Q000494:pharmacology*; D016923:Cell Death; D002460:Cell Line; D006801:Humans; D053583:Interleukin-1beta / Q000378:metabolism; D008264:Macrophages / Q000166:cytology* / Q000378:metabolism*; D009169:Mycobacterium tuberculosis / Q000235:genetics*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D053583:Interleukin-1beta; D014409:Tumor Necrosis Factor-alpha