title: [Th2 differentiation features of Mycobacterium tuberculosis heat resistant antigen-activated human γδT cells and the regulation of transcription factor T-bet/GATA-3 on differentiation].
abstract: OBJECTIVE
  To investigate Th2 differentiation features of Mycobacterium tuberculosis heat resistant antigens (MTB-HAg)-activated human γδT cells and the regulation of transcription factor T-box expression in T cells (T-bet) and GATA-binding protein 3 (GATA-3) on differentiation.
  
  
  METHODS
  Peripheral blood mononuclear cells (PBMCs) were stimulated with MTB-HAg to generate MTB-HAg-activated T cells (MTBAT) and expanded in the neutral condition or Th2 polarizing condition. After restimulation for 6 hours with phorbol myristate acetate (PMA, 10 ng/mL), ionomycin (500 ng/mL) and monensin (2.5 μmol/L), intracellular cytokines (IFN-γ, IL-4) of γδT cells and αβT cells among MTBAT were detected by four-color fluorescence mAb staining combined with flow cytometry. The highly purified γδT cells and CD4⁺ T cells were sorted by flow cytometer from MTBAT that were cultured in neutral and Th2 polarizing conditions for 28 days. The expressions of T-bet and GATA-3 mRNA in purified γδT cells and CD4⁺ T cells were determined by reverse transcription PCR (RT-PCR) technique.
  
  
  RESULTS
  γδT cells among MTBAT cultured in the neutral or Th2 polarizing condition predominantly produced IFN-γ, whereas the percentage of IFN-γ⁺ αβT cells significantly decreased in the Th2 polarizing condition as the culture time went by. Compared with the neutral condition, Th0 type (IFN-γ⁺ IL-4⁺) γδT cells significantly increased, and Th2 type (IFN-γ⁻ IL-4⁺) αβT cells also significantly increased in the Th2 polarizing condition. RT-PCR showed that mRNA expression of T-bet was still at a high level in γδT cells that were expanded in the Th2 polarizing condition, but at a low level in Th2 polarized CD4⁺ T cells. Moreover, the mRNA expressions of GATA-3 in both Th2 polarized γδT cells and CD4⁺T cells were up-regulated.
  
  
  CONCLUSIONS
  In the Th2 polarizing condition, the majority of γδT cells in MTBAT still remained Th1 profile, whereas the portion of γδT cells differentiated into Th0 type cells. Both transcription factor T-bet and GATA-3 failed to display a fully cross-regulation function in Th2 polarized γδT cells.
authors.affiliation: Department of Clinical Laboratory, First Affiliated Hospital, Bengbu Medical College, Bengbu 233004; Department of Immunology, Bengbu Medical College, Bengbu 233030, China.
authors.forename: Anyou
authors.initials: A
authors.lastname: Zhu
authors.affiliation: Department of Clinical Laboratory, First Affiliated Hospital, Bengbu Medical College, Bengbu 233004, China.
authors.forename: Hezuo
authors.initials: H
authors.lastname: Lv
authors.affiliation: Department of Clinical Laboratory, First Affiliated Hospital, Bengbu Medical College, Bengbu 233004, China.
authors.forename: Lunjun
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Department of Clinical Laboratory, First Affiliated Hospital, Bengbu Medical College, Bengbu 233004, China.
authors.forename: Jianguo
authors.initials: J
authors.lastname: Hu
authors.affiliation: Department of Clinical Laboratory, First Affiliated Hospital, Bengbu Medical College, Bengbu 233004, China.
authors.forename: Fengchao
authors.initials: F
authors.lastname: Wang
authors.affiliation: Department of Immunology, Bengbu Medical College, Bengbu 233030; Anhui Key Laboratory of Infection and Immunity, Bengbu 233030, China.
authors.forename: Baiqing
authors.initials: B
authors.lastname: Li
mesh_terms: D000328:Adult; D000942:Antigens, Bacterial / Q000276:immunology*; D002454:Cell Differentiation; D016764:Cell Polarity; D005260:Female; D050990:GATA3 Transcription Factor / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007371:Interferon-gamma / Q000276:immunology; D015847:Interleukin-4 / Q000276:immunology; D008213:Lymphocyte Activation; D008297:Male; D008875:Middle Aged; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000276:immunology*; D020825:T-Box Domain Proteins / Q000235:genetics* / Q000276:immunology; D016176:T-Lymphocyte Subsets / Q000276:immunology*; D018418:Th2 Cells / Q000166:cytology* / Q000276:immunology; D014376:Tuberculosis / Q000235:genetics / Q000276:immunology* / Q000382:microbiology; D000098815:T-bet Transcription Factor
chemical_list: D000942:Antigens, Bacterial; D050990:GATA3 Transcription Factor; C494749:GATA3 protein, human; D020825:T-Box Domain Proteins; D000098815:T-bet Transcription Factor; D015847:Interleukin-4; D007371:Interferon-gamma