title: Effect of cell density and HLA-DR incompatibility on T-cell proliferation and forkhead box P3 expression in human mixed lymphocyte reaction.
abstract: BACKGROUND
  The proliferation rates of human T cells in vitro are affected by some factors such as initial T-cell number, dose of stimulating cells, and duration of culture. The transcription factor forkhead box P3 (FoxP3) has been used to identify regulatory T cells in humans and is thought to correlate with tolerance to allogeneic organ transplant. Thus, it is important to optimize conditions to expand FoxP3 cell proliferation to improve engraftment of allogeneic organ transplants.
  
  
  METHODS
  We studied proliferative responses and FoxP3 expression in divided T cells with the use of flow cytometric analysis of Ki-67 in culture of different concentrations of responding cells (6 × 10(6), 4 × 10(6), 2 × 10(6), 1 × 10(6), and 0.5 × 10(6)cells/mL), different types of stimulating cells (lymphocytes and low density cells), and different numbers of HLA mismatches.
  
  
  RESULTS
  The proportion of CD3(+) cells, CD4(+)CD25(+) cells, and CD4(+)CD25(+)FoxP3(+) cells among mononuclear cells were highest at initial cell concentration of 2 × 10(6) responder cells/mL with lymphocytes as stimulators at day-5 mixed lymphocyte reaction (MLR). They were highest at a concentration of 4 × 10(6) responder cells/mL with low density cells as stimulators. The recovery (%), proportion of CD3(+) cells, CD4(+)CD25(+) cells, and CD4(+)CD25(+)FoxP3(+) cells with 2 HLA-DR incompatibility were significantly higher than those of 1 HLA-DR incompatibility at day-5 MLR.
  
  
  CONCLUSIONS
  Initial cell concentration and HLA-DR incompatibility can affect the generation of FoxP3+ T cells in human MLR. These factors could be considered for efficient generation of Tregs for clinical trials in the future.
authors.affiliation: Department of Pathology, University of California, San Diego, La Jolla, California; Department of Laboratory Medicine, Seoul National University College of Medicine, Seoul, Korea; Department of Molecular Medicine and Biopharmaceutical Sciences, Graduate School of Convergence Science and Technology and College of Medicine, Medical Research Center, Seoul National University, Seoul, Korea. Electronic address: eysong1@snu.ac.kr.
authors.forename: E Y
authors.initials: EY
authors.lastname: Song
authors.affiliation: Department of Pathology, University of California, San Diego, La Jolla, California.
authors.forename: S
authors.initials: S
authors.lastname: Han
authors.affiliation: Department of Pathology, University of California, San Diego, La Jolla, California.
authors.forename: B
authors.initials: B
authors.lastname: Yang
authors.affiliation: Department of Pathology, University of California, San Diego, La Jolla, California.
authors.forename: G P
authors.initials: GP
authors.lastname: Morris
authors.affiliation: Department of Pathology, University of California, San Diego, La Jolla, California.
authors.forename: J D
authors.initials: JD
authors.lastname: Bui
mesh_terms: D015415:Biomarkers / Q000378:metabolism; D002452:Cell Count; D049109:Cell Proliferation; D005434:Flow Cytometry; D051858:Forkhead Transcription Factors / Q000378:metabolism*; D006684:HLA-DR Antigens / Q000276:immunology*; D006801:Humans; D019394:Ki-67 Antigen / Q000378:metabolism; D008213:Lymphocyte Activation; D007959:Lymphocyte Culture Test, Mixed; D050378:T-Lymphocytes, Regulatory / Q000276:immunology* / Q000502:physiology
chemical_list: D015415:Biomarkers; C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors; D006684:HLA-DR Antigens; D019394:Ki-67 Antigen