title: Impact of culture conditions on the proliferative lifespan of human T cells in vitro.
abstract: BACKGROUND
  In human T cells, telomerase is transiently expressed upon activation and stimulation and, as shown previously, telomerase levels are able to control the lifespan of T cells. To improve T-cell expansion it is of critical importance to understand the effects of culture parameters on telomerase activity and lifespan.
  
  
  METHODS
  We investigated the influence of culture condition (FCS, human AB serum and autologous serum) and stimulation (PHA/feeder cells, anti-CD3/CD28 beads) on the lifespan, clonogenicity (number of positive wells), cell cycle, telomerase activity and telomere length of T cells in vitro.
  
  
  RESULTS
  The proliferative lifespan of T cells expanded with PHA/feeder cells and autologous serum from different donors was doubled compared with stimulation with PHA/feeder cells and AB serum. No or only a small difference was found for T cells expanded with anti-CD3/CD28 beads and autologous or AB serum. The use of autologous serum also increased the clonogenicity to about three-fold compared with the use of AB serum or FCS, without any signs of differences in the fractions of cycling cells. Interestingly, T cells cultured with autologous serum exhibited a significantly higher telomerase activity at day 6 after stimulation and a reduced decline of telomerase activity compared with cultures with AB serum.
  
  
  CONCLUSIONS
  The use of autologous serum combined with PHA stimulation and feeder cells remarkably extends the proliferative lifespan and clonogenicity and increases the telomerase activity of human T cells in vitro. This might be useful for applications where large numbers of specific T cells are required.
authors.affiliation: Department of Hematology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany. alexander.roeth@uni-due.de
authors.forename: A
authors.initials: A
authors.lastname: Röth
authors.forename: L
authors.initials: L
authors.lastname: Schneider
authors.forename: H
authors.initials: H
authors.lastname: Himmelreich
authors.forename: G M
authors.initials: GM
authors.lastname: Baerlocher
authors.forename: U
authors.initials: U
authors.lastname: Dührsen
mesh_terms: D000818:Animals; D002417:Cattle; D018929:Cell Culture Techniques / Q000379:methods; D002453:Cell Cycle / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D003470:Culture Media / Q000737:chemistry / Q000494:pharmacology*; D005312:Fetal Blood / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008934:Mitogens / Q000494:pharmacology; D010835:Phytohemagglutinins / Q000494:pharmacology; D044967:Serum / Q000378:metabolism; D013601:T-Lymphocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D019098:Telomerase / Q000378:metabolism; D016615:Telomere / Q000235:genetics / Q000378:metabolism
chemical_list: D003470:Culture Media; D008934:Mitogens; D010835:Phytohemagglutinins; D019098:Telomerase