title: Influence of IL-3 functional fragment on cord blood stem cell ex vivo expansion and differentiation.
abstract: BACKGROUND
  Recombinant human interleukin-3 (rhIL-3) is a multiple hematopoietic growth factor, which enhances stem cell expansion and hematopoiesis regeneration in vitro and in vivo, when administrated in combination with other cytokines. However, the structure-function study of rhIL-3 remains rarely studied, so far. The purpose of this study was to recognize the short peptide with similar function as rhIL-3, and assess the hematopoietic efficacy in umbilical cord blood (UCB) stem cell culture as well.
  
  
  METHODS
  Two novel monoclonal antibodies (mAb) (C1 and E1) were generated against rhIL-3 using hybridoma technique. Eleven short peptides were depicted and synthesized to overlap covering the full length sequence of rhIL-3. ELISA was employed to distinguish the antibody-binding peptide from the negative peptides. In addition, the multi-potential hematopoiesis capabilities of the positive peptides were evaluated by adding 25 ng/mL of each peptide to the culture medium of hematopoietic stem cells (HSCs) derived from UCB. Total nucleated cell number and the CD34(+) cell number from each individual treatment group were calculated on day 7. Correlated antibodies at 0.5 or 2 molar fold to each peptide were also tested in the stem cell expansion experiment, to further confirm the bioactivity of the peptides.
  
  
  RESULTS
  Two peptides were recognized by the novel generated antibodies, using ELISA. Peptide 3 and 8 exhibited comparable hematopoiesis potentials, with 25.01±0.14 fold, and 19.89±0.12 fold increase of total nucleated cell number on day 7, respectively, compared with the basal medium control (4.93±0.55 fold). These biological effects were neutralized by adding the corresponding mAb at a dose dependent manner.
  
  
  CONCLUSIONS
  Our results identified two specific regions of rhIL-3 responsible for HSC proliferation and differentiation, which were located from 28 to 49 amino acids (P3), and 107 to 127 amino acids (P8), respectively. The short peptide 3 and 8 might act synergistically, which could serve as an economic substitute to rhIL-3 in research laboratory.
authors.affiliation: 1 Biopharmaceutical R&D Center, Chinese Academy of Medical Sciences & Peking Union Medical College, Suzhou 215126, China ; 2 Biopharmagen Corp., Suzhou 215126, China.
authors.forename: Zhihua
authors.initials: Z
authors.lastname: Ren
authors.affiliation: 1 Biopharmaceutical R&D Center, Chinese Academy of Medical Sciences & Peking Union Medical College, Suzhou 215126, China ; 2 Biopharmagen Corp., Suzhou 215126, China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: 1 Biopharmaceutical R&D Center, Chinese Academy of Medical Sciences & Peking Union Medical College, Suzhou 215126, China ; 2 Biopharmagen Corp., Suzhou 215126, China.
authors.forename: Yanxi
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: 1 Biopharmaceutical R&D Center, Chinese Academy of Medical Sciences & Peking Union Medical College, Suzhou 215126, China ; 2 Biopharmagen Corp., Suzhou 215126, China.
authors.forename: Wenhong
authors.initials: W
authors.lastname: Jiang
authors.affiliation: 1 Biopharmaceutical R&D Center, Chinese Academy of Medical Sciences & Peking Union Medical College, Suzhou 215126, China ; 2 Biopharmagen Corp., Suzhou 215126, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Dai
authors.affiliation: 1 Biopharmaceutical R&D Center, Chinese Academy of Medical Sciences & Peking Union Medical College, Suzhou 215126, China ; 2 Biopharmagen Corp., Suzhou 215126, China.
authors.forename: Xinxin
authors.initials: X
authors.lastname: Ding
authors.affiliation: 1 Biopharmaceutical R&D Center, Chinese Academy of Medical Sciences & Peking Union Medical College, Suzhou 215126, China ; 2 Biopharmagen Corp., Suzhou 215126, China.
authors.forename: Yongping
authors.initials: Y
authors.lastname: Jiang
keywords: CD34+; expansion, human recombinant interleukin-3 (rhIL-3); monoclonal antibody (mAb); peptide; umbilical cord blood stem cell (UCB stem cell)