Surface-immobilization of adhesion peptides on substrate for ex vivo expansion of cryopreserved umbilical cord blood CD34+ cells.
The interaction between integrins and extracellular matrix proteins play an important role in the regulation of hematopoiesis. Human hematopoietic progenitor cells express very late antigen-4 (VLA-4) and VLA-5, which mediate their interaction with fibronectin by recognizing the connecting segment-1 (CS-1 and RGD motifs, respectively. In this study, we investigated the ex vivo expansion of human umbilical cord blood (UCB) CD34+ cells on synthetic substrates surface-immobilized with peptides containing the CS-1 binding motif (EILDVPST) and the RGD motif (GRGDSPC). These peptides were covalently conjugated to poly(ethylene terephthalate) (PET) film at a surface density of 2.0-2.3 nmol/cm2. UCB CD34+ cells were cultured for 10 days in serum-free medium supplemented with recombinant human thrombopoietin, stem cell factor, flt3-ligand and interleukin 3. The highest cell expansion fold was observed on the CS-1 peptide-modified surface, where total nucleated cells, total colony forming unit, and long-term culture initiating cells were expanded by 589.6+/-58.6 (mean+/-s.d.), 76.5+/-8.8, and 3.2+/-0.9-fold, respectively, compared to unexpanded cells. All substrates surface-immobilized with peptides, including the control peptides, were more efficient in supporting the expansion of CD34+, CFU-GEMM and LTC-ICs than tissue culture polystyrene surface. Nevertheless, after 10-days of ex vivo expansion from 600 CD34+ cells, only cells cultured on CS-1-immobilized surface yielded positive engraftment, even though the frequency was low. PET surface immobilized with RGD peptide was less efficient than that with CS-1 peptide. Our results suggest that covalently immobilized adhesion peptides can significantly influence the proliferation characteristics of cultured UCB CD34+ cells.
Jiang
Xue-Song
XS
Department of Chemistry, Wuhan University, Wuhan 430072, P. R. China.
Chai
Chou
C
Zhang
Yue
Y
Zhuo
Ren-Xi
RX
Mao
Hai-Quan
HQ
Leong
Kam W
KW
D000818:Animals; D018952:Antigens, CD34 / Q000276:immunology*; D002448:Cell Adhesion / Q000187:drug effects; D015815:Cell Adhesion Molecules / Q000737:chemistry / Q000494:pharmacology; D018929:Cell Culture Techniques / Q000379:methods*; D049109:Cell Proliferation / Q000187:drug effects; D020099:Coated Materials, Biocompatible / Q000737:chemistry / Q000494:pharmacology*; D015925:Cryopreservation / Q000379:methods*; D018380:Hematopoietic Stem Cell Transplantation / Q000379:methods; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000276:immunology; D006801:Humans; D051379:Mice; D016513:Mice, SCID; D009842:Oligopeptides / Q000737:chemistry / Q000494:pharmacology*; D011485:Protein Binding; D013499:Surface Properties; D014470:Umbilical Cord / Q000166:cytology* / Q000187:drug effects / Q000502:physiology
D018952:Antigens, CD34; D015815:Cell Adhesion Molecules; D020099:Coated Materials, Biocompatible; D009842:Oligopeptides; C047981:arginyl-glycyl-aspartic acid