Jiang Xue-Song, Chai Chou, Zhang Yue, Zhuo Ren-Xi, Mao Hai-Quan, Leong Kam W
Department of Chemistry, Wuhan University, Wuhan 430072, P. R. China.
Biomaterials. 2006 May;27(13):2723-32. doi: 10.1016/j.biomaterials.2005.12.001. Epub 2006 Jan 10.
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.
整合素与细胞外基质蛋白之间的相互作用在造血调控中发挥着重要作用。人类造血祖细胞表达极晚期抗原-4（VLA-4）和VLA-5，它们分别通过识别连接段-1（CS-1）和RGD基序介导与纤连蛋白的相互作用。在本研究中，我们调查了人脐带血（UCB）CD34+细胞在表面固定有含CS-1结合基序（EILDVPST）和RGD基序（GRGDSPC）的肽的合成底物上的体外扩增情况。这些肽以2.0 - 2.3 nmol/cm2的表面密度共价偶联到聚对苯二甲酸乙二酯（PET）膜上。UCB CD34+细胞在补充有重组人血小板生成素、干细胞因子、flt3配体和白细胞介素3的无血清培养基中培养10天。在CS-1肽修饰的表面上观察到最高的细胞扩增倍数，与未扩增的细胞相比，总核细胞、总集落形成单位和长期培养起始细胞分别扩增了589.6±58.6（平均值±标准差）、76.5±8.8和3.2±0.9倍。所有表面固定有肽的底物，包括对照肽，在支持CD34+、CFU-GEMM和LTC-ICs的扩增方面都比组织培养聚苯乙烯表面更有效。然而，从600个CD34+细胞进行10天的体外扩增后，只有在固定有CS-1的表面上培养的细胞产生了阳性植入，尽管频率较低。固定有RGD肽的PET表面比固定有CS-1肽的表面效率低。我们的结果表明，共价固定的黏附肽可显著影响培养的UCB CD34+细胞的增殖特性。
