Harnessing endogenous growth factor activity modulates stem cell behavior
Harnessing endogenous growth factor activity modulates stem cell behavior
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DOI：
10.1039/c1ib00021g
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发表时间：
2011-01-01
影响因子：
2.5
通讯作者：
Murphy, William L.
中科院分区：
文献类型：
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作者：
Hudalla, Gregory A.;Kouris, Nicholas A.;Murphy, William L.
在传统的细胞培养中，由于血清生物分子吸附的随机性和非特异性，特定的血清生物分子(如肝素)对生长因子依赖的细胞行为的影响往往很难阐明。我们假设化学定义良好的细胞培养底物可用于研究隔离肝素对人间充质干细胞(HMSC)行为的影响。具体地说，我们使用生物惰性自组装单层(SAM)作为干细胞培养底物，这些单层膜经过生物灵感肝素结合肽(称为HEPpep)和整合素结合肽(RGDSP)化学修饰。我们的结果表明，纯化的肝素以剂量依赖的方式与HEPpep自组装膜结合，而血清携带的肝素以剂量依赖的方式与HEPpep自组装膜特异性结合。这些肝素隔离型自组装膜通过放大内源性成纤维细胞生长因子信号促进hMSC增殖，并通过放大内源性骨形态发生蛋白信号促进hMSC成骨分化。肝素隔离的效果类似于重组成纤维细胞生长因子-2的超生理浓度的效果。HMSC在生长介质中的肝素隔离底物上通过多次群体倍增保持表型，而在成骨诱导介质中培养过程中，hMSC在相同的底物上以骨形态发生蛋白依赖的方式促进成骨分化。综上所述，这些观察表明，基质对干细胞表型的影响对培养基配方很敏感。我们的结果还表明，通过构图HEPpep在底物上的位置，可以在空间上定位增强的hMSC的增殖。重要的是，在这项研究中使用的化学定义明确的自组装膜消除了随机的、非特异性的生物分子吸附的混杂因素，并确定血清携带的肝素是hMSC对内源性生长因子反应的关键介质。
The influence of specific serum-borne biomolecules (e. g. heparin) on growth factor-dependent cell behavior is often difficult to elucidate in traditional cell culture due to the random, non-specific nature of biomolecule adsorption from serum. We hypothesized that chemically well-defined cell culture substrates could be used to study the influence of sequestered heparin on human mesenchymal stem cell (hMSC) behavior. Specifically, we used bio-inert self-assembled monolayers (SAMs) chemically modified with a bioinspired heparin-binding peptide (termed "HEPpep") and an integrin-binding peptide (RGDSP) as stem cell culture substrates. Our results demonstrate that purified heparin binds to HEPpep SAMs in a dose-dependent manner, and serum-borne heparin binds specifically and in a dose-dependent manner to HEPpep SAMs. These heparin-sequestering SAMs enhance hMSC proliferation by amplifying endogenous fibroblast growth factor (FGF) signaling, and enhance hMSC osteogenic differentiation by amplifying endogenous bone morphogenetic protein (BMP) signaling. The effects of heparin-sequestering are similar to the effects of supraphysiologic concentrations of recombinant FGF-2. hMSC phenotype is maintained over multiple population doublings on heparin-sequestering substrates in growth medium, while hMSC osteogenic differentiation is enhanced in a bone morphogenetic protein-dependent manner on the same substrates during culture in osteogenic induction medium. Together, these observations demonstrate that the influence of the substrate on stem cell phenotype is sensitive to the culture medium formulation. Our results also demonstrate that enhanced hMSC proliferation can be spatially localized by patterning the location of HEPpep on the substrate. Importantly, the use of chemically well-defined SAMs in this study eliminated the confounding factor of random, non-specific biomolecule adsorption, and identified serum-borne heparin as a key mediator of hMSC response to endogenous growth factors.