Bioengineering, University of California, Berkeley, Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley Nanoscience and Nanoengineering Institute, Berkeley, California 94720, United States.
Biomacromolecules. 2011 Apr 11;12(4):987-96. doi: 10.1021/bm1013475. Epub 2011 Feb 23.
Specific biochemical and physical cues in tissue extracellular matrices play a critical role in regulating cellular growth processes and their fate. We report initial responses of bone stem cells induced by collagen-derived DGEA-peptides on nanofibrous M13 phage tissue matrices. We constructed genetically engineered M13 phage with DGEA-peptide displayed in high density on the major coat proteins and biomimetic nanofibrous tissue-like matrices in two and three dimensions. We investigated the effects of biochemical cues, specifically DGEA-peptides on preosteoblast (MC3T3) morphologies. The preosteoblasts grown on the top of the DGEA-incorporated phage matrices exhibited significant outgrown morphology with early bone cell marker protein expression. Through soluble peptide competition assays and control experiments, we verified that the observed cellular morphologies and osteogenic protein marker expression were specifically caused by the DGEA-peptides. We confirmed that the outgrown morphologies are linked with the early phase of osteogenic protein expression through mRNA quantification and bone cell protein marker expression. Additionally, we demonstrated that the phage-based tissue matrix systems could work as a good cell culture platform to investigate the specific effect of biochemical cues, which can be tuned precisely at a single amino acid level with little change in other physical and chemical properties of the environment. Our study advances the understanding of osteogenic differentiation and our phage-based tissue matrices have the potential for future bone regeneration therapy and systemic investigation of specific cellular responses to biochemical ligand stimulation.
组织细胞外基质中的特定生化和物理线索在调节细胞生长过程及其命运方面起着关键作用。我们报告了胶原衍生的 DGEA-肽对纳米纤维 M13 噬菌体组织基质中骨干细胞的初始反应。我们构建了基因工程 M13 噬菌体，在主要衣壳蛋白上高度展示 DGEA-肽，并在二维和三维空间中仿生组织样纳米纤维基质。我们研究了生化线索，特别是 DGEA-肽对成骨前体细胞（MC3T3）形态的影响。在掺入 DGEA 的噬菌体基质顶部生长的成骨前体细胞表现出明显的向外生长形态，并表达早期骨细胞标记蛋白。通过可溶性肽竞争实验和对照实验，我们证实了观察到的细胞形态和成骨蛋白标志物表达是由 DGEA-肽特异性引起的。我们通过 mRNA 定量和骨细胞蛋白标志物表达证实了向外生长的形态与成骨蛋白表达的早期阶段有关。此外，我们证明了噬菌体组织基质系统可以作为一个很好的细胞培养平台来研究生化线索的特定影响，这些影响可以在不改变环境其他物理和化学性质的情况下，在单个氨基酸水平上进行精确调整。我们的研究推进了对成骨分化的理解，我们的噬菌体组织基质具有未来骨再生治疗和系统研究特定细胞对生化配体刺激反应的潜力。
