Naqvi Syeda Masooma, Buckley Conor Timothy
1 Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute, Trinity College Dublin , Dublin, Ireland .
Tissue Eng Part A. 2015 Jan;21(1-2):288-99. doi: 10.1089/ten.TEA.2013.0719. Epub 2014 Sep 8.
Cell-based therapies may hold significant promise for the treatment of early stage degeneration of the intervertebral disc (IVD). Given their propensity to proliferate and ability to form multiple tissue types, mesenchymal stem cells (MSCs) have been proposed as a potential cell source to promote repair of the nucleus pulposus (NP). However, for any successful cell-based therapy, a carrier biomaterial may be essential for targeted delivery providing key biophysical and biochemical cues to facilitate differentiation of MSCs. Two widely used biomaterials for NP regeneration are chitosan and alginate. The primary objective of this study was to assess the influence of alginate and chitosan hydrogels on bone marrow stem cells (BM) and NP cells in isolation or in coculture. A secondary objective of this study was to investigate coculture seeding density effects of BM and NP cells and simultaneously explore which cell type is responsible for matrix formation in a cocultured environment. Porcine NP and BM cells were encapsulated in alginate and chitosan hydrogels separately at two seeding densities (4×10(6) and 8×10(6) cells/mL) or in coculture (1:1, 8×10(6) cells/mL). Constructs (diameter=5 mm, height=3 mm) were maintained under IVD-like conditions [low-glucose, low (5%) oxygen] with or without transforming growth factor-β3 (TGF-β3) supplementation for 21 days. Results demonstrated differential viability depending on hydrogel type. NP cells remained viable in both biomaterial types whereas BM viability was diminished in chitosan. Further, hydrogel type was found to regulate sulfated glycosaminoglycan (sGAG) and collagen accumulation. Specifically, alginate better supports sGAG accumulation and collagen type II deposition for both NP and BM cell types compared with chitosan. Having identified that alginate more readily supports cell viability and matrix accumulation, we further explored additional effects of seeding density ratios (NP:BM--1:1, 1:2) for coculture studies. Interestingly, in coculture conditions, the BM cell population declined in number while NP cells increased, indicating that MSCs may in fact be signaling NP cells to proliferate rather than contributing to matrix formation. These findings provide exciting new insights on the potential of MSCs for NP tissue regeneration strategies.
基于细胞的疗法可能对治疗早期椎间盘退变具有重大前景。鉴于间充质干细胞（MSCs）具有增殖倾向和形成多种组织类型的能力，它们已被提议作为促进髓核（NP）修复的潜在细胞来源。然而，对于任何成功的基于细胞的疗法，载体生物材料对于靶向递送可能至关重要，它能提供关键的生物物理和生化线索以促进MSCs的分化。两种广泛用于NP再生的生物材料是壳聚糖和藻酸盐。本研究的主要目的是评估藻酸盐和壳聚糖水凝胶对骨髓干细胞（BM）和NP细胞单独培养或共培养时的影响。本研究的次要目的是研究BM和NP细胞的共培养接种密度效应，并同时探索在共培养环境中哪种细胞类型负责基质形成。将猪NP细胞和BM细胞分别以两种接种密度（4×10⁶和8×10⁶个细胞/毫升）封装在藻酸盐和壳聚糖水凝胶中，或进行共培养（1:1，8×10⁶个细胞/毫升）。构建体（直径 = 5毫米，高度 = 3毫米）在类似椎间盘的条件下[低糖、低（5%）氧]培养，添加或不添加转化生长因子-β3（TGF-β3），培养21天。结果表明，细胞活力因水凝胶类型而异。NP细胞在两种生物材料类型中均保持活力，而壳聚糖中的BM活力则降低。此外，发现水凝胶类型可调节硫酸化糖胺聚糖（sGAG）和胶原蛋白的积累。具体而言，与壳聚糖相比，藻酸盐能更好地支持NP细胞和BM细胞类型的sGAG积累和II型胶原蛋白沉积。在确定藻酸盐更易于支持细胞活力和基质积累后，我们进一步探索了共培养研究中接种密度比（NP:BM - 1:1、1:2）的其他影响。有趣的是，在共培养条件下，BM细胞数量减少而NP细胞增加，这表明MSCs实际上可能是在向NP细胞发出增殖信号，而不是参与基质形成。这些发现为MSCs在NP组织再生策略中的潜力提供了令人兴奋的新见解。
