Department of Anatomy and Neurobiology, Northeastern Ohio Universities Colleges of Medicine and Pharmacy, Rootstown, Ohio 44272, USA.
Tissue Eng Part A. 2010 Jul;16(7):2183-96. doi: 10.1089/ten.TEA.2009.0717.
Osteoarthritis (OA) is a prevalent age-associated disease involving altered chondrocyte homeostasis and cartilage degeneration. The avascular nature of cartilage and the altered chondrocyte phenotype characteristic of OA severely limit the capacity for in vivo tissue regeneration. Cell- and tissue-based repair has the potential to revolutionize treatment of OA, but those approaches have exhibited limited clinical success to date. In this study, we test the hypothesis that bovine and human chondrocytes in a collagen type I scaffold will form hyaline cartilage ex vivo with immunohistochemical, biochemical, and magnetic resonance (MR) endpoints similar to the original native cartilage. Chondrocytes were isolated from 1- to 3-week-old calf knee cartilage or from cartilage obtained from human total knee arthroplasties, suspended in 2.7 mg/mL collagen I, and plated as 300 microL spot cultures with 5 x 10(6) each. Medium formulations were varied, including the amount of serum, the presence or absence of ascorbate, and treatments with cytokines. Bovine chondrocytes generated metachromatic territorial and interstitial matrix and accumulated type II collagen over time. Type VI collagen was confined primarily to the pericellular region. The ex vivo-formed bovine cartilage contained more chondroitin sulfate per dry weight than native cartilage. Human chondrocytes remained viable and generated metachromatic territorial matrix, but were unable to support interstitial matrix accumulation. MR analysis of ex vivo-formed bovine cartilage revealed evidence of progressively maturing matrix, but MR-derived indices of tissue quality did not reach those of native cartilage. We conclude that the collagen-spot culture model supports formation and maturation of three-dimensional hyaline cartilage from active bovine chondrocytes. Future studies will focus on determining the capacity of human chondrocytes to show comparable tissue formation.
骨关节炎（OA）是一种与年龄相关的常见疾病，涉及到软骨细胞稳态的改变和软骨退化。软骨的无血管特性和 OA 特征性的软骨细胞表型改变严重限制了体内组织再生的能力。基于细胞和组织的修复有可能彻底改变 OA 的治疗方法，但迄今为止，这些方法的临床效果有限。在这项研究中，我们检验了这样一个假设，即在 I 型胶原支架中的牛和人软骨细胞将形成具有类似于原始天然软骨的免疫组织化学、生化和磁共振（MR）终点的透明软骨。从 1 至 3 周大的小牛膝关节软骨或从全膝关节置换术中获得的软骨中分离出软骨细胞，悬浮在 2.7mg/mL 的 I 型胶原中，并以 300μL 的斑点培养物接种，每个培养物中含有 5×10(6)个细胞。调整了培养基配方，包括血清的含量、是否存在抗坏血酸以及细胞因子的处理。牛软骨细胞随着时间的推移生成了变色的基质和间质基质，并积累了 II 型胶原。VI 型胶原主要局限于细胞周区域。体外形成的牛软骨的每干重软骨素硫酸盐含量高于天然软骨。人软骨细胞保持活力并生成变色的基质，但无法支持间质基质的积累。对体外形成的牛软骨的 MR 分析显示了基质逐渐成熟的证据，但 MR 衍生的组织质量指数尚未达到天然软骨的水平。我们得出结论，胶原斑点培养模型支持从活跃的牛软骨细胞形成和成熟三维透明软骨。未来的研究将集中于确定人软骨细胞形成类似组织的能力。
