Schulz Ronny M, Haberhauer Marcus, Zernia Göran, Pösel Claudia, Thümmler Christian, Somerson Jeremy S, Huster Daniel
Department of Cell Techniques and Applied Stem Cell Biology, Center of Biotechnology and Biomedicine, University of Leipzig, Deutscher Platz 5, D-04103, Leipzig, Germany; Translational Centre for Regenerative Medicine, University of Leipzig, Philipp-Rosenthal-Str. 55, D-04105, Leipzig, Germany.
J Tissue Eng Regen Med. 2014 Jul;8(7):566-77. doi: 10.1002/term.1561. Epub 2012 Jul 4.
Many synthetic polymers and biomaterials have been used as matrices for 3D chondrocyte seeding and transplantation in the field of cartilage tissue engineering. To develop a fully autologous carrier for chondrocyte cultivation, we examined the feasibility of allogeneic plasma and whole blood-based matrices and compared them to agarose constructs. Primary articular chondrocytes isolated from 12-month-old pigs were embedded into agarose, plasma and whole blood matrices and cultivated under static-free swelling conditions for up to four weeks. To evaluate the quality of the synthesized extracellular matrix (ECM), constructs were subjected to weekly examinations using histological staining, spectrophotometry, immunohistochemistry and biochemical analysis. In addition, gene expression of cartilage-specific markers such as aggrecan, Sox9 and collagen types I, II and X was determined by RT-PCR. Chondrocyte morphology was assessed via scanning electron microscopy and viability staining, including proliferation and apoptosis assays. Finally, (13) C NMR spectroscopy provided further evidence of synthesis of ECM components. It was shown that chondrocyte cultivation in allogeneic plasma and whole-blood matrices promoted sufficient chondrocyte viability and differentiation behaviour, resulting in neo-formation of a hyaline-like cartilage matrix.
在软骨组织工程领域，许多合成聚合物和生物材料已被用作三维软骨细胞接种和移植的基质。为了开发一种用于软骨细胞培养的完全自体载体，我们研究了同种异体血浆和全血基质的可行性，并将它们与琼脂糖构建体进行比较。从12月龄猪分离的原代关节软骨细胞被嵌入琼脂糖、血浆和全血基质中，并在无静态肿胀条件下培养长达四周。为了评估合成细胞外基质（ECM）的质量，每周使用组织学染色、分光光度法、免疫组织化学和生化分析对构建体进行检查。此外，通过逆转录聚合酶链反应（RT-PCR）测定软骨特异性标志物如聚集蛋白聚糖、Sox9以及I、II和X型胶原蛋白的基因表达。通过扫描电子显微镜和活力染色评估软骨细胞形态，包括增殖和凋亡分析。最后，碳-13核磁共振波谱（¹³C NMR）为ECM成分的合成提供了进一步的证据。结果表明，在同种异体血浆和全血基质中培养软骨细胞可促进足够的软骨细胞活力和分化行为，从而导致类透明软骨基质的新形成。
