School of Bioscience and Engineering, Jadavpur University, Kolkata, India.
J Mater Chem B. 2021 Jun 23;9(24):4873-4894. doi: 10.1039/d1tb00314c.
The use of decellularized native allogenic or xenogenic cartilaginous extracellular matrix (ECM) biomaterials is widely expanding in the fields of tissue engineering and regenerative medicine. In this study, we aimed to develop an acellular, affordable, biodegradable, easily available goat conchal cartilaginous ECM derived scaffolding biomaterial for repair and regeneration of osteochondral defects in rabbits. Cartilages harvested from freshly collected goat ears were decellularized using chemical agents, namely, hypotonic-hypertonic (HH) buffer and Triton X-100 solution, separately. The morphologies and ultrastructure orientations of the decellularized cartilages remained unaltered in spite of complete cellular loss. Furthermore, when the acellular cartilaginous ECMs were cultured with murine mesenchymal stem cells (MSCs) (C3H10T1/2 cells), cellular infiltration and proliferation were thoroughly monitored using SEM, DAPI and FDA stained images, whereas the MTT assay proved the biocompatibility of the matrices. The increasing amounts of secreted ECM proteins (collagen and sGAG) indicated successful chondrogenic differentiation of the MSCs in the presence of the treated cartilage samples. In vivo biocompatibility studies showed no significant immune response or tissue rejection in the treated samples but tissue necrosis in control samples after 3 months. Upon implantation of the constructs in rabbits' osteochondral defects for 3 months, the histological and micro-CT evaluation revealed significant enhancement and regeneration of neocartilage and subchondral bony tissues. The IGF-1 loaded cartilaginous constructs showed comparatively better healing response after 3 months. Our results showed that decellularized xenogenic cartilaginous biomaterials preserved the bioactivity and integrity of the matrices that also favored in vitro stem cell proliferation and chondrogenic differentiation and enabled osteochondral regeneration, thus paving a new way for articular cartilage reconstruction.
脱细胞化的天然同种异体或异种软骨细胞外基质 (ECM) 生物材料在组织工程和再生医学领域的应用正在广泛扩展。在这项研究中，我们旨在开发一种无细胞、经济实惠、可生物降解、易于获得的山羊耳壳软骨细胞外基质衍生支架生物材料，用于修复和再生兔的骨软骨缺损。使用化学剂（低渗-高渗 (HH) 缓冲液和 Triton X-100 溶液）分别对从新鲜采集的山羊耳软骨中提取的软骨进行脱细胞处理。尽管完全去除了细胞，但脱细胞软骨的形态和超微结构取向保持不变。此外，当无细胞软骨 ECM 与鼠间充质干细胞 (MSCs)（C3H10T1/2 细胞）共培养时，使用 SEM、DAPI 和 FDA 染色图像彻底监测细胞浸润和增殖，而 MTT 测定证明了基质的生物相容性。分泌的 ECM 蛋白（胶原和 sGAG）的数量增加表明 MSC 在处理过的软骨样本存在的情况下成功地向软骨分化。体内生物相容性研究表明，在处理过的样本中没有明显的免疫反应或组织排斥，但在对照组样本中 3 个月后出现组织坏死。在兔骨软骨缺损中植入构建体 3 个月后，组织学和 micro-CT 评估显示新软骨和软骨下骨组织的显著增强和再生。IGF-1 负载的软骨构建体在 3 个月后显示出相对更好的愈合反应。我们的结果表明，脱细胞化的异种软骨生物材料保留了基质的生物活性和完整性，这也有利于体外干细胞增殖和软骨分化，并促进骨软骨再生，为关节软骨重建开辟了新途径。
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