Service of Orthopaedic Surgery and Traumatology, Lausanne University Hospital, University of Lausanne, Switzerland.
Regenerative Therapy Unit, Lausanne University Hospital, University of Lausanne, Switzerland.
Cartilage. 2021 Dec;13(1_suppl):509S-518S. doi: 10.1177/19476035211035433. Epub 2021 Jul 30.
A pivotal aspect of cartilage tissue engineering resides in cell culture medium supplementation, in view of maximizing cell proliferation and preserving cellular functionality. Autologous human serum (aHS) is commonly used as an inducive supplement for safe human articular chondrocyte (HAC) proliferation prior to clinical implantation. However, practical clinical use of aHS is hindered by constraining manufacturing requirements and quality assurance-driven downstream processing. The present study investigated potential alternative use of commercial human platelet lysate (hPL) supplements in HAC manufacturing workflows related to clinical therapeutic pathways.
Differential effects of hPL, aHS, and fetal bovine serum were assessed on primary cultured HAC parameters (viability, proliferative rates, and morphology) in 2-dimensional systems. A 3-dimensional HAC pellet model served for postexpansion assessment of cellular functionality, by visualizing proteoglycan production (Alcian blue staining), and by using qRT-PCR relative quantification of chondrogenic marker (, and ) genetic expression.
We found that monolayer HAC culture with hPL or aHS supplements presented similar characteristics (elongated cell morphology and nearly identical growth kinetics). Chondrogenic activity appeared as conserved in HACs expanded with human or bovine supplements, wherein histologic analysis indicated a progressive sGAG accumulation and gene expression was upregulated in 3-dimensional HAC pellet models.
This study therefore supports the use of hPL as a functional equivalent and alternative to aHS for cultured HAC batch preparation, with the potential to effectively alleviate pressure on clinical and manufacturing bottlenecks in cell therapy approaches for cartilage regeneration.
软骨组织工程的一个关键方面在于细胞培养基的补充，以最大限度地促进细胞增殖并保持细胞功能。自体人血清（aHS）通常用作临床植入前安全的人关节软骨细胞（HAC）增殖的诱导补充剂。然而，aHS 的实际临床应用受到限制制造要求和质量保证驱动的下游加工的限制。本研究调查了商业人血小板裂解液（hPL）补充剂在与临床治疗途径相关的 HAC 制造工作流程中的潜在替代用途。
评估 hPL、aHS 和胎牛血清对原代培养 HAC 参数（活力、增殖率和形态）的二维系统中的影响。三维 HAC 微球模型用于细胞功能的扩展后评估，通过可视化糖胺聚糖（GAG）产生（阿利新蓝染色），并通过 qRT-PCR 相对定量软骨形成标志物（、和）的基因表达。
我们发现，使用 hPL 或 aHS 补充剂的单层 HAC 培养具有相似的特征（伸长的细胞形态和几乎相同的生长动力学）。用人类或牛补充剂扩展的 HAC 中的软骨形成活性似乎保持不变，组织学分析表明 sGAG 积累逐渐增加，3 维 HAC 微球模型中的 基因表达上调。
因此，本研究支持使用 hPL 作为 aHS 的功能等效物和替代品，用于培养 HAC 批次制备，这有可能有效地缓解细胞治疗方法在软骨再生方面的临床和制造瓶颈的压力。
