a Department of Physiology, Faculty of Medicine , University of Malaya , Kuala Lumpur , Malaysia.
b Tissue Engineering Group (TEG), National Orthopaedic Centre of Excellence in Research and Learning (NOCERAL), Department of Orthopaedic Surgery, Faculty of Medicine , University of Malaya , Kuala Lumpur , Malaysia.
Platelets. 2019;30(1):66-74. doi: 10.1080/09537104.2017.1371287. Epub 2017 Nov 1.
Platelet-rich concentrate (PRC), used in conjunction with other chondroinductive growth factors, have been shown to induce chondrogenesis of human mesenchymal stromal cells (hMSC) in pellet culture. However, pellet culture systems promote cell hypertrophy and the presence of other chondroinductive growth factors in the culture media used in previous studies obscures accurate determination of the effect of platelet itself in inducing chondrogenic differentiation. Hence, this study aimed to investigate the effect of PRC alone in enhancing the chondrogenic differentiation potential of human mesenchymal stromal cells (hMSC) encapsulated in three-dimensional alginate constructs. Cells encapsulated in alginate were cultured in serum-free medium supplemented with only 15% PRC. Scanning electron microscopy was used to determine the cell morphology. Chondrogenic molecular signature of hMSCs was determined by quantitative real-time PCR and verified at protein levels via immunohistochemistry and enzyme-linked immunosorbent assay. Results showed that the cells cultured in the presence of PRC for 24 days maintained a chondrocytic phenotype and demonstrated minimal upregulation of cartilaginous extracellular matrix (ECM) marker genes (SOX9, TNC, COL2, ACAN, COMP) and reduced expression of chondrocyte hypertrophy genes (Col X, Runx2) compared to the standard chondrogenic medium (p < 0.05). PRC group had correspondingly higher levels of glycosaminoglycan and increased concentration of chondrogenic specific proteins (COL2, ACAN, COMP) in the ECM. In conclusion, PRC alone appears to be very potent in inducing chondrogenic differentiation of hMSCs and offers additional benefit of suppressing chondrocyte hypertrophy, rendering it a promising approach for providing abundant pool of chondrogenic MSCs for application in cartilage tissue engineering.
富血小板浓缩物（PRC）与其他软骨诱导生长因子联合使用已被证明可在微球培养中诱导人间充质基质细胞（hMSC）的软骨生成。然而，微球培养系统促进细胞肥大，并且先前研究中使用的培养介质中存在其他软骨诱导生长因子，这掩盖了血小板本身在诱导软骨分化中的准确作用。因此，本研究旨在研究 PRC 单独在增强包裹在三维藻酸盐构建体中的人间充质基质细胞（hMSC）的软骨分化潜能方面的作用。将包裹在藻酸盐中的细胞在补充有 15% PRC 的无血清培养基中培养。扫描电子显微镜用于确定细胞形态。通过定量实时 PCR 确定 hMSC 的软骨发生分子特征，并通过免疫组织化学和酶联免疫吸附测定法在蛋白质水平上进行验证。结果表明，在 PRC 存在下培养 24 天的细胞保持软骨细胞表型，并显示软骨细胞外基质（ECM）标志物基因（SOX9、TNC、COL2、ACAN、COMP）的最小上调和软骨细胞肥大基因（Col X、Runx2）的表达降低，与标准软骨形成培养基相比（p < 0.05）。PRC 组的 ECM 中糖胺聚糖和软骨特异性蛋白（COL2、ACAN、COMP）的浓度相应增加。总之，PRC 单独似乎非常有效地诱导 hMSC 的软骨分化，并提供抑制软骨细胞肥大的额外益处，使其成为为软骨组织工程应用提供丰富的软骨生成 MSC 来源的有前途的方法。
