Department of Orthopaedic Surgery, CAPHRI school for public health and primary care, Maastricht University Medical Center, P.O. Box 5800, 6202 AZ, Maastricht, The Netherlands.
Bone. 2013 Mar;53(1):94-102. doi: 10.1016/j.bone.2012.11.032. Epub 2012 Dec 4.
Previous studies have shown that human articular chondrocytes in vitro are osmolarity-responsive and increase matrix synthesis under cartilage-specific physiological osmolarity. The effects of increased osmolarity on chondrogenesis of progenitor cells in vitro are largely unknown. We therefore aimed to elucidate whether hyperosmolarity facilitates their chondrogenic differentiation and whether Nfat5 is involved.
ATDC5 cells and human bone marrow stem cells (hBMSCs) were differentiated in the chondrogenic lineage in control and increased osmolarity conditions. Chondrogenic outcome was measured by gene- and protein expression analysis. RNAi was used to determine the role of Nfat5 in chondrogenic differentiation under normal and increased osmolarity.
Increasing the osmolarity of differentiation medium with 100mOsm resulted in significantly increased chondrogenic marker expression (Col2a1, Col10a1, Acan, Sox9, Runx2 and GAGs) during chondrogenic differentiation of the two chondroprogenitors, ATDC5 and hBMSCs. Nfat5 knockdown under both control and increased osmolarity affected chondrogenic differentiation and suppressed the osmolarity-induced chondrogenic induction. Knockdown of Nfat5 in early differentiation significantly decreased early Sox9 expression, whereas knockdown of Sox9 in early differentiation did not affect early Nfat5 expression.
Increasing the osmolarity of chondrogenic culture media by 100mOsm significantly increased chondrogenic gene expression during the course of chondrogenic differentiation of progenitor cells. Nfat5 may be involved in regulating chondrogenic differentiation of these cells under both normal and increased osmolarities and might regulate chondrogenic differentiation through influencing early Sox9 expression.
先前的研究表明，体外培养的人关节软骨细胞对渗透压有反应，在软骨特异性生理渗透压下增加基质合成。增加渗透压对体外前体细胞的软骨形成的影响在很大程度上尚不清楚。因此，我们旨在阐明渗透压是否有利于其软骨分化，以及 Nfat5 是否参与其中。
在对照和渗透压增加条件下，将 ATDC5 细胞和人骨髓基质干细胞（hBMSCs）分化为软骨谱系。通过基因和蛋白质表达分析来测量软骨形成的结果。使用 RNAi 来确定 Nfat5 在正常和增加渗透压下的软骨分化中的作用。
在分化培养基中增加 100mOsm 的渗透压导致两种软骨前体细胞（ATDC5 和 hBMSCs）的软骨形成过程中软骨形成标志物的表达（Col2a1、Col10a1、Acan、Sox9、Runx2 和 GAGs）显著增加。在对照和渗透压增加条件下，Nfat5 的敲低均影响软骨分化，并抑制渗透压诱导的软骨诱导。早期分化时 Nfat5 的敲低显著降低了早期 Sox9 的表达，而早期分化时 Sox9 的敲低并不影响早期 Nfat5 的表达。
在祖细胞的软骨形成过程中，将软骨形成培养基的渗透压增加 100mOsm 可显著增加软骨形成基因的表达。Nfat5 可能参与调节这些细胞在正常和增加渗透压下的软骨分化，并可能通过影响早期 Sox9 的表达来调节软骨分化。
