Signal transmission of TGF-beta superfamily in chondrocyte

从关节重建的临床角度出发，探讨了关节面修复的方法。利用人工培养的软骨细胞进行组织工程是修复关节的方法之一。利用人工培养的软骨细胞修复关节，重要的问题是软骨细胞的特性，特别是其作为关节软骨细胞的表型。换句话说，希望使用的软骨细胞不会骨化。考虑到软骨细胞的这些特性，我们研究了tgf - β超家族在软骨细胞中的信号传递。在本研究中，我们研究了phorbol， 12-肉豆蔻酸酯，13-乙酸酯（PMA），一种有效的特异性蛋白激酶C（PKC）激活剂和Staurosporine，一种有效的非选择性PKC抑制剂，对13日龄鸡胚胸骨软骨细胞分化的影响，以及PKC是否作为其作用的中介。以硫酸酸化蛋白多糖（PG）和I型和II型胶原作为分化标志物。免疫印迹法检测PKC表达。PMA对软骨形成的抑制呈剂量依赖性（1-50 ng/ml），其作用可能是由PKC从胞浆转移到膜上介导的。Staurosporine促进软骨形成（1-80nM），并以剂量依赖性的方式引起总PKC的消耗。一氧化氮合酶（NOS）特异性抑制剂n -单甲基，l -精氨酸（NMA）对Staurosporine（4和80 nM）的协同刺激PG合成，并相应抑制NO合成。PMA对PG合成的影响不受NMA的调节。我们认为NO可能不参与PMA对鸡胸骨软骨细胞分化的抑制，但Staurosporine可能至少部分通过NO依赖机制促进软骨细胞分化

From a clinical aspect of joint reconstruction, we have investigated how to repair the joint surface. One of the methods to repair the joint is tissue engineering using cultured chondrocytes which has developed markedly.To repair the joint using cultured chondrocytes, important problem is character of chondrocyte, especially their phenotype as articuler chondrocytes. In other words, it is desirable that the used chondrocytes never ossifies. Considering these character of chondrocytes, we investigated signal transmission of TGF-beta superfamily in chondrocytes. In the present study we examined the effect of phorbol, 12-myristate, 13-acetate (PMA), a potent and specific protein kinase C(PKC) activator and Staurosporine, a potent nonselective PKC inhibitor, on the differentiation of sternal chondrocytes from 13-day-old chick embryo, and whether PKC functions as a mediator of their effects. Sulfated proteoglycan (PG) and type I and type II collagen were used as markers of differentiation. PKC expression was assayd by immunoblotting. PMA inhibited chondrogenesis dose-dependently (1-50 ng/ml), and its effects were possibly mediated by the translocation of PKC from the cytozol to the membrane. Staurosporine enhanced chondrogenesis (1-80nM) and caused the depletion of total PKC in a dose-dependent fashion. A synergistic stimulation of PG synthesis by N-monomethyl, L-arginine (NMA), a specific inhibitor of nitric oxide synthase (NOS), in response to Staurosporine (4 and 80 nM), with a corresponding suppression of NO synthesis, has been observed. The effect PMA on PG synthesis was not modulated by NMA.We suggest that NO may no be involved in the inhibition of chick sternal chondrocyte differentiation by PMA,but that Staurosporine may promote chondrogenic differentiation at least in part through a NO-dependent mechanism