Adipocyte differentiation of ST-13 cells induced by fibronectin fragment

研究了纤维连接蛋白(Fib1)结构域片段(Fib1)诱导ST-13前脂肪细胞分化的相关蛋白。首先，尝试鉴定Fib1的受体。亲和层析法和亲和标记法这两种不同的分析系统通常检测到一个30 kDa的蛋白质。这个30 kDa的蛋白被认为是一种不同于整合素的特异性Fib1受体。接下来，研究了一种能够诱导ST-13细胞分化的自分泌因子，该因子是由Fib1刺激的ST-13细胞释放出来的。从经Fib1片段刺激的ST-13细胞条件培养液中纯化出一种300-500 kDa的4-硫酸软骨素蛋白多糖(PG)。该PG在极低浓度(10-50 ng/ml)下诱导ST-13细胞向脂肪细胞分化。该活性PG含有一个70 kDa的二硫键连接的蛋白质亚基，其氨基端氨基酸为D-T-H-K-S-E-I-A-H-R-F-K-D-L-G-E-E。最后，从这些因子参与ST-13细胞FN基质形成的角度，探讨了Fib1和活性PG诱导脂肪细胞分化的机制。ST-13细胞表面积累的FN基质的定量数据表明，Fib1通过抑制FN基质的积聚促进脂肪细胞的分化，活性PG以一种不依赖于FN基质的方式诱导脂肪细胞分化。FIB-1和活性PG可通过不同途径相加刺激脂肪细胞分化。

Proteins which are involved in the adipocyte differentiation of ST-13 preadipocytes induced by the fibronectin (FN) fragment (Fib 1) from the Fib 1 domain, were investigated. First, an attempt to identify the receptor for Fib 1 was performed. A 30-kDa protein was commonly detected by the two different analysis systems, affinity chromato-graphy and affinity labelling methods. The 30-kDa protein was expected to be a specific Fib 1 receptor distinct from integrins. Next, an autocrine factor capable of inducing the adipocyte differentiation of ST-13 cells, which is released from ST-13 cells stimulated with Fib 1, was studied. A 300-500-kDa chondroitin 4-sulfate type proteoglycan (PG) was purified from the conditioned medium of ST-13 cells stimulated with Fib 1 fragment. This PG induced the adipocyte differentiation of ST-13 cells at an extremely low concentrations (10-50ng as glucuronolactone/ml). The differentiation-inducing activity was diminished by treating with chondroitinase ABC.This active PG was shown to contain a 70-kDa protein subunit linked with disulfide bond, and amino-terminal amnino acids of the subunit was D-T-H-K-S-E-I-A-H-R-F-K-D-L-G-E-E.Finally, the mechanism by which Fib 1 and the active PG induce the adipocyte differentiation was studied, in terms of participation of these factors in FN matrix formation of ST-13 cells. The data of quantification of FN matrix accumulated in ST-13 cell surface indicated that Fib 1 promotes the adipocyte differentiation by inhibiting the accumulation of FN matrix and that active PG induces the differentiation in a way independent on the FN matrix asembly. Fib 1 and the active PG would stimulate additively the adipocyte differentiation in different pathways.