Yosimichi G, Nakanishi T, Nishida T, Hattori T, Takano-Yamamoto T, Takigawa M
Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine and Dentistry, Okayama, Japan.
Eur J Biochem. 2001 Dec;268(23):6058-65. doi: 10.1046/j.0014-2956.2001.02553.x.
Connective tissue growth factor/hypertrophic chondrocyte specific gene product 24 (CTGF/Hcs24) promotes proliferation and differentiation of chondrocytes in culture. We investigated the roles of two major types of mitogen activated protein kinase (MAPK) in the promotion of proliferation and differentiation by CTGF/Hcs24. Here we report the effects of the MAPKK/MEK 1/2 inhibitor, PD098059, and p38 MAPK inhibitor, SB203580, in a human chondrosarcoma-derived chondrocytic cell line (HCS-2/8) and rabbit growth cartilage (RGC) cells treated with CTGF/Hcs24. In the proliferation phase, CTGF/Hcs24 induced a approximately fivefold increase in the phosphorylation of p44/42 MAPK/ERK and a approximately twofold increase in that of p38 MAPK in an in vivo kinase assay. These inhibitors of MAPKK and MAPK suppressed phosphorylation of ets-like gene-1 (Elk-1) and nuclear activating transcription factor-2 (Atf-2) induced by CTGF/Hcs24 in a dose-dependent manner, respectively. Western blot analysis showed that phosphorylation of ERK was induced from 30 to 60 min and phosphorylation of p38 MAPK from 10 to 15 min after the addition of CTGF/Hcs24 in confluence HCS-2/8 cells. PD098059 suppressed the DNA synthesis of HCS-2/8 cells and RGC cells, while SB203580 did not. On the other hand, the p38 MAPK inhibitor, SB203580, completely inhibited the CTGF/Hcs24-induced synthesis of proteoglycans in HCS-2/8 cells and RGC cells but the MEK1/2 inhibitor, PD098059, did not. These results suggest that ERK mediates the CTGF/Hcs24-induced proliferation of chondrocytes, and that p38 MAPK mediates the CTGF/Hcs24-induced differentiation of chondrocytes.
结缔组织生长因子/肥大软骨细胞特异性基因产物24（CTGF/Hcs24）可促进培养的软骨细胞增殖和分化。我们研究了两种主要类型的丝裂原活化蛋白激酶（MAPK）在CTGF/Hcs24促进增殖和分化过程中的作用。在此，我们报告了MAPKK/MEK 1/2抑制剂PD098059和p38 MAPK抑制剂SB203580对人软骨肉瘤来源的软骨细胞系（HCS-2/8）和经CTGF/Hcs24处理的兔生长软骨（RGC）细胞的影响。在增殖阶段，CTGF/Hcs24在体内激酶试验中使p44/42 MAPK/ERK的磷酸化增加约五倍，使p38 MAPK的磷酸化增加约两倍。这些MAPKK和MAPK抑制剂分别以剂量依赖的方式抑制CTGF/Hcs24诱导的ets样基因-1（Elk-1）和核激活转录因子-2（Atf-2）的磷酸化。蛋白质印迹分析表明，在汇合的HCS-2/8细胞中加入CTGF/Hcs24后，ERK的磷酸化在30至60分钟诱导，p38 MAPK的磷酸化在10至15分钟诱导。PD098059抑制HCS-2/8细胞和RGC细胞的DNA合成，而SB203580则无此作用。另一方面，p38 MAPK抑制剂SB203580完全抑制CTGF/Hcs24诱导的HCS-2/8细胞和RGC细胞中蛋白聚糖的合成，但MEK1/2抑制剂PD098059则无此作用。这些结果表明，ERK介导CTGF/Hcs24诱导的软骨细胞增殖，而p38 MAPK介导CTGF/Hcs24诱导的软骨细胞分化。
