The role of hcs-24, a newly isolated hypertrophic chondrocyte-specific gene, in endochondral ossification

1)克隆了肥大软骨细胞特异性基因hcs-24的cDNA。hcs-24编码区核苷酸序列与结缔组织生长因子（CTGF）完全相同。2）hcs-24/CTGF在培养的兔生长软骨细胞中的表达在肥大期最高。TGF β和BMP-2刺激软骨细胞基因表达。3）免疫组化技术显示，抗CTGF抗体染色显示小鼠肋软骨连接处肥大的软骨细胞和内皮细胞。在小鼠胚胎发育过程中，hcs-24/ctgf的mRNA水平在E7时达到最高，随后逐渐下降，到E17时又开始上升。转染hcs-24表达载体的HCS-2/8细胞生长速度快于未转染细胞。转染反义寡核苷酸的血管内皮细胞增殖和迁移能力的研究 关于我们 6）纯化的CTGF和重组CTGF能促进软骨细胞增殖、蛋白多糖合成及软骨细胞和成骨细胞碱性磷酸酶的合成。该生长因子可刺激血管内皮细胞的增殖和迁移。7）建立了Hcs-24/CTGF双抗体夹心ELISA检测方法; 8）<125>在HCS-2/8细胞上发现了两种不同的rCTGF特异性结合位点。9<125>）OCNT/CTGF转基因小鼠出现骨骼疾病，提示肥大软骨细胞合成的Hcs-24/CTGF刺激增殖软骨细胞的增殖和成熟，促进成熟软骨细胞的肥大，并诱导骨血管生成进入软骨，从而促进软骨内成骨。该因子也可能参与胚胎的器官发生。少

1) cDNA of hypertrophic chondrocyte specific gene hcs-24 was isolated. The nucleotide sequence of coding region of hcs-24 was completely the same as that of connective tissue growth factor (CTGF).2) Expression of hcs-24/ctgf in rabbit growth cartilage cells in culture was highest in hypertrophic stage. The gene expression in chondrocytic cells was stimulated by TGFbeta and BMP-2.3) Immunohistochemical techniques revealed that hypertrophic chondrocytes and endothelial cells in cost-chondral junctions of mouse ribs were stained with anti-CTGF antibody in vivo. Surface of and chondrocyte clusters in articular cartilge of arthritis were also stained with the antibody.4) During development of mouse embryos, mRNA level of hcs-24/ctgf reached a maximum at E7, decreased gradually and then increased again at E17.HCS-2/8 cells transfected with an hcs-24 expression vector grew rapidly than non-transfected cells. The abilities to proliferate and migrate of vascular endothelial cells transfected wi … More th expression vectors that generate anti-sense RNA of CTGF cDNA were markedly lower than those of control.6) Purified CTGF and recombinant CTGF stimulated the proliferation and proteoglycan synthesis of chondrocytes and alkaline phosphatase in chondrocytes and osteoblasts. The growth factor simulated the proliferation and migration of vascular endothelial cells. These effects were inhibited by anti-CTGF antibody.7) An ELISA system to measure Hcs-24/CTGF was established.8) Two types of specific binding sites of ^<125>I-rCTGF were identified on HCS-2/8 cells. The binding of ^<125>I-rCTGF to rabbit growth cartilage cells in culture was maximal in growth phase and decreased as they differentiated.9) Transgenic mice of OCNT/CTGF had skeletal disorder.These findings suggest that Hcs-24/CTGF synthesized by hpertrophic chondrocytes stimulates the proliferation and maturation of proliferative chondrocytes and hypertrophy of mature chondrocytes and induces angiogenesis into cartilage from bone, resulting in promotion of endochondral ossification. The factor may also be involved in organogenesis in embryos. Less