Barak-Shalom T, Schickler M, Knopov V, Shapira R, Hurwitz S, Pines M
Institute of Animal Science, Agricultural Research Organization, Volcani Center, Bet Dagan, Israel.
Comp Biochem Physiol C Pharmacol Toxicol Endocrinol. 1995 May;111(1):49-59. doi: 10.1016/0742-8413(95)00021-x.
The regulation of synthesis and phosphorylation of osteopontin in relation to avian epiphyseal growth-plate chondrocyte differentiation was studied in situ and in culture. Osteopontin gene expression was evaluated in the tibia growth-plate of 3-week-old chickens by in situ hybridization. The gene was expressed mainly at the lower hypertrophic zone where cartilage matrix is calcified and endochondral bone formation is initiated. Within the hypertrophic region, a poorly labeled area separated the layer of osteopontin-positive hypertrophic chondrocytes from those associated with endochondral bone formation. In culture, proliferative chondrocytes show no alkaline phosphatase activity in contrast to ascorbic acid-treated chondrocytes which display the enzyme activity. Chondrocytes not treated with ascorbic acid, exhibited lower levels of osteopontin mRNA than the treated cells. The phorbol ester TPA--an activator of protein kinase C--and to a lesser extent FGF but not EGF, stimulated osteopontin gene expression. Chondrocytes secreted low levels of phosphorylated osteopontin to the medium. EGF treatment resulted in the appearance of phosphorylated osteopontin in the medium, without affecting the synthesis of other proteins. FGF and TGF beta, but not IGF-I or IGF-II, also caused phosphorylation of osteopontin. Ascorbic acid-treated chondrocytes secreted higher levels of phosphorylated osteopontin than the non-treated cells, but addition of FGF or TPA did not stimulate osteopontin phosphorylation any further. Parathyroid hormone caused a dose-dependent attenuation of osteopontin phosphorylation and inhibited the EGF-dependent osteopontin phosphorylation. The results suggest that osteopontin gene expression and phosphorylation in chondrocytes are regulated by separate mechanisms. The response to the various controlling agents varies with the state of differentiation. Both processes--the synthesis and phosphorylation of osteopontin--are under the control of local growth factors which are involved in bone growth and calcification.
对骨桥蛋白的合成与磷酸化调节与禽类骨骺生长板软骨细胞分化的关系进行了原位和体外培养研究。通过原位杂交评估3周龄鸡胫骨生长板中的骨桥蛋白基因表达。该基因主要在软骨基质钙化和软骨内骨形成起始的较低肥大区表达。在肥大区内，一个标记较弱的区域将骨桥蛋白阳性肥大软骨细胞层与参与软骨内骨形成的细胞层分隔开来。在培养中，增殖性软骨细胞无碱性磷酸酶活性，而经抗坏血酸处理的软骨细胞则显示该酶活性。未用抗坏血酸处理的软骨细胞，其骨桥蛋白mRNA水平低于处理过的细胞。佛波酯TPA（蛋白激酶C的激活剂）以及程度较轻的FGF而非EGF，刺激了骨桥蛋白基因表达。软骨细胞向培养基中分泌低水平的磷酸化骨桥蛋白。EGF处理导致培养基中出现磷酸化骨桥蛋白，而不影响其他蛋白质的合成。FGF和TGFβ，但不是IGF-I或IGF-II，也导致骨桥蛋白磷酸化。经抗坏血酸处理的软骨细胞比未处理的细胞分泌更高水平的磷酸化骨桥蛋白，但添加FGF或TPA并未进一步刺激骨桥蛋白磷酸化。甲状旁腺激素导致骨桥蛋白磷酸化呈剂量依赖性减弱，并抑制EGF依赖性骨桥蛋白磷酸化。结果表明，软骨细胞中骨桥蛋白基因表达和磷酸化受不同机制调节。对各种调控因子的反应随分化状态而变化。骨桥蛋白的合成和磷酸化这两个过程均受参与骨生长和钙化的局部生长因子控制。
