Benayahu D, Fried A, Shamay A, Cunningham N, Blumberg S, Wientroub S
Department of Histology and Cell Biology, Sackler Faculty of Medicine, Tel Aviv University, Israel.
J Cell Biochem. 1994 Sep;56(1):62-73. doi: 10.1002/jcb.240560111.
The effects of retinoic acid (RA) on the expression of osteoblastic-related cell markers was examined. A marrow stromal osteogenic cell line, MBA-15, was analyzed by Northern blotting for the expression of bone matrix proteins. These cells constitutively express mRNA encoding for procollagen alpha 2 (I), osteonectin, osteopontin, biglycan, and alkaline phosphatase (ALK-P). Gene expression was unchanged in response to RA triggering for 24 hr. Furthermore, cell growth and enzymatic activities of ALK-P and neutral endopeptidase (CD10/NEP) were studied. These parameters were examined in MBA-15 and clonal populations representing different stages of differentiation. The cell's growth rate was unchanged, while ALK-P activity was greatly increased during the culture period under RA treatment in MBA-15 and in the clonal cell lines examined while CD10/NEP activity displayed a different pattern. MBA-15.4, a preosteoblast cell line, exhibited an inhibition in CD10/NEP activity at the beginning of the culture period, reaching basal level with time. This activity was greatly increased over control level in MBA-15.6, a mature stage of osteoblasts. Furthermore, the response of cell lines to various growth factors was tested subsequent to priming the cultures with RA. A synergistic effect was monitored for ALK-P activity in MBA-15.4 and MBA-15.6 cells under rh-bone morphogenic protein (BMP-2) and purified osteogenin (BMP-3), and an antagonist effect was measured when cells were exposed to transforming growth factor beta (TGF beta). Contrarily, BMP-2 and BMP-3 inhibited the CD10/NEP activity that had remained unchanged following priming of the cell with RA. Insulin-like growth factor I (IGF-I) and basic fibroblast growth factors (bFGF) did not affect either ALK-P nor CD10/NEP activities in both cloned cells. Cellular response to bone-seeking hormone, parathyroid hormone (PTH), and prostaglandin E2 (PGE2) was monitored by activation of intracellular cAMP. Treatment with RA caused a dramatic decrease in MBA-15.6 cell responses to PTH and PGE2, but no significant effects could be observed in other clonal lines.
研究了视黄酸（RA）对成骨细胞相关细胞标志物表达的影响。采用Northern印迹法分析骨髓基质成骨细胞系MBA - 15中骨基质蛋白的表达。这些细胞组成性表达编码前胶原α2（I）、骨连接蛋白、骨桥蛋白、双糖链蛋白聚糖和碱性磷酸酶（ALK - P）的mRNA。在RA刺激24小时后，基因表达未发生变化。此外，还研究了细胞生长以及ALK - P和中性内肽酶（CD10/NEP）的酶活性。在MBA - 15和代表不同分化阶段的克隆群体中检测了这些参数。细胞生长速率未发生变化，而在MBA - 15以及所检测的克隆细胞系中，在RA处理的培养期间，ALK - P活性显著增加，而CD10/NEP活性呈现出不同的模式。前成骨细胞系MBA - 15.4在培养初期CD10/NEP活性受到抑制，随时间推移达到基础水平。在成骨细胞成熟阶段的MBA - 15.6中，该活性比对照水平大幅增加。此外，在用RA预处理培养物之后，测试了细胞系对各种生长因子的反应。在rh - 骨形态发生蛋白（BMP - 2）和纯化的骨生成素（BMP - 3）作用下，监测到MBA - 15.4和MBA - 15.6细胞中ALK - P活性具有协同效应，而当细胞暴露于转化生长因子β（TGFβ）时，检测到拮抗效应。相反，BMP - 2和BMP - 3抑制了在用RA预处理细胞后保持不变的CD10/NEP活性。胰岛素样生长因子I（IGF - I）和碱性成纤维细胞生长因子（bFGF）对两种克隆细胞中的ALK - P和CD10/NEP活性均无影响。通过细胞内cAMP的激活监测细胞对趋骨性激素甲状旁腺激素（PTH）和前列腺素E2（PGE2）的反应。RA处理导致MBA - 15.6细胞对PTH和PGE2的反应显著降低，但在其他克隆系中未观察到明显影响。
