Regulation and function of retinol binding protein in cartilage

(1)甲状旁腺激素(PTH)和二丁基环腺苷((Bu)2cAMP)诱导兔生长板软骨细胞条件培养液中视黄醇结合蛋白(RBP)的表达。(2)Northern印迹分析显示，PTH、甲状旁腺激素相关肽(PTHrP)和(Bu)2cAMP使培养的软骨细胞RBP的mRNA水平升高。此外，PTH和(Bu)2cAMP均显著诱导RBP mRNA的表达，提示翻译前调节。PTH、PTHrP和(Bu)2cAMP诱导的mRNA表达水平与维甲酸(RA)一样高。(3)RBP和PTH/PTHrP均抑制RA对生长板软骨细胞的去分化作用。这些结果表明，软骨细胞在体内和体外都能合成和分泌RBP，提示PTH/PTHrP通过在软骨细胞中产生RBP来调节RA的作用。(4)RA和骨形态发生蛋白-2(BMP-2)促进了印度刺猬(IHH)的mRNA表达，而PTH/PTH相关肽(PTHrP)显著抑制了IHH的表达。(5)放线菌酮阻断了RA对IHH的上调，提示这种刺激需要从头合成蛋白质。这些结果表明，RA参与了软骨内成骨过程中IHH的上调。(6)PTH可迅速抑制IHH的mRNA表达。PTH(1-84)抑制IHH的表达不需要从头合成蛋白质。PTH/PTHrP对IHH的直接抑制作用可能是一种负反馈机制，阻止了PTHrP在软骨中的过度积聚。

The following results were obtained to clarify the role of retinoic acid and PTH as the modulators in the growth and development of chondrocytes.(1) Parathyroid hormone (PTH) and dibutyryl cAMP ((Bu) 2cAMP) induced the expression of plasma retinol-binding protein (RBP) in the conditioned media of rabbit growth plate chondrocyte cultures.(2) Northern blot analysis showed that PTH, parathyroid hormone-related peptide (PTHrP) and (Bu) 2cAMP increased the RBP mRNA level in chondrocyte cultures. Further, both PTH and (Bu) 2cAMP markedly induced the expression of RBP mRNA indicating a pretranslational regulation. The level of the mRNA expression induced by PTH, PTHrP and (Bu) 2cAMP was as high as that by retinoic acid (RA).(3) Both RBP and PTH/PTHrP inhibited the dedifferentiative activity of RA on growth plate chondrocytes. These results demonstrate that chondrocytes synthesize and secrete RBP in vivo and in vitro, and suggest that PTH/PTHrP naodulates the effect of RA by means of RBP production in chondrocytes.(4) RA and bone morphogenetic protein-2 (BMP-2) enhanced Indian hedgehog (Ihh) mRNA expression, whereas PTH/PTH-related peptide (PTHrP) markedly suppressed Ihh expression.(5) Cycloheximide blocked the up-regulation of Ihh by RA, indicating the requirement of de novo protein synthesis for this stimulation. These findings suggest that RA is involved in the up-regulation of Ihh during endochondral bone formation.(6) PTH rapidly abolished the mRNA expression of Ihh. The inhibition of Ihh expression by PTH (1-84) did not require de novo protein synthesis. The direct inhibitory action of PTH/PTHrP on Ihh appears to be a negative feedback mechanism that prevents excess PTHrP accumulation in cartilage.