Schöfl C, Cuthbertson K S, Gallagher J A, Pennington S R, Cobbold P H, Brabant G, Hesch R D, von zur Mühlen A
Abteilung für Klinische Endokrinologie, Medizinische Hochschule Hannover, Federal Republic of Germany.
Biochem J. 1991 Feb 15;274 ( Pt 1)(Pt 1):15-20. doi: 10.1042/bj2740015.
It is known that parathyroid hormone (PTH) activates the cyclic AMP (cAMP) signalling pathway in osteoblasts. In recent years it has been suggested that an elevation of the intracellular free Ca2+ concentration ([Ca2+]i) may also be involved in the regulation of osteoblast function by PTH. However, this remains controversial. Here we investigated the effect of PTH on the [Ca2+]i of ROS 17/2.8 cells and normal human osteoblasts. The [Ca2+]i was measured in single aequorin-injected cells and in suspensions of cells loaded with fura-2. Human PTH-(1-38)-peptide (1-300 nM) had no effect on the [Ca2+]i in single aequorin-injected ROS 17/2.8 cells (n = 17) measured at various times after injection (1-20 h), or in suspensions of fura-2-loaded ROS 17/2.8 cells (n = 9). Ionomycin (1 microM) increased the [Ca2+]i in fura-2-loaded and single aequorin-injected ROS 17/2.8 cells by 285 +/- 60 nM (n = 9) and 312 +/- 99 nM (n = 6) respectively, indicating that both methods detect changes in [Ca2+]i with equal sensitivity. In contrast, human PTH-(1-38) (10-100 nM) markedly stimulated cAMP accumulation in ROS 17/2.8 cells. In single aequorin-injected normal human osteoblasts there was no change in the [Ca2+]i in response to 100 nM human PTH-(1-38) or 100 nM bovine PTH-(1-84) (n = 18). In contrast, in suspensions of normal human osteoblasts loaded with fura-2, an increase in [Ca2+]i in response to human PTH-(1-38) (100 nM) was found (60 +/- 28 nM; n = 6). Considerable variation in the magnitude of the response was observed between individual preparations and donors. These data indicate that PTH activates cAMP accumulation without affecting [Ca2+]i in ROS 17/2.8 cells and that PTH causes a rise in [Ca2+]i only in a small subset of normal human osteoblasts. We suggest that the Ca2+ response to PTH in osteoblasts is limited by the state of differentiation of the cells, and may be due either to the presence of a distinct Ca2(+)-mobilizing receptor or to a cAMP-mediated Ca2+ response.
已知甲状旁腺激素（PTH）可激活成骨细胞中的环磷酸腺苷（cAMP）信号通路。近年来，有人提出细胞内游离钙离子浓度（[Ca2+]i）的升高也可能参与PTH对成骨细胞功能的调节。然而，这一点仍存在争议。在此，我们研究了PTH对ROS 17/2.8细胞和正常人成骨细胞[Ca2+]i的影响。[Ca2+]i在单个注射水母发光蛋白的细胞以及装载fura-2的细胞悬液中进行测量。人PTH-（1-38）肽（1-300 nM）对注射后不同时间（1-20小时）测量的单个注射水母发光蛋白的ROS 17/2.8细胞（n = 17）的[Ca2+]i没有影响，对装载fura-2的ROS 17/2.8细胞悬液（n = 9）的[Ca2+]i也没有影响。离子霉素（1 microM）使装载fura-2的和单个注射水母发光蛋白的ROS 17/2.8细胞的[Ca2+]i分别增加了285±60 nM（n = 9）和312±99 nM（n = 6），这表明两种方法检测[Ca2+]i变化的灵敏度相同。相比之下，人PTH-（1-38）（10-100 nM）显著刺激了ROS 17/2.8细胞中cAMP的积累。在单个注射水母发光蛋白的正常人成骨细胞中，100 nM人PTH-（1-38）或100 nM牛PTH-（1-84）刺激后[Ca2+]i没有变化（n = 18）。相反，在装载fura-2的正常人成骨细胞悬液中，发现人PTH-（1-38）（100 nM）刺激后[Ca2+]i增加（60±28 nM；n = 6）。在不同的制备物和供体之间观察到反应幅度存在相当大的差异。这些数据表明，PTH在不影响ROS 17/2.8细胞[Ca2+]i的情况下激活cAMP积累，并且PTH仅在一小部分正常人成骨细胞中引起[Ca2+]i升高。我们认为，成骨细胞对PTH的钙离子反应受细胞分化状态的限制，可能是由于存在独特的钙离子动员受体，或者是由于cAMP介导的钙离子反应。
