THE METABOLISM OF LEAD IN BONE

骨骼是人体铅（Pb）的主要贮存库; 并且现在基于体内和体外数据认识到， 骨铅的一个骨骼亚室是容易交换的 并以类似于钙（Ca）的方式进行调节。 骨架是CaNa_2EDTA螯合Pb的部位; 最近的儿童临床观察表明， 骨骼是铅毒性作用的靶组织。 在这 在这种情况下，我们实验室的主要工作是测试 假设细胞钙稳态的扰动， 铅在相对低的浓度下产生的，是一个早期的， 铅毒性在细胞水平的离散表达。 等 可能是一个很大的障碍，可能是一个很大的障碍。 调节多个细胞过程的生理 通过细胞内离子钙变化的正常对照范围 浓度. 我们在小学进行的实验 单层培养分离的骨细胞（OC）和 成骨细胞（OB）骨细胞，在前一个资助期， 进一步证实并扩展这一假设， 现在可以直接验证。 提案的具体目标 是：1）表征稳态动力学分布， 通过去饱和技术调节OB中的Pb和Ca; 2） 通过19 F NMR直接测量胞质游离的 在有和没有铅的情况下，OC和OB中同时存在Ca和Pb（以及 促钙激素）在培养基中; 3）确定特定的 膜水平的铅钙相互作用机制 运输，其中铅对钙运输的影响，在孤立的 膜将在静息状态和神经刺激状态下进行检查 OC和OB; 4）确定Pb对Ca介导的细胞凋亡的影响 功能和cAMP的产生和降解; 5） 进一步表征铅对综合功能的影响， OC和OB单独和一起在共培养实验中。 总之，建议的定量，机械和 OB和OC的功能研究具有真实的潜力来定义， 这是第一次，早期铅毒性的分子基础， 细胞水平。

Bone is the major reservoir of body lead (Pb) stores in humans; and it is now recognized, based upon in vivo and in vitro data, that one skeletal subcompartment of bone Pb is readily exchangeable and modulated in a manner similar to that of calcium (Ca). Moreover, the skeleton is the site of Pb chelation by CaNa2EDTA; and recent clinical observations in children indicate that the skeleton is a target tissue for Pb's toxic effects. Within this context, our laboratory's primary efforts are directed at testing the postulate that pertubations in cellular Ca homeostasis, produced by Pb at relatively low concentrations, is an early and discrete expression of Pb toxicity at the cellular level. Such pertubations are likely to be far reaching and may place the regulation of multiple cellular processes out of the physiological range of normal control through changes in intracellular ionic Ca concentration. Experiments carried out by us in primary monolayer cultures of separated osteoclastic (OC) and osteoblastic (OB) bone cells, during the previous grant period, further confirm and extend this postulate to the point where it can now be verified directly. The specific aims of the proposal are to: 1) characterize the steady state kinetic distribution and modulation of Pb and Ca in OB by desaturation techniques; 2) measure directly by 19F NMR the concentration of cytosolic free Ca and Pb concurrently in OC and OB with and without Pb (and calciotropic hormones) in the medium; 3) determine the specific mechanisms of Pb-Ca interactions at the level of membrane transport, where Pb effects on Ca transport in isolated membranes will be examined in resting and hormonally stimulated OC and OB; 4) define the effects of Pb on Ca-mediated cell functions and the generation and degradation of cAMP; 5) characterize further Pb influences on the integrated functions of OC and OB separately and together in co-culture experiments. Taken together, the proposed quantitative, mechanistic and functional studies in OB and OC have the real potential to define, for the first time, the molecular basis of early Pb toxicity at the cellular level.