Signal transduction pathways provide post-receptor mechanisms that allow multiple responses to a hormone such as parathyroid hormone (PTH), for which there is a single receptor. Knowledge of which actions of the hormone are mediated by which pathways can lead to the design of selective therapies to enhance or decrease specific responses. The focus of the current proposal is the role of the phospholipid/diacylglycerol/protein kinase C pathway in parathyroid hormone actions on bone. Evidence suggests that several important actions of parathyroid hormone are likely to be mediated, completely or in part, through this pathway. These actions include the stimulation of IL-6 production, the down-regulation of cAMP production, and the inhibition of collagen synthesis. Thus, activation of this pathway by parathyroid hormone in bone would result in less anabolic action and more resorptive activity. Recent studies in other tissues reveal that the phospholipid/diacylglycerol/PKC pathway is highly diverse. There are several potential phospholipid sources for the diacylglycerol, and 12 recognized PKC isozymes. Methodologies are now available to define the PKC pathway in bone much more precisely than in the past. The hypotheses to be tested are that a) PTH actions to increase IL-6, down-regulate cAMP production, and decrease collagen synthesis are mediated through the PKC pathway and involve activation of specific PKCs, b) that the specificity in PKC activation arises through the specific diacylglycerols being produced, and c) that these specific diacylglycerols are generated through the hydrolysis of different phospholipids, with the sustained effects of PTH being due to hydrolysis of phosphatidylcholine.
The specific aims are thus 1) to characterize phosphatidylcholine hydrolysis in osteoblastic cells in response to PTH, 2) to characterize the changes in PKC isozymes in osteoblastic cells in response to PTH, and 3) to demonstrate the role of specific PKC pathways in actions of PTH on osteoblastic cells. Selective amplification or depression of the proposed pathways and isozymes would be used to test the hypothesis that they are the pathways for the end responses under consideration. The studies would be carried out in UMR-106 osteoblastic osteosarcoma cells and osteoblastic cells obtained from neonatal mouse calvaria. Effects of bPTH(1-34) will be compared to those of bPTH(3-34) and bPTH(7-34)NH/2, which are also agonists on PKC production but have little or no agonist effects on cAMP.
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