Secretion of a surface-active substance, termed surfactant, by the alveolar type II epithelial cell has been demonstrated to be induced by a variety of agents, but the mechanism of this stimulation is not known. In general, agonist binding to a receptor leads to production of second messengers and activation of cellular processes. Three of such second messengers are the phosphatidylinositol 4,5-bisphosphate (PIP-2) hydrolysis products, diacylglycerol (DAG) and inositol trisphosphate (IP-3) and cAMP produced by adenylate cyclase. In other systems, DAG activates a calcium and phospholipid-dependent protein kinase, termed protein kinase C. The phorbol ester (TPA) and the DAG analog, 1-oleoyl-2-acetyl glycerol, activators of protein kinase C, stimulated surfactant secretion in cultured type II cells. This suggests a role for this mechanism as an activator. In other systems, IP-3 mobilizes intracellular calcium. In cultured type II cells, the calcium ionophore A23187 activates secretion but its mechanism of activation is unknown. (Arg-8)Vasopressin has been shown to stimulate surfactant secretion and phosphatidylinositol (PI) hydrolysis in cultured type II cells. Studies outlined in this proposal will determine if PIP-2 is the specific PI being hydrolyzed and if IP-3 and DAG are the hydrolysis products. The role of IP-3 as an activator of intracellular calcium mobilization and DAG as an activator of protein kinase C are proposed. The receptor class activated by (Arg-8)vasopressin will be investigated. Preliminary studies demonstrated that surfactant secretion could be stimulated by histamine as well. Studies are proposed to determine if histamine acts via PIP-2 hydrolysis and if it acts via the same receptor as vasopressin. Previous studies have shown that surfactant secretion can be stimulated in a cAMP-dependent manner. Various studies also suggest that adenylate cyclase and cAMP-dependent protein kinase are sensitive to calcium. These enzymes will be fractionated to identify the subunits that are calcium dependent. Dependency on the calcium regulating protein, calmodulin, as a modulator will also be examined. Previous studies have shown that surfactant secretion can be inhibited by pretreatment of the cells with compound 48/80, the neuropeptide substance P and vasopressin. The mechanism of inhibition will be investigated. In type II cells, the cytoskeletal protein, tubulin has been suggested to play a role in secretion and its polymerization and phosphorylation will be studied.
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| Dubrovin, L C; Brown, L A (1992) Stimulation of phosphatidylcholine hydrolysis in type II alveolar epithelial cells. Am J Physiol 263:L42-50 |
| Brown, L A (1991) Influence of streptozotocin-induced diabetes on adenylate cyclase activity in cultured type II pneumocytes. Am J Respir Cell Mol Biol 4:108-14 |
| Chen, W; Yang, V C (1991) Versatile, non-clotting-based heparin assay requiring no instrumentation. Clin Chem 37:832-7 |
| Chen, M; Brown, L A (1990) Histamine stimulation of surfactant secretion from rat type II pneumocytes. Am J Physiol 258:L195-200 |
| Brown, L A; Chen, M (1990) Vasopressin signal transduction in rat type II pneumocytes. Am J Physiol 258:L301-7 |
| Brown, L A; Wood, L H (1989) Stimulation of surfactant secretion by vasopressin in primary cultures of adult rat type II pneumocytes. Biochim Biophys Acta 1001:76-81 |
