The long term goal of this project is to understand how platelets regulate monocyte function. This issue is particularly important since platelets interact with monocytes in a variety of atherosclerotic disorders. The applicant recently demonstrated that activated platelets, but not unstimulated platelets, induce immediate- early (IE) gene expression in monocytes. Among the IE genes expressed, interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1) were synthesized and secreted by monocytes exposed to activated platelets. Other IE genes, including tumor necrosis factor-2 (TNF-2), were not generated. Initial results indicate that P-selectin and RANTES (Regulated upon Activation Normal T Cell Expressed presumed Secreted), two platelet-derived molecules, are required for both IL-8 and MCP-1 synthesis in monocytes through P-selectin and RANTES. Signaling in monocytes through P-selectin and RANTES will be compared to signaling induced by LPS, a potent monocyte agonist. In the first Specific Aim, the applicant will further characterize how P-selectin and RANTES regulate monokine synthesis. These studies will initially be defined in a reduced system where different forms of purified P-selectin and RANTES are presented to monocytes; and cellular MRNA, cellular retained protein, and secreted protein for IL-8, MCP-1, and TNF-2 will be measured. A more complex system involving platelet-monocyte interactions to determine if P-selectin and RANTES are required for monokine production will be examined.
Specific Aim 3 will determine if P-selectin and RANTES regulate NF-kappaB activity in monocytes. NF- kappaB is a transcription factor that is required for maximal IL-8, MCP-1, and TNF-2 synthesis, and initial results developed by the applicant indicate that NF-kappaB, and its inhibitory factor IkappaB-2, are activated by P-selectin and RANTES. The third Specific Aim will determine if P-selectin and RANTES regulate p70 S6 kinase activity in monocytes. p70 S6 kinase is known to translationally regulate many proteins and other evidence also indicates that p70 S6 kinase can regulate NF-kappaB family members. Initial results suggest that p70 S6 kinase activity is increased in monocytes following exposure to P- selectin and RANTES or activated platelets. Moreover, inhibition of p70 S6 kinase activity by rapamycin attenuates MCP-1 secretion by monocytes exposed to activated platelets. In the final Specific Aim, correlative studies to determine if P-selectin and RANTES are localized in ruptured carotid arterial plaques will be coordinated. Together, these results will begin defining how activated platelets, through P-selectin and RANTES, induce monokine synthesis.
| Weyrich, A S; Schwertz, H; Kraiss, L W et al. (2009) Protein synthesis by platelets: historical and new perspectives. J Thromb Haemost 7:241-6 |
| Mahoney, T S; Weyrich, A S; Dixon, D A et al. (2001) Cell adhesion regulates gene expression at translational checkpoints in human myeloid leukocytes. Proc Natl Acad Sci U S A 98:10284-9 |
