Role of PKC Epsilon in Atheroprotection

PROJECT SUMMARY Cardiovascular disease afflicts approximately 50% of older Americans and is the major cause of death worldwide. It is a chronic inflammatory disease in which macrophages (MØ) play a central role; their deletion reduces plaque size and composition. Usually, inflammation is self-limiting, with resolution following initiation. In atherosclerosis this yin and yang is out of balance, with MØ failing to shift towards resolution. While many of the players in the inflammation (cytokines, chemokines) and resolution (specialized pro-resolving lipids including RvD1) phases have been identified, the intracellular signaling network that links uptake of oxLDL to resolution, and the roadblocks to that resolution in atherosclerosis, remain to be defined. Preliminary data implicate PKCe in both inflammation and resolution by restricting excessive inflammation while promoting release of specialized pro-resolving lipid mediators (SPM). However, how PKCe achieves this is a fundamental unanswered question. We generated a PKCeloxP/loxP mouse that was crossed with LysM Cre to generate a novel strain in which PKCe is selectively deleted in the myeloid lineage (meKO). When made hypercholesterolemic, meKO mice have significantly more plaque than WT animals. This result implicates myeloid PKCe as a novel atheroprotective gene. Using our meKO mice we will test the hypothesis that myeloid PKCe contributes to inflammation but also signals in the production of SPM, thus promoting resolution; its absence sustains inflammation and reduces resolution thereby exacerbating atherosclerosis. The hypothesis will be tested in vitro and in vivo. In vivo experiments include zymosan peritonitis, a self-resolving inflammation that will enable us to determine where PKCe acts along the inflammation"resolution axis. Cells and peritoneal lavage fluid will be recovered over time (4-72 h). Cell metrics include number and ratio of neutrophils to macrophages (MØ) and the polarization state of the MØ. Lavage fluid will be assayed for cytokines, chemokines, and SPMs. Secondly, archived aortic roots from hypercholesterolemic WT and meKO mice will be analyzed for histological differences (eg, localization and polarization state of MØ, amount and location of collagen, amount of necrosis and apoptosis.) Finally, in vitro studies will define the differences in the responses of WT and meKO MØ to oxidized LDL and during efferocytosis. Understanding how PKCe acts in inflammation and resolution may reveal novel nodes of regulation that can be therapeutically targeted to promote resolution in the context of hyperlipidemia. In conclusion, our findings will provide the foundation upon which to build a more mechanistic grant to understand how PKCe signals during the shift from inflammation to resolution. Translationally, the results may be applicable to those diseases in which PKCe is dysregulated and MØ play a prominent role (eg, cancer, scleroderma, Alzheimer’s, atherosclerosis, etc.).