Role of macrophage-specific calpain-2 in hypercholesterolemia induced atherosclerosis .

The objective of my proposal is to understand and elucidate the functional key role of calpains in atherosclerosis development. Calpains are calcium-dependent cysteine proteases that are unique from other intracellular proteolytic enzymes in substrate recognition and proteolytic process. Human atherosclerosis is associated with prolonged activation of calpains, however, surprisingly few cellular substrates of calpains have been identified. Recently, I have provided novel evidence that pharmacological inhibition of calpains attenuates atherosclerosis in LDLr-/- mice. The two major isoforms, calpain-1 and -2, are ubiquitously expressed whereas the other isoforms are tissue specific. Using calpain-1 deficient mice, I have determined that calpain-1 deficiency has no effect on hypercholesterolemia-induced atherosclerosis. This infers that the activated calpain-2 not calp-1 is involved in atherosclerosis development. Since the inactivation of the calpain-2 gene results in early embryonic lethality in mice due to defects in cardiac development, the role of calpain-2 in atherosclerosis is unknown. Preliminary studies demonstrates that: 1) protein abundance of calpain-1 and -2 was increased in lipid-loaded macrophages 2) calpain-2, not calpain-1 deficient macrophages had increased protein of ABCA1 3) calpain-2 deficiency prevented macrophage foam cell formation in the presence of apoA-I 4) immunostaining of mouse atherosclerotic lesions showed increased calpain-2 protein that co-localizes with macrophages 5) Calpain-2 deficiency prevented NF-kB activation in macrophages. Collectively, these results infer a novel role of calpain-2 in atherosclerosis. The availability of macrophage specific and tamoxifen-inducible whole body calpain-2 deficient mice provides an excellent model to study the role of calpain-2 in atherosclerosis development Based on background and preliminary findings, my central hypothesis is that calpain-2 deficiency in macrophages will attenuate hypercholesterolemia-induced atherosclerosis by promoting macrophage reverse cholesterol transport and suppressing NF-kB mediated inflammation. To test this hypothesis, I propose the following aims: Aim 1: Determine the role of calpain-2 in macrophage cholesterol efflux and inflammation. Aim 2: Determine the contribution of calpain-2 to in vivo macrophage reverse cholesterol transport. Aim 3: Determine the contribution of calpain-2 to hypercholesterolemia-induced atherosclerosis