Introduction: Granzymes A and B (GrA/GrB) are cytotoxic proteases that are released by immune cells to eliminate target cells. GrB is present in human vessels and its expression corresponds to increased disease severity. In addition to their cytotoxic roles, in conditions of chronic inflammation, GrA and GrB may contribute to the degradation of extracellular matrix (ECM) proteins leading to tissue degeneration.
Hypothesis: GrA and GrB levels are elevated in atherosclerotic patients and play a key role in the degradation of ECM and atherosclerosis.
Methods: ApoE −/− x GrB −/− double knockout (apoE/GrB-DKO) mice were generated. C57-wt, GrB-KO, apoE-KO or apoE/GrB-DKO mice were fed a high fat diet for 30 weeks, sacrificed, tissues processed and lesion morphology and morphometry assessed. GrA and GrB levels were also measured in human plasma from patients with or without clinically evident atherosclerosis by ELISA. GrA/GrB-mediated ECM proteolytic activity was also assessed.
Results: GrB deficiency significantly reduced the development of xanthomatosis, hair loss and atherosclerosis in the apoE-KO mice. GrB deficiency resulted in a marked reduction of elastin degradation in both the skin and blood vessels. GrB was found to co-localize to elastin fibres in atherosclerotic plaques and was capable of binding to, and cleaving elastin in vitro. ECM cleavage assays demonstrated that GrA and GrB cleave other ECM such as fibronectin. Preliminary studies indicate that GrB deficiency increases the lifespan of the apoE-KO mice as the apoE/GrB-DKO mice can survive up to 60 weeks of age on a high fat diet with no external indication of disease, skin degeneration or hair loss. In humans, preliminary evidence indicates that GrA/GrB levels are elevated in atherosclerosis. Smoking also appears to elevate plasma granzyme levels.
Conclusions: During chronic inflammation, granzymes accumulate extracellularly and exhibit proteolytic activity towards extracellular matrix proteins that results in vascular remodeling and atherosclerosis with age. Attenuation of GrB resulted not only in a significant reduction of atherosclerosis, but also prolonged the lifespan of apoE-KO mice.
Concurrent exercise improves insulin resistance and nonalcoholic fatty liver disease by upregulating PPAR-γ and genes involved in the beta-oxidation of fatty acids in ApoE-KO mice fed a high-fat diet