Abstract 615: Absence of RIP3 Reduces Atherosclerotic Progression in Type 1 Diabetes

Abstract

Introduction: Patients suffering from Type 1 diabetes (T1D) have increased risk of developing atherosclerosis. T1D is a chronic autoimmune metabolic disease characterized by a deficiency in insulin secretion and hyperglycemia, promoting oxidative stress and low-grade inflammation in part via advanced glycation end products (AGE). Recently, we and others showed that necroptosis, a newly defined pro-inflammatory programmed cell death, drives advanced atherosclerosis. Given that diabetes accelerates inflammation and atherosclerosis, we hypothesized that hyperglycemia may exacerbate macrophage necroptosis to drive atherosclerotic lesion formation. Methods/Results: Male Ldlr-/- mice were lethally irradiated, transplanted with bone-marrow from WT or Rip3 -/- mice and then administered streptozotocin (STZ) or vehicle daily for 5 days to induce T1D, prior to being fed a western diet for 6 wks. As expected, STZ-treated WT BMT Ldlr -/- mice had a marked increased in atherosclerotic aortic lesion area compared to non-diabetic mice (1.08%±0.30 vs 2.13%±0.45). which was blunted in Rip3 -/- BMT Ldr -/- mice (arch: 49% reduction, sinus: 54% reduction, p<0.05). Bone marrow derived macrophages (BMDMs) cultured in high glucose (HG) increased cell death in response to necroptotic stimuli (oxidized LDL ± caspase inhibitor zVAD) relative to those cultured in low glucose (LG). Further, the addition of recombinant s100A8/9, a soluble AGE, markedly amplified oxLDL-induced death in HG cultured BMDMs with minimal effects on LG cultured BMDMs, suggesting that pro-inflammatory AGEs released in diabetes can augment macrophage cell death. Further, the increased cell death induced by oxLDL+S100A8/9 was inhibited by a necroptotic inhibitor, Nec-1, or in Rip3 -/- BMDMs cultured in HG. Mechanistically, treatment of BMDMs cultured in HG with oxLDL+s100A8/9 increased the expression of Rip3 and Mlkl (~5-fold and ~2-fold respectively), which may drive necroptotic cell death during hyperglycemia. Conclusion: Deletion of Rip3 markedly halts atherosclerotic progression in a STZ-induced T1D mouse model by inhibiting macrophage necroptosis induced by both atherogenic ligands (oxLDL) and soluble AGE products (s100A8/9).