Abstract 328: Viral Anti-Atherogenic Proteins Alter Monocyte and Neutrophil Invasion in Mice and Bag3 Gene Expression

Abstract

Aim Atherosclerosis is characterized by chronic inflammation and cell death (apoptosis). Serine protease inhibitors, or serpins, regulate inflammatory, thrombotic and apoptotic pathways. Poxviruses encode cross-class serpins that prevent host cell apoptosis. Serp-2, from Myxoma, reduces plaque, inflammation and apoptosis in animal models; CrmA, from Vaccinia, does not. Both serpins target Caspase 1 and Granzyme B, but the reasons for the differing effects in vivo are unknown. In prior research three Myxoma viral proteins, including Serp-2, reduced monocyte invasion, plaque growth, and aneurysm formation in ApoE-/- mice after angioplasty. These same proteins alter expression of a shared cohort of 48 apoptosis-related genes in human monocytes treated with camptothecin. This study assesses the effects of Serp-2 and CrmA on apoptotic gene expression in a mouse peritoneal inflammation model. Methods Mouse strains deficient for Granzyme B (GzmB, N=13) and Caspase 1 (Casp1, N=15) were compared to their respective background mice (C57Bl/6 and Nod, N=15 each) 18 hours after treatment with PMA and either Serp-2 or CrmA. RNA was isolated and analyzed by RT-PCR and normalized to GAPDH, then to the PMA-only treated control. Results Compared to human monocytes, mouse peritoneal exudates from knockout mice displayed differential alteration of BCL2-associated athanogene 3 (BAG3) in response to treatment with Serp-2 or CrmA. Serp-2 treatment reduced expression levels compared to CrmA treatment in GzmB (p=0.0267) and C57Bl/6 mice (p=0.0280). Casp1-/- mice treated with Serp-2 downregulate BAG3, expressing 9.7-fold less than Nod mice (p=0.0006), but CrmA had no significant effect in either strain. An associated differential migration of Ly6Chi and Ly6Ghi cells was also discovered in knockout mice with serpin treatments. Discussion One candidate gene found in human monocytes, BAG3, has reduced gene expression after Serp-2 treatment in mouse peritoneal exudates but increased by CrmA. BAG3 is known to alter cell migration and apoptosis, important to atherosclerotic progression. BAG3 is regulated by 3 myxomaviral proteins in human and mouse cells, underscoring a potential role as a lynchpin in viral protein anti-inflammatory and anti-apoptotic pathways.