Atherosclerosis of native coronary arteries and graft arteriosclerosis in transplanted hearts are characterized by activation of innate and adaptive immune responses. Nucleic acids generated by infections or cell death have been detected within arteriosclerotic lesions, and it is known that microbial and synthetic nucleic acids evoke inflammatory responses in cultured vascular cells. In this study, we report that model RNA, but not DNA, instigated robust cytokine and chemokine production from intact human coronary arteries containing both intrinsic vascular cells and resident/infiltrating leukocytes. An ssRNA analog induced TNF-alpha and IFN-gamma-induced protein of 10 kDa secretion by isolated human PBMCs, but not vascular cells. Conversely, synthetic dsRNA induced these inflammatory mediators by vascular cells, but not PBMCs. IFN-gamma, a cytokine linked to atherosclerosis and graft arteriosclerosis, potentiated the inflammatory responses of intact arteries and cultured vascular smooth muscle cells (VSMCs) to polyinosinic:polycytidylic acid [poly(I:C)] and was necessary for inflammatory responses of VSMC to self-RNA derived from autologous cells. IFN-gamma also induced the expression of TLR3, melanoma differentiation-associated gene 5, and retinoic acid-inducible gene I dsRNA receptors. Small interfering RNA knockdown revealed that TLR3 mediated VSMC activation by poly(I:C), whereas melanoma differentiation-associated gene 5 was more important for VSMC stimulation by self-RNA. IFN-gamma-mediated induction of dsRNA receptors and priming for inflammatory responses to poly(I:C) was confirmed in vivo using immunodeficient mice bearing human coronary artery grafts. These findings suggest that IFN-gamma, and by inference adaptive immunity, sensitizes the vasculature to innate immune activators, such as RNA, and activation of IFN-gamma-primed vascular cells by exogenous or endogenous sources of RNA may contribute to the inflammatory milieu of arteriosclerosis.