Kaposi's sarcoma (KS) is strongly associated with KS herpes virus infection, and inflammation plays an important role in this disease. We have shown that human KS biopsy-derived SLK cells, which are of endothelial origin and form KS-like tumors in nude mice, express the viral RNA pattern recognition receptors Toll-like receptor 3 (TLR3), retinoic acid-inducible gene-I (RIG-I), and melanoma-differentiation-associated gene 5 (MDA5). Furthermore, SLK cells have enhanced release of IL-6, IL-8 (CXCL8), RANTES (CCL5), and IP-10 (CXCL10) proteins in response to the synthetic viral RNA analog poly(I:C). SiRNA knockdowns demonstrated that TLR3 mediates this inflammatory response to poly(I:C) in SLK cells. Furthermore, knockdown of the RNA receptor RIG-I resulted in enhanced chemokine release, in a TLR3 pathway-dependent manner. Thus, exposure of KS cells to viral RNA ligands can result in a TLR3-mediated increase in the secretion of inflammatory proteins associated with KS cell growth that may contribute to disease.