title: Potentiation and tolerance of toll-like receptor priming in human endothelial cells.
abstract: Repeated challenge of lipopolysaccharide (LPS) alters the response to subsequent LPS exposures via modulation of toll-like receptor 4 (TLR4). Whether activation of other TLRs can modulate TLR4 responses, and vice versa, remains unclear. Specifically with regards to endothelial cells, a key component of innate immunity, the impact of TLR cross-modulation is unknown. We postulated that TLR2 priming (via Pam3Csk4) would inhibit TLR4-mediated responses while TLR3 priming (via Poly I:C) would enhance subsequent TLR4-inflammatory signaling. We studied human umbilical vein endothelial cells (HUVECs) and neonatal human dermal microvascular endothelial cells (HMVECs). Cells were primed with a combination of Poly I:C (10 μg/ml), Pam3Csk4 (10 μg/ml), or LPS (100 ng/ml), then washed and allowed to rest. They were then rechallenged with either Poly I:C, Pam3Csk4 or LPS. Endothelial cells showed significant tolerance to repeated LPS challenge. Priming with Pam3Csk4 also reduced the response to secondary LPS challenge in both cell types, despite a reduced proinflammatory response to Pam3Csk4 in HMVECs compared to HUVECs. Poly I:C priming enhanced inflammatory and interferon producing signals upon Poly I:C or LPS rechallenge, respectively. Poly I:C priming induced interferon regulatory factor 7, leading to enhancement of interferon production. Finally, both Poly I:C and LPS priming induced significant changes in receptor-interacting serine/threonine-protein kinase 1 activity. Pharmacological inhibition of receptor-interacting serine/threonine-protein kinase 1 or interferon regulatory factor 7 reduced the potentiated phenotype of TLR3 priming on TLR4 rechallenge. These results demonstrate that in human endothelial cells, prior activation of TLRs can have a significant impact on subsequent exposures and may contribute to the severity of the host response.
authors: Department of Pediatrics, Vanderbilt University Medical Center, Nashville, Tenn.
authors: Stephen R
authors: SR
authors: Koch
authors: Department of Pediatrics, Vanderbilt University Medical Center, Nashville, Tenn.
authors: Fred S
authors: FS
authors: Lamb
authors: Department of Anesthesiology and Perioperative Care, University of California San Francisco, San Francisco, Calif.
authors: Judith
authors: J
authors: Hellman
authors: Department of Anesthesiology, Vanderbilt University Medical Center, Nashville, Tenn.
authors: Edward R
authors: ER
authors: Sherwood
authors: Department of Pediatrics, Vanderbilt University Medical Center, Nashville, Tenn. Electronic address: ryan.stark@vanderbilt.edu.
authors: Ryan J
authors: RJ
authors: Stark
mesh_terms: D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007108:Immune Tolerance / Q000187:drug effects; D050839:Interferon Regulatory Factor-7 / Q000378:metabolism; D007372:Interferons / Q000378:metabolism; D015850:Interleukin-6 / Q000096:biosynthesis; D055666:Lipopeptides / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D028861:Nuclear Pore Complex Proteins / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011070:Poly I-C / Q000494:pharmacology; D016601:RNA-Binding Proteins / Q000378:metabolism; D051193:Toll-Like Receptors / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects
chemical_list: C119724:AGFG1 protein, human; C494234:IRF7 protein, human; D050839:Interferon Regulatory Factor-7; D015850:Interleukin-6; D055666:Lipopeptides; D008070:Lipopolysaccharides; D028861:Nuclear Pore Complex Proteins; C508075:Pam(3)CSK(4) peptide; D016601:RNA-Binding Proteins; D051193:Toll-Like Receptors; D007372:Interferons; D048049:Extracellular Signal-Regulated MAP Kinases; D011070:Poly I-C