Potentiation and tolerance of toll-like receptor priming in human endothelial cells

• Published in: Translational research : the journal of laboratory and clinical medicine 2017-02, Vol.180, p.53-67.e4
• Main Authors: Koch, Stephen R, Lamb, Fred S, Hellman, Judith, Sherwood, Edward R, Stark, Ryan J
• Format: Article
• Language: English
• Subjects: Endothelial Cells - drug effects; Endothelial Cells - metabolism; Extracellular Signal-Regulated MAP Kinases - metabolism; Human Umbilical Vein Endothelial Cells - drug effects; Human Umbilical Vein Endothelial Cells - metabolism; Humans; Immune Tolerance - drug effects; Interferon Regulatory Factor-7 - metabolism; Interferons - metabolism; Interleukin-6 - biosynthesis; Internal Medicine; Lipopeptides - pharmacology; Lipopolysaccharides - pharmacology; Nuclear Pore Complex Proteins - metabolism; Phosphorylation - drug effects; Poly I-C - pharmacology; RNA-Binding Proteins - metabolism; Toll-Like Receptors - metabolism; Up-Regulation - drug effects
• ISSN: 1931-5244; 1878-1810
• DOI: 10.1016/j.trsl.2016.08.001

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.