Integrin-substrate interactions underlying shear-induced inhibition of the inflammatory response of endothelial cells

 

 Buy Article Permissions and Reprints

Summary

Conditioning of endothelial cells by shear stress suppresses their response to inflammatory cytokines. We questioned whether signalling through different integrin-matrix interactions, previously associated with the pathogenic effects of disturbed flow, supported the anti-inflammatory action of steady shear. Primary human endothelial cells were cultured on different substrates and exposed to shear stress (2.0Pa) for varying periods before stimulation with tumour necrosis factor-α (TNF). Shear-conditioning inhibited cytokine-induced recruitment of flowing neutrophils. However, the effect was similar for culture on collagen, laminin or fibronectin, even when seeding was reduced to 2hours, and shear to 3hours before TNF treatment (to minimise deposition of endothelial matrix). Nevertheless, in short- or longer- term cultures, reduction in expression of β₁-integrin (but not β₃-integrin) using siRNA essentially ablated the effect of shear-conditioning on neutrophil recruitment. Studies of focal adhesion kinase (FAK) phosphorylation, siRNA against FAK and a FAK-inhibitor (PF573228) indicated that FAK activity was an essential component downstream of β₁-integrin. In addition, MAP-kinase p38 was phosphorylated downstream of FAK and also required for functional modification. Mechanotransduction through β₁-integrins, FAK and p38 is required for anti-inflammatory effects of steady shear stress. Separation of the pathways which underlie pathological versus protective responses of different patterns of flow is required to enable therapeutic modification or mimicry, respectively.

^* Current address: Centre for Biological Engineering, Wolfson School of Mechanical and Manufacturing Engineering, Loughborough University, Loughborough, LE11 3TU, UK.

• References

• 1 Li S, Huang NF, Hsu S. Mechanotransduction in endothelial cell migration. J Cell Biochem 2005; 96: 1110-1126.
  CrossrefPubMedGoogle Scholar
• 2 Matharu NM, Rainger GE, Vohra R. et al. Effects of disturbed flow on endothelial cell function: Pathogenic implications of modified leukocyte recruitment. Biorheol 2006; 43: 31-44.
  PubMedGoogle Scholar
• 3 Chappell DC, Varner SE, Nerem RM. et al. Oscillatory shear stress stimulates adhesion molecule expression in cultured human endothelium. Circ Res 1998; 82: 532-539.
  CrossrefPubMedGoogle Scholar
• 4 Tzima E, Irani-Tehrani M, Kiosses WB. et al. A mechanosensory complex that mediates the endothelial cell response to fluid shear stress. Nature 2005; 437: 426-431.
  CrossrefPubMedGoogle Scholar
• 5 Surapisitchat J, Hoefen RJ, Pi X. et al. Fluid shear stress inhibits TNF-α activation of JNK but not ERK1/2 or p38 in human umbilical vein endothelial cells: Inhibitory crosstalk among MAPK family members. PNAS 2001; 98: 6476-6481.
  CrossrefPubMedGoogle Scholar
• 6 Sheikh S, Rainger GE, Gale Z. et al. Exposure to fluid shear stress modulates the ability of endothelial cells to recruit neutrophils in response to tumor necrosis factor-alpha: a basis for local variations in vascular sensitivity to inflammation. Blood 2003; 102: 2828-2834.
  CrossrefPubMedGoogle Scholar
• 7 Yamawaki H, Lehoux S, Berk BC. Chronic physiological shear stress inhibits tumor necrosis factor-induced proinflammatory responses in rabbit aorta perfused ex vivo. Circulation 2003; 108: 1619-1625.
  CrossrefPubMedGoogle Scholar
• 8 Chiu JJ, Lee PL, Chen CN. et al. Shear stress increases ICAM-1 and decreases VCAM-1 and E-selectin expressions induced by tumor necrosis factor-[alpha] in endothelial cells. Arterioscler Thromb Vasc Biol 2004; 24: 73-79.
  CrossrefPubMedGoogle Scholar
• 9 Luu N-T, Rahman M, Stone PC. et al. Responses of endothelial cells from different vessels to inflammatory cytokines and to shear stress: evidence for the pliability of endothelial phenotype. J Vasc Res 2010; 47: 451-461.
  CrossrefPubMedGoogle Scholar
• 10 Sheikh S, Rainger GE, Gale Z. et al. Differing mechanisms of leukocyte recruitment and sensitivity to conditioning by shear stress for endothelial cells treated with tumour necrosis factor-α or interleukin-1. Brit J Pharmacol 2005; 145: 1052-1061.
  CrossrefPubMedGoogle Scholar
• 11 Takahashi M, Ishida T, Traub O. et al. Mechanotransduction in endothelial cells: temporal signaling events in response to shear stress. J Vasc Res 1997; 34: 212-219.
  CrossrefPubMedGoogle Scholar
• 12 Chen K-D, Li Y-S, Kim M. et al. Mechanotransduction in response to shear stress. Roles of receptor tyrosine kinases, integrins, and Shc. J Biol Chem 1999; 274: 18393-18400.
  CrossrefPubMedGoogle Scholar
• 13 Shyy JY, Chien S. Role of integrins in endothelial mechanosensing of shear stress. Circ Res 2002; 91: 769-775.
  CrossrefPubMedGoogle Scholar
• 14 Orr AW, Sanders JM, Bevard M. et al. The subendothelial extracellular matrix modulates NF-kappaB activation by flow: a potential role in atherosclerosis. J Cell Biol 2005; 169: 191-202.
  CrossrefPubMedGoogle Scholar
• 15 Orr AW, Ginsberg MH, Shattil SJ. et al. Matrix-specific suppression of integrin activation in shear stress signaling. Mol Biol Cell 2006; 17: 4686-4697.
  CrossrefPubMedGoogle Scholar
• 16 Li S, Kim M, Hu YL. et al. Fluid shear stress activation of focal adhesion kinase. Linking to mitogen-activated protein kinases. J Biol Chem 1997; 272: 30455-30462.
  CrossrefPubMedGoogle Scholar
• 17 Petzold T, Orr AW, Hahn C. et al. Focal adhesion kinase modulates activation of NF-kappaB by flow in endothelial cells. Am J Physiol Cell Physiol 2009; 297: C814-C822.
  CrossrefPubMedGoogle Scholar
• 18 Chretien ML, Zhang M, Jackson MR. et al. Mechanotransduction by endothelial cells is locally generated, direction-dependent, and ligand-specific. J Cell Physiol 2010; 224: 352-361.
  CrossrefPubMedGoogle Scholar
• 19 Glen K, Luu NT, Ross E. et al. Modulation of functional responses of endothelial cells linked to angiogenesis and inflammation by shear stress: Differential effects of the mechanotransducer CD31. J Cell Physiol 2012; 227: 2710-2721.
  CrossrefPubMedGoogle Scholar
• 20 Cooke BM, Usami S, Perry I. et al. A simplified method for culture of endothelial cells and analysis of adhesion of blood cells under conditions of flow. Microvasc Res 1993; 45: 33-45.
  CrossrefPubMedGoogle Scholar
• 21 Sheikh S, Gale Z, Rainger GE. et al. Methods for exposing multiple cultures of endothelial cells to different fluid shear stresses and to cytokines, for subsequent analysis of inflammatory function. J Immunol Methods 2004; 288: 35-46.
  CrossrefPubMedGoogle Scholar
• 22 Luu NT, Rainger GE, Nash GB. Kinetics of the different steps during neutrophil migration through cultured endothelial monolayers treated with tumour necrosis factor-alpha. J Vasc Res 1999; 36: 477-485.
  CrossrefPubMedGoogle Scholar
• 23 Bahra P, Rainger GE, Wautier JL. et al. Each step during transendothelial migration of flowing neutrophils is regulated by the stimulatory concentration of tumour necrosis factor-alpha. Cell Ad Commun 1998; 06: 491-501.
  PubMedGoogle Scholar
• 24 SenBanerjee S, Lin Z, Atkins GB. et al. KLF2 Is a Novel Transcriptional Regulator of Endothelial Proinflammatory Activation. J Exp Med 2004; 199: 1305-1315.
  CrossrefPubMedGoogle Scholar
• 25 Butler LM, Rainger GE, Rahman M. et al. Prolonged culture of endothelial cells and deposition of basement membrane modify the recruitment of neutrophils. Exp Cell Res 2005; 310: 22-32.
  CrossrefPubMedGoogle Scholar