Montelukast (MLK) is a cysteinyl leukotriene receptor-1 (cysLT(1)R) antagonist with inhibitory effects on eosinophils, key proinflammatory cells in asthma. We assessed the effect of MLK on resting and GM-CSF-stimulated eosinophil adhesion to recombinant human (rh)VCAM-1 at different flow rates using our novel microflow system. At 1 or 2 dyn cm(-2), shear-stress unstimulated eosinophils tethered immediately to rhVCAM-1, "rolled" along part of the channel until they tethered, or rolled without tethering. At flow rates greater than 2 dyn cm(-2), adherent eosinophils began to be displaced from rhVCAM-1. MLK (10 nM and 100 nM) gave partial ( approximately 40%) but significant (P<0.05) inhibition of unstimulated eosinophil adhesion to rhVCAM-1 at 1 or 2 dyn cm(-2) shear stress. Once adhered, unstimulated eosinophils did not exhibit morphological changes, and GM-CSF-stimulated eosinophil adhesion under flow was characterized by greater cell flattening with significant (P<0.05) inhibition of adherent cell numbers by 100 nM MLK observed. This effect appeared specific for MLK, as the analog (E)-3-[[[3-[2-(7-chloro-2-quinolinyl)ethenyl]phenyl]-[[3-dimethylamino)-3-oxopropyl]thio]methyl]thio]-propanoic acid, sodium salt, had no significant effect on eosinophil adhesion to VCAM-1. The possibility that LTC(4), released from unstimulated or GM-CSF-treated eosinophils, contributed to their adhesion to VCAM-1 was excluded as the LT biosynthesis inhibitor 3-[1-(p-Chlorobenzyl)-5-(isopropyl)-3-t-butylthioindol-2-yl]-2,2-dimethylpropanoic acid had no inhibitory effect, and exogenously added LTC(4) did not enhance eosinophil adhesion. In contrast, LTD(4) enhanced eosinophil adhesion to VCAM-1, an effect blocked by MLK (10 and 100 nM). These findings demonstrate that MLK-mediated inhibition of unstimulated and GM-CSF-stimulated eosinophil adhesion to VCAM-1 under shear-stress conditions appears independent of cysLT(1)R antagonism.

Vascular endothelium is a potential target for therapeutic intervention in diverse pathological processes, including inflammation, atherosclerosis, and thrombosis. By virtue of their intravascular topography, endothelial cells are exposed to dynamically changing mechanical forces that are generated by blood flow. In the present study, we investigated the interactions of negatively charged 2.7 nm and 4.7 nm CdTe quantum dots and 50 nm silica particles with cultured endothelial cells under regulated shear stress (SS) conditions. Cultured cells within the engineered microfluidic channels were exposed to nanoparticles under static condition or under low, medium, and high SS rates (0.05, 0.1, and 0.5 Pa, respectively). Vascular inflammation and associated endothelial damage were simulated by treatment with tumor necrosis factor-α (TNF-α) or by compromising the cell membrane with the use of low Triton X-100 concentration. Our results demonstrate that SS is critical for nanoparticle uptake by endothelial cells. Maximal uptake was registered at the SS rate of 0.05 Pa. By contrast, endothelial exposure to mild detergents or TNF-α treatment had no significant effect on nanoparticle uptake. Atomic force microscopy demonstrated the increased formation of actin-based cytoskeletal structures, including stress fibers and membrane ruffles, which have been associated with nanoparticle endocytosis. In conclusion, the combinatorial effects of SS rates, vascular endothelial conditions, and nanoparticle physical and chemical properties must be taken into account for the successful design of nanoparticle-drug conjugates intended for parenteral delivery. Keywords: endothelium, shear stress, quantum dots, membrane ruffling, stress fibers, atomic force microscopy, microfluidics Nanoparticle (NP) technologies are significantly affecting the development of both therapeutic and diagnostic agents. Although enormous progress in the field of nanotechnology has been achieved, basic discoveries have not yet translated into effective targeted therapies. NPs can potentially improve the pharmacokinetics and pharmacodynamics of drugs; however, the complexity of in vivo systems imposes multiple barriers that severely inhibit efficiency, which must be overcome to fully exploit the theoretical potential of NPs. Endothelial cells (ECs) that line the interior of the entire vascular system represent a major barrier for therapeutic agents traveling from the bloodstream to the target tissues. Recent studies have focused on targeting the endothelium with NPs as therapeutic agents for a variety of pathological conditions in the vascular system because of the large population of ECs and their proximity to the blood flow.ECs in vivo are exposed to a variety of hemodynamic forces that are created by blood flow and by the pulse wave dictated by the cardiac cycle. Shear stress (SS) is the dragging mechanical force that acts at the interface between flowing blood and Dovepress submit your manuscript | www.dovepress.com the vessel wall. ECs recognize SS a...

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Inhibition of eosinophil adhesion to ICAM-1 by fluvastatin and lovastatin under physiological shear stress represent novel actions by these drugs that may inform the development of anti-inflammatory therapy for allergic disease.

Several studies have recognised the benefit of patients receiving statin therapy in reducing the incidence of sepsis in ICU patients. The potential protective effect of statin therapy reducing the incidence of sepsis is probably due to the immunomodulatory mechanisms of these compounds that are capable of inhibiting the inflammatory pathways. LFA‐1 is an integrin that is critical for T‐cell adhesion, T‐cell priming and cytokine secretion. Statins inhibit LFA‐1 interaction with ICAM by either directly binding to a novel site that is distant from the ICAM binding site (not pravastatin) or inhibit the prenylation of small GTPases. Our results have shown that physiologically relevant concentrations of fluva‐, meva‐, lova‐, simva‐ or pravastatin markedly inhibit T‐cell adhesion to ICAM equally, irrespective of direct LFA‐1 binding. Therefore, the reported clinical outcome of septic patients receiving statin therapy can be at least be partially associated with T‐cell‐ICAM binding activities. In conclusion the Cellix Microfluidic Platform allows for T‐cell adhesion to be accurately determined using physiological shear stress with physiologically relevant concentrations of statins.

Leukocyte adhesion to endothelial cell bound proteins, such as ICAM-1 and VCAM-1, is an initial step of the inflammatory response. We have developed an in vitro microfluidic system which mimics conditions found in blood vessels in vivo during an immune response. Using this system, we can record leukocyte adhesion levels under physiologically relevant flow conditions (e.g. 0–5 dynes/cm2). The adhesion profiles of resting and PMA-stimulated peripheral blood lymphocytes (PBLs) were recorded, with respect to VCAM-1, ICAM-1, and BSA. Images at each shear stress level were captured using a digital camera, and analysed using our in-house Ducocell software package. Distinct morphological changes in PMA-stimulated PBLs, compared to non-stimulated cells, can be observed. These include a less rounded appearance of the PMA-stimulated PBLs, and evidence of “uropod” formation, which anchor the T cell to the endothelium as part of the migration process. Levels of adhesion to VCAM-1 are high (80–90%, compared to control), but there appears to be little difference between the adhesion profiles of non-stimulated and PMA-stimulated PBLs. However, there is a distinct difference between the adhesion levels of non-stimulated and PMA-stimulated PBLs to ICAM-1, with PMA-stimulated cells showing a higher affinity for ICAM-1 than non-stimulated cells (approx. 70% and 40%, respectively). PBL adhesion to BSA is negligible.