Epidemiological studies suggest a correlation between increased airborne particulate matter (PM) and adverse health effects. The mechanisms of PM-health effects are believed to involve oxidative stress and inflammation. To evaluate the ability of PM promoting skin tissue damage, one of the main organs exposed to outdoor pollutants, we analyzed the effect of concentrated ambient particles (CAPs) in a reconstructed human epidermis (RHE) model. RHE tissues were exposed to 25 or 100 µg/ml CAPs for 24 or 48 h. Data showed that RHE seems to be more susceptible to CAPs-induced toxicity after 48 h exposure than after 24 h. We found a local reactive O(2) species (ROS) production increase generated from metals present on the particle, which contributes to lipids oxidation. Furthermore, as a consequence of altered redox status, NFkB nucleus translocation was increase upon CAPs exposure, as well as cyclooxygenase 2 and cytochrome P450 levels, which may be involved in the inflammatory response initiated by PM. CAPs also triggered an apoptotic process in skin. Surprisingly, by transition electron microscopy analysis we showed that CAPs were able to penetrate skin tissues. These findings contribute to the understanding of the cutaneous pathophysiological mechanisms initiated by CAPs exposure, where oxidative stress and inflammation may play predominant roles.

Rationale CD40L figures prominently in chronic inflammatory diseases such as atherosclerosis. However, since CD40L potently regulates immune function and haemostasis by interaction with CD40 receptor and the platelet integrin GPIIb/IIIa, its global inhibition compromises host defense and generated thromboembolic complications in clinical trials. We recently reported that CD40L mediates atherogenesis independently of CD40 and proposed Mac-1 as an alternate receptor. Objective Here, we molecularly characterized the CD40L-Mac-1 interaction and tested whether its selective inhibition by a small peptide modulates inflammation and atherogenesis in vivo. Methods and Results CD40L concentration-dependently bound to Mac-1 I-domain in solid phase binding assays, and a high affinity interaction was revealed by surface-plasmon-resonance analysis. We identified the motif EQLKKSKTL, an exposed loop between the α1 helix and the β-sheet B, on Mac-1 as binding site for CD40L. A linear peptide mimicking this sequence, M7, specifically inhibited the interaction of CD40L and Mac-1. cM7, a cyclisized version optimized for in vivo use, decreased peritoneal inflammation and inflammatory cell recruitment in vivo. Finally, LDLr-/- mice treated with intraperitoneal injections of cM7 developed smaller, less inflamed atherosclerotic lesions featuring characteristics of stability. However, cM7 did not interfere with CD40L-CD40 binding in vitro and CD40L-GPIIb/IIIa-mediated thrombus formation in vivo. Conclusions We present the novel finding that CD40L binds to the EQLKKSKTL motif on Mac-1 mediating leukocyte recruitment and atherogenesis. Specific inhibition of CD40L-Mac-1 binding may represent an attractive anti-inflammatory treatment strategy for atherosclerosis and other inflammatory conditions, potentially avoiding the unwanted immunologic and thrombotic effects of global inhibition of CD40L.

BACKGROUND: Platelets store large amounts of serotonin that they release during thrombus formation or acute inflammation. This facilitates hemostasis and modulates the inflammatory response. METHODS:Infarct size, heart function, and inflammatory cell composition were analyzed in mouse models of myocardial reperfusion injury with genetic and pharmacological depletion of platelet serotonin. These studies were complemented by in vitro serotonin stimulation assays of platelets and leukocytes in mice and men, and by measuring plasma serotonin levels and leukocyte activation in patients with acute coronary syndrome. RESULTS:Platelet-derived serotonin induced neutrophil degranulation with release of myeloperoxidase and hydrogen peroxide (H 2 O 2 ) and increased expression of membrane-bound leukocyte adhesion molecule CD11b, leading to enhanced inflammation in the infarct area and reduced myocardial salvage. In patients hospitalized with acute coronary syndrome, plasmatic serotonin levels correlated with CD11b expression on neutrophils and myeloperoxidase plasma levels. Longterm serotonin reuptake inhibition-reported to protect patients with depression from cardiovascular events-resulted in the depletion of platelet serotonin stores in mice. These mice displayed a reduction in neutrophil degranulation and preserved cardiac function. In line, patients with depression using serotonin reuptake inhibition, presented with suppressed levels of CD11b surface expression on neutrophils and lower myeloperoxidase levels in blood. CONCLUSIONS:Taken together, we identify serotonin as a potent therapeutic target in neutrophil-dependent thromboinflammation during myocardial reperfusion injury.