Neutrophils play an important role in host defense. However, deregulation of neutrophils contributes to tissue damage in severe systemic inflammation. In contrast to complications mediated by an overactive neutrophil compartment, severe systemic inflammation is a risk factor for development of immune suppression and as a result, infectious complications. The role of neutrophils in this clinical paradox is poorly understood, and in this study, we tested whether this paradox could be explained by distinct neutrophil subsets and their functionality. We studied the circulating neutrophil compartment immediately after induction of systemic inflammation by administering 2 ng/kg Escherichia coli LPS i.v. to healthy volunteers. Neutrophils were phenotyped by expression of membrane receptors visualized by flow cytometry, capacity to interact with fluorescently labeled microbes, and activation of the NADPH-oxidase by oxidation of Amplex Red and dihydrorhodamine. After induction of systemic inflammation, expression of membrane receptors on neutrophils, such as CXCR1 and -2 (IL-8Rs), C5aR, FcgammaRII, and TLR4, was decreased. Neutrophils were also refractory to fMLF-induced up-regulation of membrane receptors, and suppression of antimicrobial function was shown by decreased interaction with Staphylococcus epidermis. Simultaneously, activation of circulating neutrophils was demonstrated by a threefold increase in release of ROS. The paradoxical phenotype can be explained by the selective priming of the respiratory burst. In contrast, newly released, CD16(dim) banded neutrophils display decreased antimicrobial function. We conclude that systemic inflammation leads to a functionally heterogeneous neutrophil compartment, in which newly released refractory neutrophils can cause susceptibility to infections, and activated, differentiated neutrophils can mediate tissue damage.

While ex vivo lipopolysaccharide tolerance quickly resolves, in vivo lipopolysaccharide tolerance persists for at least 2 wks. These findings strengthen the notion that the in vivo response to lipopolysaccharide is mediated by tissue-resident macrophages and that ex vivo stimulation does not accurately reflect the in vivo innate immune response. Intervention studies utilizing the human endotoxemia model should be performed using parallel groups rather than a crossover design.

Adequate responses by our innate immune system toward invading pathogens were of vital importance for surviving infections, especially before the antibiotic era. Recently, a polymorphism in Mal (Ser180Leu, TIRAP rs8177374), an important adaptor protein downstream of the Toll-like receptor (TLR) 2 and 4 pathways, has been described to provide protection against a broad range of infectious pathogens. We assessed the functional effects of this polymorphism in human experimental endotoxemia, and we demonstrate that individuals bearing the TIRAP 180L allele display an increased, innate immune response to TLR4 and TLR2 ligands, but not to TLR9 stimulation. This phenotype has been related to an increased resistance to infection. However, an overshoot in the release of proinflammatory cytokines by TIRAP 180L homozygous individuals suggests a scenario of balanced evolution. We have also investigated the worldwide distribution of the Ser180Leu polymorphism in 14 populations around the globe to correlate the genetic makeup of TIRAP with the local infectious pressures. Based on the immunological, clinical, and genetic data, we propose that this mutation might have been selected in West Eurasia during the early settlement of this region after the out-of-Africa migration of modern Homo sapiens. This combination of functional and genetic data provides unique insights to our understanding of the pathogenesis of sepsis.Innate immunity ͉ TLR4 ͉ TLR2 ͉ evolution ͉ cytokines T he nonsynonymous single nucleotide polymorphism Ser180Leu (S180L) of the TIR domain-coding adaptor protein (TIRAP) gene, also known as adaptor protein Mal (1), was found to be protective in heterozygous individuals for a broad range of infectious diseases such as malaria, tuberculosis, bacteremia, and pneumococcal disease (1). Mal is involved in the signaling pathways of Toll-like receptors (TLR) 2 and 4, which are important innate immune receptors for the recognition of a broad range of pathogenic Gram-negative and Gram-positive bacteria (2). Recognition by TLRs eventually leads to activation of NF-B and the transcription of proinflammatory genes that activate the mechanisms of host defense and the clearance of the pathogens. However, little is known about the in vivo functional consequences of the S180L polymorphism or the possible differences in the geographic distribution of the S180L polymorphism in relation to infectious pressure.In this study we hypothesized that the difference in the frequency of TIRAP 180L between the different geographical regions was due to the effects of the mutation on the function of TLR signaling, and was shaped by the pressure exerted by different infections, either during the out-of-Africa migration of early modern humans during the early Upper Paleolithic period (60-40kya) (3) or the Neolithic demic expansion from the Near East into Europe (Ϸ10,000kya) (4, 5). This pressure may have resulted in an increase of the frequency of TIRAP S180L heterozygous individuals and thereby a strong rise of the frequency of the TIRAP 180L al...

Background-Nonlethal ischemia and reperfusion reduce ischemia-reperfusion-induced cell death, a phenomenon called ischemic preconditioning. In animal models, this potent endogenous protection is mimicked in vivo by administration of adenosine. In humans, exploitation of ischemic preconditioning is hindered by the lack of an appropriate in vivo model to study this phenomenon. To solve this problem, we aimed to set up an easy-to-use human in vivo model to study ischemic or pharmacological preconditioning. Methods and Results-Healthy male volunteers performed unilateral ischemic handgrip. At reperfusion, we intravenously injected technetium-99m-labeled Annexin A5, a presumed marker of ischemic injury, and we imaged both forearms and hands simultaneously with a gamma camera. Region of interest analysis (counts per pixel) and subsequent calculation of the percentage difference in radioactivity between experimental and control hands (thenar muscle; meanϮSE) revealed significant uptake to the ischemically exercised tissue (26Ϯ3% at 4 hours after reperfusion; PϽ0.05). This selective localization of Annexin A5 was reduced by ischemic preconditioning (10 minutes of ischemia plus reperfusion before ischemic exercise) or by infusion of adenosine into the brachial artery to 6Ϯ1% and 10Ϯ3%, respectively (PϽ0.05 versus ischemic exercise alone), resembling observations in animal models with infarct size as an end point.Appropriate control experiments supported our conclusion. Conclusions-Annexin

Caffeine abolishes IP in 2 human models at a dose equivalent to the drinking of 2 to 4 cups of coffee. (The Effect of Caffeine on Ischemic Preconditioning; http://clinicaltrials.gov/ct/show/NCT00184912?order=1; NCT00184912).