Rationale: Acute lung injury (ALI) causes high mortality, but its molecular mechanisms and therapeutic options remain ill-defined. Gram-negative bacterial infections are the main cause of ALI, leading to lung neutrophil infiltration, permeability increases, deterioration of gas exchange, and lung damage. Platelets are activated during ALI, but insights into their mechanistic contribution to neutrophil accumulation in the lung are elusive. Objectives: To determine mechanisms of platelet-mediated neutrophil recruitment in ALI. Methods: Interference with platelet–neutrophil interactions using antagonists to P-selectin and glycoprotein IIb/IIIa or a small peptide antagonist disrupting platelet chemokine heteromer formation in mouse models of ALI. Measurements and Main Results: In a murine model of LPS-induced ALI, we uncover important roles for neutrophils and platelets in permeability changes and subsequent lung damage. Furthermore, platelet depletion abrogated lung neutrophil infiltration, suggesting a sequential participation of platelets and neutrophils. Whereas antagonists to P-selectin and glycoprotein IIb/IIIa had no effects on LPS-mediated ALI, antibodies to the platelet-derived chemokines CCL5 and CXCL4 strongly diminished neutrophil eflux and permeability changes. The two chemokines were found to form heteromers in human and murine ALI samples, positively correlating with leukocyte influx into the lung. Disruption of CCL5-CXCL4 heteromers in LPS-, acid-, and sepsis-induced ALI abolished lung edema, neutrophil infiltration, and tissue damage, thereby revealing a causal contribution. Conclusions: Taken together, our data identify a novel function of platelet-derived chemokine heteromers during ALI and demonstrate means for therapeutic interference.

Prematurely born infants who require oxygen therapy often develop bronchopulmonary dysplasia (BPD), a debilitating disorder characterized by pronounced alveolar hypoplasia. Hyperoxic injury is believed to disrupt critical signaling pathways that direct lung development, causing BPD. We investigated the effects of normobaric hyperoxia on transforming growth factor (TGF)-β and bone morphogenetic protein (BMP) signaling in neonatal C57BL/6J mice exposed to 21% or 85% O₂between postnatal days P1 and P28. Growth and respiratory compliance were significantly impaired in pups exposed to 85% O₂, and these pups also exhibited a pronounced arrest of alveolarization, accompanied by dysregulated expression and localization of both receptor (ALK-1, ALK-3, ALK-6, and the TGF-β type II receptor) and Smad (Smads 1, 3, and 4) proteins. TGF-β signaling was potentiated, whereas BMP signaling was impaired both in the lungs of pups exposed to 85% O₂as well as in MLE-12 mouse lung epithelial cells and NIH/3T3 and primary lung fibroblasts cultured in 85% O₂. After exposure to 85% O₂, primary alveolar type II cells were more susceptible to TGF-β-induced apoptosis, whereas primary pulmonary artery smooth muscle cells were unaffected. Exposure of primary lung fibroblasts to 85% O₂significantly enhanced the TGF-β-stimulated production of the α₁subunit of type I collagen (Iα₁), tissue inhibitor of metalloproteinase-1, tropoelastin, and tenascin-C. These data demonstrated that hyperoxia significantly affects TGF-β/BMP signaling in the lung, including processes central to septation and, hence, alveolarization. The amenability of these pathways to genetic and pharmacological manipulation may provide alternative avenues for the management of BPD.

Despite the immune-privileged status of the male genital tract, infection and inflammation of the male genital tract are important etiological factors in male infertility. A common observation in clinical and experimental orchitis as well as in systemic infection and inflammation are decreased levels of testosterone. Emerging data point to an immunosuppressive role of testosterone. In our study, we substituted testosterone levels in experimental autoimmune orchitis (EAO) in rat by s.c. testosterone implants. EAO development was reduced to 17% when animals were treated with low-dose testosterone implants (3 cm long, EAO+T3) and to 33% when rats were supplied with high-dose testosterone implants (24 cm, EAO+T24) compared with 80% of animals developing disease in the EAO control group. In the testis, testosterone replacement in EAO animals prevented the accumulation of macrophages and significantly reduced the number of CD4+ T cells with a strong concomitant increase in the number of regulatory T cells (CD4+CD25+Foxp3+) compared with EAO control. In vitro testosterone treatment of naive T cells led to an expansion of the regulatory T cell subset with suppressive activity and ameliorated MCP-1–stimulated chemotaxis of T lymphocytes in a Transwell assay. Moreover, expression of proinflammatory mediators such as MCP-1, TNF-α, and IL-6 in the testis and secretion of Th1 cytokines such as IFN-γ and IL-2 by mononuclear cells isolated from testicular draining lymph nodes were decreased in the EAO+T3 and EAO+T24 groups. Thus, our study shows an immunomodulatory and protective effect of testosterone substitution in the pathogenesis of EAO and suggests androgens as a new factor in the differentiation of regulatory T cells.

Cell surface–associated proteolysis plays a crucial role in the migration of mononuclear phagocytes to sites of inflammation. The glycolytic enzyme enolase-1 (ENO-1) binds plasminogen at the cell surface, enhancing local plasmin production. This study addressed the role played by ENO-1 in lipopolysaccharide (LPS)–driven chemokine-directed monocyte migration and matrix invasion in vitro, as well as recruitment of monocytes to the alveolar compartment in vivo. LPS rapidly up-regulated ENO-1 cell-surface expression on human blood monocytes and U937 cells due to protein translocation from cytosolic pools, which increased plasmin generation, enhanced monocyte migration through epithelial monolayers, and promoted matrix degradation. These effects were abrogated by antibodies directed against the plasminogen binding site of ENO-1. Overexpression of ENO-1 in U937 cells increased their migratory and matrix-penetrating capacity, which was suppressed by overexpression of a truncated ENO-1 variant lacking the plasminogen binding site (ENO-1ΔPLG). In vivo, intratracheal LPS application in mice promoted alveolar recruitment of monocytic cells that overexpressed ENO-1, but not of cells overexpressing ENO-1ΔPLG. Consistent with these data, pneumonia-patients exhibited increased ENO-1 cell-surface expression on blood monocytes and intense ENO-1 staining of mononuclear cells in the alveolar space. These data suggest an important mechanism of inflammatory cell invasion mediated by increased cell-surface expression of ENO-1.

It is evident that components of the extracellular matrix (ECM) act as danger-associated molecular patterns (DAMPs) through direct interactions with pattern recognition receptors (PRRs) including Toll-like receptors (TLRs) and inflammasomes. Through these interactions, ECM-derived DAMPs autonomously trigger sterile inflammation or prolong pathogen-induced responses through the production of proinflammatory mediators and the recruitment of leukocytes to sites of injury and infection. Recent research, however, suggests that ECM-derived DAMPs are additionally involved in the resolution and fine-tuning of inflammation by orchestrating the production of anti-inflammatory mediators that are required for the resolution of tissue inflammation and the transition to acquired immunity. Thus, in this review, we discuss the current knowledge of the interplay between ECM-derived DAMPs and the innate immune signaling pathways that are activated to provide temporal control of innate immunity.