Functional Genomics of Chlorine-induced Acute Lung Injury in Mice

Acute lung injury can be induced indirectly (e.g., sepsis) or directly (e.g., chlorine inhalation). Because treatment is still limited to supportive measures, mortality remains high (∼74,500 deaths/yr). In the past, accidental (railroad derailments) and intentional (Iraq terrorism) chlorine exposures have led to deaths and hospitalizations from acute lung injury. To better understand the molecular events controlling chlorine-induced acute lung injury, we have developed a functional genomics approach using inbred mice strains. Various mouse strains were exposed to chlorine (45 ppm × 24 h) and survival was monitored. The most divergent strains varied by more than threefold in mean survival time, supporting the likelihood of an underlying genetic basis of susceptibility. These divergent strains are excellent models for additional genetic analysis to identify critical candidate genes controlling chlorine-induced acute lung injury. Gene-targeted mice then could be used to test the functional significance of susceptibility candidate genes, which could be valuable in revealing novel insights into the biology of acute lung injury

MALT1 Protease Activation Triggers Acute Disruption of Endothelial Barrier Integrity via CYLD Cleavage

Microvascular endothelial cells maintain a tight barrier to prevent passage of plasma and circulating immune cells into the extravascular tissue compartment, yet endothelial cells respond rapidly to vasoactive substances, including thrombin, allowing transient paracellular permeability. This response is a cornerstone of acute inflammation, but the mechanisms responsible are still incompletely understood. Here, we demonstrate that thrombin triggers MALT1 to proteolytically cleave cylindromatosis (CYLD). Fragmentation of CYLD results in microtubule disruption and a cascade of events leading to endothelial cell retraction and an acute permeability response. This finding reveals an unexpected role for the MALT1 protease, which previously has been viewed mostly as a driver of pro-inflammatory NF-κB signaling in lymphocytes. Thus, MALT1 not only promotes immune cell activation but also acutely regulates endothelial cell biology, actions that together facilitate tissue inflammation. Pharmacologic inhibition of MALT1 may therefore have synergistic impact by targeting multiple disparate steps in the overall inflammatory response

Surfactant-Associated Protein B Is Critical to Survival in Nickel-Induced Injury in Mice

The etiology of acute lung injury is complex and associated with numerous, chemically diverse precipitating factors. During acute lung injury in mice, one key event is epithelial cell injury that leads to reduced surfactant biosynthesis. We have previously reported that transgenic mice that express transforming growth factor α (TGFA) in the lung were protected during nickel-induced lung injury. Here, we find that the mechanism by which TGFA imparts protection includes maintenance of surfactant-associated protein B (SFTPB) transcript levels and epidermal growth factor receptor–dependent signaling in distal pulmonary epithelial cells. This protection is complex and not accompanied by a diminution in inflammatory mediator transcripts or additional stimulation of antioxidant transcripts. In mouse lung epithelial (MLE-15) cells, microarray analysis demonstrated that nickel increased transcripts of genes enriched in MTF1, E2F-1, and AP-2 transcription factor–binding sites and decreased transcripts of genes enriched in AP-1–binding sites. Nickel also increased Jun transcript and DNA-binding activity, but decreased SFTPB transcript. Expression of SFTPB under the control of a doxycycline-sensitive promoter increased survival during nickel-induced injury as compared with control mice. Together, these findings support the idea that maintenance of SFTPB expression is critical to survival during acute lung injury

Functional Genomic Assessment of Phosgene-Induced Acute Lung Injury in Mice

In this study, a genetically diverse panel of 43 mouse strains was exposed to phosgene and genome-wide association mapping performed using a high-density single nucleotide polymorphism (SNP) assembly. Transcriptomic analysis was also used to improve the genetic resolution in the identification of genetic determinants of phosgene-induced acute lung injury (ALI). We prioritized the identified genes based on whether the encoded protein was previously associated with lung injury or contained a nonsynonymous SNP within a functional domain. Candidates were selected that contained a promoter SNP that could alter a putative transcription factor binding site and had variable expression by transcriptomic analyses. The latter two criteria also required that ≥10% of mice carried the minor allele and that this allele could account for ≥10% of the phenotypic difference noted between the strains at the phenotypic extremes. This integrative, functional approach revealed 14 candidate genes that included Atp1a1, Alox5, Galnt11, Hrh1, Mbd4, Phactr2, Plxnd1, Ptprt, Reln, and Zfand4, which had significant SNP associations, and Itga9, Man1a2, Mapk14, and Vwf, which had suggestive SNP associations. Of the genes with significant SNP associations, Atp1a1, Alox5, Plxnd1, Ptprt, and Zfand4 could be associated with ALI in several ways. Using a competitive electrophoretic mobility shift analysis, Atp1a1 promoter (rs215053185) oligonucleotide containing the minor G allele formed a major distinct faster-migrating complex. In addition, a gene with a suggestive SNP association, Itga9, is linked to transforming growth factor β1 signaling, which previously has been associated with the susceptibility to ALI in mice