© 2003 American Thoracic Society
Genomics Made Functional in Ventilator-associated Lung InjuryCenter for Translational Respiratory Medicine Division of Pulmonary and Critical Care Medicine Johns Hopkins University Baltimore, Maryland Mechanical ventilation is now recognized as potentially directly harmful to patients with acute lung injury. The benefits of lower tidal volumes to achieve lower airway pressures (and thus reduce lung cell stretch) have been clearly confirmed by the landmark ARDSnet findings (1). Each additional day on the ventilator further increases the development of malnutrition, nosocomial-, and ventilator-associated pneumonia. Important studies by Parker and coworkers (2, 3) and Webb and Tierney (4) have demonstrated changes in microvascular permeability in isolated lung and intact animal models exposed to increased airway pressures, implicating the effects of mechanical stimuli on various cell-signaling pathways (5, 6). Clearly, extensive understanding of ventilator-induced lung injury is needed and the identification of novel therapeutic targets for acute lung injury is essential if there is to be continued progress in this devastating disorder. In this issue of the Journal (pp. 1051–1059), Copland and colleagues (7) report a series of studies designed to address the contribution of mechanical stress to ventilator-associated lung injury. A functional genomic approach was utilized with measurements of lung gene expression to identify genes differentially expressed in a rat model of high tidal volume ventilation (25 ml/kg, 30 minutes) compared with control animals (nonventilated rats). Despite the relatively brief duration of ventilator challenge, as well as the absence of ultrastructural evidence of injury, robust alterations in gene expression employing spotted cDNA microarrays were noted. Thus, one key observation of this work is that gene expression alterations in response to mechanical ventilation, in the absence of additional inflammatory stimuli, are easily detected by microarray techniques and may therefore provide a powerful framework to characterize normal lung responses to mechanotransduction stresses. Consistent with current concepts of the role of the ventilator in acute lung injury, the studies by Copland and colleagues (7) identified upregulation of interleukin-1ß and HSP70 genes as well as Egr-1 and c-Jun transcription factor genes. Cluster analysis confirmed linked expression of these genes (HSP70 clustering separately but closely to the others). Furthermore, temporal analysis demonstrated coincident increases in Egr-1 and c-Jun mRNA expression within 15 minutes of the onset of ventilation with high tidal volumes although significant increases in interleukin-1ß and HSP70 mRNA were not evident until 30 minutes, suggesting a potential cause-and-effect relationship. These genes, particularly interleukin-1ß, are of obvious significance in stretch-induced lung injury (5–7) and acute respiratory distress syndrome (8–10), again consistent with very early signal amplification that begins to evolve a mechanically-stimulated inflammatory phenotype. Importantly, this work utilized multiple approaches to validate the expression profiling data (Northern blot analysis) with immunohistochemical localization of interleukin-1ß to the bronchiolar epithelium (laser capture microdissection) and Western blot analysis to confirm increased interleukin-1ß protein following brief high tidal volume ventilation. This study underscores the fact that genomic data and techniques are increasingly applied to the study of biological problems. Significant genomic challenges, however, remain that include the characterization of identified gene products, understanding the role of these gene products in lung physiology, and delineating their association with human diseases. A more pragmatic challenge is to identify the most appropriate experimental model. Given the expense of the functional genomic approach, it remains unclear as to how many samples and how many chips are necessary and what time points are most informative.
There is little doubt, however, that the most difficult challenge is how best to analyze the unprecedented quantities of data generated by these genomic approaches. In September 2000, the National Heart, Blood, and Lung Institute launched the Programs for Genomic Applications, funding eleven centers to generate novel data and resources for the research community at large to advance functional genomic research related to heart, lung, blood, and sleep disorders. These resources include state-of-the-art software programs for array analysis and normalization, single nucleotide polymorphism analysis, phenotyping of animal models of disease, and a rich array of analytical tools (summarized and updated on http://www.nhlbi.nih.gov/resources/pga). Several programs are working to discover and model the associations between single nucleotide sequence differences in the genes and pathways that underlie interindividual variation in inflammatory responses and their relationship to disease risk, outcome, and treatments in common human disorders such as asthma, chronic obstructive pulmonary disease, cystic fibrosis, and acute lung injury. For example, the HopGene website (http://www.hopkins-genomics.org/) contains extensive array data for rat, murine, and canine models of acute lung injury, a list of candidate genes with preliminary evaluation for relevant single nucleotide polymorphisms and genotyping of these genes. Relevant to the article by Copland and colleagues (7), array data are posted on lungs from rats subjected to mechanical ventilation in vivo (5 hours) and human endothelial cells subjected to cyclic stretch in vitro. In the rat studies, after 5 hours of mechanical ventilation with 12 ml/kg tidal volume, animals were killed and lungs used for histologic examination and RNA for hybridization to an Affymetrix Rat RG-U34A chip ( The genetic determinants, which render patients susceptible to the adverse effects of mechanical ventilation in the setting of acute lung injury, are unknown. The identification of novel therapeutic targets is essential for progress in the understanding of ventilator-associated lung injury. The delineation of genetic susceptibility loci for ventilator-associated lung injury will lead to novel molecular targets and as yet unsuspected pathophysiologic mechanisms pertinent to this injury. Ultimately, the functional genomic approach will accelerate the development of novel therapies for this devastating human disorder. FOOTNOTES Conflict of Interest Statement: J.R.J. has no declared conflict of interest. J.G.N.G. has no declared conflict of interest. REFERENCES
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8,000 genes). Similar to the results of Copland and coworkers (
