Dataset Information

Diagnosis of Acute Lung Rejection by Gene Expression Profiling of Bronchoalveolar Lavage Cells

ABSTRACT: Acute lung rejection is a risk factor for chronic rejection, jeopardizing the long-term survival of lung transplant recipients. At present, acute rejection is diagnosed by transbronchial lung biopsies, which are invasive, expensive, and subject to significant sampling error. In this study, we sought to identify groups of genes whose collective expression in BAL cells best classifies acute rejection versus no-rejection. BAL samples were analyzed from 32 unique subjects whose concurrent histology showed acute rejection (n=14) or no rejection (n=18). Global BAL cell gene expression was measured using Affymetrix U133A microarrays. The nearest shrunken centroid method with 10-fold cross validation was used to define the classification model. 250 runs of the algorithm were performed to determine the range of misclassification error and the most influential genes in determining classifiers. The estimated overall misclassification rate was below 20%. Seven transcripts were present in every classifier and 52 transcripts were present in at least 70% of classifiers; these transcripts were notable for involvement with T-cell function, cytotoxic CD8 activity, and granulocyte degranulation. The proportions of both lymphocytes and neutrophils in BAL samples increased with increasing probability of acute rejection; this trend was more pronounced with neutrophils. We conclude that there is a prominent acute rejection-associated signature in BAL cells characterized by increased T-cell, CD8+ cytotoxic cell, and neutrophil gene expression; this is consistent with established mechanistic concepts of the acute rejection response. Keywords: Disease State Analysis

ORGANISM(S): Mus musculus Homo sapiens

PROVIDER: GSE6095 | GEO | 2008/01/01

SECONDARY ACCESSION(S): PRJNA97655

REPOSITORIES: GEO

ACCESS DATA

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Project description:Acute lung rejection is a risk factor for chronic rejection, jeopardizing the long-term survival of lung transplant recipients. At present, acute rejection is diagnosed by transbronchial lung biopsies, which are invasive, expensive, and subject to significant sampling error. In this study, we sought to identify groups of genes whose collective expression in BAL cells best classifies acute rejection versus no-rejection. BAL samples were analyzed from 32 unique subjects whose concurrent histology showed acute rejection (n=14) or no rejection (n=18). Global BAL cell gene expression was measured using Affymetrix U133A microarrays. The nearest shrunken centroid method with 10-fold cross validation was used to define the classification model. 250 runs of the algorithm were performed to determine the range of misclassification error and the most influential genes in determining classifiers. The estimated overall misclassification rate was below 20%. Seven transcripts were present in every classifier and 52 transcripts were present in at least 70% of classifiers; these transcripts were notable for involvement with T-cell function, cytotoxic CD8 activity, and granulocyte degranulation. The proportions of both lymphocytes and neutrophils in BAL samples increased with increasing probability of acute rejection; this trend was more pronounced with neutrophils. We conclude that there is a prominent acute rejection-associated signature in BAL cells characterized by increased T-cell, CD8+ cytotoxic cell, and neutrophil gene expression; this is consistent with established mechanistic concepts of the acute rejection response. Experiment Overall Design: Lung transplant recipients surviving at least 30 days after transplantation at the University of Minnesota were eligible for enrollment in the study. The study was approved by the Universityâs Institutional Review Board, and all patients provided written informed consent. Subjects underwent scheduled surveillance (n = 22) and clinically indicated (decreased pulmonary function tests or new radiographic abnormalities) bronchoscopies (n = 10), in which 100-200 cc of sterile saline was instilled into a single anatomic location (right middle lobe or lingula), and recovered using gentle suction. 4-6 transbronchial biopsies (TBB) were obtained from the same subsegmental location as the BAL, and 4-6 additional TBB were obtained from the lower lobe of the same lung. After sending samples for clinical tests, approximately 50 ml of the recovered BAL effluent was immediately placed on ice. BAL cell counts and differentials were determined with a hemocytometer. Specimens from subjects with active bronchopulmonary infections (as determined by history, exam, chest radiograph, and the results of routine laboratory tests and cultures) were excluded from analysis. Experiment Overall Design: All biopsy specimens were graded according to standard International Society for Heart and Lung Transplantation criteria8. Vascular and airway cellular infiltrate were scored separately on A (vascular) and B (airway) scales, each score ranging from 0 to 48;23;24. For the purposes of this analysis, acute rejection was defined as a combined A+B score greater than or equal to 2; and no-rejection was defined as a combined A+B score less than 2. Experiment Overall Design: The BAL samples used in this study were selected according to strict criteria that were designed to maximize potential differences in gene expression between acute rejection and non-acute rejection BAL samples; and to minimize bias from confounding factors. Each subject (n=32) was represented by one BAL sample in this study. Control subjects and samples (n=14) were selected according to the following criteria: 1) A+B score was 0 or 1 for all biopsies collected from each subject during his/her post-transplant course; 2) at least three biopsies had been graded from each subject; 3) the subject had survived at least one year post-transplant; 4) the selected BAL sample was culture negative for pathogenic bacteria, fungi, and viruses; and 5) the selected sample was from a patient that had not developed BOS or was collected at least 6 months prior to the development of BOS. Rejection subjects and samples (n=18) were selected according to the following criteria: 1) the BAL sample was culture negative for pathogenic bacteria, fungi, and viruses; 2) for subjects with more than one acute rejection sample, we used the first acute rejection sample (A+B score > 1); 3) BOS grade was 0 at the time of BAL sampling (although some subjects subsequently developed BOS).

Project description:Acute lung injury (ALI), a major cause of acute respiratory failure with high morbidity and mortality, isare characterized by significant pulmonary inflammation and both alveolar and vascular barriers dysfunction. In Pprior studies have highlighted the role of nonmuscle myosin light chain kinase (nmMLCK) as an essential element of inflammatory response with MYLK polymorphisms associated withwhich alters ALI susceptibility. In the present study we sought to further define nmMLCK in acute inflammatory syndromes and examined We examined nmMLCK as a molecular target involved in increase of lung epithelial and endothelial barrier permeability. We utilized in two muirine models of inflammatory lung injury: intratracheal administration of endotoxin/lipopolysaccharide (LPS, 2.5 mg/kg) and VILI (ventilator-induced lung injury, tidal volume 40ml/kg). Two complementary strategies were used to reduce nmMLCK activity or expression. We found that membrane permeant oligopeptide, PIK, inhibited MLC kinase activity in vitro in aand displayed dose-dependent mannerinhibition of MLC kinase activity.. Intravenous delivery of PIK significantly attenuated LPS-induced lung inflammation reflected by decreasing accumulation of bronchoalveolar lavage (BAL) albumin (~ 50% reduction) as well as reduction in BAL cells, tissue MPO activity and tissue albumin in lung homogenates. A second regulatory approach explored targeting murine nmMLCK by administration of siRNA (5mg/kg) 3 days prior to LPS challenge. siRNA decreased of nmMLCK expression in lungs (~ 70% reduction) and resulted in significant attenuation LPS-induced lung inflammation (~ 40% reduction) as reflected by decreased BAL protein level and BAL cells. For targeting pulmonary vessels nmMLCK we used ACE antibody-conjugated liposomes with nmMLCK siRNA in murine ventilator-induced lung injury (VILI) model. Protein silencing of nmMLCK was evident by immunohistochemical analysis with a decrease in relative intensity of fluorescence in lung vessels compared with control animals. Furthermore, the inhibition of nmMLCK expression by siRNA in vessels significantly attenuated VILI lung injury as reflected by decreased BAL protein level (40% reduction). Finally, MLCK knockout mice were significantly protected (reduced BAL protein and albumin) when exposed to a model of severe VILI (4h, 40ml/kg tidal volume). Conclusion: the MLCK gene KO and chemical biology results indicate that the targeting of nmMLCK in vivo attenuate the severity of LPS-induced or VILI acute lung injury. We used microarrays to detail the global programme of gene expression induced by VILI in Wild type and nmMLCK-/- mouse. Experiment Overall Design: four group (n=3) of animals were treated by SB (Spontaneouse breathing) or VILI (4 hours, 30 ml/kg tidal volume) in Wild type or nmMLCK-/- animals;

Dataset Information

Diagnosis of Acute Lung Rejection by Gene Expression Profiling of Bronchoalveolar Lavage Cells

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets