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Gene expression of diesel exhaust inhalation in peripheral blood mononuclear cells from healthy human volunteers

ABSTRACT: Background: Diesel exhaust (DE) is the primary source of urban fine particulate matter, which has been associated with cardiovascular disease in epidemiological studies. These effects may be related to oxidative stress and systemic inflammation with resulting perturbation of vascular homeostasis. Peripheral leukocytes are involved in both inflammation and control of vascular homeostasis. Objectives: We conducted an exploratory study using microarray techniques to analyze whether global gene expression in peripheral blood mononuclear cells (PBMCs) can inform on potential mechanisms of effect of DE inhalation. Methods: In a double-blind, crossover, controlled exposure study, healthy adult volunteers were exposed in randomized order to filtered air (FA) and diluted DE in two-hour sessions. We isolated RNA (Trizol/Qiagen method) form PBMCs before, and two times after each exposure. RNA samples were arrayed using the Affymetrix® platform (GeneChip® Human Genome U133 Plus 2.0 Array). Results: Microarray analyses were conducted on five subjects with available RNA sample form exposures to FA and to the highest DE inhalation (200 µg/m³ of fine particulate matter). Following data normalization and statistical analysis, a total of 1290 out of 54,675 probe sets with significant evidence for differential expression (more than 1.5-fold up or down regulated with p < 0.05) were identified. These include genes involved in inflammatory response (e.g., IL8RA, TNFAIP6, FOS), oxidative stress (e.g., HMOX1, BAX, PRDX1,), and in biochemical pathways like mitogen-activated protein kinases (MAPK) and tight junction pathways. Conclusions: These data suggest that DE may exert time-dependent changes in gene expression in PBMCs in healthy individuals. Genes that may be affected by DE inhalation are involved in inflammatory and oxidative stress processes. Keywords: time course

ORGANISM(S): Homo sapiens

PROVIDER: GSE7462 | GEO | 2008/01/03

SECONDARY ACCESSION(S): PRJNA100207

REPOSITORIES: GEO

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Project description:Background: Diesel exhaust (DE) is the primary source of urban fine particulate matter, which has been associated with cardiovascular disease in epidemiological studies. These effects may be related to oxidative stress and systemic inflammation with resulting perturbation of vascular homeostasis. Peripheral leukocytes are involved in both inflammation and control of vascular homeostasis. Objectives: We conducted an exploratory study using microarray techniques to analyze whether global gene expression in peripheral blood mononuclear cells (PBMCs) can inform on potential mechanisms of effect of DE inhalation. Methods: In a double-blind, crossover, controlled exposure study, healthy adult volunteers were exposed in randomized order to filtered air (FA) and diluted DE in two-hour sessions. We isolated RNA (Trizol/Qiagen method) form PBMCs before, and two times after each exposure. RNA samples were arrayed using the AffymetrixÂ® platform (GeneChipÂ® Human Genome U133 Plus 2.0 Array). Results: Microarray analyses were conducted on five subjects with available RNA sample form exposures to FA and to the highest DE inhalation (200 Âµg/mÂ³ of fine particulate matter). Following data normalization and statistical analysis, a total of 1290 out of 54,675 probe sets with significant evidence for differential expression (more than 1.5-fold up or down regulated with p < 0.05) were identified. These include genes involved in inflammatory response (e.g., IL8RA, TNFAIP6, FOS), oxidative stress (e.g., HMOX1, BAX, PRDX1,), and in biochemical pathways like mitogen-activated protein kinases (MAPK) and tight junction pathways. Conclusions: These data suggest that DE may exert time-dependent changes in gene expression in PBMCs in healthy individuals. Genes that may be affected by DE inhalation are involved in inflammatory and oxidative stress processes. Experiment Overall Design: We conducted a crossover, double-blind experiment, randomized to order of DE and filtered-air (FA) exposure with each participant exposed on four different days to each of four conditions: FA and DE calibrated to 50Âµg/m3 (DE50), 100Âµg/m3 (DE100), and 200Âµg/m3 (DE200) of fine particulate matter (PM2.5- particles with aerodynamic diameter 2.5Âµm or less). Exposure sessions were conducted at least 2 weeks apart.

Project description:Background: Epidemiologic associations between acutely increased cardiorespiratory morbidity and mortality and particulate air pollution are well-established, but the effects of acute pollution exposure on human gene expression changes are not well understood. Objectives: In order to identify potential mechanisms underlying epidemiologic associations between air pollution and morbidity, we explored changes in gene expression in humans following inhalation of fresh diesel exhaust (DE), a model for particulate air pollution. Methods: 14 ethnically homogeneous (white males), young, healthy subjects underwent sixty minute inhalation exposures on 2 separate days with clean air (CA) or freshly generated and diluted DE at a concentration of 300 Î¼g/m3 PM2.5. Prior to and 24 hours following each session, whole blood was sampled and fractionated for PBMC isolation, RNA extraction, and generation of cDNA, followed by hybridizationÂ withÂ Agilent Whole Human Genome (4X44K) arrays. Results: Oxidative stress and the ubiquitin proteasome pathway, as well as the coagulation system, were among hypothesized pathways identified by analysis of differentially expressed genes. Nine genes from these pathways were validated using real-time PCR comparing fold change in expression between DE exposed and clean (CA) days. Quantitative gene fold changes generated by real-time PCR were consistent with the directional fold changes from the microarray analysis. Conclusions: Changes in gene expression connected with key oxidative stress, protein degradation, and coagulation pathways are likely to underlie observed physiologic and clinical outcomes and suggest specific avenues and sensitive time points for further physiologic exploration. Diesel exhaust inhalation exposure-induced human gene expression changes were measured in peripheral blood mononuclear cells. 14 white males subjects were seated in a controlled environment facility for 2 separate 60 minute inhalation exposures which occurred at least 1 week apart to either clean air and freshly generate or diluted diesel exhaust (300ug/m3 PM2.5) .Blood collection occurred immediately prior to and 24 hours following exposures. Two microarrays per subject were completed using Agilent Whole Human Genome (4X44K) arrays. Genes that were of interest to our research group were verified using Real Time PCR analysis.

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Gene expression of diesel exhaust inhalation in peripheral blood mononuclear cells from healthy human volunteers

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