Unknown,Transcriptomics,Genomics,Proteomics

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Regulatory T cell-mediated resolution of lung injury: Identification of potential target genes via expression profiling

ABSTRACT: We examined early and late gene expression changes using the IT LPS model of Acute Lung Injury (ALI). In this model, injury peaks at day 4 and is almost completely resolved by day 10 in wild type (WT) C57BL/6 mice. In contrast to the pattern in WT mice, lymphocyte-deficient Rag-1 -/- mice exhibit strikingly delayed resolution despite similar initial injury. Total RNA was isolated from mouse lung at time 0, 1, 4, and 10 days following LPS treatment of wild type and Rag-1 null mice using TRIzol reagent (Invitrogen Life Technologies), then purified using RNAeasy columns (Qiagen) with DNase I treatment according to the manufacturerâ??s recommendations. The quality of RNA was assessed using an Agilent 2100 Bioanalyzer (Agilent Technologies).

ORGANISM(S): Mus musculus

SUBMITTER: Neil Aggarwal

PROVIDER: E-GEOD-17355 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Regulatory T cell-mediated resolution of lung injury: identification of potential target genes via expression profiling.

Aggarwal Neil R NR D'Alessio Franco R FR Tsushima Kenji K Sidhaye Venkataramana K VK Cheadle Christopher C Grigoryev Dmitry N DN Barnes Kathleen C KC King Landon S LS

Physiological genomics 20091222 2

In animal models of acute lung injury (ALI), gene expression studies have focused on the acute phase of illness, with little emphasis on resolution. In this study, the acute phase of intratracheal lipopolysaccharide (IT LPS)-induced lung injury was similar in wild-type (WT) and recombinase-activating gene-1-deficient (Rag-1(-/-)) lymphocyte-deficient mice, but resolution was impaired and resolution-phase lung gene expression remained different from baseline only in Rag-1(-/-) mice. By focusing o ...[more]

PMID: 20028937

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Project description:The common research strain S. aureus NCTC 8325-4 was subjected to serial passage in increasing concentrations of ramoplanin. The resulting strain, RSPA16, had reduced susceptibility to not only ramoplanin but vancomycin and nisin as well. Electron microscopy revealed that the cell wall of RSPA16 was double the thickness of the susceptible progenitor strain, a phenotype commonly observed in vancomycin intermediate resistant S. aureus (VISA) strains. RSPA16 was also less susceptible to lysis induced by Triton X-100 than its progenitor strain. Transcriptional profiling experiments were performed with NCTC 8325-4 and RSPA16 without antibiotic exposure and exposed to ramoplanin. Increased expression of genes associated with cell wall stress was observed when either strain was treated with ramoplanin. We also observed that treatment with ramoplanin altered of the expression levels of numerous genes encoding proteins involved with amino acid biosynthesis, central metabolic pathways, nucleotide biosynthesis, iron acquisition, ABC transporters and regulation of transcription. Comparison of the transcriptional profiles of RSPA16 and NCTC 8325-4 not exposed to ramoplanin revealed alterations in the expression of levels of several genes involved with biosynthesis of teichoic acids, biosynthesis of peptidoglycan, central metabolism, DNA replication, nucleotide biosynthesis, iron acquisition, ABC transporters, and regulation of transcription. These transcriptional profiles provide insights into the possible sources of the reduced susceptibility of RSPA16 to peptide antibiotics. Mid log phase (OD620 = 0.4) batch TSB cultures of ramoplanin susceptible (NCTC 8325-4) and resistant (RSPA16) strains of S. aureus were exposed to ramoplanin for 30 minutes at 37 C. Untreated controls were performed for each strain as well. The transcriptional response was determined using Affymetrix Gene Chip Arrays. This study also examines the transcriptional alterations which confer reduced susceptibility to ramoplanin as the transcriptional profiles of NCTC 8325-4 and RSPA16 each untreated were compared.

Project description:Acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) is an inflammatory process of the lungs characterized by increased permeability of the alveolar-capillary membrane with subsequent interstitial/alveolar edema and diffuse alveolar damage. ALI/ARDS can be the results of either direct or indirect lung injury, with pneumonia being the most common direct pulmonary insult and sepsis the most common extra-pulmonary cause. In this study, we employed the murine lipopolysaccharide (LPS)-induced direct and indirect lung injury model to explore the pathogenic mechanisms of pulmonary and extra-pulmonary ARDS, using an unbiased, discovery and quantitative proteomic approach. A total of 1,017 proteins were both identified and quantified in bronchoalveolar lavage fluid (BALF) from control, intratracheal LPS (I.T. LPS, 0.1 mg/kg) and intraperitoneal LPS (I.P. LPS, 5 mg/kg) treated mice. The two LPS groups shared 13 up-regulated and 22 down-regulated proteins compared to the control group. Among them, molecules related to bronchial and type II alveolar epithelial cell functions including cell adhesion molecule 1 and surfactant protein B were reduced, whereas lactotransferrin and resistin like alpha involved in lung innate immunity were upregulated in both LPS groups. Proteomic profiling also identified significant differences in BALF proteins between I.T. and I.P. LPS groups. Ingenuity pathway analysis revealed that acute-phase response signaling was activated by both I.T. and I.P. LPS, however, the magnitude of activation is much greater in I.T. LPS group compared to I.P. LPS group. Intriguingly, two canonical signaling pathways, liver X receptor/retinoid X receptor activation and the production of nitric oxide and reactive oxygen species in macrophages, were activated by I.T. LPS but suppressed by I.P. LPS. In addition, CXCL15 (also known as lungkine) was also up-regulated by I.T LPS but down-regulated by I.P. LPS. In conclusion, our quantitative discovery-based proteomic approach identified commonalities as well as significant differences in BALF protein expression profiles in LPS-induced direct and indirect lung injury, and importantly, LPS-induced indirect lung injury results in suppression of select components of lung innate immunity, which could contribute to the so-called “immunoparalysis” in sepsis patients.

Dataset Information

Regulatory T cell-mediated resolution of lung injury: Identification of potential target genes via expression profiling

Publications

Regulatory T cell-mediated resolution of lung injury: identification of potential target genes via expression profiling.

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