Unraveling networks of co-regulated genes on the sole basis of genome sequences

With the growing number of available microbial genome sequences, regulatory signals can now be revealed as conserved motifs in promoters of orthologous genes (phylogenetic footprints). A next challenge is to unravel genome-scale regulatory networks. Using as sole input genome sequences, we predicted cis-regulatory elements for each gene of the yeast Saccharomyces cerevisiae by discovering over-represented motifs in the promoters of their orthologs in 19 Saccharomycetes species. We then linked all genes displaying similar motifs in their promoter regions and inferred a co-regulation network including 56 919 links between 3171 genes. Comparison with annotated regulons highlights the high predictive value of the method: a majority of the top-scoring predictions correspond to already known co-regulations. We also show that this inferred network is as accurate as a co-expression network built from hundreds of transcriptome microarray experiments. Furthermore, we experimentally validated 14 among 16 new functional links between orphan genes and known regulons. This approach can be readily applied to unravel gene regulatory networks from hundreds of microbial genomes for which no other information is available except the sequence. Long-term benefits can easily be perceived when considering the exponential increase of new genome sequences

Role of P2Y2 nucleotide receptor in the physiopathology of inflammatory lung diseases

Amongst respiratory diseases, inflammatory lung diseases constitute a major part of public health problem. As a consequence, investigating the immune mechanisms that contribute to the pathogenesis of these diseases is essential to identify candidate targets for the development of new therapeutic drugs. Furthermore, over the past 20 years, the growing awareness that purinergic signalling events shape the immune and inflammatory responses to infection and allergic reactions warranted the development of animal models to assess their importance in vivo in acute lung injury and chronic airway diseases. The field of purinergic inflammation formulated the unifying concept that ATP is released as a «danger signal» to induce inflammatory responses upon binding purinergic receptors.According to these elements, we began in 2007 to evaluate lung inflammation in mice deficient for the P2Y2 purinergic receptor in TH2 and TH1 models. The most convincing evidence that the P2Y2receptor is engaged during alarm situations comes from studies related to cystic fibrosis and asthma. Indeed, chronic respiratory diseases are commonly associated with elevated airway ATP concentrations, as reported in cystic fibrosis, but also in idiopathic pulmonary fibrosis and chronic obstructive pulmonary disease (COPD) patients, and they are raised by allergens in asthmatic patients.First, we demonstrated a significant role of the P2Y2R in a TH2-ovalbumin(OVA)-induced asthma model. We observed that eosinophil accumulation, a distinctive feature of lung allergic inflammation, was defective in OVA-treated P2Y2-deficient mice compared with OVA-treated wild type animals. Interestingly, the upregulation of VCAM-1 was lower on lung endothelial cells of OVA-treated P2Y2 knockout mice compared with OVA-treated wild type animals. Adhesion assays demonstrated that the action of UTP on leukocyte adhesion through the regulation of endothelial VCAM-1 was abolished in P2Y2-deficient lung endothelial cells. Additionally, the level of soluble VCAM-1, reported as an inducer of eosinophil chemotaxis, was strongly reduced in the bronchoalveolar lavage fluid of P2Y2-deficient mice.Secondly, we studied the consequences of P2Y2R loss in lung inflammation initiated after pneumonia virus of mice (PVM) infection in collaboration with the group of Pr. Daniel Desmecht (ULg). We demonstrated here that P2Y2-/-mice display a severe increase in morbidity and mortality rate in response to PVM. Lower survival of P2Y2-/-mice was not correlated with excessive inflammation despite the higher level of neutrophil recruiters in their broncho-alveolar fluids. Interestingly, we observed lower numbers of dendritic cells, CD4+T cells and CD8+T cells in P2Y2-/-mice compared to P2Y2+/+infected lungs. Lower level of IL-12 and higher level of IL-6 in broncho-alveolar fluid support an inhibition of Th1 response in P2Y2-/-mice. Quantification of DC recruiter expression revealed comparable IP-10 and MIP-3&Doctorat en Sciences biomédicales et pharmaceutiquesinfo:eu-repo/semantics/nonPublishe

P2Y2 receptor regulates VCAM-1 membrane and soluble forms and eosinophil accumulation during lung inflammation.

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Harnessing the Potential of Enzymes as Inhaled Therapeutics in Respiratory Tract Diseases: A Review of the Literature

Respiratory tract diseases (RTDs) are a global cause of mortality and affect patient well-being and quality of life. Specifically, there is a high unmet need concerning respiratory tract infections (RTIs) due to limitations of vaccines and increased antibiotic resistance. Enzyme therapeutics, and in particular plant-based enzymes, represent an underutilised resource in drug development warranting further attention. This literature review aims to summarise the current state of enzyme therapeutics in medical applications, with a focus on their potential to improve outcomes in RTDs, including RTIs. We used a narrative review approach, searching PubMed and clinicaltrials.gov with search terms including: enzyme therapeutics, enzyme therapy, inhaled therapeutics, botanical enzyme therapeutics, plant enzymes, and herbal extracts. Here, we discuss the advantages and challenges of enzyme therapeutics in the setting of RTDs and identify and describe several enzyme therapeutics currently used in the respiratory field. In addition, the review includes recent developments concerning enzyme therapies and plant enzymes in (pre-)clinical stages. The global coronavirus disease 2019 (COVID-19) pandemic has sparked development of several promising new enzyme therapeutics for use in the respiratory setting, and therefore, it is timely to provide a summary of recent developments, particularly as these therapeutics may also prove beneficial in other RTDs

Network Analysis Tools: from biological networks to clusters and pathways (vol 3, pg 1616, 2008)

International audienceno abstrac

P2Y2 Receptor Regulates VCAM-1 Membrane and Soluble Forms and Eosinophil Accumulation during Lung Inflammation.

ATP has been defined as a key mediator of asthma. In this study, we evaluated lung inflammation in mice deficient for the P2Y(2) purinergic receptor. We observed that eosinophil accumulation, a distinctive feature of lung allergic inflammation, was defective in OVA-treated P2Y(2)-deficient mice compared with OVA-treated wild type animals. Interestingly, the upregulation of VCAM-1 was lower on lung endothelial cells of OVA-treated P2Y(2)(-/-) mice compared with OVA-treated wild type animals. Adhesion assays demonstrated that the action of UTP on leukocyte adhesion through the regulation of endothelial VCAM-1 was abolished in P2Y(2)-deficient lung endothelial cells. Additionally, the level of soluble VCAM-1, reported as an inducer of eosinophil chemotaxis, was strongly reduced in the bronchoalveolar lavage fluid (BALF) of P2Y(2)-deficient mice. In contrast, we observed comparable infiltration of macrophages and neutrophils in the BALF of LPS-aerosolized P2Y(2)(+/+) and P2Y(2)(-/-) mice. This difference could be related to the much lower level of ATP in the BALF of LPS-treated mice compared with OVA-treated mice. Our data define P2Y(2) as a regulator of membrane and soluble forms of VCAM-1 and eosinophil accumulation during lung inflammation.JOURNAL ARTICLESCOPUS: ar.jinfo:eu-repo/semantics/publishe

Streptococcus pneumoniae colonization is required to alter the nasal microbiota in cigarette smokeexposed mice

Smokers have nasal microbiota dysbiosis, with an increased frequency of colonizing bacterial pathogens. It is possible that cigarette smoke increases pathogen acquisition by perturbing the microbiota and decreasing colonization resistance. However, it is difficult to disentangle microbiota dysbiosis due to cigarette smoke exposure from microbiota changes caused by increased pathogen acquisition in human smokers. Using an experimental mouse model, we investigated the impact of cigarette smoke on the nasal microbiota in the absence and presence of nasal pneumococcal colonization. We observed that cigarette smoke exposure alone did not alter the nasal microbiota composition. The microbiota composition was also unchanged at 12 h following low-dose nasal pneumococcal inoculation, suggesting that the ability of the microbiota to resist initial nasal pneumococcal acquisition was not impaired in smoke-exposed mice. However, nasal microbiota dysbiosis occurred as a consequence of established high-dose nasal pneumococcal colonization at day 3 in smoke-exposed mice. Similar to clinical reports on human smokers, an enrichment of potentially pathogenic bacterial genera such as Fusobacterium, Gemella, and Neisseria was observed. Our findings suggest that cigarette smoke exposure predisposes to pneumococcal colonization independent of changes to the nasal microbiota and that microbiota dysbiosis observed in smokers may occur as a consequence of established pathogen colonization.</p

Identification of Drug Candidates to Suppress Cigarette Smoke-induced Inflammation via cMap Analyses

Cigarette smoking is the main risk factor for COPD, and to date, existing pharmacologic interventions have been ineffective at controlling inflammatory processes associated with the disease. To address this issue, we used the Connectivity Map (cMap) database to identify drug candidates with the potential to attenuate cigarette smoke-induced inflammation. We queried cMap using three independent, in house cohorts of healthy non-smokers and smokers. Potential drug candidates were validated against four publicly available human datasets, as well as six independent data sets from cigarette smoke-exposed mice. Overall, these analyses yielded two potential drug candidates: kaempferol and bethanechol. Subsequently, the efficacy of each drug was validated in vivo in a model of cigarette smoke-induced inflammation. BALB/c mice were exposed to room air or cigarette smoke and treated with each of the two candidate drugs either prophylactically or therapeutically. We found that kaempferol, but not bethanechol, was able to reduce cigarette smoke-induced neutrophilia both when administered prophylactically and therapeutically. Mechanistically, kaempferol decreased expression of IL-1α and CXCL-5 levels in the lung. Our data suggest that cMap analyses may serve as a useful tool to identify novel drug candidates against cigarette smoke-induced inflammation

Higher mortality rate in PVM-infected P2Y₂-deficient mice.

<p>Following intranasal inoculation of PVM (1000 PFUs), P2Y₂^+/+ and P2Y₂^−/− mice were monitored daily for survival (<b>A</b>) and weight loss (<b>B</b>). Weight curves (mean ± SEM) are relative to initial body weight. The displayed data result from the pooling of four independent experiments.</p

Quantification of neutrophils and macrophages, and their recruiters in the lungs of PVM-infected P2Y₂^+/+ and P2Y₂^−/− mice.

<p><b>A–D.</b> The level of the chemokines KC/CXCL-1 (A), MIP-2/CXCL-2 (B), MIP-1α/CCL3 (C) and MCP-1/CCL2 (D) was determined by ELISA in the BALFs of PVM-infected P2Y₂^+/+ and P2Y₂^−/− mice. (N = 9) <b>E.</b> Flow cytometry quantification of neutrophils and macrophages in the BALFs of PVM-infected P2Y₂^+/+ and P2Y₂^−/− mice at day 8 and 10 post-inoculation (N = 12). <b>F.</b> Cytospin preparations were made from BALFs of P2Y₂^+/+ and P2Y₂^−/− mice at day 8 post-infection using a Shandon III cytocentrifuge and were stained using Diff-Quick staining. Magnification: ×400.</p