Heterologous matrix metalloproteinase gene promoter activity allows In Vivo real-time imaging of Bleomycin-induced Lung fibrosis in transiently transgenized mice

Idiopathic pulmonary fibrosis is a very common interstitial lung disease derived from chronic inflammatory insults, characterized by massive scar tissue deposition that causes the progressive loss of lung function and subsequent death for respiratory failure.Bleomycin is used as the standard agent to induce experimental pulmonary fibrosis in animal models for the study of its pathogenesis. However, to visualize the establishment of lung fibrosis after treatment, the animal sacrifice is necessary. Thus, the aim of this study was to avoid this limitation by using an innovative approach based on a double bleomycin treatment protocol, along with the in vivo images analysis of bleomycintreated mice. A reporter gene construct, containing the luciferase open reading frame under the matrix metalloproteinase-1 promoter control region, was tested on doublebleomycin-treated mice to investigate, in real time, the correlation between bleomycin treatment, inflammation, tissue remodeling and fibrosis. Bioluminescence emitted by the lungs of bleomycin-treated mice, corroborated by fluorescent molecular tomography, successfully allowed real time monitoring of fibrosis establishment. The reporter gene technology experienced in this work could represent an advanced functional approach for real time non-invasive assessment of disease evolution during therapy, in a reliable and translational living animal model.Fil: Stellari, Fabio Franco. Chiese Farmaceutici; ItaliaFil: Ruscitti, Francesca. Chiese Farmaceutici; ItaliaFil: Pompilio, Daniela. Chiese Farmaceutici; ItaliaFil: Ravanetti, Francesca. Università di Parma. Dipartimento di Scienze Medico Veterinarie; ItaliaFil: Tebaldi, Giulia. Università di Parma. Dipartimento di Scienze Medico Veterinarie; ItaliaFil: Macchi, Francesca. Università di Parma. Dipartimento di Scienze Medico Veterinarie; ItaliaFil: Verna, Andrea Elizabeth. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Chiese Farmaceutici; ItaliaFil: Villetti, Gino. Chiese Farmaceutici; ItaliaFil: Donofrio, Gaetano. Università di Parma. Dipartimento di Scienze Medico Veterinarie ; Itali

Mroh1, a lysosomal regulator localized by WASH-generated actin

The steps leading to constitutive exocytosis are poorly understood. In Dictyostelium WASH complex mutants, exocytosis is blocked, so cells that take up fluorescent dextran from the medium retain it and remain fluorescent. Here, we establish a FACS-based method to select cells that retain fluorescent dextran, allowing identification of mutants with disrupted exocytosis. Screening a pool of random mutants identified members of the WASH complex, as expected, and multiple mutants in the conserved HEAT-repeat-containing protein Mroh1. In mroh1 mutants, endosomes develop normally until the stage where lysosomes neutralize to postlysosomes, but thereafter the WASH complex is recycled inefficiently, and subsequent exocytosis is substantially delayed. Mroh1 protein localizes to lysosomes in mammalian and Dictyostelium cells. In Dictyostelium, it accumulates on lysosomes as they mature and is removed, together with the WASH complex, shortly before the postlysosomes are exocytosed. WASH-generated F-actin is required for correct subcellular localization; in WASH complex mutants, and immediately after latrunculin treatment, Mroh1 relocalizes from the cytoplasm to small vesicles. Thus, Mroh1 is involved in a late and hitherto undefined actin-dependent step in exocytosis

In Vivo Imaging of Transiently Transgenized Mice with a Bovine Interleukin 8 (CXCL8) Promoter/Luciferase Reporter Construct

One of the most remarkable properties of interleukin 8 (CXCL8/IL-8), a chemokine with known additional functions also in angiogenesis and tissue remodeling, is the variation of its expression levels. In healthy tissues, IL-8 is barely detectable, but it is rapidly induced by several folds in response to proinflammatory cytokines, bacterial or viral products, and cellular stress. Although mouse cells do not bear a clear homologous IL-8 gene, the murine transcriptional apparatus may well be capable of activating or repressing a heterologous IL-8 gene promoter driving a reporter gene. In order to induce a transient transgenic expression, mice were systemically injected with a bovine IL-8 promoter–luciferase construct. Subsequently mice were monitored for luciferase expression in the lung by in vivo bioluminescent image analysis over an extended period of time (up to 60 days). We demonstrate that the bovine IL-8 promoter–luciferase construct is transiently and robustly activated 3–5 hours after LPS and TNF-α instillation into the lung, peaking at 35 days after construct delivery. Bovine IL-8 promoter–luciferase activation correlates with white blood cell and neutrophil infiltration into the lung. This study demonstrates that a small experimental rodent model can be utilized for non-invasively monitoring, through a reporter gene system, the activation of an IL-8 promoter region derived from a larger size animal (bovine). This proof of principle study has the potential to be utilized also for studying primate IL-8 promoter regions

Driving green: Financial benefits of carbon emission reduction in companies

This paper explores the relationship between carbon emissions reduction and corporate financial performance, leveraging a rich dataset of 14,866 observations from 2768 companies across 36 countries and regions, and 35 industries over the period 2002–2022. We find that carbon emissions reductions improve company financial performance, as measured by return on assets and return on equity, with this effect being even more pronounced for companies with higher carbon intensity. Additionally, country carbon regulations are positively associated with a company's financial performance, while higher ESG scores negatively impact it. Notably, we find no significant role for CSR reporting in driving financial gains. Overall, our findings suggest that companies can enhance financial performance by intensifying their carbon emission reduction efforts while also contributing to environmental stewardship. We recommend that businesses adopt tailored sustainability strategies that account for country, firm, and industry-specific factors to maximize these benefits across different regional and sectoral contexts.publishedVersio

Comprehensive analysis of aging-related genes and immune infiltration landscape in ischemic cardiomyopathy

BackgroundIschemic cardiomyopathy (ICM), which arises from obstructive coronary artery diseases, is a leading cause of heart failure. Aging is a major risk factor for cardiovascular diseases, yet its connection to ICM remains unclear. This study aimed to investigate the role of aging-related gene expression in the context of ICM.MethodsHuman microarray data (GSE116250) were retrieved from the GEO database and aging-related differentially expressed genes (ARDEGs) were identified using the Aging Atlas database. Functional enrichment and protein-protein interaction (PPI) network analyses were performed to elucidate the functions and interactions of these ARDEGs, leading to the identification of hub genes. The immune infiltration landscape in ICM was further characterized. Subsequently, integrated regulatory networks involving the hub genes were constructed through microRNA-mRNA-transcription factor interaction and GeneMANIA analyses, while their biological functions were inferred via gene set enrichment analysis (GSEA). The predictive value of the hub genes was validated using receiver operating characteristic (ROC) analysis based on both the identification dataset (GSE116250) and an independent validation cohort (GSE1145) Finally,the expression patterns of these genes were verified by RT-qPCR on mouse disease model.ResultsA total of 50 ARDEGs (42 upregulated and 8 downregulated) were identified,which are primarily involved in the response to inflammation. Ten types of immune cells showed significant alterations in the ICM heart tissues. Among these cells, CD56dim NK cells exhibited extensive and significant correlations with other immune cells. JUN was identified as a key transcription factor regulating the top five hub ARDEGs: TNF, PTGS2, IL6, IL1B, and CXCL8. ROC analysis demonstrated that TNF, CXCL8, and IL6 serve as potential biomarkers for ICM, and the combination of the three markers further improved the predictive value. RT-qPCR analysis subsequently confirmed the upregulation of these hub inflammatory ARDEGs in mouse heart tissues.ConclusionAging-related genes play a significant role in ICM and targeting these genes may pave the way for ICM diagnostic and therapeutic strategies

PSEUDOMONAS AERUGINOSA INTERACTIONS WITH HOST AND BACTERIAL NEIGHBORS IN CYSTIC FIBROSIS LUNG DISEASE

Cystic fibrosis (CF) lung infection is a complex condition where opportunistic pathogens and defective immune system cooperate in developing a constant cycle of infection and inflammation. Despite the polymicrobial nature of this pathology, Pseudomonas aeruginosa is considered the major pathogen. Several mechanisms contribute to its successful colonization of CF airways; among these, virulence factors are important players. This thesis includes two studies aimed to evaluate aspects of P. aeruginosa interaction with both host and other microorganisms, like the emerging pathogen Achromobacter xylosoxidans