Dataset Information

Lung proteome of the AAV-DTR/DT mouse model of acute epithelial lung injury

ABSTRACT: Injury of the lung epithelium is one of the initial events driving acute lung diseases such as acute lung injury and acute respiratory distress syndrome. Repeated epithelial injury followed by aberrant repair leads to fibroblast activation and extracellular matrix deposition and is therefore considered as the major cause of chronic pulmonary diseases, such as idiopathic pulmonary fibrosis. In vitro models as well as in vivo animal models are necessary to investigate early disease mechanisms and altered signalling pathways of epithelial cells. One way to target specific cells is the diphtheria toxin receptor/diphtheria toxin (DTR/DT) system. The human DTR (hDTR) is sensitive to DT, thereby inducing cell death by blocking protein synthesis, while the murine DTR is at least 10^5 times more resistant to DT. Consequently, expression of the hDTR and application of DT lead to targeted cell depletion. Therefore, a novel and flexible DTR/DT model of acute epithelial lung injury was established and described recently, which is driven by adeno-associated virus (AAV) variant 6.2 mediated hDTR expression. The AAV6.2 vector transduces specifically into bronchial epithelial and alveolar epithelial type II cells leading to their depletion after DT administration. In this study we analysed the bulk lung proteome of the recently established AAV-DTR/DT mouse model of acute epithelial lung injury, providing a detailed insight into proteomic changes upon injury. Frozen pulverized lung tissue samples (~10 mg per sample) from AAV-stuffer control (1 E11 viral genome, n=7) and AAV-hDTR (0.3 E11 viral genome, n=7) treated mice 24 h after intratracheal application of 100 ng DT were analysed in a TMTpro-based workflow. TMTpro-labelled peptides were fractionated into 24 fractions using off-line high pH reversed phase chromatography. Fractions were analysed on an Orbitrap Eclipse Tribrid mass spectrometer in combination with the FAIMS Pro Interface (Thermo Scientific) using a SPS-MS3 based method including real-time search.

INSTRUMENT(S): Orbitrap Eclipse

ORGANISM(S): Mus Musculus (ncbitaxon:10090)

SUBMITTER: Wolfgang Rist

PROVIDER: MSV000088510 | MassIVE | Sat Dec 04 09:14:00 GMT 2021

SECONDARY ACCESSION(S): PXD030177

REPOSITORIES: MassIVE

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Publications

Whole lung proteome of an acute epithelial injury mouse model in comparison to spatially resolved proteomes.

Griesser Eva E Gesell Martin M Veyel Daniel D Lamla Thorsten T Geillinger-Kästle Kerstin K Rist Wolfgang W

Proteomics 20230126 10

Epithelial injury is one of the major drivers of acute pulmonary diseases. Recurring injury followed by aberrant repair is considered as the primary cause of chronic lung diseases, such as idiopathic pulmonary fibrosis (IPF). Preclinical in vivo models allow studying early disease-driving mechanisms like the recently established adeno-associated virus-diphtheria toxin receptor (AAV-DTR) mouse model of acute epithelial lung injury, which utilises AAV mediated expression of the human DTR. We perfo ...[more]

PMID: 36641648

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Project description:Acute lung injury (ALI) is characterized by acute respiratory failure in the setting of non-cardiogenic pulmonary edema, causing acute respiratory distress syndrome (ARDS) in patients and contributes significantly to mortality of critically illness. The main goal of our study is to elucidate the role of miRNAs in neutrophil-epithelial communication during pulmonary inflammation and thereby identifying novel targets for therapy of acute lung injury (ALI). In our studies we identified a miR-223-dependent neutrophil-epithelial crosstalk during ALI. Activated neutrophils (PMN) and pulmonary epithelial cells come into a close spatial relationship during ALI. And, since previous studies had indicated the possibility that inflammatory cell-dependent release of miRNA-containing microvesicles could function as a means of exchanging genetic information from a donor to a target cell, we assessed PMN-elicited alterations of pulmonary epithelial miRNA expression in an experimental co-culture setup. This approach provided a selective and extremely robust readout: While other miRNAs were not or only moderately altered in their expression, pulmonary-epithelial-expressed miR-223 was significantly induced after 4 or 6h of co-incubation. Additional in vitro and in vivo studies clearly demonstrate that this increase of epithelial miR-223 is not due to miR-223 transcriptional induction, but instead PMN-dependent and caused by shuttling miR-223 from PMN into pulmonary epithelial cells. To address the functional role of miR-223-dependent neutrophil-epithelial crosstalk during ALI, we exposed mice to ventilator-induced ALI and observed robust induction of pulmonary miR-223 during ALI, while increases of miR-223 were completely abolished after antibody-depletion of PMN. Moreover, studies of alveolar epithelial cells isolated from mice with ALI showed robust increases of miR-223, indicating that miR-223 is shuttled from PMN towards alveolar epithelia during ALI in vivo. Functional studies revealed that gene-targeted mice for miR-223 experience a more severe phenotype during ALI as compared to controls, while their phenotype could be resuscitated by nanoparticle-mediated overexpression of miR-223 in the lungs. In summary, these studies reveal a novel role of miR-223-dependent neutrophil-epithelial crosstalk representing an anti-inflammatory pathway that can be targeted for ALI treatment.

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Dataset Information

Lung proteome of the AAV-DTR/DT mouse model of acute epithelial lung injury

Publications

Whole lung proteome of an acute epithelial injury mouse model in comparison to spatially resolved proteomes.

Similar Datasets

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