Project description:The tips of secondary alveolar septae in day 6 neonatal mouse lung tissue were isolated using laser capture microscopy. RNA was isolated from pooled secondary alveolar tips and also from pooled neonatal day 6 whole lung tissue. The isolated RNAs were then amplified in parallel. Gene array profiling of the two RNA samples was performed. Gene expression in the secondary alveolar septal tips was compared to gene expression in the whole lung tissue. In this way, the genes that are expressed in the tip of secondary alveolar septae were identified as well as the genes that are enriched in the alveolar septal tips versus in whole lung tissue. Keywords: Comparison of gene profiles in the tips of secondary alveolar septae versus in whole lung tissue of neonatal mice
Project description:The tips of secondary alveolar septae in day 6 neonatal mouse lung tissue were isolated using laser capture microscopy. RNA was isolated from pooled secondary alveolar tips and also from pooled neonatal day 6 whole lung tissue. The isolated RNAs were then amplified in parallel. Gene array profiling of the two RNA samples was performed. Gene expression in the secondary alveolar septal tips was compared to gene expression in the whole lung tissue. In this way, the genes that are expressed in the tip of secondary alveolar septae were identified as well as the genes that are enriched in the alveolar septal tips versus in whole lung tissue. Experiment Overall Design: We performed an experiment in which we used laser capture microscopy to collect 10,000 secondary alveolar tips from frozen sections of lung tissue obtained from many different day 6 neonatal mice obtained from different litters. In the same experiment, we also isolated RNA from pooled day 6 neonate whole lung tissue. The isolated RNAs were amplified in parallel and then hybridized to microarrays in parallel in order to profile and compare gene expression in the two samples. The entire experiment was performed twice with tissue from different litters. In addition, the amplified RNAs were hybridized to two different microarrays, at different times.
Project description:The importance of unanchored Ub in innate immunity has been shown only for a limited number of unanchored Ub-interactors. We investigated what additional cellular factors interact with unanchored Ub and whether unanchored Ub plays a broader role in innate immunity. To identify unanchored Ub-interacting factors from murine lungs, we used His-tagged recombinant poly-Ub chains as bait. These chains were mixed with lung tissue lysates and protein complexes were isolated with Ni-NTA beads. Sample elutions were subjected to mass spectrometry (LC-MSMS) analysis.
Project description:To describe the protein profile in hippocampus, colon and ileum tissue’ changing after the old faeces transplants, we adopted a quantitative label free proteomics approach.
Project description:SILAC based protein correlation profiling using size exclusion of protein complexes derived from Mus musculus tissues (Heart, Liver, Lung, Kidney, Skeletal Muscle, Thymus)
Project description:SILAC based protein correlation profiling using size exclusion of protein complexes derived from seven Mus musculus tissues (Heart, Brain, Liver, Lung, Kidney, Skeletal Muscle, Thymus)
Project description:Alveologenesis, the final stage in lung development, substantially remodels the distal lung, expanding the alveolar surface area for efficient gas exchange. Secondary crest myofibroblasts (SCMF) exist transiently in the neonatal distal lung and are critical for alveologenesis. However, the pathways that regulate SCMF function, proliferation, and temporal identity remain poorly understood. To address this, we purified SCMFs from reporter mice, performed bulk RNA-sequencing, and found dynamic changes in Hippo-signaling components during alveologenesis. We deleted Hippo effectors, Yap/Taz, from Acta2-expressing SCMFs at the onset of alveologenesis, causing a significant arrest in alveolar development. Using scRNA-seq, we identified a distinct cluster of cells in mutant lungs with altered expression of marker genes associated with proximal mesenchymal cell types, airway smooth muscle (ASM), and alveolar duct myofibroblasts (DMF). Using lineage tracing, we show that neonatal Acta2-expressing SCMFs give rise to adult DMFs and that Yap/Taz mutants have an increase of persisting DMF-like cells in the alveolar ducts. Our findings identify aberrant differentiation of neonatal lung myofibroblasts and demonstrate that Yap/Taz are critical for maintaining lineage commitment.
Project description:Alveologenesis, the final stage in lung development, substantially remodels the distal lung, expanding the alveolar surface area for efficient gas exchange. Secondary crest myofibroblasts (SCMF) exist transiently in the neonatal distal lung and are critical for alveologenesis. However, the pathways that regulate SCMF function, proliferation, and temporal identity remain poorly understood. To address this, we purified SCMFs from reporter mice, performed bulk RNA-sequencing, and found dynamic changes in Hippo-signaling components during alveologenesis. We deleted Hippo effectors, Yap/Taz, from Acta2-expressing SCMFs at the onset of alveologenesis, causing a significant arrest in alveolar development. Using scRNA-seq, we identified a distinct cluster of cells in mutant lungs with altered expression of marker genes associated with proximal mesenchymal cell types, airway smooth muscle (ASM), and alveolar duct myofibroblasts (DMF). Using lineage tracing, we show that neonatal Acta2-expressing SCMFs give rise to adult DMFs and that Yap/Taz mutants have an increase of persisting DMF-like cells in the alveolar ducts. Our findings identify aberrant differentiation of neonatal lung myofibroblasts and demonstrate that Yap/Taz are critical for maintaining lineage commitment.
Project description:PURPOSE: To provide a detailed gene expression profile of the normal postnatal mouse cornea. METHODS: Serial analysis of gene expression (SAGE) was performed on postnatal day (PN)9 and adult mouse (6 week) total corneas. The expression of selected genes was analyzed by in situ hybridization. RESULTS: A total of 64,272 PN9 and 62,206 adult tags were sequenced. Mouse corneal transcriptomes are composed of at least 19,544 and 18,509 unique mRNAs, respectively. One third of the unique tags were expressed at both stages, whereas a third was identified exclusively in PN9 or adult corneas. Three hundred thirty-four PN9 and 339 adult tags were enriched more than fivefold over other published nonocular libraries. Abundant transcripts were associated with metabolic functions, redox activities, and barrier integrity. Three members of the Ly-6/uPAR family whose functions are unknown in the cornea constitute more than 1% of the total mRNA. Aquaporin 5, epithelial membrane protein and glutathione-S-transferase (GST) omega-1, and GST alpha-4 mRNAs were preferentially expressed in distinct corneal epithelial layers, providing new markers for stratification. More than 200 tags were differentially expressed, of which 25 mediate transcription. CONCLUSIONS: In addition to providing a detailed profile of expressed genes in the PN9 and mature mouse cornea, the present SAGE data demonstrate dynamic changes in gene expression after eye opening and provide new probes for exploring corneal epithelial cell stratification, development, and function and for exploring the intricate relationship between programmed and environmentally induced gene expression in the cornea. Keywords: other