hyperoxia chamber  (BioSpherix)

Journal: bioRxiv

Article Title: Functional interrogation uncovers a critical role for a high-plasticity cell state in lung adenocarcinoma

doi: 10.1101/2025.10.03.680232

Figure Lengend Snippet: a, Heatmap of Pearson correlations calculated between the HPCS and recurrent pan-cancer cell states (AC: astrocyte-like cell state, OPC: oligodendrocyte progenitor cell-like, NPC: neural progenitor cell-like) , divided by cancer type (BC: breast cancer, CRC: colorectal cancer, HNC: head and neck cancer, LUAD: lung adenocarcinoma, OC: ovarian cancer, PCa: prostate adenocarcinoma, PDAC: pancreatic ductal adenocarcinoma, SCC: squamous cell carcinoma of the skin, ALL: acute lymphoblastic leukemia, MM: multiple myeloma). Arrow indicates the Stress -associated cell state from ref. . A full list of studies is listed in Supplementary Table 5 . b, Pearson correlation between the HPCS signature score ( x -axis) and plastic cell states ( y -axis) from either colorectal cancer (OF/HRC: oncofetal/core high relapse , ) ( top ), and pancreatic ductal adenocarcinoma (scBasal ) ( middle ), or castration-resistant prostate cancer adenocarcinoma (Plasticity ) ( bottom ). Two-tailed p value calculated from the exact distribution of Pearson correlation coefficient. c , Heatmaps demonstrating mean gene signature score in either SLC4A11 or PLAUR negative and positive cells scaled using a z-score generated per program to compare gene signatures across different cancer types in human scRNA-seq datasets ( Supplementary Table 5 ). For each dataset gene pair, significance is calculated by comparing positive vs negative cells and depicted in the positive cell column. Four human LUAD datasets are shown [LungM 5 ; LungK ; LungB ; LungD ] along with plastic cell states [OF/HRC , , lowerSpike , Plasticity ] in other cancers [CRC ; SCC ; PCa ; PDAC ; HNC_C , HNC_K , OC ]. t-test. d, Immunofluorescence of human LUAD stained for (top) uPAR (green) and integrin α2 (red) or (bottom) pan-cytokeratin (white). Arrows point to uPAR and integrin α2 double positive cells. Scale bar: 25 μm. e, Heatmaps demonstrating mean gene signature score in either Slc4a11 or Plaur negative and positive cells scaled using a z-score generated per program to compare gene signatures across different injury models in mouse single cell datasets ( Supplementary Table 5 ). Heatmaps of signature scores represented as z-scores of the indicated regenerative programs in Lung (DATP: damage associated transient progenitor ; ADI: KRT8 + alveolar differentiation intermediate ; PATS: pre-alveolar type-1 transitional cell state ), Pancreas (scBasal cell state ), Intestine (revSC: revival stem cell ; SSC2: singly profiled stem cell cluster 2 ) and Skin (Wound ). For each dataset gene pair, significance is calculated by comparing positive vs negative cells and depicted in the positive cell column. t-test. f, Representative multiplex FISH staining of Slc4a11 (red) and Sftpc (green) in normal lungs vs. 7 days following hyperoxia-induced lung injury. Scale bar: 10 µm.

Article Snippet: Alveolar injury was induced in young C57BL/6 mice by a 32 hour exposure to >85% O 2 in a hyperoxia chamber (BioSpherix), with FiO 2 concentration maintained at a constant flow of ∼3 L O 2 /minute and monitored by an in-line oxygen analyzer.

Techniques: Two Tailed Test, Generated, Immunofluorescence, Staining, Multiplex Assay

Journal: eLife

Article Title: Impaired myofibroblast proliferation is a central feature of pathologic post-natal alveolar simplification

doi: 10.7554/eLife.94425

Figure Lengend Snippet: ( A ) Wildtype C57BL/6 mice were treated in 75% hyperoxia versus normoxia from P0-P10 and recovered in room air until harvest at P40 for analysis by either histology or lung physiology. ( B ) H&E sections of representative lungs from ( A ) harvested at P40 (left) with quantification of mean linear intercept (right). ( C ) Mice treated as in ( A ) and harvested for lung physiology measurements of compliance, elastance and lung capacity. ( D ) Tgfbr2 F/F and Tgfbr2 F/F ;Nkx2.1-cre littermates were treated in hyperoxia versus normoxia from P0-P10 and recovered in room air until harvest at P40 for analysis by histology. ( E ) H&E sections of representative lungs from ( D ) harvested at P40 (left) with quantification of mean linear intercept (right). ( F ) Normoxia cohort treated as in ( D ) and harvested for lung physiology measurements of compliance, elastance and lung capacity. Data in ( B ), ( C ), and ( F ) compared by two-tailed unpaired Student’s t-test. Data in ( E ) compared by ANOVA with Fisher’s post hoc test. Error bars depict mean ± SEM. **p<0.01, ***p<0.001, ****p<0.0001. Scale bars = 100 μm.

Article Snippet: The hyperoxia animal chamber (BioSpherix) was attached to a medical oxygen source controlled by a ProOx single gas controller (BioSpherix) set to maintain 75% oxygen under normobaric conditions.

Techniques: Two Tailed Test

Journal: eLife

Article Title: Impaired myofibroblast proliferation is a central feature of pathologic post-natal alveolar simplification

doi: 10.7554/eLife.94425

Figure Lengend Snippet: ( A ) Wildtype C57BL/6 mice were treated in 75% hyperoxia versus normoxia from P0-P10 and harvested on P10 for analysis by flow cytometry. Graph on right shows total cells per left lung as quantified by flow cytometry. ( B ) Representative flow cytometry plots of the lung mesenchyme (live, CD45-, CD31-, and Epcam-) with gates depicting PDGFRα+ cells (left). Major cell populations of the lung were defined by the indicated cell surface markers and shown as either a percentage of all cells (top) or as absolute number (bottom). ( C ) Tgfb2 F/F and Tgfbr2 F/F ;Nkx2.1-cre littermates were maintained in normoxic conditions from P0-P10 and harvested on P10 for analysis by flow cytometry. Graph on right shows total cells per left lung as quantified by flow cytometry. ( D ) Flow cytometry plots for ( C ) using the same gating strategy as described above. Data in ( A–D ) analyzed by two-tailed unpaired Student’s t-test. Error bars depict mean ± SEM. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001.

Article Snippet: The hyperoxia animal chamber (BioSpherix) was attached to a medical oxygen source controlled by a ProOx single gas controller (BioSpherix) set to maintain 75% oxygen under normobaric conditions.

Techniques: Flow Cytometry, Two Tailed Test

Journal: eLife

Article Title: Impaired myofibroblast proliferation is a central feature of pathologic post-natal alveolar simplification

doi: 10.7554/eLife.94425

Figure Lengend Snippet: ( A ) Schematic of scRNA-seq project showing CTRL ( Tgfbr2 F/F ) and cKO ( Tgfbr2 F/F ;Nkx2.1-cre) littermates treated in 75% hyperoxia versus normoxia from P0-P10 and harvested on P7 or P14 for FACS-purification and analysis by scRNA-seq. n=2 mice harvested for each genotype, treatment condition and timepoint. ( B ) Venn-diagram depicting strategy to compare these two models to identify shared features of alveolar simplification. ( C ) UMAP projection of mesenchymal cells from scRNA-seq as outlined in ( A ). ( D ) Bar graphs depicting the frequency of each cell type by treatment condition, genotype, and timepoint. ( E ) The frequency of alveolar and ductal myofibroblasts within the mesenchyme as depicted in ( D ) with each data point representing either a biologic or technical replicate. Because the data depicts both biologic and technical replicates, no statistics were performed on this data set. ( F ) Differentially expressed genes in myofibroblasts were identified by comparing either CTRL RA vs O2 cells or RA CTRL vs cKO cells. These lists were subsequently analyzed by Qiagen IPA to identify predicted upstream regulators for each comparison. The Venn-diagram on left depicts the number of overlapping predicted upstream regulators with z-score >1.5 (upregulated), while the table on right lists these 32 shared upstream regulators. Blue = TGFβ signaling, red = inhibitors of cell cycle, green = Wnt signaling.

Article Snippet: The hyperoxia animal chamber (BioSpherix) was attached to a medical oxygen source controlled by a ProOx single gas controller (BioSpherix) set to maintain 75% oxygen under normobaric conditions.

Techniques: Purification, Comparison

Journal: eLife

Article Title: Impaired myofibroblast proliferation is a central feature of pathologic post-natal alveolar simplification

doi: 10.7554/eLife.94425

Figure Lengend Snippet: ( A ) Hurskainen et al. treated C57BL/6 wildtype mice with 85% hyperoxia versus normoxia from P0-P14 and analyzed the lungs by scRNA-seq at P3, P7, and P14 . The Seurat object used for publication was provided by the authors. UMAP projection shows the mesenchymal populations as defined by Hurskainen et al. ( B ) By using metadata within the Seurat object, we graphed the frequency of each mesenchymal population by treatment condition and time point. ( C ) The frequency of myofibroblasts within the mesenchyme as depicted in ( B ). ( D ) Xia et al. treated C57BL/6 wildtype mice with 85% hyperoxia versus normoxia from P0-P14 and analyzed the lungs by scRNA-seq at P14 . The Seurat object used for publication was provided by the authors. UMAP projection shows the mesenchymal populations as defined by Xia et al. ( E ) By using metadata within the Seurat object, we graphed the frequency of each mesenchymal population by treatment condition and time point. ( F ) The frequency of alveolar and ductal myofibroblasts within the mesenchyme as depicted in ( E ). Graphs in ( C ) and ( F ) depict one value for each condition because we were unable to extract replicate values from the data provided.

Article Snippet: The hyperoxia animal chamber (BioSpherix) was attached to a medical oxygen source controlled by a ProOx single gas controller (BioSpherix) set to maintain 75% oxygen under normobaric conditions.

Techniques:

Journal: eLife

Article Title: Impaired myofibroblast proliferation is a central feature of pathologic post-natal alveolar simplification

doi: 10.7554/eLife.94425

Figure Lengend Snippet: scRNA-seq data was analyzed using the NicheNet software package. ( A ) Alveolar epithelial clusters were pooled to define sender population and myofibroblast clusters were pooled to define receiver population. By analyzing differential expression of ligands, receptors, and downstream gene expression changes in myofibroblasts, NicheNet predicted increased (left) or decreased (right) ligand-receptor signaling in each comparison of interest. Gray boxes show ligands expressed on the epithelium and the purple-shaded boxes indicate the corresponding receptors expressed on myofibroblasts. Upper panels compare hyperoxia versus normoxia in CTRL cells. Lower panels compare CTRL versus cKO cells in normoxia. ( B ) List of ligand-receptor pairs predicted to be increased in both injury models. Tgfb1-Tgfbr2 pairings highlighted in blue. ( C ) List of ligand-receptor pairs predicted to be decreased in both injury models. Pdgfa-Pdgfra and Pdgfb-Pdgfrb pairings highlighted in red.

Article Snippet: The hyperoxia animal chamber (BioSpherix) was attached to a medical oxygen source controlled by a ProOx single gas controller (BioSpherix) set to maintain 75% oxygen under normobaric conditions.

Techniques: Software, Expressing, Comparison

Journal: eLife

Article Title: Impaired myofibroblast proliferation is a central feature of pathologic post-natal alveolar simplification

doi: 10.7554/eLife.94425

Figure Lengend Snippet: ( A ) Wildtype C57BL/6 mice were injected every other day from P2-P10 with PBS or 1D11 (pan- TGFβ-blocking antibody), treated in 75% hyperoxia treatment versus normoxia from P0-P10, and recovered in room air until harvest at P40 for analysis by either histology or lung physiology. ( B ) H&E sections of representative lungs from ( A ) harvested at P40 (left). Images shown are from PBS and 30 mg/kg 1D11 treatment groups. Mean linear intercepts calculated for all treatment groups (right). ( C ) PBS- and 30 mg/kg 1D11-treated mice treated in normoxic conditions as in ( A ) and harvested for lung physiology measurements of compliance, elastance and lung capacity at P40. Data in ( B ) compared by ANOVA with Fisher’s post hoc test; for readability and limitations of graphing, only the statistical significance values within normoxia or hyperoxia cohorts are plotted. Data in ( C ) compared by two-tailed unpaired Student’s t-test. Error bars depict mean ± SEM. *p<0.05, **p<0.01, ****p<0.0001. Scale bars = 100 μm.

Article Snippet: The hyperoxia animal chamber (BioSpherix) was attached to a medical oxygen source controlled by a ProOx single gas controller (BioSpherix) set to maintain 75% oxygen under normobaric conditions.

Techniques: Injection, Blocking Assay, Two Tailed Test

Journal: eLife

Article Title: Impaired myofibroblast proliferation is a central feature of pathologic post-natal alveolar simplification

doi: 10.7554/eLife.94425

Figure Lengend Snippet: ( A ) Either Gli1-CreERT2 or Pdgfra-CreERT2 mice and their cre-negative littermates were injected with tamoxifen on P2 and P4, treated in 75% hyperoxia versus normoxia from P0-P10, and recovered in room air until harvest at P40 for analysis by histology. ( B ) Mean linear intercepts of Gli1 CreER/+ and Gli1 +/+ mice treated as outlined in ( A ) and harvested at P40. ( C ) Mean linear intercepts of Pdgfra CreER/+ and Pdgfra +/+ mice treated as outlined in ( A ) and harvested at P40. Data compared by ANOVA with Fisher’s post hoc test. Error bars depict mean ± SEM. ***p<0.001, ****p<0.0001.

Article Snippet: The hyperoxia animal chamber (BioSpherix) was attached to a medical oxygen source controlled by a ProOx single gas controller (BioSpherix) set to maintain 75% oxygen under normobaric conditions.

Techniques: Injection

Journal: eLife

Article Title: Impaired myofibroblast proliferation is a central feature of pathologic post-natal alveolar simplification

doi: 10.7554/eLife.94425

Figure Lengend Snippet: ( A ) Tgfbr2 F/F and Tgfbr2 F/F ;Gli1-CreERT2 littermates were injected with tamoxifen on P2 and P4, treated in 75% hyperoxia versus normoxia from P0-P10, and recovered in room air until harvest at P40 for analysis by histology. ( B ) H&E sections of representative lungs from ( A ) harvested at P40 (left). Mean linear intercepts calculated for all treatment groups (right). ( C ) Itgb6 F/F and Itgb6 F/F ;Nkx2.1-cre littermates were treated in 75% hyperoxia versus normoxia from P0-P10, and recovered in room air until harvest at P40 for analysis by histology ( D ) H&E sections of representative lungs from ( C ) harvested at P40 (left). Mean linear intercepts calculated for all treatment groups (right). Data compared by ANOVA with Fisher’s post hoc test. Error bars depict mean ± SEM. *p<0.05, ***p<0.001, ****p<0.0001. Scale bars = 100 μm.

Article Snippet: The hyperoxia animal chamber (BioSpherix) was attached to a medical oxygen source controlled by a ProOx single gas controller (BioSpherix) set to maintain 75% oxygen under normobaric conditions.

Techniques: Injection

Journal: eLife

Article Title: Impaired myofibroblast proliferation is a central feature of pathologic post-natal alveolar simplification

doi: 10.7554/eLife.94425

Figure Lengend Snippet: ( A ) Itgav F/F and Itgav F/F ;Gli1-CreERT2 littermates were injected with tamoxifen on P2 and P4, treated in 75% hyperoxia versus normoxia from P0-P10, and recovered in room air until harvest at P40 for analysis by either histology or lung physiology. ( B ) H&E sections of representative lungs from ( A ) harvested at P40 (left). Mean linear intercepts calculated for all treatment groups (right). ( C ) Normoxia cohort treated as in ( A ) and harvested for lung physiology measurements of compliance, elastance and lung capacity. Data in ( B ) compared by ANOVA with Fisher’s post hoc test. Data in ( C ) compared by two-tailed unpaired Student’s t-test. Error bars depict mean ± SEM. *p<0.05, **p<0.01, ****p<0.0001. Scale bars = 100 μm.

Article Snippet: The hyperoxia animal chamber (BioSpherix) was attached to a medical oxygen source controlled by a ProOx single gas controller (BioSpherix) set to maintain 75% oxygen under normobaric conditions.

Techniques: Injection, Two Tailed Test

Journal: eLife

Article Title: Impaired myofibroblast proliferation is a central feature of pathologic post-natal alveolar simplification

doi: 10.7554/eLife.94425

Figure Lengend Snippet: ( A ) Wildtype C57BL/6 mice were treated in 75% hyperoxia versus normoxia from P0-P10 and recovered in room air until indicated timepoints for analysis. Mice were injected with EdU 24 hr prior to each analysis timepoint. ( B ) Flow cytometry plots of lung mesenchyme (CD45-, CD31-, Epcam-, MCAM-) show gating of PDGFRα+ cells (upper panels) and subsequent identification of EdU+/PDGFRα+ cells (lower panels). Panels on far-right show an EdU-untreated littermate used to define EdU + gate. ( C ) Time course graphs showing indicated populations of the lung as percent of lung (top), total cells in left lung (middle), and percent EdU + cells (bottom). Data graphed as mean ± with exception of EdU + panel in which each animal is plotted individually. Data compared by two-tailed unpaired Student’s t-test. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001.

Article Snippet: The hyperoxia animal chamber (BioSpherix) was attached to a medical oxygen source controlled by a ProOx single gas controller (BioSpherix) set to maintain 75% oxygen under normobaric conditions.

Techniques: Injection, Flow Cytometry, Two Tailed Test

Journal: eLife

Article Title: Impaired myofibroblast proliferation is a central feature of pathologic post-natal alveolar simplification

doi: 10.7554/eLife.94425

Figure Lengend Snippet: ( A ) Mean fluorescence intensity (geometric mean, MFI) of PDGFRα antibody staining on MCAM-negative mesenchymal cells (CD45-, CD31-, Epcam-) and PDGFRa + cells (CD45-, CD31-, Epcam-, MCAM-, PDGFRα+) as quantified by flow cytometry. Each column represents an experiment shown earlier in this study: normoxia vs hyperoxia at P10 , Tgfbr2 F/F vs Tgfbr2 F/F ;Nkx2.1-cre at P10 , PBS vs 1D11 at P10 , Ect2 F/F vs Ect2 F/F ;Pdgfra-CreERT2 at P14 . To compare values across multiple experiments, MFI’s were normalized to the control condition within each experiment. Data compared by ANOVA with Fisher’s post hoc test. Error bars depict mean ± SEM. ***p<0.001, ****p<0.0001.

Article Snippet: The hyperoxia animal chamber (BioSpherix) was attached to a medical oxygen source controlled by a ProOx single gas controller (BioSpherix) set to maintain 75% oxygen under normobaric conditions.

Techniques: Fluorescence, Staining, Flow Cytometry, Control