krt5  (Servicebio Inc)

Journal: International Journal of Molecular Sciences

Article Title: The Impact of Alternate-Day Fasting on the Salivary Gland Ductal Compartments and the Differentiation Potential of Keratin 5 + Salivary Gland Progenitor Cells in an Induced Mouse Model of Sjögren’s-like Hyposalivation

doi: 10.3390/ijms27094080

Figure Lengend Snippet: Immunization with salivary gland proteins induces hyposalivation, accompanied by elevated P16 expression in the submandibular glands (SMGs) of KRT5 CreERT2 ; R26 tdTomato mice. Eight-week-old female KRT5 CreERT2 ; R26 tdTomato mice were immunized with total salivary gland proteins following tamoxifen treatment. ( A ) Experimental timeline of tamoxifen induction, immunizations and analyses. ( B ) Measurement of salivary flow rate (n = 6–8). ( C ) Representative images of immunohistochemical staining of the SMG sections for P16 (scale bar = 100 μm, 400× magnification). The bar graph shows the average percentage of positively stained areas (n = 4–5). Error bars indicate the standard error of the mean (SEM).

Article Snippet: Female KRT5 CreERT2 mice (Cat# 029155), R26 tdTomato reporter mice (Cat# 007914) and C57BL/6 mice (Cat# 000664) were obtained from the Jackson Laboratory (Bar Harbor, ME, USA) and maintained under specific pathogen-free conditions at the ADA Forsyth Institute.

Techniques: Expressing, Immunohistochemical staining, Staining

Journal: International Journal of Molecular Sciences

Article Title: The Impact of Alternate-Day Fasting on the Salivary Gland Ductal Compartments and the Differentiation Potential of Keratin 5 + Salivary Gland Progenitor Cells in an Induced Mouse Model of Sjögren’s-like Hyposalivation

doi: 10.3390/ijms27094080

Figure Lengend Snippet: ADF increases salivary flow rate and reduces cellular senescence in the SMGs of KRT5 CreERT2 ; R26 tdtomato mice with induced hyposalivation. Eight-week-old female KRT5 CreERT2 ; R26 tdTomato mice were immunized with SMG proteins following tamoxifen treatment and then subjected to alternate-day fasting (ADF) or fed with standard chow ad libitum (AL), as shown in ( A ) experimental timeline. ( B ) Measurement of salivary flow rate (n = 6–8). ( C ) Representative images of immunohistochemical staining of the SMG sections for P16 (scale bar = 100 μm, 400× magnification). The bar graph shows the average percentage of positively stained areas (n = 5–6). Error bars indicate the standard error of the mean (SEM).

Article Snippet: Female KRT5 CreERT2 mice (Cat# 029155), R26 tdTomato reporter mice (Cat# 007914) and C57BL/6 mice (Cat# 000664) were obtained from the Jackson Laboratory (Bar Harbor, ME, USA) and maintained under specific pathogen-free conditions at the ADA Forsyth Institute.

Techniques: Immunohistochemical staining, Staining

Journal: International Journal of Molecular Sciences

Article Title: The Impact of Alternate-Day Fasting on the Salivary Gland Ductal Compartments and the Differentiation Potential of Keratin 5 + Salivary Gland Progenitor Cells in an Induced Mouse Model of Sjögren’s-like Hyposalivation

doi: 10.3390/ijms27094080

Figure Lengend Snippet: ADF reduces the expression of anti-apoptotic proteins BCL-2, BCL-XL and MCL-1 in the SMGs of KRT5 CreERT2 ; R26 tdtomato mice with induced hyposalivation. Eight-week-old female KRT5 CreERT2 ; R26 tdTomato mice were immunized with SMG proteins following tamoxifen treatment and then subjected to ADF or fed with standard chow AL , as outlined in A. Representative images of immunohistochemical staining of the SMG sections for BCL-2, BCL-XL or MCL-1 (scale bar = 100 µm, 400× magnification). The bar graphs display the average percentage of positively stained areas (n = 5–6). Error bars indicate the standard error of the mean (SEM).

Article Snippet: Female KRT5 CreERT2 mice (Cat# 029155), R26 tdTomato reporter mice (Cat# 007914) and C57BL/6 mice (Cat# 000664) were obtained from the Jackson Laboratory (Bar Harbor, ME, USA) and maintained under specific pathogen-free conditions at the ADA Forsyth Institute.

Techniques: Expressing, Immunohistochemical staining, Staining

Journal: International Journal of Molecular Sciences

Article Title: The Impact of Alternate-Day Fasting on the Salivary Gland Ductal Compartments and the Differentiation Potential of Keratin 5 + Salivary Gland Progenitor Cells in an Induced Mouse Model of Sjögren’s-like Hyposalivation

doi: 10.3390/ijms27094080

Figure Lengend Snippet: ADF reduces the expression levels of NLRP3 and their downstream products IL-1β and IL-18 in the SMGs of KRT5 CreERT2 ; R26 tdTomato mice with induced hyposalivation. Eight-week-old female KRT5 CreERT2 ; R26 tdTomato mice were immunized with SMG proteins following tamoxifen treatment and then subjected to ADF or fed with standard chow AL , as outlined in A. Representative images of immunohistochemical staining of the SMG protein sections for NLRP3, IL-1β and IL-18 are shown (scale bar = 100 µm, 400× magnification). The bar graphs display the average percentage of positively stained areas (n = 5–6). Error bars indicate the standard error of the mean (SEM).

Article Snippet: Female KRT5 CreERT2 mice (Cat# 029155), R26 tdTomato reporter mice (Cat# 007914) and C57BL/6 mice (Cat# 000664) were obtained from the Jackson Laboratory (Bar Harbor, ME, USA) and maintained under specific pathogen-free conditions at the ADA Forsyth Institute.

Techniques: Expressing, Immunohistochemical staining, Staining

Journal: International Journal of Molecular Sciences

Article Title: The Impact of Alternate-Day Fasting on the Salivary Gland Ductal Compartments and the Differentiation Potential of Keratin 5 + Salivary Gland Progenitor Cells in an Induced Mouse Model of Sjögren’s-like Hyposalivation

doi: 10.3390/ijms27094080

Figure Lengend Snippet: ADF moderately promotes acinar cell differentiation from KRT5 + cells in the SMGs of KRT5 CreERT2 ; R26 tdTomato mice with induced hyposalivation. ( A ) Flow cytometric analysis of KRT5 and tdTomato expression in SMG cells from KRT5 CreERT2 ; R26 tdTomato mice 2 weeks after corn oil or tamoxifen treatment. ( B ) Eight-week-old female KRT5 CreERT2 ; R26 tdTomato mice were immunized with SMG proteins following tamoxifen treatment and then subjected to ADF or fed with standard chow AL , as outlined in A. Representative images of immunofluorescence staining of the SMG sections for tdTomato (red) and AQP5 (acinar cell marker, green) are shown. DAPI: blue. Scale bar = 100 µm, 400× magnification (n = 9). The arrow indicates cell aggregates co-expressing tdTomato and AQP5.

Article Snippet: Female KRT5 CreERT2 mice (Cat# 029155), R26 tdTomato reporter mice (Cat# 007914) and C57BL/6 mice (Cat# 000664) were obtained from the Jackson Laboratory (Bar Harbor, ME, USA) and maintained under specific pathogen-free conditions at the ADA Forsyth Institute.

Techniques: Cell Differentiation, Expressing, Immunofluorescence, Staining, Marker

Journal: bioRxiv

Article Title: Comparative analysis of varicella-zoster virus and herpes simplex virus 1 interaction with epidermal terminal differentiation in primary human keratinocytes models of differentiation

doi: 10.64898/2026.04.08.717198

Figure Lengend Snippet: A. Schematic of plan of infection with HSV-1 at MOI 0.005 of undifferentiated (Undiff.), early differentiated (Early diff.) and late differentiated (Late diff.) NHEKs. “Undiff.”, “Early diff.” and “Late diff.” labelling refers to the state of NHEKs differentiation at the time of infection (day 0). The “Undiff.” condition includes the NHEKs undergoing spontaneous asynchronous differentiation after infection (Differentiating_async.) B. Analysis by qPCR of cell-associated HSV-1 genome copy number normalised to number of cells, upon infection with HSV-1 of Undiff., Early diff. and Late diff. NHEKs, and evaluated at 24h, 48h and 72h p.i. The data are generated from quantification of the HSV-1 UL27 gene (gB) normalised to RPP30 gene (host internal control) and are reported as mean of n=3 independent experiments ± SEM. C. Analysis by qPCR of cell-associated HSV-1 genome copy number normalised to number of cells, upon infection with HSV-1 of Late diff. NHEKs, and reported at 72h and 120h p.i. The data are generated from quantification of the HSV-1 UL27 gene (gB) normalised to RPP30 gene (host internal control) and are reported as the mean of three technical replicates. D. ICP0 (pink) immunofluorescence staining of Undiff., Early diff. and Late diff. NHEKs infected with HSV-1 and analysed at 4h, 24h and 72h p.i. The reported images include GFP (green) signal deriving from the GFP protein tagged to the late HSV-1 tegument protein UL46 and DAPI (grey) staining of nuclei. E. Analysis by qRT-PCR analysis of RL2 gene (encoding the ICP0 protein) and UL48 gene (encoding the VP16 protein) expression in HSV-1 infection of Undiff., Early diff. and Late diff. NHEKs, and reported at 24h, 48h and 72h p.i. The data are reported as mean of 2 -dCT of n=3 independent experiments ± SEM, where the normalisation was performed against GAPDH . F. Western Blotting analysis of ICP0 and VP16 expression at 24h, 48h and 72h p.i. following infection of Undiff., Early diff. and Late diff. NHEKs with HSV-1. GAPDH was used as loading control. Data are representative of n=4 independent experiments (Undiff. condition), n=3 independent experiments (Early diff. condition) and n=2 independent experiments (Late diff. condition). G. Schematic of the model of HSV-1 infection implemented in undifferentiated NHEKs, which either remained undifferentiated throughout the infection and until the last time point (72h) (Undifferentiated) or started undergoing spontaneous differentiation during the last time points of infection (Differentiating_async). H. Analysis by qRT-PCR of KRT5 and KRT10 mRNAs in uninfected NHEKs at 72h confirming the undifferentiated and differentiating status of the keratinocytes. The data are reported as fold change (FC) (2 -ddCT ) to the undifferentiated condition and are the mean of three technical replicates, where the normalisation was performed against GAPDH . I. Keratinocytes where infected when undifferentiated and then either maintained their undifferentiated status throughout the infection (Undifferentiated) or were allowed to spontaneously differentiate by 72h p.i. (Differentiating). Cell-associated HSV-1 genome copy number was analysed at 72h p.i. in both conditions. The data are generated from quantification of the HSV-1 UL27 gene (gB) normalised to RPP30 gene (host internal control), are reported as the mean of three technical replicates and are representative of n=3 independent experiments. J. Anlysis by qRT-PCR of RL2 and UL48 mRNAs in HSV-1 infected Undifferentiated and Differentiating NHEKs at 72h p.i. The data are reported as mean of 2 -dCT of three technical replicates, where the normalisation was performed against GAPDH . Data are representative of n=3 independent experiments. Statistical significance was evaluated in B. and E. by 2way ANOVA with Tukey’s multiple comparisons test and indicated as *P< 0.05, **P< 0.01, ***P< 0.001, ****P< 0.0001. In D. , dotted lines mark nuclei at 4h p.i. Insets contain zoomed-in areas of the ICP0 signal inside the nuclei at 4h p.i. Scale bar, 10 μm. Undiff., undifferentiated; Early diff., early differentiated; Late diff., late differentiated; p.i., post infection; NHEKs, normal human epidermal keratinocytes.

Article Snippet: The primers and probes used for host genes were purchased from Thermofisher Scientific, cat n 4448489 (VIC dye): GAPDH (ASSAY ID Hs02786624_g1) and cat n 4331182 (FAM dye): KRT10 (assay ID Hs00166289_m1), KRT1 (assay ID Hs00196158_m1), KRT5 (assay ID Hs00361185_m1), LOR (assay ID Hs01894962_s1), FLG (assay ID Hs00856927_g1), LCE3D (assay ID Hs00754375_s1), DSC1 (assay ID Hs00245189_m1).

Techniques: Infection, Generated, Control, Immunofluorescence, Staining, Quantitative RT-PCR, Expressing, Western Blot

Journal: Advanced Science

Article Title: Activation of ERBB4 Pathway Inhibits Pathological Transdifferentiation of Lung Epithelial Progenitors into CD66c + Basal Cells in Severe Lung Injury

doi: 10.1002/advs.202519151

Figure Lengend Snippet: Emergence of CD66c + basal cells (BCs) in distal lung organoids and their pro‐fibrotic properties in vitro and in vivo. (a) Schematic diagram of human distal lung organoid (DLO) cultures generating alveolar type 2 (AT2) organoids, secretory cell organoids (SCOs), and basal cell organoids (BCOs). (b–e) Flow cytometry analysis of EPCAM + epithelial cells from uncultured distal lung tissue (b, c) and day 14 DLOs (d, e), showing proportions of AT2 cells, secretory cells, CD66c − BCs, and CD66c + BCs ( n = 3). (f) Morphology of DLOs. Bright‐field (left) and H&E staining (right) show representative BCO structures. Scale bars, 50 µm. (g) Immunofluorescence (IF) staining of BCOs showing CD66c (green) and KRT5 (magenta) double‐positive cells. Boxed region is enlarged on the right. Scale bars, 50 µm. (h) RT‐qPCR analysis of ERBB ligands and fibrosis‐related genes in FACS‐sorted CD66c + BCs compared with secretory cells ( n = 3). (i, j) IF staining of BCOs showing HBEGF (i) or MUC1 (j) co‐expressed with KRT5. Nuclei counterstained with DAPI. Scale bars, 50 µm. (k) Representative bright‐field images and immunofluorescence staining for αSMA (magenta) and Ki67 (green) of MRC‐5 fibroblasts cultured alone or co‐cultured with CD66c + BCs. (l) Percentage of Ki67 + MRC‐5 fibroblasts cultured alone or co‐cultured with CD66c + BCs ( n = 4). (m) Relative mRNA expression of MKI67 of MRC‐5 fibroblasts cultured alone or co‐cultured with CD66c + BCs. Data are shown as mean ± SEM ( n = 4). (n) RT‐qPCR of MRC‐5 fibroblasts treated with conditioned medium from CD66c + BC organoids, showing upregulation of fibroblast activation‐associated genes ( n = 3–5). (o–q) In vivo fibrosis assay. (o) Experimental scheme of subcutaneous transplantation of CD66c − or CD66c + BCs into NSG mice. (p) Representative Masson's trichrome and IF staining showing collagen deposition and αSMA + myofibroblasts in CD66c + BC grafts. Scale bars, 250 µm (Masson), 100 µm (IF). (q) Quantification of the stromal fibrosis index (SFI). Data represent mean ± SEM ( n = 3). Data are shown as mean ± SEM. Statistical significance was determined by unpaired two‐tailed Student's t ‐test. n represents independent biological replicates. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Primary antibodies included: HT2‐280 (1:200, Terrace Biotechnologies, TB‐27AHT2‐280), KRT5 (1:500, ABclonal, 9661), CD66c (1:50, BioLegend, 365002), HBEGF (1:200, Bioss, bs‐3576R), MUC1 (1:200, BD Pharmingen, 555925), αSMA (1:500, Sigma, C6198), Ki67 (1:500, BD Biosciences, 550609), HER2 (1:200, Abcam, ab134182), SCGB1A1 (1:500, R&D Systems, MAB4218‐SP), PDPN (1:200, BioLegend, 337001), SFTPB (1:200, Santa Cruz, sc‐133143), SFTPC (1:200, Millipore, AB3786; alternatively, Abcam, ab312850), ERBB4 (1:50, Invitrogen, MA1‐861), and CDH1 (1:400, BD Biosciences, 610181).

Techniques: In Vitro, In Vivo, Flow Cytometry, Staining, Immunofluorescence, Quantitative RT-PCR, Cell Culture, Expressing, Activation Assay, Transplantation Assay, Two Tailed Test

Journal: Advanced Science

Article Title: Activation of ERBB4 Pathway Inhibits Pathological Transdifferentiation of Lung Epithelial Progenitors into CD66c + Basal Cells in Severe Lung Injury

doi: 10.1002/advs.202519151

Figure Lengend Snippet: Distal lung progenitors generate CD66c + basal cells (BCs) in organoid cultures. (a) Experimental workflow: FACS isolation of secretory cells (HT2‐280 − NGFR − ITGA6 − CD66c + ) from DLOs, followed by approximately 14 days of three‐dimensional culture. (b) Flow cytometry analysis of epithelial populations in secretory cell‐derived organoids at day 14, highlighting the proportion of CD66c + basal cells. (c) Bright‐field (left) and H&E staining (right) of secretory‐derived organoids. Scale bar, 50 µm. (d) Immunofluorescence showing DAPI (blue), CD66c (green), KRT5 (magenta), and CC10 (gray). Scale bar, 50 µm. (e) Serial sections of secretory‐derived organoids showing CD66c with SFTPB (top) and KRT5 with PDPN (bottom). Arrowheads indicate squamous basal cells; asterisks denote nonspecific staining. Scale bars, 50 µm. (f, g) RT‐qPCR analysis of secretory cells before and after culture, showing loss of secretory markers (f) and induction of basal markers ( KRT5 , TP63 ) and the SPB‐associated marker KRT16 (g) ( n = 3). (h) Experimental workflow: FACS isolation of AT2 cells (HT2‐280 + NGFR − ITGA6 − ) from DLOs, followed by approximately 14 days of culture. (i) Flow cytometry of AT2‐derived organoids showing the emergence of CD66c + BCs. (j) Bright‐field (left) and H&E staining (right) of AT2‐derived organoids, showing compact spheroid structures. Scale bar, 50 µm. (k) Immunofluorescence showing DAPI (blue), CD66c (green), KRT5 (magenta), and the AT2 marker SFTPC (gray). Scale bar, 50 µm. (l, m) RT‐qPCR analysis of AT2 cells before and after culture, showing decreased expression of AT2 markers (l) and increased expression of basal markers and the SPB‐associated marker KRT16 (m) ( n = 3). (n) Quantification of cell type composition in BC‐, SC‐, and AT2‐derived organoids by flow cytometry. Populations include ungated cells, CD66c + BCs, CD66c − BCs, secretory cells, and AT2 cells, with CD66c + BC proportions highlighted ( n = 3). CD66c + BC proportions differed significantly across groups (one‐way ANOVA with Tukey's multiple comparisons test). Data are shown as mean ± SEM. Statistical significance was determined by unpaired two‐tailed Student's t ‐test for comparisons between two groups and by one‐way ANOVA with Tukey's multiple comparisons test for multiple group comparisons. n represents independent biological replicates. * p < 0.05, ** p < 0.01, *** p < 0.001. [Correction added on 21 April 2026 after first online publication: Figure 3m has been updated.]

Article Snippet: Primary antibodies included: HT2‐280 (1:200, Terrace Biotechnologies, TB‐27AHT2‐280), KRT5 (1:500, ABclonal, 9661), CD66c (1:50, BioLegend, 365002), HBEGF (1:200, Bioss, bs‐3576R), MUC1 (1:200, BD Pharmingen, 555925), αSMA (1:500, Sigma, C6198), Ki67 (1:500, BD Biosciences, 550609), HER2 (1:200, Abcam, ab134182), SCGB1A1 (1:500, R&D Systems, MAB4218‐SP), PDPN (1:200, BioLegend, 337001), SFTPB (1:200, Santa Cruz, sc‐133143), SFTPC (1:200, Millipore, AB3786; alternatively, Abcam, ab312850), ERBB4 (1:50, Invitrogen, MA1‐861), and CDH1 (1:400, BD Biosciences, 610181).

Techniques: Isolation, Flow Cytometry, Derivative Assay, Staining, Immunofluorescence, Quantitative RT-PCR, Marker, Expressing, Two Tailed Test

Journal: Advanced Science

Article Title: Activation of ERBB4 Pathway Inhibits Pathological Transdifferentiation of Lung Epithelial Progenitors into CD66c + Basal Cells in Severe Lung Injury

doi: 10.1002/advs.202519151

Figure Lengend Snippet: CHIR99021 treatment preserves secretory and AT2 cell identities and suppresses their transdifferentiation Into CD66c + basal cells. (a) Bright‐field (top) and H&E (bottom) images of distal lung organoids (DLOs) cultured without (DLO–CHIR) or with CHIR99021 (DLO+CHIR). CHIR promoted a lumenized organoid morphology and reduced the formation of solid spheroids. Scale bars, 50 µm. (b) Immunofluorescence staining of DLOs showing nuclei (DAPI, blue), SFTPB (green), and KRT5 (magenta). Enlarged views of boxed regions are shown below. Scale bars, 50 µm. (c) Flow cytometry of EPCAM + epithelial cells from DLO–CHIR and DLO+CHIR showing reduced proportions of CD66c + basal cells (BCs) upon CHIR treatment. (d) Pie charts summarizing epithelial composition in −CHIR versus +CHIR DLOs ( n = 3). (e) Bright‐field (top) and immunofluorescence (bottom) of DLOs after long‐term CHIR exposure (>60 days) versus CHIR withdrawal, showing reappearance of CD66c + BCs upon CHIR removal. Scale bars, 50 µm. (f) Bright‐field (top) and immunofluorescence (bottom) of secretory‐derived organoids cultured 14 days without (SC–CHIR) or with CHIR (SC+CHIR). SC–CHIR organoids formed solid spheroids enriched in KRT5 + BCs, whereas SC+CHIR organoids formed hollow structures enriched for SFTPB + and CC10 + secretory cells. Scale bars, 50 µm. (g) Representative FACS plots of EPCAM + cells from SC‐derived organoids (−CHIR vs. +CHIR). (h) Quantification of CD66c + BCs within EPCAM + cells showing a significant reduction in the CHIR‐treated group ( n = 3). (i) RT‐qPCR analysis showing reduced KRT5 expression in SC+CHIR organoids compared with SC–CHIR controls ( n = 3). (j) Bright‐field (top) and immunofluorescence (bottom) of AT2‐derived organoids cultured 14 days without or with CHIR. −CHIR organoids formed dense spheroids expressing CD66c (green) and KRT5 (magenta), whereas +CHIR organoids retained AT2 identity, marked by SFTPC (gray). Scale bars, 50 µm. (k) UMAP of scRNA‐seq data from SC‐derived organoids (−CHIR vs. +CHIR) integrated with the published AT2‐derived dataset (Kathiriya et al.) to assess CHIR‐induced shifts in epithelial cell states, showing AT2, alveolar–basal intermediates (ABIs), secretory cells, CD66c − BCs, and CD66c + BCs. (l) Dot plot of representative marker genes for each population. Dot size indicates the percentage of positive cells; color intensity reflects average expression levels. Data are shown as mean ± SEM. Statistical significance was determined by unpaired two‐tailed Student's t ‐test. n represents independent biological replicates. ** p < 0.01, *** p < 0.001.

Article Snippet: Primary antibodies included: HT2‐280 (1:200, Terrace Biotechnologies, TB‐27AHT2‐280), KRT5 (1:500, ABclonal, 9661), CD66c (1:50, BioLegend, 365002), HBEGF (1:200, Bioss, bs‐3576R), MUC1 (1:200, BD Pharmingen, 555925), αSMA (1:500, Sigma, C6198), Ki67 (1:500, BD Biosciences, 550609), HER2 (1:200, Abcam, ab134182), SCGB1A1 (1:500, R&D Systems, MAB4218‐SP), PDPN (1:200, BioLegend, 337001), SFTPB (1:200, Santa Cruz, sc‐133143), SFTPC (1:200, Millipore, AB3786; alternatively, Abcam, ab312850), ERBB4 (1:50, Invitrogen, MA1‐861), and CDH1 (1:400, BD Biosciences, 610181).

Techniques: Cell Culture, Immunofluorescence, Staining, Flow Cytometry, Derivative Assay, Quantitative RT-PCR, Expressing, Marker, Two Tailed Test

Journal: Advanced Science

Article Title: Activation of ERBB4 Pathway Inhibits Pathological Transdifferentiation of Lung Epithelial Progenitors into CD66c + Basal Cells in Severe Lung Injury

doi: 10.1002/advs.202519151

Figure Lengend Snippet: NRG1 signaling inhibits transdifferentiation of secretory and AT2 CELLS Into CD66c + basal cells. (a) Experimental workflow: Secretory cells were isolated by fluorescence‐activated cell sorting (FACS) from distal lung organoids (DLOs) and cultured for ∼14 days in control medium or recombinant NRG1. (b) Representative flow cytometry plots of EPCAM + epithelial cells from secretory cell‐derived organoids showing reduced CD66c + basal cells (BCs) upon NRG1 treatment. (c) Quantification of epithelial subsets in secretory cell‐derived organoids cultured ± NRG1 ( n = 3). (d) Bright‐field and immunofluorescence (IF) images of secretory‐derived organoids ± NRG1. NRG1 suppressed solid organoid formation and preserved expression of secretory markers (SFTPB, CC10), while reducing KRT5 + BCs. Nuclei were counterstained with DAPI. Scale bars, 50 µm. (e) Representative flow cytometry plots showing phenotypic changes of secretory cell‐derived basal cells following siRNA transfection and NRG1 treatment, with siCtrl (−NRG1) included as the untreated control. (f, g) Flow cytometric quantification of CD66c + basal cells among EPCAM + epithelial cells under the indicated conditions ( n = 3). (f) Comparison between siCtrl (−NRG1) and siCtrl (+NRG1), and (g) comparison between siERBB4 or siERBB2 under NRG1 treatment. (h) Experimental workflow for AT2‐derived organoids: FACS‐sorted AT2 cells (HT2‐280 + NGFR − ITGA6 − ) were cultured ± NRG1 for ∼14 days. (i) Flow cytometry plots of EPCAM + cells from AT2‐derived organoids, showing decreased CD66c + BCs after NRG1 treatment. (j) Quantification of epithelial subsets in AT2‐derived organoids cultured ± NRG1 ( n = 3). (k) Bright‐field and IF images of AT2‐derived organoids cultured ± NRG1 for 14 days. NRG1 promoted cystic morphology, preserved AT2 marker HT2‐280 (gray), and reduced CD66c + KRT5 + BCs. Nuclei were counterstained with DAPI (blue). Scale bars, 50 µm. (l, m) Flow cytometric quantification of CD66c + basal cells derived from AT2 cells under the indicated conditions ( n = 3). (l) Comparison between siCtrl (−NRG1) and siCtrl (+NRG1), and (m) comparison between siERBB4 or siERBB2 under NRG1 treatment. Data are shown as mean ± SEM. Statistical significance was determined by unpaired two‐tailed Student's t ‐test for comparisons between two groups and by one‐way ANOVA with appropriate multiple comparisons tests for analyses involving three groups. n represents independent biological replicates. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Primary antibodies included: HT2‐280 (1:200, Terrace Biotechnologies, TB‐27AHT2‐280), KRT5 (1:500, ABclonal, 9661), CD66c (1:50, BioLegend, 365002), HBEGF (1:200, Bioss, bs‐3576R), MUC1 (1:200, BD Pharmingen, 555925), αSMA (1:500, Sigma, C6198), Ki67 (1:500, BD Biosciences, 550609), HER2 (1:200, Abcam, ab134182), SCGB1A1 (1:500, R&D Systems, MAB4218‐SP), PDPN (1:200, BioLegend, 337001), SFTPB (1:200, Santa Cruz, sc‐133143), SFTPC (1:200, Millipore, AB3786; alternatively, Abcam, ab312850), ERBB4 (1:50, Invitrogen, MA1‐861), and CDH1 (1:400, BD Biosciences, 610181).

Techniques: Isolation, Fluorescence, FACS, Cell Culture, Control, Recombinant, Flow Cytometry, Derivative Assay, Immunofluorescence, Expressing, Transfection, Comparison, Marker, Two Tailed Test

Journal: Advanced Science

Article Title: Activation of ERBB4 Pathway Inhibits Pathological Transdifferentiation of Lung Epithelial Progenitors into CD66c + Basal Cells in Severe Lung Injury

doi: 10.1002/advs.202519151

Figure Lengend Snippet: Loss of ERBB4 expression in epithelial cells of IPF lungs. (a) Multiplex immunofluorescence (IF) staining of lung tissue from patients with idiopathic pulmonary fibrosis (IPF) showing nuclei (DAPI, blue), CD66c (green), KRT5 (magenta), and αSMA (gray). The white dashed box marks the region shown in panel (b). Scale bars, 50 µm. (b) Higher‐magnification views of the boxed region in panel (a) show CD66c + KRT5 + basal cells (BCs; arrows) adjacent to αSMA + myofibroblasts in IPF lungs (lower row). Representative healthy lung tissue is shown in the upper row for comparison. Scale bars, 50 µm. (c) Spatial proximity of epithelial subsets to αSMA + regions in IPF lungs. Violin plots show the distribution of nearest‐neighbor distances from epithelial cells to αSMA + myofibroblasts for CD66c + BCs, CD66c − BCs, and CD66c + secretory cells (SCs; CD66c + KRT5 − ). White squares and thick lines denote medians and interquartile ranges, respectively. Median distances (µm) are shown above. Data were obtained from multiplex IF images of two IPF patients ( n = 2). (d) Multiplex IF of normal alveoli shows strong ERBB4 (magenta) colocalized with the AT2 marker SFTPC (white), with minimal or no KRT5 + BCs (green). Nuclei: DAPI (blue). Scale bars, 50 µm. (e) In normal airways, ERBB4 (magenta) is expressed in CC10 + SCs (green) but is minimal or absent in KRT5 + BCs (white). Nuclei: DAPI (blue). Scale bars, 50 µm. (f) Low‐magnification view of IPF parenchyma reveals honeycomb cysts labeled for DAPI (blue), KRT5 (green), ERBB4 (magenta), and SFTPC (gray). Scale bars, 50 µm. (g) Enlarged view of the boxed region in panel (f) highlights honeycomb cysts densely lined with KRT5 + BCs and reduced ERBB4 expression, with few SFTPC + AT2 cells. Scale bars, 50 µm. (h) Residual alveolar regions in IPF lungs exhibit reduced ERBB4 expression in SFTPC + AT2 cells. Scale bars, 50 µm. (i) UMAP plots of scRNA‐seq data comparing normal distal lung epithelium (left, this study) and IPF lung tissue (right, Kathiriya et al., 2022). ERBB4 expression is enriched in normal epithelium, particularly in AT2 and SC clusters (outlined), but is markedly reduced in IPF lungs. (j) Proposed model of ERBB4 signaling in regulating epithelial plasticity during lung injury. Distal lung epithelial progenitors (AT2 cells, secretory cells, and CD66c − BCs) exhibit lineage plasticity and can converge toward a CD66c + basal state under injury‐associated conditions. During physiological repair, NRG1‐ERBB4‐MAPK signaling maintains epithelial identity and restrains aberrant basalization, in association with balanced BMP‐ and TGFβ‐related programs. In IPF, attenuation of ERBB4 signaling coincides with increased generation of pro‐fibrotic CD66c + BCs within disease‐associated niches enriched for TGFβ‐ and YAP‐related activity, contributing to pathological remodeling and impaired regeneration.

Article Snippet: Primary antibodies included: HT2‐280 (1:200, Terrace Biotechnologies, TB‐27AHT2‐280), KRT5 (1:500, ABclonal, 9661), CD66c (1:50, BioLegend, 365002), HBEGF (1:200, Bioss, bs‐3576R), MUC1 (1:200, BD Pharmingen, 555925), αSMA (1:500, Sigma, C6198), Ki67 (1:500, BD Biosciences, 550609), HER2 (1:200, Abcam, ab134182), SCGB1A1 (1:500, R&D Systems, MAB4218‐SP), PDPN (1:200, BioLegend, 337001), SFTPB (1:200, Santa Cruz, sc‐133143), SFTPC (1:200, Millipore, AB3786; alternatively, Abcam, ab312850), ERBB4 (1:50, Invitrogen, MA1‐861), and CDH1 (1:400, BD Biosciences, 610181).

Techniques: Expressing, Multiplex Assay, Immunofluorescence, Staining, Comparison, Marker, Labeling, Activity Assay

Journal: Nature Communications

Article Title: Tuft cells shape airway remodeling by eliciting OXGR1- and SOX9-dependent stem cell programs

doi: 10.1038/s41467-026-70763-y

Figure Lengend Snippet: a Representative images of DCLK1 and BCAM staining in tracheal sections from the indicated Sox9-tdT reporter strains at day 14. Sox9-tdT + BCAM + cells (white arrow). Scale bars: 50 µm. b Quantification of Sox9-tdT + BCAM + cells in the SAE from the indicated strains at day 14. Group sizes: n = 5, 6, 12, 5 mice, from left to right. Two-tailed unpaired Mann-Whitney U test. c , d Combined in situ hybridization (ISH) for Oxgr1, Krt5 and Krt8 , and immunofluorescence for SOX9 and α-SMA in tracheal sections from WT mice with or without ALT treatment ( Krt5 for BCs, Krt8 for differentiated cells, SOX9 for SMG progenitor cells, and α-SMA for SMG myoepithelial cells). Oxgr1 + SOX9 + cells (green arrows). Scale bars: 20 μm. SMG Submucosal gland. ( e ) Quantification of Oxgr1 + cells in the specified EpC subsets. Group sizes: n = 7, 16, 8, 13, 6, 10, 7, 8 mice, from left to right. Kruskal–Wallis test followed by Dunn’s multiple-comparisons test with Holm–Bonferroni correction. f Quantification of Oxgr1 + cells among the Krt5 + or Krt8 + cells. Group sizes: n = 12, 11, 12, 7 mice, from left to right. Two-tailed unpaired Mann-Whitney U test. g Experimental schema. h , i Representative images ( h ) and quantification ( i ) of BCAM and SOX9 staining in tracheal sections from the indicated strains at day 14. Scale bars: 20 μm. Group sizes: n = 5, 8, 7, 5 mice, from left to right. Kruskal–Wallis test followed by Dunn’s multiple-comparisons test with Holm–Bonferroni correction. The dashed line marks the border between SAE and SM. Box plots ( b , e , f , i ) show median (center line), 25th–75th percentiles (box), and min–max values (whiskers). Each data point represents the average of 3–4 images from one mouse. See “Methods”. Panel g was created in BioRender. Lee, M. ( https://BioRender.com/dg3i7rm ). Source data are provided as a Source Data file.

Article Snippet: Female (12 weeks- to 14 weeks) and male (10 weeks- to 12 weeks) mice of the following genotypes and strains were used: C57BL/6 (Charles River Laboratories, CRL# 027), Oxgr1 -/- (KOMP, stock number 048933-UCD), Pou2f3 -/- (The Jackson Laboratory, stock number 037040), Cysltr1 -/- (kindly provided by Dr. Lora Bankova, BWH, Boston), Cysltr2 -/- (kindly provided by Dr. Joshua Boyce, BWH, Boston), Krt5 CreERT2 (The Jackson Laboratory, stock number 029155), Pou2f3 CreERT2 (The Jackson Laboratory, stock number 037511), Sox9 CreERT2 (The Jackson Laboratory, stock number 035092), Sox9 fl/fl (The Jackson Laboratory, stock number 013106), Oxgr1 fl/fl (Barrett Lab), Ltc4s fl/fl (kindly provided by Dr. Lora Bankova, BWH, Boston), and Ai9 (RCL-tdT) (The Jackson Laboratory, stock number 007909).

Techniques: Staining, Two Tailed Test, MANN-WHITNEY, In Situ Hybridization, Immunofluorescence

Journal: Regenerative Therapy

Article Title: Airway basal stem cell derived extracellular vesicles promote lung repair in chronic obstructive pulmonary disease

doi: 10.1016/j.reth.2026.101068

Figure Lengend Snippet: Lineage tracing of BC–mediated repair in the tracheal epithelium after LPS&PPE injury. (a) Schematic of the Krt5 Cre ERT2 -Gt (ROSA)26 Sor tm4(ACTB-tdTomato-EGFP) reporter system used for lineage tracing of airway basal stem cells. (b) Experimental timeline illustrating tamoxifen induction, LPS&PPE-induced emphysema injury, and subsequent analysis at designated time points. (c) Representative immunofluorescence images of tracheal sections at different time points before and after injury. GFP (green) marks lineage-labeled basal cells and their progeny. Co-staining with Krt5 (red) and p63 (red) identifies basal cells, with CC10 (red) and Foxj1 (red) marking Club and ciliated cells, respectively. Scale bar, 20 μm. (d) Quantification of lineage-labeled cells categorized as basal, Club, or ciliated cells at the indicated time points before and after injury. Data are presented as mean ± SEM (n = 1–2 technical replicates from 3 mice). (e) Representative immunofluorescence images of lung tissue sections showing GFP + (green) lineage-labeled cells and Krt5 (red) basal cells distributed along proximal–distal airways at different time points before and after injury. Scale bar, 100 μm.

Article Snippet: Krt5 Cre ERT2 -Gt (ROSA)26Sor tm4(ACTB-tdTomato-EGFP) mice (The Jackson Laboratory, USA) were used to perform lineage tracing.

Techniques: Immunofluorescence, Labeling, Staining

Journal: Regenerative Therapy

Article Title: Airway basal stem cell derived extracellular vesicles promote lung repair in chronic obstructive pulmonary disease

doi: 10.1016/j.reth.2026.101068

Figure Lengend Snippet: Intratracheal transplantation of BCs promotes alveolar repair in emphysematous mice. (a) Representative bright-field image of clonogenic BCs cultured on feeder layers. Scale bar, 100 μm. (b) Immunofluorescence staining of clonogenic BCs using Krt5 (green) and p63 (red) as basal cell markers. Scale bars, 100 μm. (c) Schematic diagram illustrating the experimental design for intratracheal transplantation of BCs into mice pre-treated with BLM&PPE or LPS&PPE. (d) Representative direct fluorescence image of recipient mouse lung 7 days after intratracheal delivery of 10 6 GFP-labeled BCs. (e) Immunofluorescence staining of lung sections showing engraftment and distribution of transplanted GFP + BCs (green), some of which retained Krt5 expression (red). Scale bar, 200 μm. (f) Representative H&E staining of lung sections from healthy control mice and emphysematous mice treated with PBS or BCs. Emphysema was induced by intratracheal administration of LPS and PPE. Scale bar 50 μm. (g) Quantification of average alveolar count, mean alveolar area, and percentage of lung injury across the three groups. Data are presented as mean ± SEM (n = 3 mice). Statistical analysis was performed using a one-way ANOVA with Tukey's multiple comparisons test. ∗∗ P < 0.01, ∗∗∗ P < 0.001, ∗∗∗∗ P < 0.0001; ns, not significant.

Article Snippet: Krt5 Cre ERT2 -Gt (ROSA)26Sor tm4(ACTB-tdTomato-EGFP) mice (The Jackson Laboratory, USA) were used to perform lineage tracing.

Techniques: Transplantation Assay, Cell Culture, Immunofluorescence, Staining, Fluorescence, Labeling, Expressing, Control