Journal: Cell reports

Article Title: Inhibition of FGF10-ERK signal activation suppresses intraductal papillary neoplasm of the bile duct and its associated carcinomas.

doi: 10.1016/j.celrep.2021.108772

Figure Lengend Snippet: Figure 5. Fgf10-induced IPNB shows stepwise carcinogenesis (A) Schematic representation of iFGF10: LSL-KrasG12D:Alb-Cre or Pdx1-Cre (iFGF10KA or iFGF10KP, respectively) mice. (B) H&E images of IPNB lesion in iFGF10KA (Kras+) and iFGF10A (Kras) mice following 0.002% Dox administration for 6 weeks (N = 5 each). Insets indicate high magnification of the rectangles.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Mouse Monoclonal Anti-Human Cytokeratin (clones AE1/AE3) DAKO Cat# M3515 LOT: 10066159 Rabbit Polyclonal Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) Cell Signaling Technology Cat# 4370 RRID:AB_2315112 Rabbit polyclonal Anti-alpha smooth muscle Actin [1A4] Abcam Cat# ab7817 Rabbit polyclonal Anti-S100 antibody Abcam Cat# ab166649 Rabbit polyclonal Anti-FGF10 antibody (for mouse) Santa Cruz Cat# sc-293208 Rabbit polyclonal Anti-FGF10 antibody (for human) Abcam Cat# ab80064 Rabbit polyclonal Anti-FGF10 antibody Sigma-Aldrich Cat#ABN44 Mouse monoclonal Anti-MUC5AC antibody Abcam Cat# ab3649 Mouse monoclonal Anti-MUC6 antibody Leica Biosystems Cat# 130307 Rabbit polyclonal Anti-MUC2 antibody Novus Biologicals Cat# NBP1-31231 Rabbit monoclonal Anti-MUC1 antibody Invitrogen Cat# MA5-35250 Rabbit polyclonal Anti-Phospho-FGFR Pan (Tyr653, Tyr654) antibody Invitrogen Cat# PA5-105938 Rabbit polyclonal Anti-Src (phospho Y418) antibody Abcam Cat# ab4816 Rabbit Polyclonal RFP Antibody Preadsorbed Rockland Immunochemicals Cat# 600-401-379 Goat Anti-Mouse IgG H&L (Alexa Fluor 488) preadsorbed Abcam Cat# ab150117 Goat Anti-Rabbit IgG H&L (Alexa Fluor 594) preadsorbed Abcam Cat# ab150084 Rabbit IgG (Control Antibody) Vector Laboratories Cat# I-1000 Mouse IgG (Control Antibody) Vector Laboratories Cat# I-2000 Bacterial and virus strains Escherichia coli strain SW105 Frederick National Laboratory Cancer Research, NCI NA Biological samples Human biliary tumor samples This paper NA Chemicals, peptides, and recombinant proteins Tamoxifen Sigma Cat# T5648 Recommbinant mouse FGF10 R&D Systems Cat# 6224-FG Recommbinant human FGF10 R&D Systems Cat# 345-FG-025 thioacetamide Sigma Cat# 163678 Doxycycline hyclate Sigma Cat# D9891 Polyinosinic-Polycytidylic acid Sigma Cat# P1530 - Cellstain - DAPI solution DOJINDO Cat# D523 PD0325901 Selleck Chemicals Cat# S1036 L-arabinose Sigma Cat# A3256 Sucrose FUJIFILM Cat# 192-00017 dimethyl sulfoxide FUJIFILM Cat# 043-07211 (Continued on next page) Cell Reports 34, 108772, February 23, 2021 e1

Techniques:

Journal: Cell reports

Article Title: Inhibition of FGF10-ERK signal activation suppresses intraductal papillary neoplasm of the bile duct and its associated carcinomas.

doi: 10.1016/j.celrep.2021.108772

Figure Lengend Snippet: Figure 6. Fgf10 is required for maintaining papillary structure defined as IPNB (A) Experimental design of the withdrawal of Dox and/or TAA administration to iFGF10 mice: (1) Dox-only (Dox+) administration and (2) TAA+Dox+ administration followed by the withdrawal (OFF) (N = 5 each). ‘‘IPNB establishment time’’ and ‘‘endpoint time’’ are indicated.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Mouse Monoclonal Anti-Human Cytokeratin (clones AE1/AE3) DAKO Cat# M3515 LOT: 10066159 Rabbit Polyclonal Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) Cell Signaling Technology Cat# 4370 RRID:AB_2315112 Rabbit polyclonal Anti-alpha smooth muscle Actin [1A4] Abcam Cat# ab7817 Rabbit polyclonal Anti-S100 antibody Abcam Cat# ab166649 Rabbit polyclonal Anti-FGF10 antibody (for mouse) Santa Cruz Cat# sc-293208 Rabbit polyclonal Anti-FGF10 antibody (for human) Abcam Cat# ab80064 Rabbit polyclonal Anti-FGF10 antibody Sigma-Aldrich Cat#ABN44 Mouse monoclonal Anti-MUC5AC antibody Abcam Cat# ab3649 Mouse monoclonal Anti-MUC6 antibody Leica Biosystems Cat# 130307 Rabbit polyclonal Anti-MUC2 antibody Novus Biologicals Cat# NBP1-31231 Rabbit monoclonal Anti-MUC1 antibody Invitrogen Cat# MA5-35250 Rabbit polyclonal Anti-Phospho-FGFR Pan (Tyr653, Tyr654) antibody Invitrogen Cat# PA5-105938 Rabbit polyclonal Anti-Src (phospho Y418) antibody Abcam Cat# ab4816 Rabbit Polyclonal RFP Antibody Preadsorbed Rockland Immunochemicals Cat# 600-401-379 Goat Anti-Mouse IgG H&L (Alexa Fluor 488) preadsorbed Abcam Cat# ab150117 Goat Anti-Rabbit IgG H&L (Alexa Fluor 594) preadsorbed Abcam Cat# ab150084 Rabbit IgG (Control Antibody) Vector Laboratories Cat# I-1000 Mouse IgG (Control Antibody) Vector Laboratories Cat# I-2000 Bacterial and virus strains Escherichia coli strain SW105 Frederick National Laboratory Cancer Research, NCI NA Biological samples Human biliary tumor samples This paper NA Chemicals, peptides, and recombinant proteins Tamoxifen Sigma Cat# T5648 Recommbinant mouse FGF10 R&D Systems Cat# 6224-FG Recommbinant human FGF10 R&D Systems Cat# 345-FG-025 thioacetamide Sigma Cat# 163678 Doxycycline hyclate Sigma Cat# D9891 Polyinosinic-Polycytidylic acid Sigma Cat# P1530 - Cellstain - DAPI solution DOJINDO Cat# D523 PD0325901 Selleck Chemicals Cat# S1036 L-arabinose Sigma Cat# A3256 Sucrose FUJIFILM Cat# 192-00017 dimethyl sulfoxide FUJIFILM Cat# 043-07211 (Continued on next page) Cell Reports 34, 108772, February 23, 2021 e1

Techniques:

Journal: Cell reports

Article Title: Niche-mediated repair of airways is directed in an occupant-dependent manner

doi: 10.1016/j.celrep.2022.111863

Figure Lengend Snippet: (A) Schematic representation of experimental design: mice were placed on tamoxifen chow for 3 weeks starting at 8 weeks of age. After a 3 week washout period, mice were intraperitoneally (i.p.) injected with NPT at 20 weeks of age, and lungs were isolated at 3, 7, or 21 days after injury. (B) Immunostaining for club and basal cell markers Scgb1a1 and Krt5, respectively, on control, Gli1 CreERT2 ; Fgf10 f/f , Acta2 CreERT2 ; Fgf10 f/f , Lgr6 CreERT2 ; Fgf10 f/f , and Sox2 CreERT2 ; Fgfr2b f/f mice 21 days after naphthalene injury. Higher-magnification panels on the right of proximal (P) versus distal (D) parts of the airway. (C) Schematic representation of data presented in (B). (D–F) Quantification of the percentage of the airway epithelium covered by club or basal cells in control, Gli1 CreERT2 ; Fgf10 f/f , Acta2 CreERT2 ; Fgf10 f/f , Lgr6 CreERT2 ; Fgf10 f/f , and Sox2 CreERT2 ; Fgfr2b f/f mice 3, 7, and 21 days after naphthalene injury. (G–K) Relative mRNA expression of Fgf10 , Scgb1a1 , Krt5 , Tp63 , and Muc5b in control, Gli1 CreERT2 ; Fgf10 f/f , Acta2 CreERT2 ; Fgf10 f/f , Lgr6 CreERT2 ; Fgf10 f/f , and Sox2 CreERT2 ; Fgfr2b f/f mice 3, 7, and 21 days after naphthalene injury. **p < 0.01; *p < 0.05. n ≥ 6.; error bars mean ± SEM. Scale bars, 500 μm. Two-sided t test and ANOVA used to determine statistical significance.

Article Snippet: Rabbit anti-FGF10 , Acris Antibodies , Cat# AP14882PU-N; RRID: AB_1752406.

Techniques: Injection, Isolation, Immunostaining, Control, Expressing

Journal: Cell reports

Article Title: Niche-mediated repair of airways is directed in an occupant-dependent manner

doi: 10.1016/j.celrep.2022.111863

Figure Lengend Snippet: (A and B) Immunostaining for club and ciliated cell markers Scgb1a1 and Foxj1, respectively, on control lungs without injury or at 3 days post naphthalene injury. (C) Quantification of the number of ciliated cells per 1 mm of proximal or distal airway in non-injured lungs versus lungs 3 days after naphthalene injury. (E) Immunostaining for GFP and club cell marker Scgb1a1 on Scgb1a1 CreERT ; mTmG , Piezo2 Cre ; mTmG and Krt5 CreERT2 ; mTmG mice in which we lineage traced club cells, neuroendocrine cells, and basal cells, respectively, without injury or up to 21 days after naphthalene injury. (F and G) Quantification of the percentage of club cells that are GFP labeled in each of the lineage-tracing models represented in (C) in lung and trachea. (H) Immunostaining for Muc5b on control Gli1 CreERT2 ; Fgf10 f/f , Acta2 CreERT2 ; Fgf10 f/f , Lgr6 CreERT2 ; Fgf10 f/f , and Sox2 CreERT2 ; Fgfr2b f/f mice. **p < 0.01; *p < 0.05. n ≥ 6.; error bars mean ± SEM. Scale bars, 500 μm. Two-sided t test and ANOVA used to determine statistical significance.

Article Snippet: Rabbit anti-FGF10 , Acris Antibodies , Cat# AP14882PU-N; RRID: AB_1752406.

Techniques: Immunostaining, Control, Marker, Labeling

Journal: Cell reports

Article Title: Niche-mediated repair of airways is directed in an occupant-dependent manner

doi: 10.1016/j.celrep.2022.111863

Figure Lengend Snippet: (A and J) Immunostaining for club and basal cell markers Scgb1a1 and Krt5, respectively, on control, Gli1 CreERT2 ; Smo f/f , Gli1 CreERT2 ; Smo f/f , Fgf10 f/f , Lgr6 CreER2 ; Ctnnb1 f/f , Sox2 CreERT2 ; Shh f/f , Sox2 CreERT2 ; Shh f/f ; Fgfr2b f/f , and Sox2 CreERT2 ; Wnt7b f/f mice 21 days after naphthalene injury. (B, C, G–I, K, L, and P–R) Relative mRNA expression of Fgf10 , Scgb1a1 , Krt5 , Tp63 , and Muc5b in control, Gli1 CreERT2 ; Smo f/f , Gli1 CreERT2 ; Smo f/f , Fgf10 f/f , Lgr6 CreERT2 ; Ctnnb1 f/f , Sox2 CreERT2 ; Shh f/f , Sox2 CreERT2 ; Shh f/f ; Fgfr2b f/f , and Sox2 CreERT2 ; Wnt7b f/f mice 3, 7 and 21 days after naphthalene injury. (D–F and M–O) (D–F) Quantification of the percent of the airway epithelium covered by club or basal cells in control, Gli1 CreERT2 ; Smo f/f , Gli1 CreERT2 ; Smo f/f , Fgf10 f/f , Lgr6 CreERT2 ; Ctnnb1 f/f , Sox2 CreERT2 ; Shh f/f , Sox2 CreERT2 ; Shh f/f ; Fgfr2b f/f , and Sox2 CreERT2 ; Wnt7b f/f mice. (S) Schematic representation of data presented in (A) and (J). **p < 0.01; *p < 0.05. n ≥ 6.; error bars mean ± SEM. Scale bars, 500 μm. Two-sided t test and ANOVA used to determine statistical significance.

Article Snippet: Rabbit anti-FGF10 , Acris Antibodies , Cat# AP14882PU-N; RRID: AB_1752406.

Techniques: Immunostaining, Control, Expressing

Journal: Cell reports

Article Title: Niche-mediated repair of airways is directed in an occupant-dependent manner

doi: 10.1016/j.celrep.2022.111863

Figure Lengend Snippet: (A–I) Relative mRNA expression of Wnt7b , Bmp4 , or Shh in control, Gli1 CreERT2 ; Smo f/f , Gli1 CreERT2 ; Smo f/f , Fgf10 f/f , Gli1 CreERT2 ; Fgf10 f/f , Acta2 CreERT2 ; Fgf10 f/f , Lgr6 CreERT2 ; Fgf10 f/f , Lgr6 CreERT2 ; Ctnnb1 f/f , Sox2 CreERT2 ; Shh f/f , Sox2 CreERT2 ; Shh f/f ; Fgfr2b f/f , Sox2 CreERT2 ; Fgfr2b f/f , Sox2 CreERT2 ; Wnt7b f/f , Scgb1a1 Cre ; Shh f/f , Scgb1a1 Cre ; Shh f/f ; Fgfr2b f/f , and Scgb1a1 Cre ; Smo f/f mice 3, 7, and 21 days after naphthalene injury. **p < 0.01; *p < 0.05. n ≥ 6.; error bars mean ± SEM. Two-sided t test and ANOVA used to determine statistical significance.

Article Snippet: Rabbit anti-FGF10 , Acris Antibodies , Cat# AP14882PU-N; RRID: AB_1752406.

Techniques: Expressing, Control

Journal: Cell reports

Article Title: Niche-mediated repair of airways is directed in an occupant-dependent manner

doi: 10.1016/j.celrep.2022.111863

Figure Lengend Snippet: (A) Immunostaining of human submucosal glands for Fgf10, Fgfr2b, and basal cell marker Krt5 (note the Fgf10 antibody detects Fgf10 bound to receptor). (B) Immunostaining for GFP and myoepithelial cell markers Krt5 and Acta2 on Nkx2.1 Flpo ; Acta2-Frt-STOP-Frt-Cre ERT2 ; Rosa26 mTmG mice. (C) Immunostaining for GFP and myoepithelial cell markers Krt5 and Acta2 on submucosal glands from Nkx2.1 Flpo ; Acta2-Frt-STOP-Frt-Cre ERT2 ; Rosa26 mTmG , Nkx2.1 Flpo ; Acta2-Frt-STOP-Frt-Cre ERT2 ; Rosa26 mTmG ; Fgfr2b f/f , and Nkx2.1 Flpo ; Acta2-Frt-STOP-Frt-Cre ERT2 ;Rosa26 mTmG ; Fgf10 f/f mice that were injured twice with naphthalene with a 21 day interval and were harvested 60 days after the first or 39 days after the second injury. Higher-magnification panels are found to the right. (D) Schematic representation of experimental design: mice were placed on tamoxifen chow for 3 weeks starting at 8 weeks of age. After a 3 week washout period, mice were i.p. injected with NPT at 14 weeks of age and again 3 weeks later at 17 weeks of age, and lungs were isolated 3 weeks after the second injury at 20 weeks of age. (E) Top: quantification of the percentage of basal cells that are GFP positive and therefore myoepithelial cell derived in the surface airway epithelium of the tracheas and lungs from Nkx2.1 Flpo ; Acta2-Frt-STOP-Frt-Cre ERT2 ; Rosa26 mTmG , Nkx2.1 Flpo ; Acta2-Frt-STOP-Frt-Cre ERT2 ; Rosa26 mTmG ; Fgfr2b f/f mice that were injured twice with naphthalene with a 21 day interval and were harvested 60 days after the first or 39 days after the second injury. Bottom: quantification of the percentage of the airway epithelium that is covered by basal cells in Nkx2.1 Flpo ; Acta2-Frt-STOP-Frt-Cre ERT2 ; Rosa26 mTmG , Nkx2.1 Flpo ; Acta2-Frt-STOP-Frt-Cre ERT2 ; Rosa26 mTmG ;Fgfr2b f/f mice that were injured twice with naphthalene with a 21 day interval and were harvested 60 days after the first or 39 days after the second injury. (F) Immunostaining for GFP and club and basal cell markers Scgb1a1 and Krt5, respectively, on lungs from Nkx2.1 Flpo ; Acta2-Frt-STOP-Frt-Cre ERT2 ; Rosa26 mTmG and Nkx2.1 Flpo ; Acta2-Frt-STOP-Frt-Cre ERT2 ; Rosa26 mTmG ; Fgfr2b f/f mice that were injured twice with naphthalene with a 21 day interval and were harvested 60 days after the first or 39 days after the second injury. **p < 0.01; *p < 0.05. n ≥ 6; error bars mean ± SEM. Scale bars, 200 μm. Two-sided t test and ANOVA used to determine statistical significance.

Article Snippet: Rabbit anti-FGF10 , Acris Antibodies , Cat# AP14882PU-N; RRID: AB_1752406.

Techniques: Immunostaining, Marker, Injection, Isolation, Derivative Assay

Journal: Cell reports

Article Title: Niche-mediated repair of airways is directed in an occupant-dependent manner

doi: 10.1016/j.celrep.2022.111863

Figure Lengend Snippet: Here, we show how spatiotemporal expression of Fgf10 by two niche-maintaining cell types is primarily orchestrated by the niche’s epithelial occupants—both those that reside prior to and following injury. Prior to injury, differentiated airway epithelial cells secrete Shh to inhibit Fgf10 expression by Gli1 + peribronchial mesenchymal cells of the niche. After injury, remaining epithelial cells produce Wnt7b to induce Fgf10 expression in airway smooth muscle cells of the niche. Complete induction of Fgf10 expression in the niche requires loss of an inhibitory Shh signal from the prior epithelial occupant (club cell) as well as induction of an activating Wnt7b signal by the surviving or new epithelial occupant (ciliated and basal cells). We find that this reliance on a common activator of airway epithelial stem cells allows for the recruitment of remote stem cell populations when local populations have been destroyed or exhausted.

Article Snippet: Rabbit anti-FGF10 , Acris Antibodies , Cat# AP14882PU-N; RRID: AB_1752406.

Techniques: Expressing

Journal: Cell reports

Article Title: Niche-mediated repair of airways is directed in an occupant-dependent manner

doi: 10.1016/j.celrep.2022.111863

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit anti-FGF10 , Acris Antibodies , Cat# AP14882PU-N; RRID: AB_1752406.

Techniques: Recombinant, Plasmid Preparation, Expressing, Software