Journal: Scientific Reports

Article Title: Exocrine tissue-driven TFF2 prevents apoptotic cell death of endocrine lineage during pancreas organogenesis

doi: 10.1038/s41598-018-38062-9

Figure Lengend Snippet: Elastase-Cre-mediated Pdx1 inactivation reduces acinar TFF2 in embryonic and neonatal pancreas. ( A ) The expression of TFF2 was detected by RT-PCR in control mice pancreas from E16.5. The original data are shown in Supplementary Fig. . ( B ) Expression of TFF2 is significantly less in Pdx1cKO mice (red) than in control mice (blue). (control mice: n = 7 at E14.5, n = 5 at E16.5, n = 5 at E18.5, and n = 7 at P1; Pdx1cKO mice: n = 5 at E14.5, n = 6 at E16.5, n = 6 at E18.5, and n = 7 at P1; p = N.D at E14.5, p = 0.041 at E16.5, p = 0.0065 at E18.5 and p = 0.0040 at P1). Note that the expression of TFF2 in the mutant stomach is equivalent to that in control stomach at P1 (right panel) (control mice, n = 3, Pdx1cKOmice, n = 3, p = 0.68122). ( C ) Immunostaining of TFF2. TFF2 expression was detected in exocrine cells including the proximal (dotted lines) and distal ducts and acinar cells, but not in islets (arrows) in control mice (upper panels). In Pdx1cKO mice, TFF2 expression was hardly detectable except in the proximal ducts (dotted lines), which were not recombined by Elastase-Cre (bottom panels). These expression patterns were confirmed in at least three individual mice for both genotypes. Scale bars, 100 μm. Bars represent the mean value ± SE. *p < 0.05, **p < 0.01.

Article Snippet: Recombinant mouse TFF2 (#RPA748MU01) was purchased from Uscn Life Science (Texas, USA).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Control, Mutagenesis, Immunostaining

Journal: Scientific Reports

Article Title: Exocrine tissue-driven TFF2 prevents apoptotic cell death of endocrine lineage during pancreas organogenesis

doi: 10.1038/s41598-018-38062-9

Figure Lengend Snippet: TFF2 prevents apoptosis of embryonic insulin-expressing cells through CXCR4. Explant culture of E16.5 pancreatic tissue for 48 hours. ( A ) Immunostaining of insulin (red), beta-catenin (green) and DAPI (blue) for the cultured explant without reagents (left panel), with rTFF2 (middle panel) and with rTFF2 plus AMD3100 (right panel). ( B ) Quantitative analysis of the number of Insulin+ cells. ( C ) Immunostaining of pHH3 (green), insulin (red) and DAPI (blue) for cultured control explant (left panel) and for Pdx1cKO explant without and with rTFF2 (middle and right panels, respectively). The yellow arrowhead shows pHH3-positive Insulin+ cells. ( D ) Quantitative analyses of Insulin+ cell proliferation. ( E ) Immunostaining of TUNEL (green), insulin (red) and DAPI (blue) for cultured Pdx1cKO explant without reagents (left panel), with rTFF2 (middle panel) and with rTFF2 plus AMD3100 (right panel). Yellow arrowheads show TUNEL-positive Insulin+ cells. ( F ) Quantitative analysis of Insulin+ cell apoptosis in Pdx1cKO explant. In control mice pancreas, CXCR4 protected Insulin+ cells from apoptosis. ( G ) Immunostaining of TUNEL (green), insulin (red) and DAPI (blue) for the control explant without and with AMD3100 (left and right panels, respectively). Yellow arrowheads show TUNEL-positive Insulin+ cells. ( H ) Quantitative analyses of Insulin+ cell apoptosis. Scale bars, 100 μm. Bars represent the mean value ± SE. *p < 0.05, **p < 0.01.

Article Snippet: Recombinant mouse TFF2 (#RPA748MU01) was purchased from Uscn Life Science (Texas, USA).

Techniques: Expressing, Immunostaining, Cell Culture, Control, TUNEL Assay

Journal: Scientific Reports

Article Title: Exocrine tissue-driven TFF2 prevents apoptotic cell death of endocrine lineage during pancreas organogenesis

doi: 10.1038/s41598-018-38062-9

Figure Lengend Snippet: Nkx6.1-positive trunk cells are affected by TFF2. Analyses of Nkx6.1+/Insulin- trunk cells in explant culture experiments. ( A ) Immunostaining of Nkx6.1 (green) and insulin (red) for cultured Pdx1cKO explant without reagents (left panel), with rTFF2 (middle panel) and with rTFF2 plus AMD3100 (right panel). ( B ) Quantitative analyses of the number of Nkx6.1+/Insulin- trunk cells. ( C ) TUNEL staining of cultured Pdx1cKO explant without and with rTFF2 (left and right panels, respectively). ( D ) Quantitative analyses of Nkx6.1+/Insulin- trunk cell apoptosis. Scale bars, 100 μm. Bars represent the mean value ± SE. *p < 0.05.

Article Snippet: Recombinant mouse TFF2 (#RPA748MU01) was purchased from Uscn Life Science (Texas, USA).

Techniques: Immunostaining, Cell Culture, TUNEL Assay, Staining

Journal: Mucosal immunology

Article Title: Macrophages promote epithelial proliferation following infectious and non-infectious lung injury through a Trefoil factor 2-dependent mechanism

doi: 10.1038/s41385-018-0096-2

Figure Lengend Snippet: ( a ) Schematic illustration of TFF2-flox/reporter (Tre-Tom) generation. ( b-d ) Confocal microscopy images showing baseline expression of TFF2 in agaroseinflated lung tissues of ( b ) naïve and ( c, d ) N.b. -infected TFF2-TdTomato fluorescent reporter (Tre-Tom) mice on d4 at ( c ) 200× and ( d ) 400×. ( e, f ) Quantitative Tff2 mRNA levels in sorted ( e ) epithelial cells and ( f ) AM from naïve (open bars) or d4 N.b. -infected (grey bars) lungs of CD11c Cre , CD11c Cre TFF2 flox and TFF2 −/− mice. ( g ) Percentages of BrdU + AT2 cells in CD11c Cre vs CD11c Cre TFF2 flox lungs at indicated time-points after N.b. infection. ( h ) Change of blood oxygen levels from baseline at d3 and d9 following N.b. infection in CD11c Cre and CD11c Cre TFF2 flox mice. ( i ) Representative lung pathology of naïve (left) and d9 following N.b. infection (right) in CD11c Cre and CD11c Cre TFF2 flox mice. Arrowheads indicate emphysematous areas. 20× magnification. *p<0.05, **p<0.01 determined by ANOVA or Student’s t-test. Mean ±SEM are shown.

Article Snippet: Mouse TFF2 ELISA kit was purchased from United States Biological (Salem, MA).

Techniques: Confocal Microscopy, Expressing, Infection

Journal: Mucosal immunology

Article Title: Macrophages promote epithelial proliferation following infectious and non-infectious lung injury through a Trefoil factor 2-dependent mechanism

doi: 10.1038/s41385-018-0096-2

Figure Lengend Snippet: ( a ) BAL TFF2 levels from PBS-treated or at d9 or d16 after Bleomycin (BLM) treatment in CD11c Cre and CD11c Cre -TFF2flox mice. ( b ) Weight changes over two weeks after BLM treatment in CD11c Cre (black symbols) and CD11c Cre TFF2flox (grey symbols) mice. ( c ) Total protein levels in BAL from PBS-treated or d9 BLM-treated mice. ( d ) Percentage of BrdU+EpCAM+ cells in the lungs from PBS controls or on d9 after BLM treatment. Each symbol represents individual mouse. ( e , f ) Message RNA levels of ( e ) SpC and ( f ) Cc10 from lungs of PBStreated or d9/d16 BLM-treated CD11c Cre (open bars) or CD11c Cre -TFF2flox (grey bars) mice. Mean ±SEM are shown. *p<0.05 and **p< 0.01 as determined by ANOVA or Student’s t-test. Representative results from three independent experiments were shown. N=3–6/group unless specified otherwise.

Article Snippet: Mouse TFF2 ELISA kit was purchased from United States Biological (Salem, MA).

Techniques:

Journal: Mucosal immunology

Article Title: Macrophages promote epithelial proliferation following infectious and non-infectious lung injury through a Trefoil factor 2-dependent mechanism

doi: 10.1038/s41385-018-0096-2

Figure Lengend Snippet: ( a ) Schematic showing MERA conceptual design. ( b ) Trans-epithelial resistance (TER) values of transwell inserts containing primary tracheal epithelia or bone marrow macrophages (BMMφ) cultured alone. ( c ) TER values following scratch wounding of epithelia in the presence or absence of WT BMMφ. ( d,e ) TER values following epithelial injury in the presence of ( d ) WT vs. IL-4Rα −/− BMMφ and ( e ) WT vs. TFF2 −/− BMMφ. ( f ) Levels of TFF2 in MERA cultures with no BMMφ, WT BMMφ, TFF2 −/− BMMφ or IL-4Rα −/− BMMφ. Symbols represent individual wells. ( g ) Gating strategy to identify live epithelial cells recovered from MERA wells that express junctional adhesion molecule (JAM)-1 (R1). ( h-l ) Representative flow plots showing BrdU incorporation in R1 from “ g ” on d4 postscratch wounding ( h ) in the absence of Mφ or in the presence of ( i ) WT BMMφ, ( j ) TFF2 −/− BMMφ, and ( k ) IL-4Rα −/− BMMφ. ( l ) BrdU incorporation on d4 postscratch wounding following treatment with 0.5 ng/ml rTFF2 in the absence of BMMφ. ( m ) Quantification of BrdU + epithelial cells as identified in “ h ” to “ l ”. N=36/group. ( n ) TER values after scratch wounding of epithelia in the presence of rTFF2 (0.5 or 5 ng/ml). ( o ) TER values following scratch wounding of WT vs. TFF2KO epithelia in the presence of WT or TFF2KO alveolar macrophages. ( p, q ) MERA using AT2 co-cultured without BMM or with WT vs. TFF2 −/− BMM. ( p ) Flow plots and ( q ) quantification of EdU + cells. ( r ) TER values following scratch in wells with WT vs. TFF2 −/− BMM. Data represent 2–4 independent experiments. *, p< 0.05; **, p< 0.01 and ***, p< 0.005 as determined by one-way ANOVA or t-test.

Article Snippet: Mouse TFF2 ELISA kit was purchased from United States Biological (Salem, MA).

Techniques: Cell Culture, BrdU Incorporation Assay

Journal: Mucosal immunology

Article Title: Macrophages promote epithelial proliferation following infectious and non-infectious lung injury through a Trefoil factor 2-dependent mechanism

doi: 10.1038/s41385-018-0096-2

Figure Lengend Snippet: ( a,b ) Volcano plots showing results of a cDNA-based screen for Wnt pathway target genes expressed as the fold-difference between ( a ) WT Mφ exposed to damaged or quiescent epithelia and ( b ) between WT Mφ vs. TFF2 −/− Mφ exposed to damaged epithelia. BMMφ were recovered at d4 of MERA for analysis. Each point represents mean of three biological replicates. ( c ) TER values during MERA with WT Mφ in the presence of anti-Wnt4a mAb or control IgG. ( d, e ) Flow-sorted AM, IM and CD103 + DC from WT or TFF2 KO lungs at d4 after infection were analyzed for mRNA levels of ( d ) Wnt4 and ( e ) Wnt16 . ( f ) Representative flow plots and ( g ) quantification of BrdU + SpC + epithelia from distal lung digest cells pre-gated on the live, CD45 − , EpCAM + population for each designated genotype following i.n. administration of rWnt4/16+ R-Spondin 1 cocktail (1μg/mouse, 2 doses) or saline and analyzed by flow cytometry at d4 following N.b. infection (N=5/group). Graphs show Mean±SEM. *, p<0.05; **, p<0.01 and ***, p< 0.005 as determined by ANOVA or Student’s t-test.

Article Snippet: Mouse TFF2 ELISA kit was purchased from United States Biological (Salem, MA).

Techniques: Control, Infection, Saline, Flow Cytometry