Journal: Nature Communications

Article Title: Colonic epithelial-derived FGF1 drives intestinal stem cell commitment toward goblet cells to suppress inflammatory bowel disease

doi: 10.1038/s41467-025-58644-2

Figure Lengend Snippet: A Heatmap of differentially expressed Atoh1 and its downstream target genes in PBS and rFGF1-treated UC mice ( n = 4). The mRNA level of Atoh1 in PBS or rFGF1-treated UC mice (upper panel, n = 4) and CD mice (lower panel, n = 5) was detected by qRT-PCR. B , C IF staining of ATOH1 in distal colon sections of Fgf1 fl/fl and VilCre Fgf1 fl/fl mice challenged with DSS ( B ) or TNBS ( C ), and its semi-quantitation of IF intensity ( n = 4, the mean value of 2 fields in each mouse). D , E IF staining of ATOH1 in distal colon sections of DSS ( D ) or TNBS ( E )-induced two IBD mouse models, followed by PBS or rFGF1 treatment and its semi-quantitation of IF intensity ( n = 4, the mean value of 2 fields in each mouse). F Representative images (upper panel) and IF staining of Muc2 (lower panel) in sh Atoh1 or shNC-transfected colonic organoids stimulated with vehicle or rFGF1 ( n = 6). G , H Deletion of Atoh1 in VilCre ERT2 Atoh1 fl/fl mice was confirmed by qRT-PCR ( G ) and IF staining of colon tissues ( H ) ( n = 4). I – M VilCre ERT2 Atoh1 fl/fl mice were injected with tamoxifen for five consecutive days and then given drinking water containing 1.5% DSS to induce acute colitis, followed by vehicle or rFGF1 administration for 7 days. At end of the experiment, the distal colon tissues were harvested and examined. The schematic diagram shows the strategy of tamoxifen injection, DSS challenge and rFGF1 administration ( n = 4). J – L Weight loss ( J ), disease activity index ( K ), colonic length ( L ) were monitored (n = 4). M H&E (upper panel), PAS-AB staining (middle panel) and Muc2 IF staining (lower panel) of distal colon sections ( n = 4). Data was presented as mean ± SEM. ( A , D – E , H ) two-tailed unpaired t -test; ( B , C , F , J – M ) ordinary two-way ANOVA, followed by Sidak; ( G ) Non-parametric statistical method, two-tailed Mann-Whitney test. ns, not significance; nd, not detectable.

Article Snippet: The primary antibodies are listed below: rabbit anti-FGF1 (1:200, 17400, Proteintech), mouse anti-FGF1 (1:100, sc-55520, Santa Cruz Biotechnology), EpCAM (1:800, CST42515, Cell Signaling Technology), Col1α1 (1:100, CST72026, Cell Signaling Technology), ATOH1 (1:400, 21215-1-AP, Proteintech), TCF4 (1:100, ab217668, Abcam), Muc2 (1:400, ab272692, Abcam), Dclk1 (1:300, CST62257, Cell Signaling Technology), ChgA (1:200, sc-393941, Santa Cruz Biotechnology), Fabp1 (1:50, CST13368, Cell Signaling Technology), Ki67 (1:400, ab15580, Abcam), FGFR2 (1:200, CST23328S, Cell Signaling Technology) and Lgr5 (1:100, UM870104, ORIGENE).

Techniques: Quantitative RT-PCR, Staining, Quantitation Assay, Transfection, Injection, Activity Assay, Two Tailed Test, MANN-WHITNEY

Journal: Nature Communications

Article Title: Colonic epithelial-derived FGF1 drives intestinal stem cell commitment toward goblet cells to suppress inflammatory bowel disease

doi: 10.1038/s41467-025-58644-2

Figure Lengend Snippet: A Workflow to find the rFGF1-regulated potential transcription factors of Atoh1 , and heatmap of expression level of potential transcription factors ( n = 4). B The expression levels of potential transcription factors of Atoh1 ( Tcf4 , Nr4a1 , Nfya , Tec , Hoxa13 , Ncaph2 , Kdm1a ) in distal colon sections of PBS or rFGF1-treated UC mice were analyzed by qRT-PCR ( n = 4). C , D IF staining of TCF4 in distal colon sections of Fgf1 fl/fl and VilCre Fgf1 fl/fl mice challenged with DSS ( C ) or TNBS ( D ) and its semi-quantitation of IF intensity ( n = 4, the mean value of 2 fields in each mouse). E , F The expression levels of TCF4 in distal colon sections of DSS-treated UC mice ( E ) and TNBS-treated CD mice ( F ) after PBS or rFGF1 administration and IF staining ( n = 8). G Predicted TCF4 binding sites on the promoter of Atoh1 by the JASPAR database (left panel). Six binding sites with the most significant prediction scores were selected for pairwise deletion mutation analysis (right panel; M1, in purple; M2, in green; M3, in red). H , I Luciferase activity in HT-29 cells transfected with a human Atoh1 promoter reporter plasmid or the predicted TCF4 binding site mutant reporter along with a Tcf4 overexpression plasmid ( n = 4). J ChIP experiments. The data were normalized to the corresponding IgG ( n = 4). K Images of murine colonic organoids transfected small hairpin RNA targeting Tcf4 (sh Tcf4 ) or negative control hairpin (shNT) followed by treated with rFGF1 or vehicle ( n = 6). L IF staining of ATOH1 (upper panel) and Muc2 (lower panel) in colonic organoids in different groups and its semi-quantitation of fluorescent intensity ( n = 6). Data was presented as mean ± SEM. ( B , right panel of E – F , J ) two-tailed unpaired t -test ( Tec and Ncaph2 in penal B using non-parametric statistical method, two-tailed Mann-Whitney Test); ( H – I ) ordinary one-way ANOVA, followed by Dunnett; (right panel of C – D and L , lower panel of K ) ordinary two-way ANOVA, followed by Sidak. ns, no significance.

Article Snippet: The primary antibodies are listed below: rabbit anti-FGF1 (1:200, 17400, Proteintech), mouse anti-FGF1 (1:100, sc-55520, Santa Cruz Biotechnology), EpCAM (1:800, CST42515, Cell Signaling Technology), Col1α1 (1:100, CST72026, Cell Signaling Technology), ATOH1 (1:400, 21215-1-AP, Proteintech), TCF4 (1:100, ab217668, Abcam), Muc2 (1:400, ab272692, Abcam), Dclk1 (1:300, CST62257, Cell Signaling Technology), ChgA (1:200, sc-393941, Santa Cruz Biotechnology), Fabp1 (1:50, CST13368, Cell Signaling Technology), Ki67 (1:400, ab15580, Abcam), FGFR2 (1:200, CST23328S, Cell Signaling Technology) and Lgr5 (1:100, UM870104, ORIGENE).

Techniques: Expressing, Quantitative RT-PCR, Staining, Quantitation Assay, Binding Assay, Mutagenesis, Luciferase, Activity Assay, Transfection, Plasmid Preparation, Over Expression, Negative Control, Two Tailed Test, MANN-WHITNEY

Journal: Nature Communications

Article Title: Colonic epithelial-derived FGF1 drives intestinal stem cell commitment toward goblet cells to suppress inflammatory bowel disease

doi: 10.1038/s41467-025-58644-2

Figure Lengend Snippet: A Single-cell RNA sequencing analysis (GSM4983265) showed that Fgfr2 is highly expressed in stem cells. B Representative image of colon distal stained with Lgr5 (green), FGFR2 (red) and DAPI (blue) from the C57BL/6 J mice ( n = 3). C Deletion of Fgfr2 in Lgr5-EGFP-Cre ERT2 Fgfr2 fl/fl mice treated with vehicle or tamoxifen was confirmed by IF staining ( n = 5). D Murine colonic organoids derived from vehicle or tamoxifen-treated Lgr5-EGFP-Cre ERT2 Fgfr2 fl/fl mice were stimulated with vehicle or rFGF1. Representative images, IF staining of TCF4, ATOH1 and Muc2 in colonic organoids were shown (n = 6). E – I Vehicle or tamoxifen-treated Lgr5-EGFP-Cre ERT2 Fgfr2 fl/fl mice were challenged with drinking water containing 2.5% DSS to induce UC, followed by colon tissues were harvested and examined (n = 4 for TAM + PBS group, n = 5 for other groups). Weight loss ( E ), disease activity index ( F ) and colon length ( G ) of different groups were monitored. H H&E staining of distal colon sections and histology scores (upper panel), PAS-AB staining of distal colon sections and mucin granules-positive cells per crypt (lower panel). I IF staining of TCF4 (upper panel), ATOH1 (middle panel) and Muc2 (lower panel) in distal colon sections, and its semi-quantification of IF intensity. Data was presented as mean ± SEM. (right panel of C ) two-tailed unpaired t -test; (right panel of D – F , right panel of G – I ) ordinary two-way ANOVA, followed by Sidak. ns, no significance.

Article Snippet: The primary antibodies are listed below: rabbit anti-FGF1 (1:200, 17400, Proteintech), mouse anti-FGF1 (1:100, sc-55520, Santa Cruz Biotechnology), EpCAM (1:800, CST42515, Cell Signaling Technology), Col1α1 (1:100, CST72026, Cell Signaling Technology), ATOH1 (1:400, 21215-1-AP, Proteintech), TCF4 (1:100, ab217668, Abcam), Muc2 (1:400, ab272692, Abcam), Dclk1 (1:300, CST62257, Cell Signaling Technology), ChgA (1:200, sc-393941, Santa Cruz Biotechnology), Fabp1 (1:50, CST13368, Cell Signaling Technology), Ki67 (1:400, ab15580, Abcam), FGFR2 (1:200, CST23328S, Cell Signaling Technology) and Lgr5 (1:100, UM870104, ORIGENE).

Techniques: RNA Sequencing, Staining, Derivative Assay, Activity Assay, Two Tailed Test

Journal: Bioengineered

Article Title: Small activating RNA activation of ATOH1 promotes regeneration of human inner ear hair cells.

doi: 10.1080/21655979.2022.2045835

Figure Lengend Snippet: Figure 2. Screening of ATOH1 small activating RNAs in 293 T cells and mesenchymal stem cells. Cells were transfected with the indicated small activating RNAs using RNAiMAX at the indicated concentrations for 72 h. Mock samples were transfected in the absence of duplex RNA. Plasmid DNA was transfected at 1 µg/µl using Lipofectamine 3000. The transfected cells were subjected to RT–qPCR after RNA isolation, an RT reaction to generate cDNA and a Western blotting assay after protein isolation. A and B. Relative mRNA expression levels of the ATOH1 gene in 293 T cells (a) and mesenchymal stem cells (b) 72 h after transfection. C and D. ATOH1 protein levels assessed by Western blotting in 293 T cells (c) and mesenchymal stem cells (d) at 72 h after transfection. E and F. Relative ATOH1 mRNA expression levels in response to different small activating RNA concentrations.

Article Snippet: Induction of hair cell-like cells The induced hair cell progenitor cells were maintained in 6-well plates (2 × 105 cells per well) and in a mixed medium consisting of serum-free DMEM/F12 and N2/B27 and were transfected with ATOH1 small activating RNAs using RNAiMAX or an ATOH1 plasmid (ORIGENE, #RC210192) using Lipofectamine 3000 for 10 days.

Techniques: Transfection, Plasmid Preparation, Quantitative RT-PCR, Isolation, Western Blot, Expressing