Journal: Science advances

Article Title: Hedgehog-interacting protein orchestrates alveologenesis and protects against bronchopulmonary dysplasia and emphysema.

doi: 10.1126/sciadv.adu2958

Figure Lengend Snippet: Fig. 2. Hhip deletion induces the persistence of myofibroblasts. (A) IF analysis of SMA in the alveoli of Hhip-deleted (HHIP CKO) and control mice at P14. (B) Number of myofibroblasts per unit alveolar area of Hhip-deleted and control mice. (C and D) UMAP showing cell clusters in the lung fibroblasts of Hhip-deleted and control mice at P14. (E) Violin plots showing the expression of Pdgfra, Acta2, Hhip, and Cdh4 in alveolar myofibroblasts (ALMF), ductal myofibroblasts (DMF), peribronchial fibroblasts (Perib), adventitial fibroblasts (Adv), and alveolar fibroblasts (Alv). (F) Expression of Acta2, Myh11, Tagln, and Igf1 in ALMFs and DMFs of Hhip-deleted and control mice. (G) qPCR analysis of Igf1 expression in the lung stromal cells isolated from Hhip-deleted and control mice. (H) Analysis of Igf1 (RNA in situ) and SMA expression in the alveoli. (I) Number of SMA+ Igf1+ cells per unit alveolar area of Hhip-deleted and control mice at P14. (J) Percentage of SMA+ Igf1+ cells in total Igf1+ cells. (K) Top 10 activated pathways in Hhip-deleted, relative to control myofibroblasts, analyzed with IPA. (L) qPCR analysis of Gli1, Igf1, and Acta2 expression in the lung stromal cells treated with PBS, SHH, and SHH plus HHIP. (M) qPCR analysis of Acta2 expression in SHH-stimulated lung stromal cells treated with vehicle or IGF1R inhibitor. (N) IF analysis of SMA expression in the alveoli of Hhip-deleted mice administered with vehicle or IGF1R inhibitor. (O) Number of myofibroblasts per unit alveolar area of Hhip-deleted mice administered with vehicle or IGF1R inhibitor. All in vitro experiments have been repeated at least one time with consistent results for validation. Each data point represents one mouse [(B), (G), (I), (J), and (O)] of an individual experiment. Data are expressed as mean ± SD. *P < 0.05, **P < 0.005, ***P < 0.0005, and ****P < 0.0001.

Article Snippet: The following primary antibodies were used for mouse tissue: goat anti- HHIP (catalog no. AF1568; R&D Systems; used 1: 250), rat anti- CDH4 (catalog no. MRCD5; DSHB; used 1: 20), rabbit anti- CDH4 (catalog no. BS40099; Bioworld; used 1: 100), rabbit anti- SMA (catalog no. ab5694; Abcam; used 1:400), mouse anti- SMA (catalog no. AB7817; Abcam; used 1:500), goat anti- tdTomato (catalog no. AB8181- 200; OriGene; used 1:500), rabbit anti- RFP (catalog no. 600- 401- 379; Rockland; used 1:500), rabbit anti- SPC (catalog no. AB3786; Millipore Sigma; used 1:2000), rat anti- Ki67 (catalog no. 14- 5698- 82; Thermo Fisher Scientific; used 1:100), rabbit anti- γH2A (catalog no. 9718T; Cell Signaling Technology; used at 1:250), rat anti- p21 (catalog no. ab107099; Abcam; used 1:100), rat anti– DC- LAMP (catalog no. DDX0191P- 100; Novus; used 1:250), goat anti- PDGFRα (catalog no. AF1062; R&D Systems; used 1:200), and rabbit anti- PDGFRα (catalog no. 3174; Cell Signaling Technology; used 1:200).

Techniques: Control, Expressing, Isolation, In Situ, In Vitro, Biomarker Discovery

Journal: Science advances

Article Title: Hedgehog-interacting protein orchestrates alveologenesis and protects against bronchopulmonary dysplasia and emphysema.

doi: 10.1126/sciadv.adu2958

Figure Lengend Snippet: Fig. 3. Deletion of Hhip induces cell senescence. (A) Violin plots showing the expression of Cdkn1a in ALMFs and DMFs of Hhip-deleted and control mice. (B) Senescence β-galactosidase staining of Hhip-deleted and control lungs. (C) IF analysis of SMA, p21, and CDH4 in Hhip-deleted and control lungs. Arrow: p21+ DMFs; arrowhead: p21+ ALMFs. (D and E) Number of p21+ ALMFs (D) and DMFs (E) per unit alveolar area of Hhip-deleted and control mice at P14. (F) IF analysis of SPC and p21 in Hhip-deleted and control lungs. Arrow: p21+ SPC+ cells. (G) Percentage of p21+ cells in AT2s. (H) Activation of senescence pathways in the AT2s of Hhip-deleted mice, relative to control AT2s, and analyzed with IPA. (I) Top 5 upstream regulators in the AT2s of Hhip-deleted mice, relative to control AT2s, analyzed with IPA. (J) AT2 organoids cocultured with lung stromal cells (R26RSmoM2/+) pre-infected with adenovirus-empty and adenovirus-Cre, treated with anti-IGF1 antibody and IgG. (K and L) Quantification of colony- forming efficiency (CFE) and organoid size. (M) IF analysis of SPC and p21 in AT2 organoids. Arrow: p21+ SPC+ cells. (N) Percentage of p21+ cells in AT2s in the organoid assay. All in vitro experiments have been repeated at least one time with consistent results for validation. Each data point represents one mouse [(D), (E), and (G)] of an individual experiment. Data are expressed as Mean ± SD. *P < 0.05, **P < 0.005, ***P < 0.0005, and ****P < 0.0001.

Article Snippet: The following primary antibodies were used for mouse tissue: goat anti- HHIP (catalog no. AF1568; R&D Systems; used 1: 250), rat anti- CDH4 (catalog no. MRCD5; DSHB; used 1: 20), rabbit anti- CDH4 (catalog no. BS40099; Bioworld; used 1: 100), rabbit anti- SMA (catalog no. ab5694; Abcam; used 1:400), mouse anti- SMA (catalog no. AB7817; Abcam; used 1:500), goat anti- tdTomato (catalog no. AB8181- 200; OriGene; used 1:500), rabbit anti- RFP (catalog no. 600- 401- 379; Rockland; used 1:500), rabbit anti- SPC (catalog no. AB3786; Millipore Sigma; used 1:2000), rat anti- Ki67 (catalog no. 14- 5698- 82; Thermo Fisher Scientific; used 1:100), rabbit anti- γH2A (catalog no. 9718T; Cell Signaling Technology; used at 1:250), rat anti- p21 (catalog no. ab107099; Abcam; used 1:100), rat anti– DC- LAMP (catalog no. DDX0191P- 100; Novus; used 1:250), goat anti- PDGFRα (catalog no. AF1062; R&D Systems; used 1:200), and rabbit anti- PDGFRα (catalog no. 3174; Cell Signaling Technology; used 1:200).

Techniques: Expressing, Control, Staining, Activation Assay, Infection, In Vitro, Biomarker Discovery

Journal: Molecular Cancer

Article Title: Activation of the hedgehog pathway in advanced prostate cancer

doi: 10.1186/1476-4598-3-29

Figure Lengend Snippet: Detection of HIP in human cancer specimens. By Western blotting, HIP antibodies (R&D systems Cat# AF1568) recognized one band between 75 and100 KD ( A ). Expression of endogenous HIP was detected in two GI cancer tissues, which were known to contain activated hedgehog signaling (data not shown here), but not in the matched normal tissue ( B ). Immunohistostaining of HIP I prostate cancer showed a similar pattern to PSA ( C , 200×)

Article Snippet: A standard avidin-biotin immunostaining technique was performed using a kit from Vector laboratories using specific antibodies to Su(Fu) (Santa Cruz Biotechnology Cat# 10933), PTCH1 (Santa Cruz Biotechnology Cat# 6149), HIP (R&D systems Cat# AF1568) and Shh (Santa Cruz Biotechnology Cat# 9024) and PSA (Vector laboratories).

Techniques: Western Blot, Expressing

Journal: bioRxiv

Article Title: Human alveolar Type 2 epithelium transdifferentiates into metaplastic KRT5+ basal cells during alveolar repair

doi: 10.1101/2020.06.06.136713

Figure Lengend Snippet: (A) IPA upstream regulator analysis of hAEC2s co-cultured with AHLM. (B) Bulk RNA-seq analysis of MRC5 and AHLM with differential gene expression analysis. (C) Treatment of hAEC2s+AHLM organoids with BMP4 increases SFTPC and decreases KRT5 mRNA expression at D14 of culture. (D) BMP treatment increases SFTPC+ cells and decreases KRT5+ cells in D14 organoids. (E) hAEC2s from IPF lungs demonstrates decreased SFTPC and HHIP expression when compared to hAEC2s from normal donors. (F) Treatment of hAEC2s+AHLM organoids with HHIP increases SFTPC and decreases KRT5 mRNA expression at D14 of culture. (G) HHIP increases SFTPC+ cells and decreases KRT5+ cells in D14 organoids. (H) HHIP-treated AHLM isolated from D14 organoid culture have decreased GLI1 and increased BMP3 and BMP4 mRNA levels. (I) HHIP-treated hAEC2s+AHLM organoids at D14 show significantly higher fraction of pSMAD1/5/8+ cells. See also

Article Snippet: Where applicable, recombinant BMP4 (Cat#314-BP-010; R&D Systems; used 50 ng/mL) and HHIP (Cat#9280-HP-050; R&D Systems; used 5 μg/mL) were added to the media after 48 hours and replenished in every media change.

Techniques: Cell Culture, RNA Sequencing, Gene Expression, Expressing, Isolation

Journal: Oncogenesis

Article Title: Teriflunomide modulates the PD-1/PD-L1 axis and enhances antitumor immunity in colorectal cancer

doi: 10.1038/s41389-026-00607-3

Figure Lengend Snippet: A The viability of hPD-1 Jurkat-T cells and hPD-L1 CHO cells following treatment with the indicated concentrations of TER for 24 h. B Luciferase activity measured using a PD-1/PD-L1 blockade bioassay. hPD-1 Jurkat-T cells (effector cells) were co-cultured with hPD-L1-expressing aAPC/CHO-K1 cells (target cells) in the presence of indicated concentrations of TER. The luminescence signal indicates the level of TCR signaling activation. αPD-L1 was used as a positive control. * <0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 compared with the respective control.

Article Snippet: Recombinant hPD-L1 (#71104, BPS Bioscience) was coated onto 96-well plates (Corning Inc., New York, NY, USA) at a concentration of 1 mg/mL in PBS and incubated overnight.

Techniques: Luciferase, Activity Assay, Bioassay, Cell Culture, Expressing, Activation Assay, Positive Control, Control

Journal: Oncogenesis

Article Title: Teriflunomide modulates the PD-1/PD-L1 axis and enhances antitumor immunity in colorectal cancer

doi: 10.1038/s41389-026-00607-3

Figure Lengend Snippet: A The viability of hPD-L1 MC38 cells following treatment with the indicated concentrations of TER for 72 h. B CD8 + T cells were isolated from tumors of hPD-1 knock-in mice bearing hPD-L1 MC38 tumors. These tumor-infiltrating CD8 + T cells were co-cultured with hPD-L1 MC38 cells as target cells in the presence of TER for 72 h. Cell viability measured using the CCK assay is depicted. C PD-L1 expression in hPD-L1 MC38 cells, as assessed by western blot analysis using protein lysates from co-culture conditions. GAPDH was used as a loading control. D The levels of immune-related factors, including GrB, IL-2, and IFN-γ, measured in the co-culture supernatant by ELISA. * <0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 compared with the respective control.

Article Snippet: Recombinant hPD-L1 (#71104, BPS Bioscience) was coated onto 96-well plates (Corning Inc., New York, NY, USA) at a concentration of 1 mg/mL in PBS and incubated overnight.

Techniques: Isolation, Knock-In, Cell Culture, Expressing, Western Blot, Co-Culture Assay, Control, Enzyme-linked Immunosorbent Assay

Journal: Oncogenesis

Article Title: Teriflunomide modulates the PD-1/PD-L1 axis and enhances antitumor immunity in colorectal cancer

doi: 10.1038/s41389-026-00607-3

Figure Lengend Snippet: A Body weight of hPD-1 knock-in mice during the treatment period. The mice were treated with vehicle or TER (10 or 30 mpk) for the indicated time. B Spleen weight of mice at the endpoint of the experiment. C Tumor volume was measured over time in hPD-1 knock-in mice bearing hPD-L1 MC38 tumors treated with vehicle or TER (10 or 30 mpk). Representative images of excised tumors from each group are shown. D Tumor weight at the endpoint of the experiment. E Flow cytometry analysis of CD8 + T-cell populations in tumors from each treatment group. F PD-L1 expression in tumors from each group, as assessed by western blot analysis. GAPDH was used as a loading control. G IHC staining of tumor sections for immune-related markers, including CD8 + T cells and GrB. Representative images from each group are shown, and the quantitation of marker-positive cells per field is presented. * <0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 compared with the respective control.

Article Snippet: Recombinant hPD-L1 (#71104, BPS Bioscience) was coated onto 96-well plates (Corning Inc., New York, NY, USA) at a concentration of 1 mg/mL in PBS and incubated overnight.

Techniques: Knock-In, Flow Cytometry, Expressing, Western Blot, Control, Immunohistochemistry, Quantitation Assay, Marker

Journal: Oncogenesis

Article Title: Teriflunomide modulates the PD-1/PD-L1 axis and enhances antitumor immunity in colorectal cancer

doi: 10.1038/s41389-026-00607-3

Figure Lengend Snippet: A Body weight of hPD-1 knock-in mice during the treatment period. The mice were treated with vehicle or TER (30 mpk) and received either an isotype control or a CD8 depletion antibody. B Spleen weight of mice at the endpoint of the experiment. C Tumor volume was measured in hPD-1 knock-in mice bearing hPD-L1 MC38 tumors over time following treatment with vehicle or TER (30 mpk) with or without CD8 depletion. Representative images of excised tumors from each group are shown. D Tumor weight at the endpoint of the experiment. E Flow cytometry analysis confirming CD8 + T-cell depletion in tumors from each treatment group. The proportion of CD8 + cells among total live cells was quantified. F IHC staining of tumor sections for CD8 + T cells and GrB. Representative images from each treatment group are shown, and the quantitation of marker-positive cells per field is presented. * <0.05, ** p < 0.01, and **** p < 0.0001 compared with the respective control.

Article Snippet: Recombinant hPD-L1 (#71104, BPS Bioscience) was coated onto 96-well plates (Corning Inc., New York, NY, USA) at a concentration of 1 mg/mL in PBS and incubated overnight.

Techniques: Knock-In, Control, Flow Cytometry, Immunohistochemistry, Quantitation Assay, Marker

Journal: Oncogenesis

Article Title: Teriflunomide modulates the PD-1/PD-L1 axis and enhances antitumor immunity in colorectal cancer

doi: 10.1038/s41389-026-00607-3

Figure Lengend Snippet: A CRC cell lines were treated with the indicated concentrations of TER for 72 h. Cell viability assessed using the CCK assay is shown. B Human CRC cell lines were co-cultured with hPD-1 Jurkat-T cells and treated with the indicated concentrations of TER for 72 h. C PD-L1 protein expression in CRC cells co-cultured with hPD-1 Jurkat-T cells and treated with TER for 72 h. GAPDH was used as a loading control. The results are shown as the mean ± SEM. * <0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 compared with the respective control.

Article Snippet: Next, 5 μL of 0.5 mg/mL biotinylated hPD-1 (#71109, BPS Bioscience) was added, and the plates were incubated for 2 h at room temperature.

Techniques: Cell Culture, Expressing, Control

Journal: Oncogenesis

Article Title: Teriflunomide modulates the PD-1/PD-L1 axis and enhances antitumor immunity in colorectal cancer

doi: 10.1038/s41389-026-00607-3

Figure Lengend Snippet: A The viability of hPD-1 Jurkat-T cells and hPD-L1 CHO cells following treatment with the indicated concentrations of TER for 24 h. B Luciferase activity measured using a PD-1/PD-L1 blockade bioassay. hPD-1 Jurkat-T cells (effector cells) were co-cultured with hPD-L1-expressing aAPC/CHO-K1 cells (target cells) in the presence of indicated concentrations of TER. The luminescence signal indicates the level of TCR signaling activation. αPD-L1 was used as a positive control. * <0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 compared with the respective control.

Article Snippet: Next, 5 μL of 0.5 mg/mL biotinylated hPD-1 (#71109, BPS Bioscience) was added, and the plates were incubated for 2 h at room temperature.

Techniques: Luciferase, Activity Assay, Bioassay, Cell Culture, Expressing, Activation Assay, Positive Control, Control

Journal: Oncogenesis

Article Title: Teriflunomide modulates the PD-1/PD-L1 axis and enhances antitumor immunity in colorectal cancer

doi: 10.1038/s41389-026-00607-3

Figure Lengend Snippet: A The viability of hPD-L1 MC38 cells following treatment with the indicated concentrations of TER for 72 h. B CD8 + T cells were isolated from tumors of hPD-1 knock-in mice bearing hPD-L1 MC38 tumors. These tumor-infiltrating CD8 + T cells were co-cultured with hPD-L1 MC38 cells as target cells in the presence of TER for 72 h. Cell viability measured using the CCK assay is depicted. C PD-L1 expression in hPD-L1 MC38 cells, as assessed by western blot analysis using protein lysates from co-culture conditions. GAPDH was used as a loading control. D The levels of immune-related factors, including GrB, IL-2, and IFN-γ, measured in the co-culture supernatant by ELISA. * <0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 compared with the respective control.

Article Snippet: Next, 5 μL of 0.5 mg/mL biotinylated hPD-1 (#71109, BPS Bioscience) was added, and the plates were incubated for 2 h at room temperature.

Techniques: Isolation, Knock-In, Cell Culture, Expressing, Western Blot, Co-Culture Assay, Control, Enzyme-linked Immunosorbent Assay

Journal: Oncogenesis

Article Title: Teriflunomide modulates the PD-1/PD-L1 axis and enhances antitumor immunity in colorectal cancer

doi: 10.1038/s41389-026-00607-3

Figure Lengend Snippet: A Body weight of hPD-1 knock-in mice during the treatment period. The mice were treated with vehicle or TER (10 or 30 mpk) for the indicated time. B Spleen weight of mice at the endpoint of the experiment. C Tumor volume was measured over time in hPD-1 knock-in mice bearing hPD-L1 MC38 tumors treated with vehicle or TER (10 or 30 mpk). Representative images of excised tumors from each group are shown. D Tumor weight at the endpoint of the experiment. E Flow cytometry analysis of CD8 + T-cell populations in tumors from each treatment group. F PD-L1 expression in tumors from each group, as assessed by western blot analysis. GAPDH was used as a loading control. G IHC staining of tumor sections for immune-related markers, including CD8 + T cells and GrB. Representative images from each group are shown, and the quantitation of marker-positive cells per field is presented. * <0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 compared with the respective control.

Article Snippet: Next, 5 μL of 0.5 mg/mL biotinylated hPD-1 (#71109, BPS Bioscience) was added, and the plates were incubated for 2 h at room temperature.

Techniques: Knock-In, Flow Cytometry, Expressing, Western Blot, Control, Immunohistochemistry, Quantitation Assay, Marker

Journal: Oncogenesis

Article Title: Teriflunomide modulates the PD-1/PD-L1 axis and enhances antitumor immunity in colorectal cancer

doi: 10.1038/s41389-026-00607-3

Figure Lengend Snippet: A Body weight of hPD-1 knock-in mice during the treatment period. The mice were treated with vehicle or TER (30 mpk) and received either an isotype control or a CD8 depletion antibody. B Spleen weight of mice at the endpoint of the experiment. C Tumor volume was measured in hPD-1 knock-in mice bearing hPD-L1 MC38 tumors over time following treatment with vehicle or TER (30 mpk) with or without CD8 depletion. Representative images of excised tumors from each group are shown. D Tumor weight at the endpoint of the experiment. E Flow cytometry analysis confirming CD8 + T-cell depletion in tumors from each treatment group. The proportion of CD8 + cells among total live cells was quantified. F IHC staining of tumor sections for CD8 + T cells and GrB. Representative images from each treatment group are shown, and the quantitation of marker-positive cells per field is presented. * <0.05, ** p < 0.01, and **** p < 0.0001 compared with the respective control.

Article Snippet: Next, 5 μL of 0.5 mg/mL biotinylated hPD-1 (#71109, BPS Bioscience) was added, and the plates were incubated for 2 h at room temperature.

Techniques: Knock-In, Control, Flow Cytometry, Immunohistochemistry, Quantitation Assay, Marker