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

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: bioRxiv

Article Title: The neonatal Fc receptor and DPP4 are human astrovirus receptors

doi: 10.1101/2024.07.12.603331

Figure Lengend Snippet: ( A ) Overview of the pooled CRISPR screen. The genome-scale Brunello library was introduced into Cas9 expressing Caco2 cells, followed by selection of transduced cells. After 10 days, pooled Caco2 cells were infected with HAstV1 for 24h, followed by staining using anti-HAstV capsid antibody. Uninfected cells were sorted to determine the sgRNA counts by next-generation sequencing. ( B ) FcRn protein levels in Cas9-Caco2 cells disrupted for FCGRT or DPP4 using independent sgRNAs per gene or targeted with a control anti-GFP sgRNA. Two independent replicates are shown. ( C ) Representative histogram for surface expression of DPP4 in naïve Caco2 cells stained with anti-DPP4 antibody or isotype control antibody. ( D ) Representative histogram showing DPP4 expression in fixed, permeabilized Caco2 cells disrupted for FCGRT or DPP4 or B2M using gene specific sgRNAs or targeted with a control anti-GFP sgRNA. ( E ) Percentage of DPP4 expressing Caco2 cells disrupted for DPP4 using sgRNAs or targeted with a control anti-GFP sgRNA. (n=3) ( F ) Percentage of anti-HAstV capsid antibody stained Caco2 cells treated with PBS (n=5) or isotype antibody (n=5) or various concentrations of anti-DPP4 antibody (n=6) prior to HAstV1 infection. ( G ) Percentage of anti-HAstV capsid antibody-stained control (n=6), FCGRT (n=6) or DPP4 (n=6) knockout Caco2 cells transfected with HAstV1 RNA at 48h post-transfection. Results were analyzed using Kruskal-Wallis test with Dunn’s post-test (F and G) from two to three independent experiments. *P<0.05; **P<0.01; ***P<0.001; ****P<0.0001. ns=not significant. Bars indicate mean of all data points.

Article Snippet: To assess the neutralization of HAstV infection by soluble DPP4 and soluble FcRn, 4×10 4 FFU/ml of HAstV1 were incubated with 5μg/ml recombinant human DPP4/CD26 protein (Acro Biosystems; DP4-H5266) or 5μg/ml recombinant FCGRT/B2M complex (BPS Bioscience, 71283-1) in growth media at 37°C.

Techniques: CRISPR, Expressing, Selection, Infection, Staining, Next-Generation Sequencing, Control, Knock-Out, Transfection

Journal: bioRxiv

Article Title: The neonatal Fc receptor and DPP4 are human astrovirus receptors

doi: 10.1101/2024.07.12.603331

Figure Lengend Snippet: ( A ) Enrichment [-log10 Robust Rank Aggregation (RRA)] scores of positively-selected sgRNAs in HAstV1-negative cells sorted 24hpi of a genome-wide CRISPR-Cas9 Caco2 cell library compared to HAstV1-positive cells, calculated by MAGeCK. ( B,C ) Cas9-Caco2 cells disrupted for FCGRT (n=6-9) or B2M (n=9) using two independent sgRNAs per gene or targeted with a control anti-GFP sgRNA (n=9) were infected with HAstV1or HAstV8, then stained with anti-HAstV capsid antibody at 24hpi. ( D ) Mean fluorescent intensity (MFI) of DPP4 in negative and positive cell populations of Caco2 cells infected with HAstV1 (n=6) or HAstV8 (n=6) infected at 24hpi. ( E ) Cas9-Caco2 cells were disrupted for DPP4 (n=9) using two independent sgRNAs per gene or targeted with a control anti-GFP sgRNA (n=9), then assessed for infection using anti-HAstV capsid antibody 24hpi with HAstV1 or HAstV8. (F ) Percentage of anti-HAstV capsid antibody-stained Caco2 cells treated with PBS (n=4-5) or isotype control (n=4-6) or anti-DPP4 polyclonal antibody (n=6-7) for 12h hours prior to infection with HAstV1 or HAstV8. Results from three independent experiments were analyzed using the Kruskal-Wallis test with Dunn’s post-test (B to F). *P<0.05; **P<0.01; ***P<0.001; ****P<0.0001. ns=not significant. Bars indicate mean of all data points.

Article Snippet: To assess the neutralization of HAstV infection by soluble DPP4 and soluble FcRn, 4×10 4 FFU/ml of HAstV1 were incubated with 5μg/ml recombinant human DPP4/CD26 protein (Acro Biosystems; DP4-H5266) or 5μg/ml recombinant FCGRT/B2M complex (BPS Bioscience, 71283-1) in growth media at 37°C.

Techniques: Genome Wide, CRISPR, Control, Infection, Staining

Journal: bioRxiv

Article Title: The neonatal Fc receptor and DPP4 are human astrovirus receptors

doi: 10.1101/2024.07.12.603331

Figure Lengend Snippet: ( A ) Schematic of the CRISPR activation surfaceome screen. The human surfaceome library was introduced into dCas9 expressing Caco2 cells, followed by selection of transduced cells. After 10 days, pooled Caco2 cells were infected with HAstV1 or HAstV8 for 24h, followed by staining using anti-HAstV capsid antibody. We sorted top 3% of the HAstV capsid positive cells to determine the sgRNA counts by next-generation sequencing. ( B ) Representative histogram showing expression of DPP4 in dCas9-Caco2 cells transduced with sgRNAs for DPP4 overexpression. ( C ) FcRn protein levels in dCas9-Caco2 transduced with sgRNA for overexpressing FcRn. Two independent replicates are shown. ( D ) FcRn protein levels in 293T and 293T-FCGRT cells. Two independent replicates are shown. ( E ) Representative histogram showing surface expression of DPP4 in 293T cells stained with anti-DPP4 antibody or isotype control antibody. ( F ) Abundance of DPP4 and HAstV capsid positive 293T and 293T-DPP4 cells infected with HAstV1. ( G, H ) HEK293T or HEK293T-DPP4 cells were transfected with FCGRT (n=5) , DPP4 (n=5) and/or B2M (n=5) plasmids then 48h later were infected and stained with anti-HAstV capsid antibody at 24 hpi. Results were analyzed using Kruskal-Wallis test with Dunn’s post-test (G and H) from three independent experiments. *P<0.05; **P<0.01; ***P<0.001; ****P<0.0001. ns=not significant. Bars indicate mean of all data points.

Article Snippet: To assess the neutralization of HAstV infection by soluble DPP4 and soluble FcRn, 4×10 4 FFU/ml of HAstV1 were incubated with 5μg/ml recombinant human DPP4/CD26 protein (Acro Biosystems; DP4-H5266) or 5μg/ml recombinant FCGRT/B2M complex (BPS Bioscience, 71283-1) in growth media at 37°C.

Techniques: CRISPR, Activation Assay, Expressing, Selection, Infection, Staining, Next-Generation Sequencing, Transduction, Over Expression, Control, Transfection