Journal: Frontiers in Pharmacology

Article Title: BAY 60-6583 Enhances the Antitumor Function of Chimeric Antigen Receptor-Modified T Cells Independent of the Adenosine A2b Receptor

doi: 10.3389/fphar.2021.619800

Figure Lengend Snippet: CAR T cell enhancement by BAY 60-6583 in CD133 CAR T cells seems not to be dependent on the adenosine A2b receptor. (A, B) Blocking of the adenosine A2b receptor cannot inhibit the enhanced antitumor activities induced by BAY 60-6583. In the anti-CD133 CAR T co-culture system (E:T ratio of 4:1), the adenosine A2b receptor was blocked by 1 μM PSB603 1 h before 10 μM BAY 60-6583 or 1 μM NECA was added. 24 h later, TNF-α production (A) and tumor lysis (B) were analyzed (C) The expression of the adenosine A2b receptor on conventional (WT) and A2b receptor knockout (4 + 6) anti-CD133 CAR T cells was determined by Western blot. Data from a representative experiment of n = 4 experiments (D, E) BAY 60-6583-induced enhancement of CAR T cell-mediated antitumor activities were not influenced by adenosine A2b receptor deficiency. Conventional and A2b receptor knockout anti-CD133 CAR T cells were co-cultured with U251-CD133OE luc cells at an E:T ratio of 4:1. After treatment with vehicle control or BAY 60-6583 for 24 h, cytokine secretion in the supernatant was detected (D) ; 48 h later, cytotoxicity was assessed (E) . * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 by 1-way (A, B) or 2-way ANOVA (D, E) . Data are represented as the mean ± SD from a representative experiment of n = 3 experiments (A, B, D, E) .

Article Snippet: After blocking with 5% milk for 1 h, the membranes were incubated with anti-adenosine A2b receptor (AAR-003, alomone labs) or anti-GAPDH (sc-32233, Santa Cruz Biotechnology) antibodies at 4°C overnight.

Techniques: Blocking Assay, Co-Culture Assay, Lysis, Expressing, Knock-Out, Western Blot, Cell Culture, Control

Journal: Cell Communication and Signaling : CCS

Article Title: STING-STAT1-ZBP1 axis orchestrates PANoptotic signaling in ischemia-reperfusion induced acute kidney injury

doi: 10.1186/s12964-026-02830-2

Figure Lengend Snippet: STING and IFNAR Pathways Cooperatively Promote STAT1 Activation Drives PANoptosis in Renal Tubular Epithelial Cells . A - B Primary renal tubular epithelial cells (PRTCs) were treated with DMSO (control), diABZI (STING agonist), or diABZI and Anifrolumab (IFNAR inhibitor). Representative Western blot analysis and quantification of PANoptosis markers (pyroptosis: GSDMD, N-GSDMD; apoptosis: CAS3, c-CAS3, CAS7, c-CAS7; necroptosis: p-RIP1, p-MLKL) and ZBP1 in PRTCs (n=3 α-Tubulin loading control). Red separator lines indicate different experimental conditions. C Primary renal tubular epithelial cells (PRTCs) were treated with DMSO, diABZI, or diABZI and Anifrolumab. Flow cytometry quantification of cell death (PI⁺, V610). D - E PRTCs were treated with DMSO, diABZI, or diABZI (10 μM) and IFN-α (1 ng/ml). Western blot analysis and quantification of PANoptosis markers (pyroptosis: GSDMD, N-GSDMD; apoptosis: CAS3, c-CAS3, CAS7, c-CAS7; necroptosis: p-RIP1, p-MLKL) in each group (n=3; α-Tubulin loading control). Red separator lines indicate different experimental conditions. F Proposed mechanism of the STING-STAT1-ZBP1 axis in renal tubular epithelial cells promoting pan-apoptosis in IRI-AKI. Following IRI-AKI, STING interacts with and activates STAT1, facilitating its nuclear translocation. STAT1 binds to the promoter region of the ZBP1 gene, initiating its transcription. The upregulated ZBP1 subsequently triggers the activation of PANoptosis, which drives the progression of AKI

Article Snippet: Primary cells were treated with diABZI (medchemexpress-HY-112921A), Cocl2 (sigma-232696), Fludarabine (medchemexpress-HY-B0069), Anifrolumab (medchemexpress-HY- P99168 ), IFN-α (medchemexpress-HY-P7023), Necrostatin-1 (medchemexpress-HY-15760), Z-VAD-FMK (medchemexpress-HY-16658B), MCC950 (medchemexpress-HY-12815), 2’3’-cGAMP (c-GAMP: tlrl-nacga23-02) for 24–48 h.

Techniques: Activation Assay, Control, Western Blot, Flow Cytometry, Translocation Assay

Journal: Disease Models & Mechanisms

Article Title: Adenosine and hyaluronan promote lung fibrosis and pulmonary hypertension in combined pulmonary fibrosis and emphysema

doi: 10.1242/dmm.038711

Figure Lengend Snippet: Cardiovascular physiology, metabolic profile, and hyaluronan synthesis and deposition in Ada −/− . (A-C) Right ventricular systolic pressure (RVSP; A), left ventricular systolic pressure (LVSP; B) and Fulton index ratio (C) determined by the measurement of right ventricle and left ventricle with septum from Ada + (white bars), Ada −/− (gray bars) and Ada −/− +4MU-treated (black bars) mice. (D-I) Transcript expression levels from lung tissue for Adora2b (D), Pparg (E), Sdha (F), Has1 (G), Has2 (H), Has3 (I) by RT-PCR from Ada + (white bars), Ada −/− (gray bars) and Ada −/− +4MU-treated (black bars) mice. (J) Representative dual-IHC staining in fibrotic areas of Ada −/− or Ada −/− +4MU treatment groups for αSMA (blue) and hyaluronan (magenta). Data are mean+s.e.m. ( n =5 per group). * P <0.05 (comparisons between Ada + and Ada −/− treatment groups), # P <0.05 (comparisons between Ada −/− and Ada −/− +4MU treatment groups); two-way ANOVA with the Benjamin, Krieger and Yekutieli post hoc test. Scale bars: 200 µm.

Article Snippet: The following antibodies were used: ADORA2B (rabbit polyclonal, Novus Biologicals, NBP2-41312; 1:2000); HAS3 (rabbit polyclonal, Proteintech, 15609-1-AP; 1:1000); GAPDH (mouse monoclonal, Invitrogen, AM4300; 1:500).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Immunohistochemistry

Journal: Disease Models & Mechanisms

Article Title: Adenosine and hyaluronan promote lung fibrosis and pulmonary hypertension in combined pulmonary fibrosis and emphysema

doi: 10.1242/dmm.038711

Figure Lengend Snippet: Response of macrophages to an ADORA2B agonist. (A-D) Transcript levels of Has1 (A), Has2 (B), Has3 (C) and IL6 (D) of MH-S cells exposed to DMSO (black bars), the ADORA2B antagonist GS-6201 (gray bars), the ADORA2B agonist BAY60-6583 (blue bars) or under the presence of both GS-6201 and BAY60-6583 (red bars). * P <0.05 (comparisons between DMSO and BAY60-6583 treatment groups). # P <0.05 (comparisons between BAY60-6583 and BAY60-6583+GS-6201 treatment groups); two-way ANOVA with the Benjamin, Krieger and Yekutieli post hoc test. Data are mean+s.e.m. ( n =5 for all groups).

Article Snippet: The following antibodies were used: ADORA2B (rabbit polyclonal, Novus Biologicals, NBP2-41312; 1:2000); HAS3 (rabbit polyclonal, Proteintech, 15609-1-AP; 1:1000); GAPDH (mouse monoclonal, Invitrogen, AM4300; 1:500).

Techniques:

Journal: Disease Models & Mechanisms

Article Title: Adenosine and hyaluronan promote lung fibrosis and pulmonary hypertension in combined pulmonary fibrosis and emphysema

doi: 10.1242/dmm.038711

Figure Lengend Snippet: The adenosinergic response in CPFE. (A-H) Lung transcript levels for CD39 (A), CD73 (B), ADA (C), ENT2 (D), ADORA1 (E), ADORA2A (F), ADORA2B (G) and ADORA3 (H) from upper lobes from normal lungs (white bars), upper lobes from CPFE lungs (light blue bars), lower lobes from normal lungs (gray bars) and lower lobes from CPFE lungs (dark blue bars). Data are mean+s.e.m. ( n =3 per group). * P <0.05 (comparisons between normal and CPFE groups within upper or lower lung lobes); unpaired two-tailed Student's t -test with a Welch correction.

Article Snippet: The following antibodies were used: ADORA2B (rabbit polyclonal, Novus Biologicals, NBP2-41312; 1:2000); HAS3 (rabbit polyclonal, Proteintech, 15609-1-AP; 1:1000); GAPDH (mouse monoclonal, Invitrogen, AM4300; 1:500).

Techniques: Two Tailed Test

Journal: Disease Models & Mechanisms

Article Title: Adenosine and hyaluronan promote lung fibrosis and pulmonary hypertension in combined pulmonary fibrosis and emphysema

doi: 10.1242/dmm.038711

Figure Lengend Snippet: ADORA2B and HAS3 are elevated in CPFE. (A) Western blot for ADORA2B, HAS3 and GAPDH from upper and lower lobes from normal or CPFE lungs. (B,C) Densitometries for HAS3 (B) and ADORA2B (C) from upper lobes from normal lungs (white bars), upper lobes from CPFE lungs (light blue bars), lower lobes from normal lungs (gray bars) and lower lobes from CPFE lungs (dark blue bars). (D) Dual-IHC for αSMA (red/brown) and hyaluronan (blue) in normal (upper panels) or CPFE (lower panels) lungs. The red V denotes vessels and the green asterisks indicate myofibroblasts. (E) IHC for CD68 (blue) and HAS3 (red) from lower lobes from normal (left) and CPFE (right) lungs. Red arrowheads represent CD68-positive (HAS3-negative) signals. Black arrowheads indicate dual HAS3 and CD68 signals. (F) Double HAS3- and CD68-positive cells from ten micropictographs from the upper lobes from normal lungs (white bars), upper lobes from CPFE lungs (light blue bars), lower lobes from normal lungs (gray bars) and lower lobes from CPFE lungs (dark blue bars). Data are mean+s.e.m. ( n =3 per group). * P <0.05 (comparisons between normal and CPFE groups within upper or lower lung lobes); unpaired two-tailed Student's t -test with a Welch correction. Scale bars: 50 µm.

Article Snippet: The following antibodies were used: ADORA2B (rabbit polyclonal, Novus Biologicals, NBP2-41312; 1:2000); HAS3 (rabbit polyclonal, Proteintech, 15609-1-AP; 1:1000); GAPDH (mouse monoclonal, Invitrogen, AM4300; 1:500).

Techniques: Western Blot, Two Tailed Test