Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: Adenosine A1and Prostaglandin E Receptor 3 Receptors Mediate Global Airway Contraction after Local Epithelial Injury

doi: 10.1165/rcmb.2012-0174oc

Figure Lengend Snippet: Figure 1. Adenosine released from a wounded single epithelial cell activates A1 receptors to induce airway contraction. (A) Bright-field images of a small airway embedded in a lung tissue slice before and z40 seconds after laser ablation demonstrate that damage of a single epithelial cell induces global airway contraction. Blue arrow points to the ablated epithelial cell, the apical membrane of which was ruptured by a pulsed femtosecond laser. Red dashed line outlines the lumen cross- sectional area before laser ablation. (B) Inhibition of A1 receptors with 50 mM PSB36 blocked local epithelial injury–induced airway contraction. (C) Local epithelial injury induced airway contraction, which leads to a roughly 30% reduction of the original cross-sectional area in control air- ways (n ¼ 33 airways from 12 rats). Statistical tests demonstrate that inhibition of adenosine receptors with 2 mM 9-chloro-2-(2-furanyl)- [1,2,4]triazolo[1,5-c]quinazolin-5-amine (CGS15943) (CGS, a nonselec- tive adenosine receptor antagonist; n ¼ 7 airways from 5 rats), inhibition of adenosine A1 receptors with 5–50 mM PSB36 (50 mM for 15 min incubation time, or 5 mM for 60 min; n ¼ 5 airways from 3 rats for 50 mM PSB36 treatment) or 4 mM SLV320 (n ¼ 9 airways from 4 rats), or deletion of adenosine with 5 units/ml adenosine deaminase (ADA; n ¼ 4 airways from 3 rats) significantly blocked the airway contraction induced by local epithelial injury, whereas inhibition of adenosine A2A receptors with 5 mM ZM241385 (n ¼ 4 airways from 2 rats), A2B receptors with 10 mM PSB603 (n ¼ 6 airways from 5 rats), or A3 receptors with 5 mM MRS1334 (n ¼ 8 airways from 6 rats) did not. (D) Statistical tests demonstrate that CGS15943 significantly blocked airway con- traction induced by 10 mM adenosine (ADO, a nonselective adenosine receptor agonist; n ¼ 4 airways from 4 rats for N6- cyclopentyladenosine [CPA] and n ¼ 4 airways from 4 rats for CPA 1 CGS treatment) or 10 mM ATP (n ¼ 5 airways from 5 rats for ATP and n ¼ 4 airways from 4 rats for ATP 1 CGS), but not 10 mM ATP-g-S (n ¼ 6 airways from 4 rats for ATP-g-S and n ¼ 5 airways from 4 rats for ATP-g-S 1 CGS). As a positive control, 25 mM KCl or 1 mM acetylcholine was used to verify the viability and contractility of airway smooth muscle cells after laser ablation experiments for each treatment. The ratio of lumen area was defined as the minimum cross- sectional area of airways after treatment divided by initial cross- sectional area. Statistical tests of significance of the ratio of lumen area between control airways and each treatment condition were per- formed with one-way ANOVA, and there are similar numbers of control airways for each treatment condition. *P , 0.05 was considered statis- tically significant.

Article Snippet: Adenosine, 9-chloro-2-(2-furanyl)-[1,2,4] triazolo[1,5-c]quinazolin-5-amine (CGS15943), 1-butyl-8-(hexahydro2,5-methanopentalen-3a(1H)-yl)-3,7-dihydro-3-(3-hydroxypropyl)-1Hpurine-2,6-dione (PSB36), 8-[4-[4-(4-chlorophenzyl)piperazide-1-sulfonyl) phenyl]]-1-propylxanthine (PSB603), SLV320, ZM241385, and MRS1334 were purchased from Tocris Bioscience (Ellisville, MO).

Techniques: Membrane, Inhibition, Control, Incubation, Positive Control

Journal: Scientific Reports

Article Title: Synergistic effects of HDAC inhibitor tucidinostat and ENT inhibitor dipyridamole in T-cell malignancies

doi: 10.1038/s41598-026-43642-1

Figure Lengend Snippet: Synergistic effect of dipyridamole and tucidinostat involves adenosine receptor activation. ( A ) MT-1 cells were treated with adenosine (250 µM), tucidinostat (Tuc; 360 nM), or their combination for 72 h. Cell viability was measured using the CCK-8 assay. Data are shown as means ± SD. *** p < 0.001, **** p < 0.0001. The synergy score was calculated using the SynergyFinder R package. (B-E) MT-1 cells were pretreated for 2 h with the indicated concentrations of selective adenosine receptor antagonists: DPCPX for A1 ( B ), ZM241385 for A2a ( C ), PSB-603 for A2b ( D ), or MRS1523 for A3 ( E ), followed by treatment with dipyridamole (Dip; 16 µM), tucidinostat (Tuc; 360 nM), or their combination for 48 h. Cell viability was assessed by CCK-8 assay. Data are presented as means ± SD. * p < 0.05, *** p < 0.001. Experiments were independently performed three times, and representative results are shown.

Article Snippet: DPCPX was purchased from Abcam, MRS1523 from Cayman Chemical Co. Ltd., and PSB-603 from MedChem Express.

Techniques: Activation Assay, CCK-8 Assay

Journal: International Journal of Molecular Sciences

Article Title: Ivy Leaf Dry Extract EA 575 ® Has an Inhibitory Effect on the Signalling Cascade of Adenosine Receptor A 2B

doi: 10.3390/ijms241512373

Figure Lengend Snippet: Influence of EA 575 ® ( A , B ) or the antagonists PSB-603 and SCH 442416 ( C ) on the CRE activation in transiently transfected HEK cells mediated by stimulation with adenosine ( A , C ) or BAY 60-6583 ( B ). Pre-incubation with 40–240 µg/mL EA 575 ® was conducted for 16 h ( A , B ) or with 0.1–10 µM of the antagonists for 2 h ( C ) before cells were stimulated by adding 100 µM adenosine ( A , C ) or 10 µM BAY 60-6583 ( B ). The non-specifically mediated CRE activation was significantly inhibited by pre-incubation with 80–240 µg/mL EA 575 ® ( A ) or 1 µM PSB-603 ( C ), compared to stimulated control cells not pre-incubated with EA 575 ® (SC). SCH 442416 also significantly reduced the CRE activation at a concentration of 10 µM, whereas this effect could not be observed at lower concentrations of both antagonists. Instead, 0.1 µM SCH 442416 slightly increased the CRE activation ( C ). The inhibition of the specific A 2B AR-mediated CRE activation was achieved by pre-incubation with 80–240 µg/mL EA 575 ® ( B ). Influence of the A 2A AR agonist CGS 21680 and, in comparison, BAY 60-6583 and adenosine on the CRE activation in transiently transfected HEK cells ( D ). Stimulation was performed with 1–10 µM CGS 21680, 10 µM BAY 60-6583, or 100 µM adenosine. The A 2A AR-mediated CRE activation was significantly increased by 10 µM CGS 21680 compared with completely untreated control cells (UTC), but to a lesser extent than that mediated by A 2B AR or non-specifically using adenosine ( D ). Data are shown as AUC of the fold change (FC) after stimulation normalised to stimulated control cells not pre-incubated with EA 575 ® (SC) ( A – C ) or completely untreated control cells (UTC) ( D ). Results represent the mean and SEM ( n = 3 independent experiments performed in triplicate, * p < 0.05).

Article Snippet: PSB-603 and CGS 21680 were obtained from Biomol (Hamburg, Germany).

Techniques: Activation Assay, Transfection, Incubation, Concentration Assay, Inhibition

Journal: International Journal of Molecular Sciences

Article Title: Ivy Leaf Dry Extract EA 575 ® Has an Inhibitory Effect on the Signalling Cascade of Adenosine Receptor A 2B

doi: 10.3390/ijms241512373

Figure Lengend Snippet: Influence of EA 575 ® ( A , B ) or the antagonists PSB-603 and SCH 442416 ( C ) on the IL-6 release of Calu-3 cells mediated by stimulation with adenosine ( A , C ) or BAY 60-6583 ( B ). Pre-incubation with 40–240 µg/mL EA 575 ® was conducted for 16 h ( A , B ) or with 0.1–10 µM of the antagonists for 1 h ( C ) before cells were stimulated by adding 100 µM adenosine ( A , C ) or 10 µM BAY 60-6583 ( B ) for another 24 h. The non-specifically mediated IL-6 release was significantly and dose-dependently inhibited by pre-incubation with 80–240 µg/mL EA 575 ® (A) or 1 µM PSB-603 ( C ), compared to stimulated control cells not pre-incubated with EA 575 ® (SC). SCH 442416, however, increased the concentration of IL-6 even further. ( C ). The inhibition of the specific A 2B AR-mediated IL-6 release was achieved by pre-incubation with 40–240 µg/mL EA 575 ® ( B ). Influence of the A 2A AR agonist CGS 21680 and, in comparison, BAY 60-6583 on the IL-6 release of Calu-3 cells ( D ). Stimulation was performed with 1–10 µM CGS 21680 or 10 µM BAY 60-6583 for 24 h. The A 2A AR-mediated IL-6 release was slightly, but neither significantly nor dose-dependently, increased compared to completely untreated control cells (UTC) ( D ). Results represent the mean normalised to stimulated control cells not pre-incubated with EA 575 ® (SC) ( A – C ) or completely untreated control cells (UTC) ( D ) and SEM ( n = 3 independent experiments performed in triplicate, * p < 0.05).

Article Snippet: PSB-603 and CGS 21680 were obtained from Biomol (Hamburg, Germany).

Techniques: Incubation, Concentration Assay, Inhibition