rat amp activated protein kinase alpha 1  (R&D Systems)

Journal: Frontiers in Immunology

Article Title: Putative α7-selective ligands interact with α9-containing nicotinic acetylcholine receptors and modulate immune functions of human mononuclear phagocytes

doi: 10.3389/fimmu.2026.1773637

Figure Lengend Snippet: The putative α7-selective nicotinic acetylcholine receptor (nAChR) agonists S24795 (S24) and PNU-282987 (PNU) inhibit the BzATP-mediated interleukin (IL)-1β release. Monocytic THP-1 cells and THP-1 cell-derived M1-like macrophages were primed for 5 h with lipopolysaccharide (LPS; 1 µg/ml). The P2X7 receptor agonist BzATP was added for another 40 min to trigger IL-1β release, which was quantified by ELISA (A,C) and by measuring IL-1α/β-equivalent bioactive concentrations using HEK-Blue™ IL-1R cells in a QUANTI-Blue™ assay (B, D) . The BzATP- (100 µM) induced release of IL-1β was investigated in the presence and absence of the nAChR agonists S24 (50 µM) and PNU (10 µM), acetylcholine (ACh, 10 µM) or phosphocholine (PC, 200 µM). The amount of IL-1β released in response to BzATP was calculated by subtracting the IL-1β concentrations of cells treated with LPS alone. In each experiment, the IL-1β concentrations obtained after stimulation with BzATP were set to 100% and all other values were calculated accordingly. Data are presented as individual data points, bars represent median, whiskers percentiles 25 and 75. *p ≤ 0.05, different from LPS-primed cells stimulated with BzATP alone. Friedman test followed by the Wilcoxon signed-rank test.

Article Snippet: BzATP [2’(3’)-O-(4-benzoyl-benzoyl)ATP trieethylammonium salt] was purchased from Jena Bioscience (Jena, Germany), S24795 (2-[2-(4-bromophenyl)-2-oxoethyl]-1-methylpyridinium iodide, Cat# 3518) and PNU-282987 (N-(3R)-1-azabicyclo[2.2.2]oct-3-yl-4-chlorobenzamide, Cat# 2303) from Tocris (Bio-Techne, Wiesbaden-Nordenstadt, Germany) and recombinant human interferon (INF)-γ from R&D Systems (Minneapolis, MN, United States; Cat# 285-IF-100).

Techniques: Derivative Assay, Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Immunology

Article Title: Putative α7-selective ligands interact with α9-containing nicotinic acetylcholine receptors and modulate immune functions of human mononuclear phagocytes

doi: 10.3389/fimmu.2026.1773637

Figure Lengend Snippet: The effect of α7 nicotinic acetylcholine receptor (nAChR) agonists S24795 (S24) and PNU-282987 (PNU) on the BzATP-mediated release of interleukin (IL)-1β is sensitive to methyllycaconitine (MLA). Monocytic THP-1 cells (A) and THP-1 cell-derived M1-like macrophages (B) were primed for 5 h with LPS (LPS; 1 µg/ml). The P2X7 receptor agonist BzATP was added for another 40 min to trigger IL-1β release, which was measured by ELISA. The BzATP- (100 µM) induced release of IL-1β was investigated in the presence and absence of the nAChR agonists S24 (50 µM) and PNU (10 µM), or the ligands acetylcholine (ACh, 10 µM) and phosphocholine (PC, 200 µM). In addition, MLA was co-applied. The amount of IL-1β released in response to BzATP was calculated by subtracting the IL-1β concentrations measured in supernatants of cells treated with LPS alone. In each experiment, the IL-1β concentrations obtained after stimulation with BzATP were set to 100% and all other values were calculated accordingly. Data are presented as individual data points, bars represent median, whiskers percentiles 25 and 75. *p ≤ 0.05, different from LPS-primed cells stimulated with BzATP alone; #p ≤ 0.05, different from LPS-primed cells stimulated with BzATP plus an agonist. Friedman test followed by the Wilcoxon signed-rank test.

Article Snippet: BzATP [2’(3’)-O-(4-benzoyl-benzoyl)ATP trieethylammonium salt] was purchased from Jena Bioscience (Jena, Germany), S24795 (2-[2-(4-bromophenyl)-2-oxoethyl]-1-methylpyridinium iodide, Cat# 3518) and PNU-282987 (N-(3R)-1-azabicyclo[2.2.2]oct-3-yl-4-chlorobenzamide, Cat# 2303) from Tocris (Bio-Techne, Wiesbaden-Nordenstadt, Germany) and recombinant human interferon (INF)-γ from R&D Systems (Minneapolis, MN, United States; Cat# 285-IF-100).

Techniques: Derivative Assay, Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Immunology

Article Title: Putative α7-selective ligands interact with α9-containing nicotinic acetylcholine receptors and modulate immune functions of human mononuclear phagocytes

doi: 10.3389/fimmu.2026.1773637

Figure Lengend Snippet: The effect of the putative α7-selective nicotinic acetylcholine receptor (nAChR) agonists S24795 (S24) and PNU-282987 (PNU) on the BzATP-mediated cytokine release is sensitive to the α-conopeptides [V11L,V16D]ArIB and RgIA4. Monocytic THP-1 cells and THP-1 cell-derived M1-like macrophages were primed for 5 h with LPS (LPS; 1 µg/ml). The P2X7 receptor agonist BzATP was added for another 40 min to trigger the release of IL-1β (A, C) and IL-18 (B, D) , which was measured by ELISA. The BzATP- (100 µM) induced release of IL-1β and IL-18 was investigated in the presence and absence of the α7 nAChR agonists S24 (50 µM) and PNU (10 µM). To test for the involvement of nAChR subunits the conopeptides Rg1A4 (RgIA; 200 nM) or [V11L,V16D]ArIB (500 nM) were co-applied. The amount of IL-1β and IL-18 released in response to BzATP was calculated by subtracting the IL-1β concentrations measured in supernatants of cells treated with LPS alone. In each experiment, the IL-1β concentrations obtained after stimulation with BzATP were set to 100% and all other values were calculated accordingly. Data are presented as individual data points, bars represent median, whiskers percentiles 25 and 75. *p ≤ 0.05, different from LPS-primed cells stimulated with BzATP alone; #p ≤ 0.05, different from LPS-primed cells stimulated with BzATP plus an agonist. Friedman test followed by the Wilcoxon signed-rank test.

Article Snippet: BzATP [2’(3’)-O-(4-benzoyl-benzoyl)ATP trieethylammonium salt] was purchased from Jena Bioscience (Jena, Germany), S24795 (2-[2-(4-bromophenyl)-2-oxoethyl]-1-methylpyridinium iodide, Cat# 3518) and PNU-282987 (N-(3R)-1-azabicyclo[2.2.2]oct-3-yl-4-chlorobenzamide, Cat# 2303) from Tocris (Bio-Techne, Wiesbaden-Nordenstadt, Germany) and recombinant human interferon (INF)-γ from R&D Systems (Minneapolis, MN, United States; Cat# 285-IF-100).

Techniques: Derivative Assay, Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Immunology

Article Title: Putative α7-selective ligands interact with α9-containing nicotinic acetylcholine receptors and modulate immune functions of human mononuclear phagocytes

doi: 10.3389/fimmu.2026.1773637

Figure Lengend Snippet: The putative α7-selective agonists S24795 and PNU-282987 interact with α9α10 nAChRs. Xenopus laevis oocytes heterologously expressing human α9α10 nicotinic acetylcholine receptor (nAChR) were subjected to two-electrode voltage-clamp experiments and exposed to the α7-selective agonists S24795 or PNU-282987. (A, B) Oocytes expressing α9α10 nAChRs were stimulated with 1 s pulses of acetylcholine (ACh; 60 µM) until steady baseline responses were observed, then stimulated with S24795 (A) or PNU-282987 (B) and the responses compared to those evoked by ACh. The asterisks indicate 1 s pulses of S24795 (red) or PNU-282987 (green). (C, D) S24795 and PNU-282987 were also tested for antagonist activity by perfusion of the compounds during stimulation with ACh. Oocytes were stimulated with 1 s pulses of ACh until steady baseline responses were observed, then the control solution was switched to one containing S24795 (C) ; 10 min perfusion indicated by the horizontal red bar above the current traces) or PNU-282987 (D) ; 5 min perfusion, horizontal green bar). Representative current traces are shown (A-D) from 5–8 oocytes each.

Article Snippet: BzATP [2’(3’)-O-(4-benzoyl-benzoyl)ATP trieethylammonium salt] was purchased from Jena Bioscience (Jena, Germany), S24795 (2-[2-(4-bromophenyl)-2-oxoethyl]-1-methylpyridinium iodide, Cat# 3518) and PNU-282987 (N-(3R)-1-azabicyclo[2.2.2]oct-3-yl-4-chlorobenzamide, Cat# 2303) from Tocris (Bio-Techne, Wiesbaden-Nordenstadt, Germany) and recombinant human interferon (INF)-γ from R&D Systems (Minneapolis, MN, United States; Cat# 285-IF-100).

Techniques: Expressing, Activity Assay, Control

Journal: Frontiers in Immunology

Article Title: Putative α7-selective ligands interact with α9-containing nicotinic acetylcholine receptors and modulate immune functions of human mononuclear phagocytes

doi: 10.3389/fimmu.2026.1773637

Figure Lengend Snippet: Cartoon highlighting the extracellular domain with two chains of α7, α9, and α10 of homomeric nicotinic acetylcholine receptor (nAChR). Docked ligands in the α7 [shown in grey; (A, B) ], α9 [shown in cyan; (C, D) ], and α10 [shown in pink; (E, F) ] nAChR are shown. Docked poses of representative compounds S24795 (green), PNU-282987 (red), methyllycaconitine (MLA) (grey), acetylcholine (yellow), and nicotine (Pink), demonstrate binding within the C-loop region and its vicinity. Electrostatic surface potentials for complementary chain of a7, a9 and a10 are displayed in (B, D, F) respectively.

Article Snippet: BzATP [2’(3’)-O-(4-benzoyl-benzoyl)ATP trieethylammonium salt] was purchased from Jena Bioscience (Jena, Germany), S24795 (2-[2-(4-bromophenyl)-2-oxoethyl]-1-methylpyridinium iodide, Cat# 3518) and PNU-282987 (N-(3R)-1-azabicyclo[2.2.2]oct-3-yl-4-chlorobenzamide, Cat# 2303) from Tocris (Bio-Techne, Wiesbaden-Nordenstadt, Germany) and recombinant human interferon (INF)-γ from R&D Systems (Minneapolis, MN, United States; Cat# 285-IF-100).

Techniques: Binding Assay

Journal: Frontiers in Immunology

Article Title: Putative α7-selective ligands interact with α9-containing nicotinic acetylcholine receptors and modulate immune functions of human mononuclear phagocytes

doi: 10.3389/fimmu.2026.1773637

Figure Lengend Snippet: Binding mode and ligand interaction diagram for putative α7-selective nicotinic acetylcholine receptor (nAChR) agonists in the extracellular domain of homomeric α9 and α10 nAChRs. Ligand interaction diagrams for S24795 (A, D) , PNU-282987 (B, E) , and methyllycaconitine (MLA; (C, F) in the α9 (A–C) and the α10 (D-F) nAChR subunit. Ligand and residues are shown in sticks and distances and interactions are shown in lines: blue lines indicate hydrogen bonds, gray dotted lines indicate hydrophobic interactions, yellow dotted lines indicate salt-bridges interactions, green dotted lines indicate (π-stacking or π-cation), and green lines represent halogen-bonds interaction.

Article Snippet: BzATP [2’(3’)-O-(4-benzoyl-benzoyl)ATP trieethylammonium salt] was purchased from Jena Bioscience (Jena, Germany), S24795 (2-[2-(4-bromophenyl)-2-oxoethyl]-1-methylpyridinium iodide, Cat# 3518) and PNU-282987 (N-(3R)-1-azabicyclo[2.2.2]oct-3-yl-4-chlorobenzamide, Cat# 2303) from Tocris (Bio-Techne, Wiesbaden-Nordenstadt, Germany) and recombinant human interferon (INF)-γ from R&D Systems (Minneapolis, MN, United States; Cat# 285-IF-100).

Techniques: Binding Assay

Journal: Biomolecules

Article Title: Beyond Iron Solubility: Particle Size as a Determinant of Cell Survival and Iron-Induced COX-2 Expression in Human Intestinal Cells

doi: 10.3390/biom16030388

Figure Lengend Snippet: COX-2 (% of control, left-hand Y axes) and ferritin levels (ng/mg total protein, right-hand Y -axes) in ( a ) Hutu-80 and ( b ) Caco-2, measured with ELISA. Data are presented as means of nine cell replicates ± SEM, n = 3 separate occasions. Significant differences ( p ≤ 0.05) in COX-2 levels from control cells, no added Fe, are indicated with asterisks (*). Baseline ferritin levels in untreated Caco-2 cells were 2 ± 1.3 ng/mg protein (mean ± Sdev) and were 34 ± 8 ng/mg protein (mean ± Sdev) in untreated Hutu-80. All iron treatments significantly increased ferritin levels from the control ( p ≤ 0.05). The red dashed line indicates the level of COX-2 in controls (100%).

Article Snippet: Cellular levels of COX-2 (#DYC4198-2, R&D Systems, Minnesota, MN, USA) were quantified using enzyme-linked immunosorbent assay (ELISA) kits, following the manufacturers’ protocols.

Techniques: Control, Enzyme-linked Immunosorbent Assay