Journal: Journal of cellular signaling

Article Title: Chronic IL-1 Exposed AR + PCa Cell Lines Show Conserved Loss of IL-1 Sensitivity and Evolve Both Conserved and Unique Differential Gene Expression Profiles

doi:

Figure Lengend Snippet: Parental MDA-PCa-2b (MDA2b) and chronic IL-1 subline cells (MDA-αs1, MDA-αs2, MDA-βs1, MDA-βs2) were treated acutely for 3 days (A, C) or 6 days (B) with vehicle control (V), 25 ng/ml IL-1α (a), or 25 ng/ml IL-1β (b) and analyzed for cell viability using MTT (A, B) or protein accumulation by western blot (C). Acute IL-1 exposure reduces cell viability and proliferation, reduces full-length PARP (indicative of cell death activation), induces SOD2 and LCN2 protein accumulation (canonical IL-1-induced genes), and reduces AR and NKX3.1 (canonical AR target gene) protein accumulation in MDA-PCa-2b parental cells, but has little to no effect on the chronic IL-1 sublines. Thus, the IL-1 sublines evolved insensitivity to IL-1. Error bars, ± STDEV of 4 biological replicates; p-value, *≤ 0.05, **≤ 0.005, ***≤ 0.005, NS = not significant. Fold MTT optical density (OD) is normalized to treatment control. β-actin is the western blot loading control.

Article Snippet: MDA-PCa-2b cells were maintained in HPC1/20% FB Essence (FBE) containing 0.5 ng/ml IL-1α (Gold Bio, St. Louis, MO; 1110–01A-10) or IL-1β (Gold Bio, St. Louis, MO; 1110–01B-10) for approximately 4 months; and the proliferative colonies that emerged were expanded and termed MDA-PCa-2b IL-1α subline (MDA-αs) and MDA-PCa-2b IL-1β subline (MDA-βs).

Techniques: Western Blot, Activation Assay

Journal: Journal of cellular signaling

Article Title: Chronic IL-1 Exposed AR + PCa Cell Lines Show Conserved Loss of IL-1 Sensitivity and Evolve Both Conserved and Unique Differential Gene Expression Profiles

doi:

Figure Lengend Snippet: (A) MDA-PCA-2b parental (MDA2b) and chronic IL-1 subline (MDA-αs1, MDA-αs2, MDA-βs1, MDA-βs2) cells were treated acutely for 3 days with vehicle control, 25 ng/ml IL-1α, or 25 ng/ml IL-1β and analyzed by RT-qPCR for mRNA levels of the IL-1 receptor, IL-1R1 . Acute IL-1 exposure does not increase IL-1 receptor ( IL-1R1 ) mRNA levels in parental cells, suggesting basal IL-1R1 levels are sufficient to mediate IL-1 signaling. Furthermore, IL-1 does not show a differential effect on IL-1R1 mRNA levels in parental versus subline cells, suggesting subline insensitivity is independent of IL-1R1 levels. (B) Vehicle control treated cells were compared for basal mRNA levels of IL-1R1 and of canonical IL-1-induced genes, LCN2 , NOX1 , and SOD2 . IL-1R1, LCN2 , NOX1 , and SOD2 basal mRNA levels are comparable across the parental and subline cells, suggesting chronic IL-1 exposure does not induce constitutive activation of canonical IL-1 intracellular signaling. These data suggest that MDA-PCa-2b cell lines evolve insensitivity to exogenous chronic IL-1 exposure independent of IL-1R1 levels or constitutive activation of intracellular IL-1 signaling. Error bars, ± STDEV of 3 biological replicates; p-value, *≤ 0.05, **≤ 0.005, ***≤ 0.005, NS = not significant. For IL-1-treated cells, mRNA levels are normalized to vehicle control for each cell line. For basal expression, mRNA levels are normalized to the parental cell line.

Article Snippet: MDA-PCa-2b cells were maintained in HPC1/20% FB Essence (FBE) containing 0.5 ng/ml IL-1α (Gold Bio, St. Louis, MO; 1110–01A-10) or IL-1β (Gold Bio, St. Louis, MO; 1110–01B-10) for approximately 4 months; and the proliferative colonies that emerged were expanded and termed MDA-PCa-2b IL-1α subline (MDA-αs) and MDA-PCa-2b IL-1β subline (MDA-βs).

Techniques: Quantitative RT-PCR, Activation Assay, Expressing

Journal: Journal of cellular signaling

Article Title: Chronic IL-1 Exposed AR + PCa Cell Lines Show Conserved Loss of IL-1 Sensitivity and Evolve Both Conserved and Unique Differential Gene Expression Profiles

doi:

Figure Lengend Snippet: Parental (MDA-PCA-2b (MDA2b), LNCaP) and subline (MDA-αs1, MDA-αs2, MDA-βs1, MDA-βs2, LNas1, LNbs1) cells were treated acutely for 3 days with vehicle control, 25 ng/ml IL-1α, or 25 ng/ml IL-1β and analyzed by RT-qPCR for mRNA levels (A, B, C). Acute IL-1 exposure increases LCN2 , NOX1 , and SOD2 mRNA levels in parental MDA-PCa-2b and LNCaP cells, but acute IL-1 exposure has attenuated or no effect on mRNA levels in the subline cells. Thus, both LNCaP and MDA-PCa-2b cell lines show conserved intracellular response to acute IL-1-induced changes mRNA levels and evolve chronic IL-1 insensitivity independent of constitutive canonical IL-1 intracellular signaling. Error bars, ± STDEV of 3 biological replicates; p-value, *≤ 0.05, **≤ 0.005, ***≤ 0.005, NS = not significant. For IL-1-treated cells, mRNA levels are normalized to vehicle control for each cell line.

Article Snippet: MDA-PCa-2b cells were maintained in HPC1/20% FB Essence (FBE) containing 0.5 ng/ml IL-1α (Gold Bio, St. Louis, MO; 1110–01A-10) or IL-1β (Gold Bio, St. Louis, MO; 1110–01B-10) for approximately 4 months; and the proliferative colonies that emerged were expanded and termed MDA-PCa-2b IL-1α subline (MDA-αs) and MDA-PCa-2b IL-1β subline (MDA-βs).

Techniques: Quantitative RT-PCR

Journal: iScience

Article Title: A double-edged sword role of IFN-γ-producing iNKT cells in sepsis: Persistent suppression of Treg cell formation in an Nr4a1-dependent manner

doi: 10.1016/j.isci.2024.111462

Figure Lengend Snippet:

Article Snippet: Mouse IL-4 Recombinant Protein , MedChemExpress , Cat# HY-P70653.

Techniques: Control, Virus, Recombinant, Staining, Activation Assay, Cell Isolation, Software

Journal: Diabetes Care

Article Title: Islet Interleukin-1β Immunoreactivity Is an Early Feature of Cystic Fibrosis That May Contribute to β-Cell Failure

doi: 10.2337/dc17-1387

Figure Lengend Snippet: IHC for insulin, IL-1β, and amyloid in subjects with and without CF. Representative staining of pancreas specimens from subjects in non-CF control (A–D), CF-no DM (E–H), and CFRD (I–L) groups, identifying islet β-cells using IHC for insulin (brown in A, E, and I and red in D, H, and L), IL-1β (brown; images shown from two subjects per group in B and C, F and G, and J and K), and amyloid (visualized by thioflavin S [ThioS] histochemistry; green in D, H, and L). Quantitation of the proportion of islets positive for IL-1β (M) and amyloid (N). Islet IL-1β positivity was increased in CF-no DM and CFRD subjects compared with age-matched non-CF control subjects, whereas islet amyloid deposition was only increased in CFRD subjects. Scale bar = 50 µm.

Article Snippet: The last subject was a non-CF (younger) subject who had been classified as IL-1β positive with the ProSci antibody but showed no as IL-1β immunoreactivity with the Abcam antibody.

Techniques: Staining, Control, Quantitation Assay

Journal: Diabetes Care

Article Title: Islet Interleukin-1β Immunoreactivity Is an Early Feature of Cystic Fibrosis That May Contribute to β-Cell Failure

doi: 10.2337/dc17-1387

Figure Lengend Snippet: Subject characteristics and islet morphometric analyses in individuals with CF, subdivided according to diabetes and lung transplant status

Article Snippet: The last subject was a non-CF (younger) subject who had been classified as IL-1β positive with the ProSci antibody but showed no as IL-1β immunoreactivity with the Abcam antibody.

Techniques:

Journal: Experimental Biology and Medicine

Article Title: Mechanisms of AGE-induced VSMC phenotypic switching and macrophage modulation in human abdominal aortic aneurysms

doi: 10.3389/ebm.2025.10527

Figure Lengend Snippet: AGEs Activate the NF-κB Pathway and NLRP3 Inflammasome via the RAGE/RhoA/ROCK Pathway. (A) The TransAM NF-κB Transcription Factor Assay detected the effect of the RAGE/RhoA/ROCK inhibitor on the nuclear translocation of c-Rel, p52, and p65; (B) The NF-κB dual luciferase reporter gene system assessed the impact of RAGE/RhoA/ROCK inhibitors on AGE-induced NF-κB signaling; (C) Western blot analysis showed that AGEs regulate the expression of NLRP3 via the RAGE/RhoA/ROCK/NF-κB signaling pathway; (D) The effects of the inhibitors on caspase-1 activation were assessed using a caspase-1 assay kit; (E) An LDH release assay showed that AGE promoted VSMC cell pyroptosis, but it could be reversed by RAGE, RhoA, and ROCK inhibitors; (F) An ELISA assay demonstrated that AGE promoted the release of the inflammatory factor IL-1β through RAGE/RhoA/ROCK. Data are presented as the mean ± SD from three independent experiments and were analyzed with a one-way ANOVA followed by a Dunnett’s multiple comparisons test. Asterisks indicate significant differences (*P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001) between the treatment group and the control group. Pound signs indicate a significant difference (#P < 0.05, ##P < 0.01, ###P < 0.001) between the treatment group and the AGE group.

Article Snippet: The enzymatic activity of caspase-1 was assayed using a Caspase-1 Colorimetric Assay Kit (MCE, Cat. No. HY-K2610-100T) according to the manufacturer’s protocol.

Techniques: Transcription Factor Assay, Translocation Assay, Luciferase, Western Blot, Expressing, Activation Assay, Lactate Dehydrogenase Assay, Enzyme-linked Immunosorbent Assay, Control