Journal: The FASEB Journal

Article Title: Monosodium urate crystals alter the circadian clock in macrophages leading to loss of NLRP3 inflammasome repression: Implications for timing of the gout flare

doi: 10.1096/fj.202202035r

Figure Lengend Snippet: FIGURE 4 The effect of monosodium urate (MSU) crystals on Caspase 1 activity and IL-1β secretion is time-of-day dependent. (A) Schematic of the experimental design to assess the time-of-day differences in NLRP3 inflammasome activity. Following 18 h serum starvation, cells were media changed to serum-replete media and either treated immediately with MSU crystals (500 μg/mL) or rested for 12 h before media-changing again with/without MSU crystal addition. Caspase 1 activity was measured following 12 h of MSU crystal exposure and normalized for differences in cell number as determined by CyQuant assay with reference to a standard curve. (B) The magnitude of Caspase 1 activity induced by MSU crystal exposure in cells at the 0–12 h compared with 12–24 h timepoints. (C) The magnitude of increase in secreted IL-1β levels induced by MSU crystal exposure in THP-1 macrophages at the 0–12 h compared with 12–24 h timepoints. Data shown are mean ± SEM. Data were analyzed by t-test with p < .05 considered statistically significant.

Article Snippet: Secreted IL- 1β protein levels were measured in the cell culture supernatant using a Duoset Human IL- 1β ELISA kit (R&D Systems, Minneapolis, MN, USA) following the manufacturer's protocol.

Techniques: Activity Assay, CyQUANT Assay

Journal: The FASEB Journal

Article Title: Monosodium urate crystals alter the circadian clock in macrophages leading to loss of NLRP3 inflammasome repression: Implications for timing of the gout flare

doi: 10.1096/fj.202202035r

Figure Lengend Snippet: FIGURE 6 BMAL1 represses expression of pro-IL-1β and CASP1 but not NLRP3 in THP-1 macrophages BMAL1 was knocked down or over-expressed in THP-1 macrophages using adenoviral-mediated gene delivery. Following gene transduction, cells were serum starved for 18 h then media changed to serum-replete media and either treated immediately with MSU crystals (500 μg/mL) (0–12 h timepoint) or rested for 12 h before media-changing again with/without MSU crystal addition (12–24 h timepoint). NLRP3 expression measured by RT-qPCR following BMAL1 knockdown at (A) the 0–12 h timepoint and (B) 12–24 h timepoint. NLRP3 expression measured by RT-qPCR following BMAL1 overexpression at (C) the 0–12 h timepoint and (D) 12–24 h timepoint. Pro-IL-1β expression measured by RT-qPCR following BMAL1 knockdown at (E) the 0–12 h timepoint and (F) 12–24 h timepoint. Pro-IL-1β expression measured by RT-qPCR following BMAL1 overexpression at (G) the 0–12 h timepoint and (H) 12–24 h timepoint. CASP1 expression measured by RT-qPCR following BMAL1 knockdown at (I) the 0–12 h timepoint and (J) 12–24 h timepoint. CASP1 expression measured by RT-qPCR following BMAL1 overexpression at (K) the 0–12 h timepoint and (L) 12–24 h timepoint. Data shown are mean ± SEM for 3 experimental replicates. Data were analyzed by one-way ANOVA (post-hoc Tukey) with p < .05 considered statistically significant.

Article Snippet: Secreted IL- 1β protein levels were measured in the cell culture supernatant using a Duoset Human IL- 1β ELISA kit (R&D Systems, Minneapolis, MN, USA) following the manufacturer's protocol.

Techniques: Expressing, Transduction, Quantitative RT-PCR, Knockdown, Over Expression

Journal: The FASEB Journal

Article Title: Monosodium urate crystals alter the circadian clock in macrophages leading to loss of NLRP3 inflammasome repression: Implications for timing of the gout flare

doi: 10.1096/fj.202202035r

Figure Lengend Snippet: FIGURE 7 REV-ERBα represses NLRP3 expression and inflammasome activity in THP-1 macrophages. Following 18 h serum starvation, cells were re-fed with serum-replete media and either treated immediately with/without MSU crystals (500 μg/mL) and heme (30 μM)), SR8278 (1 μM), heme+SR8278 or vehicle (0.1M NaOH + 0.1% DMSO) for 12 h (0–12 h timepoint) or rested for 12 h and then media-changed to fresh serum-replete media with/without MSU crystals and heme/SR8278/vehicle (12–24 h timepoint). (A) Caspase-1 activity normalized to cell number as measured at the 0–12 h timepoint and (B) 12–24 h timepoint. (C) Levels of secreted IL-1β measured by ELISA at the 0–12 h and (D) 12–24 h timepoint. To ensure all cells were exposed to the same vehicles, SR8278 vehicle (0.1%) DMSO) was added to the heme-only treatment and heme vehicle (0.1M NaOH) added to the SR8278-only treatment. Data shown are mean ± SEM for 3 experimental replicates. All data were analyzed by one-way ANOVA (post-hoc Tukey) p < .05 was considered statistically significant.

Article Snippet: Secreted IL- 1β protein levels were measured in the cell culture supernatant using a Duoset Human IL- 1β ELISA kit (R&D Systems, Minneapolis, MN, USA) following the manufacturer's protocol.

Techniques: Expressing, Activity Assay, Enzyme-linked Immunosorbent Assay

Journal: Nutrients

Article Title: The Clinical Effect of a Propolis and Mangosteen Extract Complex in Subjects with Gingivitis: A Randomized, Double-Blind, and Placebo-Controlled Clinical Trial

doi: 10.3390/nu16173000

Figure Lengend Snippet: Biomarkers in gingival crevicular fluid by measurement time in the PP set.

Article Snippet: The concentrations of IL-1β, PGE2, MMP-8, and MMP-9 in gingival crevicular fluid (GCF) were quantified using enzyme-linked immunosorbent assay (ELISA) kits (R&D Systems, Minneapolis, USA; IL-1β: DY201; PGE2: PKGE004B; MMP-8: DY908; MMP-9: DY911) following the manufacturer’s protocols.

Techniques: Control

Journal: Nutrients

Article Title: The Clinical Effect of a Propolis and Mangosteen Extract Complex in Subjects with Gingivitis: A Randomized, Double-Blind, and Placebo-Controlled Clinical Trial

doi: 10.3390/nu16173000

Figure Lengend Snippet: Biomarkers in gingival crevicular fluid by measurement time in the FA set.

Article Snippet: The concentrations of IL-1β, PGE2, MMP-8, and MMP-9 in gingival crevicular fluid (GCF) were quantified using enzyme-linked immunosorbent assay (ELISA) kits (R&D Systems, Minneapolis, USA; IL-1β: DY201; PGE2: PKGE004B; MMP-8: DY908; MMP-9: DY911) following the manufacturer’s protocols.

Techniques: Control

Journal: Nutrients

Article Title: The Clinical Effect of a Propolis and Mangosteen Extract Complex in Subjects with Gingivitis: A Randomized, Double-Blind, and Placebo-Controlled Clinical Trial

doi: 10.3390/nu16173000

Figure Lengend Snippet: Proposed mechanism of propolis–mangosteen extract complex (PMEC) in mitigating periodontal tissue destruction. PMEC exhibits inhibitory effects on the expression of pro-inflammatory cytokines (IL-1β and PGE2) and metalloproteases (MMP-8 and MMP-9) in oral bacterial-infected periodontal tissue cells, including neutrophils, macrophages, and gingival fibroblasts. This inhibition potentially leads to decreased osteoclastogenesis and enhanced osteogenesis, consequently reducing alveolar bone destruction. However, PMEC does not prevent the direct invasion of periodontal pathogens into gingival tissue cells or the subsequent release of lipopolysaccharides (LPSs) and exotoxins into gingival capillaries. This limitation may explain the discrepancy between clinical parameter outcomes in PMEC treatment and placebo groups, despite significant reductions in gingival crevicular fluid biomarkers (IL-1β, PGE2, MMP-8, and MMP-9) observed in PMEC-treated subjects compared to controls.

Article Snippet: The concentrations of IL-1β, PGE2, MMP-8, and MMP-9 in gingival crevicular fluid (GCF) were quantified using enzyme-linked immunosorbent assay (ELISA) kits (R&D Systems, Minneapolis, USA; IL-1β: DY201; PGE2: PKGE004B; MMP-8: DY908; MMP-9: DY911) following the manufacturer’s protocols.

Techniques: Expressing, Infection, Inhibition