Journal: Turkish Journal of Biology

Article Title: Ultrasound-assisted enzymatic extraction improves the antiinflammatory activity of red dragon fruit peel pectin

doi: 10.55730/1300-0152.2797

Figure Lengend Snippet: Effects of RDFP-U and RDFP-UAE extracted from red dragon fruit on cell viability and NO production in RAW 264.7 cells. (A,B) Cell viability of RAW 264.7 cells treated with different concentrations (100, 200, 400, and 800 μg/mL) of RDFP-U and RDFP-UAE for 24 h. (C,D) Nitric oxide (NO) production in RAW 264.7 cells. (E,F) mRNA expression of iNOS in RAW 264.7 cells pretreated with RDFP-U and RDFP-UAE at different concentrations (100, 200, 400, and 800 μg/mL) for 4 h, followed by cotreatment with lipopolysaccharide (LPS, 1 μg/mL) for an additional 12 h. Data are presented as mean ± standard error of the mean (SEM) (n = 3). p < 0.001 versus control; ** p < 0.01; *** p < 0.001 versus LPS.

Article Snippet: The RAW 264.7 murine macrophage cell line was obtained from CLS Cell Lines Service GmbH (400319; Köln, Germany).

Techniques: Expressing, Control

Journal: Turkish Journal of Biology

Article Title: Ultrasound-assisted enzymatic extraction improves the antiinflammatory activity of red dragon fruit peel pectin

doi: 10.55730/1300-0152.2797

Figure Lengend Snippet: Effects of RDFP-U and RDFP-UAE extracted from red dragon fruit on (A) PGE 2 production and (B) COX-2 expression in RAW 264.7 cells treated with different concentrations (100, 200, 400, and 800 μg/mL) for 24 h. Data are presented as mean ± SEM (n = 3). ns, not significant; ### p < 0.001 versus control; * p < 0.05; ** p < 0.01; *** p < 0.001 versus LPS.

Article Snippet: The RAW 264.7 murine macrophage cell line was obtained from CLS Cell Lines Service GmbH (400319; Köln, Germany).

Techniques: Expressing, Control

Journal: Turkish Journal of Biology

Article Title: Ultrasound-assisted enzymatic extraction improves the antiinflammatory activity of red dragon fruit peel pectin

doi: 10.55730/1300-0152.2797

Figure Lengend Snippet: Effects of RDFP-U and RDFP-UAE extracted from red dragon fruit on proinflammatory cytokines (IL-6, IL-1β, and TNF-α) in RAW 264.7 macrophages. (A–D) Protein levels and mRNA expression of IL-6. (E–H) Protein levels and mRNA expression of IL-1β. (I–L) Protein levels and mRNA expression of TNF-α. Data are presented as mean ± SEM (n = 3). ns, not significant; ### p < 0.001 versus control; * p < 0.05; ** p < 0.01; *** p < 0.001 versus LPS.

Article Snippet: The RAW 264.7 murine macrophage cell line was obtained from CLS Cell Lines Service GmbH (400319; Köln, Germany).

Techniques: Expressing, Control

Journal: PLoS Pathogens

Article Title: Capsule carbohydrate structure determines virulence in Acinetobacter baumannii

doi: 10.1371/journal.ppat.1009291

Figure Lengend Snippet: (2A) RAW 264.7 cells were co-incubated with NIH1 (left) and ATCC 17978 (right) isogenic wild-type strains and Δ gtr6 mutants. (2B) RAW 264.7 cells were co-incubated with ATCC 17978, the HUMC1:: gtr6 mutant strain with repaired gtr6 , or wild-type HUMC1. (2C) RAW 264.7 cells were co-incubated with ATCC 17978 wild type, Δ gtr6 , Δ gtr6 /pSC1a (the knockout mutant with a plasmid-borne functional copy) in the presence of complement-active serum, and Δ gtr6 /pSC1a in the presence of heat-inactivated serum. *p < 0.001. (2D) Gentamicin protection assay with RAW 264.7 cells and wild-type ATCC 17978 (black bars) or ATCC 17978 Δ gtr6 (white bars). Cytochalasin D was added as an inhibitor of phagocytosis. Total bacteria plated for CFUs and expressed as a proportion of initial bacterial inoculum. * = significant vs. bacteria-only group, ⸸ = significant vs. bacteria + RAW 264.7 cell group. *,⸸ = p < 0.01 (2E) RAW 264.7 cells were incubated with ATCC 17978, HUMC1, ATCC 17978 Δ gtr6 , and HUMC1:: gtr6 . Stained with Wright-Giemsa stain, total magnification is 1000x. Results are from two repeat experiments with duplicate samples in each. White arrows denote adherent or internalized bacteria.

Article Snippet: Bacterial strains were grown in Tryptic Soy Broth (TSB) (VWR, Radnor, PA USA #90000–372) overnight at 37°C with shaking at 200 rpm, sub-cultured to logarithmic phase, washed three times in PBS, diluted to 2×10 8 CFUs/mL based on OD 600 measurements, and added to RAW 264.7 cells at a multiplicity of infection of 20:1 in Hanks’ Balanced Salt Solution (HBSS) (VWR, #45001–101) supplemented with 10% complement-active CD-1 mouse serum (Innovative Research Inc., Novi, MI USA).

Techniques: Incubation, Mutagenesis, Knock-Out, Plasmid Preparation, Functional Assay, Staining, Giemsa Stain

Journal: PLoS Pathogens

Article Title: Capsule carbohydrate structure determines virulence in Acinetobacter baumannii

doi: 10.1371/journal.ppat.1009291

Figure Lengend Snippet: ( 4A ) 2.0×10 8 CFU of ATCC 1778 and HUMC1 had total capsule carbohydrate capsule extracted in parallel and total carbohydrate content measured via phenol-sulfuric acid colorimetry. ( 4B ) Incubation of macrophages and bacteria with purified capsule from gtr6 + (ATCC 17978, 15827) and gtr6 - (HUMC1, ATCC 17978 Δ gtr6 ) strains. Extract-free uptake was used as a control. * p < 0.0001 (4C) RAW 264.7 cells were pre-incubated with soluble mannan (0.5mg/mL), laminarin (0.5mg/mL), and dextran sulfate (0.1mg/mL) or an untreated control prior to co-incubation with ATCC 17978. *p < 0.0001. Two biological replicates for in vitro . Wide bars denote median, error bars denote IQR.

Article Snippet: Bacterial strains were grown in Tryptic Soy Broth (TSB) (VWR, Radnor, PA USA #90000–372) overnight at 37°C with shaking at 200 rpm, sub-cultured to logarithmic phase, washed three times in PBS, diluted to 2×10 8 CFUs/mL based on OD 600 measurements, and added to RAW 264.7 cells at a multiplicity of infection of 20:1 in Hanks’ Balanced Salt Solution (HBSS) (VWR, #45001–101) supplemented with 10% complement-active CD-1 mouse serum (Innovative Research Inc., Novi, MI USA).

Techniques: Colorimetric Assay, Incubation, Purification, In Vitro

Journal: PLoS Pathogens

Article Title: Capsule carbohydrate structure determines virulence in Acinetobacter baumannii

doi: 10.1371/journal.ppat.1009291

Figure Lengend Snippet: (5A) RAW 264.7 cells were pre-incubated with anti-Dectin-1, anti-CR3, anti-MR neutralizing monoclonal antibodies or an isotype control prior to co-incubation with ATCC 17978. *p < 0.0005, * *p < 0.0001 (5B) Knockdown of Dectin-1 and/or CR3 in RAW 264.7 cells followed by incubation with ATCC 17978. *p < 0.0001 (5C) Primary peritoneally-elicited macrophages from C57BL/6 mice followed by phagocytosis assays with ATCC 17978. *p < 0.05, * *p < 0.0001 (5D) Phagocytosis assays of ATCC 17978 with peritoneal neutrophils from wild-type mice with disruption of phagocytosis upon the addition of heat-inactivated serum (HI-S) or complement-active serum (CA-S). *p < 0.0001 (5E) Phagocytosis assays with RAW 264.7 macrophages with gtr6 + and capsule-free strains (ATCC 17978 WT, 15827, ATCC 17978 ΔitrA ), and gtr6 - strains (ATCC 17978 Δgtr6 , HUMC1), with complement active (CA-S) or heat-inactivated (HI-S) serum. *p < 0.0001. Experiments repeated once with two biological replicates. Wide bars denote median, error bars denote IQR.

Article Snippet: Bacterial strains were grown in Tryptic Soy Broth (TSB) (VWR, Radnor, PA USA #90000–372) overnight at 37°C with shaking at 200 rpm, sub-cultured to logarithmic phase, washed three times in PBS, diluted to 2×10 8 CFUs/mL based on OD 600 measurements, and added to RAW 264.7 cells at a multiplicity of infection of 20:1 in Hanks’ Balanced Salt Solution (HBSS) (VWR, #45001–101) supplemented with 10% complement-active CD-1 mouse serum (Innovative Research Inc., Novi, MI USA).

Techniques: Incubation

Journal: PLoS Pathogens

Article Title: Capsule carbohydrate structure determines virulence in Acinetobacter baumannii

doi: 10.1371/journal.ppat.1009291

Figure Lengend Snippet: (6A) Incubation of RAW 264.7 cells with ATCC 17978 in the presence of 100μg/mL GlcNAc (NAG), a CR3 lectin domain inhibitor. (6B) Serial two-fold dilutions of complement-active mouse serum in a RAW 264.7 cell phagocytosis assay with ATCC 17978. *p < 0.0001 (6C) Male C57BL/6 mice aged 10 weeks were infected intravenously with 2.0×10 8 CFUs of 15827, with or without administration of 15μg cobra venom factor (CVF) 48 h prior to infection. *p < 0.001. Experiments repeated once, n = 5 per group for in vivo and two technical replicates for in vitro .

Article Snippet: Bacterial strains were grown in Tryptic Soy Broth (TSB) (VWR, Radnor, PA USA #90000–372) overnight at 37°C with shaking at 200 rpm, sub-cultured to logarithmic phase, washed three times in PBS, diluted to 2×10 8 CFUs/mL based on OD 600 measurements, and added to RAW 264.7 cells at a multiplicity of infection of 20:1 in Hanks’ Balanced Salt Solution (HBSS) (VWR, #45001–101) supplemented with 10% complement-active CD-1 mouse serum (Innovative Research Inc., Novi, MI USA).

Techniques: Incubation, Phagocytosis Assay, Infection, Combined Bisulfite Restriction Analysis Assay, In Vivo, In Vitro