Journal: PLoS ONE

Article Title: Hydrogen Peroxide Produced by Oral Streptococci Induces Macrophage Cell Death

doi: 10.1371/journal.pone.0062563

Figure Lengend Snippet: Construction of S. oralis spxB deletion mutant. (A) Black arrow indicates the gene encoding pyruvate oxidase (SMSK23_0092 spxB). A targeted deletion mutant lacking this region was constructed by allelic exchange using the temperature-sensitive shuttle vector pSET4s. (B) S. oralis ATCC35037 wild-type (WT), spxB-deletion mutant (KO), or reverse mutant (Rev) was cultured in BHI broth or 5% RPMI1640 medium at 37°C for 18 h in a 5% CO 2 atmosphere. Concentrations of H 2 O 2 in culture supernatants were quantitatively determined using a hydrogen peroxide colorimetric detection kit. Data are shown as the mean ± SD of triplicate samples. *p

Article Snippet: Cell culture The human monocyte cell line THP-1 cells were purchased from RIKEN Bioresource Center and cultured in RPMI1640 medium (Invitrogen, Carlsbad, CA, USA) supplemented with 5% fetal bovine serum (FBS) (Invitrogen) (5% FBS RPMI1640), penicillin (100 U/ml), and streptomycin (100 µg/ml) at 37°C in a 5% CO2 atmosphere.

Techniques: Mutagenesis, Construct, Plasmid Preparation, Cell Culture

Journal: The Journal of Biological Chemistry

Article Title: Analyses of Group III Secreted Phospholipase A2 Transgenic Mice Reveal Potential Participation of This Enzyme in Plasma Lipoprotein Modification, Macrophage Foam Cell Formation, and Atherosclerosis * Transgenic Mice Reveal Potential Participation of This Enzyme in Plasma Lipoprotein Modification, Macrophage Foam Cell Formation, and Atherosclerosis * S⃞

doi: 10.1074/jbc.M804628200

Figure Lengend Snippet: Foam cell formation from macrophages by PLA2G3, PLA2G5 and PLA2G10. A , LDL (2 mg/ml) was incubated with or without recombinant human PLA2G3 (sPLA 2 -III) and PLA2G5 (sPLA 2 -V) for 24 h at 37 °C. Then the NEFA released was quantified ( n = 2). B and C , J774 cells were cultured for 24 h in the presence or absence of LDL (200 μg/ml) that had been incubated for 24 h with or without PLA2G3 or PLA2G5. Then the cells were homogenized, and cholesteryl ester contents were quantified ( n = 2) ( B ). Replicate cells were fixed and stained with oil red O ( C ). Ac-LDL (50 μg/ml) was used as a positive control for foam cell formation. Magnification was ×200. D , peritoneal macrophages obtained from PLA2G3 Tg ( III-Tg ) mice and PLA2G10 Tg ( X-Tg ) mice as well as WT mice were placed in RPMI1640 containing 10% lipoprotein-deficient fetal bovine serum and were incubated with or without 200 μg/ml LDL or 50 μg/ml Ac-LDL. After 24 h, the cells were stained with oil red O (magnification, ×200). Magnified views of individual macrophages are shown in the inset. E , PGE 2 production by macrophages from PLA2G3 Tg ( left ) and PLA2G10 Tg ( right ) mice in comparison with WT mice; macrophages were incubated for 24 h in the presence (+) or absence (-) of 200 μg/ml LDL (mean ± S.E., n = 3–5; * , p

Article Snippet: Peritoneal cells isolated from mice were seeded in 2 × 106 cells/well in 12-well plates (Iwaki) for 2 h in RPMI1640 containing 10% lipoprotein-deficient fetal bovine serum (Sigma), and adherent cells were used as primary macrophages.

Techniques: Incubation, Recombinant, Cell Culture, Staining, Positive Control, Mouse Assay

Journal: BMC Oral Health

Article Title: Effects of salivary protein flow and indigenous microorganisms on initial colonization of Candida albicans in an in vivo model

doi: 10.1186/1472-6831-12-36

Figure Lengend Snippet: Observation of C. albicans yeast and hyphal forms, and colony numbers after inoculation. C. albicans yeast and hyphal forms were taken pictures in microscope after incubation of them in YPD and RPMI1640 with 5% FBS ( A ). The colonization was observed on the YPD agar and BHI agar plates poured swab samples from oral cavity of NOD/SCID. e2f1 −/− mice fed 1% sucrose water at 60 min after the inoculation ( B ). White and black arrow indicated C. albicans and indigenous microorganism colony respectively. These colonies were taken pictures in the stereoscopic microscope. Data were representative in three independent assays. Samples were swabbed from the oral cavities of four 4-month-old female NOD/SCID. e2f1 +/+ and NOD/SCID. e2f1 −/− mice at 30, 60, and 90 minutes after inoculation using the yeast form. Samples swabbed in mice fed water ( C ) or 1% sucrose water ( D ) overnight before inoculation were poured on YPD agar. Samples swabbed in mice fed water ( E ) or 1% water ( F ) before inoculation were poured on BHI agar. CFU data were obtained from three independent experiments with four mice from each strain. Values are expressed as the means ± standard deviations (SDs) of the data. Asterisks show significant differences (* P

Article Snippet: The yeast form of C. albicans was predominant in culture with YPD overnight; whereas the mycelial form occurred in RPMI1640 (Gibco-Invitrogen, Grand Island, NY) with 5% FBS in overnight culture (Figure A).

Techniques: Microscopy, Incubation, Mouse Assay