Journal: Stem Cells (Dayton, Ohio)

Article Title: Antibacterial Effect of Human Mesenchymal Stem Cells Is Mediated in Part from Secretion of the Antimicrobial Peptide LL-37

doi: 10.1002/stem.544

Figure Lengend Snippet: Antimicrobial activity of MSC CM is mediated by LL-37 secretion. (A): Synthetic LL-37 showed significant antimicrobial activity against E. coli in a dose-dependent manner. Data are mean ± SD; *, p < .001 versus RPMI by analysis of variance (ANOVA; Bonferroni) n = 3. (B): Synthetic LL-37 displayed dose-dependent antimicrobial effect against P. aeruginosa. Data are mean ± SD; *, p < .001 versus RPMI by ANOVA (Bonferroni) n = 3. Preincubation of MSC CM with anti-LL-37 antibody (1 μ;g/ml), but not with mouse IgG (1 μg/ml), significantly reduced the antimicrobial effect of MSC CM against E. coli (C) and P. aeruginosa (D). Data are mean ± SD; *, p < .0002 versus MSC CM + anti-LL-37 by ANOVA (Bonferroni), n = 5–7. Abbreviations: CFU, colony-forming unit; MSC CM, mesenchymal stem cell conditioned medium; RPMI, RPMI-1640 medium.

Article Snippet: Mouse monoclonal antibody to human LL37/CAP18 clone 3D11 and mouse isotype IgG1 antibody were purchased from Hycult Biotechnology (Netherlands), goat Alexa-Fluor 488-labeled anti-mouse-IgG from Invitrogen, synthetic human LL-37 from AnaSpec (CA) and fetal bovine serum (FBS) from HyClone Laboratories Inc. (Utah).

Techniques: Activity Assay

Journal: Stem Cells (Dayton, Ohio)

Article Title: Antibacterial Effect of Human Mesenchymal Stem Cells Is Mediated in Part from Secretion of the Antimicrobial Peptide LL-37

doi: 10.1002/stem.544

Figure Lengend Snippet: Intratracheal administration of anti-LL 37 antibody reduced the therapeutic effect of MSC in E. coli pneumonia. Coadministration of MSC together with an anti-LL 37 neutralizing antibody (10 μg), but not with mouse IgG isotype antibody, inhibited the therapeutic effect of MSC in bacterial clearance in lung homogenates (A) and BAL fluid (B). Values are mean CFU ± SD; *, p < .002 versus MSC + anti-Ll-37 antibody treated mice; √, p < .005 versus MSC + anti-LL-37 antibody-treated mice by analysis of variance (ANOVA; Bonferroni), n = 5. (C): MSC administration enhanced antimicrobial activity of mouse BAL. BAL samples were incubated with E. coli (105 CFU/ml) for 2 hours, and CFU growth was counted. Data are mean ± SD; *, p < .002 versus BAL of PBS-treated mice by ANOVA (Bonferroni), n = 8–9. Abbreviations: BAL, bronchoalveolar lavage; CFU, colony-forming unit; LH, lung homogenates; MSC, mesenchymal stem cell; PBS, phosphate buffered saline.

Article Snippet: Mouse monoclonal antibody to human LL37/CAP18 clone 3D11 and mouse isotype IgG1 antibody were purchased from Hycult Biotechnology (Netherlands), goat Alexa-Fluor 488-labeled anti-mouse-IgG from Invitrogen, synthetic human LL-37 from AnaSpec (CA) and fetal bovine serum (FBS) from HyClone Laboratories Inc. (Utah).

Techniques: Activity Assay, Incubation

Journal: The Journal of Immunology Author Choice

Article Title: The Pore-Forming Toxin Listeriolysin O Is Degraded by Neutrophil Metalloproteinase-8 and Fails To Mediate Listeria monocytogenes Intracellular Survival in Neutrophils

doi: 10.4049/jimmunol.1301302

Figure Lengend Snippet: L. monocytogenes induces exocytosis of secondary and primary granules. (A) Neutrophils were incubated for 30 min at 37°C with L. monocytogenes or L. innocua at MOI of 10, washed, and labeled on ice with FITC-conjugated anti-CD66b, anti-CD63, or isotype control Abs. Cells were fixed postlabeling. (B) Neutrophils were incubated for 15 min at 37°C with LLO (0.5–5 nM) or LLOpL (50 nM), fixed, and labeled with FITC-conjugated anti-CD66b, anti-CD63, or isotype control Abs. (A and B) Neutrophil MFI was measured by flow cytometry. Data are expressed as MFI relative to untreated cells. Data from a representative experiment, of three, are shown.

Article Snippet: Neutrophils were then transferred to ice, washed, incubated with blocking solution for 30 min, and labeled with FITC-conjugated anti-CD63, anti-CD66b (clone CLB-B13.9; Accurate Chemical and Scientific Corporation, Westbury, NY), or FITC-conjugated control isotype (clone MOPC-21; Accurate Chemical and Scientific Corporation) Abs for 30 min at 4°C.

Techniques: Incubation, Labeling, Flow Cytometry