Journal: Scientific Reports

Article Title: Macrophage modulation of dental pulp stem cell activity during tertiary dentinogenesis

doi: 10.1038/s41598-020-77161-4

Figure Lengend Snippet: Macrophage reduction impairs reparative dentine secretion. Clodrosome and Encapsome injection time frame. Injections (syringe) and collection date (mouse). ( A – D ) Masson trichrome staining of molars capped with either MTA or 50 nM BIO in CS post 4 weeks of damage having gone through either Clodrosome or Encapsome injections. Mice injected with Encapsome showed normal repair in all treatments (MTA and 50 nM BIO in CS). The reparative dentine of mice injected with Clodrosome was affected as both MTA and 50 nM BIO in CS capping showed sparse reparative dentine secreted. ( A′ – D′ ) μCT Cross section image confirming through radiological image the difference of reparative dentine secretion between molars that had Encapsome injections and Clodrosome injections. ( E ) μCT mineral content analysis on the damage site showed significant decrease of mineral in MTA and 50 nM BIO capped molars of animals treated with Clodrosome, in comparison to the mineral content of the control animals (Encapsome). Unpaired t-test analysis: MTA **P = 0.0018; 50 nM BIO in CS ***P = 0.0003. 100 μm scale bars. D dentine, RD reparative dentine.

Article Snippet: CD1 wild-type and TCF/Lef:H2B-GFP reporter mice mice were injected with a macrophage depletion kit Clodrosome and Encapsome (Encapsula NanoSciences).

Techniques: Injection, Staining, Comparison, Control

Journal: Scientific Reports

Article Title: Macrophage modulation of dental pulp stem cell activity during tertiary dentinogenesis

doi: 10.1038/s41598-020-77161-4

Figure Lengend Snippet: Neutrophils in repair. ( A – C ) Immunofluorescence for Ly6G in CD1 wild-type upper first molar. ( A ) Non-damaged molars did not show resident Ly6G+ cells in the dental pulp. ( B , C ) Damaged molars capped with either MTA or 50 nM BIO in CS, and collected 1 day after damage presented Ly6G+ neutrophils at damage site. ( D – G ) Ly6G antibody injected mice 1 day after damage. ( D , E ) MTA or 50 nM BIO in CS capping showed reduced Ly6G+ cells at the molar damage site following neutrophil depletion. ( F , G ) F4/80 + macrophages were present in neutrophil depleted dental pulp. ( H ) Flow cytometry for Ly6G in bone marrow of mice femur (Ly6G antibody treated [blue] and non-treated [red]) showed clear decrease of Ly6G + cell number. ( I , J ) 4 weeks after capping—Ly6G Antibody treated CD1 wild-type. Neutrophil depleted molars capped with either MTA or 50 nM BIO in CS capping showed robust reparative dentine secretion. ( K , L ) Macrophage depletion effect on neutrophils. Immunohistochemistry for Ly6G in damaged upper first molars, capped with 50 nM BIO in CS and injected with Clodrosome or Encapsome, showed increase of Ly6G + neutrophils at damage site when macrophages are depleted. Fluorescence: 25 μm scale bars; Yellow line delineates dental pulp—red line delineates MTA area; immunohistochemistry & histology: 100 μm scale bars.

Article Snippet: CD1 wild-type and TCF/Lef:H2B-GFP reporter mice mice were injected with a macrophage depletion kit Clodrosome and Encapsome (Encapsula NanoSciences).

Techniques: Immunofluorescence, Injection, Flow Cytometry, Immunohistochemistry, Fluorescence

Journal: Scientific Reports

Article Title: Macrophage modulation of dental pulp stem cell activity during tertiary dentinogenesis

doi: 10.1038/s41598-020-77161-4

Figure Lengend Snippet: F4/80+ macrophages and Wnt responsive cells activity. ( A ) TCF/Lef:H2B-GFP mice upper first molar with no damage shows Wnt responsive cells (Green) relation to F4/80+ macrophages (Red). ( B , B′ ) One day after damage in TCF/Lef:H2B-GFP mice upper molars capped with 50 nM BIO in CS showed that F4/80+ macrophages (Red) do not colocalise with Wnt responsive cells (Green), however they are adjacent to Wnt receiving cells (arrow heads). ( C – E ) Comparison of TCF/Lef:H2B-GFP mice upper molars capped with either MTA or 50 nM BIO in CS showed that there was a significant increase in Wnt responsive cells at the damage site when teeth were caped with a GSK-3 inhibitor small molecule, increasing physical proximity with F4/80+ macrophages (Red). ( F ) Time frame of Clodrosome and Encapsome injection in TCF/Lef:H2B-GFP mice. Injections (syringe) and collection date (mouse). ( G , H ) Spleen sections of TCF/Lef:H2B-GFP mice injected with either Clodrosome or Encapsome and immunohistochemistry for F4/80 showing decrease of macrophage population in the spleen. 250 μm scale bars. ( I – K ) One day after damage in TCF/Lef:H2B-GFP mice upper molars capped with 50 nM BIO in CS, injected with Clodrosome or Encapsome. Immunofluorescence for F4/80+ macrophages (Red) and GFP+ cells (Green) showed decrease of macrophages and Wnt receiving cells numbers at the damaged dental pulp when mice were injected with Clodrosome . ( B ) Yellow dashed line delineates areas in the pulp where cells were counted. Unpaired t-test analysis: ( E ) ***P = 0.0004. ( I – K ) **P = 0.0038. ( A – D ) 75 μm scale bars, ( G – J ) 25 μm scale bars.

Article Snippet: CD1 wild-type and TCF/Lef:H2B-GFP reporter mice mice were injected with a macrophage depletion kit Clodrosome and Encapsome (Encapsula NanoSciences).

Techniques: Activity Assay, Comparison, Injection, Immunohistochemistry, Immunofluorescence

Journal: Immunity

Article Title: The chemokine receptor CCR8 promotes the migration of dendritic cells into the lymph node parenchyma to initiate the allergic immune response.

doi: 10.1016/j.immuni.2018.07.012

Figure Lengend Snippet: Key Resources Table

Article Snippet: Macrophage depletion kit of Clodrosome and Encapsome , Encapsula Nano Sciences , Cat #: 8901.

Techniques: Enzyme-linked Immunosorbent Assay, Cell Isolation, Recombinant, Blocking Assay, Plasmid Preparation, Red Blood Cell Lysis, Reverse Transcription, Software

Journal: Frontiers in Microbiology

Article Title: Combining flagellin and human β-defensin-3 to combat bacterial infections

doi: 10.3389/fmicb.2014.00673

Figure Lengend Snippet: In vitro activity of F and FBD3 proteins-secretion of IL-8. (A,B) FACS analysis of Toll-Like-Receptor-5 on Colo-205 and Bjab cell lines. (C,D) In vitro activity of the F protein on Colo-205 and Bjab cells. Colo-205 or Bjab cells (10 5 cells/ml) were incubated with 20 ng (protein) of F for 4 h. The medium separated from the various samples was tested and the amount of the IL-8 was measured as described in Methods. One representative experiment out of two performed is shown (C) . Variations were ±10%. Colo-205 cells (10 5 cells/ml) were incubated with different concentrations of F (1.2 and 12 ng protein), FBD3 (2 or 20 ng fusion protein; comprising about 1.8 and 18 ng, respectively, of the F protein) and GnRH-Cherry for 4 h (20 ng protein). The medium from the different samples was analyzed and the amount of the IL-8 was measured by an Elisa assay as described in the “Methods.” One representative experiment out of four performed is shown (D) .

Article Snippet: After 4 h incubation the cell-free supernatants were analyzed according to the manufacturer’s instructions for human IL-8 content, using Human IL-8 ELIZA KIT II (BD Biosciences San Diego, CA, USA).

Techniques: In Vitro, Activity Assay, Incubation, Enzyme-linked Immunosorbent Assay

Journal: Medical mycology

Article Title: A fucose specific lectin from Aspergillus flavus induced interleukin-8 expression is mediated by mitogen activated protein kinase p38.

doi: 10.1093/mmy/myw066

Figure Lengend Snippet: Figure 3. Interleukin-8 expression profile upon FFL treatment: Cells were treated with indicated concentrations of FFL for different time period. Total RNA was isolated and cDNA was synthesized as mentioned, mRNA expression level was determined by Q-PCR method. (A) Cell viability assay: Cells were treated with varying concentrations of FFL for 24 hours. Cell viability was determined as explained in methods. The untreated cells were considered as 100% viable. (B) Cell number assay: Cells were stimulated with varying concentrations of FFL for 24 hours. The cell number in untreated sample was taken as 100%. Trypan blue exclusion assay was used to count the cells. (C) Time dependent expression of IL-8 mRNA in L-132 cells. Cells without treatment were used as control. (D) Dose dependent expression of IL-8 mRNA in L-132 cells. (E) Inhibition of IL-8 mRNA expression by L-fucose: The 40 ng/ml of FFL was preincubated with different concentrations of L-fucose (0.1 mM, 0.2 mM, and 0.3 mM) for 1 hour prior treatment to the cells for indicated time. RNA was isolated and mRNA expression level was determined as explained earlier. (F, G) Expression of IL-8 mRNA in L-132, U-937, and PBMCs: Cells were treated with FFL for 6 and 12 hour. The mRNA expression was determined as explained earlier. (H) Measurements of IL-8 production by sandwich ELISA in L-132, U-937, and PBMCs: Cells were treated with 40 ng/ml of FFL for indicated time. After each time course supernatant was collected and IL-8 concentration was determined as mentioned in kit. The cells treated with 100 nM TPA for 4 hours was used as positive control and untreated cells considered as negative control. The data are presented as mean ± SD of three individual experiments that gave similar results. ∗∗P < .01, ∗∗∗P < .001 versus control. #P < .05, ##P < .01, ###P < .001 versus FFL (40 ng/ml).

Article Snippet: Enzyme-linked immunosorbent assay (ELISA) Interleukin-8 concentration in cell culture supernatant was determined by sandwich ELISA using human IL-8 ELISA kit from Elabscience, Beijing (E-EL-H0048), following manufacturers protocol.

Techniques: Expressing, Isolation, Synthesized, Viability Assay, Trypan Blue Exclusion Assay, Control, Inhibition, Sandwich ELISA, Concentration Assay, Positive Control, Negative Control

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Neutrophils from chronic lymphocytic leukemia patients exhibit an increased capacity to release extracellular traps (NETs)

doi: 10.1007/s00262-016-1921-7

Figure Lengend Snippet: IL-8 depletion impairs the priming activity of CLL plasma to release NETs. a IL-8 levels in plasma before (whole plasma) and after depletion with a specific Ab (n = 6). Significance was determined using Wilcoxon matched-pairs test. b Neutrophils from HD were incubated for 40 min with CLL plasma (IL-8 depleted or not), washed twice and stimulated with PMA for 4 h. Elastase activity and DNA concentration were quantified in supernatants (mean ± SEM, n = 10, Friedman test followed by Dunn’s multiple comparison posttest). c Correlation between IL-8 levels in plasma from CLL patients and the capacity of PMA-activated neutrophils to release NETs. Spearman’s correlation coefficients are shown. d Addition of recombinant IL-8 (0.15 ng/ml) to HD plasma primes neutrophils to release NETs (mean ± SEM, n = 5)

Article Snippet: TNFα was from R&D Systems (Minneapolis, MN), PE-anti-human CD69, FITC-anti-human CD80, and human IL-8 ELISA Kit were purchased from BD Biosciences (Franklin Lakes, NJ).

Techniques: Activity Assay, Incubation, Concentration Assay, Comparison, Recombinant

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Neutrophils from chronic lymphocytic leukemia patients exhibit an increased capacity to release extracellular traps (NETs)

doi: 10.1007/s00262-016-1921-7

Figure Lengend Snippet: NETs delay spontaneous apoptosis and increase the expression of activation markers in CLL-B cells. a Shown are representative dot plots and the percentage of Annexin-V-positive cells after 48 h with or without ionomycin-induced NETs. Significance was determined using Wilcoxon matched-pairs test, n = 13. b Expression of activation markers in CLL cells incubated with PMA-induced NETs, supernatants from PFA-fixed neutrophils or medium (mean ± SEM, n = 6–9 CLL). Significance was determined using Friedman test followed by Dunn’s multiple comparison posttest. c Apoptosis and activation markers in CLL cells incubated with medium, supernatants from unstimulated neutrophils or NETs induced by IL-8 + LPS (mean ± SEM, n = 6)

Article Snippet: TNFα was from R&D Systems (Minneapolis, MN), PE-anti-human CD69, FITC-anti-human CD80, and human IL-8 ELISA Kit were purchased from BD Biosciences (Franklin Lakes, NJ).

Techniques: Expressing, Activation Assay, Incubation, Comparison

Journal: Canadian Journal of Veterinary Research

Article Title: Does antimicrobial therapy improve outcomes in horses with severe equine asthma and a positive tracheal wash bacterial culture?

doi:

Figure Lengend Snippet: There was no difference in interleukin-8 (IL-8) concentration of the bronchoalveolar lavage (BAL) fluid supernatant between the principal (n = 6) and control (n = 5) groups. Each horse is represented by a letter on the graph and the medians are shown as a line.

Article Snippet: Bronchoalveolar lavage fluid (BALF) supernatant was thawed in 1-mL aliquots for use in commercially available enzyme-linked immunosorbent assays (ELISA) for TNF-α [DuoSet ELISA Equine TNF-α (R&D Systems, Minneapolis, Minnesota, USA)] and IL-8 (Horse IL-8 ELISA Kit; Cusabio, College Park, Maryland, USA).

Techniques: Concentration Assay