drug s100a12 en rage r d systems  (R&D Systems)

Journal: Biology of reproduction

Article Title: The alarmin S100A12 causes sterile inflammation of the human chorioamniotic membranes as well as preterm birth and neonatal mortality in mice†.

doi: 10.1093/biolre/ioab188

Figure Lengend Snippet: Figure 1. Incubation with S100A12 upregulates the expression of inflammatory and pattern recognition receptor genes in the human chorioamniotic membranes. (A) Experimental design. The chorioamniotic membranes from women who delivered at term without labor were incubated with S100A12 (black dots) or PBS (control, white dots) for 24 h (n = 5–8 per group). Messenger RNA abundance of NFKB1 (B), TNF (C), IL6 (D), IL1A (E), IL1B (F), IL18 (G), RAGE (H), TLR2 (I), and TLR4 (J). Relative gene expression is presented as –CT values. The p-values were determined by either paired student’s t-tests (NFKB1, IL6, RAGE, and TLR2) or Wilcoxon signed-rank tests (TNF), based on their normality of distribution.

Article Snippet: Following mapping, an ultrasound-guided intraamniotic injection of S100A12 (Cat #1052-ER-050, R&D Systems) at concentrations of 100 ng (n = 5), 200 ng (n = 6), 300 ng (n = 4), 400 ng (n = 8), or 850 ng (n = 9) per 25 μL of sterile 1X PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Incubation, Expressing, Control, Gene Expression

Journal: Biology of reproduction

Article Title: The alarmin S100A12 causes sterile inflammation of the human chorioamniotic membranes as well as preterm birth and neonatal mortality in mice†.

doi: 10.1093/biolre/ioab188

Figure Lengend Snippet: Figure 3. Incubation with S100A12 induces the transcriptional priming of inflammasomes in the human chorioamniotic membranes. The chorioamniotic membranes from women who delivered at term without labor were incubated with S100A12 (black dots) or PBS (control, white dots) for 24 h (n = 5–8 per group). Messenger RNA abundance of NLRP1 (A), NLRP3 (B), NLRC4 (C), AIM2 (D), NOD1 (E), and NOD2 (F). Relative gene expression is presented as –CT values. The P-values were determined by paired student’s t-tests.

Article Snippet: Following mapping, an ultrasound-guided intraamniotic injection of S100A12 (Cat #1052-ER-050, R&D Systems) at concentrations of 100 ng (n = 5), 200 ng (n = 6), 300 ng (n = 4), 400 ng (n = 8), or 850 ng (n = 9) per 25 μL of sterile 1X PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Incubation, Control, Gene Expression

Journal: Biology of reproduction

Article Title: The alarmin S100A12 causes sterile inflammation of the human chorioamniotic membranes as well as preterm birth and neonatal mortality in mice†.

doi: 10.1093/biolre/ioab188

Figure Lengend Snippet: Figure 2. Incubation with S100A12 increases the concentrations of inflammatory cytokines released by the human chorioamniotic membranes. The chorioam- niotic membranes from women who delivered at term without labor were incubated with S100A12 (black dots) or PBS (control, white dots) for 24 h (n = 12 per group). The concentrations of IL-6 (A) and IL-8 (B) in culture supernatants were evaluated by specific ELISA kits. The P-values were determined by paired student’s t-tests.

Article Snippet: Following mapping, an ultrasound-guided intraamniotic injection of S100A12 (Cat #1052-ER-050, R&D Systems) at concentrations of 100 ng (n = 5), 200 ng (n = 6), 300 ng (n = 4), 400 ng (n = 8), or 850 ng (n = 9) per 25 μL of sterile 1X PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Incubation, Control, Enzyme-linked Immunosorbent Assay

Journal: Biology of reproduction

Article Title: The alarmin S100A12 causes sterile inflammation of the human chorioamniotic membranes as well as preterm birth and neonatal mortality in mice†.

doi: 10.1093/biolre/ioab188

Figure Lengend Snippet: Figure 4. Incubation with S100A12 increases the protein expression of inflammasome sensor molecules in the human chorioamniotic membranes. The chorioamniotic membranes from women who delivered at term without labor were incubated with S100A12 (black dots) or PBS (control, white dots) for 24 h (n = 10–14 per group). The concentrations of NLRP1 (A), NLRP3 (B), NLRC4 (C), AIM2 (D), NOD1 (C), and NOD2 (F) in the tissue lysates of human chorioamniotic membrane samples were evaluated by specific ELISA kits. The P-values were determined by Wilcoxon signed-rank tests.

Article Snippet: Following mapping, an ultrasound-guided intraamniotic injection of S100A12 (Cat #1052-ER-050, R&D Systems) at concentrations of 100 ng (n = 5), 200 ng (n = 6), 300 ng (n = 4), 400 ng (n = 8), or 850 ng (n = 9) per 25 μL of sterile 1X PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Incubation, Expressing, Control, Membrane, Enzyme-linked Immunosorbent Assay

Journal: Biology of reproduction

Article Title: The alarmin S100A12 causes sterile inflammation of the human chorioamniotic membranes as well as preterm birth and neonatal mortality in mice†.

doi: 10.1093/biolre/ioab188

Figure Lengend Snippet: Figure 5. Incubation with S100A12 can activate caspase-1 and promote the maturation of IL-1β in the human chorioamniotic membranes. The chorioamniotic membranes from women who delivered at term without labor were incubated with S100A12 (black dots) or PBS (control, white dots) for 24 h (n = 6–15 per group). (A) The mRNA expression of CASP1. Relative gene expression is presented as a –CT value. The P-value was determined by paired student’s t-test. (B) The concentrations of caspase-1 in the tissue lysates of human chorioamniotic membrane samples were evaluated by a specific ELISA kit. The P-values were determined by Wilcoxon signed-rank tests. (C and D) Immunoblotting of pro-caspase-1, caspase-1 p20, caspase-1 p10, and GAPDH in the human chorioamniotic membranes. The P-values were determined by Wilcoxon signed-rank tests. (E) The concentrations of mature IL-1β in culture supernatants as evaluated by a specific ELISA kit. The P-values were determined by Wilcoxon signed-rank tests.

Article Snippet: Following mapping, an ultrasound-guided intraamniotic injection of S100A12 (Cat #1052-ER-050, R&D Systems) at concentrations of 100 ng (n = 5), 200 ng (n = 6), 300 ng (n = 4), 400 ng (n = 8), or 850 ng (n = 9) per 25 μL of sterile 1X PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Incubation, Control, Expressing, Gene Expression, Membrane, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Biology of reproduction

Article Title: The alarmin S100A12 causes sterile inflammation of the human chorioamniotic membranes as well as preterm birth and neonatal mortality in mice†.

doi: 10.1093/biolre/ioab188

Figure Lengend Snippet: Figure 6. Incubation with S100A12 can trigger the activation of human chorioamniotic membranes. The chorioamniotic membranes from women who delivered at term without labor were incubated with S100A12 (black dots) or PBS (control, white dots) for 24 h (n = 6 per group). Zymography showing the activation of (A) MMP-2 and (B) MMP-9 in human chorioamniotic membranes. (C) Trichrome staining of human chorioamniotic membranes incubated with S100A12 or PBS (control). Magnification = ×20, scale bar = 100 μm. The P-value was determined by a Wilcoxon signed-rank test.

Article Snippet: Following mapping, an ultrasound-guided intraamniotic injection of S100A12 (Cat #1052-ER-050, R&D Systems) at concentrations of 100 ng (n = 5), 200 ng (n = 6), 300 ng (n = 4), 400 ng (n = 8), or 850 ng (n = 9) per 25 μL of sterile 1X PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Incubation, Activation Assay, Control, Zymography, Staining

Journal: Biology of reproduction

Article Title: The alarmin S100A12 causes sterile inflammation of the human chorioamniotic membranes as well as preterm birth and neonatal mortality in mice†.

doi: 10.1093/biolre/ioab188

Figure Lengend Snippet: Figure 7. Intra-amniotic injection of S100A12 induces preterm birth and adverse neonatal outcomes in mice. (A) Amniotic fluid concentrations of S100A12 from women with preterm labor without intra-amniotic inflammation (blue dots) and preterm labor with sterile intra-amniotic inflammation (red dots). Lines indicate the median of each group. The median and maximum values of S100A12 concentrations in women with preterm labor and sterile intra-amniotic inflammation are shown. The P-value was determined by a Mann–Whitney U-test. (B) Experimental design for the intra-amniotic administration of S100A12. Pregnant C57BL/6 mice were intra-amniotically injected with S100A12 (100 ng/25 μL, n = 5; 200 ng/25 μL, n = 6; 300 ng/25 μL, n = 4; 400 ng/25 μL, n = 8; 850 ng/25 μL, n = 9) or PBS (control, n = 6) in each amniotic sac under ultrasound guidance on 16.5 days post coitum (dpc). Pregnancy and neonatal outcomes were monitored until delivery. (C) Gestational length and rate of preterm birth of dams intra-amniotically injected with S100A12 or PBS. (D) Mortality of neonates born to dams intra-amniotically injected with S100A12 or PBS. The P-value was determined by a Mann–Whitney U-test.

Article Snippet: Following mapping, an ultrasound-guided intraamniotic injection of S100A12 (Cat #1052-ER-050, R&D Systems) at concentrations of 100 ng (n = 5), 200 ng (n = 6), 300 ng (n = 4), 400 ng (n = 8), or 850 ng (n = 9) per 25 μL of sterile 1X PBS (Fisher Scientific Bioreagents, Fair Lawn, NJ, USA) was performed in each amniotic sac using a 30-gauge needle (BD PrecisionGlide Needle, Becton Dickinson, Franklin Lakes, NJ, USA).

Techniques: Injection, Sterility, MANN-WHITNEY, Control

Journal: Scientific Reports

Article Title: Serum proteins may facilitate the identification of Kawasaki disease and promote in vitro neutrophil infiltration

doi: 10.1038/s41598-020-72695-z

Figure Lengend Snippet: The values of the concentrations in samples.

Article Snippet: In this study, neutrophils and HCAECs were treated with 20% serum, IVIG (3740501374, TBSF), recombinant S100A12 protein (1052-ER-050, R&D), antibody against S100A12 (PA5-76712, Invitrogen) and/or IgG control (UB276978, eBioscience).

Techniques:

Journal: Scientific Reports

Article Title: Serum proteins may facilitate the identification of Kawasaki disease and promote in vitro neutrophil infiltration

doi: 10.1038/s41598-020-72695-z

Figure Lengend Snippet: The variations of S100A12 gene expression and the promoter methylation assay. ( a ) We used qPCR to examine the relative mRNA levels of S100A12 in total WBCs as suggested by a previous study . The variation tendency of the S100A12 gene in the WBCs was consistent with that of the S100A12 protein in serum. Data was presented as 2 -ΔΔCt with 18S gene as internal control. ( b ) Compared with empty pGL3 vector, pGL3-S100A12 demonstrated promoter activity. ( c ) The construct carrying the S100A12 promoter and luciferase was treated with M.SssI (totally methylated) or untreated (non-methylated), followed by transfection and luciferase assays. Data were presented as the mean ± SD. ** and *** denoted p-values < 0.01 and 0.001 according to t-tests, respectively.

Article Snippet: In this study, neutrophils and HCAECs were treated with 20% serum, IVIG (3740501374, TBSF), recombinant S100A12 protein (1052-ER-050, R&D), antibody against S100A12 (PA5-76712, Invitrogen) and/or IgG control (UB276978, eBioscience).

Techniques: Gene Expression, Methylation, Control, Plasmid Preparation, Activity Assay, Construct, Luciferase, Transfection

Journal: Scientific Reports

Article Title: Serum proteins may facilitate the identification of Kawasaki disease and promote in vitro neutrophil infiltration

doi: 10.1038/s41598-020-72695-z

Figure Lengend Snippet: The variations of the surface adhesion molecules anchored on the surfaces of leukocytes. We used flow cytometry to determine the intensities (geo-means) of the surface adhesion molecules of leukocytes. ( a ) Neutrophils were treated with S100A12 or not for 24 h and then measured with flow cytometry. ( b ) Neutrophils were treated with KD serum (evenly pooled serum samples) or HC serum (evenly pooled serum samples) for 24 h and then measured with flow cytometry. Data were presented as mean ± SD. *, ** and *** denoted p-values < 0.05, 0.01 and 0.001 according to t-tests, respectively.

Article Snippet: In this study, neutrophils and HCAECs were treated with 20% serum, IVIG (3740501374, TBSF), recombinant S100A12 protein (1052-ER-050, R&D), antibody against S100A12 (PA5-76712, Invitrogen) and/or IgG control (UB276978, eBioscience).

Techniques: Flow Cytometry

Journal: Scientific Reports

Article Title: Serum proteins may facilitate the identification of Kawasaki disease and promote in vitro neutrophil infiltration

doi: 10.1038/s41598-020-72695-z

Figure Lengend Snippet: The results of LTEM assays with neutrophils manipulated. ( a , b ) Clinically freshly isolated neutrophils were left untreated or treated with S100A12, followed by LTEM assays. Compared with the control, S100A12 treatment allowed more neutrophils to penetrate the endothelial layer (2,857 vs. 2,051). ( c ) By three independent assays, S100A12 treatment significantly promoted freshly isolated neutrophils (donated by a healthy male adult) to infiltrate and to migrate through the endothelial layer. ( d ) KD serum promoted neutrophils (cell line) infiltration by approximately 1.7-fold, which allowed us to mimic KD disease by using serum treatment (n = 3). ( e ) KD serum-treated neutrophils (cell line) were simultaneously treated with a control IgG, S100A12 antibody or IVIG, followed by LTEM assays (n = 3). Data were presented as the mean ± SD. *, **, *** and **** denoted p-values < 0.05, 0.01, 0.001 and 0.0001 according to t-tests, respectively.

Article Snippet: In this study, neutrophils and HCAECs were treated with 20% serum, IVIG (3740501374, TBSF), recombinant S100A12 protein (1052-ER-050, R&D), antibody against S100A12 (PA5-76712, Invitrogen) and/or IgG control (UB276978, eBioscience).

Techniques: Isolation, Control

Journal: Clinical Epigenetics

Article Title: Multiomics analyses identified epigenetic modulation of the S100A gene family in Kawasaki disease and their significant involvement in neutrophil transendothelial migration

doi: 10.1186/s13148-018-0557-1

Figure Lengend Snippet: The expression variations of S100A family genes and methylation variations of the S100A-related CpG markers. a There were five, four, and one significant CpG markers (FDR < 0.05 and variation ratio > 1.1) located within the promoters of S100A8, S100A9, and S100A12, respectively. The locations of the ten CpG markers were indicated by the corresponding star signs. For each CpG marker, the Y axis of the box plot is the beta value (methylation percentage) determined with the M450K assays on 12 HC, 12 KD1, and 12 KD3 samples. b We used qPCR assays to detect gene expressions of S100A family genes on 24 HC, 21 FC, 17 KD1, and 18 KD3 samples. One KD1 sample failed the qPCR assay. The Y axis denoted the 2 −ΔΔCt values. The data was presented as the average ± S.D. The values of the HC set were normalized to one. * and **** denoted p values < 0.05 and < 0.0001, respectively

Article Snippet: Neutrophil-like cells were first starved for 4 h and then cultured in serum-free culture medium with 10 g/ml of S100A8/A9 (8226-S8-050, R&D), 8 g/ml of S100A9 (9254-S9-050, R&D), or 4 g/ml of S100A12 (1052-ER-050, R&D) recombinant proteins for 24 h. On the day of the migration assay, the S100A-treated neutrophil cells were washed with serum-free culture medium.

Techniques: Expressing, Methylation, Marker