Journal: Frontiers in Nutrition

Article Title: Oral Administration of Flavonifractor plautii , a Bacteria Increased With Green Tea Consumption, Promotes Recovery From Acute Colitis in Mice via Suppression of IL-17

doi: 10.3389/fnut.2020.610946

Figure Lengend Snippet: The effect of LTA on interleukin (IL)-17 levels in mouse splenocytes. IL-17 production in mouse splenocytes was determined in the presence and absence of IL-6/transforming growth factor (TGF)-β and various sources of LTA. Cellular supernatants were collected, and IL-17 levels were measured with ELISA. Values represent means, and error bars indicate the SD of three independent experiments. p < 0.01 by one-way analysis of variance followed by Tukey's multiple comparison test. At least three independent experiments were conducted in triplicate. HK, heat killed; SA, Staphylococcus aureus , LTA, lipoteichoic acid.

Article Snippet: The supernatant fluid was collected, and IL-17 levels were measured by ELISA (Mouse DuoSet ELISA kit, R&D Systems, Minneapolis, MN, USA) according to the manufacturer's instructions.

Techniques: Enzyme-linked Immunosorbent Assay, Comparison

Journal: JCI Insight

Article Title: DUOX1 mediates persistent epithelial EGFR activation, mucous cell metaplasia, and airway remodeling during allergic asthma

doi: 10.1172/jci.insight.88811

Figure Lengend Snippet: (A) Quantification of total and differential cell counts in BAL fluids. (B) Analysis of mucous metaplasia by histochemical analysis of PAS staining and quantification of positive staining areas using Metamorph software. (C) Quantification of Muc5ac protein in BAL fluids by ELISA. (D and E) Evaluation of subepithelial fibrosis by Masson’s trichrome staining (D) or α-SMA immunostaining (E), as quantified by staining score or percentage of staining around airways. Scale bars: 50 μm. (F) Analysis of airways central airway resistance (Rn) in response to methacholine challenge. Representative histochemical images are shown, and dot plots represent mean ± SD of 8–12 replicates from 3 independent experiments. *P < 0.05 compared with corresponding controls, using 2-way ANOVA.

Article Snippet: Moreover, HDM-induced mucus production was also demonstrated by increased levels of Muc5ac protein in the BAL, which were also significantly attenuated in Duox1 –/– mice ( ). fig ft0 fig mode=article f1 fig/graphic|fig/alternatives/graphic mode="anchored" m1 Open in a separate window Figure 3 caption a7 caption a8 DUOX1 contributes to house dust mite–induced neutrophilic inflammation, mucous metaplasia, subepithelial fibrosis, and airways hyperresponsiveness. ( A ) Quantification of total and differential cell counts in BAL fluids. ( B ) Analysis of mucous metaplasia by histochemical analysis of PAS staining and quantification of positive staining areas using Metamorph software. ( C ) Quantification of Muc5ac protein in BAL fluids by ELISA. ( D and E ) Evaluation of subepithelial fibrosis by Masson’s trichrome staining ( D ) or α-SMA immunostaining ( E ), as quantified by staining score or percentage of staining around airways.

Techniques: Staining, Software, Enzyme-linked Immunosorbent Assay, Immunostaining

Journal: JCI Insight

Article Title: DUOX1 mediates persistent epithelial EGFR activation, mucous cell metaplasia, and airway remodeling during allergic asthma

doi: 10.1172/jci.insight.88811

Figure Lengend Snippet: (A) C57BL/6J mice were subjected to house dust mite–induced (HDM-induced) inflammation and were subsequently targeted with DUOX1 siRNA. (B) Western blot analysis of EGFR and Src activation and cysteine oxidation, as in Figure 2. (C) Quantitation of BAL total cell counts and neutrophils. (D) ELISA analysis of BAL type 2 cytokines and AREG. (E and F) Evaluation of mucous metaplasia by PAS staining (E) and Muc5ac ELISA of BAL fluids (F). (G) Evaluation and quantification of α-smooth muscle actin (SMA) immunoreactivity. Scale bars: 50 μm. White and black dots represent analysis at day 22 (3 days after final HDM challenge), and red dots reflect analysis at day 25 (6 days after final HDM and 3 days after siRNA instillation). Dot plots represent mean ± SD of 6–10 replicates from 2 separate experiments. *P < 0.05 compared with corresponding controls, by 2-way ANOVA.

Article Snippet: Moreover, HDM-induced mucus production was also demonstrated by increased levels of Muc5ac protein in the BAL, which were also significantly attenuated in Duox1 –/– mice ( ). fig ft0 fig mode=article f1 fig/graphic|fig/alternatives/graphic mode="anchored" m1 Open in a separate window Figure 3 caption a7 caption a8 DUOX1 contributes to house dust mite–induced neutrophilic inflammation, mucous metaplasia, subepithelial fibrosis, and airways hyperresponsiveness. ( A ) Quantification of total and differential cell counts in BAL fluids. ( B ) Analysis of mucous metaplasia by histochemical analysis of PAS staining and quantification of positive staining areas using Metamorph software. ( C ) Quantification of Muc5ac protein in BAL fluids by ELISA. ( D and E ) Evaluation of subepithelial fibrosis by Masson’s trichrome staining ( D ) or α-SMA immunostaining ( E ), as quantified by staining score or percentage of staining around airways.

Techniques: Western Blot, Activation Assay, Quantitation Assay, Enzyme-linked Immunosorbent Assay, Staining

Journal: JCI Insight

Article Title: DUOX1 mediates persistent epithelial EGFR activation, mucous cell metaplasia, and airway remodeling during allergic asthma

doi: 10.1172/jci.insight.88811

Figure Lengend Snippet: (A) Western blot analysis of activation and cysteine oxidation of EGFR and Src in NECs from 2 healthy (C1, C2) and 3 asthmatic subjects (A1–A3), after 30 minutes of preincubation with the NOX inhibitors DPI (1 μM) or ML171 (5 μM). (B) C57BL/6J mice were subjected to house dust mite–induced (HDM-induced) allergic inflammation and subsequently administered DPI or ML171 and harvested at day 25 (6 days after final HDM challenge). (C) Western blot analysis of activation and cysteine oxidation of EGFR and Src in lung tissues collected at day 25. (D) ELISA analysis of type 2 cytokines in BAL fluids, harvested at day 25. (E) Evaluation of mucous metaplasia markers in lung tissues by RT-PCR and Muc5ac ELISA of BAL fluids, harvested at day 25. Representative Western blots are shown. Dot plots represent mean ± SD of 6–10 replicates from 2 separate experiments. *P < 0.05 compared with corresponding controls, by 2-way ANOVA.

Article Snippet: Moreover, HDM-induced mucus production was also demonstrated by increased levels of Muc5ac protein in the BAL, which were also significantly attenuated in Duox1 –/– mice ( ). fig ft0 fig mode=article f1 fig/graphic|fig/alternatives/graphic mode="anchored" m1 Open in a separate window Figure 3 caption a7 caption a8 DUOX1 contributes to house dust mite–induced neutrophilic inflammation, mucous metaplasia, subepithelial fibrosis, and airways hyperresponsiveness. ( A ) Quantification of total and differential cell counts in BAL fluids. ( B ) Analysis of mucous metaplasia by histochemical analysis of PAS staining and quantification of positive staining areas using Metamorph software. ( C ) Quantification of Muc5ac protein in BAL fluids by ELISA. ( D and E ) Evaluation of subepithelial fibrosis by Masson’s trichrome staining ( D ) or α-SMA immunostaining ( E ), as quantified by staining score or percentage of staining around airways.

Techniques: Western Blot, Activation Assay, Enzyme-linked Immunosorbent Assay, Reverse Transcription Polymerase Chain Reaction

Journal: JCI Insight

Article Title: DUOX1 mediates persistent epithelial EGFR activation, mucous cell metaplasia, and airway remodeling during allergic asthma

doi: 10.1172/jci.insight.88811

Figure Lengend Snippet: (A) Experimental design of house dust mite–induced (HDM-induced) allergic inflammation. (B) Analysis of DUOX1 and DUOX2 mRNA in lung tissues. (C and D) Analysis of DUOX1 protein expression in lung tissues using immunofluorescence (C; nuclei counterstained in blue; original magnification ×10) or Western blot analysis (D). (E) Analysis of protein cysteine sulfenylation by DCP-Bio1 labeling of lung tissue homogenates and streptavidin blotting. (F and G) Analysis of EGFR activation by IHC for phosphorylated EGFR (Y1068) (F) and Western blot analysis of lung tissue homogenates for tyrosine phosphorylation and cysteine oxidation (-S-OH, using DCP-Bio1 labeling) within EGFR or Src (G). Scale bars: 100 μm. (H) Quantitation of lung tissue Areg mRNA expression and Areg levels in BAL fluids. Qualitative data are representative of 3 separate experiments. Dot plots indicate mean ± SD of 6–10 replicates from 3 independent experiments. *P < 0.05 compared with corresponding controls, by 2-way ANOVA.

Article Snippet: Moreover, HDM-induced mucus production was also demonstrated by increased levels of Muc5ac protein in the BAL, which were also significantly attenuated in Duox1 –/– mice ( ). fig ft0 fig mode=article f1 fig/graphic|fig/alternatives/graphic mode="anchored" m1 Open in a separate window Figure 3 caption a7 caption a8 DUOX1 contributes to house dust mite–induced neutrophilic inflammation, mucous metaplasia, subepithelial fibrosis, and airways hyperresponsiveness. ( A ) Quantification of total and differential cell counts in BAL fluids. ( B ) Analysis of mucous metaplasia by histochemical analysis of PAS staining and quantification of positive staining areas using Metamorph software. ( C ) Quantification of Muc5ac protein in BAL fluids by ELISA. ( D and E ) Evaluation of subepithelial fibrosis by Masson’s trichrome staining ( D ) or α-SMA immunostaining ( E ), as quantified by staining score or percentage of staining around airways.

Techniques: Expressing, Immunofluorescence, Western Blot, Labeling, Activation Assay, Phospho-proteomics, Quantitation Assay

Journal: Biomolecules

Article Title: Diesel Exhaust Particulates Enhances Susceptibility of LPS-Induced Acute Lung Injury through Upregulation of the IL-17 Cytokine-Derived TGF-β 1 /Collagen I Expression and Activation of NLRP3 Inflammasome Signaling in Mice

doi: 10.3390/biom11010067

Figure Lengend Snippet: ( A – D ) Cellular changes in the bronchoalveolar lavage (BAL) fluid and ( E ) representative hematoxylin and eosin (H&E) and Masson’s trichrome-stained images of the lung sections from the vehicle control (VC), DEP, LPS, and DEP pre-exposed and LPS-instilled (DEP + LPS) groups. Black, red, and blue arrows indicate particle-pigmented alveolar macrophages, inflammatory infiltration, and collagen deposition, respectively. Scale bars = 50 μm. Data represent means ± SD (n = 5 per group). # p < 0.05 vs. VC, * p < 0.05 vs. LPS group.

Article Snippet: The levels of IL-1β, IL-6, tumor necrosis factor (TNF)-α, and IL-17 in the BAL fluid were quantified by ELISA using commercial kits (Thermo Fisher Scientific), according to the manufacturer’s protocol.

Techniques: Staining, Control

Journal: Biomolecules

Article Title: Diesel Exhaust Particulates Enhances Susceptibility of LPS-Induced Acute Lung Injury through Upregulation of the IL-17 Cytokine-Derived TGF-β 1 /Collagen I Expression and Activation of NLRP3 Inflammasome Signaling in Mice

doi: 10.3390/biom11010067

Figure Lengend Snippet: Protein levels of ( A ) IL-1β, ( B ) IL-6, and ( C ) TNF-α in the bronchoalveolar (BAL) fluid from mice in the vehicle control (VC), DEP, LPS, and DEP pre-exposed and LPS-instilled (DEP+LPS) groups. Data represent means ± SD (n = 5 per group). # p < 0.05 vs. VC, *** p < 0.001 vs. LPS group.

Article Snippet: The levels of IL-1β, IL-6, tumor necrosis factor (TNF)-α, and IL-17 in the BAL fluid were quantified by ELISA using commercial kits (Thermo Fisher Scientific), according to the manufacturer’s protocol.

Techniques: Control

Journal: Biomolecules

Article Title: Diesel Exhaust Particulates Enhances Susceptibility of LPS-Induced Acute Lung Injury through Upregulation of the IL-17 Cytokine-Derived TGF-β 1 /Collagen I Expression and Activation of NLRP3 Inflammasome Signaling in Mice

doi: 10.3390/biom11010067

Figure Lengend Snippet: Protein levels of ( A ) IL-17 in the bronchoalveolar (BAL) fluid and ( B ) representative images of immunohistochemistry (IHC) for IL-17 (brown color) in the lung tissue sections from mice in the VC, DEP, LPS, and DEP pre-exposed and LPS-instilled (DEP+LPS) groups. Scale bars = 200 nm. Data represent means ± SD (n = 5 per group) *** p < 0.001 vs. LPS group.

Article Snippet: The levels of IL-1β, IL-6, tumor necrosis factor (TNF)-α, and IL-17 in the BAL fluid were quantified by ELISA using commercial kits (Thermo Fisher Scientific), according to the manufacturer’s protocol.

Techniques: Immunohistochemistry

Journal: European Journal of Human Genetics

Article Title: Human beta defensin (HBD) gene copy number affects HBD2 protein levels: impact on cervical bactericidal immunity in pregnancy

doi: 10.1038/s41431-017-0061-7

Figure Lengend Snippet: Beta defensin copy number, HBD2 protein expression and cervical antimicrobial activity. HBD2 protein concentration and antimicrobial activity (%kill/total protein) are presented on logarithmic scales. Beta defensin copy numbers are displayed as weighted mean PRT copy numbers. Red lines and grey-shaded regions on scatterplots represent regression lines and 95% confidence intervals. Copy numbers were determined by triplex PRT and protein levels were determined by ELISA. Histograms a , c and f display the distribution of values for beta defensin copy number, HBD2 protein concentration and antimicrobial activity, respectively. A significant positive correlation was observed between copy number and HBD2 protein levels (Plot b ; Spearman r = 0.21, p = 0.0032). A weak but significant correlation was observed between copy number and antimicrobial activity (Plot d Spearman r = 0.17, p = 0.028. There was a strong correlation between HBD2 protein level and antimicrobial activity (Plot e ; Spearman r = 0.49, p < 0.0001)

Article Snippet: Cervicovaginal fluid HBD2 levels were determined by ELISA (Peprotech ELISA Development Kit, according to the manufacturer’s instructions).

Techniques: Expressing, Activity Assay, Protein Concentration, Enzyme-linked Immunosorbent Assay

Journal: European Journal of Human Genetics

Article Title: Human beta defensin (HBD) gene copy number affects HBD2 protein levels: impact on cervical bactericidal immunity in pregnancy

doi: 10.1038/s41431-017-0061-7

Figure Lengend Snippet: Beta defensin copy number, HBD2 protein expression and gestational age at sample collection. HBD2 protein concentration is presented on a logarithmic scale. Beta defensin copy numbers are displayed as weighted mean PRT copy numbers. Red lines and grey-shaded regions on scatterplots represent regression lines and 95% confidence intervals. Copy numbers were determined by triplex PRT and protein levels were determined by ELISA. Histograms a , c and f display the distribution of values for beta defensin copy number, HBD2 protein concentration and gestational age at sampling (in days), respectively. HBD2 protein levels decreased significantly with increasing gestational age at sample collection (Plot e Spearman r = −0.25, p = 0.0003); there was no correlation between copy number and gestational age at sample collection (Plot d ). (Plots b and c are as described in Fig. )

Article Snippet: Cervicovaginal fluid HBD2 levels were determined by ELISA (Peprotech ELISA Development Kit, according to the manufacturer’s instructions).

Techniques: Expressing, Protein Concentration, Enzyme-linked Immunosorbent Assay, Sampling

Journal: European journal of pharmacology

Article Title: Chromofungin, a chromogranin A-derived peptide, protects against sepsis-induced acute lung injury by inhibiting LBP/TLR4-dependent inflammatory signaling.

doi: 10.1016/j.ejphar.2023.176043

Figure Lengend Snippet: Fig. 1. Effects of CHR pretreatment on pulmonary inflammation in septic mice. Mice were subjected to CHR pretreatment and CLP administration as described in the “Materials and Methods” section. (A) Structure diagram of CHR. (B) Survival rates of mice in the Sham (n = 18), CLP (n = 18), CHR + Sham (n = 17), and CHR + CLP groups (n = 16). Eighty mice were randomly divided into four groups with twenty mice in each group, excluding mice that died during anesthesia and resuscitation. (C–E) Protein levels of IL-1β, IL-4, IL-10 in BALF (C), mRNA levels of TNF-α, IL-1β, IL-4 and IL-10 in lung tissues (D), images of H&E-stained lung tissues (200x) and semiquantitative histological scores of lung injury (E) from mice with or without treatment 30 min after intraperitoneal CHR injection, followed by sham or CLP administration. One-way ANOVA followed by multiple comparison tests were used to analyze the data, and the data represent the mean ± SEM (n = 3). The sig nificance level was adjusted at 0.05. Scale bar = 50 μM *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.

Article Snippet: Cytokine (IL-1β, interleukin-4 [IL-4] and interleukin-10 [IL-10]) levels in mouse bronchoalveolar lavage fluid (BALF) or cell culture supernatants were measured by a mouse ELISA detection kit (Proteintech).

Techniques: Staining, Injection, Comparison