Journal: Communications biology

Article Title: Low-dose ethanol consumption inhibits neutrophil extracellular traps formation to alleviate rheumatoid arthritis.

doi: 10.1038/s42003-023-05473-y

Figure Lengend Snippet: Fig. 3 Ethanol treatment may be associated with the inhibition of ERS and upregulation of IRE1-NETs. a Immunofluorescence staining of MPO (green) and p-IRE1 (red) in the ankle joints of mice. MPO and p-IRE1, which indicate NETs and ERS, respectively, are co-localized in the ankle joints of CIA mice (Scale Bar: 200 μm and 50 μm). b Immunohistochemical staining of MPO in the joint synovium of patients with OA and RA (Scale Bar: 100 μm and 50 μm). c Western blot detection of neutrophils in peripheral blood of healthy volunteers and RA patients. Data are representative of three independent experiments. d Immunofluorescence staining of MPO (green) and p-IRE1 (red) in the synovium of OA and RA patients. ERS is observed in neutrophils of the synovium of RA patients (Scale Bar: 25 μm).

Article Snippet: Slides were stained overnight at 4 °C with the following primary antibodies: MPO (66177-1-lg 90, Proteintech) and/or p-IRE1 (Ser724) (R24754, Zen BioScience) and/or H3cit (ab219407, Abcam).

Techniques: Inhibition, Staining, Immunohistochemical staining, Western Blot

Journal: Communications biology

Article Title: Low-dose ethanol consumption inhibits neutrophil extracellular traps formation to alleviate rheumatoid arthritis.

doi: 10.1038/s42003-023-05473-y

Figure Lengend Snippet: Fig. 6 Acetate reduces ERS in neutrophils and inhibits the formation of NETs. a Scanning electron microscopy observation of treated neutrophils (Scale Bar: 2 μm). i: Neutrophil-like differentiation of HL-60 (dHL-60) cells that developed NETs after 4 h of PMA treatment in vitro. ii: There were no NETs in dHL-60 cells treated with acetate and PMA. b Immunofluorescence staining of Sytox-green (green) and MPO (red) in dHL-60 cells, showing that NETs are reduced under sodium acetate treatment (Scale Bar: 50 μm and 25 μm). c Immunofluorescence staining of MPO (green) and p-IRE1 (red) in dHL-60 cells, showing that MPO and p-IRE1 co-localization is reduced under sodium acetate treatment (Scale Bar: 10 μm and 50 μm). d The effects of GRP78/p-IRE1/ IRE1/CHOP/ATF6/XBP1/H3cit/MPO in dHL-60 cells exposed to different concentrations of acetate. Data are representative of three independent experiments. e The effects of H3cit/MPO in dHL-60 cells exposed to different concentrations of ethanol. Data are representative of three independent experiments. f The effects of acetate receptor GPR43 and its downstream proteins in dHL-60 cells after stimulated by TG. Data are representative of three independent experiments. g The effects of GRP78/p-IRE1/IRE1/CHOP/ATF6/XBP1/H3cit/MPO in dHL-60 cells after stimulated by TG. Data are representative of three independent experiments. h Immunoprecipitation analysis of ubiquitination of endogenous GRK2 in dHL-60 cells. Data are representative of three independent experiments.

Article Snippet: Slides were stained overnight at 4 °C with the following primary antibodies: MPO (66177-1-lg 90, Proteintech) and/or p-IRE1 (Ser724) (R24754, Zen BioScience) and/or H3cit (ab219407, Abcam).

Techniques: Electron Microscopy, In Vitro, Staining, Immunoprecipitation, Ubiquitin Proteomics

Journal: Communications biology

Article Title: Low-dose ethanol consumption inhibits neutrophil extracellular traps formation to alleviate rheumatoid arthritis.

doi: 10.1038/s42003-023-05473-y

Figure Lengend Snippet: Fig. 8 Response process after low-dose ethanol treatment. Low-does ethanol treatment alters the gut microbiota, among which Muribaculaceae increases significantly, and the levels of SCFAs especially acetate in serum increases accordingly. Acetate affects the activity of neutrophils by acting on GPR43 on neutrophils, inhibits the formation of NETs by inhibiting the activation of IRE1 phosphorylation and reducing ERS in neutrophils, thereby alleviating the severity of RA.

Article Snippet: Slides were stained overnight at 4 °C with the following primary antibodies: MPO (66177-1-lg 90, Proteintech) and/or p-IRE1 (Ser724) (R24754, Zen BioScience) and/or H3cit (ab219407, Abcam).

Techniques: Activity Assay, Activation Assay, Phospho-proteomics

Journal: Biomolecules

Article Title: Investigation of Uterine Fluid Extracellular Vesicles’ Proteomic Profiles Provides Novel Diagnostic Biomarkers of Bovine Endometritis

doi: 10.3390/biom14060626

Figure Lengend Snippet: Potential biomarker for diagnosis of subclinical endometritis (SE). Only one protein, Serine peptidase HtrA1 (HTRA1), was unique with significantly enriched ( p ≤ 0.05) in UF-EV samples of SE compared to samples acquired from healthy cows (H) and cows with clinical endometritis (CLE), which therefore could potentially be a biomarker for SE ( a ). The HTRA1 results were validated using Western blotting (WB) showing similarly higher HTRA1 protein intensities in SE compared to H ( p = 0.03) and CLE ( p = 0.07). The equal loading of total protein per sample was confirmed using β-actin ( b ).

Article Snippet: The primary antibody against HTRA1 (cat. PA5-23395, Thermo Fisher Scientific, Waltham, MA, USA) and β-actin (cat. 20536-1-AP, Proteintech Group Inc., Rosemont, IL, USA), in the concentrations of 1:1000 and 1:10,000, respectively, were used.

Techniques: Biomarker Discovery, Western Blot

Journal: Stem Cells International

Article Title: Mesenchymal Stem Cells Inhibit Epithelial-to-Mesenchymal Transition by Modulating the IRE1 α Branch of the Endoplasmic Reticulum Stress Response

doi: 10.1155/2023/4483776

Figure Lengend Snippet: MSCs attenuated EMT via the IRE1 α /XBP1 pathway. (a) The protein expression levels of IRE1 α and p-IRE1 α were measured using western blotting and quantified using densitometry in ImageJ software ( n = 4, one-way ANOVA with Duncan's post hoc test). (b) The mRNA expression level of IRE1 α was measured using Q-PCR ( n = 3, one-way ANOVA with Duncan's post hoc test). (c) A549 cells were treated with 4 μ 8c (5 and 10 μ M) for 48 hr, and the protein expression levels of IRE1 α and XBP-1s were measured using western blotting and quantified using densitometry in ImageJ software ( n = 4, one-way ANOVA with Duncan's post hoc test). (d) A549 cells were treated with 10 ng/ml TGF- β 1 in the presence or absence of 4 μ 8c for 72 hr. The protein expression levels of p-IRE1 α , XBP-1s, E-cadherin and vimentin were measured using western blotting and quantified using densitometry in ImageJ software ( n = 4, one-way ANOVA with Duncan's post hoc test). The data are shown as the means ± SEMs ( ∗∗∗ P < 0.001, ∗∗ P < 0.01, ∗ P < 0.05 vs. the control group; ### P < 0.001, ## P < 0.01, # P < 0.05 vs. the TGF- β 1 group).

Article Snippet: A549 cells were treated with TGF- β 1 (10 ng/ml, 100-21, PeproTech, Rocky Hill, NJ, USA) for 72 hr with or without pretreatment with the ER stress inhibitor TUDCA (100 μ M, abs816166; Absin, Shanghai, China) or IRE1 α -specific inhibitor 4 μ 8c (10 μ M, T6363, Topscience, Shanghai, China) for 1 hr.

Techniques: Expressing, Western Blot, Software, Control

Journal: Stem Cells International

Article Title: Mesenchymal Stem Cells Inhibit Epithelial-to-Mesenchymal Transition by Modulating the IRE1 α Branch of the Endoplasmic Reticulum Stress Response

doi: 10.1155/2023/4483776

Figure Lengend Snippet: MSCs attenuated ER stress and EMT in the lungs of mice with lung fibrosis. (a) The mRNA expression levels of E-cadherin and Vimentin in lung tissues were measured using Q-PCR ( n = 5, one-way ANOVA with Duncan's post hoc test). (b) The protein expression levels of E-cadherin and vimentin in lung tissues were measured using western blotting, and the results were quantified via densitometry by using ImageJ software ( n = 3, one-way ANOVA with Duncan's post hoc test). (c) The mRNA expression levels of Bip , Atf6 , Atf4 , Xbp-1s , Ire1α and Chop in lung tissues were measured using Q-PCR ( n = 5, one-way ANOVA with Duncan's post hoc test). (d) The protein expression levels of ATF6, ATF4, IRE1 α , p-IRE1 α , XBP-1s, XBP-1u, BiP and CHOP in lung tissues were measured using western blotting, and the results were quantified via densitometry by using ImageJ software ( n = 3, one-way ANOVA with Duncan's post hoc test). (e) Images of immunofluorescence staining of BiP (green) and vimentin (red) in the lung tissues of mice. Scale bar, 100 μ m. The data are shown as the means ± SEMs ( ∗∗ P < 0.01, ∗ P < 0.05 vs. the control group; # P < 0.05, ## P < 0.01 vs. the BLM group).

Article Snippet: A549 cells were treated with TGF- β 1 (10 ng/ml, 100-21, PeproTech, Rocky Hill, NJ, USA) for 72 hr with or without pretreatment with the ER stress inhibitor TUDCA (100 μ M, abs816166; Absin, Shanghai, China) or IRE1 α -specific inhibitor 4 μ 8c (10 μ M, T6363, Topscience, Shanghai, China) for 1 hr.

Techniques: Expressing, Western Blot, Software, Immunofluorescence, Staining, Control