Journal: Antioxidants & Redox Signaling

Article Title: ATF3 Protects Pulmonary Resident Cells from Acute and Ventilator-Induced Lung Injury by Preventing Nrf2 Degradation

doi: 10.1089/ars.2014.5987

Figure Lengend Snippet: Absence of ATF3 results in decreased expression of tight junction protein occludin. (A) Quantitative real-time PCR for occludin gene expression in whole lungs from chimeric mice. Bar graphs show fold change relative to GAPDH. Data are expressed as SEM (n=4–8); *p<0.05 for the comparison between control and LPS treated, and #p<0.05 for the comparison between WTWT and the ATF3 chimera (ATF3ATF3, ATF3WT, and WTATF3). (B) Representative Western blots showing decreased expression of occludin in the negative control chimera (ATF3ATF3) compared with the positive control chimera (WTWT) and experimental chimera (ATF3WT or WTATF3) exposed to intra-tracheal LPS plus MV (two-hit model). (C) Bar graphs represent densitometry analysis from three independent experiments (n=3). Levels of occludin expression are lowest in chimera that are deficient for ATF3 in parenchymal cells (ATF3WT). For all graphs, the expression of occludin was normalized to β-actin. Data are presented as means±SEM. *p<0.05 versus corresponding WTWT saline control, $p<0.05 versus ATF3ATF3 saline control, #p<0.05 versus corresponding experimental chimera exposed to the two-, and &p<0.05 versus the one-hit model. (D) Representative photomicrographs of immunohistochemistry for occludin (Mag.×40). Brown color (black arrows) indicates occludin protein expression in vascular endothelial and bronchoalveolar epithelial pulmonary cells (IgG negative control not shown). Chimera that do not express ATF3 (negative control, ATF3ATF3) or that lack ATF3 in resident cells only (ATF3WT) show the most pronounced decrease in occludin protein expression after co-exposure to LPS and MV.

Article Snippet: Occludin Rabbit anti-human Polyclonal antibody (#LS-B2187) was from LifeSpan BioSciences.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Western Blot, Negative Control, Positive Control, Immunohistochemistry

Journal: Journal of Enzyme Inhibition and Medicinal Chemistry

Article Title: Exposure of human intestinal epithelial cells and primary human hepatocytes to trypsin-like serine protease inhibitors with potential antiviral effect

doi: 10.1080/14756366.2021.1886093

Figure Lengend Snippet: Influence of inhibitors MI-1900 and MI-1907 on occludin expression. (A) An immunofluorescence staining of occludin in controls and in inhibitor-treated cells (50 µM) for 24 hours. HIEC-6 cells were cultured on membrane inserts for eight days and subsequently exposed to inhibitors MI-1900 and MI-1907. Cells were labelled for occludin by incubating them with an anti-occludin rabbit polyclonal primary antibody followed by treatment with Alexa-Fluor 546-conjugated anti-rabbit IgG secondary antibodies. Scale bar shows 20 µm. (B) Quantitative determination of occludin levels after 24 hours treatment of HIEC-6 with inhibitors MI-1900 and MI-1907 at 50 µM. The data show mean occludin concentration values in ng/mL units ± SDs. There were no differences in occludin levels between control and MI-treated HIEC-6 ( p >.05, n = 6). (C) An immunofluorescence staining of occludin in controls and in inhibitor-treated cells (50 µM) for 24 hours. PHH cells were exposed to inhibitors MI-1900 and MI-1907. Cells were labelled for occludin by incubating them with an anti-occludin rabbit polyclonal primary antibody followed by treatment with Alexa-Fluor 546-conjugated anti-rabbit IgG secondary antibodies. Scale bar shows 30 µm. (D) Quantitative determination of occludin levels after 24 hours treatment of PHH with inhibitors MI-1900 and MI-1907 at 50 µM. The data show mean occludin concentration values in ng/mL units ± SDs. There were no differences in occludin levels between control and MI-treated PHH ( p >.05, n = 6).

Article Snippet: Sections were incubated for 1 h at room temperature in presence of anti-occludin rabbit polyclonal primary antibody (1:200, Merck, Darmstadt, Germany).

Techniques: Expressing, Immunofluorescence, Staining, Cell Culture, Membrane, Concentration Assay, Control