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

Journal: Animals : an Open Access Journal from MDPI

Article Title: Changes in Tight Junction Protein Expression Levels but Not Distribution in Commercial White and Brown Laying Hens Supplemented with Chondrus crispus or Ascophyllum nodosum Seaweed

doi: 10.3390/ani14050777

Figure Lengend Snippet: Tight junction regulatory protein immunoreactivity in hen jejunum. ( a ) Occludin (OCC) reactivity (red) is seen throughout the intestinal epithelium, appearing more intense in the deeper cryptal areas adjacent to the muscularis. ( b ) At higher magnification, OCC staining can be seen across the cell surface of the epithelial layer, often concentrated at the apicolateral cell surfaces. ( c ) OCC negative control. ( d ) Expression of zonula occludens-1 (ZO-1) protein in jejunal epithelium, appearing less pronounced in cryptal areas than the luminal villi. ( e ) Higher magnification shows ZO-1 reactivity is most pronounced at the apico-lateral cell surface. ( f ) ZO-1 negative control. v—villus; c—crypt; m—muscularis.

Article Snippet: Separated proteins were transferred to polyvinylidene difluoride membranes (BioRad) in Towbin buffer (25 mM Tris, 192 mM glycine, pH 8.4, 20% v / v methanol) at 100 V for 120 min. Nonspecific binding sites were blocked with a solution of non-fat dry milk (3%, w / v ) containing 0.1% Tween 20 in tris-buffered saline (0.05-M Tris-HCl, 0.15-M NaCl, pH 7.6; TTBS) for one hour; then, the membrane was incubated with 12.5 mg/mL primary rabbit polyclonal antibody against occludin (Cat.# 711500, Fisher Scientific), 8.3 mg/mL rabbit polyclonal antibody ZO-1 (Cat.#617300, Fisher Scientific) or control rabbit IgG (Cat.#026102, Fisher Scientific) at the same concentration as the primary antibody.

Techniques: Staining, Negative Control, Expressing

Journal: Animals : an Open Access Journal from MDPI

Article Title: Changes in Tight Junction Protein Expression Levels but Not Distribution in Commercial White and Brown Laying Hens Supplemented with Chondrus crispus or Ascophyllum nodosum Seaweed

doi: 10.3390/ani14050777

Figure Lengend Snippet: Western blot results of occludin (OCC) and zonula occludens-1 (ZO-1) protein expression in laying hen jejunum. Graphed results are least squares means ± standard error following Tukey’s means comparisons of square-root transformed relative band intensities. ( a ) Strain and seaweed supplement effects; within effect grouping, means with the same letter were not different ( p < 0.05). ( b ) Strain x seaweed interaction effect on ZO-1 expression. Co—control diet; CC— Chondrus crispus supplement; AN— Ascophyllum nodosum supplement (See for original western blot information of ).

Article Snippet: Separated proteins were transferred to polyvinylidene difluoride membranes (BioRad) in Towbin buffer (25 mM Tris, 192 mM glycine, pH 8.4, 20% v / v methanol) at 100 V for 120 min. Nonspecific binding sites were blocked with a solution of non-fat dry milk (3%, w / v ) containing 0.1% Tween 20 in tris-buffered saline (0.05-M Tris-HCl, 0.15-M NaCl, pH 7.6; TTBS) for one hour; then, the membrane was incubated with 12.5 mg/mL primary rabbit polyclonal antibody against occludin (Cat.# 711500, Fisher Scientific), 8.3 mg/mL rabbit polyclonal antibody ZO-1 (Cat.#617300, Fisher Scientific) or control rabbit IgG (Cat.#026102, Fisher Scientific) at the same concentration as the primary antibody.

Techniques: Western Blot, Expressing, Transformation Assay, Control

Journal: Animals : an Open Access Journal from MDPI

Article Title: Changes in Tight Junction Protein Expression Levels but Not Distribution in Commercial White and Brown Laying Hens Supplemented with Chondrus crispus or Ascophyllum nodosum Seaweed

doi: 10.3390/ani14050777

Figure Lengend Snippet: Tight junction regulatory protein immunoreactivity in hen jejunum. ( a ) Occludin (OCC) reactivity (red) is seen throughout the intestinal epithelium, appearing more intense in the deeper cryptal areas adjacent to the muscularis. ( b ) At higher magnification, OCC staining can be seen across the cell surface of the epithelial layer, often concentrated at the apicolateral cell surfaces. ( c ) OCC negative control. ( d ) Expression of zonula occludens-1 (ZO-1) protein in jejunal epithelium, appearing less pronounced in cryptal areas than the luminal villi. ( e ) Higher magnification shows ZO-1 reactivity is most pronounced at the apico-lateral cell surface. ( f ) ZO-1 negative control. v—villus; c—crypt; m—muscularis.

Article Snippet: Sections were incubated for 45 min with 1.25 mg/mL primary rabbit polyclonal antibody against occludin (Cat.# 711500, Fisher Scientific), 1.25 mg/mL rabbit polyclonal antibody ZO-1 (Cat.#617300, Fisher Scientific) or 1.25 mg/mL control rabbit IgG (Cat #026102, Fisher Scientific).

Techniques: Staining, Negative Control, Expressing

Journal: Animals : an Open Access Journal from MDPI

Article Title: Changes in Tight Junction Protein Expression Levels but Not Distribution in Commercial White and Brown Laying Hens Supplemented with Chondrus crispus or Ascophyllum nodosum Seaweed

doi: 10.3390/ani14050777

Figure Lengend Snippet: Western blot results of occludin (OCC) and zonula occludens-1 (ZO-1) protein expression in laying hen jejunum. Graphed results are least squares means ± standard error following Tukey’s means comparisons of square-root transformed relative band intensities. ( a ) Strain and seaweed supplement effects; within effect grouping, means with the same letter were not different ( p < 0.05). ( b ) Strain x seaweed interaction effect on ZO-1 expression. Co—control diet; CC— Chondrus crispus supplement; AN— Ascophyllum nodosum supplement (See for original western blot information of ).

Article Snippet: Sections were incubated for 45 min with 1.25 mg/mL primary rabbit polyclonal antibody against occludin (Cat.# 711500, Fisher Scientific), 1.25 mg/mL rabbit polyclonal antibody ZO-1 (Cat.#617300, Fisher Scientific) or 1.25 mg/mL control rabbit IgG (Cat #026102, Fisher Scientific).

Techniques: Western Blot, Expressing, Transformation Assay, Control