Structured Review

Abcam rabbit anti occludin
Adipose tissue extracts enable the EMT of CCA cells. Human cholangiocarcinoma (CCA) cell line RBE was incubated with complete growth medium or medium containing human adipose tissue extracts for 24 h. The cell morphology was observed under the invert microscopy (40×) ( a ). Protein expression of MMP2, MMP9, TGF-β1, claudin 1, <t>occludin,</t> E-cadherin, β-catenin, SNAIL, and Smad3 were analyzed by immunoblot analysis in response to adipose extracts incubation, and the band densitometry analysis was carried out relative to the loading control β-actin or GAPDH ( b , c ). To determine fatty acid metabolism, the culture medium with different adipose components prior to or after incubation with RBE cells for 24 h were collected for extracellular glycerol and nonestesterified fatty acid (NEFA) detection using colorimetric assays, with the data presented as the absorbance at 490 nm( d , e ). The intracellular lipids in adipose cultured RBE cells were visualized by the fluorescence dye Bodipy (20 μg/mL) using confocal microscopy analysis, using Hochest33342 (0.5 μg/mL) for nuclear staining ( f ). All the samples were prepared in triplicate, and all experiments were repeated at least three times. * P
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1) Product Images from "Adipocytes promote cholangiocarcinoma metastasis through fatty acid binding protein 4"

Article Title: Adipocytes promote cholangiocarcinoma metastasis through fatty acid binding protein 4

Journal: Journal of Experimental & Clinical Cancer Research : CR

doi: 10.1186/s13046-017-0641-y

Adipose tissue extracts enable the EMT of CCA cells. Human cholangiocarcinoma (CCA) cell line RBE was incubated with complete growth medium or medium containing human adipose tissue extracts for 24 h. The cell morphology was observed under the invert microscopy (40×) ( a ). Protein expression of MMP2, MMP9, TGF-β1, claudin 1, occludin, E-cadherin, β-catenin, SNAIL, and Smad3 were analyzed by immunoblot analysis in response to adipose extracts incubation, and the band densitometry analysis was carried out relative to the loading control β-actin or GAPDH ( b , c ). To determine fatty acid metabolism, the culture medium with different adipose components prior to or after incubation with RBE cells for 24 h were collected for extracellular glycerol and nonestesterified fatty acid (NEFA) detection using colorimetric assays, with the data presented as the absorbance at 490 nm( d , e ). The intracellular lipids in adipose cultured RBE cells were visualized by the fluorescence dye Bodipy (20 μg/mL) using confocal microscopy analysis, using Hochest33342 (0.5 μg/mL) for nuclear staining ( f ). All the samples were prepared in triplicate, and all experiments were repeated at least three times. * P
Figure Legend Snippet: Adipose tissue extracts enable the EMT of CCA cells. Human cholangiocarcinoma (CCA) cell line RBE was incubated with complete growth medium or medium containing human adipose tissue extracts for 24 h. The cell morphology was observed under the invert microscopy (40×) ( a ). Protein expression of MMP2, MMP9, TGF-β1, claudin 1, occludin, E-cadherin, β-catenin, SNAIL, and Smad3 were analyzed by immunoblot analysis in response to adipose extracts incubation, and the band densitometry analysis was carried out relative to the loading control β-actin or GAPDH ( b , c ). To determine fatty acid metabolism, the culture medium with different adipose components prior to or after incubation with RBE cells for 24 h were collected for extracellular glycerol and nonestesterified fatty acid (NEFA) detection using colorimetric assays, with the data presented as the absorbance at 490 nm( d , e ). The intracellular lipids in adipose cultured RBE cells were visualized by the fluorescence dye Bodipy (20 μg/mL) using confocal microscopy analysis, using Hochest33342 (0.5 μg/mL) for nuclear staining ( f ). All the samples were prepared in triplicate, and all experiments were repeated at least three times. * P

Techniques Used: Incubation, Microscopy, Expressing, Cell Culture, Fluorescence, Confocal Microscopy, Staining

2) Product Images from "A transgenic toolkit for visualizing and perturbing microtubules reveals unexpected functions in the epidermis"

Article Title: A transgenic toolkit for visualizing and perturbing microtubules reveals unexpected functions in the epidermis

Journal: eLife

doi: 10.7554/eLife.29834

Spastin OE does not impair tight junction localization or function. ( A ) ZO-1 localization in control and K10-rtTA; TRE-spastin epidermis. Scale-25µm. ( B ) Region where spastin-positive cells are next to spastin-negative cells in K10-rtTA; TRE-spastin tissue. Note that ZO-1 is still cortically localized in spastin-positive cells. Scale-10µm. ( C ) Localization of occludin at the cell cortex is maintained in K10-rtTA; TRE-spastin epidermis. Scale-10µm. ( D ) Biotin diffusion is blocked by occludin in K10-rtTA; TRE-spastin epidermis. Scale-10µm.
Figure Legend Snippet: Spastin OE does not impair tight junction localization or function. ( A ) ZO-1 localization in control and K10-rtTA; TRE-spastin epidermis. Scale-25µm. ( B ) Region where spastin-positive cells are next to spastin-negative cells in K10-rtTA; TRE-spastin tissue. Note that ZO-1 is still cortically localized in spastin-positive cells. Scale-10µm. ( C ) Localization of occludin at the cell cortex is maintained in K10-rtTA; TRE-spastin epidermis. Scale-10µm. ( D ) Biotin diffusion is blocked by occludin in K10-rtTA; TRE-spastin epidermis. Scale-10µm.

Techniques Used: Diffusion-based Assay

3) Product Images from "Progressive Hearing Loss in Mice Carrying a Mutation in Usp53"

Article Title: Progressive Hearing Loss in Mice Carrying a Mutation in Usp53

Journal: The Journal of Neuroscience

doi: 10.1523/JNEUROSCI.1965-15.2015

USP53 interacts with TJP1 and TJP2 via its C-terminal tail. A , GFP–USP53 fusion constructs, depicted schematically in C , were injectoporated into cochlear explants from P4 wild-type (WT) mice. GFP fusion proteins containing the C-terminal tail of USP53 localized to TJs. B , GFP and GFP–USP53 fusion constructs were expressed in HEK293T cells and immunoprecipitated with anti-GFP antibody (Abcam). Extracts were directly analyzed for the expression of GFP-tagged USP53 constructs using anti-GFP antibody (Sigma). An asterisk marks a nonspecific cross-reacting band at ∼80 kDa used as a loading control. Native TJ proteins TJP1 and TJP2, but not occludin (OCLN), coprecipitated specifically with GFP fusion proteins that contain the C-terminal tail of USP53. IB, Immunoblot; IP, immunoprecipitation. C , Schematic diagram of the GFP–USP53 fusion proteins and TJPs used for biochemical experiments and their predicted molecular sizes. D , HEK293T cells were transfected with the constructs indicated on top of each panel. Immunoprecipitations were performed with anti-GFP antibody (Abcam) that recognizes GFP–USP53 fusion proteins followed by Western blotting with anti-FLAG antibody or anti-GFP antibody (Abcam). 2xFLAG–TJP2 coprecipitated with TJP2–GFP (TJP2 homodimerization), as well as with GFP–USP53 constructs containing a stretch of 328 aa adjacent to the catalytic domain. Neither the catalytic domain nor GFP alone precipitated 2xFLAG–TJP2. E , Schematic representation of C-terminal deletion constructs of USP53 used in D . The region containing the TJP2 interaction site is highlighted in blue. Scale bar: A , 5 μm.
Figure Legend Snippet: USP53 interacts with TJP1 and TJP2 via its C-terminal tail. A , GFP–USP53 fusion constructs, depicted schematically in C , were injectoporated into cochlear explants from P4 wild-type (WT) mice. GFP fusion proteins containing the C-terminal tail of USP53 localized to TJs. B , GFP and GFP–USP53 fusion constructs were expressed in HEK293T cells and immunoprecipitated with anti-GFP antibody (Abcam). Extracts were directly analyzed for the expression of GFP-tagged USP53 constructs using anti-GFP antibody (Sigma). An asterisk marks a nonspecific cross-reacting band at ∼80 kDa used as a loading control. Native TJ proteins TJP1 and TJP2, but not occludin (OCLN), coprecipitated specifically with GFP fusion proteins that contain the C-terminal tail of USP53. IB, Immunoblot; IP, immunoprecipitation. C , Schematic diagram of the GFP–USP53 fusion proteins and TJPs used for biochemical experiments and their predicted molecular sizes. D , HEK293T cells were transfected with the constructs indicated on top of each panel. Immunoprecipitations were performed with anti-GFP antibody (Abcam) that recognizes GFP–USP53 fusion proteins followed by Western blotting with anti-FLAG antibody or anti-GFP antibody (Abcam). 2xFLAG–TJP2 coprecipitated with TJP2–GFP (TJP2 homodimerization), as well as with GFP–USP53 constructs containing a stretch of 328 aa adjacent to the catalytic domain. Neither the catalytic domain nor GFP alone precipitated 2xFLAG–TJP2. E , Schematic representation of C-terminal deletion constructs of USP53 used in D . The region containing the TJP2 interaction site is highlighted in blue. Scale bar: A , 5 μm.

Techniques Used: Construct, Mouse Assay, Immunoprecipitation, Expressing, Transfection, Western Blot

4) Product Images from "The toxic effects of microcystin-LR on mouse lungs and alveolar type II epithelial cells"

Article Title: The toxic effects of microcystin-LR on mouse lungs and alveolar type II epithelial cells

Journal: Toxicon : official journal of the International Society on Toxinology

doi: 10.1016/j.toxicon.2016.03.007

Alveolar type II epithelial cells (ATII cells) were treated with MC-LR at 0, 0.5, 5, 50 or 500 nM for 48 h. (A) The expression of occludin was measured by immunofluorescent microscopy (× 600). (B) The expression of CK18, CK19 and SP-C in ATII
Figure Legend Snippet: Alveolar type II epithelial cells (ATII cells) were treated with MC-LR at 0, 0.5, 5, 50 or 500 nM for 48 h. (A) The expression of occludin was measured by immunofluorescent microscopy (× 600). (B) The expression of CK18, CK19 and SP-C in ATII

Techniques Used: Expressing, Microscopy

Mice were administered with MC-LR at 1 μg/L, 10 μg/L, and 40 μg/L in drinking water for 6 consecutive months. Mice were sacrificed and lungs were obtained. (A) Lung tissue expression of occludin was measured by immunofluorescent
Figure Legend Snippet: Mice were administered with MC-LR at 1 μg/L, 10 μg/L, and 40 μg/L in drinking water for 6 consecutive months. Mice were sacrificed and lungs were obtained. (A) Lung tissue expression of occludin was measured by immunofluorescent

Techniques Used: Mouse Assay, Expressing

5) Product Images from "Nonsteroidal Anti-inflammatory Drugs Alter the Microbiota and Exacerbate Clostridium difficile Colitis while Dysregulating the Inflammatory Response"

Article Title: Nonsteroidal Anti-inflammatory Drugs Alter the Microbiota and Exacerbate Clostridium difficile Colitis while Dysregulating the Inflammatory Response

Journal: mBio

doi: 10.1128/mBio.02282-18

Indomethacin promotes relocalization of TJ-associated protein ZO1 and perturbs colonic epithelial cell junctions of C. difficile -infected mice. (A) Transmission electron micrographs showing lateral views of colonic mucosa from untreated control mice (Mock) or mice treated with the following: cefoperazone alone (Abx); cefoperazone and indomethacin (Abx + Indo); cefoperazone and C. difficile (Abx + CD); or cefoperazone, indomethacin, and C. difficile (Abx + Indo + CD). Arrows point to intact tight junctions (TJ). Red arrowheads point to TJ unzipping or separation. (B and C) Mouse colonic tissues from the groups described above were stained for TJ-protein occludin (B) and TJ-associated protein ZO1 (C). Occludin and ZO1 stain are pseudocolored in red. DAPI (blue) was used to stain DNA. Yellow arrowheads and white arrowheads indicate cytoplasmic relocalization of occludin and ZO1, respectively (see also Fig. S1 ).
Figure Legend Snippet: Indomethacin promotes relocalization of TJ-associated protein ZO1 and perturbs colonic epithelial cell junctions of C. difficile -infected mice. (A) Transmission electron micrographs showing lateral views of colonic mucosa from untreated control mice (Mock) or mice treated with the following: cefoperazone alone (Abx); cefoperazone and indomethacin (Abx + Indo); cefoperazone and C. difficile (Abx + CD); or cefoperazone, indomethacin, and C. difficile (Abx + Indo + CD). Arrows point to intact tight junctions (TJ). Red arrowheads point to TJ unzipping or separation. (B and C) Mouse colonic tissues from the groups described above were stained for TJ-protein occludin (B) and TJ-associated protein ZO1 (C). Occludin and ZO1 stain are pseudocolored in red. DAPI (blue) was used to stain DNA. Yellow arrowheads and white arrowheads indicate cytoplasmic relocalization of occludin and ZO1, respectively (see also Fig. S1 ).

Techniques Used: Infection, Mouse Assay, Transmission Assay, Staining

6) Product Images from "Intestinal Epithelial Cell-Specific Deletion of PLD2 Alleviates DSS-Induced Colitis by Regulating Occludin"

Article Title: Intestinal Epithelial Cell-Specific Deletion of PLD2 Alleviates DSS-Induced Colitis by Regulating Occludin

Journal: Scientific Reports

doi: 10.1038/s41598-017-01797-y

PLD2 is involved in DSS-induced downregulation of occludin. ( A ) Western blots showing the effect of DSS treatment on the level of PLD2 in HT-29 colon epithelial cells at the indicated time points. Quantitative data showing the relative PLD2 and occludin expression at 8, 12, and 24 hours. ( B ) Effects of siRNA-mediated PLD2 knockdown in HT-29 colon epithelial cells ( C ) HT-29 colon epithelial cells were transfected with control, or PLD2 vector and PLD2, and occludin expression was analysed at 48 and 72 hours. Graph showing relative protein expression. Panel A–C: PLD2 expression of untreated (NT) vs. the indicated time point, or specified treatment group (*p
Figure Legend Snippet: PLD2 is involved in DSS-induced downregulation of occludin. ( A ) Western blots showing the effect of DSS treatment on the level of PLD2 in HT-29 colon epithelial cells at the indicated time points. Quantitative data showing the relative PLD2 and occludin expression at 8, 12, and 24 hours. ( B ) Effects of siRNA-mediated PLD2 knockdown in HT-29 colon epithelial cells ( C ) HT-29 colon epithelial cells were transfected with control, or PLD2 vector and PLD2, and occludin expression was analysed at 48 and 72 hours. Graph showing relative protein expression. Panel A–C: PLD2 expression of untreated (NT) vs. the indicated time point, or specified treatment group (*p

Techniques Used: Western Blot, Expressing, Transfection, Plasmid Preparation

Mechanism of epithelial barrier integrity regulation by PLD2. DSS treatment upregulates the expression of PLD2 in colon epithelial cells, which activates c-SRC. c-SRC is a tyrosine kinase that interacts with the tight junction protein occludin. Activated c-Src phosphorylates occludin at the tyrosine residues and targets it for proteasome-mediated degradation. Dissociation of occludin from the membrane causes an increase in barrier permeability, which leads to increased inflammation. Graphical illustration was drawn using the images from Servier Medical Art by Servier with slight modifications ( http://www.servier.com/Powerpoint-image-bank , https://creativecommons.org/licenses/by/3.0/ ).
Figure Legend Snippet: Mechanism of epithelial barrier integrity regulation by PLD2. DSS treatment upregulates the expression of PLD2 in colon epithelial cells, which activates c-SRC. c-SRC is a tyrosine kinase that interacts with the tight junction protein occludin. Activated c-Src phosphorylates occludin at the tyrosine residues and targets it for proteasome-mediated degradation. Dissociation of occludin from the membrane causes an increase in barrier permeability, which leads to increased inflammation. Graphical illustration was drawn using the images from Servier Medical Art by Servier with slight modifications ( http://www.servier.com/Powerpoint-image-bank , https://creativecommons.org/licenses/by/3.0/ ).

Techniques Used: Expressing, Permeability

Intestine-specific knockdown of PLD2 decreases occludin expression in mice. ( A ) In-vivo permeability assay in control and IEC KO mice, conducted after 4 hours of treatment with FITC-dextran, administered by oral gavage. FITC concentration in the serum was measured using a spectrofluorometer and is shown as the mean ± SEM. ( B ) Immunohistochemistry assessing the expression of occludin in the paraformaldehyde-fixed paraffin-embedded sections of colon tissue from control and IEC KO mice. Scale bar is 50 µm. ( C ) Western blots of occludin in isolated colon epithelial cells from the control and IEC KO mice. ( D ) Associated western blot quantification. All data are shown as the mean ± SEM. *p
Figure Legend Snippet: Intestine-specific knockdown of PLD2 decreases occludin expression in mice. ( A ) In-vivo permeability assay in control and IEC KO mice, conducted after 4 hours of treatment with FITC-dextran, administered by oral gavage. FITC concentration in the serum was measured using a spectrofluorometer and is shown as the mean ± SEM. ( B ) Immunohistochemistry assessing the expression of occludin in the paraformaldehyde-fixed paraffin-embedded sections of colon tissue from control and IEC KO mice. Scale bar is 50 µm. ( C ) Western blots of occludin in isolated colon epithelial cells from the control and IEC KO mice. ( D ) Associated western blot quantification. All data are shown as the mean ± SEM. *p

Techniques Used: Expressing, Mouse Assay, In Vivo, Permeability, Concentration Assay, Immunohistochemistry, Western Blot, Isolation

PLD2 regulates occludin levels by mediating the proteasome-mediated degradation of occludin via a Src kinase-dependent pathway. ( A ) HT-29 cells transfected with PLD2 siRNA were treated with 2% DSS and 10 nM MG132 to inhibit protein degradation; PLD2 and occludin expression were analysed by western blotting at the indicated time points. ( B ) HT-29 cells, transfected with control PLD2 siRNA, were treated with 2% DSS and 10 µg/mL cycloheximide (CHX) to inhibit protein degradation; PLD2 and occludin expression were analysed by western blotting at the indicated time points. ( C ) Occludin was immunoprecipitated after HT-29 colon epithelial cells were treated with 2% DSS; cell lysates were immunoblotted for phospho-tyrosine to assess the level of phospho-occludin at the indicated time points. PLD2 and occludin expression is shown. ( D ) HT-29 cells were treated with 2% DSS and the expression of PLD2, occludin, c-Src kinase, and phosphor c-Src kinase, at the specified time points, was analysed using western blotting. PLD2 expression: *p
Figure Legend Snippet: PLD2 regulates occludin levels by mediating the proteasome-mediated degradation of occludin via a Src kinase-dependent pathway. ( A ) HT-29 cells transfected with PLD2 siRNA were treated with 2% DSS and 10 nM MG132 to inhibit protein degradation; PLD2 and occludin expression were analysed by western blotting at the indicated time points. ( B ) HT-29 cells, transfected with control PLD2 siRNA, were treated with 2% DSS and 10 µg/mL cycloheximide (CHX) to inhibit protein degradation; PLD2 and occludin expression were analysed by western blotting at the indicated time points. ( C ) Occludin was immunoprecipitated after HT-29 colon epithelial cells were treated with 2% DSS; cell lysates were immunoblotted for phospho-tyrosine to assess the level of phospho-occludin at the indicated time points. PLD2 and occludin expression is shown. ( D ) HT-29 cells were treated with 2% DSS and the expression of PLD2, occludin, c-Src kinase, and phosphor c-Src kinase, at the specified time points, was analysed using western blotting. PLD2 expression: *p

Techniques Used: Transfection, Expressing, Western Blot, Immunoprecipitation

7) Product Images from "Bacteroides fragilis Protects Against Antibiotic-Associated Diarrhea in Rats by Modulating Intestinal Defenses"

Article Title: Bacteroides fragilis Protects Against Antibiotic-Associated Diarrhea in Rats by Modulating Intestinal Defenses

Journal: Frontiers in Immunology

doi: 10.3389/fimmu.2018.01040

Bacteroides fragilis ZY-312 improves gut barrier integrity in antibiotic-associated diarrhea (AAD) rats. (A) Alcian blue staining for detecting goblet cells numbers were shown in normal rats, AAD rats treated with normal saline, ZY-312 [10 9 colony-forming units (CFU)], Bifico (70 mg/day), or Bifico (70 mg/day) combined with ZY-312 (10 8 CFU) on days 11 and 17. n = 8/group. (B) Immunohistochemistry of Muc2 protein located in colonic tissues was detected in all groups as mentioned above. (C) Immunofluorescence staining for ZO-1 and ZO-1 ( TJP1 ) mRNA level in all the groups was shown as mentioned above. The magnifications of the above figures are 10× and 40×. (D) Colon protein level and mRNA expression of occludin in all groups on days 11 and 17 were shown. (E) qPCR analysis of AQP1, AQP3, and AQP8 gene in colonic tissues were detected in all groups as indicated in Figure 3 , with mRNA fold changes normalized to those of normal control group. One-way ANOVA, * P
Figure Legend Snippet: Bacteroides fragilis ZY-312 improves gut barrier integrity in antibiotic-associated diarrhea (AAD) rats. (A) Alcian blue staining for detecting goblet cells numbers were shown in normal rats, AAD rats treated with normal saline, ZY-312 [10 9 colony-forming units (CFU)], Bifico (70 mg/day), or Bifico (70 mg/day) combined with ZY-312 (10 8 CFU) on days 11 and 17. n = 8/group. (B) Immunohistochemistry of Muc2 protein located in colonic tissues was detected in all groups as mentioned above. (C) Immunofluorescence staining for ZO-1 and ZO-1 ( TJP1 ) mRNA level in all the groups was shown as mentioned above. The magnifications of the above figures are 10× and 40×. (D) Colon protein level and mRNA expression of occludin in all groups on days 11 and 17 were shown. (E) qPCR analysis of AQP1, AQP3, and AQP8 gene in colonic tissues were detected in all groups as indicated in Figure 3 , with mRNA fold changes normalized to those of normal control group. One-way ANOVA, * P

Techniques Used: Staining, Immunohistochemistry, Immunofluorescence, Expressing, Real-time Polymerase Chain Reaction

8) Product Images from "Characterization of the intestinal graft in a swine hypotensive after brain death model 1"

Article Title: Characterization of the intestinal graft in a swine hypotensive after brain death model 1

Journal: Acta Cirúrgica Brasileira

doi: 10.1590/s0102-865020190110000007

The expression of ZO-1 and occludin proteins in small bowel demonstrated by western blot analysis. Levels of the indicated protein at the baseline, T4, T8 and T12.
Figure Legend Snippet: The expression of ZO-1 and occludin proteins in small bowel demonstrated by western blot analysis. Levels of the indicated protein at the baseline, T4, T8 and T12.

Techniques Used: Expressing, Western Blot

9) Product Images from "Placental ischemia in pregnant rats impairs cerebral blood flow autoregulation and increases blood–brain barrier permeability"

Article Title: Placental ischemia in pregnant rats impairs cerebral blood flow autoregulation and increases blood–brain barrier permeability

Journal: Physiological Reports

doi: 10.14814/phy2.12134

Expression of tight junction proteins in the (A) anterior cerebrum and (B) posterior cerebrum of placental ischemic rats. Quantitative analysis of Western blot for Claudin‐1, Occludin, and ZO‐1 normalized to beta‐actin or GAPDH. Blots are representative samples for respective groups – order of blots match the order of the bars. Bars represent Mean ± SEM. * P
Figure Legend Snippet: Expression of tight junction proteins in the (A) anterior cerebrum and (B) posterior cerebrum of placental ischemic rats. Quantitative analysis of Western blot for Claudin‐1, Occludin, and ZO‐1 normalized to beta‐actin or GAPDH. Blots are representative samples for respective groups – order of blots match the order of the bars. Bars represent Mean ± SEM. * P

Techniques Used: Expressing, Western Blot

10) Product Images from "Structural alterations of the intestinal epithelial barrier in Parkinson’s disease"

Article Title: Structural alterations of the intestinal epithelial barrier in Parkinson’s disease

Journal: Acta Neuropathologica Communications

doi: 10.1186/s40478-015-0196-0

Expression of TJs proteins in colonic biopsies from patients with Parkinson’s disease (PD) and control subjects (CTRL). Biopsies lysates (20 μg of protein per sample) were subjected to immunoblot analysis using antibodies against occludin and ZO-1 (A) . Beta-actin was used as a loading control. The optical densities of occludin (B) and ZO-1 (C) immunoreactive bands were measured, normalized to the optical densities of beta-actin immunoreactive bands in the same samples and expressed as percentages of controls. Data correspond to mean ± SEM of 11 samples for control subjects (CTRL) and 31 samples for Parkinson’s disease (PD) patients. Patients versus control, *: p
Figure Legend Snippet: Expression of TJs proteins in colonic biopsies from patients with Parkinson’s disease (PD) and control subjects (CTRL). Biopsies lysates (20 μg of protein per sample) were subjected to immunoblot analysis using antibodies against occludin and ZO-1 (A) . Beta-actin was used as a loading control. The optical densities of occludin (B) and ZO-1 (C) immunoreactive bands were measured, normalized to the optical densities of beta-actin immunoreactive bands in the same samples and expressed as percentages of controls. Data correspond to mean ± SEM of 11 samples for control subjects (CTRL) and 31 samples for Parkinson’s disease (PD) patients. Patients versus control, *: p

Techniques Used: Expressing

Localization of TJs proteins in the colonic mucosa of healthy controls (CTRL) and patients with Parkinson’s disease (PD). Representative photomicrographs of the colonic mucosa labeled with antibodies against ZO-1 (A, B) and occludin (C, D) in the colonic mucosa of control and PD patients; scale bar: 100 μm. High-magnification image of each area marked by red square; scale bar: 10 μm.
Figure Legend Snippet: Localization of TJs proteins in the colonic mucosa of healthy controls (CTRL) and patients with Parkinson’s disease (PD). Representative photomicrographs of the colonic mucosa labeled with antibodies against ZO-1 (A, B) and occludin (C, D) in the colonic mucosa of control and PD patients; scale bar: 100 μm. High-magnification image of each area marked by red square; scale bar: 10 μm.

Techniques Used: Labeling

11) Product Images from "Recombinant Human Sonic Hedgehog Protein Regulates the Expression of ZO-1 and Occludin by Activating Angiopoietin-1 in Stroke Damage"

Article Title: Recombinant Human Sonic Hedgehog Protein Regulates the Expression of ZO-1 and Occludin by Activating Angiopoietin-1 in Stroke Damage

Journal: PLoS ONE

doi: 10.1371/journal.pone.0068891

Shh Treatment Upregulates the Expression of ZO-1 and Occludin In the Brain Tissue of Ischemic Rats. After establishment of pMCAO, rats were intracerebroventricularly injected 3 µL of PBS, 3 µg of Shh, and Shh (3 µg) plus cyclopamine (Cyc, 30 µg). The brain slices were staining by using a 2% TTC solution, and we selected the ischemic penumbra for the ensuing experiments. The expression of ZO-1, occludin and claudin-5 from ischemic penumbra was detected by Western blotting and real-time RT-PCR at indicated time points (1 d, 3 d, and 7 d). (A) Representative photographs of TTC stained brain slice. Brain samples were selected from ischemic penumbra. (B-D) The mRNA levels of ZO-1 (B), occludin (C) and claudin-5 (D). (E) Representative photographs of Western Blotting bands. (F-G) The IDV of ZO-1, occludin, and claudin-5. Grouping was the same as above. Δ P
Figure Legend Snippet: Shh Treatment Upregulates the Expression of ZO-1 and Occludin In the Brain Tissue of Ischemic Rats. After establishment of pMCAO, rats were intracerebroventricularly injected 3 µL of PBS, 3 µg of Shh, and Shh (3 µg) plus cyclopamine (Cyc, 30 µg). The brain slices were staining by using a 2% TTC solution, and we selected the ischemic penumbra for the ensuing experiments. The expression of ZO-1, occludin and claudin-5 from ischemic penumbra was detected by Western blotting and real-time RT-PCR at indicated time points (1 d, 3 d, and 7 d). (A) Representative photographs of TTC stained brain slice. Brain samples were selected from ischemic penumbra. (B-D) The mRNA levels of ZO-1 (B), occludin (C) and claudin-5 (D). (E) Representative photographs of Western Blotting bands. (F-G) The IDV of ZO-1, occludin, and claudin-5. Grouping was the same as above. Δ P

Techniques Used: Expressing, Injection, Staining, Western Blot, Quantitative RT-PCR, Slice Preparation

Shh Treatment Upregulates the Expression of ZO-1 and Occludin in BMECs under OGD. BMECs were pre-treated with PBS, Shh and/or cyclopamine (Cyc) for 30 min, and then subjected to the OGD for 4 h. (A-C) The mRNA levels of ZO-1 (A), occludin (B) and claudin-5 (C) were determined by real-time RT-PCR. Ctrl: cells were pre-treated with PBS and then subjected to normal oxygen condition. Cyc: cells were pre-treated with Cyc and then subjected to normal oxygen condition. OGD: cells were pre-treated with PBS and then subjected to OGD. Shh+OGD: cells were pre-treated with Shh and then subjected to OGD. Cyc+Shh+OGD: cells were pre-treated with Shh plus Cyc and then subjected to OGD. Δ P
Figure Legend Snippet: Shh Treatment Upregulates the Expression of ZO-1 and Occludin in BMECs under OGD. BMECs were pre-treated with PBS, Shh and/or cyclopamine (Cyc) for 30 min, and then subjected to the OGD for 4 h. (A-C) The mRNA levels of ZO-1 (A), occludin (B) and claudin-5 (C) were determined by real-time RT-PCR. Ctrl: cells were pre-treated with PBS and then subjected to normal oxygen condition. Cyc: cells were pre-treated with Cyc and then subjected to normal oxygen condition. OGD: cells were pre-treated with PBS and then subjected to OGD. Shh+OGD: cells were pre-treated with Shh and then subjected to OGD. Cyc+Shh+OGD: cells were pre-treated with Shh plus Cyc and then subjected to OGD. Δ P

Techniques Used: Expressing, Quantitative RT-PCR

Treatment with Ang-1-neutralizing Antibody Suppresses Shh-up-regulated ZO-1 and Occludin in BMECs under OGD. BMECs were pre-treated with Ang-1-neutralizing antibody (AngAb), and/or PBS, Shh for 30 min, and then subjected to the OGD for 4 h. (A-C) The mRNA levels of ZO-1 (A), occludin (B) and claudin-5 (C) were determined by real-time RT-PCR. Ctrl: cells were pre-treated with PBS and then subjected to normal oxygen condition. AngAb: cells were pre-treated with AngAb and then subjected to normal oxygen condition. OGD: cells were pre-treated with PBS and then subjected to OGD. Shh+OGD: cells were pre-treated with Shh and then subjected to OGD. AngAb+Shh+OGD: cells were pre-treated with Shh plus AngAb and then subjected to OGD. Δ P
Figure Legend Snippet: Treatment with Ang-1-neutralizing Antibody Suppresses Shh-up-regulated ZO-1 and Occludin in BMECs under OGD. BMECs were pre-treated with Ang-1-neutralizing antibody (AngAb), and/or PBS, Shh for 30 min, and then subjected to the OGD for 4 h. (A-C) The mRNA levels of ZO-1 (A), occludin (B) and claudin-5 (C) were determined by real-time RT-PCR. Ctrl: cells were pre-treated with PBS and then subjected to normal oxygen condition. AngAb: cells were pre-treated with AngAb and then subjected to normal oxygen condition. OGD: cells were pre-treated with PBS and then subjected to OGD. Shh+OGD: cells were pre-treated with Shh and then subjected to OGD. AngAb+Shh+OGD: cells were pre-treated with Shh plus AngAb and then subjected to OGD. Δ P

Techniques Used: Quantitative RT-PCR

12) Product Images from "Blood-brain barrier disruption induced by diagnostic ultrasound combined with microbubbles in mice"

Article Title: Blood-brain barrier disruption induced by diagnostic ultrasound combined with microbubbles in mice

Journal: Oncotarget

doi: 10.18632/oncotarget.23527

Representative blots ( A ) and relative quantitative analysis ( B , C , D ) of TJ related proteins ZO-1, occludin and claudin-5 expression in each group. Data were shown as the mean ± SEM, * P
Figure Legend Snippet: Representative blots ( A ) and relative quantitative analysis ( B , C , D ) of TJ related proteins ZO-1, occludin and claudin-5 expression in each group. Data were shown as the mean ± SEM, * P

Techniques Used: Expressing

Distribution and expression level of TJ related proteins ZO-1 ( A ), occludin ( C ) and claudin-5 ( E ) observed via immunohistofluorescence staining in each group. Relative fluorescence intensity of ZO-1 ( B ), occludin ( D ) and claudin-5 ( F ) compare with the control group. Data were shown as the mean ± SEM, * P
Figure Legend Snippet: Distribution and expression level of TJ related proteins ZO-1 ( A ), occludin ( C ) and claudin-5 ( E ) observed via immunohistofluorescence staining in each group. Relative fluorescence intensity of ZO-1 ( B ), occludin ( D ) and claudin-5 ( F ) compare with the control group. Data were shown as the mean ± SEM, * P

Techniques Used: Expressing, Immunohistofluorescence, Staining, Fluorescence

13) Product Images from "Noncentrosomal microtubules and type II myosins potentiate epidermal cell adhesion and barrier formation"

Article Title: Noncentrosomal microtubules and type II myosins potentiate epidermal cell adhesion and barrier formation

Journal: The Journal of Cell Biology

doi: 10.1083/jcb.201206143

Tight junction barrier activity is increased by cortical microtubules. (A) Transepithelial resistance (TER) measurements were taken of wild-type cells, cells treated with taxol, cells treated with E-cadherin inhibitory DECMA antibodies (with and without taxol), and in blebbistatin-treated cells (with and without taxol). P = 0.0037 for control vs. taxol treatment; n = 9. (B) TER levels in myosin IIA WT and null cells (with and without taxol). P = 0.007. (C and D) In vivo epidermal barrier assay. DMSO or nocodazole was injected subcutaneously with a biotin tracer. After 30 min the skin was removed, embedded, and sectioned for analysis. Streptavidin-FITC (green) allowed visualization of the diffusion of the biotin, and occludin (red) puncta mark the tight junctions in the granular layer of the epidermis. Bar, 10 µm (5 µm for insets).
Figure Legend Snippet: Tight junction barrier activity is increased by cortical microtubules. (A) Transepithelial resistance (TER) measurements were taken of wild-type cells, cells treated with taxol, cells treated with E-cadherin inhibitory DECMA antibodies (with and without taxol), and in blebbistatin-treated cells (with and without taxol). P = 0.0037 for control vs. taxol treatment; n = 9. (B) TER levels in myosin IIA WT and null cells (with and without taxol). P = 0.007. (C and D) In vivo epidermal barrier assay. DMSO or nocodazole was injected subcutaneously with a biotin tracer. After 30 min the skin was removed, embedded, and sectioned for analysis. Streptavidin-FITC (green) allowed visualization of the diffusion of the biotin, and occludin (red) puncta mark the tight junctions in the granular layer of the epidermis. Bar, 10 µm (5 µm for insets).

Techniques Used: Activity Assay, In Vivo, Injection, Diffusion-based Assay

Loss of myosin IIA and B results in tight junction defects in the epidermis. (A) Barrier assay in E18.5 control littermate (left) and myosin IIA/B double cKO epidermis. Note the barrier defects on digits, eyes, and ears, and normal barrier function over the rest of the body. (B and C) Normal expression of differentiation markers: K5/14 (green) and K1 (red) in control (B) and myosin II dKO epidermis (C). (D and E) Normal expression of granular layer marker loricrin (green) in control (D) and myosin II dKO epidermis (E). β4-integrin (red) marks the basement membrane in these images. (F and G) Abnormal expression of the stress marker keratin 6 (K6, red) in myosin II dKO epidermis (G). Asterisks mark nonspecific cornified envelope staining in F and G. (H and I) Biotin diffusion assay in control (H) and myosin II dKO (I) epidermis. Biotin is detected with streptavidin (green) and tight junctions are marked with occludin (red). (J and K) Adherens junction components E-cadherin (green) and α-catenin (red) in control (J) and myosin II dKO epidermis (K). (L) Extracts from Myosin IIA/B dKO epidermis or littermate controls were immunoprecipitated with anti–β-catenin antibodies. Bound proteins were analyzed by Western blotting with vinculin and β-catenin. Bars, 10 µm.
Figure Legend Snippet: Loss of myosin IIA and B results in tight junction defects in the epidermis. (A) Barrier assay in E18.5 control littermate (left) and myosin IIA/B double cKO epidermis. Note the barrier defects on digits, eyes, and ears, and normal barrier function over the rest of the body. (B and C) Normal expression of differentiation markers: K5/14 (green) and K1 (red) in control (B) and myosin II dKO epidermis (C). (D and E) Normal expression of granular layer marker loricrin (green) in control (D) and myosin II dKO epidermis (E). β4-integrin (red) marks the basement membrane in these images. (F and G) Abnormal expression of the stress marker keratin 6 (K6, red) in myosin II dKO epidermis (G). Asterisks mark nonspecific cornified envelope staining in F and G. (H and I) Biotin diffusion assay in control (H) and myosin II dKO (I) epidermis. Biotin is detected with streptavidin (green) and tight junctions are marked with occludin (red). (J and K) Adherens junction components E-cadherin (green) and α-catenin (red) in control (J) and myosin II dKO epidermis (K). (L) Extracts from Myosin IIA/B dKO epidermis or littermate controls were immunoprecipitated with anti–β-catenin antibodies. Bound proteins were analyzed by Western blotting with vinculin and β-catenin. Bars, 10 µm.

Techniques Used: Expressing, Marker, Staining, Diffusion-based Assay, Immunoprecipitation, Western Blot

14) Product Images from "Akkermansia muciniphila-derived extracellular vesicles influence gut permeability through the regulation of tight junctions"

Article Title: Akkermansia muciniphila-derived extracellular vesicles influence gut permeability through the regulation of tight junctions

Journal: Experimental & Molecular Medicine

doi: 10.1038/emm.2017.282

A. muciniphila -derived EVs improve gut permeability in high-fat diet (HFD)-fed mice. Mice were fed a normal chow diet (NCD) or a HFD for 12 weeks, followed by oral administration of AmEVs (10 μg per mouse) every other day for 2 weeks. ( a ) Body weight changes were measured at the indicated time points. The graph shows body weight change from the day that EV feeding started. ( b ) In vivo intestinal permeability assay in NCD, NCD+AmEV, HFD and HFD+AmEV mice conducted after 4 h of treatment with FITC-dextran. The serum FITC concentration was measured using blood collected by retro-orbital bleeding, and fluorescence was measured using a spectrofluorometer. Data are shown as the mean±s.e.m. ( c ) Gross imaging of the colons dissected from NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. ( d ) The colon length of NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. ( e ) Hematoxylin and eosin staining of colon sections from NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. Red arrows indicate the recruitment of immune cells. Scale bar is 50 μm. ( f ) Immunohistochemical images showing occludin expression (occludin—red, nucleus—blue) in NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. Scale bar is 50 μm. All data are presented as the mean±s.e.m. of 3 experiments; n =5–7 per group; * P
Figure Legend Snippet: A. muciniphila -derived EVs improve gut permeability in high-fat diet (HFD)-fed mice. Mice were fed a normal chow diet (NCD) or a HFD for 12 weeks, followed by oral administration of AmEVs (10 μg per mouse) every other day for 2 weeks. ( a ) Body weight changes were measured at the indicated time points. The graph shows body weight change from the day that EV feeding started. ( b ) In vivo intestinal permeability assay in NCD, NCD+AmEV, HFD and HFD+AmEV mice conducted after 4 h of treatment with FITC-dextran. The serum FITC concentration was measured using blood collected by retro-orbital bleeding, and fluorescence was measured using a spectrofluorometer. Data are shown as the mean±s.e.m. ( c ) Gross imaging of the colons dissected from NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. ( d ) The colon length of NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. ( e ) Hematoxylin and eosin staining of colon sections from NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. Red arrows indicate the recruitment of immune cells. Scale bar is 50 μm. ( f ) Immunohistochemical images showing occludin expression (occludin—red, nucleus—blue) in NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. Scale bar is 50 μm. All data are presented as the mean±s.e.m. of 3 experiments; n =5–7 per group; * P

Techniques Used: Derivative Assay, Permeability, Mouse Assay, In Vivo, Concentration Assay, Fluorescence, Imaging, Staining, Immunohistochemistry, Expressing

A. muciniphila -derived EVs reduce LPS-induced intestinal permeability through AMPK phosphorylation. ( a ) Trans-epithelial electrical resistance in Caco-2 cells 4 h after treatment with LPS (5 μg ml −1 ), LPS (5 μg ml −1 )+EcEVs (1 μg ml −1 ) and LPS (5 μg ml −1 )+AmEVs (1 μg ml −1 ). ( b ) In vitro permeability assay in Caco-2 cells 4 h after LPS (5 μg ml −1 ), LPS (5 μg ml −1 )+EcEV (1 μg ml −1 ) and LPS (5 μg ml −1 )+AmEV (1 μg ml −1 ) treatment. FITC-dextran was added to the upper chamber in the Caco-2 transwell chamber assay, and the transport of FITC-labeled dextran across the Caco-2 monolayer culture was measured by fluorescence spectrofluorometry. ( c ) Expression of tight junction proteins in Caco-2 cells after LPS, LPS+EcEV and LPS+AmEV treatments. A graph showing the relative occludin expression (normalized to actin) is also shown (* P
Figure Legend Snippet: A. muciniphila -derived EVs reduce LPS-induced intestinal permeability through AMPK phosphorylation. ( a ) Trans-epithelial electrical resistance in Caco-2 cells 4 h after treatment with LPS (5 μg ml −1 ), LPS (5 μg ml −1 )+EcEVs (1 μg ml −1 ) and LPS (5 μg ml −1 )+AmEVs (1 μg ml −1 ). ( b ) In vitro permeability assay in Caco-2 cells 4 h after LPS (5 μg ml −1 ), LPS (5 μg ml −1 )+EcEV (1 μg ml −1 ) and LPS (5 μg ml −1 )+AmEV (1 μg ml −1 ) treatment. FITC-dextran was added to the upper chamber in the Caco-2 transwell chamber assay, and the transport of FITC-labeled dextran across the Caco-2 monolayer culture was measured by fluorescence spectrofluorometry. ( c ) Expression of tight junction proteins in Caco-2 cells after LPS, LPS+EcEV and LPS+AmEV treatments. A graph showing the relative occludin expression (normalized to actin) is also shown (* P

Techniques Used: Derivative Assay, Permeability, In Vitro, Transwell Chamber Assay, Labeling, Fluorescence, Expressing

15) Product Images from "The toxic effects of microcystin-LR on mouse lungs and alveolar type II epithelial cells"

Article Title: The toxic effects of microcystin-LR on mouse lungs and alveolar type II epithelial cells

Journal: Toxicon : official journal of the International Society on Toxinology

doi: 10.1016/j.toxicon.2016.03.007

Alveolar type II epithelial cells (ATII cells) were treated with MC-LR at 0, 0.5, 5, 50 or 500 nM for 48 h. (A) The expression of occludin was measured by immunofluorescent microscopy (× 600). (B) The expression of CK18, CK19 and SP-C in ATII
Figure Legend Snippet: Alveolar type II epithelial cells (ATII cells) were treated with MC-LR at 0, 0.5, 5, 50 or 500 nM for 48 h. (A) The expression of occludin was measured by immunofluorescent microscopy (× 600). (B) The expression of CK18, CK19 and SP-C in ATII

Techniques Used: Expressing, Microscopy

Mice were administered with MC-LR at 1 μg/L, 10 μg/L, and 40 μg/L in drinking water for 6 consecutive months. Mice were sacrificed and lungs were obtained. (A) Lung tissue expression of occludin was measured by immunofluorescent
Figure Legend Snippet: Mice were administered with MC-LR at 1 μg/L, 10 μg/L, and 40 μg/L in drinking water for 6 consecutive months. Mice were sacrificed and lungs were obtained. (A) Lung tissue expression of occludin was measured by immunofluorescent

Techniques Used: Mouse Assay, Expressing

16) Product Images from "Akkermansia muciniphila-derived extracellular vesicles influence gut permeability through the regulation of tight junctions"

Article Title: Akkermansia muciniphila-derived extracellular vesicles influence gut permeability through the regulation of tight junctions

Journal: Experimental & Molecular Medicine

doi: 10.1038/emm.2017.282

A. muciniphila -derived EVs improve gut permeability in high-fat diet (HFD)-fed mice. Mice were fed a normal chow diet (NCD) or a HFD for 12 weeks, followed by oral administration of AmEVs (10 μg per mouse) every other day for 2 weeks. ( a ) Body weight changes were measured at the indicated time points. The graph shows body weight change from the day that EV feeding started. ( b ) In vivo intestinal permeability assay in NCD, NCD+AmEV, HFD and HFD+AmEV mice conducted after 4 h of treatment with FITC-dextran. The serum FITC concentration was measured using blood collected by retro-orbital bleeding, and fluorescence was measured using a spectrofluorometer. Data are shown as the mean±s.e.m. ( c ) Gross imaging of the colons dissected from NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. ( d ) The colon length of NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. ( e ) Hematoxylin and eosin staining of colon sections from NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. Red arrows indicate the recruitment of immune cells. Scale bar is 50 μm. ( f ) Immunohistochemical images showing occludin expression (occludin—red, nucleus—blue) in NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. Scale bar is 50 μm. All data are presented as the mean±s.e.m. of 3 experiments; n =5–7 per group; * P
Figure Legend Snippet: A. muciniphila -derived EVs improve gut permeability in high-fat diet (HFD)-fed mice. Mice were fed a normal chow diet (NCD) or a HFD for 12 weeks, followed by oral administration of AmEVs (10 μg per mouse) every other day for 2 weeks. ( a ) Body weight changes were measured at the indicated time points. The graph shows body weight change from the day that EV feeding started. ( b ) In vivo intestinal permeability assay in NCD, NCD+AmEV, HFD and HFD+AmEV mice conducted after 4 h of treatment with FITC-dextran. The serum FITC concentration was measured using blood collected by retro-orbital bleeding, and fluorescence was measured using a spectrofluorometer. Data are shown as the mean±s.e.m. ( c ) Gross imaging of the colons dissected from NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. ( d ) The colon length of NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. ( e ) Hematoxylin and eosin staining of colon sections from NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. Red arrows indicate the recruitment of immune cells. Scale bar is 50 μm. ( f ) Immunohistochemical images showing occludin expression (occludin—red, nucleus—blue) in NCD-, NCD+AmEV-, HFD- and HFD+AmEV-fed mice. Scale bar is 50 μm. All data are presented as the mean±s.e.m. of 3 experiments; n =5–7 per group; * P

Techniques Used: Derivative Assay, Permeability, Mouse Assay, In Vivo, Concentration Assay, Fluorescence, Imaging, Staining, Immunohistochemistry, Expressing

A. muciniphila -derived EVs reduce LPS-induced intestinal permeability through AMPK phosphorylation. ( a ) Trans-epithelial electrical resistance in Caco-2 cells 4 h after treatment with LPS (5 μg ml −1 ), LPS (5 μg ml −1 )+EcEVs (1 μg ml −1 ) and LPS (5 μg ml −1 )+AmEVs (1 μg ml −1 ). ( b ) In vitro permeability assay in Caco-2 cells 4 h after LPS (5 μg ml −1 ), LPS (5 μg ml −1 )+EcEV (1 μg ml −1 ) and LPS (5 μg ml −1 )+AmEV (1 μg ml −1 ) treatment. FITC-dextran was added to the upper chamber in the Caco-2 transwell chamber assay, and the transport of FITC-labeled dextran across the Caco-2 monolayer culture was measured by fluorescence spectrofluorometry. ( c ) Expression of tight junction proteins in Caco-2 cells after LPS, LPS+EcEV and LPS+AmEV treatments. A graph showing the relative occludin expression (normalized to actin) is also shown (* P
Figure Legend Snippet: A. muciniphila -derived EVs reduce LPS-induced intestinal permeability through AMPK phosphorylation. ( a ) Trans-epithelial electrical resistance in Caco-2 cells 4 h after treatment with LPS (5 μg ml −1 ), LPS (5 μg ml −1 )+EcEVs (1 μg ml −1 ) and LPS (5 μg ml −1 )+AmEVs (1 μg ml −1 ). ( b ) In vitro permeability assay in Caco-2 cells 4 h after LPS (5 μg ml −1 ), LPS (5 μg ml −1 )+EcEV (1 μg ml −1 ) and LPS (5 μg ml −1 )+AmEV (1 μg ml −1 ) treatment. FITC-dextran was added to the upper chamber in the Caco-2 transwell chamber assay, and the transport of FITC-labeled dextran across the Caco-2 monolayer culture was measured by fluorescence spectrofluorometry. ( c ) Expression of tight junction proteins in Caco-2 cells after LPS, LPS+EcEV and LPS+AmEV treatments. A graph showing the relative occludin expression (normalized to actin) is also shown (* P

Techniques Used: Derivative Assay, Permeability, In Vitro, Transwell Chamber Assay, Labeling, Fluorescence, Expressing

17) Product Images from "Effects of Fullerenols on Mouse Brain Microvascular Endothelial Cells"

Article Title: Effects of Fullerenols on Mouse Brain Microvascular Endothelial Cells

Journal: International Journal of Molecular Sciences

doi: 10.3390/ijms18081783

Effect of fullerenols on the amount of junctional proteins in mouse brain microvascular endothelial cell (MBMEC) cultures under basal and inflammatory conditions. Western blot analysis and densitometric quantification (normalized to basal or inflamed untreated control cell cultures) of the amount of occludin ( A – C ), claudin 3 ( D – F ) and claudin 5 ( G – I ) proteins in MBMEC after exposure to interferon-γ and tumor necrosis factor-α (I + T; 100 IU each) and treatment with fullerenol (F; 1, 10 and 100 µg/mL; n = 4–5) for 18 h ( C , F , I ) compared to cultures under basal (homeostatic milieu) conditions ( B , E , H ). β-Actin was used as loading control. Ctrl, untreated control; Veh, vehicle treatment. * p
Figure Legend Snippet: Effect of fullerenols on the amount of junctional proteins in mouse brain microvascular endothelial cell (MBMEC) cultures under basal and inflammatory conditions. Western blot analysis and densitometric quantification (normalized to basal or inflamed untreated control cell cultures) of the amount of occludin ( A – C ), claudin 3 ( D – F ) and claudin 5 ( G – I ) proteins in MBMEC after exposure to interferon-γ and tumor necrosis factor-α (I + T; 100 IU each) and treatment with fullerenol (F; 1, 10 and 100 µg/mL; n = 4–5) for 18 h ( C , F , I ) compared to cultures under basal (homeostatic milieu) conditions ( B , E , H ). β-Actin was used as loading control. Ctrl, untreated control; Veh, vehicle treatment. * p

Techniques Used: Western Blot

Effect of fullerenols on the relative gene expression levels of tight junctions of mouse brain microvascular endothelial cell (MBMEC) cultures under basal and inflammatory conditions. Real time PCR analysis of the expression levels of occludin ( A ), claudin 3 ( B ), and claudin 5 ( C ) in MBMEC after exposure to interferon-γ and tumor necrosis factor-α (inflamed; I + T; 100 IU each) and treatment with fullerenol (F; 1, 10 and 100 µg/mL; n = 3−4) for 18 h compared to cultures under basal (homeostatic milieu) conditions. GAPDH was used as a housekeeping gene. Ctrl, untreated control; Veh, vehicle treatment.
Figure Legend Snippet: Effect of fullerenols on the relative gene expression levels of tight junctions of mouse brain microvascular endothelial cell (MBMEC) cultures under basal and inflammatory conditions. Real time PCR analysis of the expression levels of occludin ( A ), claudin 3 ( B ), and claudin 5 ( C ) in MBMEC after exposure to interferon-γ and tumor necrosis factor-α (inflamed; I + T; 100 IU each) and treatment with fullerenol (F; 1, 10 and 100 µg/mL; n = 3−4) for 18 h compared to cultures under basal (homeostatic milieu) conditions. GAPDH was used as a housekeeping gene. Ctrl, untreated control; Veh, vehicle treatment.

Techniques Used: Expressing, Real-time Polymerase Chain Reaction

18) Product Images from "Progressive Hearing Loss in Mice Carrying a Mutation in Usp53"

Article Title: Progressive Hearing Loss in Mice Carrying a Mutation in Usp53

Journal: The Journal of Neuroscience

doi: 10.1523/JNEUROSCI.1965-15.2015

USP53 interacts with TJP1 and TJP2 via its C-terminal tail. A , GFP–USP53 fusion constructs, depicted schematically in C , were injectoporated into cochlear explants from P4 wild-type (WT) mice. GFP fusion proteins containing the C-terminal tail of USP53 localized to TJs. B , GFP and GFP–USP53 fusion constructs were expressed in HEK293T cells and immunoprecipitated with anti-GFP antibody (Abcam). Extracts were directly analyzed for the expression of GFP-tagged USP53 constructs using anti-GFP antibody (Sigma). An asterisk marks a nonspecific cross-reacting band at ∼80 kDa used as a loading control. Native TJ proteins TJP1 and TJP2, but not occludin (OCLN), coprecipitated specifically with GFP fusion proteins that contain the C-terminal tail of USP53. IB, Immunoblot; IP, immunoprecipitation. C , Schematic diagram of the GFP–USP53 fusion proteins and TJPs used for biochemical experiments and their predicted molecular sizes. D , HEK293T cells were transfected with the constructs indicated on top of each panel. Immunoprecipitations were performed with anti-GFP antibody (Abcam) that recognizes GFP–USP53 fusion proteins followed by Western blotting with anti-FLAG antibody or anti-GFP antibody (Abcam). 2xFLAG–TJP2 coprecipitated with TJP2–GFP (TJP2 homodimerization), as well as with GFP–USP53 constructs containing a stretch of 328 aa adjacent to the catalytic domain. Neither the catalytic domain nor GFP alone precipitated 2xFLAG–TJP2. E , Schematic representation of C-terminal deletion constructs of USP53 used in D . The region containing the TJP2 interaction site is highlighted in blue. Scale bar: A , 5 μm.
Figure Legend Snippet: USP53 interacts with TJP1 and TJP2 via its C-terminal tail. A , GFP–USP53 fusion constructs, depicted schematically in C , were injectoporated into cochlear explants from P4 wild-type (WT) mice. GFP fusion proteins containing the C-terminal tail of USP53 localized to TJs. B , GFP and GFP–USP53 fusion constructs were expressed in HEK293T cells and immunoprecipitated with anti-GFP antibody (Abcam). Extracts were directly analyzed for the expression of GFP-tagged USP53 constructs using anti-GFP antibody (Sigma). An asterisk marks a nonspecific cross-reacting band at ∼80 kDa used as a loading control. Native TJ proteins TJP1 and TJP2, but not occludin (OCLN), coprecipitated specifically with GFP fusion proteins that contain the C-terminal tail of USP53. IB, Immunoblot; IP, immunoprecipitation. C , Schematic diagram of the GFP–USP53 fusion proteins and TJPs used for biochemical experiments and their predicted molecular sizes. D , HEK293T cells were transfected with the constructs indicated on top of each panel. Immunoprecipitations were performed with anti-GFP antibody (Abcam) that recognizes GFP–USP53 fusion proteins followed by Western blotting with anti-FLAG antibody or anti-GFP antibody (Abcam). 2xFLAG–TJP2 coprecipitated with TJP2–GFP (TJP2 homodimerization), as well as with GFP–USP53 constructs containing a stretch of 328 aa adjacent to the catalytic domain. Neither the catalytic domain nor GFP alone precipitated 2xFLAG–TJP2. E , Schematic representation of C-terminal deletion constructs of USP53 used in D . The region containing the TJP2 interaction site is highlighted in blue. Scale bar: A , 5 μm.

Techniques Used: Construct, Mouse Assay, Immunoprecipitation, Expressing, Transfection, Western Blot

19) Product Images from "Netrin‐1 Preserves Blood‐Brain Barrier Integrity Through Deleted in Colorectal Cancer/Focal Adhesion Kinase/RhoA Signaling Pathway Following Subarachnoid Hemorrhage in Rats"

Article Title: Netrin‐1 Preserves Blood‐Brain Barrier Integrity Through Deleted in Colorectal Cancer/Focal Adhesion Kinase/RhoA Signaling Pathway Following Subarachnoid Hemorrhage in Rats

Journal: Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease

doi: 10.1161/JAHA.116.005198

Changes in expressions of downstream signaling pathway of Focal Adhesion Kinase ( FAK )/RhoA after exogenous Netrin‐1 ( NTN ‐1) treatment and NTN ‐1 si RNA pretreatment at 24 hours post–subarachnoid hemorrhage ( SAH ). Exogenous NTN ‐1 significantly further enhanced the FAK phosphorylation (A), which resulted in the suppression of RhoA activity (B). While the endogenous NTN ‐1 was depleted by use of NTN ‐1 si RNA , the p‐ FAK level decreased (A), which augmented the RhoA activation (B). The RhoA activity was negatively related to the levels of endothelial tight junction proteins including ZO ‐1 (C) and Occludin (D) in the ipsilateral hemisphere. Relative densities of each protein have been normalized against the sham group. n=6 for each group. * P
Figure Legend Snippet: Changes in expressions of downstream signaling pathway of Focal Adhesion Kinase ( FAK )/RhoA after exogenous Netrin‐1 ( NTN ‐1) treatment and NTN ‐1 si RNA pretreatment at 24 hours post–subarachnoid hemorrhage ( SAH ). Exogenous NTN ‐1 significantly further enhanced the FAK phosphorylation (A), which resulted in the suppression of RhoA activity (B). While the endogenous NTN ‐1 was depleted by use of NTN ‐1 si RNA , the p‐ FAK level decreased (A), which augmented the RhoA activation (B). The RhoA activity was negatively related to the levels of endothelial tight junction proteins including ZO ‐1 (C) and Occludin (D) in the ipsilateral hemisphere. Relative densities of each protein have been normalized against the sham group. n=6 for each group. * P

Techniques Used: Activity Assay, Activation Assay

20) Product Images from "Dietary leonurine hydrochloride supplementation attenuates lipopolysaccharide challenge-induced intestinal inflammation and barrier dysfunction by inhibiting the NF-κB/MAPK signaling pathway in broilers"

Article Title: Dietary leonurine hydrochloride supplementation attenuates lipopolysaccharide challenge-induced intestinal inflammation and barrier dysfunction by inhibiting the NF-κB/MAPK signaling pathway in broilers

Journal: Journal of Animal Science

doi: 10.1093/jas/skz078

Expression of ZO-1 and Occludin proteins in the jejunal mucosa of broilers fed diets with or without 120 mg/kg leonurine hydrochloride after intraperitoneal administration of lipopolysaccharide or saline on day 21. (A) Representative western blots and bar diagrams showing densitometric analyses of (B) the ZO-1/β-actin ratio and (C) the Occludin/β-actin ratio. a,b Mean values with different letters differed significantly ( P
Figure Legend Snippet: Expression of ZO-1 and Occludin proteins in the jejunal mucosa of broilers fed diets with or without 120 mg/kg leonurine hydrochloride after intraperitoneal administration of lipopolysaccharide or saline on day 21. (A) Representative western blots and bar diagrams showing densitometric analyses of (B) the ZO-1/β-actin ratio and (C) the Occludin/β-actin ratio. a,b Mean values with different letters differed significantly ( P

Techniques Used: Expressing, Western Blot

21) Product Images from "A Selected Lactobacillus rhamnosus Strain Promotes EGFR-Independent Akt Activation in an Enterotoxigenic Escherichia coli K88-Infected IPEC-J2 Cell Model"

Article Title: A Selected Lactobacillus rhamnosus Strain Promotes EGFR-Independent Akt Activation in an Enterotoxigenic Escherichia coli K88-Infected IPEC-J2 Cell Model

Journal: PLoS ONE

doi: 10.1371/journal.pone.0125717

Western blot detection of tight junction proteins. (A) Representative panels of zonula occludens-1 (ZO-1) and occludin proteins in IPEC-J2 cells collected from the indicated cultures at 3 h after F4 + ETEC challenge. Expression of GAPDH was measured as an internal control. Results are presented as the ratio of the (B) ZO-1 band intensity and (C) occludin band intensity to the GAPDH band intensity. Data are presented as means ± SEM of three independent experiments. * P
Figure Legend Snippet: Western blot detection of tight junction proteins. (A) Representative panels of zonula occludens-1 (ZO-1) and occludin proteins in IPEC-J2 cells collected from the indicated cultures at 3 h after F4 + ETEC challenge. Expression of GAPDH was measured as an internal control. Results are presented as the ratio of the (B) ZO-1 band intensity and (C) occludin band intensity to the GAPDH band intensity. Data are presented as means ± SEM of three independent experiments. * P

Techniques Used: Western Blot, Expressing

22) Product Images from "Long-term exposure to ethanol downregulates tight junction proteins through the protein kinase Cα signaling pathway in human cerebral microvascular endothelial cells"

Article Title: Long-term exposure to ethanol downregulates tight junction proteins through the protein kinase Cα signaling pathway in human cerebral microvascular endothelial cells

Journal: Experimental and Therapeutic Medicine

doi: 10.3892/etm.2017.5180

Effect of ethanol treatment on the expression of claudin-5, occludin and ZO-1. Protein expression levels were measured using western blotting, and mRNA expression levels were determined using reverse transcription-quantitative polymerase chain reaction. (A) Western blotting analysis of claudin-5, occludin, ZO-1 and β-actin expression following ethanol treatment at (A) 24, (B) 48 and (C) 72 h. Quantitative analysis of western blotting results, which were normalized to β-actin expression at (D) 24, (E) 48 and (F) 72 h. Relative expression of claudin-5, occludin and ZO-1 mRNA at (G) 24, (H) 48 and (I) 72 h. The results (n=5) are presented as the mean ± standard deviation. *P
Figure Legend Snippet: Effect of ethanol treatment on the expression of claudin-5, occludin and ZO-1. Protein expression levels were measured using western blotting, and mRNA expression levels were determined using reverse transcription-quantitative polymerase chain reaction. (A) Western blotting analysis of claudin-5, occludin, ZO-1 and β-actin expression following ethanol treatment at (A) 24, (B) 48 and (C) 72 h. Quantitative analysis of western blotting results, which were normalized to β-actin expression at (D) 24, (E) 48 and (F) 72 h. Relative expression of claudin-5, occludin and ZO-1 mRNA at (G) 24, (H) 48 and (I) 72 h. The results (n=5) are presented as the mean ± standard deviation. *P

Techniques Used: Expressing, Western Blot, Real-time Polymerase Chain Reaction, Standard Deviation

Effect of the ethanol-induced increase in p-PKCα expression on claudin-5, occludin and ZO-1 expression. (A) Western blotting analysis of p-PKCα and β-actin protein expression. (B) Quantitative analysis of p-PKCα protein expression relative to β-actin expression. (C) Western blotting analysis of claudin-5, occludin and ZO-1 protein expression. (D) Quantitative analysis of claudin-5, occludin and ZO-1 protein expression relative to β-actin expression. The results (n=5) are presented as the mean ± standard deviation. *P
Figure Legend Snippet: Effect of the ethanol-induced increase in p-PKCα expression on claudin-5, occludin and ZO-1 expression. (A) Western blotting analysis of p-PKCα and β-actin protein expression. (B) Quantitative analysis of p-PKCα protein expression relative to β-actin expression. (C) Western blotting analysis of claudin-5, occludin and ZO-1 protein expression. (D) Quantitative analysis of claudin-5, occludin and ZO-1 protein expression relative to β-actin expression. The results (n=5) are presented as the mean ± standard deviation. *P

Techniques Used: Expressing, Western Blot, Standard Deviation

23) Product Images from "Effects of biomechanical forces on signaling in the cortical collecting duct (CCD)"

Article Title: Effects of biomechanical forces on signaling in the cortical collecting duct (CCD)

Journal: American Journal of Physiology - Renal Physiology

doi: 10.1152/ajprenal.00634.2013

Immunolocalization of occludin, ZO-1, and F-actin in murine cortical collecting duct (CCD; mpkCCD) cells exposed to fluid shear stress (FSS; 0.4 dyne/cm 2 , 30 min) or circumferential stretch (CS; 10% equibiaxial stretch, 30 min). The tight junction protein
Figure Legend Snippet: Immunolocalization of occludin, ZO-1, and F-actin in murine cortical collecting duct (CCD; mpkCCD) cells exposed to fluid shear stress (FSS; 0.4 dyne/cm 2 , 30 min) or circumferential stretch (CS; 10% equibiaxial stretch, 30 min). The tight junction protein

Techniques Used:

24) Product Images from "Endothelial Adora2a Activation Promotes Blood–Brain Barrier Breakdown and Cognitive Impairment in Mice with Diet-Induced Insulin Resistance"

Article Title: Endothelial Adora2a Activation Promotes Blood–Brain Barrier Breakdown and Cognitive Impairment in Mice with Diet-Induced Insulin Resistance

Journal: The Journal of Neuroscience

doi: 10.1523/JNEUROSCI.2506-18.2019

Progressive changes in BBB integrity with increasing durations of obesogenic diet consumption. A , Weight gain with exposure to a HFD or LFD in male mice. B , Fasting hyperglycemia (left) and fasting hyperinsulinemia (right) were evident after 16 weeks consumption of HFD. C , Increased penetration of the low molecular weight tracer NaFl was observed in clarified hippocampal lysates after 12 or 16 weeks HFD (left graph). Right graph shows increases in hippocampal Evans blue after 16 weeks HFD. D ). Scale bar: (in LFD 8WK ) for all panels, 10 μm. E , Reductions in hippocampal claudin-5 (left) and occludin (right) protein were detected by Western blotting after 16 weeks of HFD. F , Micrographs show representative images of tomato lectin labeling for the indicated diets and time points. Scale bar: (in LFD 12WK ) for all micrographs, 100 μm. G , Increases in vascular area (left) and tortuosity (right) in the dentate molecular layer with increasing durations of HFD. For all graphs, bar or symbol height represents the mean of ( n = 4–6) mice per diet at each time point and error bars represent SEM. *Indicates significant differences ( p
Figure Legend Snippet: Progressive changes in BBB integrity with increasing durations of obesogenic diet consumption. A , Weight gain with exposure to a HFD or LFD in male mice. B , Fasting hyperglycemia (left) and fasting hyperinsulinemia (right) were evident after 16 weeks consumption of HFD. C , Increased penetration of the low molecular weight tracer NaFl was observed in clarified hippocampal lysates after 12 or 16 weeks HFD (left graph). Right graph shows increases in hippocampal Evans blue after 16 weeks HFD. D ). Scale bar: (in LFD 8WK ) for all panels, 10 μm. E , Reductions in hippocampal claudin-5 (left) and occludin (right) protein were detected by Western blotting after 16 weeks of HFD. F , Micrographs show representative images of tomato lectin labeling for the indicated diets and time points. Scale bar: (in LFD 12WK ) for all micrographs, 100 μm. G , Increases in vascular area (left) and tortuosity (right) in the dentate molecular layer with increasing durations of HFD. For all graphs, bar or symbol height represents the mean of ( n = 4–6) mice per diet at each time point and error bars represent SEM. *Indicates significant differences ( p

Techniques Used: Mouse Assay, Molecular Weight, Western Blot, Labeling

Inducible ablation of Adora2a on endothelial cells maintains BBB structure and function in mice with diet-induced insulin resistance. A , Schematic depiction of Tg mouse model with inducible ablation of Adora2a in Tek/Tie2-expressing endothelial cells. B , Amplification of genomic DNA from BVECs, microglia, astrocytes, or whole hippocampus with primers directed against the Adora2a deleted sequence reveals selective deletion in BVECs following doxycycline (Dox) administration. C , Weight gain on HFD or LFD is similar in Tie2 rtTA /tetO cre /Adora2a fl/fl Tg mice and nTg littermates, with no differences either before and after induction during Week 12. D , Intraperitoneal glucose tolerance testing revealed comparable deficits in glycemic control in Tg/HFD mice and nTg/HFD littermates. C , D , Symbol height represents the average of ( n = 14–18) mice per condition. E , Inducible ablation of endothelial Adora2a restores hippocampal expression of the tight junction proteins claudin-5 (left) and occludin (right). Bar height represents the average of ( n = 4–6) mice per condition. F , Increases in NaFl (left) and Evans blue (right) extravasation were eliminated in Tg mice on HFD. Bar height shows the mean of ( n = 8–12) mice per condition. G , Tg mice are protected against extravascular IgG accumulation with HFD. Micrographs (right) show immunofluorescence detection of extravascular IgG and tomato lectin after background subtraction. Scale bar, 25 μm. H , Ablation of endothelial Adora2a eliminates obesity-induced pericyte regression (left). Right graph shows perivascular IgG deposits in microvascular regions with or without PDGFRb+ pericytes. Micrographs (far right) show labeling for PDGFRb, tomato lectin, and mouse IgG. Scale bar, 5 μm. G , H , Bar height represents the average of ( n = 4–6) mice per condition. For all graphs, error bars represent SEM and asterisk (*) indicates significance at p
Figure Legend Snippet: Inducible ablation of Adora2a on endothelial cells maintains BBB structure and function in mice with diet-induced insulin resistance. A , Schematic depiction of Tg mouse model with inducible ablation of Adora2a in Tek/Tie2-expressing endothelial cells. B , Amplification of genomic DNA from BVECs, microglia, astrocytes, or whole hippocampus with primers directed against the Adora2a deleted sequence reveals selective deletion in BVECs following doxycycline (Dox) administration. C , Weight gain on HFD or LFD is similar in Tie2 rtTA /tetO cre /Adora2a fl/fl Tg mice and nTg littermates, with no differences either before and after induction during Week 12. D , Intraperitoneal glucose tolerance testing revealed comparable deficits in glycemic control in Tg/HFD mice and nTg/HFD littermates. C , D , Symbol height represents the average of ( n = 14–18) mice per condition. E , Inducible ablation of endothelial Adora2a restores hippocampal expression of the tight junction proteins claudin-5 (left) and occludin (right). Bar height represents the average of ( n = 4–6) mice per condition. F , Increases in NaFl (left) and Evans blue (right) extravasation were eliminated in Tg mice on HFD. Bar height shows the mean of ( n = 8–12) mice per condition. G , Tg mice are protected against extravascular IgG accumulation with HFD. Micrographs (right) show immunofluorescence detection of extravascular IgG and tomato lectin after background subtraction. Scale bar, 25 μm. H , Ablation of endothelial Adora2a eliminates obesity-induced pericyte regression (left). Right graph shows perivascular IgG deposits in microvascular regions with or without PDGFRb+ pericytes. Micrographs (far right) show labeling for PDGFRb, tomato lectin, and mouse IgG. Scale bar, 5 μm. G , H , Bar height represents the average of ( n = 4–6) mice per condition. For all graphs, error bars represent SEM and asterisk (*) indicates significance at p

Techniques Used: Mouse Assay, Expressing, Amplification, Sequencing, Immunofluorescence, Labeling

25) Product Images from "Bile salt dependent lipase promotes intestinal adaptation in rats with massive small bowel resection"

Article Title: Bile salt dependent lipase promotes intestinal adaptation in rats with massive small bowel resection

Journal: Bioscience Reports

doi: 10.1042/BSR20180077

The expressions of TJ proteins in ileum ( A ) Representative claudin-1 immunofluorescence images in ileum of the sham group, the SBS group, and the SBS-BSDL group (crypt vertical section, magnification 200×). ( B ) Western blot representing total occludin and claudin-1 in ileum of the sham group, the SBS group, and the SBS-BSDL group. ( C ) Densitometric quantitation of relative expression levels normalized to GAPDH. Data are expressed as mean ± S.E.M.; rats n =4 per group. * P
Figure Legend Snippet: The expressions of TJ proteins in ileum ( A ) Representative claudin-1 immunofluorescence images in ileum of the sham group, the SBS group, and the SBS-BSDL group (crypt vertical section, magnification 200×). ( B ) Western blot representing total occludin and claudin-1 in ileum of the sham group, the SBS group, and the SBS-BSDL group. ( C ) Densitometric quantitation of relative expression levels normalized to GAPDH. Data are expressed as mean ± S.E.M.; rats n =4 per group. * P

Techniques Used: Immunofluorescence, Western Blot, Quantitation Assay, Expressing

26) Product Images from "FRAP Analysis Reveals Stabilization of Adhesion Structures in the Epidermis Compared to Cultured Keratinocytes"

Article Title: FRAP Analysis Reveals Stabilization of Adhesion Structures in the Epidermis Compared to Cultured Keratinocytes

Journal: PLoS ONE

doi: 10.1371/journal.pone.0071491

ZO-1 GFP exhibits low mobility in epidermis. A–D) Localization of ZO-1-GFP in skin sections taken from a ZO-1-GFP knock-in mouse. Scale bar 10 µm. A) ZO-1-GFP co-localizes with the tight junction protein occludin in kidney tissue sections of adult mouse. DNA is stained blue. B) Tissue section of lung taken from adult mouse. C) Whole mount epidermis of embryonic day 17.5 mouse. Note the ZO-1 signal in distinctive cobblestone pattern at cell-cell junctions in the granular layer. D) Whole mount small intestine taken from adult mouse. E) Mobile fractions from FRAP experiments are plotted. The box represents the 25 th to 75 th percentile and the whiskers represent the 10 th and 90 th percentiles. ** p
Figure Legend Snippet: ZO-1 GFP exhibits low mobility in epidermis. A–D) Localization of ZO-1-GFP in skin sections taken from a ZO-1-GFP knock-in mouse. Scale bar 10 µm. A) ZO-1-GFP co-localizes with the tight junction protein occludin in kidney tissue sections of adult mouse. DNA is stained blue. B) Tissue section of lung taken from adult mouse. C) Whole mount epidermis of embryonic day 17.5 mouse. Note the ZO-1 signal in distinctive cobblestone pattern at cell-cell junctions in the granular layer. D) Whole mount small intestine taken from adult mouse. E) Mobile fractions from FRAP experiments are plotted. The box represents the 25 th to 75 th percentile and the whiskers represent the 10 th and 90 th percentiles. ** p

Techniques Used: Knock-In, Staining

27) Product Images from "Cell-Cell Adhesions and Cell Contractility Are Upregulated upon Desmosome Disruption"

Article Title: Cell-Cell Adhesions and Cell Contractility Are Upregulated upon Desmosome Disruption

Journal: PLoS ONE

doi: 10.1371/journal.pone.0101824

Tight junctions are altered upon loss of desmoplakin. (A-B′) Calcium was added to WT and DP-null keratinocytes, and cells were fixed at various time points and stained for tight junction proteins occludin (red) and ZO-1 (green). Scale bar, 10 µm. (C,D) ZO-1 staining of WT and DP-null keratinocytes at 3 hour after calcium switch. (E) Co-stain for ZO-1 (red) and E-cadherin (green) in DP null keratinocytes 3 hours after calcium switch. (F) RNA was isolated from WT and DP-null keratinocytes, and RT-PCR for several claudins was performed. ***, p
Figure Legend Snippet: Tight junctions are altered upon loss of desmoplakin. (A-B′) Calcium was added to WT and DP-null keratinocytes, and cells were fixed at various time points and stained for tight junction proteins occludin (red) and ZO-1 (green). Scale bar, 10 µm. (C,D) ZO-1 staining of WT and DP-null keratinocytes at 3 hour after calcium switch. (E) Co-stain for ZO-1 (red) and E-cadherin (green) in DP null keratinocytes 3 hours after calcium switch. (F) RNA was isolated from WT and DP-null keratinocytes, and RT-PCR for several claudins was performed. ***, p

Techniques Used: Staining, Isolation, Reverse Transcription Polymerase Chain Reaction

28) Product Images from "Rivaroxaban does not influence hemorrhagic transformation in a diabetes ischemic stroke and endovascular thrombectomy model"

Article Title: Rivaroxaban does not influence hemorrhagic transformation in a diabetes ischemic stroke and endovascular thrombectomy model

Journal: Scientific Reports

doi: 10.1038/s41598-018-25820-y

Effects of Rivaroxaban on occludin expression and MMP-9 activity in rats. ( A ) A representative western blot showing changes in occludin expression (n = 6 in each group). Both the Control and Rivaroxaban groups have significantly lower occludin expression level than the Sham group (* P
Figure Legend Snippet: Effects of Rivaroxaban on occludin expression and MMP-9 activity in rats. ( A ) A representative western blot showing changes in occludin expression (n = 6 in each group). Both the Control and Rivaroxaban groups have significantly lower occludin expression level than the Sham group (* P

Techniques Used: Expressing, Activity Assay, Western Blot

29) Product Images from "Combination Compositions Composed of l-Glutamine and Si-Jun-Zi-Tang Might Be a Preferable Choice for 5-Fluorouracil-Induced Intestinal Mucositis: An Exploration in a Mouse Model"

Article Title: Combination Compositions Composed of l-Glutamine and Si-Jun-Zi-Tang Might Be a Preferable Choice for 5-Fluorouracil-Induced Intestinal Mucositis: An Exploration in a Mouse Model

Journal: Frontiers in Pharmacology

doi: 10.3389/fphar.2020.00918

Effect of G-SJZ on the expression of occludin, claudin-1, and ZO-1 in mice administered continuous 5-Fu: representative images of immunohistochemical staining under a microscope of 200 and 400 magnifications (A) , AOD analyzed by Image-Pro Plus (B) . * Significant difference ( P
Figure Legend Snippet: Effect of G-SJZ on the expression of occludin, claudin-1, and ZO-1 in mice administered continuous 5-Fu: representative images of immunohistochemical staining under a microscope of 200 and 400 magnifications (A) , AOD analyzed by Image-Pro Plus (B) . * Significant difference ( P

Techniques Used: Expressing, Mouse Assay, Immunohistochemistry, Staining, Microscopy

30) Product Images from "Distinct role of heme oxygenase-1 in early- and late-stage intracerebral hemorrhage in 12-month-old mice"

Article Title: Distinct role of heme oxygenase-1 in early- and late-stage intracerebral hemorrhage in 12-month-old mice

Journal: Journal of Cerebral Blood Flow & Metabolism

doi: 10.1177/0271678X16655814

Effect of HO-1 on blood–brain barrier after collagenase-induced ICH. (a) Evans blue (EB) extravasation on day 3 after ICH. (b, c) Western blot analysis of tight junction proteins ZO-1 (b) and occludin (c) on day 3 after collagenase-induced ICH. (d) Gelatin gel zymography analysis shows activation of matrix metalloproteinase (MMP)-9 and MMP-2 on day 3 after ICH. n = 5 mice/group, * p
Figure Legend Snippet: Effect of HO-1 on blood–brain barrier after collagenase-induced ICH. (a) Evans blue (EB) extravasation on day 3 after ICH. (b, c) Western blot analysis of tight junction proteins ZO-1 (b) and occludin (c) on day 3 after collagenase-induced ICH. (d) Gelatin gel zymography analysis shows activation of matrix metalloproteinase (MMP)-9 and MMP-2 on day 3 after ICH. n = 5 mice/group, * p

Techniques Used: Western Blot, Zymography, Activation Assay, Mouse Assay

31) Product Images from "Integrative Proteomic and Phosphoproteomic Profiling of Testis from Wip1 Phosphatase-Knockout Mice: Insights into Mechanisms of Reduced Fertility*"

Article Title: Integrative Proteomic and Phosphoproteomic Profiling of Testis from Wip1 Phosphatase-Knockout Mice: Insights into Mechanisms of Reduced Fertility*

Journal: Molecular & Cellular Proteomics : MCP

doi: 10.1074/mcp.RA117.000479

Schematic model of the proposed mechanisms responsible for the subfertility and spermatogenesis defects induced by Wip1 deficiency. Proinflammatory cytokines impairs the BTB and apical ectoplasmic specialization dynamics by decreasing the expression of junction-associated proteins (occludin, ZO-1, N-cadherin, and pβ-catenin (S552)), which effect could be partially responsible for the spermatid sloughing and oligospermia and subfertility in Wip1 -deficient mice.
Figure Legend Snippet: Schematic model of the proposed mechanisms responsible for the subfertility and spermatogenesis defects induced by Wip1 deficiency. Proinflammatory cytokines impairs the BTB and apical ectoplasmic specialization dynamics by decreasing the expression of junction-associated proteins (occludin, ZO-1, N-cadherin, and pβ-catenin (S552)), which effect could be partially responsible for the spermatid sloughing and oligospermia and subfertility in Wip1 -deficient mice.

Techniques Used: Expressing, Mouse Assay

Wip1 deficiency disrupts the expression of junction-associated proteins and the integrity of BTB. ( A , a-d) Immunofluorescence images of testis sections from 10-week-old WT and Wip1 -KO mice stained with anti-WT1 and anti-β-catenin body. ( A , e-j) Immunoreactivity of junction-associated proteins (occludin, N-cadherin, ZO-1) was altered in the seminiferous tubules of Wip1 -deficient mice. Distribution of occludin (red arrows; e-f), N-cadherin (red arrows; g-h), and ZO-1 (red arrows; i-j) in the seminiferous epithelium of WT and Wip1 -KO mice ( n = 4 per group) counterstained with DAPI. Scale bars: 50 μm (a-b); 20 μm (c-j). B , Relative mRNA expression levels of junction-associated components in WT and Wip1 -KO mouse testes. C , Relative protein expression levels of junction-associated components in WT and Wip1 -KO mouse testes. β-actin served as a loading control. D , Quantified relative band intensity ratio of C. E , In vivo functional assay (FITC-labeled dextran tracer) to determine BTB integrity in WT and Wip1 -KO mouse testes. In testes sections from WT mice, FITC green fluorescence was only visible in the interstitial spaces and basal compartment, whereas in sections of testes from Wip1 -KO mice, FITC green fluorescence was observed in the interstitial spaces and basal compartment, and in the lumen of the seminiferous tubules. Scale bars: 50 μm. Data are expressed as mean ± S.E. * p
Figure Legend Snippet: Wip1 deficiency disrupts the expression of junction-associated proteins and the integrity of BTB. ( A , a-d) Immunofluorescence images of testis sections from 10-week-old WT and Wip1 -KO mice stained with anti-WT1 and anti-β-catenin body. ( A , e-j) Immunoreactivity of junction-associated proteins (occludin, N-cadherin, ZO-1) was altered in the seminiferous tubules of Wip1 -deficient mice. Distribution of occludin (red arrows; e-f), N-cadherin (red arrows; g-h), and ZO-1 (red arrows; i-j) in the seminiferous epithelium of WT and Wip1 -KO mice ( n = 4 per group) counterstained with DAPI. Scale bars: 50 μm (a-b); 20 μm (c-j). B , Relative mRNA expression levels of junction-associated components in WT and Wip1 -KO mouse testes. C , Relative protein expression levels of junction-associated components in WT and Wip1 -KO mouse testes. β-actin served as a loading control. D , Quantified relative band intensity ratio of C. E , In vivo functional assay (FITC-labeled dextran tracer) to determine BTB integrity in WT and Wip1 -KO mouse testes. In testes sections from WT mice, FITC green fluorescence was only visible in the interstitial spaces and basal compartment, whereas in sections of testes from Wip1 -KO mice, FITC green fluorescence was observed in the interstitial spaces and basal compartment, and in the lumen of the seminiferous tubules. Scale bars: 50 μm. Data are expressed as mean ± S.E. * p

Techniques Used: Expressing, Immunofluorescence, Mouse Assay, Staining, In Vivo, Functional Assay, Labeling, Fluorescence

32) Product Images from "Integrative Proteomic and Phosphoproteomic Profiling of Testis from Wip1 Phosphatase-Knockout Mice: Insights into Mechanisms of Reduced Fertility*"

Article Title: Integrative Proteomic and Phosphoproteomic Profiling of Testis from Wip1 Phosphatase-Knockout Mice: Insights into Mechanisms of Reduced Fertility*

Journal: Molecular & Cellular Proteomics : MCP

doi: 10.1074/mcp.RA117.000479

Schematic model of the proposed mechanisms responsible for the subfertility and spermatogenesis defects induced by Wip1 deficiency. Proinflammatory cytokines impairs the BTB and apical ectoplasmic specialization dynamics by decreasing the expression of junction-associated proteins (occludin, ZO-1, N-cadherin, and pβ-catenin (S552)), which effect could be partially responsible for the spermatid sloughing and oligospermia and subfertility in Wip1 -deficient mice.
Figure Legend Snippet: Schematic model of the proposed mechanisms responsible for the subfertility and spermatogenesis defects induced by Wip1 deficiency. Proinflammatory cytokines impairs the BTB and apical ectoplasmic specialization dynamics by decreasing the expression of junction-associated proteins (occludin, ZO-1, N-cadherin, and pβ-catenin (S552)), which effect could be partially responsible for the spermatid sloughing and oligospermia and subfertility in Wip1 -deficient mice.

Techniques Used: Expressing, Mouse Assay

Wip1 deficiency disrupts the expression of junction-associated proteins and the integrity of BTB. ( A , a-d) Immunofluorescence images of testis sections from 10-week-old WT and Wip1 -KO mice stained with anti-WT1 and anti-β-catenin body. ( A , e-j) Immunoreactivity of junction-associated proteins (occludin, N-cadherin, ZO-1) was altered in the seminiferous tubules of Wip1 -deficient mice. Distribution of occludin (red arrows; e-f), N-cadherin (red arrows; g-h), and ZO-1 (red arrows; i-j) in the seminiferous epithelium of WT and Wip1 -KO mice ( n = 4 per group) counterstained with DAPI. Scale bars: 50 μm (a-b); 20 μm (c-j). B , Relative mRNA expression levels of junction-associated components in WT and Wip1 -KO mouse testes. C , Relative protein expression levels of junction-associated components in WT and Wip1 -KO mouse testes. β-actin served as a loading control. D , Quantified relative band intensity ratio of C. E , In vivo functional assay (FITC-labeled dextran tracer) to determine BTB integrity in WT and Wip1 -KO mouse testes. In testes sections from WT mice, FITC green fluorescence was only visible in the interstitial spaces and basal compartment, whereas in sections of testes from Wip1 -KO mice, FITC green fluorescence was observed in the interstitial spaces and basal compartment, and in the lumen of the seminiferous tubules. Scale bars: 50 μm. Data are expressed as mean ± S.E. * p
Figure Legend Snippet: Wip1 deficiency disrupts the expression of junction-associated proteins and the integrity of BTB. ( A , a-d) Immunofluorescence images of testis sections from 10-week-old WT and Wip1 -KO mice stained with anti-WT1 and anti-β-catenin body. ( A , e-j) Immunoreactivity of junction-associated proteins (occludin, N-cadherin, ZO-1) was altered in the seminiferous tubules of Wip1 -deficient mice. Distribution of occludin (red arrows; e-f), N-cadherin (red arrows; g-h), and ZO-1 (red arrows; i-j) in the seminiferous epithelium of WT and Wip1 -KO mice ( n = 4 per group) counterstained with DAPI. Scale bars: 50 μm (a-b); 20 μm (c-j). B , Relative mRNA expression levels of junction-associated components in WT and Wip1 -KO mouse testes. C , Relative protein expression levels of junction-associated components in WT and Wip1 -KO mouse testes. β-actin served as a loading control. D , Quantified relative band intensity ratio of C. E , In vivo functional assay (FITC-labeled dextran tracer) to determine BTB integrity in WT and Wip1 -KO mouse testes. In testes sections from WT mice, FITC green fluorescence was only visible in the interstitial spaces and basal compartment, whereas in sections of testes from Wip1 -KO mice, FITC green fluorescence was observed in the interstitial spaces and basal compartment, and in the lumen of the seminiferous tubules. Scale bars: 50 μm. Data are expressed as mean ± S.E. * p

Techniques Used: Expressing, Immunofluorescence, Mouse Assay, Staining, In Vivo, Functional Assay, Labeling, Fluorescence

33) Product Images from "Rivaroxaban does not influence hemorrhagic transformation in a diabetes ischemic stroke and endovascular thrombectomy model"

Article Title: Rivaroxaban does not influence hemorrhagic transformation in a diabetes ischemic stroke and endovascular thrombectomy model

Journal: Scientific Reports

doi: 10.1038/s41598-018-25820-y

Effects of Rivaroxaban on occludin expression and MMP-9 activity in rats. ( A ) A representative western blot showing changes in occludin expression (n = 6 in each group). Both the Control and Rivaroxaban groups have significantly lower occludin expression level than the Sham group (* P
Figure Legend Snippet: Effects of Rivaroxaban on occludin expression and MMP-9 activity in rats. ( A ) A representative western blot showing changes in occludin expression (n = 6 in each group). Both the Control and Rivaroxaban groups have significantly lower occludin expression level than the Sham group (* P

Techniques Used: Expressing, Activity Assay, Western Blot

34) Product Images from "Chronic administration of AMD3100 increases survival and alleviates pathology in SOD1G93A mice model of ALS"

Article Title: Chronic administration of AMD3100 increases survival and alleviates pathology in SOD1G93A mice model of ALS

Journal: Journal of Neuroinflammation

doi: 10.1186/s12974-016-0587-6

AMD3100 reduces BSCB permeability and increases tight junction proteins levels. a Evans blue permeability assay. Fifty-day-old SOD1 G93A mice were treated with AMD3100 ( n = 3) or PBS ( n = 3). One hundred ten-day-old mice were IP injected with Evans blue dye (50 ug/g body weight). Three hours post injection mice were sacrificed and perfused, and dye presence in the spinal cord was measured as ratio of tissue weight. b Regional analysis of hemosiderin deposits. c IgG leakage in the lumbar cord anterior horn of SOD1 G93A mice treated with AMD3100 ( n = 3) or PBS ( n = 3) from 50 to 110 days old. Fifteen nonadjacent sections were examined for each treatment group. d Occludin levels of SOD1 G93A mice. e Claudin-5 levels of SOD1 G93A mice. f ZO-1 levels of SOD1 G93A mice. g MCP-1 levels of SOD1 G93A mice. For tight junction proteins, SOD1 G93A mice were treated with AMD3100 or PBS and sacrificed at 110 days old, and protein levels were measured using western blot. Five mice in each treatment group of SOD1 G93A mice were tested. Results are mean ± S.E.M., * p
Figure Legend Snippet: AMD3100 reduces BSCB permeability and increases tight junction proteins levels. a Evans blue permeability assay. Fifty-day-old SOD1 G93A mice were treated with AMD3100 ( n = 3) or PBS ( n = 3). One hundred ten-day-old mice were IP injected with Evans blue dye (50 ug/g body weight). Three hours post injection mice were sacrificed and perfused, and dye presence in the spinal cord was measured as ratio of tissue weight. b Regional analysis of hemosiderin deposits. c IgG leakage in the lumbar cord anterior horn of SOD1 G93A mice treated with AMD3100 ( n = 3) or PBS ( n = 3) from 50 to 110 days old. Fifteen nonadjacent sections were examined for each treatment group. d Occludin levels of SOD1 G93A mice. e Claudin-5 levels of SOD1 G93A mice. f ZO-1 levels of SOD1 G93A mice. g MCP-1 levels of SOD1 G93A mice. For tight junction proteins, SOD1 G93A mice were treated with AMD3100 or PBS and sacrificed at 110 days old, and protein levels were measured using western blot. Five mice in each treatment group of SOD1 G93A mice were tested. Results are mean ± S.E.M., * p

Techniques Used: Permeability, Mouse Assay, Injection, Western Blot

35) Product Images from "In Vitro Modeling of the Neurovascular Environment by Coculturing Adult Human Brain Endothelial Cells with Human Neural Stem Cells"

Article Title: In Vitro Modeling of the Neurovascular Environment by Coculturing Adult Human Brain Endothelial Cells with Human Neural Stem Cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0106346

Cytoarchitectural characterization of vessel-like structures. (A) Astrocytes (monoclonal GFAP+ cells) form a layer of support around endothelial cells that organize into tubular structures readily identified by phalloidin. (B) Tubular structures are tightly packed with endothelial cells expressing intercellular adhesion molecule-2 (ICAM-2), although there is a degree of inconsistency in along the VLS indicating different stages of development/maturity. (C) Endothelial cells organized into VLS are polarizing as indicated by the expression of the apical marker podocalyxin that influences astrocytes positioning of endfeet. (D) Endothelial cells within the VLS also express markers indicative of tight junctions, such as Claudin-5 and Occludin.
Figure Legend Snippet: Cytoarchitectural characterization of vessel-like structures. (A) Astrocytes (monoclonal GFAP+ cells) form a layer of support around endothelial cells that organize into tubular structures readily identified by phalloidin. (B) Tubular structures are tightly packed with endothelial cells expressing intercellular adhesion molecule-2 (ICAM-2), although there is a degree of inconsistency in along the VLS indicating different stages of development/maturity. (C) Endothelial cells organized into VLS are polarizing as indicated by the expression of the apical marker podocalyxin that influences astrocytes positioning of endfeet. (D) Endothelial cells within the VLS also express markers indicative of tight junctions, such as Claudin-5 and Occludin.

Techniques Used: Expressing, Marker

36) Product Images from "Endothelial Adora2a Activation Promotes Blood–Brain Barrier Breakdown and Cognitive Impairment in Mice with Diet-Induced Insulin Resistance"

Article Title: Endothelial Adora2a Activation Promotes Blood–Brain Barrier Breakdown and Cognitive Impairment in Mice with Diet-Induced Insulin Resistance

Journal: The Journal of Neuroscience

doi: 10.1523/JNEUROSCI.2506-18.2019

Progressive changes in BBB integrity with increasing durations of obesogenic diet consumption. A , Weight gain with exposure to a HFD or LFD in male mice. B , Fasting hyperglycemia (left) and fasting hyperinsulinemia (right) were evident after 16 weeks consumption of HFD. C , Increased penetration of the low molecular weight tracer NaFl was observed in clarified hippocampal lysates after 12 or 16 weeks HFD (left graph). Right graph shows increases in hippocampal Evans blue after 16 weeks HFD. D ). Scale bar: (in LFD 8WK ) for all panels, 10 μm. E , Reductions in hippocampal claudin-5 (left) and occludin (right) protein were detected by Western blotting after 16 weeks of HFD. F , Micrographs show representative images of tomato lectin labeling for the indicated diets and time points. Scale bar: (in LFD 12WK ) for all micrographs, 100 μm. G , Increases in vascular area (left) and tortuosity (right) in the dentate molecular layer with increasing durations of HFD. For all graphs, bar or symbol height represents the mean of ( n = 4–6) mice per diet at each time point and error bars represent SEM. *Indicates significant differences ( p
Figure Legend Snippet: Progressive changes in BBB integrity with increasing durations of obesogenic diet consumption. A , Weight gain with exposure to a HFD or LFD in male mice. B , Fasting hyperglycemia (left) and fasting hyperinsulinemia (right) were evident after 16 weeks consumption of HFD. C , Increased penetration of the low molecular weight tracer NaFl was observed in clarified hippocampal lysates after 12 or 16 weeks HFD (left graph). Right graph shows increases in hippocampal Evans blue after 16 weeks HFD. D ). Scale bar: (in LFD 8WK ) for all panels, 10 μm. E , Reductions in hippocampal claudin-5 (left) and occludin (right) protein were detected by Western blotting after 16 weeks of HFD. F , Micrographs show representative images of tomato lectin labeling for the indicated diets and time points. Scale bar: (in LFD 12WK ) for all micrographs, 100 μm. G , Increases in vascular area (left) and tortuosity (right) in the dentate molecular layer with increasing durations of HFD. For all graphs, bar or symbol height represents the mean of ( n = 4–6) mice per diet at each time point and error bars represent SEM. *Indicates significant differences ( p

Techniques Used: Mouse Assay, Molecular Weight, Western Blot, Labeling

Inducible ablation of Adora2a on endothelial cells maintains BBB structure and function in mice with diet-induced insulin resistance. A , Schematic depiction of Tg mouse model with inducible ablation of Adora2a in Tek/Tie2-expressing endothelial cells. B , Amplification of genomic DNA from BVECs, microglia, astrocytes, or whole hippocampus with primers directed against the Adora2a deleted sequence reveals selective deletion in BVECs following doxycycline (Dox) administration. C , Weight gain on HFD or LFD is similar in Tie2 rtTA /tetO cre /Adora2a fl/fl Tg mice and nTg littermates, with no differences either before and after induction during Week 12. D , Intraperitoneal glucose tolerance testing revealed comparable deficits in glycemic control in Tg/HFD mice and nTg/HFD littermates. C , D , Symbol height represents the average of ( n = 14–18) mice per condition. E , Inducible ablation of endothelial Adora2a restores hippocampal expression of the tight junction proteins claudin-5 (left) and occludin (right). Bar height represents the average of ( n = 4–6) mice per condition. F , Increases in NaFl (left) and Evans blue (right) extravasation were eliminated in Tg mice on HFD. Bar height shows the mean of ( n = 8–12) mice per condition. G , Tg mice are protected against extravascular IgG accumulation with HFD. Micrographs (right) show immunofluorescence detection of extravascular IgG and tomato lectin after background subtraction. Scale bar, 25 μm. H , Ablation of endothelial Adora2a eliminates obesity-induced pericyte regression (left). Right graph shows perivascular IgG deposits in microvascular regions with or without PDGFRb+ pericytes. Micrographs (far right) show labeling for PDGFRb, tomato lectin, and mouse IgG. Scale bar, 5 μm. G , H , Bar height represents the average of ( n = 4–6) mice per condition. For all graphs, error bars represent SEM and asterisk (*) indicates significance at p
Figure Legend Snippet: Inducible ablation of Adora2a on endothelial cells maintains BBB structure and function in mice with diet-induced insulin resistance. A , Schematic depiction of Tg mouse model with inducible ablation of Adora2a in Tek/Tie2-expressing endothelial cells. B , Amplification of genomic DNA from BVECs, microglia, astrocytes, or whole hippocampus with primers directed against the Adora2a deleted sequence reveals selective deletion in BVECs following doxycycline (Dox) administration. C , Weight gain on HFD or LFD is similar in Tie2 rtTA /tetO cre /Adora2a fl/fl Tg mice and nTg littermates, with no differences either before and after induction during Week 12. D , Intraperitoneal glucose tolerance testing revealed comparable deficits in glycemic control in Tg/HFD mice and nTg/HFD littermates. C , D , Symbol height represents the average of ( n = 14–18) mice per condition. E , Inducible ablation of endothelial Adora2a restores hippocampal expression of the tight junction proteins claudin-5 (left) and occludin (right). Bar height represents the average of ( n = 4–6) mice per condition. F , Increases in NaFl (left) and Evans blue (right) extravasation were eliminated in Tg mice on HFD. Bar height shows the mean of ( n = 8–12) mice per condition. G , Tg mice are protected against extravascular IgG accumulation with HFD. Micrographs (right) show immunofluorescence detection of extravascular IgG and tomato lectin after background subtraction. Scale bar, 25 μm. H , Ablation of endothelial Adora2a eliminates obesity-induced pericyte regression (left). Right graph shows perivascular IgG deposits in microvascular regions with or without PDGFRb+ pericytes. Micrographs (far right) show labeling for PDGFRb, tomato lectin, and mouse IgG. Scale bar, 5 μm. G , H , Bar height represents the average of ( n = 4–6) mice per condition. For all graphs, error bars represent SEM and asterisk (*) indicates significance at p

Techniques Used: Mouse Assay, Expressing, Amplification, Sequencing, Immunofluorescence, Labeling

37) Product Images from "Adult human dental pulp stem cells promote blood–brain barrier permeability through vascular endothelial growth factor-a expression"

Article Title: Adult human dental pulp stem cells promote blood–brain barrier permeability through vascular endothelial growth factor-a expression

Journal: Journal of Cerebral Blood Flow & Metabolism

doi: 10.1177/0271678X15608392

Immunocytochemistry of BMEC monolayers stained for the tight junction protein occludin (Cy2/green) and nuclei (DAPI/blue). (a) Untreated co-culture of BMECs and astrocytes. (b) BMEC and astrocyte co-culture negative control for the occludin primary antibody.
Figure Legend Snippet: Immunocytochemistry of BMEC monolayers stained for the tight junction protein occludin (Cy2/green) and nuclei (DAPI/blue). (a) Untreated co-culture of BMECs and astrocytes. (b) BMEC and astrocyte co-culture negative control for the occludin primary antibody.

Techniques Used: Immunocytochemistry, Staining, Co-Culture Assay, Negative Control

Immunocytochemistry of BMEC monolayers stained for the tight junction protein occludin (Cy2/green) and nuclei (DAPI/blue). (a) BMEC and astrocyte co-culture negative control for the occludin primary antibody. (b) Positive control for the occludin antibody,
Figure Legend Snippet: Immunocytochemistry of BMEC monolayers stained for the tight junction protein occludin (Cy2/green) and nuclei (DAPI/blue). (a) BMEC and astrocyte co-culture negative control for the occludin primary antibody. (b) Positive control for the occludin antibody,

Techniques Used: Immunocytochemistry, Staining, Co-Culture Assay, Negative Control, Positive Control

38) Product Images from "Mono-(2-Ethylhexyl) Phthalate-Induced Disruption of Junctional Complexes in the Seminiferous Epithelium of the Rodent Testis Is Mediated by MMP2 1"

Article Title: Mono-(2-Ethylhexyl) Phthalate-Induced Disruption of Junctional Complexes in the Seminiferous Epithelium of the Rodent Testis Is Mediated by MMP2 1

Journal: Biology of Reproduction

doi: 10.1095/biolreprod.109.080374

MEHP exposure decreases expression levels of junction proteins in vitro and in vivo. The protein levels of claudin11, occludin, ZO1, laminin-γ3, and β1-integrin in whole testis homogenates ( A and B ) from 28-day-old C57BL/6J mice and in
Figure Legend Snippet: MEHP exposure decreases expression levels of junction proteins in vitro and in vivo. The protein levels of claudin11, occludin, ZO1, laminin-γ3, and β1-integrin in whole testis homogenates ( A and B ) from 28-day-old C57BL/6J mice and in

Techniques Used: Expressing, In Vitro, In Vivo, Mouse Assay

Alterations in the expression and localization of claudin11, occludin, and ZO1 in the seminiferous epithelium are observed by immunohistochemical analysis. Twenty-eight-day-old C57BL/6J mice are pretreated with SB-3CT (0 and 25 mg/kg) for 6 h and then
Figure Legend Snippet: Alterations in the expression and localization of claudin11, occludin, and ZO1 in the seminiferous epithelium are observed by immunohistochemical analysis. Twenty-eight-day-old C57BL/6J mice are pretreated with SB-3CT (0 and 25 mg/kg) for 6 h and then

Techniques Used: Expressing, Immunohistochemistry, Mouse Assay

39) Product Images from "Comparative benefits of simvastatin and exercise in a mouse model of vascular cognitive impairment and dementia"

Article Title: Comparative benefits of simvastatin and exercise in a mouse model of vascular cognitive impairment and dementia

Journal: The FASEB Journal

doi: 10.1096/fj.201901002R

Neuroinflammatory responses in the cortex were not exacerbated by HCD. A , B ) Activation of cortical astrocytes and microglia as quantified with GFAP ( A ) and Iba-1 ( B ) immunostaining was significantly reduced by unlimited EX but not by SV (cohort 1). C – F ) When EX was limited (cohort 2), significant decreases were observed in cortical GFAP ( C ) but not in Iba-1 ( D ). E , F ) HCD did not exacerbate these pathologies that are already present in TGF mice in either cohort. IL levels of TNF-α, IL-1β, and IL-6 measured by ELISA were not different between groups ( E ), and no evidence of BBB damage was found when quantifying the tight junction protein occludin by Western blot ( F ) ( n = 4–5 mice/group). Scale bars, 100 μm for GFAP and Iba-1 (10 μm, inset). * P
Figure Legend Snippet: Neuroinflammatory responses in the cortex were not exacerbated by HCD. A , B ) Activation of cortical astrocytes and microglia as quantified with GFAP ( A ) and Iba-1 ( B ) immunostaining was significantly reduced by unlimited EX but not by SV (cohort 1). C – F ) When EX was limited (cohort 2), significant decreases were observed in cortical GFAP ( C ) but not in Iba-1 ( D ). E , F ) HCD did not exacerbate these pathologies that are already present in TGF mice in either cohort. IL levels of TNF-α, IL-1β, and IL-6 measured by ELISA were not different between groups ( E ), and no evidence of BBB damage was found when quantifying the tight junction protein occludin by Western blot ( F ) ( n = 4–5 mice/group). Scale bars, 100 μm for GFAP and Iba-1 (10 μm, inset). * P

Techniques Used: Activation Assay, Immunostaining, Mouse Assay, Enzyme-linked Immunosorbent Assay, Western Blot

Related Articles

Western Blot:

Article Title: MST1 Suppression Reduces Early Brain Injury by Inhibiting the NF-κB/MMP-9 Pathway after Subarachnoid Hemorrhage in Mice
Article Snippet: .. Western Blotting The ipsilateral/right cortex was isolated and homogenized for Western blotting as previously described [ ] with the following primary antibodies: anti-MST1 (1 : 1000, Cell Signaling Technology, Danvers, MA, USA), anti-phosphorylated MST1 (1 : 1000, Cell Signaling Technology, Danvers, MA, USA), anti-ZO-1 (1 : 500 Invitrogen, Grand Island, NY, USA), anti-occludin (1 : 1000 Abcam, Shanghai, China), anti-claudin-5 (1 : 1000, Abcam, Shanghai, China), anti-NF-κ B (1 : 1000, Cell Signaling Technology, Danvers, MA, USA), and anti-matrix metallopeptidase- (MMP-) 9 (1 : 1000, Cell Signaling Technology, Danvers, MA, USA). .. GAPDH (1 : 10,000, Proteintech, Rosemont, IL, USA) was used as an internal loading control.

Incubation:

Article Title: 1, 25-D3 Protects From Cerebral Ischemia by Maintaining BBB Permeability via PPAR-γ Activation
Article Snippet: .. The slices were first incubated with the blocking solution (0.1% Triton X-100 in 0.1 M PBS, 10% bovine serum albumin, pH 7.6) for 2 h and then overnight with anti-occludin (1:100, Abcam, USA) and anti-claudin-5 (1:100, Thermo Fisher Scientific, Rockford, IL, USA) antibodies at 4°C. .. After washing with PBS, the tissues were incubated with the secondary antibodies and counterstained with 4′,6-diamidino-2-phenylindole (DAPI, Beyotime, China).

Article Title: Comparison of Melatonin, Hypertonic Saline, and Hydroxyethyl Starch for Resuscitation of Secondary Intra-Abdominal Hypertension in an Animal Model
Article Snippet: .. The membranes were incubated with anti-phospho-Akt (1:2000; Cell Signaling Technology, Danvers, MA, USA), anti-Akt (1:1000; Cell Signaling Technology, Danvers, MA, USA), anti- zonula occludens-1(ZO-1) (1:50; Abcam, Cambridge, MA, USA), and anti-occludin (1:100; Abcam, Cambridge, MA, USA) antibodies overnight at 4°C. .. The membrane-bound antibodies were visualized using IRDye 800 infrared-labeled donkey anti-rabbit secondary antibodies (1:15 000;Li-Cor Bioscience, Lincoln, NE,USA)and the Odyssey Infrared Imaging System (Li-Cor Bioscience, Bad Homburg, Germany).

Article Title: Attaching-and-Effacing Pathogens Exploit Junction Regulatory Activities of N-WASP and SNX9 to Disrupt the Intestinal Barrier
Article Snippet: .. After incubation with anti–E-cadherin (Cell Signaling Technology), anti-occludin (Abcam), or anti–zonula occludens-1 (ZO-1) (Abcam) primary antibodies, all at 1:400, slides were washed and incubated with Alexa 594–conjugated goat anti-rabbit or Alexa 488–conjugated goat anti-rabbit secondary antibodies and DAPI (Life Technologies, Grand Island, NY). .. Standard epifluorescence microscopy was performed using an AX70 upright microscope (Olympus, Tokyo, Japan).

Article Title: A Role of Exopolysaccharide Produced by Streptococcus thermophilus in the Intestinal Inflammation and Mucosal Barrier in Caco-2 Monolayer and Dextran Sulphate Sodium-Induced Experimental Murine Colitis
Article Snippet: .. Then the proteins were electro-transferred to PVDF member at 200 mA for 2 h. The membranes were incubated with primary antibodies: anti-β-actin (1:2500, Bioss, Beijing, China), Anti-Occludin (1:50,000, Abcam, Cambridge, MA, USA), anti-E-cadherin (1:1000, Abcam, USA), anti-Claudin1 (1:2000, Abcam, USA) for 12 h at 4 °C. .. After 5 times washed with 0.1 g/100 mL Tween-20 in PBS, then incubated with secondary antibodies (anti-rabbit and anti-mouse, 1:1000, Beyotime, Shanghai, China) for 1 h at room temperature.

Article Title: Analysis of differential expression of tight junction proteins in cultured oral epithelial cells altered by Porphyromonas gingivalis, Porphyromonas gingivalis lipopolysaccharide, and extracellular adenosine triphosphate
Article Snippet: .. The nitrocellulose membranes were then incubated with rabbit polyclonal antibodies to human occludin (ab31721), JAM-A (ab106114), claudin-1 (ab15098), claudin-4 (ab53156), ZO-1 (ab59720), claudin-15 (ab215354; all 1 μg·mL−1 , Abcam, Cambridge, UK), and β-actin (0.1 μg·mL−1 , GenTex, Zeeland, MI, USA) in 0.05% Tween20/TBS for 4 h. β-actin was used as a loading control. .. After the incubation, the membranes were washed three times and subsequently incubated with alkaline phosphatase (AP)-conjugated secondary antibody (goat-anti rabbit IgG, DAKO) diluted 1:1 500 in Tween20/TBS for 2 h. Bound antibodies were displayed with AP substrate (Bio-Rad) after the development of reactivity for proteins.

Blocking Assay:

Article Title: 1, 25-D3 Protects From Cerebral Ischemia by Maintaining BBB Permeability via PPAR-γ Activation
Article Snippet: .. The slices were first incubated with the blocking solution (0.1% Triton X-100 in 0.1 M PBS, 10% bovine serum albumin, pH 7.6) for 2 h and then overnight with anti-occludin (1:100, Abcam, USA) and anti-claudin-5 (1:100, Thermo Fisher Scientific, Rockford, IL, USA) antibodies at 4°C. .. After washing with PBS, the tissues were incubated with the secondary antibodies and counterstained with 4′,6-diamidino-2-phenylindole (DAPI, Beyotime, China).

Isolation:

Article Title: MST1 Suppression Reduces Early Brain Injury by Inhibiting the NF-κB/MMP-9 Pathway after Subarachnoid Hemorrhage in Mice
Article Snippet: .. Western Blotting The ipsilateral/right cortex was isolated and homogenized for Western blotting as previously described [ ] with the following primary antibodies: anti-MST1 (1 : 1000, Cell Signaling Technology, Danvers, MA, USA), anti-phosphorylated MST1 (1 : 1000, Cell Signaling Technology, Danvers, MA, USA), anti-ZO-1 (1 : 500 Invitrogen, Grand Island, NY, USA), anti-occludin (1 : 1000 Abcam, Shanghai, China), anti-claudin-5 (1 : 1000, Abcam, Shanghai, China), anti-NF-κ B (1 : 1000, Cell Signaling Technology, Danvers, MA, USA), and anti-matrix metallopeptidase- (MMP-) 9 (1 : 1000, Cell Signaling Technology, Danvers, MA, USA). .. GAPDH (1 : 10,000, Proteintech, Rosemont, IL, USA) was used as an internal loading control.

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  • 92
    Abcam rabbit anti occludin
    Adipose tissue extracts enable the EMT of CCA cells. Human cholangiocarcinoma (CCA) cell line RBE was incubated with complete growth medium or medium containing human adipose tissue extracts for 24 h. The cell morphology was observed under the invert microscopy (40×) ( a ). Protein expression of MMP2, MMP9, TGF-β1, claudin 1, <t>occludin,</t> E-cadherin, β-catenin, SNAIL, and Smad3 were analyzed by immunoblot analysis in response to adipose extracts incubation, and the band densitometry analysis was carried out relative to the loading control β-actin or GAPDH ( b , c ). To determine fatty acid metabolism, the culture medium with different adipose components prior to or after incubation with RBE cells for 24 h were collected for extracellular glycerol and nonestesterified fatty acid (NEFA) detection using colorimetric assays, with the data presented as the absorbance at 490 nm( d , e ). The intracellular lipids in adipose cultured RBE cells were visualized by the fluorescence dye Bodipy (20 μg/mL) using confocal microscopy analysis, using Hochest33342 (0.5 μg/mL) for nuclear staining ( f ). All the samples were prepared in triplicate, and all experiments were repeated at least three times. * P
    Rabbit Anti Occludin, supplied by Abcam, used in various techniques. Bioz Stars score: 92/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit anti occludin/product/Abcam
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    99
    Abcam primary rabbit anti occludin antibody
    Western blot analysis of <t>occludin</t> protein with or without acrolein exposure. Data represent Mean ± SE, n = 6, p = 0.337 as compared to controls. “C” represents control group and “A” represents acrolein treated group.
    Primary Rabbit Anti Occludin Antibody, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam monoclonal rabbit anti occludin
    Fluoxetine downregulated MMP-9 expression and prevented degradation of the tight junction proteins in the ipsilateral cortex at 24 h after SAH. a Representative western blot bands of MMP-9, <t>occludin,</t> claudin-5, and ZO-1. b Densitometric quantification of MMP-9. ** p
    Monoclonal Rabbit Anti Occludin, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Adipose tissue extracts enable the EMT of CCA cells. Human cholangiocarcinoma (CCA) cell line RBE was incubated with complete growth medium or medium containing human adipose tissue extracts for 24 h. The cell morphology was observed under the invert microscopy (40×) ( a ). Protein expression of MMP2, MMP9, TGF-β1, claudin 1, occludin, E-cadherin, β-catenin, SNAIL, and Smad3 were analyzed by immunoblot analysis in response to adipose extracts incubation, and the band densitometry analysis was carried out relative to the loading control β-actin or GAPDH ( b , c ). To determine fatty acid metabolism, the culture medium with different adipose components prior to or after incubation with RBE cells for 24 h were collected for extracellular glycerol and nonestesterified fatty acid (NEFA) detection using colorimetric assays, with the data presented as the absorbance at 490 nm( d , e ). The intracellular lipids in adipose cultured RBE cells were visualized by the fluorescence dye Bodipy (20 μg/mL) using confocal microscopy analysis, using Hochest33342 (0.5 μg/mL) for nuclear staining ( f ). All the samples were prepared in triplicate, and all experiments were repeated at least three times. * P

    Journal: Journal of Experimental & Clinical Cancer Research : CR

    Article Title: Adipocytes promote cholangiocarcinoma metastasis through fatty acid binding protein 4

    doi: 10.1186/s13046-017-0641-y

    Figure Lengend Snippet: Adipose tissue extracts enable the EMT of CCA cells. Human cholangiocarcinoma (CCA) cell line RBE was incubated with complete growth medium or medium containing human adipose tissue extracts for 24 h. The cell morphology was observed under the invert microscopy (40×) ( a ). Protein expression of MMP2, MMP9, TGF-β1, claudin 1, occludin, E-cadherin, β-catenin, SNAIL, and Smad3 were analyzed by immunoblot analysis in response to adipose extracts incubation, and the band densitometry analysis was carried out relative to the loading control β-actin or GAPDH ( b , c ). To determine fatty acid metabolism, the culture medium with different adipose components prior to or after incubation with RBE cells for 24 h were collected for extracellular glycerol and nonestesterified fatty acid (NEFA) detection using colorimetric assays, with the data presented as the absorbance at 490 nm( d , e ). The intracellular lipids in adipose cultured RBE cells were visualized by the fluorescence dye Bodipy (20 μg/mL) using confocal microscopy analysis, using Hochest33342 (0.5 μg/mL) for nuclear staining ( f ). All the samples were prepared in triplicate, and all experiments were repeated at least three times. * P

    Article Snippet: The primary antibodies used were rabbit anti-FABP4 (EPR3579, Abcam, Cambridge, MA, USA, 1:1000), rabbit anti-claudin 1 (Abcam, Cambridge, MA, USA, 1:1000), rabbit anti-occludin (EPR8208, Abcam, Cambridge, MA, USA, 1:50,000), rabbit anti-E-cadherin (EP700Y, Abcam, Cambridge, MA, USA, 1:10,000), rabbit anti-SNAIL (Abcam, Cambridge, MA, USA, 1:1000), rabbit anti-Smad3 (EP568Y, Abcam, Cambridge, MA, USA, 1:1000), rabbit anti-β-catenin (Cell Signaling Technology, Danvers, MA, USA, 1:1000), rabbit anti-MMP2 (Cell Signaling Technology, Danvers, MA, USA,1:1000), rabbit anti-MMP9 (Cell Signaling Technology, Danvers, MA, USA,1:1000), and anti-β-actin (Beyotime Biotechnology, Haimen, China).

    Techniques: Incubation, Microscopy, Expressing, Cell Culture, Fluorescence, Confocal Microscopy, Staining

    Spastin OE does not impair tight junction localization or function. ( A ) ZO-1 localization in control and K10-rtTA; TRE-spastin epidermis. Scale-25µm. ( B ) Region where spastin-positive cells are next to spastin-negative cells in K10-rtTA; TRE-spastin tissue. Note that ZO-1 is still cortically localized in spastin-positive cells. Scale-10µm. ( C ) Localization of occludin at the cell cortex is maintained in K10-rtTA; TRE-spastin epidermis. Scale-10µm. ( D ) Biotin diffusion is blocked by occludin in K10-rtTA; TRE-spastin epidermis. Scale-10µm.

    Journal: eLife

    Article Title: A transgenic toolkit for visualizing and perturbing microtubules reveals unexpected functions in the epidermis

    doi: 10.7554/eLife.29834

    Figure Lengend Snippet: Spastin OE does not impair tight junction localization or function. ( A ) ZO-1 localization in control and K10-rtTA; TRE-spastin epidermis. Scale-25µm. ( B ) Region where spastin-positive cells are next to spastin-negative cells in K10-rtTA; TRE-spastin tissue. Note that ZO-1 is still cortically localized in spastin-positive cells. Scale-10µm. ( C ) Localization of occludin at the cell cortex is maintained in K10-rtTA; TRE-spastin epidermis. Scale-10µm. ( D ) Biotin diffusion is blocked by occludin in K10-rtTA; TRE-spastin epidermis. Scale-10µm.

    Article Snippet: The following primary antibodies were used in this study: rat anti-HA (11867423001, Sigma-Aldrich), rat anti-α-tubulin (sc-53029, Santa Cruz), rabbit anti-keratin 6 (PRB-169P, Covance), chicken anti-keratin 5/14 (generated in the Lechler lab), rabbit anti-keratin 10 (905401, Covance), rabbit anti-filaggrin (905801, Biolegend), rabbit anti-loricrin (kind gift from Colin Jamera), rat anti-BrdU (ab6326, Abcam), rabbit anti-active-caspase-3 (AF835, R and D systems), rat anti-β4 integrin (553745, BD Biosciences), rat anti-ECCD2 (kind gift from Colin Jamora), rabbit anti-keratin 1 (kind gift from Colin Jamora), mouse anti-desmoplakin (CBL173, Chemicon/Millipore), mouse anti-desmocollin-2/3 (clone 7G6, Santa Cruz), mouse anti-desmoglein-1 (610273, BD Biosciences), rabbit anti-phospho-myosin light chain 2 (Thr18/Ser19) (3674, Cell Signaling), rabbit anti-occludin (ab3172, Abcam), rabbit anti-ZO-1 (61–7300, Zymed/Invitrogen), rabbit anti-centrin1 (ab101332, Abcam), mouse anti-acetylated tubulin (T7451, Sigma-Aldrich), Rhodamine Ulex Europaeus Agglutinin 1 (RL-1062, Vector Laboratories).

    Techniques: Diffusion-based Assay

    Western blot analysis of occludin protein with or without acrolein exposure. Data represent Mean ± SE, n = 6, p = 0.337 as compared to controls. “C” represents control group and “A” represents acrolein treated group.

    Journal: PLoS ONE

    Article Title: Acute Acrolein Exposure Induces Impairment of Vocal Fold Epithelial Barrier Function

    doi: 10.1371/journal.pone.0163237

    Figure Lengend Snippet: Western blot analysis of occludin protein with or without acrolein exposure. Data represent Mean ± SE, n = 6, p = 0.337 as compared to controls. “C” represents control group and “A” represents acrolein treated group.

    Article Snippet: Primary rabbit anti-occludin antibody (ab31721) and primary mouse anti-4 HNE antibody (ab48506) were purchased from Abcam (Cambridge, MA); anti-rabbit IgG-HRP (goat) and anti-mouse IgG-HRP (goat) from Santa Cruz Biotechnology (sc-2004, sc-2005, Dallas, TX); enhanced chemiluminescene reagent (ECL) from Pierce Endogen (Rockford, IL); Alexa Fluor 488 goat anti-rabbit IgG (H+L) antibody (A-11034) from Thermo Scientific (Rockford, IL); and cyanine Cy™ 3 Goat Anti-Mouse IgG (H+L) (115-165-166) from Jackson ImmunoResearch Inc (West Grove, PA).

    Techniques: Western Blot

    Expression levels of mRNAs encoding typical tight junctional proteins. Tissues were incubated with acrolein as the concentration indicated for 3 hours. The mRNA levels of occludin (A) and claudin3 (B) were determined by qPCR. Data represent Mean ± SE, n = 6 (p = 0.259 for occludin and p = 0.556 for claudin3) as compared to controls.

    Journal: PLoS ONE

    Article Title: Acute Acrolein Exposure Induces Impairment of Vocal Fold Epithelial Barrier Function

    doi: 10.1371/journal.pone.0163237

    Figure Lengend Snippet: Expression levels of mRNAs encoding typical tight junctional proteins. Tissues were incubated with acrolein as the concentration indicated for 3 hours. The mRNA levels of occludin (A) and claudin3 (B) were determined by qPCR. Data represent Mean ± SE, n = 6 (p = 0.259 for occludin and p = 0.556 for claudin3) as compared to controls.

    Article Snippet: Primary rabbit anti-occludin antibody (ab31721) and primary mouse anti-4 HNE antibody (ab48506) were purchased from Abcam (Cambridge, MA); anti-rabbit IgG-HRP (goat) and anti-mouse IgG-HRP (goat) from Santa Cruz Biotechnology (sc-2004, sc-2005, Dallas, TX); enhanced chemiluminescene reagent (ECL) from Pierce Endogen (Rockford, IL); Alexa Fluor 488 goat anti-rabbit IgG (H+L) antibody (A-11034) from Thermo Scientific (Rockford, IL); and cyanine Cy™ 3 Goat Anti-Mouse IgG (H+L) (115-165-166) from Jackson ImmunoResearch Inc (West Grove, PA).

    Techniques: Expressing, Incubation, Concentration Assay, Real-time Polymerase Chain Reaction

    Immunohistochemical study of lipid peroxidation marker with or without acrolein exposure. (A). A typical confocal image of vocal fold epithelia. 4-HNE was stained in red on the left panel; occludin was stained in green on the middle panel; and the merged signals in yellow was present on the right panel. (B). Quantification of signal intensities of 4-HNE and occludin. Data represent Mean ± SE, n = 4, *: p

    Journal: PLoS ONE

    Article Title: Acute Acrolein Exposure Induces Impairment of Vocal Fold Epithelial Barrier Function

    doi: 10.1371/journal.pone.0163237

    Figure Lengend Snippet: Immunohistochemical study of lipid peroxidation marker with or without acrolein exposure. (A). A typical confocal image of vocal fold epithelia. 4-HNE was stained in red on the left panel; occludin was stained in green on the middle panel; and the merged signals in yellow was present on the right panel. (B). Quantification of signal intensities of 4-HNE and occludin. Data represent Mean ± SE, n = 4, *: p

    Article Snippet: Primary rabbit anti-occludin antibody (ab31721) and primary mouse anti-4 HNE antibody (ab48506) were purchased from Abcam (Cambridge, MA); anti-rabbit IgG-HRP (goat) and anti-mouse IgG-HRP (goat) from Santa Cruz Biotechnology (sc-2004, sc-2005, Dallas, TX); enhanced chemiluminescene reagent (ECL) from Pierce Endogen (Rockford, IL); Alexa Fluor 488 goat anti-rabbit IgG (H+L) antibody (A-11034) from Thermo Scientific (Rockford, IL); and cyanine Cy™ 3 Goat Anti-Mouse IgG (H+L) (115-165-166) from Jackson ImmunoResearch Inc (West Grove, PA).

    Techniques: Immunohistochemistry, Marker, Staining

    Fluoxetine downregulated MMP-9 expression and prevented degradation of the tight junction proteins in the ipsilateral cortex at 24 h after SAH. a Representative western blot bands of MMP-9, occludin, claudin-5, and ZO-1. b Densitometric quantification of MMP-9. ** p

    Journal: Journal of Neuroinflammation

    Article Title: Fluoxetine attenuates neuroinflammation in early brain injury after subarachnoid hemorrhage: a possible role for the regulation of TLR4/MyD88/NF-κB signaling pathway

    doi: 10.1186/s12974-018-1388-x

    Figure Lengend Snippet: Fluoxetine downregulated MMP-9 expression and prevented degradation of the tight junction proteins in the ipsilateral cortex at 24 h after SAH. a Representative western blot bands of MMP-9, occludin, claudin-5, and ZO-1. b Densitometric quantification of MMP-9. ** p

    Article Snippet: Then, we blocked the membranes with a nonfat dry milk buffer for 2 h, followed by incubation overnight with the following primary antibodies: polyclonal rabbit anti-NF-κB p65 (1:1000, ab16502, Abcam), monoclonal mouse anti-TLR4 (1:200, sc-293,072, Santa Cruz Biotechnology), polyclonal rabbit anti-MyD88 (1:1000, ab2064, Abcam), monoclonal rabbit anti-MMP9 (1:5000, ab137867, Abcam), polyclonal rabbit anti-ZO-1 (1:2000, ab96587, Abcam); polyclonal goat anti-claudin-5 (1:500, sc-17,668, Santa Cruz Biotechnology), monoclonal rabbit anti-occludin (1:50000, ab167161, Abcam).

    Techniques: Expressing, Western Blot