rabbit anti-cx37  (Alpha Diagnostics)

Journal: Cell Communication and Signaling : CCS

Article Title: NO, via its target Cx37, modulates calcium signal propagation selectively at myoendothelial gap junctions

doi: 10.1186/1478-811X-12-33

Figure Lengend Snippet: NO inhibited myoendothelial signal propagation in co-cultured HUVEC and HUVSMC. A/B depicts the immunohistochemical staining of Cx37 (EC, red), CD31 (EC, blue) and α-smooth-muscle actin (SMC, red) in a co-culture of endothelial and smooth muscle cells ( A , scale bar 30 μm) and an area with endothelial cells only within the same co-culture ( B , scale bar 30 μm). C depicts the number of responding cells in co-cultures of HUVEC and HUVSMC. Treatment with NO (15 min, 2 μM SNAP) significantly reduced the Ca 2+ i signal transfer from SMC to EC and also from EC to SMC whereas it did not affect the signal transfer from EC to EC and from SMC to SMC (n = 13-21, w = C = 5; p < 0.05). In the remaining responding cells, the time delay (D) was significantly increased from SMC to EC whereas the signal spread faster from SMC to SMC after exposure to NO. The amplitude of the Ca 2+ i increase (in the remaining responding cells) was unchanged in all cells (n = 17-106, w = 14-21, C = 6; *: p < 0.05, con vs. NO, NG). E . The decrease of the mechanically induced calcium rise in the initial stimulated cells was reduced by incubation with SNAP (15 min, 2 μM).

Article Snippet: The cells were incubated with primary antibodies: rabbit polyclonal anti-Cx37 (1.25 μg/ml, Invitrogen, Cat. No. 42–4400, or 2.5 μg/ml Alpha Diagnostics, Cat. No. Cx37A11-A) and mouse monoclonal anti-CD31 (3 μg/ml, Abcam, Cat. No. ab9409), all diluted in PBS containing 1% BSA overnight.

Techniques: Cell Culture, Immunohistochemical staining, Staining, Co-Culture Assay, Incubation

Journal: Cell Communication and Signaling : CCS

Article Title: NO, via its target Cx37, modulates calcium signal propagation selectively at myoendothelial gap junctions

doi: 10.1186/1478-811X-12-33

Figure Lengend Snippet: Distribution of Cx37 within the internal elastic lamina. A . Representative 2-photon-image of a small resistance artery for visualization of the internal elastic lamina and the connexins. Cx37 (red) located in the small holes (dark dots) within the internal elastic lamina (autofluorescence, green). Arrows indicate some of the holes in which Cx37 could be detected; scale bar: 25 μm. B depicts the summary of n = 7-10 experiments (3 vessels each) revealing the percentage of holes in the internal elastic lamina that contain the different vascular Cx (*: p < 0.05 vs. Cx40; #: p < 0.05 vs. Cx45, NG).

Article Snippet: The cells were incubated with primary antibodies: rabbit polyclonal anti-Cx37 (1.25 μg/ml, Invitrogen, Cat. No. 42–4400, or 2.5 μg/ml Alpha Diagnostics, Cat. No. Cx37A11-A) and mouse monoclonal anti-CD31 (3 μg/ml, Abcam, Cat. No. ab9409), all diluted in PBS containing 1% BSA overnight.

Techniques:

Journal: Cell Communication and Signaling : CCS

Article Title: NO, via its target Cx37, modulates calcium signal propagation selectively at myoendothelial gap junctions

doi: 10.1186/1478-811X-12-33

Figure Lengend Snippet: Location of vascular Cx across the vessel wall. A . Confocal images of triple (Cx, α-actin, CD31) immunohistochemical stainings of Cx40, Cx45, Cx37, and Cx43 in small resistance arteries. Left panel: Overlay of a z-series in xy-direction, right panel: Cross section (slice) of the z-stack in yz-direction along the yellow lines in the z-stack. The arrows depict the Cx expression in ECL (yellow) or beyond EC and within SMC (SMCL, white), scale bars: 10 μm. B . Summary of the Cx distribution within the ECL and SMCL for all Cx (n = 4-8, at least 3 vessels each; *: p < 0.001).

Article Snippet: The cells were incubated with primary antibodies: rabbit polyclonal anti-Cx37 (1.25 μg/ml, Invitrogen, Cat. No. 42–4400, or 2.5 μg/ml Alpha Diagnostics, Cat. No. Cx37A11-A) and mouse monoclonal anti-CD31 (3 μg/ml, Abcam, Cat. No. ab9409), all diluted in PBS containing 1% BSA overnight.

Techniques: Immunohistochemical staining, Expressing

Journal: Animal Cells and Systems

Article Title: Beneficial effect of simvastatin on human umbilical vein endothelial cells gap junctions induced by TNF-α

doi: 10.1080/19768354.2021.2023037

Figure Lengend Snippet: List of oligonucleotide primers used for real-time PCR analysis.

Article Snippet: The membranes were blocked with with blocking buffer (PBS containing 3%BSA and 0.1% sodium azide), for the reason that the molecular weight of Cx isoforms (37, 40, 43) and β-actin (42KD) are very close, then incubated for 1 h at room temperature with the following primary antibodies: rabbit polyclonal anti-Cx37 antibody (rabbit polyclonal Cx37 antibody raised against human; Cat. No. SAB4501180, Sigma-Aldrich), anti-Cx40 antibody (rabbit polyclonal Cx40 antibody raised against human; Cat. No. SAB1304973, Sigma-Aldrich) and anti-Cx43 antibody (rabbit polyclonal Cx43 antibody raised against human; Cat. No. C6219, Sigma-Aldrich) (1:1000) and anti-β actin antibody (rabbit polyclonal antibody raised against human; Cat. No. SAB5500001, Sigma-Aldrich) (1:1000).

Techniques: Real-time Polymerase Chain Reaction, Sequencing

Journal: Animal Cells and Systems

Article Title: Beneficial effect of simvastatin on human umbilical vein endothelial cells gap junctions induced by TNF-α

doi: 10.1080/19768354.2021.2023037

Figure Lengend Snippet: Immunofluorescence results showing effect of simvastatin on GJIC targeting Cx37, Cx40 and Cx43. Cx37 (A) and Cx40 (B) expression was downregulated in gap junctions of HUVECs treated with TNF-α relative to the control group. Treatment of HUVECs with simvastatin upregulated Cx37 and Cx40 expression. Similarly, HUVECs treated with TNF-α exhibited Cx43 up-regulation (C) although simvastatin reversed this phenomenon. D: Quantitative comparison of FITC staining intensity normalized to control. n = 6, * P < 0.05, ** P < 0.01 vs. Control; # P < 0.05, ## P < 0.01 vs. TNF-α (+) Simvastain (−) group. Bar = 30 μm.

Article Snippet: The membranes were blocked with with blocking buffer (PBS containing 3%BSA and 0.1% sodium azide), for the reason that the molecular weight of Cx isoforms (37, 40, 43) and β-actin (42KD) are very close, then incubated for 1 h at room temperature with the following primary antibodies: rabbit polyclonal anti-Cx37 antibody (rabbit polyclonal Cx37 antibody raised against human; Cat. No. SAB4501180, Sigma-Aldrich), anti-Cx40 antibody (rabbit polyclonal Cx40 antibody raised against human; Cat. No. SAB1304973, Sigma-Aldrich) and anti-Cx43 antibody (rabbit polyclonal Cx43 antibody raised against human; Cat. No. C6219, Sigma-Aldrich) (1:1000) and anti-β actin antibody (rabbit polyclonal antibody raised against human; Cat. No. SAB5500001, Sigma-Aldrich) (1:1000).

Techniques: Immunofluorescence, Expressing, Staining

Journal: Animal Cells and Systems

Article Title: Beneficial effect of simvastatin on human umbilical vein endothelial cells gap junctions induced by TNF-α

doi: 10.1080/19768354.2021.2023037

Figure Lengend Snippet: Differential expression of Cx37, Cx40 and Cx43 mRNAs in HUVECs across groups. Cx37 and Cx40 had a 2-fold downregulation, whereas Cx43 was upregulated relative to the control group. Cx37 and Cx40 were significantly upregulated in HUVECs treated with TNF-α in combination with simvastatin relative to those treated with TNF-α alone. Cx43 was significantly downregulated. Data presented are means ± SEM, n = 6, ** P < 0.01 versus only TNF-α treatment group.

Article Snippet: The membranes were blocked with with blocking buffer (PBS containing 3%BSA and 0.1% sodium azide), for the reason that the molecular weight of Cx isoforms (37, 40, 43) and β-actin (42KD) are very close, then incubated for 1 h at room temperature with the following primary antibodies: rabbit polyclonal anti-Cx37 antibody (rabbit polyclonal Cx37 antibody raised against human; Cat. No. SAB4501180, Sigma-Aldrich), anti-Cx40 antibody (rabbit polyclonal Cx40 antibody raised against human; Cat. No. SAB1304973, Sigma-Aldrich) and anti-Cx43 antibody (rabbit polyclonal Cx43 antibody raised against human; Cat. No. C6219, Sigma-Aldrich) (1:1000) and anti-β actin antibody (rabbit polyclonal antibody raised against human; Cat. No. SAB5500001, Sigma-Aldrich) (1:1000).

Techniques: Expressing

Journal: Animal Cells and Systems

Article Title: Beneficial effect of simvastatin on human umbilical vein endothelial cells gap junctions induced by TNF-α

doi: 10.1080/19768354.2021.2023037

Figure Lengend Snippet: Cx37, Cx40 and Cx43 proteins were differentially expressed in HUVECs in different treatment groups. A: Western blots. 1: Control, 2: TNF-α (+)/Simvastatin (−), 3: TNF-α (+)/Simvastatin (+). B: qRT-PCR results showing relative transcript levels. Data presented are means ± SEM, n = 6, ** P < 0.01 versus only TNF-α treatment group.

Article Snippet: The membranes were blocked with with blocking buffer (PBS containing 3%BSA and 0.1% sodium azide), for the reason that the molecular weight of Cx isoforms (37, 40, 43) and β-actin (42KD) are very close, then incubated for 1 h at room temperature with the following primary antibodies: rabbit polyclonal anti-Cx37 antibody (rabbit polyclonal Cx37 antibody raised against human; Cat. No. SAB4501180, Sigma-Aldrich), anti-Cx40 antibody (rabbit polyclonal Cx40 antibody raised against human; Cat. No. SAB1304973, Sigma-Aldrich) and anti-Cx43 antibody (rabbit polyclonal Cx43 antibody raised against human; Cat. No. C6219, Sigma-Aldrich) (1:1000) and anti-β actin antibody (rabbit polyclonal antibody raised against human; Cat. No. SAB5500001, Sigma-Aldrich) (1:1000).

Techniques: Western Blot, Quantitative RT-PCR

Journal: bioRxiv

Article Title: GATA2 controls lymphatic endothelial cell junctional integrity and lymphovenous valve morphogenesis through miR-126

doi: 10.1101/660068

Figure Lengend Snippet: E12.0 Wild type and Lyve1-Cre;Gata2 f/f littermates or Tg(Prox1-tdTomato) and Tg(Prox1-tdTomato); Lyve1-Cre;Gata2 f/f littermates were frontally sectioned and IHC was performed for LVV-EC markers (PROX1, tdTomato, FOXC2, ITGA5, ITGA9), ECM components (FN1, LAMA5), and cell junction proteins (CD31, VE-Cadherin, GJA4, CLDN5). No obvious differences were observed between control and mutant samples. Abbreviations: A, artery; LS, lymph sac. Measuring bar: (A-P) 100 µm. Statistics: n=3 embryos and 6 LVV complexes per genotype per antibody.

Article Snippet: Primary antibodies for immunohistochemistry: rabbit anti-PROX1 (Cat# 11-002, Angiobio, San Diego, CA, USA), goat anti-human PROX1 (Cat# AF2727, R&D Systems, Minneapolis, MN, USA), sheep anti-mouse FOXC2 (Cat# AF6989, R&D Systems, Minneapolis, MN, USA), goat anti-mouse VEGRF3 (Cat# AF743, R&D Systems, Minneapolis, MN, USA), rat anti-mouse CD31 (Cat# 553370, BD Pharmingen, San Jose, CA, USA), goat anti-mouse ITGA9 (Cat # AF3827, R&D Systems, Minneapolis, MN, USA), rat anti-mouse VE-Cadherin (Cat# 550548, BD Pharmingen, San Jose, CA, USA), hamster anti-mouse PDPN (Cat# 127401, Biolegend, San Diego, CA, USA), rat anti-mouse ITGA5 (Cat #553319, BD Pharmingen, San Jose, CA, USA), goat anti-mouse GATA2 (Cat #AF2046, R&D Systems, Minneapolis, MN, USA), rabbit anti-mouse CX37 (Cat #40-4200, Life technologies, Grand Island, NY, USA), rabbit anti-mouse LAMA5 (Abcam, Cambridge, MA, USA), rabbit anti-human Fibronectin (Cat# ab2413, Abcam, Cambridge, MA, USA), Goat anti-human ANGPT2 (Cat# AF623, R&D Systems, Minneapolis, MN, USA), goat anti-mouse EGFL7 (Cat # AF3089, R&D Systems, Minneapolis, MN, USA), rabbit anti-mouse CLDN5 (Cat #34-1600, Thermo Fisher Scientific, Rockford, IL, USA), rabbit anti-mouse LYVE-1 (Cat # 11-034, Angiobio, San Diego, CA, USA).

Techniques: Mutagenesis