polyclonal rabbit anti-connexin 43 cx43 Search Results


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  • 94
    Thermo Fisher polyclonal rabbit anti connexin43 cx43
    HUVEC and cFB acquired the cardiac phenotype through cell fusion with cardiomyocytes. (A) LacZ -expressing cardiomyocytes of neonatal rats were cocultured with GFP + HUVEC (a–d) or GFP + cFB (e–h) and stained with mouse monoclonal anti-cTnT (red) and rabbit <t>polyclonal</t> anti-β-gal antibodies (blue). Merged images were obtained from the same confocal plane. GFP + HUVEC and GFP + cFB (a and e, arrow) expressed cTnT (b and f, arrow) and β-gal (c and g, arrow) in the same cell (merged on d and h). Arrowheads indicate the nonfused cardiomyocytes. Bars, 50 μm. (B) LacZ -expressing cardiomyocytes of neonatal rats were cocultured with GFP + HUVEC (a–f) or GFP + cFB (g–l) and stained with mouse monoclonal anti-cTnT (red) and goat polyclonal anti-GATA4 or rabbit polyclonal anti-ANF or <t>anti-connexin43</t> antibodies (blue). cTnT-expressing GFP + HUVEC (a, c, and e) and cTnT-expressing GFP + cFB (g, i, and k) expressed GATA4 (b and h, arrowhead), ANF (d and j, arrowhead), and connexin43 (f and l, arrowhead). Bars, 50 μm.
    Polyclonal Rabbit Anti Connexin43 Cx43, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore polyclonal rabbit anti connexin 43 cx43
    rAdVax immunotherapy recovered the injured heart tissue of chronically T. cruzi -infected mice. ( A ) Chronically Colombian T. cruzi strain-infected mice were evaluated for heart injury markers pre-therapy (120 dpi) or primed-boosted with 2 × 10 8 plaque-forming units (PFU) of rAdCtrl or a mixture of 10 8 PFU of each adenovirus vaccine preparation (rAdASP2+rAdTS; rAdVax). Mortality was recorded until 230 dpi (110 days post-therapy; dpt), when the surviving mice were analyzed for heart injury markers. ( B ) Kaplan-Meier curve representing the percentages of surviving mice (14–20 mice/group in two independent experiments). ( C ) Relative spleen weight (mg of spleen/g of body) and quantitative immunohistochemical staining (IHS) data for T. cruzi parasitism (nests/100 microscopic fields) in the heart tissue of chronically infected mice (120 and 230 dpi, respectively, pre- and post-therapy). ( D ) IHS showing fibronectin (FN)-stained areas in representative cardiac tissue sections of noninfected (NI) controls and chronically T. cruzi -infected mice pre- (120 dpi) and post-therapy (230 dpi; 110 dpt) with rAdVax. ( E ) Quantification of the FN-stained area (%) and <t>connexin</t> 43 <t>(Cx43)-containing</t> gap junction distances detected using IHS staining of heart tissue sections of NI controls or T. cruzi -infected mice pre- and post-therapy with rAdVax. ( F ) Evaluation of CK-MB activity in the serum of NI controls and T. cruzi -infected mice pre- and post-therapy with rAdVax. The data are presented as the means ± SD. ** P
    Polyclonal Rabbit Anti Connexin 43 Cx43, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 171 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Abcam polyclonal rabbit anti connexin 43 cx43 primary antibody
    rAdVax immunotherapy recovered the injured heart tissue of chronically T. cruzi -infected mice. ( A ) Chronically Colombian T. cruzi strain-infected mice were evaluated for heart injury markers pre-therapy (120 dpi) or primed-boosted with 2 × 10 8 plaque-forming units (PFU) of rAdCtrl or a mixture of 10 8 PFU of each adenovirus vaccine preparation (rAdASP2+rAdTS; rAdVax). Mortality was recorded until 230 dpi (110 days post-therapy; dpt), when the surviving mice were analyzed for heart injury markers. ( B ) Kaplan-Meier curve representing the percentages of surviving mice (14–20 mice/group in two independent experiments). ( C ) Relative spleen weight (mg of spleen/g of body) and quantitative immunohistochemical staining (IHS) data for T. cruzi parasitism (nests/100 microscopic fields) in the heart tissue of chronically infected mice (120 and 230 dpi, respectively, pre- and post-therapy). ( D ) IHS showing fibronectin (FN)-stained areas in representative cardiac tissue sections of noninfected (NI) controls and chronically T. cruzi -infected mice pre- (120 dpi) and post-therapy (230 dpi; 110 dpt) with rAdVax. ( E ) Quantification of the FN-stained area (%) and <t>connexin</t> 43 <t>(Cx43)-containing</t> gap junction distances detected using IHS staining of heart tissue sections of NI controls or T. cruzi -infected mice pre- and post-therapy with rAdVax. ( F ) Evaluation of CK-MB activity in the serum of NI controls and T. cruzi -infected mice pre- and post-therapy with rAdVax. The data are presented as the means ± SD. ** P
    Polyclonal Rabbit Anti Connexin 43 Cx43 Primary Antibody, supplied by Abcam, used in various techniques. Bioz Stars score: 86/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/polyclonal rabbit anti connexin 43 cx43 primary antibody/product/Abcam
    Average 86 stars, based on 6 article reviews
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    93
    Santa Cruz Biotechnology connexin 43 cx43 polyclonal rabbit antibodies
    rAdVax immunotherapy recovered the injured heart tissue of chronically T. cruzi -infected mice. ( A ) Chronically Colombian T. cruzi strain-infected mice were evaluated for heart injury markers pre-therapy (120 dpi) or primed-boosted with 2 × 10 8 plaque-forming units (PFU) of rAdCtrl or a mixture of 10 8 PFU of each adenovirus vaccine preparation (rAdASP2+rAdTS; rAdVax). Mortality was recorded until 230 dpi (110 days post-therapy; dpt), when the surviving mice were analyzed for heart injury markers. ( B ) Kaplan-Meier curve representing the percentages of surviving mice (14–20 mice/group in two independent experiments). ( C ) Relative spleen weight (mg of spleen/g of body) and quantitative immunohistochemical staining (IHS) data for T. cruzi parasitism (nests/100 microscopic fields) in the heart tissue of chronically infected mice (120 and 230 dpi, respectively, pre- and post-therapy). ( D ) IHS showing fibronectin (FN)-stained areas in representative cardiac tissue sections of noninfected (NI) controls and chronically T. cruzi -infected mice pre- (120 dpi) and post-therapy (230 dpi; 110 dpt) with rAdVax. ( E ) Quantification of the FN-stained area (%) and <t>connexin</t> 43 <t>(Cx43)-containing</t> gap junction distances detected using IHS staining of heart tissue sections of NI controls or T. cruzi -infected mice pre- and post-therapy with rAdVax. ( F ) Evaluation of CK-MB activity in the serum of NI controls and T. cruzi -infected mice pre- and post-therapy with rAdVax. The data are presented as the means ± SD. ** P
    Connexin 43 Cx43 Polyclonal Rabbit Antibodies, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/connexin 43 cx43 polyclonal rabbit antibodies/product/Santa Cruz Biotechnology
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    95
    Thermo Fisher polyclonal rabbit anti cx43
    MMC induces alteration in <t>Cx43</t> association with ZO-1. Cells were treated with prewarmed media containing 5 μM MMC for 60 minutes or media alone (CTRL). ( A ) Coimmunoprecipitation with a rabbit <t>polyclonal</t> anti-Cx43 was performed followed by immunoblotting
    Polyclonal Rabbit Anti Cx43, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 31 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    98
    Cell Signaling Technology Inc rabbit polyclonal anti connexin 43
    Results from in-cell Western analyses showing effects of kinase inhibition on total <t>Cx43</t> expression in rat primary VSMCs following incubation in the presence of (A) 8Br-cAMP (100 μM), (B) 8Br-cGMP (100 μM), (C) BAY (100 nM), and (D) DADS (50 μM) for 270 min . (A) Inhibition of PKA (by PKI), PKG (by DT2), or PKC (by CalC) each independently subdued 8Br-cAMP-induced increase in Cx43 expression. (B,C) Only PKG and PKC inhibition reversed the effects of 8Br-cGMP and BAY, respectively. (D) Stimulation of Cx43 expression in the presence of DADS was not dependent on PKA, PKG, or PKC. (E) Treatment with any individual kinase inhibitor did not affect total basal Cx43 expression compared with vehicle controls. Data are mean ± SE from n = 4–7 for each treatment. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p
    Rabbit Polyclonal Anti Connexin 43, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 98/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti connexin 43/product/Cell Signaling Technology Inc
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    92
    Thermo Fisher rabbit anti phospho cx43
    Results from in-cell Western analyses showing effects of kinase inhibition on total <t>Cx43</t> expression in rat primary VSMCs following incubation in the presence of (A) 8Br-cAMP (100 μM), (B) 8Br-cGMP (100 μM), (C) BAY (100 nM), and (D) DADS (50 μM) for 270 min . (A) Inhibition of PKA (by PKI), PKG (by DT2), or PKC (by CalC) each independently subdued 8Br-cAMP-induced increase in Cx43 expression. (B,C) Only PKG and PKC inhibition reversed the effects of 8Br-cGMP and BAY, respectively. (D) Stimulation of Cx43 expression in the presence of DADS was not dependent on PKA, PKG, or PKC. (E) Treatment with any individual kinase inhibitor did not affect total basal Cx43 expression compared with vehicle controls. Data are mean ± SE from n = 4–7 for each treatment. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p
    Rabbit Anti Phospho Cx43, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Thermo Fisher rabbit anti phospho cx43 ser373
    Schematic showing potential roles of astrocytic <t>Cx43,</t> hemichannels, and GJIC during OGD/R injury. Under normal conditions, astrocytic Cx43 is expressed in the plasma membrane and assembled into hemichannels that are normally closed. Hemichannel-hemichannel interactions induce the formation of GJIC between adjacent astrocytes, which permits the exchange of ions and small molecules; also, plasma membrane’s Cx43 was phosphorylated at Ser368 site. In such circumstances, astrocytes, together with those resting microglia, function as a supportive assistant for healthy neurons. OGD/R injury caused abnormal hemichannel opening and consequent substantial astrocytic ATP release. It also induced microglial activation with a predominance of the pro-inflammatory cytokine-releasing M1 subtype. Extracellular ATP induced further microglial activation and pro-inflammatory cytokine release, and these pro-inflammatory cytokines induced further opening of astrocytic hemichannels. SalB reversed these effects and thus provided protection against OGD/R injury. This suggests the existence of a vicious cycle in which astrocytic hemichannel opening and pro-inflammatory microglial activation reinforce each other following OGD/R injury. This vicious cycle may account for secondary injury and extended damage after OGD/R injury; OGD/R injury caused gap junction internalization, which may account for the astrocytic uncoupling events. It also decreased plasma membrane levels of Ser368-phosphorylated Cx43 while increasing plasma membrane levels of <t>Ser373-phosphorylated</t> Cx43, Ser265-phosphorylated Cx43, and Src’s Tyr416-phosphorylated activated form. The activated Src may well have phosphorylated Cx43 at Tyr265 and further induced gap junction internalization or autophagy. SalB directly inhibits Src, which may allow it to exert protective effects by attenuating Cx43 internalization. CBX, a non-selective hemichannel and GJIC inhibitor, did not apparently affect Cx43 phosphorylation, but it inhibited PKC and Src activity
    Rabbit Anti Phospho Cx43 Ser373, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    HUVEC and cFB acquired the cardiac phenotype through cell fusion with cardiomyocytes. (A) LacZ -expressing cardiomyocytes of neonatal rats were cocultured with GFP + HUVEC (a–d) or GFP + cFB (e–h) and stained with mouse monoclonal anti-cTnT (red) and rabbit polyclonal anti-β-gal antibodies (blue). Merged images were obtained from the same confocal plane. GFP + HUVEC and GFP + cFB (a and e, arrow) expressed cTnT (b and f, arrow) and β-gal (c and g, arrow) in the same cell (merged on d and h). Arrowheads indicate the nonfused cardiomyocytes. Bars, 50 μm. (B) LacZ -expressing cardiomyocytes of neonatal rats were cocultured with GFP + HUVEC (a–f) or GFP + cFB (g–l) and stained with mouse monoclonal anti-cTnT (red) and goat polyclonal anti-GATA4 or rabbit polyclonal anti-ANF or anti-connexin43 antibodies (blue). cTnT-expressing GFP + HUVEC (a, c, and e) and cTnT-expressing GFP + cFB (g, i, and k) expressed GATA4 (b and h, arrowhead), ANF (d and j, arrowhead), and connexin43 (f and l, arrowhead). Bars, 50 μm.

    Journal: The Journal of Cell Biology

    Article Title: Cardiomyocytes fuse with surrounding noncardiomyocytes and reenter the cell cycle

    doi: 10.1083/jcb.200312111

    Figure Lengend Snippet: HUVEC and cFB acquired the cardiac phenotype through cell fusion with cardiomyocytes. (A) LacZ -expressing cardiomyocytes of neonatal rats were cocultured with GFP + HUVEC (a–d) or GFP + cFB (e–h) and stained with mouse monoclonal anti-cTnT (red) and rabbit polyclonal anti-β-gal antibodies (blue). Merged images were obtained from the same confocal plane. GFP + HUVEC and GFP + cFB (a and e, arrow) expressed cTnT (b and f, arrow) and β-gal (c and g, arrow) in the same cell (merged on d and h). Arrowheads indicate the nonfused cardiomyocytes. Bars, 50 μm. (B) LacZ -expressing cardiomyocytes of neonatal rats were cocultured with GFP + HUVEC (a–f) or GFP + cFB (g–l) and stained with mouse monoclonal anti-cTnT (red) and goat polyclonal anti-GATA4 or rabbit polyclonal anti-ANF or anti-connexin43 antibodies (blue). cTnT-expressing GFP + HUVEC (a, c, and e) and cTnT-expressing GFP + cFB (g, i, and k) expressed GATA4 (b and h, arrowhead), ANF (d and j, arrowhead), and connexin43 (f and l, arrowhead). Bars, 50 μm.

    Article Snippet: The following antibodies were used for immunostaining: mouse monoclonal anti-cTnT (RV-C2, DSMZ-Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH), goat polyclonal anti-cTnT, goat polyclonal anti-GATA4 (Santa Cruz Biotechnology, Inc.), rabbit polyclonal anti-ANF (Peninsula Laboratories), rabbit polyclonal anti-connexin43 (Zymed Laboratories), rabbit polyclonal anti-desmin, rabbit polyclonal anti-vWF, mouse monoclonal anti–rat Ki67, mouse monoclonal anti–human Ki67 (Dako Cytomation), mouse monoclonal anti-β-gal, rabbit polyclonal anti-β-gal (CHEMICON International, Inc.), mouse monoclonal anti-vimentin, mouse monoclonal anti-Cre (Sigma-Aldrich), rabbit polyclonal anti-PH3 (Upstate Biotechnology), mouse monoclonal anti-cyclinB1 (Neomarkers), and rabbit polyclonal anti-RFP (MBL International Corporation).

    Techniques: Expressing, Staining

    rAdVax immunotherapy recovered the injured heart tissue of chronically T. cruzi -infected mice. ( A ) Chronically Colombian T. cruzi strain-infected mice were evaluated for heart injury markers pre-therapy (120 dpi) or primed-boosted with 2 × 10 8 plaque-forming units (PFU) of rAdCtrl or a mixture of 10 8 PFU of each adenovirus vaccine preparation (rAdASP2+rAdTS; rAdVax). Mortality was recorded until 230 dpi (110 days post-therapy; dpt), when the surviving mice were analyzed for heart injury markers. ( B ) Kaplan-Meier curve representing the percentages of surviving mice (14–20 mice/group in two independent experiments). ( C ) Relative spleen weight (mg of spleen/g of body) and quantitative immunohistochemical staining (IHS) data for T. cruzi parasitism (nests/100 microscopic fields) in the heart tissue of chronically infected mice (120 and 230 dpi, respectively, pre- and post-therapy). ( D ) IHS showing fibronectin (FN)-stained areas in representative cardiac tissue sections of noninfected (NI) controls and chronically T. cruzi -infected mice pre- (120 dpi) and post-therapy (230 dpi; 110 dpt) with rAdVax. ( E ) Quantification of the FN-stained area (%) and connexin 43 (Cx43)-containing gap junction distances detected using IHS staining of heart tissue sections of NI controls or T. cruzi -infected mice pre- and post-therapy with rAdVax. ( F ) Evaluation of CK-MB activity in the serum of NI controls and T. cruzi -infected mice pre- and post-therapy with rAdVax. The data are presented as the means ± SD. ** P

    Journal: PLoS Pathogens

    Article Title: A Human Type 5 Adenovirus-Based Trypanosoma cruzi Therapeutic Vaccine Re-programs Immune Response and Reverses Chronic Cardiomyopathy

    doi: 10.1371/journal.ppat.1004594

    Figure Lengend Snippet: rAdVax immunotherapy recovered the injured heart tissue of chronically T. cruzi -infected mice. ( A ) Chronically Colombian T. cruzi strain-infected mice were evaluated for heart injury markers pre-therapy (120 dpi) or primed-boosted with 2 × 10 8 plaque-forming units (PFU) of rAdCtrl or a mixture of 10 8 PFU of each adenovirus vaccine preparation (rAdASP2+rAdTS; rAdVax). Mortality was recorded until 230 dpi (110 days post-therapy; dpt), when the surviving mice were analyzed for heart injury markers. ( B ) Kaplan-Meier curve representing the percentages of surviving mice (14–20 mice/group in two independent experiments). ( C ) Relative spleen weight (mg of spleen/g of body) and quantitative immunohistochemical staining (IHS) data for T. cruzi parasitism (nests/100 microscopic fields) in the heart tissue of chronically infected mice (120 and 230 dpi, respectively, pre- and post-therapy). ( D ) IHS showing fibronectin (FN)-stained areas in representative cardiac tissue sections of noninfected (NI) controls and chronically T. cruzi -infected mice pre- (120 dpi) and post-therapy (230 dpi; 110 dpt) with rAdVax. ( E ) Quantification of the FN-stained area (%) and connexin 43 (Cx43)-containing gap junction distances detected using IHS staining of heart tissue sections of NI controls or T. cruzi -infected mice pre- and post-therapy with rAdVax. ( F ) Evaluation of CK-MB activity in the serum of NI controls and T. cruzi -infected mice pre- and post-therapy with rAdVax. The data are presented as the means ± SD. ** P

    Article Snippet: Other antibodies included an anti-F4/80 polyclonal antibody (Caltag, USA), polyclonal rabbit anti-connexin 43 (Cx43) (Sigma, USA), polyclonal rabbit anti-mouse fibronectin (FN) (Gibco-BRL, USA), biotinylated anti-rat immunoglobulin (Dako, Denmark) and biotinylated anti-rabbit immunoglobulin and peroxidase-streptavidin complex (Amersham, UK).

    Techniques: Infection, Mouse Assay, Immunohistochemistry, Staining, Activity Assay

    Mouse RBCs express pannexin 1 (Panx1). A : topological schematic of Panx1 indicates the location of the epitopes targeted by the three antibodies used. Scissors indicate caspase cleavage site on COOH terminus. B : analysis of protein content in red blood cell (RBC) ghosts via Western blotting with carboxy-terminus (CT) and second extracellular loop (EL2) antibodies shows Panx1 expression in wild-type (WT) but not Panx1 −/− (KO) RBCs. No connexin 43 (Cx43) expression was detected. Far left Cx43 band is 20 µg mouse lung lysate, used as a positive control. NS, nonspecific bands. C : confocal immunofluorescence microscopy of RBCs using CT and EL2 antibodies shows expression of Panx1 in RBCs. Scale bar, 20 µm. D : 3,3′-diaminobenzidine (DAB) staining of RBCs trapped in afferent arteriole kidney tissue slices using amino-terminus (NT) antibody confirms Panx1 expression on both endothelium and RBCs, stained dark brown. Arrows denote RBCs. No DAB staining was detected when sections were probed with Cx43 antibody, IgG antibody, or secondary antibody only. Scale bar, 20 µm.

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Possible roles for ATP release from RBCs exclude the cAMP-mediated Panx1 pathway

    doi: 10.1152/ajpcell.00178.2017

    Figure Lengend Snippet: Mouse RBCs express pannexin 1 (Panx1). A : topological schematic of Panx1 indicates the location of the epitopes targeted by the three antibodies used. Scissors indicate caspase cleavage site on COOH terminus. B : analysis of protein content in red blood cell (RBC) ghosts via Western blotting with carboxy-terminus (CT) and second extracellular loop (EL2) antibodies shows Panx1 expression in wild-type (WT) but not Panx1 −/− (KO) RBCs. No connexin 43 (Cx43) expression was detected. Far left Cx43 band is 20 µg mouse lung lysate, used as a positive control. NS, nonspecific bands. C : confocal immunofluorescence microscopy of RBCs using CT and EL2 antibodies shows expression of Panx1 in RBCs. Scale bar, 20 µm. D : 3,3′-diaminobenzidine (DAB) staining of RBCs trapped in afferent arteriole kidney tissue slices using amino-terminus (NT) antibody confirms Panx1 expression on both endothelium and RBCs, stained dark brown. Arrows denote RBCs. No DAB staining was detected when sections were probed with Cx43 antibody, IgG antibody, or secondary antibody only. Scale bar, 20 µm.

    Article Snippet: Connexin 43 (Cx43) rabbit polyclonal antibody was purchased from Sigma Aldrich (no. C6219).

    Techniques: Western Blot, Expressing, Positive Control, Immunofluorescence, Microscopy, Staining

    Expression of meningeal markers in E13.5 ch mice. Corresponding merged images of immunohistochemical staining (A–D) of CD31 (red) and Cx43 (green) with Hoechst stained nuclei (blue). In situ hybridization of Cx43 (E–H), Bmp7 (I–L),

    Journal:

    Article Title: Impaired meningeal development in association with apical expansion of calvarial bone osteogenesis in the Foxc1 mutant

    doi: 10.1111/j.1469-7580.2008.00893.x

    Figure Lengend Snippet: Expression of meningeal markers in E13.5 ch mice. Corresponding merged images of immunohistochemical staining (A–D) of CD31 (red) and Cx43 (green) with Hoechst stained nuclei (blue). In situ hybridization of Cx43 (E–H), Bmp7 (I–L),

    Article Snippet: The following primary antibodies were used: rat anti-mouse CD31 (PECAM-1) monoclonal antibody (BD Pharmingen, USA) and rabbit anti-connexin43 (Cx43) antibody (Sigma-Aldrich, USA).

    Techniques: Expressing, Mouse Assay, Immunohistochemistry, Staining, In Situ Hybridization

    Expression of meningeal markers in E14.5 ch mice. The corresponding merged images of immunohistochemical staining (A–D) of CD31 (red) and Cx43 (green) with Hoechst stained nuclei (blue). In situ hybridization of Cx43 (E–H), Bmp7 (I–L),

    Journal:

    Article Title: Impaired meningeal development in association with apical expansion of calvarial bone osteogenesis in the Foxc1 mutant

    doi: 10.1111/j.1469-7580.2008.00893.x

    Figure Lengend Snippet: Expression of meningeal markers in E14.5 ch mice. The corresponding merged images of immunohistochemical staining (A–D) of CD31 (red) and Cx43 (green) with Hoechst stained nuclei (blue). In situ hybridization of Cx43 (E–H), Bmp7 (I–L),

    Article Snippet: The following primary antibodies were used: rat anti-mouse CD31 (PECAM-1) monoclonal antibody (BD Pharmingen, USA) and rabbit anti-connexin43 (Cx43) antibody (Sigma-Aldrich, USA).

    Techniques: Expressing, Mouse Assay, Immunohistochemistry, Staining, In Situ Hybridization

    Induction of Cx43 by inhibition of IPDE3 and IPDE4. Mesangial cells were treated with IPDE3, IPDE4, IPDE3 plus IPDE4, IBMX or IPDE2 plus IPDE5 for 24 h and subjected to Western blot analysis of Cx43 (a). Densitometric analysis of the data is shown

    Journal: British Journal of Pharmacology

    Article Title: Profiling of functional phosphodiesterase in mesangial cells using a CRE-SEAP-based reporting system

    doi: 10.1038/sj.bjp.0706785

    Figure Lengend Snippet: Induction of Cx43 by inhibition of IPDE3 and IPDE4. Mesangial cells were treated with IPDE3, IPDE4, IPDE3 plus IPDE4, IBMX or IPDE2 plus IPDE5 for 24 h and subjected to Western blot analysis of Cx43 (a). Densitometric analysis of the data is shown

    Article Snippet: After blocking with 3% bovine serum albumin in phosphate-buffered saline (PBS), the membranes were incubated with anti-vasodilator-stimulated phosphoprotein (VASP) antibody (dilution 1 : 1000; Chemicon International, Temecula, CA, U.S.A.), anti-connexin43 (Cx43) antibody (dilution 1 : 2000; Sigma), anti-caspase-3 antibody (dilution 1 : 1000; Cell Signaling, Beverly, MA, U.S.A.), or anti-phospho-mitogen-activated protein (MAP) kinase antibody (dilution 1 : 1000; Cell Signaling).

    Techniques: Inhibition, Western Blot

    Olfactory bulb astrocyte cultures respond to iodomelatonin. Top: control cultures treated with fresh medium. Bottom: cultures treated with 3 nM iodomelatonin in fresh medium. Connexin43 immunoreactivity increased in the treated cultures compared to controls.

    Journal: Neuroscience

    Article Title: Melatonin in the mammalian olfactory bulb

    doi: 10.1016/j.neuroscience.2013.12.033

    Figure Lengend Snippet: Olfactory bulb astrocyte cultures respond to iodomelatonin. Top: control cultures treated with fresh medium. Bottom: cultures treated with 3 nM iodomelatonin in fresh medium. Connexin43 immunoreactivity increased in the treated cultures compared to controls.

    Article Snippet: Primary antisera used in this study were the following: rabbit anti-MT1R (1:1000 in 1% milk/TBST; Abbiotec, 250761), rabbit anti-MT1R (1:1000 in 4% milk/TBST; 1:3000 in Ni-DAB IHC; Novus, NBP1-71113), goat anti-MT2R (1:500 in 4% milk/TBST; Santa Cruz, sc-13177), rabbit anti-MT2R (1:1000 in 4% milk/TBST, 1:10,000 in Ni-DAB IHC, 1:500 in fluorescent IHC (FIHC) using streptavidin and a biotinylated secondary antibody; Novus, NLS932), mouse antityrosine hydroxylase (1:30,000 in FIHC; Chemicon, MAB318), mouse anti-glutamic acid decarboxylase (GAD)-65 (1:10,000 in FIHC; Abcam, ab26113), mouse anti-GAD67 (1:10,000 in FIHC; Chemicon, MAB5406), goat anti-calretinin (1:30,000 in FIHC; Chemicon, AB1550), goat anti-parvalbumin (1:2000 in FIHC; Swant, PVG-214), rabbit monoclonal anti-β-III-tubulin (1:2000 in 4% milk/TBST; Cell Signaling Technology, 5568), and rabbit anti-connexin43 (1:1000 in immunocytochemistry; Millipore AB1728).

    Techniques:

    Immunofluorescent identification of connexin expression in transfected HeLa cells. HeLa cells expressing Cx43 (A) and Cx26 (B) stained with antibodies to Cx43 and Cx26, respectively, show typical punctate staining of Cx43 and Cx26 gap junction plaques at cell–cell contact areas, as well as Cx43 and Cx26 localization in the cell membranes of single cells. Green labeling represents staining of connexin proteins, while blue shows DAPI staining of cell nuclei. The right panels show bright field images. Scale bar, 10 μm.

    Journal: Frontiers in Pharmacology

    Article Title: Cyclic nucleotide permeability through unopposed connexin hemichannels

    doi: 10.3389/fphar.2013.00075

    Figure Lengend Snippet: Immunofluorescent identification of connexin expression in transfected HeLa cells. HeLa cells expressing Cx43 (A) and Cx26 (B) stained with antibodies to Cx43 and Cx26, respectively, show typical punctate staining of Cx43 and Cx26 gap junction plaques at cell–cell contact areas, as well as Cx43 and Cx26 localization in the cell membranes of single cells. Green labeling represents staining of connexin proteins, while blue shows DAPI staining of cell nuclei. The right panels show bright field images. Scale bar, 10 μm.

    Article Snippet: Commercially available anti-connexin43 (Sigma) and anti-connexin26 (Zymed Labs) antibodies were used for immunostaining.

    Techniques: Expressing, Transfection, Staining, Labeling

    MMC induces alteration in Cx43 association with ZO-1. Cells were treated with prewarmed media containing 5 μM MMC for 60 minutes or media alone (CTRL). ( A ) Coimmunoprecipitation with a rabbit polyclonal anti-Cx43 was performed followed by immunoblotting

    Journal: Investigative Ophthalmology & Visual Science

    Article Title: Rapid Changes in Connexin-43 in Response to Genotoxic Stress Stabilize Cell-Cell Communication in Corneal Endothelium

    doi: 10.1167/iovs.11-7272

    Figure Lengend Snippet: MMC induces alteration in Cx43 association with ZO-1. Cells were treated with prewarmed media containing 5 μM MMC for 60 minutes or media alone (CTRL). ( A ) Coimmunoprecipitation with a rabbit polyclonal anti-Cx43 was performed followed by immunoblotting

    Article Snippet: The beads were discarded and then cleared lysates were incubated with 1 μg antibody per 500 μg of lysate using a rabbit polyclonal anti-Cx43 antibody (Invitrogen) overnight at 4°C.

    Techniques:

    Results from in-cell Western analyses showing effects of kinase inhibition on total Cx43 expression in rat primary VSMCs following incubation in the presence of (A) 8Br-cAMP (100 μM), (B) 8Br-cGMP (100 μM), (C) BAY (100 nM), and (D) DADS (50 μM) for 270 min . (A) Inhibition of PKA (by PKI), PKG (by DT2), or PKC (by CalC) each independently subdued 8Br-cAMP-induced increase in Cx43 expression. (B,C) Only PKG and PKC inhibition reversed the effects of 8Br-cGMP and BAY, respectively. (D) Stimulation of Cx43 expression in the presence of DADS was not dependent on PKA, PKG, or PKC. (E) Treatment with any individual kinase inhibitor did not affect total basal Cx43 expression compared with vehicle controls. Data are mean ± SE from n = 4–7 for each treatment. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p

    Journal: Frontiers in Physiology

    Article Title: Control of Vascular Smooth Muscle Cell Growth by Connexin 43

    doi: 10.3389/fphys.2012.00220

    Figure Lengend Snippet: Results from in-cell Western analyses showing effects of kinase inhibition on total Cx43 expression in rat primary VSMCs following incubation in the presence of (A) 8Br-cAMP (100 μM), (B) 8Br-cGMP (100 μM), (C) BAY (100 nM), and (D) DADS (50 μM) for 270 min . (A) Inhibition of PKA (by PKI), PKG (by DT2), or PKC (by CalC) each independently subdued 8Br-cAMP-induced increase in Cx43 expression. (B,C) Only PKG and PKC inhibition reversed the effects of 8Br-cGMP and BAY, respectively. (D) Stimulation of Cx43 expression in the presence of DADS was not dependent on PKA, PKG, or PKC. (E) Treatment with any individual kinase inhibitor did not affect total basal Cx43 expression compared with vehicle controls. Data are mean ± SE from n = 4–7 for each treatment. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p

    Article Snippet: Fixed cells were permeabilized with 0.1% Triton, blocked, and incubated with IR-tagged rabbit antibodies against Cx43 or the phosphorylated forms of Cx43 at Serines 255, 262, 279, and 368 (Cell Signaling) along with mouse α-tubulin (Sigma) as the housekeeping control at 1:500 dilutions overnight at 4°C.

    Techniques: In-Cell ELISA, Inhibition, Expressing, Incubation

    In-cell Western analyses of site-specific Cx43 phosphorylation in rat primary VSMCs in the presence of (A) 8Br-cAMP and (B) DADS . (A) In the presence of 8Br-cAMP, phosphorylation of the MAPK-sensitive Ser279 was significantly increased compared to vehicle controls. (B) The presence of DADS significantly increased phosphorylation at the MAPK sensitive Ser255 and Ser279 sites, p34 cdc2 kinase-sensitive Ser262, and the PKC-sensitive Ser368. Data are mean ± SE from three independent experiments with an n = 4 for Ser279 phosphorylation, and n = 3–4 for Ser255, 262, and 368 phosphorylation. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p

    Journal: Frontiers in Physiology

    Article Title: Control of Vascular Smooth Muscle Cell Growth by Connexin 43

    doi: 10.3389/fphys.2012.00220

    Figure Lengend Snippet: In-cell Western analyses of site-specific Cx43 phosphorylation in rat primary VSMCs in the presence of (A) 8Br-cAMP and (B) DADS . (A) In the presence of 8Br-cAMP, phosphorylation of the MAPK-sensitive Ser279 was significantly increased compared to vehicle controls. (B) The presence of DADS significantly increased phosphorylation at the MAPK sensitive Ser255 and Ser279 sites, p34 cdc2 kinase-sensitive Ser262, and the PKC-sensitive Ser368. Data are mean ± SE from three independent experiments with an n = 4 for Ser279 phosphorylation, and n = 3–4 for Ser255, 262, and 368 phosphorylation. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p

    Article Snippet: Fixed cells were permeabilized with 0.1% Triton, blocked, and incubated with IR-tagged rabbit antibodies against Cx43 or the phosphorylated forms of Cx43 at Serines 255, 262, 279, and 368 (Cell Signaling) along with mouse α-tubulin (Sigma) as the housekeeping control at 1:500 dilutions overnight at 4°C.

    Techniques: In-Cell ELISA

    Effects of cyclic nucleotide or Cx43 stimulation on VSMC DNA synthesis . In rat primary VSMCs in the presence of 10% serum for 6 h, DADS (50 μM) significantly reduced the rate of DNA synthesis by 20% compared with vehicle controls estimated by BrdU incorporation. Data are mean ± SE from three independent experiments each with an n = 6. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p

    Journal: Frontiers in Physiology

    Article Title: Control of Vascular Smooth Muscle Cell Growth by Connexin 43

    doi: 10.3389/fphys.2012.00220

    Figure Lengend Snippet: Effects of cyclic nucleotide or Cx43 stimulation on VSMC DNA synthesis . In rat primary VSMCs in the presence of 10% serum for 6 h, DADS (50 μM) significantly reduced the rate of DNA synthesis by 20% compared with vehicle controls estimated by BrdU incorporation. Data are mean ± SE from three independent experiments each with an n = 6. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p

    Article Snippet: Fixed cells were permeabilized with 0.1% Triton, blocked, and incubated with IR-tagged rabbit antibodies against Cx43 or the phosphorylated forms of Cx43 at Serines 255, 262, 279, and 368 (Cell Signaling) along with mouse α-tubulin (Sigma) as the housekeeping control at 1:500 dilutions overnight at 4°C.

    Techniques: DNA Synthesis, BrdU Incorporation Assay

    Total Cx43 expression in VSMCs . (A) In-cell Western blotting for total Cx43 expression in rat primary VSMC homogenates. Incubation of rat primary VSMCs for 270 min in the presence of 8Br-cAMP (100 μM), 8Br-cGMP (100 μM), BAY (100 nM), or DADS (50 μM) significantly stimulated total Cx43 expression normalized to α-tubulin. Data are from two independent experiments each performed in triplicate. (B) After 24-h incubation the stimulation of Cx43 was significantly increased only after DADS treatment with no observable changes for the 8Br-cAMP, 8Br-cGMP, or BAY treatment groups compared with vehicle controls. For these experiments n = 6 for each treatment. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p

    Journal: Frontiers in Physiology

    Article Title: Control of Vascular Smooth Muscle Cell Growth by Connexin 43

    doi: 10.3389/fphys.2012.00220

    Figure Lengend Snippet: Total Cx43 expression in VSMCs . (A) In-cell Western blotting for total Cx43 expression in rat primary VSMC homogenates. Incubation of rat primary VSMCs for 270 min in the presence of 8Br-cAMP (100 μM), 8Br-cGMP (100 μM), BAY (100 nM), or DADS (50 μM) significantly stimulated total Cx43 expression normalized to α-tubulin. Data are from two independent experiments each performed in triplicate. (B) After 24-h incubation the stimulation of Cx43 was significantly increased only after DADS treatment with no observable changes for the 8Br-cAMP, 8Br-cGMP, or BAY treatment groups compared with vehicle controls. For these experiments n = 6 for each treatment. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p

    Article Snippet: Fixed cells were permeabilized with 0.1% Triton, blocked, and incubated with IR-tagged rabbit antibodies against Cx43 or the phosphorylated forms of Cx43 at Serines 255, 262, 279, and 368 (Cell Signaling) along with mouse α-tubulin (Sigma) as the housekeeping control at 1:500 dilutions overnight at 4°C.

    Techniques: Expressing, In-Cell ELISA, Incubation

    Effects of cyclic nucleotide or Cx43 stimulation on VSMC proliferation . (A) In rat primary VSMCs in the presence of 10% serum for 72 h and using the MTT assay, 8Br-cAMP (100 μM), the sGC stimulator BAY (100 nM) or the garlic-derived Cx inducer DADS (50 μM) significantly reduced rat primary VSMC numbers compared with vehicle controls. Data are mean ± SEM with an n = 6. (B) Using hemocytometry after 72 h, both 8Br-cAMP and BAY significantly reduced cell numbers by ∼40%, and the presence of DADS reduced cell numbers by ∼20% ( p = 0.06). Data are mean ± SEM of two independent experiments each with n = 3–4. (C) MTT assay performed on VSMCs after 12 h shows no observable changes in any treatment group. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p

    Journal: Frontiers in Physiology

    Article Title: Control of Vascular Smooth Muscle Cell Growth by Connexin 43

    doi: 10.3389/fphys.2012.00220

    Figure Lengend Snippet: Effects of cyclic nucleotide or Cx43 stimulation on VSMC proliferation . (A) In rat primary VSMCs in the presence of 10% serum for 72 h and using the MTT assay, 8Br-cAMP (100 μM), the sGC stimulator BAY (100 nM) or the garlic-derived Cx inducer DADS (50 μM) significantly reduced rat primary VSMC numbers compared with vehicle controls. Data are mean ± SEM with an n = 6. (B) Using hemocytometry after 72 h, both 8Br-cAMP and BAY significantly reduced cell numbers by ∼40%, and the presence of DADS reduced cell numbers by ∼20% ( p = 0.06). Data are mean ± SEM of two independent experiments each with n = 3–4. (C) MTT assay performed on VSMCs after 12 h shows no observable changes in any treatment group. Student–Newman–Keuls method for multiple comparisons following one-way ANOVA was used. * p

    Article Snippet: Fixed cells were permeabilized with 0.1% Triton, blocked, and incubated with IR-tagged rabbit antibodies against Cx43 or the phosphorylated forms of Cx43 at Serines 255, 262, 279, and 368 (Cell Signaling) along with mouse α-tubulin (Sigma) as the housekeeping control at 1:500 dilutions overnight at 4°C.

    Techniques: MTT Assay, Derivative Assay

    Schematic showing potential roles of astrocytic Cx43, hemichannels, and GJIC during OGD/R injury. Under normal conditions, astrocytic Cx43 is expressed in the plasma membrane and assembled into hemichannels that are normally closed. Hemichannel-hemichannel interactions induce the formation of GJIC between adjacent astrocytes, which permits the exchange of ions and small molecules; also, plasma membrane’s Cx43 was phosphorylated at Ser368 site. In such circumstances, astrocytes, together with those resting microglia, function as a supportive assistant for healthy neurons. OGD/R injury caused abnormal hemichannel opening and consequent substantial astrocytic ATP release. It also induced microglial activation with a predominance of the pro-inflammatory cytokine-releasing M1 subtype. Extracellular ATP induced further microglial activation and pro-inflammatory cytokine release, and these pro-inflammatory cytokines induced further opening of astrocytic hemichannels. SalB reversed these effects and thus provided protection against OGD/R injury. This suggests the existence of a vicious cycle in which astrocytic hemichannel opening and pro-inflammatory microglial activation reinforce each other following OGD/R injury. This vicious cycle may account for secondary injury and extended damage after OGD/R injury; OGD/R injury caused gap junction internalization, which may account for the astrocytic uncoupling events. It also decreased plasma membrane levels of Ser368-phosphorylated Cx43 while increasing plasma membrane levels of Ser373-phosphorylated Cx43, Ser265-phosphorylated Cx43, and Src’s Tyr416-phosphorylated activated form. The activated Src may well have phosphorylated Cx43 at Tyr265 and further induced gap junction internalization or autophagy. SalB directly inhibits Src, which may allow it to exert protective effects by attenuating Cx43 internalization. CBX, a non-selective hemichannel and GJIC inhibitor, did not apparently affect Cx43 phosphorylation, but it inhibited PKC and Src activity

    Journal: Journal of Neuroinflammation

    Article Title: Roles of astrocytic connexin-43, hemichannels, and gap junctions in oxygen-glucose deprivation/reperfusion injury induced neuroinflammation and the possible regulatory mechanisms of salvianolic acid B and carbenoxolone

    doi: 10.1186/s12974-018-1127-3

    Figure Lengend Snippet: Schematic showing potential roles of astrocytic Cx43, hemichannels, and GJIC during OGD/R injury. Under normal conditions, astrocytic Cx43 is expressed in the plasma membrane and assembled into hemichannels that are normally closed. Hemichannel-hemichannel interactions induce the formation of GJIC between adjacent astrocytes, which permits the exchange of ions and small molecules; also, plasma membrane’s Cx43 was phosphorylated at Ser368 site. In such circumstances, astrocytes, together with those resting microglia, function as a supportive assistant for healthy neurons. OGD/R injury caused abnormal hemichannel opening and consequent substantial astrocytic ATP release. It also induced microglial activation with a predominance of the pro-inflammatory cytokine-releasing M1 subtype. Extracellular ATP induced further microglial activation and pro-inflammatory cytokine release, and these pro-inflammatory cytokines induced further opening of astrocytic hemichannels. SalB reversed these effects and thus provided protection against OGD/R injury. This suggests the existence of a vicious cycle in which astrocytic hemichannel opening and pro-inflammatory microglial activation reinforce each other following OGD/R injury. This vicious cycle may account for secondary injury and extended damage after OGD/R injury; OGD/R injury caused gap junction internalization, which may account for the astrocytic uncoupling events. It also decreased plasma membrane levels of Ser368-phosphorylated Cx43 while increasing plasma membrane levels of Ser373-phosphorylated Cx43, Ser265-phosphorylated Cx43, and Src’s Tyr416-phosphorylated activated form. The activated Src may well have phosphorylated Cx43 at Tyr265 and further induced gap junction internalization or autophagy. SalB directly inhibits Src, which may allow it to exert protective effects by attenuating Cx43 internalization. CBX, a non-selective hemichannel and GJIC inhibitor, did not apparently affect Cx43 phosphorylation, but it inhibited PKC and Src activity

    Article Snippet: Western blot analysis Protein concentration was evaluated using the BCA method, and 20–40 μg of total extract was separated on SDS polyacrilamide gels, and then the gel-separated proteins were transferred onto polyvinylidene fluoride (PVDF) membranes and probed using the indicated primary antibodies: overnight at 4 °C with Rabbit anti-pan Cx43(1:1000, Cell signaling technology, Cat#3512); Rabbit anti-phospho-Cx43/ GJA1(Ser 368)(1:1000, Abcam,Cat#ab30559); Rabbit anti-phospho-Cx43 (Ser373) (1:1000, Thermo Fisher Scientific, Cat#PA5-64670); Rabbit anti-phospho-Cx43(Ser265)(1:500, Thermo Fisher Scientific, Cat#PA5-37584); Rabbit anti-PKCε (1:200, Santa Cruz Biotechnology, Cat#sc-214); Goat anti-phospho-PKCε (Ser729) (1: 200, Santa Cruz Biotechnology, Cat#sc-12355); Rabbit anti-Akt(PKB) (1:1000, Cell Signaling Technology, Cat#4691); Rabbit anti-phospho-Akt(Thr308) antibody (1:1000, Cell Signaling Technology, Cat#13038); Rabbit anti-Src (1:1000, Cell Signaling Technology, Cat#2108); Rabbit anti-Phospho-Src(Tyr527) (1:1000, Cell Signaling Technology, Cat#2105); Rabbit anti-Phospho-Src(Tyr416) (1:1000, Cell Signaling Technology, Cat#6943); Rabbit anti-GAPDH antibody (1/5000, Abcam, Cat#ab37168); mouse anti-β-actin(1:1000, Abcam, Cat#ab6276); Rabbit anti-Arginase-1 (1:1000, Cell Signaling Technology, Cat#93668).

    Techniques: Activation Assay, Activity Assay

    OGD/R injury with SalB or CBX application influenced astrocytic phosphorylated Cx43 and corresponding kinases. a1 , a2 OGD/R injury had no significant effect on cytoplasmic PKCε levels but significantly increased plasma membrane levels of PKCε and its Ser729-phosphorylated activated state. SalB and CBX reduced PKCε levels in the plasma membrane and increased them in the cytoplasm. Conversely, the OGD/R group’s Ser368-phosphorylated Cx43 levels were decreased in the plasma membrane and increased in the cytoplasm. SalB reversed these effects, but CBX reduced Ser368-phosphorylated Cx43 levels. b1 , b2 The OGD/R group exhibited decreased cytoplasmic levels of Thr308-phosphorylated PKB, though SalB reversed this effect. There were no significant changes in the plasma membrane levels. Furthermore, the OGD/R group exhibited elevated cytoplasmic and plasma membrane levels of Ser373-phosphorylated Cx43. SalB reduced the plasma membrane levels but further increased the cytoplasmic levels. CBX had little effect on the levels of Ser373-phosphorylated Cx43 and Thr308-phosphorylated PKB. c1 , c2 The OGD/R group exhibited elevated cytoplasmic and plasma membrane levels of Tyr416-phosphorylated Src. The OGD/R group also exhibited elevated plasma membrane levels of Src, which may be related to the elevated Tyr416-phosphorylated Src levels. SalB increased the plasma membrane levels of Src’s Tyr527-phosphorylated deactivated form but did not significantly affect plasma membrane levels of Tyr416-phosphorylated Src. CBX significantly reduced cytoplasmic and plasma membrane levels of Tyr416-phosphorylated Src. Furthermore, the OGD/R group exhibited increased cytoplasmic and plasma membrane levels of Tyr265-phosphorylated Cx43. SalB reversed this effect, but CBX achieved only non-significant reduction of the plasma membrane levels. We evaluated the statistical significance with ANOVA and Duncan’s multiple comparisons test. * p

    Journal: Journal of Neuroinflammation

    Article Title: Roles of astrocytic connexin-43, hemichannels, and gap junctions in oxygen-glucose deprivation/reperfusion injury induced neuroinflammation and the possible regulatory mechanisms of salvianolic acid B and carbenoxolone

    doi: 10.1186/s12974-018-1127-3

    Figure Lengend Snippet: OGD/R injury with SalB or CBX application influenced astrocytic phosphorylated Cx43 and corresponding kinases. a1 , a2 OGD/R injury had no significant effect on cytoplasmic PKCε levels but significantly increased plasma membrane levels of PKCε and its Ser729-phosphorylated activated state. SalB and CBX reduced PKCε levels in the plasma membrane and increased them in the cytoplasm. Conversely, the OGD/R group’s Ser368-phosphorylated Cx43 levels were decreased in the plasma membrane and increased in the cytoplasm. SalB reversed these effects, but CBX reduced Ser368-phosphorylated Cx43 levels. b1 , b2 The OGD/R group exhibited decreased cytoplasmic levels of Thr308-phosphorylated PKB, though SalB reversed this effect. There were no significant changes in the plasma membrane levels. Furthermore, the OGD/R group exhibited elevated cytoplasmic and plasma membrane levels of Ser373-phosphorylated Cx43. SalB reduced the plasma membrane levels but further increased the cytoplasmic levels. CBX had little effect on the levels of Ser373-phosphorylated Cx43 and Thr308-phosphorylated PKB. c1 , c2 The OGD/R group exhibited elevated cytoplasmic and plasma membrane levels of Tyr416-phosphorylated Src. The OGD/R group also exhibited elevated plasma membrane levels of Src, which may be related to the elevated Tyr416-phosphorylated Src levels. SalB increased the plasma membrane levels of Src’s Tyr527-phosphorylated deactivated form but did not significantly affect plasma membrane levels of Tyr416-phosphorylated Src. CBX significantly reduced cytoplasmic and plasma membrane levels of Tyr416-phosphorylated Src. Furthermore, the OGD/R group exhibited increased cytoplasmic and plasma membrane levels of Tyr265-phosphorylated Cx43. SalB reversed this effect, but CBX achieved only non-significant reduction of the plasma membrane levels. We evaluated the statistical significance with ANOVA and Duncan’s multiple comparisons test. * p

    Article Snippet: Western blot analysis Protein concentration was evaluated using the BCA method, and 20–40 μg of total extract was separated on SDS polyacrilamide gels, and then the gel-separated proteins were transferred onto polyvinylidene fluoride (PVDF) membranes and probed using the indicated primary antibodies: overnight at 4 °C with Rabbit anti-pan Cx43(1:1000, Cell signaling technology, Cat#3512); Rabbit anti-phospho-Cx43/ GJA1(Ser 368)(1:1000, Abcam,Cat#ab30559); Rabbit anti-phospho-Cx43 (Ser373) (1:1000, Thermo Fisher Scientific, Cat#PA5-64670); Rabbit anti-phospho-Cx43(Ser265)(1:500, Thermo Fisher Scientific, Cat#PA5-37584); Rabbit anti-PKCε (1:200, Santa Cruz Biotechnology, Cat#sc-214); Goat anti-phospho-PKCε (Ser729) (1: 200, Santa Cruz Biotechnology, Cat#sc-12355); Rabbit anti-Akt(PKB) (1:1000, Cell Signaling Technology, Cat#4691); Rabbit anti-phospho-Akt(Thr308) antibody (1:1000, Cell Signaling Technology, Cat#13038); Rabbit anti-Src (1:1000, Cell Signaling Technology, Cat#2108); Rabbit anti-Phospho-Src(Tyr527) (1:1000, Cell Signaling Technology, Cat#2105); Rabbit anti-Phospho-Src(Tyr416) (1:1000, Cell Signaling Technology, Cat#6943); Rabbit anti-GAPDH antibody (1/5000, Abcam, Cat#ab37168); mouse anti-β-actin(1:1000, Abcam, Cat#ab6276); Rabbit anti-Arginase-1 (1:1000, Cell Signaling Technology, Cat#93668).

    Techniques: