Journal: International Journal of Molecular Sciences

Article Title: Cx43 Promotes Endothelial Cell Migration and Angiogenesis via the Tyrosine Phosphatase SHP-2

doi: 10.3390/ijms23010294

Figure Lengend Snippet: Downregulation of Cx43 reduces endothelial cell migration. ( a ) Western blot analysis of Cx43, Cx40, and Cx37 revealed that Cx43 is expressed in both human umbilical vein endothelial cells (HUVEC) and human microvascular endothelial cells (HMEC), whereas Cx40 and Cx37 are not expressed in HMEC. Cx43 (Cx43), Cx40 (Cx), and Cx37 (Cx) expressing HeLa cells were used as the positive controls. CTL: Wildtype HeLa cell lysate. ( b ) Western blot analysis of Cx43 in HMEC transfected with 100 nM siRNA against Cx43 or non-silencing control siRNA (Ctrl) for 48 h. The Cx43 expression was analysed using a polyclonal Cx43 antibody. Immunostaining for GAPDH was used as the loading control. ( c ) Serum-induced cell migration of HMEC transfected with a Cx43-siRNA or a control siRNA (Ctrl) in response to 10% FCS was assessed in a wound assay. Downregulation of Cx43 significantly decreased cell migration, represented as accumulated distance (*** p < 0.001 Ctrl-siRNA vs. Cx43-siRNA, n = 6, 3 independent cell cultures). ( d ) Representative single cell traces of migrated HMEC.

Article Snippet: For immunoprecipitation, the supernatants were incubated over night at 4 °C with a mouse anti-SHP-2 antibody (Santa Cruz) or a rabbit anti-Cx43 antibody (Sigma Aldrich), and were magnetic beads coated with protein A (Cx43) or G (SHP-2) (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Migration, Western Blot, Expressing, Transfection, Control, Immunostaining

Journal: International Journal of Molecular Sciences

Article Title: Cx43 Promotes Endothelial Cell Migration and Angiogenesis via the Tyrosine Phosphatase SHP-2

doi: 10.3390/ijms23010294

Figure Lengend Snippet: Downregulation of Cx43 impairs vessel sprouting ex vivo. ( a ) Downregulation of Cx43 in the mouse aortae was achieved by transfection with Cx43 siRNA (500 nM) or control (Ctrl) siRNA, as assessed by Western blot 48 h later. ( b ) Representative images of aortic sprouts in matrigel. Scale bar represents 200 µm. ( c ) The vessel sprouting area (** p < 0.01 Ctrl-siRNA vs. Cx43-siRNA, n = 2 aortae with eight aortic segments each at 3 days; n = 3–4 aortae with 6–8 aortic segments each at 6 days) and ( d ) the number of sprout bifurcations (** p < 0.01 Ctrl-siRNA vs. Cx43-siRNA, n = 2 aortae with 8 aortic segments each at 3 days; *** p < 0.001 Ctrl-siRNA vs. Cx43-siRNA n = 3–4 aortae with 6–8 aortic segments each at 6 days) were significantly reduced upon downregulation of Cx43, as assessed 3 days and 6 days after transfection in a matrigel assay.

Article Snippet: For immunoprecipitation, the supernatants were incubated over night at 4 °C with a mouse anti-SHP-2 antibody (Santa Cruz) or a rabbit anti-Cx43 antibody (Sigma Aldrich), and were magnetic beads coated with protein A (Cx43) or G (SHP-2) (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Ex Vivo, Transfection, Control, Western Blot, Matrigel Assay

Journal: International Journal of Molecular Sciences

Article Title: Cx43 Promotes Endothelial Cell Migration and Angiogenesis via the Tyrosine Phosphatase SHP-2

doi: 10.3390/ijms23010294

Figure Lengend Snippet: Interaction of Cx43 and SHP-2 induces SHP-2 phosphatase activity. ( a ) Upon immunoprecipitation (IP) of SHP-2 from HeLa cells expressing only Cx43, Cx43 was co-immunoprecipitated. As a control, cells transfected with an empty vector (control (CTL)) were used. L: Whole cell lysates. ( b ) SHP-2 was detected upon immunoblotting after immunoprecipitation of Cx43 from HeLa cells expressing only Cx43 in contrast to the control cells (CTL). L: Whole cell lysates. Images originates from the same blot, which was cropped; ( c ) SHP-2 was immunoprecipitated together with Cx43 in HMEC; ( d ) Co-localization of SHP-2 and Cx43 was detected by immunofluorescent staining of HMEC followed by confocal microscopic imaging. Scale bar represents 20 µm; ( e ) SHP-2 phosphatase activity was significantly increased in HeLa cells expressing Cx43 compared to HeLa cells transfected with an empty vector (* p < 0.05, n = 4 independent cell cultures), as assessed by detection of dephosphorylation of a phosphate analogue in SHP-2 immunoprecipitates.

Article Snippet: For immunoprecipitation, the supernatants were incubated over night at 4 °C with a mouse anti-SHP-2 antibody (Santa Cruz) or a rabbit anti-Cx43 antibody (Sigma Aldrich), and were magnetic beads coated with protein A (Cx43) or G (SHP-2) (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Activity Assay, Immunoprecipitation, Expressing, Control, Transfection, Plasmid Preparation, Western Blot, Staining, Imaging, De-Phosphorylation Assay

Journal: International Journal of Molecular Sciences

Article Title: Cx43 Promotes Endothelial Cell Migration and Angiogenesis via the Tyrosine Phosphatase SHP-2

doi: 10.3390/ijms23010294

Figure Lengend Snippet: Inactivation of SHP-2 reduces endothelial cell migration. ( a ) Overexpression of a dominant negative substrate trapping mutant SHP-2 (SHP-2 CS) in HMEC significantly reduced serum (10%) induced endothelial cell migration, as assessed by the accumulated distance in µm in an in vitro wound assay (* p < 0.05, n = 3 independent cell cultures) compared to SHP-2 WT. Additional knock-down of Cx43 (Cx43 siRNA) did not further decrease migration (ns: not significant, n = 3 independent cell cultures). ( b ) Representative single cell traces of migrated HMEC.

Article Snippet: For immunoprecipitation, the supernatants were incubated over night at 4 °C with a mouse anti-SHP-2 antibody (Santa Cruz) or a rabbit anti-Cx43 antibody (Sigma Aldrich), and were magnetic beads coated with protein A (Cx43) or G (SHP-2) (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Migration, Over Expression, Dominant Negative Mutation, Mutagenesis, In Vitro, Knockdown

Journal: International Journal of Molecular Sciences

Article Title: Cx43 Promotes Endothelial Cell Migration and Angiogenesis via the Tyrosine Phosphatase SHP-2

doi: 10.3390/ijms23010294

Figure Lengend Snippet: The Cx43 and SHP-2 interaction is vital for endothelial migration. ( a ) Treatment of HMEC overexpressing a constitutively active mutant SHP-2 (SHP-2 EA) with Cx43 siRNA did not rescue the reduced migratory response caused by Cx43 knock-down (Cx43 siRNA) (ns: not significant, n = 3 independent cell cultures) compared to SHP-2 EA cells treated with control siRNA (Ctrl) (* p < 0.05, n = 3 independent cell cultures), as assessed by the accumulated distance in µm in an in vitro wound assay (* p < 0.05, n = 3 independent cell cultures). ( b ) Representative single cell traces of migrated HMEC.

Article Snippet: For immunoprecipitation, the supernatants were incubated over night at 4 °C with a mouse anti-SHP-2 antibody (Santa Cruz) or a rabbit anti-Cx43 antibody (Sigma Aldrich), and were magnetic beads coated with protein A (Cx43) or G (SHP-2) (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Migration, Mutagenesis, Knockdown, Control, In Vitro

Journal: Heliyon

Article Title: Differentiation of cardiomyocyte-like cells from human amniotic fluid mesenchymal stem cells by combined induction with human platelet lysate and 5-azacytidine

doi: 10.1016/j.heliyon.2020.e04844

Figure Lengend Snippet: The primers for RT–qPCR and products size.

Article Snippet: After fixation, the cells were blocked in 10% AB-serum in 1% BSA-PBS for 30 min at room temperature, and then incubated with mouse anti-human Cx43 primary antibodies (Sigma-aldrich, USA) for 2 h at 37 °C.

Techniques:

Journal: Heliyon

Article Title: Differentiation of cardiomyocyte-like cells from human amniotic fluid mesenchymal stem cells by combined induction with human platelet lysate and 5-azacytidine

doi: 10.1016/j.heliyon.2020.e04844

Figure Lengend Snippet: A: hPL ability to up-regulate cardiomyogenic specific genes expression. Each graph displays the expression levels of GATA4 , cTnT , Cx4 3 and Nkx2.5 , which were normalized to GAPDH and were relative to the control group. Data are presented as mean ± S.E. values. ∗ statistically significant versus control. # statistically significant between group. B: hPL combined with 5-aza ability to up-regulate cardiomyogenic specific genes expression. Each graph displays the expression levels of GATA4 , cTnT , Cx43 and Nkx2.5 , which were normalized to GAPDH and were relative to the control group. Data are presented as mean ± S.E. values. ∗ statistically significant versus control. # statistically significant between group.

Article Snippet: After fixation, the cells were blocked in 10% AB-serum in 1% BSA-PBS for 30 min at room temperature, and then incubated with mouse anti-human Cx43 primary antibodies (Sigma-aldrich, USA) for 2 h at 37 °C.

Techniques: Expressing

Journal: Heliyon

Article Title: Differentiation of cardiomyocyte-like cells from human amniotic fluid mesenchymal stem cells by combined induction with human platelet lysate and 5-azacytidine

doi: 10.1016/j.heliyon.2020.e04844

Figure Lengend Snippet: Ability of hPL to promote the differentiation of hAF-MSCs into cardiomyocyte-like cells. Each graph displays the expression levels of GATA4 , cTnT , Cx43 and Nkx2. 5 that were normalized to GAPDH and were relative to the control group. Data are presented as mean ± S.E. values. ∗ statistically significant versus control. # statistically significant between group.

Article Snippet: After fixation, the cells were blocked in 10% AB-serum in 1% BSA-PBS for 30 min at room temperature, and then incubated with mouse anti-human Cx43 primary antibodies (Sigma-aldrich, USA) for 2 h at 37 °C.

Techniques: Expressing

Journal: Heliyon

Article Title: Differentiation of cardiomyocyte-like cells from human amniotic fluid mesenchymal stem cells by combined induction with human platelet lysate and 5-azacytidine

doi: 10.1016/j.heliyon.2020.e04844

Figure Lengend Snippet: Detection of cardiomyogenic specific proteins; immunofluorescence staining FITC (495 nm/519 nm), green color for all cardiomyogenic specific proteins (A–I) and immunoenzymatic staining (J–L); GATA4 (localized in nucleus) staining (A) control group, (B) 10 μM 5-aza induced group, (C) 10 μM 5-aza with 20% hPL induced group; cTnT (localized in cytoplasm) staining (D) control group, (E) 10 μM 5-aza induced group, (F) 10 μM 5-aza 20% hPL induced group; Nkx2.5 (localized in nucleus) staining (G) control group, (H) 10 μM 5-aza induced group, (I) 10 μM 5-aza with 20% hPL induced group; Cx43 (localized in cell membrane) staining (J) control group, (K) 10 μM 5-aza induced group (black arrow), (L) 10 μM 5-aza with 20% hPL induced group (black arrow). (A-C and G-I) insets without nuclear counterstain showing no nuclear staining of two key core cardiac transcription factors. Scale bar = 100 μm.

Article Snippet: After fixation, the cells were blocked in 10% AB-serum in 1% BSA-PBS for 30 min at room temperature, and then incubated with mouse anti-human Cx43 primary antibodies (Sigma-aldrich, USA) for 2 h at 37 °C.

Techniques: Immunofluorescence, Staining

Journal: Heliyon

Article Title: Differentiation of cardiomyocyte-like cells from human amniotic fluid mesenchymal stem cells by combined induction with human platelet lysate and 5-azacytidine

doi: 10.1016/j.heliyon.2020.e04844

Figure Lengend Snippet: Image J analysis showing the results of CTCF (the expression levels of GATA4, cTnT, Nkx2.5 and Cx43 proteins signal). Data are presented as mean ± S.E. values. ∗ statistically significant versus control.

Article Snippet: After fixation, the cells were blocked in 10% AB-serum in 1% BSA-PBS for 30 min at room temperature, and then incubated with mouse anti-human Cx43 primary antibodies (Sigma-aldrich, USA) for 2 h at 37 °C.

Techniques: Expressing

Journal: bioRxiv

Article Title: Targeting the Cx43 Carboxyl Terminal H2 Domain Preserves Left Ventricular Function Following Ischemia-Reperfusion Injury

doi: 10.1101/668509

Figure Lengend Snippet: A) Schematics of full length Cx43 and αCT1 peptide. B) αCT1 interaction with ZO-1 PDZ domains as indicated by EDC zero-length cross-linking to GST fusion PDZ1, PDZ2 and PDZ3 polypeptides and neutravidin labeling of biotin-tagged peptide at concentrations of 5, 25 and 50 μM. The deletion of the CT Isoleucine (I) in αCT1-I renders this peptide incompetent to interact with the ZO-1 PDZ2 domain. C) Coomassie blue gel of EDC cross-linked products of kinase reaction mixtures containing GST-Cx43 CT and PKC-ε, with (αCT1) and without (Vehicle) αCT1. The fainter band above GST-Cx43 bands (indicated by lines) in the αCT1 lanes were cut from gels and analyzed by Tandem Mass Spectrometry (MS/MS). The boxes to right of gel show Cx43 CT peptides identified by MS/MS as being cross-linked to αCT1. D) Tandem mass spectrum of a quintuply charged crosslinked peptide (m/z: 674.1) between Cx43 345-366 (a-chain) and αCT1 peptide through Cx43 K346 and E8 in αCT1 (b-chain). Only the b-and y-sequence specific ions are labeled. Arrow indicates ion (b a5 2+ ) consistent with cross-linkage between Cx43 CT lysine K346 and the glutamic acid (E) residue of αCT1 at position −1.

Article Snippet: Phospho-Connexin43 (Ser368) (Cell Signaling, 3511S, Danvers, MA), anti-Cx43 produced in rabbit (Sigma: C6219, St. Louis, MO), anti-GST produced in goat (GE, 27457701, Little Chalfont, UK).

Techniques: Labeling, Mass Spectrometry, Tandem Mass Spectroscopy, Sequencing

Journal: bioRxiv

Article Title: Targeting the Cx43 Carboxyl Terminal H2 Domain Preserves Left Ventricular Function Following Ischemia-Reperfusion Injury

doi: 10.1101/668509

Figure Lengend Snippet: Blots of EDC cross-linked products of kinase reaction mixtures containing GST-Cx43 CT, GST-Cx43 CT QQ/KK in which the lysine (K) residues were mutated to neutral glutamines (Q), PKC-ε and αCT1 (at 5, 10 and 25 μM) and a scrambled αCT1 (M4 scr) variant at the same concentrations. Only αCT1 is seen to be covalently linked by EDC to Cx43 CT in a concentration-dependent manner.

Article Snippet: Phospho-Connexin43 (Ser368) (Cell Signaling, 3511S, Danvers, MA), anti-Cx43 produced in rabbit (Sigma: C6219, St. Louis, MO), anti-GST produced in goat (GE, 27457701, Little Chalfont, UK).

Techniques: Variant Assay, Concentration Assay

Journal: bioRxiv

Article Title: Targeting the Cx43 Carboxyl Terminal H2 Domain Preserves Left Ventricular Function Following Ischemia-Reperfusion Injury

doi: 10.1101/668509

Figure Lengend Snippet: A) Schematics of Cx43 and the secondary structure of Cx43 CT from amino acid residues Glycine252 (G252) through to Isoleucine 382 (I382). The depiction of secondary structure in 2A has been modified from a diagram originally provided by Sosinsky and co-workers . B) ZDOCK and C) Schrodinger molecular modeling software analysis of the structure of a proposed αCT1-Cx43 CT complex. The protonated structure of αCT1 peptide and Cx43 CT (PDB:1r5s), constrained by a salt-bridge interaction between K346 in the Cx43 CT and the glutamic acid (E) at position −1 of αCT1. The αCT1-Cx43 interaction shown represents that based on the lowest energy minimization score determined in the model. D) Schrodinger molecular modeling software, a 2D map of αCT1-Cx43 CT in anti-parallel orientation showing location of amino acids predicted to bond to each other and the type of bond that is predicted to occur.

Article Snippet: Phospho-Connexin43 (Ser368) (Cell Signaling, 3511S, Danvers, MA), anti-Cx43 produced in rabbit (Sigma: C6219, St. Louis, MO), anti-GST produced in goat (GE, 27457701, Little Chalfont, UK).

Techniques: Modification, Software

Journal: bioRxiv

Article Title: Targeting the Cx43 Carboxyl Terminal H2 Domain Preserves Left Ventricular Function Following Ischemia-Reperfusion Injury

doi: 10.1101/668509

Figure Lengend Snippet: SPR was used to analyze interactions of biotin-αCT1 and biotin-αCT1 variant peptides, immobilized to streptavidin-coated chips, with the Cx43 CT (Cx43-CT: amino acids 255 to 382) and Cx43 CT-KK/QQ as analytes, respectively. The mean of three runs is plotted for each analyte concentration. The exposure of the sensor chip to the specific analyte is indicated by the gray area. Sensorgrams obtained for: A) Cx43 CT and biotin-αCT1. B) Cx43 CT-KK/QQ and biotin-αCT1. C) Cx43 CT and biotin-M1 AALAI. D) Cx43 CT-KK/QQ and biotin-M1 AALAI. E) Cx43 CT and biotin-M3 DDLAI. F) Cx43 CT-KK/QQ and biotin-M3 DDLAI.

Article Snippet: Phospho-Connexin43 (Ser368) (Cell Signaling, 3511S, Danvers, MA), anti-Cx43 produced in rabbit (Sigma: C6219, St. Louis, MO), anti-GST produced in goat (GE, 27457701, Little Chalfont, UK).

Techniques: Variant Assay, Concentration Assay

Journal: bioRxiv

Article Title: Targeting the Cx43 Carboxyl Terminal H2 Domain Preserves Left Ventricular Function Following Ischemia-Reperfusion Injury

doi: 10.1101/668509

Figure Lengend Snippet: A) Melt curves (top) and first derivative of melt curves (bottom) for ZO-1 PDZ2 at 500 μg/mL in combination αCT1 at concentrations of 25, 50 and 100 μM. B) Temperature maxima (Tm) from Boltzman curves from left-to-right of Cx43 CT (Cx43-CT: amino acids 255 to 382) alone, Cx43 CT in combination with αCT1, and the αCT1 variants including: M1 AALAI, M2 AALEI, M3 DDLAI, M4 scrambled, αCT-I and αCT11. αCT1, αCT1-I and αCT11 show similar abilities to destabilize (i.e., significantly decrease the Tm of) Cx43 CT. **p<0.01, *** p<0.002, N=6. C) Temperature maxima (Tm) from Boltzman curves from left-to-right of PDZ2 alone, and PDZ2 in combination with αCT1 and αCT1variants including αCT1 variants including: M1 AALAI, M2 AALEI, M3 DDLAI, M4 scrambled, αCT-I and αCT11. M3 DDLAI, αCT1, and αCT11 show similar abilities to stabilize (i.e., significantly increase the Tm of) PDZ2. **p<0.01, ***p<0.002, N=6

Article Snippet: Phospho-Connexin43 (Ser368) (Cell Signaling, 3511S, Danvers, MA), anti-Cx43 produced in rabbit (Sigma: C6219, St. Louis, MO), anti-GST produced in goat (GE, 27457701, Little Chalfont, UK).

Techniques:

Journal: bioRxiv

Article Title: Targeting the Cx43 Carboxyl Terminal H2 Domain Preserves Left Ventricular Function Following Ischemia-Reperfusion Injury

doi: 10.1101/668509

Figure Lengend Snippet: A) Blots of Cx43-pS368 (top) and total Cx43 (bottom) in kinase reactions mixtures including no-kinase controls with substrate (Cx43-CT: amino acids 255 to 382), but no PKC-ε (PKC-minus); Cx43-CT substrate with PKC-ε (PKC-plus); and mixtures containing PKC-ε, Cx43 CT, and biotin-tagged αCT1, biotin-tagged αCT1 mutant peptides with alanine substitutions (M1 AALAI, M2 AALEI, M3 DDLAI) and biotin-tagged M4 scrambled. Peptides are at 20 μM. B) Blots of Cx43-pS368 (top) and total Cx43 (bottom) in kinase reactions mixtures including no-kinase controls with Cx43 CT substrate, but no PKC-ε (PKC-minus); Cx43-CT substrate with PKC-ε (PKC-plus); and mixtures containing PKC-ε, Cx43 CT, and biotin-αCT1, biotin-αCT1-I or biotin-αCT11 (RPRPDDLEI with no antennapedia sequence at peptide NT) and biotin-M4 scrambled peptide. Peptides are at 20 μM. C) Chart showing that the ability of unmodified αCT1 and the Cx43 CT interaction-competent peptides biotin-αCT1-I or biotin-αCT11 to induce S368 phosphorylation was 3-5 fold greater than that of non-Cx43 CT interacting peptides. * p<0.05, ** p<0.01, *** p<0.002, N=5 αCT1 and M4, other peptides N=3.

Article Snippet: Phospho-Connexin43 (Ser368) (Cell Signaling, 3511S, Danvers, MA), anti-Cx43 produced in rabbit (Sigma: C6219, St. Louis, MO), anti-GST produced in goat (GE, 27457701, Little Chalfont, UK).

Techniques: Mutagenesis, Sequencing

Journal: bioRxiv

Article Title: Targeting the Cx43 Carboxyl Terminal H2 Domain Preserves Left Ventricular Function Following Ischemia-Reperfusion Injury

doi: 10.1101/668509

Figure Lengend Snippet: Langendorff ischemia-reperfusion (I/R) injury protocols were performed on adult mouse hearts instrumented to monitor LV contractility (protocol in ). LV Systolic responses are shown in 7A-C : (A) Plots of left ventricular (LV) systolic developed pressure against balloon volume ; (B) LV maximal rate of tension development (+dP/dt) against balloon volume; (C) Maximal systolic elastance (E max ) – i.e., the slope from (A) ; (D) Plots of LV end diastolic pressure (EDP) against balloon volume; (E) Maximal rate of relaxation (-dP/dt) against balloon volume; (F) Stiffness, the reciprocal of the slope from (D) ; ( G) Percentage of LV contractile function recovery post-ischemia relative to baseline level. Data shown are mean ± S.E. N=4-8. *p<0.05, ***p<0.001, N=4-8 hearts/group. H) Blots of Cx43-pS368 (top) and total Cx43 (bottom) of LV samples infused with peptide for 20 minutes according to the protocol in . For hearts used in Western blots, the protocol did not proceed to the ischemia and reperfusion phases, being terminated after the peptide infusion step. Only those peptides competent to interact with Cx43 CT increase pS368 levels relative to total Cx43 above vehicle control.

Article Snippet: Phospho-Connexin43 (Ser368) (Cell Signaling, 3511S, Danvers, MA), anti-Cx43 produced in rabbit (Sigma: C6219, St. Louis, MO), anti-GST produced in goat (GE, 27457701, Little Chalfont, UK).

Techniques: Western Blot

Journal: bioRxiv

Article Title: Targeting the Cx43 Carboxyl Terminal H2 Domain Preserves Left Ventricular Function Following Ischemia-Reperfusion Injury

doi: 10.1101/668509

Figure Lengend Snippet: Langendorff I/R protocols were performed on adult mouse hearts instrumented to monitor LV contractility. Protocol in , except that a 20-minute peptide infusion was begun after ischemic injury at the initiation of reperfusion. (A) Plots of left ventricular (LV) developed pressure against balloon volume; (B) Maximal systolic elastance (E max ), the slope from (A); (C) Maximal rate of tension development (+dP/dt) against balloon volume; (D) Plots of end diastolic pressure (EDP) against balloon volume; (E) Stiffness, the reciprocal of the slope from (D); (F) Maximal rate of relaxation (-dP/dt) against balloon volume. * p<0.05, *** p<0.001, N=4-8. G) Laser scanning confocal microscopic fields from sections of Vehicle control, αCT1, and αCT11 group hearts stained for Cx43 (green), nuclei (DAPI-blue), and Alexa647-conjugated streptavidin (red). H) Average intensities of biotinylated peptide (indicated by streptavidin Alexa647 fluorescence intensity level relative to background) in Vehicle control, αCT1, and αCT11 groups. ** p<0.05; not significant (ns) N=5 hearts/group. Scale bar = 5 μm.

Article Snippet: Phospho-Connexin43 (Ser368) (Cell Signaling, 3511S, Danvers, MA), anti-Cx43 produced in rabbit (Sigma: C6219, St. Louis, MO), anti-GST produced in goat (GE, 27457701, Little Chalfont, UK).

Techniques: Staining, Fluorescence