rabbit anti cx43 primary antibody  (Cell Signaling Technology Inc)

Journal: Advanced healthcare materials

Article Title: Glucocorticoids Alter Bone Microvascular Barrier via MAPK/Connexin43 Mechanisms.

doi: 10.1002/adhm.202404302

Figure Lengend Snippet: Figure 4. Glucocorticoids control endothelial and osteoblasts interactions via Cx43. A) RNA was isolated from 3D devices in the presence of Hyd, Pred, and Psl after 18 days. The expression of cell-cell adhesion genes including zonula occludens-1 (ZO-1), vascular endothelial cadherin (VE-cad), and connexin43 (Cx43) was examined by RT-qPCR. The transcript levels were normalized to 18S. N = 3. B) The protein expression of Cx43 in the presence of GCs was analyzed by immunoblotting. The below panel shows the quantification of Cx43 levels compared to control. N = 5. C) Representative images of immunofluorescent signals for Cx43 (green), IB4 (red), and DAPI (blue) in bone sections from a mouse implanted with Psl pellets for 60 days or corresponding placebo control. Scale bars, 20 μm and 5 μm (zoom in images); Red arrows indicate the presence of Cx43 in the cell membrane area. Quantitative analysis of the Cx43-positive membrane intensity in sections from each mouse stain. n = 10. D) Immunostaining for Cx43 (red) for co-culture system of ECs and EGFP-labeled HOBs (green). The nuclei were stained with DAPI (blue). Membrane localization of Cx43 is indicated by yellow arrows. Scale bars, 20 μm. E) Representative endothelial integrity images on the devices treated with GAP19 (5 μm; 30 min) using fluorescence labeled 70 kDa dextran. Scale bars, 100 μm. The below graph demonstrates the endothelial leakiness by diffusive permeability coefficient (Pd) for control and GAP19 treated. The quantitative data are expressed as means ± SD. N = 3, n = 6; significant differences: *, p-value < 0.05, **, p-value < 0.01, ***, p-value < 0.001.

Article Snippet: Primary antibody against rabbit anti-human Cx43 (1:100 dilution in blocking solution; Cell Signaling Technology) was incubated overnight at 4 °C.

Techniques: Control, Isolation, Expressing, Quantitative RT-PCR, Western Blot, Membrane, Staining, Immunostaining, Co-Culture Assay, Labeling, Permeability

Journal: Advanced healthcare materials

Article Title: Glucocorticoids Alter Bone Microvascular Barrier via MAPK/Connexin43 Mechanisms.

doi: 10.1002/adhm.202404302

Figure Lengend Snippet: Figure 5. Glucocorticoids regulate microvascular barrier function via MAPK/Cx43. A) Left, Western blot analysis showing GCs-induced activation of Phospho-Cx43Ser282, and Cx43 in co-cultures of HOBs and ECs following stimulation with Hyd, Pred, and Psl for 30 min, 60 min, and 120 min. Inhibitor treatment was applied for 30 min. Right, Quantitative analysis of the Western blot results. N = 3. B) Left, Western blot analysis showing GCs-induced activation of MAPKs (ERK, p38, and JNK) in co-cultures of HOBs and ECs following stimulation with Hyd, Pred, and Psl for 30 min, 60 min, and 120 min. Inhibitor treatment was applied for 30 min. Right, Quantitative analysis of the Western blot results. N = 3. C) Representative images showing endothelial integrity in control and inhibitor-treated group exposed to GCs on the devices for 30 min, using fluorescence-labeled 70 kDa Texas Red dextran. Scale bar, 100 μm. Right panel, Graph demonstrates the endothelial leakiness measured by the diffusive permeability coefficient (Pd) for control and inhibitor- treated groups with GCs. The quantitative data are expressed as means ± SD. N = 3, n = 3; significant differences: *, p-value < 0.05, **, p-value < 0.01, ***, p-value < 0.001.

Article Snippet: Primary antibody against rabbit anti-human Cx43 (1:100 dilution in blocking solution; Cell Signaling Technology) was incubated overnight at 4 °C.

Techniques: Western Blot, Activation Assay, Control, Labeling, Permeability

Journal: Cellular & Molecular Biology Letters

Article Title: Acquired drug resistance interferes with the susceptibility of prostate cancer cells to metabolic stress

doi: 10.1186/s11658-022-00400-1

Figure Lengend Snippet: Combined DCX/MET treatment enhances invasive potential of DCX-resistant PC-3 cells. A Displacement of PC-3 WT and DCX-resistant PC-3_DCX20 cells was estimated with time-lapse videomicroscopy immediately after the administration of DCX (2.5 nM) and/or MET (10 mM; cf. Additional file : Figs. S1 and S4, left panel) or after 48-h-long treatment (right panel). B Invasiveness of DCX and/or MET treated cells estimated with Transwell migration assay. C Snail-1 expression and localization in PC-3 WT/PC-3_DCX20 cells cultivated in the presence of (2.5 nM) and/or MET (10 mM) [Snail-1 (green), DNA (blue)]. D , E Cx43 levels and intracellular localization in DCX- and/or MET-treated cells estimated with immunoblotting ( D , β-tubulin was used as a reference protein) and immunofluorescence ( E ; Cx43: yellow, DNA: blue). F GJIC estimated with calcein transfer assay in the absence/presence of AGA (α-glycyrrhetinic acid; 25 μM). Coupling index/ratio represents the percentage of coupled donor cells. The statistical significance of the differences was tested with Student’s t -test; # P ≤ 0.05 versus untreated control; * P ≤ 0.05 as indicated in the charts. All results are representative of at least three independent experiments ( N ≥ 3). Error bars represent SEM. Note increased invasiveness of Snail-1/Cx43 high cells after DCX/MET

Article Snippet: Blocking of unspecific staining (skimmed milk/TBST solution) was followed by the application of primary monoclonal rabbit anti-Cx43 (no. C6219; 1:3000, Sigma-Aldrich), washing, and administration of HRP conjugated goat anti-rabbit IgG (no. 31466, Thermo Fisher Scientific).

Techniques: Transwell Migration Assay, Expressing, Western Blot, Immunofluorescence

Journal: Frontiers in Pain Research

Article Title: Role of Connexin 43 in an Inflammatory Model for TMJ Hyperalgesia

doi: 10.3389/fpain.2021.715871

Figure Lengend Snippet: (A) Expression of glial fibrillary acidic protein (GFAP) and connexin43 (Cx43) in the trigeminal ganglion of an OvXE (a–c) and male rat (d–f) 10 days after intra-TMJ injection of CFA. Scale = 30 μm. (B) Densitometry values expressed as percentage of positive area for Cx43 in male, OvX and OvXE rats under sham conditions and at 4 and 10 days after intra-TMJ injection of CFA based on an average of immunostaining as shown by the examples in panel (A) . * p < 0.05, ** p < 0.01 vs. sham; a p < 0.05 vs. males. Sample size = 4 rats per group, average of 5–7 images per rat.

Article Snippet: Free-floating sections were incubated in blocking buffer (PBS, 0.1% Triton X-100, 1% secondary serum) for 1 h and then incubated with anti-mouse primary antibody for glial fibrillary acidic protein (GFAP, Abnova MAB107670, Walnut, CA) and anti-rabbit primary antibody for Cx43 (Cell Signaling 3512, Danvers, MA) at 1:1,000 in PBS with 0.1% Triton X-100 overnight at 4°C.

Techniques: Expressing, Injection, Immunostaining

Journal: Frontiers in Pain Research

Article Title: Role of Connexin 43 in an Inflammatory Model for TMJ Hyperalgesia

doi: 10.3389/fpain.2021.715871

Figure Lengend Snippet: siRNA for Cx43 inhibits intra-TMJ ATP-evoked MMemg activity in (A) male, (B) OvX and (C) OvXE females treated with CFA 10 days prior to recording. Note that responses to TMJ stimuli in sham (PBS-injected) rats were not affected. * p < 0.05, ** p < 0.01 vs. PBS stimulation; a p < 0.05, b p < 0.01 siRNA treated vs. untreated rats. Sample size = 5–6 rats per treatment group.

Article Snippet: Free-floating sections were incubated in blocking buffer (PBS, 0.1% Triton X-100, 1% secondary serum) for 1 h and then incubated with anti-mouse primary antibody for glial fibrillary acidic protein (GFAP, Abnova MAB107670, Walnut, CA) and anti-rabbit primary antibody for Cx43 (Cell Signaling 3512, Danvers, MA) at 1:1,000 in PBS with 0.1% Triton X-100 overnight at 4°C.

Techniques: Activity Assay, Injection

Journal: Frontiers in Pain Research

Article Title: Role of Connexin 43 in an Inflammatory Model for TMJ Hyperalgesia

doi: 10.3389/fpain.2021.715871

Figure Lengend Snippet: Western blots for Cx43 of TG tissue from OvXE (A) and male rats (B) at 10 days after CFA and treated with siRNA for Cx43 or with PBS by intra-TG injection 3 days prior to tissue collection. (C) Summary of the effects of siRNA for Cx43 on protein levels in the TG of male, OvX, and OvXE females. * p < 0.05; ** p < 0.01 vs sham controls; a p < 0.05, b p < 0.01 vs males vs. sham controls. Sample size = 4 rats per group.

Article Snippet: Free-floating sections were incubated in blocking buffer (PBS, 0.1% Triton X-100, 1% secondary serum) for 1 h and then incubated with anti-mouse primary antibody for glial fibrillary acidic protein (GFAP, Abnova MAB107670, Walnut, CA) and anti-rabbit primary antibody for Cx43 (Cell Signaling 3512, Danvers, MA) at 1:1,000 in PBS with 0.1% Triton X-100 overnight at 4°C.

Techniques: Western Blot, Injection