Journal: Redox Biology

Article Title: RND3 attenuates oxidative stress and vascular remodeling in spontaneously hypertensive rat via inhibiting ROCK1 signaling

doi: 10.1016/j.redox.2021.102204

Figure Lengend Snippet: RND3 mRNA and protein expressions in WKY and SHR. A, RND3 mRNA and protein expressions in aorta and mesentery artery (MA). B, representative images of immunohistochemistry for RND3 staining (brown color) in aorta. C, bar graph showing the relative density of RND3 staining in media and adventitia of aorta. D, RND3 mRNA and protein expressions in VSMCs of WKY and SHR. Values are mean ± SE. *P < 0.05 vs WKY; †P < 0.05 vs media. n = 4 per group. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: The primary anti-RND3 antibody (1:100) and primary anti-PCNA antibody (1:100) were obtained from Protein Tech Group Inc (Chicago, IL, USA).

Techniques: Immunohistochemistry, Staining

Journal: Redox Biology

Article Title: RND3 attenuates oxidative stress and vascular remodeling in spontaneously hypertensive rat via inhibiting ROCK1 signaling

doi: 10.1016/j.redox.2021.102204

Figure Lengend Snippet: Effects of RND3 overexpression (OE) on superoxide production and NOX expressions in VSMCs of WKY and SHR. The VSMCs of WKY and SHR were treated with PBS, negative control (NC) or RND3 overexpressed plasmid (50 nM) for 24 h. A, superoxide level. B, relative DHE fluorescence intensity. C, representative images of DHE fluorescence staining. D, H2O2 level. E, NOX activity. F, NOX1, NOX2 and NOX4 mRNA levels. G, NOX1, NOX2 and NOX4 protein expressions. Values are mean ± SE. *P < 0.05 vs WKY; †P < 0.05 vs PBS or NC. n = 6 per group in A-E; n = 3 per group in F. n = 4 per group in G.

Article Snippet: The primary anti-RND3 antibody (1:100) and primary anti-PCNA antibody (1:100) were obtained from Protein Tech Group Inc (Chicago, IL, USA).

Techniques: Over Expression, Negative Control, Plasmid Preparation, Fluorescence, Staining, Activity Assay

Journal: Redox Biology

Article Title: RND3 attenuates oxidative stress and vascular remodeling in spontaneously hypertensive rat via inhibiting ROCK1 signaling

doi: 10.1016/j.redox.2021.102204

Figure Lengend Snippet: Roles of RND3 overexpression (OE) in VSMCs migration and proliferation of WKY and SHR. The VSMCs of WKY and SHR were treated with PBS, negative control (NC) or RND3 overexpressed plasmid (50 nM) for 24 h. A, VSMCs migration evaluated with wound healing assay. B, VSMCs migration evaluated with Boyden chamber assay. C, PCNA protein expression. D, percentage of EdU-positive cells. E, representative images of EdU-positive cells. The red fluorescence represents proliferating cells, and the blue fluorescence represents total cells. Values are mean ± SE. *P < 0.05 vs WKY; †P < 0.05 vs PBS or NC. n = 4 per group in C; n = 6 per group in others. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: The primary anti-RND3 antibody (1:100) and primary anti-PCNA antibody (1:100) were obtained from Protein Tech Group Inc (Chicago, IL, USA).

Techniques: Over Expression, Migration, Negative Control, Plasmid Preparation, Wound Healing Assay, Boyden Chamber Assay, Expressing, Fluorescence

Journal: Redox Biology

Article Title: RND3 attenuates oxidative stress and vascular remodeling in spontaneously hypertensive rat via inhibiting ROCK1 signaling

doi: 10.1016/j.redox.2021.102204

Figure Lengend Snippet: Comparing the effects of ML171 (a NOX1 inhibitor), GSK2795039 (a NOX2 inhibitor) and PEG-SOD with the effects of RND3 overexpression (RND3 OE) on VSMCs migration and proliferation in VSMCs of SHR. The VSMCs migration and proliferation were respectively evaluated with wound healing assay (A) and EdU incorporation assay (B). The VSMCs of SHR were treated with PBS, 1% DMSO (vehicle), RND3 overexpressed plasmid (50 nM), ML171 (10 μM), GSK2795039 (25 μM) or PEG-SOD (25 U/mL) for 24 h. Values are mean ± SE.. *P < 0.05 vs PBS or DMSO; †P < 0.05 vs RND3; ‡P < 0.05 vs ML171. #P < 0.05 vs GSK2795039. n = 6 per group.

Article Snippet: The primary anti-RND3 antibody (1:100) and primary anti-PCNA antibody (1:100) were obtained from Protein Tech Group Inc (Chicago, IL, USA).

Techniques: Over Expression, Migration, Wound Healing Assay, Plasmid Preparation

Journal: Redox Biology

Article Title: RND3 attenuates oxidative stress and vascular remodeling in spontaneously hypertensive rat via inhibiting ROCK1 signaling

doi: 10.1016/j.redox.2021.102204

Figure Lengend Snippet: ROCK1 expression and phosphorylation of MYPT1 in the VSMCs of WKY and SHR treated with RND3 overexpression. The VSMCs were treated with PBS, negative control (NC) or RND3 overexpressed plasmid (50 nM) for 24 h. A, ROCK1 protein expression. B, RhoA protein expression. C. ROCK1 activity evaluated with the phosphorylation of MYPT1. D, RND3 protein expression. Values are mean ± SE. *P < 0.05 vs WKY; †P < 0.05 vs PBS or NC. n = 4 per group.

Article Snippet: The primary anti-RND3 antibody (1:100) and primary anti-PCNA antibody (1:100) were obtained from Protein Tech Group Inc (Chicago, IL, USA).

Techniques: Expressing, Phospho-proteomics, Over Expression, Negative Control, Plasmid Preparation, Activity Assay

Journal: Redox Biology

Article Title: RND3 attenuates oxidative stress and vascular remodeling in spontaneously hypertensive rat via inhibiting ROCK1 signaling

doi: 10.1016/j.redox.2021.102204

Figure Lengend Snippet: Effects of ROCK1 OE on the roles of RND3 OE in inhibiting VSMCs oxidative stress. The VSMCs were treated with PBS, negative control (NC) or RND3 OE plasmid (50 nmol/L) and ROCK1 OE plasmid (50 nmol/L) for 24 h. A, RND3 and ROCK1 protein expressions. B, DHE fluorescence staining for detecting ROS in VSMCs C, Superoxide level. D, H2O2 level. E, NOX activity. F, NOX1 and NOX2 protein expressions. Values are mean ± SE. *P < 0.05 vs WKY; †P < 0.05 vs Ctrl; ‡P < 0.05 vs PBS or NC. n = 3 per group in A and G. n = 6 per group in others.

Article Snippet: The primary anti-RND3 antibody (1:100) and primary anti-PCNA antibody (1:100) were obtained from Protein Tech Group Inc (Chicago, IL, USA).

Techniques: Negative Control, Plasmid Preparation, Fluorescence, Staining, Activity Assay

Journal: Redox Biology

Article Title: RND3 attenuates oxidative stress and vascular remodeling in spontaneously hypertensive rat via inhibiting ROCK1 signaling

doi: 10.1016/j.redox.2021.102204

Figure Lengend Snippet: Effects of ROCK1 OE on the roles of RND3 OE in inhibiting VSMCs migration and proliferation. The VSMCs were treated with PBS, negative control (NC) or RND3 OE plasmid (50 nmol/L) and ROCK1 OE plasmid (50 nmol/L) for 24 h. A & B, VSMCs migration evaluated with wound healing assay (A) and Boyden chamber assay (B). C & D, VSMCs proliferation were evaluated with PCNA protein expression (C) and EdU-positive cells (D). Values are mean ± SE. *P < 0.05 vs WKY; †P < 0.05 vs Ctrl; ‡P < 0.05 vs PBS or NC. n = 6 per group.

Article Snippet: The primary anti-RND3 antibody (1:100) and primary anti-PCNA antibody (1:100) were obtained from Protein Tech Group Inc (Chicago, IL, USA).

Techniques: Migration, Negative Control, Plasmid Preparation, Wound Healing Assay, Boyden Chamber Assay, Expressing

Journal: Redox Biology

Article Title: RND3 attenuates oxidative stress and vascular remodeling in spontaneously hypertensive rat via inhibiting ROCK1 signaling

doi: 10.1016/j.redox.2021.102204

Figure Lengend Snippet: Therapeutic effects of adenoviral (Ad)-mediated RND3 overexpression on hypertension and vascular remodeling. The rat in WKY or SHR group was subjected to intravenous administration of PBS, Ad-scrambled RND3 (NC) or Ad-RND3 (RND3 OE, 1 × 107 plaque forming units). The blood pressure was measured every week in awake state. The vascular remodeling was examined 4 weeks after the intravenous injection. A, systolic blood pressure (SBP), diastolic blood pressure (DBP), mean arterial pressure (MAP) and heart rate (HR) in WKY and SHR. B, bar graph showing the Masson's staining analysis for media thickness, lumen diameter and the ratio of medium thickness to lumen diameter in aorta and mesentery artery (MA). C, representative images of Masson's staining of aorta and MA. D, bar graph showing the relative values of the PCNA immunohistochemistry analyses. E, representative images of PCNA immunohistochemistry (brown color) in aorta and mesentery artery (MA). Values are mean ± SE. *P < 0.05 vs WKY; †P < 0.05 vs. PBS or NC. n = 6 per group. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: The primary anti-RND3 antibody (1:100) and primary anti-PCNA antibody (1:100) were obtained from Protein Tech Group Inc (Chicago, IL, USA).

Techniques: Over Expression, Injection, Staining, Immunohistochemistry

Journal: Redox Biology

Article Title: RND3 attenuates oxidative stress and vascular remodeling in spontaneously hypertensive rat via inhibiting ROCK1 signaling

doi: 10.1016/j.redox.2021.102204

Figure Lengend Snippet: Roles of adenoviral (Ad)-mediated RND3 overexpression in WKY and SHR in PCNA and ROCK1 protein expressions, phosphorylation of MYPT1 and oxidative stress. The rat was subjected to intravenous administration of PBS, Ad-scrambled RND3 (NC) or Ad-RND3 (RND3 OE, 1 × 107 plaque forming units). The measurements were performed 4 weeks after the intravenous injection. A, ROCK1 protein expression. B, MYPT1 phosphorylation. C, bar graph showing the relative values of the DHE fluorescence staining analyses. D, representative images of DHE fluorescence staining, a marker of ROS production in aorta and MA. E, NOX1, NOX2 and NOX4 mRNA levels. Values are mean ± SE. *P < 0.05 vs WKY; †P < 0.05 vs. PBS or NC. n = 6 per group.

Article Snippet: The primary anti-RND3 antibody (1:100) and primary anti-PCNA antibody (1:100) were obtained from Protein Tech Group Inc (Chicago, IL, USA).

Techniques: Over Expression, Phospho-proteomics, Injection, Expressing, Fluorescence, Staining, Marker

Journal: BMC cancer

Article Title: RhoB regulates prostate cancer cell proliferation and docetaxel sensitivity via the PI3K-AKT signaling pathway.

doi: 10.1186/s12885-025-13762-4

Figure Lengend Snippet: Fig. 8 RhoB activates the PI3K-AKT signaling pathway upon DTXL treatment in prostate cancer cells. (A) In the presence of DTXL, RhoB OE can activate AKT (p-308) while, in the absence of DTXL, RhoB OE can inhibit AKT (p-308). (B) Immunofluorescence co-staining of p-FAK and F-actin in prostate cancer cells with RhoB different expression status after DTXL treatment. F-actin and p-FAK immunostaining. Representative microphotographs are shown, objec tive 20×, F-actin (red), p-FAK (green), nucleus staining (blue)

Article Snippet: Immunohistochemical (IHC) staining was used to detect the expression of RhoB (14326-1-AP, Proteintech), cleaved caspase-3 (GB11532100), Ki67 (GB121141-100) on the tumor cells.

Techniques: Immunofluorescence, Staining, Expressing, Immunostaining

Journal: BMC cancer

Article Title: RhoB regulates prostate cancer cell proliferation and docetaxel sensitivity via the PI3K-AKT signaling pathway.

doi: 10.1186/s12885-025-13762-4

Figure Lengend Snippet: Fig. 9 Prostate cancer xenograft to study the RhoB effects on cancer progression. (A, B) The growth curve displays that RhoB OE could inhibit cancer growth. (C, D) TCGA data showed the RhoB expression in normal prostate tissues and prostate adenocarcinoma (PRAD) tissues, while the survival curve demonstrated that the RhoB expression affected the survival time of prostate cancer patients. (E) H&E staining of tumor tissues dissected from the pros tate cancer xenograft. (F) TUNEL staining of tumor tissues with RhoB overexpression or knockout. (G, H, I) IHC staining of Ki67, cleaved caspase 3 and RhoB proteins in the tumor tissues with RhoB overexpression or knockout, objective 20×. (J) Quantification of IHC staining for Ki67, cleaved caspase 3, and RhoB proteins in prostate cancer xenograft. *P < 0.05, when compared with indicated group, one way ANOVA

Article Snippet: Immunohistochemical (IHC) staining was used to detect the expression of RhoB (14326-1-AP, Proteintech), cleaved caspase-3 (GB11532100), Ki67 (GB121141-100) on the tumor cells.

Techniques: Expressing, Staining, TUNEL Assay, Over Expression, Knock-Out, Immunohistochemistry

Journal: Cells

Article Title: Hyperglycemic Stress Induces Expression, Degradation, and Nuclear Association of Rho GDP Dissociation Inhibitor 2 (RhoGDIβ) in Pancreatic β-Cells

doi: 10.3390/cells13030272

Figure Lengend Snippet: RhoGDI expression in INS-1 832/13 cells, MIN6 cells, rat islets, and human islets. Western blot data depicting the expression of RhoGDIα, RhoGDIβ, and RhoGDIγ in INS1 832/13 cells, MIN6 cells, rat islets, and human islets.

Article Snippet: Antibodies specific for RhoGDIα (sc-373724), RhoGDIβ (sc-271108), RhoGDIγ (sc-393690), cleaved RhoGDIβ (sc-52936), and Lamin B (sc-56144) were obtained from Santa Cruz Biotechnology (Dallas, TX, USA).

Techniques: Expressing, Western Blot

Journal: Cells

Article Title: Hyperglycemic Stress Induces Expression, Degradation, and Nuclear Association of Rho GDP Dissociation Inhibitor 2 (RhoGDIβ) in Pancreatic β-Cells

doi: 10.3390/cells13030272

Figure Lengend Snippet: Expression levels of RhoGDIα, RhoGDIβ, and RhoGDIγ in INS-1 832/13 cells exposed to either basal or chronic hyperglycemic conditions. ( A ): Representative Western blots from multiple studies showing the effect of high-glucose conditions on the expression of RhoGDIα (n = 3), RhoGDIβ (n = 7), and RhoGDIγ (n = 3). Cells were cultured in the presence of either basal (LG; 2.5 mM) or high-glucose (HG; 20 mM) treatment for 24 h and the degree of expression of RhoGDIα, RhoGDIβ, and RhoGDIγ was determined by Western blotting. β-actin was used as a loading control. ( B ): Densitometric quantification of RhoGDIα, RhoGDIβ, and RhoGDIγ using Image Studio Lite v3.1 (Li-COR; Lincoln, Nebraska). Data are presented as mean ± SEM with values shown as fold change from LG; significance is considered for p -values < 0.05 (**: p -value < 0.01) ns = not significant.

Article Snippet: Antibodies specific for RhoGDIα (sc-373724), RhoGDIβ (sc-271108), RhoGDIγ (sc-393690), cleaved RhoGDIβ (sc-52936), and Lamin B (sc-56144) were obtained from Santa Cruz Biotechnology (Dallas, TX, USA).

Techniques: Expressing, Western Blot, Cell Culture, Control

Journal: Cells

Article Title: Hyperglycemic Stress Induces Expression, Degradation, and Nuclear Association of Rho GDP Dissociation Inhibitor 2 (RhoGDIβ) in Pancreatic β-Cells

doi: 10.3390/cells13030272

Figure Lengend Snippet: Expression levels of RhoGDIα, RhoGDIβ, and RhoGDIγ in rat and human islets exposed to either basal or chronic hyperglycemic conditions. Representative Western blots from rodents and human islets showing the expression of RhoGDIα, RhoGDIβ, and RhoGDIγ when exposed to 24 h basal (LG; 2.5 mM) or high glucose (HG; 20 mM). β-actin was used as a loading control. Rat islet data shown are from a single study (n = 1), while human islet data are from 2 separate studies.

Article Snippet: Antibodies specific for RhoGDIα (sc-373724), RhoGDIβ (sc-271108), RhoGDIγ (sc-393690), cleaved RhoGDIβ (sc-52936), and Lamin B (sc-56144) were obtained from Santa Cruz Biotechnology (Dallas, TX, USA).

Techniques: Expressing, Western Blot, Control

Journal: Cells

Article Title: Hyperglycemic Stress Induces Expression, Degradation, and Nuclear Association of Rho GDP Dissociation Inhibitor 2 (RhoGDIβ) in Pancreatic β-Cells

doi: 10.3390/cells13030272

Figure Lengend Snippet: Chronic hyperglycemic conditions increase the cleavage of RhoGDIβ in INS-1 832/13 cells. ( A ): A representative Western blot displaying cleaved RhoGDIβ expression in INS-1 832/13 lysates exposed to basal (LG; 2.5 mM) or high-glucose (HG; 20 mM) conditions for 24 h is shown here. β-actin was used as a loading control. ( B ): Densitometric quantification of expression levels of cleaved RhoGDIβ in INS-1 832/13 cells following exposure to either basal (LG; 2.5 mM) or high glucose (HG; 20 mM) for 24 h. Data are presented as mean ± SEM. Results are represented as fold change from LG (n = 5; ***: p < 0.001).

Article Snippet: Antibodies specific for RhoGDIα (sc-373724), RhoGDIβ (sc-271108), RhoGDIγ (sc-393690), cleaved RhoGDIβ (sc-52936), and Lamin B (sc-56144) were obtained from Santa Cruz Biotechnology (Dallas, TX, USA).

Techniques: Western Blot, Expressing, Control

Journal: Cells

Article Title: Hyperglycemic Stress Induces Expression, Degradation, and Nuclear Association of Rho GDP Dissociation Inhibitor 2 (RhoGDIβ) in Pancreatic β-Cells

doi: 10.3390/cells13030272

Figure Lengend Snippet: Confocal imaging analysis of the expression of RhoGDIα, RhoGDIγ, and RhoGDIβ (full-length and cleaved forms) in INS-1 832/13 cells. Representative confocal images displaying the expression and subcellular distribution of RhoGDIα ( A ), RhoGDIγ ( B ), full-length RhoGDIβ ( C ), and cleaved RhoGDIβ ( D ) using FITC conjugated secondary antibody (green) in INS-1 832/13 cells exposed to either basal (LG; 2.5 mM) or high-glucose (HG; 20 mM) conditions for 24 h, as described in the text. DAPI is shown as a marker for the nuclear association (blue). Images shown are representative of 2 independent confocal studies.

Article Snippet: Antibodies specific for RhoGDIα (sc-373724), RhoGDIβ (sc-271108), RhoGDIγ (sc-393690), cleaved RhoGDIβ (sc-52936), and Lamin B (sc-56144) were obtained from Santa Cruz Biotechnology (Dallas, TX, USA).

Techniques: Imaging, Expressing, Marker

Journal: Cells

Article Title: Hyperglycemic Stress Induces Expression, Degradation, and Nuclear Association of Rho GDP Dissociation Inhibitor 2 (RhoGDIβ) in Pancreatic β-Cells

doi: 10.3390/cells13030272

Figure Lengend Snippet: Non-nuclear and nuclear association of RhoGDIα, RhoGDIγ, RhoGDIβ (full-length and cleaved) in INS-1 832/13 cells exposed to basal or hyperglycemic conditions. A representative Western blot depicting the association of RhoGDIα ( A ) and RhoGDIγ ( C ). Densitometric analysis of cytosol (CYT) and nuclear (NUC) expression for RhoGDIα ( B ) and RhoGDIγ ( D ) in INS-1 832/13 cells exposed to either basal (LG; 2.5 mM) or high-glucose (HG; 20 mM) conditions for 24 h from 3 independent studies (n = 3; nd: not detected). Representative Western blots showing the expression of full-length RhoGDIβ ( E ) and cleaved RhoGDIβ ( G ) in INS-1 832/13 cells exposed to either basal (LG; 2.5 mM) or high-glucose (HG; 20 mM) conditions for 24 h are shown here. Densitometric analysis of pooled data are presented for full-length RhoGDIβ (( F ), n = 3) and cleaved RhoGDIβ (( H ), n = 3). *: p -value < 0.05, **: p -value < 0.01. GAPDH and Lamin B were used as loading and purity controls for the cytosolic and nuclear fractions, respectively. Data are expressed as fold change from LG; significance considered when p < 0.05. ns = not significant; nd = not detected.

Article Snippet: Antibodies specific for RhoGDIα (sc-373724), RhoGDIβ (sc-271108), RhoGDIγ (sc-393690), cleaved RhoGDIβ (sc-52936), and Lamin B (sc-56144) were obtained from Santa Cruz Biotechnology (Dallas, TX, USA).

Techniques: Western Blot, Expressing

Journal: Cells

Article Title: Hyperglycemic Stress Induces Expression, Degradation, and Nuclear Association of Rho GDP Dissociation Inhibitor 2 (RhoGDIβ) in Pancreatic β-Cells

doi: 10.3390/cells13030272

Figure Lengend Snippet: Examination of non-nuclear and nuclear association of full length RhoGDIβ in INS-1 832/13 cells exposed acutely to either basal or stimulatory glucose. Representative Western blots depicting relative abundance of full-length RhoGDIβ ( A ) in INS-1 832/13 cells exposed to either basal (2.5 mM) or stimulatory glucose (20 mM) for 45 min are shown here. Densitometric analyses of data from multiple experiments highlighting abundance in the cytosolic (CYT) and nuclear (NUC) fractions for full-length RhoGDIβ ( B ) are shown. GAPDH and Lamin B were used as markers for the cytosolic and nuclear fractions, respectively. Data are expressed as fold change from LG; *: p < 0.05 (n = 3). nd = not detected.

Article Snippet: Antibodies specific for RhoGDIα (sc-373724), RhoGDIβ (sc-271108), RhoGDIγ (sc-393690), cleaved RhoGDIβ (sc-52936), and Lamin B (sc-56144) were obtained from Santa Cruz Biotechnology (Dallas, TX, USA).

Techniques: Western Blot

Journal: Scientific Reports

Article Title: Hindlimb Ischemia Impairs Endothelial Recovery and Increases Neointimal Proliferation in the Carotid Artery

doi: 10.1038/s41598-017-19136-6

Figure Lengend Snippet: miR-16 upregulation leads to increased activation of RhoA by targeting RhoGDIα in endothelial cell. ( a ) Expression levels of RhoGDIα in ECs from rat carotid artery of BI and FL-BI groups at 14 days after balloon injury. * P < 0.01 versus control group; n = 5. ( b ) Cultured ECs were transfected with miR-16 Mimic, miR-16 Inhibitor, Mimic-NC, or Inhibitor-NC, showing modulation in RhoGDIα levels 48 hours after manipulation of miR-16 levels (real-time RT-PCR). * P < 0.05 versus cells transfected with Mimic-NC; n = 4. ( c ) Representative immunoblotting (upper) and quantification (lower) of RhoGDIα protein levels in HUVEC transfected with miR-16 Mimic, miR-16 Inhibitor, Mimic-NC or Inhibitor-NC. * P < 0,01 versus cells transfected with Mimic-NC; n = 4 ( d ) Representative immunoblotting (upper) and quantification (lower) of p-AKT in ECs transfected with miR-16 Inhibitor. * P < 0,01 versus cells transfected with Inhibitor-NC; n = 5. ( e ) Assay of RhoA activation in ECs transfected with miR-16 Inhibitor and stimulated with TNF. * P < 0.05 versus cells tranfected with Inhibitor-NC and treated with TNFα; n = 4.

Article Snippet: The antibodies used were the following: polyclonal rabbit anti-eNOS antibody (Cell Signaling, catalog number #9572), goat polyclonal anti-phospho-eNOS (Ser 1177) antibody (Santa Cruz Biotech., sc-12972), AKT antibody (Cell Signaling, catalog number #9272), rabbit Phospho-Akt (Ser473) antibody (Cell Signaling, catalog number #193H12), mouse monoclonal RhoGDIα antibody (Santa Cruz Biotech., sc-373724) and rabbit polyclonal anti-GAPDH (FL-335) antibody (Santa Cruz Biotech., sc-25778).

Techniques: Activation Assay, Expressing, Control, Cell Culture, Transfection, Quantitative RT-PCR, Western Blot

Journal: Scientific Reports

Article Title: Hindlimb Ischemia Impairs Endothelial Recovery and Increases Neointimal Proliferation in the Carotid Artery

doi: 10.1038/s41598-017-19136-6

Figure Lengend Snippet: Systemic delivery of antagomiR-16 promotes endothelial recovery and inhibits neointima formation in the carotid artery of rats with hindlimb ischemia. ( a ) Schematic model of the experimental setup. ( b ) Expression levels of miR-16 in vascular endothelium. Total RNAs were obtained from vascular endothelium of rat carotid artery 14 days after injury. * P < 0.01 versus control group; n = 6. ( c ) Relative expression of RhoGDIα and eNOS mRNA transcripts in vascular endothelium of rat carotid artery 14 days after injury. * P < 0.05 versus control group; n = 6. ( d ) Left: Representative images of Haematoxylin and eosin staining in balloon-injured carotid arteries at 14 days in rats treated with or without Antago-16. Scale bars, 100 µm. Right: Bar graphs represent the morphometric analysis of arterial sections. Neointima /media ratio of arteries in differently treated groups is shown. * P < 0.05 versus NC group; n = 7. ( e ) Left: Representative sections of carotid arteries immunostained for the specific endothelial cell marker CD31. Nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI). Carotid arteries were explanted from experimental groups at 14 days after balloon injury. Scale bars = 50 µm. Right: Bar graphs represents the percentage of re-endothelializated circumference of the common carotid artery. * P < 0.05 versus rat treated with antagomir scrambled; n = 6 for group. ( f ) Left: Representative sections of carotid arteries stained for the macrophage (brown) marker CD68. Carotid arteries were explanted from rats at 14 days after balloon injury. Right: Quantitative data derived from arterial sections stained with CD68 positive cells. * P < 0.05 versus control; n = 5.

Article Snippet: The antibodies used were the following: polyclonal rabbit anti-eNOS antibody (Cell Signaling, catalog number #9572), goat polyclonal anti-phospho-eNOS (Ser 1177) antibody (Santa Cruz Biotech., sc-12972), AKT antibody (Cell Signaling, catalog number #9272), rabbit Phospho-Akt (Ser473) antibody (Cell Signaling, catalog number #193H12), mouse monoclonal RhoGDIα antibody (Santa Cruz Biotech., sc-373724) and rabbit polyclonal anti-GAPDH (FL-335) antibody (Santa Cruz Biotech., sc-25778).

Techniques: Expressing, Control, Staining, Marker, Derivative Assay