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Results: 7

1.
Figure 7

Figure 7. From: Coupling of Fc?RI to Fc?RIIB by Src Kinase Mediates C-reactive Protein Impairment of Endothelial Function.

The impairment of reendothelialization by CRP requires FcγRIIB. Carotid artery reendothelialization was evaluated in FcγRIIB+/+, FcγRIIB+/+;TG-CRP, FcγRIIB−/− and FcγRIIB−/−;TG-CRP littermates. A. The area of remaining denudation 96h post-injury that has incorporated Evans blue dye is shown for representative studies. B. The area of denudation was quantified and expressed in arbitrary units. Values are mean±SEM (n=10–12 mice/group), *p<0.05 vs no CRP, †p<0.05 vs FcγRIIB+/+;TG-CRP.

Nathan C. Sundgren, et al. Circ Res. ;109(10):1132-1140.
2.
Figure 6

Figure 6. From: Coupling of Fc?RI to Fc?RIIB by Src Kinase Mediates C-reactive Protein Impairment of Endothelial Function.

The impairment of reendothelialization by CRP requires the γ chain of FcγRI. Carotid artery reendothelialization was evaluated in FcRγ+/+, FcRγ+/+;TG-CRP, FcRγ−/− and FcRγ−/−;TG-CRP littermates. A. The area of remaining denudation 96h post-injury that has incorporated Evans blue dye is shown for representative studies. B. The area of denudation was quantified and expressed in arbitrary units. Values are mean±SEM (n=8–14 mice/group), *p<0.05 vs no CRP, †p<0.05 vs FcRγ+/+;TG-CRP.

Nathan C. Sundgren, et al. Circ Res. ;109(10):1132-1140.
3.
Figure 1

Figure 1. From: Coupling of Fc?RI to Fc?RIIB by Src Kinase Mediates C-reactive Protein Impairment of Endothelial Function.

CRP inhibits eNOS activation via FcγRI. BAEC were preincubated for 2h in the presence of control IgG (10μg/mL) or blocking antibodies to FcγRI (monoclonal 10.1 or polyclonal H-250 Abs) or FcγRIII (10μg/mL), and further preincubated for 20 min with CRP (25μg/mL). eNOS activation by insulin (500nM) was then evaluated by quantifying 14C-L-arginine conversion to 14C-L-citrulline during 15 min incubations. Values are mean±SEM, n=6, *p<0.05 vs no CRP, †p<0.05 vs Control.

Nathan C. Sundgren, et al. Circ Res. ;109(10):1132-1140.
4.
Figure 2

Figure 2. From: Coupling of Fc?RI to Fc?RIIB by Src Kinase Mediates C-reactive Protein Impairment of Endothelial Function.

CRP inhibits eNOS activation via Src kinase. eNOS activation by acetylcholine (Ach, 10μM), in the absence or presence of CRP (5μg/mL), was measured in intact BAEC by quantifying 14C-L-arginine conversion to 14C-L-citrulline during 15 min incubations. Incubations were performed in the absence or presence of the Src kinase inhibitor PP2 (10μM) (A) or the Syk inhibitor piceatannol (Pice, 10μM) (B). Values are mean±SEM, n=6–8, *p<0.05 vs Basal, †p<0.05 vs no CRP.

Nathan C. Sundgren, et al. Circ Res. ;109(10):1132-1140.
5.
Figure 3

Figure 3. From: Coupling of Fc?RI to Fc?RIIB by Src Kinase Mediates C-reactive Protein Impairment of Endothelial Function.

CRP stimulates Src through FcγRI. A. BAEC were treated with CRP (25μg/mL) for 0–30 min and Src activation was analyzed by immunoblot analysis for phospho-Src (Tyr416) and total Src. HDL (50μg/mL) treatment for 10 min provided a positive control. Summary data for the ratio of pSrc to Src normalized to time 0 is provided. B. BAEC were treated with 0–25μg/mL CRP for 10 min, and Src activation was analyzed. Summary data for the ratio of pSrc to Src normalized to time 0 is provided. C. Mouse primary aortic endothelial cells from FcγRI+/+ or FcγRI−/− mice were treated with HDL (50μg/mL) for 10min or CRP (25μg/mL) for 15 min, and Src activation was analyzed. Summary data for the ratio of pSrc to Src normalized to basal is provided. Values are mean±SEM, n=4–6, *p<0.05 vs no CRP.

Nathan C. Sundgren, et al. Circ Res. ;109(10):1132-1140.
6.
Figure 4

Figure 4. From: Coupling of Fc?RI to Fc?RIIB by Src Kinase Mediates C-reactive Protein Impairment of Endothelial Function.

Src activation by CRP is required for FcγRIIB activation and subsequent SHIP-1 activation. A. BAEC were transfected with cDNA encoding human FcγRIIB and 48h later treated with CRP (100μg/ml, 15 min) in the presence of vehicle control (DMSO) or PP2 (10μM). Cell lysates were immunoblotted with anti-phospho-FcγRIIB (Tyr292) antibody and anti-FcγRIIB antibody. Summary data for P-FcγRIIB to total FcγRIIB ratio is normalized to basal conditions. B. BAEC were treated with CRP (25μg/ml, 30 min) in the absence or presence of PP2, and cell lysates were immunoblotted with anti-phospho-SHIP-1 (Tyr1020) and anti-SHIP-1 antibodies. Summary data for P-SHIP-1 to total SHIP-1 ratio is normalized to basal conditions. In A and B, values are mean±SEM, n=9, *p<0.05 vs no CRP, †p<0.05 vs no PP2.

Nathan C. Sundgren, et al. Circ Res. ;109(10):1132-1140.
7.
Figure 5

Figure 5. From: Coupling of Fc?RI to Fc?RIIB by Src Kinase Mediates C-reactive Protein Impairment of Endothelial Function.

Immune complex activation of Src causes FcγRIIB and SHIP-1 activation and eNOS antagonism. Heat-aggregated IgG (aIgG) was used to mimick immune complex. A. eNOS activation in response to acetylcholine (Ach, 10μM) was evaluated in the absence or presence of aIgG (5μg/mL), with and without 10μM PP2 added. Values are mean±SEM, n=8, *p<0.05 vs Basal, †p<0.05 vs no aIgG. B. BAEC were transfected with cDNA encoding human FcγRIIB and 48h later treated with aIgG (100μg/mL) for 15 min in the presence of vehicle control (DMSO) or PP2 (10μM). Cell lysates were immunoblotted with anti-phospho-FcγRIIB (Tyr292) antibody and anti-FcγRIIB antibody. Summary data for pFcγRIIB to total FcγRIIB ratio is normalized to basal conditions. C. BAEC were treated with aIgG (100μg/mL) for 30 min in the absence or presence of PP2, and cell lysates were immunoblotted with anti-phospho-SHIP-1 (Tyr1020) and anti-SHIP-1 antibodies. Summary data for pSHIP-1 to total SHIP-1 ratio is normalized to basal conditions. In B and C, values are mean±SEM, n=9, *p<0.05 vs Basal, †p<0.05 vs no PP2.

Nathan C. Sundgren, et al. Circ Res. ;109(10):1132-1140.

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