J de Bono

RGS1 regulates myeloid cell accumulation in atherosclerosis and aortic aneurysm rupture through altered chemokine signalling

Chemokine signalling drives monocyte recruitment in atherosclerosis and aortic aneurysms. The mechanisms that lead to retention and accumulation of macrophages in the vascular wall remain unclear. Regulator of G-Protein Signalling-1 (RGS1) deactivates G-protein signalling, reducing the response to sustained chemokine stimulation. Here we show that Rgs1 is upregulated in atherosclerotic plaque and aortic aneurysms. Rgs1 reduces macrophage chemotaxis and desensitizes chemokine receptor signalling. In early atherosclerotic lesions, Rgs1 regulates macrophage accumulation and is required for the formation and rupture of Angiotensin II-induced aortic aneurysms, through effects on leukocyte retention. Collectively, these data reveal a role for Rgs1 in leukocyte trafficking and vascular inflammation and identify Rgs1, and inhibition of chemokine receptor signalling as potential therapeutic targets in vascular disease.

Mammalian γ2 AMPK regulates intrinsic heart rate

AMPK is a conserved serine/threonine kinase whose activity maintains cellular energy homeostasis. Eukaryotic AMPK exists as αβγ complexes, whose regulatory γ subunit confers energy sensor function by binding adenine nucleotides. Humans bearing activating mutations in the γ2 subunit exhibit a phenotype including unexplained slowing of heart rate (bradycardia). Here, we show that γ2 AMPK activation downregulates fundamental sinoatrial cell pacemaker mechanisms to lower heart rate, including sarcolemmal hyperpolarization-activated current (If) and ryanodine receptor-derived diastolic local subsarcolemmal Ca2+ release. In contrast, loss of γ2 AMPK induces a reciprocal phenotype of increased heart rate, and prevents the adaptive intrinsic bradycardia of endurance training. Our results reveal that in mammals, for which heart rate is a key determinant of cardiac energy demand, AMPK functions in an organ-specific manner to maintain cardiac energy homeostasis and determines cardiac physiological adaptation to exercise by modulating intrinsic sinoatrial cell behavior.AMPK regulates cellular energy balance using its γ subunit as an energy sensor of cellular AMP and ADP to ATP ratios. Here, the authors show that γ2 AMPK activation lowers heart rate by reducing the activity of pacemaker cells, whereas loss of γ2 AMPK increases heart rate and prevents the adaptive bradycardia of endurance training in mice.

YIA5 RGS-1 Regulates Leukocyte Trafficking in Atherosclerosis and Aortic Aneurysm Formation through Chemokine Receptor Desensitisation

The regulation of macrophage recruitment and retention into the vascular wall is critical in the progression of atherosclerosis and aortic aneurysm formation. This can be mediated by chemokine activation of multiple G-protein coupled receptors. The Regulator of G-Protein Signalling-1 (RGS-1) acts to deactivate the intracellular response to sustained chemokine stimulation. We have found that RGS-1 is up-regulated with atherosclerotic plaque progression and with monocyte-macrophage activation. However little is known about the role of RGS-1 in macrophage function in vivo . Rgs-1 deficient macrophages have significantly enhanced migratory responses to atherogenic chemokines (p < 0.05) and have impaired desensitisation to repeated chemokine re-stimulation (p < 0.001). In vivo , RGS-1 has a role in the accumulation of macrophages in atherosclerotic lesions and during Angiotensin-II aortic aneurysm rupture. In the absence of RGS1, atherosclerosis is attenuated in early lesions in the aortic root and aortas of ApoE-/- mice (p < 0.001) which is accompanied by fewer macrophages in the aortic root. Rgs1-/-ApoE-/- mice are protected from Angiotensin-II induced aneurysm rupture compared to ApoE-/- mice with 94% survival vs. 56% (p = 0.0147). Rgs1-/-ApoE-/- mice have significantly fewer CD11b+ myeloid cells and CD14+ macrophages in aortas than ApoE-/- mice (p < 0.05) after 5 days of Angiotensin-II infusion. Following bone marrow transplantation, recipient mice receiving ApoE-/- bone marrow were more susceptible to aortic aneurysm rupture (p = 0.0124), indicating bone marrow-derived RGS-1 is required for aneurysm rupture. Furthermore, Angiotensin-II treatment increased systolic blood pressure to a greater extent in Rgs1-/-ApoE-/- mice than ApoE-/- mice suggesting aneurysm formation in these mice is independent of Angiotensin-II induced hypertension and this is mediated by vascular-derived RGS-1.To gain insight into the mechanism by which RGS1 regulates trafficking, we selectively labelled inflammatory monocytes in vivo to track their movement into aortas following Angiotensin-II infusion. We found an accumulation of labelled CD45+ cells in the aortas of ApoE-/- mice from day 3 to day 5 but not in Rgs1-/-ApoE-/- mice indicating RGS-1 as a regulator of macrophage retention in aortic aneurysms. These findings identify a novel role for RGS-1 in leukocyte function and vascular inflammation and identifies RGS-1 as a potential target for the treatment of cardiovascular disease.

138 Catheter ablation of atrial fibrillation whilst taking therapeutic warfarin: a british experience: Abstract 138 Table 1

Introduction Peri-procedural anticoagulation of patients undergoing catheter ablation of atrial fibrillation (AF) reduces thromboembolism risk but with potential haemorrhagic complications. In the UK, warfarin is generally discontinued pre-procedure and low molecular weight heparin (LMWH) used to bridge the sub-therapeutic period. In some institutions around the world ablation is performed with a therapeutic International Normalised Ratio (INR) and problems with LMWH administration are reduced, but the trans-septal puncture is usually performed under intracardiac echo (ICE) guidance. In the UK, cost prevents routine ICE use but we aimed to see if this could still be done safely within our system. Methods Fifty-six consecutive AF ablation patients were studied prospectively (warfarin group). A target INR of 2–3 was used for the procedure. A double trans-septal puncture technique was used. Unfractionated heparin was still used to maintain ACT between 300 and 350. Wide area circumferential ablation was performed along with any other ablation clinically indicated. If the INR was less than two at the time of ablation, LMWH was used afterwards until it was two or more. A group of 56 patients prior (bridging LMWH group) were compared. Endpoints were minor bleeding (haematomas), major bleeding (transfusion requiring), pericardial effusions, and stroke. Results In two patients the INR was >3 so the procedure was delayed. In 11 the INR was <2. In 43 the INR was 2–3 and they were all successfully ablated. The endpoint results are shown in the Abstract 138 table 1. The patient who had a pericardial effusion and tamponade during the procedure had ablation completed after reversal of the heparin and drainage, and warfarin was omitted for 24 h only. There were no pseudoaneurysms in the warfarin group and one in the bridging LMWH group. Abstract 138 Table 1 Minor bleeding Major bleeding Pericardial effusion Stroke Warfarin 6 0 1 0 Bridging LMWH 11 0 0 0 In the warfarin group LMWH had to be used for <24 h in the 9/11 patients with an INR <2. In the bridging LMWH group duration of treatment was 3±1 days. Conclusions AF ablation can be performed safely with therapeutic warfarin in the UK, with a low rate of procedure cancellation. Although the number of patients studied was relatively small, there was a lower incidence of minor bleeding with this approach than with bridging LMWH and a low rate of other complications in both groups. This strategy also reduces the need for LMWH after ablation which has both cost and patient comfort implications.

134 High incidence of acute circumflex artery injury following mitral isthmus ablation

Background Mitral isthmus ablation is technically challenging and often requires epicardial ablation in the coronary sinus (CS) for success. The circumflex artery lies in close proximity to the CS and mitral annulus and may potentially be injured during radiofrequency ablation. This study investigates the incidence of coronary artery injury following mitral isthmus ablation. Methods This is a single-centre, prospective study of 34 patients who underwent mitral isthmus ablation for atrial fibrillation (AF) and/or atrial tachycardia. Irrigation ablation catheters were used with the following settings: endocardial surface (max power: 40/50 W at the annular end; max temperature: 48°C; flow rate: 20 mls/min); CS (max power: 25/30 W; max temperature: 48°C; flow rate: 20 mls/min). Left coronary angiography was performed pre- and post-ablation. The angiograms were analysed off-line by an independent interventional cardiologist blinded to the ablation procedure. Quantitative Coronary Angiography (QCA) was performed for proximal, mid and distal circumflex artery (Cx), the obtuse marginal branch (OM) and the proximal left anterior descending artery (LAD). Results The mean age is 57±11 years, 73% are males, 81% have persistent AF, 35% have structural heart disease and mean left atrial size is 44 mm. Successful mitral isthmus block was achieved in 88% and CS ablation was performed in 68%. The mean ablation times were 686 s (total), 508 s (endocardial) and 170 s (CS). The mean diameters of the vessels are presented in Abstract 134 table 1. Abstract 134 Table 1 Pre-ablation diameter, cm (±SD) Post-ablation diameter, cm (±SD) p Value Proximal Cx 3.31±0.74 3.39±0.79 0.09 Mid Cx 2.42±0.85 2.16±0.88 0.04 Distal Cx 1.97±0.92 1.78±1.07 0.03 OM 2.49±0.71 2.55±0.66 3.3 Proximal LAD 3.66±0.80 3.60±0.72 0.08 Seven out of 34 patients (21%) had mild atheromatous disease at the start. Ten patients (29%) had new angiographic changes following ablation: three had distal occlusion of very small distal Cx; five had stenosis of mid Cx (40–85% stenosis); One had stenosis of mid Cx (60%) and proximal OM (40%); One had stenosis of proximal Cx (25%) and proximal OM (35%). The significant stenoses resolved with intracoronary Glycerin Trinitrate (GTN). Hence, they were likely to represent spasm. Small Cx (<1.5 cm) was associated with a higher risk of injury (50% vs 22%, p<0.05). All significant stenoses were associated with CS ablation. Two cases of small distal Cx occlusion occurred in patients who had endocardial ablation only (40 W). Patients who had Cx artery injury tended to have longer mean ablation times but this did not reach significance (total: 818 s vs 627 s; CS: 240 s vs 139 s; endocardial: 578 s vs 477 s). These patients did not report any clinical symptoms. Mild stenosis 1 prox Cx, 1 prox OM Moderate stenosis 4 mid Cx, 1 prox OM Severe stenosis 2 mid Cx Occlusion 3 distal Cx Conclusions Twenty nine per cent of patients had acute angiographic changes following mitral isthmus ablation. In many cases, they represent coronary spasm. These did not result in clinical symptoms. Further studies are needed to investigate the long term outcome of these patients.