Our main aims are to demonstrate novel mechanisms of initiation of inflammation, and to define links between inflammation and formation of blood clots in arteries. We have shown that both CRP (C-reactive protein) and SAA (Serum Amyloid A), which are inflammation markers predictive of death or heart attack in both normal populations and those with CAD, can stimulate blood monocytes to produce tissue factor, the most powerful initiator of clotting. There is a non-specific upregulation of monocyte responsiveness to both CRP and SAA in patients with CAD, so the higher concentrations of these proteins seen in patients, who have events, indicates that both CRP and SAA are not just markers of inflammation but may play a role in blood clotting which precipitates adverse events.
We have further investigated the mechanism whereby SAA is pro-inflammatory and found very early and potent up-regulation of many pro-inflammatory molecules by SAA and monocytes are the principle source. This is even greater in monocyte-derived macrophages present in atherosclerotic coronary artery vessel walls, and appears to be mediated by the nuclear switch NF-κB, and may be an important amplifier of both inflammation and thrombosis in acute coronary syndromes (including heart attack) and sudden coronary death. We have also shown that SAA is released into the coronary circulation in patients with CAD but not in controls. Our recent investigations have focused on the role of SAA in initiating vascular dysfunction. We have shown that exposure of blood vessel lining cells (endothelial cells) to pathological SAA induces endothelial dysfunction by affecting Ca2+ signal transduction pathways, stimulating the production of reactive oxygen species (e.g. O2-•), up-regulating the expression of pro-inflammatory genes and genes involved in the coagulation cascade, and decreasing cGMP which is involved in the synthesis of the blood vessel vasodilator nitric oxide. Presently, we are using an apoE deficient mouse model to assess the pro-atherogenic activity of SAA.
We have also demonstrated an important protective effect of HDL, part of the ‘good cholesterol’ that has other protective actions against atherosclerosis. We are beginning to study the protective effect of HDL on SAA-induced endothelial abnormalities, under high-glucose conditions to simulate diabetes. The group continues its longstanding collaboration with Prof Carolyn Geczy (UNSW) in these projects, and in other projects to ascertain the role of S100 proteins in atherosclerosis.
Platelet and Leukocyte Activation and Interactions within CAD
Our aims are to examine the extent and mechanisms of platelet and leukocyte activation occurring due to the presence of CAD both systemically and in the intracoronary tree. We also aim to examine interactions occurring between the platelets and leukocytes that may be contributing to CAD. By observing the response of the platelets and leukocytes in the intracoronary tree we have an in vivo model of activation demonstrating the effect of plaque volume on platelet and leukocyte activation.
Recent work involving a novel platelet marker – extracellular matrix metalloproteinase inducer (EMMPRIN or CD147) – shows that despite the lack of response of traditional platelet activation markers, p-selectin and PAC-1 (conformational change of GPIIb/IIIa), EMMPRIN on the platelet is upregulated systemically. This has been found to be due to the inflammatory state and age of the patient rather than CAD specifically. EMMPRIN expression has been found to be upregulated on monocytes in patients with acute myocardial infarction (Schmidt et al 2006 Circulation), our results concur with this finding in the systemic circulation; however, we observed upregulation due to CAD on both the monocyte and granulocyte populations (Pennings et al 2010 JTH). Upregulation of platelet activation markers across coronary stenoses has been observed. Current work will include continued examination of what occurs within the coronary tree with respect to platelet and leukocyte mechanisms of activation and studies of the regulation in GPVI in collaboration with Drs Elizabeth Gardiner and Robert Andrews (MonashUniversity). Dr Andy Yong will be extending his work on intracoronary platelet activation by relating this to the functional severity of coronary stenoses (Eur Heart J 2010), and computationally modeled shear stress in coronary arteries in collaboration with Professor M Behnia from the University of Sydney.
Dr Witting and his staff and students interests include ischemic injury to the heart (heart attack) and brain (Stroke) and effects of myoglobinurea on kidney function.The main research thrust involves design and testing of potential (synthetic antioxidant) inhibitors of damage to myocardial, renal and neuronal tissues in the setting of acute insult to these organs.
Current work has seen the study in neuronal protection evolve into gene targeting of a novel haem protein called neuroglobin.This protein is expressed in the brain during acute ischaemic insult and is believed to play a role in protecting the brain during stroke.Studies using a neuroglobin-traansfected neuronal cell line have recently been published in the highest ranking journal in the field of oxidative stress (Antao, et al. Antioxidant and Redox Sig. 2010 in press).
Importantly, the lab continues to support young undergraduate and postgraduate researchers with Miss Hong Duong graduating with a PhD from the University of Sydney and Mr Ben Rayner submitting his PhD for assessment in the last 12 months.
Our focus is gene expression and modification of vascular and myocardial injury. Specifically, the contexts of neointimal formation following vein graft and native coronary injury, and myocardial ischemia-reperfusion injury are being examined. Using a number of in vitro and in vivo animal models we are investigating the use of DNAzyme and other novel gene-targeting approaches to inhibit injury responses.
We are particularly interested in diabetic heart disease and are currently investigating how diabetic heart muscle responds to ischemia-reperfusion injury compared with non-diabetic heart muscle. We have established a novel model of heart attack in diabetes, which is allowing us to examine both early and late responses to injury, both in recent onset and in established diabetes.
Egr-1-targeting DNAzymes are sequence-specific enzymes that cleave Egr-1 mRNA, thus preventing expression. Using in vitro models of diabetes and myocardial ischemia, and our own animal model of myocardial ischemia in diabetes, we are examining the effects of DNAzyme delivery on Egr-1 expression, molecular pathways of ischemic injury, and infarct size. This work has the potential to provide novel “diabetes specific” therapies for heart attack.
Coronary bypass grafts (saphenous vein grafts or SVGs) have a tendency to develop narrowings in the years following coronary surgery, leading to further symptoms of heart attack. The nature of these narrowings remains poorly understood, but we believe it is distinct from the atheroma that develops in native coronary vessels. We are trying to determine the nature of atheroma in SVGs and define the similarities and differences from atheroma in native coronaries. This will help us understand, and possibly tackle, the atherosclerosis process in these conduits in a better way.
Our focus is on the utility of novel measures of haemostatic (blood clotting) function in characterising therapeutic responses to anti-platelet drugs in patients with coronary disease. Patients undergoing invasive cardiac procedures like stenting receiving anti-platelet therapies have their overall haemostatic function determined by assays include the Overall Haemostatic Potential (OHP) Assay and calibrated automated thrombogram (CAT). These have been developed in collaboration with the Haematology Departments at ConcordHospital and RoyalNorthShoreHospitals. The impact of different anti-platelet agents and combinations of these agents on these assays are being evaluated and compared against conventional functional studies of platelet function.
Echocardiographic Evaluation of Cardiac Dysfunction
Patients with clots in the lungs- pulmonary embolism- or high blood pressure in the lungs- pulmonary hypertension commonly have problems with the right side of the heart. We are investigating the acute and long term effects of these diseases on the heart, and whether we can detect improvements more accurately using new ultrasound techniques. Dr Wei Zhao is a highly trained echocardiographer who is applying new speckle-tracking ultrasound techniques to the measurement of the contraction of the right ventricle. She has developed a new measurement which detects recovery of cardiac function before conventional imaging studies can detect improvement. Dr Vincent Chow, an Echocardiography fellow in our Department and APA Masters Student at the University of Sydney Dr Austin Ng and Dr Tommy Chung, are evaluating the incidence of late pulmonary hypertension in patients who have previous clots in the lungs. There are very few studies systematically recording the long term effects of pulmonary embolism on the heart and his work will establish important baseline data for the elderly Australian population.
In collaboration with Professor Handelsman, Dr Zhao will be investigating the effects of androgen receptor expression and androgen deficiency on myocardial function in mice. She has established high frequency echocardiography of mice at the ARI and will use this technique to assess the heart response to injury and to treatments that may promote its recovery.
Infective endocarditis is a life-threatening condition, the incidence of which has remained unchanged over the last 20 years. As part of his PhD thesis at the University of Sydney, Dr Ray Sy has completed a series of studies investigating risk stratification, population incidence and the effect of timing on prognostic benefit in patients with infective endocarditis in NSW. Ray found that thrombocytopenia is an important predictor of outcome, and that it remains a predictor even after two weeks of treatment whereas some other factors lose their predictive power over time (J Am Coll Cardiol 2009, European Heart J 2009). Ray found in a linkage study covering the state of NSW that the mortality of infective endocarditis remains alarmingly high and that this condition is affecting an increasingly elederly population (European Heart J 2010). He also found that the prognostic benefits assigned to treatment must take into account the timing of the treatment if they are not to introduce systematic biases (Circ Cardiovasc Outcomes 2010).
Choice of Health Options In Preventing Cardiac Events (CHOICE Study)
Patients who survive a heart attack have a high risk of death or recurrent heart attack, which can be reduced by effective programs of secondary prevention like cardiac rehabilitation. Unfortunately, 70% of survivors do not access formal cardiac rehabilitation, and yet their risk factor levels are much higher than those who do attend. We have developed a simple program called CHOICE (Choice of Health Options In preventing Cardiac Events), which allows patients to choose which risk factor(s) they will lower and how they will lower it (them). The program was extremely effective and results persisted for 1 year, and our follow up of the original cohort indicated the effect was long lasting and little diminished at 4 years. Current studies will extend the program much more widely to show it can be scaled up, and examine whether a brief intervention is sufficient to produce long term changes over 3 years, or whether a longer program will be better in maintaining the results.
In our current study across 4 hospitals we have enrolled almost 300 patients who have survived a coronary event but elected not to participate in traditional cardiac rehabilitation. Following these patients will tell us whether a brief intervention will have long lasting effects on multiple risk factors.
