Animal Models of Heart Disease

Appropriate animal models of human heart disease are an attractive alternative for proteomic investigations as human diseased tissue samples can often be compromised by factors such as the disease stage, tissue heterogeneity, genetic variability, and the patient s medical history/therapy. Avoiding any of the above complications when working with human samples can prove to be extremely difficult 

Identifying altered canine and bovine proteins has proved to be particularly challenging since these species are poorly represented in current genomic databases. As a result of this, new bioinformatic tools (Multildent, http //expasy.org) have had to be developed to facilitate cross-species protein identification (Wilkins et al., 1998). The most significant change observed for bovine DCM was a sevenfold increase in the enzyme ubiquitin carboxyl-terminal hydrolase (UCH) (Weekes 

More recently we have investigated whether the ubiquitin-proteosome system is perturbed in the heart of human DCM patients (Weekes et al, 2003). As in bovine DCM, expression of the enzyme UCH was elevated more than 8-fold at the protein level and elevated more than 5-fold at the mRNA level in human DCM. Moreover, this increased expression of UCH was shown by immuno-cytochemistry to be associated with the myocytes, which do not exhibit detectable staining in control hearts. Overall protein ubiquitination was increased 5-fold in DCM relative to control hearts. Using a selective affinity purification method we were able to demonstrate enhanced ubiquitination of a number of distinct proteins in DCM hearts. We have identified a number of these proteins by mass spectrometry. Interestingly many of these proteins were the same proteins previously found to be present at reduced abundance in DCM hearts (Corbett et al, 1998). This new evidence strengthens our hypothesis that inappropriate ubiquitin conjugation leads to proteolysis and depletion of certain proteins in the DCM heart and may contribute to loss of normal cellular function in the diseased heart. 

---

There are four gel protein databases of cardiac proteins, established by three independent groups, that can be accessed via the World Wide Web (Table 16.1). These databases facilitate proteomic research into heart diseases containing information on several hundred cardiac proteins that have been identified by protein chemical methods. They all conform to the rules for federated 2-D protein databases (Appel et al., 1996). In addition, 2-D protein databases for other mammals, such as the mouse, rat (li et al., 1999), dog (Dimn et al., 1997), pig and cow, are also under construction to support work on animal models of heart disease and heart failure. 

Pharmacologists are often able to develop tissue and whole animal models of human disease. In some instances, studies on isolated tissues, such as blood vessels, heart muscle or brain slices, will... 

The disturbance of balance between superoxide and nitric oxide occurs in a variety of common disease states. For example, altered endothelium-dependent vascular relaxation due to a decrease in NO formation has been shown in animal models of hypertension, diabetes, cigarette smoking, and heart failure [21]. Miller et al. [22] suggested that a chronic animal model atherosclerosis closely resembles the severity of atherosclerosis in patients. On the whole, the results obtained in humans, for example, in hypertensive patients [23] correspond well to animal experiments. It is important that endothelium-dependent vascular relaxation in patients may be improved by ascorbic acid probably through the reaction with superoxide. 

The transdifferentiation of HSCs into a mature hematopoietic fate (e.g., endothelium) in the heart is less controversial [148]. In animal models of stem cell therapy in ischemic heart disease, the evidence points toward increased neovascularization (with reduced myocardial ischemia) and consequent improvement in cardiac function [149-151]. Bone marrow stem cells may directly contribute to an increase in contractility or, more likely, may passively limit infarct expansion and remodeling. Unfortunately, the limitations of the present animal models leave this question unanswered. 

Of these, omapatrilat is at the most advanced stage of clinical development. It lowers blood pressure in animal models of hypertension as well as in hypertensive patients and improves cardiac function in patients with heart failure. Unfortunately, omapatrilat causes a significant incidence of angioedema in addition to cough and dizziness. Nevertheless, combined inhibition of neutral endopeptidase NEP 24.11 and ACE with this new class of drugs may be a promising approach to treat cardiovascular disease and further clinical trials are underway. 

Tacrolimus causes acute reversible renal dysfunction in renal [661-663,667], hver [290,664-666,679,680], heart [681-683] and pulmonary [684, 685] transplant recipients and in patients with immunologically mediated diseases [686]. Tacrolimus-induced GFR and RBF decrease is associated with an important increase in renal vascular resistance, both in humans and rodents [63,679,687-692]. Calcium channel blockers improved renal function in TAC-treated liver transplant recipients [693] and in animal models of TAC nephrotoxicity [689,694-6%]. Tacrohmus acute nephrotoxicity, similar to CSA, shows normal renal histology or non-specific changes such as isometric cytoplasmic vacuolation in tubular epithelial cells, microcalcification, giant mitochondria and lysosomes, and necrosis and early hyahnosis of individual smooth muscle cells in the afferent arterioles, which revert with drug reduction or discontinuation [697-699]. 

A possible novel therapeutic strategy for heart failure following myocardial infarction may be to increase the number of functional myocytes within the diseased area by the implantation of exogenous myogenic cells. Early studies used neonatal rat cardiomyocytes for transplantation, as these cells have cardiac phenotype and still retain some proliferation capacity [2-4]. Fetal cardiomyocyte cell grafts showed the formation of cell-to-cell contacts, complete with gap junction proteins [4]. Moreover, cultured human fetal cardiomyocytes were shown to survive, and fetal rat cardiomyocytes were shown to be present in the infarcted rats hearts for up to 6 months after transplantation [5]. Further studies in animal models of myocardial infarction showed that grafting of cardiomyocytes from fetal and neonatal sources was asso-... 

In this chapter we will review proteomic investigations of cardiac proteins and focus on their application to the study of heart disease in the human and in animal models of cardiac dysfunction. The majority of these studies of the cardiac proteome have involved protein separation, visualisation and quantitation using the traditional 2-DE approach combined with protein identification by mass spectrometry. These essential technologies will be briefly described. However, there is increasing interest in using alternative gel-free techniques based on mass spectrometry or protein arrays for high throughput proteomics. These alternative approaches will be introduced, but further details can be found in Chapter 2 of this volume by Michel Faupel. 

To study CAVD in a controlled system, animal models of valvular dysfunction are utilized in the laboratory setting. The same principles used to diagnose AS in humans may be applied in the study of the progression of this disease in a wide array of animal models. Valvular cells extracted from bovine sources have been widely used due to the ease of extraction and because the structure of the bovine aortic valve is similar to the human aortic valve [65-67]. Porcine models have been utilized to a large extent for studies of the aortic valve, since the heart and cardiovascular system are nearer in size to the human equivalents [68-70]. A relevant porcine model for aortic valve calcification has been developed by Balachandran et al [71] in which the effects of cyclic stretch were studied using porcine aortic valve leaflets in a unidirectional stretch bioreactor. Calcification was significantly... 

Animal models of disease that might be considered for use in SP shonld be dictated by whether or not they improve predictive value of adverse reactions in patients, bnt of course they may also be valuable/preferred in studies on mechanism of action and efficacy. Scientists may be guided by selection of specific models that are well known to manifest increased sensitivity for adverse reactions in the clinic (e.g., obesity, senility, heart failure, hypertrophy, diabetes). Clinicians already know, and consider in design of clinical trials, many features of subjects (e.g., age, geographical location, sex, somatotype, strain) that inflnence the outcome of smdies. It is likely that diversity of subjects in preclinical smdies is as important as diversity of snbjects in clinical studies. My suspicion is that smdies conducted on one mouse, one rat, one rabbit, one gninea pig, one dog, and one monkey per each of four groups (vehicle, low dose, mid... 

Animal models of diseases, as well as studies in patients with congestive heart failure, liver cirrhosis or renal insufficiency, have shown that prostanoids contribute to normal renal function by regulating vascular tone and normal blood flow (Harris, 2002 Parente and Perretti, 2003). PGE is involved in the regulation of sodium reabsorption, and its inhibition results in sodium retention that can manifest clinically in a variety of ways, such as peripheral edema, increased blood pressure, weight gain, and occasionally deterioration or heart failure. PGI increases potassium secretion and, since it has vasodilatory properties, it increases renal blood flow and glomerular filtration rate (Harris, 2002). 

---