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Various cell-based therapies are currently investigated in an attempt to tackle the high morbidity and mortality associated with heart failure. The need for these therapies to move towards the clinic is pressing. Therefore, preclinical large animal studies that use non-autologous cells are needed to evaluate their potential. However, non-autologous cells are highly immunogenic and trigger immune rejection responses resulting in potential loss of efficacy. To overcome this issue, adequate immunosuppressive regimens are of imminent importance but clear guidelines are currently lacking. In this review, we assess the immunological barriers regarding non-autologous cell transplantation and immune modulation with immunosuppressive drugs. In addition, we provide recommendations with respect to immunosuppressive regimens in preclinical cardiac cell-replacement studies.

Preclinical testing in animal model is a required stage of vascular device development. Among small animal models, rabbits provide vasculature with relative larger caliber for anastomotic implantation of vascular grafts as preclinical testing before conducting large animal studies. Rabbits have similar hemostatic mechanism with human and can accommodate vascular grafts with various diameters at different locations, and thus provide a valid model to assess small-diameter vascular grafts. This chapter will describe the procedures and materials required to conduct survival surgery in rabbit carotid artery models for implantation of small-diameter tubular grafts with an end-to-side and end-to-end anastomotic technique.

Stem cell therapy has an unparalleled potential to treat blood cancers, cardiovascular diseases and neurodegenerative conditions, among others. However, stem cell therapeutics must overcome multiple requirements before reaching clinical trials, including large animal safety and efficacy studies. In cardiovascular diseases swine models are the most widely adopted due to its great translational potential to humans. In this chapter, we will describe several protocols to induce iPSC dedifferentiation in swine fibroblasts, as well as conditioning treatments that may help in the reprogramming process.

Recent evidence has provided exciting proof of concepts for the use of pluripotent stem cell-derived cardiomyocytes (PSC-CMs) for cardiac repair; however, large animal studies, which better reflect human disease, are required for clinical application. Here, we describe how to create myocardial infarction in cynomolgus monkey followed by transplantation of PSC-CMs. This method ensures the establishment of a myocardial infarction model and enables reliable PSC-CM transplantation.

Myocardial infarction is a major clinical challenge for interventional, pharmacological, and potential molecular treatment of the ischemic insult. A large animal model with clinic-derived instrumentation allows for detailed imitation of interventional catheterization routines and application routes, whereas similar anatomy and heart proportions raise the possibility to precisely evaluate the efficacy of application modes, e.g., antegrade or retrograde intracoronary application of locally acting pharmaceutical agents, viruses, and cells. Here, we describe the techniques of left ventricular catheterization and induction of ischemia and reperfusion, as well as hemodynamic monitoring and regional application of therapeutic agents in pigs.

The development of new advances in understanding the role of neutrophils in inflammation requires effective procedures for isolating and purifying neutrophils. Methods for isolating human neutrophils are fairly standard, and some are covered in other chapters of this volume and previous editions. However, procedures for isolating neutrophils from nonhuman species used to model human diseases vary from those used in isolating human neutrophils and are not as well developed. Since neutrophils are highly reactive and sensitive to small perturbations, the methods of isolation are important to avoid isolation technique-induced alterations in cell function. We present methods here for reproducibly isolating highly purified neutrophils from large animal models (bovine, equine, ovine), small animal models (murine and rabbit), and nonhuman primates (cynomolgus macaques) and describe optimized details for obtaining the highest cell purity, yield, and viability.

This chapter describes various sheep models that faithfully and consistendy reproduce major manifestations of adverse postinfarction left ventricular remodeling following acute myocardial infarctions in patients. The models are produced by simply occluding coronary arteries that supply 21 to 24 percent of the left ventricular mass in different, specific locations. The advantages of these large animal models are illustrated by studies of early and late reperfusion of ischemic myocardium and studies restraining infarct or mitral annular expansion on the remodeling process.

Myocarditis in humans is often subclinical and difficult to recognize,1 limiting our full appreciation of the natural history and potential progression of the disease to chronic cardiomyopathy. Searching for a suitable animal model we studied the cardiovascular effects of infection of young pigs with a cardiotropic strain of encephalomyocarditis virus. This model was found to closely resemble viral myocarditis in humans in the acute as well as subacute and chronic phases. The pig was selected because of similarities in cardiac anatomy and physiology as well as the technical ease of making clinical observations, including noninvasive echocardiography and electrocardiography. This chapter presents an overview of our experience with this model establishing its usefulness for further studies of the natural history and pathophysiology of viral myocarditis

The Batista operation for dilated cardiomyopathy (DCM) has not, thus far, yielded satisfactory results. One of the explanations for disappointing results is the absence of a suitable animal model of dilated cardiomyopathy. Scientific research on volume reduction surgery needs to be carried out in an appropriate experimental model. This chapter reviews the few studies of volume reduction surgery using animal models of DCM published thus far, including our studies.

Dogs develop spontaneous brain tumors as in humans and serve as a large animal model for evaluation of BNCT for treatment of brain tumor. In this study, 13 dogs with spontaneous brain tumors were treated surgery (SX) followed by BNCT and the median survival of was determined. The median survival was also determined for a group of dogs with spontaneous brain tumors previously treated with BNCT alone (n = 20). All dogs had histologic confirmation of tumor type either at the time of surgery or at necropsy. Of the dogs treated surgically, BNCT followed 3 to 6 weeks after surgery. Dogs of each group had various tumor types, however, the majority were meningiomas (13/20 BNCT; 9/13 SX + BNCT). Of the dogs in the surgery group, all had incomplete resections of the tumor primarily due to tumor location or infiltration of surrounding brain. In both groups, BSH or p-BPA were used for the boron agent. Peak brain dose was 10 to 12.5 Gy- equivalent.

Transgenic murine models have been employed with increasing frequency over the past several years for the elucidation of mechanisms of normal and abnormal cardiovascular function. While these models have been used extensively for biochemical and cellular physiological studies, their use in studies of in vivo cardiovascular physiology has until recently been limited by technical considerations. Recent advances in surgical and catheterization techniques, imaging modalities and other technologies have allowed for detailed in vivo cardiovascular studies in mice. These developments recall the technical advances made several decades ago that allowed for detailed study of cardiovascular physiology in large animals. These investigations in large animal models demonstrated not only the importance of in vivo studies for understanding complex physiological relationships, but also the profound effects of anesthesia on measures of cardiovascular function 1-9. Ideally, studies of transgenic murine cardiovascular physiology would similarly be carried out in conscious, chronically instrumented animals. Limitations in current technology, however, have confined conscious studies to only a few techniques. This chapter reviews the currently available methods for the measurement of cardiovascular parameters in conscious mice, including ranges for normal values, and discusses the effects of anesthesia on cardiovascular function in the mouse which make the need for further progress on conscious measures so compelling.

Boron neutron capture therapy (BNCT) treatments utilizing epithermal neutron beams and available boron compounds result in a mixed irradiation field of low and high linear energy components (LET). Previous studies attempting to measure the repair of the low LET component, comparing a multifraction study to single fraction study failed to identify sufficient repair in normal tissue tolerance studies of the dog brain utilizing borocaptate sodium (BSH) and epithermal neutron irradiation.1 Many current patients in clinical trials relapse approximately 6 months after their initial BNCT treatment. Many have considered the possibility of retreatment due to the theoretical dose localization in BNCT. A six-month interval would not be sufficient to allow retreatment of a full course of conventional irradiation. Therefore, a study was undertaken to give purpose-bred laboratory dogs an initial BNCT to near tolerance levels followed in six-month by a second identical dose.

Purpose While most patients who undergo bladder exstrophy closure develop sufficient bladder capacity, others do not. To discover the factors which lead to these differences, we compare bladder biopsies from normal neonatal sheep to those of sheep which have undergone creation of bladder exstrophy. We then compared these results with those of a previous study in which we compared bladder biopsies from human neonatal bladders to newborn exstrophy bladders.

Originally, the term ischemic preconditioning described the observation that four 5 minute episodes of coronary artery occlusion in dogs followed by prolonged ischemia (40 minutes) and reperfusion resulted in a marked reduction in the rate of ATP depletion and limitation of myocardial infarction (1). Sustained ischemia alone produces about 30% myocardial infarction of the area at risk. These observations have been reproduced in small and large animal models of coronary artery occlusion and reperfusion whereby a single episode (or multiple episodes) of ischemia and reperfusion protected the myocardium against infarction, but failed to augment recovery of ventricular function in the same model.

The goal of encapsulated cell therapy research is to develop implants containing living xenogeneic cells to treat serious and disabling human conditions. The concept is straightforward: cells or small clusters of tissue are surrounded by a selective membrane barrier which admits oxygen and required metabolites, releases bioactive cell secretions but restricts the transport of the larger cytotoxic agents of the body’s immune defense system. Use of a selective membrane both eliminates the need for chronic immuno-suppression in the host and allows cells to be obtained from non-human sources, thus avoiding the cell-sourcing constraints which have limited the clinical application of generally successful investigative trials of unencapsulated cell transplantation for chronic pain1, Parkinson’s disease2, and type I diabetes.3–5 Cross-species immunoisolated cell therapy has been validated in small and large animal models of chronic pain6,7, Parkinson’s disease8,9, and Type 1 diabetes10–12, and is under active investigation by several groups in animal models of Huntington’s13, ALS14 and Alzheimer’s15–19. In addition, the first encapsulated therapy using xenografts in humans has been performed in chronic nain20 and ALS21.

Animal models are useful to study the relative effects of BNCT on normal tissues and tumors. True efficacy studies of the modality on glioblastoma or other human malignancies requires human clinical trials. The use of large animals, primarily dogs, to study the effects of BNCT has been and continues to be of major interest. The use of large animals permits the study of normal tissue tolerance of the tissues of the head at an acceptable total body dose. These studies have been extended from normal dogs to dogs with induced and spontaneous tumors. While induced tumors in rodents are often used to study tumor response, the total body dose accompanying BNCT severely limits their use for normal tissue tolerance studies, especially with the epithermal-neutron beams.

The INEL BNCT program has been involved in normal tissue dose tolerance studies using dogs as the large animal model1. In order to accurately predict when to begin an irradiation and then determine the length of the radiation required to achieve the desired radiological dose at a specified mean boron concentration, a rapid technique for the determination of boron is required. It has previously been shown that after administration and distribution of Na2B12H11SH (BSH), the elimination phase has a T½ of ~ 300 minutes2 and one should be able to roughly predict blood B concentrations at specified times during the elimination. However, the physiology can be expected to vary somewhat from animal to animal and the blood B concentrations are expected to vary as well. These variations mean that target B concentrations can easily be missed.

Single dose RBEs for the radiations encountered in BNCT have been determined1 and can provide useful guidance for the tolerance to be expected. The difficulty of microdosimetric determination of dose2, the rapidly varying mix of constituent high and low LET radiations with depth, the possible synergistic interaction between high and low LET radiations3, and the strong dependence of biological efficacy on the distribution of boron within the cell are all factors limiting the usefulness of RBEs. Large animal studies arenecessary to determine late effect tolerance. To design, interpret, compare, and apply these studies to clinical use, it has been proposed to divide factors influencing “RBE” into two groups4; 1) estimated tolerance dose (ETD), which can be inferred from other experience reasonably well; and 2) compound factor (CF), which is much more variable as a function of microdosimetry and boron compound.

Spontaneous tumors, in comparison to transplantable tumors, in animals offer several advantages for investigating new therapeutic modalities prior to human clinical trials. The main advantages are “typical” tumor vasculature, normal anatomical sites, slow growth rates and long cell-cycle times, and a nonimmunogenic nature.

Boron Neutron Capture Therapy (BNCT) offers a potential form of therapy for highly malignant brain tumours such as glioblastoma multiforme1,2. The High Flux Reactor of the Commission of the European Communities in Petten, The Netherlands, will serve as the neutron source for this treatment3. Before first clinical trials can be performed on patients, the tolerance of healthy tissue exposed to the radiation fleld has to be determined. Due to the characteristics of epithermal neutrons a large animal model has to be used for this task. Since the canine model has proved to be an excellent tool to answer this issue4, it has been chosen for the healthy tissue tolerance studies at Petten. The paper here describes the experimental set up and the initial results from the first two canine experiments which were irradiated in July 1991.

An epithermal neutron beam is needed to treat relatively deep seated tumors. The scattering characteristics of neutrons in this energy range dictate that in vivo experiments be conducted in a large animal to prevent unacceptable total body irradiation. The canine species has proven an excellent model to evaluate the various problems of boron neutron capture utilizing an epithermal neutron beam. This paper discusses three major components of our study: I) the pharmacokinetics of borocaptate sodium (NA2B12H11SH or BSH) in dogs with spontaneously occurring brain tumors, II) the radiation tolerance of normal tissues in the dog using an epithermal beam alone and in combination with borocaptate sodium, and III) initial treatment of dogs with spontaneously occurring brain tumors utilizing borocaptate sodium and an epithermal neutron beam.

The Power Burst Facility/Boron Neutron Capture Therapy (PBF/BNCT) Program has been funded since 1988 to evaluate brain treatment using Na2B12H11SH (borocaptate sodium or BSH) and epithermal neutrons. The PBF/BNCT Program pursues this goal as a comprehensive, multidisciplinary, multiorganizational endeavor applying modern program management techniques. The initial focus was to: (1) establish a representative large animal model and (2) develop the generic analytical and measurement capabilities required to control treatment repeatability and determine critical treatment parameters independent of tumor type and body location. This paper will identify the PBF/BNCT Program elements and summarize the status of some of the developed capabilities.

The selection of spontaneously hypercholesterolemic rats in a large animal breeding unit and the maintenance of this characteristic over numerous successive generations were performed by Zucker1, Kolestsky2, Imai et al.3 and Muller4. We were particularly interested by this last strain: by as early as 1979, RICO rats (rats with increased cholesterol) were 100% hypercholesterolemic after the fifteenth generation of consanguineous breeding. These animals are normotriglyceridemic and non-obese, in contrast with “Zucker” rats. They were isolated from heterozygous normocholesterolemic rats from this animal breeding unit (CIBA-GEIGY) which we shall refer to as SW. A single study, published in 1979, reported some of the characteristics of the cholesterol metabolism of these animals4. Since 1983, we have therefore conducted an exhaustive study of the characteristics of the cholesterol metabolism of this strain of rats.

It is now well established that extracorporeal shock wave lithotripsy (ESWL†) produces significant acute renal side effects. There is strong evidence that chronic effects may also accompany ESWL therapy. The clinical and experimental data that document tissue injury from ESWL are compelling but do not adequately address the factors that are responsible for adverse acute side effects or the conditions that may lead to serious long-term health problems. In order to resolve these issues, it is necessary to determine the precise mechanism by which cells are damaged by ESWL. To accomplish this goal, animal experimentation is required so that the time course and severity of acute and chronic alterations can be followed. Animal models allow whole organ analysis of changes in both structural and functional features. These studies must be designed to give the best approximation of the treatment conditions that are applied to patients. To do this requires an animal model with a kidney that has similar structure and function to that of man. The kidneys of most small animals and many large animals, including the dog, do not mimic human renal anatomy and physiology. The laboratory pig appears to be the best large animal subject for ESWL research. The pig offers a model in which kidney size, renal/urinary anatomy, and characteristics of renal function closely resemble the human. Moreover, the methodology for correlative structure and function analysis in the pig is well established. Preliminary data on the mini-pig suggest that it is an excellent model to study the bioeffects of ESWL in that acute traumatic injury, similar to that reported for patients, is induced at therapeutic levels of shock wave delivery.

Studying lipid metabolism in large animals has sense as the results either can be of help in elucidating the etiology of metabolic disorders that affect economic loss in large animal husbandry or can demonstrate how large animals can be used as models in fundamental research in the field of lipid metabolism.