Transplantation of the heart

Transplantation of the Heart JOHN S. VASKO, M.D., Columbus,

On December 3, 1967 the first human heart transplant was performed by Barnard and his associates [1] and the announcement of this medical milestone electrified the world. This initial clinical trial was immediately followed by several others at various medical centers throughout the world and at the time of this writing (September 7, 1968) forty-one human cardiac homotransplants and two heterotransplants have been performed and it has been predicted that more than one hundred will have been carried out by the end of the year. It is not surprising that these pioneering efforts have appeared almost simultaneously when one considers the evolutionary development of experimental technics which have resulted in consistently successful cardiac transplants in animals and the improved methods of immunosuppression and histocompatibility typing which have been largely a product of the vast clinical experiences with renal transplantation. The surgical technic utilized in the clinical trials was developed in 1960 by Lower and Shumway [Z]. In 1964 Hardy and associates [3] contemplated homotransplantation of the human heart in a potential recipient dying of severe coronary atherosclerosis. For lack of a suitable human donor and because of a patient whose condition was rapidly deteriorating, they utilized the heart of a chimpanzee as a heterotransplant. The heart resumed a forceful beat but was unable to effectively accommodate the circulation. Although the technical aspects of cardiac transplantation have been well developed, long-term survival in animals has not been impressive. It should be emphasized, however, that experiences with immunosuppression in the laboratory animal have yielded considerably less effective results than those obtained in clinical renal transplantation.

From the Department of Surgery, Division of Thoracic and Cardiovascular Surgery, The Ohio State University, College of Medicine, Columbus, Ohio 43210. This work was supported in part by U. S. Public Health Service Grant No. Hi11057-01. 344

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The initial human heart transplants were associated with a high mortality and morbidity which shrouded the procedure in controversy. Nevertheless, the operative mortality, complication rate, and early survival in the last twenty-five patients have already remarkably improved and the future of cardiac transplantation appears secure. Many clinical, moral, ethical, legal, theologic, and psychologic questions have been raised, which have motivated several organizational efforts aimed at solving important issues, such as the definition of lega death and the selection of donors and recipients. In view of these developments it seems likely that human cardiac transplantation will survive and grow, possibly more rapidly than has renal transplantation, thus providing a great stimulus to research in organ storage, transplantation immunology, assisted circulation, and the development of an implantable artificial heart. In recognition of the new base line provided by the recent clinical trials it seems pertinent to review comprehensively the investigative efforts that have made the concept of clinical cardiac transplantation possible. Heterotopic Transplantation The first attempt at cardiac transplantation is credited to Carrel and Guthrie [&I, who in 1905 homotransplanted dog hearts by re-establishing their circulation by means of anastomosis to the carotid and jugular vessels in the neck of recipients. Little detail was provided in their report and no additional attempts were recorded until 1933 when Mann and associates [5] reported a similar technic (Fig. 1) and found that the transplanted hearts beat vigorously for one to eight days and then ceased to function. Histologically these hearts were markedly infiltrated with lymphocytes, and polymorphonuclear large mononuclears, leukocytes. In 1948 Sinitsyn [6] reported essentially identical findings, utilizing a similar preparation. (Fig. 1.) Marcus, Wong, and The American

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Fig. 1. Composite summarv of the most commonly topic cardiac. transplants.

Luisada [7] in 1951 employed cross circulation to maintain the donor heart during transplantation to the neck of recipient animals and found that protection from temporary ischemia did not alter survival of the donor heart. In [8] trans1953 Wesolowski and Fennessey planted puppy hearts into the necks of adult dogs and observed that the homografts functioned similarly to skin homografts reported by Medawar [9], with an initial “take period” characterized by a forceful beat and electrical activity, followed by apparent death of the graft with microscopic evidence of necrosis and marked cellular infiltration. Marcus, Wong, and Luisada [lo] in 1953 homotransplanted dog hearts and heart-lung complexes into various sites in recipient animals and attempted unsuccessfully to delay rejection by crossmatching the bloods of the donor and recipient. In 1953 Dowie [11] transplanted puppy hearts into the necks of adult dogs and found less cellular infiltration histologically, suggesting that humeral factors may be important in the destruction of the transplant. Luisada and Marcus [12] in 1954 employed several variations of the cervical cardiac homotransplant and compared the survival of working and nonworking grafts. The results were equivocal, but suggested that the working heart survived slightly longer. They also observed that the transplanted heart had a remarkably stable rate and was hypersensitive to various pharmaVol.117.March 1969

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cologic agents. In addition they attempted unsuccessfully to alter the rejection phenomenon with adrenocorticosteroids and cross matching of the blood of both donor and recipient. In 1957 Sayegh and Creech [A?] transplanted puppy and fetal hearts into the necks of adult dogs and found them to be equally antigenic. Lee and Webb [14] in 1969 found that coronary blood flow averaged 128 cc/min./l00 gm. and that left ventricular weight and myocardial oxygen consumption averaged 6.4 cc./min. in cervical homografts. Metabolic studies showed myocardial utilization of glucose, lactate, and pyruvate, although an early negative balance of pyruvate was frequent. In 1960 Reemtsma, Delgado, and Creech [IS] studied cervical puppy heart homografts in adult recipients and found wide variations in oxygen consumption and relatively stable myocardial blood flow and lactate production. Demikov [16] in 1961 described twenty-four different technics of heterotopic cardiac transplantation including an ingenious technic of assisting the circulation with an intrathoracic cardiac homograft. This method was further studied by Reemtsma [17] in 1964 and later modified by McGough, Brewer, and Reemtsma [18] in 1966. DePasquale et al. [19] in 1965 found electrocardiographic alterations in the hearts of hosts with orthotopic cardiac homografts which did not occur when immunosuppressive drugs were used or in response to renal homografts, sug345

Vasko gesting an organ-specific reaction in which the recipient’s heart was the target organ. In 1962 Reemtsma and co-workers [ZO] transplanted puppy hearts into the necks of adult dogs and employed a folic acid antagonist (amethopterin) with limited success in an attempt to prolong homograft survival. Chiba and associates [21] in 1962 extensively studied the metabolism and histopathology or cervical canine heart homografts and found that they released pyruvate, lactate, malic dehydrogenase, and adolase, although glucose extraction usually remained positive. As rejection accelerated, pyruvate and lactate release was more pronounced and glucose concentrations increased in coronary venous blood. Usually the respiratory quotient and the glucose-oxygen extraction ratio were elevated and the authors concluded that these changes were the result of conversion of carbohydrates to fat. They suggested that metabolic blocks in the transplanted heart are produced by a diminution of intracellular enzymes and co-enzymes due to increased cellular permeability. The redox potential across the transplanted heart was positive, indicating no significant hypoxia and glycolysis advanced in the presence of oxygen. Histopathologic observations demonstrated early perivascular lymphoeytic infiltration and vascular endothelial swelling followed by increased cellular infiltration evolving into a granulomatous myocarditis and ultimate tissue necrosis. Histochemical studies showed a significant reduction of myocardial malic dehydrogenase which was concurrent with increases of this enzyme in the serum. They concluded that cardiac tissue is extremely antigenie and produces an immune response rapidly when implanted as a homograft. The great majority of studies concerning heterotopic cardiac transplants were carried out prior to the development of the pump oxygenator and hypothermic technics which made orthotopic cardiac transplantation possible. It is significant that the information obtained from investigations of heterotopic grafts provided a firm foundation for our understanding of cardiac transplantation and that subsequent investigations of orthotopic grafts have largely confirmed these previous observations. Certainly the heterotopic cardiac graft possesses many advantages as an experimental model and it is likely that it will continue to be employed in transplantation studies. As a re346

sult of these studies, the stage was set for the development of experimental orthotopic cardiac transplantation and associated studies of graft function.

Orthotopic Transplantation Autotransplantation. Before the feasibility of total cardiac transplantation could be ascertained, it was apparent that orthotopic autotransplantation was necessary in order to completely separate immunologic influences from other factors associated with the technics of transplantation and to determine the functional characteristics of the totally denervated heart. In 1957 Webb and Howard [22] autotransplanted the heart and lungs in dogs with the aid of cardiopulmonary bypass and obtained survival as long as four hours. They concluded that heart autotransplants were able to maintain the circulation satisfactorily. Cass and Brock [23] in 1959 employed several technics of orthotopic autotransplantation of the dog heart with survival measured in minutes. They emphasized the technical problems associated with anastomosis of the aorta and pulmonary veins which resulted in uncontrollable hemorrhage and suggested retaining a cuff of recipient left atria1 wall with attached pulmonary veins for simplicity of reanastomosis. Lower, Stofer, and Shumway [2,24] made a monumental contribution in 1960, when they developed a practical surgical technic for orthotopic transplantation of the heart. (Fig. 2.) Their method produced consistent operative survivals in dogs with cardiac homografts and autografts and demonstrated that the transplanted heart can function in a relatively normal fashion until graft rejection intercedes. Their method has survived the test of time with only minor modifications and is basically that used in the recent clinical trials. The important features of their technic include the use of total cardiopulmonary bypass to support the recipient, retention of a dish of the recipient’s left and right atrium containing the entrance of the pulmonary veins and the venae cavae, thus permitting more rapid and precise suture anastomosis, and preservation of the donor heart by rapid cooling in isotonic saline solution at 4’~. during transfer from the donor to recipient. In 1962 this same group [25] reported 80 per cent operative survival in canine cardiac autotransplants and hemodynamic The

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studies in long-term survivors [,26,27,28], indicating that the denervated autograft was physiologically altered but capable of maintaining the circulation. In these dogs cardiac outputs were essentially normal and the responses to inotropic and vasopressor drugs were delayed but similar in magnitude to those observed in control animals. They also found early electrocardiographic ST-T wave depression, which disappeared approximately five hours after res’toration of the coronary circulation, and observed that ventricular conductivity and activation occurred in normal patterns. [29,30] in 1962 autoWillman and co-workers transplanted dog hearts, employing a different operative technic and reported a high operative mortality and limited long-term survival. In their operative survivors congestive heart failure characteristically developed, which frequently resulted in death in spite of vigorous therapy; they suggested that these deleterious effects were the consequence of extrinsic cardiac denervation or interrupted lymph drainage. The varying results obtained by these two groups was of great concern to workers in this [27] suggested field and Dong and associates that the differences were the result of variations in operative technics and postoperative management. Donald and Shepherd [S1] and Dong et al. [27’] in studies of denervated hearts obtained essentially identical results, indicating that function was satisfactory. In 1964 Dong and co-workers [B8] studied canine cardiac autografts eighteen to twenty-three months after transplantation and observed that atria1 pacemakers governed cardiac rhythm and vagal reinnervation was evidenced by sinus arrhythmia, reflex bradycardia with arterial hypertension, and bradycardia with direct vagal stimulation. Sympathetic reinnervation was indicated by positive chronotropic responses to Tyramine. Cardiac output and right heart pressure were normal at rest and during exercise, and heart rate increased physiologically during exercise. They concluded that the major physiologic determinants of cardiac control and performance operate relatively normally in the transplanted heart and that cardiac homotransplantation in man should be successful if graft rejection could be averted. Lower [SZ] in 1966 suggested that the posterior atria1 walls which are retained in the recipient of cardiac transplants are rich in baroVol. 117, March 1%)

Method of orthotopic cardiac transplantaFig. 2. tion introduced by Lower and Shumway which retains the right and left atrial posterior walls for anastomosis with the donor heart. This technic enables more rapid anastomosis and preserves the conduction mechanisms.

receptors which may play a significant role in circulatory homeostasis. In 1963 Willman et al. [SS] found that orthotropic cardiac autotransplants were characterized by an immediate loss of sympathetic and parasympathetic neuroregulation, depletion of myocardial catecholamines, and cardiac hypersensitivity to exogenous l-norepinephrine. These findings persisted for several months until approximately a year after operation there was evidence of cardiac reinnervation and a return of normal responses to exogenous catecholamines. Further studies in 1964 by this same group [S4] showed that the autotransplanted heart had enhanced responses to directly administered catecholamines and altered or absent responses to many other sympathomimetric drugs, although the response to digitalis was normal. In addition chronic studies [35] of twelve to thirty month survivors of cardiac autotransplants, reinnervation usually began approximately eleven months after transplantation and was accompanied by normal responses to sympathetic and parasympathetic stimulation and the return of myocardial catecholamine concentrations to normal. In 1964 Willman et al. [36] found no alterations in acetylcholinesterase activity and a rise in cardiac glycogen and hexokinase activity in autotransplanted hearts, which con347

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firmed the previous observations of Chiba and associates [al] in heterotopic cardiac homotransplants. Potter and associates [U] in 1965 found that after cardiac autotransplantation and degeneration of postganglionic sympathetic nerve fibers, average myocardial catecholamine content declined to 1 per cent of normal, radioactive CF-Dopamine uptake and norepinephrine synthesis were markedly reduced, uptake and retention of H3-norepinephrine was decreased, and the ultrastructure of the transplanted tissue was barely distinguishable from normal. These studies demonstrated the important role of adrenergic innervation of the heart in normal synthesis, storage, and inactivation of norepinephrine. Stone, Bishop, and Dong [88] in 1967 observed that the completely denervated heart alters its output solely by changes in stroke volume, whereas hearts subjected to beta-adrenergic blockade alone can alter cardiac output by increasing heart rate due to decreased parasympathetic activity. In 1967 Daggett and associates [39] characterized the work capacity and efficiency of the autotransplanted heart. Left ventricular oxygen consumption was similar to that of control animals, although left ventricular weights were significantly greater than normal. Oxygen consumption was greater in autotransplants during the performance of comparable levels of external work and at similar states of contractility. No significant alterations in the handling of glucose, lactate, or pyruvate were observed, and although normal dogs showed no net change in circulating catecholamines across the coronary vascular bed, those with autotransplants consistently showed a net uptake of catecholamines. Electron microscopy showed enlarged myocardial mitrochondria in the autotransplants. These investigators concluded that the totally denervated cardiac autotransplant can achieve a level of performance similar to that of the normal heart, but is less efficient in doing so. On the basis of the information obtained from these studies of cardiac autotransplants, it became apparent that the consequences of cardiac denervation do not preclude relatively normal heart function, and successful homotransplantation would largely depend upon the modification of immunologic factors controlling graft rejection. Homotramplantation. The investigations of 348

orthotopic cardiac autotransplants demonstrated that cardiac transplantation could be accomplished with consistent success from a technical standpoint and that the denervated heart performs satisfactorily in maintaining the circulation. With this evidence it became apparent that successful homotransplantation of the heart was largely dependent upon the control of immunologic mechanisms, and problems associated with procurement, intercorporeal preservation, and storage of donor hearts. The earliest attempts at orthotopic transplantation of the homologous heart are credited to Neptune and his associates [40] who in 1953 transplanted the heart and lungs of dogs with the aid of hypothermia, circulatory arrest, and plastic connectors for rapid vessel anastomosis. They obtained survival for as long as six hours after operation with the transplanted heart maintaining the circulation. Webb and Howard [41] in 1957 were the first to utilize a pump oxygenator to support recipient animals during homologous transplantation of the heart. They found that donor hearts could be preserved successfully by perfusion with Ringer’s lactate solution and refrigeration at 4%. in Tyrode’s solution and 10 per cent serum or by maintaining the heart and lungs as a functioning unit during preparation of the recipient and graft implantation. Successful restoration of relatively normal cardiac function with acceptable electrocardiographic patterns was common and survival for up to twenty-two hours was achieved. In 1958 Golberg, Berman, and Akman [42,4S] transplanted dog hearts with the aid of cardiopulmonary bypass. The donor dog heart was arrested with potassium citrate in an effort to reduce its metabolism during transfer and they describe for the first time the use of a left atria1 cuff to avoid individual reanastomosis of pulmonary veins. The transplanted heart quickly resumed a forceful beat after restoration of coronary blood flow, and when cardiopulmonary bypass was discontinued, the transplanted heart maintained the circulation for several hours before decompensation occurred. In 1959 Webb, Howard, and Neely [44] expanded their experiences with cardiac homotransplants utilizing their previously reported technics and obtained short-term survivals with acceptable cardiac function. One of the most important milestones in the development of cardiac transThe American

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Heart Transplantation plantation occurred in 1960 when Lower and Shumway [8] reported their technics (Fig. 2) and a series of successful cardiac homotransplants in dogs. Their animals lived for six to twenty-one days and appeared entirely normal until death occurred as a consequence of homograft rejection. Serial electrocardiograms demonstrated ST-T wave changes compatible with postoperative pericarditis and in some instances the P waves were abnormal. Tracings obtained a few hours prior to death showed no evidence of arrhythmia or conduction defects. At autopsy the hearts were ecchymotic and edematous and microscopic examination revealed severe myocarditis with massive round cell infiltration, patchy necrosis, interstitial hemorrhage, and edema. The regional lymph nodes were grossly enlarged and microscopy showed a specific increase in plasma cells and histiocytes. In 1962 Lower and associates [45] successfully homotransplanted canine hearts after hypothermic preservation at 4%. for as long as seven hours. Attempted storage by this technic for longer periods was unsuccessful although adequate function could often be temporarily restored after twelve to fourteen hours. They concluded that with rapid cooling of the heart to 4%., six or seven hours is the maximum duration of hypoxia from which the normal heart can completely recover. In 1963, Blumenstock and his collaborators [SS] administered methotrexate to suppress rejection in dogs with orthotopic cardiac homotransplants and found a decreased rate in rejection with maximum survival of forty-two days. Swelling of vascular endothelium and perivascular infiltration of lymphocytes and plasma cells usually seen during the first day were not seen in animals that died at one, three, and four days. Moderate perivascular mononuclear infiltration and calcification were observed in longer survivors and extensive myocardial necrosis occurred in one animal treated late with azathioprine. In 1965 Lower, Dong, and Shumway [47] reported their experience with long-term survival of orthotopic cardiac homografts in dogs treated with methyl prednisolone and 6-mercaptopurine or azathioprine for immunosuppression. In a control group of twenty untreated dogs with orthotopic cardiac homotransplants, the duration of survival varied from four to twenty-one days with a mean survival of seven days. Twenty-five animals were treated with immunosuppressive drugs with Vol.117,March 1969

survival ranging from six to 260 days. Those treated with immunosuppressant agents were divided into three groups: one received continuous therapy, beginning on the day of operation ; the second received continuous treatment from the time of operation until eight to eighteen days, when the drugs were discontinued abruptly to observe the pattern of rejection; and in the third group intermittent therapy was employed during episodes of threatened rejection as determined by decreases in the electrocardiographic voltage and was observed to reverse the electrocardiographic voltage decline. Several complications of drug therapy were observed including infections, delayed wound healing, and poor nutrition, which were of least magnitude in the group intermittently treated. Microscopic examination of the hearts from dogs with prolonged survival demonstrated diminished congestion, edema, and cellular infiltration of the myocardium. There was, however, extensive focal myocardial degeneration and scarring which presumably resulted from previous episodes of rejection, and advanced arteriolar degeneration occasionally resulted in areas of fresh infarction and necrosis. They concluded that although prolonged survival was achieved with intermittent immunosuppressive therapy, a degree of irreversible myocardial coverage occurs which must be attributed to rejection and that a multiplicity of treated rejection crises may result in accumulative myocardial changes which could be of major physiologic importance. It is of additional significance that in all animals surviving more than three months, proliferative intimal lesions in the coronary vessels were present, indicating chronic low grade rejection which may not be appreciated until it reaches advanced and irreversible stages. Kondo, Gradel, and Kantrowitz [48,49] in 1965 successfully homotransplanted puppy hearts with the aid of profound hypothermia and obtained prolonged survival without immunosuppressive treatment. Twentyfour of forty puppies survived more than one day, thirteen lived more than seven days, and one dog was alive and well at 112 days. Later the same year these same investigators [SO] resuccessful homotransplantation of ported puppy hearts stored in cold Tyrode’s solution for sixty minutes or preserved by refrigeration at 4%. and hyperbaric oxygen (three to four atmospheres absolute) for twelve hours. In 1966 Hardy et al. [51] reported their experi349

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ences with heart transplantation in dogs, in which they utilized the technics described by Lower and Shumway, and reviewed the intraoperative probIems and postoperative compIications which commonly interfered with success. The results of these investigations established conclusively that cardiac homotransplantation was technically and physiologically possible and established certain base lines for the use of immunosuppressive therapy. At this point it became apparent that trials in human subjects were in the near future.

Heterotransplantation The significance of developments which would permit successful organ heterotransplantation can be readily appreciated when one considers the following comments of Medawar [52] : “A general solution of the problems of how to make human tissues and organs acceptable to someone other than the original owners would be a discovery of great practical importance, only to be compared with those other few discoveries (anesthesia and asepsis, for example) which have made it possible for surgical science to move forward as a whole. But the scale of its usefulness must not be exaggerated beyond reason. Homografting will never occupy more than a small fraction of surgical practice; the point is that when homografts are needed at all they ‘are needed urgently, often with mortal urgency. Homografts owe their surgical promise not to the number but to the gravity of the occasion on which they might be used.” For these and many other reasons it is evident that human homotransplantation is limited by many factors which are likely to prevent its development into a practical clinical tool. The use of organs from animal species would circumvent most of these problems and biologic organ banks could be maintained. In attempts to transplant tissues between different species, however, such as cat to dog, turkey to chicken, rat to mouse, and primate to man, rejection usually occurs at a remarkably rapid rate and the graft quickly fails[48]. As a result of these experiences considerable pessimism exists concerning heterotransplants, which may be best summarized by the state350

ments of Billingham [53] : “As a general rule heterografts are destroyed rapidly and with many species combinations rarely enjoy even a transient phase of apparent well being. There are good grounds for belief that with most donor to host species combinations, heterografts are destroyed principally if not exclusively through the agency of readily detectable humeral antibodies as opposed to the immunologically activated bloodborne mononuclear cells of the lymphatic series responsible for homograft rejection. Furthermore there is evidence that the antigens involved in heterograft reactivity are more stable and powerful than those responsible for provoking destruction of homografts. Unless one is prepared to deny the mutational basis of evolution, it seems inconceivable that a heterograft from any non-human primate donor can ever be as compatible immunologically as a homograft from a well matched human donor. Apart from immunologic considerations there are others that must be taken into account. In the first place, the supply of suitable healthy primate donors is restricted. These animals are slow breeders and do not breed well in captivity. Secondly, the organ mass that the single donor can supply may be inadequate for a human being. And finally, there is the risk of introducing oncogenic viruses into patients who may be abnormally susceptible.” Nevertheless the concept of a readily available source for human transplantation is appealing and many of the earliest attempts at organ transplants involved the use of heterografts. Although attempts at extremity heterotransplants date back to antiquity, most of our present understanding of heterografts is the result of experiences with renal transplants in experimental animals and a few clinical trials. In many instances the early results of heterotransplants were encouraging, but failure ultimately I esulted and interest in this approach waned. The development of immunosuppressive technics and successes with renal homotransplants provided a new stimulus to explore heterotransplantation, and in 1964 chimpanzee [54] and baboon kidneys [55] were again tried in man with disappointing results. Most The American

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heterografts failed within two months and the longest survival was nine months. It was observed, however, that initial graft function, occurrence of rejection, and reversal of rejection were qualitatively similar to those seen in homografts. It is somewhat surprising that the literature concerning cardiac transplantation is almost devoid of information about heterografts. To our knowledge the first attempt at cardiac heterotransplantation was a clinical trial in 1964 when Hardy et al. [S] transplanted the heart of a chimpanzee into a male patient dying of severe coronary artery atherosclerosis. The transplanted heart was readily resuscitated but failed shortly thereafter. The only other report [56] concerning cardiac heterografts was published in 1966 and demonstrated that immunologic enhancement was not effective in rabbit-to-dog heterotopic cardiac heterografts. It is generally appreciated that successful heterotransplantation is presently precluded because of many obstacles which appear insurmountable, although it seems reasonable to predict that the recent clinical trials with cardiac homotransplants will provide a new stimulus to accomplishments which could tax the imagination. The interesting suggestion of Lederberg [57] to develop a new primate type evolved to function as an organ donor may not be as far-fetched as it may seem. In any case it appears that we may be standing at the threshold of a revolutionary period of organ transplantation, which has been triggered by successful transplantation of the human heart.

Organ Procurement and Preservation Practical clinical transplantation of vital unpaired organs, such as the heart, will depend upon the availability of suitable donors. Use of the human cadaver heart as presently defined involves two main problems. Can such a heart be adequately resuscitated after transplantation so that it is able to support fully and immediately the host’s circulation, and what is the legal and clinical definition of death? In 1953, Baker [58] resuscitated fetal hearts obtained 45 to 150 minutes after removal from terminated pregnancies by means of perfusion technics and demonstrated that forceful cardiac activity could be consistently restored. Kuliabko [59] and Ossinowski [SO] have revived the hearts of children up to Vol.117,hlarchlN9

thirty hours after death. Webb and Howard [ZZ] in 1957 perfused donor hearts with cold Ringer’s lactate solution and stored them in Tyrode’s solution with 10 per cent serum at 4’~. during preparation of recipient animals for homologous heart transplants. They found that this technic provided adequate protection of the donor heart during the time necessary for the performance of cardiac transplantation. These same authors [41] in 1957 homografted hearts into the necks of host dogs, utilizing this technic for donor heart storage, and demonstrated a consistent return of adequate funetion, whereas control hearts, which had not been perfused or refrigerated, would not maintain the limited circulation. Golberg and Akman [42], in their early experiments with homologous cardiac transplants, employed potassium-induced cardioplegia to reduce cardiac metabolism during transplantation ; and although the heart was able to temporarily maintain the circulation in many of the animals, cardiac function was generally inadequate. In 1959, Webb, Howard, and Neely [44] showed that donor hearts refrigerated in a nutrient medium at 4%. could be preserved for eight hours and return to relatively normal function. Barsamian et al. [61] in 1959 found that puppy hearts dehydrated to 55 per cent could be resuscitated after rehydration and heterotopic transplantation within a maximum safe period of twenty-four hours. Rapid freezing of puppy hearts followed by slow thawing produced irreversible injury, whereas a combination of mechanical dehydration and glyceryol saturation permitted super cooling to -18%. without freezing. Although these hearts could be resuscitated after rehydration and warming, they were transplanted into the neck of recipients and not required to assume the load of the circulation. In 1960 Lower and Shumway [2] reported their surgical technics which were consistently successful in homotransplantation of the canine heart. The hearts were rapidly excised from donor animals, immediately immersed in normal saline solution at ~OC., and implanted in recipient animals which were maintained on cardiopulmonary bypass. Webb, DeGuzman, and Hoopes [62] in 1961 summarized the status of cardiac preservation at that time and stated that the normothermic heart can survive total anoxia for ninety minutes without evidence of damage and upon transplantation these hearts can maintain 351

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physiologic work loads if the coronary cireulation is flushed of all blood. When refrigeration in a nutrient medium was added, relatively normal function returned after intercorporeal storage up to eight hours. After eight hours, however, the preparation showed steady deterioration of function. Lower and associates [45] in 1962 successfully transplanted normal canine hearts after anoxic preservation for as long as seven hours by immersion in saline solution at 2 to 4’~. These hearts electrically defibrillated with ease but required a period of support with a pump-oxygenator, during which their contractions progressively improved and cardiopulmonary bypass could be discontinued. When donor hearts were maintained anoxic in a cold saline bath for twelve to fourteen hours prior to transplantation, defibrillation and the return of a beat were obtained, although the electrocardiogram showed consistent T-wave inversion and rapid ventricular failure ensued when cardiopulmonary bypass was withdrawn. Hearts anoxic for twentyfour hours would not remain defibrillated for any significant length of time and the gross and microscopic appearance was that of edema with scattered areas of interstitial myocardial hemorrhage. Efforts to perfuse the coronary vessels of the cold arrested heart were disappointing, and adequate pressures to maintain competence of the aortic valve damaged the small myocardial vessels, resulting in interstitial hemorrhages. Retrograde myocardial perfusion through the coronary sinus and hypothermia was suggested by Hardy [S,Sl,SS,SS]. However, its value for long-term perfusion remains to be evaluated. As a result of these studies it became apparent that hypothermic technics, although satisfactory for short-term organ preservation, were generally disappointing and the solution to storage of complex organs was sought in other directions. Bloch and associates [65] in 1964 stored dog hearts in vitro for twenty-four hours, employing a combination of hypothermia and hyperbaric oxygenation, and these hearts regained a forceful beat when transplanted in the neck of recipient animals. Eyal and associates [66] in 1965 added chlorpromazine to hypothermia and hyperbaric oxygenation and obtained improved cardiac preservation. Hearts stored in this fashion could be resuscitated when transplanted to the neck of host animals after twenty-four to forty-eight hours of storage. 352

They attributed the benefits of chlorpromazine to stabilization of cellular membranes and inhibition of mitochondrial swelling. Webb and co-workers [67] in 1966 demonstrated that magnesium sulfate or magnesium fluoride and adrenochrome were effective in suspending animation of the rat heart. Viability of anoxic hearts at 37”~. was increased significantly by these agents when compared to those with anoxic or cardioplegic arrest. They suggested that magnesium has its protective effect in maintaining membrane potentials or diverting available energy into essential functions. Shumway, Lower, and Stofer [68] employed the technics described by Bloch et al. [65] and found that although these hearts would readily defibrillate and maintain a coordinated beat after twenty-four to forty-eight hours of hypothermic and hyperbaric preservation, they failed to handle small pumping requirements after orthotopic transplantation. They also observed that the isolated heart continued to beat for several hours when coronary perfusion was maintained with a pump and either a mechanical or biological oxygenator, although interstitial bleeding and edema gradually appeared and cardiac function deteriorated. Shumway, Angell, and Wuerflein [69] found that two to three hours of biologic perfusion will restore cadaver heart function sufficiently to permit later successful orthotopic transplantation. Cadaver status of one hour’s duration could be reversed by placing the heart in conduit with the femoral vessels of an intermediate host. These same workers have employed storage in an intermediate host after resuscitation by biologic perfusion [70]. The graft is implanted into the neck of the intermediate host which is treated vigorously with immunosuppressive drugs. These hearts are then removed after varying periods and transplanted to an orthotopic locus. This method has produced successful orthotopic transplants after four days of storage, but resuscitation of the organ is mandatory before heterotopic placement. Wuerflein and Shumway [71] in 1967 demonstrated that cadaver dog hearts, anoxic for ninety minutes at normothermia, could be resuscitated with 70 per cent success by perfusion technics. An occasional success occurred after ninety to 120 minutes of anoxia but after two hours none could be revived. After resuscitation the cadaver hearts were maintained as a functional heart-lung preparation for peThe American

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riods up to thirty hours. Some of these hearts were then transplanted in the orthotopic position and, although there were no long-term survivors, all were able to maintain the circulation without supportive drugs for eight to twenty-eight hours. Robicsek and co-workers [72] in 1967 reported a stabilized heart-lung preparation which made possible experimental transplantation of a fully beating heart without the assistance of parabiosis or extracorporeal circuits. A similar technic was developed by Whiffen, Boake, and Gott [78] and, although both of these methods are ingenious, they would appear to have little practical significance at this time. Copeland, Kosek, and Hurley [74] in 1968 produced anoxic cardiac arrest in canine hearts and observed the time course of morphologic changes at cadaver temperatures for periods up to two hours post mortem and morphologic and functional response during reperfusion with oxygenated blood for as long as seven hours. Functional recovery after reperfusion was evaluated by contractile force measurements and recovery was more rapid in those hearts subjected to the shortest periods of anoxia. Satisfactory recovery followed anoxia of fifteen to twentyfive minutes, decreased functional and morphologic recovery followed thirty to forty-five minutes of anoxia, and poor results ensued after sixty minutes of anoxia. Structural alterations were evident after fifteen minutes and progressed in a predictable manner as the duration of anoxia increased. Changes occurring during anoxic periods of up to thirty minutes were for the most part reversed after one hour of perfusion and characteristic nuclear, cytoplasmic, and mitochondrial recovery patterns were observed. Anoxic arrest of fortyfive minutes or more produced ultrastructural changes that were consistently reversed by reperfusion. At this time progress in organ preservation and storage is lagging substantially behind other transplantation developments and may significantly retard advances in organ replacement. Several avenues of investigation are being explored and perhaps the solution may be found in some unusua1 area such as the mechanisms controlling hibernation in animals. Metabolic inhibitors in conjunction with hypothermia and hyperbaric oxygenation may provide a “preservation triad” which will prove satisfactory for short-term organ storage, alVot.117.March 1969

though long-term storage will probably the development of low-flow continuous fusion technics. In any case, at this moment the most tical and effective method of managing hearts appears to be premortem donor thermia followed by rapid removal of the after death and immediate transplantation the recipient with the aid of donor organ thermia.

await perpracdonor hypoorgan into hypo-

Rejection Phenomenon and Its Modification In 1962 Bing and associates [75] reviewed the status of cardiac transplantation and concluded that transplantation immunity is the common basic obstacle to success in all transplants except those between identical twins. They summarized the rejection process wasthe consequence of antigens originating in the graft reaching regional lymph nodes, stimulating the production of immunologically activated cells which then infiltrate the graft and destroy it. This explanation, of course, is a marked oversimplification and it is now generally appreciated that &any complex immunologic mechanisms are involved and substantial controversy still exists concerning the precise mechanisms by which rejection occurs [76]. The general histologic pattern observed in the homografted heart is characterized by an initial perivascular infiltration of lymphoid cells, and as rejection proceeds, extensive cellular infiltration occurs with associated myocardial edema and necrosis until by the time cardiac contractions cease, very little myocardium appears viable by microscopy. Homograft rejection has been considered to be predominantly a cellular process, although Ramos et al. [77] in 1963 demonstrated that humeral factors are present which increase vascular permeability in the homograft as early as one hour after transplantation and before a cellular response is observed. Chiba et al. [.21] described the serial histopathologic alterations accompanying rejection of canine cardiac homografts and stated that the earliest changes were an accumulation of lymphocytes about blood vessels within three hours after transplantation. Swelling of vascular endothelium follows and by five hours a cellular infiltrate of lymphocytes, plasma cells, macrophages, and histiocytes is evident. At nineteen hours Ashoffand Anitschklow-like cells appear and a granulomatous myocarditis develops, advancing from 353

Vasko perivascular to interstitial involvement with lymphocytic and histiocytic invasion of the myocardium. Days later vascular endothelial and myocardial granulomatoses are pronounced and necrosis of the myocardium is prominent. In the presence of accelerated rejection, these changes were amplified and necrosis began as early as four hours after grafting. Leandri [78] in 1967 observed that the cellular reaction in cardiac homografts occurred mainly in the perivascular areas and in the subepicardial and subendocardial layers. All valvular structures were involved by cellular infiltration but aortic and pulmonary cusps were involved less severely. The vascular changes were typically the most severe, although occasionally some vascular segments and myocardial areas were remarkably spared. Intermittent immunosuppressive therapy with azathioprine diminished these changes but did not completely prevent the rejection process, suggesting that continuous immunosuppressive therapy might better control the host versus graft reaction. Although the specific nature of the antigens that produce transplantation immunity is unknown, it is recognized that antigenic substances are especially concentrated in the nuclear fractions of cells. Also it is generally appreciated that transplantation immunity is individual and not organ- or species-specific and the involved antigens are under genetic control. Successful transplantation then depends upon identical dominant genes in both donor and recipient, as demonstrated by successful transplantation of organs between identical twins. In addition, it should be emphasized that for practical purposes genetic compatibility between unrelated individuals is so rare that it can be disregarded. In view of these factors, it is evident that if tissue transplantation is to be consistently successful, methods must be found to attenuate the immunologic reaction of the host and induce graft tolerance. Several observations have provided important clues which have contributed substantially to our present understanding of transplantation immunology and the development of technics for the control of rejection. Electrophoretic studies have demonstrated that many antibodies reside in gamma globulins and patients with agammaglobulinemia occasionally accept homografts. Immunologic inadequacy may also be found in patients with uremia, lymphomas, advanced malignancies, 354

cirrhosis of the liver, scleroderma, and several other conditions. In animals, tolerance may be induced by introducing reticuloendothelial cells, such as those contained in the spleen, from the prospective donor into the prospective recipient in utero or at the time of birth, before the immune apparatus of the recipient has “learned” to distinguish its own tissue from those of the donor. Later, by virtue of this induced tolerance, grafts from the original donor may be accepted. There is also some evidence that transplantation immunity may be largely mediated by nucleic acid fractions. Suppression of the immune reaction in animals and man has been achieved by host irradiation, radiomimetic alkylating agents, antimetabolites such as 6-mercaptopurine, methotrexate or azathioprine, adrenocorticosteroids, and certain antibiotics. The objective, of course, is to develop potent methods which will suppress graft rejection without producing dangerous side effects in the host. These methods have been used alone and in various combinations but heavy total body irradiation is being used less frequently because of the related high mortality and morbidity. In 1962 Reemtsma et al. [20] employed a folic acid antagonist (amethopterin) in an effort to delay rejection of puppy hearts grafted into the necks of adult animals. Survival of the grafts was increased to a maximum of twenty-six days, which was substantially greater than the ten day survival in untreated animals. Blumenstock et al. in 1963 [46] treated dogs with orthotopic cardiac homotransplants with methotrexate and obtained prolonged survival and a reduction in the magnitude of graft rejection. Moderate perivascular mononuclear infiltration and calcification were seen in the myocardium of animals surviving longer than threeorfourdays. Advances in renal transplantation produced significant refinements in immunosuppressive therapy and presently the combination of azathioprine and prednisone are most widely used clinically. Lo-wer and associates 14.51 compared continuous, temporary, and intermittent immunosuppressive treatment with azathioprine combined with methylprednisolone in dogs with orthotopic cardiac homografts using the electrocardiographic voltage as a monitor of impending rejection. Prolonged survival of cardiac homografts was obtained with intermittent immunosuppression during periods of The

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threatened rejection, whereas continuous immunosuppressive therapy resulted in a high rate of serious complications, including drug toxicity, infections, and gastrointestinal bleeding, and failed consistently to prevent rejection. Discontinuation of temporary therapy was followed by the rapid onset of rejection. Although intermittent therapy resulted in the greatest numbers of survivors, microscopic studies of these hearts showed arteriolar and myocardial fiber degeneration, which may be irreversible features of chronic rejection and may ultimately produce significant physiologic impairment of the graft. Kondo et al. [48,49, 501 homotransplanted puppy hearts into the orthotopic focus with the aid of profound hypothermia. Thirteen of forty animals survived more than seven days, and one animal survived 112 days postoperatively without the use of immunosuppressive agents. The prolonged survivals are difficult to explain, although unpredicted histocompatibility may be the common denominator. It should be emphasized that in all reported experiments very few animals have survived for longer than three months, although occasionally an animal has survived a year or longer after homotransplantation and treatment with immunosuppressive agents. Considerable enthusiasm is developing concerning the potential immunosuppressive value globulin antilymphocyte of heterologous (ALG), which has been used recently with substantial success in renal transplantation [79,80]. The use of ALG has reduced the early mortality in clinical renal homotransplantation and has permitted the use of reduced dosages of other immunosuppressive drugs, particularly steroids, which have contributed heavily to post-transplantation morbidity and mortality. Various other technics of immunosuppression have been developed, including local irradiation of the graft, radiation of circulating blood, and thoracic duct cannulation and drainage. All appear to have some value in specific cases, but are generally considered as adjuvants to the use of immunosuppressive drugs. Histocompatibility testing has been employed with encouraging results in renal homografting [80] and it seems reasonable to expect that these technics will contribute significantly to improved survival in cardiac transplantation. One of the basic principles of transVol.117.March 1969

plantation immunology is that the intensity of the rejection reaction depends upon the extent to which the donor and recipient share important genetically determined transplantation antigens. When they differ at major histocompatibility loci, reactions of near maximal strength are provoked which are extremely difficult to suppress by available technics without severely impairing the well-being of the host. When donor and host are similar with respect to major transplantation antigens, histoincompatibilities are relatively weak, graft acceptance is more likely to occur with reduced immunosuppressive therapy, and immunologic tolerance in the host can be more easily achieved. Most surgeons agree that chance matching of donors and recipients with respect to histocompatibility factors probably explains a large proportion of successful longterm renal homografts in man. There is little evidence that genetic determinants of histocompatibility express themselves on the erythrocytes. However, some types of red-cell antigens may be important since there is evidence that there is little success for renal homografts between persons whose blood groups are incompatible. Presently the preponderance of attention to histocompatibility testing has been directed at leukocytes, which are known to express transplantation antigens in a highly effective form. Pioneer workers in the area include Dausset [al], Payne [82], Van Rood and Van Lecuwen [83], and Terasaki, Marchioro, and Star-21 [84]. Conclusion In any case, it is apparent that the immunologic obstacles to successful organ transplantation are being attacked from many standpoints and this has been productive of significant improvements in clinical results and it is likely that immediate advances in transplantation will be the result of a combination of several new therapeutic technics. The ultimate resolution of the rejection problem, however, probably awaits intensive investigation at cellular and subcellular levels. Of particular importance to immunosuppressive treatment in heart transplantation is the ability to monitor the progress of rejection. An extensive search for hemodynamic indicators of early rejection has not been fruitful and the only available index has been a decline in electrocardiographic voltage, which 355

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has proved to be relatively insensitive. Unlike renal transplantation, in which renal function is a relatively sensitive indicator of rejection by which immunosuppressive therapy can be regulated and in which hemodialysis is available to support the host, late rejection of the heart results in death since there is presently no available technic for long-term support of the circulation. Until some reliable indicator of early cardiac rejection becomes available to guide immunosuppressive therapy, it is likely that graft failure will be a significant problem. In view of present evidence it seems reasonable to conclude that, although cardiac transplantation is technically and physiologically feasible, the hazards of immunosuppression, the paucity of parameters to monitor rejection, and the major problem of chronic rejection constitute a significant deterrent to prolonged survival.

References 1.

BARNARD,C. N. A human cardiac transplant: an interim report of a successful operation performed at Groote Schuur Hospital, Cape Town. South African M. J., 41:1271,1967. 2. LOWER, R. R. and SHUMWAY, N. E. Studies on orthotopic homotransplantation of the canine heart. S. Forum, 11:18, 1960. 3. HARDY,J. D., CHAVEZ,C. M., KURRUS,F. D., NEELY,W. A., ERASLAN,S., TURNER,M. D., FABIAN, L. W., and LABECKI,T. D. Heart transplantation in man: developmental studies and report of a case. J.A.M.A., 188: 1132,1964. 4. CARREL,A. and GUTHRIE, C. C. The transplantation of veins and organs. Am. Med., 10:1101,1905. 5. MANN, F. C., PRIESTLEY,J. T., MARKOWITZ, J., and YATER, W. M. Transplantation of the intact mammalian heart. Arch. Surg., 26 :219, 1933. 6. SINITSYN, N. Peresadka serdtsa kak noviyi metod v eksperimental’noe. Biologii i Meditsine, 1948, Moscow. 7. MARCUS, S., WONG, S. N. T., and LUISADA, A. A. Homologous heart grafts: transplantation of the heart in dogs. S. Forum, 2: 212,195l. 8. WESOLOWSKI,S. A. and FENNESSEY,J. F. Pattern of failure of the homografted canine heart. Circulation. 8 :750.1953. 9. MEDAWAR,P. B. Behavior and fate in skin autografts in rabbits. J. Anatomy, 78:176, 1944. 10. MARCUS,E., WONG, S. N. T., and LUISADA, A. A. Homologous heart grafts. I. Tech356

11. 1.2.

13. 14.

15.

16.

17.

18.

19.

20.

21.

22.

23.

24.

25.

nique of interim parabiotic perfusion. II. Transplantation of the heart in dogs. Arch. Surg., 66:179,1953. DOWNIE, H. G. Homotransplantation of the dog heart. Arch. Surg., 66:624, 1953. LUISADA,A. A. and MARCUS,E. The behavior of a transplanted heart. Cardiologia, 25 :16, 1954. SAYEGH, S. F. and CREECH, O., JR. Transplantation of the homologous canine heart. J. Thoracic Surg., 34 :692,1957. LEE, S. S. and WEBB, W. R. Cardiac metabolism as influenced by ischemia, refrigeration and enzyme precursors. Am. Surgeon, 25 ~776, 1959. REEMTSMA,K., DELGADO,J. P., and CREECH, O., JR. Transplantation of the homologous canine heart: serial studies of myocardial blood flow, oxygen consumption, and carbohydrate metabolism. Surgery, 46:292, 1906. DEMIKOV,V. P. Experimental Transplantation of Vital Organs. New York, 1962. Consultants Bureau. REEMTSMA,K. The heart as a test organ in transnlantation studies. Ann. New York Acad: Sci., 120 ~778, 1964. MCGOUGH, E. C., BREWER, P. L., and REEMTSMA,K. The parallel heart: studies in intrathoracic auxiliary cardiac transplants. Surgery, 60:153, 1966. DEPASQUALE,N. P., BURCH, G. E., PENA, E., and REEMTSMA, K. Electrocardiographic studies in homologous canine heart transplantations. Circulation Res., 16 :197, 1965. REEMTSMA, K., WILLIAMSON, W. E., JR., IGLESIAS,F., PENA, E., SAYEGH,S. F., and CREECH,O., JR. Studies in homologous canine heart transplantation : prolongation of survival with a folic acid antagonist. Surgery, 52 :127,1962. CHIBA, C., WOLF, P. L., GUDBJARNASON, S., CHRYSOHOU,A., RAMOS, H., PEARSON,B., and BING, R. J. Studies on the transplanted heart. J. Exper. Med., 115:853,1962. WEBB, W. R. and HOWARD,H. S. Cardio-pulmonary transplantation. S. Forum., 8 :313, 1957. CASS, M. H. and BROCK, R. Heart excision and replacement. Guy’s Hosp. Rep., 108: 285,1959. LOWER,R. R., STOFER,R. C., and SHUMWAY, N. E. Homovital transplantation of the heart. J. Thoraoic & Cardiovas. Surg., 41: 196, 1961. HURLEY,E. J., DONG, E., JR., STOFER,R. C.,. and SHUMWAY,N. E. Isotopic replacement of the totally excised canine heart. J. S. Res., 2:90, 1962. The American

Journal

of Surgery

Heart Transplantation 26.

27.

28.

29.

30.

31.

32.

33.

34.

35.

36.

37.

38.

39.

DONG, E., JR., LOWER, R. R., HANCOCK,E. W., STOF~, R. C., and SHUMWAY, N. E. An approach to extracorporeal surgery of the heart. .J. Thormk & Cardiouccs. Surg,, 44:776,1962. DONG, E., JR., HURLEY,E. J., LOWER, R. R., and SHUMWAY, N. E. Performance of the heart two years after autotransplantation. Surgemd, 66:270,1964. DON% E., JR., FOWKES,W. C., HURLEY,E. J., HANCOCK, E. W., and PILLSBURY, R. C. Hemodynamic effect of cardiac autotransplantation. Circ&~&n, 29 :77, 1964. WILLMAN, V. L., COOPER,T., CIAN, L. G., and HANLON, C. R. Autotransplantation of the canine heart. Surg. Ggnec. & Obst., 115: 299,1962. WILLMAN, V. L., COOPER,T., CIAN, L. G., and HANLON, C. R. Mechanmismof cardiac failure after excision and reimplantation of the canine heart. S. Fomm., 13:93, 1962. DONALD,D. E. and SHEPHERD,J. T. Response to exercise in dogs with cardiac denervaLion. Am. J. Phyeiol., 205:393, 1963. LOWER, R. R. Cardiac transplantation without complete oardiac denervation: the role of the afFerent receptors. Am. Heart J., 72: 841, 1966. WILLMAN, V. L., COOPER,T., CIAN, L. G., HANLON, C. R. Neural responses following autotransplantation of the canine heart. Ciroulation, 27 :713, 1963. COOPER,T., WILLMAN, V. L., and HANLON, C. R. Drug responses of the transplanted heart. Dis. Chest, 45:284, 1964. WILLMAN, V. L., COOPER,T., and HANLON, C. R. Return of neural responses after autotransplantation of the heart. Am. J. Physiol., 207 : 187, 1964. WILLMAN, V. L., JEZLINEK, M., COOPER,T., TSUNEKAWA,T., KAISER, G. C., and HANLON, C. R. Metabolism of the transplanted heati: effect of excision and reimpIantation on myocardid glycogen, hexokinase, and acetylcholine esteraze. Surgery, 56 : 266, 1964. POTTER, I,. T., COOPER,T., ‘WILLMAN, V. L., and WOLFE, D. E. Synthesis, binding, release and metabolism of norepinephrine in normal and transplanted dog hearts. Circulation Res., 16:468, 1965. STONE, H. L., BISHOP, V. S., and DONG, E., JR. Ventricular function in cardiac-denervated and cardiac-sympa&hectomized consciom dogs. Circulation Res., 20 :687, 1967. DAGGETT, W. M., WILLMAN, V. L., Coop% T., anti HANLON, C. R. Work capacity and efftgency of the antotransplanlted heart circulation, 35 :96, 1967.

HmLEy, E. J.,

Vol. 117, March1969

40.

41.

42.

NEPTUNE, W. B., COOKSON,B. A., BAILEY, C. P., APPLER,R., and RAJKOWSKI,F. &,QMete hommlogous heart transplantation. Arch. Surg., 66:173, 1953. WEBB, W. R. and HOWARD,H. S. Restoration of function of the refrigerated heart. S. Forum, 8 :302,1957. GOLBERG,M., BERMAN, E. F., and AKMAN, L. C. Homologous transplantation of the canine heart. J. Internat. Coil. Surge~m, 30 :576,1958.

43.

44.

45.

46.

BERMAN,E. F., GOLBERG,M., and AKMAN, L. Experimental replacement of the heart in the dog. Transplant Bull., 5 :iO, 1958. WEBB, W. R., HOWARD,H. S., and NEELY, W. A. Practical methods of homologous cardiac transplantation. J. Thoracic Sung., 37 :361, 1959. LOWER, R. R., STOFER, R. C., HURLEY, E. J., DONG, E., JR., COHN, R. B., and SRUMWAI-, N. E. Successful homotraneplantation of the canine heart after anoxic preserv&ion for seven hours. Am. J. Surg., 104:302, 1962. BWMENSTOCX, D, A., HECHTMAN, Ii. B., Cob LINS, J. A., JARETZKI, A., III, HOSBEIN, J. D., ZINGG, W., and POWERS,J. H. Prolonged

survival of orthotopic homotransplants of the heart in animals treated with methotrexate. J. Thoracic & Cardiovas. Smg., 46:616, 1963. 47. LOWER, R. R., DONG, E., JR., and SHWMWAY, N. E. Long-term survival of cardiac homografts. Surgev, 58:110,1965. 48. KONDO, Y., GFUDEL, F., KANTXOWITZ, A. Homotransplanta,tion of the heart in puppies under profound hypothermia: long survival without immunosuppressive treatment. Ann. Surg., 162:837, 1965. 49. KONDO,Y., GRADEL, F., and KANTROWXTZ, A. Heart, homotransplantation in puppies: long survival without immunosuppressive therapy. Circulation, 31: 181, 1965. 50. KONDO, Y., GRADEL, F. O., CHAPTAX+ PA., MEYER, W., COTTLE, H. R., and KANTROWITZ, A. Immediate and delayed orthotopic homotranslplantation of the heart. J. Thma& & Cardiovm. Surp., 60 : 781,1%% 51. HARDY, J. D., CHAVEZ, (2. M., EmSuN, S-9 ADKINS, J, R., and WILLIAMS, R. D. Heart transplantation in dogs. Procedures, physiologic problems, and results in 142 eXPetimerits. Szcrgerg, 60 :361, 19%. 52. MEDA~AR, P. B. The immunology of tray* plantation. In : The Harvey Lectures 1966 57, p. 144. New York, 1968. Academic Press. 53. BILLINGHAM, R, E. SOme problems in OrfF~ transplantation. Surgery, 121:27% 1965. 35?

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REEMTSMA, K. Renal heterotransplantation in man. Ann. Surg., 160:409, 1964. 55. STARZL, T. E. Renal heterotransplantation from baboon-to-man : experience with six cases. Transplantation, 2 : 752, 1964. 56, KENNEDY,J. H. and ALOUSI,M. The effect of enhancement on survival of heart heterotransp!ants in the dog. J. Cardiovas. Surg., 7 :156,1966. J. Biological future of man. In: 57. LEDERBERG, Man and His Future. Edited by Wolstenholme, G. E. W. Boston, 1963. Little, Brown & co. 58. BAKER, J. B. E. Some observations upon isolated perfused human foetal hearts. J. Physiol., 120 :122, 1953. 59. KULIABKO, M. A. Sur la reviviscence du coeur. Rappel, des battements du coeur human en trente heures apres la mort. Compt. rend. Acad. Sci. Paris, 136:63, 1903. 60. OSSINOWSKI,N. I. Versuche, evier, wiederbelebung des isoliecteu kinderherzens. Acta paediat., 29:233, 1940. 61. BARSAMIAN,E. M., JACOB,S. W., COLLINS,S. C., and OWEN, 0. E. The transplantation of dehydrated and supercooled hearts. S. Forum, 10 :lOO, 1959. 62. WEBB, W. R., DEGUZMAN,V., and HOOPES,J. E. Cardiopulmonary transplantation : Experimental study of current problems. Am. Surgeon, 27 :236,1961. 63. HARDY, J. D. The transplantation of organs. Surgery, 56 :685, 1964. 64. HARDY, J. D., KURRUS,F. D., CHAVEZ,C. M., and WEBB, W. R. Heart transplantation in infant calves: evaluation of coronary perfusion to preserve organ during transfer. Ann. New York Acad. Sci., 120:786, 1964. 65. BLOCH, J. H., MANAX, W. G., EYAL, Z., LILLEHEI,R. C. Heart preservation in vitro with hyperbarci oxygenation and hypothermia. J. Thoracic & Cardiovas. Surg., 48 :969, 1964. 66. EYAL, Z., MANAX, W. G., BLOCH, J. H., and LILLEHEI, R. C. Utilization of chlorpromazine in heart storage and its combined use with hyperbaric oxygen and hypothermia. Surgev, 120 :1237,1965. WEBB, W. R., DODDS, R. P., UNAL, M. O., 67. KAROW, A. M., COOK, W. A., and DANIEL, C. R. Suspended animation of the heart with metabolic inhibitors: effect of magnesium sulfate or fluoride and adrenochrome in rats. Ann. Surg., 164 :343,1966. SHUMWAY, N. E., LOWER,R. R., and STOFER, 68. R. C. Advances in Surgery: Transplantation of the heart, vol. 2, p. 265. Chicago, 1966. Year Book Medical Publishers, Inc. SHUMWAY,N. E., ANGELL,W. W., and WUER69.

54.

358

FLEIN, R. D. Recent advances in cardiac replacement. Surgew, 62 : 794, 1967. 70. ANGELL, W. W. and SHUMWAY, N. E. Resuscitative storage of the cadaver heart transplant. S. Forum, 17 :224,1966. 71. WUERFLEIN,R. D. and SHUMWAY, N. E. ReJuscitation and function of the cadaver heart. Circulation, 35 :92,1967. 72. ROBICSEK,F., LESAGE, A., SANGER, P. W., DAUGHERTY, H. K., GALLUCCI, V., and BAGBY, E. Experimental studies: transplantation of “live” hearts. Am. J. Cardiol., 20 :803, 1967. 73. WHIFFEN, J. D., BOAKE, W. C., and GOTT, V. L. Normothermic orthotopic canine heart homotransplantation. J. S. Res., 7:421, 1967. 74. COPELAND, J., KOSEK,J. C., and HURLEY,E. J. Early functional and ultrastructural recovery of canine cadaver hearts. Circulation, 37:188, 1968. 75. BING, R. J., CHIBA, C., CHRYSOHOU, A., WOLF, P. L., and GUDBJARNASON, S. Transplantation of the heart. Circulation: 25,273, 1973. 76. CHIBA, C., KONDO,M., ROSENBLATT, M., WOLF, P. L., and BING, R. J. The selective irradiation of canine lymph nodes by means of intralymphatic injection of “PI. Transplantation, 5 :232, 1967. P., 77. RAMOS, H. R., CHIBA, C., SCHOLLMEYER, WOLF, P. L., PEARSON,B., and BING, R. J. The presence of humeral factors in homograft rejection of the transplanted heart. Transplantation, 1:284, 196’ 78. LEANDRI,J. Some histological features of canine cardiac transplants. Thoraz, 22 :397, 1967. 79. RAPPAPORT,F. T. and DAUSSET, J. Human Transplantation, p. 357. New York, 1968. Grune and Stratton. 80. STARZL,T. E., GROTH,C. G., TERASAKI,P. I., PUTNAM, C. W., BRETTSCHNEIDER, L., and MARCHIORO, T. L. Heterologous antilymphocyte globulin, histoincompatibility matching, and human renal homotransplantation. Surg. Gynec. & Obst., 126:1023, 1968. 81. DAUSSET,J. Leucocytes, platelets and human homografts. VOX Sang., 7 :257,1962. 82. PAYNE, R. The development and persistence of laucoagglutinins in parous women. Blood, 19:411, 1962. 83. VAN ROOD,J. J. and VAN LECUWEN,A. Leucocyte grouping: a method and its application. J. Clin. Invest., 42:1382, 1963. 84. TERASAKI, P. I., MARCHIORO,T. L., and STARZL, T. E. Serotyping of human leucocyte antigens: preliminary trials on long term kidney homograft survivors. In: Histocompatibility Testing. Monograph NO. 83. National Academy of Science. The

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