World J. Surg. ... Recipients and Concordant and Discordant Donor Species: Foundation .... third-party pig's lung (or both) were used to adsorb preexisting.
World J. Surg. 21, 943–950, 1997
WORLD Journal of
SURGERY © 1997 by the Socie´te´ Internationale de Chirurgie
Survival following Orthotopic Cardiac Xenotransplantation between Juvenile Baboon Recipients and Concordant and Discordant Donor Species: Foundation for Clinical Trials Leonard L. Bailey, M.D., Steven R. Gundry, M.D. Department of Surgery, Division of Cardiothoracic Surgery, Loma Linda University Medical Center, Loma Linda, California 92354, USA
Abstract. It has been more than a decade since the last clinical trial of cardiac xenotransplantation in a newborn infant. Since that event, laboratory research at Loma Linda University has focused on survival studies of orthotopically xenografted juvenile baboon recipients. Both concordant and discordant donor species have been used. Transgenic donors have not been explored at Loma Linda. Instead, simplified host immunoregulative protocols, consistent with those used in neonatal cardiac allografting, have been adapted to xenotransplant research. Xenograft bridge to alloengraftment was evaluated in a series of five juvenile baboon recipients. Heterotopically implanted cardiac xenografts stimulated host production of xenoreactive antibody. Orthotopic cardiac allografting was then carried out. Xenoantibody appeared to play little role in immediate or chronic survival of experimental hosts. A clinical protocol of xenobridging to allotransplantation would likely succeed. Two consecutive series of orthotopically xenotransplanted hosts using rhesus monkey cardiac donors demonstrated unprecedented long-term survival. Splenectomy combined with maintenance therapy consisting of FK-506 and methotrexate contributed to survival of up to 502 days in one series of xenografted baboon hosts selected for ABO blood grouping, mixed lymphocyte culture, and crossmatch compatibility. Survival beyond a year (maximum 515 days) among three consecutive juvenile baboon recipients of orthotopically implanted rhesus monkey hearts, in which splenectomy was omitted and cyclosporine was substituted for FK-506, represents a benchmark achievement. Commencing maintenance immunosuppression several weeks prior to transplantation appeared to improve chronic survival significantly. Investigation of discordant (pig-to-baboon) host survival has focused on adsorption of naturally occurring xenoreactive antibody at the time of transplantation. This strategy, combined with pretransplant total lymphoid irradiation and both pre- and posttransplant immunosuppression, succeeded in preventing hyperacute rejection and resulted in survival of up to 24 days, thereby permitting observation of the delayed xenograft rejection phase. Data support consideration of additional clinical trials of concordant neonatal cardiac xenotransplantation and offer promise for the development of discordant xenotransplantation as an ultimate therapeutic resource.
Mortality among neonates and young infants awaiting heart transplantation in North America ranges from 15% to 40%. These figures do not include hundreds of others who could potentially benefit from transplantation but die annually because they never reach the waiting list. Fewer than 200 American infants achieve cardiac alloengraftment during any one year. This donor-dependent figure has not increased over the past 5 years, indicating that Correspondence to: L.L. Bailey, M.D.
altruistic infant organ donation either will not or cannot support more. It is for this reason and others that worldwide xenotransplantation research is at an all-time high. Orthotopic cardiac xenotransplantation at Loma Linda University has focused on both concordant and discordant models. The purpose of this report is to summarize and illustrate data generated in these laboratories as it relates to cardiac xenotransplantation in juvenile baboon recipients. Concordant xenograft donors have included African green and rhesus monkeys, and piglets have provided organs for discordant transplantation. Concordant Xenotransplantation Human neonatal cross-species transplantation evolved around the idea of xenograft bridging to cardiac allografting. The important question relating to this approach was whether the bridged recipient would develop an antibody response to the initial xenograft that would be cross-reactive with the allograft donor. Sensitization of this nature might preclude successful secondary allotransplantation. This question was initially explored by Alonso de Begona [1], who employed a model in which African green monkey hearts were transplanted into the necks of five immunosuppressed juvenile baboons using a technique previously illustrated by the Columbia University group. These grafts rejected over a period of 5 to 65 days. Lymphocytotoxic xenoantibody was identified in recipient blood samples. The rejected xenografts were removed, and the recipient circulating xenoantibody titers were observed to peak over 24 to 48 hours. Using cardiopulmonary bypass primed without blood, the immature baboon recipients then underwent orthotopic cardiac allotransplantation. All survived the secondary allotransplant procedure without evidence of hyperacute, antibody-mediated rejection. They were immunosuppressed to varying degrees using a cyclosporine (CSA) protocol. The last two recipients were permitted to survive chronically. Table 1 documents outcomes among the five recipient animals. Despite a high titer of circulating xenoantibody in each of the host baboons (xenoantibody that was cross-reactive to donor baboon lymphocytes), orthotopic alloengraftment was possible in all five recipients. Each was immunosuppressed with gradations of CSAbased therapy. Survival to 5 and 6 months of the last two
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Table 1. Survival of xenografts and allografts and host therapy employed in a xenograft bridge to allograft model using an immature baboon recipient. Cardiac heterotopic xenograft (African green monkey)
Cardiac orthotopic allograft (common olive baboon)
Experiment
Therapya
Survival (days)
Therapya
Survival (days)
Allograft rejection
1 2 3 4 5
A A A A; B 1 Crescue A1B1C
11 5 6 13 65
A A; A; A; A;
10 58 65 198b 164b
Severe Moderate to severe Moderate to severe None None
Brescue Brescue B 1 Crescue Binduction
a Immunosuppression. A: cyclosporine 1 azathioprine 1 solumedrol; B: goat anti-human T cell IgG; C: monoclonal antibody; rescue: therapy for acute rejection; induction: therapy for induction only. b Electively euthanized.
Table 2. Results of orthotopic cardiac xenotransplantation between immature baboon recipients and rhesus monkey donors. Animal Group 1 (controls)a 1 2 3 4 5 6 Group 2a 1 2 3 4 5 Group 3a 1 2 3 4 5 6 7
Survival (days)
Pathologic condition of autopsied xenograft
IgM deposit
Cause of death
6 7 8 8 9 10
Moderate cellular rejection Moderate cellular rejection Severe cellular rejection Severe cellular rejection Severe cellular rejection Severe cellular rejection
2 6 2 1 1 6
Rejection Rejection Rejection Rejection Rejection Rejection
25
CMV infection, no rejection
2
32 53 57 75
CMV infection Mild cellular rejection Mild cellular rejection No rejection
2 2 6 6
CMV infection (heart, lung, pancreas, liver, kidney) CMV infection (heart) CMV infection (lung) During rejection treatment During rejection treatment
Mild cellular rejection Cellular infiltration to coronary artery Mild cellular rejection No rejection Patchy fibrosis in septum and inferior wall Toxoplasmosis No rejection
2 2 6 2 2 2 2
Klebsiella pneumoniae CMV infection (lung, kidney) Pulmonary embolism Renal failure CMV infection (lung, kidney) Toxoplasmosis CMV infection
35 43 74 96 123 234 502
2: absent; 1: present; 6: indeterminate; CMV: cytomegalovirus. Group 1: controls. Group 2: antilymphocyte globulin induction, FK-506 maintenance immunosuppression. Rescue therapy using methotrexate and methylprednisolone. Group 3: same immunosuppressive strategy as for group 2, except methotrexate was utilized as chronic maintenance therapy. Group 2 and 3 subjects had pretransplant splenectomy. a
consecutive animals (which were ultimately euthanized) was not unlike that expected for allotransplanted hosts. Xenoantibody did not appear to alter acute or chronic survival of baboon recipients managed with a clinically applicable regimen of immune regulation. The two chronic survivors had well-functioning allografts that were free of significant rejection injury. These findings have subsequently been confirmed and elaborated on by Michler et al. [2]. Kawauchi et al. [3] completed a series of 18 orthotopic cardiac xenotransplants using immature olive baboon recipients and rhesus monkey donors, selected for ABO blood grouping, mixed lymphocyte culture, and crossmatch compatibility. On an immunosuppressive regimen that included pretransplant splenectomy and the use of FK-506 and methotrexate for chronic management, prolonged survival was observed. Table 2 catalogs outcomes in this series, which includes one of the longest known survivors of orthotopic cardiac xenotransplantation (a baboon recipient that
lived 502 days). Two important features emerged from this series of preclinical transplants. First, most deaths were due to either infection or the logistics of maintaining reliable chronic immunosuppression and surveillance in this difficult model. Second, a subgroup of four hosts, subjected to a period of pretransplant maintenance immunosuppression [4], was among the longest surviving recipients, including the animal that lived 502 days. Hayashi et al. [5] have confirmed this observation in a hamsterto-rat model. The concept of pretransplant immunosuppression was subsequently applied to a second set of animals (described later). A closer look at the longest surviving recipient is of interest. Figures 1, 2, and 3 each relate to Max, the baboon recipient of an orthotopically implanted rhesus monkey heart. Illustrations represent findings in this animal 12 months after transplantation. He was, and remained, a healthy, active, growing baboon whose cardiac function studies and coronary arteriograms were inter-
Bailey and Gundry: Survival after Cardiac Xenotransplantation
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Fig. 3. Coronary arteriograms (CAG) obtained on routine evaluation of Max, an immature baboon recipient of a rhesus monkey heart orthotopically implanted 1 year prior to these contrast studies. Coronary arteries appear normal in size and distribution. Fig. 1. Max, an immature baboon recipient of an orthotopic cardiac xenotransplant acquired from a donor rhesus monkey. He was an active, growing, healthy baboon photographed 1 year after transplantation. He went on to live 502 days and died of cytomegalovirus disease. His xenograft was free of rejection, but coronary disease was in evidence at autopsy.
Fig. 2. Contrast left ventriculograms (LVG) obtained on routine evaluation of Max, an immature baboon recipient of a rhesus monkey heart. Systolic ejection and diastolic compliance were normal in the xenograft 1 year after orthotopic transplantation.
preted as normal. His cause of death points out some of the difficulties inherent in the use of chronic primate experimental models. In an effort to convert him to oral medications, his level of immunosuppression fluctuated widely. Unreliable maintenance immune regulation led to a late, powerful rejection response. This xenograft rejection was reversed successfully using corticosteroids and T cell antibody. The additional bolus immunosuppression,
however, permitted the development of generalized cytomegalovirus (CMV) disease and eventual bacterial sepsis from which Max ultimately died. The animal’s autopsied xenograft was free of rejection. Management of CMV infection in this splenectomized series of baboon recipients proved to be at least as difficult as controlling the immune response toward their cardiac xenografts. The cellular immune response to these xenografts has been characterized by Fukushima et al. [6, 7], who also examined the incidence of accelerated graft atherosclerosis. Two of the twelve (17%) study recipients showed necropsy evidence of coronary intimal hyperplasia producing partial coronary obstruction. Matsumiya et al. [8 –10] developed a similar series of six xenotransplanted baboons in which the concept of pretransplant immunosuppression was utilized and splenectomy was omitted. Cyclosporine and methotrexate were administered in a chronic maintenance protocol comparable to that utilized for infant cardiac allografting at Loma Linda University. Three consecutive recipients lived for more than 1 year, the longest period being 515 days. There was limited morbidity and mortality relating to xenograft rejection (30%). Most morbidity and mortality related to viral infection and the vicissitudes of chronic maintenance immunoregulation in juvenile baboons. Graft surveillance was based on noninvasive echocardiographic findings. Table 3 and Figure 4 detail outcomes of this series of xenotransplanted baboons. A key feature of these primate xenotransplants was documentation of xenograft growth [11]. Consecutive long-term survival in an orthotopically xenoengrafted primate model is an important benchmark on which to consider further clinical trials of concordant neonatal cardiac xenotransplantation. Discordant Xenotransplantation The feasibility of transplanting across discordant xenogenic barriers in an orthotopic newborn pig-to-juvenile baboon model was first explored in the Loma Linda laboratories during the early 1990s. Because hyperacute rejection was at that time the single
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Table 3. Summary of survival data among three consecutive baboon recipients of orthotopic xenotransplantation using rhesus monkey donor hearts.a Animal
Survival (days)
No. of rejection episodesb
1 2
413 515
11 2
3
371
8
Cause of death Disseminated herpesvirus Right heart fibrosis (coronary obstruction) Encephalopathy (probably papova virus)
a Baboon recipients were treated prior to transplantation with a 2- to 4-week course of cyclosporine and methotrexate administration followed by a 7- to 10-day drug-free interval, then immunosuppressed chronically with maintenance doses of cyclosporine and methotrexate. Methylprednisolone was utilized in bolus doses to reverse rejection episodes identified on daily echocardiography. Animal 1 died during a presumed rejection episode for which total lymphoid irradiation was being employed. He had been a somewhat aggressive immune responder (11 rejection episodes treated) throughout his course. Necropsy revealed no rejection or graft coronary disease. Instead, the animal was found to have suffered widespread herpesvirus infection. b Rejections were detected mainly by echocardiography.
most important factor in limiting discordant xenotransplantation, early strategies were directed toward eliminating or reducing baboon preformed xenoantibody to swine sugar antigens. Bouchart and Fukushima performed a series of orthotopic transplants of pig hearts into juvenile baboons. Donor pig or a large, third-party pig’s lung (or both) were used to adsorb preexisting anti-pig antibody from the recipient baboon’s blood during cardiopulmonary bypass and subsequent pig heart xenotransplantation. Animals were treated with a perioperative immunosuppressive regimen of splenectomy plus FK-506, deoxyspregoline, and nefamstat mesilate (FUT), an anticomplement agent. All animals survived transplantation. Examination and echocardiography revealed no evidence of hyperacute rejection. Survival ranged from 3 to 16 days, with the longest survivor succumbing to pneumonia without clinical or microscopic evidence of rejection. A variable amount of delayed xenograft rejection was observed histologically among the other recipient baboons [12, 13]. A second series of discordant orthotopic transplants was accomplished by Matsumiya et al. [14]. Splenectomy was omitted, and the baboon recipients were preoperatively immunosuppressed for 1 month using CSA and methotrexate. Total lymphoid irradiation (TLI) was administered to the baboon recipients for 1 week prior to transplantion. Immunoadsorption was performed with a third-party pig’s lung during cardiopulmonary bypass. Baboons received postoperative maintenance doses of CSA and methotrexate. All animals survived the transplantation procedure. Nearly complete adsorption of anti-pig xenoantibody was documented, and no signs of hyperacute rejection were observed. Discordant graft function was monitored noninvasively by echocardiography done on a daily basis in each recipient. Delayed xenograft rejection occurred almost uniformly between postoperative days 10 and 14. Rejection was successfully reversed in two animals using bolus infusions of methylprednisolone, but the two animals succumbed to exacerbation of delayed xenograft rejection at 19 and 24 days, respectively. Cellular infiltrates included mononuclear cells, macrophages, and natural killer (NK) cells, suggesting that delayed discordant xenograft rejection occurs by
Fig. 4. Baboon recipient of orthotopic rhesus monkey cardiac xenotransplantation. This recipient lived 515 days on a maintenance immunosuppressive regimen of cyclosporine and methotrexate. The animal required treatment with methylprednisolone boluses for only two episodes of graft rejection. She grew and developed well. She ultimately died of right coronary artery obstruction and selective right ventricular fibrosis.
mechanisms other than classic allograft cellular or humoral pathways. To determine if the newborn would be a more suitable host for discordant xenotransplantation, a series of heterotopic pig heartto-newborn goat transplants was performed by Xu et al. in the Loma Linda laboratories. Newborn goats, like newborn humans and baboons [15], exhibit no preformed swine-specific xenoantibody. Xu et al. observed pig graft survival up to 7 days in untreated newborn goat recipients [16]. There was no suggestion of hyperacute rejection. Interestingly, pig xenoantibody was rapidly produced by the newborn goats. Subsequent experiments have shown the presence of anti-pig immunoglobulin M (IgM) within splenocytes of newborn goats. Splenectomy, splenic irradiation, or both may be required, even in newborns, to permit prolonged discordant xenoengraftment. Discussion Solid-organ allografting is an everyday, worldwide event in 1997. Renal, hepatic, and cardiac allografting has achieved outcomes that could never have been predicted early in transplantation history. Strategies of immune regulation promise even brighter results in the future, possibly leading to host tolerance. Despite this picture, allograft organ donations have remained woefully static and inadequate. With more than 50,000 potential recipients filling the transplantation waiting lists in North America alone, the time is ripe for expanding donor resources. Many experimental biologists believe the ultimate answer lies in the ability to engineer pigs genetically in such a way as to “humanize” antigenpresenting elements of graft endothelium or to counter antigen–
Bailey and Gundry: Survival after Cardiac Xenotransplantation
antibody interactions on these endothelial surfaces. This multifaceted process is well under way, as evidenced by several of the reports in this issue. There are many compelling reasons, however, why carefully planned and executed clinical trials of concordant solid-organ xenotransplantation ought soon to begin among selected neonatal recipients. These trials would make use of immature baboon donors selected on the basis of ABO blood grouping compatibility, major histocompatibility (MHC) homology, the absence of potentially harmful infectious diseases, and host freedom from preformed lymphocytotoxic antibody. Engineering a “universal” baboon donor is under way at Loma Linda University [17]. Data concerning outcomes of experimental concordant xenotransplantation in primates coupled with Loma Linda University’s experience with a previous human newborn cardiac xenotransplant [18] have set the stage for carefully controlled additional clinical trials of baboon-to-newborn infant heart transplants. As was the case historically, this approach takes advantage of the newborn recipient’s relatively immature immune response, discussed by Nehlsen-Cannarella and Chang [19] and well characterized by Michler et al. [20] in a discordant model. This neonatal “window of opportunity” has translated into a greater than 10% survival advantage among nearly 100 newborn allograft recipients compared to all other age groups having cardiac transplantation at Loma Linda University [21]. Countering this effort are the speculative, yet understandable concerns relating to the transmission of infectious agents from baboons to humans, so-called xenozoonoses [22]. These concerns are not peculiar to the use of baboons but are valid with respect to any crossing of species during transplantation therapy [23]. On the one hand, the inability of species-specific viruses to infect and produce disease in a closely related species is the principle underlying two relatively recent baboon-to-human liver transplants [24] and a novel case of baboon “bone marrow” transplantation to an individual with end-stage acquired immunodeficiency syndrome (AIDS) [25]. These experiences are recorded and updated elsewhere in this special issue of the Journal. As best we know, none of these three human subjects developed xenozoonoses. Nor have subjects of any previous baboon or chimpanzee organ transplants or those with intentional medical exposure to animal organs (monkey lung oxygenators, baboon and pig extracorpeal liver perfusion, baboon corneal transplants) developed xenozoonoses. Despite mankind’s limited medical exposure to the potential for disease transmission, no known xenozoonotic public health crisis has emerged. An additional report of a healthy animal caretaker in whom simian foamy virus was isolated after exposure 20 years earlier suggests that “in contrast to active infection in monkeys, foamy virus persists in a state of latency following accidental infection of humans” [26]. On the other hand, circumstantial data leading to the assumption that human immunodeficiency viruses (HIV-1 and HIV-2) are really mutations of their simian (SIV-1 and SIV-2) counterparts fuels the fear of creating yet another public health risk through xenotransplantation. There is no direct proof that this simian retroviral mutation into HIV-1 or HIV-2 has occurred. SIV and HIV share some homology of amino acid sequencing, but it does not prove that one retrovirus mutated to become the other. Simian behavior (particularly that observed in captivity) parallels that of humans, giving apes and monkeys (particularly baboons) virtually equal opportunity to develop their own species-specific
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viral infections, be they latent and endogenously transferred from one generation to the next or some form of disease process such as immunodeficiency. SIV has been isolated in at least one asymptomatic laboratory worker (handling infected macacques) [27]. The virus has retained its SIV identifying characteristics (rather than mutating into HIV) and has not been transmitted to other individuals or produced illness in its human host. Clearly, animal-to-human transmission of catastrophic infectious disease has and does occur and is distressing. Some of the infectious agents acquired by humans from animals sources have the capability of lateral transmission (epidemic potential) and others do not. Ebola, Hanta, Marburg, Bunyavirus, and the herpesvirus known as B-virus are zoonoses contracted largely by accidental human exposure. Each is capable of producing a rapidly lethal outcome in humans. Fortunately, each can be isolated, identified, and (where pertinent) eliminated during the course of selecting a donor animal. Unfortunately, stories relating to these viruses have made them the substrate of frightening novels and motion pictures, which have set the stage for social hysteria. An intelligent, healthy respect for the potential of xenozoonotic illness is, and always has been, a vital part of planning for clinical trials of xenotransplantation. One superb report in this symposium is devoted to this topic, and infectious disease is a principal agenda item for any commission or regulatory body charged with review and oversight of xenotransplantation. Be that as it may, historical observations and the ability to select baboon donors for the absence of threatening viruses and other infectious diseases are reassuring. A healthy suspicion is prudent with regard to the potential for development of xenozoonoses by means of xenotransplantation. Impassioned pleas to inhibit clinical trials of xenotransplantation based on virologic speculation and fear is entirely inappropriate. Research experience with orthotopic discordant xenotransplantation at Loma Linda University is also unique in terms of host survival patterns. Preformed primate xenoantibody directed toward donor pigs in unmodified solid organ transplants results in immediate graft destruction. Using a simplified strategy of antibody adsorption and conventional host maintenance immunosuppression, hyperacute graft loss was avoided in a series of orthotopic piglet-to-juvenile baboon cardiac xenotransplants, and recipient survival averaged a week and in one instance ranged to 16 days (Fig. 5). Subsequent investigations, which included pretransplant host immunosuppression and total lymphoid irradiation, extended survival up to 24 days. This unprecedented survival of orthotopically transplanted hosts using discordant cardiac grafts is equal to or exceeds that observed using donor pigs transgenic for expression of human complement regulatory factors. Survival among the Loma Linda series of piglet-to-baboon xenografts also compares favorably with kidney graft survival observed in a strategy of “tolerance” through mixed chimerism following bone marrow transplantation. Clearly, there are several mechanisms in existence for overcoming the initial, hyperacute immune response typically encountered with discordant xenotransplantation. These various manipulations have finally permitted observation and initial characterization of subsequent phases of the powerful immune response observed between disparate species. Several distinguished authors address these issues elsewhere in this anthology. As these subsequent xenograft rejection phases become better understood, it is likely they may eventually be controlled. Pretransplant modification of
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seriously. These agencies have assumed a preemptive role to protect public health and welfare. Each proposal or protocol to initiate additional clinical trials of xenotransplantation will be reviewed by these agencies with an eye to scientific merit and appropriate safeguards for both the experimental subjects and for the public health. Keeping in mind the daily loss of human life that occurs during the exasperating wait for human donor organs, institutions poised to commence or recommence clinical trials of xenotransplantation will do so only after appropriate evaluation by designated institutional review boards and federal oversight agencies whose mandate is to ensure that patient and public benefit clearly outweigh potential risks.
Re´sume´
Fig. 5. Vigorous, healthy juvenile baboon recipient of orthotopically implanted piglet heart photographed 10 days after transplantation. She went on to live 16 days, dying suddenly of delayed xenograft rejection.
both the donor graft and the recipient immune response suggests a basis for optimism about eventual clinical application of discordant xenotransplantation. Laboratory data infer that clinical trials should ultimately occur in a neonatal recipient first. There are vexing social issues relating to the appropriate care and use of animals that cloud the future of clinical xenotransplantation. Vociferous, even militant so-called animal rights groups, whose opinions (though not widely held) ring out in protest, require some level of attention. Few among the society in which we live seriously protest the compassionate use of an animal’s organs to save a human life. Clinical documentation of human life improved or saved (as may be the case in the most recent baboon-to-human bone marrow transplant) can serve to place limited social protest in context. The balance of nature supports the concept of preserving one’s own species, even if regularly at the expense of another. Social protest, on the constructive side, serves to remind us to be sensitive to the trade-off that inevitably occurs in the balance of nature. Aggressive, terrorist-like group behavior within the human species is an issue for law enforcement, not for medical science. Relevant philosophy and ethics pertaining to xenotransplantation are outlined specifically elsewhere in this series of reports. Conclusions Data from the experimental laboratories of Loma Linda University and elsewhere during the past decade indicate that the momentum toward clinical xenotransplantation is both timely and justified. Federal oversight agencies are taking this research
La dernie`re xe´notransplantation cardiaque chez le nouveau-ne ´ date de plus de 10 ans. Depuis, notre recherche de laboratoire `a l’Universite´ de Loma Linda s’est consacre´e `a la survie des babouins juve´niles apre `s une xe´nogreffe orthotopique. On a re´alise´ des expe´riences avec des donneurs concordants et discordants. On n’a pas, par contre, fait d’expe´rience avec des donneurs transge´niques. Pour la recherche sur les xe ´nogreffes, des protocoles immunore´gulateurs simplifie´s, similaires `a ceux utilise´s pour les allogreffes cardiaques du nouveau-ne´, ont ´ete´ ´elabore´s. L’expe´rimentation a consiste ´ en la re´alisation d’une xe ´nogreffe suivie d’une allogreffe dans une se´rie de cinq receveurs babouins juve´niles. La xe ´nogreffe cardiaque he ´te´rotopique a ´ete´ utilise´e pour stimuler la production d’anticorps xe´nore´actifs. Une allogreffe cardiaque orthotopique a ensuite ´ete´ re´alise´e. Les xe ´noanticorps ont apparemment joue´ peu ou pas de ro ˆle dans la survie `a court ou `a long terme des ho ˆtes dans ces expe´riences. Un protocole clinique de «xe´norelais» avant l’allogreffe aurait pu re´ussir. Deux se´ries successives d’ho ˆtes xe´notransplante ´s orthotopiquement, en utilisant des singes rhe´sus comme donneurs cardiaques, ont donne´ lieu `a une survie `a long terme sans pre´ce´dent. La sple´nectomie combine ´e `a une the´rapie de maintien consistant en le FK-506 et le me´thotrexate ont permis une survie de 502 jours dans une se´rie de xe´nogreffes chez les ho ˆtes babouins, se´lectionne ´s pour leur compatabilite ´ ABO, MLC, et croise´e. La survie au dela` d’un an (max. 515 jours) chez trois babouins juve´niles, receveurs de coeur de singe rhe´sus orthotopique, chez lesquels la sple´nectomie a ´ete´ omise et chez lesquels la cyclosporine a remplace´ le FK-506, repre´sente un succe`s qui fait re´fe´rence. La survie `a long terme est nettement ame´liore´e en commenc¸ant le traitement immunosuppresseur plusieurs semaines avant la transplantation. Les investigations portant sur la survie de l’ho ˆte discordant (cochon-babouin) se sont surtout concentre´es sur l’absorption d’anticorps xe´nore´actifs naturels au moment de la transplantation. Cette strate´gie, combine ´e `a l’irradiation totale lymphoı¨de pre´transplantation et l’immunosuppresion pre ´- et posttransplantation, a re´ussi `a pre´venir le rejet hyper-aigu et a permis une survie de 24 jours. De cette fac¸on, on a pu observer une phase de rejet de xe´nogreffe retarde ´. Ces re´sultats sont en faveur de la poursuite d’essais cliniques supple´mentaires sur la xe ´notransplantation cardiaque ne´onatale concordante, et offrent une promesse de de´veloppement de xe ´notransplantation discordante comme une ultime ressource the´rapeutique.
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Resumen
References
Ya ha pasado ma´s de un decenio desde el ensayo clı´nico de xenotrasplante cardı´aco en un recie´n nacido. A partir de tal evento, la investigacio ´n de laboratorio en la Universidad de Loma Linda se ha enfocado hacia la supervivencia de mandriles jo ´venes sometidos a xenotrasplantes ortoto ´picos. Se han utilizado tanto especies discordantes como concordantes, aunque no se ha trabajado con donantes transge´nicos. Ma´s bien, se han adaptado protocolos immunoregulatorios simplificados en el hue´sped a la investigacio ´n en xenotrasplante neonatal. El xenotrasplante como puente para el alotrasplante cardı´aco fue evaluado en una serie de cinco mandriles jo ´venes receptores. Los xenotrasplantes cardı´acos implantados en forma heteroto ´pica estimularon la produccio ´n de anticuerpo xenoractivo en el hue´sped. El aloinjerto cardı´aco fue luego practicado. El xenoanticuerpo parecio ´ jugar un papel menor en cuanto a la supervivencia inmediata o cro ´nica del hue´sped experimental, y se piensa que podrı´a haber sido exitoso un protocolo clı´nico de un xenotrasplante puente para llegar al alotrasplante. Dos series consecutivas de hue´spedes con xenotrasplantes ortoto ´picos utilizando corazones donantes de monos rhesus, demostraron una supervivencia sin precendente a largo plazo. La esplenectomı´a combinada con una terapia de mantenimiento consistente en FK-506 y metotrexato contribuyo ´ a una supervivencia hasta de 502 dı´as en una serie de mandriles que fueron seleccionados por compatibilidad ABO, Ma C y pruebas cruzadas. La supervivencia de un an ˜o (ma´ximo 515 dı´as) entre tres mandriles jo ´venes receptores de corazones de monos rhesus implantados en forma ortoto ´pica en los cuales se omitio ´ la esplenectomı´a y la ciclosporina fue sustituida por FK-506, representa un verdadero logro experimental. El comenzar la inmunosupresio ´n de mantenimiento con anterioridad de varias semanas al trasplante parecio ´ mejorar en forma significativa la supervivencia cro ´nica. La investigacio ´n sobre la supervivencia de hue´s pedes discordantes (cerdo-a-mandril) ha sido enfocada a la adsorcio ´n del anticuerpo xenoreactivo natural en el momento del trasplante. Tal estrategia, combinada con irradiacio ´n linfoide total pretrasplante y con inmunosupresio ´n pre y post-trasplante, probo ´ ser exitosa en la prevencio ´n del rechazo hiperagudo y resulto ´ en supervivencia hasta de 24 dı´as. Esto ha posibilitado la observacio ´n de la fase tardı´a de rechazo al xenotrasplante. Nuestros resultados dan apoyo a la consideracio ´n de ensayos clı´nicos adicionales de xenotrasplante cardı´aco neonatal concordante, y son promisorios en lo referente al desarrollo del xenotrasplante cardı´aco discordante como un recurso terape´utico final.
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Acknowledgments This research has been spearheaded by a litany of brilliant Fellows from Japan, France, Spain, China, and America and has spanned more than a decade. Dr. Sandra Nehlsen-Cannarella, director of the Loma Linda University Immunology Center, and her capable staff, particularly Omar Fagoaga, deserve special recognition for immense support and counsel. This work could not have been accomplished without the loyal support of the surgical research laboratory staff, including Geoffrey Visbal, Joyce Ivey, Jan Smallwood, and Nancy Holmquist.
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