Bruno Reichart’s research while affiliated with Ludwig-Maximilians-Universität in Munich and other places

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Publications (40)


Ultrasound Assessment of Pleural Effusions After Orthotopic Pig-to-Baboon Cardiac Xenotransplantation
  • Article

November 2024

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9 Reads

Xenotransplantation

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Bruno Reichart

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Background Pleural effusions develop frequently after cardiac surgery in humans. Lung ultrasound is an essential non‐invasive tool in the diagnosis and treatment of these effusions. Pleural effusions also develop regularly after preclinical cardiac xenotransplantation experiments. Unlike in the human setting, modern ultrasound devices lack pre‐installed tools for calculating the volume of pleural effusions in baboons. The aim of this study was to analyze ultrasound examinations of pleural effusions after orthotopic pig‐to‐baboon cardiac xenotransplantation experiments in order to develop a formula for calculating the effusion volume based on ultrasound measurements. Methods Hearts from seven genetically modified (GGTA1‐KO, hCD46/hTBM transgenic) juvenile pigs were orthotopically transplanted into male baboons. Postoperatively, the baboons were tested regularly for the development of pleural effusions using ultrasound. When thoracocentesis was required, the drained effusion volume (EV) was compared to ultrasound‐derived calculations using various formulas. These calculations were based on measuring the distance between lung and diaphragm at the effusions’ maximum height (H max ). Subsequently, the most promising formula was used to describe the interobserver variability between trained and untrained staff members to predict effusion volumes based on ultrasound measurements. Results Ultrasound measurement correlated very strongly with the absolute EV ( r = 0.9156, p < 0.0001), with EV indexed to total body weight ( r = 0.9344, p < 0.0001) and with EV indexed to body surface area (BSA) ( r = 0.9394, p < 0.0001). The ratio between H max and EV increased with total body weight and BSA and also depended on the baboon species. The sonographic measurements taken by an experienced and an inexperienced observer showed only low interobserver variability. A Bland–Altman plot of both observers’ measurements showed an overall bias of –2.39%. Conclusion Ultrasound imaging provides a simple and non‐invasive tool for measuring pleural effusion quantity in baboons. This facilitates simple and efficient monitoring even in the hands of untrained personnel and may guide the decision‐making to perform thoracocentesis.


FIGURE 1 | Survival after orthotopic pig-to-baboon cardiac xenotransplantations from 1994 to 2023. Data taken from [1, 2, 15-21, 23-27].
Progress in Orthotopic Pig Heart Transplantation in Nonhuman Primates
  • Literature Review
  • Full-text available

September 2024

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15 Reads

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1 Citation

Transplant International

Xenotransplantation of porcine hearts has become a promising alternative to human allotransplantation, where organ demand still greatly surpasses organ availability. Before entering the clinic, however, feasibility of cardiac xenotransplantation needs to be proven, ideally in the life supporting orthotopic pig-to-nonhuman primate xenotransplantation model. In this review, we shortly outline the last three decades of research and then discuss in detail its most recent advances. These include the genetic modifications of donor pigs to overcome hyperacute rejection and coagulation dysregulation, new organ preservation methods to prevent perioperative xenograft dysfunction, experimental immunosuppressive and immunomodulatory therapies to inhibit the adaptive immune system and systemic inflammation in the recipient, growth control concepts to avoid detrimental overgrowth of the porcine hearts in nonhuman primates, and lastly, the avoidance of porcine cytomegalovirus infections in donor pigs. With these strategies, consistent survival of 6–9 months was achieved in the orthotopic xenotransplantation model, thereby fulfilling the prerequisites for the initiation of a clinical trial.

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Combination of Anti-CD40 and Anti-CD40L Antibodies as Co-Stimulation Blockade in Preclinical Cardiac Xenotransplantation

August 2024

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98 Reads

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2 Citations

The blockade of the CD40/CD40L immune checkpoint is considered essential for cardiac xenotransplantation. However, it is still unclear which single antibody directed against CD40 or CD40L (CD154), or which combination of antibodies, is better at preventing organ rejection. For example, the high doses of antibody administered in previous experiments might not be feasible for the treatment of humans, while thrombotic side effects were described for first-generation anti-CD40L antibodies. To address these issues, we conducted six orthotopic pig-to-baboon cardiac xenotransplantation experiments, combining a chimeric anti-CD40 antibody with an investigational long-acting PASylated anti-CD40L Fab fragment. The combination therapy effectively resulted in animal survival with a rate comparable to a previous study that utilized anti-CD40 monotherapy. Importantly, no incidence of thromboembolic events associated with the administration of the anti-CD40L PAS-Fab was observed. Two experiments failed early because of technical reasons, two were terminated deliberately after 90 days with the baboons in excellent condition and two were extended to 120 and 170 days, respectively. Unexpectedly, and despite the absence of any clinical signs, histopathology revealed fungal infections in all four recipients. This study provides, for the first time, insights into a combination therapy with anti-CD40/anti-CD40L antibodies to block this immune checkpoint.


An Approach to Controlling Inflammation and Coagulation in Pig-to-Baboon Cardiac Xenotransplantation

July 2024

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96 Reads

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3 Citations

Xenotransplantation

Introduction Inflammatory responses and coagulation disorders are a relevant challenge for successful cardiac xenotransplantation on its way to the clinic. To cope with this, an effective and clinically practicable anti‐inflammatory and anti‐coagulatory regimen is needed. The inflammatory and coagulatory response can be reduced by genetic engineering of the organ‐source pigs. Furthermore, there are several therapeutic strategies to prevent or reduce inflammatory responses and coagulation disorders following xenotransplantation. However, it is still unclear, which combination of drugs should be used in the clinical setting. To elucidate this, we present data from pig‐to‐baboon orthotopic cardiac xenotransplantation experiments using a combination of several anti‐inflammatory drugs. Methods Genetically modified piglets (GGTA1‐KO, hCD46/hTBM transgenic) were used for orthotopic cardiac xenotransplantation into captive‐bred baboons ( n = 14). All animals received an anti‐inflammatory drug therapy including a C1 esterase inhibitor, an IL‐6 receptor antagonist, a TNF‐α inhibitor, and an IL‐1 receptor antagonist. As an additive medication, acetylsalicylic acid and unfractionated heparin were administered. The immunosuppressive regimen was based on CD40/CD40L co‐stimulation blockade. During the experiments, leukocyte counts, levels of C‐reactive protein (CRP) as well as systemic cytokine and chemokine levels and coagulation parameters were assessed at multiple timepoints. Four animals were excluded from further data analyses due to porcine cytomegalovirus/porcine roseolovirus (PCMV/PRV) infections ( n = 2) or technical failures ( n = 2). Results Leukocyte counts showed a relevant perioperative decrease, CRP levels an increase. In the postoperative period, leukocyte counts remained consistently within normal ranges, CRP levels showed three further peaks after about 35, 50, and 80 postoperative days. Analyses of cytokines and chemokines revealed different patterns. Some cytokines, like IL‐8, increased about 2‐fold in the perioperative period, but then decreased to levels comparable to the preoperative values or even lower. Other cytokines, such as IL‐12/IL‐23, decreased in the perioperative period and stayed at these levels. Besides perioperative decreases, there were no relevant alterations observed in coagulation parameters. In summary, all parameters showed an unremarkable course with regard to inflammatory responses and coagulation disorders following cardiac xenotransplantation and thus showed the effectiveness of our approach. Conclusion Our preclinical experience with the anti‐inflammatory drug therapy proved that controlling of inflammation and coagulation disorders in xenotransplantation is possible and well‐practicable under the condition that transmission of pathogens, especially of PCMV/PRV to the recipient is prevented because PCMV/PRV also induces inflammation and coagulation disorders. Our anti‐inflammatory regimen should also be applicable and effective in the clinical setting of cardiac xenotransplantation.


Perioperative changes of hyaluronan (a,b), heparan sulfate (c,d) and syndecan-1 (e,f) in absolute values (ng/mL) and as fold increases compared to the preoperative values (left and right, respectively). (b,d,f) Mean values ± SD (n = 4). CPB, cardiopulmonary bypass; POD1, first postoperative day; XTx, xenotransplantation.
Perioperative courses of serum lactate in absolute values (ng/mL) and as fold increases compared to start of surgery (a,b). Correlation between perioperative lactate levels and changes of hyaluronan (c), heparan sulfate (d) and syndecan-1 (e). (b), mean values ± SD (n = 4). CPB, cardiopulmonary bypass; POD1, first postoperative day; XTx, xenotransplantation.
Postoperative courses of hyaluronan (a), heparan sulfate (b) and syndecan-1 (c) plasma concentrations. Group I, experiments deliberately terminated after 90 postoperative days with the baboons in excellent clinical condition, mean values ± SEM (n = 2); Group II, baboons with PCMV/PRV infections, mean values ± SEM (n = 2). PCMV/PRV, porcine cytomegalovirus/roseolovirus; POD, postoperative day.
Overview of the study group. F, female; M, male; PCMV/PRV, porcine cytomegalovirus/ porcine roseolovirus.
The Endothelial Glycocalyx in Pig-to-Baboon Cardiac Xenotransplantation—First Insights

June 2024

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62 Reads

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2 Citations

Cardiac xenotransplantation has seen remarkable success in recent years and is emerging as the most promising alternative to human cardiac allotransplantation. Despite these achievements, acute vascular rejection still presents a challenge for long-term xenograft acceptance and new insights into innate and adaptive immune responses as well as detailed characterizations of signaling pathways are necessary. In allotransplantation, endothelial cells and their sugar-rich surface—the endothelial glycocalyx—are known to influence organ rejection. In xenotransplantation, however, only in vitro data exist on the role of the endothelial glycocalyx so far. Thus, in the current study, we analyzed the changes of the endothelial glycocalyx components hyaluronan, heparan sulfate and syndecan-1 after pig-to-baboon cardiac xenotransplantations in the perioperative (n = 4) and postoperative (n = 5) periods. These analyses provide first insights into changes of the endothelial glycocalyx after pig-to-baboon cardiac xenotransplantation and show that damage to the endothelial glycocalyx seems to be comparable or even less pronounced than in similar human settings when current strategies of cardiac xenotransplantation are applied. At the same time, data from the experiments where current strategies, like non-ischemic preservation, growth inhibition or porcine cytomegalovirus (a porcine roseolovirus (PCMV/PRV)) elimination could not be applied indicate that damage of the endothelial glycocalyx also plays an important role in cardiac xenotransplantation.


Transthoracic echocardiography is a simple tool for size matching in cardiac xenotransplantation

May 2024

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12 Reads

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2 Citations

Xenotransplantation

Background Preoperative size matching is essential for both allogeneic and xenogeneic heart transplantation. In preclinical pig‐to‐baboon xenotransplantation experiments, porcine donor organs are usually matched to recipients by using indirect parameters, such as age and total body weight. For clinical use of xenotransplantation, a more precise method of size measurement would be desirable to guarantee a “perfect match.” Here, we investigated the use of transthoracic echocardiography (TTE) and described a new method to estimate organ size prior to xenotransplantation. Methods Hearts from n = 17 genetically modified piglets were analyzed by TTE and total heart weight (THW) was measured prior to xenotransplantation into baboons between March 2018 and April 2022. Left ventricular (LV) mass was calculated according to the previously published method by Devereux et al. and a newly adapted formula. Hearts from n = 5 sibling piglets served as controls for the determination of relative LV and right ventricular (RV) mass. After explantation, THW and LV and RV mass were measured. Results THW correlated significantly with donor age and total body weight. The strongest correlation was found between THW and LV mass calculated by TTE. Compared to necropsy data of the control piglets, the Devereux formula underestimated both absolute and relative LV mass, whereas the adapted formula yielded better results. Combining the adapted formula and the relative LV mass data, THW can be predicted with TTE. Conclusions We demonstrate reliable LV mass estimation by TTE for size matching prior to xenotransplantation. An adapted formula provides more accurate results of LV mass estimation than the generally used Devereux formula in the xenotransplantation setting. TTE measurement of LV mass is superior for the prediction of porcine heart sizes compared to conventional parameters such as age and total body weight.


Xenotransplantation von OrganenXenotransplantation of solid organs

May 2024

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66 Reads

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1 Citation

Die Chirurgie

Zusammenfassung Die Transplantation genetisch veränderter Schweineherzen und -nieren kann in den nächsten Jahren eine Lösung für den bestehenden Mangel an Organspendern darstellen. Fortschritte im Bereich des „Genetic Engineering“, aber auch verbesserte Organpräservationstechniken, eine Immunsuppression mit Kostimulationsblockade (Anti-CD40/CD40L-mAb) sowie eine verbesserte virologische Diagnostik, um eine Übertragung von pathogenen Schweineviren auf den Empfänger zu verhindern, haben hierzu beigetragen. Da Landrasse-Schweineorgane auch im Transplantatempfänger ihre Originalgröße erreichen, werden nun Schweinerassen verwendet, die entweder ein für den Menschen passendes Endgewicht erreichen (z. B. Auckland Island-Schweine) oder deren Wachstumshormonrezeptor genetisch inaktiviert wurde (z. B. in 10fach genetisch veränderten Schweinen der Fa. Revivicor/United Therapeutics, USA). Mit der ersten klinischen Pilotstudie an terminal Herzkranken wird in Deutschland in ca. 2 Jahren gerechnet. Graphic abstract


Genetic diversity, growth and heart function of Auckland Island pigs, a potential source for organ xenotransplantation

April 2024

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111 Reads

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5 Citations

Xenotransplantation

One of the prerequisites for successful organ xenotransplantation is a reasonable size match between the porcine organ and the recipient's organ to be replaced. Therefore, the selection of a suitable genetic background of source pigs is important. In this study, we investigated body and organ growth, cardiac function, and genetic diversity of a colony of Auckland Island pigs established at the Center for Innovative Medical Models (CiMM), LMU Munich. Male and female Auckland Island pig kidney cells (selected to be free of porcine endogenous retrovirus C) were imported from New Zealand, and founder animals were established by somatic cell nuclear transfer (SCNT). Morphologically, Auckland Island pigs have smaller body stature compared to many domestic pig breeds, rendering their organ dimensions well‐suited for human transplantation. Furthermore, echocardiography assessments of Auckland Island pig hearts indicated normal structure and functioning across various age groups throughout the study. Single nucleotide polymorphism (SNP) analysis revealed higher runs of homozygosity (ROH) in Auckland Island pigs compared to other domestic pig breeds and demonstrated that the entire locus coding the swine leukocyte antigens (SLAs) was homozygous. Based on these findings, Auckland Island pigs represent a promising genetic background for organ xenotransplantation.




Citations (25)


... Längin et al. highlighted the progress in orthotopic pig heart transplantation in non-human primates [4]. Since the first study in 1994, it has been possible to increase the survival time of orthotopically transplanted pig hearts from 39 to 59 to 195 and finally to 264 days. ...

Reference:

Role of a Porcine Herpesvirus, PCMV/PRV, in Xenotransplantation
Progress in Orthotopic Pig Heart Transplantation in Nonhuman Primates

Transplant International

... Various PASylated biologics have been produced in diverse commercially established expression platforms such as E. coli [123], P. fluorescens [59], Gram-positive bacteria like C. glutamicum [121], yeast [122] as well as mammalian cell culture (CHO and HEK cells) [124]. PASylation is safe and well tolerated in animals as tested in many different species from rodents to cynomolgus monkeys [125], rhesus macaques [126] and baboons [127]. Applications of PASylation technology published so far range from the PK optimization of tumor imaging agents [128][129][130] to the half-life extension of wellknown biologics, including cytokines [131][132][133], enzymes [124,134], antibody fragments [58,135,136], alternative binding proteins [137][138][139], peptides [140,141] and various peptide/protein hormones [123,142,143], to the shielding of nanoparticles [144,145] and liposomes [146]. ...

Combination of Anti-CD40 and Anti-CD40L Antibodies as Co-Stimulation Blockade in Preclinical Cardiac Xenotransplantation

... This can be addressed by generating source pigs lacking the major carbohydrate antigens and expressing one or several human complement pathway regulatory proteins (see below). However, coagulation dysregulation can still occur after xenogeneic organ transplantation due to incompatibilities between membranebound factors on porcine endothelial cells and soluble components in human/NHP blood [3]. An example is the thrombin-thrombomodulin (THBD) interaction. ...

An Approach to Controlling Inflammation and Coagulation in Pig-to-Baboon Cardiac Xenotransplantation

Xenotransplantation

... September 2024 | Volume 37 | Article 13607 complement inhibitory drugs (cobra venom factor or C1 esterase inhibitor) and various immunomodulatory drugs (interleukin (IL) 1 and IL6 receptor blockers, tumor necrosis factor (TNF) α inhibitors) have been added to existing immunosuppressive regimens [23,[25][26][27]62]. Furthermore, glycocalyx shedding, a surrogate parameter for endothelial dysfunction and inflammation, was only marginal under anti-inflammatory therapy [63]. It remains unclear, however, to what extend each of these anti-inflammatory substances contribute: for instance, IL6 receptor blockers have been shown to bind to baboon but not to pig IL6 receptors. ...

The Endothelial Glycocalyx in Pig-to-Baboon Cardiac Xenotransplantation—First Insights

... With the rise in pig-to-human surgeries, developing specific guidelines for these procedures is crucial to ensuring the efficacy of xenotransplantation [68]. Researchers have identified that transthoracic echocardiography may become increasingly valuable for size matching in cardiac xenotransplantation [69]. ...

Transthoracic echocardiography is a simple tool for size matching in cardiac xenotransplantation
  • Citing Article
  • May 2024

Xenotransplantation

... Auckland Island pigs are ideal organ donors for xenotransplantation because of their optimal organ size for humans [23]. Therefore, there is no need for a knock-out of the porcine growth factor receptor as is performed with other pig breeds [24]. ...

Genetic diversity, growth and heart function of Auckland Island pigs, a potential source for organ xenotransplantation

Xenotransplantation

... However, successful transplantation in patients who are too sick to be listed on a regular waitlist will be difficult. Given the current results, designing trials in xenotransplantation presents immense challenges and multidisciplinary collaborative efforts are needed to overcome these hurdles and develop an ethical path forward towards first studies [25]. The patients who willingly accept the risks associated with participating in xenotransplant trials, which could potentially include accelerated death, will be true heroes. ...

Research Opportunities and Ethical Considerations for Heart and Lung Xenotransplantation Research: A report from a National Heart, Lung, and Blood Institute workshop

American Journal of Transplantation

... 17 Given the cohabitation of humans and pigs for 10 000 years, the transmission of unknown pathogens is considered unlikely. 18 Lastly, methods for screening animals and organs, monitoring recipients, and antimicrobial prophylaxis have improved. 19 These developments underscore the need for a critical reassessment of recommendations by regulatory and ethical bodies. ...

Does Xenotransplantation Offer a Large Benefit for Human Patients?—A Reply
  • Citing Article
  • February 2024

The Thoracic and Cardiovascular Surgeon

... Byrne and McGregor assumed that an increased sensitivity of the porcine heart to ischemia/reperfusion injury was a major contributor to PCXD [36]. Although PCXD was overcome in the heterotopic thoracic xHTx model [43] and in singular cases of orthotopic xHTx experiments [21], consistent survival could not be achieved using this conservation technique. In 2016, Steen et al. developed a cold, non-ischemic preservation method with an oxygenated, hyperoncotic, erythrocyte containing cardioplegic solution, with which pig hearts were successfully preserved for up to 24 h [44]. ...

Hemodynamics in pig‐to‐baboon heterotopic thoracic cardiac xenotransplantation: Recovery from perioperative cardiac xenograft dysfunction and impairment by cardiac overgrowth

Xenotransplantation

... The message they took home (cf. Kamla et al., 2023;Schmoeckel et al., 2023) from this meeting was that researchers expressed optimism about imminent breakthroughs in xenotransplantation. They discussed plans for another pig-heart-to-human transplant in Germany, with more to follow in the USA. ...

Current status of cardiac xenotransplantation

The Thoracic and Cardiovascular Surgeon