Ameliorating Atopy by Compensating Micronutritional Deficiencies in Immune Cells: A Double-Blind Placebo-Controlled Pilot Study.

BACKGROUND: Functional iron deficiency facilitates allergy development and amplifies the symptom burden in people experiencing allergies. Previously we selectively delivered micronutrients to immune cells with ß-lactoglobulin as carrier (holoBLG), resulting in immune resilience and allergy prevention. OBJECTIVE: The clinical efficacy of a food for special medical purposes-lozenge containing ß-lactoglobulin with iron, polyphenols, retinoic acid, and zinc (holoBLG lozenge) was assessed in allergic women. METHODS: In a randomized, double-blind, placebo-controlled pilot study, grass- and/or birch pollen-allergic women (n = 51) were given holoBLG or placebo lozenges over 6 months. Before and after dietary supplementation, participants were nasally challenged and the blood was analyzed for immune and iron parameters. Daily symptoms, medications, pollen concentrations, and well-being were recorded by an electronic health application. RESULTS: Total nasal symptom score after nasal provocations improved by 42% in the holoBLG group versus 13% in the placebo group. The combined symptom medication score during the birch peak and entire season as well as the entire grass pollen season improved in allergic subjects supplemented with the holoBLG lozenge by 45%, 31%, and 40%, respectively, compared with the placebo arm. Participants ingesting the holoBLG lozenge had improved iron status with increased hematocrit values, decreased red cell distribution width, and higher iron levels in circulating CD14+ cells compared with the placebo group. CONCLUSIONS: Targeted micronutrition with the holoBLG lozenge seemed to be effective in elevating the labile iron levels in immune cells and reducing the symptom burden in allergic women in this pilot study. The underlying allergen-independent mechanism provides evidence that dietary nutritional supplementation of the immune system is one of the ways to combat atopy.

Micronutritional supplementation with a holoBLG-based FSMP (food for special medical purposes)-lozenge alleviates allergic symptoms in BALB/c mice: Imitating the protective farm effect.

BACKGROUND: Previously, the protective farm effect was imitated using the whey protein beta-lactoglobulin (BLG) that is spiked with iron-flavonoid complexes. Here, we formulated for clinical translation a lozenge as food for special medical purposes (FSMP) using catechin-iron complexes as ligands for BLG. The lozenge was tested in vitro and in a therapeutical BALB/c mice model. METHODS: Binding of iron-catechin into BLG was confirmed by spectroscopy and docking calculations. Serum IgE binding of children allergic or tolerating milk was assessed to loaded (holo-) versus empty (apo-) BLG and for human mast cell degranulation. BLG and Bet v 1 double-sensitized mice were orally treated with the holoBLG or placebo lozenge, and immunologically analysed after systemic allergen challenge. Human PBMCs of pollen allergic subjects were flow cytometrically assessed after stimulation with apoBLG or holoBLG using catechin-iron complexes as ligands. RESULTS: One major IgE and T cell epitope were masked by catechin-iron complexes, which impaired IgE binding of milk-allergic children and degranulation of mast cells. In mice, only supplementation with the holoBLG lozenge reduced clinical reactivity to BLG and Bet v 1, promoted Tregs, and suppressed antigen presentation. In allergic subjects, stimulation of PBMCs with holoBLG led to a significant increase of intracellular iron in circulating CD14+ cells with significantly lower expression of HLADR and CD86 compared to their stimulation with apoBLG. CONCLUSION: The FSMP lozenge targeted antigen presenting cells and dampened immune activation in human immune cells and allergic mice in an antigen-non-specific manner, thereby conferring immune resilience against allergic symptoms.

First evaluation of a symbiotic food supplement in an allergen exposure chamber in birch pollen allergic patients.

BACKGROUND: Allergic rhinitis/rhinoconjunctivitis is the most common immune disease worldwide, but still largely underestimated, underdiagnosed, and undertreated. Dysbiosis and reduced microbial diversity is linked to the development of allergies, and the immunomodulatory effects of pro- and prebiotics might be used to counteract microbiome dysbiosis in allergy. Adequate symbiotic (multi-strain pro-, plus prebiotic) supplementation can be suggested as a complementary approach in the management of allergic rhinitis. OBJECTIVE: The effects of the daily intake of a symbiotic food supplement (combination of Lactobacillus acidophilus NCFM and Bifidobacterium lactis BL-04 with Fructo-Oligosaccharides) for 4 months in birch pollen allergic rhinoconjunctivitis patients were investigated for the first time in an allergen exposure chamber (AEC) allowing standardised, reproducible pollen exposure before and after intake. METHODS: Eligible patients were exposed to birch pollen (8000 pollen/m³ for 120 min) at the GA2LEN AEC, at baseline (V1) and final visit (V3) outside the season. The Total Symptom Score (TSS) and the scores for nose, eye, bronchial system, and others were evaluated every 10 min during exposure. Other secondary endpoints were the changes in well-being, Peak Nasal Inspiratory Flow (PNIF), lung function parameters, and safety. Co-primary endpoints were differences in Total Nasal Symptom Score (TNSS) and TSS after 120 min of exposure between both visits. Temporal evolution of symptom scores were analysed in an exploratory way using linear mixed effects models. RESULTS: 27 patients (mean age 45 years, 15% male) completed the study. Both co-primary endpoints showed significant improvement after intake of the symbiotic. Median TNSS and TSS were decreased 50% and 80% at 120 min (adjusted p-value = 0.025 and p < 0.01 respectively).All four symptom scores and the personal well-being, improved to a clinically relevant extent over time, visible by a weaker increase in symptoms during 120 min of the final birch pollen exposure. No relevant differences were observed for PNIF, PEF, and spirometry. There were no airway obstructions or lung restrictions before and after both exposures. Late phase reactions after exposure were reduced after V3, documenting a better birch pollen tolerability of the patients. The safety and tolerability profile of the symbiotic food supplement was excellent, no adverse events (AEs) were observed. CONCLUSIONS: This first evaluation of a symbiotic food supplement in an AEC in rhinoconjunctivitis patients with or without asthma induced by birch pollen revealed a significant beneficial effect, harnessing significant improvements of symptoms and well-being while maintaining an excellent safety and tolerability profile.

Shortened up-dosing with sublingual immunotherapy drops containing tree allergens is well tolerated and elicits dose-dependent clinical effects during the first pollen season.

BACKGROUND: This study compared a rapid home-based up-dosing schedule for sublingual immunotherapy (SLIT) drops containing tree pollen allergens with two previously established schedules. Furthermore, the clinical effect of the SLIT was investigated with respect to patients' first pollen season under treatment. METHODS: In this open-label, prospective, patient-preference, non-interventional study, local and systemic reactions were compared between three up-dosing groups using a SLIT formulation containing birch, alder, and hazel pollen extracts (ORALVAC® Compact Bäume). Clinical improvement after patients' first season under treatment was analysed using symptom scores, ARIA classification, symptom control, and the use of symptomatic medication and was compared with data from the previous, pre-treatment pollen season. As the real-life study design allowed no placebo group, the late-treated patients (co-seasonal) served as a control, and crowd-sourced symptom data from persons with hay fever were used from a free web-based online diary. RESULTS: In 33 study centres in Germany and Austria, 164 patients were included. The treatment was well tolerated, without difference between the groups during the up-dosing phase. At the end of the assessment, 96.1% rated the tolerability of the treatment as good or very good. Local reactions were mostly mild in severity and no serious adverse events occurred. Symptom scores decreased from the 2016 pollen season to the 2017 pollen season. As for the ARIA classification, 79.0% of patients had persistent, moderate-to-severe rhinitis before treatment, but only 18.6% had the same classification after treatment. In all, 62.4% of patients achieved symptom control, and 34.3% of patients required no symptomatic medication after treatment. The rhinoconjunctivitis score was 34.4% lower for pre-seasonal treatment initiation than for the control group. Crowd-sourced symptom load indices showed that the 2016 season caused slightly more symptoms; however, it is assumed that this difference of 0.3-0.5 (score range 0-10) was of less clinical relevance. CONCLUSION: The treatment administered using the rapid home-based up-dosing schedule was safe and well tolerated. Symptom relief and reduction in medication use were observed during the first pollen season with SLIT. TRIAL REGISTRATION NUMBER: NCT03097432 (clinicaltrials.gov).

Author Correction: Consistent success in life-supporting porcine cardiac xenotransplantation.

In this Letter, Mayuko Kurome and Valeri Zakhartchenko have been added to the author list (affiliated with Institute of Molecular Animal Breeding and Biotechnology, Gene Center, LMU Munich, Munich, Germany). The author list and 'Author contributions' section have been corrected online; see accompanying Amendment.

Consistent success in life-supporting porcine cardiac xenotransplantation.

Heart transplantation is the only cure for patients with terminal cardiac failure, but the supply of allogeneic donor organs falls far short of the clinical need1-3. Xenotransplantation of genetically modified pig hearts has been discussed as a potential alternative4. Genetically multi-modified pig hearts that lack galactose-α1,3-galactose epitopes (α1,3-galactosyltransferase knockout) and express a human membrane cofactor protein (CD46) and human thrombomodulin have survived for up to 945 days after heterotopic abdominal transplantation in baboons5. This model demonstrated long-term acceptance of discordant xenografts with safe immunosuppression but did not predict their life-supporting function. Despite 25 years of extensive research, the maximum survival of a baboon after heart replacement with a porcine xenograft was only 57 days and this was achieved, to our knowledge, only once6. Here we show that α1,3-galactosyltransferase-knockout pig hearts that express human CD46 and thrombomodulin require non-ischaemic preservation with continuous perfusion and control of post-transplantation growth to ensure long-term orthotopic function of the xenograft in baboons, the most stringent preclinical xenotransplantation model. Consistent life-supporting function of xenografted hearts for up to 195 days is a milestone on the way to clinical cardiac xenotransplantation7.

Virus-like particles (VLP) in prophylaxis and immunotherapy of allergic diseases.

BACKGROUND: Apart from active allergen avoidance, immunotherapy is regarded as the most effective form of treatment available for type I allergies. Such treatments involve the administration of allergen preparations in various forms and by various routes. Virus-like particles (VLPs) offer a very effective platform for immunization with the allergen and are characterized by high immunogenicity, low allergenicity and high clinical efficacy. Formulations that include Toll-like receptor ligands, T cell stimulatory epitopes and/or depot-forming adjuvants appear to enhance activation of the relevant immune cells. Short nucleotide sequences including CpG motifs have also been intensively explored as potent stimulators of dendritic cells and B cells. METHODS: The present paper is based on a systematic literature search in PubMed and MEDLINE, and focuses on the pertinent immunological processes and on clinical data relating to use of VLPs and CpG motifs for the treatment of allergic rhinitis (AR). RESULTS: Many published studies have reported positive clinical results following administration of VLPs, either alone or in combination with CpG motifs and, in some cases, even in the absence of the allergen-specific allergen. CONCLUSIONS: These results indicate that VLPs modulate immune responses in ways which underline their exceptional promise as a platform for the immunotherapy of allergic disorders. However, clinical evaluations remain limited, and further large-scale and longer-term studies will be necessary to substantiate the efficacy and safety of these novel therapies.

Distribution of Porcine Cytomegalovirus in Infected Donor Pigs and in Baboon Recipients of Pig Heart Transplantation.

The porcine cytomegalovirus (PCMV) is a herpesvirus that may pose a risk for xenotransplantation using pig cells, tissues, or organs. Here, three orthotopic pig heart transplantations into baboons were studied. To detect PCMV, a real-time PCR and a Western blot assay based on four PCMV protein sequences, including two tegument proteins, were used. The transmission of PCMV from the donor pig to the recipient baboon was found in two cases, despite PCMV not being detected in the blood of the donor pigs by real-time PCR. Although it was not in the blood, PCMV was detected in different organs of the donor pigs, and in sibling animals. Immunohistochemistry using an antiserum that is specific for PCMV detected virus protein-expressing cells in all of the organs of the recipient baboon, most likely representing disseminated pig cells. Therefore, for the first time, the distribution of PCMV in organs of the donor pigs and the recipient baboons was described. In addition, baboon cytomegalovirus (BaCMV) was found activated in the recipient, and a screening for hepatitis E virus (HEV) and porcine lymphotropic herpesviruses (PLHV) was performed. For the first time, a cross-reactivity between antibodies directed against PCMV and BaCMV was found.

Large-Animal Biventricular Working Heart Perfusion System with Low Priming Volume-Comparison between in vivo and ex vivo Cardiac Function.

BACKGROUND: Existing large-animal, ex vivo, cardiac perfusion models are restricted in their ability to establish an ischemia/reperfusion condition as seen in cardiac surgery or transplantation. Other working heart systems only challenge one ventricle or require a substantially larger priming volume. We describe a novel biventricular cardiac perfusion system with reduced priming volume. METHODS: Juvenile pig hearts were cardiopleged, explanted, and reperfused ex vivo after 150 minutes of cold ischemia. Autologous whole blood was used as perfusate (minimal priming volume 350 mL). After 15 minutes of Langendorff perfusion (LM), the system was switched into a biventricular working mode (WM) and studied for 3 hours. RESULTS: During reperfusion, complete unloading of both ventricles and constant-pressure coronary perfusion was achieved. During working mode perfusion, the preload and afterload pressure of both ventricles was controlled within the targeted physiologic range. Functional parameters such as left ventricular work index were reduced in ex vivo working mode (in vivo: 787 ± 186 vs. 1 h WM 498 ± 66 mm Hg·mL/g·min; p < 0.01), but remained stable throughout the following study period (3 h WM 517 ± 103 mm Hg·mL/g·min; p = 0.63). Along with the elevated workload during WM, myocardial metabolism and oxygen consumption increased compared with LM (0.021 ± 0.08 vs. 0.06 ± 0.01 mL/min/g; 1 h after reperfusion). Histologic examination of the myocardium revealed no structural damage. CONCLUSION: In the ex vivo perfusion system, stable hemodynamic and metabolic conditions can be established for a period of 3 hours while functional and blood parameters are easily accessible. Moreover, because of the minimal priming volume, the novel ex vivo cardiac perfusion circuit allows for autologous perfusion, using the limited amount of blood available from the organ donating animal.

Hemodynamic and perioperative management in two different preclinical pig-to-baboon cardiac xenotransplantation models.

BACKGROUND: The perioperative phase of preclinical cardiac xenotransplantations significantly affects the experimental outcome. Moderate or even severe hemodynamic and respiratory impairment occurs frequently in baboons after receiving a cardiac transplant. The perioperative management of such postoperative instability is very demanding, especially in the experimental setting. We compared perioperative changes of hemodynamic and laboratory findings during orthotopic and heterotopic thoracic cardiac xenotransplantations and describe our monitoring, treatment and intensive care. METHODS: Twenty-eight pig-to-baboon cardiac xenotransplantations were performed using either the orthotopic (oHTx, n=5) or heterotopic thoracic (htHTx; n=23) technique. In both techniques, cardioplegia and an intraoperative cardiopulmonary bypass (CPB) were required. Preoperatively, intensive care (eg, transfusions, catecholamine therapy) was provided and fast extubation was targeted. A central venous catheter, a femoral arterial thermodilution catheter, a telemetric pressure transmitter and transthoracic echocardiography were used to monitor the animal. Baboon jackets with a tethering system were used to continuously apply medication postoperatively and permit blood sampling, also after extubation of the animal and transfer into the cage. Perioperative survival, hemodynamics, catecholamine doses, respiratory function and weaning from respirator were compared. Perioperative organ damage was evaluated based on laboratory findings 12 hours after transplantation. RESULTS: Recipients could be weaned from CPB in the 20 htHTx and all five oHTx experiments, and three htHTx procedures were terminated during the operation. The time of cardiopulmonary bypass was significantly lower in the heterotopic group (oHTx median 171 [157-193] minutes; htHTx median 144 [100-190] minutes; P=.02). In 17 htHTx procedures, no inotropics were used, whereas epinephrine had to be administered in four of the five oHTx experiments; the mean time of catecholamine support was longer in the oHTx group (oHTx 972±348 minutes vs htHTx 111±92 minutes; P<.01). After htHTx, weaning off the respirator was possible in 19 of 20 cases (one died due to pneumothorax). After oHTx, three of the five baboons could be weaned off the respirator; in these cases, the arterial saturation was higher compared with the extubated baboons after htHTx (oHTx 99±1% vs htHTx 91±4%, P=.01). Intraoperative blood loss was similar between the two groups, and hemostasis was impaired after all procedures, but relevant postoperative bleeding never occurred. CONCLUSION: Intensive intra- and postoperative monitoring and care is required in both transplantation techniques as a requirement for successful weaning from CPB and respirator. After htHTx, the animals needed less catecholamines and were hemodynamically more stable. Even though pulmonary function was often impaired after htHTx, weaning from the respirator and extubation was more successful in this group.

Complement C3 inhibitor Cp40 attenuates xenoreactions in pig hearts perfused with human blood.

BACKGROUND: The complement system plays a crucial role in acute xenogeneic reactions after cardiac transplantation. We used an ex vivo perfusion model to investigate the effect of Cp40, a compstatin analog and potent inhibitor of complement at the level of C3. METHODS: Fifteen wild-type pig hearts were explanted, cardiopleged, and reperfused ex vivo after 150 minutes of cold ischemia. Hearts were challenged in a biventricular working heart mode to evaluate cardiac perfusion and function. In the treatment group (n=5), the complement cascade was blocked at the level of C3 using Cp40, using diluted human blood. Untreated human and porcine blood was used for controls. RESULTS: Throughout the perfusion, C3 activation was inhibited when Cp40 was used (mean of all time points: 1.11 ± 0.34% vs 3.12 ± 0.48% control activation; P<.01). Compared to xenoperfused controls, the cardiac index improved significantly in the treated group (6.5 ± 4.2 vs 3.5 ± 4.8 mL/min/g; P=.03, 180 minutes perfusion), while the concentration of lactate dehydrogenase as a maker for cell degradation was reduced in the perfusate (583 ± 187 U/mL vs 2108 ± 1145 U/mL, P=.02). Histological examination revealed less hemorrhage and edema, and immunohistochemistry confirmed less complement fragment deposition than in untreated xenoperfused controls. CONCLUSIONS: Cp40 efficiently prevents C3 activation of the complement system, resulting in reduced cell damage and preserved function in wild-type porcine hearts xenoperfused ex vivo. We suggest that this compstatin analog, which blocks all main pathways of complement activation, could be a beneficial perioperative treatment in preclinical and in future clinical xenotransplantation.

Ascending Aortic Disease is Associated with Earlier Menopause and Shorter Reproductive Life Span.

OBJECTIVES: The incidence of cardiovascular morbidity and mortality in premenopausal women is comparatively low, but increases sharply after menopause. The principal aim of this study was to determine whether women with ascending aortic disease (AAD) have a different reproductive history from that of an age-matched control group. METHODS: In this retrospective study, women who had undergone ascending aortic aneurysm (AscAA) repair between 2000 and 2010 were asked to complete a questionnaire concerning risk factors and reproductive history. Data from 142 women with AAD were evaluated, and a subgroup (n = 64) with AscAA ≥5 cm was analyzed and compared to an age-matched random control group without known aortic diseases. RESULTS: Almost all women were menopausal at the time of the questionnaire (98.4% vs. 90.6%, AscAA ≥5 cm subcohort vs. control, p = 0.12) and all subjects presented with a comparable age of menarche (13.7 ± 2.6 years vs. 14.2 ± 1.8 years, AscAA ≥5 cm subcohort vs. control, log-rank 0.04, p = 0.84). However, mean menopausal age was significantly lower in the case subcohort than in controls (48.1 ± 4.8 years vs. 50.6 ± 5.8 years, AscAA ≥5 cm subcohort vs. control, log-rank 8.35, p = 0.004), and reproductive life span was correspondingly shorter (34.2 ± 5.2 years vs. 36.2 ± 5.7 years, p = 0.04). Furthermore, hypertension was more prevalent in women with AscAA ≥5 cm compared to controls (89.1% vs. 61.9%, AscAA ≥5 cm subcohort vs. control, p < 0.001). CONCLUSION: Women who experience menopause at an earlier age than the regional mean could profit from screening for cardiovascular disease in general and particularly for AAD. Screening would enable early aneurysm detection and could, therefore, reduce morbidity and mortality.

Ribonuclease (RNase) Prolongs Survival of Grafts in Experimental Heart Transplantation.

BACKGROUND: Cell damage, tissue and vascular injury are associated with the exposure and release of intracellular components such as RNA, which promote inflammatory reactions and thrombosis. Based on the counteracting anti-inflammatory and cardioprotective functions of ribonuclease A (RNase A) in this context, its role in an experimental model of heart transplantation in rats was studied. METHODS AND RESULTS: Inbred BN/OrlRj rat cardiac allografts were heterotopically transplanted into inbred LEW/OrlRj rats. Recipients were intravenously treated every other day with saline or bovine pancreatic RNase A (50 µg/kg). Toxic side effects were not found (macroscopically and histologically). Heart tissue flow cytometry and quantitative morphological analyses of explanted hearts at postoperative day 1 or postoperative day 4 showed reduced leukocyte infiltration, edema, and thrombus formation in RNase A-treated rats. In allogeneic mixed lymphocyte reactions, RNase A decreased the proliferation of effector T cells. RNase A treatment of rats resulted in prolonged median graft survival up to 10.5 days (interquartile range 1.8) compared to 6.5 days (interquartile range 1.0) in saline treatment (P=0.001). Treatment of rats with a new generated (recombinant) human pancreatic RNase 1 prolonged median graft survival similarly, unlike treatment with (recombinant) inactive human RNase 1 (each 50 µg/kg IV every other day, 11.0 days, interquartile range 0.3, versus 8.0 days, interquartile range 0.5, P=0.007). CONCLUSIONS: Upon heart transplantation, RNase administration appears to present a promising and safe drug to counteract ischemia/reperfusion injury and graft rejection. Furthermore, RNase treatment may be considered in situations of critical reperfusion after percutaneous coronary interventions or in cardiac surgery using the heart-lung machine.

Pre-clinical heterotopic intrathoracic heart xenotransplantation: a possibly useful clinical technique.

BACKGROUND: As a step towards clinical cardiac xenotransplantation, our experimental heterotopic intrathoracic xenotransplantation model offers a beating and ejecting donor heart while retaining the recipient's native organ as a backup in case of graft failure. Clinically applicable immunosuppressive regimens (IS) were investigated first, then treatments known to be effective in hypersensitized patients or those with recalcitrant rejection reactions. METHODS: Consecutive experiments were carried out between 2009 and 2013. Twenty-one genetically modified pigs (GGTA1-knockout/hCD46/± thrombomodulin, in one case HLA-E instead) were used as donors. In all experiments, two cycles of immunoabsorption reduced preformed antibodies. Recipient baboons were divided into two groups according to IS regimen: In group one (n = 10), pre-treatment started either one (anti-CD20) or four weeks (anti-CD20 plus the proteasome inhibitor bortezomib) prior to transplantation. The extended conventional (as for allotransplantation) immunosuppressive maintenance regimen included anti-thymocyte globuline, tacrolimus, mycophenolate mofetil, methylprednisolone and weekly anti-CD20. In group two (n = 11), myeloablative pre-treatment as in multiple myeloma patients (long and short regimens) was added to extended conventional IS; postoperative total thoracic and abdominal lymphoid irradiation (TLI; single dose of 600 cGY) was used to further reduce antibody-producing cells. RESULTS: In the perioperative course, the surgical technique was safely applied: 19 baboons were weaned off extracorporeal circulation and 17 extubated. Nine animals were lost in the early postoperative course due to causes unrelated to surgical technique or IS regimen. Excluding these early failures, median graft survival times of group 1 and 2 were 18.5 (12-50) days and 16 (7-35) days. Necropsy examination of group 1 donor organs revealed hypertrophy of the left ventricular wall in the six longer-lasting grafts; myocardial histology confirmed pre-clinical suspicion of humoral rejection, which was not inhibited by the extended conventional IS including intensified treatments, and signs of thrombotic microangiopathy. Grafts of group 2 presented with only mild-to-moderate features of humoral rejection and thrombotic microangiopathy, except in one case of delayed rejection on day 17. The other experiments in this group were terminated because of untreatable pulmonary oedema, recurring ventricular fibrillation, Aspergillus sepsis, as well as a combination of a large donor organ and late toxic side effects due to TLI. CONCLUSIONS: Longer-term results were difficult to achieve in this model due to the IS regimens used. However, we conclude that heterotopic intrathoracic heart transplantation may be an option for clinical xenotransplantation.

Engraftment and reversal of diabetes after intramuscular transplantation of neonatal porcine islet-like clusters.

BACKGROUND: Intraportal infusion is currently the method of choice for clinical islet cell transplantation but suffers from poor efficacy. As the liver may not represent an optimal transplantation site for Langerhans islets, we examined the potential of neonatal porcine islet-like clusters (NPICCs) to engraft in skeletal muscle as an alternative transplantation site. METHODS: Neonatal porcine islet-like clusters were isolated from 2- to 5-day-old piglets and either transplanted under the kidney capsule (s.k.) or injected into the lower hindlimb muscle (i.m.) of streptozotocin-diabetic NOD-SCID IL2rγ(-/-) (NSG) mice. Survival, vascularization, maturation, and functional activity were analyzed by intraperitoneal glucose tolerance testing and immunohistochemical analyses. RESULTS: Intramuscular transplantation of NPICCs resulted in development of normoglycemia and restored glucose homeostasis. Time to reversal of diabetes and glucose tolerance (AUC glucose and AUC insulin) did not significantly differ as compared to s.k. transplantation. Intramuscular grafts exhibited rapid neovascularization and graft composition with cytokeratin-positive ductal cells and beta cells at post-transplant weeks 2 and 8 and after establishment of normoglycemia was comparable in both groups. CONCLUSIONS: Intramuscular injection represents a minimally invasive but efficient alternative for transplantation of NPICCs and, thus, offers an attractive alternative site for xenotransplantation approaches. These findings may have important implications for improving the outcome and the monitoring of pig islet xenotransplantation.

Xenotransplantation of Cells, Tissues, Organs and the German Research Foundation Transregio Collaborative Research Centre 127.

Human organ transplantation is the therapy of choice for end-stage organ failure. However, the demand for organs far exceeds the donation rate, and many patients die while waiting for a donor. Clinical xenotransplantation using discordant species, particularly pigs, offers a possible solution to this critical shortfall. Xenotransplantation can also increase the availability of cells, such as neurons, and tissues such as cornea, insulin producing pancreatic islets and heart valves. However, the immunological barriers and biochemical disparities between pigs and primates (human) lead to rejection reactions despite the use of common immunosuppressive drugs. These result in graft vessel destruction, haemorrhage, oedema, thrombus formation, and transplant loss. Our consortium is pursuing a broad range of strategies to overcome these obstacles. These include genetic modification of the donor animals to knock out genes responsible for xenoreactive surface epitopes and to express multiple xenoprotective molecules such as the human complement regulators CD46, 55, 59, thrombomodulin and others. We are using (new) drugs including complement inhibitors (e.g. to inhibit C3 binding), anti-CD20, 40, 40L, and also employing physical protection methods such as macro-encapsulation of pancreatic islets. Regarding safety, a major objective is to assure that possible infections are not transmitted to recipients. While the aims are ambitious, recent successes in preclinical studies suggest that xenotransplantation is soon to become a clinical reality.

De novo sirolimus with low-dose tacrolimus versus full-dose tacrolimus with mycophenolate mofetil after heart transplantation--8-year results.

BACKGROUND: Although acute cellular rejection after heart transplantation (HTX) can be controlled by full-dose calcineurin inhibitor (CNI)-based immunosuppressive regimens, cardiac allograft vasculopathy (CAV), nephrotoxicity, and malignancy remain ongoing problems. To evaluate the potential beneficial effects of sirolimus and CNI reduction, we compared de novo low-dose tacrolimus and sirolimus with standard tacrolimus and mycophenolate mofetil (MMF)-based immunosuppression after HTX. METHODS: We analyzed a long-term follow-up cohort of 126 patients who underwent HTX during the period 1998-2005 and received either de novo low-dose tacrolimus/sirolimus (lowTAC/SIR; n = 61) or full-dose tacrolimus/MMF (TAC/MMF; n = 64). RESULTS: Freedom from treatment switch was less in the low TAC/SIR group than in the TAC/MMF group (51.7% vs 73.0%, p = 0.038) 8 years after HTX. Freedom from acute rejection was 90.6% in the low TAC/SIR group vs 80.3% in the TAC/MMF group (p = 0.100). There was no difference in freedom from International Society for Heart and Lung Transplantation CAV grade ≥ 1 (55.4% vs 60.0%, p = 0.922), time until CAV diagnosis (4.2 ± 2.0 years vs 3.2 ± 2.4 years, p = 0.087), and CAV severity (p = 0.618). The benefit of reduced early maximum creatinine for low TAC/SIR treatment (1.8 ± 0.9 mg/dl vs 2.4 ± 1.1 mg/dl in TAC/MMF group, p < 0.001) did not continue 5 years and 8 years after HTX (1.4 ± 0.4 mg/dl vs 1.7 ± 1.2 mg/dl, p = 0.333, and 1.6 ± 1.1 mg/dl vs 1.6 ± 0.8 mg/dl, p = 0.957). The trend for superior survival at 5 years with low TAC/SIR treatment (93.1% vs 81.3% in TAC/MMF group, p = 0.051) could not be confirmed after 8 years (84.7% vs 75.0%, p = 0.138). Multivariate analysis at 8 years did not reveal any benefit of low TAC/SIR treatment. CONCLUSIONS: Reduction of de novo CNI did not result in superior long-term renal function. Low-dose mechanistic target of rapamycin inhibition did not achieve any benefit in CAV prevention compared with full-dose TAC/MMF after HTX.

Applying the Gender Lens to Risk Factors and Outcome after Adult Cardiac Surgery.

BACKGROUND: Applying the gender lens to risk factors and outcome after adult cardiac surgery is of major clinical interest, as the inclusion of sex and gender in research design and analysis may guarantee more comprehensive cardiovascular science and may consecutively result in a more effective surgical treatment as well as cost savings in cardiac surgery. METHODS: We have reviewed classical cardiovascular risk factors (diabetes, arterial hypertension, hyperlipidemia, smoking) according to a gender-based approach. Furthermore, we have examined comorbidities such as depression, renal insufficiency, and hormonal influences in regard to gender. Gender-sensitive economic aspects have been evaluated, surgical outcome has been analyzed, and cardiovascular research has been considered from a gender perspective. RESULTS: The influence of typical risk factors and outcome after cardiac surgery has been evaluated from a gender perspective, and the gender-specific distribution of these risk factors is reported on. The named comorbidities are listed. Economic aspects demonstrated a gender gap. Outcome after coronary and valvular surgeries as well as after heart transplantation are displayed in this regard. Results after postoperative use of intra-aortic balloon pump are shown. Gender-related aspects of clinical and biomedical cardiosurgical research are reported. CONCLUSIONS: Female gender has become an independent risk factor of survival after the majority of cardiosurgical procedures. Severely impaired left ventricular ejection fraction independently predicts survival in men, whereas age does in females.