Alexander Horke

Decellularized fresh homografts for pulmonary valve replacement: a decade of clinical experience

OBJECTIVES Decellularized homografts have shown auspicious early results when used for pulmonary valve replacement (PVR) in congenital heart disease. The first clinical application in children was performed in 2002, initially using pre-seeding with endogenous progenitor cells. Since 2005, only non-seeded, fresh decellularized allografts have been implanted after spontaneous recellularization was observed by several groups. METHODS A matched comparison of decellularized fresh pulmonary homografts (DPHs) implanted for PVR with cryopreserved pulmonary homografts (CHs) and bovine jugular vein conduits (BJVs) was conducted. Patients’ age at implantation, the type of congenital malformation, number of previous cardiac operations and number of previous PVRs were considered for matching purposes, using an updated contemporary registry of right ventricular outflow tract conduits (2300 included conduits, >12 000 patient-years). RESULTS A total of 131 DPHs were implanted for PVR in the period from January 2005 to September 2015. Of the 131, 38 were implanted within prospective trials on DPH from October 2014 onwards and were therefore not analysed within this study. A total of 93 DPH patients (58 males, 35 females) formed the study cohort and were matched to 93 CH and 93 BJV patients. The mean age at DPH implantation was 15.8 ± 10.21 years (CH 15.9 ± 10.4, BJV 15.6 ± 9.9) and the mean DPH diameter was 23.9 mm (CH 23.3 ± 3.6, BJV 19.9 ± 2.9). There was 100% follow-up for DPH, including 905 examinations with a mean follow-up of 4.59 ± 2.76 years (CH 7.4 ± 5.8, BJV 6.4 ± 3.8), amounting to 427.27 patient-years in total (CH 678.3, BJV 553.0). Tetralogy-of-Fallot was the most frequent malformation (DPH 50.5%, CH 54.8%, BJV 68.8%). At 10 years, the rate of freedom of explantation was 100% for DPH, 84.2% for CH (P = 0.01) and 84.3% for BJV (P= 0.01); the rate of freedom from explantation and peak trans-conduit gradient ≥50 mmHg was 86% for DPH, 64% for CH (n.s.) and 49% for BJV (P < 0.001); the rate of freedom from infective endocarditis (IE) was 100% for DPH, 97.3 ± 1.9% within the matched CH patients (P = 0.2) and 94.3 ± 2.8% for BJV patients (P = 0.06). DPH valve annulus diameters converged towards normal Z-values throughout the observation period, in contrast to other valve prostheses (BJV). CONCLUSIONS Mid-term results of DPH for PVR confirm earlier results of reduced re-operation rates compared with CH and BJV.

Elastogenesis at the onset of human cardiac valve development

Semilunar valve leaflets have a well-described trilaminar histoarchitecture, with a sophisticated elastic fiber network. It was previously proposed that elastin-containing fibers play a subordinate role in early human cardiac valve development; however, this assumption was based on data obtained from mouse models and human second and third trimester tissues. Here, we systematically analyzed tissues from human fetal first (4-12 weeks) and second (13-18 weeks) trimester, adolescent (14-19 years) and adult (50-55 years) hearts to monitor the temporal and spatial distribution of elastic fibers, focusing on semilunar valves. Global expression analyses revealed that the transcription of genes essential for elastic fiber formation starts early within the first trimester. These data were confirmed by quantitative PCR and immunohistochemistry employing antibodies that recognize fibronectin, fibrillin 1, 2 and 3, EMILIN1 and fibulin 4 and 5, which were all expressed at the onset of cardiac cushion formation (~week 4 of development). Tropoelastin/elastin protein expression was first detectable in leaflets of 7-week hearts. We revealed that immature elastic fibers are organized in early human cardiovascular development and that mature elastin-containing fibers first evolve in semilunar valves when blood pressure and heartbeat accelerate. Our findings provide a conceptual framework with the potential to offer novel insights into human cardiac valve development and disease.

A Single-Institution Experience With the Ross Operation Over 11 Years

BACKGROUND Although the Ross operation requires double-valve replacement for aortic valve pathology, it is the only autologous, aortic valve replacement available. We report a single-units 11-year experience. METHODS Before August 2006, 467 patients (mean age, 41 +/- 15 years; 358 males) underwent a Ross operation. The right ventricular outflow tract was repaired with a cryopreserved pulmonary homograft. Follow-up was 94.4% complete. RESULTS The 30-day mortality was 0.6%. The Kaplan-Meier survival estimate at 120 months was 94.4% +/- 2.9% (standard error [SE], 0.0146). Reoperation was due to autograft failure in 15 patients (7 repairs, 8 replacements), with a Kaplan-Meier freedom from autograft failure measured as reoperation or regurgitation exceeding grade II at 120 months of 94.2% +/- 2.8% (SE, 0.0142). Homograft replacement, mostly due to stenosis, occurred in 11 patients. Freedom from homograft dysfunction, defined as homograft reoperation or peak homograft gradient of 30 mm Hg or more, at 120 months was 79.3% +/- 7.3% (SE, 0.0372). Freedom from all autograft- and homograft-related reoperations at 120 months was 85.9% +/- 6.3% (SE, 0.0321). Autograft or homograft endocarditis occurred in 8 patients, and 1 patient had simultaneous endocarditis of both valves. CONCLUSIONS Patient survival and freedom from prostheses-related events over 11 years still compares favorably with conventional heart valve prostheses. Mortality and morbidity remain low. Reoperation for autograft or homograft failure is higher than our previous reports, and endocarditis is also evident, 1.9% (9 of 467). Homograft dysfunction is higher in younger recipients.

Decellularized aortic homografts for aortic valve and aorta ascendens replacement

OBJECTIVES The choice of valve prosthesis for aortic valve replacement (AVR) in young patients is challenging. Decellularized pulmonary homografts (DPHs) have shown excellent results in pulmonary position. Here, we report our early clinical results using decellularized aortic valve homografts (DAHs) for AVR in children and mainly young adults. METHODS This prospective observational study included all 69 patients (44 males) operated from February 2008 to September 2015, with a mean age of 19.7 ± 14.6 years (range 0.2–65.3 years). In 18 patients, a long DAH was used for simultaneous replacement of a dilated ascending aorta as an extended aortic root replacement (EARR). Four patients received simultaneous pulmonary valve replacement with DPH. RESULTS Thirty-nine patients (57%) had a total of 62 previous operations. The mean aortic cross-clamp time in isolated cases was 129 ± 41 min. There was 1 conduit-unrelated death. The mean DAH diameter was 22.4 ± 3.7 mm (range, 10–29 mm), the average peak gradient was 14 ± 15 mmHg and the mean aortic regurgitation grade (0.5 = trace, 1 = mild) was 0.6 ± 0.5. The mean effective orifice area (EOA) of 25 mm diameter DAH was 3.07 ± 0.7 cm2. DAH annulus z-values were 1.1 ± 1.1 at implantation and 0.7 ± 1.3 at the last follow-up. The last mean left ventricle ejection fraction and left ventricle end diastolic volume index was 63 ± 7% and 78 ± 16 ml/m2 body surface area, respectively. To date, no dilatation has been observed at any level of the graft during follow-up; however, the observational time is short (140.4 years in total, mean 2.0 ± 1.8 years, maximum 7.6 years). One small DAH (10 mm at implantation) had to be explanted due to subvalvular stenosis and developing regurgitation after 4.5 years and was replaced with a 17 mm DAH without complication. No calcification of the explanted graft was noticed intraoperatively and after histological analysis, which revealed extensive recellularization without inflammation. CONCLUSIONS DAHs withstand systemic circulation, provide outstanding EOA and appear as an alternative to conventional grafts for AVR in young patients. EARR using DAH is a further option in aortic valve disease associated with aorta ascendens dilatation as it avoids the use of any prosthetic material.

Bovine Jugular Veins versus Homografts in the Pulmonary Position: An Analysis across Two Centers and 711 Patients-Conventional Comparisons and Time Status Graphs as a New Approach.

BACKGROUND Various diseases and diversity in implantation ages, together with evolving diagnostic and therapeutic options, hinder comparative evaluations of long-term outcomes for valved conduits used for reconstruction of the right ventricular outflow tract (RVOT). We combined two common evaluation methods to optimally use information obtained by pooling the raw data from two high volume centers, each with very regular follow-up procedures, with the aim of analyzing durability differences between conventional homografts and bovine jugular veins. PATIENTS AND METHODS In the period 1985 to 2012, a total of 444 bovine jugular veins and 267 homografts were implanted, and 6,738 postoperative examinations took place. Evaluations included age-stratified Kaplan-Meier analyses, Cox regression models, and time status graphs, the third showing age-group stratified, time-related frequencies of intact, insufficient, stenotic, both insufficient and stenotic, and postinterventional conduits below the freedom from explantation curve. They take into account interventions, explantations, and the nonterminal character of echocardiographic findings. RESULTS The durability of intact bovine jugular veins in children and young adults is not inferior to that of homografts. Averaged over the first 12 years after implantation, the age groups < 25 years in fact showed advantages for bovine jugular vein recipients. The average fraction of patients younger than 25 years whose conduits were not explanted, postinterventional, stenotic, insufficient, or stenotic and insufficient was at least 10% higher in recipients of bovine jugular veins than in homograft recipients. CONCLUSION According to the time status graphs, the use of bovine jugular veins for RVOT in patients younger than 25 years appears to lead to superior results when compared with cryopreserved homografts.

Systemic inflammatory response syndrome after pediatric congenital heart surgery: Incidence, risk factors, and clinical outcome

Systemic inflammatory response syndrome (SIRS) is frequent after cardiac surgery, but data on its incidence and perioperative risk factors are scarce for children with congenital heart disease.

Decellularization of aortic valves: only time will tell.

Which is the best procedure for aortic valve replacement for a 33-year old female patient and which is most suitable for a 15-year old male athlete? One choice could be a mechanical valve, with the aim of providing lifetime durability. However, the era of purely artificial valve prostheses composed of rigid components is drawing to a close. In Germany, for example, their share of the total of implanted aortic valve prostheses implanted in 2014 amounted to a mere 14%. Glutaraldehyde-fixed xenografts still have limited durability, which precludes their use in very young patients. (Paediatric) Cardiac surgeons therefore have turned to the concept of tissue engineering, and a number of groups have pursued the idea of decellularizing existing biological valve resources to create a biological aortic valve prosthesis, which, in the best case, would regenerate and be remodelled as the patient’s own valve or, at the very least, would be less prone to degeneration and calcification. Several protocols have been evaluated and decellularization is usually achieved by cell lysis due to changes of osmotic pressure induced using buffer solutions such as tris(hydroxymethyl)-aminomethane [1]. Bile components (sodium deoxycholate, sodium dodecyl sulphate), TRITON-100 (polyethylene glycol p-(1,1,3,3-tetramethylbutyl)-phenyl ether) [2] or 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulphonate) [2] are then used to wash out the cell remnants from the collagen, while preserving the basal membrane and scaffold integrity. Artificial scaffolds created via tissue engineering offer several advantages, such as the possibility of creating custom-made scaffolds with consistent quality and the feasibility of off-the-shelf usage. But despite their comparative strengths in theory, only three protocols [3–5], out of the numerous groups working in the field of tissue-engineered heart valves have been successfully transferred to the clinic, all of which use decellularized allogenic matrices to generate aortic valve substitutes. The use of human aortic valves remains challenging and allograft procurement remains an unsolved problem [6]. Decellularized aortic valves have shown remarkable initial results. Surgical handling resembles that in native aortic valve surgery and leads to superior effective valve areas in comparison with other valve prostheses [7–9], while avoiding the two-valve procedure of a Ross operation. However, on the basis of current preclinical experience, little is known about the factors surrounding the desired recellularization process, which is key to creating an individualized, functioning graft. Cultivated mesenchymal stem cells either derived from bone marrow [10] or de-differentiated from circulating blood [11] cells have been used in conjunction with endothelial cells for in vitro reseeding of the decellularized matrix. Reseeding the scaffold under quasi-physiological conditions in an appropriately equipped pulse duplicator favours and accelerates the reseeding process, which also was shown to take place after direct implantation of the unseeded scaffold [12]. Ex vivo reseeding, however, has the disadvantage of being both timeand resource-intensive, and infections may further reduce the already small number of available conduits, or put the patient at risk if an infection is discovered only after implantation. In vivo recellularization has been demonstrated in large animal studies on aortic valve replacement [12], and there is evidence for recellularization of decellularized grafts in humans (Fig. 1). Recellularization is crucial and therefore research has also addressed factors which may enhance invasion of recipient’s cell populations. These ‘growth factors’, however, have thus far not shown satisfying results and have therefore not yet been viable for clinical application. From a surgical point of view, I would emphasize the importance of perfect implantation, even a normal aortic valve may fail in the setting of a relevant subaortic stenosis due to jet-damaged leaflets. Although recellularization may also occur from the adventitial side, as shown by the Padua group [13], recellularization from the luminal side is certainly important for cusps. It seems very clear that this is much more likely if surgery is able to establish a laminar flow across the implanted graft. From my experience, it is of utmost importance to avoid any distortion of the graft, as these cusps are soft and susceptible to mechanical alterations. As we have found no evidence for dilatation using the Hannover protocol for decellularization, full root replacement without any augmentation can maintain graft compliance and possibility of growth. Since individual predispositions (especially in congenital heart defects, e.g. after LeCompte manoeuvre in transposition) are prevalent among our patients, it is quite difficult to simulate these factors in animal models, meaning that we will have to wait for the mid-term and long-term results of the first clinical implantation series to fully assess these aspects. There are numerous technical aspects surrounding the implantation of a decellularized aortic valve which also need to be

RNA isolation from fetal and adult human tissues for transcriptional profiling.

Investigations involving rare human tissues that are difficult to acquire due to their scarcity are highly challenging. The need to verify microarray analysis data by additional methods such as immunohistochemical staining and quantitative PCR creates an even greater demand for these valuable tissues. Furthermore, since rare human tissues may come from different sources and may have been processed by variable methods, the comparability of these samples must be verified. The aim of this study was to determine and validate a processing method that allows the analysis of human fetal and adult cardiovascular tissues from different sources that were preserved using varying methods. Due to restricted access to fresh human tissues and the need to accumulate these samples over an extended period of time, we used formalin-fixed paraffin-embedded tissues for gene expression analyses. We analyzed RNA levels from four different age groups: fetal first and second trimester, adolescents, and adults. In this study, we present an improved standard processing procedure for tissue sample processing and analysis of rare human cardiovascular tissues.

Long-term results after repair of anomalous origin of left coronary artery from the pulmonary artery: Takeuchi repair versus coronary transfer

OBJECTIVES: We evaluated long-term results of two different repair strategies of anomalous origin of left coronary artery from pulmonary artery (ALCAPA) in two German surgical centres. METHODS: We performed a retrospective review of patients who underwent ALCAPA repair between November 1980 and October 2012. Ventricular function was assessed by standardized transthoracic echocardiographic studies. RESULTS: A total of 30 patients with a median age of 0.4 years (range 0.05–37.6 years) underwent ALCAPA repair. Nineteen patients received coronary transfer, 9 underwent Takeuchi repair (creation of an intrapulmonary tunnel) and 2 were treated by closure of the ALCAPA. There was one concomitant mitral valve repair. The total follow-up was 244.8 years with a mean follow-up of 8.16 ± 6.7 years (range 0.06–24.3 years). There was no late mortality. None of the patients treated with coronary transfer or Takeuchi repair died. One moribund patient who received coronary ligation in the 1980s died perioperatively. The 10-year rate of freedom from reoperation was 94.1% for patients treated with coronary transfer. For patients with Takeuchi repair, the 10-year rate of freedom from reoperation was 71.1% (P = 0.146). At last followup, left ventricular end-diastolic diameter (LVEDD) and function measured by fractional shortening was normal in all patients. The mean shortening fraction improved from 20.25 ± 10.86% to 35.7 ± 4.8%, with the most remarkable change taking place in the first year, and the mean LVEDD indexed to normal improved from 42 ± 34% above normal values preoperatively to 3 ± 12% at the last follow-up (P < 0.01). The number of patients with moderate-to-severe mitral regurgitation (MR) improved from 50% to 6.7% (2 of 30) at the last followup. In all children receiving ALCAPA repair under the age of 6 months, MR was absent or trivial at the last follow-up. Eight years after Takeuchi repair, 79.9% of these patients suffered from at least moderate pulmonary regurgitation, in contrast to 0% after coronary transfer (P < 0.001). CONCLUSIONS: Survival rates and long-term left ventricular function were excellent for both surgical strategies. Mitral regurgitation resolved in all patients operated during their first year of life independent of the type of repair. Takeuchi repair, however, led to significant pulmonary regurgitation.

NT-proBNP Indicates Left Ventricular Impairment and Adverse Clinical Outcome in Patients With Tetralogy of Fallot and Pulmonary Regurgitation

BACKGROUND The goal of this study was to interrelate N-terminal B-type natriuretic peptide (NT-proBNP) levels and cardiac magnetic resonance imaging-derived ventricular function, mass, and volumes in adults with pulmonary regurgitation after Fallot repair and to evaluate the prognostic relevance of these parameters regarding adverse clinical outcome. METHODS Eighty-one patients (aged 26.3 ± 7.4 years; male sex, 45.7%; New York Heart Association class I, 72.8%; pulmonary valve velocity, < 3 m/s) were included. At baseline cardiac magnetic resonance imaging and NT-proBNP measurements were performed. RESULTS During a mean observation time of 6.9 ± 2.6 years, 13 patients (16.1%) had sustained supraventricular arrhythmias or heart failure (2.4 per 100 patient-years). Multivariate Cox analysis identified NT-proBNP, left ventricular (LV) end-systolic volume index and LV ejection fraction, right ventricular (RV) end-diastolic volume index, and tricuspid regurgitation as independent predictors of adverse events. NT-proBNP correlated with LV but not with RV parameters. In receiver operating characteristic curve analysis using significant variables of the multivariate analysis, NT-proBNP was superior to all other parameters to detect patients at risk (area under the curve [AUC], 0.873; 95% confidence interval, 0.772-0.974). LV end-systolic volume index (AUC, 0.734), RV end-diastolic volume index (AUC, 0.645) und tricuspid regurgitation (AUC, 0.747) showed lower diagnostic accuracy. CONCLUSIONS Even in mildly symptomatic patients with pulmonary regurgitation after Fallot repair NT-proBNP is a strong predictor of adverse outcome. It is rather associated with LV but not with RV impairment. In severe pulmonary regurgitation an increase in the level of NT-proBNP and LV impairment seem to provide additional useful information for the timing of pulmonary valve replacement.