Characterization of new mutations in the FOLR1-gene

In der vorliegenden experimentell durchgeführten Dissertation wurden neue Mutationen im FOLR1-Gen molekular charakterisiert. Die untersuchten Mutationen führten zu Veränderungen im Folatrezeptor α, einem von Zellen des Plexus choroideus exprimierten Protein, welches den Haupttransporter von  5-MTHF über die Blut-Liquor-Schranke darstellt. Mutationen im FOLR1-Gen sind kürzlich als Ursache einer zerebralen Folattransport-Defizienz identifiziert worden. Diese Erkrankung gehört zu einer Gruppe neurologischer Störungen, die sich durch isoliert niedrige Liquorfolatwerte  auszeichnen und zusammenfassend als zerebrale Folatdefizienz bezeichnet werden. Die zerebrale Folattransportdefizienz manifestiert sich typischerweise im frühen Kleinkindesalter und äußert sich klinisch in Form einer chronisch-progredienten psychomotorischen Regression mit zerebralen Krampfanfällen und einer durch MRT nachweisbaren Myelinisierungsstörung. Eine Substitution mit  5‘-Formyltetrahydrofolat konnte bei der Mehrzahl der identifizierten Patienten eine partielle Remission der Symptome bewirken.  Im Rahmen dieser Arbeit wurden vier Mutationen im FOLR1-Gen hinsichtlich ihrer molekularen Auswirkungen auf die Proteinxpression, die Funktionalität bzw. Rezeptor-Bindungsfähigkeit und die Lokalisation des Proteins untersucht. Bei den Mutationen handelt es sich um zwei neue, aus Patienten-DNS identifizierte Punktmutationen, p.C169Y und p.N222S, sowie die bereits beschriebenen FRα-Mutanten p.C105R und FRα p.K44_P49dup.  Die heterologe Expression der mutanten Folatrezeptoren zeigte in Westernblot-Analysen keine signifikanten Veränderungen der Protein-Expressionsrate, verglichen mit dem Wildtyp-Protein. Allerdings bestand eine stark verminderte Rezeptor-Folsäurebindung in radioaktiven Bindungsassays. Ein funktioneller Unterschied zwischen den einzelnen Mutanten konnte im Verlauf der Experimente identifiziert werden. Die FRα-Proteinmutanten p.N222S und p.K44_P49dup zeigten verglichen mit den anderen Mutanten eine höhere Folsäure-Restbindung von ca. 20 % des Wildtypproteins. Im Rahmen von Immunfluoreszenzmikroskopien konnte gezeigt werden, dass die FRα-Mutante p.K44_P49dup partiell zellmembranständig, entsprechend dem FRα-Wildtyp, exprimiert wurde. Die übrigen untersuchten Proteinmutanten zeigten in intrazellulären Kompartimenten zumindest teilweise eine Kolokalisation mit dem Marker des endoplasmatischen Retikulums. Alle Untersuchungen wurden mit transfizierten CHO-K1-Zellen durchgeführt und konnten in zwei polaren Zelllinien, immortalisierten Epithelzellen des Plexus choroideus (Z310) und humanen Leberzellkarzinomzellen (HepG2) bestätigt werden. Die Ergebnisse dieser Dissertation sind Bestandteil einer kürzlich in BRAIN veröffentlichten Arbeit (Grapp et al. 2012) und tragen zum besseren Verständnis der molekularen Grundlagen der zerebralen Folattransport-Defizienz, einer neuerkannten, behandelbaren neuropädiatrischen Erkrankung, bei. Die  Pathogenität der untersuchten FOLR1-Mutationen wird auf molekularer Ebene belegt. Die Bedeutung dieser molekulargenetischen Untersuchungen besteht darin, dass eine frühzeitige Folat-Behandlung erkrankter Kinder zu einer deutlichen Verbesserung der Symptome führt

Mechanical unloading by miniature axial flow pumps in late cardiac allograft failure due to acute rejection

Allograft failure secondary to rejection commonly requires a multimodal treatment, ultimately including mechanical circulatory support. A few case reports have demonstrated the use of Impella-devices due to its assumed favorable safety profile in this fragile cohort. However, this treatment option does not play a role in choice of anti-rejective therapy in clinical routine up to date. We summarize our institutional experiences and literature mini-review on Impella-based treatment strategies in allograft rejection after heart transplantation. In all seven cases, three from our institution and four reported in the literature, Impella-based therapies led to hemodynamic stabilization in allograft failure secondary to rejection. Adverse events included hemolysis, non-fatal bleeding and in one patient a relevant aortic valve insufficiency occurred. All patients showed an improvement of allograft function. Two patients died in context of severe immunosuppression or late secondary organ failure. Based on the limited available data, we propose that Impella-mediated mechanical unloading represents a valuable option for hemodynamic stabilization in severe allograft failure due to rejection, enabling an initiation of causal therapy and thereby potentially representing an opportunity to prevent mortality. Furthermore, we hypothesize it might add to the traditional therapeutic approaches by facilitating recovery by decompressing the myocardium in allograft rejection

Missing the blind spot in a HeartMate 3 outflow graft obstruction caused by fungal infection

A patient was admitted in cardiogenic shock and a constant decrease of pump flow requiring combined inotropic support. To evaluate the cause, echocardiography and a ramp test were performed. The results suggested a LVAD related problem - particularly a suspected outflow graft obstruction. Wether CT scan nor angiography confirmed the assumption. However, a post-mortem LVAD examination revealed an outflow obstruction caused by a fungal thrombus formation invisible for standard imaging procedures.ISSN:1525-1594ISSN:0160-564

Fluorescence in situ hybridization and polymerase chain reaction to detect infections of cardiac implantable electronic devices

Aims In patients with infections of cardiac implantable electronic devices (CIEDs), the identification of causative pathogens is complicated by biofilm formations and previous antibiotic therapy. In this work, the impact of an additional fluorescence in situ hybridization (FISH), in combination with polymerase chain reaction and sequencing (FISHseq) was investigated. Methods and results In 36 patients with CIED infections, FISHseq of explanted devices was performed and compared with standard microbiological cultivation of preoperative and intraoperative samples. The mean age was 61.9 (+/- 16.2) years; 25 (69.4%) were males. Most patients (62.9%) had heart failure with reduced ejection fraction. Infections occurred as endoplastits (n = 26), isolated local generator pocket infection (n = 8), or both (n = 2); CIED included cardiac resynchronization therapy defibrillator (n = 17), implantable cardioverter defibrillator (n = 11), and pacemaker (n = 8) devices. The overall positive FISHseq detection rate was 97%. Intraoperatively, pathogens were isolated in 42 vs. 53% in standard cultivation vs. FISHseq, respectively. In 16 of 17 FISHseq-negative patients, the nucleic acid strain DAPI (4 ',6-diamidino-2-phenylindole) indicated inactive microorganisms, which were partially organized in biofilms (n = 4) or microcolonies (n = 2). In 13 patients in whom no pathogen was identified preoperatively, standard cultivation and FISHseq identified pathogens in 3 (23%) vs. 8 (62%), respectively. For the confirmation of preoperatively known bacteria, a combined approach was most efficient. Conclusion Fluorescence in situ hybridization sequencing is a valuable tool to detect causative microorganisms in CIED infections. The combination of FISHseq with preoperative cultivation showed the highest efficacy in detecting pathogens. Additional cultivation of intraoperative tissue samples or swabs yielded more confirmation of pathogens known from preoperative culture.ISSN:1099-5129ISSN:1532-209

Movement therapy in advanced heart failure assisted by a lightweight wearable robot: a feasibility pilot study

AIMS The aim of this pilot study was to investigate the safety, feasibility, tolerability, and acceptability of an assisted mobilization of advanced heart failure patients, using a lightweight, exoskeleton-type robot (Myosuit, MyoSwiss AG, Zurich, Switzerland). METHODS AND RESULTS Twenty patients in functional NYHA class III performed activities of daily life (ADL, n = 10) or participated in a single, standardized, 60 min rehabilitation exercise unit (REU, n = 10) with and without the Myosuit. The outcome assessment included the evaluation of vital signs, adverse events, rates of perceived exertion and dyspnoea (RPE, RPD), the ability to perform ADL or REU, and the individual acceptability. The mean age of the subjects was 49.4 (±11.0) years; 80% were male. The mean left ventricular ejection fraction was 22.1% (±7.4%) and the median NT-proBNP 2054 pg/mL (IQR 677, 3270 pg/mL). In all patients, mobilization with the Myosuit was feasible independently or with minor support. The mean individual difference in the total walking distance of the patients without and with robotic assistance was -26.5 m (95% confidence interval (CI) -142 to 78 m, P = 0.241). No adverse events occurred. RPE and RPD showed no significant difference with or without the device (ADL: RPE -0.1 m, 95% CI -1.42 to 1.62, P = 0.932 and RPD -0.95 m, 95% CI -0.38 to 2.28, P = 0.141; REU: RPE 1.1 m, 95% CI -2.90 to 0.70, P = 0.201 and RPD 0.5 m, 95% CI -2.02 to 1.02, P = 0.435). All median responses in the acceptability questionnaire were positive. The patients felt safe and enjoyed the experience; 85% would be interested in participating in robot-assisted training on a regular basis. CONCLUSION This feasibility pilot trial provides first indications that a robotic exoskeleton-assisted mobilization of patients with advanced heart failure is safe, feasible, well-tolerated, and well-accepted. The results are highly encouraging to further pursue this innovative approach in rehabilitation programmes. This trial was registered at ClinicalTrials.gov: NCT04839133

Movement therapy in lung transplantation candidates assisted by a lightweight wearable robot

The aim of this pilot-study was to investigate the safety, feasibility and tolerability of an assisted mobilization of patients with advanced pulmonary diseases, using a lightweight, exoskeleton-type robot (Myosuit, MyoSwiss AG, Zurich, Switzerland). Ten patients performed activities of daily life (ADL) both with and without the device. The mean age was 53.6 (±5.6) years; 70% were male. The assessment of outcome included the evaluation of vital signs, adverse events, rates of perceived exertion and dyspnea (PRE, PRD), the ability to perform ADL and the individual acceptability. Robotic-assisted mobilization was feasible in all patients. No adverse events occurred. RPE and RPD showed no significant difference with or without the Myosuit (mean difference in RPE -1.7, 95%-confidence interval (CI) -1.16, 4.49; p = 0.211; mean difference in RPD 0.00, 95%-CI -1.88, 1.88; p = 0.475). 80% of patients were interested to participate in a robotic-assisted training on a regular basis. A robotic exoskeleton-assisted mobilization is safe, feasible, well-tolerated and well-accepted. The results are highly encouraging to further pursue this highly innovative approach

Acute Kidney Injury After Heart Transplantation: Risk Factors and Clinical Outcomes

Objective: Acute kidney injury (AKI) requiring renal-replacement therapy (RRT) after heart transplantation (OHT) is common and impairs outcomes. This study aimed to identify independent donor and recipient risk factors associated with RRT after OHT. Design: A retrospective data analysis. Setting: Data were collected from clinical routines in a maximum-care university hospital. Participants: Patients who underwent OHT. Interventions: The authors retrospectively analyzed data from 264 patients who underwent OHT between 2012 and 2021; 189 patients were eligible and included in the final analysis. Measurements and Main Results: The mean age was 48.0 ± 12.3 years, and 71.4% of patients were male. Ninety (47.6%) patients were on long-term mechanical circulatory support (lt-MCS). Posttransplant AKI with RRT occurred in 123 (65.1%) patients. In a multivariate analysis, preoperative body mass index >25 kg/m² (odds ratio [OR] 4.74, p < 0.001), elevated preoperative creatinine levels (OR for each mg/dL increase 3.44, p = 0.004), administration of red blood cell units during transplantation procedure (OR 2.31, p = 0.041) and ischemia time (OR for each hour increase 1.77, p = 0.004) were associated with a higher incidence of RRT. The use of renin-angiotensin-aldosterone system blockers before transplantation was associated with a reduced risk of RRT (OR 0.36, p = 0.013). The risk of mortality was 6.9-fold higher in patients who required RRT (hazard ratio 6.9, 95% CI: 2.1-22.6 p = 0.001). Previous lt-MCS, as well as donor parameters, were not associated with RRT after OHT. Conclusions: The implementation of guideline-directed medical therapy, weight reduction, minimizing ischemia time (ie, organ perfusion systems, workflow optimization), and comprehensive patient blood management potentially influences renal function and outcomes after OHT

Impact of Muscle Mass as a Prognostic Factor for Failed Waiting Time Prior to Heart Transplantation

Objectives: Clinical deterioration during the waiting time impairs the prognosis of patients listed for heart transplantation. Reduced muscle mass increases the risk for mortality after cardiac surgery, but its impact on resilience against deterioration during the waiting time remains unclear. Methods: We retrospectively analyzed data from 93 patients without a VAD who were listed in Eurotransplant status “high urgent (HU)” for heart transplantation between January 2015 and October 2020. The axial muscle area of the erector spinae muscles at the level of thoracic vertebra 12 indexed to body surface area (TMESA/BSA) measured in the preoperative thoracic computed tomography scan was used to measure muscle mass. Results: Forty patients (43%) underwent emergency VAD implantation during the waiting time and four patients (4%) died during the waiting time. The risk of emergency VAD implantation/death during the waiting time decreased by 10% for every cm2/m2 increase in muscle area [OR 0.901 (95% CI: 0.808–0.996); p = 0.049]. After adjusting for gender [OR 0.318 (95% CI: 0.087–1.073); p = 0.072], mean pulmonary artery pressure [OR 1.061 (95% CI: 0.999–1.131); p = 0.060], C-reactive protein [OR 1.352 (95% CI: 0.986–2.027); p = 0.096], and hemoglobin [OR 0.862 (95% CI: 0.618–1.177); p = 0.360], TMESA/BSA [OR 0.815 (95% CI: 0.698–0.936); p = 0.006] remained an independent risk factor for emergency VAD implantation/death during the HU waiting time. Conclusion: Muscle area of the erector spinae muscle appears to be a potential, easily identifiable risk factor for emergency VAD implantation or death in patients on the HU waiting list for heart transplantation. Identifying patients at risk could help optimize the outcome and the timing of VAD support.ISSN:2297-055