Conference Agenda

Introduction

Infective endocarditis is an increasingly prevalent, often fatal disease. When affecting the mitral or tricuspid valve, repair should be performed whenever possible, using a synthetic annuloplasty ring. Currently used rings do not offer any antibacterial features. Therefore, systemic antibiosis remains the only antimicrobial treatment option. In this study we developed a fully absorbable magnesium (Mg)-based annuloplasty ring covered with silver (Ag) nanoparticle (AgNP)-loaded electrospun polycaprolactone (PCL) providing local antibacterial activity. Initial in vitro and in vivo biocompatibility studies of the Mg alloy ZX00 and the AgNP-PCL coverage were performed before long-term functionality and safety were assessed in a sheep model.

Methods

In vitro biocompatibility and hemocompatibility assays were performed. AgNP-PCL was tested for in vitro antibacterial activity. ZX00 was implanted subcutaneously in rat and rabbit models for histopathological, µCT, inflammatory and mechanical bending assessments. To ensure orthotopic implant functionality, the rings were implanted in sheep hearts in the mitral position. Widely used Edwards mitralrings were used as controls. Echocardiography, hemograms, blood chemistry, organ histopathological assessments, immunohistochemistry and mass spectrometry (ICP-MS) studies were conducted.

Results

Excellent in vitro and in vivo biocompatibility of ZX00 and AgNP-PCL as well as significant in vitro antibacterial activity of AgNP-PCL was observed (>99% reduction of bacteria). Exceptional suture retention of the newly developed Mg ring was demonstrated through successful implantation in sheep hearts. Implants degraded homogeneously (55 ± 6% implant volume reduction after one year of implantation) and showed remarkable ingrowth behavior comparable to clinically available rings. Echocardiography showed preserved mitral valve function (mean pressure gradient: 1.37 ± 0.52 mmHg) during the entire one-year follow-up period. Toxicological safety could be demonstrated through monthly hematological assessments as well as histopathological analysis of parenchymatous organs. Finally, mass spectrometric screenings of Ag revealed no significant accumulation of AgNP in parenchymatous organs compared to control animals. The relevant patent of the invention was published in August 2025 (Publication Number: WO/2025/176798).

Conclusion

This newly developed, antibacterial and biodegradable annuloplasty ring presents a promising new option for the surgical treatment of infective endocarditis, whenever repair of the valve is possible.

Normothermic machine perfusion (NMP) allows liver preservation under near-physiological conditions and thus may, serve as a platform for ex situ therapeutic interventions. Liver-directed adeno-associated viruses (AAV) hold promise for treating genetic liver diseases, yet highly specific vectors remain limited and are lacking. We applied Oxford Nanopore Technology (ONT) for direct sequencing of vector DNA and RNA in an NMP model to optimize AAV-based gene therapy protocols.

Porcine livers (n=4) were subjected to NMP (organOx Metra) for 48h, and eYFP-labeled AAV3B was administered to the perfusate. Serial perfusate samples were evaluated for organ function. Tissue biopsies were collected before vector administration and after 3h, 24h, and 48h for real-time confocal microscopy and immunofluorescent staining to assess eYFP protein, or cryopreserved for ONT-based quantification of AAV3B cDNA and gDNA.
ONT sequencing confirmed high-quality AAV3B vector preparation. All livers were successfully perfused for 48h with preserved organ function, as indicated by lactate clearance and low liver transaminase and LDH levels.

At a dose of 1×10¹³ vg/kg, AAV transduction was successfully detected by eYFP fluorescence in tissue biopsies after 24h and 48h, but not as early as 3h after vector application. The addition of a poloxamer (Pluronic) enhanced AAV3B transduction efficiency, while a reduced dose of 2×10^12 vg/kg with Pluronic achieved comparable efficiency to the high-dosage protocol without it.

In conclusion, AAV administration during liver NMP is feasible, well tolerated, and establishes NMP as a powerful platform for the development of AAV-based gene-directed therapies.

Background: Normothermic machine perfusion (NMP) has emerged as a promising strategy to evaluate liver graft viability prior to transplantation and may serve as a tool for future organ repair strategies. As a prerequisite, reliable experimental models to study and quantify liver injury are warranted.

Methods: To identify a model in which a notable but not detrimental degree of damage is inflicted on livers prior to NMP in a standardized setting, porcine livers (Sus domesticus, 70–95 kg, total n=7) underwent 24 h of cold ischemia time (CIT) at 4 °C (n=3). Biopsies were collected at baseline and after 24 h of storage, formalin-fixed, and histologically graded for structural injury (H&E). Markers of apoptosis, proliferation, and immune activation were analyzed using immunohistochemistry (IHC). Functional outcomes were measured during subsequent NMP (OrganOx® Metra), comparing livers after 24h vs. 0h of CIT.

Results: After 24 h of CIT, histological evaluation revealed only discrete increase of vacuolization (score 0h: 1±1.5 vs. 24h 2±1; n.s.), and an absence of graft necrosis, architecture destruction, and apoptosis (all p>0.05, respectively). Likewise, IHC displayed no significant alterations in markers indicating apoptosis (Caspase-3, Bcl-2), proliferation (Ki-67), or endothelial activation (ICAM-1, TLR4, VEGFRII), concerning both staining intensity and the proportion of positive cells (all p>0.05). Furthermore, lymphocyte subset composition (CD3, CD25, CD4, FoxP3) remained unaltered. Reperfusion of livers after 24h of CIT resulted in elevated lactate levels following NMP initiation (24h: 109.8 ± SD vs 0h: 65 ± SD, p>0.05), which gradually declined over 24h of NMP. Pronounced hepatocyte damage was indicated by high ALT (24h CIT: 734.6U/L ± SD vs 0h CIT: 43U/L ± SD, p<0.058) and AST (24h CIT: 10224.3 U/L ±SD vs 0h CIT: 111 U/L ±SD, p<0.058) levels.

Conclusion: A 24-h period of cold ischemia at 4 °C, although twice the duration typically applied clinically, did not induce major morphologically detectable damage. Functional assessment during NMP indicated moderate but not extensive impairment, suggesting a reproducible and suitable experimental model for NMP research. These findings highlight the value of NMP-based viability testing with functional readouts to evaluate grafts subjected to prolonged CIT.

Background: A promising strategy to meet the growing demand for organ donations is the allocation of extended criteria or marginal donor grafts. However, the use of suboptimal liver allografts poses a significant risk for serious postoperative complications. While hypothermic oxygenated machine perfusion (HOPE) has been shown to significantly improve postoperative outcomes by ameliorating ischemia reperfusion injury (IRI), the reduced metabolism in the cold limits the possibility to assess organ quality during machine perfusion, i.e. prior to transplantation. Mitochondrial flavin mononucleotide (FMN) and glycocalyx component syndecan-1 (Sdc-1) have previously been investigated as potential biomarkers of liver viability and have shown promising success in prediction of postoperative early allograft dysfunction (EAD) and other outcome parameters.

Methods: FMN and Sdc-1 concentrations in HOPE perfusates of 40 liver allografts were measured via fluorescence spectroscopy and enzyme-linked immunosorbent essay (ELISA), respectively. Previously reported cut-off values of 10.65 ng/mL after five minutes of perfusion for FMN and 808 ng/mL after 60 minutes for Sdc-1 were used for risk stratification. With a sequential combination of FMN and Sdc-1, refined risk groups could be investigated in order to further improve the prediction of postoperative outcomes in liver transplantation.

Results: Early allograft dysfunction (EAD) was largely absent in the low-risk cohort, occurring in only 9% of patients with FMN concentrations below the threshold (2/22) and 27% of those with sub-threshold Sdc-1 levels (7/26). In contrast, among high-risk individuals, EAD was observed in 52% of cases with elevated FMN (13/25) and 58% with elevated Sdc-1 (7/12). Intriguingly, no instances of EAD were documented when both biomarkers remained below their respective cut-off values (0/12). Conversely, simultaneous elevation of FMN and Sdc-1 was associated with a markedly higher incidence of EAD, affecting 67% of patients (6/9). When only one biomarker exceeded the threshold, FMN demonstrated superior predictive performance, with EAD occurring in 58% of such cases, compared with 33% for isolated Sdc-1 elevation.

Conclusion: FMN and Sdc-1 levels assessed during HOPE serve as reliable biomarkers for predicting EAD following liver transplantation. Their combined evaluation markedly enhances predictive specificity and may provide additional support for clinical decision making.

Background:

Orthotopic liver transplantation (OLT) remains the only curative treatment for end-stage liver disease, but the shortage of suitable organs and limitations of current donor assessment tools restrict optimal allocation. Traditional laboratory markers lack predictive accuracy for donor organ quality. The hepatomiR® P-score, based on circulating microRNAs (miR-122-5p, miR-151a-5p, miR-192-5p), has shown prognostic value in hepatic surgery, but its role in donor evaluation for OLT remains unexplored.

Methods:

Between 2016 and 2021, 89 brain-dead donors and 60 corresponding recipients at the Medical University of Vienna were prospectively studied. Donor blood samples were collected before procurement, and recipient samples pre-OLT, 30 minutes post-reperfusion, and on postoperative day 1. Plasma microRNA levels were quantified using the hepatomiR® kit to calculate P-scores. Donor histology, laboratory parameters, and clinical outcomes were analyzed. Early allograft dysfunction (EAD) was defined by a model of early allograft function (MEAF) score ≥8.

Results:

Donors with higher P-scores were more frequently associated with non-utilization (75% vs. 31.1%, p=0.014) and grafts leading to EAD (33.3% vs. 3.9%, p=0.031). Median P-scores were significantly higher in recipients versus donors (0.515 vs. 0.245, p<0.001), particularly in non-malignant indications. Perioperative dynamics revealed a rapid P-score increase as early as 30 minutes after reperfusion, correlating with MEAF (r=0.413, p=0.001). At reperfusion, P-score predicted EAD with an AUC of 0.833, outperforming routine laboratory tests. No correlation was observed between donor P-score and histological steatosis or fibrosis.

Conclusion:

The hepatomiR® P-score identifies high-risk donor grafts and predicts EAD more accurately than conventional biomarkers at early time points. Its rapid responsiveness to ischemia-reperfusion injury and standardized measurement support its utility as a complementary tool in donor assessment. Incorporation of hepatomiR® into clinical practice may refine graft selection and improve OLT outcomes.