@unpublished{ThausHofrichterLubnowetal., author = {Thaus, Christopher and Hofrichter, Elena and Lubnow, Matthias and Krenkel, Lars and Lehle, Karla}, title = {Hemocompatibility of Membrane Lung Components from Extracorporeal Membrane Oxygenation with Different Antithrombogenic Coatings}, doi = {10.20944/preprints202412.0615.v1}, pages = {1 -- 11}, abstract = {Thrombus formation within extracorporeal membrane oxygenation (ECMO) devices remained a critical complication. One reason seems to be the contact of blood with large artificial surfaces within the membrane lung (ML). The aim was to test the hemocompatibility of different na{\"i}ve ECMO materials. Blood and platelets from five healthy volunteers were incubated with gas exchange (GF) and heat exchange fibers (HE) from four different commercial available new MLs representing different antithrombogenic coatings. Adherent platelets were stained with rhodamine-phalloidin. Surface coverage was quantified with ImageJ. Non-adherent platelets were stained with antibodies (CD62P, PAC-1, CD61) and fibrinogen to detect platelet activation with flow cytometry. Hemolysis of red blood cells after material contact was detected. All ECMO-materials were non-hemolytic and did not induce platelet activation. However, platetelet adhesion (median (IQR)) was significantly elevated on uncoated GFs made of polymethylpentene (GF-PMP; 12 (7-19)\%) and on GFs from the Hilite-MLs (GF-Hilite; 13 ((8-19)\%) compared to the other materials. In vitro testing of platelet adhesion disclosed significant differences of ECMO-materials with different antithrombogenic surface coatings. Instead, circulating platelets remained non-activated. ECMO-materials and its coatings were non-hemolytic. Finally, this study confirmed the good hemocompatibility of GFs and HEs from commerciall available MLs.}, language = {en} } @article{WagnerKranzKrenkeletal., author = {Wagner, Maria Stella and Kranz, Michael and Krenkel, Lars and Pointner, Daniel and Foltan, Maik and Lubnow, Matthias and Lehle, Karla}, title = {Computer based visualization of clot structures in extracorporeal membrane oxygenation and histological clot investigations for understanding thrombosis in membrane lungs}, series = {Frontiers in Medicine}, journal = {Frontiers in Medicine}, number = {11}, editor = {Becatti, Matteo}, publisher = {Frontiers}, doi = {10.3389/fmed.2024.1416319}, abstract = {Extracorporeal membrane oxygenation (ECMO) was established as a treatment for severe cardiac or respiratory disease. Intra-device clot formation is a common risk. This is based on complex coagulation phenomena which are not yet sufficiently understood. The objective was the development and validation of a methodology to capture the key properties of clots deposed in membrane lungs (MLs), such as clot size, distribution, burden, and composition. One end-oftherapy PLS ML was examined. Clot detection was performed using multidetector computed tomography (MDCT), microcomputed tomography (μCT), and photography of fiber mats (fiber mat imaging, FMI). Histological staining was conducted for von Willebrand factor (vWF), platelets (CD42b, CD62P), fibrin, and nucleated cells (4′, 6-diamidino-2-phenylindole, DAPI). The three imaging methods showed similar clot distribution inside the ML. Independent of the imaging method, clot loading was detected predominantly in the inlet chamber of the ML. The μCT had the highest accuracy. However, it was more expensive and time consuming than MDCT or FMI. The MDCT detected the clots with low scanning time. Due to its lower resolution, it only showed clotted areas but not the exact shape of clot structures. FMI represented the simplest variant, requiring little effort and resources. FMI allowed clot localization and calculation of clot volume. Histological evaluation indicated omnipresent immunological deposits throughout the ML. Visually clot-free areas were covered with leukocytes and platelets forming platelet-leukocyte aggregates (PLAs). Cells were embedded in vWF cobwebs, while vWF fibers were negligible. In conclusion, the presented methodology allowed adequate clot identification and histological classification of possible thrombosis markers such as PLAs.}, language = {en} } @article{LingelHausPaschkeetal., author = {Lingel, Maximilian P. and Haus, Moritz and Paschke, Lukas and Foltan, Maik and Lubnow, Matthias and Gruber, Michael and Krenkel, Lars and Lehle, Karla}, title = {Clinical relevance of cell-free DNA during venovenous extracorporeal membrane oxygenation}, series = {Artificial organs}, volume = {47}, journal = {Artificial organs}, number = {11}, publisher = {Wiley}, issn = {1525-1594}, doi = {10.1111/aor.14616}, pages = {1720 -- 1731}, abstract = {BACKGROUND: Thrombosis remains a critical complication during venovenous extracorporeal membrane oxygenation (VV ECMO). The involvement of neutrophil extracellular traps (NETs) in thrombogenesis has to be discussed. The aim was to verify NETs in the form of cell-free DNA (cfDNA) in the plasma of patients during ECMO. METHODS: A fluorescent DNA-binding dye (QuantifFluor®, Promega) was used to detect cell-free DNA in plasma samples. cfDNA concentrations from volunteers (n = 21) and patients (n = 9) were compared and correlated with clinical/technical data before/during support, ECMO end and time of a system exchange. RESULTS: Before ECMO, patients with a median (IQR) age of 59 (51/63) years, SOFA score of 11 (10/15), and ECMO run time of 9.0 (7.0/19.5) days presented significantly higher levels of cfDNA compared to volunteers (6.4 (5.8/7.9) ng/μL vs. 5.9 (5.4/6.3) ng/μL; p = 0.044). Within 2 days after ECMO start, cfDNA, inflammatory, and hemolysis parameters remained unchanged, while platelets decreased (p = 0.005). After ECMO removal at the end of therapy, cfDNA, inflammation, and coagulation data (except antithrombin III) remained unchanged. The renewal of a system resulted in known alterations in fibrinogen, d-dimers, and platelets, while cfDNA remained unchanged. CONCLUSION: Detection of cfDNA in plasma of ECMO patients was not an indicator of acute and circuit-induced thrombogenesis.}, language = {en} } @article{PaschkeFoltanWagneretal., author = {Paschke, Lukas and Foltan, Maik and Wagner, Maria S. and Lubnow, Matthias and Gruber, Michael and Krenkel, Lars and Lehle, Karla}, title = {Clinical Relevance of Platelet-Leukocyte Aggregates and Platelet P-Selectin Expression During Venovenous Extracorporeal Membrane Oxygenation}, series = {ASAIO Journal}, journal = {ASAIO Journal}, number = {April 03}, publisher = {Wolters Kluwer}, issn = {1058-2916}, doi = {10.1097/MAT.0000000000002421}, abstract = {Thrombosis continues to be a significant complication during venovenous extracorporeal membrane oxygenation (V-V ECMO). Platelet activation markers might serve as indicators of inflammation and thrombogenesis. The aim was to identify these markers in ECMO patients. Blood from 10 ECMO patients (before, during, after ECMO) and 11 healthy volunteers were collected to determine platelet-neutrophil-aggregates (PNAs), platelet-monocyte-aggregates (PMAs), fibrinogen-binding, and P-selectin-expression on platelets by flow cytometry. Critical illness was associated with significantly elevated levels of PNAs and PMAs, increased P-selectin expression, reduced fibrinogen-binding, and restricted activation of platelets. Although PNAs and PMAs decreased significantly within 2 hours after the initiation of ECMO and remained at those levels, ECMO did not affect basal P-selectin expression and fibrinogen-binding. These results correlated with coagulation activation. Platelet markers before ECMO were not indicators for an imminent system exchange and end of therapy. In conclusion, platelet dysfunction during ECMO was mainly attributed to the critical illness. Extracorporeal membrane oxygenation support strengthened the restricted response of platelets to exogenous agonists (P-selectin). Furthermore, a decrease in PNAs/PMAs after ECMO started identified a reduced inflammatory response. There was no correlation of analyzed platelet parameters with the incidence of thrombotic complications.}, language = {en} } @article{LehlePhilippKrenkeletal., author = {Lehle, Karla and Philipp, Alois and Krenkel, Lars and Gruber, Michael and Hiller, Karl-Anton and M{\"u}ller, Thomas and Lubnow, Matthias}, title = {Thrombocytopenia During Venovenous Extracorporeal Membrane Oxygenation in Adult Patients With Bacterial, Viral, and COVID-19 Pneumonia}, series = {ASAIO Journal}, journal = {ASAIO Journal}, publisher = {Wolters Kluwer}, issn = {1058-2916}, doi = {10.1097/MAT.0000000000002383}, abstract = {Contact of blood with artificial surfaces triggers platelet activation. The aim was to compare platelet kinetics after venovenous extracorporeal membrane oxygenation (V-V ECMO) start and after system exchange in different etiologies of acute lung failure. Platelet counts and coagulation parameters were analyzed from adult patients with long and exchange-free (≥8 days) ECMO runs (n = 330) caused by bacterial (n = 142), viral (n = 76), or coronavirus disease 2019 (COVID-19) (n = 112) pneumonia. A subpopulation requiring a system exchange and with long, exchange-free runs of the second oxygenator (≥7 days) (n = 110) was analyzed analogously. Patients with COVID-19 showed the highest platelet levels before ECMO implantation. Independent of the underlying disease and ECMO type, platelet counts decreased significantly within 24 hours and reached a steady state after 5 days. In the subpopulation, at the day of a system exchange, platelet counts were lower compared with ECMO start, but without differences between underlying diseases. Subsequently, platelets remained unchanged in the bacterial pneumonia group, but increased in the COVID-19 and viral pneumonia groups within 2-4 days, whereas D-dimers decreased and fibrinogen levels increased. Thus, overall platelet counts on V-V ECMO show disease-specific initial dynamics followed by an ongoing consumption by the ECMO device, which is not boosted by new artificial surfaces after a system exchange.}, language = {en} } @article{KranzWagnerPointneretal., author = {Kranz, Michael and Wagner, Maria Stella and Pointner, Daniel and Haus, Moritz and Lubnow, Matthias and Lehle, Karla and Krenkel, Lars}, title = {Polymer embedding of membrane lungs for histological investigations of intra-device clot formation}, series = {Cardiovascular Medicine}, volume = {13}, journal = {Cardiovascular Medicine}, publisher = {Frontiers}, address = {Lausanne}, doi = {10.3389/fcvm.2026.1650978}, pages = {21}, abstract = {Extracorporeal membrane oxygenation (ECMO) is an invasive but potentially lifesaving treatment option for severe cardiac or respiratory failure. Despite its beneficial effect, coagulation-related complications, mainly due to clot formation, excessive bleeding and the accumulation of deposits in the membrane lung (ML) remain common, causing higher mortality. In this context, the formation of clots and other deposits in the ML is of particular interest. Previous histological examinations of the polymethylpentene fiber mats inside the ML could only be performed in a top view, prohibiting valid quantification and examination of the multi-layered deposits or fiber mat spanning structures. Our objective was the establishment of a polymer embedding to increase the mechanical stability of the deposits and thus enable cross-sectional microtome cutting through the ML hollow-fibers. Clinically used MLs (PLS, Getinge, Rastatt, Germany) were stabilized with a polymer resin (HistoCURE 8100). Specimens were cut out of the embedded MLs and microtome sections with a thickness of 10 µm were performed. In addition to standard histological staining with hematoxylin-eosin (HE) and Pappenheim (May-Grunwald-Giemsa), fluorescence DNA staining for nucleated cells with 4′,6-diamidino-2-phenylindole (DAPI) and SYTOX™ Green as well as immunohistochemical and immunofluorescence staining for the lysosomal enzyme myeloperoxidase (MPO) and von Willebrand factor (vWF) were established. The protocol provides a method for large volume embedding (400 mL). The cellular and extracellular deposits were securely fixed by the polymer scaffold allowing the examination of clots in MLs in native position which was not possible with conventional paraffin embedding. Multi-layered deposits and fiber mat spanning structures are no longer disrupted during specimen extraction and can now be quantified. Staining with HE, Pappenheim, DAPI, SYTOX™ Green, MPO, and vWF was successfully tested with this protocol. This method may be the foundation for new insights into the complex clotting phenomena observed in MLs}, language = {en} }