@misc{BirkenmaierKrenkelLehle, author = {Birkenmaier, Clemens and Krenkel, Lars and Lehle, Karla}, title = {Linking flow conditions in membrane oxygenators to arrangements of multimeric von-Willebrand-factor as indication for coagulation}, series = {World Congress of Biomechanics 2018, Convention Centre Dublin, 8.-12. Juli 2018}, journal = {World Congress of Biomechanics 2018, Convention Centre Dublin, 8.-12. Juli 2018}, abstract = {Introduction Shear induced multimerisation of von-Willebrand-factor (vWF) is supposed to play an important role in coagulation inside extracorporeal membrane oxygenators. However, there is no proof that links observed vWF structures to computed or measured flow conditions. Methods The structures of multimeric vWF fibers, observed in clinically used membrane oxygenators is examined using immunofluorescence microscopy (IFM) using Carstairs' staining method (positive ethics committee vote). The flow around the membrane fibres inside the oxygenator is investigated in terms of shear rate, wall shear velocity and streamlines by using CFD (RANS, Carreau-Yasuda viscosity, geometry remodelled after high-resolution µCT-scans). By interpreting the histological and numerical results in this common context, indications for shear induced coagulation mechanisms can be identified. Results The fibre structures of multimeric vWF build regular but not exactly symmetric formations around the contact face (CF) between the crosswise stacked oxygenator fibres (OF), see fig.1B, vWF marked red. Annular around the CF arranged, cells are likely to be found, see fig.1B, nuclei marked blue. The computed streamlines around the OF show attached flow around the circular fibres. However, the irregular arrangement of real OF produce considerable cross flow between the interconnected neighbouring channels, in contrast to previous 2D-simulations. Thus, the CF are washed around closely by blood, also from neighbouring channels. The wall shear velocity streamlines form regular, slightly asymmetric shapes around the contact faces. The occurring maximum shear rates are in the range of 1,000 1/s. Discussion The shapes of vWF structures found in clinically used oxygenators match the computational results in terms of wall shear velocity and streamlines well. The accumulation of cells close to the CF can also be explained by fluid mechanics, as there are small shear gradients and slow velocities. However, occurring shear rates between OFs are too low to trigger multimerisation of vWF. That raises the question where in the circuit the actual activation of vWF is started and how, at least partly chained, vWF multimeres are attracted towards the OF surface. A next step will be the investigation of the actual shear rate triggered (or mediated) multimerisation of vWF. Towards this end, microfluidic experiments with shear triggered coagulation will be performed. Also of big interest is the computation of the flow situation in the oxygenator in proximity to chaining threads, which have been ignored in computations so far. However, first a realistic representation of the effective viscosity in computations is needed, which is not available yet.}, language = {en} } @article{PhilippdeSomerFoltanetal., author = {Philipp, Alois and de Somer, Filip and Foltan, Maik and Bredthauer, Andre and Krenkel, Lars and Zeman, Florian and Lehle, Karla}, title = {Life span of different extracorporeal membrane systems for severe respiratory failure in the clinical practice}, series = {PLOS ONE}, volume = {13}, journal = {PLOS ONE}, number = {6}, publisher = {PLOS}, doi = {10.1371/journal.pone.0198392}, pages = {1 -- 10}, abstract = {Over the past decade, veno-venous extracorporeal membrane oxygenation (vvECMO) has been increasingly utilized in respiratory failure in patients. This study presents our institution´s experience focusing on the life span of ECMO systems reflecting the performance of a particular system. A retrospective review of our ECMO database identified 461 adult patients undergoing vvECMO (2010-2017). Patients that required more than one system and survived the first exchange >24 hours (n = 139) were included. Life span until the first exchange and exchange criteria were analyzed for all systems (PLS, Cardiohelp HLS-set, both Maquet Cardiopulmonary, Rastatt, Germany; Deltastream/Hilite7000LT, iLA-activve, Xenios/NovaLung, Heilbronn, Germany; ECC.O5, LivaNova, Mirandola, Italy). At our ECMO center, the frequency of a system exchange was 30\%. The median (IQR) life span was 9 (6-12) days. There was no difference regarding the different systems (p = 0.145 and p = 0.108, respectively). However, the Deltastream systems were exchanged more frequently due to elective technical complications (e. g. worsened gas transfer, development of coagulation disorder, increased bleedings complications) compared to the other exchanged systems (p = 0.013). In summary, the used ECMO systems are safe and effective for acute respiratory failure. There is no evidence for the usage of a specific system. Only the increased predictability of an imminent exchange preferred the usage of a Deltastream system. However, the decision to use a particular system should not depend solely on the possible criteria for an exchange.}, language = {en} } @techreport{SteigerFoltanPhilippetal., type = {Working Paper}, author = {Steiger, Tamara and Foltan, Maik and Philipp, Alois and M{\"u}ller, Thomas and Gruber, Michael and Bredthauer, Andre and Krenkel, Lars and Birkenmaier, Clemens and Lehle, Karla}, title = {Accumulations of von Willebrand factor within ECMO oxygenators: Potential indicator of coagulation abnormalities in critically ill patients?}, abstract = {Clot formation within membrane oxygenators (MOs) remains a critical problem during extracorporeal membrane oxygenation (ECMO). The composition of the clots—in particular, the presence of von Willebrand factor (vWF)—may be an indicator for prevalent nonphysiological flow conditions, foreign body reactions, or coagulation abnormalities in critically ill patients. Mats of interwoven gas exchange fibers from randomly collected MOs (PLS, Maquet, Rastatt, Germany) of 21 patients were stained with antibodies (anti-vWF and anti-P-selectin) and counterstained with 4′,6-diamidino-2-phenylindole. The extent of vWF-loading was correlated with patient and technical data. While 12 MOs showed low vWF-loadings, 9 MOs showed high vWF-loading with highest accumulations close to crossing points of adjacent gas fibers. The presence and the extent of vWF-fibers/"cobwebs," leukocytes, platelet-leukocyte aggregates (PLAs), and P-selectin-positive platelet aggregates were independent of the extent of vWF-loading. However, the highly loaded MOs were obtained from patients with a significantly elevated SOFA score, severe thrombocytopenia, and persistent liver dysfunction. The coagulation abnormalities of these critically ill patients may cause an accumulation of the highly thrombogenic and elongated high-molecular-weight vWF multimers in the plasma which will be trapped in the MOs during the ECMO therapy.}, language = {en} } @inproceedings{BirkenmaierSteigerPhilippetal., author = {Birkenmaier, Clemens and Steiger, Tamara and Philipp, Alois and Lehle, Karla and Krenkel, Lars}, title = {Flow-induced accumulations of von Willebrand factor inside oxygenators during extracorporeal life support therapy}, series = {Proceedings of 12th International Conference BIOMDLORE 2018, June 28-30, 2018, Białystok, Poland}, booktitle = {Proceedings of 12th International Conference BIOMDLORE 2018, June 28-30, 2018, Białystok, Poland}, publisher = {IEEE}, address = {Piscataway, NJ}, isbn = {978-1-5386-2396-1}, doi = {10.1109/BIOMDLORE.2018.8467205}, pages = {3}, abstract = {BACKGROUND: Shear-induced conformational changes of von Willebrand factor (vWF) may be responsible for coagulation disorder and clot formation inside membrane oxygenators (MOs) during extracorporeal membrane oxygenation (ECMO) therapy. OBJECTIVE: The aim was to identify vWF structures inside clinically used MOs and employ computational fluid dynamics to verify the corresponding flow conditions. METHODS: Samples from gas exchange membranes (GEM) from MOs were analysed for accumulations of vWF and P-selectin-positive platelets using immunofluorescence techniques. Streamlines and shear rates of the flow around GEMs were computed using a laminar steady Reynolds-Averaged-Navier-Stokes approach. RESULTS: Most samples were colonized with equally distributed leukocytes, integrated in thin cobweb-like vWF-structures. Only 25 \% of the samples showed extended accumulations of vWF. Computed streamlines showed considerable cross flow between interconnected neighbouring channels. Stagnation points were non-symmetric and contact faces were washed around closely. The occurring maximum shear rates ranged from 2,500 to 3,000 1/s. CONCLUSIONS: If pronounced vWF structures are present, shape and extent match the flow computations well. Computed shear rates bear a critical degree of uncertainty due to the improper viscosity model. If flow conditions inside the MO were sufficient to affect vWF, a more consistent distribution of vWF across the samples should be present.}, language = {en} } @article{BirkenmaierDorniaLehleetal., author = {Birkenmaier, Clemens and Dornia, Christian and Lehle, Karla and M{\"u}ller, Thomas and Gruber, Michael and Philipp, Alois and Krenkel, Lars}, title = {Analysis of Thrombotic Deposits in Extracorporeal Membrane Oxygenators by High-resolution Microcomputed Tomography: A Feasibility Study}, series = {ASAIO Journal / American Society for Artificial Internal Organs}, volume = {66}, journal = {ASAIO Journal / American Society for Artificial Internal Organs}, number = {8}, publisher = {Lippincott Williams \& Wilkins}, issn = {1538-943X}, doi = {10.1097/MAT.0000000000001089}, pages = {922 -- 928}, abstract = {Coagulative disorders, especially clotting during extracorporeal membrane oxygenation, are frequent complications. Direct visualization and analysis of deposits in membrane oxygenators using computed tomography (CT) may provide an insight into the underlying mechanisms causing thrombotic events. However, the already established multidetector CT1 (MDCT) method shows major limitations. Here, we demonstrate the feasibility of applying industrial micro-CT (μCT) to circumvent these restrictions. Three clinically used membrane oxygenators were investigated applying both MDCT and μCT. The scans were analyzed in terms of clot volume and local clot distribution. As validation, the clot volume was also determined from the fluid volume, which could be filled into the respective used oxygenator compared to a new device. In addition, cross-sectional CT images were compared with crosscut oxygenators. Based on the μCT findings, a morphological measure (sphericity) for assessing clot structures in membrane oxygenators is introduced. Furthermore, by comparing MDCT and μCT results, an augmentation of the MDCT method is proposed, which allows for improved clot volume determination in a clinical setting.}, language = {en} } @misc{BirkenmaierDorniaLehleetal., author = {Birkenmaier, Clemens and Dornia, Christian and Lehle, Karla and Krenkel, Lars}, title = {Feasibility of detecting thrombotic deposits in membrane oxygenators using micro computed tomography}, series = {25th Congress of the European Society of Biomechanics, July 7-10, 2019, Vienna, Austria}, journal = {25th Congress of the European Society of Biomechanics, July 7-10, 2019, Vienna, Austria}, language = {en} } @article{ObermaierLehleSchmidetal., author = {Obermaier, Lisa and Lehle, Karla and Schmid, Stefanie and Schmid, Christof and Schratzenstaller, Thomas}, title = {Introduction of a new ex vivo porcine coronary artery model: Evaluation of the direct vascular injury after stent implantation with and without dogbone effect}, series = {European Surgical Research}, volume = {63}, journal = {European Surgical Research}, number = {4}, publisher = {Karger}, address = {Basel}, issn = {1421-9921}, doi = {10.1159/000527883}, pages = {285 -- 293}, abstract = {Introduction: Neointimal hyperplasia after percutaneous coronary intervention remains a major determinant of in-stent restenosis (ISR). The extent of mechanical vessel injury correlates with ISR. A new ex vivo porcine stent model was introduced and evaluated comparing different stent designs. Methods: Coronary arteries were prepared from pig hearts from the slaughterhouse and used for ex vivo implantations of coronary stents. One basic stent design in two configurations (dogbone, DB; non-dogbone, NDB) was used. Vascular injury was determined according to a modified injury score (IS). Results: Standardized experimental conditions ensured comparable vessel dimensions and overstretch data. DB stents caused more severe IS compared to NDB stents. The mean IS and the IS at the distal end of all stents were significantly reduced for NDB stents (ISMean, DB, 1.16 ±0.12; NDB, 1.02 ±0.12; p=0.018; ISDist, DB, 1.39 ±0.28; NDB, 1.13 ±0.24; p=0.03). Discussion/Conclusion: The introduced ex-vivo model allowed the evaluation of different stent designs exclude unfavorable stent designs.}, language = {en} } @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{HoenickaLehleJacobsetal., author = {Hoenicka, Markus and Lehle, Karla and Jacobs, V. R. and Dendorfer, Sebastian and Kostorz, A. and Schmid, F. X. and Birnbaum, D. E.}, title = {Mechanical and seeding properties of human umbilical vein - a potential scaffold for a tissue-engineered vessel graft}, series = {The Thoracic and Cardiovascular Surgeon}, volume = {55}, journal = {The Thoracic and Cardiovascular Surgeon}, number = {S 1}, publisher = {Thieme}, doi = {10.1055/s-2007-967592}, pages = {P_37}, abstract = {Objectives: The mechanical properties and seeding with endothelial cells were investigated in fresh and cryopreserved human umbilical vein. Methods: Human umbilical veins (HUV) were frozen in Euro-Collins/1M DMSO at -1°C/min and stored in liquid nitrogen. Stress-strain relationships of fresh and thawed veins were determined in an uniaxial tension-testing rig. HUV endothelial cells (HUVEC) were seeded onto denuded HUV under static conditions and grown for 3d. Luminal surfaces were analyzed by scanning electron microscopy. Calcein-stained cells were seeded hyperconfluently to determine the cell retention capacity of fresh and cryopreserved veins. Results: The stress-strain relationships of HUV followed a biphasic pattern typical for natural vessels. Neither the failure stress (2.71±0.36 vs. 3.25±0.97 N, n=3) nor the displacement required to achieve failure (9.73±0.9 vs. 7.43±2.07mm, n=3) were altered by cryopreservation. The burst pressure was estimated as approx. 1000mm Hg within the limitations of the uniaxial model. HUVEC seeded onto denuded HUV formed patches (at 9E3 cells per cm2) or an almost confluent endothelium (at 3E4 cells per cm2) within three days. The capacity to retain seeded HUVEC of denuded HUV was not altered by cryopreservation (1.15±0.08E5 vs. 1.26±0.14E5 cells per cm2, n=6). Conclusions: The burst pressure of HUV seems to be sufficiently high for the human arterial circulation and is not altered by cryopreservation. HUVEC can establish a confluent endothelium on denuded HUV. Therefore HUV appears to be a suitable storable scaffold for vascular tissue engineering.}, subject = {Nabelvene}, 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{FoltanDinhGruberetal., author = {Foltan, Maik and Dinh, D. and Gruber, Michael and M{\"u}ller, Thomas and Hart, C. and Krenkel, Lars and Schmid, C. and Lehle, Karla}, title = {Incidence of neutrophil extracellular traps (NETs) in different membrane oxygenators: pilot in vitro experiments in commercially available coated membranes}, series = {Journal of artificial organs : the official journal of the Japanese Society for Artificial Organs}, journal = {Journal of artificial organs : the official journal of the Japanese Society for Artificial Organs}, doi = {10.1007/s10047-024-01486-4}, abstract = {Neutrophil extracellular traps (NETs) were detected in blood samples and in cellular deposits of oxygenator membranes during extracorporeal membrane oxygenation (ECMO) therapy and may be responsible for thrombogenesis. The aim was to evaluate the effect of the base material of gas fiber (GF, polymethylpentene) and heat exchange (HE) membranes and different antithrombogenic coatings on isolated granulocytes from healthy volunteers under static culture conditions. Contact of granulocytes with membranes from different ECMO oxygenators (with different surface coatings) and uncoated-GFs allowed detection of adherent cells and NETotic nuclear structures (normal, swollen, ruptured) using nuclear staining. Flow cytometry was used to identify cell activation (CD11b/CD62L, oxidative burst) of non-adherent cells. Uncoated-GFs were used as a reference. Within 3 h, granulocytes adhered to the same extent on all surfaces. In contrast, the ratio of normal to NETotic cells was significantly higher for uncoated-GFs (56-83\%) compared to all coated GFs (34-72\%) (p < 0.001) with no difference between the coatings. After material contact, non-adherent cells remained vital with unchanged oxidative burst function and the proportion of activated cells remained low. The expression of activation markers was independent of the origin of the GF material. In conclusion, the polymethylpentene surfaces of the GFs already induce NET formation. Antithrombogenic coatings can already reduce the proportion of NETotic nuclei. However, it cannot be ruled out that NET formation can induce thrombotic events. Therefore, new surfaces or coatings are required for future ECMO systems and long-term implantable artificial lungs.}, 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{SteigerFoltanPhilippetal., author = {Steiger, Tamara and Foltan, Maik and Philipp, Alois and M{\"u}ller, Thomas and Gruber, Michael and Bredthauer, Andre and Krenkel, Lars and Birkenmaier, Clemens and Lehle, Karla}, title = {Accumulations of von Willebrand factor within ECMO oxygenators: Potential indicator of coagulation abnormalities in critically ill patients?}, series = {Artificial Organs}, volume = {43}, journal = {Artificial Organs}, number = {11}, publisher = {Wiley}, address = {Hoboken}, issn = {1525-1594}, doi = {10.1111/aor.13513}, pages = {1065 -- 1076}, abstract = {Clot formation within membrane oxygenators (MOs) remains a critical problem during extracorporeal membrane oxygenation (ECMO). The composition of the clots-in particular, the presence of von Willebrand factor (vWF)-may be an indicator for prevalent nonphysiological flow conditions, foreign body reactions, or coagulation abnormalities in critically ill patients. Mats of interwoven gas exchange fibers from randomly collected MOs (PLS, Maquet, Rastatt, Germany) of 21 patients were stained with antibodies (anti-vWF and anti-P-selectin) and counterstained with 4 ',6-diamidino-2-phenylindole. The extent of vWF-loading was correlated with patient and technical data. While 12 MOs showed low vWF-loadings, 9 MOs showed high vWF-loading with highest accumulations close to crossing points of adjacent gas fibers. The presence and the extent of vWF-fibers/"cobwebs," leukocytes, platelet-leukocyte aggregates (PLAs), and P-selectin-positive platelet aggregates were independent of the extent of vWF-loading. However, the highly loaded MOs were obtained from patients with a significantly elevated SOFA score, severe thrombocytopenia, and persistent liver dysfunction. The coagulation abnormalities of these critically ill patients may cause an accumulation of the highly thrombogenic and elongated high-molecular-weight vWF multimers in the plasma which will be trapped in the MOs during the ECMO therapy.}, language = {en} } @article{HausFoltanPhilippetal., author = {Haus, Moritz and Foltan, Maik and Philipp, Alois and M{\"u}ller, Thomas and Lingel, Maximilian P. and Krenkel, Lars and Gruber, Michael and Lehle, Karla}, title = {Neutrophil extracellular traps -a potential trigger for the development of thrombocytopenia during extracorporeal membrane oxygenation}, volume = {15}, publisher = {frontiers}, doi = {10.3389/fimmu.2024.1339235}, abstract = {Neutrophil extracellular traps (NETs) have recently emerged as a potential link between inflammation, immunity, and thrombosis, as well as other coagulation disorders which present a major challenge in the context of extracorporeal membrane oxygenation (ECMO). By examining blood from ECMO patients for NETs and their precursors and correlating them with clinical and laboratory biomarkers of coagulation and inflammation, this study aims to evaluate the association between the presence of NETs in the bloodstream of ECMO patients and the development of potentially severe coagulation disorders during ECMO therapy. Therefore, blood samples were collected from healthy volunteers (n=13) and patients receiving veno-venous (VV) ECMO therapy (n=10). To identify NETs and their precursors, DNA and myeloperoxidase as well as granulocyte marker CD66b were visualized simultaneously by immunofluorescence staining in serial blood smears. Differentiation of DNA-containing objects and identification of NETs and their precursors was performed semiautomatically by a specific algorithm using the shape and size of DNA staining and the intensity of MPO and CD66b signal. Neutrophil extracellular traps and their precursors could be detected in blood smears from patients requiring VV ECMO. Compared to volunteers, ECMO patients presented significantly higher rates of NETs and NET precursors as well as an increased proportion of neutrophil granulocytes in all detected nucleated cells. A high NET rate prior to the initiation of ECMO therapy was associated with both increased iL-6 and TNF-α levels as an expression of a high cytokine burden. These patients with increased NET release also presented an earlier and significantly more pronounced decrease in platelet counts and ATIII activity following initiation of therapy compared with patients with less elevated NETs. These findings provide further indications for the development of immune-mediated acquired thrombocytopenia in ECMO patients.}, 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} } @inproceedings{KranzPointnerWagneretal., author = {Kranz, Michael and Pointner, Daniel and Wagner, Maria Stella and Lubnow, Matthias and Lehle, Karla and Krenkel, Lars}, title = {High-Resolution Flow Investigations in Membrane-Lungs for Understanding Shear-Induced Blood Clot Formation}, series = {New Results in Numerical and Experimental Fluid Mechanics XV : Contributions to the 24th STAB/DGLR Symposium, Regensburg, Germany, 2024}, booktitle = {New Results in Numerical and Experimental Fluid Mechanics XV : Contributions to the 24th STAB/DGLR Symposium, Regensburg, Germany, 2024}, editor = {Dillmann, Andreas and Heller, Gerd and Kr{\"a}mer, Ewald and Breitsamter, Christian and Wagner, Claus and Krenkel, Lars}, publisher = {Springer}, address = {Cham}, doi = {10.1007/978-3-032-11115-9_12}, pages = {125 -- 134}, abstract = {Complex blood flow phenomena in membrane lungs (MLs) play a crucial role in intra-device clot formation and the occurrence of thromboembolic events. At present, however, the local flow conditions within an ML are not yet sufficiently known. The aim was to gain a deeper understanding of local flow regimes inside MLs by performing highly resolved computational fluid dynamics (CFD) of generic and native fiber mat bundles. Straight cylinders with a diameter of 380 μm in parallel arrangement were the foundation of the generic model. For validation, a method for reconstructing a native geometry from a microcomputed tomography (μCT) scan was established, with both models used for CFD. While the generic model showed a symmetrical flow regime without indicating any pathological flow, the native model did show an irregular fiber arrangement and no symmetrical flow regime. In conclusion, the fiber arrangement significantly affects the local flow regimes inside MLs.}, language = {en} } @misc{KranzPointnerLehleetal., author = {Kranz, Michael and Pointner, Daniel and Lehle, Karla and Lubnow, Matthias and Krenkel, Lars}, title = {High-resolution flow field investigations in membrane lungs, considering the complex blood rheology}, series = {1st European Fluid Dynamics Conference (EFDC1), 16-20.September 2024, Aachen}, journal = {1st European Fluid Dynamics Conference (EFDC1), 16-20.September 2024, Aachen}, doi = {10.35096/othr/pub-8921}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:898-opus4-89214}, pages = {2}, abstract = {Despite major improvements over the last years, coagulative disorders and clotting phenomena in membrane lungs (MLs) are still considerable complications in extracorporeal membrane oxygenation (ECMO). ECMO is an increasingly used treatment for patients with severe respiratory failure or cardiac arrest [1]. For both, evaluation of therapeutic decisions and fundamental research on patient specific intra-device clotting phenomena, the direct visualization and analysis of clot formation in combination with a detailed flow field correlation is highly desirable and therefore an intensively followed research topic. Modelling blood flow and shear induced coagulation in MLs is challenging. The relevant geometry of oxygenator fibers and chaining threads is complex and spans several length scales. In relevant scales and regimes, blood shows several significant non-Newtonian effects. Viscosity impacts shear rate, which is important in several coagulation mechanisms. Additionally, coagulation processes are influencing fluid properties and geometry significantly. Existing approaches of previous research work are only able to consider some, but not all relevant effects and geometrical details. Due to the enormous size of the discretized geometries, highly detailed viscosity and coagulations models are not applicable. Our goal is to develop a model for combined viscosity and coagulation properties of blood flow in MLs. In our work, we compare the influence of different levels of detail of the ML geometry as well as the influence of considering realistic blood flow behavior (viscosity change by considering the local hematocrit distribution within the F{\aa}hraeus-Lindqvist-Effect) on the resulting flow field in relevant subsections of a ML. High-resolution micro-CT geometry reconstructions [1] are compared to idealized generic fiber representations. For realistic blood flow modelling, Newtonian representation is compared to the established Carreau-Yasuda and a multiphase Euler-Euler approach. Results are presented for relevant subsections as well as for the complete ML.}, language = {en} } @misc{KranzWagnerPointneretal., author = {Kranz, Michael and Wagner, Maria Stella and Pointner, Daniel and Waldbauer, Selina and M{\"u}ller, Thomas and Lubnow, Matthias and Foltan, Maik and Krenkel, Lars and Lehle, Karla}, title = {Polymeric Embedding of Membrane Lungs: A Novel Method for Histological Investigations of Intra-Device Clot Formation}, series = {12th EuroELSO Congress, 24-27. April 2024, Krakow}, journal = {12th EuroELSO Congress, 24-27. April 2024, Krakow}, language = {en} } @misc{KranzWagnerKrenkeletal., author = {Kranz, Michael and Wagner, Maria Stella and Krenkel, Lars and M{\"u}ller, Thomas and Lubnow, Matthias and Philipp, Alois and Lehle, Karla}, title = {Clot Localization within Membrane Lungs using different Imaging Methods and Histological Clot Characterization as a way to prevent Thrombosis in Extracorporeal Membrane Oxygenation}, volume = {2023}, language = {en} } @misc{KoesterKranzWagneretal., author = {K{\"o}ster, Leonie and Kranz, Michael and Wagner, Maria Stella and Foltan, Maik and M{\"u}ller, Thomas and Lubnow, Matthias and Krenkel, Lars and Lehle, Karla}, title = {Histological Investigations of Intra-Device Clot Formation in ECMO Pumps}, series = {12th EuroELSO Congress, 24-27. April 2024, Krakow}, journal = {12th EuroELSO Congress, 24-27. April 2024, Krakow}, language = {en} } @article{PointnerKranzWagneretal., author = {Pointner, Daniel and Kranz, Michael and Wagner, Maria Stella and Haus, Moritz and Lehle, Karla and Krenkel, Lars}, title = {Automated deep learning based detection of cellular deposits on clinically used ECMO membrane lungs}, series = {Frontiers in Bioinformatics}, volume = {6}, journal = {Frontiers in Bioinformatics}, publisher = {Frontiers}, doi = {10.3389/fbinf.2026.1771574}, pages = {18}, abstract = {Introduction: Despite the promising application of extracorporeal membrane oxygenation (ECMO) in the treatment of critically ill patients, coagulation-associated technical complications, primarily clot formation and critical bleeding, remain a major challenge during ECMO therapy. The deposition of nucleated cells on the surface has been shown, yet the role of these cells towards complication development is still matter of ongoing research. In particular, the membrane lung (MemL) is prone to clot formation. Therefore, the investigation of nuclear deposits on its hollow-fibers may provide insights for a better understanding of the cellular mechanisms involved in the development of ECMO complications. Methods: To support current research, this study aimed to develop a deep learning-based tool for the automated detection and quantitative analysis of nuclear depositions on MemL hollow-fiber mats. A customized fluorescence microscopy workflow, combined with a semi-automated iterative labeling strategy, was used to generate a high-quality dataset for model training. Results: Six configurations of instance segmentation models were evaluated, with a Mask R-CNN with ResNet 101 backbone using dilated convolution providing the most balanced performance in both nuclei count and area accuracy. Compared with U-Net-based approaches such as Cellpose or StarDist, the proposed model demonstrated superior segmentation of overlapping and low-intensity nuclei, maintaining accuracy even in densely packed cellular regions. Discussion: We present an automated image analysis tool for clinically used MemLs, which exhibit complex three-dimensional hollow-fiber architectures and irregular cellular deposits that challenge conventional tools. A dedicated graphical user interface enables streamlined detection, morphometric analysis, and spatial clustering of nuclei, establishing a reproducible workflow for high-throughput analysis of fluorescence microscopy images. This approach eliminates labor-intensive manual counting and facilitates large-scale studies on cell-fiber interactions and disease-related correlations.}, language = {en} }