Biology subjects

Groll, J.

Publications and source records attributed to Groll, J..

6 recordsLinked to original sources

bioRxiv2026

Generation of Self-Organizing Macrovascular Constructs by Bioprinting human iPSC-Derived Mesodermal Progenitor Cells

Vascularization remains a major obstacle in tissue engineering. Here, we introduce a developmentally inspired bioprinting strategy to generate centimeter-scale, self-organising "mother vessel" constructs from iPSC-derived human mesodermal progenitor cells (hiMPCs). By systematically optimizing the bioink composition, we identified a formulation that combines high print fidelity, mechanical stability and cell compatibility within a single-step bioprinting process. Within the first week after printing, hiMPCs in the "mother vessel" constructs underwent spontaneous differentiation and morphogenesis, forming intima-, media-, and adventitia-like layers containing CD31 endothelial, SMA mural and CD34/CD150 progenitor cells. Remarkably, Iba1 macrophage-like cells appeared despite their absence in the initial population, indicating intrinsic differentiation into both vascular and non-vascular lineages essential for angiogenesis, remodeling and tissue homeostasis. Surrounding the newly formed vessel wall-like structure was a broad, vascularized mesodermal tissue compartment that also contained the above-mentioned progenitors. Co-culture with prevascularized mesodermal organoids resulted in early structural interconnection of microvessels with the printed wall, representing a prerequisite for subsequent hierarchical vascular network formation. As a proof-of-concept, the mother vessel withstood controlled flow conditions in a bioreactor without detectable leakage, demonstrating its principal suitability for perfusion analyses. Together, these findings establish a biologically driven platform that bridges macro- and microvascularization. This may pave the way toward perfusable, vascularized larger tissue constructs, a major bottleneck in regenerative biofabrication.

bioengineering↗

bioRxiv2024

Biofabrication of a filtration barrier by integrating electrospun membranes and flow in a glomerular co-culture

The glomerulus is the functional unit of the kidney, where urine is filtered from blood. This process happens through the glomerular filtration barrier (GFB) which is composed of glomerular endothelial cells, podocytes, and glomerular basement membrane (GBM). Damage to any component of GFB results in failure of the barriers function, causing proteinuria that can lead to end-stage kidney failure. There is a high need for reliable in vitro models of the GFB to study pathological conditions and to test potential novel therapeutic options. We established an artificial GBM generated by electrospinning of poly-L-lactic acid fibers that were coated with polydopamine and gelatin and seeded with human glomerular endothelial cells and podocytes onto different sides. The orientation of fibers in the artificial GBM, their surface chemistry and effects on glomerular cells were characterized in depth. Glomerular endothelial cells and podocytes, including hiPSC-derived podocytes formed monolayers on the artificial GBM and revealed cell type-specific marker expression and morphology. Cell-cell communication was possible between podocytes and glomerular endothelial cells in both directions though the artificial membrane. Different molecular dextrans showed size selective permeability of the ex vivo GFB model. Introduction of shear stress by applying flow with a 3D printed micro-bioreactor improved cellular ultrastructure with formation of glomerular endothelial cell fenestrae-like structures and long podocyte foot process-like protrusions that both are usually absent in other in vitro models. Personalized hiPSC-derived podocytes within our model will allow to study the role of patient-specific podocyte mutations or individual treatment response ex vivo in the future.

bioengineering↗

bioRxiv2023

Volumetric Printing across Melt Electrowritten Scaffolds Fabricates Multi-Material Living Constructs with Tunable Architecture and Mechanics

Major challenges in biofabrication revolve around capturing the complex, hierarchical composition of native tissues. However, individual 3D printing techniques have limited capacity to produce composite biomaterials with multi-scale resolution. Volumetric bioprinting recently emerged as a paradigm-shift in biofabrication. This ultra-fast, light-based technique sculpts cell-laden hydrogel bioresins into three-dimensional structures in a layerless fashion, providing unparalleled design freedom over conventional bioprinting. However, it yields prints with low mechanical stability, since soft, cell-friendly hydrogels are used. Herein, for the first time, the possibility to converge volumetric bioprinting with melt electrowriting, which excels at patterning microfibers, is shown for the fabrication of tubular hydrogel-based composites with enhanced mechanical behavior. Despite including non-transparent melt electrowritten scaffolds into the volumetric printing process, high-resolution bioprinted structures were successfully achieved. Tensile, burst and bending mechanical properties of printed tubes were tuned altering the electrowritten mesh design, resulting in complex, multi-material tubular constructs with customizable, anisotropic geometries that better mimic intricate biological tubular structures. As a proof-of-concept, engineered vessel-like structures were obtained by building tri-layered cell-laden vessels, and features (valves, branches, fenestrations) that could be resolved only by synergizing these printing methods. This multi-technology convergence offers a new toolbox for manufacturing hierarchical and mechanically tunable multi-material living structures.

bioengineering↗

bioRxiv2022

Single-molecule fluorescence microscopy demonstrates fast dynamics of the variant surface glycoprotein coat on living trypanosomes

The fluidity of Trypanosoma bruceis dense coat of GPI-anchored variant surface glycoproteins (VSGs) is fundamental for the survival of the parasite. In order to maintain the integrity of the coat, it is recycled on the time scale of a few minutes. This is surprisingly fast as endo- and exocytosis take place in the same small membrane invagination called the flagellar pocket. Here, we present measurements of VSG dynamics on the single-molecule level in living trypanosomes. A large number of short protein trajectories sampling the parasites surface were analysed in two distinct scenarios: diffusion and directed motion. To this end, we employed a previously published algorithm and implemented two extensions to consider rim effects as well as localisations errors inherent to single-mole tracking. Neglect of the latter can have a significant distortive effect on the measured diffusion coefficient; in our case resulting in an underestimation by 20 %. We found large heterogeneity in the local diffusion coefficients and velocities with a surprisingly high average value of [Formula] and [Formula], respectively. To decide on the locally dominant motion mode, we present a guideline based on random walk simulations. We find that VSG dynamics is indeed dominated by diffusion. Complementary simulations on long time scales not accessible in the experiment showed that passive VSG randomisation is fast enough to prevent re-endocytosis newly exocytosed VSGs and to accomplish turnover of the full VSG coat within a few minutes. Author summarySingle-molecule tracking in biological systems often suffers from trajectories being too short to obtain statistically robust decisions on the present motion mode. We faced this issue when investigating the dynamics of the protein surface coat of African trypanosomes. To address the question whether diffusion or directed motion governs coat dynamics, we have adopted an algorithm based on temporal decomposition and spatial binning of an ensemble of single-molecule trajectories. We introduced several extensions to the original approach, including the consideration of localisation errors inherent to single-molecule tracking. This improvement alone already prevented the diffusion coefficient from being underestimated by 20 %. We analysed the coat dynamics in two scenarios, diffusion and directed motion, and offer a decision guideline to identify the locally dominating motion mode. Our extended algorithm is available to the scientific community via GitHub. For the trypanosome surface coat we found that the motion is indeed mainly characterised by diffusion with a surprisingly high diffusion coefficient. This finding solves a long-standing question how the parasite maintains its protein coat.

biophysics↗

bioRxiv2021

Human iPSC-derived mesodermal progenitor cells preserve their vasculogenesis potential after extrusion and form hierarchically organized blood vessels

Post-fabrication formation of a proper vasculature remains an unresolved challenge in bioprinting. Established strategies focus on the supply of the fabricated structure with nutrients and oxygen and either rely on the mere formation of a channel system using fugitive inks, or additionally use mature endothelial cells and/or peri-endothelial cells such as smooth muscle cells for the formation of blood vessels in vitro. Functional vessels, however, exhibit a hierarchical organization and multilayered wall structure that is important for their function. Human induced pluripotent stem cell-derived mesodermal progenitor cells (hiMPCs) have been shown to possess the capacity to form blood vessels in vitro, but have so far not been assessed for their applicability in bioprinting processes. Here, we demonstrate that hiMPCs, after formulation into an alginate / collagen type 1 bioink and subsequent extrusion, retain their ability to give rise to the formation of complex vessels that display a hierarchical network in a process that mimicks the embryonic steps of vessel formation by vasculogenesis. Histological evaluations at different time points of extrusion revealed initial formation of spheres, followed by lumen formation and further structural maturation as evidenced by building a multilayered vessel wall and a vascular network. These findings are supported by immunostainings for endothelial and peri-endothelial cell markers as well as electron microscopic analyses at the ultrastructural level. Moreover, capillary-like vessel structures deposited a basement membrane-like matrix structure at the basal side between the vessel wall and the alginate-collagen matrix. These results evidence the applicability and great potential of hiMPCs for the bioprinting of vascular structures mimicking the basic morphogenetic steps of de novo vessel formation during embryogenesis.

bioengineering↗

bioRxiv2020

Differential cellular responses to adhesive interactions with galectin-8 and fibronectin coated substrates

The mechanisms underlying the cellular response to extracellular matrices (ECM), consisting of multiple adhesive ligands, each with distinct properties, are still poorly understood. Here we address this topic by monitoring the cellular responses to two very different extracellular adhesion molecules - fibronectin and galectin-8 - and to mixtures of the two. Fibronectin is one of the major integrin ligands, inducing cell spreading and development of focal adhesions associated with contractile stress fibers. Galectin-8 is a mammalian lectin, which specifically binds to {beta}-galactoside residues present on some integrins, as well as to other cell surface receptors. We found marked differences in HeLa-JW cell spreading, assembly of focal adhesions and actomyosin stress fibers, and formation of adherent filopodia, on rigid flat substrates functionalized by fibronectin or galectin-8 alone, or by mixtures of these two proteins. Spreading on galectin-8 resulted in a larger projected cell area compared to that on fibronectin, by more extensive formation of filopodia, coupled with an inability to activate focal adhesion and stress fiber assembly. These differences could be partially reversed by experimental manipulations of small G-proteins of the Rho family and their downstream targets, such as formins, the Arp2/3 complex, and Rho kinase. Another factor affecting the spreading process was shown to be the enhanced physical adhesion of the cells to galectin-8, as compared to fibronectin. Notably, at least one process, the formation of adherent filopodia, was synergistically upregulated by both ligands, so filopodia development on the substrate coated with a mixture of fibronectin and galectin-8 was far more prominent than on each ligand alone.

cell biology↗