TY - JOUR A1 - Sales, Adria A1 - Picart, Catherine A1 - Kemkemer, Ralf T1 - Age-dependent migratory behavior of human endothelial cells revealed by substrate microtopography JF - Experimental cell research : ECR N2 - Cell migration is part of many important in vivo biological processes and is influenced by chemical and physical factors such as substrate topography. Although the migratory behavior of different cell types on structured substrates has already been investigated, up to date it is largely unknown if specimen's age affects cell migration on structures. In this work, we investigated age-dependent migratory behavior of human endothelial cells from young (≤ 31 years old) and old (≥ 60 years old) donors on poly(dimethylsiloxane) microstructured substrates consisting of well-defined parallel grooves. We observed a decrease in cell migration velocity in all substrate conditions and in persistence length perpendicular to the grooves in cells from old donors. Nevertheless, in comparison to young cells, old cells exhibited a higher cell directionality along grooves of certain depths and a higher persistence time. We also found a systematic decrease of donor age dependent responses of cell protrusions in orientation, velocity and length, all of them decreased in old cells. These observations lead us to hypothesize a possible impairment of actin cytoskeleton network and affected actin polymerization and steering systems, caused by aging. KW - endothelial cells KW - microtopography KW - aging KW - migration KW - protrusions Y1 - 2019 U6 - http://dx.doi.org/10.1016/j.yexcr.2018.10.008 SN - 0014-4827 VL - 374 IS - 1 SP - 1 EP - 11 PB - Elsevier ER - TY - JOUR A1 - Tendulkar, Gauri A1 - Sreekumar, Vrinda A1 - Rupp, Frank A1 - Teotia, Arun A1 - Athanasopulu, Kiriaki A1 - Kemkemer, Ralf T1 - Characterisation of porous knitted titanium for replacement of intervertebral disc nucleus pulposus JF - Scientific reports N2 - Effective restoration of human intervertebral disc degeneration is challenged by numerous limitations of the currently available spinal fusion and arthroplasty treatment strategies. Consequently, use of artificial biomaterial implant is gaining attention as a potential therapeutic strategy. Our study is aimed at investigating and characterizing a novel knitted titanium (Ti6Al4V) implant for the replacement of nucleus pulposus to treat early stages of chronic intervertebral disc degeneration. Specific knitted geometry of the scaffold with a porosity of 67.67 ± 0.824% was used to overcome tissue integration failures. Furthermore, to improve the wear resistance without impairing original mechanical strength, electro-polishing step was employed. Electro-polishing treatment changed a surface roughness from 15.22 ± 3.28 to 4.35 ± 0.87 μm without affecting its wettability which remained at 81.03 ± 8.5°. Subsequently, cellular responses of human mesenchymal stem cells (SCP1 cell line) and human primary chondrocytes were investigated which showed positive responses in terms of adherence and viability. Surface wettability was further enhanced to super hydrophilic nature by oxygen plasma treatment, which eventually caused substantial increase in the proliferation of SCP1 cells and primary chondrocytes. Our study implies that owing to scaffolds physicochemical and biocompatible properties, it could improve the clinical performance of nucleus pulposus replacement. Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bsz:rt2-opus4-17520 SN - 2045-2322 VL - 7 IS - article nr. 16611 SP - 1 EP - 12 PB - Nature Publishing Group CY - London ER - TY - JOUR A1 - Kutuzova, Larysa A1 - Athanasopulu, Kiriaki A1 - Schneider, Markus A1 - Kandelbauer, Andreas A1 - Kemkemer, Ralf A1 - Lorenz, Günter T1 - In vitro bio-stability screening of novel implantable polyurethane elastomers : morphological design and mechanical aspects JF - Current directions in biomedical engineering N2 - A series of novel biomedical TPCUs with different percentages of hard segment and a silicone component in the soft segment were synthesized in a multi stage one-pot method. The kinetic profiles of the urethane formation in TPCU-based copolymer systems were monitored by rheological, in line FTIR spectroscopic (React IR) and real-time calorimetric (RC1) methods. This process-analytically monitored multi step synthesis was successfully used to optimize the production of medical-grade TPCU elastomers on preparative scale (in lots of several kg) with controlled molecular structure and mechanical properties. Various surface and bulk analytical methods as well as systematic studies of the mechanic response of the elastomer end-products towards compression and tensile loading were used to estimate the bio-stability of the prepared TPCUs in vitro after 3 months. The tests suggested that high bio-stability of all polyurethane formulations using accelerating in vitro test can be attributed to the synthetic design as well as to the specific techniques used for specimen preparation, namely: (1) the annealing for reducing residual polymer surface stress and preventing IES, (2) stabilization of the morphology by long time storage of the specimens after processing before being immersed in the test liquids, (3) purification by extraction to remove the shot chain oligomers which are the most susceptible to degradation. All mechanical tests were performed on cylindrical and circular disc specimens for modelling the thickness of the meniscus implants under application-relevant stress conditions. KW - in vitro KW - long-term implants KW - fatigue KW - bio-stable polyurethane KW - morphological design KW - large scale production Y1 - 2018 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bsz:rt2-opus4-21695 SN - 2364-5504 VL - 4 IS - 1 SP - 535 EP - 538 PB - De Gruyter CY - Berlin ER - TY - JOUR A1 - Frey, Kerstin A1 - Fischer, Alena A1 - Krastev, Rumen A1 - Kemkemer, Ralf T1 - CurvChip - chip platform for investigating cell responses to curved surface features JF - Current directions in biomedical engineering N2 - Surface topographies are often discussed as an important parameter influencing basic cell behavior. Whereas most in vitro studies deal with microstructures with sharp edges, smooth, curved microscale topographies might be more relevant concerning in-vivo situations. Addressing the lack of highly defined surfaces with varying curvature, we present a topography chip system with 3D curved features of varying spacing, curvature radii as well as varying overall dimensions of curved surfaces. The CurvChip is produced by low-cost photolithography with thermal reflow, subsequent (repetitive) PDMS molding and hot embossing. The platform facilitates the systematic in-vitro investigation of the impact of substrate curvature on cell types like epithelial, endothelial, smooth muscle cells, or stem cells. Such investigations will not only help to further understand the mechanism of curvature sensation but may also contribute to optimize cell-material interactions in the field of regenerative medicine. KW - photolithography KW - thermal reflow KW - topography with defined 3D curvature KW - chip platform Y1 - 2018 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bsz:rt2-opus4-21505 SN - 2364-5504 VL - 4 IS - 1 SP - 453 EP - 456 PB - De Gruyter CY - Berlin ER - TY - JOUR A1 - Sales, Adria A1 - Wang, Fanlu A1 - Chen, Hao A1 - Kemkemer, Ralf A1 - Greiner, Alexandra T1 - Cell-age and cell type-dependent behavior of human vascular cells on micro-structured or soft polymer substrates JF - Biomedizinische Technik / Biomedical Engineering N2 - Increasing number of studies are focused on how adherent cells respond, in vitro, to different properties of a material. Typical properties are the surface chemistry, topographical cues (at the nano- and micro-scale) of the surface, and the substrate stiffness. Cell Response studies are of importance for designing new biomaterials with applications in cell culture technologies, regenerative medicine, or for medical implants. However, only very few studies take the cell age factor, respectively the donor age, into account. In this work, we tested two types of human vascular cells (smooth muscle and endothelial cells) from old and young donors on (a) micro-structured surfaces made of pol (dimethylsiloxane) or on (b) flat polyacrylamide hydrogels with varying stiffnesses. These experiments reveal age-dependent and cell typedependent differences in the cell response to the topography and stiffness, and may establish the Basis for further studies focusing on cell age-dependent responses. Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bsz:rt2-opus4-801 SN - 1862-278X VL - 59 IS - s1 : Track D - Cellular, Tissue and Bioengineering SP - S286 EP - S289 PB - de Gruyter CY - Berlin ER - TY - JOUR A1 - Nisenholz, Noam A1 - Rajendran, Kavitha A1 - Dang, Quynh A1 - Chen, Hao A1 - Kemkemer, Ralf A1 - Krishnan, Ramaswamy A1 - Zemel, Assaf T1 - Active mechanics and dynamics of cell spreading on elastic substrates JF - Soft matter N2 - The spreading area of cells has been shown to play a central role in the determination of cell fate and tissue morphogenesis; however, a clear understanding of how spread cell area is determined is still lacking. The observation that cell area and force generally increase with substrate rigidity suggests that cell area is dictated mechanically, by means of a force-balance between the cell and the substrate. A simple mechanical model, corroborated by experimental measurements of cell area and force is presented to analyze the temporal force balance between the cell and the substrate during spreading. The cell is modeled as a thin elastic disc that is actively pulled by lamellipodia protrusions at the cell front. The essential molecular mechanisms of the motor activity at the cell front, including, actin polymerization, adhesion kinetics, and the actin retrograde flow, are accounted for and used to predict the dynamics of cell spreading on elastic substrates; simple, closed-form expressions for the evolution of cell size and force are derived. Time-resolved, traction force microscopy, combined with measurements of cell area are performed to investigate the simultaneous variations of cell size and force. We find that cell area and force increase simultaneously during spreading but the force develops with an apparent delay relative to the increase in cell area. We demonstrate that this may reflect the strain-stiffening property of the cytoskeleton. We further demonstrate that the radial cell force is a concave function of spreading speed and that this may reflect the strengthening of cell–substrate adhesions during spreading. Y1 - 2015 U6 - http://dx.doi.org/10.1039/C4SM00780H SN - 1744-683X VL - 10 IS - 37 SP - 7234 EP - 7246 PB - Royal Society of Chemistry CY - London ER - TY - JOUR A1 - Thies, Christian A1 - Khachaturyan, Galina A1 - Zemel, Assaf A1 - Kemkemer, Ralf T1 - Image analysis of self-organized multicellular patterns : multicellular pattern formation on compliant elastomer surfaces as model system JF - Current directions in biomedical engineering N2 - Analysis of multicellular patterns is required to understand tissue organizational processes. By using a multi-scale object oriented image processing method, the spatial information of cells can be extracted automatically. Instead of manual segmentation or indirect measurements, such as general distribution of contrast or flow, the orientation and distribution of individual cells is extracted for quantitative analysis. Relevant objects are identified by feature queries and no low-level knowledge of image processing is required. KW - cell segmentation KW - image analysis KW - object extraction KW - phase contrast micrographs KW - spatial information KW - tissue organization Y1 - 2016 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bsz:rt2-opus4-12348 VL - 2 IS - 1 SP - 523 EP - 527 PB - De Gruyter CY - Berlin ER - TY - JOUR A1 - Grützmacher, Simon A1 - Kemkemer, Ralf A1 - Thies, Christian A1 - Curio, Cristóbal T1 - Detecting lamellipodia in epithelial cell clusters using a fully convolutional neural network for phase contrast microscopy images JF - Current directions in biomedical engineering N2 - We present an approach for segmenting individual cells and lamellipodia in epithelial cell clusters using fully convolutional neural networks. The method will set the basis for measuring cell cluster dynamics and expansion to improve the investigation of collective cell migration phenomena. The fully learning-based front-end avoids classical feature engineering, yet the network architecture needs to be designed carefully. Our network predicts how likely each pixel belongs to one of the classes and, thus, is able to segment the image. Besides characterizing segmentation performance, we discuss how the network will be further employed. KW - lamellipodia KW - convolutional neural network Y1 - 2018 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bsz:rt2-opus4-21877 SN - 2364-5504 VL - 4 IS - 1 SP - 449 EP - 452 PB - De Gruyter CY - Berlin ER - TY - JOUR A1 - Mellert, Kevin A1 - Lechner, Stefan A1 - Lüdeke, Manuel A1 - Lamla, Markus A1 - Möller, Peter A1 - Kemkemer, Ralf A1 - Scheffzek, Klaus A1 - Kaufmann, Dieter T1 - Restoring functional neurofibromin by protein transduction JF - Scientific reports N2 - In Neurofibromatosis 1 (NF1) germ line loss of function mutations result in reduction of cellular neurofibromin content (NF1+/−, NF1 haploinsufficiency). The Ras-GAP neurofibromin is a very large cytoplasmic protein (2818 AA, 319 kDa) involved in the RAS-MAPK pathway. Aside from regulation of proliferation, it is involved in mechanosensoric of cells. We investigated neurofibromin replacement in cultured human fibroblasts showing reduced amount of neurofibromin. Full length neurofibromin was produced recombinantly in insect cells and purified. Protein transduction into cultured fibroblasts was performed employing cell penetrating peptides along with photochemical internalization. This combination of transduction strategies ensures the intracellular uptake and the translocation to the cytoplasm of neurofibromin. The transduced neurofibromin is functional, indicated by functional rescue of reduced mechanosensoric blindness and reduced RasGAP activity in cultured fibroblasts of NF1 patients or normal fibroblasts treated by NF1 siRNA. Our study shows that recombinant neurofibromin is able to revert cellular effects of NF1 haploinsuffiency in vitro, indicating a use of protein transduction into cells as a potential treatment strategy for the monogenic disease NF1. Y1 - 2018 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bsz:rt2-opus4-21493 SN - 2045-2322 VL - 8 IS - Aufsatz 6171 SP - 1 EP - 9 PB - Macmillan Publishers Limited CY - London ER - TY - JOUR A1 - Greiner, Alexandra A1 - Sales, Adria A1 - Chen, Hao A1 - Biela, Sarah A1 - Kaufmann, Dieter A1 - Kemkemer, Ralf T1 - Nano- and microstructured materials for in vitro studies of the physiology of vascular cells JF - Beilstein journal of nanotechnology N2 - The extracellular environment of vascular cells in vivo is complex in its chemical composition, physical properties, and architecture. Consequently, it has been a great challenge to study vascular cell responses in vitro, either to understand their interaction with their native environment or to investigate their interaction with artificial structures such as implant surfaces. New procedures and techniques from materials science to fabricate bio-scaffolds and surfaces have enabled novel studies of vascular cell responses under well-defined, controllable culture conditions. These advancements are paving the way for a deeper understanding of vascular cell biology and materials–cell interaction. Here, we review previous work focusing on the interaction of vascular smooth muscle cells (SMCs) and endothelial cells (ECs) with materials having micro- and nanostructured surfaces. We summarize fabrication techniques for surface topographies, materials, geometries, biochemical functionalization, and mechanical properties of such materials. Furthermore, various studies on vascular cell behavior and their biological responses to micro- and nanostructured surfaces are reviewed. Emphasis is given to studies of cell morphology and motility, cell proliferation, the cytoskeleton and cell-matrix adhesions, and signal transduction pathways of vascular cells. We finalize with a short outlook on potential interesting future studies. KW - fabrication methods KW - materials selection KW - nano- and micro-topography KW - vascular endothelial cells KW - vascular smooth muscle cells Y1 - 2016 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bsz:rt2-opus4-10710 SN - 2190-4286 VL - 7 SP - 1620 EP - 1641 PB - Beilstein-Institut zur Förderung der Chemischen Wissenschaften CY - Frankfurt am Main ER -