Translate this page to:
Choose Language…
In JoVE (3)
- Chip-based Three-dimensional Cell Culture in Perfused Micro-bioreactors
- Microfabrication of Chip-sized Scaffolds for Three-dimensional Cell cultivation
- Interview: Bioreactors and Surfaced-Modified 3D-Scaffolds for Stem Cell Research
Other Publications (4)
Articles by Karl-Friedrich Weibezahn in JoVE
JoVE General
Chip-based Three-dimensional Cell Culture in Perfused Micro-bioreactors
Institute for Biological Interfaces, Forschungszentrum Karlsruhe
We describe a chip-based platform for the three-dimensional cultivation of cells in micro-bioreactors. One chip can house up to 10 Mio. cells that can be cultivated under precisely defined conditions with regard to fluid flow, oxygen tension etc. in a sterile, closed circulation loop.
JoVE General
Microfabrication of Chip-sized Scaffolds for Three-dimensional Cell cultivation
1Institute for Biological Interfaces, Karlsruhe Research Centre, 2Institute for BioMedical Technology, University of Twente, 3Department of Materials Research, Institute for Heavy Ion Research, 4Institute of Microstructure Technology, Karlsruhe Research Centre, 5Institute for Micro Process Engineering, Karlsruhe Research Centre
We present two processes for the microfabrication of porous polymer chips for three-dimensional cell cultivation. The first one is hot embossing combined with a solvent vapour welding process. The second one uses a recently developed microthermoforming process combined with ion track technology leading to a significant simplification of manufacture.
JoVE General
Interview: Bioreactors and Surfaced-Modified 3D-Scaffolds for Stem Cell Research
Institute for Biological Interfaces, Karlsruhe Institute of Technology
In the past many in vitro culture systems -- mainly monolayer cultures -- often suffered from the disadvantage that differentiated primary cells had a relatively short life-span and de-differentiated during culture. As a consequence, most of their organ-specific functions were lost rapidly. Thus, in order to reproduce better conditions for these cells in vitro, modifications and adaptations have been made to conventional monolayer cultures.
Other articles by Karl-Friedrich Weibezahn on PubMed
Microstructured Scaffolds for Liver Tissue Cultures of High Cell Density: Morphological and Biochemical Characterization of Tissue Aggregates
Very high cell densities and optimal vascularization characterize among others organs and tissues in vivo. In order to study organ-specific functions in vitro or to make use of them in medical devices/treatments in the future, this natural architecture should be rebuilt. An important aspect in this context is the appropriate ratio of medium to cell volume being so far not optimally reestablished in most of the currently available in vitro systems. To improve such culture conditions, we constructed a microstructure to culture hepatocytes and (without any addition of extracellular matrix material) characterized liver tissue in the form of evenly sized aggregates. The liver-specific differentiation status of such aggregates was monitored by their ability to perform CYP450 dependent xenobiotic metabolism along with the measurement of albumin secretion. Freshly isolated adult rat hepatocytes show an initial loss of total CYP450 content and of associated activities (mixed function oxidases). However, in the aggregate system, this level did not decrease further but remained stable or even increased throughout the culture period of 10-13 days. The CYP450 dependent metabolism of the hepatocytes is able to respond to classic inducing agents. The described culture efficiently supports liver-specific functions of adult rat hepatocytes and seems to be suited not only for use in an extracorporeal liver device but also for the formation of evenly sized small aggregates to be of use in transplantation of differentiated liver tissue. Moreover, after design variations, the microstructure can be applied for functional analysis of metabolically active hepatocytes as well as for toxicological and pharmacological validation.
Expression of HSP72 After ELF-EMF Exposure in Three Cell Lines
It has been reported that magnetic fields with flux densities ranging from microT to mT are able to induce heat shock factor, HSP72 mRNA or heat shock proteins in various cells. In this study we investigated changes in the HSP72 mRNA transcription level in three cell lines (HL-60, H9c2, and Girardi heart cells) and in the intracellular HSP72 protein content in two cell lines (HL-60 and Girardi heart cells) after treatment schemes using electromagnetic fields with a flux density of 2 microT to 4 mT, a frequency of 50 Hz and exposure times from 15 to 30 min. None of the treatments or modalities showed any significant effect on the HSP72 protein level, although HSP72 mRNA could be induced, at least to some extent, with one of the parameter combinations in all cell lines tested. Obviously, HSP72 mRNA transcription and translation are not necessarily coupled in certain cells. This leads to the conclusion that electromagnetic field effects on HSP72 mRNA levels are not indicative for downstream effects unless increased mRNA levels can be correlated with increased HSP72 protein levels as well.
A Chip-based Platform for the in Vitro Generation of Tissues in Three-dimensional Organization
We describe a multi-purpose platform for the three-dimensional cultivation of tissues. The device is composed of polymer chips featuring a microstructured area of 1-2 cm(2). The chip is constructed either as a grid of micro-containers measuring 120-300 x 300 x 300 microm (h x l x w), or as an array of round recesses (300 microm diameter, 300 microm deep). The micro-containers may be separately equipped with addressable 3D-micro-electrodes, which allow for electrical stimulation of excitable cells and on-site measurements of electrochemically accessible parameters. The system is applicable for the cultivation of high cell densities of up to 8 x 10(6) cells and, because of the rectangular grid layout, allows the automated microscopical analysis of cultivated cells. More than 1000 micro-containers enable the parallel analysis of different parameters under superfusion/perfusion conditions. Using different polymer chips in combination with various types of bioreactors we demonstrated the principal suitability of the chip-based bioreactor for tissue culture applications. Primary and established cell lines have been successfully cultivated and analysed for functional properties. When cells were cultured in non-perfused chips, over time a considerable degree of apoptosis could be observed indicating the need for an active perfusion. The system presented here has also been applied for the differentiation analysis of pluripotent embryonic stem cells and may be suitable for the analysis of the stem cell niche.
Tissue Reconstruction in 3D-spheroids from Rodent Retina in a Motion-free, Bioreactor-based Microstructure
While conventional rotation culture-based retinal spheroids are most useful to study basic processes of retinogenesis and tissue regeneration, they are less appropriate for an easy and inexpensive mass production of histotypic 3-dimensional tissue spheroids, which will be of utmost importance for future bioengineering, e.g. for replacement of animal experimentation. Here we compared conventionally reaggregated spheroids derived from dissociated retinal cells from neonatal gerbils (Meriones unguiculatus) with spheroids cultured on a novel microscaffold cell chip (called cf-chip) in a motion-free bioreactor. Reaggregation and developmental processes leading to tissue formation, e.g. proliferation, apoptosis and differentiation were observed during the first 10 days in vitro (div). Remarkably, in each cf-chip micro-chamber, only one spheroid developed. In both culture systems, sphere sizes and proliferation rates were almost identical. However, apoptosis was only comparably high up to 5 div, but then became negligible in the cf-chip, while it up-rose again in the conventional culture. In both systems, immunohistochemical characterisation revealed the presence of Müller glia cells, of ganglion, amacrine, bipolar and horizontal cells at a highly comparable arrangement. In both systems, photoreceptors were detected only in spheroids from P3 retinae. Benefits of the chip-based 3D cell culture were a reliable sphere production at enhanced viability, the feasibility of single sphere observation during cultivation time, a high reproducibility and easy control of culture conditions. Further development of this approach should allow high-throughput systems not only for retinal but also other types of histotypic spheroids, to become suitable for environmental monitoring and biomedical diagnostics.
