10 research outputs found
Automated analysis of acetaminophen toxicity on 3D HepaRG cell culture in microbioreactor
Real-time monitoring of bioanalytes in organotypic cell cultivation devices is a major research challenge in establishing stand-alone diagnostic systems. Presently, no general technical facility is available that offers a plug-in system for bioanalytics in diversely available organotypic culture models. Therefore, each analytical device has to be tuned according to the microfluidic and interface environment of the 3D in vitro system. Herein, we report the design and function of a 3D automated culture and analysis device (3D-ACAD) which actively perfuses a custom-made 3D microbioreactor, samples the culture medium and simultaneously performs capillary-based flow ELISA. A microstructured MatriGrid ® has been explored as a 3D scaffold for culturing HepaRG cells, with albumin investigated as a bioanalytical marker using flow ELISA. We investigated the effect of acetaminophen (APAP) on the albumin secretion of HepaRG cells over 96 h and compared this with the albumin secretion of 2D monolayer HepaRG cultures. Automated on-line monitoring of albumin secretion in the 3D in vitro mode revealed that the application of hepatotoxic drug-like APAP results in decreased albumin secretion. Furthermore, a higher sensitivity of the HepaRG cell culture in the automated 3D-ACAD system to APAP was observed compared to HepaRG cells cultivated as a monolayer. The results support the use of the 3D-ACAD model as a stand-alone device, working in real time and capable of analyzing the condition of the cell culture by measuring a functional analyte. Information obtained from our system is compared with conventional cell culture and plate ELISA, the results of which are presented herein
MatriGrid® based biological morphologies: tools for 3D cell culturing
Recent trends in 3D cell culturing has placed organotypic tissue models at another level. Now, not only is the microenvironment at the cynosure of this research, but rather, microscopic geometrical parameters are also decisive for mimicking a tissue model. Over the years, technologies such as micromachining, 3D printing, and hydrogels are making the foundation of this field. However, mimicking the topography of a particular tissue-relevant substrate can be achieved relatively simply with so-called template or morphology transfer techniques. Over the last 15 years, in one such research venture, we have been investigating a micro thermoforming technique as a facile tool for generating bioinspired topographies. We call them MatriGrid ® s. In this research account, we summarize our learning outcome from this technique in terms of the influence of 3D micro morphologies on different cell cultures that we have tested in our laboratory. An integral part of this research is the evolution of unavoidable aspects such as possible label-free sensing and fluidic automatization. The development in the research field is also documented in this account
Measles Virus Interacts with and Alters Signal Transduction in T-Cell Lipid Rafts
By a contact-dependent surface interaction, the measles virus (MV) glycoprotein complex induces a pronounced inhibition of T-cell proliferation. We now show that MV directly interacts with glycosphingolipid-enriched membrane microdomains on human primary T cells and alters recruitment and segregation of membrane proximal signaling components. Contact-dependent interference with T-cell receptor-stimulated tyrosine phosphorylation and Ca mobilization is a late event seen 24 h after MV treatment. In contrast, stimulated recruitment of pleckstrin homology domain-containing proteins such as Akt and Vav is inhibited early after MV contact, as is segregation of the activated Akt kinase from rafts. Tyrosine phosphorylation of the regulatory subunit of the phosphatidylinositol 3-kinase (PI3K), p85, is apparently normal then, yet this protein fails to partition to the lipid raft fraction, and this is associated with stable expression of its negative regulator Cbl-b. Thus, by interaction with lipid rafts, MV contact initially targets recruitment of PI3K by preventing stimulated Cbl-b degradation and activation of PI3K-dependent signaling components
PEDOT Coated Thick Film Electrodes for In Situ Detection of Cell Adhesion in Cell Cultures
Low temperature cofired ceramics (LTCC) provide a technology for the 3-dimensional integration of sensor arrays into bioreactors covering dimensions of several hundred micrometers. Since optical control in such assemblies is not possible, the in situ detection of cell adhesion on impedance electrodes with high spatial resolution would deliver crucial information. A current limitation is the increasing impedance of microelectrodes with decreasing diameter. This study evaluates the suitability of thick film gold electrodes, pristine and coated with electropolymerized poly(3,4-ethylenedioxythiophene) (PEDOT), for the detection of cell adhesion on the electrode surface. The impedance as criterion for cell attachment is measured with a recording system for electroactive cells with the aim of improving usability. Two cell cultures with different adhesion characteristic are used for adhesion assessment on planar test chips. The impedance increase measured on individual PEDOT coated electrodes due to tight contact of cells reaches a factor of 6.8 in cultures of well-adherent HepG2 cells. Less adhered NG108-15 cells produce a maximum impedance increase by a factor of 2.6. Since the electrode impedance is significantly reduced by PEDOT coating, a reduction of the electrode diameter to values below 100 µm and spatially resolved detection is possible. The results encourage further studies using PEDOT coated thick film electrodes as bio-electronic-interfaces. We presume that such miniaturized electrodes are suitable for 3-dimensional recordings in electroactive cell cultures, providing information of local cell adhesion at the same time
ZwiPhos - Entwicklung eines Lagerungskonzeptes für Klärschlammmonoverbrennungsaschen für Deutschland mit dem Ziel einer späteren Phosphorrückgewinnung
Author Correction: Consistent success in life-supporting porcine cardiac xenotransplantation
Consistent success in life-supporting porcine cardiac xenotransplantation
Heart transplantation is the only cure for patients with terminal cardiac failure, but the supply of allogeneic donor organs falls far short of the clinical need1–3. Xenotransplantation of genetically modified pig hearts has been discussed as a potential alternative4. Genetically multi-modified pig hearts that lack galactose-α1,3-galactose epitopes (α1,3-galactosyltransferase knockout) and express a human membrane cofactor protein (CD46) and human thrombomodulin have survived for up to 945 days after heterotopic abdominal transplantation in baboons5. This model demonstrated long-term acceptance of discordant xenografts with safe immunosuppression but did not predict their life-supporting function. Despite 25 years of extensive research, the maximum survival of a baboon after heart replacement with a porcine xenograft was only 57 days and this was achieved, to our knowledge, only once6. Here we show that α1,3-galactosyltransferase-knockout pig hearts that express human CD46 and thrombomodulin require non-ischaemic preservation with continuous perfusion and control of post-transplantation growth to ensure long-term orthotopic function of the xenograft in baboons, the most stringent preclinical xenotransplantation model. Consistent life-supporting function of xenografted hearts for up to 195 days is a milestone on the way to clinical cardiac xenotransplantation7