Development and characterization of a small-scale bioreactor based on a bioartificial hepatic culture model for predictive pharmacological in vitro screenings

Abstract

A vast majority of pharmacons are beset by possible interactions and side effects which have usually been tested in laboratory animals. However, better methods are needed to reduce the number of animal experiments and interspecies differences with respect to drug metabolism, as well as to provide a faster and more cost-effective way of analysis. These facts have led to the development of in vitro models based on isolated primary hepatocytes to better assess drug metabolism, interactions, and toxicity. A new small-scale bioreactor with the hepatic sandwich model and a gas-permeable membrane at the bottom allowing a definable oxygen exchange, has been constructed and compared with the conventional well plates. Compared to hepatocytes cultured in conventional systems, the cells exhibited a stronger liver-specific capacity and remained in a differentiated state in the small-scale bioreactor over a cultivation period of 17 days. This in vitro model could serve as a tool to predict the liver response to newly developed drugs.

Nanostructured self-assembling peptides as a defined extracellular matrix for long-term functional maintenance of primary hepatocytes in a bioartificial liver modular device

“…The cells were seeded at a density of 2.5 × 10 5 cells/cm 2 in a RAD peptides coated bioreactor under a defined combination of growth factors and cytokines (Williams' E medium + Activin A + WNT 3a + HGF + FGF-4 + EGF + Oncostatin M + FGF-10 + BMP-4 + Dex + RA all from Sigma-Aldrich Chemie Gmbh [Munich, Germany] to mimic the in vivo hepatic microenvironment) 28 and were incubated at 37°C in a humidified atmosphere containing 5% CO 2 and 20% O 2 (v/v). The fundamental design of our flat membrane bioreactor 11 and the minibioreactor 13 is similar. Funtional maintenance of primary hepatocytes was conducted three times.…”

“…In our construction, our bioreactor offers a maximal oxygen supply of 90 µmol per 1.77 cm 2 via direct delivery of oxygen to the cells from the bottom of the device. 13 Our bioreactor also allows direct contact of every individual liver cell with the oxygen supply, for enhanced oxygenation that is closer to the in vivo liver. 5,11,12 While the available image data was obtained from CLSM, basically providing the image series for our image analysis, we have restricted ourselves to 2D image-based assessments on single images taken centrally from the stacks.…”

“…However, most experimental approaches aimed at the long-term functional maintenance of primary hepatocytes in a culture from 1 week to several weeks (42 days) have been reported by various investigators, [2][3][4] including us. [11][12][13] However, to the best of our knowledge, there is no report for the long-term functional maintenance of primary hepatocytes up to 90 days within a defined hepatic microenvironment to provide a better strategy as an alternative to in vivo animal experimentation. US Food and Drug Administration (FDA) rules require the survival potential of primary hepatocytes of a minimum of 14 days and a maximum of 90 days for in vivo toxicological experiments.…”

Real-time in situ viability assessment in a 3D bioreactor with liver cells using resazurin assay

“…Alternative cell culture systems are based on 3D cell culture techniques mimicking the microenvironment of tissues or organs and allow long term maintenance of the cells. These cultivation systems include gel-based systems imitating extracellular matrix (Sodunke et al 2007), spheroids (AbuAbsi et al 2002;Mueller et al 2011a), micro scaffolds (Bokhari et al 2007) or various types of hollow fiber bioreactors (Schmitmeier et al 2006;Schmelzer et al 2010). Hollow fiber 3D bioreactors were applied as bioartificial liver for clinical applications (Gerlach et al 2003) and were down-scaled for analytical purposes.…”

Orientation and capturing of antibody affinity ligands: Applications to surface plasmon resonance biochips

“…Human serum albumin is a common model protein and also a critical biomarker for hepatocyte differentiation and function often used in vitro [23,24]. Troponin T is an important clinical biomarker for myocardial states, and is one of the endpoints for in vitro cardiotoxicity testing [25,26].…”