Joerg C. Gerlach

Adsorption of xenobiotics to plastic tubing incorporated into dynamic in vitro systems used in pharmacological research--limits and progress.

Commonly used materials incorporated into dynamic culture systems typically show the feature of adsorption of lipophilic xenobiotics. Yet, this phenomenon is strongly limiting the use of dynamic culture models and ex vivo organ perfusions in pharmacological and toxicological research. The aim of the study was to characterize different materials with respect to their capacity for drug adsorption and to find methods or materials to reduce the loss of substrate by adsorption in order to improve the use of dynamic in vitro systems. The adsorption of different xenobiotics (lidocaine, midazolam, lormetazepam, phenobarbital, testosterone, ethoxyresoroufine) to tubes used in dynamic in vitro systems (polyvinyl-chloride, silicone) were investigated and compared to a new material (silicone-caoutchouc-mixture). In addition, the role of protein deposition onto the tubing was studied and it was investigated whether it was possible to reach saturation of the inner tube surface by pre-loading it with the test compound. We found that silicone tubes provided the highest comfort with respect to handling and reusability, but they also demonstrated the highest capacity for substrate adsorption. Polyvinyl-chloride was the second best in handling but also demonstrated a high complexity in its adsorption behavior. The silicone-caoutchouc-mixture reached acceptable experimental results with respect to its handling and demonstrated a very low capacity for substrate adsorption.

Concept for modular extracorporeal liver support for the treatment of acute hepatic failure

Acute liver failure has a poor prognosis. The introduction of liver transplantation as a therapeutic option reduced mortality to 20–40%. With the growing disparity between the number of organ donations and the number of patients waiting for liver transplantation, efforts have been made to optimize the allocation of organs and to design extracorporeal methods to support the failing liver. The modular extracorporeal liver support is a concept for the treatment of hepatic failure. The CellModule is a multicompartment bioreactor for extracorporeal liver support therapy. The construction provides efficient integrated oxygenator functions and decentralized mass transfer is effected by a woven array of capillary systems. The bioreactor promotes primary human liver cells to spontaneous neo-formation of liver sinusoidal structures in vitro. Small capillary subunits, in which interwoven membrane links represent the liver lobuli, are simultaneously perfused. The used cell mass of 400–600 g enabled the clinical application of a liver lobe equivalent hybrid organ. The DetoxModule enables albumin-dialysis for removal of albumin-bound toxins; a DialysisModule for continuous veno-venous hemofiltration can be added to the system, in the case of hepato-renal failure.

Comparative analysis of metabolism of medium- and plasma perfused primary pig hepatocytes cultured around a 3-D membrane network.

Culture media are frequently used in the evaluation of metabolical functions of hepatocytes in hybrid liver support systems (hLSS). However, media compositions differ substantially from those of plasma. Therefore, our study was designed to investigate whether current in vitro studies with medium are suitable to assess the metabolical competence of hLSS-cultures during clinical application as well as to explore whether the cell nutrition with medium provides a suitable modus operandi for stand by cultivation. Paired bioreactor cultures were perfused with either Williams’ Medium E (MPB) or human plasma (PPB). About 6x108 primary pig hepatocytes (>97% viability) were cultured in three laboratory scale bioreactors designed according to Gerlachs bioreactor-concept. Different perfusion protocols were initiated after a standardised period allowing for cell attachment and reorganisation in aggregates. Whereas patterns of enzyme release were similar in both protocols the metabolical behaviour was different between MPB (anabolic state) and PPB (catabolic state). Furthermore, compared to MPB the lidocaine-MEGX-tests for PPB demonstrated lower MEGX-concentrations and a different reaction pattern. We conclude that the nutrition of hepatocytes with medium during the stand by period itself might influence the cell function and subsequently the efficacy of the hLSS-treatment during clinical application. (Int J Artif Organs 2000; 23: 104–10)

Network Analysis of the Kinetics of Amino Acid Metabolism in a Liver Cell Bioreactor

The correlation of the kinetics of 18 amino acids, ammonia and urea in 18 liver cell bioreactor runs was analyzed and described by network structures. Three kinds of networks were investigated: i) correlation networks, ii) Bayesian networks, and iii) dynamic networks that obtain their structure from systems of differential equations. Three groups of liver cell bioreactor runs with low, medium and high performance, respectively, were investigated. The aim of this study was to identify patterns and structures of the amino acid metabolism that can characterize different performance levels of the bioreactor.

Dynamic model of amino acid and carbohydrate metabolism in primary human liver cells

Human liver cell bioreactors are used in extracorporeal liver support therapy. To optimize bioreactor operation with respect to clinical application an understanding of the central metabolism is desired. A two-compartment model consisting of a system of 48 differential equations was fitted to time series data of the concentrations of 18 amino acids, ammonia, urea, glucose, galactose, sorbitol and lactate, measured in the medium outflow of seven liver cell bioreactor runs. Using the presented model, the authors predict an amino acid secretion from proteolytic activities during the first day after inoculation of the bioreactor with primary liver cells. Furthermore, gluconeogenetic activites from amino acids and/or protein were predicted.

Prediction of the performance of human liver cell bioreactors by donor organ data

Human liver cell bioreactors are used in extracorporeal liver support therapy. To optimize bioreactor operation with respect to clinical application an early prediction of the long-term bioreactor culture performance is of interest. Data from 70 liver cell bioreactor runs labeled by low (n=18), medium (n=34) and high (n=18) performance were analyzed by statistical and machine learning methods. 25 variables characterizing donor organ properties, organ preservation, cell isolation and cell inoculation prior to bioreactor operation were analyzed with respect to their importance to bioreactor performance prediction. Results obtained were compared and assessed with respect to their robustness. The inoculated volume of liver cells was found to be the most relevant variable allowing the prediction of low versus medium/high bioreactor performance with an accuracy of 84 %.

Hepatozyten-Tissue Engineering und Bioreaktoren für hybride Leberunterstützungssysteme

Ein auf Kapillarmembranen basierender Bioreaktor wurde entwickelt. Die verwendeten Kapillarsysteme bilden dreidimensionale Strukturen, an denen und zwischen denen sich die Zellen organisieren konnen. Wie im naturlichen Organ werden viele kleine identische Kapillareinheiten fur wenige Zellen geschaffen, die naherungsweise der physiologischen Einheit des Leberlobulus entsprechen. Zur Gewinnung von Hepatozyten in KoKultur mit primaren Endothelzellen wurde eine kombinierte Isolationsmethode fur beide Zelltypen entwickelt. Die Zellen (ca. 500 g pro Bioreaktor) formieren sich im Bioreaktor zu lebergewebsahnlichen Strukturen. Das System wurde fur klinische Studien entwickelt und an acht Patienten erfolgreich bei Patienten im Leberausfallkoma im Sinne des Bridging der Wartezeit zur Verfugbarkeit eines Transplantates bzw. zur Lebertransplantation eingesetzt.