Development of a collagen gel sandwich hepatocyte bioreactor for detecting hepatotoxicity of drugs and chemicals

Understanding the hepatotoxicity of drugs and chemicals is essential for progress in academic research, medical science and in the development of new pharmaceuticals. Studying hepatotoxicity in vitro is a challenging task because hepatocytes, the metabolically active cells of the liver, are very difficult to maintain in culture. After just 24 hours, the cells detach from the plate and die, and even if they survive they usually do not express the metabolic functions which they have in vivo. It has been observed by others that culturing hepatocytes between two layers of collagen type I maintains in vivo-like morphology and also many drug metabolizing enzymes for weeks. In spite of the research examining drug metabolism in collagen sandwiches, there are very few studies evaluating this system for investigating hepatotoxicity. We cultured primary rat hepatocytes in the collagen sandwich configuration and our goal was to optimize this system for long-term studies and to examine toxicity of a variety of hepatotoxins. By measuring secretions of urea and albumin, and P4501A activity, we determined the optimal cell density to be 50,000 cells/cm2. We also evaluated the need for epidermal growth factor (EGF) in our cultures, by comparing urea and albumin secretions in cultures grown with and without EGF. The cultures without EGF had significantly less secretion of both urea and albumin just two days after plating. Therefore, we decided to include EGF in the medium. The toxins we examined were aflatoxin B1, acetaminophen, carbon tetrachloride, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), methyl methane sulfonate (MMS), cadmium, vinyl acetate and dimethylformamide (DMF). The cells were sensitive to aflatoxin B1, MMS, MNNG and cadmium. However, they were

Department dc:contributor.department

Massachusetts Institute of Technology. Biological Engineering Division.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2004

Author dc:creator

• Farkas, Dóra, 1976-

Advisor dc:contributor.advisor

• Steven R. Tannenbaum.

• Biological Engineering Division.

Statement dc:rights

• MIT theses are protected by copyright. They may be viewed, downloaded, or printed from this source but further reproduction or distribution in any format is prohibited without written permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/18040

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/18040