A Microscale In Vitro Physiological Model of the Liver: Predictive Screens for Drug Metabolism and Enzyme Induction

Abstract

In vitro models of the liver using isolated primary hepatocytes have been used as screens for measuring the metabolism, toxicity and efficacy of xenobiotics, for studying hepatocyte proliferation, and as bioartificial liver support systems. Yet, primary isolated hepatocytes rapidly lose liver specific functions when maintained under standard in vitro cell culture conditions. Many modifications to conventional culture methods have been developed to foster retention of hepatocyte function. Still, not all of the important functions -- especially the biotransformation functions of the liver -- can as yet be replicated at desired levels, prompting continued development of new culture systems. In the first part of this article, we review primary hepatocyte in vitro systems used in metabolism and enzyme induction studies. We then describe a scalable microreactor system that fosters development of 3D-perfused micro-tissue units and show that primary rat cells cultured in this system are substantially closer to native liver compared to cells cultured by other in vitro methods, as assessed by a broad spectrum of gene expression, protein expression and biochemical activity metrics. These results provide a foundation for extension of this culture model to other applications in drug discovery -- as a model to study drug-drug interactions, as a model for the assessment of acute and chronic liver toxicity arising from exposure to drugs or environmental agents; and as a disease model for the study of viral hepatitis infection and cancer metastasis.

Chromosome conformation and gene expression patterns differ profoundly in human fibroblasts grown in spheroids versus monolayers

“…3D human-cell models may also provide more effective and economical screening of new drugs than the use of animal models. 8,9 Furthermore, in vitro 3D modeling of native tissue provides tools for regenerative medicine. However, understanding the fundamental cell biology is critical in translating in vitro discoveries into clinical applications, e.g., functional replacement of damaged tissue.…”

Predictive toxicology using systemic biology and liver microfluidic “on chip” approaches: Application to acetaminophen injury

“…Bioartificial organs can now take advantage of recent developments in microtechnology to produce systems on a very small scale (Griffith and Naughton 2002;Powers et al, 2002;Sivaraman et al, 2005;Chao et al, 2010;Prot et al, 2011a;Baudoin et al, 2007;Novik et al, 2010;Baudoin et al, 2011). The cellular organization brought about by the micro-topography of these systems and their dynamic microfluidic culture conditionsappear to be key features for reproducing in vivo environments.…”

Exploitation of physical and chemical constraints for three-dimensional microtissue construction in microfluidics

“…Microfluidic platforms for 3D cell culture are rapidly gaining importance in biomedical applications ranging from tissue engineering to drug toxicity or metabolism studies 170,171 and other aspects of the drug discovery process. 155 Cell immobilization in a microfluidic system is becoming increasingly important as a way of creating artificial tissues.…”

“…6,172 In vitro culture of liver cells ͑hepatocytes͒ is important because many drug candidates fail in clinical studies; increasingly due to toxicity issues. 171,172 Many studies focus on trapping and culturing hepatocytes in microfluidic platforms. Powers et al 173,174 described a 3D microarray bioreactor where they cultured primary hepatocytes for two weeks, forming viable tissue-like structures showing constant albumin secretion and urea genesis; and ultrastructure analysis of hepatocytes revealed bile canaliculi, numerous tight junctions, and glycogen storage.…”

“…Prokop et al 179 presented a NanoLiterBioReactor in which hepatocytes were trapped by 3 m sievelike features. Sivaraman et al 171 used a 3D microfabricated bioreactor system to make microtissue units of dimensions ϳ2 ϫ 10 −5 cm 3 , which showed better gene expression, protein expression, and other biochemical activities as compared to 2D cultures. Co-cultures of hepatocytes with 3T3-J2 fibroblasts in a 64 ͑8 ϫ 8͒ element array 170 of microfluidic wells or on a microgrooved glass substrate 180 exhibited steady levels of albumin and urea production.…”