WHAT IS THE BIOARTIFICIAL LIVER?
The bioartificial liver (AMC-BAL), created thanks to the collaboration with the Academisch Medisch Centrum of Amsterdam, is a hepatic hybrid supporting system composed of polyester fibers. This system supports the cellsimplantation in the form of little aggregates and capillaries permeable to the gases, as an oxygenation system able to furnish to the hepatocytes the adequate contribution of oxygen. The role of the hepatic supporting systems is to substitute the catabolic and synthetic functions peculiar to the hepatic cell, which are compromised in some pathological conditions, such as the acute liver failure.
Before human tests, functional capability of AMC-BAL has been evaluated in several animal models of hepatic failure like:
- Total hepatic ischemia, in the rat;
- Hepatic devascularization, in the pig;
- Total hepatectomy, in the pig.
There have been significant results, from a statistic point of view, in the animals exposed to the BAL treatment. For example:
- Increase of survival;
- Reduction of plasmatic levels of ammonium and bilirubin;
- An effective and safe device.
From 2000 to 2003 the Centre of Biotechnology has coordinated a Multicentric Trial joined by 14 patients affected with fulminant hepatitis of different aetiology, that have been supported with a BAL loaded with about 10 billion pig hepatocytes “Specific Pathogen Free” (SPF), as a bridging therapy before the orthotopic liver transplantation. The patient waiting for the organ was connected to the BAL with a plasmapheresis system. Thus, the BAL was able to support the vital functions for 24 hours and more, with a survival percentage of the patients under treatment that goes from 50% to 72%.
Which kind of cells have to be used in the BAL?
In 2004, due to a Moratory emanated by the European Community about the clinical applications of xenotransplantations, the clinical experimentation on human with bioartificial liver was interrupted, so we started using the bioreactor looking for alternative cells and innovative applications. For this type of experimentation, the bioreactor was loaded with human, fetal and adult immortalized hepatocytes, the latter taken from tumor resections or discarded livers because not suitable for transplantation. Results of a comparative study between the use of fetal hapatocytes and the use of adult hepatocytes demonstrated that the adult ones are, at the moment, the first choice in the sector.
The BAL as cellular biosensor
The development of an effective supporting system for the three-dimensional cultivation of human hepatocytes, induce us to explore innovative applications of the bioreactor. The hepatotoxicity evaluation in pharmaceutical, cosmetic and food products, is mandatory for the trade of new products. This is particularly restricted in the pharmaceutical sector, in which the development of new drugs costs about ten million euros. In most cases these costs ought to be multiplied for hundreds molecules that are at the end pulled back for pharmacokinetic and/or pharmacodynamic profile lacking. The failure of the new molecule can depend to the different types of pharmaceutical interactions or to the undesirable toxic effects appearing during post-marketing monitoring.
The need to solve the problem of the reduced hepatocytes availability in experimentation convinced us to plan and realize, in collaboration with RanD ( a company specialized in realizing medical devices), BAL miniaturization. So, a “mini BAL” was built and it is capable of containing up to 300 million human cells. MiniBAL is a system for in vitro analysis of new molecules hepatotoxicity, that have direct effect to synthetic and/or metabolic capability of the cell, produced by the pharmaceutical, cosmetic and food industry, in a premature phase of the developing process. Basically, this device can be considered as a cellular biosensor that is capable of verifying the hepatotoxicity of pharmaceutical, cosmetic and food products using cellular systems that not only are related to the organ functions, but also to the peculiarity of species. The reduction of the number of hepatocytes to use would permit us to realize multiple toxicological profiles, using a single hepatocytic sample and to get important information for toxicological and pharmacologic screening of a big number of molecules in the pre-clinic development phase. This would allow to one of miniBAL’s application to be an intermediate between in vitro and clinic experimentation. At the same time this miniBAL can be used as an alternative method to animal experimentation, satisfying the common demand of validating systems that would lead to a reduced usage of animals. In our Centre we studied the effects of amphetamine’s administration, a substance with strong hepatocytic toxicity, on functionality and on hepatic vitality indexes during 7 days using three different doses. The preliminary results showed a reduced capability for human hepatocytes to convert ammonium in urea, that is directly related to the dose administrated and reversible with the treatment suspension . Currently, an international leader company in the field of immunoderivativesproducts commissioned the analysis, with the support of miniBAL system, of a molecule’s hepatotoxicity that will be an innovative and promising therapy in some types of tumor. Preliminary results showed a reduction of hepatic function parameters related to the concentrations that were used.
The Centre of Biotechnology is planning to realize, in collaboration with Bio TekNet consortium company, a bioartificial micro liver, with a reduced tridimensional structure. A further miniaturization will permit to realize a system in which several bioartificial liver, arranged in parallel, will examine the hepatotoxicity in more compounds or in several concentrations of the same compound. It will decrease all the issues connected to biological variability and contributing in reducing the costs and the time spent in clinical experimentations. Moreover, in the Centre will be realized a bank for cryopreserved human hepatocytes, where functional and vital hepatic cells will be available, without waiting for the episodic resection, making the whole process more effective.