Cytotoxicity of Bile in Human Hep G2 Cells and in Primary Cultures of Rat Hepatocytes

Abstract

There has been increasing interest in the development of a hepatocyte bioreactor for the treatment of acute hepatic failure; however, little is known about the effect of hepatocyte byproducts on the viability of the cells in the bioreactor environment. We investigated the effects of increasing concentrations of bile on the growth and viability of the human hepatoma cell line Hep G2 and on the cytochrome P-450 content and dependent mixed function oxidase (MFO) activities, reduced glutathione (GSH) content, and glutathione S-transferase (GST) activity of primary cultures of rat hepatocytes. Our purpose was to determine whether or not it would be necessary to pretreat the plasma from patients with acute liver failure to remove elevated bile concentrations which might be toxic to the hepatocytes in an artificial liver device. Bile was found to inhibit Hep G2 cell growth at concentrations as low as 0.1% and to decrease viability at concentrations above 0.5%. The cytochrome P-450 and GSH contents and the activities of the MFO system and of GST were decreased in the primary cultures of hepatocytes following 24 h treatment with concentrations of bile at and above 0.5%. The MFO activities associated with different cytochrome P-450 isoenzymes decreased to different extents in the presence of bile with the O-dealkylation of pentoxyresorufin being more labile than that of ethoxyresorufin. Our data indicate that elevated bile concentrations are cytotoxic to liver cells, and it may be necessary to pretreat patient plasma to decrease its bile content to protect the cells during the clinical operation of a hepatocyte bioreactor device.

Alleviating liver failure conditions using an integrated hybrid cryogel based cellular bioreactor as a bioartificial liver support

“…All these toxins could exert a detrimental effect on the cells used within a BAL device, such as loss of cell-cell contact, cell death due to apoptosis and necrosis, and loss in functionality282930.…”

Modulation of Hepatocyte Function in an Immortalized Human Hepatocyte Cell Line Following Exposure to Liver‐Failure Plasma

“…The decrease may be a result of the conjugation of reactive intermediates in the plasma samples with GSH, thus leading to a depletion of cellular GSH stores. GSH can also react directly with reactive oxygen free‐radicals (8), and this can also contribute to GSH depletion. A consequence of reduced GSH levels could also be a reduction of cellular GSSG levels.…”

“…Hollow fiber cartridges with specific molecular weight cut‐off points can remove unwanted molecules such as complement (24), endotoxins, and proinflammatory cytokines (21). Hepatocyte functions that provide protection against toxic effects could be upregulated by culturing hepatocytes in three‐dimensional configurations, by coculture, or by the addition of matrix (31,32) while maintenance of critical hepatocyte metabolites such as GSH levels could also be controlled by supplementation with precursor amino acids (8) or specific inducers (10). If functions were absent but were deemed of critical importance, then cultures could be transfected with the genes of interest.…”

“…Only limited studies have investigated the effect of normal plasma on liver cells (2,4), and there is a paucity of data examining the effect of liver‐failure plasma (LFP) (5,6). These studies have identified two major challenges: first, the selection of an appropriate cell that maintains the critical functions of hepatocytes in a BAL; and second, because LFP contains many factors such as cytokines that inhibit hepatocyte function (7) and may cause hepatocyte necrosis (8), the performance of cells in LFP (2) needs to be elucidated.…”

Effects of acute‐liver‐failure‐plasma exposure on hepatic functionality of HepaRG‐AMC‐Bioartificial Liver

“…To achieve optimal performance of the HepaRG‐AMC‐BAL in a clinical setting, it is essential that its functionality is maintained during exposure to the toxic plasma of patients suffering from ALF or acute‐on‐chronic liver failure. Particularly ALF plasma contains high levels of hepatotoxic compounds that might affect BAL functionality , including constituents of necrotic cells, cytokines produced by a deranged immune response, lipopolysaccharides and compounds that are normally detoxified by the liver, such as ammonia, lactate and bile acids . All these toxins can exert detrimental effects to the cells applied in the BAL, including loss of cell‐cell contacts, a decrease in DNA and protein synthesis, a decrease in glutathione production, apoptosis and necrosis, and, most importantly, a decrease in hepatic functionality .…”