sis or apoptosis. Necrosis is characterized by cell swelling Toxic bile salts cause hepatocyte necrosis at high conand autolysis with breakdown of the plasma membrane percentrations and apoptosis at lower concentrations. Almeability barrier. 4 In contrast, apoptosis is characterized by though fructose prevents bile salt-induced necrosis, the cell shrinkage, chromatin condensation, nuclear and DNA effect of fructose on bile salt-induced apoptosis is unfragmentation, and ultimately fragmentation of the cell into clear. Our aim was to determine if fructose also protects membrane-defined cell bodies referred to as apoptotic bodies. 5 against bile salt-induced apoptosis. Fructose inhibited Recently, we have investigated the cellular mechanisms of glycochenodeoxycholate (GCDC)-induced apoptosis in a hepatocellular injury by toxic bile salts. 6-8 We have demonconcentration-dependent manner with a maximum inhistrated directly that toxic bile salts at high concentrations bition of 72% { 10% at 10 mmol/L. First, we determined (Β’250 mmol/L) cause cell death by necrosis whereas lower if fructose inhibited apoptosis by decreasing adenosine concentrations of toxic bile salts (Β°100 mmol/L) cause cell triphosphate (ATP) and intracellular pH (pH i ). Although death by apoptosis. 6,7 However, information regarding the fructose decreased ATP to Γ΅25% of basal values, oligosimilar and dissimilar mechanisms responsible for bile saltmycin (an ATP synthase inhibitor) did not inhibit induced necrosis vs. apoptosis is lacking. Identification of apoptosis despite decreasing ATP to similar values. potentially common mechanisms responsible for both bile Fructose (10 mmol/L) decreased intracellular pH (pH i ) salt-induced necrosis and apoptosis would be of scientific by 0.2 U. However, extracellular acidification (pH 6.8), and clinical importance. Not only would an understanding of which decreased hepatocyte pH i 0.35 U and is known to the common mechanisms responsible for bile salt-induced inhibit necrosis, actually potentiated apoptosis 1.6-fold.
necrosis and apoptosis provide basic information on the Fructose cytoprotection also could not be explained by mechanisms of cell death, but the information may also geninduction of bcl-2 transcription or metal chelation. Beerate strategies for the treatment of cholestatic liver diseases.
cause we could not attribute fructose cytoprotection to
Cytoprotective agents provide a unique paradigm with metabolic effects, alterations in the expression of bclwhich to help identify the common mechanisms responsible 2, or metal chelation, we next determined if the poorly for cell death. Cytoprotection by an agent against multiple metabolized ketohexoses, tagatose and sorbose, also inmodels or modes of cell death suggests common mechanisms hibited apoptosis; unexpectedly, both ketohexoses inare involved in the death process. Conversely, cytoprotection hibited apoptosis. Because bile salt-induced apoptosis in one model and a lack of protection in another model would and necrosis are inhibited by fructose, these data sugsuggest dissimilar mechanisms of cell death. We have demongest that similar processes initiate bile salt-induced hestrated that fructose, a cytoprotective agent in many models patocyte necrosis and apoptosis. In contrast, acidosis, of cell necrosis, prevents bile salt-induced necrosis of hepatowhich inhibits necrosis, potentiates apoptosis. Thus, ketohexose-sensitive pathways appear to initiate both bile cytes. 6 Fructose cytoprotection is thought to be mediated by salt-induced cell apoptosis and necrosis, whereas disglycolytic generation of adenosine triphosphate (ATP) or insimilar, pH-sensitive, effector mechanisms execute these tracellular acidification. 9,10 However, the effect of fructose two different cell death processes. (HEPATOLOGY and its metabolic effects on cellular ATP and pH during bile 1997;25:81-86.)
salt-induced apoptosis is unknown. Thus, the overall objective of this study was to determine if fructose also protects against bile salt-induced apoptosis. Our specific aims were The impairment of bile flow in chronic cholestatic liver to answer the following questions during apoptosis of culdiseases results in the retention and accumulation of toxic tured rat hepatocytes by 100 mmol/L glycochenodeoxycholate bile salts within the hepatocyte. 1,2 The increase in the (GCDC): Is fructose cytoprotective? Is fructose cytoprotection intracellular accumulation of toxic bile salts contributes to mediated by changes in cellular ATP, changes in intracellular hepatocellular injury in these liver diseases. 3 Although it is pH (pH i ), alterations of bcl-2 expression, or metal chelation? clinically important, the mechanisms of bile salt-induced-Are other ketohexoses cytoprotective? We chose GCDC as the hepatocellular injury remain incompletely understood. Cell toxic bile salt for these studies because it is a primary bile death may occur by one of two fundamental processes: necrosalt, the concentrations of which increase ΓΊ20-fold during cholestasis. 11 In addition, we have demonstrated previously that GCDC causes hepatocyte apoptosis. 12 Abbreviations: ATP, adenosine triphosphate; BCECF, 2-7-bis(carboxy-ethyl)-5,6-carboxyfluorescein; DAPI, 4-6-diamidino-2-phenylindole dihydrochloride; GCDC, glycocheno-
Methods
deoxycholate; pHi, intracellular pH.