We investigated cell death during glucose deprivation in rat cardiomyocyte-derived H9c2 cells. Electron microscopic analysis revealed accumulation of autophagic vacuoles during glucose deprivation. The addition of 3-methyladenine or LY294002, which are known to inhibit autophagosome formation, reduced cell death while Z-VAD-FMK, a caspase inhibitor, slightly affected cell death. Thus, cell death during glucose deprivation is not type I programmed cell death (apoptotic cell death) but type II programmed cell death (autophagic cell death). Moreover, we found that both insulin-like growth factor-I and the adenovirus-mediated overexpression of wild-type class I PI 3-kinase accelerated cell death as well as accumulation of autophagic vacuoles during glucose deprivation while dominant-negative PI 3-kinase reduced these phenomena. The results indicate that IGF-I/PI 3-kinase accelerates the accumulation of autophagic vacuoles and subsequent autophagic cell death during glucose deprivation, revealing the opposing role of IGF-I/ PI 3-kinase in two distinct types of programmed cell death (apoptotic and autophagic cell death).

It is postulated that the hepatotoxicity of valproic acid (VPA) results from the mitochondrial beta-oxidation of its cytochrome P450 metabolite, 2-propyl-4-pentenoic acid (4-ene VPA), to 2-propyl-(E)-2,4-pentadienoic acid ((E)-2,4-diene VPA) which, in the CoA thioester form, either depletes GSH or produces a putative inhibitor of beta-oxidation enzymes. In order to test this hypothesis, 2-fluoro-2-propyl-4-pentenoic acid (alpha-fluoro-4-ene VPA) which was expected to be inert to beta-oxidative metabolism was synthesized and its effect on rat liver studied in comparison with that of 4-ene VPA. Similarly, the known hepatotoxicant 4-pentenoic acid (4-PA) and 2,2-difluoro-4-pentenoic acid (F2-4-PA) were compared. Male Sprague-Dawley rats (150-180 g, 4 rats per group) were dosed ip with 4-ene VPA (0.7 mmol/kg per day), 4-PA (1.0 mmol/kg per day), or equivalent amounts of their alpha-fluorinated analogues for 5 days. Both 4-ene VPA and 4-PA induced severe hepatic microvesicular steatosis ( > 85% affected hepatocytes), and 4-ene VPA produced mitochondrial alterations. By contrast, alpha-fluoro-4-ene VPA and F2-4-PA were not observed to cause morphological changes in the liver. The major metabolite of 4-ene VPA in the rat urine and serum was the beta-oxidation product (E)-2,4-diene VPA. The N-acetylcysteine (NAC) conjugate of (E)-2,4-diene VPA was also found in the urine. Neither (E)-2,4-diene VPA nor the NAC conjugate could be detected in the rats administered alpha-fluoro-4-ene VPA. In a second set of rats (3 rats per group), total liver GSH levels were determined to be depleted to 56% and 72% of control following doses of 4-ene VPA (1.4 mmol/kg) and equivalent alpha-fluoro-4-ene VPA, respectively. Mitochondrial GSH remained unchanged in the alpha-fluoro-4-ene VPA treated group but was reduced to 68% of control in the rats administered 4-ene VPA. These results strongly support the theory that hepatotoxicity of 4-ene VPA, and possibly VPA itself, is mediated largely through beta-oxidation of 4-ene VPA to reactive intermediates that are capable of depleting mitochondrial GSH.

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Ethanol metabolism-induced lipogenesis may trigger the SNS-activated HSCs feedback loop, and then induct the activated HSCs and the activated HSCs-derived TNF-α, the mediator of lipogenesis, overproduction. Carvedilol may block this feedback loop via antisympathetic activity and demonstrate its preventive role on the development of hepatosteatosis in rat with AFLD.

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