PGE1 abolishes the mitochondrial-independent cell death pathway induced by D-galactosamine in primary culture of rat hepatocytes
Corresponding Author
EMILIO SIENDONES
Clinical Unit of Digestive Apparatus, University Hospital Reina Sofía and
Emilio S Castillo, Unidad de Investigación, Unidad Clínica Aparato Digestivo, Hospital Universitario Reina Sofía, Av. Menéndez Pidal s/n, E-14004 Córdoba, Spain. Email: [email protected]Search for more papers by this authorYOLANDA JIMÉNEZ-GÓMEZ
Clinical Unit of Digestive Apparatus, University Hospital Reina Sofía and
Search for more papers by this authorJOSÉ LUÍS MONTERO
Clinical Unit of Digestive Apparatus, University Hospital Reina Sofía and
Search for more papers by this authorCONSUELO GÓMEZ-DÍAZ
Department of Cellular Biology, Physiology and Immunology, Faculty of Sciences, University of Cordoba, Cordoba, Spain
Search for more papers by this authorJOSÉ MANUEL VILLALBA
Department of Cellular Biology, Physiology and Immunology, Faculty of Sciences, University of Cordoba, Cordoba, Spain
Search for more papers by this authorJORDI MUNTANÉ
Clinical Unit of Digestive Apparatus, University Hospital Reina Sofía and
Search for more papers by this authorCorresponding Author
EMILIO SIENDONES
Clinical Unit of Digestive Apparatus, University Hospital Reina Sofía and
Emilio S Castillo, Unidad de Investigación, Unidad Clínica Aparato Digestivo, Hospital Universitario Reina Sofía, Av. Menéndez Pidal s/n, E-14004 Córdoba, Spain. Email: [email protected]Search for more papers by this authorYOLANDA JIMÉNEZ-GÓMEZ
Clinical Unit of Digestive Apparatus, University Hospital Reina Sofía and
Search for more papers by this authorJOSÉ LUÍS MONTERO
Clinical Unit of Digestive Apparatus, University Hospital Reina Sofía and
Search for more papers by this authorCONSUELO GÓMEZ-DÍAZ
Department of Cellular Biology, Physiology and Immunology, Faculty of Sciences, University of Cordoba, Cordoba, Spain
Search for more papers by this authorJOSÉ MANUEL VILLALBA
Department of Cellular Biology, Physiology and Immunology, Faculty of Sciences, University of Cordoba, Cordoba, Spain
Search for more papers by this authorJORDI MUNTANÉ
Clinical Unit of Digestive Apparatus, University Hospital Reina Sofía and
Search for more papers by this authorAbstract
Background and Aim: PGE1 reduces in vivo and in vitro D-galactosamine (D-GalN)-induced cell death in hepatocytes. The present study was undertaken to elucidate the intracellular pathway by which D-GalN induces cell death in cultured hepatocytes. In addition, we evaluated if PGE1 was able to modulate different parameters related to D-GalN-induced apoptosis in cultured rat hepatocytes.
Methods: Hepatocytes were isolated from male Wistar rats (225–275 g) by the classical collagenase procedure. PGE1 (1 µM) was administered 2 h before D-GalN (5 mM) in primary culture of rat hepatocytes. Apoptosis was determined by DNA fragmentation and caspase-3, -6, -8 and -9 activation in hepatocytes. Caspase activation was evaluated by the detection of the related cleaved product and its associated activity. Cell necrosis was determined by the measurement of lactate dehydrogenase (LDH) activity in culture medium. To elucidate the role of mitochondria, we measured neutral (nSMase) and acid (aSMase) sphingomyelinase, as well as the expression of cytochrome c in mitochondria and cytoplasm fractions from D-GalN treated hepatocytes.
Results: D-GalN induced caspase-3 activation and DNA fragmentation in hepatocytes. This apoptotic response was not associated with the activation of caspase-6, -8 or -9. The use of specific inhibitors confirmed that only caspase-3 was involved in D-GalN-induced apoptosis. D-GalN did not modify nSMase and aSMase activities, nor mitochondrial cytochrome c release in hepatocytes.
Conclusions: D-GalN induced apoptosis through caspase-3 activation but without modification of the activity of caspase-6, -8, -9, SMases or cytochrome c release. PGE1 appears to prevent D-GalN-induced apoptosis by a mitochondria-independent mechanism.
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