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Published on: March 24, 2019
Altered protein expression and protein nitration pattern during d-galactosamine-induced cell death in human
Antonio Rodríguez-Ariza1, Laura M López-Sánchez, Raul González
1Liver Research Unit, Hospital Universitario Reina Sofía, Córdoba, Spain. antonio.rodriguez.exts@juntadeandalucia.es
Background/Aims:
Hepatic injury by d-galactosamine (d-GalN) is a suitable experimental model of hepatocellular injury. The induction of oxidative and nitrosative stress participates during d-GalN-induced cell death in cultured rat hepatocytes. This study aimed to identify protein expression changes during the induction of apoptosis and necrosis by d-GalN in cultured human hepatocytes.
Methods:
A proteomic approach was used to identify the proteins involved and those altered by tyrosine nitration. A high dose of d-GalN (40 mM) was used to induce apoptosis and necrosis in primary culture of human hepatocytes. Cellular lysates prepared at different times after addition of d-GalN were separated by two-dimensional electrophoresis. Gel spots with an altered expression and those matching nitrotyrosine-immunopositive proteins were excised and analyzed by mass spectrometry.
Results:
d-GalN treatment upregulated microsomal cytochrome b5, fatty acid binding protein and manganese superoxide dismutase, and enhanced annexin degradation. d-GalN increased tyrosine nitration of four cytosolic (Hsc70, Hsp70, annexin A4 and carbonyl reductase) and three mitochondrial (glycine amidinotransferase, ATP synthase beta chain, and thiosulfate sulfurtransferase) proteins in human hepatocytes.
Conclusions:
The results provide evidences that oxidative stress and nitric oxide-derived reactive oxygen intermediates induce specific alterations in protein expression that may be critical for the induction of apoptosis and necrosis by d-GalN in cultured human hepatocytes.
