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Kulhanek-Heinze, Stefanie (2004): Characterisation of Anti-Apoptotic Signalling Pathways in Hepatocytes activated by alpha-Lipoic Acid and Atrial Natriuretic Peptide. Dissertation, LMU München: Faculty of Chemistry and Pharmacy
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Abstract

Both, the R-enantiomer of the antioxidant alpha-lipoic acid (R-LA) and the hormone atrial natriuretic peptide (ANP) are known to exert potent hepatoprotective action. The present work characterises alpha-lipoic acid- and ANP-mediated signal transduction pathways involved in the regulation of apoptotic cell death in two different models: primary hepatocytes and ischemic isolated perfused rat livers. alpha-lipoic acid was shown to protect isolated hepatocytes from TNF-alpha-/ActinomycinD-induced apoptosis. Astonishingly, this effect did not seem to be governed neither by its well described antioxidative nor its Fe-chelating properties. In fact, the LA-mediated activation of the PI3-K/Akt survival pathway seemed to be responsible for the antiapoptotic properties of alpha-lipoic acid. Consequently, incubation with a specific PI3-K-inhibitor significantly reduced both, R-LA-mediated decrease in caspase activity and R-LA-induced BAD phosphorylation. Thus, PI3-K-mediated Akt activation and subsequent phosphorylation of the proapoptotic protein BAD at Ser136 are causally involved in the antiapoptotic signalling mediated by R-LA. Perfusion with ANP 20 min prior to the ischemic period is known to reduce apoptotic cell death occurring at the end of the ischemic period. We could previously show that this preconditioning of rat livers leads to a marked activation of p38 MAPK. Since ANP reduces apoptotic cell death, the potential connection between this ANP-induced p38 MAPK activation and apoptosis reduction was investigated. Astonishingly, liver perfusion with an p38 MAPK inhibitor even decreased apoptotic cell death, supporting a detrimental role of this kinase. PKA-specific inhibitors demonstrated the involvement of PKA in this ANP-mediated protection. Interestingly, it also turned out that PKA phosphorylates the proapoptotic protein BAD at Ser112, an effect known to contribute to the inhibition of apoptosis. In summary, the present data show for the first time that phosphorylation of BAD at either Ser136 or Ser112 turns out to be a central protective mechanism to defend from hepatocyte apoptosis.