Biliverdin Reductase-A correlates with inducible nitric oxide synthasein in atorvastatin treated aged canine brain

Alzheimer's disease is a neurodegenerative disorder characterized by progressive cognitive impairment and neuropathology. Recent preclinical and epidemiological studies proposed statins as a possible therapeutic drug for Alzheimer's disease, but the exact mechanisms of action are still unknown. Biliverdin reductase-A is a pleiotropic enzyme involved in cellular stress responses. It not only transforms biliverdin-IX alpha into the antioxidant bilirubin-IX alpha but its serine/threonine/tyrosine kinase activity is able to modulate cell signaling networks. We previously reported the beneficial effects of atorvastatin treatment on biliverdin reductase-A and heme oxygenase-1 in the brains of a well characterized pre-clinical model of Alzheimer's disease, aged beagles, together with observed improvement in cognition. Here we extend our knowledge of the effects of atorvastatin on inducible nitric oxide synthase in parietal cortex, cerebellum and liver of the same animals. We demonstrated that atorvastatin treatment (80 mg/day for 14.5 months) to aged beagles selectively increased inducible nitric oxide synthase in the parietal cortex but not in the cerebellum. In contrast, inducible nitric oxide synthase protein levels were significantly decreased in the liver. Significant positive correlations were found between biliverdin reductase-A and inducible nitric oxide synthase as well as heme oxygenase-1 protein levels in the parietal cortex. The opposite was observed in the liver. Inducible nitric oxide synthase up-regulation in the parietal cortex was positively associated with improved biliverdin reductase-A functions, whereas the oxidative-induced impairment of biliverdin reductase-A in the liver negatively affected inducible nitric oxide synthase expression, thus suggesting a role for biliverdin reductase-A in atorvastatin-dependent inducible nitric oxide synthase changes. Interestingly, increased inducible nitric oxide synthase levels in the parietal cortex were not associated with higher oxidative/nitrosative stress levels. We hypothesize that biliverdin reductase-A-dependent inducible nitric oxide synthase regulation strongly contributes to the cognitive improvement observed following atorvastatin treatment.

The Gamma-Secretase-Mediated Proteolytic Processing of APP C-Terminal Fragments as a Therapeutic Target for Alzheimer's Disease

Jemila Houacine

Alzheimer's disease (AD) is a devastating neurodegenerative disorder which severely impairs cognitive functions by triggering neuronal cell death and synaptic loss, and finally leads the patients to death. Two main histopathological hallmarks can be found in the brain tissue of AD patients: the amyloid plaques composed of aggregated Amyloid-β peptides (Aβ) and the neurofibrillary tangles (NFTs) constituted of hyperphosphorylated tau protein. To explain the underlying molecular mechanisms of AD pathogenesis, the amyloid cascade hypothesis states that early accumulation of Aβ peptides (especially the Aβ42 specie identified as the most toxic one) triggers their assembly into oligomers and their subsequent extracellular aggregation into dense fibrillar deposits. The oligomeric Aβ assemblies further initiate a succession of neurotoxic events including disruptions at the synaptic level, inflammation, neuritic injury, altered calcium homeostasis, oxidative stress, and altered phosphorylation activity leading to the hyperphosphorylation of tau and its aggregation into NFTs. Finally, this cascade of neuronal deleterious effects induces cognitive dysfuntions leading to dementia. The Aβ peptides originate from the amyloidogenic processing of the amyloid precursor protein (APP), a type I membrane protein that undergoes a first cleavage by β-secretase to liberate the soluble APP domain (sAPPβ) in the extracellular space, and the membrane bound APP-C99 fragment. Then, APP-C99 is processed by the intramembrane aspartyl-protease gamma-secretase (γ-secretase) composed of four subunits (presenilin (PS), nicastrin (NCT), anterior pharynx-defective protein 1 (APH1), and presenilin enhancer 2 (PEN2)) to release the toxic Aβ peptides in the lumen, and the APP intracellular domain (AICD) in the cytosol. Alternatively, APP can undergo the non-amyloidogenic processing, in which it is first shedded by α-secretase to generate the sAPPα domain and the APP-C83 fragment. The latter is further cleaved by γ-secretase into the non toxic p3 peptide and the AICD. AD is the most frequent form of dementia in elderly humans. Despite this evidence, no treatment is currently available to prevent or cure this disease. Thus, multiple potential drugs are tested in clinical trials or are still being developed in preclinical studies. In this work, a new therapeutic approach to lower Aβ production by specifically targeting the γ-secretase-mediated processing of APP-C99 is presented. Monoclonal antibodies against APP-C99 were generated and characterized following mice immunization with the active recombinant substrate APP-C99. When tested in cells, these antibodies decreased the γ-secretase-dependent processing of APP-C99, thus lowering Aβ production. Furthermore, the intracerebroventricular injection of anti-APP-C99 antibodies in a mouse model of AD decreased total soluble Aβ levels. These results validated this new approach as a potential immunotherapeutic strategy to prevent and/or delay the neurotoxic effects caused by Aβ in AD pathogenesis. Next, the γ-secretase-mediated processing of the APP-carboxy-terminal fragments (APP-CTFs) regulating the toxic versus non-toxic pathways was further investigated with the help of in vitro activity assays. This comparative study of APP-C99 and APP-C83 processing by γ-secretase revealed that both substrates were identically processed by the enzyme, with the same AICDs being produced. In addition, the effects of familial AD (FAD) mutations in the PS subunit of highly pure and homogeneous γ-secretase complexes were also investigated, and showed a drastic loss-of-function phenotype linked to these mutants. Overall, in vitro studies of the APP-CTFs processing by γ-secretase underlined particular functional aspects of this processing, which could be related to the cellular pathways involved in AD pathogenesis such as the competition between the amyloidogenic and non-amyloidogenic pathways and the FAD-linked impaired metabolism of APP.

EPFL2011

The Gamma-Secretase-Mediated Proteolytic Processing of APP C-Terminal Fragments as a Therapeutic Target for Alzheimer's Disease

Jemila Houacine

EPFL2011