Molecular mechanisms of neurodegenerative diseases

Delio1 (Calissano) (sml)
The interruption of NGF signal activates amyloidogenesis. (Right) Immunofluorescence analysis performed with anti-Aβ antibody against amino acid residues 1–17 (MAb 6E10). Arrows mark varicosities forming along neurites after 6 h of NGF removal. (Left) Hoechst staining of nuclei.

Delio2 (Calissano) (sml)
Western blot analysis performed with APP C-terminal antibody of lysates from hippocampal neurons of controls (+NGF) or of neurons exposed to anti-NGF antibodies (αNGF) in a time ranging from 30 min to 48 h.

The hallmarks of Alzheimer disease are intracellular formation of neurofibrillay tangles NFT) and extracellular formation of senile plaques(SP). Formation of these structures is due to altered processing, conformation or post traslational modification of two proteins: tau and amyloid precursor protein(APP).

A widely accepted hypothesis is that APP altered processing leads to activation of the so called amyloidogenic route. This, in turn, initiates a cascade of events with excessive production and release of 1-40;1-42 peptides. These peptides, with special reference to 1-42, exert a toxic action on healthy neurons and aggregate to form senile plaques. Such altered processing, in turn, causes an hyperphosphorylation of tau which eventually leads to microtubule collapse and all subsequent intracellular events connected with an altered function of microtubules. The end result would be, according to this "cascade hypothesis", formation of NFT and neuronal death.

Our research group has hypothesized that, at least part of the above mentioned molecular events, are due to an unproper activation of apoptosis in specific, strategic neuronal populations such as those operating in hyppocampus and cerebral cortex. This hypothesis has been tested in vitro cultures of cerebellar granule cells, hyppocampal and cortical neurons as well as in a clonal cell line (PC12 cells) which, in the presence of NGF, cease dividing and acquire a neuronal phenotype., The bulk of these studies demonstrate that indeed apoptosis induced in these cultures is accompanied by the most relevant molecular events described (see selected list of publications).

We have found that a deficit of NGF or BDNF in hyppocampal cultures induces a marked increase of APP, PS1, beta and gamma secretase and a concomitant increased production of Aß, both intracellularly and in the culture medium, accompanied by cell death. Inhibition of Aß production by α and ß secretase inhibitors or by an anti Aß antibody prevents neuronal death. Exogenous administration of Aß to hyppocampal cultures induces events similar to those occurring after NGF or BDNF removal, suggesting that a deficit in one of these neurotrophins activates a sort of autocrine toxic loop whereby Aß released further worsens cell viability. These data demonstrate a link between the interruption of neurotrophin signalling and the activation of the amyloidogenic route.

Furthermore, we have shown that adenovirus-mediated overexpression of the NH2 26-230aa tau fragment which is formed during neuronal apoptosis evokes a potent neurotoxic effect in primary neuronal cultures. This NH2-terminally cleaved tau fragment is also specifically detected in vivo in a model of progressive AD-like neurodegeneration and its expression correlates with a significant up-regulation of caspase(s) activity. Presently our research group is continuing the experimental work along these lines of investigations.