Roger M. Nitsch

Regulation of proteolytic processing of the amyloid beta-protein precursor of Alzheimer's disease in transfected cell lines and in brain slices.

beta A4 is the principal component of Alzheimers disease brain amyloid. It is derived from proteolytic processing of amyloid beta-protein precursors (APP), a family of transmembrane glycoproteins. Secretion of APPs, a secreted proteolytic derivative that is cleaved within the beta A4 domain of APP, is increased many-fold by the activation of cell-surface receptors, like the muscarinic m1 and m3 receptor subtypes, which are coupled to protein kinase C. Concomitantly, their activation decreases the formation of both secreted soluble beta A4 and of endosomal-lysosomal C-terminal APP derivatives. These data suggest that muscarinic m1 and m3 receptors accelerate non-amyloidogenic APP processing and depress the formation of potentially amyloidogenic derivatives. Other receptors that stimulate APPs secretion include those for bradykinin, vasopressin, and interleukin-1 receptors. A similar control mechanism is present in rat brain tissue slices, in which the release of both APPs and endogenous neurotransmitters is increased by electrical depolarization. This increase is tetrodotoxin-sensitive and frequency-dependent, suggesting that APPs release may normally depend on neuronal activity. Taken together, our findings suggest that specific receptor agonists might be effective in reducing the formation of potentially amyloidogenic APP derivatives in vivo.

Regulation of APP Processing Potential for the Therapeutical Reduction of Brain Amyloid Burdena

The role of brain amyloid in the pathogenesis of Alzheimers disease (AD) is discussed controversially, but combined genetic and biochemical evidence points to a central role of the gene encoding the amyloid precursor APP in at least some forms of AD. This article proposes that preventing brain amyloid formation is a rational concept for drug treatment of AD. We suggest that pharmacologically active ligands for specific cell surface receptor subtypes—normally stimulated by neurotransmitters, growth factors, and cytokines—constitute a class of chemicals that might be useful to accelerate processing of APP into non‐amyloidogenic, and biologically active, derivatives. This class of agents includes muscarinic m1 and m3 agonists, serotoninergic 5‐HT2a and 5‐HT2c agonists, glutamatergic mGluR1 agonists, as well as agonists for bradykinin and vasopressin receptors.

Regulation of Amyloid Precursor Protein Release By Protein Kinase C in Swiss 3T3 Fibroblastsa

Release of the amyloid precursor protein (APP) of Alzheimers disease from Swiss 3T3 fibroblasts was stimulated in a concentration‐dependent manner by phorbol 12‐myristate 13‐acetate. In fibroblasts overexpressing protein kinase Co (PKCα), the EC50 for this response was 7 nM, while in control cells the EC50 was 63 nM. The effect of PMA was inhibited by the PKC antagonist H‐7 in control cells, but not in cells that overexpressed PKCα. Basal release of APP was higher in cells that overexpressed PKCα, and was not affected by the phosphatase inhibitor okadaic acid, although this compound doubled APP release from control cells. The results suggest that PKCα regulates APP processing in mammalian cells. Alterations in the activity of PKC have been reported to occur in Alzheimers disease and might potentially contribute to abnormalities of APP metabolism characteristic of this disorder.

Phospholipid metabolite levels are altered in cerebral cortex of patients with dominantly inherited olivopontocerebellar atrophy

We measured metabolic precursors and breakdown products of phosphatidylcholine (choline, glycerophosphocholine (GPC) and phosphatidylethanolamine (ethanolamine, glycerophosphoethanolamine (GPE)) as well as the amino acid serine, a precursor of phosphatidylserine, in four morphologically unaffected cerebral cortical areas obtained at autopsy from 14 patients with dominantly inherited olivopontocerebellar atrophy (OPCA) and 13 controls matched for age and postmortem interval. As compared with the controls, mean GPE levels were elevated by 49-57% in frontal and parietal cortices of OPCA brains whereas concentrations of ethanolamine were significantly reduced in temporal, occipital and parietal cortex (-40 to -54%). This resulted in increased GPE/ethanolamine ratios (+80 to +146%). GPC levels were significantly increased (by 53%) in the frontal cortex of OPCA patients relative to controls. Free serine levels were reduced by 20 to 28% in frontal, parietal, temporal, and occipital cortices. These abnormalities in phospholipid metabolite levels in OPCA resemble those seen in Alzheimers disease, although the changes in GPC are less pronounced. These changes in phospholipid metabolism in OPCA cerebral cortex, a brain area spared from neurodegenerative changes, points to generalized disturbances in cellular membrane function in this disease.

Regulation of APP Processing by First Messengers

Brain amyloid deposits are invariant neuropathological hallmarks of Alzheimer’s disease (AD) and Down’s syndrome, and are sometimes also found in lesser amounts in brains of non-demented aged human subjects. Alzheimer disease-type brain amyloid consists of aggregated amyloid s- (As) peptides which are self-aggregating molecules of 39–43 residues in length. As is derived, by proteolytic processing, from a larger amyloid s-protein precursor (APP). The hydrophobic C-terminal region of the amyloidogenic As domain is located within the single transmembrane domain of APP, and its N-terminus extends 28 residues into the ectodomain (for review, see Selkoe, 1994). The APP gene is expressed at remarkably high levels in brain but is also expressed in many peripheral tissues. The biological function of APP is unclear and it is possible that individual proteolytic derivatives of APP have distinct biological consequences. For instance, accumulating evidence suggests that the full-length protein and its secreted N-terminal derivatives can promote cell adhesion (Schubert et al., 1989), stimulate neunte outgrowth (Milward et al., 1992), and protect cultured neurons from excitotoxic damage (Mattson et al., 1993). In contrast, As and its aggregates can be cytotoxic and induce cell death in cultured neurons (Yankner et al., 1990; Loo et al., 1993). Moreover, As inhibits the normal function of a potassium channel which appears to be impaired in fibroblasts obtained from AD patients (Etcheberrigaray et al., 1994). These initial data support the concept that large portions of the N-terminal ectodomain have trophic functions whereas the As domain can be cytotoxic. The biochemical mechanisms involved in the regulation of local brain tissue concentrations of particular proteolytic APP derivatives may thus play an important role in determining the actual functions of APP.

Choline Metabolism, Membrane Phospholipids, and Alzheimer’s Disease

Alzheimer’s Disease (AD) and even apparently-normal aging are associated with defects in brain choline metabolism. This chapter describes these abnormalities as well as some of the experimental systems used to study their mechanisms and effects. Cell culture, superfused brain slices, in vivo microdialysis, postmortem tissue analysis, and in vivo magnetic resonance spectroscopy (MRS) studies have been employed. Available data suggest that these defects provide a biochemical basis for the special vulnerability of cholinergic neurons in AD.

Modulation of APP Processing by Neurotransmission

Brain amyloid deposits are invariant neuropathological hallmarks of Alzheimer’s disease (AD) and Down’s syndrome, and are sometimes also found in lesser amounts in brains of neuropsychologically normal, aged human subjects. AD-type brain amyloid consists of aggregated As peptides which are 39–43 amino acid residues in length. As is derived, by proteolytic processing, from a larger amyloid s-protein precursor (APP), which is a transmembrane glycoprotein that contains a single membrane spanning domain, a large N-terminal ectodomain and a short cytoplasmic C-terminal tail. The As domain is located within the ectodomain and extends with its hydrophobic C-terminal region 11-15 residues into the membrane. APP exists in various forms generated by alternative splicing of mRNA derived from a single gene on chromosome 21 (for review, see Kosik, 1992). APP is highly conserved and expressed at high levels in brain and, at lower levels, in many peripheral tissues. The biological function of APP is unclear but accumulating evidence suggests roles in cell adhesion (Schubert et al., 1989), in neurite outgrowth (Milward et al., 1992), as well as excitoprotective functions via the regulation of intracellular calcium concentrations (Mattson et al., 1993). Mature APP is rapidly degraded by various alternative proteolytic processing pathways. Proteolytic derivatives are secreted into the extracellular space and are found at high concentrations in human cerebrospinal fluid. Secreted APP derivatives include the large N-terminal ectodomain, termed APPs (Esch et al., 1990; Sisodia et al., 1990) and ~4KDa As-peptides (Haass et al., 1992; Shoji et al., 1992) that potentially can aggregate into amyloid. In addition to the secretory processing pathways, full-length APP can be internalized from the cell surface and targeted to the endosomal-lysosomal system (Haass et al., 1992), where multiple cleavage products are generated. Some of these contain the intact sA4 domain and thus are also potentially amyloidogenic (Golde et al., 1992; Estus et al., 1992). As is neurotoxic in some experimental systems (Yankner et al., 1990) and may induce apoptosis (Loo et al., 1993). Thus, APP processing pathways yielding either As or APPs are likely to have distinct cellular consequences: processing events that generate As may be toxic and are potentially amyloidogenic, whereas APP processing to yield APPs generate trophic and precludes APP’s role as an amyloidogenic molecule. It thus becomes important to understand the cellular mechanisms involved in the regulation of APP processing pathways.

Accelerated Cell Membrane Degradation in Alzheimer’s Disease Brain: Relationship to Amyloid Formation?

Alzheimer’s disease is a degenerative brain disease first described by the German psychiatrist Alois Alzheimer (Alzheimer, 1907). His initial report was based upon the histological examination of brain tissue from a 51 year old female patient who experienced a rapid progressive deterioration of memory and died four and a half years after the onset of the first symptoms with total loss of mental functions. The brain of this woman showed generalized atrophy without macroscopic lesions. Using a novel silver staining method developed by Bielschowsky, Alzheimer found that more than a quarter of the cortical neurons contained fibrillary tangles, and that this abnormality was accompanied by the extracellular deposition of “multiple miliary lesions”, containing a “unique substance” which was later recognized to be amyloid. In his initial report, Alzheimer also described severe loss of nerve cells, which was most prominent in the upper layers of the brain cortex. More recently, Terry and colleagues used computerized image processing to confirm the loss of cortical neurons, and showed that large cortical neurons are particularly affected (Terry et al., 1981). Neuronal atrophy and the presence of degenerating neurites in the immediate vicinity of amyloid plaques has generated two controversial hypotheses: First, that amyloid is toxic to neurons, and, second, that amyloid is a byproduct of neuronal atrophy formed by neurons during their degeneration.