ABSTRACT
Amyloids have a number of structural features in common, that is, the cross-β structure, the binding to characteristic dyes, like Congo Red (CR) and Thioflavine T (ThT), the fibrillar morphology observed by transmission electronic microscopy (TEM) or atomic force microscopy (AFM), and the green birefringence under polarized light upon staining with CR (Nilsson, 2004). Table 15.2 shows the intracellular aggregates known as inclusion bodies. At least one of these aggregates, the neurofibrillary tangles, has the same structural features than the amyloids. The tangles are thus “intracellular amyloid.” Other inclusions may have some, but not all, of these properties, for example, inclusion bodies within muscle fibers of inclusion body myositis, which are stained by CR, exhibit green birefringence and are immunoreactive with anti-β-amyloid. Table 15.2 Intracellular inclusions with known biochemical composition, with or without amyloid properties (Sipe at al., 2010)
Inclusion name Site
Protein nature
Examples of associate diseaseLewy bodies Neurons intracytoplasmic a-synuclein Parkinson’s diseaseHuntington bodies Neurons intranuclear PolyQ expanded huntingtin Huntington’s diseaseHirano bodies Neurons Actin Neurodegenerative disordersCollins bodies Neurons Neuroserpin Familial presenile dementiaNot specified Neurons, many different cells Ferritin Familial neurodegenerative disorders Most of conformational diseases share a striking number of common pathological features, such as evidence of membrane damage, oxidative stress, mitochondrial dysfunction, up-regulation of autophagy, and cell death (Glabe, 2006). In this way, amyloids from different diseases may share a common pathway for fibril formation. The initiating event is a protein misfolding, which results in the acquisition of the ability to aggregate in an infinitely propagating
fashion. Quasistable intermediate aggregates ranging from dimers up to particles of a million Dalton or greater have been observed by a variety of methods. Soluble spherical aggregates have been observed for many different types of amyloids and it is assumed that these spherical oligomers, usually called amyloid-β-derived diffusible ligands (ADDLs) appear to represent intermediates in the pathway of fibril formation. Recent evidence suggest that amyloid oligomers, which represent intermediates in the fibril formation progress, may be primarily responsible for amyloid pathogenesis, rather than the mature fibrils that accumulate as large aggregates (Hardy and Selkoe, 2002; Lambert et al., 1998), even when the oligomers are formed from proteins that are not normally related to degenerative diseases (Bucciantini, 2002). Amyloid oligomers and fibrils have in common the structural features indicated above. Amyloid oligomers are generically toxic to cells. This fact and the common structure shared by the amyloid oligomers imply that the primary mechanism of toxicity in these diseases must be the same or, said in other words, that the oligomers must act on the same primary target. As some amyloids arise from cytosolic proteins, while others are derived from extracellular proteins, the most obvious target that is accessible to both cytosolic and extracellular compartments is the plasma membrane that forms the interface between the two compartments. A growing body of evidence suggests that membrane permeabilization by amyloid oligomers may represent the common, primary mechanism of pathogenesis of amyloidosis (Glabe, 2006). Concerning the neurodegenerative diseases, Parkinson’s disease (PD) and Alzheimer’s disease (AD) are the diseases with a greater incidence on the world population. Both diseases lack definite diagnostic approaches and effective cure at the present. Moreover, the currently available diagnostic tools are not sufficient for an early screening of AD/PD in order to start preventive approaches. However, the emerging field of nanotechnology has promised new techniques to solve some of the AD/PD challenges. Nanotechnology uses engineered nanomaterials or devices that can interact with biological systems at molecular levels with a high degree of specificity (Silva, 2006; Editorial, 2003). Thus they can stimulate, respond to, and interact with target cells and tissues in controlled ways to induce desired physiological responses, while minimizing
undesirable effects. With all this potential, nanotechnology could have a revolutionary impact on diagnosis and therapy of PD and AD (Morris, 2004). In this chapter, we present the promises that nanotechnology brings in research on the PD/AD diagnosis and therapy. 15.2 Alzheimer’s DiseaseAD is one of a number of diseases in which proteins form amyloid aggregates. AD is a progressive, neurodegenerative disease, which inevitably leads to dementia and death. The brains of patients with AD contain a large number of amyloid deposits in the form of senile plaques. The amyloid core of these plaques is formed by interwoven fibrils that are composed by variants of the b-amyloid (Ab) peptide. These fibrils are surrounded by dead neurones. Ab varies from 39 to 43 amino acids in length, the most abundant forms being 40 and 42 amino acids (Glenner and Wong, 1984). Ab is generated from the proteolytic processing of the amyloid precursor protein (APP) by β-and γ-secretases. Although a causal relationship between Ab and the development of AD has not been conclusively demonstrated, considerable experimental data suggest that Aβ aggregates are important in the etiology of AD (Harper and Lanbury, 1997; Murphy, 2002; Soto, 2003; Gorman and Chakrabarty, 2001). Whereas early evidence suggested that Ab fibrils initiate a cascade of events that result in neuronal cell death(Yankner, 1996), a number of investigators recently proposed that soluble aggregates of Ab (also called oligomers or protofibrils), rather than monomers or insoluble amyloid fibrils, may be responsible for synaptic dysfunction in AD (Hardy and Selkoe, 2002; Hartley et al., 1999; Klein et al., 2001; Westerman et al., 2002; Kawarabayashi et al., 2004). Nevertheless, Aβ fibrillogenesis is still thought to play a critical role in the development of AD. On the other hand, the conversion of Aβ, which in native form is random, into fibrils rich in β-form, involves major structural changes, leading to the partial or complete disruption of the native fold (Klein et al., 2004). In vitro studies have suggested that Ab fibrillogenesis occurs by a multistep, nucleation-dependent process (Jarret and Lansbury, 1993). Formation of the nucleation seed is rate limiting,
so in the absence of preformed seed fibrils, there is a significant lag period for the formation of the Aβ fibrils, followed by a rapid fibril elongation phase once seed fibrils, or nuclei, have been generated. Therefore, the fibrillogenesis can be considered to occur in two distinct stages, controlled by two key parameters, nucleation rate (kn) and elongation rate (ke) constants.This model is similar to a nucleation-dependent polymerization process that characterizes crystal growth. The kinetics of such a fibrillization process can be explained in terms of an autocatalytic reaction mediating the transition from the monomer to the aggregate species (Sabaté et al., 2003).Aβ fibrillogenesis can be completed by considering the presence of oligomers (Haass and Selkoe, 2007), as well as of micelles (Sabaté and Estelrich, 2005a). As in other amyloidoses, one mechanism by which oligomers can damage the neurons in AD is the formation of pores or ion channels through the cell membrane. Early work in this area showed that Ab can insert into planar lipid bilayers and allow a calcium current upon insertion and further that these channels can be blocked (Arispe et al., 1993), suggesting that the calcium current is really due to channel formation, not just bilayer permeabilization of the peptide. More recent work has been done using AFM, showing that peptide monomers oligomerize after insertion into the bilayer. Furthermore, in the presence of these oligomers, current can flow (Lin et al., 2001). However, other work has indicated that soluble oligomers specifically increase lipid bilayer conductance, while fibrils and soluble low-molecular-weight species have no effect (Kayed et al., 2004). Microglia, the resident macrophage cells of the central nervous system (CNS), play a controversial role in AD: some types of reactive microglia seem detrimental, whereas others can actually be beneficial (Weitz and Town, 2012). A large part of microglial cell’s role in the brain is maintaining homeostasis in noninfected regions and promoting inflammation in infected or damaged tissue. Microglia respond to different pathological agents with a reaction termed “microglia activation.” Once activated, microglia release both neuroprotective and cytotoxic molecules. One of such molecules, interleukin-1β, when overexpressed, leads to the hyperphosphorilation of tau protein). Activated microglia synthesize APP and they are found to be associated with amyloid deposits in the brains of AD patients.
