Chapter 5 Current concepts on the etiology and pathogenesis of Parkinson disease
Anatomical and biochemical pathology of PD
Historical introduction
Parkinson disease (PD) was first described in 1817 with the publication by James Parkinson of a book entitled An Essay on the Shaking Palsy (Parkinson, 1817). In it, he described six individuals with the clinical features that have come to be recognized as a disease entity. One of the people was followed in detail over a long period of time; the other five consisted of brief descriptions, including two of whom he had met walking in the street, and another whom he had observed at a distance. Such distant observations without a medical examination demonstrate how easily distinguishable the condition is. The physical appearance of flexed posture, resting tremor, and shuffling gait are readily recognizable. Parkinson’s opening description has his key essentials: “Involuntary tremulous motion, with lessened muscular power, in parts not in action and even when supported; with a propensity to bend the trunk forward, and to pass from a walking to a running pace: the senses and intellects being uninjured.” In the small monograph, Parkinson provided a detailed description of the symptoms and also discussed the progressive worsening of the disorder, which he called “the shaking palsy” and also its Latin term “paralysis agitans.”
After the publication of Parkinson’s Essay, the disease was widely accepted in the medical community. It took 70 years for the name of the disorder to be referred to as “Parkinson’s disease,” as recommended by the French neurologist Charcot (1879) who argued against the term “paralysis agitans” (see Goetz, 1987, for English translation) and recommended the disorder be named after James Parkinson. Charcot argued that there is no true paralysis, but rather the “lessened muscular power” is what is today called akinesia, hypokinesia, or bradykinesia; all three terms often being used interchangeably by clinicians, although these three terms specifically refer to lack of movement, small movement, and slow movement, respectively. These terms represent a paucity of movement not due to weakness or paralysis. Similarly, Charcot emphasized that tremor need not be present in the disorder, so “agitans” and “shaking” are not appropriate as part of the name of the disorder.
The clinical features of PD and its differential diagnosis are presented in Chapter 4.
Incidence, prevalence, and mortality
Although PD can develop at any age, it begins most commonly in older adults, with a peak age at onset around 60 years. The likelihood of developing PD increases with age, with a lifetime risk of about 2% (Elbaz et al., 2002). A positive family history doubles the risk of developing PD to about 4%. A summation from seven population-based studies in various European countries found the overall prevalence of PD in people aged over 65 to be 1.8%, with an increase from 0.6% for persons aged 65–69 years to 2.6% for people aged 85–89 years (de Rijk et al., 2000). Twin studies indicate that PD with an onset under the age of 50 years is more likely to have a genetic relationship than for patients with an older age at onset (Tanner et al., 1999). Males have higher prevalence (male-to-female ratio of 3:2) and incidence rates than females (Fig. 5.1) (Bower et al., 2000), but the age-specific incidence rates have not varied over the past 70 years (Fig. 5.2) (Rocca et al., 2001). The incidence rates vary among studies, but average between 11.0 and 13.9/100 000 population per year (Van Den Eeden et al., 2003). In a northern California study, the incidence rates varied among ethnic groups, being highest in Hispanics, then non-Hispanic whites, then Asians, and lowest in African-Americans (Van Den Eeden et al., 2003).
The prevalence rates of PD have varied in different studies and in different countries. The figure of 187 per 100 000 population given by Kurland (1958) is a reasonable estimate in the US. But if the population is restricted to adults older than 39 years of age, the prevalence rate is 347 per 100 000 (Schoenberg et al., 1985) since both prevalence and incidence rates increase with age. At age 70, the prevalence is approximately 550 per 100 000, and the incidence is 120 per 100 000/year. At the present time, approximately 850 000 individuals in the US have PD, although one recent estimate is less than this (Hirtz et al., 2007). The number is expected to grow as the population ages (Dorsey et al., 2007). Advancing age is the single greatest risk factor for developing sporadic PD. Approximately 2% of the population will have PD by the time they reach the age of 80 years.
In the pre-levodopa era, mortality was reported to be three-fold greater in patients with PD (Hoehn and Yahr, 1967). The mortality rate was reduced to 1.6-fold greater than age-matched non-PD individuals after the introduction of levodopa (Yahr, 1976; Elbaz et al., 2003). Today, patients with PD can live 20 or more years, depending on the age at onset (Kempster et al., 2007). Death in PD is usually due to some concurrent unrelated illness or due to the effects of decreased mobility, aspiration, or increased falling with subsequent physical injury. The Parkinson-plus syndromes typically progress at a faster rate and often cause death within 9 years. Thus, the diagnosis of PD versus other forms of parkinsonism is of prognostic importance, as well as of therapeutic significance because it almost always responds to at least a moderate degree with levodopa therapy, whereas the Parkinson-plus disorders usually do not.
Anatomical pathology of PD
It was many years after Parkinson’s original description before the basal ganglia were recognized by Meynert in 1871 as being involved in disorders of abnormal movements. And it was not until 1895 that the substantia nigra (SN) was suggested to be affected in PD. Brissaud (1895) suggested this on the basis of a report by Blocq and Marinesco (1893) of a tuberculoma in that site that was associated with a contralateral hemiparkinsonian tremor. These authors were careful to point out that the pyramidal tract and the brachium conjunctivum above and below the level of the lesion contained no degenerating fibers. The importance of the SN was emphasized by Tretiakoff in 1919. He studied the SN in nine cases of PD, one case of hemiparkinsonism, and three cases of postencephalitic parkinsonism, finding neuronal loss in this nucleus in all cases. With the hemiparkinsonian case, Tretiakoff found a lesion in the nigra on the opposite side, concluding that the nucleus served the motor activity on the contralateral side of the body. The SN, so named because of its normal content of neuromelanin pigment, was noted to show depigmentation, loss of nerve cells, and gliosis. These findings remain the principal and essential histopathologic features of the disease. In his study, Tretiakoff also found Lewy bodies in the SN, expanding the earlier observation of Lewy (1912, 1914), who had discovered the presence of these cytoplasmic inclusions in the substantia innominata and the dorsal vagus nucleus in PD. Lewy bodies (Fig. 5.3) are now widely recognized as the major pathologic hallmark of the disorder. Lewy bodies have since been seen in autonomic ganglia, the peripheral nervous system and in other regions of the central nervous system, including the cerebral cortex (Jellinger, 2009a).
Foix and Nicolesco (1925) made a detailed study of the pathology of PD in 1925 and found that the most constant and severe lesions are in the substantia nigra. Since then many workers, including Hassler (1938) and Greenfield and Bosanquet (1953), have confirmed these findings and added other observations, including involvement of other brainstem nuclei such as the locus coeruleus and the raphe nuclei. The pigmented cells of the locus coeruleus contain neuromelanin; these cells are also lost in PD, with many of those remaining containing Lewy bodies. The asymmetry of clinical signs in PD is reflected by the asymmetrical and more severe contralateral loss of substantia nigra pars compacta (SNc) neurons. Neuronal loss extends beyond loss in SNc, locus coeruleus, and raphe, with loss of neurons in the dorsal motor vagal nucleus, hypothalamus, the nucleus basalis of Meynert, and sympathetic ganglia (Forno, 1982; Jellinger, 1987). There are also losses of glutamatergic projection neurons from thalamus to the basal ganglia (Henderson et al., 2000) and glutamatergic projection neurons from the presupplementary motor cortex to the premotor cortices (Halliday et al., 2005).
PD and the Parkinson-plus syndromes have in common a degeneration of SNc dopaminergic neurons, with a resulting deficiency of striatal dopamine due to loss of these nigrostriatal neurons. Accompanying this neuronal loss is an increase in glial cells in the nigra and a loss of the neuromelanin normally contained in the dopaminergic neurons. There is a reduction of nigral neurons and striatal dopamine with aging also (Carlsson and Winblad, 1976; McGeer et al., 1977; Fearnley and Lees, 1991) (Fig. 5.4), and although PD is associated with increasing age, the greater rate (Fig. 5.5) and the pattern of cell loss in the SN differs between that in aging and that in PD (Figs 5.6, 5.7) (Fearnley and Lees, 1991). Clinical features begin to emerge when approximately 80% of striatal dopamine content (or 60% of nigral dopaminergic neurons) are lost (Bernheimer et al., 1973). The course of clinical decline is associated with the progressive reduction of striatal dopamine (Riederer and Wuketich, 1976).
Figure 5.5 Total pigmented nigral neurons with disease duration.
From Fearnley JM, Lees AJ. Ageing and Parkinson’s disease: substantia nigra regional selectivity. Brain 1991;114:2283–2301.
Pathologically, almost all patients with PD have Lewy bodies in the SN and locus coeruleus. Juvenile PD (Dwork et al., 1993; Hattori et al., 2000), now recognized as being due mainly to homozygous parkin mutations, is a major exception. Another exception is with PD due to the R1441G mutation in the LRRK2 gene (Martí-Massó et al., 2009).The pathology has not yet been described in juvenile PD patients due to homozygous mutations in DJ-1 genes, but in one case of young-onset PD with PINK1 mutation, the autopsy showed the presence of Lewy bodies (Samaranch et al., 2010). There are no Lewy bodies in the Parkinson-plus syndromes or in postencephalitic parkinsonism (Jellinger, 2009b).
The presence of Lewy bodies in the SNc and the locus coeruleus plus the clinical features of PD (without the characteristic clinical features of some other form of parkinsonism) are usually used to make the pathologic diagnosis of PD, but there is no complete agreement among neuropathologists about the pathologic criteria for the diagnosis of PD (Forno, 1982, 1996). Some patients with clinical PD die with nigral degeneration without Lewy bodies (Rajput et al., 1991; Hughes et al., 1992, 2002). In fact, as mentioned above, patients with juvenile PD usually do not have Lewy bodies, especially those with homozygous parkin mutations (Dwork et al., 1993; Hattori et al., 2000). Thus, Lewy bodies confirm the diagnosis and are a critical pathologic marker to confirm the diagnosis, but they are not necessary and their presence is not pathognomonic for PD (Forno 1982). They are found in 4–6% of routine autopsies (Forno, 1982), the incidence rate increases with age as in PD (Fearnley and Lees, 1991) (Table 5.1), and people dying with such incidental Lewy bodies are considered to have a presymptomatic state of PD. The site of regional nigral neuronal loss in incidental Lewy body disease is the lateral region in the ventral tier, the same as for PD (Fig. 5.6) (Fearnley and Lees, 1991). These brains show a reduction in striatal dopaminergic markers (e.g., tyrosine hydroxylase and vesicular monoamine transporter 2), but not as severe as those with clinical PD (DelleDonne et al., 2008; Dickson et al., 2008). Cortical Lewy bodies in patients with dementia and no parkinsonism could be a separate disease or a variant in the presentation of the same disorder that causes PD. Lewy bodies consist of a dense inner core surrounded by a radiating filamentous outer zone (Fig. 5.8) (Duffy and Tennyson, 1965; Forno and Norville, 1976).
| Age | No. | Percent |
|---|---|---|
| <20 | 0/2 | |
| 20–29 | 0/6 | |
| 30–39 | 0/6 | |
| 40–49 | 1/7 | 3.7 |
| 50–59 | 0/20 | |
| 60–69 | 2/51 | 3.9 |
| 70–79 | 3/56 | 5.4 |
| 80–89 | 6/49 | 12.2 |
| 90–99 | 1/8 | 12.5 |
Data from Fearnley JM, Lees AJ. Ageing and Parkinson’s disease: substantia nigra regional selectivity. Brain 1991;114:2283–2301.
Biochemical pathology of PD
The pigmented neurons of both the SNc and the ventral tegmental area (medial to the SN in the midbrain) contain dopamine. The former neurons project to the neostriatum, the latter to the limbic system and the neocortex. In PD, the mesolimbic and mesocortical neurons are relatively spared, whereas the nigrostriatal neurons are progressively lost. As a result, there is a corresponding decrease of dopamine content in both the nigra and the striatum, with the innervation of the posterior putamen affected first and most severely, as can be detected in FDOPA PET scans (Fig. 5.9). Whereas dopamine is reduced initially in the posterior striatum in PD, over time, as the disease progresses, all striatal subregions are affected to a similar degree (Nandhagopal et al., 2009).
The pigmented neurons of the locus coeruleus contain norepinephrine, and these neurons project widely in the CNS. A third set of monoaminergic neurons are those containing serotonin (5-HT), located in the raphe of the pons and medulla. In PD there is a progressive loss of all three types of monoaminergic cells, particularly the dopaminergic cells. So, in addition to a depletion of striatal dopamine, there is also a reduction in brain norepinephrine and 5-HT (Ehringer and Hornykiewicz, 1960; Hornykiewicz, 1966) (Table 5.2). There is also a reduction in other neurotransmitters (Agid et al., 1987) as well as enzyme activities for the synthesis of other neurotransmitters (Table 5.2), indicating that the biochemical changes in PD extend beyond the loss of only the monoamines.
Table 5.2 Concentration of neurotransmitters, their metabolites, and synthesizing enzymes in selected regions of brain of controls and PD
In addition to the motor features of PD, the symptoms that define the clinical diagnosis, there are also a host of nonmotor symptoms, some occurring before the motor symptoms and some occurring after (see Chapter 8). The early motor symptoms of bradykinesia and rigidity and tremor are associated with monoaminergic cell and chemical loss. The later motor symptoms of flexed posture, loss of postural reflexes, and the freezing phenomenon appear to correlate poorly with dopaminergic deficit. The nonmotor symptoms probably are the result of loss of neuronal function other than dopamine. Catecholamine reduction in PD is seen in the autonomic nervous system and accounts for the reduction in MIBG SPECT scan labeling in the heart in PD due to loss of postganglionic myocardial sympathetic nerve fibers (Rascol and Schelosky, 2009).
Among the neurotransmitter changes is the reduction of brain acetylcholine (Bohnen and Albin, 2010). Acetylcholinesterase (AChE), which serves as a marker for cholinergic neurons, can be measured by PET scanning (Shimada et al., 2009). A reduction of AChE begins early in PD (Bohnen and Albin, 2009). Reduced thalamic AChE activity correlates with falling in PD (Bohnen et al., 2009), and in part represents decreased cholinergic output of the pedunculopontine nucleus (PPN), which appears to be important for gait. Cortical loss of acetylcholine probably contributes to the dementia seen in PD (Shimada et al., 2009; Bohnen and Albin, 2010). Overall, reduced AChE is more widespread and profound both in PD with dementia and in dementia with Lewy bodies (Shimada et al., 2009).
There are compensatory changes, such as supersensitivity of dopamine receptors, so that symptoms of PD are first encountered only when there is about an 80% reduction of dopamine concentration in the putamen (or a loss of 60% of nigral dopaminergic neurons) (Bernheimer et al., 1973). Another compensatory mechanism is an increase in neurotransmitter turnover, as detected by an increased HVA/DA ratio. In a major review, Hornykiewicz (1966) correlated loss of dopamine concentration in the striatum with severity of bradykinesia and rigidity in PD. With further loss of dopamine concentration, parkinsonian bradykinesia becomes more severe. The progressive loss of the dopaminergic nigrostriatal pathway can be detected during life using PET and SPECT scanning (Fig. 5.9); these show a continuing reduction of FDOPA and dopamine transporter ligand binding in the striatum that correlates with the bradykinesia score in the Unified Parkinson’s Disease Rating Scale (Brooks et al., 1990; Snow et al., 1994; Seibyl et al., 1995; Morrish et al., 1996; Vingerhoets et al., 1997; Broussolle et al., 1999; Benamer et al., 2000). Using special statistical techniques, FDG PET also shows a correlation between worsening bradykinesia and increase of lentiform metabolism (Eidelberg et al., 1995). In fact, using FDG PET demonstrates a metabolic network characteristic of PD compared to other forms of parkinsonism (Hirano et al., 2009; Tang et al., 2010). While postsynaptic dopamine receptors have been thought to be preserved in PD, a recent study suggested a downregulation of the D3 receptors in drug-naive PD patients (Boileau et al., 2009).
Interconnected pathogenic mechanisms of PD
The best correlation of symptoms with progressive loss of striatal dopamine are those of bradykinesia and rigidity, which relate to striatal dopamine deficiency and loss of SNc dopaminergic neurons and can be correlated with a progressive decrease of dopaminergic imaging by PET or SPECT (see above). Since these symptoms are two of the cardinal features of the disease, research efforts have concentrated on the pathogenic mechanisms that cause loss of the nigrostriatal dopamine system, and this will be reflected in this review. Similar mechanisms might involve the other monoaminergic systems (noradrenergic and serotonergic). But there is little knowledge of the pathogenesis of neuronal loss for nonaminergic neurons. It seems likely that loss of these other monoamines might be instrumental in the high rate of depression (Mayeux et al., 1984) and anxiety in patients with PD. Little is known about the anatomical or biochemical associations for the other clinical features of the disease, including motoric features of tremor, freezing, flexed posture and loss of postural reflexes, and the multitude of nonmotor features.
A variety of pathogenic mechanisms have been uncovered for the loss of dopamine neurons, and probably more will be uncovered. The reader is referred to reviews on this topic for details (Dauer and Przedborski, 2003; Jenner and Olanow, 2006; Gupta et al., 2008). With the development of genetic causes of PD, a multiple hit hypothesis was proposed by Sulzer (2007), discussed later in this chapter. Evidence has accumulated over decades from pathologic and biochemical findings that implicate oxidative stress, mitochondrial dysfunction, excitotoxicity, inflammation, and apoptosis as taking place in the SNc (Fig. 5.10). More recently protein aggregation in the form of Lewy bodies and Lewy neurites has placed a major emphasis on the accumulation of toxic protein as perhaps the most important pathogenic factor. Each of these factors cross-interact with the others to add to the pathogenesis of cell death. Toxic proteins accumulate because of insufficient degradation or an excess synthesis that the normal degradation process cannot handle.
Oxidative stress
A key source of oxidative stress in monoaminergic neurons is via monoamine metabolism and auto-oxidation (discussed in more detail below). Since most research in PD neurodegeneration has been applied to dopaminergic neurons, we will focus on dopamine. Antioxidant defenses protect cells, and one of the leading antioxidants is reduced glutathione (GSH), and this is reduced in the SNc of PD patients at postmortem (Perry et al., 1982; Sian et al., 1994). This reduction of GSH is specific to PD brains, and is not seen in atypical parkinsonisms with nigral degeneration. The reduction of GSH likely reflects an excess utilization of this reducing agent, implying a high degree of oxidative stress taking place there. The decrease in GSH in the SNc occurs in incidental Lewy bodies as well as in PD, suggesting that oxidative stress has occurred prior to nerve cell loss. Some evidence suggests that GSH depletion itself may play an active role in PD pathogenesis (Martin and Teismann, 2009). Iron accumulation in the nigra also contributes to oxidative stress (Dexter et al., 1989a). Oxidized products of lipids, DNA, and protein are also seen in the PD nigra (Dexter et al.,1989b; Sanchez-Ramos et al., 1994), providing postmortem evidence of oxidative stress. The formation of neuromelanin in dopamine neurons is derived from the condensation of oxidized dopamine products, and thus represents a protective mechanism by the cell to defend itself against oxidative stress (Sulzer et al., 2000). One of the mutant genes that can cause PD, DJ-1, functions normally to protect against oxidative stress and is discussed below.
It is of interest that an endogenous substance, uric acid, which has antioxidant properties, has been correlated with a reduction in developing PD (Weisskopf et al., 2007). Subsequent studies evaluating urate levels in plasma in the PRECEPT (Schwarzschild et al., 2008) and CSF in the DATATOP (Ascherio et al., 2009) clinical trials found that higher urate levels in men were associated with a slower rate of progression of PD.
Mitochondrial dysfunction
Mitochondria appear to play an important role in the pathogenesis of PD (Banerjee et al., 2009). The finding that 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) intoxication causes parkinsonism (Langston et al., 1983), and the discovery that MPTP selectively destroys dopamine neurons and impairs complex I activity in the mitochondria (Nicklas et al., 1987) led to the study of mitochondria in PD patients. Decreased complex I activity is seen in the SNc of PD brains and not elsewhere, nor in other forms of parkinsonism (Mizuno et al., 1989; Schapira et al., 1990). Another complex I toxin, rotenone, which is a commonly used pesticide, also damages SNc neurons in animals (Betarbet et al., 2000). We will see below that two genes related to PD, namely parkin and PINK1, help maintain the integrity and function of the mitochondria, and loss of function of these genes results in PD. Mitochondrial dysfunction impairs ATP production, which hinders energy-dependent mechanisms of degradation of misfolded proteins by the ubiquitin-proteasomal system. Mitochondrial dysfunction is both a cause and a consequence of oxidative stress. Deregulation of mitochondrial respiration leads to generation of reactive oxygen species, contributing to oxidative stress, while oxidative and nitrosative stress deteriorates mitochondrial function. An early event in MPTP toxicity is oxidative stress, which is the consequence of an inability to transport electrons due to the inhibition of mitochondrial complex I. The accumulating electrons are a source of oxidative stress (Zhou et al., 2008). Besides their role in electron transport and oxidative phosphorylation, mitochondria are a major cellular source of free radicals, affect calcium homeostasis, and instigate cell-death pathways via apoptosis (Henchcliffe and Beal, 2008; Schapira, 2008).
Other mechanisms
Excitotoxicity from excessive glutamatergic activity results in an increase in calcium and can damage mitochondria; this has been implicated in PD (Beal, 1998). Nitrosative stress is induced by nitric oxide forming peroxynitrite, leading to protein nitration, and has also been suggested as a pathogenic factor (Tsang and Chung, 2009). Inflammation is seen in PD SN (McGeer et al., 1988a, 1988b), but is not considered an early event (Zhou et al., 2008). Rather, inflammation appears to augment the continuing degeneration (Tansey et al., 2007; Hirsch and Hunot, 2009; Saijo et al., 2009; Przedborski, 2010). Apoptosis is thought to represent the cellular death mechanism in PD (Mochizuki et al., 1996; Hirsch et al., 1999).
Accumulation of toxic proteins
The accumulation of too much protein in the cell has invoked the concept that these proteins are toxic and can lead to cell death. The concept of accumulated unwanted protein derived from genetic studies with the discovery that the protein α-synuclein is present in Lewy bodies (discussed below). Proteins are commonly damaged by misfolding or some other alteration, so that they are not functioning normally, and the cell attempts to repair them or eliminate them through degradation by either the ubiquitin-proteasomal system or by autophagy via the lysosome. Thus, misfolded, damaged, or altered proteins are either repaired or else removed from the cell, otherwise they would accumulate and act as toxins that would damage the cell. The first step to eliminate such proteins is to try to repair them through chaperone-mediated mechanisms. If this fails, the ubiquitin-proteasomal system attempts to remove the unwanted protein. If this is not capable of removing the protein, then autophagy via the lysosome takes place (Kopito, 2000; Pan et al., 2008). Autophagy appears to play a major role in removing α-synuclein and other unwanted proteins. This mechanism is described later in this chapter.
Genetics as an etiologic factor of PD
Familial PD
Families with PD occurring in several members have been recognized over the years, with approximately 10% of newly diagnosed patients reporting someone else in the family having PD. One of the earliest to describe a family with multiple affected members was Allen (1937). Perhaps the largest pedigree of PD described to date and with many generations affected was that reported by Mjönes (1949) of a Swedish and Swedish-American family. He found an autosomal dominant inheritance pattern in this family with 60% penetrance. But generally, since most PD patients are sporadic without a positive family history, the disease was not thought to have a genetic etiology.
Twin studies
To study the possibility of a genetic cause of PD though, Duvoisin, Eldridge and their colleagues (1981) evaluated twins with PD. They found zero concordance in 12 monozygotic twin pairs and concluded that “genetic factors appear not to play a major role in the etiology of PD.” This group (Ward et al., 1983) continued to analyze twin pairs and now in 43 monozygotic (MZ) and 19 dizygotic (DZ) pairs with the index case having definite PD, the frequency of PD in MZ twins was similar to that expected in an unrelated control group matched for age and sex. The authors again concluded that “the major factors in the etiology of PD are nongenetic.” A Finnish twin study 5 years later (Martilla et al., 1988) also found low concordance in MZ twin pairs and also concluded that PD appears to be “an acquired disease not caused by a hereditary process.” However, Bill Johnson, then at Columbia University, began to question the conclusions of the twin studies (Johnson et al., 1990), saying they were too small to be statistically conclusive and recommended that linkage studies be conducted.
The momentum toward a genetic etiology of PD
In 1990 Golbe and Duvoisin and their colleagues described a large kindred with autosomal dominant PD originating in Contoursi, Italy, with some of the family having emigrated to the US. This led Duvoisin to rethink his previous conclusions that PD is largely nongenetic (Duvoisin and Johnson, 1992). At this time there were also reports that FDOPA PET scans can detect decreased FDOPA uptake in some nonaffected relatives, including twins with an affected co-twin (Brooks, 1991). A large PET study in twins indicated that the concordance for decreased striatal FDOPA uptake in PD twins is greater than previously realized (Burn et al., 1992). With the availability of the Contoursi kindred and with the tools of linkage analysis, the stage was set for finding a gene mutation in familial PD.
Finding the first gene mutation in PD
Linkage analysis was carried out on the Contoursi kindred, and after several years of searching, linkage to chromosome 4q21–q23 was found (Polymeropoulos et al., 1996). By the next year, Polymeropoulos and colleagues (1997) identified the mutated gene, SNCA, for the protein α-synuclein. The Contoursi family actually originated in Greece and immigrated to Italy. The mutation (Ala53Thr) was also found in three small, unrelated Greek families. Subsequently, two other mutations were found in SNCA, which also caused autosomal dominant PD, namely Ala30Pro in a German family (Krüger et al., 1998) and Glu46Lys in a Spanish family (Zarranz et al., 2004). Families with SNCA mutations have a younger age at onset (usually in their forties) and a more rapidly worsening course of PD, and also with some early cognitive impairment. So, the gene mutation, on average, causes a more severe form of the disease than the typical adult-onset sporadic case. The SNCA gene was originally labeled PARK1.
Although all these SNCA mutations are rare in causing all worldwide cases of both familial and sporadic PD, the protein α-synuclein has taken on a premier and most important role as being highly likely involved in the disease’s pathogenesis, including the sporadic cases. Immediately after the gene mutation was discovered (Polymeropoulos et al., 1997), Spillantini and colleagues (1997) found that fibrillary α-synuclein is a major component of the Lewy body, a finding confirmed the following year (Mezey et al., 1998). In fact, staining microscopic brain slices with the antibody for α-synuclein has become the pathologist’s tool to detect Lewy bodies, and probably accounts for the recognition that diffuse Lewy body disease (DLBD), also known as dementia with Lewy bodies (DLB), is the second most common cause of dementia after Alzheimer disease. Prior to staining for α-synuclein, ubiquitin antibodies were used because ubiquitin is another protein found within the Lewy body. Ubiquitin is located in the central core and α-synuclein in the surrounding halo (Fig. 5.11). Autopsies on patients with mutated or excess amounts of α-synuclein show an abundance of Lewy bodies (Seidel et al., 2010).
Other gene mutations in PD
After the SNCA gene mutation was found, other investigators began collecting families with PD and conducting linkage analyses on them and then cloning the gene when linkage was obtained. Japanese investigators discovered the second gene to cause PD, which they called parkin and designated as PARK2, with the protein also called parkin (Kitada et al., 1998). The gene mutation was initially reported in a family with autosomal recessive juvenile parkinsonism without Lewy bodies that had been linked to chromosome 6q25.2–q27 (Matsumine et al., 1998). Discovery was aided because many patients had deletions of large sections of chromosome 6, where the gene resides. Parkin mutations have now been identified pan-ethnically and are thought to be the cause of approximately 50% of familial young-onset PD and 15–20% of sporadic young-onset PD (<50 years). Over 70 mutations, including exon rearrangements, point mutations, and deletions, have been identified, many of them recurrent in different populations. The identification of parkin mutations is inversely correlated with age of onset, with the earliest age of onset having the greatest association. However, PARK2 does not appear to be restricted to young-onset PD, and parkin mutations have been identified in individuals over 50 (Lücking et al., 2000). Juvenile parkin-related PD is associated with mutations in both parkin alleles (homozygotes or compound heterozygotes). Some heterozygotes have now been recognized as having adult-onset PD with Lewy bodies (Farrer et al., 2001; Pramstaller et al., 2005). A single parkin mutation (heterozygote) may be responsible for instances of later onset PD (Foroud et al., 2003), and numerous heterozygote cases have now been found (Pramstaller et al., 2005). This is a crucial finding in the search for the cause of idiopathic PD, and it is clear that parkin plays an important role. The frequency and penetrance of parkin mutations have yet to be determined. PARK2, especially in juvenile cases, has been characterized by slow clinical progression, sustained response to levodopa, high likelihood of levodopa-induced dyskinesias, dystonia, sleep benefit, and hyperreflexia. The rate of loss of FDOPA striatal uptake is much slower in symptomatic and asymptomatic parkin carriers, compatible with the slow clinical worsening in this form of PD (Pavese et al., 2009).
The locus for PARK3 has been localized by linkage to chromosome 2p13 in a small group of European families with autosomal dominant PD with incomplete penetrance (Gasser et al., 1998). Affected individuals have clinical and pathologic findings similar to sporadic late-onset PD, including age of onset. The sepiapterin reductase gene is a likely candidate for PARK3 (Karamohamed et al., 2003; Sharma et al., 2006). Sepiapterin reductase is an enzyme in the pathway for tetrahydrobiopterin synthesis, a cofactor for tyrosine hydroxylase. At present PARK3 is considered a susceptibility gene.
The evolution of the gene identification for PARK4 is a fascinating story. Waters and Miller (1994) described a kindred with autosomal dominant Lewy body parkinsonism in four generations, with an early age at onset (PARK4). The kindred was extended (Muenter et al., 1998; Farrer et al., 1999; Gwinn-Hardy et al., 2000), and action tremor similar to essential tremor and dementia were prominent within the family, and widespread Lewy bodies were seen in the cerebral cortex. Linkage was reported to chromosome 4p, but this haplotype also occurred in individuals in the pedigree who did not have clinical Lewy body parkinsonism but rather suffered from postural tremor only (Farrer et al., 1999). A second genome-wide search in this family found linkage on chromosome 4q and the mutation to be a triplication of a region of the chromosome that includes the SNCA gene (Singleton et al., 2003), the same gene that is mutated in PARK1. Instead of a mutation, though, this triplication produces a doubling of the normal, wild-type α-synuclein protein. A second PD family (this one being Swedish-American) with PD dementia with triplication of SNCA has also been reported (Farrer et al., 2004). The large Swedish family reported originally by Mjönes (1949) has been genetically analyzed and found to consist of duplications and triplications of normal SNCA (Fuchs et al., 2007). The remarkable thing about it is that the family has an α-synuclein duplication in one branch, and a triplication in the other, with a correspondingly different clinical picture (late versus early onset, respectively).
PARK5, an autosomal dominant mutation of the gene for ubiquitin-carboxy-terminal-hydrolase L1 (UCHL1), was found in a single family and was mapped to chromosome 4p14 (Leroy et al., 1998). Additional families with this mutation are necessary before concluding that UCHL1 is a definite causative gene. But since this enzyme is part of the ubiquitin-proteasomal system, it implicates this degradative pathway as being important in the pathogenesis of PD (see below). Another potential pathogenic mechanism is the association of UCH-L1 with cellular membranes, including the endoplasmic reticulum (Liu et al., 2009). The membrane-associated form of UCH-L1 has been correlated with the intracellular level of α-synuclein.
Families with autosomal recessive, young-onset PD in southern Europe were mapped to chromosome 1p35–p36, labeled as PARK6 (Valente et al., 2002), and Valente and her colleagues (2004) identified two homozygous mutations in the PTEN-induced putative kinase 1 (PINK1) gene. Novel homozygous mutations in this gene were subsequently found in six unrelated Asian families (Hatano et al., 2004). Other families around the globe have been reported, and heterozygosity appears to increase susceptibility to developing PD (Hedrich et al., 2006). PINK1 is a mitochondrial serine/threonine kinase, and appears important in combating oxidative stress. The patients with PINK1 mutations resemble those with parkin mutations. Excessive parkin can rescue the PINK1 mutant phenotype, indicating that parkin works downstream from PINK1 in a common pathway. One autopsy of a young-onset patient with PD due to PINK1 mutations showed the presence of Lewy bodies (Samaranch et al., 2010).
PARK7, the DJ-1 gene mutation, is another cause of early-onset, autosomal recessive PD and is due to mutations on chromosome 1p36 (Bonifati et al., 2003). The mutations and gene identification were found in a consanguineous Dutch family and families in Italy and Uruguay. PARK7, like parkin and PINK1, is characterized by slow progression and a good response to levodopa. No autopsies have been reported for individuals with DJ-1. The crystal structure of DJ-1 reveals a redox-reactive center. DJ-1 may be activated in the presence of reactive oxygen species under conditions of oxidative stress. DJ-1, therefore, is an important redox-reactive signaling intermediate controlling oxidative stress (Kahle et al., 2009). DJ-1 and PINK1 are both important for mitochondrial function (Cookson, 2010). Knock-out mutants of DJ-1 enhance mitochondrial oxidative stress produced by pacemaking of SNc neurons (Guzman et al., 2010).
The most common PD gene mutation found so far has been PARK8, the gene located on chromosome 12p11.2–q13.1 for the protein leucine-rich repeat kinase 2 (LRRK2) (Zimprich et al., 2004), which is also called dardarin (Paisan-Ruiz et al., 2004). Initially, linkage to chromosome 12 was reported in a large Japanese autosomal dominant family without Lewy bodies (Funayama et al., 2002). But the Zimprich and Paisan-Ruiz families included subjects with Lewy body pathology, tangle pathology, and without Lewy bodies. One LRRK2 mutation, G2019S, is the most common and is seen in more than 2% in sporadic North American and English PD patients (Gilks et al., 2005; Nichols et al., 2005). The highest frequencies are in the Portuguese (Bras et al., 2005), Ashkenazi Jewish (Clark et al., 2006; Ozelius et al., 2006), and North African Arab (Lesage et al., 2006) patients, even in the absence of a family history of PD. The clinical, pathologic, and genetic epidemiology of LRRK2 were reviewed by Haugarvoll and Wszolek (2006) and mixed pathology resembling typical PD, tauopathies (tangles), and multiple system atrophy (MSA) were seen. However, patients with typical MSA were not found to have the LRRK2 G2019S mutation (Ozelius et al., 2007). In the Chinese population the G2019S mutation is rare, but about 9% of PD patients have a heterozygous G2385R mutation (Fung et al., 2006; Farrer et al., 2007). LRRK2 mutations show the typical age at onset and are not prone to be young onset (Clark et al., 2006). Penetrance is not complete and variable rates have been reported. Penetrance increases with age. In one report, it increases from 28% at age 59 years, to 51% at 69 years, and 74% at 79 years (Healy et al., 2008), and in another report the penetrance rate was 24% at the age of 80 (Clark et al., 2006).
How mutations in LRRK2 cause PD is not clear. Increased kinase activity appears to be involved. Enhanced kinase activity (approximately three-fold) has been consistently reported for G2019S; but four disease mutants, R1441C, R1441G, Y1699C, and I2020T, have kinase activity less than two-fold of the wild-type protein (Dauer and Ho, 2010). Inhibitors of LRRK2 kinase have been found to be protective in both in vitro and in vivo models of LRRK2-induced neurodegeneration. These results establish that LRRK2-induced degeneration of neurons in vivo is kinase-dependent (Lee et al., 2010).
PARK9 is actually a Parkinson-plus disorder rather than PD. The autosomal recessive disorder has a mutation for the A form of parkinsonism-plus seen with a mutation of lysosomal type 5 ATPase, ATP13A2, on chromosome 1q36, known as PARK9. There is juvenile onset, and the disorder was originally known as the Kufor-Rakeb syndrome, after the village in Jordan where the affected families reside (Najim al-Din et al., 1994). This syndrome had features of Parkinson-plus with pyramidal signs, dystonia, and supranuclear gaze palsy. The disorder is levodopa-responsive, and some cases show putaminal and caudate iron accumulation, making it another neurodegeneration with brain iron accumulation (NBIA) (Schneider et al., 2010).
A genome-wide screen on 117 Icelandic patients with classic late-onset Parkinson disease and 168 of their unaffected relatives from 51 families resulted in linkage to chromosome 1p32 (Hicks et al., 2002). This susceptibility gene for PD was designated as PARK10.
A genome-wide screen on 150 families with PD resulted in finding linkage for a susceptibility gene on chromosome 2q36–q37, designated as PARK11 (Pankratz et al., 2003a). Mutations in the gene coding for GRB10-interacting GYF protein 2 (GIGYF2) was reported (Lautier et al., 2008), but this has been questioned by other investigators (Guo et al., 2009; Nichols et al., 2009).
A genome-wide search also found a positive LOD score for a locus on chromosome Xq21–q25 (Pankratz et al., 2003b), which has been designated as PARK12.
Using a candidate gene approach based on the phenotype of motor neuron degeneration in mice, a genome search on German PD patients found a novel mutation of a mitochondrial serine protease HTRA2 (high temperature requirement protein A2) in four patients (Strauss et al., 2005; Plun-Favreau et al., 2008). This mutation has been listed as PARK13.
Homozygosity for a missense mutation in the PLA2G6 gene was discovered in a large consanguineous Pakistani family with neurodegeneration with brain iron accumulation (NBIA2), a condition known as infantile neuroaxonal dystrophy (Morgan et al., 2006). Spasticity, cerebellar ataxia, and optic atrophy are common in these infants (Kurian et al., 2008). But the phenotypic spectrum has been found to be much broader. It can cause autosomal recessive dystonia–parkinsonism at all ages, including adults, and without evidence of brain iron accumulation (Paisan-Ruiz et al., 2009). It has been labeled PARK14.
Parkinsonism with spasticity occurring in the first decade of life has been called pallido-pyramidal syndrome (Davison, 1954). It is a recessive disorder that responds well to levodopa therapy (Horowitz and Greenberg, 1975; Bronstein and Vickrey, 1996). It has also been called parkinsonism-pyramidal syndrome, and the mutation was found in the FBXO7 gene coding for the F-box only protein 7 (Di Fonzo et al., 2009). It has been labeled PARK15.
PARK16 was discovered by a large genome-wide association study in the Japanese population on chromosome 1q32 (Satake et al., 2009). This finding was replicated in a large genome-wide association study in the European population (Simón-Sánchez et al., 2009) as did another replication study, which showed an association in ethnic Chinese but no association in caucasians of European ancestry (Vilariño-Güell et al., 2010). The nature of the abnormal gene is unknown.
A mutation in the EIF4G1 gene on chromosome 3q27 was recently found in European and American families, all with the same ancestral haplotype (Chartier-Harlin et al., 2009).
Although not designated with a PARK label, heterozygotic mutations of the lysosomal enzyme, β-glucocerebrosidase (GBA) gene on chromosome 1q21 (homozygotes are the cause of Gaucher disease) have been associated with PD (Goker-Alpan et al., 2004; Aharon-Peretz et al., 2004, 2005; Clark et al., 2005) and in other Lewy body disorders (Goker-Alpan et al., 2006). Mutations of GBA are most common in the Ashkenazi Jewish population. Between 17–30% of the Ashkenazi sporadic PD patients are heterozygotes for the GBA mutation (Aharon-Peretz et al., 2005; Clark et al., 2007). The GBA mutation is associated with onset younger than 50 years compared to the older age onset in sporadic PD (Clark et al., 2007). Heterozygous mutations have been found in patients with PD in all ethnic groups, and has been designated a risk factor for PD. Being a lysosomal enzyme, it implies a link between autophagy malfunction and PD.
Evaluating eight mutations of GBA among 420 patients with PD and 4138 controls of Israeli Ashkenazi origin, the relative frequency of mutation carriers was 18% versus 4% and severe mutations were associated with 13-fold increased risk of PD compared to two-fold risk increase in subjects with mild mutations (Gan-Or et al., 2008). In addition, 14% of the studied patients had the G2019S LRRK2 mutation. Thus more than a third of all PD Ashkenazi Jewish patients carried the GBA or the LRRK2 mutation. In one study involving a Japanese population of 534 patients with PD and 544 controls, 55 patients with PD (9.4%) and only 2 controls (0.37%), odds ratio (OR) 28.0, had at least one of 11 pathogenic heterozygous variants in the GBA gene (Mitsui et al., 2009). In another study, involving 790 British (non-Jewish) PD patients and 257 controls, GBA mutations were found in 33 (4.18%) of patients and 3 (1.17%) of controls (OR 3.7) (Neumann et al., 2009). The GBA-positive patients were younger at onset and 45% had visual hallucinations and 48% had cognitive decline or dementia. Diffuse neocortical Lewy body pathology occurred more frequently in the 17 GBA-positive patients examined at autopsy, than in controls. A large worldwide 16-center collaborative project involving 5691 patients with PD and 4898 controls revealed an OR of 5.43 for any GBA mutation in patients versus controls (Sidransky et al., 2009).
Compared to the general population, parkinsonism in patients with Gaucher disease is more frequent, occurs at an earlier age, responds less well to levodopa, and is more frequently associated with cortical dysfunction (Kraoua et al., 2009). Enzyme replacement therapy and substrate reduction therapy were ineffective. The independently increased risk of GBA mutations in patients with PD, together with the identification of mutations in a lysosomal type 5 P-type ATPase gene responsible for the PARK9 locus, suggests the link between neurodegeneration producing the PD phenotype and impaired autophagic lysosomal mechanisms (Pan et al., 2008). For those with Gaucher disease 5–7% develop PD by age 70, and 9–12% before age 80 years (Rosenbloom et al., 2011).
Families with compound heterozygosity in the DNA polymerase gamma 1 (POLG1) were discovered to have early onset PD in addition to a polyneuropathy (Davidson et al., 2006; Orsucci et al., 2011). This enzyme is involved in the replication of mitochondrial DNA, indicating that impaired mitochondrial function can lead to PD.
Perry syndrome is a rapidly progressive disease with familial parkinsonism, hypoventilation, depression, and weight loss (Perry et al., 1975; Wider and Wszolek, 2008
