REVIEW ARTICLE published: 06 July 2012 MOLECULAR NEUROSCIENCE doi: 10.3389/fnmol.2012.00079 Cystatin C in Alzheimer’s disease

Gurjinder Kaur and Efrat Levy*

Departments of Psychiatry, Biochemistry, and Molecular Pharmacology, Center for Dementia Research, Nathan S. Kline Institute, New York University School of Medicine, Orangeburg, NY, USA

Edited by: Changes in expression and secretion levels of cystatin C (CysC) in the brain in various Eva Zerovnik, Jozef Stefan Institute, neurological disorders and in animal models of neurodegeneration underscore a role for Slovenia CysC in these conditions. A polymorphism in the CysC gene (CST3)islinkedtoincreased Reviewed by: risk for Alzheimer’s disease (AD). AD pathology is characterized by deposition of oligomeric Eva Zerovnik, Jozef Stefan Institute, β β Slovenia and fibrillar forms of amyloid (A ) in the neuropil and cerebral vessel walls, neurofibrillary Ruben Vidal, Indiana University tangles composed mainly of hyperphosphorylated tau, and neurodegeneration. The School of Medicine, USA implication of CysC in AD was initially suggested by its co-localization with Aβ in Natasha Kopitar-Jerala, Jozef Stefan amyloid-laden vascular walls, and in senile plaque cores of amyloid in the brains of patients Institute, Slovenia with AD, Down’s syndrome, hereditary cerebral hemorrhage with amyloidosis, Dutch type *Correspondence: β Efrat Levy, Departments of (HCHWA-D), and cerebral infarction. CysC also co-localizes with A amyloid deposits Psychiatry, Biochemistry, and in the brains of non-demented aged individuals. Multiple lines of research show that Molecular Pharmacology, Center for CysC plays protective roles in AD. In vitro studies have shown that CysC binds Aβ and Dementia Research, Nathan S. Kline inhibits Aβ oligomerization and fibril formation. In vivo results from the brains and plasma Institute, New York University School of Medicine, 140 Old of Aβ-depositing transgenic mice confirmed the association of CysC with the soluble, Orangeburg Road, Orangeburg, non-pathological form of Aβ and the inhibition of Aβ plaques formation. The association of 10962 NY, USA. CysC with Aβ was also found in brain and in cerebrospinal fluid (CSF) from AD patients and e-mail: [email protected] non-demented control individuals. Moreover, in vitro results showed that CysC protects neuronal cells from a variety of insults that may cause cell death, including cell death induced by oligomeric and fibrillar Aβ. These data suggest that the reduced levels of CysC manifested in AD contribute to increased neuronal vulnerability and impaired neuronal ability to prevent neurodegeneration. This review elaborates on the neuroprotective roles of CysC in AD and the clinical relevance of this protein as a therapeutic agent.

Keywords: cystatin C, Alzheimer’s disease, cerebral amyloidosis, amyloid, Aβ, neurodegeneration

INTRODUCTION for a variety of diseases, disease progression, and the effect of CysC also known as γ-trace, is a basic protein (Hochwald et al., therapy. 1967), originally identified in human CSF and subsequently, also CysC is also implicated in the processes of neuronal degenera- found in all other mammalian body fluids and tissues (Bobek tion and repair of the nervous system (reviewed in Gauthier et al., and Levine, 1992; Turk et al., 2008). CysC is highly abundant 2011). CysC was originally identified as an inhibitor of cysteine in brain tissue (Hakansson et al., 1996), expressed by neurons, proteases such as cathepsins required for housekeeping function astrocytes, and microglial cells in the brains of different species during protein turnover (Turk et al., 2000). Imbalance between (Yasuhara et al., 1993; Palm et al., 1995; Miyake et al., 1996). CysC active proteases and their endogenous inhibitors may lead to plays a variety of biological roles, ranging from anti-viral and uncontrolled proteolysis, which has been associated with different anti-bacterial properties (Bobek and Levine, 1992), bone resorp- neurological diseases (Nakamura et al., 1991). The involvement of tion (Lerner and Grubb, 1992), tumor metastasis (Huh et al., proteases and their inhibitors in the processes of neuronal degen- 1999; Taupin et al., 2000), modulation of inflammatory responses eration and repair of the nervous system is reviewed in (Tizon and (Warfel et al., 1987; Bobek and Levine, 1992), cell proliferation Levy, 2006). and growth (Sun, 1989; Tavera et al., 1992), and astrocytic differ- The majority of CysC in the CSF is produced by the choroid entiation during mouse brain development (Kumada et al., 2004). plexus (Tu et al., 1992). CysC CSF levels in normal brain were Involvement of CysC has been shown in various diseases rang- found to be five times higher than plasma levels suggesting a ing from cancer to neurodegenerative disorders. Multiple studies potential physiological role of CysC in the brain (Grubb, 1992). have demonstrated changes in CysC concentrations in serum AlterationsinCysClevelsintheCSFinneurodegenerativedis- associated with a variety of conditions, such as chronic kidney eases have been documented (Pasinetti et al., 2006; Mares et al., disease, urinary infection, cancer, hypertension, cardiovascular 2009; Mori et al., 2009; Tsuji-Akimoto et al., 2009; Yang et al., disease, rheumatoid arthritis, glucocorticoid treatment, thyroid 2009; Maetzler et al., 2010). For example, CysC has shown a function, and aging (reviewed in Filler et al., 2005). CysC concen- great diagnostic potential as a biomarker for Amyotropic Lateral tration in specific tissues and body fluids can serve as a marker Sclerosis (ALS), a fatal neuromuscular disease characterized by

Frontiers in Molecular Neuroscience www.frontiersin.org July 2012 | Volume 5 | Article 79 | 1 Kaur and Levy Neuroprotection by cystatin C

progressive motor neuron degeneration (Wilson et al., 2010). walls, and in senile plaque cores of amyloid in brains of patients CysC levels in the CSF of ALS patients were significantly reduced with AD, Down’s syndrome, HCHWA-D, intracranial hemor- as compared to healthy controls (Pasinetti et al., 2006; Tsuji- rhage, cerebral infarction, and of elderly subjects without any Akimoto et al., 2009; Wilson et al., 2010). In addition, the neurological disorder (Maruyama et al., 1990; Vinters et al., 1990; direction of the longitudinal change in CSF CysC levels corre- Itoh et al., 1993; Haan et al., 1994; Levy et al., 2001). Abundant lated with the rate of ALS disease progression, and initial CSF cystatin A (CysA) and cystatin B (CysB), also called stefin B, were CysC levels were predictive of patient survival, suggesting that demonstrated in senile plaques in the brain of AD patients (Ii CysC may function as a surrogate marker of disease progres- et al., 1993; Bernstein et al., 1994). sion and survival (Wilson et al., 2010). CysC is also linked to The deposition of fibrillar protein aggregates in the walls of ALS histopathologically, as it is one of only two known proteins arteries, arterioles, and sometimes capillaries and veins of the that localize to Bunina bodies, small intraneuronal inclusions central nervous system is known as cerebral amyloid angiopa- contained in degenerating motor neurons, which are a specific thy (CAA) (Nagai et al., 2008). Hereditary cerebral hemorrhage neuropathologic feature of ALS (Okamoto et al., 2008). Similarly, with amyloidosis, Icelandic type (HCHWA-I) (Arnason, 1935; it was shown that CysC levels in the CSF of AD patients are lower Gudmundsson et al., 1972), also called hereditary cystatin C compared to non-demented individuals (Simonsen et al., 2007; amyloid angiopathy (HCCAA; Olafsson et al., 1996), is an auto- Hansson et al., 2009). Furthermore, specific neuronal cell pop- somal dominant form of CAA. Amyloid deposition in cerebral ulations in the brains of patients with AD showed an enhanced and spinal arteries and arterioles of HCHWA-I patients leads to CysC expression (Deng et al., 2001; Levy et al., 2001). Changes in recurrent hemorrhagic strokes causing serious brain damage and the levels of CysC in the CSF and brain in various neurodegen- eventually fatal stroke (Gudmundsson et al., 1972). The amy- erative diseases suggest important roles for the secreted protein loid deposited is composed mainly of a Leu68Gln variant of in these disorders. In support of such a role, CysC level dysregu- CysC (Cohen et al., 1983; Ghiso et al., 1986; Palsdottir et al., lation was also observed in animal models of neurodegenerative 1988; Levy et al., 1989; Abrahamson et al., 1990). A heterozy- conditions caused by facial nerve axotomy (Miyake et al., 1996), gous point mutation, identical to that found in the CST3 gene noxious input to the sensory spinal cord (Yang et al., 2001), of these patients, was also identified in a Croatian man with CAA perforant path transections (Ying et al., 2002), hypophysectomy and intracerebral hemorrhage (Graffagnino et al., 1995). Thus, (Katakai et al., 1997), transient forebrain ischemia (Palm et al., sporadic CAA in some patients may be associated with muta- 1995; Ishimaru et al., 1996), photothrombotic stroke (Pirttila tions in the CST3 gene (Graffagnino et al., 1995; McCarron et al., and Pitkanen, 2006), and induction of epilepsy (Aronica et al., 2000). 2001; Hendriksen et al., 2001; Lukasiuk et al., 2002). Augmented Amyloid β usually accumulates both in cerebral blood ves- CysC expression in the neurodegenerative states puts forward sels and in brain parenchyma as amyloid plaques. However, in two conflicting theories whether enhanced CysC expression is some cases Aβ deposits predominantly in the cerebral vascula- causing or further exacerbating already initiated neurodegenera- ture (Vinters, 2001). The factors leading to vascular rather than tive changes or, alternatively, it is an endogenous neuroprotective parenchymal amyloid deposition are unknown and it is unclear response to the disease (reviewed in Gauthier et al., 2011). This when CAA leads to hemorrhage. A role for CysC in CAA-related review focuses on the roles of CysC in the pathological processes hemorrhage is implicated from immunohistochemical studies of AD. that revealed co-localization of CysC and Aβ in amyloid-laden vascular walls (Maruyama et al., 1990; Vinters et al., 1990; Itoh CysC AND ALZHEIMER’S DISEASE et al., 1993; Haan et al., 1994). It was reported that only patients AD pathology is characterized by formation of amyloid deposits showing co-localization of CysC and Aβ immunoreactivity in in the brain composed mainly of Aβ, a processing product of the their diseased cerebral vessels suffered fatal subcortical hemor- amyloid β precursor protein (APP), aggregation of neurofibril- rhages (Maruyama et al., 1990). The degree of cerebrovascular lary tangles composed mainly of hyperphosphorylated tau, loss amyloid deposition in these patients was also greater than in of neurons with accelerated atrophy of specific brain areas and patients without cerebral hemorrhages. Studies were conducted decreased synapse number in surviving neurons. While it was to find out whether CysC exists as amyloid fibrils or as unpoly- demonstrated that fibrillar Aβ plays a central role in neurotoxi- merized CysC absorbed onto or trapped within the bundles of city in AD brains (for review see Butterfield and Boyd-Kimball, Aβ amyloid fibrils. ELISA analysis of crude amyloid fibrils iso- 2004), both in vitro and in vivo reports describe a potent neu- lated from cerebral blood vessels of one patient revealed that rotoxic activity for soluble, nonfibrillar, oligomeric assemblies of CysC and Aβ have been included at the ratio of about 1:100 Aβ (for reviews see Klein et al., 2001; Walsh and Selkoe, 2004). (Nagai et al., 1998). In another case of sporadic CAA, isola- In this section, we discuss the involvement of CysC in AD as tion and chemical analysis of amyloid fibril proteins from lep- suggested by immunohistochemical, genetic, and biochemical tomeningeal vessels revealed that while Aβ was fibrillar, CysC studies. was soluble (Maruyama et al., 1992). It has been suggested that CysC deposition occurs secondarily to Aβ deposition and may CysC CO-DEPOSITION WITH AMYLOID β increase the predisposition to cerebral hemorrhages (Itoh et al., The involvement of cystatins in AD was originally suggested due 1993). to their co-localization with amyloid plaques. CysC was the first CysC also co-localizes with Aβ deposits in the brains of ani- cystatin found co-localized with Aβ in amyloid-laden vascular mal models of cerebral amyloidosis. Co-localization of Aβ and

Frontiers in Molecular Neuroscience www.frontiersin.org July 2012 | Volume 5 | Article 79 | 2 Kaur and Levy Neuroprotection by cystatin C

CysC was demonstrated in vascular and parenchymal deposits in amyloidosis (familial amyloidosis, Finnish type; Kiuru et al., 1999; the brains of aged rhesus monkeys and in vascular amyloid in Kiuru-Enari et al., 2002) and familial CAA, British type (Ghiso brains of aged squirrel monkeys (Wei et al., 1996). Non-human et al., 1995). Thus, CysC may play a role in CAA and hemorrhage primates are good models to study cerebral changes that occur in a variety of diseases that involve deposition of heterogeneous through aging. Neuropathologies characteristic of AD and nor- types of amyloid proteins. mal aging in humans were also found in senescent non-human primates (Wisniewski and Terry, 1973; Walker et al., 1990; Price INTRACELLULAR CO-LOCALIZATION OF CysC WITH Aβ IN THE BRAIN et al., 1994). In aged rhesus monkeys (Macaca mulatta), amy- Immunohistochemical analyses have shown intense CysC loid deposition predominates in senile plaques with relatively immunoreactive neurons and activated glia in the cerebral minor vascular involvement. However, cerebrovascular deposits cortex of some aged human cases and of all AD patients in aged squirrel monkeys (Saimiri sciureus), usually are more (Yasuhara et al., 1993; Deng et al., 2001). High neuronal conspicuous than senile plaques (Walker et al., 1990). Sequence staining of CysC in AD brains was primarily limited to pyra- analysis of rhesus and squirrel monkey CysC cDNA revealed that midal neurons in cortical layers III and V (Deng et al., 2001; squirrel monkey has Met at position 68, which is Leu in the rhe- Levy et al., 2001). The regional distribution of CysC neuronal sus and wild-type human CysC and Gln in HCHWA-I patients immunostaining duplicated the pattern of neuronal suscep- (Wei et al., 1996). An additional difference between squirrel and tibility in AD brains: the strongest staining was found in the rhesus monkeys in CysC sequence was found at position 10, a entorhinal cortex, in the hippocampus, and in the temporal residue that was shown to affect the specificity of the inhibitor cortex; fewer pyramidal neurons were stained in the frontal, for different cysteine proteases (Lindahl et al., 1994). The species- parietal, and occipital lobes (Deng et al., 2001). Using an specific CysC sequences in humans, rhesus, and squirrel monkeys end-specific antibody to the carboxyl-terminus of Aβ42,intra- may be responsible for the variability of the amyloid deposits cellular immunoreactivity was observed in the same neuronal observed. subpopulation (Levy et al., 2001). These data showed that In order to elucidate the role of increased expression of this Aβ42 accumulates in a specific population of pyramidal neu- protein in vivo, CysC transgenic mice were generated (Pawlik rons in the brain, the same cell type in which CysC is highly et al., 2004). These mice express either human wild-type or the expressed. Leu68Gln variant CysC genes under the transcriptional con- trol of its own promoter (Levy et al., 1989), overexpressing the LOW CONCENTRATION OF CysC IN CSF AND PLASMA OF AD PATIENTS transgene along with its endogenous counterpart in the appro- Low levels of serum CysC precede clinically manifested AD in priate tissues (Pawlik et al., 2004). Lines of mice expressing elderly men free of dementia at baseline and may be a marker various levels of the transgene in the brain were selected. All of future risk of AD (Sundelof et al., 2008). Clinically diag- selected lines had very high concentrations of the transgene in nosed AD patients also showed a reduction in the CSF levels of the blood. None of the mice had amyloid deposits either in CysC compared to controls (Hansson et al., 2009). Interestingly, the vessel walls or in the neuropil. Neuropathological exami- CysC levels were positively correlated with both tau and Aβ42 nation of dead or ailing aged transgenic mice revealed some levels in the CSF, independent of age, gender, and apolipopro- mice with cerebral or subarachnoid hemorrhages (Pawlik et al., tein E (APOE) genotype (Sundelof et al., 2010). Therefore, it 2002). Conversely, no hemorrhages were observed in their non- was suggested to measure the change in CysC CSF levels as transgenic siblings. These data demonstrate that elevated brain a biomarker for AD diagnosis (Simonsen et al., 2007; Mares and/or blood levels of CysC can cause hemorrhagic strokes in et al., 2009; Zellner et al., 2009; Ndjole et al., 2010; Sundelof the absence of vascular amyloid deposits. It has been shown et al., 2010; Craig-Schapiro et al., 2011; Perrin et al., 2011). that the risk of cerebral hemorrhage in the brains of aged indi- Moreover, analysis of CysC levels in plasma revealed a signifi- viduals and AD patients increases when high levels of CysC cant tendency of conversion from mild cognitive impairment to are present in cerebrovascular Aβ deposits. These findings sug- dementia in subjects with CysC levels below the median (CysC gest that binding of CysC to Aβ in the vasculature, resulting in lower than 1067 ng/ml; Ghidoni et al., 2010). A large proportion local accumulation of the protease inhibitor, may contribute to of demented Lewy body disease patients have AD-like pathol- hemorrhages. ogy, in particular Aβ plaques. Demented Lewy body disease CysC also co-localizes with Aβ deposits in the parenchyma and patients also showed decreased CSF CysC levels (Maetzler et al., vasculature in brains of transgenic mice overexpressing human 2010). APP (Levy et al., 2001; Steinhoff et al., 2001). A striking increase in CAA with aging was found in the APP transgenic mouse LINKAGE OF CysC GENE POLYMORPHISM WITH AD line APP23 (Winkler et al., 2001). Antibodies to CysC revealed Studies were conducted to determine whether the association of appreciable staining of cerebrovascular amyloid in these mice. CysC with AD could be identified at the genetic level. The CST3 Similar to CysC staining of Aβ deposits in human brains, the CysC gene, localized on chromosome 20 (Abrahamson et al., 1989; immunoreactivity was restricted to a subpopulation of amyloid- Saitoh et al., 1989), has three genetically linked base substitu- laden vessels and was clearly less intense than Aβ staining (Levy tions in the 3 region (Balbin and Abrahamson, 1991; Balbin et al., 2001; Steinhoff et al., 2001; Winkler et al., 2001). et al., 1993). A G73A transition in exon 1 results in Ala/Thr Wild-type CysC co-localization with amyloid, other than Aβ, variation in the coding region of CST3, within the signal pep- was observed in a variety of disorders, such as hereditary gelsolin tide. While the allele containing Ala at that position was called

Frontiers in Molecular Neuroscience www.frontiersin.org July 2012 | Volume 5 | Article 79 | 3 Kaur and Levy Neuroprotection by cystatin C

theAallele,theonecontainingThrinthesamepositionwas 2010) showed that the BB genotype of the CST3 gene is associ- called B allele. Several studies have linked CST3 gene polymor- ated with reduced CSF CysC levels in patients with Lewy body phisms with an increased risk of developing AD (Crawford et al., disease with dementia. Furthermore, a study of the targeting 2000; Finckh et al., 2000; Beyer et al., 2001; Olson et al., 2002; of the Thr haplotype in cultured retinal pigment epithelial and Lin et al., 2003; Goddard et al., 2004; Cathcart et al., 2005; HeLa cells have shown that a proportion of the Thr protein Bertram et al., 2007) and a possible interaction with APOE undergoes incorrect trafficking. In contrast to the Ala haplo- genotype was noted. However, some studies have failed to show type that is targeted to the Golgi apparatus, the Thr variant was an association between CST3 and AD in a German cohort associated primarily with mitochondria, resulting in a substan- (Dodel et al., 2002), a Dutch sample with early onset AD (Roks tial reduction in the efficiency of targeting CysC for secretion et al., 2001), Japanese AD patients (Maruyama et al., 2001), a (Paraoan et al., 2004). A multicentric electroencephalographic Finnish population (Helisalmi et al., 2009), and in early onset (EEG) study analyzed the effects of CST3 haplotypes on rest- AD families (Parfitt et al., 1993). Other studies found a con- ing cortical rhythmicity in subjects with AD and mild cognitive nection between the CST3 polymorphism and AD in Caucasian impairment. A relationship between the CST3 Thr haplotype and populations, but not in Asian populations (Hua et al., 2012), global neurophysiological phenotype (i.e., cortical delta and alpha including Chinese (Wang et al., 2008). Over-all meta-analyses rhythmicity) was found. While APOE ε4 affects EEG rhythms using this polymorphism have reported CST3 as a susceptibility in AD (Lehtovirta et al., 1996, 2000; Jelic et al., 1997), the gene for AD. For update on the linkage of the CST3 polymor- effects of CST3 polymorphism were independent of APOE ε4 phism with AD, see the Alzgene Internet site of the Alzheimer co-presence (Babiloni et al., 2006). A decreased CysC secretion Research Forum (http://www.alzforum.org/res/com/gen/alzgene/ associated with a polymorphism found in the CysC gene, puts geneoverview.asp?geneid=66). forward a mechanism for the increased-risk of late-onset spo- Some groups have shown that the A allele of CST3 may be crit- radic AD conferred by this polymorphism and suggests that ical for the development of AD while others have held B allele reduced CysC brain concentration may be associated with the responsible for that. A multicentric AD population was geneti- disease. cally studied by age at onset and it was found that A allele of CST3 has an age related increased influence on onset of AD (Crawford ALTERED CysC TRAFFICKING AND FAMILIAL AD et al., 2000). A significant interaction between the homozygous Mutations in the presenilin 1 and presenilin 2 genes account AgenotypeofCST3 and age of onset of AD was found, such for the majority of familial AD (FAD) cases. Two of the muta- that in the over 80 years age group this genotype was responsi- tions in the presenilin 2 gene that are linked to FAD (PS2 M239I ble for a twofold increased risk for the disease. This interaction and T122R) alter CysC trafficking in mouse primary neurons was independent of the APOE genotype (Crawford et al., 2000). and cause reduced CysC secretion (Ghidoni et al., 2007). The Another study of large European and American populations, with primary structure of CysC is indicative of a secreted protein mean age at onset of 73.1 and 75.0 for AD and controls, respec- and accordingly, it was demonstrated that most of the CysC tively, showed linkage between the B allele and late onset AD is targeted extracellularly via the secretory pathway (Wei et al., with no synergistic association with APOE allele (Finckh et al., 1998; Paraoan et al., 2001). Moreover, it was recently shown 2000). A synergistic association between the CST3 and APOE that CysC is also secreted in association with exosomes (Ghidoni ε4 alleles was found in a Spanish sample. The CST3 B allele et al., 2011). Full-length APP and APP processing products, caused a threefold elevated risk of AD before age 70 and there including Aβ are also released in association with exosomes. was an eightfold increase in risk for APOE ε4 carriers with this Over-expression of presenilin 2 with the FAD-associated muta- allele (Beyer et al., 2001). In another genetic study the combi- tions (PS2 M239I and PS2 T122R) resulted in decreased levels nation of one or two CST3 B alleles and APOE ε4 carried a of CysC within exosomes (Ghidoni et al., 2011). Assuming a 14-fold increased risk for men and 16-fold for women. These protective role for CysC, as described below, the reduction in risks apply to a shift in risk from ages 65 and older to younger CysC levels may represent the molecular factor responsible for ages (Cathcart et al., 2005). The attempt to determine the associ- the increased risk of AD in FAD patients, the carriers of these ation between CST3 polymorphism and AD or vascular dementia mutations. resulted in the associations between CST3 BgenotypeandAD patients older than 75, or vascular dementia patients younger NEUROPROTECTION BY CysC IN AD than 75 years. A synergistic association of CST3 and APOE ε4 Intense CysC immunoreactivity was observed in specific neu- alleles was observed in predicting vascular dementia patients (Lin ronal populations in the cerebral cortex of some aged human et al., 2003). cases and of all AD patients (Yasuhara et al., 1993; Deng The amino acid exchange from Ala to Thr at the –2 position et al., 2001; Levy et al., 2001). Enhanced CysC expression in for signal peptide cleavage alters the hydrophobicity profile of specific cell populations is not limited to AD but was also the signal sequence (Finckh et al., 2000), resulting in a less effi- observed in other neurodegenerative conditions, such as epilepsy, cient cleavage of the signal peptide and thus a reduced secretion ischemia, and progressive myoclonus epilepsy (Palm et al., 1995; of CysC (Benussi et al., 2003). The CST3 gene G73A poly- Ishimaru et al., 1996; Aronica et al., 2001; Hendriksen et al., morphism functionally affects CysC plasma levels (Noto et al., 2001; Lukasiuk et al., 2002; Kaur et al., 2010). Contradictory 2005)andCysCCSFlevels(Maetzler et al., 2010; Yamamoto- conclusions were reached from multiple studies, suggesting Watanabe et al., 2010). Maetzler and colleagues (Maetzler et al., that increased CysC cellular expression in the brain is either

Frontiers in Molecular Neuroscience www.frontiersin.org July 2012 | Volume 5 | Article 79 | 4 Kaur and Levy Neuroprotection by cystatin C

associated with the neurodegenerative process, or alternatively CysC protected neuronal cells from death in a concentration is part of a neuroprotective response aimed at prevention of dependent manner. Moreover, primary cortical neurons isolated neurodegeneration. In this section of the review we provide from the brains of CysC overexpressing transgenic mice (Pawlik a description of different neuroprotective mechanisms acti- et al., 2004) were protected from spontaneous death induced by vated by CysC. We hypothesize that reduced secretion of CysC culturing and from B27-supplement-deprivation, and cells iso- into extracellular body fluids can hamper the ability of the lated from CysC knockout mice (Huh et al., 1999)weremore brain to prevent neurodegeneration in various pathological sensitive to in vitro toxicity compared to cells isolated from brains conditions. of wild-type mice (Tizon et al., 2010b). Using multiple methods, it was demonstrated that CysC induces autophagy in cells under Protection by inhibition of cysteine proteases basal conditions, and enhances the autophagic activation in cells In vitro experiments have shown that CysC inhibits cathep- exposed to nutritional deprivation and oxidative stress (Tizon sins B, H, K, L, and S (for review see Bernstein et al., et al., 2010b). The autophagic pathway consists of sequestration 1996) and is inactivated by proteolytic degradation by and turnover of organelles and cytoplasm in autophagic vacuoles cathepsin D (Abrahamson et al., 1991; Lenarcic et al., 1991). that following maturation fuse with lysosomes, leading to degra- Neuropathological observations suggest an association between dation of their content. CysC induces a fully functional autophagy CysC and cathepsins B and D in AD (Deng et al., 2001). via the mTOR pathway that includes competent proteolytic clear- Pyramidal neurons in layers III and V in the cortex of AD ance of autophagy substrates by lysosomes (Tizon et al., 2010b). patients have displayed a quantitative increase in cathepsin D Enhanced lysosomal turnover can protect against neurodegen- immunoreactivity (Cataldo et al., 1995), the same neuronal eration and CysC can serve to modulate the efficiency of the population that show increased CysC expression (Deng et al., autophagic pathway. It remains to be demonstrated that CysC 2001; Levy et al., 2001). Lysosomal cathepsins are involved in induces autophagy in vivo as a protective mechanism in brain neuronal cell death (Cataldo and Nixon, 1990). Intense cyto- injury and in neurodegenerative disorders, such as AD. plasmic labeling of cathepsin B was detected when neurons had become morphologically altered with obvious shrinkage of the Protection by inhibition of Aβ oligomerization and amyloid fibril cytoplasm (Hill et al., 1997). Enhanced expression of several formation cathepsins has been documented in response to injuries, similar The co-localization of CysC with Aβ in parenchymal and vascular to those inducing CysC expression upregulation, such as in amyloid deposits reflects the involvement of CysC in amyloidoge- transient ischemia (Nitatori et al., 1995; Yamashima et al., 1998), nesis, therefore, the association of CysC with Aβ was determined and inhibitors of cathepsins B and L reduce neuronal damage by Western blot analysis of immunoprecipitated cell lysate or in the hippocampus after ischemia (Tsuchiya et al., 1999). An medium proteins, revealing binding of CysC to full-length APP imbalance in the expression of cathepsins and their inhibitors and to secreted soluble APP (Sastre et al., 2004). Deletion mutants may cause or exacerbate existing neuropathological changes and of APP localized the CysC binding site to the Aβ region within increased localizes CysC expression may represent an attempt to APP. CysC association with APP resulted in increased secre- curb the cathepsin activity. tion of soluble APP but did not affect the levels of secreted Aβ The in vivo role of CysC as an inhibitor was observed by both in vitro (Sastre et al., 2004)andin vivo in transgenic mice deletion of CysC in knockout mice, resulting in an increased expressing the human CysC gene (Pawlik et al., 2004). The asso- cathepsin B activity (Sun et al., 2008). Another in vivo study ciation of CysC with APP was confirmed using a method for has demonstrated that CysC can mediate neuroprotection by the in vivo mapping of protein interactions in intact mouse tis- inhibition of cysteine proteases. A mouse model of an inherited sue (Bai et al., 2008). CysC does not bind only to Aβ sequences neurodegenerative disorder, the progressive myoclonic epilepsy, within APP, but also to the peptide itself (Sastre et al., 2004). has been generated by knocking-out the CysB gene (Pennacchio Analysis of the association of CysC and Aβ demonstrated a et al., 1998). CysB deficiency in these mice results in increased specific, saturable and high affinity binding between CysC and cathepsins activity (Kaur et al., 2010). CysC overexpression in both Aβ1−42 and Aβ1−40 (Sastre et al., 2004). Most importantly, CysB knockout mice decreased cathepsin B and D activities in CysC association with Aβ resulted in a concentration depen- the brain (Kaur et al., 2010). It was demonstrated that clinical dent inhibition of Aβ amyloid fibril formation (Sastre et al., symptoms and neuropathologies, including deficient motor coor- 2004). In vitro studies also demonstrated that CysC association dination, cerebellar atrophy, neuronal loss in the cerebellum and with Aβ inhibits Aβ oligomerization (Selenica et al., 2007; Tizon cerebral cortex, and gliosis caused by CysB deficiency, are res- et al., 2010a). A structural model of the human CysC/Aβ complex cued by CysC overexpression (Kaur et al., 2010). These data show using a combination of selective proteolytic excision and high- that CysC partially prevents neurodegeneration in CysB knockout resolution mass spectrometry identified a specific C-terminal mice through inhibition of cathepsins activity. epitope (residues 101–117) as the Aβ-binding region within CysC (Juszczyk et al., 2009). Protection by induction of autophagy The anti-amyloidogenic role of CysC was demonstrated in An in vitro study of the effect of CysC on cells of neuronal origin vivo in Aβ depositing APP transgenic mice overexpressing human under neurotoxic stimuli has shown that CysC protects neuronal CysC. Several lines of transgenic mice, expressing human CysC cells from death by a mechanism that does not require cathep- either under control sequences of the human CysC gene (Mi sin inhibition (Tizon et al., 2010b). Exogenously applied human et al., 2007), or specifically in cerebral neurons (Kaeser et al.,

Frontiers in Molecular Neuroscience www.frontiersin.org July 2012 | Volume 5 | Article 79 | 5 Kaur and Levy Neuroprotection by cystatin C

2007), were crossbred with mice overexpressing human APP. Protection by neurogenesis CysC bound to the soluble, non-pathological form of Aβ in the CysC can also regulate cell proliferation (Sun, 1989; Tavera et al., brains and plasma of these mice and inhibited the aggregation 1992). In rats undergoing acute hippocampal injury or sta- and deposition of Aβ plaques in the brain (Kaeser et al., 2007; Mi tus epilepticus-induced epileptogenesis, the expression of CysC et al., 2007). However, deletion of CysC in knockout mice resulted mRNA and protein are increased in the hippocampus and in in enhanced Aβ degradation (Sun et al., 2008). Unlike a complete the dentate gyrus (Aronica et al., 2001; Hendriksen et al., 2001; deletion of CysC, reduced or enhanced levels of CysC expression Lukasiuk et al., 2002). The time of increased CysC expression affect the aggregation of Aβ,notAβ levels (Kaeser et al., 2007; Mi was shown to be matching the time of prominent neurogenesis et al., 2007). (Parent et al., 1997; Nairismagi et al., 2004). Moreover, the basal Investigation of the binding between Aβ and CysC in level of neurogenesis in the subgranular layer of dentate gyrus was human central nervous system was conducted by co- decreased (Taupin et al., 2000; Pirttila et al., 2005)andthepro- immunoprecipitation of CysC and Aβ in brain and CSF liferation and migration of newborn granule cells in the dentate from AD patients and controls (Mi et al., 2009). Sequential gyruswereimpairedinCysCknockoutmice(Pirttila et al., 2005), centrifugation of brain homogenates was used to identify that supporting a role for CysC in neurogenesis. CysC was shown the cellular fraction contains Aβ/CysC complexes. While CysC to regulate glial development, as addition of human CysC into binding to soluble Aβ was observed in AD patients and con- the culture medium of primary brain cells increased the num- trols, an SDS-resistant, stable CysC/Aβ complex was detected ber of glial fibrillary acidic protein (GFAP)-positive and nestin- exclusively in brains of neuropathologically normal controls, positive cells, as well as the number of neurospheres formed from but not in AD cases. The association of CysC with Aβ in brain embryonic brain (Hasegawa et al., 2007). Thus, another mech- from control individuals and in CSF revealed an interaction of anism of neuroprotection by CysC might involve induction of these two polypeptides in their soluble form (Mi et al., 2009). neurogenesis. The association between Aβ and CysC was shown to prevent Aβ accumulation and fibrillogenesis in experimental systems, CONCLUSIONS arguing that CysC plays a protective role in the pathogenesis of Immunohistochemical, genetic, and biochemical studies suggest AD in humans and explains why a decrease in CysC concentra- the involvement of CysC in AD. Immunohistochemical studies tion caused by the CST3 polymorphism or by specific presenilin have shown that CysC co-localizes with Aβ in amyloid-laden vas- 2 mutations, can lead to the development of the disease. In cular walls, and in senile plaque cores of amyloid and that CysC addition to its anti-amyloidogenic property, CysC directly pro- and Aβ immunoreactivity co-localizes in a specific population tects neuronal cells from Aβ toxicity. The extracellular addition of pyramidal neurons that is vulnerable to neurodegeneration of human CysC together with preformed either oligomeric or in AD. Biochemical studies have shown binding of CysC to Aβ fibrillar Aβ to cultured primary hippocampal neurons and to a and that this binding prevents Aβ oligomerization, fibril for- neuronal cell line increased cell survival (Tizon et al., 2010a). The mation, and amyloid deposition. Genetic studies have shown data obtained show that subtle modifications in CysC expression linkage of a CysC gene polymorphism with AD, associated with levels in the central nervous system, or possibly in the periphery, decreased secretion of CysC. Two FAD-linked presenilin 2 gene affect amyloid deposition and protect from the toxicity of mutations alter CysC trafficking and cause reduced CysC secre- aggregated Aβ. tion. Moreover, low concentrations of CysC were measured in Aβ interacts not only with CysC (Sastre et al., 2004), but also CSF and plasma of AD patients. While some studies have shown with CysB (Skerget et al., 2010; Zerovnik et al., 2010). It was that CysC can be toxic to cells under certain conditions, there are shown that CysB binding to Aβ is oligomer specific and that reports showing that under other specific conditions, CysC can the dimers and tetramers of CysB inhibit Aβ fibril formation be protective. While in high concentrations CysC may be toxic (Skerget et al., 2010). Aβ interaction with amyloid proteins is to cells, low concentrations may not be sufficient to protect the not restricted to CysC, but include transthyretin (Schwarzman cells. We hypothesize that protection imparted by CysC is efficient et al., 1994, 2004; Choi et al., 2007; Buxbaum et al., 2008), gelsolin under a limited range of concentrations. Slightly increased CysC (Chauhan et al., 1999), α2-macroglobulin (Kuo et al., 2000), and expression activates multiple mechanisms of protection. These crystallin-αB(Wilhelmus et al., 2006). The interaction between mechanisms include inhibition of cysteine proteases, induction the amyloidogenic proteins and Aβ inhibits Aβ fibril formation of autophagy, induction of cell division, and prevention of amy- (Matsuoka et al., 2003; Sastre et al., 2004; Wilhelmus et al., 2006; loidogenesis. We hypothesize that in AD, a variety of these mech- Kaeser et al., 2007; Mi et al., 2007; Skerget et al., 2010). Wilhelmus anisms are activated. However, the reduction in CysC levels may et al. (2007) suggested that “amateur” chaperones that co-localize represent the molecular factor responsible for increased risk of with the pathological lesions of AD, such as apolipoproteins and AD. These finding propose that a therapy involving enhancing heparan sulfate proteoglycans, bind amyloidogenic proteins and CysC concentration may be developed to prevent, postpone, or may be involved in conformational changes of Aβ and in the halt the disease. clearance of Aβ from the brain via phagocytosis or active trans- port across the blood-brain barrier. Similarly, interaction between ACKNOWLEDGMENTS amyloidogenic proteins results in inhibition of amyloid formation This work was supported by the National Institutes of Health and, therefore, has a neuroprotective function in diseases such (AG017617 and AG037693) and the Alzheimer’s Association as AD. (IIRG-11-204579).

Frontiers in Molecular Neuroscience www.frontiersin.org July 2012 | Volume 5 | Article 79 | 6 Kaur and Levy Neuroprotection by cystatin C

REFERENCES Finckh,U.,Nitsch,R.M.,and for Alzheimer disease. Neurology 64, Stoppe, G., Reiss, J., Miserez, A. Abrahamson, M., Buttle, D. J., Mason, Binetti, G. (2003). Alzheimer 755–757. R.,Staehelin,H.B.,Rebeck,G.W., R.W.,Hansson,H.,Grubb,A.O., disease-associated cystatin C vari- Chauhan, V. P., Ray, I., Chauhan, A., Hyman, B. T., Binetti, G., Hock, Lilja, H., and Ohlsson, K. (1991). ant undergoes impaired secretion. and Wisniewski, H. M. (1999). C., Growdon, J. H., and Nitsch, Regulation of cystatin C activity by Neurobiol. Dis. 13, 15–21. Binding of gelsolin, a secretory R. M. (2000). Genetic association serine proteinases. Biomed. Biochim. Bernstein, H. G., Kirschke, H., protein, to amyloid β-protein. of a cystatin C gene polymor- Acta 50, 587–593. Wiederanders, B., Pollak, K. Biochem. Biophys. Res. Commun. phism with late-onset Alzheimer Abrahamson, M., Islam, M. Q., Szpirer, H.,Zipress,A.,andRinne,A. 258, 241–246. disease. Arch. Neurol. 57, J., Szpirer, C., and Levan, G. (1989). (1996). The possible place of Choi, S. H., Leight, S. N., Lee, V. M., 1579–1583. The human cystatinSgene, SSC cathepsins and cystatins in the Li,T.,Wong,P.C.,Johnson,J.A., Gauthier,S.,Kaur,G.,Mi,W.,Tizon, (CST3), mutated in hereditary puzzle of Alzheimer disease: a Saraiva, M. J., and Sisodia, S. S. B., and Levy, E. (2011). Protective cystatin C amyloid angiopathy, is review. Mol. Chem. Neuropathol. 27, (2007). Accelerated Aβ deposition mechanisms by cystatin C in neu- located on chromosome 20. Hum. 225–247. in APPswe/PS1deltaE9 mice with rodegenerative diseases. Front. Genet. 82, 223–226. Bernstein, H. G., Rinne, R., Kirschke, hemizygous deletions of TTR Biosci. (Schol Ed.) 3, 541–554. Abrahamson, M., Olafsson, I., H., Jarvinen, M., Knofel, B., and (transthyretin). J. Neurosci. 27, Gene overview of all published Palsdottir, A., Ulvsback, M., Rinne, A. (1994). Cystatin A- 7006–7010. AD-association studies for Lundwall, A., Jensson, O., and like immunoreactivity is widely Cohen, D. H., Feiner, H., Jensson, O., CST3: http://www.alzforum.org/res/ Grubb, A. O. (1990). Structure and distributed in human brain and and Frangione, B. (1983). Amyloid com/gen/alzgene/geneoverview.asp? expression of the human cystatin C accumulates in neuritic plaques of fibril in hereditary cerebral hemor- geneid=66 gene. Biochem. J. 268, 287–294. Alzheimer disease subjects. Brain rhage with amyloidosis (HCHWA) Ghidoni, R., Benussi, L., Glionna, M., Arnason, A. (1935). Apoplexie und ihre Res. Bull. 33, 477–481. is related to the gastroentero- Desenzani, S., Albertini, V., Levy, Vererbung. Acta Psychiatr. Neurol. Bertram, L., McQueen, M. B., Mullin, pancreatic neuroendocrine protein, E., Emanuele, E., and Binetti, G. Scand. Suppl. 7, 1–180. K.,Blacker,D.,andTanzi,R.E. γ trace. J. Exp. Med. 158, 623–628. (2010). Plasma cystatin C and risk Aronica, E., van Vliet, E. A., (2007). Systematic meta-analyses of Craig-Schapiro, R., Kuhn, M., Xiong, of developing Alzheimer’s disease Hendriksen, E., Troost, D., Lopes Alzheimer disease genetic associa- C.,Pickering,E.H.,Liu,J., in subjects with mild cognitive da Silva, F. H., and Gorter, J. A. tion studies: the AlzGene database. Misko, T. P., Perrin, R. J., Bales, impairment. J. Alzheimers Dis. 22, (2001). Cystatin C, a cysteine Nat. Genet. 39, 17–23. K. R., Soares, H., Fagan, A. M., 985–991. protease inhibitor, is persistently Beyer,K.,Lao,J.I.,Gomez,M.,Riutort, and Holtzman, D. M. (2011). Ghidoni, R., Benussi, L., Paterlini, up-regulated in neurons and glia N.,Latorre,P.,Mate,J.L.,andAriza, Multiplexed immunoassay panel A., Missale, C., Usardi, A., Rossi, in a rat model for mesial temporal A. (2001). Alzheimer’s disease and identifies novel CSF biomarkers for R., Barbiero, L., Spano, P., and lobe epilepsy. Eur. J. Neurosci. 14, the cystatin C gene polymorphism: Alzheimer’s disease diagnosis and Binetti, G. (2007). Presenilin muta- 1485–1491. an association study. Neurosci. Lett. prognosis. PLoS ONE 6:e18850. doi: tions alter cystatin C trafficking in Babiloni, C., Benussi, L., Binetti, G., 315, 17–20. 10.1371/journal.pone.0018850 mouse primary neurons. Neurobiol. Bosco, P., Busonero, G., Cesaretti, Bobek, L. A., and Levine, M. J. (1992). Crawford, F. C., Freeman, M. J., Aging 28, 371–376. S.,DalForno,G.,DelPercio,C., Cystatins-inhibitors of cysteine pro- Schinka, J. A., Abdullah, L. I., Ghidoni, R., Paterlini, A., Albertini, Ferri, R., Frisoni, G., Roberta, G., teinases. Crit. Rev. Oral Biol. Med. 3, Gold, M., Hartman, R., Krivian, K., V., Glionna, M., Monti, E., Rodriguez, G., Squitti, R., and 307–332. Morris, M. D., Richards, D., Duara, Schiaffonati, L., Benussi, L., Rossini, P. M. (2006). Genotype Butterfield, D. A., and Boyd- R., Anand, R., and Mullan, M. J. Levy, E., and Binetti, G. (2011). (cystatin C) and EEG phenotype Kimball, D. (2004). Amyloid (2000). A polymorphism in the Cystatin C is released in associa- in Alzheimer disease and mild β-peptide(1–42) contributes to the cystatin C gene is a novel risk factor tion with exosomes: a new tool of cognitive impairment: a mul- oxidative stress and neurodegener- for late-onset Alzheimer’s disease. neuronal communication which is ticentric study. Neuroimage 29, ation found in Alzheimer disease Neurology 55, 763–768. unbalanced in Alzheimer’s disease. 948–964. brain. Brain Pathol. 14, 426–432. Deng,A.,Irizarry,M.C.,Nitsch,R. Neurobiol. Aging 32, 1435–1442. Bai, Y., Markham, K., Chen, F., Buxbaum, J. N., Ye, Z., Reixach, N., M., Growdon, J. H., and Rebeck, Ghiso,J.,Plant,G.T.,Revesz,T., Weerasekera, R., Watts, J., Horne, Friske, L., Levy, C., Das, P., Golde, G. W. (2001). Elevation of cys- Wisniewski, T., and Frangione, B. P., Wakutani, Y., Bagshaw, R., T., Masliah, E., Roberts, A. R., tatin C in susceptible neurons in (1995). Familial cerebral amyloid Mathews, P. M., Fraser, P. E., and Bartfai, T. (2008). Transthyretin Alzheimer’s disease. Am. J. Pathol. angiopathy (British type) with non- Westaway, D., St. George-Hyslop, protects Alzheimer’s mice from the 159, 1061–1068. neuritic amyloid plaque formation P., and Schmitt-Ulms, G. (2008). behavioral and biochemical effects Dodel, R. C., Du, Y., Depboylu, C., maybeduetoanovelamy- The in vivo brain interactome of Aβ toxicity. Proc.Natl.Acad.Sci. Kurz, A., Eastwood, B., Farlow, loid protein. J. Neurol. Sci. 129, of the amyloid precursor pro- U.S.A. 105, 2681–2686. M., Oertel, W. H., Muller, U., 74–75. tein. Mol. Cell Proteomics 7, Cataldo, A. M., Barnett, J. L., Berman, and Riemenschneider, M. (2002). Ghiso, J., Pons-Estel, B., and Frangione, 15–34. S. A., Li, J., Quarless, S., Bursztajn, A polymorphism in the cystatin B. (1986). Hereditary cerebral Balbin, M., and Abrahamson, M. S.,Lippa,C.,andNixon,R.A. C promoter region is not associ- amyloid angiopathy: the amyloid (1991). SstII polymorphic sites in (1995). Gene expression and cel- ated with an increased risk of AD. fibrils contain a protein which is a the promoter region of the human lular content of cathepsin D in Neurology 58, 664. variant of cystatin C, an inhibitor cystatin C gene. Hum. Genet. 87, Alzheimer’s disease brain: evi- Filler, G., Bokenkamp, A., Hofmann, of lysosomal cysteine proteases. 751–752. dence for early up-regulation of W.,LeBricon,T.,Martinez-Bru,C., Biochem. Biophys. Res. Commun. Balbin, M., Grubb, A. O., and the endosomal- lysosomal system. and Grubb, A. (2005). Cystatin C 136, 548–554. Abrahamson, M. (1993). An Neuron 14, 671–680. as a marker of GFR–history, indi- Goddard, K. A., Olson, J. M., Payami, Ala/Thr variation in the coding Cataldo, A. M., and Nixon, R. A. cations, and future research. Clin. H., van der Voet, M., Kuivaniemi, region of the human cystatinSgene, (1990). Enzymatically active lyso- Biochem. 38, 1–8. H., and Tromp, G. (2004). Evidence SSC (CST3) detected as a SstII somal proteases are associated with Finckh, U., von der Kammer, H., of linkage and association on chro- polymorphism. Hum. Genet. 92, amyloid deposits in Alzheimer Velden, J., Michel, T., Andresen, mosome 20 for late-onset Alzheimer 206–207. brain. Proc. Natl. Acad. Sci. U.S.A. B., Deng, A., Zhang, J., Muller- disease. Neurogenetics 5, 121–128. Benussi, L., Ghidoni, R., Steinhoff, T., 87, 3861–3865. Thomsen, T., Zuchowski, Graffagnino, C., Herbstreith, M. Alberici, A., Villa, A., Mazzoli, F., Cathcart, H. M., Huang, R., Lanham, K.,Menzer,G.,Mann,U., H.,Schmechel,D.E.,Levy,E., Nicosia, F., Barbiero, L., Broglio, I. S., Corder, E. H., and Poduslo, S. Papassotiropoulos, A., Heun, Roses, A. D., and Alberts, M. J. L., Feudatari, E., Signorini, S., E. (2005). Cystatin C as a risk factor R., Zurdel, J., Holst, F., Benussi, L., (1995). Cystatin C mutation in

Frontiers in Molecular Neuroscience www.frontiersin.org July 2012 | Volume 5 | Article 79 | 7 Kaur and Levy Neuroprotection by cystatin C

an elderly man with sporadic trace proteins. Proc. Soc. Exp. Biol. Katakai, K., Shinoda, M., Kabeya, K., and kininogen by human cathepsin amyloid angiopathy and intrac- Med. 124, 961–966. Watanabe, M., Ohe, Y., Mori, M., D. FEBS Lett. 280, 211–215. erebral hemorrhage. Stroke 26, Hua,Y.,Zhao,H.,Lu,X.,Kong,Y., and Ishikawa, K. (1997). Changes Lerner, U. H., and Grubb, A. (1992). 2190–2193. and Jin, H. (2012). Meta-analysis of in distribution of cystatin C, Human cystatin C, a cysteine Grubb, A. O. (1992). Diagnostic value the cystatin C(CST3) Gene G73A apolipoprotein E and ferritin in proteinase inhibitor, inhibits bone of analysis of cystatin C and pro- polymorphism and susceptibility to rat hypothalamus after hypophy- resorption in vitro stimulated by tein HC in biological fluids. Clin. Alzheimer’s disease. Int. J. Neurosci. sectomy. J. Neuroendocrinol. 9, parathyroid hormone and parathy- Nephrol. 38(Suppl. 1), S20–S27. [Epub ahead of print]. 247–253. roid hormone-related peptide of Gudmundsson, G., Hallgrimsson, J., Huh, C. G., Hakansson, K., Nathanson, Kaur, G., Mohan, P., Pawlik, M., malignancy. J. Bone Miner. Res. 7, Jonasson, T. A., and Bjarnason, O. C. M., Thorgeirsson, U. P., Jonsson, Derosa,S.,Fajiculay,J.,Che,S., 433–440. (1972). Hereditary cerebral haem- N., Grubb, A., Abrahamson, M., Grubbs, A., Ginsberg, S., Nixon, Levy, E., Lopez-Otin, C., Ghiso, J., orrhage with amyloidosis. Brain 95, and Karlsson, S. (1999). Decreased R., and Levy, E. (2010). Cystatin Geltner, D., and Frangione, B. 387–404. metastatic spread in mice homozy- C rescues degenerating neurons (1989). Stroke in Icelandic patients Haan, J., Maat-Schieman, M. L. C., gous for a null allele of the cystatin C in a cystatin B-knockout mouse with hereditary amyloid angiopathy van Duinen, S. G., Jensson, O., protease inhibitor gene. Mol. Pathol. model of progressive myoclonus is related to a mutation in the Thorsteinsson, L., and Roos, R. 52, 332–340. epilepsy. Am. J. Pathol. 177, cystatinSgene, SSC, an inhibitor of A. C. (1994). Co-localization of Ii, K., Ito, H., Kominami, E., and 2256–2267. cysteine proteases. J. Exp. Med. 169, β/A4 and cystatin C in corti- Hirano, A. (1993). Abnormal dis- Kiuru, S., Salonen, O., and Haltia, M. 1771–1778. cal blood vessels in Dutch, but tribution of cathepsin proteinases (1999). Gelsolin-related spinal and Levy, E., Sastre, M., Kumar, A., Gallo, not in Icelandic hereditary cerebral and endogenous inhibitors (cys- cerebral amyloid angiopathy. Ann. G., Piccardo, P., Ghetti, B., and hemorrhage with amyloidosis. Acta tatins) in the hippocampus of Neurol. 45, 305–311. Tagliavini, F. (2001). Codeposition Neurol. Scand. 89, 367–371. patients with Alzheimer’s disease, Kiuru-Enari, S., Somer, H., of cystatin C with amyloid-ßprotein Hakansson,K.,Huh,C.,Grubb,A., parkinsonism-dementia complex Seppalainen, A. M., Notkola, in the brain of Alzheimer’s dis- Karlsson, S., and Abrahamson, M. on Guam, and senile dementia I. L., and Haltia, M. (2002). ease patients. J. Neuropathol. Exp. (1996). Mouse and rat cystatin C: and in the aged. Virchows Arch. Neuromuscular pathology in Neurol. 60, 94–104. Escherichia coli production, charac- A Pathol. Anat. Histopathol. 423, hereditary gelsolin amyloidosis. Lin, C., Wang, S. T., Wu, C. W., terization and tissue distribution. 185–194. J. Neuropathol. Exp. Neurol. 61, Chuo, L. J., and Kuo, Y. M. (2003). Comp.Biochem.Physiol.BBiochem. Ishimaru, H., Ishikawa, K., Ohe, Y., 565–571. The association of a cystatin C Mol. Biol. 114, 303–311. Takahashi, A., and Maruyama, Y. Klein, W. L., Krafft, G. A., and Finch, gene polymorphism with late- Hansson, S. F., Andreasson, U., Wall, (1996). Cystatin C and apolipopro- C. E. (2001). Targeting small Aβ onset alzheimer’s disease and M., Skoog, I., Andreasen, N., Wallin, tein E immunoreactivities in oligomers: the solution to an vascular dementia. Chin. J. Physiol. A., Zetterberg, H., and Blennow, K. CA1 neurons in ischemic gerbil Alzheimer’s disease conundrum? 46, 111–115. (2009). Reduced levels of amyloid- hippocampus. Brain Res. 709, Trends Neurosci. 24, 219–224. Lindahl, P., Ripoll, D., Abrahamson, β-binding proteins in cerebrospinal 155–162. Kumada, T., Hasegawa, A., Iwasaki, Y., M., Mort, J. S., and Storer, A. C. fluid from Alzheimer’s disease Itoh, Y., Yamada, M., Hayakawa, M., Baba, H., and Ikenaka, K. (2004). (1994). Evidence for the interaction patients. J. Alzheimers Dis. 16, Otomo, E., and Miyatake, T. (1993). Isolation of cystatin C via func- of valine-in cystatin C with the S2 389–397. Cerebral amyloid angiopathy: a sig- tional cloning of astrocyte differ- subsite of cathepsin B. Biochemistry Hasegawa, A., Naruse, M., Hitoshi, S., nificant cause of cerebellar as well entiation factors. Dev. Neurosci. 26, 33, 4384–4392. Iwasaki, Y., Takebayashi, H., and as lobar cerebral hemorrhage in 68–76. Lukasiuk, K., Pirttila, T. J., and Ikenaka, K. (2007). Regulation of the elderly. J. Neurol. Sci. 116, Kuo, Y. M., Kokjohn, T. A., Kalback, Pitkanen, A. (2002). Upregulation glial development by cystatin C. 135–141. W., Luehrs, D., Galasko, D. R., of cystatin C expression in J. Neurochem. 100, 12–22. Jelic, V., Julin, P., Shigeta, M., Nordberg, Chevallier, N., Koo, E. H., the rat hippocampus during Helisalmi, S., Vakeva, A., Hiltunen, A.,Lannfelt,L.,Winblad,B., Emmerling, M. R., and Roher, epileptogenesis in the amygdala M., and Soininen, H. (2009). and Wahlund, L. O. (1997). A. E. (2000). Amyloid-β peptides stimulation model of temporal lobe Flanking markers of cystatin c Apolipoprotein E epsilon4 allele interact with plasma proteins epilepsy. Epilepsia 43(Suppl. 5), (CST3) gene do not show asso- decreases functional connec- and erythrocytes: implications 137–145. ciation with Alzheimer’s disease. tivity in Alzheimer’s disease as for their quantitation in plasma. Maetzler, W., Schmid, B., Synofzik, M., Dement. Geriatr. Cogn. Disord. 27, measured by EEG coherence. J. Biochem. Biophys. Res. Commun. Schulte, C., Riester, K., Huber, H., 318–321. Neurol. Neurosurg. Psychiatr. 63, 268, 750–756. Brockmann, K., Gasser, T., Berg, Hendriksen, H., Datson, N. A., Ghijsen, 59–65. Lehtovirta, M., Partanen, J., Kononen, D., and Melms, A. (2010). The W.E.,vanVliet,E.A.,daSilva,F. Juszczyk, P., Paraschiv, G., Szymanska, M., Hiltunen, J., Helisalmi, S., CST3 BB genotype and low cys- H., Gorter, J. A., and Vreugdenhil, E. A., Kolodziejczyk, A. S., Hartikainen, P., Riekkinen, P., Sr., tatin C cerebrospinal fluid levels are (2001). Altered hippocampal gene Rodziewicz-Motowidlo, S., and Soininen, H. (2000). A lon- associated with dementia in Lewy expression prior to the onset of Grzonka, Z., and Przybylski, gitudinal quantitative EEG study body disease. J. Alzheimers Dis. 19, spontaneous seizures in the rat post- M. (2009). Binding epitopes and of Alzheimer’s disease: relation to 937–942. status epilepticus model. Eur. J. interaction structure of the neu- apolipoprotein E polymorphism. Mares, J., Kanovsky, P., Herzig, R., Neurosci. 14, 1475–1484. roprotective protease inhibitor Dement. Geriatr. Cogn. Disord. 11, Stejskal, D., Vavrouskova, J., Hill, I. E., Preston, E., Monette, R., and cystatin C with β-amyloid revealed 29–35. Hlustik, P., Vranova, H., Burval, MacManus, J. P. (1997). A compar- by proteolytic excision mass spec- Lehtovirta, M., Partanen, J., Kononen, S., Zapletalova, J., Pidrman, V., ison of cathepsin B processing and trometry and molecular docking M., Soininen, H., Helisalmi, S., Obereigneru, R., Suchy, A., Vesely, distribution during neuronal death simulation. J. Med. Chem. 52, Mannermaa, A., Ryynanen, M., J., Podivinsky, J., and Urbanek, in rats following global ischemia 2420–2428. Hartikainen, P., and Riekkinen, P. K. (2009). New laboratory mark- or decapitation necrosis. Brain Res. Kaeser, S. A., Herzig, M. C., Sr. (1996). Spectral analysis of EEG ers in diagnosis of Alzheimer 751, 206–216. Coomaraswamy, J., Kilger, E., in Alzheimer’s disease: relation to dementia. Neurol. Res. 31, Hochwald, G. M., Pepe, A. J., and Selenica, M. L., Winkler, D. T., apolipoprotein E polymorphism. 1056–1059. Thorbecke, G. J. (1967). Trace Staufenbiel, M., Levy, E., Grubb, A., Neurobiol. Aging 17, 523–526. Maruyama, H., Izumi, Y., Oda, M., proteins in biological fluids. IV. and Jucker, M. (2007). Cystatin C Lenarcic, B., Krasovec, M., Ritonja, A., Torii, T., Morino, H., Toji, H., Physicochemical properties and modulates cerebral β-amyloidosis. Olafsson, I., and Turk, V. (1991). Sasaki, K., Terasawa, H., Nakamura, sites of formation of γ trace and β Nat. Genet. 39, 1437–1439. Inactivation of human cystatin C S., and Kawakami, H. (2001). Lack

Frontiers in Molecular Neuroscience www.frontiersin.org July 2012 | Volume 5 | Article 79 | 8 Kaur and Levy Neuroprotection by cystatin C

of an association between cystatin mutations in cystatin C gene in Palm, D. E., Knuckey, N. W., Primiano, myoclonic epilepsy and cerebellar C gene polymorphisms in Japanese cerebral amyloid angiopathy with M. J., Spangenberger, A. G., and apoptosis in cystatin B-deficient patients with Alzheimer’s disease. cystatin C deposition. Mol. Chem. Johanson, C. E. (1995). Cystatin C, mice. Nat. Genet. 20, 251–258. Neurology 57, 337–339. Neuropathol. 33, 63–78. a protease inhibitor, in degenerating Perrin, R. J., Craig-Schapiro, R., Maruyama, K., Ikeda, S., Ishihara, Nagai, A., Terashima, M., Sheikh, A. M., rat hippocampal neurons following Malone, J. P., Shah, A. R., Gilmore, T., Allsop, D., and Yanagisawa, Notsu, Y., Shimode, K., Yamaguchi, transient forebrain ischemia. Brain P.,Davis,A.E.,Roe,C.M.,Peskind, N. (1990). Immunohistochemical S., Kobayashi, S., Kim, S. U., and Res. 691, 1–8. E.R.,Li,G.,Galasko,D.R.,Clark,C. characterization of cerebrovascular Masuda, J. (2008). Involvement of Palsdottir, A., Abrahamson, M., M., Quinn, J. F., Kaye, J. A., Morris, amyloid in autopsied cases using cystatin C in pathophysiology of Thorsteinsson, L., Arnason, A., J. C., Holtzman, D. M., Townsend, antibodies to β protein and cystatin CNS diseases. Front. Biosci. 13, Olafsson, I., Grubb, A. O., and R. R., and Fagan, A. M. (2011). C. Stroke 21, 397–403. 3470–3479. Jensson, O. (1988). Mutation in Identification and validation of Maruyama, K., Kametani, F., Ikeda, Nairismagi, J., Grohn, O. H., Kettunen, cystatin C gene causes hereditary novel cerebrospinal fluid biomark- S.,Ishihara,T.,andYanagisawa,N. M. I., Nissinen, J., Kauppinen, brain haemorrhage. Lancet 2, ers for staging early Alzheimer’s (1992). Characterization of amy- R. A., and Pitkanen, A. (2004). 603–604. disease. PLoS ONE 6:e16032. doi: loid fibril protein from a case of Progression of brain damage after Paraoan, L., Ratnayaka, A., Spiller, D. 10.1371/journal.pone.0016032 CAA showing immunohistochem- status epilepticus and its associa- G., Hiscott, P., White, M. R., and Pirttila,T.J.,Lukasiuk,K.,Hakansson, ical reactivity for both β protein tion with epileptogenesis: a quanti- Grierson, I. (2004). Unexpected K.,Grubb,A.,Abrahamson,M., and cystatin C. Neurosci. Lett. 144, tative MRI study in a rat model of intracellular localization of the and Pitkanen, A. (2005). Cystatin 38–42. temporal lobe epilepsy. Epilepsia 45, AMD-associated cystatin C variant. C modulates neurodegeneration Matsuoka, Y., Saito, M., LaFrancois, 1024–1034. Traffic 5, 884–895. and neurogenesis following status J.,Gaynor,K.,Olm,V.,Wang, Nakamura, Y., Takeda, M., Suzuki, H., Paraoan, L., White, M. R., Spiller, epilepticus in mouse. Neurobiol. L.,Casey,E.,Lu,Y.,Shiratori,C., Hattori, H., Tada, K., Hariguchi, D. G., Grierson, I., and Maden, Dis. 20, 241–253. Lemere, C., and Duff, K. (2003). S., Hashimoto, S., and Nishimura, B. E. (2001). Precursor cystatin Pirttila, T. J., and Pitkanen, A. (2006). Novel therapeutic approach for T. (1991). Abnormal distribution of C in cultured retinal pigment Cystatin C expression is increased the treatment of Alzheimer’s cathepsins in the brain of patients epithelium cells: evidence for in the hippocampus following pho- disease by peripheral adminis- with Alzheimer’s disease. Neurosci. processing through the secretory tothrombotic stroke in rat. Neurosci. tration of agents with an affinity Lett. 130, 195–198. pathway. Mol. Membr. Biol. 18, Lett. 395, 108–113. to β-amyloid. J. Neurosci. 23, Ndjole, A. M., Bodolea, C., Nilsen, 229–236. Price, D. L., Martin, L. J., Sisodia, S. 29–33. T., Gordh, T., Flodin, M., and Parent,J.M.,Yu,T.W.,Leibowitz,R. S., Walker, L. C., Voytko, M. L., McCarron,M.O.,Nicoll,J.A.,Stewart, Larsson, A. (2010). Determination T.,Geschwind,D.H.,Sloviter,R. Wagster, M. V., Cork, L. C., and J., Ironside, J. W., Mann, D. M., of cerebrospinal fluid cystatin C S., and Lowenstein, D. H. (1997). Koliatsos, V. E. (1994). “The aged Love, S., Graham, D. I., and on Architect ci(8200). J. Immunol. Dentate granule cell neurogen- nonhuman primate. A model for Grubb, A. (2000). Absence of Methods 360, 84–88. esis is increased by seizures and the behavioral and brain abnormal- cystatin C mutation in sporadic Nitatori, T., Sato, N., Waguri, S., contributes to aberrant network ities occurring in aged humans,” in cerebral amyloid angiopathy- Karasawa, Y., Araki, H., Shibanai, reorganization in the adult rat Alzheimer’s Disease, edsR.D.Terry, related hemorrhage. Neurology 54, K., Kominami, E., and Uchiyama, hippocampus. J. Neurosci. 17, R.Katzman,andK.L.Blick(New 242–244. Y. (1995). Delayed neuronal death 3727–3738. York, NY: Raven Press), 231–245. Mi, W., Jung, S. S., Yu, H., Schmidt, S. in the CA1 pyramidal cell layer Parfitt, M., Crook, R., Roques, P., Roks,G.,Cruts,M.,Slooter,A.J., D.,Nixon,R.A.,Mathews,P.M., of the gerbil hippocampus follow- Rossor, M., and Chartier-Harlin, M. Dermaut, B., Hofman, A., van Tagliavini, F., and Levy, E. (2009). ing transient ischemia is apoptosis. C. (1993). The cystatin-C gene is Broeckhoven, C., and van Duijn, C. Complexes of amyloid-β and cys- J. Neurosci. 15, 1001–1011. not linked to early onset familial M. (2001). The cystatin C polymor- tatin C in the human central ner- Noto, D., Cefalu, A. B., Barbagallo, C. Alzheimer’s disease. Neurosci. Lett. phism is not associated with early vous system. J. Alzheimers Dis. 18, M.,Pace,A.,Rizzo,M.,Marino,G., 154, 81–83. onset Alzheimer’s disease. Neurology 273–280. Caldarella, R., Castello, A., Pernice, Pasinetti,G.M.,Ungar,L.H.,Lange, 57, 366–367. Mi, W., Pawlik, M., Sastre, M., Jung, S. V., Notarbartolo, A., and Averna, D. J., Yemul, S., Deng, H., Yuan, Saitoh, E., Sabatini, L. M., Eddy, R. L., S.,Radvinsky,D.S.,Klein,A.M., M. R. (2005). Cystatin C levels X.,Brown,R.H.,Cudkowicz, Shows, T. B., Azen, E. A., Isemura, Sommer,J.,Schmidt,S.D.,Nixon, are decreased in acute myocar- M. E., Newhall, K., Peskind, E., S., and Sanada, K. (1989). The R.A.,Mathews,P.M.,andLevy,E. dial infarction: effect of cystatin Marcus, S., and Ho, L. (2006). human cystatinSgene, SSC (CST3) (2007). Cystatin C inhibits amyloid- C G73A gene polymorphism on Identification of potential CSF is a member of the cystatin gene β deposition in Alzheimer’s dis- plasma levels. Int. J. Cardiol. 101, biomarkers in ALS. Neurology 66, family which is localized on chro- ease mouse models. Nat. Genet. 39, 213–217. 1218–1222. mosome 20. Biochem. Biophys. Res. 1440–1442. Okamoto, K., Mizuno, Y., and Fujita, Pawlik, M., Danilov, V., Mancevska, K., Commun. 162, 1324–1331. Miyake, T., Gahara, Y., Nakayama, Y. (2008). Bunina bodies in Morbin, M., Tagliavini, F., and Levy, Sastre, M., Calero, M., Pawlik, M., M., Yamada, H., Uwabe, K., and amyotrophic lateral sclerosis. E. (2002). Hemorrhages caused by Mathews, P. M., Kumar, A., Danilov, Kitamura, T. (1996). Up-regulation Neuropathology 28, 109–115. overexpression of cystatin C in V., Schmidt, S. D., Nixon, R. A., of cystatin C by microglia in the Olafsson, I., Thorsteinsson, L., and transgenic mice. Neurobiol. Aging Frangione, B., and Levy, E. (2004). rat facial nucleus following axo- Jensson, O. (1996). The molecu- 23, S242. Binding of cystatin C to Alzheimer’s tomy. Brain Res. Mol. Brain Res. 37, lar pathology of hereditary cys- Pawlik, M., Sastre, M., Calero, M., amyloid b inhibits amyloid fib- 273–282. tatin C amyloid angiopathy causing Mathews, P. M., Schmidt, S. D., ril formation. Neurobiol. Aging 25, Mori, F., Tanji, K., Miki, Y., and brain hemorrhage. Brain Pathol. 6, Nixon, R. A., and Levy, E. (2004). 1033–1043. Wakabayashi, K. (2009). Decreased 121–126. Overexpression of human cystatin C Schwarzman, A. L., Gregori, L., Vitek, cystatin C immunoreactivity Olson, J. M., Goddard, K. A., and in transgenic mice does not affect M. P., Lyubski, S., Strittmatter, in spinal motor neurons and Dudek, D. M. (2002). A second levels of endogenous brain amy- W. J., Enghilde, J. J., Bhasin, astrocytes in amyotrophic lateral locus for very-late-onset Alzheimer loid β peptide. J. Mol. Neurosci. 22, R., Silverman, J., Weisgraber, K. sclerosis. J. Neuropathol. Exp. disease: a genome scan reveals link- 13–18. H.,Coyle,P.K.,Zagorski,M. Neurol. 68, 1200–1206. age to 20p and epistasis between Pennacchio, L. A., Bouley, D. M., G., Talafous, J., Eisenberg, M., Nagai, A., Kobayashi, S., Shimode, K., 20p and the amyloid precursor pro- Higgins, K. M., Scott, M. P., Saunders, A. M., Roses, A. D., and Imaoka, K., Umegae, N., Fujihara, tein region. Am. J. Hum. Genet. 71, Noebels, J. L., and Myers, R. Goldgaber, D. (1994). Transthyretin S., and Nakamura, M. (1998). No 154–161. M. (1998). Progressive ataxia, sequesters amyloid β protein

Frontiers in Molecular Neuroscience www.frontiersin.org July 2012 | Volume 5 | Article 79 | 9 Kaur and Levy Neuroprotection by cystatin C

and prevents amyloid formation. Sundelof, J., Sundstrom, J., Hansson, of cystatin C gene in choroid protein toxicity. Brain Res. 1089, Proc. Natl. Acad. Sci. U.S.A. 91, O., Eriksdotter-Jonhagen, M., plexus. Am. J. Physiol. 263, 67–78. 8368–8372. Giedraitis, V., Larsson, A., R195–R200. Wilhelmus, M. M., de Waal, R. M., and Schwarzman, A. L., Tsiper, M., Degerman-Gunnarsson, M., Turk, B., Turk, D., and Turk, V. Verbeek, M. M. (2007). Heat shock Wente, H., Wang, A., Vitek, M. Ingelsson, M., Minthon, L., (2000). Lysosomal cysteine pro- proteins and amateur chaperones in P., Vasiliev, V., and Goldgaber, Blennow, K., Kilander, L., Basun, teases: more than scavengers. amyloid-β accumulation and clear- D. (2004). Amyloidogenic and H., and Lannfelt, L. (2010). Cystatin Biochim. Biophys. Acta 1477, ance in Alzheimer’s disease. Mol. anti-amyloidogenic properties of C levels are positively correlated 98–111. Neurobiol. 35, 203–216. recombinant transthyretin variants. with both Aβ42 and tau levels Turk,V.,Stoka,V.,andTurk,D. Wilson, M. E., Boumaza, I., Lacomis, Amyloid 11, 1–9. in cerebrospinal fluid in persons (2008). Cystatins: biochemical and D., and Bowser, R. (2010). Cystatin Selenica, M. L., Wang, X., Ostergaard- with Alzheimer’s disease, mild structural properties, and medi- C: a candidate biomarker for Pedersen, L., Westlind-Danielsson, cognitive impairment, and healthy cal relevance. Front. Biosci. 13, amyotrophic lateral sclero- A., and Grubb, A. (2007). Cystatin C controls. J. Alzheimers Dis. 21, 5406–5420. sis. PLoS ONE 5:e15133. doi: reduces the in vitro formation of sol- 471–478. Vinters, H. V. (2001). Cerebral amy- 10.1371/journal.pone.0015133 uble Aβ1-oligomers and protofib- Taupin,P.,Ray,J.,Fischer,W.H., loid angiopathy: a microvascular Winkler, D. T., Bondolfi, L., Herzig, rils. Scand. J. Clin. Lab. Invest. 67, Suhr, S. T., Hakansson, K., Grubb, link between parenchymal and vas- M. C., Jann, L., Calhoun, M. E., 179–190. A., and Gage, F. H. (2000). FGF- cular dementia? Ann. Neurol. 49, Wiederhold, K. H., Tolnay, M., Simonsen, A. H., McGuire, J., Podust, 2-Responsive neural stem cell 691–693. Staufenbiel, M., and Jucker, M. V. N., Hagnelius, N. O., Nilsson, proliferation requires CCg, a novel Vinters,H.V.,Nishimura,G.S.,Secor, (2001). Spontaneous hemorrhagic T. K., Kapaki, E., Vassilopoulos, Autocrine/Paracrine cofactor. D. L., and Pardridge, W. M. (1990). stroke in a mouse model of cerebral D., and Waldemar, G. (2007). Neuron 28, 385–397. Immunoreactive A4 and γ-trace amyloid angiopathy. J. Neurosci. 21, A novel panel of cerebrospinal Tavera, C., Leung-Tack, J., Prevot, peptide colocalization in amy- 1619–1627. fluid biomarkers for the differ- D., Gensac, M. C., Martinez, J., loidotic arteriolar lesions in brains Wisniewski, H. M., and Terry, R. D. ential diagnosis of Alzheimer’s Fulcrand, P., and Colle, A. (1992). of patients with Alzheimer’s (1973). Morphology of the aging disease versus normal aging Cystatin C secretion by rat glomeru- disease. Am. J. Pathol. 137, brain, human and animal. Prog. and frontotemporal dementia. lar mesangial cells: autocrine loop 233–240. Brain Res. 40, 167–186. Dement. Geriatr. Cogn. Disord. 24, for in vitro growth-promoting activ- Walker, L. C., Masters, C., Beyreuther, Yamamoto-Watanabe, Y., Watanabe, 434–440. ity. Biochem. Biophys. Res. Commun. K., and Price, D. L. (1990). Amyloid M., Jackson, M., Akimoto, H., Skerget, K., Taler-Vercic, A., Bavdek, 182, 1082–1088. in the brains of aged squirrel Sugimoto, K., Yasujima, M., A.,Hodnik,V.,Ceru,S.,Tusek- Tizon, B., and Levy, E. (2006). “Protease monkeys. Acta Neuropathol. 80, Wakasaya, Y., Matsubara, E., Znidaric, M., Kumm, T., Pitsi, D., inhibitors and their involvement 381–387. Kawarabayashi, T., Harigaya, Y., Pompe-Novak, M., Palumaa, P., in neurological disorders,” in Walsh, D. M., and Selkoe, D. J. (2004). Lyndon,A.R.,andShoji,M. Soriano, S., Kopitar-Jerala, N., Turk, Handbook of Neurochemistry and Deciphering the molecular basis of (2010). Quantification of cystatin V.,Anderluh,G.,andZerovnik, Molecular Neurobiology,edA. memory failure in Alzheimer’s dis- C in cerebrospinal fluid from E. (2010). Interaction between Lajtha (New York, NY: Springer ease. Neuron 44, 181–193. various neurological disorders and oligomers of stefin B and amyloid-β Publishers), 591–624. Wang, B., Xie, Y. C., Yang, Z., Peng, D., correlation with G73A polymor- in vitro and in cells. J. Biol. Chem. Tizon, B., Ribe, E. M., Mi, W., Troy, C. Wang, J., Zhou, S., Li, S., and Ma, phism in CST3. Brain Res. 1361, 285, 3201–3210. M., and Levy, E. (2010a). Cystatin X. (2008). Lack of an association 140–145. Steinhoff, T., Moritz, E., Wollmer, M. C protects neuronal cells from amy- between Alzheimer’s disease and the Yamashima, T., Kohda, Y., Tsuchiya, K., A., Mohajeri, M. H., Kins, S., and loid β-induced toxicity. J. Alzheimers cystatin C (CST3) gene G73A poly- Ueno, T., Yamashita, J., Yoshioka, Nitsch, R. M. (2001). Increased cys- Dis. 19, 665–894. morphism in Mainland Chinese. T., and Kominami, E. (1998). tatin C in astrocytes of transgenic Tizon, B., Sahoo, S., Yu, H., Gauthier, Dement. Geriatr. Cogn. Disord. 25, Inhibition of ischaemic hippocam- mice expressing the K670N-M671L S., Kumar, A. R., Mohan, P., 461–464. pal neuronal death in primates mutation of the amyloid precur- Figliola, M., Pawlik, M., Grubb, Warfel,A.H.,Zucker-Franklin,D., with cathepsin B inhibitor CA-074, sor protein and deposition in brain A., Uchiyama, Y., Bandyopadhyay, Frangione, B., and Ghiso, J. (1987). a novel strategy for neuroprotec- amyloid plaques. Neurobiol. Dis. 8, S.,Cuervo,A.M.,Nixon,R.A., Constitutive secretion of cystatin tion based on ‘calpain-cathepsin 647–654. and Levy, E. (2010b). Induction of C(γ-trace) by monocytes and hypothesis’. Eur. J. Neurosci. 10, Sun, B., Zhou, Y., Halabisky, B., Lo, autophagy by cystatin C: a mecha- macrophages and its downregula- 1723–1733. I., Cho, S. H., Mueller-Steiner, S., nism that protects murine primary tion after stimulation. J. Exp. Med. Yang, H. T., Wilkening, S., and Devidze, N., Wang, X., Grubb, A., cortical neurons and neuronal 166, 1912–1917. Iadarola, M. J. (2001). Spinal and Gan, L. (2008). Cystatin C- cell lines. PLoS ONE 5:e9819. doi: Wei, L., Berman, Y., Castano, E. M., cord genes enriched in rat dorsal cathepsin B axis regulates amy- 10.1371/journal.pone.0009819 Cadene, M., Beavis, R. C., Devi, horn and induced by noxious loid β levels and associated neu- Tsuchiya, K., Kohda, Y., Yoshida, M., L., and Levy, E. (1998). Instability stimulation identified by sub- ronal deficits in an animal model Zhao,L.,Ueno,T.,Yamashita, of the amyloidogenic cystatin traction cloning and differential of Alzheimer’s disease. Neuron 60, J., Yoshioka, T., Kominami, C variant of hereditary cerebral hybridization. Neuroscience 103, 247–257. E., and Yamashima, T. (1999). hemorrhage with amyloidosis, 493–502. Sun, Q. (1989). Growth stimulation of Postictal blockade of ischemic Icelandic type. J. Biol. Chem. 273, Yang, Y., Liu, S., Qin, Z., Cui, Y., Qin, 3T3 fibroblasts by cystatin. Exp. Cell hippocampal neuronal death in 11806–11814. Y., and Bai, S. (2009). Alteration Res. 180, 150–160. primates using selective cathep- Wei, L., Walker, L. C., and Levy, of cystatin C levels in cerebrospinal Sundelof, J., Arnlov, J., Ingelsson, E., sin inhibitors. Exp. Neurol. 155, E. (1996). Cystatin C: Icelandic- fluid of patients with Guillain- Sundstrom, J., Basu, S., Zethelius, 187–194. like mutation in an animal model Barre syndrome by a proteomi- B., Larsson, A., Irizarry, M. C., Tsuji-Akimoto, S., Yabe, I., Niino, M., of cerebrovascular β amyloidosis. cal approach. Mol. Biol. Rep. 36, Giedraitis, V., Ronnemaa, E., Kikuchi, S., and Sasaki, H. (2009). Stroke 27, 2080–2085. 677–682. Degerman-Gunnarsson, M., Cystatin C in cerebrospinal fluid as Wilhelmus, M. M., Boelens, W. C., Yasuhara,O.,Hanai,K.,Ohkubo, Hyman,B.T.,Basun,H.,Kilander, a biomarker of ALS. Neurosci. Lett. Otte-Holler, I., Kamps, B., de Waal, I., Sasaki, M., McGeer, P. L., and L., and Lannfelt, L. (2008). Serum 452, 52–55. R. M., and Verbeek, M. M. (2006). Kimura, H. (1993). Expression cystatin C and the risk of Alzheimer Tu, G. F., Aldred, A. R., Southwell, B. Small heat shock proteins inhibit of cystatin C in rat, monkey and disease in elderly men. Neurology R., and Schreiber, G. (1992). Strong amyloid-β protein aggregation human brains. Brain Res. 628, 71, 1072–1079. conservation of the expression and cerebrovascular amyloid-β 85–92.

Frontiers in Molecular Neuroscience www.frontiersin.org July 2012 | Volume 5 | Article 79 | 10 Kaur and Levy Neuroprotection by cystatin C

Ying,G.X.,Huang,C.,Jiang,Z.H., Zerovnik, E., Staniforth, R. A., and commercial or financial relationships Mol. Neurosci. 5:79. doi: 10.3389/fnmol. Liu, X., Jing, N. H., and Zhou, Turk, D. (2010). Amyloid fib- that could be construed as a potential 2012.00079 C. F. (2002). Up-regulation of cys- ril formation by human stefins: conflict of interest. Copyright © 2012 Kaur and Levy. tatin C expression in the murine structure, mechanism and puta- This is an open-access article dis- hippocampus following perforant tive functions. Biochimie 92, tributed under the terms of the Creative path transections. Neuroscience 112, 1597–1607. Received: 12 April 2012; paper pend- Commons Attribution License,which 289–298. ing published: 03 May 2012; accepted: permits use, distribution and reproduc- Zellner, M., Veitinger, M., and Umlauf, 20 June 2012; published online: 06 July tion in other forums, provided the origi- E. (2009). The role of proteomics in Conflict of Interest Statement: The 2012. nal authors and source are credited and dementia and Alzheimer’s disease. authors declare that the research Citation: Kaur G and Levy E (2012) subject to any copyright notices concern- Acta Neuropathol. 118, 181–195. was conducted in the absence of any Cystatin C in Alzheimer’s disease. Front. ing any third-party graphics etc.

Frontiers in Molecular Neuroscience www.frontiersin.org July 2012 | Volume 5 | Article 79 | 11