Background: Synaptic and axonal loss are two major mechanisms underlying Alzheimer’s disease (AD) pathogenesis, and biomarkers reflecting changes in these cellular processes are needed for early diagnosis and monitoring the progression of AD. Contactin-2 is a synaptic and axonal membrane protein that interacts with proteins involved in the pathology of AD such as amyloid precursor protein (APP) and beta-secretase 1 (BACE1). We hypothesized that AD might be characterized by changes in contactin-2 levels in the cerebrospinal fluid (CSF) and brain tissue. Therefore, we aimed to investigate the levels of contactin-2 in the CSF and evaluate its relationship with disease pathology. Methods: Contactin-2 was measured in CSF from two cohorts (selected from the Amsterdam Dementia Cohort), comprising samples from controls (cohort 1, n = 28; cohort 2, n = 20) and AD (cohort 1, n = 36; cohort 2, n = 70) using an analytically validated commercial enzyme-linked immunosorbent assay (ELISA). The relationship of contactin-2 with cognitive decline (Mini-Mental State Examination (MMSE)) and other CSF biomarkers reflecting AD pathology were analyzed. We further characterized the expression of contactin-2 in postmortem AD human brain (n = 14) versus nondemented controls (n = 9). Results: CSF contactin-2 was approximately 1.3-fold reduced in AD patients compared with controls (p < 0.0001). Overall, contactin-2 levels correlated with MMSE scores (r = 0.35, p = 0.004). We observed that CSF contactin-2 correlated with the levels of phosphorylated tau within the control (r = 0.46, p < 0.05) and AD groups (r = 0.31, p <0. 05). Contactin-2 also correlated strongly with another synaptic biomarker, neurogranin (control: r = 0.62, p < 0.05; AD: r = 0.60, p < 0.01), and BACE1, a contactin-2 processing enzyme (control: r = 0.64, p < 0.01; AD: r = 0.46, p < 0.05). Results were further validated in a second cohort (p < 0.01). Immunohistochemical analysis revealed that contactin-2 is expressed in the extracellular matrix. Lower levels of contactin-2 were specifically found in and around amyloid plaques in AD hippocampus and temporal cortex. Conclusions: Taken together, these data reveal that the contactin-2 changes observed in tissues are reflected in CSF, suggesting that decreased contactin-2 CSF levels might be a biomarker reflecting synaptic or axonal loss. Keywords: Contactin-2, Alzheimer’s disease (AD), CSF biomarker, Tau, Neurogranin, Beta secretase 1 * Correspondence: firstname.lastname@example.org Neurochemistry Laboratory, Clinical Chemistry Department, VU University Medical Center, De Boelelaan 1117, 1081 HV Amsterdam, the Netherlands Full list of author information is available at the end of the article © The Author(s). 2018 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Chatterjee et al. Alzheimer's Research & Therapy (2018) 10:52 Page 2 of 11 Background contactin-2 as a CSF biomarker candidate reflecting synap- Alzheimer’s disease (AD) is the major cause of dementia tic and axonal dysfunction in AD and to examine its rela- worldwide . AD patients are characterized by high tionship with other important players in AD pathogenesis. levels of cerebrospinal fluid (CSF) tau reflecting tangle Moreover, we further characterized the expression of this pathology whereas the underlying amyloid beta (Aβ) protein in postmortem hippocampus to explore the poten- plaque pathology is mirrored by decreased levels of tial role of this protein in AD pathogenesis. Aβ42 in the CSF . However, about 30% of the cogni- tively normal elderly also have an AD CSF biomarker Methods profile, making AD diagnosis complex [3, 4]. Thus, add- Human CSF sample subjects itional biomarkers are needed for a better diagnosis. Fur- For the first study, we included cognitively normal con- thermore, synaptic dysfunction [5, 6] and axonal loss  trols with subjective memory complaints (n = 28) and are early events in the pathogenesis of AD [6, 8–12]. AD patients (n = 36) (Table 1) from the Amsterdam De- Synapse loss has been suggested to be related more mentia Cohort . An additional validation cohort was strongly with cognitive impairment than plaque or tan- also included to replicate the findings (controls, n = 20; gle pathology [13–16]. Therefore, biomarkers reflecting AD, n = 70) (Table 1) from the same dementia cohort. these changes might be useful to support early diagnosis Diagnoses were defined in a multidisciplinary commit- and prognosis of AD. Several synaptic biomarkers in tee according to the criteria of the National Institute of CSF have been identified, such as neurogranin [17, 18], Neurological and Communicative Disorders and Stroke– synaptotagmin , synaptosomal-associated protein Alzheimer’s Disease and Related Disorders Association (SNAP)-25 , and Ras-related protein (Rab)-3A . [39, 40]. AD cases were additionally selected based on a Neurogranin is a promising synaptic biomarker which positive AD biomarker profile (CSF total tau (tTau)/ has been found to be specifically increased in AD [17, Aβ42 ratio > 0.52 ). When patients presented with 18, 21]. So far, there are no established biomarkers for cognitive complaints and the results of clinical assess- axonal loss specific for AD. Increased tau level has been ments were within the normal range they were labeled related with axonal loss , but increased tau is a rather as subjective memory complaint, hereinafter referred to unspecific finding indicating neurodegeneration . as controls . These nondemented control cases were Contactin-2 is a soluble cell-adhesion protein primar- selected based on a negative CSF AD biomarker profile ily expressed on the axonal and synaptic membranes (CSF tTau/Aβ42 ratio < 0.52 ). CSF was collected by [23–29]. It belongs to the immunoglobulin superfamily standard lumbar puncture and stored according to pre- and consists of six members (contactin-1 to contactin-6) viously published JPND-BIOMARKAPD guidelines until [29, 30]. Contactin-2 is expressed in hippocampal pyr- analysis [18, 42]. Samples within each cohort were amidal cells, cerebellar granule cells, the juxtaparanodal matched for age. Demographic and clinical details of all regions of myelinated nerve fibers [24, 30], and frontal patients are listed in Table 1. and temporal lobes [23, 31]. Contactin-2 is a multifunc- tional protein that plays important roles in axonal guid- Postmortem brain tissue ance during development [32, 33], neuronal fasciculation Postmortem hippocampus and temporal cortex from , axonal domain organization , and neuron-glia AD patients (n = 14) and nondemented controls (n =9) interaction . Interestingly, a genome-wide associ- were obtained from the Netherlands Brain Bank. Con- ation study (GWAS) identified single nucleotide poly- sidering that contactin-2 is expressed in the hippocam- morphisms (SNPs) in the gene encoding contactin-2 pus [30, 43] and temporal cortex [23, 31], and these (CNTN2) associated with AD . Contactin-2 interacts brain areas are primarily affected in AD , we used with proteins involved in AD pathogenesis, such as homogenates of postmortem hippocampus and temporal amyloid precursor protein (APP) [37, 38] and cortex tissue. All donors or their next of kin provided beta-secretase 1 (BACE1) [37, 39, 40]. Lower levels of written informed consent for brain autopsy and the use contactin-2 correlated with higher BACE1 activity in of medical records for research purposes. Sample pro- postmortem AD tissue . Thus, the interactions be- cessing is described in detail in Additional file 1: Section tween contactin-2 and BACE1 and APP proteins may in- 1.1. Patient details such as clinical and pathological diag- fluence the production of Aβ peptide and the nosis, Braak stage, age and sex, and postmortem delay subsequent formation of amyloid plaques. Interestingly, are outlined in Additional file 1: Table S1. higher levels of contactin-2 have been reported in AD CSF pools using proteomics approaches . Enzyme-linked immunosorbent assay (ELISA) analysis We hypothesized that AD might be associated with Contactin-2 was measured in both CSF and postmortem changes in contactin-2 levels in both CSF and brain. In brain tissue homogenates with the Contactin-2 duoset this study, we aimed to evaluate the potential for ELISA kit (R&D, Minneapolis, USA; cat. nos. DY1714– Chatterjee et al. Alzheimer's Research & Therapy (2018) 10:52 Page 3 of 11 Table 1 Demographic details of cohort 1 and cohort 2 Cohort 1 Cohort 2 Controls Patients with AD Controls Patients with AD n 28 36 20 70 Gender (male:female) 13:15 15:21 14:6 29:41 Age (years) (mean ± SD) 60 ± 7 62 ± 6 62 ± 3 62 ± 5 MMSE (mean ± SD) 27 ± 3 19 ± 5*** 28 ± 2 20 ± 6*** Aβ42 (pg/ml) (median [IQR]) 915 [815–1026] 468 [395–552]*** 1063 [1009–1214] 578 [518–645]*** tTau (pg/ml) (median [IQR]) 216 [161–309] 691 [559–962]*** 274 [239–315] 734 [552–1021]*** pTau (pg/ml) (median [IQR]) 47 [33–54] 92 [77–116]*** 43 [39–50] 90 [69–107]*** Contactin-2 (ng/ml) (median [IQR]) 78 [69–110] 59 [42–74]*** 65 [54–99] 61 [39–78]* Aβ amyloid beta, AD Alzheimer’s disease, IQR interquartile range, MMSE Mini-Mental State Examination, SD standard deviation *p < 0.05, **p < 0.01, ***p < 0.001, versus controls 05 and DY008), which uses antibodies raised against the or monoclonal rabbit anti-GAPDH (1:1000, clone 14C10; secreted part of contactin-2 (Leu29-Asn1012). We vali- Cell Signaling Technology, MA, USA)—overnight at 4 °C. dated this kit both for CSF samples and tissue samples After washing with wash buffer (0.05% w/v milk in TBS-T), using previous validation guidelines [45, 46] (Additional membranes were incubated for 1 h with polyclonal goat file 1: Table S2). CSF and postmortem brain samples anti-rabbit IgG/HRP (1:2000, DAKO, Glostrup, Denmark) were diluted 1:16 and 1:100, respectively, in reagent dilu- or goat anti-mouse IgG/HRP (1:1000, DAKO) in blocking ents provided in the kit and the assay was performed ac- buffer. Protein bands were detected with the ECL Western cording to the manufacturer’s protocol. The intra-assay Blotting detection kit (GE Healthcare, Amersham, UK). percentage coefficients of variation (%CVs) for CSF and Samples were always randomly distributed within the gels brain tissue were 1.9 and 1.3, and the interassay %CVs and the researcher was unaware of the diagnosis and spe- were 8.7 (CSF) and 9 (brain tissue), respectively. Spe- cifics of the samples. Immunoblot films were scanned, and cifics about the procedure can be found in Additional signal quantification was performed using ImageJ 1.45 file 1: Section 1.2. CSF Aβ42, tTau, and phosphorylated (NIH,Bethesda, USA).Contactin-2 band signal was nor- tau (pTau) were measured as a part of routine diagnosis malized by the GAPDH signal intensity. at the Neurochemistry laboratory at VU University Med- ical Centre, Amsterdam, the Netherlands, using com- Immunohistochemistry and immunofluorescence mercially available ELISA (Fujirebio, Ghent, Belgium) as Formalin-fixed and paraffin-embedded hippocampus previously performed  (Table 1). CSF BACE1 and and temporal cortex sections (5 μm) were mounted on neurogranin were measured using commercially avail- Superfrost plus tissue slides (Menzel-Glaser, Braun- able analytically validated ELISA kits from Euroimmun schweig, Germany) and dried overnight at 37 °C. Samples (Lübeck, Germany). CSF Aβ40 was measured using the from 12 individuals (7 AD and 6 controls) were immu- V-PLEX Plus Aβ Peptide Panel 1 (6E10) Kit (MSD, nostained. Two sections from each subject were analyzed Maryland, USA). All samples were randomized and were and stainings were found to be consistent. Immunohis- measured by a single experienced technician blinded to tochemistry (IHC) and immunofluorescence (IF) proce- the clinical groups. dures are described in detail in Additional file 1: Section 1.3. The primary antibodies used were: affinity-purified Western blotting polyclonal rabbit anti-Contactin-2 (IHC: 1:400, IF: 1:25; Human hippocampus and temporal cortex tissue homoge- HPA001397, Atlas Antibodies, Stockholm, Sweden); nates (20 μg per sample) were prepared in sample buffer monoclonal mouse anti- pTau Ser202/Thr205 AT-8 (IF: (2% SDS, 0.03 M Tris, 5% 2-mercaptoethanol, 10% glycerol, 1:800, MN1020, Thermo Fisher Scientific, Landsmeer, bromophenol blue) and heated for 5 min at 95 °C. Electro- Netherlands); and monoclonal mouse anti-Aβ IC-16 (IF: phoresis was carried out using 10% SDS-PAGE minigels. 1:200, a kind gift from Dr. Korth, University of Duessel- Next, proteins were transferred to polyvinylidene fluoride dorf, Germany). For IHC, the bound primary antibody (PVDF) membranes (Millipore, Bedford, USA) that were was detected using DAKO anti-rabbit/mouse EnVision+ subsequently blocked for 30 min with blocking buffer (5% System-HRP (DAKO, 45007, Glostrup, Denmark). Nu- w/v nonfat dried milk in PBS-Tween 0.5% v/v (PBS-T)), clei were visualized by Mayer’s hematoxylin counterstain and incubated with the corresponding primary anti- (Merck, MHS1, Zwijndrecht, Netherlands). For IF, the bodies—affinity-purified polyclonal rabbit anti-Contactin-2 following secondary antibodies were used: anti-rabbit  (1:1500, SAB4200299; Sigma Aldrich, St. Louis, USA) alexa-647 (1:250, DAKO), anti-mouse alexa-488 (1:250, Chatterjee et al. Alzheimer's Research & Therapy (2018) 10:52 Page 4 of 11 DAKO), and anti-mouse alexa-594 (1:250, DAKO). cohort (p = 0.049; Fig. 1b), where contactin-2 was reduced Thioflavin-S (Merck, T1892) was added to brain tissue by 20%. A positive correlation was observed between CSF sections after incubation with primary anti-pTau and contactin-2 and the Mini-Mental State Examination anti-contactin-2 antibodies and corresponding secondary (MMSE) in the total group (r =0.35, p =0.004; Additional antibodies for 1 min with a prior 10-min acetone fixation file 1: Figure S1). The correlation between contactin-2 and at room temperature. The slides were finally incubated MMSE were not significant when AD and control groups with DAPI for 10 mins and subsequently covered using were analyzed separately. The correlation was not ob- 80% glycerol in TBS, pH 7.4. Staining and imaging was served in the second cohort (r =0.11, p =0.2). performed by two independent researchers who were un- aware of the diagnosis of the cases. IHC images were cap- CSF contactin-2 and its relationship with core AD tured with a Zeiss light microscope equipped with a biomarkers digital camera and a 10× or 25× objective (12.5× ocular). Correlations were analyzed within each diagnostic group IF images were captured with a Nikon Eclipse Ti confocal (controls and AD individually). No correlation between microscope equipped with a 60× oil (numerical aperture contactin-2 and Aβ42 was observed within the control (NA) = 1.40) objective and NisElements 4.30 software. or AD groups (Fig. 2a). Both tTau and pTau strongly correlated with contactin-2 within controls (tTau: r = Statistics 0.48, p = 0.009, Fig. 2b; pTau: r = 0.46, p = 0.01, Fig. 2c). Differences in CSF contactin-2 levels between groups Within the AD groups only pTau correlated positively were tested with analysis of covariance (ANCOVA) ad- with contactin-2 (r = 0.31, p = 0.05, Fig. 2c). Similar re- justed for age and gender when applicable. Data were sults were obtained in the second cohort, with the ex- normalized by Templeton’s two-step method  if not ception of tTau which now correlated with contactin-2 normally distributed. Correlation analyses were per- within both control and AD groups (Additional file 1: formed using Pearson or Spearman correlation for para- Figure S2). Additionally, contactin-2 also correlated with metric and nonparametric data, respectively. Group CSF Aβ40 within both groups (n = 37; controls: r = 0.64, differences between AD and controls in postmortem p = 0.008; AD: r = 0.46, p = 0.03, Fig. 2d). Since samples were evaluated by Mann-Whitney U test. contactin-2 was associated with age within the AD The statistical tests were two-tailed and values with group in the second cohort, an age correction was ap- p(two-tailed) < 0.05 were considered significant. Statis- plied (Additional file 1: Figure S2). tical analyses were performed on SPSS version 22 (IBM SPSS Statistics for Windows, Version 21.0; IBM Corp., Contactin-2 correlates with neurogranin and BACE-1 Armonk, NY, USA). Graphs were plotted using Graph- To explore the role of contactin-2 in synapse loss, we in- Pad Prism version 6.07. vestigated the relationship of contactin-2 with an estab- lished synaptic biomarker, neurogranin. Contactin-2 Results correlated strongly with CSF neurogranin within controls Contactin-2 CSF levels decreased in AD patients and AD (controls: r =0.62, p = 0.01; AD: r =0.60, p = Demographic and biomarker characteristics of all cases 0.004, Fig. 3a). Furthermore, we analyzed the correlation are listed in Table 1. CSF contactin-2 was reduced by of contactin-2 with its processing enzyme BACE1. Strong 38% in AD patients compared with controls (p < 0.0001; correlations between contactin-2 and CSF BACE1 were Fig. 1a). This result was further validated in a second present within controls and AD (controls: r = 0.64, p = Fig. 1 Contactin-2 levels in the CSF. a. Scatterplot showing CSF contactin-2 levels in nondemented controls with subjective memory complaints (controls, n = 28) and patients with Alzheimer’s disease (AD, n = 36). b. Contactin-2 levels in controls (n = 20) and AD patients (n = 70) in a second validation cohort. The values are presented as medians with interquartile ranges. Data were adjusted for age and gender. *p < 0.05, ***p < 0.0001 Chatterjee et al. Alzheimer's Research & Therapy (2018) 10:52 Page 5 of 11 Fig. 2 Correlations of CSF contactin-2 levels with a. amyloid beta (Aβ)42, b. total tau (tTau), c. phosphorylated tau (pTau), and d. Aβ40. *p < 0.05, **p < 0.01. AD Alzheimer’s disease 0.007; AD: r = 0.46, p =0.04, Fig. 3b). These results were probable plaques are shown by arrowheads). We next ana- validated in the second cohort (Additional file 1:Figure lyzed the potential relationship of contactin-2 expression S3). Since contactin-2 and BACE1 both were associated with the main hallmarks of AD (Fig. 5). Areas with re- with age within the AD group in the second cohort, an duced contactin-2 staining contained deposits of Aβ age correction was applied in the corresponding correl- (Fig. 5a–c), as well as pTau- and Thioflavin S-positive ation analyses (Additional file 1:Figure S3). structures (Fig. 5d–i), which indicates the reduction of contactin-2 staining in fibrillar neuritic plaques. Characterization of contactin-2 in postmortem human Analysis of postmortem tissue homogenates by ELISA hippocampus and temporal cortex confirmed that contactin-2 tended to be decreased in Immunohistochemistry showed that contactin-2 was AD hippocampus (n = 7) compared with controls (n =6, mainly expressed in the extracellular matrix in both con- p = 0.07) (Fig. 6a). However, Western blot analysis re- trol and AD groups in postmortem hippocampus and vealed a significant reduction in contactin-2 levels since temporal cortex (Fig. 4). Interestingly, within the AD cases the expected 113-kDa contactin-2 band decreased in AD we observed a specific reduction in contactin-2 staining in (n = 7) compared with controls (n =5, p = 0.01; Fig. 6b areas resembling amyloid plaques (Fig. 4a, c,where and Additional file 1: Figure S5). Fig. 3 Correlations of CSF contactin-2 with a. CSF neurogranin and b. CSF beta-secretase 1 (BACE1). *p < 0.05, **p < 0.01. AD Alzheimer’s disease Chatterjee et al. Alzheimer's Research & Therapy (2018) 10:52 Page 6 of 11 Fig. 4 Immunohistochemistry on postmortem human brain sections. Brain sections (a,b hippocampus; c,d temporal cortex) of subjects with Alzheimer’s disease (AD) (a,c) and control subjects (b,d) were stained with anti-contactin-2 antibody. Areas with reduced contactin-2 staining are clearly visible in AD brain sections possibly in and around areas with amyloid plaques (shown by arrowheads) Discussion . Considering that synaptic/axonal changes occur in The main finding of this study is that the levels of the very early stages of the disease, it would be of interest to synaptic/axonal protein contactin-2 in the CSF differs explore whether stronger or opposite changes are ob- between AD patients and controls, and is associated served at earlier stages of the disease, and to study its po- with other biomarkers, particularly tTau, pTau, Aβ40, tential as a diagnostic and prognostic marker for early AD. BACE1, and neurogranin. Moreover, we also performed Interestingly, similar to the changes in CSF, contactin-2 characterization of this protein in postmortem human levels were decreased in postmortem brain tissue of AD brain tissue and found areas with reduced contactin-2 cases compared with controls. Our results are supported expression in and around fibrillar neuritic plaques. by a previous study that found a reduction in contactin-2 Synaptic dysfunction and axonal loss are early events in in hippocampal brain tissue homogenates of selected AD AD preceding cognitive decline [5, 7]. Detection of patients with high BACE1 activity compared with changes related to these mechanisms may therefore con- age-matched controls . Therefore, these results not tribute to early diagnosis of the disease. Our findings in only indicate that contactin-2 is changed in the AD brain the CSF reveal that contactin-2 is reduced in AD cases but also that such changes are reflected within the CSF, compared with controls in two cohorts, which challenges highlighting the potential of this protein as a novel bio- previous proteomics findings that identified increased marker for loss of synaptic/axonal integrity. levels of this synaptic protein in three pooled AD CSF Synaptic biomarkers such as neurogranin have been samples . However, the use of specific antibody-based suggested to reflect cognitive decline [18, 51]. In this technologies detecting very specific epitopes of study, we observed a correlation of CSF contactin-2 with contactin-2 in the current study may explain the observed MMSE, suggesting a possible relationship between discrepancies. Even though CSF contactin-2 levels were contactin-2 and cognition. However, this could not be val- lower in AD patients compared with controls, there was a idated in the second cohort. Nonetheless, we found a substantial overlap between the groups in both cohorts strong correlation between contactin-2 and neurogranin, which may limit its diagnostic performance. Contactin-2 supporting the role of contactin-2 in synaptic dysfunction. levels may even be increased in the early stages of AD and CSF contactin-2 correlated with tTau and pTau within then decrease with disease severity as has been shown in the AD/control groups, being stronger within the con- longitudinal analysis of other neuronal injury markers trol group, which suggests that contactin-2 is a sensitive Chatterjee et al. Alzheimer's Research & Therapy (2018) 10:52 Page 7 of 11 Fig. 5 Immunolabeling of contactin-2, Aβ42, and pTau in hippocampal postmortem human AD brain sections. CA subiculum areas of hippocampal sections stained with anti-contactin-2 (a,d), anti-Aβ42 (b), thioflavin S (e), and anti-phosphorylated tau (f). Merged images are shown in c. (contactin-2 + IC16), g. (contactin-2 + thioflavin S), h. (contactin-2 + AT8), and i. (contactin-2 + thioflavin S + AT8). Areas with reduced contactin-2 expression (shown by white arrows) can be seen in AD brain sections in and around areas with neuritic amyloid plaques. Areas marked with blue arrows have been magnified in the inserts marker reflecting general axonal loss and changes in tau protective mechanism to avoid Aβ40 formation in those homeostasis under normal physiological conditions. Im- areas. We did not observe a strong absence of contactin-2 munohistochemical characterization of contactin-2 per- in areas with diffuse plaques (data not shown) that pri- formed in postmortem brain tissue showed a reduction in marily consist of Aβ42 . Similarly, correlation with contactin-2 expression in areas with neuritic amyloid pla- Aβ42 was lacking in the CSF. Taken together, these data ques, characterized by thioflavin S, pTau, and Aβ staining. suggest that contactin-2 can influence the homeostasis of Therefore, similar to the findings in CSF, contactin-2 ex- Aβ which may ultimately affect the formation of amyloid pression was also found to be related with tau in brain tis- deposits and the pathogenesis of AD. The decrease in sue, supporting the potential role of contactin-2 in axonal contactin-2 levels in AD might be a cellular protective loss and incipient neurodegeneration. mechanism to reduce the binding of contactin-2 with APP CSF contactin-2 strongly correlated with Aβ40 and and thus subsequently lowering production of Aβ (Fig. 7). BACE-1, suggesting an association between contactin-2 It should be noted that we observed positive correla- and Aβ production. This is supported by previous studies tions between contactin-2 and tau, BACE1, and neuro- showing that binding of contactin-2 to APP [52, 53]en- granin, which is the opposite to what can be expected hances the production of the APP intracellular domain on the basis of usually increased CSF levels of the latter (AICD) in the cytosol with concomitant Aβ peptide gener- proteins in AD. This indicates that contactin-2 is physio- ation [54, 55](Fig. 7). Interestingly, thioflavin S-positive fi- logically associated with these proteins strongly, demon- brillar plaques that show a stronger presence of Aβ40 strated by high positive correlations within controls, and than Aβ42  had lower contactin-2, probably as a that a disease pathology such as AD possibly disrupts Chatterjee et al. Alzheimer's Research & Therapy (2018) 10:52 Page 8 of 11 Fig. 6 Contactin-2 levels in postmortem hippocampus of Alzheimer’s disease (AD) patients versus controls. a. Contactin-2 concentration measured by ELISA and corrected for total protein concentration. The values are presented as medians with interquartile ranges. b. Western blot showing contactin-2 levels normalized with GAPDH in AD versus controls. Full image of the Western blot is shown in Additional file 1: Figure S5. Unpaired t test was used for group comparisons these associations making the correlations weaker in the towards negative correlation with Tau as expected (co- CSF of AD patients. In addition, these discrepancies may hort 1, r = − 0.23, p = 0.09; Additional file 1: Figure S7). occur because the correlations were analyzed within the CSF BACE was not significantly changed in AD versus AD and control groups separately rather than as a whole controls. Thus, a pattern in correlation may not be cohort. In the whole cohort, there is indeed a tendency evident. Fig. 7 Schematic summary of the hypothesis. Contactin-2 interacts with beta-secretase 1 (BACE1) and amyloid precursor protein (APP). Binding of contactin-2 with APP leads to APP processing and amyloid beta (Aβ) peptide release. Based on our data, we hypothesize that a decrease in contactin-2 levels (shown by a thick dark blue arrow) in AD might be a cellular protective mechanism to reduce the binding of contactin-2 with APP and thus subsequently lowering production of Aβ. Correlations of contactin-2 with total tau (tTau)/phosphorylated tau (pTau) and neurogranin suggest possible interactions among these molecules or their involvement in common pathogenic mechanisms. Solid single/double headed arrows indicate correlations/interactions between two proteins and dashed arrow indicates no correlation. CSF cerebrospinal fluid Chatterjee et al. Alzheimer's Research & Therapy (2018) 10:52 Page 9 of 11 One limitation of this case-control study was the rela- Authors’ contributions MC, MdW, and THJM performed the experiments. MDC prepared brain tissue tively small sample size, even though we eventually in- samples and provided various protocols. WMvdF provided CSF samples and cluded 106 AD patients and 50 controls in the total the patient database, and JJMH provided the postmortem patient samples. group. Controls with subjective memory complaints, AD MC, MDC, THJM, JHJM, and CET analyzed the patient data, and MC, MDC, WMvdF, JJMH, and CET wrote the paper. MC, MDC, JJMH, and CET designed patients, or patients with other neurodegenerative disor- the research study. All authors read and approved the final manuscript. ders often present similar clinical symptoms  which might obscure the differences between the different clin- Ethics approval and consent to participate The ethical review board of the VU Medical Center approved the study, and ical groups. However, AD patients were selected by clini- all subjects provided written informed consent. This manuscript does not cians from a specialized memory center based on the contain individual/personal details of subjects. cut-off value for CSF tau to Aβ42 ratio  ensuring a Consent for publication more reliable diagnosis based on fluid biomarkers . Not applicable as this manuscript does not contain individual/personal Another limitation of the study was that no cohort was details of subjects. used from another memory clinic. It would be interest- Competing interests ing to investigate the levels of contactin-2 in larger inde- The authors declare that they have no competing interests. pendent cohorts and in AD patients in different stages of the disease. Publisher’sNote Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Conclusions In summary, this study reveals a reduction in the axonal Author details and synaptic protein contactin-2 in two CSF cohorts and Neurochemistry Laboratory, Clinical Chemistry Department, VU University Medical Center, De Boelelaan 1117, 1081 HV Amsterdam, the Netherlands. postmortem tissue, and indicates the potential of this Department of Pathology, VU University Medical Center, Amsterdam, the protein as a novel AD CSF biomarker reflecting synap- Netherlands. Alzheimer Center, VU University Medical Center, Amsterdam, tic/axonal dysfunction. Future studies should investigate the Netherlands. Department of Epidemiology & Biostatistics, VU University Medical Center, Amsterdam, the Netherlands. how contactin-2 is changed during the course of AD in a longitudinal study design with larger patient cohorts. Received: 22 January 2018 Accepted: 9 May 2018 In addition, studies revealing a mechanistic relation be- tween contactin-2, Aβ, and tau are required to under- References stand the bigger picture of the cell signaling pathway 1. Masters CL, Bateman R, Blennow K, Rowe CC, Sperling RA, Cummings JL. underlying AD pathogenesis and to open new leads for Alzheimer’s disease. Nat Rev Dis Primers. 2015;1:15056. https://doi.org/10. 1038/nrdp.2015.56. therapy development. 2. Dubois B, Feldman HH, Jacova C, Cummings JL, DeKosky ST, Barberger-Gateau P, Delacourte A, Frisoni G, Fox NC, Galasko D, Gauthier S, Hampel H, Jicha GA, Meguro K, O’Brien J, Pasquier F, Robert P, Rossor M, Salloway S, Sarazin M, de Additional file: Souza LC, Stern Y, Visser PJ, Scheltens P. Revising the definition of Alzheimer’s disease: a new lexicon. Lancet Neurol. 2010;9:1118–27. https://doi.org/10.1016/ Additional file 1: Supplementary methods and results. (DOCX 645 kb) S1474-4422(10)70223-4. 3. Shaw LM, Vanderstichele H, Knapik-Czajka M, Clark CM, Aisen PS, Petersen RC, Blennow K, Soares H, Simon A, Lewczuk P, Dean R, Siemers E, Potter W, Abbreviations Lee VMY, Trojanowski JQ. Cerebrospinal fluid biomarker signature in AD: Alzheimer’s disease; AICD: Amyloid precursor protein intracellular alzheimer’s disease neuroimaging initiative subjects. Ann Neurol. 2009;65(4): domain; APP: Amyloid precursor protein; Aβ: Amyloid beta; BACE1: Beta- 403–13. https://doi.org/10.1002/ana.21610. secretase 1; CSF: Cerebrospinal fluid; CV: Coefficient of variation; 4. De Meyer G, Shapiro F, Vanderstichele H, Vanmechelen E, Engelborghs S, De ELISA: Enzyme-linked immunosorbent assay; IF: Immunofluorescence; Deyn PP, Coart E, Hansson O, Minthon L, Zetterberg H, Blennow K, Shaw L, IHC: Immunohistochemistry; MMSE: Mini-Mental State Examination; Trojanowski JQ. Alzheimer’s disease neuroimaging initiative, diagnosis- pTau: Phosphorylated tau; tTau: Total tau independent alzheimer disease biomarker signature in cognitively normal elderly people. Arch Neurol. 2010;67:949. https://doi.org/10.1001/archneurol.2010.179. Acknowledgements 5. Selkoe DJ. Alzheimer’s disease is a synaptic failure. Science (80- ). 2002;298: The authors thank Marije Benedictus for selecting patients for the different 789–91. https://doi.org/10.1126/science.1074069. cohorts, Johnny Aarnoutse for performing the neurogranin assay, Vera 6. Portelius E, Zetterberg H, Skillbäck T, Törnqvist U, Andreasson U, Trojanowski Wiersma for assistance with confocal microscopy, Harry Twaalfhoven for JQ, Weiner MW, Shaw LM, Mattsson N, Blennow K. Cerebrospinal fluid helping analyze contactin-2 ELISA validation data, and M.J. Koel-Simmelink neurogranin: relation to cognition and neurodegeneration in Alzheimer’s for designing the ELISA validation protocol. disease. Brain. 2015;138(Pt 11):3373–85. https://doi.org/10.1093/brain/ awv267. Funding 7. Terwel D, Dewachter I, Van Leuven F. Axonal transport, tau protein, and This project was funded by the European Neuroscience Campus Network, an neurodegeneration in Alzheimer’s disease. Neuromolecular Med. 2002;2(2): Erasmus Mundus Joint Doctoral Program (cycle 5/2014/P-04). The funding 151–65. https://doi.org/10.1385/NMM:2:2:151. sources had no role in the design of the study, the collection, analysis, and 8. Ossenkoppele R, Jansen WJ, Rabinovici GD, Knol DL, van der Flier WM, van interpretation of data, or in writing the manuscript. Berckel BNM, Scheltens P, Visser PJ, Verfaillie SCJ, Zwan MD, Adriaanse SM, Lammertsma AA, Barkhof F, Jagust WJ, Miller BL, Rosen HJ, Landau SM, Availability of data and materials Villemagne VL, Rowe CC, Lee DY, Na DL, Seo SW, Sarazin M, Roe CM, Sabri O, The datasets used and/or analyzed during the current study are available BarthelH,KoglinN,Hodges J, Leyton CE,Vandenberghe R, van Laere K,Drzezga from the corresponding author on reasonable request. A, Forster S, Grimmer T, Sánchez-Juan P, Carril JM, Mok V, Camus V, Klunk WE, Chatterjee et al. Alzheimer's Research & Therapy (2018) 10:52 Page 10 of 11 Cohen AD, Meyer PT, Hellwig S, Newberg A, Frederiksen KS, Fleisher AS, Mintun neurite outgrowth-promoting activity. Cell. 1990;61:157–70. https://doi.org/ MA,WolkDA,Nordberg A, Rinne JO,Chételat G,LleoA,Blesa R, Fortea J,Madsen 10.1016/0092-8674(90)90223-2. K, Rodrigue KM, Brooks DJ. Prevalence of amyloid PET positivity in dementia 27. Ogawa J, Lee S, Itoh K, Nagata S, Machida T, Takeda Y, Watanabe K. Neural syndromes. JAMA. 2015;313:1939. https://doi.org/10.1001/jama.2015.4669. recognition molecule NB-2 of the contactin/F3 subgroup in rat: specificity in 9. DeKosky ST, Scheff SW. Synapse loss in frontal cortex biopsies in Alzheimer’s neurite outgrowth-promoting activity and restricted expression in the brain disease: correlation with cognitive severity. Ann Neurol. 1990;27:457–64. regions. J Neurosci Res. 2001;65:100–10. http://www.ncbi.nlm.nih.gov/ https://doi.org/10.1002/ana.410270502. pubmed/11438979. 10. Hamos JE, DeGennaro LJ, Drachman DA. Synaptic loss in Alzheimer’s 28. Karagogeos D, Morton SB, Casano F, Dodd J, Jessell TM. Developmental disease and other dementias. Neurology. 1989;39(3):355–61. https://www. expression of the axonal glycoprotein TAG-1: differential regulation by ncbi.nlm.nih.gov/pubmed/2927643. central and peripheral neurons in vitro. Development. 1991;112:51–67. http://www.ncbi.nlm.nih.gov/pubmed/1769341. 11. Teunissen CE, Parnetti L. New CSF biomarkers on the block. EMBO Mol Med. 29. Frei JA, Stoeckli ET. SynCAMs extend their functions beyond the synapse. 2016;8:1118–9. https://doi.org/10.15252/emmm.201606801. Eur J Neurosci. 2014;39(11):1752–60. https://doi.org/10.1111/ejn.12544. 12. Blennow K, Zetterberg H. The past and the future of Alzheimer’sdisease CSF 30. Murai KK, Misner D, Ranscht B. Contactin supports synaptic plasticity biomarkers—a journey toward validated biochemical tests covering the whole associated with hippocampal long-term depression but not potentiation. spectrum of molecular events. Front Neurosci. 2015;9:345. https://doi.org/10.3389/ Curr Biol. 2002;12:181–90. http://www.ncbi.nlm.nih.gov/pubmed/11839269. fnins.2015.00345. 31. Gautam V, D’Avanzo C, Hebisch M, Kovacs DM, Kim DY. BACE1 activity 13. Davies CA, Mann DM, Sumpter PQ, Yates PO. A quantitative morphometric regulates cell surface contactin-2 levels. Molecular Neurodegeneration. analysis of the neuronal and synaptic content of the frontal and temporal 2014;9:4. https://doi.org/10.1186/1750-1326-9-4. cortex in patients with Alzheimer’s disease. J Neurol Sci. 1987;78(2):151-64. 32. Stoeckli ET. Neural circuit formation in the cerebellum is controlled by cell https://www.ncbi.nlm.nih.gov/pubmed/3572454. adhesion molecules of the contactin family. Cell Adhes Migr. 2010;4(4):523– 14. Blennow K, Bogdanovic N, Alafuzoff I, Ekman R, Davidsson P. Synaptic 526. https://doi.org/10.4161/cam.4.4.12733. pathology in Alzheimer’s disease: relation to severity of dementia, but not 33. Frei JA, Stoeckli ET. SynCAMs—from axon guidance to neurodevelopmental to senile plaques, neurofibrillary tangles, or the ApoE4 allele. J Neural disorders. Mol Cell Neurosci. 2017;81:41–8. https://doi.org/10.1016/j.mcn.2016.08.012. Transm (Vienna). 1996;103(5):603–18. https://www.ncbi.nlm.nih.gov/ 34. Wolman MA, Sittaramane VK, Essner JJ, Yost HJ, Chandrasekhar A, Halloran pubmed/8811505 MC. Transient axonal glycoprotein-1 (TAG-1) and laminin-alpha1 regulate 15. Sze CI, Troncoso JC, Kawas C, Mouton P, Price DL, Martin LJ. Loss of the dynamic growth cone behaviors and initial axon direction in vivo. Neural presynaptic vesicle protein synaptophysin in hippocampus correlates with Development 2008;3:6. https://doi.org/10.1186/1749-8104-3-6. cognitive decline in Alzheimer disease. J Neuropathol Exp Neurol. 1997; 35. Traka M, Dupree JL, Popko B, Karagogeos D. The neuronal adhesion protein 56(8):933–44. https://www.ncbi.nlm.nih.gov/pubmed/9258263. TAG-1 is expressed by Schwann cells and oligodendrocytes and is localized to 16. Masliah E, Mallory M, Alford M, DeTeresa R, Hansen LA, McKeel DW, Morris the juxtaparanodal region of myelinated fibers. J Neurosci. 2002;22(8):3016–24. JC. Altered expression of synaptic proteins occurs early during progression 36. Suter DM, Pollerberg GE, Buchstaller A, Giger RJ, Dreyer WJ, Sonderegger P. of Alzheimer’s disease. Neurology. 2001;56(1):127–9. https://www.ncbi.nlm. Binding between the neural cell adhesion molecules axonin-1 and Nr-CAM/ nih.gov/pubmed/11148253 Bravo is involved in neuron-glia interaction. J Cell Biol. 1995;131:1067–81. 17. Bereczki E, Francis PT, Howlett D, Pereira JB, Höglund K, Bogstedt A, Cedazo- http://www.ncbi.nlm.nih.gov/pubmed/7490283. Minguez A, Baek J-H, Hortobágyi T, Attems J, Ballard C, Aarsland D. Synaptic 37. Yin GN, Lee HW, Cho J-Y, Suk K. Neuronal pentraxin receptor in proteins predict cognitive decline in Alzheimer’s disease and Lewy body cerebrospinal fluid as a potential biomarker for neurodegenerative diseases. dementia. Alzheimers Dement. 2016;12:1149–58. https://doi.org/10.1016/j. Brain Res. 2009;1265:158–70. https://doi.org/10.1016/j.brainres.2009.01.058. jalz.2016.04.005. 18. Kester MI, Teunissen CE, Crimmins DL, Herries EM, Ladenson JH, Scheltens P, van 38. Van Der Flier WM, Pijnenburg YAL, Prins N, Lemstra AW, Bouwman FH, der Flier WM, Morris JC, Holtzman DM, Fagan AM. Neurogranin as a Teunissen CE, Van Berckel BNM, Stam CJ, Barkhof F, Visser PJ, Van Egmond cerebrospinal fluid biomarker for synaptic loss in symptomatic Alzheimer disease. E, Scheltens P. Optimizing patient care and research: The Amsterdam JAMA Neurol. 2015;72:1275–80. https://doi.org/10.1001/jamaneurol.2015.1867. dementia cohort. J Alzheimers Dis. 2014;41(1):313–27. https://doi.org/10. 19. Davidsson P, Jahn R, Bergquist J, Ekman R, Blennow K. Synaptotagmin, a 3233/JAD-132306. synaptic vesicle protein, is present in human cerebrospinal fluid. Mol Chem 39. McKhann G, Drachman D, Folstein M, Katzman R, Price D, Stadlan EM. Neuropathol. 1996;27:195–210. https://doi.org/10.1007/BF02815094. Clinical diagnosis of Alzheimer’s disease: report of the NINCDS-ADRDA Work Group under the auspices of Department of Health and Human Services 20. Brinkmalm A, Brinkmalm G, Honer WG, Frölich L, Hausner L, Minthon L, Hansson Task Force on Alzheimer’s Disease. Neurology. 1984;34:939–44. http://www. O, Wallin A, Zetterberg H, Blennow K, Öhrfelt A. SNAP-25 is a promising novel ncbi.nlm.nih.gov/pubmed/6610841. cerebrospinal fluid biomarker for synapse degeneration in Alzheimer’sdisease. 40. McKhann GM, Knopman DS, Chertkow H, Hyman BT, Jack CR, Kawas CH, Mol Neurodegener. 2014;9:53. https://doi.org/10.1186/1750-1326-9-53. Klunk WE, Koroshetz WJ, Manly JJ, Mayeux R, Mohs RC, Morris JC, Rossor 21. Wellington H, Paterson RW, Portelius E, Törnqvist U, Magdalinou N, Fox NC, MN, Scheltens P, Carrillo MC, Thies B, Weintraub S, Phelps CH. The diagnosis Blennow K, Schott JM, Zetterberg H. Increased CSF neurogranin of dementia due to Alzheimer’s disease: Recommendations from the concentration is specific to Alzheimer disease. Neurology. 2016;86(9):829–35. National Institute on Aging-Alzheimer’s Association workgroups on https://doi.org/10.1212/WNL.0000000000002423. diagnostic guidelines for Alzheimer’s disease. Alzheimers Dement. 2011;7(3): 22. Schraen-Maschke S, Sergeant N, Dhaenens C-M, Bombois S, Deramecourt V, 263–9. https://doi.org/10.1016/j.jalz.2011.03.005. Caillet-Boudin M-L, Pasquier F, Maurage C-A, Sablonnière B, Vanmechelen E, 41. Duits FH, Teunissen CE, Bouwman FH, Visser PJ, Mattsson N, Zetterberg H, Buée L. Tau as a biomarker of neurodegenerative diseases. Biomark Med. Blennow K, Hansson O, Minthon L, Andreasen N, Marcusson J, Wallin A, Rikkert 2008;2:363–84. https://doi.org/10.2217/175203220.127.116.113. MO, Tsolaki M, Parnetti L, Herukka SK, Hampel H, De Leon MJ, Schröder J, 23. Lu Z, Reddy MVVVS, Liu J, Kalichava A, Liu J, Zhang L, Chen F, Wang Y, Aarsland D, Blankenstein MA, Scheltens P, Van Der Flier WM. The cerebrospinal Holthauzen LMF, White MA, Seshadrinathan S, Zhong X, Ren G, Rudenko G. fluid “alzheimer profile”: easily said, but what does it mean? Alzheimers Molecular architecture of contactin-associated protein-like 2 (CNTNAP2) and its Dement. 2014;10(6):713–723.e2. https://doi.org/10.1016/j.jalz.2013.12.023. interaction with contactin 2 (CNTN2). J Biol Chem. 2016;291(46):24133–24147. https://doi.org/10.1074/jbc.M116.748236. 42. Bouwman FH, Schoonenboom NSM, Verwey NA, van Elk EJ, Kok A, Blankenstein MA, Scheltens P, van der Flier WM. CSF biomarker levels in 24. Masuda T. Contactin-2/TAG-1, active on the front line for three decades. Cell early and late onset Alzheimer’s disease. Neurobiol Aging. 2009;30:1895–901. Adh Migr. 2017;11(5-6):524–531. https://doi.org/10.1080/19336918.2016. https://doi.org/10.1016/j.neurobiolaging.2008.02.007. 43. Masuda T. Contactin-2/TAG-1, active on the front line for three decades 25. Ranscht B. Sequence of contactin, a 130-kD glycoprotein concentrated in contactin-2/TAG-1, active on the front line for three decades. Cell Adh Migr. areas of interneuronal contact, defines a new member of the 2017;11(5-6):524–531. https://doi.org/10.1080/19336918.2016.1269998. immunoglobulin supergene family in the nervous system. J Cell Biol. 1988 44. Serrano-Pozo A, Frosch MP, Masliah E, Hyman BT. Neuropathological Oct;107(4):1561–73. https://doi.org/10.1083/jcb.107.4.1561. alterations in Alzheimer disease. Cold Spring Harb Perspect Med. 2011;1(1): 26. Furley AJ, Morton SB, Manalo D, Karagogeos D, Dodd J, Jessell TM. The a006189. https://doi.org/10.1101/cshperspect.a006189. axonal glycoprotein TAG-1 is an immunoglobulin superfamily member with Chatterjee et al. Alzheimer's Research & Therapy (2018) 10:52 Page 11 of 11 45. Andreasson U, Perret-Liaudet A, van Waalwijk van Doorn LJC, Blennow K, Chiasserini D, Engelborghs S, Fladby T, Genc S, Kruse N, Kuiperij HB, Kulic L, Lewczuk P, Mollenhauer B, Mroczko B, Parnetti L, Vanmechelen E, Verbeek MM, Winblad B, Zetterberg H, Koel-Simmelink M, Teunissen CE. A practical guide to immunoassay method validation. Front Neurol. 2015;6:1–8. https://doi.org/10.3389/fneur.2015.00179. 46. van Waalwijk van Doorn LJC, Koel-Simmelink MJ, Haußmann U, Klafki H, Struyfs H, Linning P, Knölker H-J, Twaalfhoven H, Kuiperij HB, Engelborghs S, Scheltens P, Verbeek MM, Vanmechelen E, Wiltfang J, Teunissen CE. Validation of soluble amyloid-β precursor protein assays as diagnostic CSF biomarkers for neurodegenerative diseases. J Neurochem. 2016;137:112–21. https://doi.org/10.1111/jnc.13527. 47. Mulder C, Verwey NA, van der Flier WM, Bouwman FH, Kok A, van Elk EJ, Scheltens P, Blankenstein MA. Amyloid-beta(1-42), total tau, and phosphorylated tau as cerebrospinal fluid biomarkers for the diagnosis of Alzheimer disease. Clin Chem. 2010;56:248–53. https://doi.org/10.1373/ clinchem.2009.130518. 48. Chatterjee M, Nöding B, Willemse EAJ, Koel-Simmelink MJA, Van Der Flier WM, Schild D, Teunissen CE. Detection of contactin-2 in cerebrospinal fluid (CSF) of patients with Alzheimer’s disease using Fluorescence Correlation Spectroscopy (FCS). Clin Biochem. 2017;50(18):1061–1066. https://doi.org/10. 1016/j.clinbiochem.2017.08.017. 49. Templeton GF. A Two-Step Approach for Transforming Continuous Variables to Normal: Implications and Recommendations for IS Research. Communications of the Association for Information Systems: Vol. 28 , Article 4. http://aisel.aisnet.org/cais/vol28/iss1/4 50. C.L. Sutphen, L. McCue, E.M. Herries, C. Xiong, J.H. Ladenson, D.M. Holtzman, A.M. Fagan. Longitudinal decreases in multiple cerebrospinal fluid biomarkers of neuronal injury in symptomatic late onset Alzheimer’s disease. Alzheimers Dement. 2018. https://doi.org/10.1016/j.jalz.2018.01.012. 51. De Vos A, Struyfs H, Jacobs DI, Fransen E, Klewansky T, De Roeck E, Robberecht C, Van Broeckhoven C, Duyckaerts C, Engelborghs S, Vanmechelen E. The cerebrospinal fluid neurogranin/BACE1 ratio is a potential correlate of cognitive decline in Alzheimer’s disease. J Alzheimers Dis. 2016.https://doi.org/10.3233/JAD-160227. 52. Ma QH, Bagnard D, Xiao ZC, Dawe GS. A TAG on to the neurogenic functions of APP. Cell Adh Migr. 2008;2(1):2–8. https://doi.org/10.4161/cam.2.1.5790. 53. Mattson MP, Van Praag H. TAGing APP constrains neurogenesis. Nat Cell Biol. 2008;10(3):249–50. https://doi.org/10.1038/ncb0308-249. 54. Ma Q-H, Futagawa T, Yang W-L, Jiang X-D, Zeng L, Takeda Y, Xu R-X, Bagnard D, Schachner M, Furley AJ, Karagogeos D, Watanabe K, Dawe GS, Xiao Z-C. A TAG1-APP signalling pathway through Fe65 negatively modulates neurogenesis. Nat Cell Biol. 2008;10(3):283–94. https://doi.org/10. 1038/ncb1690. 55. Konietzko U. AICD nuclear signaling and its possible contribution to Alzheimer’s disease. Curr Alzheimer Res. 2012;9:200–16. https://www.ncbi. nlm.nih.gov/pubmed/21605035. 56. Irvine GB, El-Agnaf OM, Shankar GM, Walsh DM. Protein aggregation in the brain: the molecular basis for Alzheimer’s and Parkinson’s diseases. Mol Med. 2008;14:451–64. https://doi.org/10.2119/2007-00100.Irvine. 57. Kovacs GG, Milenkovic I, Wöhrer A, Höftberger R, Gelpi E, Haberler C, Hönigschnabl S, Reiner-Concin A, Heinzl H, Jungwirth S, Krampla W, Fischer P, Budka H. Non-Alzheimer neurodegenerative pathologies and their combinations are more frequent than commonly believed in the elderly brain: a community-based autopsy series. Acta Neuropathol. 2013; 126:365–84. https://doi.org/10.1007/s00401-013-1157-y. 58. Coart E, Barrado LG, Duits FH, Scheltens P, van der Flier WM, Teunissen CE, van der Vies SM, Burzykowski T. Alzheimer’s disease neuroimaging initiative: correcting for the absence of a gold standard improves diagnostic accuracy of biomarkers in Alzheimer’s disease. J Alzheimers Dis. 2015;46:889–99. https://doi.org/10.3233/JAD-142886.
Alzheimer's Research & Therapy – Springer Journals
Published: Jun 1, 2018
It’s your single place to instantly
discover and read the research
that matters to you.
Enjoy affordable access to
over 18 million articles from more than
15,000 peer-reviewed journals.
All for just $49/month
Query the DeepDyve database, plus search all of PubMed and Google Scholar seamlessly
Save any article or search result from DeepDyve, PubMed, and Google Scholar... all in one place.
Get unlimited, online access to over 18 million full-text articles from more than 15,000 scientific journals.
Read from thousands of the leading scholarly journals from SpringerNature, Elsevier, Wiley-Blackwell, Oxford University Press and more.
All the latest content is available, no embargo periods.
“Hi guys, I cannot tell you how much I love this resource. Incredible. I really believe you've hit the nail on the head with this site in regards to solving the research-purchase issue.”Daniel C.
“Whoa! It’s like Spotify but for academic articles.”@Phil_Robichaud
“I must say, @deepdyve is a fabulous solution to the independent researcher's problem of #access to #information.”@deepthiw
“My last article couldn't be possible without the platform @deepdyve that makes journal papers cheaper.”@JoseServera