SNAP-25 Research Articles

Plasma extracellular vesicle synaptic proteins as biomarkers of clinical progression in patients with Parkinson’s disease

Synaptic dysfunction plays a key role in Parkinson’s disease (PD), and plasma extracellular vesicle (EV) synaptic proteins are emerging as biomarkers for neurodegenerative diseases. Assessment of plasma EV synaptic proteins for their efficacy as biomarkers in PD and their relationship with disease progression was conducted. In total, 144 participants were enrolled, including 101 people with PD (PwP) and 43 healthy controls (HCs). The changes in plasma EV synaptic protein levels between baseline and 1-year follow-up did not differ significantly in both PwP and HCs. In PwP, the changes in plasma EV synaptic protein levels were significantly associated with the changes in Unified Parkinson’s Disease Rating Scale (UPDRS)-II and III scores. Moreover, PwP with elevated levels (first quartile) of any one plasma EV synaptic proteins (synaptosome-associated protein 25, growth-associated protein 43 or synaptotagmin-1) had significantly greater disease progression in UPDRS-II score and the postural instability and gait disturbance subscore in UPDRS-III than did the other PwP after adjustment for age, sex, and disease duration. The promising potential of plasma EV synaptic proteins as clinical biomarkers of disease progression in PD was suggested. However, a longer follow-up period is warranted to confirm their role as prognostic biomarkers.

Correlation of Presynaptic and Postsynaptic Proteins with Pathology in Alzheimer's Disease.

Synaptic transmission is essential for nervous system function and the loss of synapses is a known major contributor to dementia. Alzheimer's disease dementia (ADD) is characterized by synaptic loss in the mesial temporal lobe and cerebral neocortex, both of which are brain areas associated with memory and cognition. The association of synaptic loss and ADD was established in the late 1980s, and it has been estimated that 30-50% of neocortical synaptic protein is lost in ADD, but there has not yet been a quantitative profiling of different synaptic proteins in different brain regions in ADD from the same individuals. Very recently, positron emission tomography (PET) imaging of synapses is being developed, accelerating the focus on the role of synaptic loss in ADD and other conditions. In this study, we quantified the densities of two synaptic proteins, the presynaptic protein Synaptosome Associated Protein 25 (SNAP25) and the postsynaptic protein postsynaptic density protein 95 (PSD95) in the human brain, using enzyme-linked immunosorbent assays (ELISA). Protein was extracted from the cingulate gyrus, hippocampus, frontal, primary visual, and entorhinal cortex from cognitively unimpaired controls, subjects with mild cognitive impairment (MCI), and subjects with dementia that have different levels of Alzheimer's pathology. SNAP25 is significantly reduced in ADD when compared to controls in the frontal cortex, visual cortex, and cingulate, while the hippocampus showed a smaller, non-significant reduction, and entorhinal cortex concentrations were not different. In contrast, all brain areas showed lower PSD95 concentrations in ADD when compared to controls without dementia, although in the hippocampus, this failed to reach significance. Interestingly, cognitively unimpaired cases with high levels of AD pathology had higher levels of both synaptic proteins in all brain regions. SNAP25 and PSD95 concentrations significantly correlated with densities of neurofibrillary tangles, amyloid plaques, and Mini Mental State Examination (MMSE) scores. Our results suggest that synaptic transmission is affected by ADD in multiple brain regions. The differences were less marked in the entorhinal cortex and the hippocampus, most likely due to a ceiling effect imposed by the very early development of neurofibrillary tangles in older people in these brain regions.

Chiral Supramolecular Nanofibers Regulated Tumor-Derived Exosomes Secretion for Constructing an Anti-Tumor Extracellular Microenvironment.

Tumor-derived exosomes (TDEs) induced extracellular microenvironment has recently been validated to be critical for tumor progression and metastasis, however, remodeling it for oncotherapy still remains a major challenge due to difficulty in regulation of TDEs secretion. Herein, the supramolecular chiral nanofibers, composed of L/D-phenylalanine derivates (L/D-Phe) and linear hyaluronic acid (HA), are successfully employed to construct TDEs induced anti-tumor extracellular microenvironment. The left-handed L-Phe @HA nanofibers significantly inhibit TDEs secretion into extracellular microenvironment, which results in suppression of tumor proliferation and metastasis in vitro and vivo. Biological assays and theoretical modeling reveal that these results are mainly attributed to strong adsorption of the key exosomes transporters (Ras-related protein Rab-27A and synaptosome-associated protein 23) on left-handed L-Phe @HA nanofibers via enhanced stereoselective interaction, leading to degradation and phosphorylated dropping of exosomes transporters. Subsequently, transfer function of exosomes transporters is limited, which causes remarkable inhibition of TDEs secretion. These findings provide a promising novel insight of chiral functional materials to establish an anti-tumor extracellular microenvironment via regulation of TDEs secretion.

The use of synaptic biomarkers in cerebrospinal fluid to differentiate behavioral variant of frontotemporal dementia from primary psychiatric disorders and Alzheimer’s disease

BackgroundLack of early molecular biomarkers in sporadic behavioral variants of frontotemporal dementia (bvFTD) and its clinical overlap with primary psychiatric disorders (PPD) hampers its diagnostic distinction. Synaptic dysfunction is an early feature in bvFTD and identification of specific biomarkers might improve its diagnostic accuracy. Our goal was to understand the differential diagnostic potential of cerebrospinal fluid (CSF) synaptic biomarkers in bvFTD versus PPD and their specificity towards bvFTD compared with Alzheimer’s disease (AD) and controls. Additionally, we explored the association of CSF synaptic biomarkers with social cognition, cognitive performance, and disease severity in these clinical groups.MethodsParticipants with probable bvFTD (n = 57), PPD (n = 71), AD (n = 60), and cognitively normal controls (n = 39) with available CSF, cognitive tests, and disease severity as frontotemporal lobar degeneration-modified clinical dementia rating scale (FTLD-CDR) were included. In a subset of bvFTD and PPD cases, Ekman 60 faces test scores for social cognition were available. CSF synaptosomal-associated protein 25 (SNAP25), neurogranin (Ng), neuronal pentraxin 2 (NPTX2), and glutamate receptor 4 (GluR4) were measured, along with neurofilament light (NfL), and compared between groups using analysis of covariance (ANCOVA) and logistic regression. Diagnostic accuracy was assessed using ROC analyses, and biomarker panels were selected using Wald’s backward selection. Correlations with cognitive measures were performed using Pearson’s partial correlation analysis.ResultsNPTX2 concentrations were lower in the bvFTD group compared with PPD (p < 0.001) and controls (p = 0.003) but not compared with AD. Concentrations of SNAP25 (p < 0.001) and Ng (p < 0.001) were elevated in patients with AD versus those with bvFTD and controls. The modeled panel for differential diagnosis of bvFTD versus PPD consisted of NfL and NPTX2 (AUC = 0.96, CI: 0.93–0.99, p < 0.001). In bvFTD versus AD, the modeled panel consisted of NfL, SNAP25, Ng, and GluR4 (AUC = 0.86, CI: 0.79–0.92, p < 0.001). In bvFTD, lower NPTX2 (Pearson’s r = 0.29, p = 0.036) and GluR4 (Pearson’s r = 0.34, p = 0.014) concentrations were weakly associated with worse performance of total cognitive score. Lower GluR4 concentrations were also associated with worse MMSE scores (Pearson’s r = 0.41, p = 0.002) as well as with worse executive functioning (Pearson’s r = 0.36, p = 0.011) in bvFTD. There were no associations between synaptic markers and social cognition or disease severity in bvFTD.ConclusionOur findings of involvement of NTPX2 in bvFTD but not PPD contribute towards better understanding of bvFTD disease pathology.

The clarithromycin-binding proteins NIPSNAP1 and 2 regulate cytokine production through mitochondrial quality control

The mechanism underlying the anti-inflammatory effect of macrolide antibiotics, such as clarithromycin (CAM), remains to be clarified. The CAM-binding proteins 4-nitrophenylphosphatase domain and non-neuronal synaptosomal associated protein 25 (SNAP25)-like protein homolog (NIPSNAP) 1 and 2 are involved in the immune response and mitochondrial homeostasis. However, the axis between CAM-NIPSNAP-mitochondria and Toll-like receptor (TLR) and their molecular mechanisms remain unknown. In this study, we sought to elucidate the relationship between mitochondrial homeostasis mediated by NIPSNAP1 and 2 and the immunomodulatory effect of CAM. NIPSNAP1 or 2 knockdown (KD) by RNA interference impaired TLR4-mediated interleukin-8 (IL-8) production. Similar impairment was observed upon treatment with mitochondrial function inhibitors. However, IL-8 secretion was not impaired in NIPSNAP1 and 2 individual knockout (KO) and double KO (DKO) cells. Moreover, the oxygen consumption rate (OCR) in mitochondria measured using a flex analyzer was significantly reduced in NIPSNAP1 or 2 KD cells, but not in DKO cells. CAM also dose-dependently reduced the OCR. These results indicate that CAM suppresses the IL-8 production via the mitochondrial quality control regulated by temporary functional inhibition of NIPSNAP1 and 2. Our findings provide new insight into the mechanisms underlying cytokine production, including the TLR-mitochondria axis, and the immunomodulatory effects of macrolides.

VAMP5 promotes Fcγ receptor-mediated phagocytosis and regulates phagosome maturation in macrophages.

Phagosome formation and maturation reportedly occur via sequential membrane fusion events mediated by synaptosomal-associated protein of 23 kDa (SNAP23), a plasma membrane-localized soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) family. Vesicle-associated membrane protein 5 (VAMP5), also a plasmalemma SNARE, interacts with SNAP23; however, its precise function in phagocytosis in macrophages remains elusive. To elucidate this aspect, we investigated the characteristics of macrophages in the presence of VAMP5 overexpression or knockdown and found that VAMP5 participates in Fcγ receptor-mediated phagosome formation, although not directly in phagosome maturation. Overexpressed VAMP5 was localized to the early phagosomal membrane but no longer localized to the lysosomal-associated membrane protein 1-positive maturing phagosomal membrane. Analyses using compound-based selective inhibitors demonstrated that VAMP5 dissociation from early phagosomes occurs in a clathrin- and dynamin-dependent manner and is indispensable for SNAP23 function in subsequent membrane fusion during phagosome maturation. Accordingly, to the best of our knowledge, we demonstrate, for the first time, that VAMP5 exerts an immunologically critical function during phagosome formation and maturation via SNARE-based membrane trafficking in macrophages.

Putative role of endothelin receptor B in the development and maintenance of taste buds within the circumvallate papillae

This study aimed to achieve a better understanding of taste receptor cell development relative to endothelin receptor B (ETB) in circumvallate papillae (CVP). ETB localization was assessed by immunohistochemistry during tongue development of the mouse. Co-localization of ETB with taste receptor type III cell marker, Synaptosomal-Associated Protein 25 kDa (SNAP25), was evident in both the developing and adult CVP. ETB was strongly localized in the stromal core region. As development progressed, ETB became localized in the CVP mesenchyme and partially in the epithelium. ETB and SNAP25 co-localization indicates that ETB may regulate innervation from the CVP mesenchyme to taste buds.

Insights into early pathogenesis of sporadic Alzheimer's disease: role of oxidative stress and loss of synaptic proteins.

Oxidative stress, induced by impaired insulin signaling in the brain contributes to cognitive loss in sporadic Alzheimer's disease (sAD). This study evaluated early hippocampal oxidative stress, pre- and post-synaptic proteins in intraperitoneal (IP) and intracerebroventricular (ICV) streptozotocin (STZ) models of impaired insulin signaling. Adult male Wistar rats were injected with STZ, IP, or ICV, and sacrificed 1-, 3-, or 6-weeks post injection. Rat's cognitive behavior was assessed using Morris water maze (MWM) tests at weeks 3 and 6. Hippocampal synaptosomal fractions were examined for oxidative stress markers and presynaptic [synapsin I, synaptophysin, growth-associated protein-43 (GAP-43), synaptosomal-associated protein-25 (SNAP-25)] and postsynaptic [drebrin, synapse-associated protein-97 (SAP-97), postsynaptic density protein-95 (PSD-95)] proteins. IP-STZ and ICV-STZ treatment impaired rat's cognition, decreased the levels of reduced glutathione (GSH) and increased the levels of thiobarbituric acid reactive species (TBARS) in a time dependent manner. In addition, it reduced the expression of pre- and post-synaptic proteins in the hippocampus. The decline in cognition is significantly correlated with the reduction in synaptic proteins in the hippocampus. In conclusion, impaired insulin signaling in the brain is deleterious in causing early synaptosomal oxidative damage and synaptic loss that exacerbates with time and correlates with cognitive impairments. Our data implicates oxidative stress and synaptic protein loss as an early feature of sAD and provides insights into early biochemical and behavioral changes during disease progression.

SNAP-25, but not SNAP-23, is essential for photoreceptor development, survival, and function in mice

SNARE-mediated vesicular transport is thought to play roles in photoreceptor glutamate exocytosis and photopigment delivery. However, the functions of Synaptosomal-associated protein (SNAP) isoforms in photoreceptors are unknown. Here, we revisit the expression of SNAP-23 and SNAP-25 and generate photoreceptor-specific knockout mice to investigate their roles. Although we find that SNAP-23 shows weak mRNA expression in photoreceptors, SNAP-23 removal does not affect retinal morphology or vision. SNAP-25 mRNA is developmentally regulated and undergoes mRNA trafficking to photoreceptor inner segments at postnatal day 9 (P9). SNAP-25 knockout photoreceptors develop normally until P9 but degenerate by P14 resulting in severe retinal thinning. Photoreceptor loss in SNAP-25 knockout mice is associated with abolished electroretinograms and vision loss. We find mistrafficked photopigments, enlarged synaptic vesicles, and abnormal synaptic ribbons which potentially underlie photoreceptor degeneration. Our results conclude that SNAP-25, but not SNAP-23, mediates photopigment delivery and synaptic functioning required for photoreceptor development, survival, and function.

Theobromine improves hyperactivity, inattention, and working memory via modulation of dopaminergic neural function in the frontal cortex of spontaneously hypertensive rats.

Attention-deficit/hyperactivity disorder (ADHD) is a developmental disorder and dopaminergic dysfunction in the prefrontal cortex (PFC) may play a role. Our previous research indicated that theobromine (TB), a methylxanthine, enhances cognitive function in rodents via the PFC. This study investigates TB's effects on hyperactivity and cognitive function in stroke-prone spontaneously hypertensive rats (SHR), an ADHD animal model. Male SHRs (6-week old) received a diet containing 0.05% TB for 40 days, while control rats received normal diets. Age-matched male Wistar-Kyoto rats (WKY) served as genetic controls. During the TB administration period, we conducted open-field tests and Y-maze tasks to evaluate hyperactivity and cognitive function, then assessed dopamine concentrations and tyrosine hydroxylase (TH), dopamine receptor D1-5 (DRD1-5), dopamine transporter (DAT), vesicular monoamine transporter-2 (VMAT-2), synaptosome-associated protein-25 (SNAP-25), and brain-derived neurotrophic factor (BDNF) expressions in the PFC. Additionally, the binding affinity of TB for the adenosine receptors (ARs) was evaluated. Compared to WKY, SHR exhibited hyperactivity, inattention and working memory deficits. However, chronic TB administration significantly improved these ADHD-like behaviors in SHR. TB administration also normalized dopamine concentrations and expression levels of TH, DRD2, DRD4, SNAP-25, and BDNF in the PFC of SHR. No changes were observed in DRD1, DRD3, DRD5, DAT, and VMAT-2 expression between SHR and WKY rats, and TB intake had minimal effects. TB was found to have affinity binding to ARs. These results indicate that long-term TB supplementation mitigates hyperactivity, inattention and cognitive deficits in SHR by modulating dopaminergic nervous function and BDNF levels in the PFC, representing a potential adjunctive treatment for ADHD.

A sensitive cell-based assay for testing potency of botulinum neurotoxin type A

Botulinum neurotoxin type A (BoNT/A) is a biopharmaceutic widely used for the treatment of neu­rological diseases and in aesthetic medicine to achieve months-long paralysis of target muscles and glands. Large numbers of mice are used in the mouse bioassay (MBA) for various botulinum-related applications including batch release potency testing, antitoxin testing, countermeasure development, and basic research. BoNT/A intoxication causes severe suffering to the mice used for testing, and application-specific, non-animal alternatives are urgently needed. Cell-based assays (CBA) need to replicate all the physiological steps of botulinum intoxication, comprising neuronal binding, internali­zation, endosomal escape, and cleavage of synaptosomal-associated protein of 25 kDa (SNAP25). However, the CBA currently in use have limitations. In this study we show that LAN5 cells, a human neuroblastoma-derived cell line, are sensitive to BoNT/A and can be engineered to express a recom­binant NanoLuciferase (NanoLuc)-tagged SNAP25 reporter molecule. On exposure, the reporter molecule is cleaved and releases a NanoLuc-SNAP25 fragment that can be captured on a 96-well plate for quantitative luminometry using a cleavage-specific SNAP25 antibody. We demonstrate, using purified BoNT/A and a commercial BoNT/A product, that the sensitivity of this new cell-based assay is in the fM range, comparable to that of the MBA. The new assay could replace the MBA in research and commercial testing of BoNT/A, benefiting both scientific progress and animal welfare.

Allosteric inhibition of phosphodiesterase 4D induces biphasic memory-enhancing effects associated with learning-activated signaling pathways.

Phosphodiesterase 4D negative allosteric modulators (PDE4D NAMs) enhance memory and cognitive function in animal models without emetic-like side effects. However, the relationship between increased cyclic adenosine monophosphate (cAMP) signaling and the effects of PDE4D NAM remains elusive. To investigate the roles of hippocampal cAMP metabolism and synaptic activation in the effects of D159687, a PDE4D NAM, under baseline and learning-stimulated conditions. At 3mg/kg, D159687 enhanced memory formation and consolidation in contextual fear conditioning; however, neither lower (0.3mg/kg) nor higher (30mg/kg) doses induced memory-enhancing effects. A biphasic (bell-shaped) dose-response effect was also observed in a scopolamine-induced model of amnesia in the Y-maze, whereas D159687 dose-dependently caused an emetic-like effect in the xylazine/ketamine anesthesia test. At 3mg/kg, D159687 increased cAMP levels in the hippocampal CA1 region after conditioning in the fear conditioning test, but not in the home-cage or conditioning cage (i.e., context only). By contrast, 30mg/kg of D159687 increased hippocampal cAMP levels under all conditions. Although both 3 and 30mg/kg of D159687 upregulated learning-induced Fos expression in the hippocampal CA1 30min after conditioning, 3mg/kg, but not 30mg/kg, of D159687 induced phosphorylation of synaptic plasticity-related proteins such as cAMP-responsive element-binding protein, synaptosomal-associated protein 25kDa, and the N-methyl-D-aspartate receptor subunit NR2A. Our findings suggest that learning-stimulated conditions can alter the effects of a PDE4D NAM on hippocampal cAMP levels and imply that a PDE4D NAM exerts biphasic memory-enhancing effects associated with synaptic plasticity-related signaling activation.

SNAP-25 Polymorphisms in Autism Spectrum Disorder: A Pilot Study towards a Possible Endophenotype.

While there is substantial agreement on the diagnostic criteria for autism spectrum disorder, it is also acknowledged that it has a broad range of clinical presentations. This can complicate the diagnostic process and aggravate the choice of the most suitable rehabilitative strategy for each child. Attentional difficulties are among the most frequently reported comorbidities in autism spectrum disorder. We investigated the role of SNAP-25 polymorphisms. Synaptosome-associated protein 25 (SNAP25) is a presynaptic membrane-binding protein; it plays a crucial role in neurotransmission and has already been studied in numerous psychiatric disorders. It was also seen to be associated with hyperactivity in children with autism spectrum disorder. We collected clinical, behavioral and neuropsychological data on 41 children with a diagnosis of autism spectrum disorder, and then genotyped them for five single-nucleotide polymorphisms of SNAP-25. Participants were divided into two groups according to the Autism Diagnostic Observation Schedule (ADOS-2) Severity Score. In the group with the highest severity score, we found significant associations of clinical data with polymorphism rs363050 (A/G): children with the GG genotype had lower total IQ, more severe autistic functioning and more attentional difficulties. Our research could be the starting point for outlining a possible endophenotype among patients with autism spectrum disorder who are clinically characterized by severe autistic functioning and significant attentional difficulties.

Botulinum toxin type a antinociceptive activity in trigeminal regions involves central transcytosis

ObjectiveBotulinum toxin type A (BoNT-A) provides lasting pain relief in patients with craniofacial pain conditions but the mechanisms of its antinociceptive activity remain unclear. Preclinical research revealed toxin axonal transport to the central afferent terminals, but it is unknown if its central effects involve transsynaptic traffic to the higher-order synapses. To answer this, we examined the contribution of central BoNT-A transcytosis to its action in experimental orofacial pain. Material and methods:Male Wistar rats, 3–4 months old, were injected with BoNT-A (7 U/kg) unilaterally into the vibrissal pad. To investigate the possible contribution of toxin's transcytosis, BoNT-A-neutralizing antiserum (5 IU) was applied intracisternally. Antinocicepive BoNT-A action was assessed by duration of nocifensive behaviors and c-Fos activation in the trigeminal nucleus caudalis (TNC) following bilateral or unilateral formalin (2.5%) application into the vibrissal pad. Additionally, cleaved synaptosomal-associated protein of 25 kDa (cl-SNAP-25) immunoreactivity was analyzed in the bilateral TNC. Results:Unilaterally injected BoNT-A reduced the nocifensive behaviors and bilateral c-Fos activation induced by formalin, which was accompanied by the toxin's enzymatic activity on both sides of the TNC. BoNT-A antinociceptive or enzymatic activities were prevented by the specific neutralizing antitoxin. BoNT-A contralateral action occurred independently from ipsilateral side nociception or contralateral trigeminal nerve-mediated axonal traffic. ConclusionHerein, we demonstrate that antinociceptive action of pericranially administered BoNT-A involves transsynaptic transport to second order synapses and contralateral trigeminal nociceptive nuclei. These results reveal more complex central toxin activity, necessary to explain its clinical effectiveness in the trigeminal region-related pain states.

SIP30 involvement in vesicle exocytosis from PC12 cells

SNAP25 (synaptosome-associated protein of 25 kDa) is a core SNARE (soluble N-ethylmaleimide-sensitive factor attachment receptor) protein; and the interaction between SNAP25 and other SNARE proteins is essential for synaptic vesicle exocytosis. Identified as a SNAP25 interacting protein, SIP30 (SNAP25 interacting protein at 30 kDa) has been shown to modulate neuropathic pain behavior, and is potentially involved in the cellular process of vesicle exocytosis. Previous study demonstrated that using a vesicle secretion assay in PC12 cells, anti-SIP30 siRNA reduced vesicle exocytosis. We investigated vesicle exocytosis from PC12 cells with FM1-43 fluorescence dye, and demonstrated that anti-SIP30 siRNA reduced the pool of releasable vesicles and the rate of vesicle exocytosis, without affecting the endocytosis and recycling of the exocytosed vesicles. The results show that SIP30 is involved in vesicle exocytosis, suggesting a potential mechanism of SIP30 modulation of neuropathic pain.

SNAP25 is a potential target for early stage Alzheimer’s disease and Parkinson’s disease

BackgroundAlzheimer’s disease (AD) and Parkinson’s disease (PD), two common irreversible neurodegenerative diseases, share similar early stage syndromes, such as olfaction dysfunction. Yet, the potential comorbidity mechanism of AD and PD was not fully elucidated.MethodsThe gene expression profiles of GSE5281 and GSE8397 were downloaded from the Gene Expression Omnibus (GEO) database. We utilized a series of bioinformatics analyses to screen the overlapped differentially expressed genes (DEGs). The hub genes were further identified by the plugin CytoHubba of Cytoscape and validated in the hippocampus (HIP) samples of APP/PS-1 transgenic mice and the substantial nigra (SN) samples of A53T transgenic mice by real-time quantitative polymerase chain reaction (RT-qPCR). Meanwhile, the expression of the target genes in the olfactory epithelium/bulb was detected by RT-qPCR. Finally, molecular docking was used to screen potential compounds for the target gene.ResultsOne hundred seventy-four overlapped DEGs were identified in AD and PD. Five of the top ten enrichment pathways mainly focused on the synapse. Five hub genes were identified and further validated. As a common factor in AD and PD, the changes of synaptosomal-associated protein 25 (SNAP25) mRNA in olfactory epithelium/bulb were significantly decreased and had a strong association with those in the HIP and SN samples. Pazopanib was the optimal compound targeting SNAP25, with a binding energy of − 9.2 kcal/mol.ConclusionsOur results provided a theoretical basis for understanding the comorbidity mechanism of AD and PD and highlighted that SNAP25 in the olfactory epithelium may serve as a potential target for early detection and intervention in both AD and PD.

Blood‐based SNAP‐25 and VAMP‐2 in Alzheimer’s disease; relation to cognition, atrophy and synaptic density.

AbstractBackgroundA hallmark of neurodegenerative disorders is synaptic dysfunction and degeneration. This makes the presynaptic protein synaptosomal‐associated protein 25 (SNAP‐25) and the Vesicle‐associated membrane protein 2 (VAMP‐2) targets of interest and importance. Previously, these proteins have only been measurable in cerebrospinal fluid (CSF) and are increased in Alzheimer’s disease (AD) patients. Considering the advancement of blood‐based detection of phosphorylated tau (p‐tau), neurofilament light (NfL) and amyloid‐β (Aβ42/40), a measure of synaptic degeneration in blood would be important for disease prognosis and clinical trial outcome. In this study, we describe results from novel single molecular arrays (Simoa) for blood SNAP‐25 and VAMP‐2.MethodPrototype Simoa assays for N‐terminal SNAP‐25 and VAMP‐2 were developed as more sensitive versions of previously validated in‐house CSF assays with diluent suited for both plasma and CSF. We examined 110 participants (54 AD and 56 healthy controls) from the Swedish BioFINDER pilot study. The mean age of patients was 75.9 years (7.1 SD) and there was an even distribution of sex. We examined group differences by the Wilcoxon rank sum test and associations between these two blood biomarkers and Aβ‐PET, tau‐PET, MRI and MMSE were determined by linear regression, adjusting for age and sex.ResultSNAP‐25 was significantly increased in patients with AD (mean [SD], 0.81 pg/mL [0.27]; p &lt; 0.001) as compared to controls (0.60 pg/mL [0.22]). SNAP‐25 distinguished AD patients from controls with an area under the curve of 0.74 (95% CI = 0.65‐0.83). SNAP‐25 showed association with Aβ PET (β = 0.10, p = 0.007) but not tau PET. SNAP‐25 associated with cortical thickness (β = ‐0.05, p = 0.009) and MMSE (β = ‐6.69, p = 0.001). No significant associations were found when analysing the AD group alone. VAMP‐2 showed no significant association with any of the mentioned parameters above.ConclusionThis novel ultra‐sensitive immunoassay demonstrates an increase of plasma SNAP‐25 in AD patients, and a significant relationship with cognition and cortical atrophy. Further studies will verify these changes in independent cohorts and explore the relationship between plasma SNAP‐25 and VAMP‐2 with synaptic vesicle glycoprotein 2A (SV2A) PET in a broad range of neurodegenerative disorders.

Plasma SNAP‐25 as a potential Alzheimer’s disease biomarker

AbstractBackgroundSynaptic degeneration and dysfunction are hallmarks of several neurodegenerative diseases, including Alzheimer’s disease (AD). Several studies have shown that the presynaptic protein, synaptosomal‐associated‐protein‐25 (SNAP‐25), is increased in the cerebrospinal fluid (CSF) of AD patients. In the era of ultra‐sensitive immunoassays, the quantification of SNAP‐25 concentration in blood could provide an easily accessible measure of synaptic integrity in AD.MethodIn this pilot study, we investigated 42 participants (n = 23 cognitively unimpaired [CU]; n = 19 MCI or AD dementia patients [CI]) from Geneva University Hospital. A total of 27 participants underwent amyloid positron emission tomography (PET) scans and 14 underwent tau PET scans within 1 year. Global cognition was assessed with Mini Mental State Examination (MMSE). Mann‐Whitney tests were used to assess differences in plasma SNAP25 levels between diagnostic, amyloid (A+/‐) and tau (T+/‐) classification. Furthermore, we used Pearson correlations to assess the association between SNAP25, amyloid centiloid, Tau SUVr and MMSE.ResultThe mean age was 66.9 years +/‐ 7.6 years and 57% were female. SNAP‐25 was significantly increased in CI patients (0.87 pg/mL ±/‐ 0.19) compared to CU participants (0.59 pg/mL ±/‐ 0.11; p&gt;0.001). SNAP‐25 levels were also significantly increased in A+ (0.89 pg/mL ±/‐ 0.18) and T+ (0.92 pg/mL ±/‐ 0.19) individuals compared to A‐ (0.65 pg/mL ±/‐ 0.09; p&lt;0.001) and T‐ (0.717 pg/mL ±/‐ 0.07; p = 0.041). Moreover, we observed a significant correlation between the SNAP‐25 levels and amyloid centiloid (p = 0.016, rho = 0.48), while the correlation was not significant between SNAP‐25 and tau SUVr (p = 0.146, rho = 0.41). Finally, we observed a significant correlation between SNAP‐25 levels and MMSE score (p = 0.033, rho = ‐0.35).ConclusionThe novel plasma single molecular array (Simoa) can accurately quantify SNAP‐25 in blood plasma. In a pilot study, we observed significantly elevated plasma SNAP‐25 levels in cognitively impaired individuals and amyloid and tau positive individuals. Our findings suggest that SNAP‐25 is a potential biomarker indexing synaptic loss in AD. Further results will extend to a memory clinic cohort of &gt;350 participants to assess its diagnostic and prognostic power and relationship to amyloid, tau, MRI and CSF biomarkers.

Biomarkers of synaptic and neuronal dysfunction in relation to mild behavioral impairment: a study of dementia‐free older adults

AbstractBackgroundMild behavioral impairment (MBI) represents a high‐risk state for incident cognitive decline and dementia, capturing later‐life emergent and persistent neuropsychiatric symptoms in five domains: 1) decreased motivation; 2) affective dysregulation 3) impulse dyscontrol; 4) social inappropriateness; and 5) psychosis. We examined cross‐sectional associations between MBI and cerebrospinal fluid levels of markers of synaptic/neuronal dysfunction including 1) alpha‐synuclein (α‐syn); 2) growth‐associated protein 43 (GAP‐43); 3) neurogranin (NGRN); 4) synaptosomal‐associated protein 25 (SNAP‐25); 5) visinin‐like protein 1 (VILIP‐1); and 6) chitinase‐3‐like protein 1 (YKL‐40). Longitudinally, we determined associations between these biomarkers and incident MBI or dementia.MethodAlzheimer’s Disease Neuroimaging Initiative participants with normal cognition and mild cognitive impairment were included (n = 853). MBI scores were derived from the Neuropsychiatric Inventory using a published algorithm. Participants were classified as MBI+ if MBI symptoms were present at ≥2 consecutive visits.Cross‐sectional associations between MBI total score and each log‐transformed biomarker were modelled using multivariable‐adjusted linear regression. Biomarkers with cross‐sectional associations meeting a pre‐defined p‐value threshold (p&lt;0.1) were then incorporated as exposures into adjusted longitudinal Cox proportional hazard regression models (n = 111) of 1) MBI and 2) dementia as the outcomes, the latter of which also included interactions between MBI status and biomarker levels.ResultDemographics are summarized in Table 1. Cross‐sectionally, only MBI associations with VILIP‐1 (beta = 3.3%, 95%CI:+0.2% to +6.5%, p = 0.04) and SNAP‐25 (beta = 2.0%, 95%CI:‐0.4% to 6.3%, p = 0.09) met the pre‐defined p‐value for inclusion in Cox models. Longitudinally, higher VILIP‐1 levels were associated with a 3.02‐fold greater rate of incident MBI (95%CI:1.43‐6.40, p = 0.004) and a 3.83‐fold greater rate of incident dementia (95%CI:1.22‐12.03, p = 0.021). The relationship between VILIP‐1 and dementia did not depend on participants having MBI. MBI status predicted incident dementia with an HR = 4.75 (95%CI:2.36‐9.55, p&lt;0.001) and HR = 4.06 (95%CI:2.01‐8.21, p&lt;0.001) after adjusting for VILIP‐1.ConclusionElevated VILIP‐1 levels are associated with greater incidence of MBI and greater incidence of dementia, suggesting that synaptic dysfunction traditionally related to cognitive decline and dementia may also contribute to behavioral changes. Treatments aimed to mitigate MBI symptoms and dementia may explore VILIP‐1—a neuronal calcium‐sensor protein—as a therapeutic target, though more research is needed.

Synapse dysfunction and astrocyte reactivity are associated independently of amyloid‐β and tau pathologies

AbstractBackgroundBiomarkers of astrocyte reactivity have the potential to improve diagnostic precision, disease monitoring, and treatment efficacy. Glial fibrillary acidic protein (GFAP) protein is expressed in astrocytes, which is important to synaptic plasticity, cell communication, and reactive gliosis.MethodCerebrospinal fluid (CSF) biomarkers were measured in participants of the McGill TRIAD cohort. 75 individuals (&gt;50 years old, 44 cognitively unimpaired (CU), and 31 with cognitive impairment (CI)), had available Aβ and tau‐PET. We measured CSF GFAP and synaptic markers (growth‐associated protein 43 (GAP‐43), neurogranin (Ng), synaptotagmin 1 (SYT1), presynaptic protein synaptosomal‐associated protein 25 (SNAP‐25)). Linear regressions adjusted for age, sex, clinical diagnosis, and Aβ/tau‐PET were used to test the associations between astrocyte reactivity and synaptic function.ResultDemographic information is shown in Table 1. We found an association between CSF GFAP and presynaptic markers (GAP‐43: p&lt;0.0001, β = 0.1076; SYT1: p&lt;0.0001, β = 0.0024; SNAP‐25 long: p&lt;0.0001, β = 0.0008) as well as postsynaptic markers (Ng: p&lt;0.05, β = 0.0066) independently of Aβ and tau burden (Figure 1).ConclusionOur biomarker results support experimental literature suggesting that astrocyte reactivity plays a role in downstream synaptic dysfunction independent of the brain levels of Aβ and tau tangles pathologies. This supports in vitro literature suggesting that therapeutic interventions targeting astrocyte reactivity can contribute to halting AD progression.