The localization of amyloid precursor protein to ependymal cilia in vertebrates and its role in ciliogenesis and brain development in zebrafish.

Amyloid precursor protein (APP) is expressed in many tissues in human, mice and in zebrafish. In zebrafish, there are two orthologues, Appa and Appb. Interestingly, some cellular processes associated with APP overlap with cilia-mediated functions. Whereas the localization of APP to primary cilia of in vitro-cultured cells has been reported, we addressed the presence of APP in motile and in non-motile sensory cilia and its potential implication for ciliogenesis using zebrafish, mouse, and human samples. We report that Appa and Appb are expressed by ciliated cells and become localized at the membrane of cilia in the olfactory epithelium, otic vesicle and in the brain ventricles of zebrafish embryos. App in ependymal cilia persisted in adult zebrafish and was also detected in mouse and human brain. Finally, we found morphologically abnormal ependymal cilia and smaller brain ventricles in appa-/-appb-/- mutant zebrafish. Our findings demonstrate an evolutionary conserved localisation of APP to cilia and suggest a role of App in ciliogenesis and cilia-related functions.

Comprehensive analysis of O-glycosylation of amyloid precursor protein (APP) using targeted and multi-fragmentation MS strategy.

BACKGROUND: The aberrant proteolytic processing of amyloid precursor protein (APP) into amyloid ß peptide (Aß) in brain is a critical step in the pathogenesis of Alzheimer's disease (AD). As an O-glycosylated protein, O-glycosylation of APP is considered to be related to Aß generation. Therefore, comprehensive analysis of APP O-glycosylation is important for understanding its functions. METHODS: We developed a Targeted MS approach with Multi-Fragmentation techniques (TMMF strategy), and successfully characterized O-glycosylation profiling of APP695 expressed in HEK-293 T cells. We calculated relative abundance of glycopeptides with various O-glycosites and O-glycans, and further investigated the alteration of APP O-glycosylation upon TNF-α treatment. RESULTS: A total of 14 O-glycosites were identified on three glycopeptides of APP, and at least four O-glycans including GalNAc (Tn antigen), core 1, and mono-/di-sialylated core 1 glycans were determinant at the residues of Thr576 and Thr577. We found a dense cluster of truncated O-glycans on the region nearby beginning of E2 domain and high abundance of sialylated O-glycans on the region close to ß-cleavage site. Moreover, we also observed that TNF-α could upregulate the expression of APP and the truncated O-glycans on APP in HEK-293 T cell. CONCLUSION: Our study established an intact O-glycopeptide MS analysis strategy for APP O-glycopeptide identification with enhanced fragmentation efficiency and detection sensitivity. These results provide a comprehensive O-glycosylation map of APP expressed in HEK-293 T cell. GENERAL SIGNIFICANCE: The accurate O-glycosites and O-glycan structures on APP may lead to a better understanding of the roles O-glycosylation plays in the processing and functions of APP.

Clinical, Imaging, and Pathologic Characteristics of Patients With Right vs Left Hemisphere-Predominant Logopenic Progressive Aphasia.

OBJECTIVE: To assess and compare demographic, clinical, neuroimaging, and pathologic characteristics of a cohort of patients with right hemisphere-predominant vs left hemisphere-predominant logopenic progressive aphasia (LPA). METHODS: This is a case-control study of patients with LPA who were prospectively followed at Mayo Clinic and underwent [18F]-fluorodeoxyglucose (FDG) PET scan. Patients were classified as rLPA if right temporal lobe metabolism was ≥1 SD lower than left temporal lobe metabolism. Patients with rLPA were frequency-matched 3:1 to typical left-predominant LPA based on degree of asymmetry and severity of temporal lobe metabolism. Patients were compared on clinical, imaging (MRI, FDG-PET, ß-amyloid, and tau-PET), and pathologic characteristics. RESULTS: Of 103 prospectively recruited patients with LPA, 8 (4 female) were classified as rLPA (7.8%); all patients with rLPA were right-handed. Patients with rLPA had milder aphasia based on the Western Aphasia Battery-Aphasia Quotient (p = 0.04) and less frequent phonologic errors (p = 0.015). Patients with rLPA had shorter survival compared to typical LPA: hazard ratio 4.0 (1.2-12.9), p = 0.02. There were no other differences in demographics, handedness, genetics, or neurologic or neuropsychological tests. Compared to the 24 frequency-matched patients with typical LPA, patients with rLPA showed greater frontotemporal hypometabolism of the nondominant hemisphere on FDG-PET and less atrophy in amygdala and hippocampus of the dominant hemisphere. Autopsy evaluation revealed a similar distribution of pathologic findings in both groups, with Alzheimer disease pathologic changes being the most frequent pathology. CONCLUSIONS: rLPA is associated with less severe aphasia but has shorter survival from reported symptom onset than typical LPA, possibly related to greater involvement of the nondominant hemisphere.

Associations of AT(N) biomarkers with neuropsychiatric symptoms in preclinical Alzheimer's disease and cognitively unimpaired individuals.

The development of in vivo biomarkers of Alzheimer's disease (AD) has advanced the diagnosis of AD from a clinical syndrome to a biological construct. The preclinical stage of AD continuum is defined by the identification of AD biomarkers crossing the pathological threshold in cognitively unimpaired individuals. While neuropsychiatric symptoms (NPS) are non-cognitive symptoms that are increasingly recognized as early manifestations of AD, the associations of NPS with AD pathophysiology in preclinical AD remain unclear. Here, we review the associations between NPS and AD biomarkers amyloid-ß (Aß), tau and neurodegeneration in preclinical AD and cognitively-unimpaired individuals in 19 eligible English-language publications (8 cross-sectional studies, 10 longitudinal, 1 both cross-sectional and longitudinal). The cross-sectional studies have consistently shown that NPS, particularly depressive and anxiety symptoms, are associated with higher Aß. The longitudinal studies have suggested that greater NPS are associated with higher Aß and cognitive decline in cognitively unimpaired subjects over time. However, most of the studies have either cross-sectionally or longitudinally shown no association between NPS and tau pathology. For the association of NPS and neurodegeneration, two studies have shown that the cerebrospinal fluid total-tau is linked to longitudinal increase in NPS and that the NPS may predict longitudinal metabolic decline in preclinical AD, respectively. However, evidence for the association between atrophy and NPS in preclinical AD is less consistent. Therefore, future longitudinal studies with well-designed methodologies and NPS measurements are required not only to determine the relationship among AT(N) biomarkers, NPS and cognitive decline, but also to elucidate the contribution of comorbid pathology to preclinical AD.

Development and Technical Validation of an Immunoassay for the Detection of APP669-711 (Aß-3-40) in Biological Samples.

The ratio of amyloid precursor protein (APP)669-711 (Aß-3-40)/Aß1-42 in blood plasma was reported to represent a novel Alzheimer's disease biomarker. Here, we describe the characterization of two antibodies against the N-terminus of Aß-3-x and the development and "fit-for-purpose" technical validation of a sandwich immunoassay for the measurement of Aß-3-40. Antibody selectivity was assessed by capillary isoelectric focusing immunoassay, Western blot analysis, and immunohistochemistry. The analytical validation addressed assay range, repeatability, specificity, between-run variability, impact of pre-analytical sample handling procedures, assay interference, and analytical spike recoveries. Blood plasma was analyzed after Aß immunoprecipitation by a two-step immunoassay procedure. Both monoclonal antibodies detected Aß-3-40 with no appreciable cross reactivity with Aß1-40 or N-terminally truncated Aß variants. However, the amyloid precursor protein was also recognized. The immunoassay showed high selectivity for Aß-3-40 with a quantitative assay range of 22 pg/mL-7.5 ng/mL. Acceptable intermediate imprecision of the complete two-step immunoassay was reached after normalization. In a small clinical sample, the measured Aß42/Aß-3-40 and Aß42/Aß40 ratios were lower in patients with dementia of the Alzheimer's type than in other dementias. In summary, the methodological groundwork for further optimization and future studies addressing the Aß42/Aß-3-40 ratio as a novel biomarker candidate for Alzheimer's disease has been set.

In Vivo Visualization of Moving Synaptic Cargo Complexes within Drosophila Larval Segmental Axons.

Identifying moving synaptic vesicle complexes and isolating specific proteins present within such complexes in vivo is challenging. Here we detail a protocol that we have developed that is designed to simultaneously visualize the axonal transport of two fluorescently tagged synaptic vesicle proteins in living Drosophila larval segmental nerves in real time. Using a beam-splitter and split view software, larvae expressing GFP-tagged Synaptobrevin (Syb) and mRFP-tagged Rab4-GTPase or YFP-tagged Amyloid Precursor protein (APP) and mRFP-tagged Rab4-GTPase are imaged simultaneously using separate wavelengths. Merged kymographs from the two wavelengths are evaluated for colocalization analysis. Vesicle velocity analysis can also be done. Such analysis enables us to visualize the motility behaviors of two synaptic proteins present on a single vesicle complex and identify candidate proteins moving on synaptic vesicles in vivo, under physiological conditions.

Longitudinal Cognitive Decline in a Novel Rodent Model of Cerebral Amyloid Angiopathy Type-1.

Cerebral amyloid angiopathy (CAA) is a small vessel disease characterized by ß-amyloid (Aß) accumulation in and around the cerebral blood vessels and capillaries and is highly comorbid with Alzheimer's disease (AD). Familial forms of CAA result from mutations within the Aß domain of the amyloid ß precursor protein (AßPP). Numerous transgenic mouse models have been generated around expression of human AßPP mutants and used to study cerebral amyloid pathologies. While behavioral deficits have been observed in many AßPP transgenic mouse lines, relative to rats, mice are limited in behavioral expression within specific cognitive domains. Recently, we generated a novel rat model, rTg-DI, which expresses Dutch/Iowa familial CAA Aß in brain, develops progressive and robust accumulation of cerebral microvascular fibrillar Aß beginning at 3 months, and mimics many pathological features of the human disease. The novel rTg-DI model provides a unique opportunity to evaluate the severity and forms of cognitive deficits that develop over the emergence and progression of CAA pathology. Here, we present an in-depth, longitudinal study aimed to complete a comprehensive assessment detailing phenotypic disease expression through extensive and sophisticated operant testing. Cohorts of rTg-DI and wild-type (WT) rats underwent operant testing from 6 to 12 months of age. Non-operant behavior was assessed prior to operant training at 4 months and after completion of training at 12 months. By 6 months, rTg-DI animals demonstrated speed-accuracy tradeoffs that later manifested across multiple operant tasks. rTg-DI animals also demonstrated delayed reaction times beginning at 7 months. Although non-operant assessments at 4 and 12 months indicated comparable mobility and balance, rTg-DI showed evidence of slowed environmental interaction. Overall, this suggests a form of sensorimotor slowing is the likely core functional impairment in rTg-DI rats and reflects similar deficits observed in human CAA.

Kinesin light chain-1 serine-460 phosphorylation is altered in Alzheimer's disease and regulates axonal transport and processing of the amyloid precursor protein.

Damage to axonal transport is an early pathogenic event in Alzheimer's disease. The amyloid precursor protein (APP) is a key axonal transport cargo since disruption to APP transport promotes amyloidogenic processing of APP. Moreover, altered APP processing itself disrupts axonal transport. The mechanisms that regulate axonal transport of APP are therefore directly relevant to Alzheimer's disease pathogenesis. APP is transported anterogradely through axons on kinesin-1 motors and one route for this transport involves calsyntenin-1, a type-1 membrane spanning protein that acts as a direct ligand for kinesin-1 light chains (KLCs). Thus, loss of calsyntenin-1 disrupts APP axonal transport and promotes amyloidogenic processing of APP. Phosphorylation of KLC1 on serine-460 has been shown to reduce anterograde axonal transport of calsyntenin-1 by inhibiting the KLC1-calsyntenin-1 interaction. Here we demonstrate that in Alzheimer's disease frontal cortex, KLC1 levels are reduced and the relative levels of KLC1 serine-460 phosphorylation are increased; these changes occur relatively early in the disease process. We also show that a KLC1 serine-460 phosphomimetic mutant inhibits axonal transport of APP in both mammalian neurons in culture and in Drosophila neurons in vivo. Finally, we demonstrate that expression of the KLC1 serine-460 phosphomimetic mutant promotes amyloidogenic processing of APP. Together, these results suggest that increased KLC1 serine-460 phosphorylation contributes to Alzheimer's disease.

Brain functions and unusual ß-amyloid accumulation in the hypertensive white matter lesions of rats.

This study used Sprague Dawley (SD) rats with stroke-prone renovascular hypertension (RHRSP) to establish an animal model of hypertensive white matter lesions (WML), so as to explore the brain functions and unusual ß-amyloid (Aß) accumulation in WML. Hypertensive WML and brain dysfunctions were evaluated by measuring the caudal arterial pressure of model rats, and by observing the histomorphological deformations o f the prefrontal lobe, temporal lobe, hippocampus and corpus callosum, as well as by counting of the number of neurons using Hematoxylin and Eosin (H and E) staining, and by evaluating the changes in rat brain functions, including memory and the ability of visual space learning, using the Morris Water Maze Test. In addition, the study discussed the correlation between Aß accumulation and hypertensive WML cognitive impairment by adopting an enzyme-linked immunosorbent assay (ELISA) to detect the level of Aß 1-42, and by detecting the expression of amyloid precursor protein (APP) and Beta-secretase 1 (BACE1) using Western blot. Results of the study showed that at 4 weeks, 8 weeks, 12 weeks and 16 weeks after operation, the blood pressure and brain Aß expression in the rats of the model group notably increased (P less than 0.01), along with deformed and degenerated brain tissues, confirming that the unusual Aß accumulation may participate in the occurrence and development of hypertensive WML as well as the induction of cerebral cognitive decreases.

Liquid biopsy of cerebrospinal fluid identifies neuronal pentraxin receptor (NPTXR) as a biomarker of progression of Alzheimer's disease.

Background Alzheimer's disease (AD) is the most prevalent form of dementia. Currently, the most studied biomarkers of AD are cerebrospinal fluid (CSF) amyloid ß 1-42, total tau and phosphorylated tau. However, misdiagnosis can exceed 20%. Recently, we found that CSF amyloid ß precursor-like protein-1 (APLP1) and neuronal pentraxin receptor (NPTXR) are promising biomarkers of AD. The aim of the present study is to validate CSF APLP1 and NPTXR as biomarkers of AD severity. Methods APLP1 and NPTXR concentrations were measured in the CSF of patients with mild cognitive impairment (MCI) (n = 14), mild AD (n = 21), moderate AD (n = 43) and severe AD (n = 30) using enzyme-linked immunosorbent assays (ELISAs). Results CSF APLP1 and NPTXR were not associated with age or sex. CSF APLP1 was not different between any of the AD severity groups (p = 0.31). CSF NPTXR was significantly different between MCI and mild AD (p = 0.006), mild and moderate AD (p = 0.016), but not between moderate and severe AD (p = 0.36). NPTXR concentration progressively declined from MCI to mild, to moderate and to severe AD patients (p < 0.0001, Kruskal-Wallis test). CSF NPTXR positively correlated with the Mini-Mental Status Examination (MMSE) score (p < 0.001). Conclusions NPTXR concentration in CSF is a promising biomarker of AD severity and could inform treatment success and disease progression in clinical settings.

A study of residual lesions in horses that recovered from clinical signs of chronic equine dysautonomia.

BACKGROUND: Equine dysautonomia (ED) causes degeneration and loss of autonomic neurons. Approximately 50% of chronic cases recover, but it is unclear how they survive neuronal loss. OBJECTIVES: To assess lesions, autonomic neuron numbers, interstitial cells of Cajal (ICC), and neurodegeneration in recovered cases. ANIMALS: Thirteen cases (group ED), euthanized 10.3 ± 5.2 (1-16) years from diagnosis and 6 age-matched controls (group C). METHODS: Prospective, case control; routine post mortem examination, neuron counts in peripheral and enteric ganglia and immunohistochemical assessment of neural networks (Protein gene product [PGP] 9.5), ICC (c-kit), and neurodegeneration (beta-amyloid precursor protein and ubiquitin) in intestine. RESULTS: Postmortem findings in group ED were small intestinal dilation (4/12, 33%) and muscular hypertrophy (4/12, 33%), and gastric mucosal hypertrophy (3/11, 27%) and ulceration (4/11, 36%). Neuron density was lower in group ED (mean 39% lower for cranial cervical ganglion [P < .001], median 44% lower in celiacomesenteric ganglion [P = .01]). In intestine, neuronal depletion was worst in ileum (median 100% lower in submucosal plexus [P < .001], 91% lower in myenteric plexus [P = .004]). Group ED had less PGP 9.5 staining in ileal myenteric plexus (mean 66% lower [P = .04]) and circular muscle (median 75% lower [P = .006]). In ileum, there was less c-kit staining in myenteric plexus (median 57% lower [P = .02]) but not muscularis externa. Beta-amyloid precursor protein and ubiquitin results were not indicitive of neurodegeneration. CONCLUSIONS AND CLINICAL IMPORTANCE: Intact ICC in muscularis externa might help maintain motility after neuronal loss. Treatment supporting ICC function warrants investigation.

Diagnostic and prognostic biomarkers for HAND.

In 2007, the nosology for HIV-1-associated neurocognitive disorders (HAND) was updated to a primarily neurocognitive disorder. However, currently available diagnostic tools lack the sensitivity and specificity needed for an accurate diagnosis for HAND. Scientists and clinicians, therefore, have been on a quest for an innovative biomarker to diagnose (i.e., diagnostic biomarker) and/or predict (i.e., prognostic biomarker) the progression of HAND in the post-combination antiretroviral therapy (cART) era. The present review examined the utility and challenges of four proposed biomarkers, including neurofilament light (NFL) chain concentration, amyloid (i.e., sAPPα, sAPPß, amyloid ß) and tau proteins (i.e., total tau, phosphorylated tau), resting-state functional magnetic resonance imaging (fMRI), and prepulse inhibition (PPI). Although significant genotypic differences have been observed in NFL chain concentration, sAPPα, sAPPß, amyloid ß, total tau, phosphorylated tau, and resting-state fMRI, inconsistencies and/or assessment limitations (e.g., invasive procedures, lack of disease specificity, cost) challenge their utility as a diagnostic and/or prognostic biomarker for milder forms of neurocognitive impairment (NCI) in the post-cART era. However, critical evaluation of the literature supports the utility of PPI as a powerful diagnostic biomarker with high accuracy (i.e., 86.7-97.1%), sensitivity (i.e., 89.3-100%), and specificity (i.e., 79.5-94.1%). Additionally, the inclusion of multiple CSF and/or plasma markers, rather than a single protein, may provide a more sensitive diagnostic biomarker for HAND; however, a pressing need for additional research remains. Most notably, PPI may serve as a prognostic biomarker for milder forms of NCI, evidenced by its ability to predict later NCI in higher-order cognitive domains with regression coefficients (i.e., r) greater than 0.8. Thus, PPI heralds an opportunity for the development of a brief, noninvasive diagnostic and promising prognostic biomarker for milder forms of NCI in the post-cART era.

Azure B affects amyloid precursor protein metabolism in PS70 cells.

Alzheimer's disease (AD), the most common form of dementia, is characterized by abundant deposition of amyloid-ß (Aß) peptide that is the result of sequential cleavage of amyloid precursor protein (APP) by ß-secretase and γ-secretase. Several studies have documented that inhibition of Aß peptide synthesis or facilitating its degradation is one of the attractive therapeutic strategies in AD. Methylene blue (MethB), which has recently been investigated in Phase II clinical trials, is a prominent inhibitor in reducing Aß oligomers. Herein, we wonder whether the mitigating effects of MethB on amyloid metabolism are related to the activity of its major metabolite, azure B. The goal of this study was to investigate the effects of azure B, which is also a cholinesterase inhibitor, on APP processing by using Chinese hamster ovary cells stably expressing human wild-type APP and presenilin 1 (PS70). Azure B significantly decreased the levels of secreted APPα (sAPPα) and Aß40/42 in culture medium with a dose-dependent manner. A significant decrease was also observed in the levels of intracellular APP without affecting the cell viability. In parallel with the decrease of APP and APP metabolites, the activity of ß-secretase 1 (BACE1) was significantly attenuated compared to control. Overall, our results show that azure B has a large contribution for the pharmacological profile of MethB in APP metabolism.

One-step synthesized flower-like materials used for sensitively detecting amyloid precursor protein.

In this paper, we developed a new method to detect Alzheimer's disease (AD)-related amyloid precursor protein (APP). A composite material containing horseradish peroxidase (HRP), APP antibody, and Cu3(PO4)2 was synthesized as the biosensor by co-precipitation method. In this competitive immunoassay, APP was first conjugated onto the microplate surface with the help of poly-L-lysine as the coating reagent; the composite materials were then attached onto the microplate through the interaction of APP and antibody; the HRP can catalyze the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) and formed colored species. Therefore, the more APP in the detection solution (free form), the less composite material was combined with the immobilized APP on the microplate, resulting in the production of less colored TMB species. A series of detection parameters were studied, such as the composite material synthesis process, the concentration, and reaction time of different compounds. Our method has higher sensitivity compared with the similar immunoassay without using composite materials (the limits of detection are 0.3 and 3 ng/mL, respectively), and can be used for real samples (human serum) detection. The detection results using our method are consistent with the ELISA results, which is useful for the AD detection. Graphical abstract ᅟ.

[The detection of the biomolecular markers of the axonal damage resulting from the craniocerebral injury by the immunohistological methods]. / Immunogistokhimicheskaia detektsiia biomolekuliarnykh markerov aksonal'nogo povrezhdeniia pri cherepno-mozgovoi travme.

The objective of the present study was to define and evaluate the parameters of validity of the diagnostic signs of the diffuse axonal damage resulting from the craniocerebral injury with the use of the immunohistological reaction for the presence of amyloid precursor protein (beta-APP) in the brain tissue associated with the craniocerebral injury. Both histological and immunological methods were used. The positive beta-APP reaction was documented in 2 of the six cases. It is concluded that the positive results of the immunohistological reaction for the presence of beta-APP protein provide an additional information about the mechanism behind the damage and confirm the diagnosis of the craniocerebral injury in the cases when its macroscopic signs are either absent or unapparent.

Expression of amyloid precursor-like protein 2 (APLP2) in glioblastoma is associated with patient prognosis.

The purpose of this study was to investigate the expression status of amyloid precursor-like protein 2 (APLP2) and its clinical relevance in patients with glioblastoma. The publically available database Project Betastasis involving Repository for Molecular Brain Neoplasia Data (REMBRANDT) and The Cancer Genome Atlas (TCGA) was first utilized to analyze the expression and prognostic potential of APLP2 in glioblastoma. Compared with normal controls, the glioblastoma group from each dataset showed no significant difference of APLP2 expression (p > 0.05). However, when connected to glioblastoma patient's prognosis, a high APLP2 expression was found to be associated with short overall survival in REMBRANDT cases (p = 0.0323) but not the TCGA group (p = 0.0578). Consistently, APLP2 expression detected by immunohistochemistry in our cohort revealed an undifferentiated expression pattern between glioblastoma (n = 114) and normal brain (n = 16) (p = 0.265) and among all grade gliomas. Furthermore, univariate and multivariate analyses identified a high APLP2 expression as an independent risk factor for overall survival (hazard ratio = 1.537, p = 0.041) and progression-free survival (hazard ratio = 1.783, p = 0.037) of glioblastoma patients. In conclusion, the expression of APLP2 might correlate with tumor development and be a prognostic factor for patients with glioblastoma.

Overexpression of Amyloid Precursor Protein Promotes the Onset of Seborrhoeic Keratosis and is Related to Skin Ageing.

Seborrhoeic keratosis (SK) is an age-related skin disease. Amyloid precursor protein (APP) plays an important role in the pathogenesis of age-related Alzheimer's disease. The aim of this study was to elucidate the expression characteristics of APP in SK tissues (n = 50), and explore whether the production of APP is related to the onset of SK and skin ageing, including ultraviolet (UV)-induced ageing, as observed in normal skin (n = 79). The results of immunohistochemistry, Western blotting and quantitative real-time PCR showed that APP and its downstream products (i.e. amyloid-ß42) were more highly expressed in SK than in paired adjacent normal skin tissues. In contrast, the expression of its key secretase (i.e. ß-secretase1) was generally low. Furthermore, APP expression was higher in UV-exposed than non-exposed skin sites, and expression in the older age group (61-85 years) was greater than that in the younger age group (41-60 years) in SK tissues (p<0.05). APP expression correlated positively with age in epidermis (p<0.05), but not in dermis. These findings suggest that overexpression of APP may promote the onset of SK and is a marker of skin ageing and UV damage. Further research will elucidate whether therapeutic mitigation of increased levels of APP in the skin might delay the onset of SK and skin ageing.

Pulsatile stretch as a novel modulator of amyloid precursor protein processing and associated inflammatory markers in human cerebral endothelial cells.

Amyloid ß (Aß) deposition is a hallmark of Alzheimer's disease (AD). Vascular modifications, including altered brain endothelial cell function and structural viability of the blood-brain barrier due to vascular pulsatility, are implicated in AD pathology. Pulsatility of phenomena in the cerebral vasculature are often not considered in in vitro models of the blood-brain barrier. We demonstrate, for the first time, that pulsatile stretch of brain vascular endothelial cells modulates amyloid precursor protein (APP) expression and the APP processing enzyme, ß-secretase 1, eventuating increased-Aß generation and secretion. Concurrent modulation of intercellular adhesion molecule 1 and endothelial nitric oxide synthase (eNOS) signaling (expression and phosphorylation of eNOS) in response to pulsatile stretch indicates parallel activation of endothelial inflammatory pathways. These findings mechanistically support vascular pulsatility contributing towards cerebral Aß levels.

Translational evidence for two distinct patterns of neuroaxonal injury in sepsis: a longitudinal, prospective translational study.

BACKGROUND: Brain homeostasis deteriorates in sepsis, giving rise to a mostly reversible sepsis-associated encephalopathy (SAE). Some survivors experience chronic cognitive dysfunction thought to be caused by permanent brain injury. In this study, we investigated neuroaxonal pathology in sepsis. METHODS: We conducted a longitudinal, prospective translational study involving (1) experimental sepsis in an animal model; (2) postmortem studies of brain from patients with sepsis; and (3) a prospective, longitudinal human sepsis cohort study at university laboratory and intensive care units (ICUs). Thirteen ICU patients with septic shock, five ICU patients who died as a result of sepsis, fourteen fluid-resuscitated Wistar rats with fecal peritonitis, eleven sham-operated rats, and three human and four rat control subjects were included. Immunohistologic and protein biomarker analysis were performed on rat brain tissue at baseline and 24, 48, and 72 h after sepsis induction and in sham-treated rats. Immunohistochemistry was performed on human brain tissue from sepsis nonsurvivors and in control patients without sepsis. The clinical diagnostics of SAE comprised longitudinal clinical data collection and magnetic resonance imaging (MRI) and electroencephalographic assessments. Statistical analyses were performed using SAS software (version 9.4; SAS Institute, Inc., Cary, NC, USA). Because of non-Gaussian distribution, the nonparametric Wilcoxon test general linear models and the Spearman correlation coefficient were used. RESULTS: In postmortem rat and human brain samples, neurofilament phosphoform, ß-amyloid precursor protein, ß-tubulin, and H&E stains distinguished scattered ischemic lesions from diffuse neuroaxonal injury in septic animals, which were absent in controls. These two patterns of neuroaxonal damage were consistently found in septic but not control human postmortem brains. In experimental sepsis, the time from sepsis onset correlated with tissue neurofilament levels (R = 0.53, p = 0.045) but not glial fibrillary acidic protein. Of 13 patients with sepsis who had clinical features of SAE, MRI detected diffuse axonal injury in 9 and ischemia in 3 patients. CONCLUSIONS: Ischemic and diffuse neuroaxonal injury to the brain in experimental sepsis, human postmortem brains, and in vivo MRI suggest these two distinct lesion types to be relevant. Future studies should be focused on body fluid biomarkers to detect and monitor brain injury in sepsis. The relationship of neurofilament levels with time from sepsis onset may be of prognostic value. TRIAL REGISTRATION: ClinicalTrials.gov, NCT02442986 . Registered on May 13, 2015.

[The importance of marker proteins of the nervous tissue for morphological diagnostics of the craniocerebral injury]. / Znachenie belkov-markerov nervnoi tkani dlia morfologicheskoi diagnostiki cherepno-mozgovoi travmy.

The present review of the literature involves 50 publications concerning various substrates of importance as the biological markers of axonal damages with special reference to the secondary molecular and cellular mechanisms on which to base in vitro and in vivo modeling of the craniocerebral injury. The results of the investigations with the application of mass-spectrometry for the identification of the proteins specifically synthesized in response to the injury are presented; their biological functions are described. The use of the sequential microscopic imaging technique and the immunohistochemical methods made it possible to determine that the majority of the marker proteins are involved in the specific intracellular processes that are triggered in response to the traumatic impact including apoptosis, proliferation, formation of lamellipodia, axon regeneration, actin remodeling, cell migration and inflammation. In addition, a rise in the amount of intracellular actin-associated proteins has been observed. It is concluded that the investigations into the properties and the physiological role of beta-amyloid precursor protein (beta-APP) are of special value for the characteristic of nervous tissue damages and morphological diagnostics of the craniocerebral injury.