Early but not late conformational changes of tau in association with ubiquitination of neurofibrillary pathology in Alzheimer's disease brains.

In Alzheimer's disease, tau protein undergoes post-translational modifications including hyperphosphorylation and truncation, which promotes two major conformational changes associated with progressive N-terminal folding. Along with the development of the disease, tau ubiquitination was previously shown to emerge in the early and intermediate stages of the disease, which is closely associated with early tau truncation at aspartic acid 421, but not with a subsequently truncated tau molecule at glutamic acid 391. In the same group of cases, using multiple immunolabeling and confocal microscopy, a possible relationship between the ubiquitin-targeting of tau and the progression of conformational changes adopted by the N-terminus of this molecule was further studied. A comparable number of neurofibrillary tangles was found displaying ubiquitin, an early conformation recognized by the Alz-50 antibody, and a phosphorylation. However, a more reduced number of neurofibrillary tangles were immunoreactive to Tau-66 antibody, a late tau conformational change marker. When double-labeling profiles of neurofibrillary tangles were assessed, ubiquitination was clearly demonstrated in tau molecules undergoing early N-terminal folding, but was barely observed in late conformational changes of the N-terminus adopted by tau. The same pattern of colocalization was visualized in neuritic pathology. Overall, these results indicate that a more intact conformation of the N-terminus of tau may facilitate tau ubiquitination, but this modification may not occur in a late truncated and more compressed folding of the N-terminus of the tau molecule.

Kinase-loaded magnetic beads for sequential in vitro phosphorylation of peptides and proteins.

Post-translational modifications, including phosphorylation, greatly impact the physiological function of proteins, especially those that are natively unfolded and implicated in many neurodegenerative diseases. However, structural and functional studies of such proteins require fully defined phosphorylation, including those that are not physiological. Thus, the kinases ERK2 and GSK-3ß were immobilized to various superparamagnetic beads with carboxylic, aldehyde, Ni2+, or Co3+ functional groups, with a view to efficiently phosphorylate peptides and proteins in vitro. Full phosphorylation of specific synthetic peptides confirmed that beads were successfully loaded with kinases. Remarkably, enzymes covalently immobilized on carboxylated SeraMag beads remained active upon reuse, with residual activity after 10 uses 99.5 ± 0.34% for GSK-3ß and 36.2 ± 2.01% for ERK2. The beads were also used to sequentially phosphorylate recombinant tau, which in vivo is a biomarker of Alzheimer's disease. Thus, a system consisting of two fully active kinases immobilized to magnetic beads is demonstrated for the first time. In comparison to soluble enzymes, the beads are easier to handle, reusable, and thus low-cost. Importantly, these beads are also convenient to remove from reactions to minimize contamination of phosphorylated products or to exchange with other kinases.

Inhibitors of Acetylcholinesterase Derived from 7-Methoxytacrine and Their Effects on the Choline Transporter CHT1.

BACKGROUND: Reversible acetylcholinesterase inhibitors are used in Alzheimer disease therapy. However, tacrine and its derivatives have severe side effects. Derivatives of the tacrine analogue 7-methoxytacrine (MEOTA) are less toxic. METHODS: We evaluated new derivatives of 7-MEOTA (2 homodimers linked by 2 C4-C5 chains and 5 N-alkylated C4-C8 side chain derivatives) in vitro, using the rat hippocampal choline transporter CHT1. RESULTS: Some derivatives were effective inhibitors of rat acetylcholinesterase and comparable with 7-MEOTA. All derivatives were able to inhibit CHT1, probably via quaternary ammonium, and this interaction could be involved in the enhancement of their detrimental side effects and/or in the attenuation of their promising effects. Under conditions of disrupted lipid rafts, the unfavorable effects of some derivatives were weakened. Only tacrine was probably able to stereospecifically interact with the naturally occurring amyloid-ß isoform and to simultaneously stimulate CHT1. Some derivatives, when coincubated with amyloid ß, did not influence CHT1. All derivatives also increased the fluidity of the cortical membranes. CONCLUSION: The N-alkylated derivative of 7-MEOTA bearing from C4 side chains appears to be the most promising compound and should be evaluated in future in vivo research.

Montreal cognitive assessment (MoCA): Normative data for old and very old Czech adults.

The principal aim of our study was to present norms for old and very old Czech adults on the Czech version of the Montreal Cognitive Assessment (MoCA) and investigate the influence of social and demographic factors on MoCA performance. We analyzed 540 adults aged ≥ 60 years (5-year age categories; nationally representative sample in terms of sex and educational level), who met strict inclusion criteria for the absence of neurodegenerative disorders and performed within normal range in neuropsychological assessment. Using multiple regression model, we found that MoCA performance was affected by age and education (both p < .001) but not sex. The study provides normed percentile estimates for MoCA performance stratified by age (60-74 years; ≥ 75 years) and education lower versus higher. We also present percentile equivalents for the MoCA and Mini-Mental State Examination (MMSE) for use in clinical practice. We found age- and education-related effects on MoCA performance which support the use of culturally adapted normative data.

The role of steroids in the prediction of affective disorders in adult men.

OBJECTIVE: Anxiety and mood disorders (AMD) are the most frequent mental disorders in the human population. They have recently shown increasing prevalence, and commonly disrupt personal and working lives. The aim of our study was to analyze the spectrum of circulating steroids in order to discover differences that could potentially be markers of affective depression or anxiety, and identify which steroids could be a predictive component for these diseases. METHODS: We studied the steroid metabolome including 47 analytes in 20 men with depression (group D), 20 men with anxiety (group AN) and 30 healthy controls. OPLS and multivariate regression models were used for statistical analysis. RESULTS: Discrimination of group D from controls by the OPLS method was absolute, as was group AN from controls (sensitivity=1.000 (0.839, 1.000), specificity=1.000 (0.887, 1.000)). Relatively good predictivity was also found for discrimination between group D from AN (sensitivity=0.850 (0.640, 0.948), specificity=0.900 (0.699, 0.972)). CONCLUSION: Selected circulating steroids, including those that are neuroactive and neuroprotective, can be useful tools for discriminating between these affective diseases in adult men.

A Rat Model of Alzheimer's Disease Based on Abeta42 and Pro-oxidative Substances Exhibits Cognitive Deficit and Alterations in Glutamatergic and Cholinergic Neurotransmitter Systems.

Alzheimer's disease (AD) is one of the most serious human, medical, and socioeconomic burdens. Here we tested the hypothesis that a rat model of AD (Samaritan; Taconic Pharmaceuticals, USA) based on the application of amyloid beta42 (Abeta42) and the pro-oxidative substances ferrous sulfate heptahydrate and L-buthionine-(S, R)-sulfoximine, will exhibit cognitive deficits and disruption of the glutamatergic and cholinergic systems in the brain. Behavioral methods included the Morris water maze (MWM; long-term memory version) and the active allothetic place avoidance (AAPA) task (acquisition and reversal), testing spatial memory and different aspects of hippocampal function. Neurochemical methods included testing of the NR1/NR2A/NR2B subunits of NMDA receptors in the frontal cortex and CHT1 transporters in the hippocampus, in both cases in the right and left hemisphere separately. Our results show that Samaritan rats(™) exhibit marked impairment in both the MWM and active place avoidance tasks, suggesting a deficit of spatial learning and memory. Moreover, Samaritan rats exhibited significant changes in NR2A expression and CHT1 activity compared to controls rats, mimicking the situation in patients with early stage AD. Taken together, our results corroborate the hypothesis that Samaritan rats are a promising model of AD in its early stages.

Sex-Dependent Changes in Striatal Dopamine Transport in Preadolescent Rats Exposed Prenatally and/or Postnatally to Methamphetamine.

Methamphetamine (MA) is the most commonly used psychostimulant drug, the chronic abuse of which leads to neurodegenerative changes in the brain. The global use of MA is increasing, including in pregnant women. Since MA can cross both placental and haematoencephalic barriers and is also present in maternal milk, children of chronically abused mothers are exposed prenatally as well as postnatally. Women seem to be more vulnerable to some aspects of MA abuse than men. MA is thought to exert its effects among others via direct interactions with dopamine transporters (DATs) in the brain tissue. Sexual dimorphism of the DAT system could be a base of sex-dependent actions of MA observed in behavioural and neurochemical studies. Possible sex differences in the DATs of preadolescent offspring exposed to MA prenatally and/or postnatally have not yet been evaluated. We examined the striatal synaptosomal DATs (the activity and density of surface expressed DATs and total DAT expression) in preadolescent male and female Wistar rats (31-35-day old animals) exposed prenatally and/or postnatally to MA (daily 5 mg/kg, s.c. to mothers during pregnancy and lactation). To distinguish between specific and nonspecific effects of MA on DATs, we also evaluated the in vitro effects of lipophilic MA on the fluidity of striatal membranes isolated from preadolescent and young adult rats of both sexes. We observed similar changes in the DATs of preadolescent rats exposed prenatally or postnatally (MA-mediated drop in the reserve pool but no alterations in surface-expressed DATs). However, prenatal exposure evoked significant changes in males and postnatal exposure in females. A significant decrease in the activity of surface-expressed DATs was found only in postnatally exposed females sensitized to MA via prenatal exposure. MA applied in vitro increased the fluidity of striatal membranes of preadolescent female but not male rats. In summary, DATs of preadolescent males are more sensitive to prenatal MA exposure via changes in the reserve pool and those of preadolescent females to postnatal MA exposure via the same mechanism. The combination of prenatal and postnatal MA exposure increases the risk of dopaminergic deficits via alterations in the activity of surface-expressed DATs especially in preadolescent females. MA-mediated changes in DATs of preadolescent females could be still enhanced via nonspecific disordering actions of MA on striatal membranes.

Circulating C19 steroids and progesterone metabolites in women with acute depression and anxiety disorders.

Depression and anxiety disorders are highly prevalent in women. Although several studies have reported altered circulating steroids accompanying various mental disturbances, knowledge about alterations in the peripheral steroid pattern in such pathologies is incomplete. Therefore, we attempted to add to this knowledge using the simultaneous quantification of circulating steroids by gas chromatography mass spectrometry (GC-MS) in groups of premenopausal women in the follicular phase of the menstrual cycle (22 women with depression, 17 with anxiety disorders, 17 healthy controls). In addition to age-adjusted analysis of covariance (ANCOVA) followed by multiple comparisons, we developed models to successfully discriminate these groups from each other on the basis of steroid levels. Women with depression showed a reduced sulfoconjugation of steroids as well as lower levels of 7α-, 7ß- and 16α-hydroxy-metabolites of C19 Δ5 steroids. Women with depression have significantly lower circulating levels of 5α/ß-reduced pregnane steroids (with exception of free isopregnanolone) than women with anxiety or controls. Finally, our data indicate higher levels of estrogens in women with anxiety disorders when compared to women with depression.

Expression of Tau Produces Aberrant Plasma Membrane Blebbing in Glial Cells Through RhoA-ROCK-Dependent F-Actin Remodeling.

Abnormal aggregation of Tau in glial cells has been reported in Alzheimer's disease (AD) and other tauopathies; however, the pathological significance of these aggregates remains unsolved to date. In this study, we evaluated whether full-length Tau (Tau441) and its aspartic acid421-truncated Tau variant (Tau421) produce alterations in the normal organization of the cytoskeleton and plasma membrane (PM) when transiently expressed in cultured C6-glial cells. Forty-eight hours post-transfection, abnormal microtubule bundling was observed in the majority of the cells, which expressed either Tau441 or Tau421. Moreover, both variants of Tau produced extensive PM blebbing associated with cortical redistribution of filamentous actin (F-Actin). These effects were reverted when Tau-expressing cells were incubated with drugs that depolymerize F-Actin. In addition, when glial cells showing Tau-induced PM blebbing were incubated with inhibitors of the Rho-associated protein kinase (ROCK) signaling pathway, both formation of abnormal PM blebs and F-Actin remodeling were avoided. All of these effects were initiated upstream by abnormal Tau-induced microtubule bundling, which may release the microtubule-bound guanine nucleotide exchange factor-H1 (GEF-H1) into the cytoplasm in order to activate its major effector RhoA-GTPase. These results may represent a new mechanism of Tau toxicity in which Tau-induced microtubule bundling produces activation of the Rho-GTPase-ROCK pathway that in turn mediates the remodeling of cortical Actin and PM blebbing. In AD and other tauopathies, these Tau-induced abnormalities may occur and contribute to the impairment of glial activity.

High dietary advanced glycation end products are associated with poorer spatial learning and accelerated Aß deposition in an Alzheimer mouse model.

There is growing evidence of the involvement of advanced glycation end products (AGEs) in the pathogenesis of neurodegenerative processes including Alzheimer's disease (AD) and their function as a seed for the aggregation of Aß, a hallmark feature of AD. AGEs are formed endogenously and exogenously during heating and irradiation of foods. We here examined the effect of a diet high in AGEs in the context of an irradiated diet on memory, insoluble Aß42 , AGEs levels in hippocampus, on expression of the receptor for AGEs (RAGE), and on oxidative stress in the vasculature. We found that AD-like model mice on high-AGE diet due to irradiation had significantly poorer memory, higher hippocampal levels of insoluble Aß42 and AGEs as well as higher levels of oxidative stress on vascular walls, compared to littermates fed an isocaloric diet. These differences were not due to weight gain. The data were further supported by the overexpression of RAGE, which binds to Aß42 and regulates its transport across the blood-brain barrier, suggesting a mediating pathway. Because exposure to AGEs can be diminished, these insights provide an important simple noninvasive potential therapeutic strategy for alleviating a major lifestyle-linked disease epidemic.

Difficulties associated with the structural analysis of proteins susceptible to form aggregates: The case of Tau protein as a biomarker of Alzheimer's disease.

Mass spectrometry coupled with bioaffinity separation techniques is considered a powerful tool for studying protein interactions. This work is focused on epitope analysis of tau protein, which contains two VQIXXK aggregation motifs regarded as crucial elements in the formation of paired helical filaments, the main pathological characteristics of Alzheimer's disease. To identify major immunogenic structures, the epitope extraction technique utilizing protein fragmentation and magnetic microparticles functionalized with specific antibodies was applied. However, the natural adhesiveness of some newly generated peptide fragments devalued the experimental results. Beside presumed peptide fragment specific to applied monoclonal anti-tau antibodies, the epitope extraction repeatedly revealed inter alia tryptic fragment 299-HVPGGGSVQIVYKPVDLSK-317 containing the fibril-forming motif 306-VQIVYK-311. The tryptic fragment pro-aggregation and hydrophobic properties that might contribute to adsorption phenomenon were examined by Thioflavin S and reversed-phase chromatography. Several conventional approaches to reduce the non-specific fragment sorption onto the magnetic particle surface were performed, however with no effect. To avoid methodological complications, we introduced an innovative approach based on altered proteolytic digestion. Simultaneous fragmentation of tau protein by two immobilized proteases differing in the cleavage specificity (TPCK-trypsin and α-chymotrypsin) led to the disruption of motif responsible for undesirable adhesiveness and enabled us to obtain undistorted structural data.

Identification and characterization of natural antibodies against tau protein in an intravenous immunoglobulin product.

The latest therapeutic approaches to Alzheimer disease are using intravenous immunoglobulin (IVIG) products. Therefore, the detailed characterization of target-specific antibodies naturally occurring in IVIG products is beneficial. We have focused on characterization of antibodies isolated against tau protein, a biomarker of Alzheimer's disease, from Flebogamma IVIG product. The analysis of IgG subclass distribution indicated skewing toward IgG3 in anti-tau-enriched IgG fraction. The evaluation of their reactivity and avidity with several recombinant tau forms was performed by ELISA and blotting techniques. Truncated non-phosphorylated tau protein (amino acids 155-421) demonstrated the highest reactivity and avidity index. We provide the first detailed insight into the reactivity of isolated natural antibodies against tau protein.

Levels of 17ß-Hydroxysteroid Dehydrogenase Type 10 in Cerebrospinal Fluid of People with Mild Cognitive Impairment and Various Types of Dementias.

BACKGROUND: Overexpression of the mitochondrial enzyme 17ß-hydroxysteroid dehydrogenase type 10 (17ß-HSD10, which is also known as the intracellular amyloid-ß peptide (Aß) binding protein) is observed in cortical or hippocampal regions of patients with Alzheimer's disease (AD). It appears that 17ß-HSD10 may play a role in the pathogenesis of AD. OBJECTIVE: We investigated the possibility that levels of 17ß-HSD10 in cerebrospinal fluid could be a prospective biomarker of AD. METHODS: We estimated the enzyme levels in 161 people (15 non-demented controls, 52 people with mild cognitive impairment (MCI), 35 people with probable AD, or 59 people with other types of dementia) and compared them with those of Aß(1- 42), tau, and phospho-tau. RESULTS: We found significantly higher levels of 17ß-HSD10 in people with MCI due to AD (to 109.9% ), with AD (to 120.0% ), or with other types of dementia (to 110.9% ) when compared to the control group. The sensitivity of the new biomarker to AD was 80.0% , and the specificity was 73.3% (compared to controls) or 52.5-59.1% (compared to other types of dementia). Results of multiple linear regression and of correlation analysis revealed AD-mediated changes in links between 17ß-HSD10 and Mini Mental State Examination score. CONCLUSION: It seems that changes in 17ß-HSD10 start many years before symptom onset, analogous to those in Aß1 - 42, tau, or phospho-tau and that the levels are a relatively highly sensitive but unfortunately less specific biomarker of AD. A role of 17ß-HSD10 overexpression in AD is discussed.

Differences Between Tg2576 and Wild Type Mice in the NMDA Receptor-Nitric Oxide Pathway After Prolonged Application of a Diet High in Advanced Glycation End Products.

It has been suggested that advanced glycation end (AGE) products, via cognate receptor activation, are implicated in several diseases, including Alzheimer's disease. The NMDA receptor-nitric oxide pathway appears to be influenced by AGE products and involved in the pathogenesis of this type of dementia. In this study, C57BL/6J (WT) and transgenic (Tg2576) mice expressing human mutant amyloid precursor protein were kept on prolonged (8 months) diets containing regular or high amounts of AGE products. After the decapitation of 11-months old mice, brain tissue analyses were performed [expressions of the NR1, NR2A and NR2B subunits of NMDA receptors, activities of neuronal, endothelial and inducible nitric oxide synthase (nNOS, eNOS and iNOS)]. Moreover, levels of malondialdehyde and of human amyloid ß 1-42 were estimated. We found increased activity of nNOS in WT mice maintained on a high compared to regular AGE diet; however, no similar differences were found in Tg2576 mice. In addition, we observed an increase in NR1 expression in Tg2576 compared to WT mice, both kept on a diet high in AGE products. Correlation analyses performed on mice kept on the regular AGE diet supported close links between particular subunits (NR2A-NR2B, in WT as well as in Tg2576 mice), between subunits and synthase (NR2A/NR2B-nNOS, only in WT mice) or between particular synthases (nNOS-iNOS, only in WT). Correlation analysis also revealed differences between WT mice kept on both diets (changed correlations between NR2A/NR2B-nNOS, between nNOS-eNOS and between eNOS-iNOS). Malondialdehyde levels were increased in both Tg2576 groups when compared to the corresponding WT mice, but no effects of the diets were observed. Analogously, no significant effects of diets were found in the levels of soluble or insoluble amyloid ß 1-42 in Tg2576 mice. Our results demonstrate that prolonged ingestion of AGE products can influence the NMDA receptor-nitric oxide pathway in the brain and that only WT mice, not Tg2576 mice, are able to maintain homeostasis among subunits and synthases or among particular synthases. The prolonged application of AGE products enhanced differences between 11-months old Tg2576 and WT mice regarding this pathway. Observed differences in the pathway between WT mice kept on regular or high AGE diets suggest that the prolonged application of a diet low in AGE products could have beneficial effects in older or diabetic people and perhaps also in people with Alzheimer's disease.

Hippocampal spatial position evaluation on MRI for research and clinical practice.

In clinical practice as well as in many volumetric studies we use different reorientations of the brain position towards x and y axis on the magnetic resonance imaging (MRI) scans. In order to find out whether it has an overall effect on the resulting 2D data, manual hippocampal area measurements and rotation variability of the brain (in two reoriented axes) and the skull were performed in 23 Alzheimer's disease patients and 31 healthy controls. After the MRI scanning, native brain scans (nat) were reoriented into the two different artificial planes (anterior commissure-posterior commissure axis (AC-PC) and hippocampal horizontal long axis (hipp)). Hippocampal area and temporal horn of the lateral ventricle was measured manually using freeware Image J program. We found that 1) hippocampal area of nat images is larger compared to hipp images, area of the nat images is equal to the AC-PC images and area of the hipp images is smaller compared to AC-PC images, 2) hippocampal area together with the area of the temporal horn for nat images is larger compared to hipp images, area of the hipp images is smaller compared to the AC-PC images and area of the nat images is smaller compared to the AC-PC images. The conclusion is that the measured area of the hippocampus in the native MRI is almost the same as the area of MRI reoriented only into the AC-PC axis. Therefore, when performing 2D area studies of the hippocampus or in the clinical practice we recommend usage of not-reoriented MRI images or to reorient them into the AC-PC axis. Surprising finding was that rotation of both AC-PC and hipp line towards x-axis among patients varies up to 35° and the same is true for the skull rotation so that it is not only a matter of the brain position.

Age-related differences in NMDA receptor subunits of prenatally methamphetamine-exposed male rats.

There is accumulating evidence that methamphetamine (MA) is a widely abused drug popular among pregnant women. MA exposure is associated with changes in the function of neurotransmitter systems, namely the dopaminergic, serotonergic and glutamatergic systems. Since N-methyl-D-aspartate receptors (NMDA) are affected by MA-induced glutamate release, we assessed the expression of NMDAR subunits (NR1, NR2A, and NR2B) and postsynaptic density protein 95 (PSD-95), which is connected with NMDAR. We measured the expression of these proteins in adolescent (30 days old) and adult (60 days old) rat males exposed to MA during the entire prenatal period and compared them with the same parameters in age matched saline-exposed rats. There was a significant increase in the NR1 and NR2B subunits in the hippocampus of adult males, but not in adolescent males. We identified a significant change in adult MA-induced rats when compared to adult controls for NR2A and NR2B, while in adolescent MA rats this change was close to the boundary of significance. In summary, our study suggests that prenatal MA exposure is connected with changes in NMDAR subunit expression in adult rats but not in adolescent rats.

7-MEOTA-donepezil like compounds as cholinesterase inhibitors: Synthesis, pharmacological evaluation, molecular modeling and QSAR studies.

A novel series of 7-methoxytacrine (7-MEOTA)-donepezil like compounds was synthesized and tested for their ability to inhibit electric eel acetylcholinesterase (EeAChE), human recombinant AChE (hAChE), equine serum butyrylcholinesterase (eqBChE) and human plasmatic BChE (hBChE). New hybrids consist of a 7-MEOTA unit, representing less toxic tacrine (THA) derivative, connected with analogues of N-benzylpiperazine moieties mimicking N-benzylpiperidine fragment from donepezil. 7-MEOTA-donepezil like compounds exerted mostly non-selective profile in inhibiting cholinesterases of different origin with IC50 ranging from micromolar to sub-micromolar concentration scale. Kinetic analysis confirmed mixed-type inhibition presuming that these inhibitors are capable to simultaneously bind peripheral anionic site (PAS) as well as catalytic anionic site (CAS) of AChE. Molecular modeling studies and QSAR studies were performed to rationalize studies from in vitro. Overall, 7-MEOTA-donepezil like derivatives can be considered as interesting candidates for Alzheimer's disease treatment.

Effects of ferrofluid and phytoalexin spirobrassinin on thioflavin-T-based fluorescence in cerebrospinal fluid of the elderly and multiple sclerosis patients.

It is well known that misfolded peptides/proteins can play a role in processes of normal ageing and in the pathogenesis of many diseases including Alzheimer's disease. Previously, we evaluated samples of cerebrospinal fluid from patients with Alzheimer's disease and multiple sclerosis by means of thioflavin-T-based fluorescence. We observed attenuated effects of magnetite nanoparticles operated via anti-aggregation actions on peptides/proteins from patients with Alzheimer's disease but not from those with multiple sclerosis when compared to age-related controls. In this study, we have evaluated the in vitro effects of anti-aggregation operating ferrofluid and phytoalexin spirobrassinin in the cerebrospinal fluid of patients with multiple sclerosis and Alzheimer's disease. We have found significant differences in native fluorescence (λ excitation = 440 nm, λ emission = 485 nm) of samples among particular groups (young controls < multiple sclerosis, Alzheimer's disease < old controls). Differences among groups were observed also in thioflavin-T-based fluorescence (young controls = multiple sclerosis < Alzheimer's disease < old controls) and the most marked change from native to thioflavin-T-based fluorescence was found in young controls (28-40 years old people). Both ferrofluid and spirobrassinin evoked drops in thioflavin-T-based fluorescence; however, ferrofluid was more efficient in old controls (54-75 years old people) and spirobrassinin in multiple sclerosis patients, both compared to young controls. The results are discussed especially in relation to aggregated peptides/proteins and liposoluble fluorescent products of lipid peroxidation. Based on the significant effect of spirobrassinin in vitro, we suggest that spirobrassinin may be of therapeutic value in multiple sclerosis.

Simple method for evaluation of planum temporale pyramidal neurons shrinkage in postmortem tissue of Alzheimer disease patients.

We measured the length of the pyramidal neurons in the cortical layer III in four subregions of the planum temporale (transitions into superior temporal gyrus, Heschl's gyrus, insular cortex, and Sylvian fissure) in control group and Alzheimer disease patients. Our hypothesis was that overall length of the pyramidal neurons would be smaller in the Alzheimer disease group compared to controls and also there would be right-left asymmetry in both the control and Alzheimer disease groups. We found pyramidal neuron length asymmetry only in controls--in the transition into the Sylvian fissure--and the rest of the subregions in the control group and Alzheimer disease patients did not show size difference. However, control-Alzheimer disease group pyramidal neuron length comparison revealed (a) no length difference in superior temporal gyrus transition area, (b) reversal of asymmetry in the insular transition area with left insular transition significantly shorter in the Alzheimer disease group compared to the control group, (c) both right and left Heschl's gyrus transitions significantly shorter in the Alzheimer disease group compared to the control group, and (d) right Sylvian fissure transition significantly shorter in the Alzheimer disease group compared to the control group. This neuronal length measurement method could supplement already existing neuropathological criteria for postmortem Alzheimer disease diagnostics.

Gold nanoparticle-based immuno-PCR for detection of tau protein in cerebrospinal fluid.

Tau protein in cerebrospinal fluid (CSF) is an important biomarker of Alzheimer's disease and some other brain diseases. Enzyme-linked immunosorbent assays (ELISAs) have been mostly used for quantification of tau and other biomarkers in CSF. However, these assays do not have sufficient sensitivity and dynamic range. In this study we tested the suitability of gold nanoparticles functionalized with tau-specific monoclonal antibody and oligonucleotide template for immuno-polymerase chain reaction (Nano-iPCR) quantification of tau protein in human CSF samples and compared it with ELISA, either commercial or newly developed with tyramide signal amplification. Our data indicate that Nano-iPCR is superior in sensitivity and detection range to ELISA in tau protein detection.