Impairments of N-methyl-D-aspartate receptor (NMDAR) activity have been implicated in several neuropsychiatric disorders, with pharmacological inhibition of NMDAR-mediated currents and associated neurobehavioral changes considered as a model of schizophrenia. We analyzed the effects of brief and long-term exposure of rat cortical cultures to the most prevalent endogenous modulators of NMDAR (kynurenic acid, pregnenolone sulfate, spermidine, and zinc) on neuronal viability, stimulation-induced release of glutamate, and dendritic morphology with synaptic density. Both, glutamate release and neuronal viability studies revealed no difference between the test and control groups. No differences were also observed in the number of dendritic branching and length, or density of synaptic connections and neuronal soma size. Comparison of the extent of dendritic projections and branching patterns, however, revealed enhanced distal arborization with the expansion of the dendritic area under prolonged treatment of cultures with physiological concentrations of NMDAR modulators, with differences reaching significance in spermidine and pregnenolone sulfate tests. Measurements of the density of glutamatergic synapses showed consistency across all neuronal groups, except those treated with pregnenolone sulfate, which showed a reduction of PSD-95-positive elements. Overall, our data suggest that constitutive glutamatergic activity mediated by NMDAR controls the dendritic field expansion and can influence the integrative properties of cortical neurons.
In Alzheimer's disease (AD), two mutually exclusive amino-terminal-dependent conformations have been reported to occur during the aggregation of Tau protein into neurofibrillary tangles (NFTs). An early conformation of full-length Tau, involving the bending of the amino terminus over the third repeated domain, is recognized by the Alz-50 antibody, followed by a second conformation recognized by Tau-66 antibody that depends on the folding of the proline-rich region over the third repeated domain in a molecule partially truncated at the amino- and carboxyl-termini. α-1-antichymotrypsin (ACT) is an acute phase serum glycoprotein that accumulates abnormally in the brain of AD patients, and since it is considered to promote the in vitro and in vivo aggregation of amyloid-β, we here seek further evidence that ACT may also contribute to the abnormal aggregation of Tau in AD. By analyzing brain samples from a population of AD cases under immunofluorescence and high-resolution confocal microscopy, we demonstrate here the abundant expression of ACT in hippocampal neurons, visualized as a granular diffuse accumulation, frequently reaching the nuclear compartment. In a significant number of these neurons, intracellular NFTs composed of abnormally phosphorylated and truncated Tau at Asp421 were also observed to coexist in separated regions of the cytoplasm. However, we found strong colocalization between ACT and diffuse aggregates of Tau-66-positive granules, which was not observed with Alz-50 antibody. These results suggest that ACT may play a role during the development of Tau conformational changes facilitating its aggregation during the formation of the neurofibrillary pathology in AD.
Neurodegenerative diseases (NDDs) are associated with the accumulation of a range of misfolded proteins across the central nervous system and related autoimmune responses, including the generation of antibodies and the activation of immune cells. Both innate and adaptive immunity become mobilized, leading to cellular and humoral effects. The role of humoral immunity in disease onset and progression remains to be elucidated with rising evidence suggestive of positive (protection, repair) and negative (injury, toxicity) outcomes. In this study, we review advances in research of neuron-targeting autoantibodies in the most prevalent NDDs. We discuss their biological origin, molecular diversity and changes in the course of diseases, consider their relevance to the initiation and progression of pathology as well as diagnostic and prognostic significance. It is suggested that the emerging autoimmune aspects of NDDs not only could facilitate the early detection but also might help to elucidate previously unknown facets of pathobiology with relevance to the development of precision medicine.
The nucleus-encoded 17β-hydroxysteroid dehydrogenase type 10 (17β-HSD10) regulates cyclophilin D (cypD) in the mitochondrial matrix. CypD regulates opening of mitochondrial permeability transition pores. Both mechanisms may be affected by amyloid β peptides accumulated in mitochondria in Alzheimer's disease (AD). In order to clarify changes occurring in brain mitochondria, we evaluated interactions of both mitochondrial proteins in vitro (by surface plasmon resonance biosensor) and detected levels of various complexes of 17β-HSD10 formed in vivo (by sandwich ELISA) in brain mitochondria isolated from the transgenic animal model of AD (homozygous McGill-R-Thy1-APP rats) and in cerebrospinal fluid samples of AD patients. By surface plasmon resonance biosensor, we observed the interaction of 17β-HSD10 and cypD in a direct real-time manner and determined, for the first time, the kinetic parameters of the interaction (ka 2.0 × 105 M1s-1, kd 5.8 × 104 s-1, and KD 3.5 × 10-10 M). In McGill-R-Thy1-APP rats compared to controls, levels of 17β-HSD10-cypD complexes were decreased and those of total amyloid β increased. Moreover, the levels of 17β-HSD10-cypD complexes were decreased in cerebrospinal fluid of individuals with AD (in mild cognitive impairment as well as dementia stages) or with Frontotemporal lobar degeneration (FTLD) compared to cognitively normal controls (the sensitivity of the complexes to AD dementia was 92.9%, that to FTLD 73.8%, the specificity to AD dementia equaled 91.7% in a comparison with the controls but only 26.2% with FTLD). Our results demonstrate the weakened ability of 17β-HSD10 to regulate cypD in the mitochondrial matrix probably via direct effects of amyloid β. Levels of 17β-HSD10-cypD complexes in cerebrospinal fluid seem to be the very sensitive indicator of mitochondrial dysfunction observed in neurodegeneration but unfortunately not specific to AD pathology. We do not recommend it as the new biomarker of AD.
- MeSH
- 17-hydroxysteroidní dehydrogenasy mozkomíšní mok metabolismus MeSH
- Alzheimerova nemoc metabolismus MeSH
- amyloidový prekurzorový protein beta genetika MeSH
- kinetika MeSH
- lidé MeSH
- mitochondrie metabolismus MeSH
- mozek metabolismus MeSH
- peptidylprolylisomerasa F metabolismus MeSH
- potkani transgenní MeSH
- potkani Wistar MeSH
- povrchová plasmonová rezonance MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Závěrečná zpráva o řešení grantu Agentury pro zdravotnický výzkum MZ ČR
Nestr.
We would like to exploit the obtained preliminary data in preparation of more active hybrids combining derivative of melatonin and AChE inhibitor – huprine. Such compounds should exert multipotent profile in Alzheimer ́s disease treatment by combining beneficial effect of mainly, but not exclusively, cholinergic enhancement and antioxidant activity. Funnel-like drug development process from design and synthesis though in vitro efficacy and safety evaluation to in vivo validation involving kinetic, toxic, pharmacodynamic and behavioral examination will be applied in order to select the best drug candidate. In this project, basic research (9%) represent the synthesis of novel compounds and the structure-biological activity relationship (SAR) evaluation. Applied research (91%) represents majority of the project, thus, the funnel-like selection of a drug candidate, its in vivo validation and subsequent patent protection application with commercial utilization.
V rámci řešení tohoto projektu chceme zužitkovat získaná předběžná data při přípravě učinných konjugátů obsahující derivát melatoninu a inhibitor AChE - huprin. Tyto sloučeniny by měly disponovat multipotentním profilem v léčbě Alzheimerovy choroby zejména, ale nikoliv výlučně tím, že kombinují příznivý vliv cholinergní stimulace a antioxidační aktivitu. Vývoj léčiva zahrnuje proces od návrhu a organické syntézy i přes in vitro hodnocení účinnosti a bezpečnosti léčiva po in vivo validaci zahrnující studium kinetiky, toxicity a farmakodynamiky tak, aby bylo možné vybrat nejlepšího potenciálního kandidáta na léčivo . V rámci tohoto projektu základní výzkum (9%) je reprezentován syntézou nových sloučenin a zhodnocením vztahu mezi strukturou a biologickou aktivitou (SAR). Většinový aplikovaný výzkum (91%) zde představuje selekční mechanismus pro výběr nejlepšího kadnidáta, jeho in vivo ověření učinnosti a bezpečnosti s následnou aplikací patentové ochrany včetně potenciálního komerčního využití
- MeSH
- acetylcholinesterasa MeSH
- Alzheimerova nemoc farmakoterapie MeSH
- cholinesterasové inhibitory terapeutické užití MeSH
- intravitální mikroskopie MeSH
- melatonin terapeutické užití MeSH
- tryptofan terapeutické užití MeSH
- vyvíjení léků MeSH
- Konspekt
- Patologie. Klinická medicína
- NLK Obory
- farmacie a farmakologie
- neurologie
- NLK Publikační typ
- závěrečné zprávy o řešení grantu AZV MZ ČR
Cíl: Cílem práce bylo porovnat metody ELISA pro stanovení hladin tripletu proteinů – celkového (t-tau) a fosforylovaného tau (p-tau) proteinů a β-amyloidu (1– 42) (Aβ42) v mozkomíšním moku (MMM) od dvou výrobců Fujirebio a EUROIMMUN a zároveň vytvořit orientační vodítka pro normální koncentrace u české populace od kognitivně zdravých starších osob. Soubor a metodika: Koncentrace likvorového tripletu proteinů od 38 kognitivně zdravých osob starších 55 let (19 mužů, průměrný věk 67 ± 8 let) byly změřeny jak soupravami ELISA INNOTEST, tak soupravami EUROIMMUN AG. Výsledky: Výsledky srovnání metod pro t-tau a p-tau proteiny a Aβ42 pomocí rozdílového grafu dle Blanda a Altmana ukazují, že hodnoty rozdílů mezi metodami ležely v rozsahu limitů shody (průměr ± 1,96 SD) až na ojedinělé výjimky odlehlých hodnot. Hodnoty korelačních koeficientů svědčí pro významnou shodu pro soupravy na stanovení t-tau proteinu. Na základě našich výsledků a zkušeností navrhujeme rozdělit koncentrace likvorového tripletu na tři skupiny: 1. snížené; 2. nerozhodné a hraniční; 3. zvýšené koncentrace v ng/ l na základě našeho konsenzu takto: t-tau protein < 280– 400 >, p181-tau protein < 50– 60 > a Aβ42 < 430– 480 >. Závěr: Soupravy ELISA obou výrobců jsou vhodné ke stanovení tripletu proteinů v MMM. Pro jejich referenční rozmezí lze orientačně využít naše výsledky, založené na reálném měření vzorků pečlivě vybraných a vyšetřených zdravých starších osob z ČR, nebo normy na základě našeho společného názoru.
Aim: The aim of the study was to compare ELISA methods for determination of triplet protein levels – total (t-tau) and phosphorylated tau (p-tau) proteins and β-amyloid (1– 42) (Aβ42) in cerebrospinal fl uid (CSF) from manufacturers Fujirebio and EUROIMMUN; and to provide guidance for normal concentrations from cognitively healthy elderly people. Patients and methods: The group consisted of 38 cognitively healthy persons over 55 years of age (19 men) whose average age was 67 ± 8 years. In each CSF sample, we examined the protein triplet by both ELISA INNOTEST and EUROIMMUN AG kits. Results: A comparison of ELISA methods for determination of t-tau and p-tau proteins and Aβ42 us ing the Bland and Altman difference graph shows that the differences between methods, with the exception of isolated outliers, were within the range of compliance (mean ± 1.96 SD). Cor relation coeffi cient values suggest significant agreement for t-tau protein as say kits. Based on our results and experience, we propose to clas sify CSF concentrations into three groups: 1. decreased; 2. questionable and borderline; 3. increased concentrations in ng/ L based on our consensus as follows: t-tau protein < 280– 400 >, p181-tau protein < 50– 60 > and Aβ42 < 430– 480 >. Conclusion: ELISA kits from two producers are suitable for determination of the triplet proteins in CSF. Our results based on real measurements of samples of carefully selected and examined healthy elderly people from the Czech Republic or the norms based on our consensus recom mendation can be used as reference range of the triplet.
- MeSH
- Alzheimerova nemoc * diagnóza MeSH
- amyloidní beta-protein * analýza MeSH
- ELISA MeSH
- klinická studie jako téma MeSH
- lidé středního věku MeSH
- lidé MeSH
- mozkomíšní mok MeSH
- proteiny tau analýza MeSH
- proteiny v mozkomíšním moku * analýza MeSH
- senioři MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
AIM: We aimed to characterize the role of mitochondrial 17β-hydroxysteroid dehydrogenase type 10 (17β-HSD10) overexpression in multiple sclerosis (MS) and to evaluate its use as a biomarker. Materials & methods: We estimated levels of 17β-HSD10, amyloid β 1-42, cyclophilin D, 17β-HSD10-cyclophilin D complexes or 17β-HSD10-parkin complexes in cerebrospinal fluid (CSF) samples. RESULTS: The increase in 17β-HSD10 levels or in 17β-HSD10-parkin complexes and links to leukocytes were found only in relapsing-remitting MS. The sensitivity of the biomarker was 64%, the specificity equaled 60-63% compared with controls. CONCLUSION: Increased CSF levels of 17β-HSD10 in later stages of MS could be interpreted via its upregulation in demyelinated neuronal axons. CSF levels of 17β-HSD10 are not the valuable biomarker for the early diagnosis or for the progression of MS.
- MeSH
- 3-hydroxyacyl-CoA-dehydrogenasy MeSH
- amyloidní beta-protein MeSH
- biologické markery MeSH
- dospělí MeSH
- lidé MeSH
- peptidové fragmenty MeSH
- roztroušená skleróza MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
AIM: We aimed to characterize the role of mitochondrial 17β-hydroxysteroid dehydrogenase type 10 (17β-HSD10) overexpression in multiple sclerosis (MS) and to evaluate its use as a biomarker. Materials & methods: We estimated levels of 17β-HSD10, amyloid β 1-42, cyclophilin D, 17β-HSD10-cyclophilin D complexes or 17β-HSD10-parkin complexes in cerebrospinal fluid (CSF) samples. RESULTS: The increase in 17β-HSD10 levels or in 17β-HSD10-parkin complexes and links to leukocytes were found only in relapsing-remitting MS. The sensitivity of the biomarker was 64%, the specificity equaled 60-63% compared with controls. CONCLUSION: Increased CSF levels of 17β-HSD10 in later stages of MS could be interpreted via its upregulation in demyelinated neuronal axons. CSF levels of 17β-HSD10 are not the valuable biomarker for the early diagnosis or for the progression of MS.
- MeSH
- 3-hydroxyacyl-CoA-dehydrogenasy mozkomíšní mok MeSH
- amyloidní beta-protein mozkomíšní mok MeSH
- biologické markery mozkomíšní mok MeSH
- dospělí MeSH
- lidé MeSH
- peptidové fragmenty mozkomíšní mok MeSH
- roztroušená skleróza mozkomíšní mok MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The presence of pre-existing natural antibodies against Alzheimer's disease (AD) pathological proteins might interfere with immune responses to therapeutic vaccination with these proteins. We aimed to compare levels of antibodies in CSF and serum: We observed higher reactivity of natural tau-reactive antibodies towards phosphorylated bovine tau protein than to human recombinant (non-phosphorylated) tau protein. Males with MCI-AD had higher amounts of these antibodies than corresponding controls. Concentrations of antibodies were lower in females with the MCI-AD than in control females. These findings may have implications for tau vaccination trials.
- MeSH
- Alzheimerova nemoc krev imunologie MeSH
- autoprotilátky analýza krev MeSH
- demence krev imunologie MeSH
- druhová specificita MeSH
- fosfoproteiny chemie imunologie MeSH
- fosforylace MeSH
- kognitivní dysfunkce krev imunologie MeSH
- lidé středního věku MeSH
- lidé MeSH
- pohlavní dimorfismus * MeSH
- posttranslační úpravy proteinů MeSH
- proteiny tau chemie imunologie MeSH
- rekombinantní proteiny imunologie MeSH
- reprodukovatelnost výsledků MeSH
- senioři MeSH
- skot MeSH
- specificita protilátek MeSH
- zvířata MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- skot MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
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-β isoform and to simultaneously stimulate CHT1. Some derivatives, when coincubated with amyloid β, 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.
- MeSH
- alkylace MeSH
- amyloidní beta-protein účinky léků MeSH
- cholesterol metabolismus MeSH
- cholin metabolismus MeSH
- cholinesterasové inhibitory chemická syntéza farmakologie MeSH
- fluidita membrány účinky léků MeSH
- hipokampus účinky léků metabolismus MeSH
- krysa rodu rattus MeSH
- kvartérní amoniové sloučeniny chemie farmakologie MeSH
- membránové mikrodomény účinky léků MeSH
- mozková kůra účinky léků MeSH
- potkani Wistar MeSH
- proteiny přenášející kationty účinky léků MeSH
- stereoizomerie MeSH
- synaptozomy účinky léků metabolismus MeSH
- takrin analogy a deriváty chemická syntéza farmakologie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
