Cílem studie bylo zjistit prevalenci antinukleozomálních protilátek u pacientů se SLE (systémový lupus erythematodes), jiných autoimunitních onemocnění a u zdravých kontrol a určit optimální diagnostickou soupravu umožňující rutinní diagnostiku antinukleozomálních protilátek (anti-Ncs Ab). Dalším úkolem bylo porovnat přítomnost anti-Ncs Ab s výskytem anti-ds DNA protilátek a antihistonových protilátek. Pacienti a metody. Bylo vyšetřeno 128 pacientů, a to 52 pacientů se SLE, 14 s lupusovou nefritidou, 8 s neuropsychiatrickým lupusem (NPSLE), 39 s difuzní formou sklerodermie, 15 se Sjögrenovým syndromem (SS, primárním či sekundárním) a 51 zdravých kontrol srovnatelných distribucí k věku a pohlaví. K detekci antinukleozomálních protilátek byly použity komerční soupravy ELISA od 3 výrobců, a to Orgentec, BL Diagnostika a Euroimmun. U sledovaného souboru pacientů byly dále testovány anti-ds DNA protilátky a antihistonové protilátky. Výsledky stanovení frekvence pozitivity anti-Ncs Ab u vyšetřovaného souboru byly u jednotlivých diagnostických souprav v pořadí Orgentec, BL Diagnostika a Euroimmun následující: v souboru SLE 99,3 %, 99,3 % a 72,3 %, u lupusové nefritidy 100 %, 100 % a 79 %, u neuropsychiatrického lupusu 100 %, 100 % a 62,5%, u sklerodermie 82,1 %, 82,1 % a 17,9 %, u Sjögrenova syndromu 14%, 14% a 7 % a u zdravých kontrol 8 %, 8 % a 6 %. U pacientů se SLE byla frekvence pozitivity anti-Ncs Ab, anti-dsDNA a antihistonových protilátek 72,3 %, 63 % a 54 % v porovnání s nemocnými se SCL a SS (17,9 %, 8 %, 5 % a 7 %, 7 % a 0 %). Pacienti s lupusovou nefritidou měli statisticky zvýšenou hladinu cirkulujících antinukleozomálních protilátek ve srovnání s kontrolami i ostatními vyšetřovanými skupinami. Závěr. Vzhledem k poměrně vysokému výskytu pozitivních výsledků (82,1 %, p<0,001) získanými soupravami výrobců Orgentec a BL- Diagnostika u skupiny pacientů se SCL se autoři studie rozhodli rutinně používat k detekci antinukleozomálních protilátek ELISA soupravu firmy Euroimmun (17,9 %, NS). Tyto hodnoty také odpovídaly výsledkům publikovaných zahraničních studií. Ve shodě s literárními údaji byla zaznamenána vysoká prevalence anti-Ncs Ab u pacientů se SLE (62,3-79 %) a srovnatelná prevalence anti-Ncs Ab u kontrolního souboru zdravých (6 %).

The aim of this study is to evaluate three commercial anti-nucleosome enzyme-linked immunosorbent assays (ELISA), to find the presence anti-nucleosomal antibodies (Ncs–Ab) and to report our experience with the detection of anti-nucleosome (anti-Nuc) antibodies (Ab) in series of SLE patients, and to compare these results with those of anti-histone and anti-dsDNA Ab. Patients and methods. We selected 128 patients (106 female, 22 male, mean age 40) including 52 patients with SLE, 14 with lupus nephritis, 8 with neuropsychiatric lupus (NPSLE) and as controls 39 with systemic sclerosis (SCL), 15 with Sjögren syndrome and 51 healthy controls (40 female, 11 male, mean age 42) and to evaluate three commercial ELISA kits (Orgentec, BL Diagnostika, Euroimmun). The sera were assayed for the levels of anti-ds DNA (ELISA, Orgentec), anti-histone (INNO LIA ANA Update, Innogenetics) also. Results. The prevalence of positive anti-nucleosome was by detection Orgentec, BL Diagnostika and Euroimmun in group of patients with SLE 99,3 %, 99,3% and 72,3%, with lupus nephritis 100 %, 100 % and 79 %, with NPSLE 100 %, 100 % and 62,3%, with SCL 82,1 %, 82,1 %, 17,9 % , with SS 14%, 14% and 7% and with healthy controls 8 %, 8 % and 6 %, respectively. The frequencies of positive anti-nucleosome, anti-dsDNA and anti-histone antibodies were in group of patients with SLE 73 %, 63 % and 54%, with SCL 17,9 %, 8 %, 5 % and with SS 7 %, 7 % and 0 %, respectively. Patients with SLE have increased levels of anti-nucleosome antibodies in their sera compared with healthy controls, SCL patients and patients with SS. Conclusion. Based on frequency of positive observations of anti-Ncs Ab in the group of patients suffering with SLE (62,3-79%) and discrepancy findings in the group with SCL patiens (82,1 %, p<0,001) by using Orgentec and BL Diagnostika we decided to introduce the kit by Euroimmun (17,9 %, NS) for rutine use. The results obstained using this kit are in good corelation with results published so far. All the three autoantibodies were present more frequently in cases with lupus and this corelation was significant in statistical means.

• Klíčová slova
• Orgentec, BL Diagnostika, Euroimmun,
• MeSH
• antinukleární protilátky krev   MeSH
• difuzní sklerodermie diagnóza   MeSH
• dospělí MeSH
• ELISA metody   MeSH
• finanční podpora výzkumu jako téma MeSH
• lidé MeSH
• reagenční diagnostické soupravy statistika a číselné údaje   MeSH
• Sjögrenův syndrom diagnóza   MeSH
• systémový lupus erythematodes diagnóza   MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• srovnávací studie MeSH

OBJECTIVE: While circulating nucleosome levels are high in obese mouse models, it is unknown where these nucleosomes originate from and whether they are a marker of cardio-metabolic health in humans. Here, we aimed to determine whether an association exists between circulating nucleosomes and the risk of developing obesity, metabolic syndrome (MetS) and/or a dysfunctional cardiovascular performance. METHODS: We randomly selected 120 participants of the Kardiovize Brno 2030 study across three BMI strata: BMI 18-25, 25-30, and > 30. We assessed the association between circulating nucleosome levels and the risk of obesity, MetS, and poor cardiovascular health. We then cultured human neutrophils, adipocytes, and hepatoma cells to study nucleosome origins in a fat-rich environment. RESULTS: Circulating nucleosome levels positively correlated with BMI (R = 0.602, p < 0.05), fatty liver index (R = 0.622, p < 0.05), left ventricular mass (R = 0.457, p < 0.05), and associated with MetS (p < 0.001) and poor cardiovascular health (p < 0.001). Incubating neutrophils with 1-10 μM free fatty acids triggered nucleosome production without concomitant cell death. Nucleosomes were not produced during pre-adipocyte differentiation or upon incubation of hepatic cells with palmitic acid. CONCLUSIONS: Neutrophils are a bona fide source of circulating nucleosomes in an obesogenic environment and in overweight/obese patients. High nucleosome levels are associated with MetS and cardiovascular performance, and might represent novel candidate biomarkers for cardio-metabolic health.

• MeSH
• buňky Hep G2 MeSH
• dospělí MeSH
• index tělesné hmotnosti MeSH
• kardiovaskulární nemoci etiologie   MeSH
• kultivované buňky MeSH
• lidé středního věku MeSH
• lidé MeSH
• metabolický syndrom krev   komplikace   MeSH
• nadváha krev   komplikace   metabolismus   MeSH
• neutrofily cytologie   metabolismus   MeSH
• nukleozomy metabolismus   MeSH
• obezita krev   komplikace   metabolismus   MeSH
• senioři MeSH
• tukové buňky cytologie   metabolismus   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

OBJECTIVE: To determine the amniotic fluid nucleosome concentrations in pregnancies that are complicated by preterm prelabor rupture of membranes and their correlation to microbial invasion of the amniotic cavity (MIAC), histologic chorioamnionitis (HCA), and their association with neonatal outcomes. METHODS: Eighty-nine women with singleton pregnancies were included in this study. Amniotic fluid was collected, and nucleosome concentration in the amniotic fluid was determined using enzyme-linked immunosorbent assay. RESULT: There were no differences observed in the amniotic fluid nucleosome concentrations in women with or without MIAC. The presence of HCA (with chorioamnionitis: median 0.5; without chorioamnionitis: median 0.21; p = 0.01) and funisitis (with funisitis: median 0.85; without funisitis: median 0.22; p = 0.0008) was associated with higher nucleosome concentrations using crude analysis; however, this was not significant after adjusting for gestational age (p = 0.12 for both). A negative correlation was observed between amniotic fluid nucleosome concentrations and gestational age (ρ = -0.52; p < 0.0001). There was no association identified between amniotic fluid nucleosome concentration and neonatal morbidity. CONCLUSIONS: Amniotic fluid nucleosome concentrations remained a stable physiologic constituent in pregnancies complicated by preterm prelabor rupture of membranes, and these concentrations were gestational age dependent. Neither MIAC nor HCA significantly affected amniotic fluid nucleosome concentrations.

• MeSH
• amniocentéza MeSH
• amnion mikrobiologie   MeSH
• bronchopulmonální dysplazie diagnóza   MeSH
• chorioamnionitida mikrobiologie   patologie   MeSH
• dospělí MeSH
• ELISA MeSH
• gestační stáří MeSH
• lidé MeSH
• novorozenec MeSH
• nukleozomy chemie   ultrastruktura   MeSH
• plodová voda chemie   MeSH
• předčasný odtok plodové vody metabolismus   MeSH
• syndrom respirační tísně novorozenců diagnóza   MeSH
• těhotenství MeSH
• výsledek těhotenství * MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• novorozenec MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Chromatin adopts a diversity of regular and irregular fiber structures in vitro and in vivo. However, how an array of nucleosomes folds into and switches between different fiber conformations is poorly understood. We report the 9.7 Å resolution crystal structure of a 6-nucleosome array bound to linker histone H1 determined under ionic conditions that favor incomplete chromatin condensation. The structure reveals a flat two-start helix with uniform nucleosomal stacking interfaces and a nucleosome packing density that is only half that of a twisted 30-nm fiber. Hydroxyl radical footprinting indicates that H1 binds the array in an on-dyad configuration resembling that observed for mononucleosomes. Biophysical, cryo-EM, and crosslinking data validate the crystal structure and reveal that a minor change in ionic environment shifts the conformational landscape to a more compact, twisted form. These findings provide insights into the structural plasticity of chromatin and suggest a possible assembly pathway for a 30-nm fiber.

• MeSH
• DNA chemie   genetika   metabolismus   MeSH
• elektronová kryomikroskopie MeSH
• Escherichia coli genetika   metabolismus   MeSH
• exprese genu MeSH
• genetické vektory chemie   metabolismus   MeSH
• histony chemie   genetika   metabolismus   MeSH
• hydroxylový radikál chemie   MeSH
• interakční proteinové domény a motivy MeSH
• klonování DNA MeSH
• konformace proteinů, alfa-helix MeSH
• konformace proteinů, beta-řetězec MeSH
• krystalografie rentgenová MeSH
• lidé MeSH
• molekulární modely MeSH
• multimerizace proteinu MeSH
• nukleozomy chemie   metabolismus   ultrastruktura   MeSH
• osmolární koncentrace MeSH
• protein 1 vytvářející nukleozómy chemie   genetika   metabolismus   MeSH
• rekombinantní proteiny chemie   genetika   metabolismus   MeSH
• vazba proteinů MeSH
• vazebná místa MeSH
• Xenopus laevis MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

A-tracts are functionally important DNA sequences which induce helix bending and have peculiar structural properties. While A-tract structure has been qualitatively well characterized, their mechanical properties remain controversial. A-tracts appear structurally rigid and resist nucleosome formation, but seem flexible in DNA looping. In this work, we investigate mechanical properties of symmetric AnTn and asymmetric A2n tracts for n = 3, 4, 5 using two types of coarse-grained models. The first model represents DNA as an ensemble of interacting rigid bases with non-local quadratic deformation energy, the second one treats DNA as an anisotropically bendable and twistable elastic rod. Parameters for both models are inferred from microsecond long, atomic-resolution molecular dynamics simulations. We find that asymmetric A-tracts are more rigid than the control G/C-rich sequence in localized distortions relevant for nucleosome formation, but are more flexible in global bending and twisting relevant for looping. The symmetric tracts, in contrast, are more rigid than asymmetric tracts and the control, both locally and globally. Our results can reconcile the contradictory stiffness data on A-tracts and suggest symmetric A-tracts to be more efficient in nucleosome exclusion than the asymmetric ones. This would open a new possibility of gene expression manipulation using A-tracts.

• MeSH
• adenin chemie   MeSH
• biomechanika MeSH
• DNA chemie   MeSH
• entropie MeSH
• konformace nukleové kyseliny MeSH
• nukleozomy chemie   MeSH
• poly A chemie   MeSH
• sekvence nukleotidů MeSH
• simulace molekulární dynamiky MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

• MeSH
• DNA účinky záření   MeSH
• lidé MeSH
• molekulární modely MeSH
• nukleozomy účinky záření   MeSH
• poškození DNA MeSH
• techniky in vitro MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH

We report a series of microarray-based comparisons of gene expression in the leaf and crown of the winter barley cultivar Luxor, following the exposure of young plants to various periods of low (above and below zero) temperatures. A transcriptomic analysis identified genes which were either expressed in both the leaf and crown, or specifically in one or the other. Among the former were genes responsible for calcium and abscisic acid signalling, polyamine synthesis, late embryogenesis abundant proteins and dehydrins. In the crown, the key organ for cereal overwintering, cold treatment induced transient changes in the transcription of nucleosome assembly genes, and especially H2A and HTA11, which have been implicated in cold sensing in Arabidopsis thaliana. In the leaf, various heat-shock proteins were induced. Differences in expression pattern between the crown and leaf were frequent for genes involved in certain pathways responsible for osmolyte production (sucrose and starch, raffinose, γ-aminobutyric acid metabolism), sugar signalling (trehalose metabolism) and secondary metabolism (lignin synthesis). The action of proteins with antifreeze activity, which were markedly induced during hardening, was demonstrated by a depression in the ice nucleation temperature.

• MeSH
• aklimatizace genetika   MeSH
• GABA genetika   metabolismus   MeSH
• genetická transkripce MeSH
• histony genetika   MeSH
• ječmen (rod) genetika   metabolismus   MeSH
• lignin biosyntéza   genetika   MeSH
• listy rostlin genetika   metabolismus   MeSH
• metabolismus sacharidů genetika   MeSH
• nízká teplota MeSH
• nukleozomy metabolismus   MeSH
• proteiny tepelného šoku genetika   MeSH
• regulace genové exprese u rostlin MeSH
• roční období MeSH
• stanovení celkové genové exprese MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The histone H3 variant CENP-A marks centromeres epigenetically and is essential for mitotic fidelity. Previous crystallographic studies of the CENP-A nucleosome core particle (NCP) reconstituted with a human α-satellite DNA derivative revealed both DNA ends to be highly flexible, a feature important for CENP-A mitotic functions. However, recent cryo-EM studies of CENP-A NCP complexes comprising primarily Widom 601 DNA reported well-ordered DNA ends. Here, we report the cryo-EM structure of the CENP-A 601 NCP determined by Volta phase-plate imaging. The data reveal that one ('left') 601 DNA end is well ordered whereas the other ('right') end is flexible and partly detached from the histone core, suggesting sequence-dependent dynamics of the DNA termini. Indeed, a molecular dynamics simulation of the CENP-A 601 NCP confirmed the distinct dynamics of the two DNA extremities. Reprocessing the image data using two-fold symmetry yielded a cryo-EM map in which both DNA ends appeared well ordered, indicating that such an artefact may inadvertently arise if NCP asymmetry is lost during image processing. These findings enhance our understanding of the dynamic features that discriminate CENP-A from H3 nucleosomes by revealing that DNA end flexibility can be fine-tuned in a sequence-dependent manner.

• MeSH
• DNA chemie   MeSH
• elektronová kryomikroskopie MeSH
• lidé MeSH
• nukleozomy chemie   ultrastruktura   MeSH
• protein CENP-A chemie   MeSH
• simulace molekulární dynamiky MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Intramural MeSH

• MeSH
• DNA MeSH
• molekulární sekvence - údaje MeSH
• mutageneze cílená MeSH
• nukleozomy MeSH
• repetitivní sekvence nukleových kyselin MeSH
• sekvence nukleotidů MeSH
• tabák MeSH

• MeSH
• antinukleární protilátky MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• senioři MeSH
• systémový lupus erythematodes MeSH
• thrombomodulin MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH