Novorozenecká sepse je hlavní příčinou celkové mortality u dětí do 5 let. Časná novorozenecká sepse má mortalitu až 30%. Přesná a rychlá diagnóza a léčba je zásadní pro prevenci závažné morbidity i mortality. Klinická prezentace novorozenecké sepse je nespecifická a obecně užívané biomarkery mají nízkou prediktivní hodnotu. Morbidita a mortalita u sepse může být ovlivněna pochopením patofyziologie defenzivní zánětové odpovědi na bakteriální invazi, která porušuje integritu životních funkcí novorozence. Sepse je doprovázena zánětovou odpovědí spouštěnou endotelem. V průběhu sepse dochází k aktivaci a poškození endotelových buněk. Cílem projektu je detekce biomarkerů endoteliální aktivace a poškození při novorozenecké sepsi. Solubilní endoteliální faktory, endoteliální mikrovezikuly a microRNA budou analyzovány v krvi zralých a mírně nezralých novorozenců s podezřením na časnou neonatální sepsi. Průkaz biomarkerů endoteliálního poškození může významně zlepšit časnou diagnostiku závažných novorozeneckých infekcí a snížit novorozeneckou mortalitu a morbiditu.; Neonatal sepsis is a leading cause of mortality in children younger than 5 years. Early-onset sepsis has mortality rates up to 30%. Accurate diagnosis and treatment are crucial to prevent severe morbidity and mortality. The clinical presentation of neonatal sepsis is often subtle and nonspecific, and commonly used biomarkers have low predictive values. Mortality and morbidity can be influenced by understanding the pathophysiology of defensive inflammatory response to bacterial invasion causing impairment of health status of the newborn. Sepsis is associated with inflammatory response, which include endothelial activation and damage. The aim of our study is to explore biomarkers of endothelial activation and damage during neonatal sepsis. Soluble endothelium-specific proteins, endothelial microvesicles and microRNA will be evaluated in blood of the newborns with early-onset sepsis risk. The introduction of endothelial injury biomarkers may significantly improve early diagnosis and treatment of severe neonatal infections and potentially decrease neonatal mortality and morbidity.

• Keywords
• microRNA, microRNA, biomarkery, biomarkers, sepse, sepsis, mortalita, mortality, morbidita, morbidity, poškození endotelu, endothelial injury, novorozenec, newborn, mikrovezikuly buněčných membrán, cell-derived membrane microvesicles,

• NML Publication type
• závěrečné zprávy o řešení grantu AZV MZ ČR

Prions are responsible for a number of lethal neurodegenerative and transmissible diseases in humans and animals. Extracellular vesicles, especially small exosomes, have been extensively studied in connection with various diseases. In contrast, larger microvesicles are often overlooked. In this work, we compared the ability of large extracellular vesicles (lEVs) and small extracellular vesicles (sEVs) to spread prions in cell culture. We utilized CAD5 cell culture model of prion infection and isolated lEVs by 20,000×g force and sEVs by 110,000×g force. The lEV fraction was enriched in β-1 integrin with a vesicle size starting at 100 nm. The fraction of sEVs was partially depleted of β-1 integrin with a mean size of 79 nm. Both fractions were enriched in prion protein, but the lEVs contained a higher prion-converting activity. In addition, lEV infection led to stronger prion signals in both cell cultures, as detected by cell and western blotting. These results were verified on N2a-PK1 cell culture. Our data suggest the importance of lEVs in the trafficking and spread of prions over extensively studied small EVs.

• MeSH
• Cell Culture Techniques MeSH
• Exosomes * metabolism   MeSH
• Extracellular Vesicles * metabolism   MeSH
• Integrins metabolism   MeSH
• Humans MeSH
• Prions * metabolism   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Endothelial dysfunction (ED) is an important marker of future atherosclerosis and cardiovascular disease, especially in people with diabetes. This article summarizes the evidence on endothelial dysfunction in people with diabetes and adds different perspectives that can affect the presence and severity of ED and its consequences. We highlight that data on ED in type 1 diabetes are lacking and discuss the relationship between ED and arterial stiffness. Several interesting studies have been published showing that ED modulates microRNA, microvesicles, lipid levels, and the endoplasmatic reticulum. A better understanding of ED could provide important insights into the microvascular complications of diabetes, their treatment, and even their prevention.

• MeSH
• Atherosclerosis * complications   MeSH
• Diabetes Mellitus, Type 1 * complications   MeSH
• Cardiovascular Diseases * complications   MeSH
• Humans MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Review MeSH

In the ciliate Tetrahymena thermophila, lysosome-related organelles called mucocysts accumulate at the cell periphery where they secrete their contents in response to extracellular events, a phenomenon called regulated exocytosis. The molecular bases underlying regulated exocytosis have been extensively described in animals but it is not clear whether similar mechanisms exist in ciliates or their sister lineage, the Apicomplexan parasites, which together belong to the ecologically and medically important superphylum Alveolata. Beginning with a T. thermophila mutant in mucocyst exocytosis, we used a forward genetic approach to uncover MDL1 (Mucocyst Discharge with a LamG domain), a novel gene that is essential for regulated exocytosis of mucocysts. Mdl1p is a 40 kDa membrane glycoprotein that localizes to mucocysts, and specifically to a tip domain that contacts the plasma membrane when the mucocyst is docked. This sub-localization of Mdl1p, which occurs prior to docking, underscores a functional asymmetry in mucocysts that is strikingly similar to that of highly polarized secretory organelles in other Alveolates. A mis-sense mutation in the LamG domain results in mucocysts that dock but only undergo inefficient exocytosis. In contrast, complete knockout of MDL1 largely prevents mucocyst docking itself. Mdl1p is physically associated with 9 other proteins, all of them novel and largely restricted to Alveolates, and sedimentation analysis supports the idea that they form a large complex. Analysis of three other members of this putative complex, called MDD (for Mucocyst Docking and Discharge), shows that they also localize to mucocysts. Negative staining of purified MDD complexes revealed distinct particles with a central channel. Our results uncover a novel macromolecular complex whose subunits are conserved within alveolates but not in other lineages, that is essential for regulated exocytosis in T. thermophila.

• MeSH
• Exocytosis genetics   MeSH
• Lysosomes metabolism   MeSH
• Organelles metabolism   MeSH
• Secretory Vesicles genetics   metabolism   MeSH
• Tetrahymena thermophila * genetics   MeSH
• Tetrahymena * MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Extramural MeSH

Chorioamnionitida je jednou z nejčastějších příčin předčasného porodu, která velmi často vede k rozvoji syndromu systémové zánětlivé odpovědi plodu (fetal inflammatory response syndrome - FIRS). FIRS je vysoce rizikovým faktorem perinatální mortality a dlouhodobé neonatální morbidity (myokardiální dysfunkce, bronchopulmonální dysplázie a poškození mozku). FIRS je definován zvýšenou systémovou koncentrací cytokinů plodu. Nedávno publikované práce prokazují, že krevní destičky a destičkové mikropartikule mají důležitou úlohu při zánětové odpovědi organismu. Náš projekt je zaměřen na zjištění stavu destiček a přítomnosti destičkových mikropartikulí v pupečníkové krvi u nedonošených dětí ve vztahu k chorioamnionitidě pomocí kvantitativní průtokové cytometrie. Výsledky dostatečně reprezentativního vzorku u nezralých novorozenců budou analyzovány ve vztahu k závažným klinickým komplikacím. Cílem našeho projektu je vývoj nové diagnostické metody FIRS, která by umožňovala lepší predikci závažných klinických komplikací a vedla k individualizaci péče u předčasně narozených dětí.; The most common cause of preterm birth is chorioamnionitis which often leads to the development of fetal inflammatory response syndrome (FIRS). FIRS is a major risk factor for perinatal mortality, but also for early ad long-term neonatal morditidy, such as impaired cardiac function, bronchopulmonary dysplasia and brain injury. FIRS is defined by increased systemic inflammatory cytokine concentrations. Recently it became clear that blood platelets and platelet microparticles play an important role in many inflammatory conditions. Our project is aimed on a study of platelet status and presence of platelet microparticles in cord blood of preterm infants with and without chorioamnionitis. We plan to utilize quantitative flow cytometry for gathering representative data and correlate the results with the clinical outcome of the enrolled preterm birth infants. The goal of our project is to develop new diagnostic method of FIRS which would allow better prediction of clinical complications and individualization of the clinical care for preterm born infants.

• MeSH
• Chorioamnionitis diagnosis   etiology   MeSH
• Fetal Blood MeSH
• Pregnancy Complications, Infectious MeSH
• Humans MeSH
• Cell-Derived Microparticles pathology   MeSH
• Morbidity MeSH
• Infant, Premature MeSH
• Fetus cytology   pathology   MeSH
• Premature Birth MeSH
• Flow Cytometry MeSH
• Systemic Inflammatory Response Syndrome diagnosis   etiology   MeSH
• Platelet Function Tests MeSH
• Check Tag
• Humans MeSH

• Conspectus
• Pediatrie
• NML Fields
• perinatologie a neonatologie
• infekční lékařství
• cytologie, klinická cytologie
• NML Publication type
• závěrečné zprávy o řešení grantu AZV MZ ČR

Celý svůj profesionální život jsem se zabýval vývojovou fyziologií a je tedy logické, že jsem se musel setkat s otázkou o významu mateřského mléka (zvláště když jsem celou jednu kapitolu svého výzkumu zasvětil otázkám výživy a vývoje CNS). Nejde přitom jenom o otázky složení – kvality a kvantity významu jednotlivých komponent pro vývoj novorozence. Jde rovněž o vztahy vznikající během tohoto procesu (kojení), a to pro vývojové trendy, pro jejich optimalizaci. Jedná se pochopitelně i o takových, které mohou představovat zcela reálně rizika. Pokládám za užitečné představit čtenářům pouze určité aspekty a úseky. Vím, že v učebnicích jsou v tomto směru informace spíše povšechného charakteru. Navíc v posledních dekádách došlo ve výzkumu komponent mateřského mléka k poměrně k velkým posunům.

All my professional life I have been dealing with developmental physiology, so it is logical that I had to meet the question and importance of breast milk (especially when I gave a whole chapter of my research to the issues of nutrition and CNS development). It is not just a question of composition-quality and quantity but also the importance of individual components for the development of the newborn. Important are also the relations that arise during this process (breastfeeding) for the developmental trends, and for their optimization. Of course, these are also factors that can pose real risks. I find it useful to present readers with only certain aspects and questions. I know that common textbooks bring only more general information on this topic. In addition, in recent decades there have been comparatively large progress in research on the breast milk components.

• MeSH
• Dietary Fats MeSH
• Phospholipids physiology   MeSH
• Glycoproteins physiology   MeSH
• Lactation physiology   MeSH
• Humans MeSH
• RNA, Small Untranslated MeSH
• Fatty Acids chemistry   MeSH
• Milk, Human * chemistry   physiology   microbiology   MeSH
• Microbiota physiology   MeSH
• Mammary Glands, Human * anatomy & histology   embryology   physiology   MeSH
• Oxytocin physiology   MeSH
• Secretory Vesicles physiology   MeSH
• Transcytosis physiology   MeSH
• Check Tag
• Humans MeSH
• Female MeSH

Outer membrane vesicles (OMVs), nanoparticles released by Shiga toxin-producing Escherichia coli (STEC), have been identified as novel efficient virulence tools of these pathogens. STEC O157 OMVs carry a cocktail of virulence factors including Shiga toxin 2a (Stx2a), cytolethal distending toxin V (CdtV), EHEC hemolysin, flagellin, and lipopolysaccharide. OMVs are taken up by human intestinal epithelial and microvascular endothelial cells, the major targets during STEC infection, and deliver the virulence factors into host cells. There the toxins separate from OMVs and are trafficked via different pathways to their target compartments, i.e., the cytosol (Stx2a-A subunit), nucleus (CdtV-B subunit), and mitochondria (EHEC hemolysin). This leads to a toxin-specific host cell injury and ultimately apoptotic cell death. Besides their cytotoxic effects, STEC OMVs trigger an inflammatory response via their lipopolysaccharide and flagellin components. In this chapter, we describe methods for the isolation and purification of STEC OMVs, for the detection of OMV-associated virulence factors, and for the analysis of OMV interactions with host cells including OMV cellular uptake and intracellular trafficking of OMVs and OMV-delivered toxins.

• MeSH
• Bacterial Toxins metabolism   MeSH
• Endothelial Cells metabolism   microbiology   pathology   MeSH
• Escherichia coli O157 * metabolism   pathogenicity   MeSH
• Virulence Factors metabolism   MeSH
• Humans MeSH
• Cell-Derived Microparticles metabolism   MeSH
• Shiga Toxin 2 metabolism   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

BACKGROUND: Microparticles (MPs) are heterogeneous vesicles derived from membranes of different cells. Between 70 to 90% of MPs detected in blood originate from platelets. The release of MPs is associated with proinflammatory and procoagulant states. Elevated levels of MPs have been found in different diseases. We investigated MPs levels in patients with chronic hepatitis C (CHC) and changes in level during treatment using direct-acting antivirotics (DAA). PATIENTS AND METHODS: Thirty-six patients with CHC and forty healthy volunteers were included in the study. Concentrations of MPs were determined indirectly by measuring their procoagulant activity in plasma at baseline, end of therapy (EOT), and 12 weeks after EOT when the sustained virological response was assessed (SVR12). RESULTS: All patients achieved SVR12, which was associated with rapid improvement of markers of liver damage and function as well as liver stiffness (P=0.002). MPs levels were significantly higher in CHC patients than in healthy volunteers (P<0.001). No statistically significant decrease was found observed between baseline and SVR12 (P=0,330). Analysis of subpopulations with minimal fibrosis F0-1 (P=0.647), advanced fibrosis F2-4 (P=0.370), women(P=0.847), men (P=0.164) and genotype 1 (P=0.077) showed no significant changes during the follow-up period. CONCLUSIONS: MPs levels are higher in CHC patients and remain elevated shortly after achieving SVR. Higher concentrations of MPs in plasma are probably caused by a chronic uncontrolled exaggerated inflammatory response caused by CHC. Longer observation would probably confirm the significance of MPs levels decrease because normalization of liver function, inflammation, and structure after SVR requires more than 12 weeks.

• MeSH
• Antiviral Agents therapeutic use   MeSH
• Hepatitis C, Chronic * drug therapy   MeSH
• Fibrosis MeSH
• Liver Cirrhosis drug therapy   MeSH
• Humans MeSH
• Cell-Derived Microparticles * MeSH
• Sustained Virologic Response MeSH
• Inflammation MeSH
• Check Tag
• Humans MeSH
• Male MeSH
• Female MeSH
• Publication type
• Journal Article MeSH

BACKGROUND/AIM: This work aimed to prospectively evaluate the clinical significance of circulating microparticles (MPs) in relation to thrombotic risk factors and thrombotic complications in patients with BCR/ABL1-negative myeloproliferative neoplasms (MPN). PATIENTS AND METHODS: In a cohort of 206 patients with MPN, MPs' procoagulant activity was measured by the Zymuphen functional assay in 429 samples, while platelet- and erythrocyte-MPs were enumerated by flow cytometry in 558 samples. RESULTS: MPN patients had higher MP levels than the control group. The levels of MPs were higher in male patients, smokers, and those who were older than 60 years, and in the presence of JAK2V617F mutation, history of thrombosis, platelets >400×109/l, hematocrit >45%, or leukocytes >10×109/l. Cytoreductive treatment reduced MP levels, with anagrelide being associated with lower MP levels than hydroxyurea. CONCLUSION: The relationship with thrombotic risk factors indicates a possible role of MPs in the complex thrombotic mechanism, though cytoreductive treatment seems to affect this role through reducing MP levels.

• MeSH
• Humans MeSH
• Cell-Derived Microparticles * MeSH
• Myeloproliferative Disorders * drug therapy   genetics   MeSH
• Neoplasms * MeSH
• Blood Platelets MeSH
• Thrombosis * etiology   genetics   MeSH
• Check Tag
• Humans MeSH
• Male MeSH
• Publication type
• Journal Article MeSH

Roztroušená skleróza (RS) je chronické zánětlivé onemocnění centrálního nervového systému. Deregulace imunitního systému, migrace imunitních buněk přes hematoencefalickou bariéru a jejich podíl na vzniku demyelinizačních lézí hrají kritickou roli v patogenezi tohoto onemocnění. Přesná a spolehlivá diagnóza a sledování progrese onemocnění jsou komplikovány absencí spolehlivých a specifických laboratorních testů. Buněčné mikrovesikuly (MVs) přítomné v krvi, mozkomíšním moku a dalších tělních tekutinách představují donedávna neznámý mechanismus mezibuněčné komunikace podílející se na patologických procesech včetně RS. MVs nesou tkáňově-specifické informace a jsou tedy i nosiči specifické informace z tkání účastnících se vzniku a rozvoje onemocnění. MVs jsou proto vysoce slibným potenciálním diagnostickým nástrojem pro mnoho různých onemocnění. Podrobná molekulární charakterizace MVs z krve a mozkomíšního moku pacientů s RS a zhodnocení potenciálu MVs pro diagnostiku a monitorování RS je cílem tohoto projektu.; Multiple sclerosis is chronic inflammatory disease of central nervous system. Dysregulation of immune system, migration of immune cells across blood brain barrier and their participation in development demyelinating lesions play pivotal role in the disease pathogenesis. Diagnosis and disease progression monitoring is hindered by absence of reliable, sensitive, and specific laboratory tests reflecting accurately and specifically the disease activity. Cellular microvesicles (MVs) present in blood, CSF and other body fluids represent previously unrecognized means of cell communication which takes part in many pathological conditions, including MS. As such MVs carrying tissue-specific information are new and highly promising diagnostic targets for number of diseases and evaluation of their potential in MS is of utmost interest. Detailed molecular characterization of MVs present in CSF and blood of MS patients is the aim of the proposed project.

• MeSH
• Blood Chemical Analysis MeSH
• Biomarkers blood   cerebrospinal fluid   MeSH
• Exosomes MeSH
• Immunity MeSH
• Humans MeSH
• Cell-Derived Microparticles MeSH
• Cerebrospinal Fluid MeSH
• Multiple Sclerosis diagnosis   MeSH
• Check Tag
• Humans MeSH

• Conspectus
• Patologie. Klinická medicína
• NML Fields
• neurologie
• NML Publication type
• závěrečné zprávy o řešení grantu AZV MZ ČR