Objev inzulinu v roce 1921 znamenal revoluci v léčbě diabetes mellitus. Od počátečních intravenózních aplikací inzulinu se léčba přesunula k subkutánním injekcím a následně k využití inzulinových per. Významný pokrok přinesly inzulinové pumpy, které umožňují kontinuální subkutánní podávání inzulinu a tím napodobení fyziologické sekrece. Propojení inzulinových pump s kontinuální monitorací glukózy vedlo k vývoji hybridních uzavřených smyček. Tyto systémy využívají algoritmy pro automatickou úpravu dávek inzulinu na základě aktuálních a predikovaných hodnot glykemie. Výsledkem je zlepšení glykemické kontroly, snížení variability glykemie a snížení rizika jak hypoglykemických, tak hyperglykemických epizod.
The discovery of insulin in 1921 revolutionized the treatment of diabetes mellitus. From initial intravenous applications, treatment progressed to subcutaneous injections and subsequently to the use of insulin pens. Insulin pumps represented a significant advancement, enabling continuous subcutaneous insulin infusion and thus mimicking physiological secretion. The combination of insulin pumps with continuous glucose monitoring has led to the development of hybrid closed-loop systems. These systems utilize algorithms to automatically adjust insulin doses based on current and predicted glucose levels. The result is improved glycemic control, reduced glycemic variability, and decreased risk of both hypoglycemic and hyperglycemic episodes.
- MeSH
- Diabetes Mellitus, Type 1 drug therapy blood prevention & control MeSH
- Diabetes Mellitus drug therapy prevention & control MeSH
- Hyperglycemia blood prevention & control MeSH
- Hypoglycemia blood prevention & control MeSH
- Insulin administration & dosage pharmacology therapeutic use MeSH
- Insulin Infusion Systems * MeSH
- Continuous Glucose Monitoring methods instrumentation MeSH
- Humans MeSH
- Check Tag
- Humans MeSH
- Publication type
- Review MeSH
Bordetella pertussis is the causative agent of whooping cough in humans, a disease that has recently experienced a resurgence. In contrast, Bordetella bronchiseptica infects the respiratory tract of various mammalian species, causing a range of symptoms from asymptomatic chronic carriage to acute illness. Both pathogens utilize type III secretion system (T3SS) to deliver the effector protein BteA into host cells. Once injected, BteA triggers a cascade of events leading to caspase 1-independent necrosis through a mechanism that remains incompletely understood. We demonstrate that BteA-induced cell death is characterized by the fragmentation of the cellular endoplasmic reticulum and mitochondria, the formation of necrotic balloon-like protrusions, and plasma membrane permeabilization. Importantly, genome-wide CRISPR-Cas9 screen targeting 19,050 genes failed to identify any host factors required for BteA cytotoxicity, suggesting that BteA does not require a single nonessential host factor for its cytotoxicity. We further reveal that BteA triggers a rapid and sustained influx of calcium ions, which is associated with organelle fragmentation and plasma membrane permeabilization. The sustained elevation of cytosolic Ca2+ levels results in mitochondrial calcium overload, mitochondrial swelling, cristolysis, and loss of mitochondrial membrane potential. Inhibition of calcium channels with 2-APB delays both the Ca2+ influx and BteA-induced cell death. Our findings indicate that BteA exploits essential host processes and/or redundant pathways to disrupt calcium homeostasis and mitochondrial function, ultimately leading to host cell death.IMPORTANCEThe respiratory pathogens Bordetella pertussis and Bordetella bronchiseptica exhibit cytotoxicity toward a variety of mammalian cells, which depends on the type III secretion effector BteA. Moreover, the increased virulence of B. bronchiseptica is associated with enhanced expression of T3SS and BteA. However, the molecular mechanism underlying BteA cytotoxicity is elusive. In this study, we performed a CRISPR-Cas9 screen, revealing that BteA-induced cell death depends on essential or redundant host processes. Additionally, we demonstrate that BteA disrupts calcium homeostasis, which leads to mitochondrial dysfunction and cell death. These findings contribute to closing the gap in our understanding of the signaling cascades targeted by BteA.
- MeSH
- Bacterial Proteins * metabolism genetics MeSH
- Bordetella bronchiseptica genetics metabolism drug effects MeSH
- Bordetella pertussis genetics pathogenicity metabolism drug effects MeSH
- Cell Death * drug effects MeSH
- Endoplasmic Reticulum metabolism drug effects MeSH
- Homeostasis * MeSH
- Host-Pathogen Interactions MeSH
- Humans MeSH
- Mitochondria metabolism drug effects MeSH
- Type III Secretion Systems metabolism genetics MeSH
- Calcium * metabolism MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
Plasmacytoid dendritic cells (pDCs) are a minority subset of dendritic cells that despite their tiny quantity play an important role in the immune system, especially in antiviral immunity. They are known mostly as the major producers of type I IFN, which they secrete upon stimulation of endosomal Toll-like receptors 7 and 9 with viral RNA and DNA. However, the functionality of pDCs is more complex, as they were shown to be also involved in autoimmunity, inflammation, and cancer. In the context of the tumor microenvironment, pDCs mostly show substantial functional defects and thus contribute to establishing immunosuppressive micromilieu. Indeed, tumor-infiltrating pDCs were shown to be predominantly pro-tumorigenic, with reduced ability to produce IFNα and capacity to prime regulatory T cells via the ICOS/ICOS-L pathway. Here we describe in detail a method to assess the functional capacity of pDCs upon exposure to tumor-derived cell culture supernatants. The same technique can be implemented with minimal variations to test any soluble factor's impact on pDC phenotype and function.
- MeSH
- Dendritic Cells * immunology metabolism MeSH
- Humans MeSH
- Tumor Microenvironment immunology MeSH
- Neoplasms * immunology pathology metabolism MeSH
- Flow Cytometry methods MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
Fonticins are phage tail-like bacteriocins produced by the Gram-negative bacterium Pragia fontium from the family Budviciaceae. This bacterium produces contractile-type particles that adsorb on the surface of sensitive bacteria and penetrate the cell wall, probably during contraction, in a way similar to the type VI secretion system. We characterized the pore-forming activity of fonticins using both living cells and in vitro model membranes. Using a potassium leakage assay, we show that fonticins are able to permeabilize sensitive cells. On black lipid membranes, single-pore conductance is about 0.78 nS in 1 M NaCl and appears to be linearly dependent on the increasing molar strength of NaCl solution, which is a property of considerably large pores. In agreement with these findings, fonticins are not ion selective for Na+, K+, and Cl-. Polyethylene glycol 3350 (PEG 3350) molecules of about 3.5 nm in diameter can enter the fonticin pore lumen, whereas the larger molecules cannot pass the pore. The size of fonticin pores was confirmed by transmission electron microscopy. The terminal membrane-piercing complex of the fonticin tube probably creates a selective barrier restricting passage of macromolecules. IMPORTANCE Phage tail-like bacteriocins are now the subject of research as potent antibacterial agents due to their narrow host specificity and single-hit mode of action. In this work, we focused on the structure and mode of action of fonticins. According to some theories, related particles were initially adapted for passage of double-stranded DNA (dsDNA) molecules, but fonticins changed their function during the evolution; they are able to form large pores through the bacterial envelope of Gram-negative bacteria. As various pore-forming proteins are extensively used for nanopore sequencing and stochastic sensing, we decided to investigate the pore-forming properties of fonticin protein complexes on artificial lipid membranes. Our research revealed remarkable structural properties of these particles that may have a potential application as a nanodevice.
Bordetella pertussis is a Gram-negative, strictly human re-emerging respiratory pathogen and the causative agent of whooping cough. Similar to other Gram-negative pathogens, B. pertussis produces the type III secretion system, but its role in the pathogenesis of B. pertussis is enigmatic and yet to be elucidated. Here, we combined RNA-seq, LC-MS/MS, and co-immunoprecipitation techniques to identify and characterize the novel CesT family T3SS chaperone BP2265. We show that this chaperone specifically interacts with the secreted T3SS regulator BtrA and represents the first non-flagellar chaperone required for the secretion of an anti-sigma factor. In its absence, secretion but not production of BtrA and most T3SS substrates is severely impaired. It appears that the role of BtrA in regulating T3SS extends beyond its activity as an antagonist of the sigma factor BtrS. Predictions made by artificial intelligence system AlphaFold support the chaperone function of BP2265 towards BtrA and outline the structural basis for the interaction of BtrA with its target BtrS. We propose to rename BP2265 to BtcB for the Bordetella type III chaperone of BtrA.In addition, the absence of the BtcB chaperone results in increased expression of numerous flagellar genes and several virulence genes. While increased production of flagellar proteins and intimin BipA translated into increased biofilm formation by the mutant, enhanced production of virulence factors resulted in increased cytotoxicity towards human macrophages. We hypothesize that these phenotypic traits result indirectly from impaired secretion of BtrA and altered activity of the BtrA/BtrS regulatory node.
- MeSH
- Bacterial Proteins genetics metabolism MeSH
- Bordetella pertussis * metabolism MeSH
- Chromatography, Liquid MeSH
- Humans MeSH
- Whooping Cough * MeSH
- Gene Expression Regulation, Bacterial MeSH
- Sigma Factor genetics MeSH
- Tandem Mass Spectrometry MeSH
- Artificial Intelligence MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
Cystická fibróza (CF) je autozomálně recesivně dědičné onemocnění. Příčinou jsou mutace CFTR genu, které vedou k dysfunkci chloridového kanálu. Tím dochází ke změně složení a vlastnosti sekretů, v klinickém obraze dominuje hlavně postižení plic a pankreatu. V posledních letech se díky novým terapiím výrazně prodlužuje střední doba života pacientů s CF, ale zároveň stoupá i výskyt komplikací. Jednou z nich je diabetes mellitus vázaný na cystickou fibrózu (cystic fibrosis-related diabetes, CFRD). V současné době má po 30. roce věku problém s porušenou glukózovou tolerancí či přímo s CFRD až 50 % pacientů. Proto je indikován pravidelný screening pomocí oGTT, roli ve včasné diagnostice budou hrát i nové technologie, jako je kontinuální monitorování glukózy pomocí senzorů. Dle současných doporučení je lékem volby při CFRD jednoznačně inzulin, v pokročilých stadiích využíváme léčbu inzulinovou pumpou spolu se senzorem (hybridní uzavřená smyčka). Dieta u CFRD je odlišná od ostatních typů diabetu. Neomezujeme energetický příjem ani příjem tuků. Konzumace sacharidů je limitována pouze částečně, omezení platí zejména pro sladké nápoje.
Cystic fibrosis (CF) is an autosomal recessive disorder. It is caused by mutations in CFTR gene that lead to chloride channel dysfunction and consequent changes in composition and properties of the secrets. The clinical presentation is dominated by the involvement of the lungs and pancreas. Thanks to novel therapies, the life expectancy of CF patients has improved significantly within the past years; anyway, the complication rate has also increased. One of them is cystic fibrosis-related diabetes (CFRD). After the age of 30 years old, up to 50 % of patients are currently developing impaired glucose tolerance or overt CFRD. Therefore regular screening is provided using oGTT, and new technologies such as continuous glucose monitoring (sensor) will also be employed in the near future. According to current guidelines, early insulin treatment is recommended. In advanced stages, insulin pump treatment linked with a sensor (hybrid closed loop system) is helpful. The CFRD diet differs from diets in other types of diabetes. Neither energy nor fat intake is reduced. Carbohydrate consumption is only partially restricted, especially the pop drinks.
The classical Bordetella species infect the respiratory tract of mammals. While B. bronchiseptica causes rather chronic respiratory infections in a variety of mammals, the human-adapted species B. pertussis and B. parapertussisHU cause an acute respiratory disease known as whooping cough or pertussis. The virulence factors include a type III secretion system (T3SS) that translocates effectors BteA and BopN into host cells. However, the regulatory mechanisms underlying the secretion and translocation activity of T3SS in bordetellae are largely unknown. We have solved the crystal structure of BopN of B. pertussis and show that it is similar to the structures of gatekeepers that control access to the T3SS channel from the bacterial cytoplasm. We further found that BopN accumulates at the cell periphery at physiological concentrations of calcium ions (2 mM) that inhibit the secretion of BteA and BopN. Deletion of the bopN gene in B. bronchiseptica increased secretion of the BteA effector into calcium-rich medium but had no effect on secretion of the T3SS translocon components BopD and BopB. Moreover, the ΔbopN mutant secreted approximately 10-fold higher amounts of BteA into the medium of infected cells than the wild-type bacteria, but it translocated lower amounts of BteA into the host cell cytoplasm. These data demonstrate that BopN is a Bordetella T3SS gatekeeper required for regulated and targeted translocation of the BteA effector through the T3SS injectisome into host cells. IMPORTANCE The T3SS is utilized by many Gram-negative bacteria to deliver effector proteins from bacterial cytosol directly into infected host cell cytoplasm in a regulated and targeted manner. Pathogenic bordetellae use the T3SS to inject the BteA and BopN proteins into infected cells and upregulate the production of the anti-inflammatory cytokine interleukin-10 (IL-10) to evade host immunity. Previous studies proposed that BopN acted as an effector in host cells. In this study, we report that BopN is a T3SS gatekeeper that regulates the secretion and translocation activity of Bordetella T3SS.
- MeSH
- Bacterial Proteins metabolism MeSH
- Bordetella pertussis metabolism MeSH
- Virulence Factors metabolism MeSH
- Humans MeSH
- Whooping Cough * MeSH
- Mammals MeSH
- Type III Secretion Systems * metabolism MeSH
- Calcium MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
Acid-β-glucosidase (GCase, EC3.2.1.45), the lysosomal enzyme which hydrolyzes the simple glycosphingolipid, glucosylceramide (GlcCer), is encoded by the GBA1 gene. Biallelic mutations in GBA1 cause the human inherited metabolic disorder, Gaucher disease (GD), in which GlcCer accumulates, while heterozygous GBA1 mutations are the highest genetic risk factor for Parkinson's disease (PD). Recombinant GCase (e.g., Cerezyme® ) is produced for use in enzyme replacement therapy for GD and is largely successful in relieving disease symptoms, except for the neurological symptoms observed in a subset of patients. As a first step toward developing an alternative to the recombinant human enzymes used to treat GD, we applied the PROSS stability-design algorithm to generate GCase variants with enhanced stability. One of the designs, containing 55 mutations compared to wild-type human GCase, exhibits improved secretion and thermal stability. Furthermore, the design has higher enzymatic activity than the clinically used human enzyme when incorporated into an AAV vector, resulting in a larger decrease in the accumulation of lipid substrates in cultured cells. Based on stability-design calculations, we also developed a machine learning-based approach to distinguish benign from deleterious (i.e., disease-causing) GBA1 mutations. This approach gave remarkably accurate predictions of the enzymatic activity of single-nucleotide polymorphisms in the GBA1 gene that are not currently associated with GD or PD. This latter approach could be applied to other diseases to determine risk factors in patients carrying rare mutations.
- MeSH
- Cellulases * genetics MeSH
- Gaucher Disease * drug therapy genetics MeSH
- Heterozygote MeSH
- Humans MeSH
- Mutation MeSH
- Parkinson Disease * genetics MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
V decembri 2019 sa rozšíril do sveta z Číny nový typ koronavírusu, SARS-CoV-2, spôsobujúci ťažký akútny respiračný syndróm. Koronavírusy patria medzi obalené ssRNA vírusy a sú klasifikované do 4 rodov: Alphacoronavirus, Betacoronavirus, Gammacoronavirus a Deltacoronavirus. Predpokladá sa, že SARS-CoV-2 sa šíri primárne prostredníctvom osobného kontaktu cez väčšie respiračné kvapôčky. Tieto kvapôčky s vírusmi môžu byť priamo inhalované inými ľuďmi, resp. môžu pristáť na neživých povrchoch s možnosťou ďalšieho šírenia. Zistilo sa, že povrch oka je jednou zo vstupných brán pre infekciu. Ľudské oko má svoj vlastný renín-angiotenzínový systém s prítomnosťou ACE2 receptorov, na ktoré sa vírus viaže pomocou spike proteínu. Najbežnejšími prejavmi infekcie SARS-CoV-2 sú horúčka, kašeľ a dýchavičnosť. Viacero klinických diagnóz, ako konjunktivitída, uveitída, retinitída a neuritída boli a sú taktiež spájané s touto infekciou. Najčastejším oftalmologickým symptómom súvisiacim s ochorením COVID-19 je konjunktivitída. Niektoré štúdie naznačujú, že očné príznaky sa bežne vyskytujú u pacientov s ťažkou pneumóniou COVID-19 a že je možné detegovať vírusovú RNA zo spojivkového vaku týchto pacientov. V oftalmologickej praxi riešime nielen otázku liečby zápalov očných štruktúr v spojení s touto infekciou, ale aj celkový manažment návštev a kontrol pacientov rizikových a pozitívnych na koronavírusové ochorenie. Oftalmológovia by mohli potencionálne mať vyššie riziko vzniku infekcie SARS-CoV-2 v dôsledku osobnej komunikácie s pacientmi, častého vystavenia slzám a očnému sekrétu a používania prístrojov. Preto chceme poskytnúť pohľad na problém z pohľadu oftalmológa.
In December 2019, a novel coronavirus (CoV) epidemic, caused by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) emerged from China. Coronaviruses belong to enveloped ssRNA viruses and are classified into four genera: Alpha coronavirus, Beta coronavirus, Gamma coronavirus and Delta coronavirus. It is assumed that SARS-CoV-2 is spread primarily during a personal contact via bigger respiratory droplets. These droplets with viruses can be directly inhaled by other people or can lend on the surfaces with the possibility of further spreading. The ocular surface has been suggested as one of possible infection entries. Human eye has its own renin-angiotensin system with present ACE2 receptors, which bind the virus through spike protein. The most common symptoms of the SARS-CoV-2 infection are fever, cough and dyspnoea. Several clinical entities, such as conjunctivitis, anterior uveitis, retinitis, and optic neuritis have been associated with this infection. The most common ophthalmologic symptom associated with COVID-19 disease is conjunctivitis. Some studies indicate that eye symptoms are commonly present in patients with severe COVID-19 pneumonia and that it is possible to detect viral RNA from the conjunctival sac of these patients. In ophthalmologic praxis, we manage not only the therapy of the eye structures` inflammation in relation with this infection, but also the overall management of the visits and the supervision of the patients who are at risk and positive for coronavirus. Ophthalmologists could potentially have a higher risk of SARS-CoV-2 infection due to personal communication with the patients, frequent exposure to tears and eye secrets and the use of devices. We would like to provide an ophthalmologist`s perspective on this topic.
- MeSH
- Chloroquine adverse effects therapeutic use MeSH
- COVID-19 * etiology complications transmission MeSH
- Dexamethasone pharmacology therapeutic use MeSH
- Humans MeSH
- Eye Diseases * etiology pathology therapy virology MeSH
- Disease Transmission, Infectious prevention & control MeSH
- Conjunctivitis, Viral transmission MeSH
- Vitamin D pharmacology therapeutic use MeSH
- Health Personnel MeSH
- Check Tag
- Humans MeSH
- Publication type
- Review MeSH
Hypovitaminóza D vitaminu - deficit a insuficience, je globální zdravotní problém, který se dotýká více než miliardy dětí a dospělých. Optimální hladina vitaminu D hraje esenciální roli v regulaci kalciového a fosfátového metabolismu a je nezbytná nejen pro správný kostní metabolismus, ale má i jiné funkce v organismu, jako je vliv na funkci svalů, stimulaci diferenciace buněk, sekreci inzulinu a stimulaci imunitního systému. Hypovitaminóza D vitaminu pak bývá spojena nejen s poruchou kostního metabolismu, ale i s výskytem kardiovaskulárních onemocnění, arteriální hypertenze, dyslipidemie, diabetu mellitu II. typu, nádorových onemocnění, depresí, demencí, psychiatrických onemocnění a dalších. Hypovitaminóza D je také jedním z možných faktorů komplikovaného průběhu nemoci covid-19. Článek shrnuje poslední poznatky o jeho suplementaci.
Vitamin D hypovitaminosis - deficiency and insufficiency - is a global health problem that affects more than a billion children and adults. The optimal level of vitamin D plays an essential role in the regulation of calcium and phosphate metabolism and is necessary not only for proper bone metabolism, but also has other functions in the body, such as influencing muscle function, stimulating cell differentiation, insulin secretion and stimulating the immune system. Hypovitaminosis of vitamin D is then associated not only with a disorder of bone metabolism, but also with the occurrence of cardiovascular diseases, arterial hypertension, dyslipidemia, diabetes mellitus type II, cancer, depression, dementia, psychiatric diseases and others. Hypovitaminosis D is also one of the factors in the complicated course of the disease covid 19. The article summarizes the latest findings on its supplementation.
