Catheter ablation (CA) has become an established treatment strategy for managing recurrent ventricular tachycardias (VTs) in patients with structural heart disease. In recent years, percutaneous mechanical circulatory support (PMCS) devices have been increasingly used intra-operatively to improve the ablation outcome. One indication would be rescue therapy for patients who develop haemodynamic deterioration during the ablation. However, more efforts are focused on identifying subjects who are at high risk of such deterioration and could benefit from the pre-emptive use of the PMCS. The third reason to use PMCS could be the inability to identify diffuse substrate, especially in non-ischaemic cardiomyopathy. This paper reviews available experiences using various types of PMCS in different clinical scenarios. Although PMCS allows mapping during VT, it does not significantly influence acute outcomes and not convincingly long-term outcomes. On the contrary, the complication rate appears to be higher in PMCS cohorts. Our data suggest that even in patients with severe left ventricular dysfunction, the substrate modification can be performed without the need for general anaesthesia and risk of haemodynamic decompensation. In end-stage heart failure associated with the electrical storm, implantation of a left ventricular assist device (or PMCS with a transition to the left ventricular assist device) might be the preferred strategy before CA. In high-risk patients who are not potential candidates for these treatment options, radiotherapy could be considered as a bail-out treatment of recurrent VTs. These approaches should be studied in prospective trials.

• MeSH
• Hemodynamics MeSH
• Catheter Ablation * methods   adverse effects   MeSH
• Tachycardia, Ventricular * surgery   physiopathology   MeSH
• Humans MeSH
• Heart-Assist Devices * MeSH
• Risk Factors MeSH
• Heart Failure physiopathology   therapy   MeSH
• Treatment Outcome MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Review MeSH

The 3'-5', 3'-5' cyclic dinucleotides (3'3'CDNs) are bacterial second messengers that can also bind to the stimulator of interferon genes (STING) adaptor protein in vertebrates and activate the host innate immunity. Here, we profiled the substrate specificity of four bacterial dinucleotide synthases from Vibrio cholerae (DncV), Bacillus thuringiensis (btDisA), Escherichia coli (dgcZ), and Thermotoga maritima (tDGC) using a library of 33 nucleoside-5'-triphosphate analogues and then employed these enzymes to synthesize 24 3'3'CDNs. The STING affinity of CDNs was evaluated in cell-based and biochemical assays, and their ability to induce cytokines was determined by employing human peripheral blood mononuclear cells. Interestingly, the prepared heterodimeric 3'3'CDNs bound to the STING much better than their homodimeric counterparts and showed similar or better potency than bacterial 3'3'CDNs. We also rationalized the experimental findings by in-depth STING-CDN structure-activity correlations by dissecting computed interaction free energies into a set of well-defined and intuitive terms. To this aim, we employed state-of-the-art methods of computational chemistry, such as quantum mechanics/molecular mechanics (QM/MM) calculations, and complemented the computed results with the {STING:3'3'c-di-ara-AMP} X-ray crystallographic structure. QM/MM identified three outliers (mostly homodimers) for which we have no clear explanation of their impaired binding with respect to their heterodimeric counterparts, whereas the R2 = 0.7 correlation between the computed ΔG'int_rel and experimental ΔTm's for the remaining ligands has been very encouraging.

• MeSH
• Bacillus thuringiensis enzymology   ultrastructure   MeSH
• Cytokines chemistry   genetics   MeSH
• Escherichia coli enzymology   ultrastructure   MeSH
• Crystallography, X-Ray MeSH
• Quantum Theory MeSH
• Leukocytes, Mononuclear chemistry   enzymology   MeSH
• Humans MeSH
• Membrane Proteins chemistry   genetics   ultrastructure   MeSH
• Nucleotides biosynthesis   chemistry   genetics   MeSH
• Immunity, Innate genetics   MeSH
• Substrate Specificity MeSH
• Thermotoga maritima enzymology   ultrastructure   MeSH
• Vibrio cholerae enzymology   ultrastructure   MeSH
• Structure-Activity Relationship * MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

The bacterium Pseudomonas putida KT2440 is gaining considerable interest as a microbial platform for biotechnological valorization of polymeric organic materials, such as lignocellulosic residues or plastics. However, P. putida on its own cannot make much use of such complex substrates, mainly because it lacks an efficient extracellular depolymerizing apparatus. We seek to address this limitation by adopting a recombinant cellulosome strategy for this host. In this work, we report an essential step in this endeavor-a display of designer enzyme-anchoring protein "scaffoldins", encompassing cohesin binding domains from divergent cellulolytic bacterial species on the P. putida surface. Two P. putida chassis strains, EM42 and EM371, with streamlined genomes and differences in the composition of the outer membrane were employed in this study. Scaffoldin variants were optimally delivered to their surface with one of four tested autotransporter systems (Ag43 from Escherichia coli), and the efficient display was confirmed by extracellular attachment of chimeric β-glucosidase and fluorescent proteins. Our results not only highlight the value of cell surface engineering for presentation of recombinant proteins on the envelope of Gram-negative bacteria but also pave the way toward designer cellulosome strategies tailored for P. putida.

• MeSH
• beta-Glucosidase metabolism   MeSH
• Cellulose metabolism   MeSH
• Cellulosomes metabolism   MeSH
• Chromosomal Proteins, Non-Histone chemistry   MeSH
• Escherichia coli metabolism   MeSH
• Genome, Bacterial * MeSH
• Membrane Proteins metabolism   MeSH
• Metabolic Engineering methods   MeSH
• Protein Domains MeSH
• Cell Cycle Proteins chemistry   MeSH
• Escherichia coli Proteins metabolism   MeSH
• Pseudomonas putida genetics   metabolism   MeSH
• Recombinant Proteins metabolism   MeSH
• Bacterial Outer Membrane metabolism   MeSH
• Green Fluorescent Proteins metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

The patients with mantle cell lymphoma (MCL) have translocation t(11;14) associated with cyclin D1 overexpression. We observed that iron (an essential cofactor of dioxygenases including prolyl hydroxylases [PHDs]) depletion by deferoxamine blocked MCL cells' proliferation, increased expression of DNA damage marker γH2AX, induced cell cycle arrest and decreased cyclin D1 level. Treatment of MCL cell lines with dimethyloxalylglycine, which blocks dioxygenases involving PHDs by competing with their substrate 2-oxoglutarate, leads to their decreased proliferation and the decrease of cyclin D1 level. We then postulated that loss of EGLN2/PHD1 in MCL cells may lead to down-regulation of cyclin D1 by blocking the degradation of FOXO3A, a cyclin D1 suppressor. However, the CRISPR/Cas9-based loss-of-function of EGLN2/PHD1 did not affect cyclin D1 expression and the loss of FOXO3A did not restore cyclin D1 levels after iron chelation. These data suggest that expression of cyclin D1 in MCL is not controlled by ENGL2/PHD1-FOXO3A pathway and that chelation- and 2-oxoglutarate competition-mediated down-regulation of cyclin D1 in MCL cells is driven by yet unknown mechanism involving iron- and 2-oxoglutarate-dependent dioxygenases other than PHD1. These data support further exploration of the use of iron chelation and 2-oxoglutarate-dependent dioxygenase inhibitors as a novel therapy of MCL.

• MeSH
• Amino Acids, Dicarboxylic pharmacology   MeSH
• Iron Chelating Agents pharmacology   MeSH
• Cyclin D1 metabolism   MeSH
• Deferoxamine pharmacology   MeSH
• Iron Deficiencies MeSH
• Dioxygenases antagonists & inhibitors   metabolism   MeSH
• Down-Regulation drug effects   MeSH
• Hydroxylation MeSH
• Cell Hypoxia drug effects   MeSH
• Enzyme Inhibitors pharmacology   MeSH
• Ketoglutaric Acids pharmacology   MeSH
• Humans MeSH
• Lymphoma, Mantle-Cell enzymology   MeSH
• RNA, Messenger genetics   metabolism   MeSH
• Cell Line, Tumor MeSH
• DNA Damage MeSH
• Hypoxia-Inducible Factor-Proline Dioxygenases metabolism   MeSH
• Forkhead Box Protein O3 genetics   metabolism   MeSH
• Iron MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Lyzozomální střádavá onemocnění (lysosomal storage diseases – LSD) představují heterogenní skupinu > 60 závažných dědičných poruch metabolismu způsobených poruchou funkce jednoho nebo více lyzozomálních enzymů nebo poruchami procesování a transportu proteinů přes lyzozomální membránu. Diagnostika LSD je založena na klinickém podezření a výsledcích enzymatických a mole-kulárních analýz.Z 836 pacientů s LSD, které jsme diagnostikovali v posledních 30 letech, byly nejčastější mukopolysacharidózy (n = 150), Gaucherova nemoc (n = 57) a Fabryho nemoc (74 hemizygotních mužů a 116 heterozygotních žen). Mezi počáteční příznaky LSD, které byly zpočátku diagnosticky nespecifické, patřily otoky a omezená hybnost velkých a/nebo malých kloubů, bolesti kolen a kyčlí, kloubní defigurace a kostní deformity, hepatomegalie a/nebo splenomegalie, opakované infekty horních cest dýchacích, obstrukční a/nebo restriktivní plicní porucha, pokles renálních funkcí, anemie a u některých typů LSD i poruchy růstu, kraniofaciální dysmorfie a porucha kognitivních funkcí. Pacienti procházeli řadou odborných ambulancí včetně revmatologie, ortopedie, ORL, neurologie, nefrologie či hematologie, což vedlo ve většině případů k pozdní diagnostice. Ačkoliv jednotlivé typy LSD se vyskytují poměrně vzácně, řada z nich je léčitelná nebo alespoň léčbou ovlivnitelná. V terapii se uplatňuje enzymová substituční terapie (enzyme replacement therapy – ERT), substrát redukující terapie (SRT) a transplantace hematopoetickými kmenovými buňkami, které mohou významně zlepšit prognózu onemocnění nebo alespoň stabilizovat jeho progresi. Závěr: Účinnost léčby u pacientů s lyzozomálním střádavým onemocněním významně závisí na včasné diagnostice, na které by se měli podílet i odborníci v oboru revmatologie.

Lysosomal storage diseases (LSD) represent a heterogeneous group of 60 severe hereditary metabolic disorders due to impaired function of one or more lysosomal enzymes or disorders of protein processing and transport across the lysosomal membrane. The diagnosis of LSD is based on clinical suspicion and the results of enzymatic and molecular analyses. Of the 836 LSD patients diagnosed in the last 30 years, mucopolysaccharidoses (n = 150), Gaucher disease (n = 57), and Fabry disease (74 hemizygous men and 116 heterozygous women) were the most common. Initial symptoms of LSD that were initially diagnostically non-specific included swelling and limited mobility of large and/or small joints, knee and hip pain, joint disfiguration and bone deformity, hepatomegaly and/or splenomegaly, recurrent upper respiratory tract infections, obstructive and/or restrictive pulmonary disorder, decreased renal function, anemia and, in some types of LSD, growth disorders, craniofacial dysmorphia and cognitive impairment. Patients went through a number of specialist outpatient clinics including rheumatology, orthopedics, ENT, neurology, nephrology or hematology, which in most cases led to late diagnosis. Although individual types of LSD are rare, many are treatable or at least therapeutically modifiable. Enzyme replacement therapy (ERT), substrate reducing therapy (SRT), and hematopoietic stem cell transplantation are useful in therapy, which can significantly improve disease prognosis or at least stabilize its progression. Conclusion: The efficacy of treatment in patients with lysosomal storage diseases is significantly dependent on early diagnosis, in which rheumatology specialists should participate as well.

• MeSH
• Fabry Disease diagnosis   pathology   therapy   MeSH
• Gaucher Disease diagnosis   pathology   therapy   MeSH
• Incidence MeSH
• Humans MeSH
• Lysosomal Storage Diseases * diagnosis   genetics   classification   MeSH
• Mucopolysaccharidoses diagnosis   pathology   therapy   MeSH
• Musculoskeletal Abnormalities * etiology   physiopathology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH

Organic dye-tagged lipid analogs are essential for many fluorescence-based investigations of complex membrane structures, especially when using advanced microscopy approaches. However, lipid analogs may interfere with membrane structure and dynamics, and it is not obvious that the properties of lipid analogs would match those of non-labeled host lipids. In this work, we bridged atomistic simulations with super-resolution imaging experiments and biomimetic membranes to assess the performance of commonly used sphingomyelin-based lipid analogs. The objective was to compare, on equal footing, the relative strengths and weaknesses of acyl chain labeling, headgroup labeling, and labeling based on poly-ethyl-glycol (PEG) linkers in determining biomembrane properties. We observed that the most appropriate strategy to minimize dye-induced membrane perturbations and to allow consideration of Brownian-like diffusion in liquid-ordered membrane environments is to decouple the dye from a membrane by a PEG linker attached to a lipid headgroup. Yet, while the use of PEG linkers may sound a rational and even an obvious approach to explore membrane dynamics, the results also suggest that the dyes exploiting PEG linkers interfere with molecular interactions and their dynamics. Overall, the results highlight the great care needed when using fluorescent lipid analogs, in particular accurate controls.

• MeSH
• Diffusion MeSH
• Fluorescent Dyes chemistry   metabolism   MeSH
• Phosphatidylcholines chemistry   MeSH
• Lipid Bilayers chemistry   metabolism   MeSH
• Polyethylene Glycols chemistry   MeSH
• Molecular Dynamics Simulation MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Surface bioactivity has been under intensive study with reference to its use in medical implants. Our study is focused on coatings prepared from an electroactive material which can support bone cell adhesion. Until now, hydroxyapatite films have usually been utilized as a chemically-active surface agent. However, electrically-active films could set a new direction in hard tissue replacement. As a base for these films, it is necessary to prepare an intermediate film, which can serve as a suitable barrier against the possible diffusion of some allergens and toxic elements from the substrate. The intermediate film also improves the adaptation of the mechanical properties of the basic material to an electroactive film. The aim of our work was to select an implantable and biocompatible material for this intermediate film that is suitable for coating several widely-used materials, to check the possibility of preparing an electroactive film for use on a material of this type, and to characterize the structure and several mechanical properties of this intermediate film. TiNb was selected as the material for the intermediate film, because of its excellent chemical and mechanical properties. TiNb coatings were deposited by magnetron sputtering on various substrates, namely Ti, Ti6Al4V, stainless steel, and bulk TiNb (as standard), and important properties of the layers, e.g. surface morphology and surface roughness, crystalline structure, etc., were characterized by several methods (SEM, EBSD, X-ray diffraction, nanoindentation and roughness measurement). It was found that the structure and the mechanical properties of the TiNb layer depended significantly on the type of substrate. TiNb was then used as a substrate for depositing a ferroelectrically active material, e.g., BaTiO3, and the adhesion, viability and proliferation of human osteoblast-like Saos-2 cells on this system were studied. We found that the electroactive BaTiO3 film was not only non-cytotoxic (i.e. it did not affect the cell viability). It also enhanced the growth of Saos-2 cells in comparison with pure TiNb and with standard tissue culture polystyrene wells, and also in comparison with BaTiO3 films deposited on Ti, i.e. a material clinically used for implantation into the bone.

• MeSH
• Adhesives MeSH
• X-Ray Diffraction MeSH
• Durapatite MeSH
• Humans MeSH
• Osteoblasts MeSH
• Surface Properties MeSH
• Prostheses and Implants MeSH
• Alloys chemistry   MeSH
• Materials Testing MeSH
• Titanium MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

Ribosomes synthesizing proteins containing consecutive proline residues become stalled and require rescue via the action of uniquely modified translation elongation factors, EF-P in bacteria, or archaeal/eukaryotic a/eIF5A. To date, no structures exist of EF-P or eIF5A in complex with translating ribosomes stalled at polyproline stretches, and thus structural insight into how EF-P/eIF5A rescue these arrested ribosomes has been lacking. Here we present cryo-EM structures of ribosomes stalled on proline stretches, without and with modified EF-P. The structures suggest that the favored conformation of the polyproline-containing nascent chain is incompatible with the peptide exit tunnel of the ribosome and leads to destabilization of the peptidyl-tRNA. Binding of EF-P stabilizes the P-site tRNA, particularly via interactions between its modification and the CCA end, thereby enforcing an alternative conformation of the polyproline-containing nascent chain, which allows a favorable substrate geometry for peptide bond formation.

• MeSH
• Cryoelectron Microscopy MeSH
• Peptide Elongation Factors chemistry   genetics   metabolism   ultrastructure   MeSH
• Escherichia coli genetics   metabolism   MeSH
• Peptide Initiation Factors chemistry   metabolism   MeSH
• Nucleic Acid Conformation MeSH
• Protein Conformation MeSH
• RNA, Messenger chemistry   genetics   metabolism   MeSH
• Mutation MeSH
• Peptides chemistry   metabolism   MeSH
• RNA-Binding Proteins chemistry   metabolism   MeSH
• Escherichia coli Proteins chemistry   genetics   metabolism   ultrastructure   MeSH
• Protein Biosynthesis MeSH
• Ribosomes chemistry   metabolism   ultrastructure   MeSH
• RNA, Transfer chemistry   genetics   metabolism   MeSH
• Molecular Dynamics Simulation MeSH
• Molecular Docking Simulation MeSH
• Protein Binding MeSH
• Binding Sites MeSH
• Structure-Activity Relationship MeSH
• Publication type
• Journal Article MeSH

• MeSH
• Cells * MeSH
• Molecular Biology MeSH

• Conspectus
• Biochemie. Molekulární biologie. Biofyzika
• NML Fields
• molekulární biologie, molekulární medicína
• NML Publication type
• učebnice vysokých škol

Lysosomální střádavá onemocnění (LSDs, Lysosomal Storage Diseases) tvoří skupinu více než 70 vzácných dědičných metabolických onemocnění, jejichž příčinou je nejčastěji porucha funkce některého z lysosomálních enzymů. Cílem přehledového sdělení je představení enzymové substituční terapie (ERT, Enzyme Replacement Therapy), která si našla v terapii LSDs zásadní místo. ERT je založena na náhradě defektního enzymu pomocí rekombinantního proteinu podávaného v 2–4hodinové intravenózní infuzi jednou za 1 či 2 týdny. Tato léčba je v současné době k dispozici pro sedm LSDs – Gaucherovu chorobu, Fabryho chorobu, Pompeho chorobu a mukopolysacharidózu typu I, II, IVA a VI. ERT významně ovlivňuje viscerální manifestaci onemocnění (hepatomegalie, splenomegalie, vitální kapacita plic, svalová slabost, anémie, gastrointestinální obtíže aj.), není ovšem účinná v terapii postižení centrálního nervového systému (CNS) a pouze malý efekt je pozorován u postižení kosti, chrupavky a srdečních chlopní. Časně zahájená ERT je schopna zamezit manifestaci téměř všech symptomů onemocnění, vyjma postižení CNS. U části pacientů se na podávaný enzym může rozvinout imunitní reakce s klinickými projevy alergie a sníženého efektu terapie. Stručně jsou zmíněny i další modality léčby LSDs: transplantace hematopoetických kmenových buněk, substrát redukující terapie, použití chaperonů a genová terapie. Závěr: ERT zpomaluje progresi onemocnění a výrazně mění jeho přirozený průběh, zlepšuje kvalitu života a prodlužuje přežití pacientů. Omezení jsou daná špatným průnikem do některých tkání, možnými alergickými reakcemi a vysokou finanční náročností. Zásadním předpokladem pro úspěch ERT je její časné zahájení při rychlé diagnostice onemocnění.

Lysosomal storage diseases (LSDs) form a group of more than 70 rare inherited metabolic diseases, usually caused by a malfunction of some of the lysosomal enzymes. The aim of this review is to introduce enzyme replacement therapy (ERT) which gained an indispensable role in the therapy of LSDs. ERT is based upon replacement of the defective enzyme by a recombinant protein administred in a 2–4 hour infusion once a week or once a fortnight. This therapy is currently available for seven LSDs: Gaucher disease, Fabry disease, Pompe disease and mucopolysaccharidoses I, II, IVA and VI. ERT significantly affects visceral manifestations of the diseases (hepatomegaly, splenomegaly, vital lung capacity, muscle weakness, anaemia, gastrointestinal disorders etc.), yet it is not effective in treating the central nervous system (CNS) involvement and only a small effect is observed in treating disease manifestations in bones, cartilage and heart valves. An immune reaction with clinical presentation of an allergic reaction and therapy effectiveness decrease can occur with a portion of patients. Other modalities for LSDs therapy are concisely mentioned: haematopoietic stem cell transplantation, substrate reduction therapy, chaperons and gene therapy. Conclusion: ERT slows the progression of the disease and markedly alters its natural course, improves quality of life and prolongs patients’ lifespan. The limitations include insufficient effect in some tissues, possible allergic reactions and great financial demands. An essential presumption for effectiveness and success of ERT is its early initiation following quick diagnosis of the disease.

• Keywords
• lysosomální střádavá onemocnění,
• MeSH
• Enzyme Therapy MeSH
• Enzyme Replacement Therapy * methods   adverse effects   MeSH
• Enzymes administration & dosage   adverse effects   MeSH
• Fabry Disease drug therapy   MeSH
• Gaucher Disease drug therapy   MeSH
• Glycogen Storage Disease drug therapy   MeSH
• Infusions, Intravenous MeSH
• Drug Hypersensitivity complications   MeSH
• Humans MeSH
• Lysosomal Storage Diseases * drug therapy   physiopathology   MeSH
• Mucopolysaccharidoses drug therapy   MeSH
• Recombinant Proteins administration & dosage   adverse effects   therapeutic use   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH