Fully acetylated deoxyfluorinated hexosamine analogues and non-fluorinated 3,4,6-tri-O-acylated N-acetyl-hexosamine hemiacetals have previously been shown to display moderate anti-proliferative activity. We prepared a set of deoxyfluorinated GlcNAc and GalNAc hemiacetals that comprised both features: O-acylation at the non-anomeric positions with an acetyl, propionyl and butanoyl group, and deoxyfluorination at selected positions. Determination of the in vitro cytotoxicity towards the MDA-MB-231 breast cancer and HEK-293 cell lines showed that deoxyfluorination enhanced cytotoxicity in most analogues. Increasing the ester alkyl chain length had a variable effect on the cytotoxicity of fluoro analogues, which contrasted with non-fluorinated hemiacetals where butanoyl derivatives had always higher cytotoxicity than acetates. Reaction with 2-phenylethanethiol indicated that the recently described S-glyco-modification is an unlikely cause of cytotoxicity.

• MeSH
• Galactosamine * MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

BACKGROUND: In recent years, enzymes modifying N-acetylhexosamine substrates have emerged in numerous theoretical studies as well as practical applications from biology, biomedicine, and biotechnology. Advanced enzyme engineering techniques converted them into potent synthetic instruments affording a variety of valuable glycosides. SCOPE OF REVIEW: This review presents the diversity of engineered enzymes active with N-acetylhexosamine carbohydrates: from popular glycoside hydrolases and glycosyltransferases to less known oxidases, epimerases, kinases, sulfotransferases, and acetylases. Though hydrolases in natura, engineered chitinases, β-N-acetylhexosaminidases, and endo-β-N-acetylglucosaminidases were successfully employed in the synthesis of defined natural and derivatized chitooligomers and in the remodeling of N-glycosylation patterns of therapeutic antibodies. The genes of various N-acetylhexosaminyltransferases were cloned into metabolically engineered microorganisms for producing human milk oligosaccharides, Lewis X structures, and human-like glycoproteins. Moreover, mutant N-acetylhexosamine-active glycosyltransferases were applied, e.g., in the construction of glycomimetics and complex glycostructures, industrial production of low-lactose milk, and metabolic labeling of glycans. In the synthesis of biotechnologically important compounds, several innovative glycoengineered systems are presented for an efficient bioproduction of GlcNAc, UDP-GlcNAc, N-acetylneuraminic acid, and of defined glycosaminoglycans. MAJOR CONCLUSIONS: The above examples demonstrate that engineering of N-acetylhexosamine-active enzymes was able to solve complex issues such as synthesis of tailored human-like glycoproteins or industrial-scale production of desired oligosaccharides. Due to the specific catalytic mechanism, mutagenesis of these catalysts was often realized through rational solutions. GENERAL SIGNIFICANCE: Specific N-acetylhexosamine glycosylation is crucial in biological, biomedical and biotechnological applications and a good understanding of its details opens new possibilities in this fast developing area of glycoscience.

• MeSH
• Glycoproteins biosynthesis   MeSH
• Glycoside Hydrolases metabolism   MeSH
• Glycosylation MeSH
• Glycosyltransferases metabolism   MeSH
• Hexosamines metabolism   MeSH
• Catalysis MeSH
• Oligosaccharides biosynthesis   MeSH
• Protein Engineering * MeSH
• Sulfotransferases metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH

Diabetická retinopatie (DR) je typickou mikrovaskulární komplikací diabetu 1. i 2. typu. Ve vyspělých zemích je nejčastější příčinou slepoty u osob v produktivním věku. Základní metabolickou odchylkou, která hraje nejdůležitější roli při jejím vzniku, je hyperglykemie. Hyperglykemie zvyšuje mitochondriální produkci reaktivních kyslíkových radikálů a oxidační stres, vede k hromadění pokročilých produktů glykace, tvorbě hexosaminu, akcentaci polyolové cesty a zvýšení osmoticky aktivního sorbitolu a také ke zvýšení proteinkinázy C. Závažnost vaskulární patologie je významně ovlivněna genetickou dispozicí jedince a je modifikována dalšími epigenetickými (včetně microRNA), metabolickými a hemodynamickými faktory, jako je dyslipidemie, hypertenze, inzulinová rezistence a další. Uvedené abnormity ovlivní nejprve funkci a později i morfologii celé řady cílových buněk, které zahrnují nejen vlastní buňky retiny (endotelové buňky, pericyty, neurony, mikroglie, makroglie, pigmentový epitel), ale také imunokompetentní buňky, které infiltrují do místa patologie z cévního řečiště (non-rezidentní buňky). Aktivované buňky pak vedou k produkci řady mediátorů s vazoaktivními a růstovými vlastnostmi, které dál zhoršují oxidační stres a subklinický zánět. Zdrojem těchto mediátorů mohou být i další tkáně, např. tuková tkáň, játra, CNS, střevo, kosterní sval a další. Výsledkem je komplex, zpočátku funkčních a později strukturálních změn, které se projeví v dysregulaci (a) krevního průtoku, (b) buněčného růstu – apoptóza, proliferace, hypertrofie, a vedou k (c) proliferaci vaziva, zmnožení extracelulární hmoty ztluštění bazálních membrán. Důsledkem je pak rozvoj morfologicky fixované orgánové patologie. Léčba a prevence DR se v současné době opírá (a) o režimovou a farmakologickou léčbu ovlivnitelných rizikových faktorů, to znamená snahu o normalizaci hladin glykemie, krevního tlaku a hladin lipidů, (b) aktivní screening DR a (c) specializovanou oftalmologickou léčbu, která zahrnuje laserovou fotokoagulaci, intravitreální aplikaci farmak a vitreoretinální chirurgické postupy. Poznání patofyziologických mechanizmů rozvoje DR dává naději na možnou kauzální léčbu a prevenci.

Diabetic retinopathy (DR) develops in patients with both type 1 and type 2 diabetes and is the major cause of vision loss and blindness in the working population. The main risk factor of DR is hyperglycemia accompanied by enhanced mitochondrial production of reactive oxygen species and oxidative stress, formation of advanced glycation end products (AGE) and hexosamines, increase in polyol metabolism of glucose. The severity of vascular injury depends on the individual genetic background and is modified by other epigenetic, metabolic and haemodynamic factors, including hypertension, dyslipidemia and oxidative stress. In diabetes, damage to the retina occurs in the vasculature (endothelial cells and pericytes), neurons and glia, pigment epithelial cells and infiltrating immunocompetent cells: monocytes, granulocytes, lymfocytes. These activated cells change the production pattern of a number of mediators such as growth factors, proinflammatory cytokines, vasoactive molecules, coagulation factors and adhesion molecules resulting in increased blood flow, increased capillary permeability, proliferation of extracellular matrix and thickening of basal membranes, altered cell turnover (apoptosis, proliferation, hypertrophy), procoagulant and proaggregant pattern, and finally in angiogenesis and tissue remodelling. Brain, liver, adipose tissue, GUT, skeletal muscle and other tissues could be another source of mediators. Therapeutic approaches used for patients with or at risk for diabetic retinopathy include drug therapy to reduce modifiable risk factors, laser photocoagulation, intravitreous administration of anti-VEGF agents/steroids and intraocular surgery. Screening plays an important role in early detection and intervention to prevent the progression of diabetic retinopathy. Described insights into pathophysiological mechanisms responsible for DR, could help in the development of more targeted approach for prevention and treatment of diabetic retinopathy.

• Keywords
• aflibercept,
• MeSH
• Bevacizumab therapeutic use   MeSH
• Endothelium, Vascular MeSH
• Diabetic Retinopathy * etiology   drug therapy   physiopathology   MeSH
• Dyslipidemias complications   MeSH
• Hyperglycemia * complications   MeSH
• Hypertension complications   MeSH
• Intravitreal Injections MeSH
• Humans MeSH
• MicroRNAs MeSH
• Retinal Neurons MeSH
• Oxidative Stress MeSH
• Pericytes MeSH
• Ranibizumab therapeutic use   MeSH
• Receptors, Vascular Endothelial Growth Factor therapeutic use   MeSH
• Recombinant Fusion Proteins therapeutic use   MeSH
• Retinal Vessels physiopathology   MeSH
• Vascular Endothelial Growth Factors MeSH
• Check Tag
• Humans MeSH
• Publication type
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH

Controlled feeding of glucose has been employed previously to enhance the productivity of recombinant glycoproteins but there is a concern that low concentrations of glucose could limit the synthesis of precursors of glycosylation. Here we investigate the effect of glucose depletion on the metabolism, productivity and glycosylation of a chimeric human-llama monoclonal antibody secreted by CHO cells. The cells were inoculated into media containing varying concentrations of glucose. Glucose depletion occurred in cultures with an initial glucose ≤5.5 mM and seeded at low density (2.5 × 10(5) cells/mL) or at high cell inoculum (≥2.5 × 10(6) cells/mL) at higher glucose concentration (up to 25 mM). Glucose-depleted cultures produced non-glycosylated Mabs (up to 51%), lower galactosylation index (GI <0.43) and decreased sialylation (by 85%) as measured by mass spectrometry and HPLC. At low glucose a reduced intracellular pool of nucleotides (0.03-0.23 fmoles/cell) was measured as well as a low adenylate energy charge (<0.57). Low glucose also reduced GDP-sugars (by 77%) and UDP-hexosamines (by 90%). The data indicate that under glucose deprivation, low levels of intracellular nucleotides and nucleotide sugars reduced the availability of the immediate precursors of glycosylation. These results are important when applied to the design of fed-batch cultures.

• MeSH
• CHO Cells MeSH
• Cricetulus MeSH
• Glucose metabolism   MeSH
• Glycoproteins biosynthesis   metabolism   MeSH
• Glycosylation MeSH
• Cricetinae MeSH
• Humans MeSH
• Antibodies, Monoclonal biosynthesis   metabolism   MeSH
• Recombinant Proteins biosynthesis   genetics   MeSH
• Animals MeSH
• Check Tag
• Cricetinae MeSH
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Konečné produkty pokročilé glykace (advanced glycation end products – AGEs) hrají významnou roli v patogenezi řady chronických onemocnění a jejich komplikací, především diabetických komplikací, aterosklerózy, komplikací chronických onemocnění ledvin a neurodegenerativních onemocnění. Tyto látky vznikají neenzymatickou glykací a jejich tvorba je potencována vlivem karbonylového stresu. AGEs tvoří heterogenní skupinu látek a patří mezi ně např. karboxymetyllyzin, pentozin, metylglyoxallyzin dimer, vesperlyzin, imidazolony a další. AGEs jednak modifikují proteiny a mění jejich fyzikální a chemické vlastnosti, jednak mají účinky zprostředkované přes receptory, z nichž nejznámější, ale ne jediný, je receptor RAGE (receptor pro konečné produkty pokročilé glykace). RAGE je receptor multiligandový, váže také HMGB1 (high mobility group box protein 1), S100 proteiny či amyloidové fibrily. Vazba ligand na tento receptor má za následek aktivaci řady signálních cest včetně indukce oxidačního stresu a aktivace nukleárního faktoru κB a následnou prozánětlivou odpověď v závislosti na buněčném typu. AGEs a RAGE se spolu s dalšími mechanizmy – hexosaminovou cestou, polyolovou cestou, poruchou metabolizmu lipidů, aktivací proteinkinázy C, oxidačním stresem a zánětlivou reakcí spoluúčastní v patogenezi diabetických komplikací. Terapeuticky je možné snižovat endogenní tvorbu AGEs, ovlivnit přísun AGEs do organizmu stravou a jejich absorpci ve střevě či stimulovat jejich degradaci. Klíčová slova: AGEs – diabetes mellitus – karbonylový stres – konečné produkty pokročilé glykace – oxidační stres – RAGE – receptor pro AGEs – sRAGE – zánět

Advanced glycation end products (AGEs) play an important role in the pathogenesis of chronic diseases and their complications, especially diabetic complications, atherosclerosis, complications of chronic kidney diseases and neurodegenerative diseases. These substances are formed via non-enzymatic glycation and their formation is potentiated in case of carbonyl stress. AGEs are represented by a heterogeneous group of compounds, e.g. carboxymethyllysine, pentosine, methylglyoxallysin dimer, vesperlysine, imidazolones etc. AGEs can modify proteins and so change their physical and chemical properties and can act also via specific receptors, among them RAGE (receptor for advanced glycation end products) is the best known but not the unique one. RAGE is a multiligand receptor capable to bind also HMGB1 (high mobility group box protein 1), S100 proteins or amyloid fibrils. RAGE – ligand interactions results to activation of a variety of signaling pathways including oxidative stress and activation of nuclear factor κB and subsequent proinflammatory response depending on the cell type. AGEs and RAGE together with further mechanisms – hexosamine pathway, polyol pathway, lipid metabolism disorder, activation of proteinkinase C, oxidative stress and inflammatory reaction take part in the pathogenesis of diabetic complications. Terapeuticaly it is possible to decrease endogenous formation of AGEs, influence the AGEs intake to the organism and their absorption in the intestine or stimulate their degradation. Key words: AGEs – advanced glycation end-products – carbonyl stress – diabetes mellitus – inflammation – oxidative stress – RAGE – receptor for AGEs – sRAGE

• Keywords
• karbonylový stres, sRAGE, receptor pro AGEs,
• MeSH
• Diabetes Mellitus etiology   MeSH
• Glycated Hemoglobin physiology   MeSH
• Humans MeSH
• Oxidative Stress physiology   MeSH
• Glycation End Products, Advanced * physiology   metabolism   adverse effects   MeSH
• Receptors, Immunologic * biosynthesis   MeSH
• Inflammation MeSH
• Check Tag
• Humans MeSH
• Publication type
• Research Support, Non-U.S. Gov't MeSH

The binding of monosaccharides and short peptides to lymphocyte receptors (human CD69 and rat NKR-P1A) was first reported in 1994 and then in a number of subsequent publications. Based on this observation, numerous potentially high-affinity saccharide ligands have been synthesized over the last two decades in order to utilize their potential in antitumor therapy. Due to significant inconsistencies in their reported binding properties, we decided to re-examine the interaction between multiple ligands and CD69 or NKR-P1A. Using NMR titration and isothermal titration calorimetry we were unable to detect the binding of the tested ligands such as N-acetyl-D-hexosamines and oligopeptides to both receptors, which contradicts the previous observations published in more than twenty papers over the last fifteen years.

• MeSH
• Antigens, CD metabolism   MeSH
• Antigens, Differentiation, T-Lymphocyte metabolism   MeSH
• Rats MeSH
• Lectins, C-Type metabolism   MeSH
• Humans MeSH
• Oligopeptides chemical synthesis   pharmacology   MeSH
• Polysaccharides chemical synthesis   pharmacology   MeSH
• Receptors, Immunologic metabolism   MeSH
• Recombinant Proteins metabolism   MeSH
• Protein Binding MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

• MeSH
• Adipose Tissue, White metabolism   MeSH
• Energy Metabolism * physiology   MeSH
• Glucose metabolism   MeSH
• Hexosamines MeSH
• Adipose Tissue, Brown metabolism   MeSH
• Thyroid Hormones metabolism   MeSH
• Liver metabolism   drug effects   MeSH
• Muscle, Skeletal physiology   metabolism   MeSH
• Fatty Acids, Nonesterified metabolism   MeSH
• Anti-Obesity Agents pharmacology   classification   MeSH
• Humans MeSH
• Lipoprotein Lipase metabolism   MeSH
• Mitochondrial Proteins physiology   genetics   metabolism   MeSH
• Models, Animal MeSH
• Mice MeSH
• Obesity metabolism   MeSH
• AMP-Activated Protein Kinases physiology   metabolism   MeSH
• Body Weight physiology   drug effects   MeSH
• Thiazolidinediones pharmacology   classification   MeSH
• Adipose Tissue * physiology   metabolism   secretion   MeSH
• Check Tag
• Humans MeSH
• Mice MeSH

Publikace přináší současný pohled na obezitu jako na komplexní nemoc, která v posledních desetiletích nabyla charakteru celosvětové epidemie se závažnými zdravotními a sociálně-ekonomickými dopady. Čtenář se seznámí jak s epidemiologií a etiopatogenezí obezity, tak se současnými postupy v diagnostice, léčbě a prevenci obezity. Autorský kolektiv tvoří lékaři a vědci, kteří jsou uznávanými odborníky v oboru u nás i ve světě. Někteří z nich se významně podíleli na tvorbě evropských doporučení pro léčbu obezity.

• MeSH
• Diagnosis, Differential MeSH
• Child MeSH
• Adult MeSH
• Diabetes Complications MeSH
• Adolescent MeSH
• Obesity, Morbid diagnosis   drug therapy   surgery   immunology   physiopathology   prevention & control   therapy   MeSH
• Obesity history   diagnosis   diet therapy   epidemiology   etiology   genetics   surgery   immunology   therapy   MeSH
• Risk Factors MeSH
• Check Tag
• Child MeSH
• Adult MeSH
• Adolescent MeSH

• Conspectus
• Patologie. Klinická medicína
• Učební osnovy. Vyučovací předměty. Učebnice
• NML Fields
• vnitřní lékařství
• NML Publication type
• kolektivní monografie

Publikace přináší současný pohled na obezitu jako na komplexní nemoc, která v posledních desetiletích nabyla charakteru celosvětové epidemie se závažnými zdravotními a sociálně-ekonomickými dopady. Čtenář se seznámí jak s epidemiologií a etiopatogenezí obezity, tak se současnými postupy v diagnostice, léčbě a prevenci obezity. Autorský kolektiv tvoří lékaři a vědci, kteří jsou uznávanými odborníky v oboru u nás i ve světě. Někteří z nich se významně podíleli na tvorbě evropských doporučení pro léčbu obezity.

• Keywords
• Interna, Metabolizmus, imunita, Ostatní lékařské obory,
• MeSH
• Diagnosis, Differential MeSH
• Child MeSH
• Adult MeSH
• Diabetes Complications MeSH
• Adolescent MeSH
• Obesity, Morbid diagnosis   drug therapy   surgery   immunology   physiopathology   prevention & control   therapy   MeSH
• Obesity history   diagnosis   diet therapy   epidemiology   etiology   genetics   surgery   immunology   therapy   MeSH
• Risk Factors MeSH
• Check Tag
• Child MeSH
• Adult MeSH
• Adolescent MeSH

• NML Fields
• vnitřní lékařství

BACKGROUND: Fibrinogen-related proteins with lectin activity are believed to be part of the tick innate immune system. Several fibrinogen-related proteins have been described and characterised mainly on the basis of their cDNA sequences while direct biochemical evidence is missing. One of them, the haemolymph lectin Dorin M from the tick Ornithodoros moubata was isolated and characterised in more depth. RESULTS: Several fibrinogen-related proteins were detected in the haemolymph of ixodid ticks Dermacentor marginatus, Rhipicephalus appendiculatus, R. pulchellus, and R. sanguineus. These proteins were recognised by sera directed against the tick lectin Dorin M and the haemagglutination activity of the ticks R. appendiculatus and D. marginatus. Cross-reactivity of the identified proteins with antibodies against the fibrinogen domain of the human ficolin was also shown. The carbohydrate-binding ability of tick haemolymph was confirmed by haemagglutination activity assays, and this activity was shown to be inhibited by neuraminic acid and sialylated glycoproteins as well as by N-acetylated hexosamines. The fibrinogen-related proteins were shown to be glycosylated and they were localised in salivary glands, midguts, and haemocytes of D. marginatus. Hemelipoglycoprotein was also recognised by sera directed against the fibrinogen-related proteins in all three Rhipicephalus species as well as in D. marginatus. However, this protein does not contain the fibrinogen domain and thus, the binding possibly results from the structure similarity between hemelipoglycoprotein and the fibrinogen domain. CONCLUSIONS: The presence of fibrinogen-related proteins was shown in the haemolymph of four tick species in high abundance. Reactivity of antibodies directed against ficolin or fibrinogen-related proteins with proteins which do not contain the fibrinogen domain points out the importance of sequence analysis of the identified proteins in further studies. Previously observed expression of fibrinogen-related proteins in haemocytes together with the results of this study suggest involvement of fibrinogen-related proteins in tick immunity processes. Thus, they have potential as targets for anti-tick vaccines and as antimicrobial proteins in pharmacology. Research on fibrinogen-related proteins could reveal further details of tick innate immunity processes.

• MeSH
• Animal Structures chemistry   MeSH
• Dermacentor chemistry   MeSH
• Fibrinogen immunology   MeSH
• Glycoproteins immunology   metabolism   MeSH
• Hemagglutinins immunology   metabolism   MeSH
• Insect Proteins immunology   metabolism   MeSH
• Lectins immunology   metabolism   MeSH
• Humans MeSH
• Carbohydrate Metabolism MeSH
• Ornithodoros chemistry   MeSH
• Antibodies immunology   MeSH
• Rhipicephalus chemistry   MeSH
• Protein Binding MeSH
• Cross Reactions MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH