Regulation of H2S homeostasis in humans is poorly understood. Therefore, we assessed the importance of individual enzymes in synthesis and catabolism of H2S by studying patients with respective genetic defects. We analyzed sulfur compounds (including bioavailable sulfide) in 37 untreated or insufficiently treated patients with seven ultrarare enzyme deficiencies and compared them to 63 controls. Surprisingly, we observed that patients with severe deficiency in cystathionine β-synthase (CBS) or cystathionine γ-lyase (CSE) - the enzymes primarily responsible for H2S synthesis - exhibited increased and normal levels of bioavailable sulfide, respectively. However, an approximately 21-fold increase of urinary homolanthionine in CBS deficiency strongly suggests that lacking CBS activity is compensated for by an increase in CSE-dependent H2S synthesis from accumulating homocysteine, which suggests a control of H2S homeostasis in vivo. In deficiency of sulfide:quinone oxidoreductase - the first enzyme in mitochondrial H2S oxidation - we found normal H2S concentrations in a symptomatic patient and his asymptomatic sibling, and elevated levels in an asymptomatic sibling, challenging the requirement for this enzyme in catabolizing H2S under physiological conditions. Patients with ethylmalonic encephalopathy and sulfite oxidase/molybdenum cofactor deficiencies exhibited massive accumulation of thiosulfate and sulfite with formation of large amounts of S-sulfocysteine and S-sulfohomocysteine, increased renal losses of sulfur compounds and concomitant strong reduction in plasma total cysteine. Our results demonstrate the value of a comprehensive assessment of sulfur compounds in severe disorders of homocysteine/cysteine metabolism and provide evidence for redundancy and compensatory mechanisms in the maintenance of H2S homeostasis.

• MeSH
• cystein MeSH
• homeostáza MeSH
• homocystein MeSH
• lidé MeSH
• síra MeSH
• sulfan * metabolismus   MeSH
• sulfidy metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Cystathionine β-synthase (CBS) deficiency has a wide clinical spectrum, ranging from neurodevelopmental problems, lens dislocation and marfanoid features in early childhood to adult onset disease with predominantly thromboembolic complications. We have analysed clinical and laboratory data at the time of diagnosis in 328 patients with CBS deficiency from the E-HOD (European network and registry for Homocystinurias and methylation Defects) registry. We developed comprehensive criteria to classify patients into four groups of pyridoxine responsivity: non-responders (NR), partial, full and extreme responders (PR, FR and ER, respectively). All groups showed overlapping concentrations of plasma total homocysteine while pyridoxine responsiveness inversely correlated with plasma/serum methionine concentrations. The FR and ER groups had a later age of onset and diagnosis and a longer diagnostic delay than NR and PR patients. Lens dislocation was common in all groups except ER but the age of dislocation increased with increasing responsiveness. Developmental delay was commonest in the NR group while no ER patient had cognitive impairment. Thromboembolism was the commonest presenting feature in ER patients, whereas it was least likely at presentation in the NR group. This probably is due to the differences in ages at presentation: all groups had a similar number of thromboembolic events per 1000 patient-years. Clinical severity of CBS deficiency depends on the degree of pyridoxine responsiveness. Therefore, a standardised pyridoxine-responsiveness test in newly diagnosed patients and a critical review of previous assessments is indispensable to ensure adequate therapy and to prevent or reduce long-term complications.

• MeSH
• cystathionin-beta-synthasa nedostatek   MeSH
• dítě MeSH
• dospělí MeSH
• fenotyp MeSH
• homocystinurie diagnóza   farmakoterapie   enzymologie   MeSH
• kojenec MeSH
• lidé středního věku MeSH
• lidé MeSH
• lineární modely MeSH
• methionin krev   MeSH
• mladiství MeSH
• mladý dospělý MeSH
• opožděná diagnóza MeSH
• předškolní dítě MeSH
• pyridoxin terapeutické užití   MeSH
• registrace MeSH
• senioři MeSH
• stupeň závažnosti nemoci MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• kojenec MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Evropa MeSH

The chicken Tva cell surface protein, a member of the low-density lipoprotein receptor family, has been identified as an entry receptor for avian leukosis virus of classic subgroup A and newly emerging subgroup K. Because both viruses represent an important concern for the poultry industry, we introduced a frame-shifting deletion into the chicken tva locus with the aim of knocking-out Tva expression and creating a virus-resistant chicken line. The tva knock-out was prepared by CRISPR/Cas9 gene editing in chicken primordial germ cells and orthotopic transplantation of edited cells into the testes of sterilized recipient roosters. The resulting tva -/- chickens tested fully resistant to avian leukosis virus subgroups A and K, both in in vitro and in vivo assays, in contrast to their susceptible tva +/+ and tva +/- siblings. We also found a specific disorder of the cobalamin/vitamin B12 metabolism in the tva knock-out chickens, which is in accordance with the recently recognized physiological function of Tva as a receptor for cobalamin in complex with transcobalamin transporter. Last but not least, we bring a new example of the de novo resistance created by CRISPR/Cas9 editing of pathogen dependence genes in farm animals and, furthermore, a new example of gene editing in chicken.

• MeSH
• editace genu MeSH
• genový knockout MeSH
• kur domácí virologie   MeSH
• kuřecí embryo MeSH
• kyselina methylmalonová krev   MeSH
• posunová mutace MeSH
• ptačí proteiny genetika   fyziologie   MeSH
• virové receptory genetika   fyziologie   MeSH
• virus ptačí leukózy klasifikace   fyziologie   MeSH
• vitamin B 12 metabolismus   MeSH
• zvířata MeSH
• Check Tag
• kuřecí embryo MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Combined methylmalonic aciduria with homocystinuria (cblC type) is a rare disease caused by mutations in the MMACHC gene. MMACHC encodes an enzyme crucial for intracellular vitamin B12 metabolism, leading to the accumulation of toxic metabolites e.g. methylmalonic acid (MMA) and homocysteine (Hcy), and secondary disturbances in folate and one-carbon metabolism when not fully functional. Patients with cblC deficiency often present in the neonatal or early childhood period with a severe multisystem pathology, which comprises a broad spectrum of treatment-resistant ophthalmological phenotypes, including retinal degeneration, impaired vision, and vascular changes. To examine the potential function of MMACHC in the retina and how its loss may impact disease, we performed gene expression studies in human and mouse, which showed that local expression of MMACHC in the retina and retinal pigment epithelium is relatively stable over time. To study whether functional MMACHC is required for retinal function and tissue integrity, we generated a transgenic mouse lacking Mmachc expression in cells of the peripheral retina. Characterization of this mouse revealed accumulation of cblC disease related metabolites, including MMA and the folate-dependent purine synthesis intermediates AICA-riboside and SAICA-riboside in the retina. Nevertheless, fundus appearance, morphology, vasculature, and cellular composition of the retina, as well as ocular function, remained normal in mice up to 6 or 12 months of age. Our data indicates that peripheral retinal neurons do not require intrinsic expression of Mmachc for survival and function and questions whether a local MMACHC deficiency is responsible for the retinal phenotypes in patients.

• MeSH
• degenerace retiny genetika   metabolismus   MeSH
• dospělí MeSH
• fenotyp MeSH
• homocystein metabolismus   MeSH
• homocystinurie metabolismus   MeSH
• kyselina methylmalonová metabolismus   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mutace genetika   MeSH
• myši inbrední BALB C MeSH
• myši inbrední C57BL MeSH
• myši transgenní MeSH
• myši MeSH
• oxidoreduktasy genetika   metabolismus   MeSH
• retina metabolismus   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• vitamin B 12 metabolismus   MeSH
• vrozené poruchy metabolismu aminokyselin metabolismus   MeSH
• zvířata MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• myši MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

• Publikační typ
• abstrakt z konference MeSH

Classical homocystinuria (HCU) is an inborn error of metabolism caused by loss of cystathionine β-synthase (CBS) activity with the concomitant buildup of homocysteine. In knockout (KO) mice, a mouse model of HCU, complete lack of CBS is neonatally lethal. Administration of OT-58, an enzyme therapy for HCU, during the first 5 weeks of life rescued KO mice survival by preventing liver disease. Here, we studied the impact of a long-term uninterrupted OT-58 treatment or its absence beyond the neonatal period on liver pathology and metabolism. Plasma and liver metabolites of KO mice on OT-58 treatment were substantially improved or normalized compared with those receiving vehicle. Increased plasma activities of alanine aminotransferase and aspartate aminotransferase of vehicle-injected KO mice suggested the progression of liver damage with age and lack of treatment. At 3 months of age, liver histology showed no signs of hepatopathy in both vehicle- and OT-58-treated KO mice. However, moderate to severe liver disease, characterized by steatosis, hepatocellular necroses, disorganized endoplasmic reticulum, and swollen mitochondria, developed in 6-month-old vehicle-injected KO mice. KO mice on OT-58 treatment remained asymptomatic and were indistinguishable from age-matched healthy controls. Long-term uninterrupted OT-58 treatment was essential to prevent severe liver disease in the KO mouse model of HCU.

• MeSH
• enzymová substituční terapie * MeSH
• homocystinurie farmakoterapie   MeSH
• játra patologie   MeSH
• modely nemocí na zvířatech MeSH
• myši knockoutované MeSH
• myši MeSH
• nemoci jater prevence a kontrola   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

• MeSH
• buněčné klony MeSH
• buňky K562 MeSH
• CRISPR-Cas systémy MeSH
• genový knockout MeSH
• homozygot MeSH
• hyperhomocysteinemie farmakoterapie   genetika   MeSH
• kultivované buňky MeSH
• kyselina listová terapeutické užití   MeSH
• lidé MeSH
• megaloblastová anemie farmakoterapie   genetika   MeSH
• mladiství MeSH
• posunová mutace MeSH
• recidiva MeSH
• sekvenční delece MeSH
• sekvenování exomu MeSH
• sodíko-vodíkový výměnný transportér 1 nedostatek   genetika   MeSH
• vitamin B 12 terapeutické užití   MeSH
• Check Tag
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• Publikační typ
• dopisy MeSH
• kazuistiky MeSH
• práce podpořená grantem MeSH

BACKGROUND AND PURPOSE: Homocystinurias are rare genetic defects characterized by altered fluxes of sulfur compounds including homocysteine and cysteine. We explored whether the severely perturbed sulfur amino acid metabolism in patients with homocystinurias affects the metabolism of hydrogen sulfide. EXPERIMENTAL APPROACH: We studied 10 treated patients with a block in the conversion of homocysteine to cysteine due to cystathionine β-synthase deficiency (CBSD) and six treated patients with remethylation defects (RMD) and an enhanced flux of sulfur metabolites via transsulfuration. Control groups for CBSD and RMD patients consisted of 22 patients with phenylketonuria on a low-protein diet and of 12 healthy controls respectively. Plasma and urine concentrations of selected sulfur compounds were analysed by HPLC and LC-MS/MS. KEY RESULTS: Patients with CBSD exhibited plasma concentrations of monobromobimane-detected sulfide similar to appropriate controls. Urinary homolanthionine and thiosulfate in CBSD were increased significantly 1.9 and 3 times suggesting higher hydrogen sulfide synthesis by γ-cystathionase and detoxification respectively. Surprisingly, patients with RMD had significantly lower plasma sulfide levels (53 and 64% of controls) with lower sulfite concentrations, and higher taurine and thiosulfate levels suggesting enhanced cysteine oxidation and hydrogen sulfide catabolism respectively. CONCLUSION AND IMPLICATIONS: The results from this study suggest that severe inherited defects in sulfur amino acid metabolism may be accompanied by only moderately perturbed hydrogen sulfide metabolism and lends support to the hypothesis that enzymes in the transsulfuration pathway may not be the major contributors to the endogenous hydrogen sulfide pool. LINKED ARTICLES: This article is part of a themed section on Chemical Biology of Reactive Sulfur Species. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.4/issuetoc.

• MeSH
• dítě MeSH
• dospělí MeSH
• homocystinurie krev   metabolismus   moč   MeSH
• kojenec MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• předškolní dítě MeSH
• sloučeniny síry krev   metabolismus   moč   MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• kojenec MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The most common ureagenesis defect is X-linked ornithine transcarbamylase (OTC) deficiency which is a main target for novel therapeutic interventions. The spf ash mouse model carries a variant (c.386G>A, p.Arg129His) that is also found in patients. Male spf ash mice have a mild biochemical phenotype with low OTC activity (5%-10% of wild-type), resulting in elevated urinary orotic acid but no hyperammonemia. We recently established a dried blood spot method for in vivo quantification of ureagenesis by Gas chromatography-mass spectrometry (GC-MS) using stable isotopes. Here, we applied this assay to wild-type and spf ash mice to assess ureagenesis at different ages. Unexpectedly, we found an age-dependency with a higher capacity for ammonia detoxification in young mice after weaning. A parallel pattern was observed for carbamoylphosphate synthetase 1 and OTC enzyme expression and activities, which may act as pacemaker of this ammonia detoxification pathway. Moreover, high ureagenesis in younger mice was accompanied by elevated periportal expression of hepatic glutamine synthetase, another main enzyme required for ammonia detoxification. These observations led us to perform a more extensive analysis of the spf ash mouse in comparison to the wild-type, including characterization of the corresponding metabolites, enzyme activities in the liver and plasma and the gut microbiota. In conclusion, the comprehensive enzymatic and metabolic analysis of ureagenesis performed in the presented depth was only possible in animals. Our findings suggest such analyses being essential when using the mouse as a model and revealed age-dependent activity of ammonia detoxification.

• MeSH
• amoniak metabolismus   MeSH
• hyperamonemie genetika   metabolismus   patologie   MeSH
• játra metabolismus   patologie   MeSH
• lidé MeSH
• močovina metabolismus   MeSH
• modely nemocí na zvířatech MeSH
• myši transgenní MeSH
• myši MeSH
• nemoc z nedostatku ornithinkarbamoyltransferázy genetika   metabolismus   patologie   MeSH
• ornithinkarbamoyltransferasa genetika   MeSH
• stárnutí fyziologie   MeSH
• věkové faktory MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

Classical homocystinuria (HCU) is the most common inherited disorder of sulfur amino acid metabolism caused by deficiency in cystathionine beta-synthase (CBS) activity and characterized by severe elevation of homocysteine in blood and tissues. Treatment with dietary methionine restriction is not optimal, and poor compliance leads to serious complications. We developed an enzyme replacement therapy (ERT) and studied its efficacy in a severe form of HCU in mouse (the I278T model). Treatment was initiated before or after the onset of clinical symptoms in an effort to prevent or reverse the phenotype. ERT substantially reduced and sustained plasma homocysteine concentration at around 100 μM and normalized plasma cysteine for up to 9 months of treatment. Biochemical balance was also restored in the liver, kidney, and brain. Furthermore, ERT corrected liver glucose and lipid metabolism. The treatment prevented or reversed facial alopecia, fragile and lean phenotype, and low bone mass. In addition, structurally defective ciliary zonules in the eyes of I278T mice contained low density and/or broken fibers, while administration of ERT from birth partially rescued the ocular phenotype. In conclusion, ERT maintained an improved metabolic pattern and ameliorated many of the clinical complications in the I278T mouse model of HCU.

• MeSH
• aminokyseliny sírové krev   metabolismus   MeSH
• cystathionin-beta-synthasa aplikace a dávkování   chemie   MeSH
• enzymová substituční terapie * MeSH
• fenotyp * MeSH
• glukosa metabolismus   MeSH
• homocystinurie diagnóza   metabolismus   terapie   MeSH
• játra účinky léků   metabolismus   MeSH
• metabolismus lipidů MeSH
• modely nemocí na zvířatech MeSH
• myši MeSH
• oxidační stres MeSH
• polyethylenglykoly chemie   MeSH
• preklinické hodnocení léčiv MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH