Inhibition of rat brain glutamate decarboxylase (GAD, EC 4.1.1.15) by individual stereoisomers of 4-fluoroglutamate (4-F-Glu) and 2-fluoro-4-aminobutyrate (2-F-GABA) was studied. All stereoisomers of 4-F-Glu inhibited decarboxylation of L-glutamate catalysed by the enzyme preparation. At 1 x 10(-2) M concentration, the most potent inhibitor of GAD was D-erythro-4-F-Glu with about 70% inhibition in the presence of 1.23 x 10(-2)M L-glutamate. The inhibition by all stereoisomers was of the competitive type. Ki values ranged from 2 x 10(-3)M for the D-erythro isomer to 1.1 x 10(-2)M for the D-threo and L-erythro isomers. The influence of all stereoisomers was reversible as shown by dialysis except for a small amount in the case of the D-erythro isomer. The inhibition was independent of external pyridoxal-5'-phosphate added. No inhibition of rat brain GAD was found with 2-fluoro-4-aminobutyrate stereoisomers.

• MeSH
• GABA analogy a deriváty   farmakologie   MeSH
• glutamát dekarboxyláza antagonisté a inhibitory   MeSH
• glutamáty farmakologie   MeSH
• inhibitory enzymů farmakologie   MeSH
• krysa rodu Rattus MeSH
• mozek enzymologie   MeSH
• stereoizomerie MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• 4-fluoroglutamic acid MeSH Prohlížeč
• GABA MeSH
• glutamát dekarboxyláza MeSH
• glutamáty MeSH
• inhibitory enzymů MeSH

The inhibitory activity of the quaternary benzophenanthridine alkaloid sanguinarine on rat brain glutamate decarboxylase (GAD; EC 4.1.1.15) was studied in vitro. A value of Ki 7.10(-4) mol.1(-1) for sanguinarine was found. The inhibition was irreversible and increased during the preincubation time of sanguinarine with the GAD preparation. The results suggest that reaction of the iminium bond in the benzophenanthridine molecule with thiol groups of the enzyme participates in GAD inhibition. As GAD catalyzes the rate-limiting step of GABA synthesis, its inhibition by sanguinarine may contribute to its physiological action in vivo.

• MeSH
• alkaloidy chemie   farmakologie   MeSH
• benzofenantridiny MeSH
• GABA biosyntéza   MeSH
• glutamát dekarboxyláza antagonisté a inhibitory   MeSH
• inhibitory enzymů chemie   farmakologie   MeSH
• isochinoliny MeSH
• kinetika MeSH
• krysa rodu Rattus MeSH
• mozek účinky léků   enzymologie   MeSH
• potkani Wistar MeSH
• techniky in vitro MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• alkaloidy MeSH
• benzofenantridiny MeSH
• GABA MeSH
• glutamát dekarboxyláza MeSH
• inhibitory enzymů MeSH
• isochinoliny MeSH
• sanguinarine MeSH Prohlížeč

Glutamate decarboxylase (EC 4.1.1.15, GAD) activity was studied in the brain of 12-day-old and adult rats treated with 3-mercaptopropionic acid (3-MPA), an inhibitor of GAD competitive with glutamate. Control GAD activity in the brains of immature animals (91.8 +/- 18.2 nmol/h/mg of protein) was lower than that of the adult rats (228 +/- 37.5 nmol/h/mg of protein). Brain GAD inhibition in adult rats was 58% at the onset of seizures (9 min on the average after administration of 70 mg 3-MPA/kg). At the same time, 3-MPA-treated young rats exhibited 76% inhibition of GAD despite the fact that at 9 min these animals were not yet having seizures. At the onset of seizures (19 min after 3-MPA on the average) their GAD activity remained at the same level. The difference between the groups was not related to the presence of the coenzyme pyridoxal-5'-phosphate in the enzyme assay. The inhibition of GAD by 3-MPA in vitro in the immature and adult brains was similar (Ki at 5.1 microM and 4.8 microM concentrations of 3-MPA, respectively). Identical values were found for Km of GAD (at 4.5 mM concentration of L-glutamate). Calculations based on the results suggest that 3-MPA enters the immature brain more easily than the brain of the adult animals. While GAD inhibition by 3-MPA is the primary cause of seizures, their onset is influenced by other factors, in which the immature brain differs from the adult one and which may include less sensitivity to GABA decrease due to relative overactivity of the GABA system.

• MeSH
• glutamát dekarboxyláza antagonisté a inhibitory   MeSH
• krysa rodu Rattus MeSH
• kyselina 3-merkaptopropionová farmakologie   MeSH
• mozek enzymologie   růst a vývoj   MeSH
• potkani Wistar MeSH
• pyridoxalfosfát fyziologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• glutamát dekarboxyláza MeSH
• kyselina 3-merkaptopropionová MeSH
• pyridoxalfosfát MeSH

UNLABELLED: Tolerogenic DCs (tolDCs) are being researched as a promising intervention strategy also in autoimmune diseases including type 1 diabetes (T1D). T1D is a T-cell-mediated, organ-specific disease with several well-defined and rather specific autoantigens, i.e., proinsulin, insulin, glutamic acid decarboxylase 65 (GAD65), that have been used in animal as well as human intervention trials in attempts to achieve a more efficient, specific immunotherapy. In this study, we have tested tolerogenic DCs for their effectiveness to prevent adoptive transfer of diabetes by diabetogenic splenocytes into non-obese diabetes (NOD)-severe combined immunodeficiency (NOD-SCID) recipients. While i.p. application of tolDCs prepared from bone marrow of prediabetic NOD mice by vitamin D2 and dexamethasone significantly reduced diabetes transfer into the NOD-SCID females, this effect was completely abolished when tolDCs were loaded with the mouse recombinant GAD65, but also with a control protein-ovalbumin (OVA). The effect was not dependent on the presence of serum in the tolDC culture. Similar results were observed in NOD mice. Removal of possible bystander antigen-presenting cells within the diabetogenic splenocytes by negative magnetic sorting of T cells did not alter this surprising effect. Tolerogenic DCs loaded with an immunodominant mouse GAD65 peptide also displayed diminished diabetes-preventive effect. Tolerogenic DCs were characterized by surface maturation markers (CD40, CD80, CD86, MHC II) and the lipopolysaccharide stability test. Data from alloreactive T cell proliferation and cytokine induction assays (IFN-γ) did not reveal the differences observed in the diabetes incidence. Migration of tolDCs, tolDCs-GAD65 and tolDCs-OVA to spleen, mesenteric- and pancreatic lymph nodes displayed similar, mucosal pattern with highest accumulation in pancreatic lymph nodes present up to 9 days after the i.p. APPLICATION: These data document that mechanisms by which tolDCs operate in vivo require much better understanding for improving efficacy of this promising cell therapy, especially in the presence of an antigen, e.g., GAD65.

• Klíčová slova
• autoantigen, cell therapy, dendritic cells, glutamic acid decarboxylase 65, non-obese diabetes mice, non-obese diabetes-severe combined immunodeficiency mouse, tolerogenic, type 1 diabetes,
• MeSH
• autoantigeny imunologie   MeSH
• dendritické buňky imunologie   MeSH
• diabetes mellitus 1. typu imunologie   MeSH
• glutamát dekarboxyláza imunologie   MeSH
• imunologická tolerance imunologie   MeSH
• modely nemocí na zvířatech MeSH
• myši inbrední C57BL MeSH
• myši inbrední NOD MeSH
• myši SCID MeSH
• myši MeSH
• převzatá imunita MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• autoantigeny MeSH
• glutamát dekarboxyláza MeSH
• glutamate decarboxylase 2 MeSH Prohlížeč

OBJECTIVE: To evaluate the efficacy of aluminum-formulated intralymphatic glutamic acid decarboxylase (GAD-alum) therapy combined with vitamin D supplementation in preserving endogenous insulin secretion in all patients with type 1 diabetes (T1D) or in a genetically prespecified subgroup. RESEARCH DESIGN AND METHODS: In a multicenter, randomized, placebo-controlled, double-blind trial, 109 patients aged 12-24 years (mean ± SD 16.4 ± 4.1) with a diabetes duration of 7-193 days (88.8 ± 51.4), elevated serum GAD65 autoantibodies, and a fasting serum C-peptide >0.12 nmol/L were recruited. Participants were randomized to receive either three intralymphatic injections (1 month apart) with 4 μg GAD-alum and oral vitamin D (2,000 IE daily for 120 days) or placebo. The primary outcome was the change in stimulated serum C-peptide (mean area under the curve [AUC] after a mixed-meal tolerance test) between baseline and 15 months. RESULTS: Primary end point was not met in the full analysis set (treatment effect ratio 1.091 [CI 0.845-1.408]; P = 0.5009). However, GAD-alum-treated patients carrying HLA DR3-DQ2 (n = 29; defined as DRB1*03, DQB1*02:01) showed greater preservation of C-peptide AUC (treatment effect ratio 1.557 [CI 1.126-2.153]; P = 0.0078) after 15 months compared with individuals receiving placebo with the same genotype (n = 17). Several secondary end points showed supporting trends, and a positive effect was seen in partial remission (insulin dose-adjusted HbA1c ≤9; P = 0.0310). Minor transient injection site reactions were reported. CONCLUSION: Intralymphatic administration of GAD-alum is a simple, well-tolerated treatment that together with vitamin D supplementation seems to preserve C-peptide in patients with recent-onset T1D carrying HLA DR3-DQ2. This constitutes a disease-modifying treatment for T1D with a precision medicine approach.

• MeSH
• C-peptid MeSH
• diabetes mellitus 1. typu * farmakoterapie   MeSH
• dvojitá slepá metoda MeSH
• glutamát dekarboxyláza * MeSH
• lidé MeSH
• potravní doplňky MeSH
• vitamin D MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• klinické zkoušky, fáze II MeSH
• multicentrická studie MeSH
• práce podpořená grantem MeSH
• randomizované kontrolované studie MeSH
• Názvy látek
• C-peptid MeSH
• glutamát dekarboxyláza * MeSH
• vitamin D MeSH

• Klíčová slova
• CYANIDES/pharmacology *, DESMOLASES/metabolism *, TRANSAMINASES/metabolism *,
• MeSH
• GABA * MeSH
• glutamát dekarboxyláza * MeSH
• kyanid draselný * MeSH
• kyanidy farmakologie   MeSH
• lyasy metabolismus   MeSH
• mozek * MeSH
• transaminasy metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• GABA * MeSH
• glutamát dekarboxyláza * MeSH
• kyanid draselný * MeSH
• kyanidy MeSH
• lyasy MeSH
• transaminasy MeSH

• MeSH
• anaerobní bakterie enzymologie   izolace a purifikace   MeSH
• bakteriologické techniky MeSH
• Enterobacteriaceae izolace a purifikace   MeSH
• glutamát dekarboxyláza metabolismus   MeSH
• gramnegativní bakterie enzymologie   izolace a purifikace   MeSH
• papírová chromatografie MeSH
• Vibrionaceae izolace a purifikace   MeSH
• Publikační typ
• anglický abstrakt MeSH
• časopisecké články MeSH
• Názvy látek
• glutamát dekarboxyláza MeSH

BACKGROUND: It is not always easy to classify diabetes (DM) diagnosed in adults, with a significant group of patients initially classified and treated for type 2 diabetes mellitus (DM2T) presenting signs indicating the presence of autoimmune insulitis (AI), which is characteristic of type 1 diabetes mellitus (DM1T), or latent autoimmune diabetes mellitus in adults (LADA). GOAL: Identify the proportion of patients entered with DM2T who present AI signs, and the number of patients of that proportion, who at the same time present low insulin secretion, and what clinical and laboratory manifestations could be used to differentiate between these patients.Cohort and methods: A randomized clinical trial with a pre-determined set of assessed parameters for n = 625 patients, who were hospitalized during the first 6 months of 2016 at the National Endocrinology and Diabetology Institute (NEDU), Lubochna. Apart from the standard parameters, C-peptide (CP) and autoantibodies to glutamic acid decarboxylase (GADA) were examined for each patient. GADA positive (GADA+) patients were compared to GADA negative (GADA-) patients in the following parameters: gender, age, age at the time of dia-gnosing DM, duration of DM, HbA1c, incidence of hypoglycemia, lipidogram, fasting C-peptide levels, BMI, waist circumference, incidence of hypoglycemias, presence of microvascular and macrovascular complications, treatment of dia-betes and incidence of other endocrinopathies. GADA+ with low CP were subsequently compared to GADA+ patients with normal CP. RESULTS: Of 625 patients originally classified and treated as DM2T, 13 % were GADA+. 31 % of them had low CP (< 0.2 nmol/l) and 28 % had CP levels within the intermediary range (0.2-0.4 nmol/l). Females made up a larger proportion of GADA+ patients, with a lower BMI, smaller waist circumference, lower CP, higher HDL cholesterol levels, a greater incidence of hypoglycemias and lower total daily dose of insulin. GADA+ patients with low CP differed from GADA+ patients with normal CP in higher HDL cholesterol levels, lower triglyceride levels and earlier need of insulin thera-py. The testing for GADA and CP levels with regard to the other relevant characteristics led to re-classification, or more precisely adding of DM1T/LADA (as the main, or parallel cause of DM) for 2.9 % of all the patients included and a clinically significant proportion of AI could be assumed in 6.1 % of the patients. SUMMARY: The results of our study show that the pathogenesis of DM in patients initially diagnosed and registered with DM2T and with concurrent presence of GADA includes mechanisms characteristic of both DM2T (insulin resistance) and DM1T (autoimmune insulitis) acting in parallel, with different intensity, in differing proportions and time sequence as a fluid continuum, which also accounts for the differences between individual patients. The characteristics highlighting the presence and role of AI based on our results include high titre of GADA+, low CP levels, early need of insulin therapy, presence of thyroid disorder, higher HDL cholesterol levels and lower triglyceride levels. The characteristics highlighting the dominance of mechanisms characteristic of DM2T (insulin resistance) included higher BMI and waist circumference values, normal CP levels, low HDL cholesterol levels, higher triglyceride levels, higher blood pressure and borderline titre of GADA. KEY WORDS: autoimmune diabetes mellitus - C-peptide - GADA - HDL-cholesterol - classification.

• MeSH
• autoprotilátky krev   MeSH
• C-peptid krev   MeSH
• diabetes mellitus 1. typu farmakoterapie   epidemiologie   MeSH
• diabetes mellitus 2. typu krev   farmakoterapie   imunologie   MeSH
• dospělí MeSH
• glutamát dekarboxyláza krev   MeSH
• HDL-cholesterol krev   MeSH
• hospitalizace MeSH
• hypoglykemie farmakoterapie   epidemiologie   MeSH
• hypoglykemika terapeutické užití   MeSH
• inzulin terapeutické užití   MeSH
• inzulinová rezistence fyziologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• nemoci štítné žlázy epidemiologie   MeSH
• obvod pasu MeSH
• senioři MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• randomizované kontrolované studie MeSH
• Názvy látek
• autoprotilátky MeSH
• C-peptid MeSH
• glutamát dekarboxyláza MeSH
• HDL-cholesterol MeSH
• hypoglykemika MeSH
• inzulin MeSH

AIM OF STUDY: To assess the prevalence of markers of autoimmune destruction of pancreatic beta-cells in patients with non-insulin dependent diabetes mellitus (NIDDM). SUBJECTS: 127 hospitalized NIDDM patients subdivided to the following subgroups: non-obese with C-peptide < 0.3 nmol/l (NIDDM-(-)), non-obese with C-peptide > 0.3 nmol/l (NIDDM-(+)), obese with C-peptide < 0.3 nmol/l (NIDDM+(-)) and obese with C-peptide > 0.3 nmol/l (NIDDM2+). METHODS AND MEASURED PARAMETERS: Age, BMI, C-peptide, autoantibodies to glutamic acid decarboxylase (antiGAD-Ab), autoantibodies to islet cells (ICA), markers of specific cellular immunity CD4, CD8, CD19, CD4/CD8, CD4/CD45/RA+, CD4/CD45/RA-, NK (CD16+56), CD3/HLADR, organ specific/non-specific autoantibodies. RESULTS: AntiGAD-Ab were positive in 5/15 (33.3%) NIDDM-(-), 1/32 (3.1%) NIDDM-(+), 2/9 (22.2%) NIDDM+(-) and in 3/71 (4.2%) NIDDM2+. The positivity of antiGAD-Ab in NIDDM-(-) and NIDDM+(-) was significantly higher (p < 0.05) than in NIDDM-(+) and NIDDM2+. CONCLUSION: Some patients with manifestation of diabetes in older age initially classified and treated as having NIDDM may have in fact slowly evolving autoimmune insulin-dependent diabetes mellitus (LADA). These patients can be identified by measurement of antiGAD-Ab or other markers (ICA, IA-2) of autoimmune destruction of pancreatic beta-cells (AID). Moreover, in some patients both AID and insulin resistance may coexist in parallel.

• MeSH
• autoprotilátky analýza   MeSH
• biologické markery krev   MeSH
• C-peptid krev   MeSH
• diabetes mellitus 2. typu krev   imunologie   patologie   MeSH
• diabetes mellitus krev   imunologie   MeSH
• dospělí MeSH
• glutamát dekarboxyláza imunologie   MeSH
• Langerhansovy ostrůvky patologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• obezita MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• anglický abstrakt MeSH
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• autoprotilátky MeSH
• biologické markery MeSH
• C-peptid MeSH
• glutamát dekarboxyláza MeSH

Variability in the number of tandem repeats of the insulin gene (INS VNTR) is probably involved in the genetic regulation of insulin secretion. The aim of this study was to investigate the association of INS VNTR polymorphism with the presence of glutamic acid decarboxylase antibodies (GADA) and C-peptide levels in patients with the onset of diabetes after 35 years of age. We investigated 117 patients, median of age 63 (range 40-83) years, median of diabetes duration 8 (range 1-30) years; 31 GADA-positive and 86 GADA-negative subjects. INS VNTR polymorphism was typed indirectly using - 23HphI (T/A) polymorphism, which is in complete linkage disequilibrium with INS VNTR. The I/I, I/III and III/III genotypes were found in 22 (71 %), 8 (26 %), 1 (3 %) GADA-positive individuals and in 39 (45 %), 35 (41 %), 12 (14 %) GADA-negative individuals, respectively. The Class I allele and the genotype I/I were significantly associated with the presence of GADA (OR=2.72, CI 95 %=1.29-5.73 and OR=2.95, CI 95 %=1.22-7.13). The presence of Class III allele was significantly associated with a higher level of postprandial C-peptide in GADA-positive subjects, even when regarding the duration of diabetes. Our results of INS VNTR polymorphism in patients with the onset of diabetes after 35 years of age confirm the association of Class I INS VNTR with autoimmune diabetes and the protective effect of Class III INS VNTR on the insulin secretion in GADA-positive subjects.

• MeSH
• C-peptid krev   metabolismus   MeSH
• diabetes mellitus genetika   imunologie   metabolismus   MeSH
• DNA genetika   izolace a purifikace   MeSH
• dospělí MeSH
• frekvence genu MeSH
• genotyp MeSH
• glutamát dekarboxyláza imunologie   MeSH
• heterozygot MeSH
• homozygot MeSH
• interval spolehlivosti MeSH
• inzulin genetika   metabolismus   MeSH
• lidé středního věku MeSH
• lidé MeSH
• minisatelitní repetice genetika   MeSH
• odds ratio MeSH
• polymorfismus genetický * MeSH
• postprandiální období MeSH
• protilátky krev   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• věk při počátku nemoci MeSH
• výběr pacientů MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• C-peptid MeSH
• DNA MeSH
• glutamát dekarboxyláza MeSH
• inzulin MeSH
• protilátky MeSH