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2.
Biosci Biotechnol Biochem ; 84(1): 198-207, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31566090

RESUMO

High glycosidase-producing strains of Aspergillus luchuensis were isolated from 2-deoxyglucose (2-DG) resistant mutants. α-Amylase, exo-α-1,4-glucosidase, ß-glucosidase and ß-xylosidase activity in the mutants was ~3, ~2, ~4 and ~2.5 times higher than the parental strain RIB2604 on koji-making conditions, respectively. Citric acid production and mycelia growth of the mutants, however, approximately halved to that of the parent. Compared to the parent, the alcohol yield from rice and sweet potato shochu mash of the mutant increased ~5.7% and 3.0%, respectively. The mutant strains showed significantly low glucose assimilability despite the fructose one was almost normal, and they had a single missense or nonsense mutation in the glucokinase gene glkA. The recombinant strain that was introduced at one of the mutations, glkA Q300K, demonstrated similar but not identical phenotypes to the mutant strain. This result indicates that glkA Q300K is one of the major mutations in 2-DG resistant strains.


Assuntos
Aspergillus/genética , Aspergillus/isolamento & purificação , Separação Celular/métodos , Códon sem Sentido/genética , Genes Fúngicos/genética , alfa-Glucosidases/metabolismo , Aspergillus/classificação , Aspergillus/metabolismo , Catepsina A/metabolismo , Ácido Cítrico/metabolismo , Desoxiglucose/farmacologia , Farmacorresistência Fúngica , Etanol/metabolismo , Fermentação , Frutose/metabolismo , Glucoquinase/genética , Glucose/metabolismo , Ipomoea batatas/química , Oryza/química , Fenótipo , Saccharomyces cerevisiae/classificação , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/isolamento & purificação , Saccharomyces cerevisiae/metabolismo , Xilosidases/metabolismo , alfa-Amilases/metabolismo , beta-Glucosidase/metabolismo
3.
Int J Mol Sci ; 20(19)2019 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-31569356

RESUMO

Glucose phosphorylating enzymes are crucial in the regulation of basic cellular processes, including metabolism and gene expression. Glucokinases and hexokinases provide a pool of phosphorylated glucose in an adenosine diphosphate (ADP)- and ATP-dependent manner to shape the cell metabolism. The glucose processing enzymes from Kluyveromyces lactis are poorly characterized despite the emerging contribution of this yeast strain to industrial and laboratory scale biotechnology. The first reports on K. lactis glucokinase (KlGlk1) positioned the enzyme as an essential component required for glucose signaling. Nevertheless, no biochemical and structural information was available until now. Here, we present the first crystal structure of KlGlk1 together with biochemical characterization, including substrate specificity and enzyme kinetics. Additionally, comparative analysis of the presented structure and the prior structures of lactis hexokinase (KlHxk1) demonstrates the potential transitions between open and closed enzyme conformations upon ligand binding.


Assuntos
Glucoquinase/química , Kluyveromyces/enzimologia , Modelos Moleculares , Conformação Proteica , Glucoquinase/genética , Glucoquinase/metabolismo , Glucose/metabolismo , Cinética , Kluyveromyces/genética , Kluyveromyces/metabolismo , Especificidade por Substrato
4.
Int J Med Microbiol ; 309(6): 151341, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31451389

RESUMO

Francisella tularensis is the causative agent of the human disease referred to as tularemia. Other Francisella species are known but less is understood about their virulence factors. The role of environmental amoebae in the life-cycle of Francisella is still under discussion. Francisella sp. strain W12-1067 (F-W12) is an environmental Francisella isolate recently identified in Germany which is negative for the Francisella pathogenicity island, but exhibits a putative alternative type VI secretion system. Putative virulence factors have been identified in silico in the genome of F-W12. In this work, we established a "scatter screen", used earlier for pathogenic Legionella, to verify experimentally and identify candidate fitness factors using a transposon mutant bank of F-W12 and Acanthamoeba lenticulata as host organism. In these experiments, we identified 79 scatter clones (amoeba sensitive), which were further analyzed by an infection assay identifying 9 known virulence factors, but also candidate fitness factors of F-W12 not yet described as fitness factors in Francisella. The majority of the identified genes encoded proteins involved in the synthesis or maintenance of the cell envelope (LPS, outer membrane, capsule) or in the metabolism (glycolysis, gluconeogenesis, pentose phosphate pathway). Further 13C-flux analysis of the Tn5 glucokinase mutant strain revealed that the identified gene indeed encodes the sole active glucokinase in F-W12. In conclusion, candidate fitness factors of the new Francisella species F-W12 were identified using the scatter screen method which might also be usable for other Francisella species.


Assuntos
Acanthamoeba/microbiologia , Proteínas de Bactérias/genética , Francisella/fisiologia , Francisella/patogenicidade , Fatores de Virulência/genética , Elementos de DNA Transponíveis , Francisella/genética , Francisella/crescimento & desenvolvimento , Glucoquinase/genética , Interações Hospedeiro-Patógeno , Viabilidade Microbiana , Mutagênese Insercional , Mutação
5.
In Vitro Cell Dev Biol Anim ; 55(7): 522-532, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31264061

RESUMO

Emerging evidences exposed that long noncoding RNAs (lncRNAs) play important roles in various tumor progression including breast cancer (BC). However, the role of lncRNA ADP-dependent glucokinase antisense RNA 1 (ADPGK-AS1) in BC progression remains undiscovered. Hence, this study aimed to investigate the role of ADPGK-AS1 in BC. qRT-PCR was performed to investigate ADPGK-AS1 expression level in BC tissues and cell lines. The effect of ADPGK-AS1 knockdown on BC cellular process was assessed by loss-of-function assay. Luciferase reporter and RIP assay were performed to investigate the combination between ADPGK-AS1 and miR-3196. The combination between miR-3196 and orthodenticle homeobox 1 (OTX1) was verified by luciferase reporter assay. Finally, rescue assays were performed to confirm the effects of ADPGK-AS1/miR-3196/OTX1 axis on BC development. ADPGK-AS1 expression level was upregulated in BC tissues and cell lines. High expression of ADPGK-AS1 predicted poor prognosis for BC patients. Functionally, ADPGK-AS1 promoted cell proliferation, migration, induced epithelial-mesenchymal transition (EMT) process, and suppressed cell apoptosis. Mechanistically, ADPGK-AS1 acted as a miR-3196 sponge to release OTX1 in BC cells. Currently, ADPGK-AS1 acted as a competing endogenous RNA (ceRNA) via modulating miR-3196/OTX1 axis in BC.


Assuntos
Neoplasias da Mama/genética , Regulação Neoplásica da Expressão Gênica/genética , Glucoquinase/genética , MicroRNAs/genética , Fatores de Transcrição Otx/genética , RNA Longo não Codificante/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Feminino , Técnicas de Silenciamento de Genes , Humanos , Células MCF-7 , Pessoa de Meia-Idade , RNA Antissenso/genética
6.
Zhejiang Da Xue Xue Bao Yi Xue Ban ; 48(2): 200-203, 2019 Apr 25.
Artigo em Chinês | MEDLINE | ID: mdl-31309759

RESUMO

Maturity onset diabetes of the young (MODY) is a monogenic autosomal dominant inherited disease. Its clinical manifestations are asymptomatic with slightly elevated fasting blood glucose and few complications. This paper reports a novel mutation W257R in glucokinase (GCK) gene from a Chinese patient with MODY. Heterozygous mutation c.769T>C (p.W257R) in exon 7 of GCK gene (Chr744187343) was found in the proband, her father and brother. This W257R mutation was first reported in Chinese population.


Assuntos
Diabetes Mellitus Tipo 2 , Glucoquinase , Mutação , China , Diabetes Mellitus Tipo 2/genética , Feminino , Glucoquinase/genética , Humanos , Masculino , Linhagem
7.
BMC Complement Altern Med ; 19(1): 136, 2019 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-31215434

RESUMO

BACKGROUND: Tomato fruit (Lycopersicon esculentum Mill.) has been suggested to be useful for the prevention of diabetes. Esculeoside A is the main saponin compounds in tomatoes. This study investigated the hypoglycemic effects and the underlying mechanism of esculeoside A in C57BLKS/Leprdb (db/db) mice. METHODS: Wild-type C57BLKS (db/dm) mice were used in the db/dm mouse group and db/db mice were randomly divided into 2 groups: untreated and treated db/db mouse groups. Esculeoside A (100 mg/kg) was administered by gavage for 56 days to the treated db/db mouse group. Distilled water was administered to the db/dm mouse group and the untreated db/db mouse group. The blood and liver biochemical parameters and the expression of liver insulin signaling-related proteins were examined. RESULTS: The results showed that esculeoside A reduced the fasting blood glucose (FBG) levels and improved the glucose tolerance. Further investigation revealed that hepatic protein expressions of total AMP-activated protein kinase (T-AMPK), phosphorylated AMP-activated protein kinase (p-AMPK), insulin receptor substrate-1 (IRS-1), and glucokinase (GCK) were significantly upregulated after esculeoside A treatment. In contrast, the hepatic protein expression of phosphoenolpyruvate carboxykinase (PEPCK) was significantly downregulated by esculeoside A treatment. CONCLUSION: These findings suggested that esculeoside A has a potential of alleviating the metabolic abnormalities in db/db mice via regulation of AMPK/IRS-1 pathway. Our findings supported a possible application of esculeoside A as a functional supplement for diabetes treatment.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Diabetes Mellitus Tipo 2/tratamento farmacológico , Hipoglicemiantes/administração & dosagem , Proteínas Substratos do Receptor de Insulina/genética , Sapogeninas/administração & dosagem , Proteínas Quinases Ativadas por AMP/genética , Animais , Diabetes Mellitus Tipo 2/enzimologia , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Glucoquinase/genética , Glucoquinase/metabolismo , Humanos , Insulina/metabolismo , Proteínas Substratos do Receptor de Insulina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação/efeitos dos fármacos , Regulação para Cima
8.
Zhonghua Er Ke Za Zhi ; 57(6): 440-444, 2019 Jun 02.
Artigo em Chinês | MEDLINE | ID: mdl-31216801

RESUMO

Objective: To explore the gene mutation characteristics and detailed clinical presentations of hyperglycemia caused by GCK mutations in 10 patients. Methods: The clinical and follow-up data of 10 patients with hyperglycemia caused by mutation of GCK gene were reviewed. The patients were ascertained between January 1, 2014 and August 31, 2018 at the Department of Pediatrics, the First Affiliated Hospital of Zhejiang University and Ningbo Women & Children's Hospital. Clinical data were collected, including age, gender, main complaint, family history, fasting blood glucose, fasting blood insulin, 2-hour blood glucose, 2-hour blood insulin after oral glucose tolerance test, glycosylated hemoglobin, anti-glutamic acid decarboxylase antibody and body mass index. Mutations of GCK gene were detected by Sanger sequencing or high-throughput sequencing of diabetes-related genes in the patients and their family members. Results: There were ten patients, 8 of them were male, 2 were female.The ages at diagnosis varied between 4.7 to 12.3 years. The patients usually did not have obvious clinical symptoms of diabetes mellitus. Most of them were unexpectedly found to have hyperglycemia and with impaired glucose metabolism in three consecutive generations. The fasting blood glucose of patients was 6.8-7.7 mmol/L, 2-hour postprandial blood glucose was 7.8-11.6 mmol/L. Fasting blood insulin was 0.5-8.5 mU/L, glucose tolerance test results showed that 2 h postprondial blood insulin was 1.3-55.4 mU/L. The level of glycosylated hemoglobin was 6.1%-6.8%. Anti-glutamic acid decarboxylase antibody was negative in all patients. The GCK mutations identified in patients and one of their parents were located at exon5 (4 cases), exon9 (2 cases), exon2 (1 case), exon4 (1 case), exon6 (1 case) and exon7 (1 case). Conclusions: Most of the hyperglycemia patients caused by GCK mutations did not have typical clinical symptoms of diabetes. The fasting blood glucose was slightly elevated. Abnormal glucose tolerance test results were found in all 10 patients. Three consecutive generations of family had impaired glucose metabolism. GCK mutations located at exon 5 were common in 10 cases. There was no correlation between type of mutations and plasma glucose levels in domestic and international researches. When fasting glucose was found abnormal in clinic, a complete family history should be taken and the GCK gene should be sequenced to confirm the diagnosis in time.


Assuntos
Diabetes Mellitus Tipo 2/diagnóstico , Diabetes Mellitus Tipo 2/genética , Glucoquinase/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Hiperglicemia/genética , Glicemia , Criança , Pré-Escolar , Feminino , Marcadores Genéticos , Teste de Tolerância a Glucose , Humanos , Hiperglicemia/diagnóstico , Masculino , Mutação
9.
Bull Exp Biol Med ; 167(2): 263-266, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31243677

RESUMO

We studied the expression of genes encoding enzymes of carbohydrate and lipid metabolism ketohexokinase (Khk), glucokinase (Gck), pyruvate kinase (Pklr), acetyl-Co-carboxylase (Acaca), fatty acid synthase (Fasn), stearoyl-CoA desaturase (Scd), and their transcription regulators ChREBP (Mlxipl), SREBP-1c (Srebf1), and PPARα (Ppara) in rat liver. Control group rats received a semisynthetic ration over 20 weeks. Experimental group 1 received a semisynthetic ration and 20% fructose solution instead of drinking water. Experimental group 2 rats received a semisynthetic ration with quercetin (0.1% fodder weight) and 20% fructose solution. Consumption of 20% fructose solution (experimental group 1) led to an increase in Scd expression in comparison with the control and did not affect the expression of other genes. Addition of quercetin to the ration (experimental group 2) led to a decrease in the expression of Khk, Gck, Fasn, Scd, Mlxipl, and Ppara genes in comparison with experimental group 1. The results suggest that quercetin reduced the expression of genes of carbohydrate and lipid metabolism enzymes in the liver of rats receiving high-fructose ration.


Assuntos
Enzimas/genética , Frutose/metabolismo , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Fígado/efeitos dos fármacos , Fígado/enzimologia , Quercetina/farmacologia , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Metabolismo dos Carboidratos/efeitos dos fármacos , Metabolismo dos Carboidratos/genética , Dieta , Enzimas/metabolismo , Frutoquinases/genética , Frutoquinases/metabolismo , Glucoquinase/genética , Glucoquinase/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolismo dos Lipídeos/genética , Fígado/metabolismo , Masculino , Piruvato Quinase/genética , Piruvato Quinase/metabolismo , Ratos , Ratos Wistar , Estearoil-CoA Dessaturase/genética , Estearoil-CoA Dessaturase/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo
10.
Diabetes Res Clin Pract ; 151: 231-236, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31063852

RESUMO

We report on 134 unique GCK variants in 217 families, including 27 unpublished variants, identified in the US Monogenic Diabetes Registry in the last decade. Using ACMG guidelines, 26% were pathogenic, 56% likely pathogenic and 18% were of uncertain significance. Those with pathogenic variants had clinical features consistent with GCK-MODY.


Assuntos
Diabetes Mellitus Tipo 2/diagnóstico , Diabetes Mellitus Tipo 2/epidemiologia , Glucoquinase/genética , Adolescente , Adulto , Criança , Diabetes Mellitus Tipo 2/patologia , Feminino , Humanos , Masculino , Sistema de Registros , Estados Unidos , Adulto Jovem
11.
J Fish Biol ; 95(2): 393-400, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31017661

RESUMO

The partial cDNA sequences of eight reference genes (actb, tuba1, gapdh58, gapdh59, eef1a1, RNA 18 s, pabpc1, ube2I) were cloned from largemouth bass Micropterus salmoides. The expression levels of these eight genes were compared in the various tissues (eye, spleen, kidney, gill, muscle, brain, liver, heart, gut and gonad) of M. salmoides fed on forage fish. The results showed that the candidate genes exhibited tissue-specific expression to various degrees and the stability ranking order was eef1a1 > tuba1 > RNA 18 s > pabpc1 > ube2I > actb > gapdh58 > gapdh59 among tissue types. Four candidate genes eef1a1, tuba1, RNA 18 s and actb were used to analyse the stability in liver tissues of largemouth bass between the forage-fish group and the formulated-feed group. The candidate genes also showed some changes in expression levels in the livers, while eef1a1 and tuba1 had the most stable expression in livers of fish fed on alternative diets within 10 candidates. So eef1a1 and tuba1 were recommended as optimal reference gene in quantitative real-time PCR analysis to normalise the expression levels of target genes in tissues and lives of the M. salmoides fed on alternative diets. In livers, the expression levels of gck normalised by eef1a1 and tuba1 showed the significant up-regulation in formulated feed group (P < 0.05) than those in forage-fish group. While sex difference has no significant effects on the expression levels of gck in both groups.


Assuntos
Bass/genética , Dieta/veterinária , Animais , Bass/anatomia & histologia , Bass/fisiologia , DNA Complementar/biossíntese , DNA Complementar/química , Bases de Dados de Ácidos Nucleicos , Expressão Gênica , Instabilidade Genômica , Glucoquinase/genética , Fígado/metabolismo , RNA/análise , RNA/isolamento & purificação , RNA/normas , Reação em Cadeia da Polimerase em Tempo Real
12.
Am J Physiol Regul Integr Comp Physiol ; 316(3): R265-R273, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30649892

RESUMO

Metformin is an antidiabetic drug with a major impact on regulating blood glucose levels by decreasing hepatic gluconeogenesis, but also by affecting other pathways, including glucose transport and energy/lipid metabolism. Carnivorous fish are considered glucose intolerant, as they exhibit poor ability in using dietary carbohydrates. To increase the current knowledge about the molecular mechanisms by which metformin can improve glucose homeostasis in carnivorous fish, we addressed the effect of intraperitoneal administration of metformin, in the presence or absence of a glucose load, on metabolic rate-limiting enzymes and lipogenic factors in the liver of gilthead sea bream ( Sparus aurata). Hyperglycemia markedly upregulated the expression of glycolytic enzymes (glucokinase and 6-phosphofructo-1-kinase, PFK1) 5 h following glucose administration, while at 24 h posttreatment, it increased isocitrate dehydrogenase (IDH) activity, a key enzyme of the tricarboxylic acid cycle, and the expression of lipogenic factors (PGC1ß, Lpin1, and SREBP1). Metformin counteracted glucose-dependent effects, and downregulated glutamate dehydrogenase, alanine aminotransferase, and mammalian target of rapamycin 5 h posttreatment in the absence of a glucose load, leading to decreased long-term activity of PFK1 and IDH. The results of the present study suggest that hyperglycemia enhances lipogenesis in the liver of S. aurata and that metformin may exert specific metabolic effects in fish by decreasing hepatic transdeamination and suppressing the use of amino acids as gluconeogenic substrates. Our findings highlight the role of amino acid metabolism in the glucose-intolerant carnivorous fish model.


Assuntos
Desaminação/efeitos dos fármacos , Glucose/metabolismo , Hipoglicemiantes/farmacologia , Lipogênese/efeitos dos fármacos , Fígado/metabolismo , Metformina/farmacologia , Dourada/metabolismo , Aminoácidos/metabolismo , Animais , Desaminação/genética , Glucoquinase/genética , Glucoquinase/metabolismo , Gluconeogênese/efeitos dos fármacos , Glucose/farmacologia , Hiperglicemia/metabolismo , Lipogênese/genética , Fígado/efeitos dos fármacos , Fosfofrutoquinase-2/metabolismo
13.
Acta Diabetol ; 56(4): 413-420, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30663027

RESUMO

AIMS: Diabetes in pregnancy may be associated with monogenic defects of beta-cell function, frequency of which depends on ethnicity, clinical criteria for selection of patients as well as methods used for genetic analysis. The aim was to evaluate the contribution and molecular spectrum of mutations among genes associated with monogenic diabetes in non-obese Russian patients with diabetes in pregnancy using the next-generation sequencing (NGS). METHODS: 188 non-obese pregnant women with diabetes during pregnancy were included in the study; among them 57 subjects (30.3%) met the American Diabetes Association (ADA) criteria of preexisting pregestational diabetes (pre-GDM), whereas 131 women (69.7%) fulfilled criteria of gestational diabetes mellitus (GDM). A custom NGS panel targeting 28 diabetes causative genes was used for sequencing. The sequence variants were rated according to the American College of Medical Genetics and Genomics (ACMG) guidelines. RESULTS: In total, 23 pathogenic, 18 likely pathogenic and 16 variants of uncertain significance were identified in 59/188 patients (31.4%). The majority of variants (38/59) were found in GCK gene. No significant differences in the number of variants among the two study groups (pre-GDM and GDM) were observed. CONCLUSIONS: The study suggests that frequency of monogenic variants of diabetes might be underestimated, which warrants a broader use of genetic testing, especially in pregnancy.


Assuntos
Diabetes Mellitus Tipo 2/epidemiologia , Diabetes Mellitus Tipo 2/genética , Diabetes Gestacional/epidemiologia , Diabetes Gestacional/genética , Polimorfismo Genético , Gravidez em Diabéticas/epidemiologia , Gravidez em Diabéticas/genética , Adulto , Análise Mutacional de DNA/métodos , Diabetes Mellitus Tipo 2/complicações , Feminino , Frequência do Gene , Testes Genéticos , Glucoquinase/genética , Intolerância à Glucose/epidemiologia , Intolerância à Glucose/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Mutação , Estado Pré-Diabético/complicações , Estado Pré-Diabético/epidemiologia , Estado Pré-Diabético/genética , Gravidez , Complicações na Gravidez/epidemiologia , Complicações na Gravidez/genética , Federação Russa/epidemiologia
14.
Genet Med ; 21(4): 939-947, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30245511

RESUMO

PURPOSE: To estimate the prevalence of glucokinase variant-induced diabetes (GCK-DM) in a general population and to establish a clinical strategy for identifying GCK-DM from type 2 diabetes (T2DM). METHODS: A population-based study of diabetes in a rural region of Beijing, China, was conducted using two-stage stratified random cluster sampling. The glucokinase exons were sequenced in patients with diabetes. RESULTS: A total of 3345 subjects, including 545 patients with diabetes, participated in this study. Seven patients with GCK-DM were identified. The estimated prevalence rates of GCK-DM were 0.21% and 1.3% in the whole population and the diabetic patients, respectively. In the newly diagnosed diabetic patients (New-DM), a triglyceride cutoff ≤1.43 mmol/L (126.55 mg/dl) could discriminate GCK-DM from T2DM with 100% sensitivity and 68.4% specificity. Its effectiveness was confirmed in an additional 134 early-onset young patients with T2DM and mild hyperglycemia. A clinical criterion based on triglyceride and mild hyperglycemia could differentiate GCK-DM from T2DM in New-DM and was shown to be effective in identifying GCK-DM from 559 early-onset young patients with T2DM in the hospital. CONCLUSIONS: The prevalence of GCK-DM is approximately 1.3% in the Chinese population with diabetes, and the new clinical screening strategy is helpful for identifying GCK-DM.


Assuntos
Diabetes Mellitus Tipo 2/genética , Predisposição Genética para Doença , Glucoquinase/genética , Adulto , Idade de Início , Glicemia , China , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/patologia , Feminino , Humanos , Masculino , Mutação , Fenótipo
15.
Diabet Med ; 36(2): 252-255, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30362177

RESUMO

BACKGROUND: Glucokinase-maturity-onset diabetes of the young (GCK-MODY) is a form of diabetes caused by heterozygous inactivating mutations in the GCK gene. Affected individuals maintain their fasting glucose levels at a higher set point (5.4-8.3 mmol/l) than the general population. Hyperglycaemia in women with Type 1 or Type 2 diabetes is known to confer increased risk of fetal congenital abnormalities. The association between GCK-MODY and congenital abnormalities, however, remains uncertain. CASE REPORT: A 35-year-old woman in her third pregnancy was diagnosed with gestational diabetes at 13 weeks' gestation (fasting blood glucose 6.0 mmol/L, 1-h blood glucose 9.2 mmol/l, 2-h blood glucose 7.3 mmol/l). The morphology scan at 19+2 weeks' gestation showed a Type III sacral agenesis. The woman elected to terminate the pregnancy. Her postpartum oral glucose tolerance test was suggestive of GCK-MODY (fasting blood glucose 7.4 mmol/l, 1-h blood glucose 9.3 mmol/l, 2-h blood glucose 7.3 mmol/l). Mutation analysis of the GCK gene identified a novel heterozygous GCK missense mutation, p.V199M, classified as likely pathogenic, providing molecular confirmation of the suspected GCK-MODY diagnosis. DISCUSSION: Sacral agenesis is a rare form of sacral abnormality affecting 0.005% to 0.1% of pregnancies. It is a subtype of the caudal regression sequence, a cardinal feature of diabetic embryopathy. This case raises the question as to whether hyperglycaemia in GCK-MODY may increase the risk of fetal caudal regression syndrome as reported in women with pre-existing diabetes mellitus. Improved diagnostic rates of GCK-MODY, and MODY registers that include pregnancy outcomes, are important to further elucidate risk of congenital abnormalities in GCK-MODY.


Assuntos
Diabetes Mellitus Tipo 2/genética , Feto/anormalidades , Glucoquinase/genética , Mutação de Sentido Incorreto/genética , Gravidez em Diabéticas/genética , Sacro/anormalidades , Adulto , Feminino , Heterozigoto , Humanos , Hiperglicemia/complicações , Gravidez , Fatores de Risco
16.
J Assist Reprod Genet ; 36(2): 255-266, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30284103

RESUMO

PURPOSE: Motility of spermatozoa helps not only in planning the type of infertility treatment but also directly reflects the success rate in assisted reproductive technology (ART). Previously, biotin, a water-soluble vitamin, has been shown to increase the motility and longevity of cryopreserved human spermatozoa. The present study was designed to understand the molecular basis of the beneficial effects of presence of biotin in sperm wash medium on early embryo development. METHODS: The effect biotin supplementation to sperm wash medium on the sperm parameters were assessed in swim-up fraction of normozoospermic and asthenozoospermic ejaculates collected from infertile men. Fertilization and early embryo development was studied using Swiss albino mice. RESULTS: Even though both biotin and pentoxifylline (PTX) enhanced the motility of spermatozoa from normozoospermic and asthenozoospermic samples, biotin group exhibited higher in vitro survival. Using mouse model, we observed that presence of biotin or PTX in sperm wash medium improved the fertilization rate and blastocyst rate compared to control. Blastocysts from these groups had significantly higher total cell number (P < 0.01) and lower apoptotic index. In silico target prediction revealed that GTPase HRas (HRas), tyrosine-protein phosphatase nonreceptor type 1 (PTP1B), and glucokinase are the probable targets for biotin. Solution-state Nuclear Magnetic Resonance (NMR) studies confirmed that biotin interacts both with human HRas and PTP1B. CONCLUSION: Our results indicate that presence of biotin in sperm wash medium can improve the fertilization potential and preimplantation embryo development and can be considered as a safe alternate to PTX.


Assuntos
Astenozoospermia/tratamento farmacológico , Meios de Cultura/química , Desenvolvimento Embrionário/efeitos dos fármacos , Espermatozoides/crescimento & desenvolvimento , Animais , Astenozoospermia/patologia , Biotina/farmacologia , Blastocisto/efeitos dos fármacos , Criopreservação , Feminino , Fertilização/efeitos dos fármacos , Fertilização In Vitro/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Glucoquinase/genética , Humanos , Masculino , Camundongos , Pentoxifilina/farmacologia , Gravidez , Proteína Tirosina Fosfatase não Receptora Tipo 1/genética , Motilidade Espermática/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos
17.
Biochim Biophys Acta Mol Basis Dis ; 1865(2): 428-433, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30465894

RESUMO

Monogenic diabetes is caused by mutations that reduce ß-cell function. While Sanger sequencing is the standard method used to detect mutated genes. Next-generation sequencing techniques, such as whole exome sequencing (WES), can be used to find multiple gene mutations in one assay. We used WES to detect genetic mutations in both permanent neonatal (PND) and type 1B diabetes (T1BD). A total of five PND and nine T1BD patients were enrolled in this study. WES variants were assessed using VarioWatch, excluding those identified previously. Sanger sequencing was used to confirm the mutations, and their pathogenicity was established via the literature or bioinformatic/functional analysis. The PND and T1BD patients were diagnosed at 0.1-0.5 and 0.8-2.7 years of age, respectively. Diabetic ketoacidosis was present at diagnosis in 60% of PND patients and 44.4% of T1BD patients. We found five novel mutations in five different genes. Notably, patient 602 had a novel homozygous missense mutation c.1295C > A (T432 K) in the glucokinase (GCK) gene. Compared to the wild-type recombinant protein, the mutant protein had significantly lower enzymatic activity (2.5%, p = 0.0002) and Vmax (1.23 ±â€¯0.019 vs. 0.33 ±â€¯0.016, respectively; p = 0.005). WES is a robust technique that can be used to unravel the etiologies of genetically heterogeneous forms of diabetes. Homozygous inactivating mutations of the GCK gene may have a significant role in PND pathogenesis.


Assuntos
Diabetes Mellitus Tipo 1/enzimologia , Diabetes Mellitus Tipo 1/genética , Glucoquinase/genética , Glucoquinase/metabolismo , Mutação/genética , Sequenciamento Completo do Exoma , Feminino , Humanos , Lactente , Recém-Nascido , Cinética , Masculino
18.
Endocrinology ; 160(1): 205-219, 2019 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-30445425

RESUMO

The increased hepatic gluconeogenesis in type 2 diabetes mellitus has often been ascribed to increased transcription of phosphoenolpyruvate carboxykinase 1, cystolic form (PEPCK1), although recent evidence has questioned this attribution. To assess the metabolic role of PEPCK1, we treated regular chow fed and high-fat fed (HFF) male Sprague-Dawley rats with a 2'-O-methoxyethyl chimeric antisense oligonucleotide (ASO) against PEPCK1 and compared them with control ASO-treated rats. PEPCK1 ASO effectively decreased PEPCK1 expression in the liver and white adipose tissue. In chow fed rats, PEPCK1 ASO did not alter adiposity, plasma glucose, or insulin. In contrast, PEPCK1 ASO decreased the white adipose tissue mass in HFF rats but without altering basal rates of lipolysis, de novo lipogenesis, or glyceroneogenesis in vivo. Despite the protection from adiposity, hepatic insulin sensitivity was impaired in HFF PEPCK1 ASO-treated rats. PEPCK1 ASO worsened hepatic steatosis, although without additional impairments in hepatic insulin signaling or activation of inflammatory signals in the liver. Instead, the development of hepatic insulin resistance and the decrease in hepatic glycogen synthesis during a hyperglycemic clamp was attributed to a decrease in hepatic glucokinase (GCK) expression and decreased synthesis of glycogen via the direct pathway. The decrease in GCK expression was associated with increased expression of activating transcription factor 3, a negative regulator of GCK transcription. These studies have demonstrated that PEPCK1 is integral to coordinating cellular metabolism in the liver and adipose tissue, although it does not directly effect hepatic glucose production or adipose glyceroneogenesis.


Assuntos
Adiposidade , Diabetes Mellitus Tipo 2/enzimologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Glicogênio Hepático/biossíntese , Fígado/metabolismo , Oligonucleotídeos Antissenso/genética , Fosfoenolpiruvato Carboxiquinase (GTP)/genética , Tecido Adiposo Branco/metabolismo , Animais , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/fisiopatologia , Dieta Hiperlipídica/efeitos adversos , Glucoquinase/genética , Glucoquinase/metabolismo , Humanos , Insulina/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Lipogênese , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fosfoenolpiruvato Carboxiquinase (GTP)/metabolismo , Ratos , Ratos Sprague-Dawley
19.
Diabetes ; 68(3): 654-664, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30523024

RESUMO

We recently reported that in rodent models of type 2 diabetes (T2D), a single intracerebroventricular (icv) injection of fibroblast growth factor 1 (FGF1) induces remission of hyperglycemia that is sustained for weeks. To clarify the peripheral mechanisms underlying this effect, we used the Zucker diabetic fatty fa/fa rat model of T2D, which, like human T2D, is characterized by progressive deterioration of pancreatic ß-cell function after hyperglycemia onset. We report that although icv FGF1 injection delays the onset of ß-cell dysfunction in these animals, it has no effect on either glucose-induced insulin secretion or insulin sensitivity. These observations suggest that FGF1 acts in the brain to stimulate insulin-independent glucose clearance. On the basis of our finding that icv FGF1 treatment increases hepatic glucokinase gene expression, we considered the possibility that increased hepatic glucose uptake (HGU) contributes to the insulin-independent glucose-lowering effect of icv FGF1. Consistent with this possibility, we report that icv FGF1 injection increases liver glucokinase activity by approximately twofold. We conclude that sustained remission of hyperglycemia induced by the central action of FGF1 involves both preservation of ß-cell function and stimulation of HGU through increased hepatic glucokinase activity.


Assuntos
Fator 1 de Crescimento de Fibroblastos/uso terapêutico , Animais , Glicemia/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Glucoquinase/genética , Glucoquinase/metabolismo , Teste de Tolerância a Glucose , Humanos , Hipoglicemiantes/uso terapêutico , Insulina/metabolismo , Resistência à Insulina , Masculino , Ratos , Ratos Zucker , Reação em Cadeia da Polimerase em Tempo Real
20.
Acta Diabetol ; 56(4): 405-411, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30535721

RESUMO

AIMS: GCK-MODY is characterized by mild hyperglycemia. Treatment is not required outside of pregnancy. During pregnancy, insulin treatment is recommended if second trimester fetal ultrasound monitoring shows macrosomia, suggesting the fetus has not inherited the GCK gene. There are limited data about GCK-MODY management in pregnancy. The aim of this study was to examine clinical management and pregnancy outcomes amongst women with a known diagnosis of GCK-MODY. METHODS: In this observational, cross-sectional study, a survey was distributed via Redcap to women ≥ 18 years enrolled in the University of Chicago Monogenic Diabetes Registry (n = 94). All or part of the survey was completed by 54 women (128 pregnancies). RESULTS: There were 78 term births (61%), 15 pre-term births (12%), and 24 miscarriages (19%). Of the 39 pregnancies where insulin was given, 22 (56%) had occasional or frequent hypoglycemia including 9 with severe hypoglycemia. Average birth weight for full-term GCK-affected infants was significantly less in cases of maternal insulin treatment versus no treatment (2967 and 3725 g, p = 0.005). For GCK-unaffected infants, conclusions are limited by small sample size but large for gestational age (LGA) was common with maternal insulin treatment (56%) and no treatment (33%), p = 0.590. CONCLUSIONS: The observed miscarriage rate was comparable to the background US population rate (15-20%). Patients treated with insulin experienced a 23% incidence of severe hypoglycemia and lower birth weights were observed in the insulin-treated, GCK-affected neonates. These data support published guidelines of no treatment if the fetus is suspected to have inherited GCK-MODY and highlight the importance of additional studies to determine optimal pregnancy management for GCK-MODY, particularly among unaffected fetuses.


Assuntos
Diabetes Mellitus Tipo 2/epidemiologia , Diabetes Mellitus Tipo 2/terapia , Glucoquinase/genética , Resultado da Gravidez/epidemiologia , Gravidez em Diabéticas/epidemiologia , Gravidez em Diabéticas/terapia , Aborto Espontâneo/epidemiologia , Adolescente , Adulto , Peso ao Nascer/efeitos dos fármacos , Estudos Transversais , Diabetes Mellitus Tipo 2/genética , Feminino , Humanos , Hiperglicemia/epidemiologia , Hiperglicemia/genética , Hiperglicemia/terapia , Recém-Nascido , Insulina/uso terapêutico , Mutação , Gravidez , Gravidez em Diabéticas/genética , Sistema de Registros , Resultado do Tratamento , Adulto Jovem
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