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1.
Am J Hum Genet ; 110(10): 1769-1786, 2023 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-37729906

RESUMO

Defects in hydroxymethylbilane synthase (HMBS) can cause acute intermittent porphyria (AIP), an acute neurological disease. Although sequencing-based diagnosis can be definitive, ∼⅓ of clinical HMBS variants are missense variants, and most clinically reported HMBS missense variants are designated as "variants of uncertain significance" (VUSs). Using saturation mutagenesis, en masse selection, and sequencing, we applied a multiplexed validated assay to both the erythroid-specific and ubiquitous isoforms of HMBS, obtaining confident functional impact scores for >84% of all possible amino acid substitutions. The resulting variant effect maps generally agreed with biochemical expectations and provide further evidence that HMBS can function as a monomer. Additionally, the maps implicated specific residues as having roles in active site dynamics, which was further supported by molecular dynamics simulations. Most importantly, these maps can help discriminate pathogenic from benign HMBS variants, proactively providing evidence even for yet-to-be-observed clinical missense variants.


Assuntos
Hidroximetilbilano Sintase , Porfiria Aguda Intermitente , Humanos , Hidroximetilbilano Sintase/química , Hidroximetilbilano Sintase/genética , Hidroximetilbilano Sintase/metabolismo , Mutação de Sentido Incorreto/genética , Porfiria Aguda Intermitente/diagnóstico , Porfiria Aguda Intermitente/genética , Substituição de Aminoácidos , Simulação de Dinâmica Molecular
2.
Int J Sport Nutr Exerc Metab ; 30(6): 386-395, 2020 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-32998111

RESUMO

Fasting enhances the beneficial metabolic outcomes of exercise; however, it is unknown whether body composition is favorably modified on the short term. A baseline-follow-up study was carried out to assess the effect of an established protocol involving short-term combined exercise with fasting on body composition. One hundred seven recreationally exercising males underwent a 10-day intervention across 15 fitness centers in the Netherlands involving a 3-day gradual decrease of food intake, a 3-day period with extremely low caloric intake, and a gradual 4-day increase to initial caloric intake, with daily 30-min submaximal cycling. Using dual-energy X-ray absorptiometry analysis, all subjects substantially lost total body mass (-3.9 ± 1.9 kg; p < .001) and fat mass (-3.3 ± 1.3 kg; p < .001). Average lean mass was lost (-0.6 ± 1.5 kg; p < .001), but lean mass as a percentage of total body mass was not reduced. The authors observed a loss of -3.9 ± 1.9% android fat over total fat mass (p < .001), a loss of -2.2 ± 1.9% gynoid over total fat mass (p < .001), and reduced android/gynoid ratios (-0.05 ± 0.1; p < .001). Analyzing 15 preselected single-nucleotide polymorphisms in 13 metabolism-related genes revealed trending associations for thyroid state-related single-nucleotide polymorphisms rs225014 (deiodinase 2) and rs35767 (insulin-like growth factor1), and rs1053049 (PPARD). In conclusion, a short period of combined fasting and exercise leads to a substantial loss of body and fat mass without a loss of lean mass as a percentage of total mass.


Assuntos
Composição Corporal , Exercício Físico , Jejum , Absorciometria de Fóton , Adulto , Ingestão de Energia , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Países Baixos , Polimorfismo de Nucleotídeo Único , Adulto Jovem
3.
Genet Med ; 21(11): 2605-2613, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31073229

RESUMO

With the advent of precision and genomic medicine, a critical issue is whether a disease gene variant is pathogenic or benign. Such is the case for the three autosomal dominant acute hepatic porphyrias (AHPs), including acute intermittent porphyria, hereditary coproporphyria, and variegate porphyria, each resulting from the half-normal enzymatic activities of hydroxymethylbilane synthase, coproporphyrinogen oxidase, and protoporphyrinogen oxidase, respectively. To date, there is no public database that documents the likely pathogenicity of variants causing the porphyrias, and more specifically, the AHPs with biochemically and clinically verified information. Therefore, an international collaborative with the European Porphyria Network and the National Institutes of Health/National Center for Advancing Translational Sciences/National Institute of Diabetes and Digestive and Kidney Diseases (NIH/NCATS/NIDDK)-sponsored Porphyrias Consortium of porphyria diagnostic experts is establishing an online database that will collate biochemical and clinical evidence verifying the pathogenicity of the published and newly identified variants in the AHP-causing genes. The overall goal of the International Porphyria Molecular Diagnostic Collaborative is to determine the pathogenic and benign variants for all eight porphyrias. Here we describe the overall objectives and the initial efforts to validate pathogenic and benign variants in the respective heme biosynthetic genes causing the AHPs.


Assuntos
Porfirias/genética , Porfirias/fisiopatologia , Virulência/genética , Curadoria de Dados/métodos , Bases de Dados Factuais , Feminino , Humanos , Masculino , Patologia Molecular , Sintase do Porfobilinogênio/deficiência , Sintase do Porfobilinogênio/genética , Porfiria Aguda Intermitente/genética , Porfiria Aguda Intermitente/fisiopatologia , Porfirias Hepáticas/genética , Porfirias Hepáticas/fisiopatologia , Estados Unidos
4.
J Card Surg ; 34(4): 226-228, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30847947

RESUMO

We report implantation of a left ventricular assist device (LVAD) in a patient with Parkinson's disease. Postoperative fluid overload together with insufficient LVAD output in the setting of vasodilation through levodopa likely caused renal hypoperfusion and acute kidney injury. A patient like ours, therefore, requires the highest possible increase of HM3 RPM and LVAD flow early after surgery.


Assuntos
Insuficiência Cardíaca/cirurgia , Ventrículos do Coração/cirurgia , Coração Auxiliar , Levodopa/efeitos adversos , Doença de Parkinson/tratamento farmacológico , Vasodilatação/efeitos dos fármacos , Injúria Renal Aguda/etiologia , Injúria Renal Aguda/prevenção & controle , Insuficiência Cardíaca/complicações , Coração Auxiliar/efeitos adversos , Humanos , Isquemia/etiologia , Isquemia/prevenção & controle , Rim/irrigação sanguínea , Levodopa/administração & dosagem , Masculino , Pessoa de Meia-Idade , Doença de Parkinson/complicações , Complicações Pós-Operatórias/etiologia , Complicações Pós-Operatórias/prevenção & controle
5.
Diabetologia ; 61(11): 2386-2397, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30145664

RESUMO

AIMS/HYPOTHESIS: The aim of this study was to evaluate the effect of sitagliptin on glucose tolerance, plasma lipids, energy expenditure and metabolism of brown adipose tissue (BAT), white adipose tissue (WAT) and skeletal muscle in overweight individuals with prediabetes (impaired glucose tolerance and/or impaired fasting glucose). METHODS: We performed a randomised, double-blinded, placebo-controlled trial in 30 overweight, Europid men (age 45.9 ± 6.2 years; BMI 28.8 ± 2.3 kg/m2) with prediabetes in the Leiden University Medical Center and the Alrijne Hospital between March 2015 and September 2016. Participants were initially randomly allocated to receive sitagliptin (100 mg/day) (n = 15) or placebo (n = 15) for 12 weeks, using a randomisation list that was set up by an unblinded pharmacist. All people involved in the study as well as participants were blinded to group assignment. Two participants withdrew from the study prior to completion (both in the sitagliptin group) and were subsequently replaced with two new participants that were allocated to the same treatment. Before and after treatment, fasting venous blood samples and skeletal muscle biopsies were obtained, OGTT was performed and body composition, resting energy expenditure and [18F] fluorodeoxyglucose ([18F]FDG) uptake by metabolic tissues were assessed. The primary study endpoint was the effect of sitagliptin on BAT volume and activity. RESULTS: One participant from the sitagliptin group was excluded from analysis, due to a distribution error, leaving 29 participants for further analysis. Sitagliptin, but not placebo, lowered glucose excursion (-40%; p < 0.003) during OGTT, accompanied by an improved insulinogenic index (+38%; p < 0.003) and oral disposition index (+44%; p < 0.003). In addition, sitagliptin lowered serum concentrations of triacylglycerol (-29%) and very large (-46%), large (-35%) and medium-sized (-24%) VLDL particles (all p < 0.05). Body weight, body composition and energy expenditure did not change. In skeletal muscle, sitagliptin increased mRNA expression of PGC1ß (also known as PPARGC1B) (+117%; p < 0.05), a main controller of mitochondrial oxidative energy metabolism. Although the primary endpoint of change in BAT volume and activity was not met, sitagliptin increased [18F] FDG uptake in subcutaneous WAT (sWAT; +53%; p < 0.05). Reported side effects were mild and transient and not necessarily related to the treatment. CONCLUSIONS/INTERPRETATION: Twelve weeks of sitagliptin in overweight, Europid men with prediabetes improves glucose tolerance and lipid metabolism, as related to increased [18F] FDG uptake by sWAT, rather than BAT, and upregulation of the mitochondrial gene PGC1ß in skeletal muscle. Studies on the effect of sitagliptin on preventing or delaying the progression of prediabetes into type 2 diabetes are warranted. TRIAL REGISTRATION: ClinicalTrials.gov NCT02294084. FUNDING: This study was funded by Merck Sharp & Dohme Corp, Dutch Heart Foundation, Dutch Diabetes Research Foundation, Ministry of Economic Affairs and the University of Granada.


Assuntos
Tecido Adiposo Marrom/efeitos dos fármacos , Tecido Adiposo Marrom/metabolismo , Inibidores da Dipeptidil Peptidase IV/uso terapêutico , Sobrepeso/tratamento farmacológico , Sobrepeso/metabolismo , Estado Pré-Diabético/tratamento farmacológico , Fosfato de Sitagliptina/uso terapêutico , Tecido Adiposo Branco/efeitos dos fármacos , Tecido Adiposo Branco/metabolismo , Adulto , Glicemia/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Proteínas de Transporte/genética , Método Duplo-Cego , Metabolismo Energético/efeitos dos fármacos , Humanos , Masculino , Pessoa de Meia-Idade , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Estado Pré-Diabético/metabolismo , Proteínas de Ligação a RNA
6.
FASEB J ; 31(1): 14-28, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27729415

RESUMO

Obesity and type 2 diabetes are associated disorders that involve a multiplicity of tissues. Both fasting and physical exercise are known to counteract dyslipidemia/hyperglycemia. Skeletal muscle plays a key role in the control of blood glucose levels, and the metabolic changes and related signaling pathways in skeletal muscle induced by fasting overlap with those induced by exercise. The reduction of fat disposal has been shown to extend to the liver and to white and brown adipose tissue and to involve an increase in their metabolic activities. In recent years signal transduction pathways related to exercise and fasting/food withdrawal in muscle have been intensively studied, both in animals and in humans. Combining fasting/food withdrawal with exercise in animals as well as in humans causes changes unlike those seen during fasting/food withdrawal or exercise alone, which favor repair of muscle over autophagy. In addition, compounds that mimic exercise have been studied in combination with exercise or fasting/food withdrawal. This review addresses our current knowledge of the mechanisms that underlie the individual and combined effects of fasting/food withdrawal, endurance or resistance exercise, and their mimetics, in muscle vs other organs in rodents and humans, and highlights which combinations may improve metabolic disorders.-Jaspers, R. T., Zillikens, M. C., Friesema, E. C. H., delli Paoli, G., Bloch, W., Uitterlinden, A. G., Goglia, F., Lanni, A., de Lange, P. Exercise, fasting, and mimetics: toward beneficial combinations.


Assuntos
Exercício Físico/fisiologia , Privação de Alimentos/fisiologia , Animais , Glicemia , Diabetes Mellitus Tipo 2/metabolismo , Humanos , Lipídeos/sangue , Obesidade/metabolismo
8.
J Inherit Metab Dis ; 41(5): 809-817, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29675607

RESUMO

INTRODUCTION: A small proportion of patients with acute intermittent porphyria (AIP) suffer from recurrent porphyric attacks, with a severely diminished quality of life. In this retrospective case-control study, the burden of disease is quantified and compared among three AIP patient subgroups: cases with recurrent attacks, cases with one or occasional attacks and asymptomatic carriers. METHODS: Data from patient records and questionnaires were collected in patients between 1960 and 2016 at the Erasmus Medical Center, Rotterdam, the Netherlands. We collected symptoms related to porphyria, porphyria related complications, attack frequency, hospitalisation frequency, hospitalisation days related to acute porphyric attacks, frequency of heme infusions and medical healthcare costs based on hospitalisations and heme therapy. RESULTS: In total 11 recurrent AIP cases, 24 symptomatic AIP cases and 53 AIP carriers as controls were included. All recurrent patients reported porphyria related symptoms, such as pain, neurological and/or psychiatric disorders, and nearly all developed complications, such as hypertension and chronic kidney disease. In the recurrent cases group, the median lifelong number of hospitalisation days related to porphyric attacks was 82 days per patient (range 10-374), and they spent a median of 346 days (range 34-945) at a day-care facility for prophylactic heme therapy; total follow-up time was 243 person-years (PYRS). In the symptomatic non-recurrent group the median lifelong number of hospitalisation days related to porphyric attacks was 7 days per patient (range 1-78), total follow-up time was 528 PYRS. The calculated total medical healthcare cost for recurrent cases group was €5.8 million versus €0.3 million for the symptomatic cases group.


Assuntos
Heme/uso terapêutico , Porfiria Aguda Intermitente/complicações , Porfiria Aguda Intermitente/tratamento farmacológico , Porfiria Aguda Intermitente/economia , Adolescente , Adulto , Carcinoma Hepatocelular/etiologia , Estudos de Casos e Controles , Efeitos Psicossociais da Doença , Feminino , Custos de Cuidados de Saúde , Hospitalização/economia , Humanos , Hipertensão/etiologia , Neoplasias Hepáticas/etiologia , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Países Baixos , Qualidade de Vida , Insuficiência Renal Crônica/etiologia , Estudos Retrospectivos , Inquéritos e Questionários , Adulto Jovem
9.
Clin Sci (Lond) ; 130(14): 1209-20, 2016 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-27129187

RESUMO

ARNI [dual AT1 (angiotensin II type 1) receptor-neprilysin inhibition] exerts beneficial effects on blood pressure and kidney function in heart failure, compared with ARB (AT1 receptor blockade) alone. We hypothesized that ARNI improves cardiac and kidney parameters in diabetic TGR(mREN2)27 rats, an angiotensin II-dependent hypertension model. Rats were made diabetic with streptozotocin for 5 or 12 weeks. In the final 3 weeks, rats were treated with vehicle, irbesartan (ARB) or irbesartan+thiorphan (ARNI). Blood pressure, measured by telemetry in the 5-week group, was lowered identically by ARB and ARNI. The heart weight/tibia length ratio in 12-week diabetic animals was lower after ARNI compared with after ARB. Proteinuria and albuminuria were observed from 8 weeks of diabetes onwards. ARNI reduced proteinuria more strongly than ARB, and a similar trend was seen for albuminuria. Kidneys of ARNI-treated animals showed less severe segmental glomerulosclerosis than those of ARB-treated animals. After 12 weeks, no differences between ARNI- and ARB-treated animals were found regarding diuresis, natriuresis, plasma endothelin-1, vascular reactivity (acetylcholine response) or kidney sodium transporters. Only ARNI-treated rats displayed endothelin type B receptor-mediated vasodilation. In conclusion, ARNI reduces proteinuria, glomerulosclerosis and heart weight in diabetic TGR(mREN2)27 rats more strongly than does ARB, and this occurs independently of blood pressure.


Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/uso terapêutico , Pressão Sanguínea/fisiologia , Diabetes Mellitus Experimental/tratamento farmacológico , Nefropatias Diabéticas/prevenção & controle , Neprilisina/antagonistas & inibidores , Aminobutiratos/uso terapêutico , Animais , Fator Natriurético Atrial/sangue , Compostos de Bifenilo , Pressão Sanguínea/efeitos dos fármacos , Combinação de Medicamentos , Ratos , Ratos Sprague-Dawley , Estreptozocina , Tetrazóis/uso terapêutico , Valsartana
10.
Kidney Int ; 88(1): 109-20, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25830765

RESUMO

Neprilysin inhibitors prevent the breakdown of bradykinin and natriuretic peptides, promoting vasodilation and natriuresis. However, they also increase angiotensin II and endothelin-1. Here we studied the effects of a low and a high dose of the neprilysin inhibitor thiorphan on top of AT1 receptor blockade with irbesartan versus vehicle in TGR(mREN2)27 rats with high renin hypertension. Mean arterial blood pressure was unaffected by vehicle or thiorphan alone. Irbesartan lowered blood pressure, but after 7 days pressure started to increase again. Low- but not high-dose thiorphan prevented this rise. Only during exposure to low-dose thiorphan plus irbesartan did heart weight/body weight ratio, cardiac atrial natriuretic peptide expression, and myocyte size decrease significantly. Circulating endothelin-1 was not affected by low-dose thiorphan with or without irbesartan, but increased after treatment with high-dose thiorphan plus irbesartan. This endothelin-1 rise was accompanied by an increase in renal sodium-hydrogen exchanger 3 protein abundance, and an upregulation of constrictor vascular endothelin type B receptors. Consequently, the endothelin type B receptor antagonist BQ788 no longer enhanced endothelin-1-induced vasoconstriction (indicative of endothelin type B receptor-mediated vasodilation), but prevented it. Thus, optimal neprilysin inhibitor dosing reveals additional cardioprotective effects on top of AT1 receptor blockade in renin-dependent hypertension.


Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Compostos de Bifenilo/farmacologia , Rim/metabolismo , Miocárdio/patologia , Neprilisina/antagonistas & inibidores , Inibidores de Proteases/farmacologia , Receptor Tipo 1 de Angiotensina/efeitos dos fármacos , Tetrazóis/farmacologia , Tiorfano/farmacologia , Animais , Pressão Arterial/efeitos dos fármacos , Fator Natriurético Atrial/metabolismo , Peso Corporal , Antagonistas do Receptor de Endotelina B/farmacologia , Endotelina-1/sangue , Irbesartana , Rim/patologia , Miócitos Cardíacos/patologia , Oligopeptídeos/farmacologia , Tamanho do Órgão , Piperidinas/farmacologia , Inibidores de Proteases/administração & dosagem , Ratos , Receptor de Endotelina B/metabolismo , Sistema Renina-Angiotensina/efeitos dos fármacos , Trocador 3 de Sódio-Hidrogênio , Trocadores de Sódio-Hidrogênio/metabolismo , Tiorfano/administração & dosagem , Regulação para Cima , Vasoconstrição/efeitos dos fármacos , Vasodilatação/efeitos dos fármacos
11.
Hum Mutat ; 34(7): 1018-25, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23568789

RESUMO

SLC 16A2, the gene for the second member of the solute carrier family 16 (monocarboxylic acid transporter), located on chromosome Xq13.2, encodes a very efficient thyroid hormone transporter: monocarboxylate transporter 8, MCT8. Its loss of function is responsible in males for a continuum of psychomotor retardation ranging from severe (no motor acquisition, no speech) to mild (ability to walk with help and a few words of speech). Triiodothyronine uptake measurement in transfected cells and, more recently, patient fibroblasts, has been described to study the functional consequences of MCT8 mutations. Here, we describe three novel MCT8 mutations, including one missense variation not clearly predicted to be damaging but found in a severely affected patient. Functional studies in fibroblasts and JEG3 cells demonstrate the usefulness of both cellular models in validating the deleterious effects of a new MCT8 mutation if there is still a doubt as to its pathogenicity. Moreover, the screening of fibroblasts from a large number of patient fibroblasts and of transfected mutations has allowed us to demonstrate that JEG3 transfected cells are more relevant than fibroblasts in revealing a genotype-phenotype correlation.


Assuntos
Estudos de Associação Genética , Transportadores de Ácidos Monocarboxílicos/genética , Mutação , Transtornos Psicomotores/genética , Transtornos Psicomotores/fisiopatologia , Adolescente , Linhagem Celular Tumoral , Células Cultivadas , Criança , Pré-Escolar , Fibroblastos/metabolismo , Humanos , Masculino , Transportadores de Ácidos Monocarboxílicos/metabolismo , Índice de Gravidade de Doença , Simportadores , Hormônios Tireóideos/metabolismo
12.
bioRxiv ; 2023 Feb 06.
Artigo em Inglês | MEDLINE | ID: mdl-36798224

RESUMO

Defects in hydroxymethylbilane synthase (HMBS) can cause Acute Intermittent Porphyria (AIP), an acute neurological disease. Although sequencing-based diagnosis can be definitive, ~⅓ of clinical HMBS variants are missense variants, and most clinically-reported HMBS missense variants are designated as "variants of uncertain significance" (VUS). Using saturation mutagenesis, en masse selection, and sequencing, we applied a multiplexed validated assay to both the erythroid-specific and ubiquitous isoforms of HMBS, obtaining confident functional impact scores for >84% of all possible amino-acid substitutions. The resulting variant effect maps generally agreed with biochemical expectation. However, the maps showed variants at the dimerization interface to be unexpectedly well tolerated, and suggested residue roles in active site dynamics that were supported by molecular dynamics simulations. Most importantly, these HMBS variant effect maps can help discriminate pathogenic from benign variants, proactively providing evidence even for yet-to-be-observed clinical missense variants.

13.
JAMA Dermatol ; 156(5): 570-575, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32186677

RESUMO

Importance: The effectiveness of afamelanotide treatment in patients with erythropoietic protoporphyria (EPP) in clinical practice who experience pain after light exposure that substantially impairs quality of life is unknown. Objective: To evaluate the association of afamelanotide treatment with outcomes in patients with EPP in regular practice during longer-term follow-up. Design, Setting, and Participants: This single-center, prospective postauthorization safety and efficacy cohort study was directed and approved by the European Medicines Agency. Data were collected from patients with EPP treated with afamelanotide at Erasmus MC between June 2016 and September 2018. Analysis began October 2018. Main Outcomes and Measures: Time spent outside during treatment, number of phototoxic reactions, disease-specific quality of life, usage of protective clothing, and adverse events. Results: A total of 117 patients with EPP (59 women [50.4%]; mean [SD] age, 43.0 [15.5] years) were treated with afamelanotide. Nearly all patients continued treatment (115 [98%]) with a median (interquartile range) follow-up of 2.0 (1.3-2.1) years. Compared with baseline, mean time spent outside during treatment increased significantly by an added 6.1 hours per week (95% CI, 3.62-8.67; P < .001). Mean quality of life score improved significantly by 14.01% (95% CI, 4.53%-23.50%; P < .001). Phototoxic reactions were less painful (ß, -0.85; 95% CI, -1.43 to -0.26; P < .001), but there was no significant difference in number or duration of reactions. Minor self-limiting adverse events occurred, such as nausea, fatigue, and headache. Conclusions and Relevance: This cohort study found that afamelanotide treatment was associated with improved clinical outcomes and a good safety profile for patients with EPP. The treatment has clinically significant, sustained positive associations with quality of life, is associated with increased duration of sun exposure, and is associated with less severe phototoxic reactions.


Assuntos
Fármacos Dermatológicos/administração & dosagem , Protoporfiria Eritropoética/tratamento farmacológico , Qualidade de Vida , alfa-MSH/análogos & derivados , Adolescente , Adulto , Idoso , Estudos de Coortes , Fármacos Dermatológicos/efeitos adversos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Protoporfiria Eritropoética/fisiopatologia , Luz Solar/efeitos adversos , Resultado do Tratamento , Adulto Jovem , alfa-MSH/administração & dosagem , alfa-MSH/efeitos adversos
14.
Hum Mutat ; 30(1): 29-38, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-18636565

RESUMO

Monocarboxylate transporter 8 (MCT8; approved symbol SLC16A2) facilitates cellular uptake and efflux of 3,3',5-triiodothyronine (T3). Mutations in MCT8 are associated with severe psychomotor retardation, high serum T3 and low 3,3',5'-triiodothyronine (rT3) levels. Here we report three novel MCT8 mutations. Two subjects with the F501del mutation have mild psychomotor retardation with slightly elevated T3 and normal rT3 levels. T3 uptake was mildly affected in F501del fibroblasts and strongly decreased in fibroblasts from other MCT8 patients, while T3 efflux was always strongly reduced. Moreover, type 3 deiodinase activity was highly elevated in F501del fibroblasts, whereas it was reduced in fibroblasts from other MCT8 patients, probably reflecting parallel variation in cellular T3 content. Additionally, T3-responsive genes were markedly upregulated by T3 treatment in F501del fibroblasts but not in fibroblasts with other MCT8 mutations. In conclusion, mutations in MCT8 result in a decreased T3 uptake in skin fibroblasts. The much milder clinical phenotype of patients with the F501del mutation may be correlated with the relatively small decrease in T3 uptake combined with an even greater decrease in T3 efflux. If fibroblasts are representative of central neurons, abnormal brain development associated with MCT8 mutations may be the consequence of either decreased or increased intracellular T3 concentrations.


Assuntos
Transportadores de Ácidos Monocarboxílicos/genética , Mutação , Sequência de Bases , Células Cultivadas , Fibroblastos , Humanos , Deficiência Intelectual/genética , Deficiência Intelectual/patologia , Dados de Sequência Molecular , RNA Mensageiro/metabolismo , Simportadores , Hormônios Tireóideos/metabolismo , Transfecção , Tri-Iodotironina/metabolismo
15.
Clin Endocrinol (Oxf) ; 70(5): 742-50, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-18785993

RESUMO

OBJECTIVE: Five per cent to 30% of cases of idiopathic isolated GH deficiency (IGHD) have first-degree relatives with short stature, which is suggestive of a genetic aetiology. The HYPOPIT study aimed to obtain an overall picture of gene encoding pituitary GH (GH1) and gene encoding GH releasing hormone-receptor (GHRHR) defects in a Dutch IGHD cohort and to relate them with clinical parameters. DESIGN, PATIENTS AND MEASUREMENTS: Genetic analysis was performed of exons and exon-intron boundaries of GH1 and GHRHR in 89 Caucasian IGHD patients from 81 families, using denaturing high-performance liquid chromatography (dHPLC), DNA sequencing and multiplex ligation-dependent probe amplification. In addition, we performed functional studies on novel identified GH1 exonic variants. RESULTS: Five different heterozygous GH1 mutations were present in 5 out of 81 participating families (6.1%), whereas no mutations in GHRHR were found. Patients with IGF-I SDS < -4.0 and peak GH levels < 5.7 mU/l had a mutation frequency of 40%, in contrast to 6.8% in patients with only one criterion, and 0.0% in patients with none of these criteria (P = 0.00007). Five new GH1 and two GHRHR variants were also identified; two of them (GH1 F92L and D153H) caused a marked reduction of GH secretion in vitro. CONCLUSION: GH1 and GHRHR mutations are rare in Caucasian Dutch IGHD patients, which suggests the involvement of other genetic determinants in the aetiology of IGHD. IGF-I < -4.0 and peak GH levels < 5.7 mU/l are strong predictors of GH1 mutations in the studied population.


Assuntos
Hormônio do Crescimento Humano/deficiência , Hormônio do Crescimento Humano/genética , Animais , Estatura , Células COS , Pré-Escolar , Chlorocebus aethiops , Estudos de Coortes , Feminino , Frequência do Gene , Testes Genéticos , Variação Genética , Heterozigoto , Humanos , Lactente , Masculino , Mutação , Países Baixos , Fenótipo , Receptores de Neuropeptídeos/genética , Receptores de Hormônios Reguladores de Hormônio Hipofisário/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Transfecção , População Branca/genética
16.
Trends Endocrinol Metab ; 19(2): 50-6, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-18291666

RESUMO

Thyroid hormone (TH) is essential for the proper development of numerous tissues, notably the brain. TH acts mostly intracellularly, which requires transport by TH transporters across the plasma membrane. Although several transporter families have been identified, only monocarboxylate transporter (MCT)8, MCT10 and organic anion-transporting polypeptide (OATP)1C1 demonstrate a high degree of specificity towards TH. Recently, the biological importance of MCT8 has been elucidated. Mutations in MCT8 are associated with elevated serum T(3) levels and severe psychomotor retardation, indicating a pivotal role for MCT8 in brain development. MCT8 knockout mice lack neurological damage, but mimic TH abnormalities of MCT8 patients. The exact pathophysiological mechanisms in MCT8 patients remain to be elucidated fully. Future research will probably identify novel TH transporters and disorders based on TH transporter defects.


Assuntos
Transporte Biológico , Membrana Celular/metabolismo , Hormônios Tireóideos/metabolismo , Sistemas de Transporte de Aminoácidos Neutros/metabolismo , Sistemas de Transporte de Aminoácidos Neutros/fisiologia , Animais , Transporte Biológico/fisiologia , Humanos , Camundongos , Camundongos Knockout , Modelos Biológicos , Transportadores de Ácidos Monocarboxílicos/genética , Transportadores de Ácidos Monocarboxílicos/metabolismo , Transportadores de Ácidos Monocarboxílicos/fisiologia , Mutação , Transportadores de Ânions Orgânicos/metabolismo , Transportadores de Ânions Orgânicos/fisiologia , Simportadores , Síndrome da Resistência aos Hormônios Tireóideos/genética , Síndrome da Resistência aos Hormônios Tireóideos/metabolismo , Síndrome da Resistência aos Hormônios Tireóideos/terapia
17.
Mol Endocrinol ; 22(6): 1357-69, 2008 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-18337592

RESUMO

Cellular entry of thyroid hormone is mediated by plasma membrane transporters, among others a T-type (aromatic) amino acid transporter. Monocarboxylate transporter 10 (MCT10) has been reported to transport aromatic amino acids but not iodothyronines. Within the MCT family, MCT10 is most homologous to MCT8, which is a very important iodothyronine transporter but does not transport amino acids. In view of this paradox, we decided to reinvestigate the possible transport of thyroid hormone by human (h) MCT10 in comparison with hMCT8. Transfection of COS1 cells with hMCT10 cDNA resulted in 1) the production of an approximately 55 kDa protein located to the plasma membrane as shown by immunoblotting and confocal microscopy, 2) a strong increase in the affinity labeling of intracellular type I deiodinase by N-bromoacetyl-[(125)I]T(3), 3) a marked stimulation of cellular T(4) and, particularly, T(3) uptake, 4) a significant inhibition of T(3) uptake by phenylalanine, tyrosine, and tryptophan of 12.5%, 22.2%, and 51.4%, respectively, and 5) a marked increase in the intracellular deiodination of T(4) and T(3) by different deiodinases. Cotransfection studies using the cytosolic thyroid hormone-binding protein micro-crystallin (CRYM) indicated that hMCT10 facilitates both cellular uptake and efflux of T(4) and T(3). In the absence of CRYM, hMCT10 and hMCT8 increased T(3) uptake after 5 min incubation up to 4.0- and 1.9-fold, and in the presence of CRYM up to 6.9- and 5.8-fold, respectively. hMCT10 was less active toward T(4) than hMCT8. These findings establish that hMCT10 is at least as active a thyroid hormone transporter as hMCT8, and that both transporters facilitate iodothyronine uptake as well as efflux.


Assuntos
Sistemas de Transporte de Aminoácidos Neutros/metabolismo , Hormônios Tireóideos/metabolismo , Sequência de Aminoácidos , Sistemas de Transporte de Aminoácidos Neutros/genética , Animais , Transporte Biológico , Células COS , Células Cultivadas , Chlorocebus aethiops , Eficiência , Humanos , Dados de Sequência Molecular , Transportadores de Ácidos Monocarboxílicos/genética , Transportadores de Ácidos Monocarboxílicos/metabolismo , Filogenia , Homologia de Sequência de Aminoácidos , Simportadores , Fatores de Tempo , Transfecção , Cristalinas mu
18.
Endocrinology ; 149(5): 2184-90, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18187543

RESUMO

Loss-of-function mutations in thyroid hormone transporter monocarboxylate transporter 8 (MCT8) lead to severe X-linked psychomotor retardation and elevated serum T(3) levels. Most patients, for example those with mutations V235M, S448X, insI189, or delF230, cannot stand, walk, or speak. Patients with mutations L434W, L568P, and S194F, however, walk independently and/or develop some dysarthric speech. To study the relationship between mutation and phenotype, we transfected JEG3 and COS1 cells with wild-type or mutant MCT8. Expression and function of the transporter were studied by analyzing T(3) and T(4) uptake, T(3) metabolism (by cotransfected type 3 deiodinase), Western blotting, affinity labeling with N-bromoacetyl-T(3), immunocytochemistry, and quantitative RT-PCR. Wild-type MCT8 increased T(3) uptake and metabolism about 5-fold compared with empty vector controls. Mutants V235M, S448X, insI189, and delF230 did not significantly increase transport. However, S194F, L568P, and L434W showed about 20, 23, and 37% of wild-type activity. RT-PCR did not show significant differences in mRNA expression between wild-type and mutant MCT8. Immunocytochemistry detected the nonfunctional mutants V235M, insI189, and delF230 mostly in the cytoplasm, whereas mutants with residual function were expressed at the plasma membrane. Mutants S194F and L434W showed high protein expression but low affinity for N-bromoacetyl-T(3); L568P was detected in low amounts but showed relatively high affinity. Mutations in MCT8 cause loss of function through reduced protein expression, impaired trafficking to the plasma membrane, or reduced substrate affinity. Mutants L434W, L568P, and S194F showed significant residual transport capacity, which may underlie the more advanced psychomotor development observed in patients with these mutations.


Assuntos
Transportadores de Ácidos Monocarboxílicos/genética , Mutação , Fenótipo , Animais , Células COS , Células Cultivadas , Chlorocebus aethiops , Cristalinas/genética , Genótipo , Humanos , Transportadores de Ácidos Monocarboxílicos/metabolismo , Mutagênese Sítio-Dirigida , Polimorfismo de Nucleotídeo Único , Desempenho Psicomotor , Simportadores , Hormônios Tireóideos/metabolismo , Hormônios Tireóideos/farmacocinética , Transfecção , Cristalinas mu
19.
Endocrinology ; 149(10): 5307-14, 2008 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-18566113

RESUMO

Organic anion transporting polypeptide (OATP) 1C1 has been characterized as a specific thyroid hormone transporter. Based on its expression in capillaries in different brain regions, OATP1C1 is thought to play a key role in transporting thyroid hormone across the blood-brain barrier. For this reason, we studied the specificity of iodothyronine transport by OATP1C1 in detail by analysis of thyroid hormone uptake in OATP1C1-transfected COS1 cells. Furthermore, we examined whether OATP1C1 is rate limiting in subsequent thyroid hormone metabolism in cells cotransfected with deiodinases. We also studied the effect of genetic variation in the OATP1C1 gene: polymorphisms were determined in 155 blood donors and 1192 Danish twins and related to serum thyroid hormone levels. In vitro effects of the polymorphisms were analyzed in cells transfected with the variants. Cells transfected with OATP1C1 showed increased transport of T4 and T4 sulfate (T4S), little transport of rT3, and no transport of T3 or T3 sulphate, compared with mock transfected cells. Metabolism of T4, T4S, and rT3 by cotransfected deiodinases was greatly augmented in the presence of OATP1C1. The OATP1C1-intron3C>T, Pro143Thr, and C3035T polymorphisms were not consistently associated with thyroid hormone levels, nor did they affect transport function in vitro. In conclusion, OATP1C1 mediates transport of T4, T4S, and rT3 and increases the access of these substrates to the intracellular active sites of the deiodinases. No effect of genetic variation on the function of OATP1C1 was observed.


Assuntos
Variação Genética , Transportadores de Ânions Orgânicos/genética , Transportadores de Ânions Orgânicos/metabolismo , Tiroxina/metabolismo , Tri-Iodotironina/metabolismo , Animais , Células COS , Chlorocebus aethiops , Estudos de Coortes , Dinamarca , Haplótipos , Humanos , Radioisótopos do Iodo , Polimorfismo Genético , Sulfatos , Tiroxina/sangue , Transfecção , Tri-Iodotironina/sangue , Gêmeos
20.
Endocrinology ; 149(9): 4695-701, 2008 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18499754

RESUMO

Sulfation is an important pathway in the metabolism of thyroid hormone and estrogens. Sulfation of estrogens is reversible by estrogen sulfatase, but sulfation of thyroid hormone accelerates its degradation by the type 1 deiodinase in liver. Organic anion transporters (OATPs) are capable of transporting iodothyronine sulfates such as T4 sulfate (T4S), T3S, and rT3S or estrogen sulfates like estrone sulfate (E1S), but the major hepatic transporter for these conjugates has not been identified. A possible candidate is OATP1B1 because model substrates for this transporter include the bilirubin mimic bromosulfophthalein (BSP) and E1S, and it is highly and specifically expressed in liver. Therefore, OATP1B1-transfected COS1 cells were studied by analysis of BSP, E1S, and iodothyronine sulfate uptake and metabolism. Two Caucasian populations (155 blood donors and 1012 participants of the Rotterdam Scan Study) were genotyped for the OATP1B1-Val174Ala polymorphism and associated with bilirubin, E1S, and T4S levels. OATP1B1-transfected cells strongly induced uptake of BSP, E1S, T4S, T3S, and rT3S compared with mock-transfected cells. Metabolism of iodothyronine sulfates by cotransfected type 1 deiodinase was greatly augmented in the presence of OATP1B1. OATP1B1-Val174 showed a 40% higher induction of transport and metabolism of these substrates than OATP1B1-Ala174. Carriers of the OATP1B1-Ala174 allele had higher serum bilirubin, E1S, and T4S levels. In conclusion, OATP1B1 is an important factor in hepatic transport and metabolism of bilirubin, E1S, and iodothyronine sulfates. OATP1B1-Ala174 displays decreased transport activity and thereby gives rise to higher bilirubin, E1S, and T4S levels in carriers of this polymorphism.


Assuntos
Estrogênios/metabolismo , Fígado/metabolismo , Transportadores de Ânions Orgânicos/fisiologia , Hormônios Tireóideos/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Bilirrubina/metabolismo , Transporte Biológico/genética , Células COS , Chlorocebus aethiops , Estrona/análogos & derivados , Estrona/metabolismo , Feminino , Humanos , Transportador 1 de Ânion Orgânico Específico do Fígado , Masculino , Pessoa de Meia-Idade , Modelos Biológicos , Transportadores de Ânions Orgânicos/genética , Polimorfismo de Nucleotídeo Único/fisiologia , Sulfatos/metabolismo , Sulfobromoftaleína/farmacocinética , Tiroxina/análogos & derivados , Tiroxina/metabolismo
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