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1.
Cells ; 12(13)2023 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-37443824

RESUMO

Magnesium is an essential mediator of a vast number of critical enzymatic cellular reactions in the human body. Some clinical epidemiological studies suggest that hypomagnesemia accounts for declines in insulin secretion in patients with type 2 diabetes (T2D); however, the results of various experimental studies do not support this notion. To address this discrepancy, we assessed the short- and long-term effects of hypomagnesemia on ß-cell function and insulin secretion in primary mouse islets of Langerhans and in a mouse model of hypomagnesemia known as Trpm6Δ17 /fl;Villin1-Cre mice. We found that lowering the extracellular Mg2+ concentration from 1.2 mM to either 0.6 or 0.1 mM remarkably increased glucose-induced insulin secretion (GIIS) in primary islets isolated from C57BL/6 mice. Similarly, both the plasma insulin levels and GIIS rose in isolated islets of Trpm6Δ17 /fl;Villin1-Cre mice. We attribute these rises to augmented increases in intracellular Ca2+ oscillations in pancreatic ß-cells. However, the glycemic metabolic profile was not impaired in Trpm6Δ17 /fl;Villin1-Cre mice, suggesting that chronic hypomagnesemia does not lead to insulin resistance. Collectively, the results of this study suggest that neither acute nor chronic Mg2+ deficiency suppresses glucose-induced rises in insulin secretion. Even though hypomagnesemia can be symptomatic of T2D, such deficiency may not account for declines in insulin release in this disease.


Assuntos
Diabetes Mellitus Tipo 2 , Camundongos , Humanos , Animais , Secreção de Insulina , Diabetes Mellitus Tipo 2/metabolismo , Camundongos Endogâmicos C57BL , Insulina/metabolismo , Glucose/metabolismo
2.
J Biol Chem ; 298(6): 101974, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35469921

RESUMO

Organic cation transporter 1 (OCT1) is a membrane transporter that affects hepatic uptake of cationic and weakly basic drugs. OCT1 transports structurally highly diverse substrates. The mechanisms conferring this polyspecificity are unknown. Here, we analyzed differences in transport kinetics between human and mouse OCT1 orthologs to identify amino acids that contribute to the polyspecificity of OCT1. Following stable transfection of HEK293 cells, we observed more than twofold differences in the transport kinetics of 22 out of 28 tested substrates. We found that the ß2-adrenergic drug fenoterol was transported with eightfold higher affinity but at ninefold lower capacity by human OCT1. In contrast, the anticholinergic drug trospium was transported with 11-fold higher affinity but at ninefold lower capacity by mouse Oct1. Using human-mouse chimeric constructs and site-directed mutagenesis, we identified nonconserved amino acids Cys36 and Phe32 as responsible for the species-specific differences in fenoterol and trospium uptake. Substitution of Cys36 (human) to Tyr36 (mouse) caused a reversal of the affinity and capacity of fenoterol but not trospium uptake. Substitution of Phe32 to Leu32 caused reversal of trospium but not fenoterol uptake kinetics. Comparison of the uptake of structurally similar ß2-adrenergics and molecular docking analyses indicated the second phenol ring, 3.3 to 4.8 Å from the protonated amino group, as essential for the affinity for fenoterol conferred by Cys36. This is the first study to report single amino acids as determinants of OCT1 polyspecificity. Our findings suggest that structure-function data of OCT1 is not directly transferrable between substrates or species.


Assuntos
Proteínas da Membrana Plasmática de Transporte de Catecolaminas/química , Transportador 1 de Cátions Orgânicos , Sequência de Aminoácidos , Animais , Proteínas da Membrana Plasmática de Transporte de Catecolaminas/metabolismo , Fenoterol , Células HEK293 , Humanos , Camundongos , Simulação de Acoplamento Molecular , Transportador 1 de Cátions Orgânicos/química , Transportador 1 de Cátions Orgânicos/metabolismo
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