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1.
EBioMedicine ; 9: 257-277, 2016 Jul.
Article in English | MEDLINE | ID: mdl-27333023

ABSTRACT

Dravet syndrome is a devastating genetic brain disorder caused by heterozygous loss-of-function mutation in the voltage-gated sodium channel gene SCN1A. There are currently no treatments, but the upregulation of SCN1A healthy allele represents an appealing therapeutic strategy. In this study we identified a novel, evolutionary conserved mechanism controlling the expression of SCN1A that is mediated by an antisense non-coding RNA (SCN1ANAT). Using oligonucleotide-based compounds (AntagoNATs) targeting SCN1ANAT we were able to induce specific upregulation of SCN1A both in vitro and in vivo, in the brain of Dravet knock-in mouse model and a non-human primate. AntagoNAT-mediated upregulation of Scn1a in postnatal Dravet mice led to significant improvements in seizure phenotype and excitability of hippocampal interneurons. These results further elucidate the pathophysiology of Dravet syndrome and outline a possible new approach for the treatment of this and other genetic disorders with similar etiology.


Subject(s)
Brain/metabolism , Epilepsies, Myoclonic/pathology , NAV1.1 Voltage-Gated Sodium Channel/metabolism , RNA, Long Noncoding/metabolism , Alleles , Animals , Base Sequence , Behavior, Animal , Brain/diagnostic imaging , Cell Line , Chlorocebus aethiops , Disease Models, Animal , Electroencephalography , Epilepsies, Myoclonic/genetics , Epilepsies, Myoclonic/metabolism , Gene Expression , Gene Knock-In Techniques , Hippocampus/physiology , Humans , In Vitro Techniques , Interneurons/metabolism , Mice , Mice, Transgenic , Molecular Sequence Data , NAV1.1 Voltage-Gated Sodium Channel/chemistry , NAV1.1 Voltage-Gated Sodium Channel/genetics , Nucleic Acid Conformation , Oligonucleotides, Antisense/metabolism , Patch-Clamp Techniques , Phenotype , RNA, Long Noncoding/antagonists & inhibitors , RNA, Long Noncoding/genetics , Real-Time Polymerase Chain Reaction , Sequence Alignment , Sequence Analysis, RNA , Temperature , Up-Regulation , Vero Cells , Video Recording
2.
Eur J Endocrinol ; 163(4): 509-13, 2010 Oct.
Article in English | MEDLINE | ID: mdl-20621957

ABSTRACT

OBJECTIVE: Despite the contribution of national registries and population-based reports, data concerning the epidemiology of acromegaly is scanty. In addition, the role of the environmental context has not been investigated. DESIGN: Epidemiology of acromegaly was studied in the province of Messina (Sicily, Italy), focusing on the influence of environmental factors. METHODS: Four zones, characterized by different degrees of exposition to environmental toxins due to industrial pollution, were identified in the province: area A (76,338 inhabitants), area B (287,328 inhabitants), area C (243,381 inhabitants), and area D (47,554 inhabitants) at low, middle-low, middle, and high industrial density respectively. We identified all acromegalics who were born and resided in the province of Messina, among patients either referred to our endocrine unit or referred elsewhere but recorded in the archives of the provincial healthcare agency. RESULTS: In the province of Messina, we found 64 patients (2 in area A, 24 in area B, 28 in area C, and 10 in area D). Macroadenomas were 60%, the male/female ratio was 1, and mean age at diagnosis (±s.e.m.) was 45.4±1.6 years. Overall, prevalence was 97 c.p.m. in the province (26 c.p.m. in area A, 84 c.p.m. in area B, 115 c.p.m. in area C, and 210 c.p.m. in area D). Risk ratio (RR), calculated in every area assuming area A as a reference, showed an increased risk of developing acromegaly in people residing in area D (RR=8.03; P<0.0014). CONCLUSION: This study confirms the prevalence of acromegaly reported recently. The increased risk of developing this disease in area D suggests that the pathogenetic role of environmental context needs to be better evaluated.


Subject(s)
Acromegaly/epidemiology , Environmental Pollution/adverse effects , Acromegaly/etiology , Adult , Female , Geography , Humans , Italy/epidemiology , Male , Middle Aged , Prevalence
3.
Diabetologia ; 49(7): 1567-77, 2006 Jul.
Article in English | MEDLINE | ID: mdl-16752175

ABSTRACT

AIMS/HYPOTHESIS: ATP-sensitive K(+) (K(ATP)) channels located on the beta cell plasma membrane play a critical role in regulating insulin secretion and are targets for the sulfonylurea class of antihyperglycaemic drugs. Recent reports suggest that these channels may also reside on insulin-containing dense-core vesicles and mitochondria. The aim of this study was to explore these possibilities and to test the hypothesis that vesicle-resident channels play a role in the control of organellar Ca(2+) concentration or pH. METHODS: To quantify the subcellular distribution of the pore-forming subunit Kir6.2 and the sulfonylurea binding subunit SUR1 in isolated mouse islets and clonal pancreatic MIN6 beta cells, we used four complementary techniques: immunoelectron microscopy, density gradient fractionation, vesicle immunopurification and fluorescence-activated vesicle isolation. Intravesicular and mitochondrial concentrations of free Ca(2+) were measured in intact or digitonin-permeabilised MIN6 cells using recombinant, targeted aequorins, and intravesicular pH was measured with the recombinant fluorescent probe pHluorin. RESULTS: SUR1 and Kir6.2 immunoreactivity were concentrated on dense-core vesicles and on vesicles plus the endoplasmic reticulum/Golgi network, respectively, in both islets and MIN6 cells. Reactivity to neither subunit was detected on mitochondria. Glibenclamide, tolbutamide and diazoxide all failed to affect Ca(2+) uptake into mitochondria, and K(ATP) channel regulators had no significant effect on intravesicular free Ca(2+) concentrations or vesicular pH. CONCLUSIONS/INTERPRETATION: A significant proportion of Kir6.2 and SUR1 subunits reside on insulin-secretory vesicles and the distal secretory pathway in mouse beta cells but do not influence intravesicular ion homeostasis. We propose that dense-core vesicles may serve instead as sorting stations for the delivery of channels to the plasma membrane.


Subject(s)
ATP-Binding Cassette Transporters/metabolism , Insulin-Secreting Cells/metabolism , Multidrug Resistance-Associated Proteins/metabolism , Potassium Channels, Inwardly Rectifying/metabolism , Potassium/metabolism , ATP-Binding Cassette Transporters/physiology , Animals , Calcium/metabolism , Cells, Cultured , Homeostasis , Hydrogen-Ion Concentration , Insulin-Secreting Cells/ultrastructure , Ion Transport , Membrane Potential, Mitochondrial , Mice , Multidrug Resistance-Associated Proteins/physiology , Organelles/metabolism , Potassium Channels, Inwardly Rectifying/physiology , Receptors, Drug , Secretory Vesicles/metabolism , Sulfonylurea Receptors , Tissue Distribution
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