title: Down-regulation of ZnT8 expression in INS-1 rat pancreatic beta cells reduces insulin content and glucose-inducible insulin secretion.
abstract: The SLC30A8 gene codes for a pancreatic beta-cell-expressed zinc transporter, ZnT8. A polymorphism in the SLC30A8 gene is associated with susceptibility to type 2 diabetes, although the molecular mechanism through which this phenotype is manifest is incompletely understood. Such polymorphisms may exert their effect via impacting expression level of the gene product. We used an shRNA-mediated approach to reproducibly downregulate ZnT8 mRNA expression by >90% in the INS-1 pancreatic beta cell line. The ZnT8-downregulated cells exhibited diminished uptake of exogenous zinc, as determined using the zinc-sensitive reporter dye, zinquin. ZnT8-downregulated cells showed reduced insulin content and decreased insulin secretion (expressed as percent of total insulin content) in response to hyperglycemic stimulus, as determined by insulin immunoassay. ZnT8-depleted cells also showed fewer dense-core vesicles via electron microscopy. These data indicate that reduced ZnT8 expression in cultured pancreatic beta cells gives rise to a reduced insulin response to hyperglycemia. In addition, although we provide no direct evidence, these data suggest that an SLC30A8 expression-level polymorphism could affect insulin secretion and the glycemic response in vivo.
authors.affiliation: Division of Nephrology & Hypertension, Department of Medicine, Oregon Health & Science University, Portland, Oregon, United States of America.
authors.forename: Yi
authors.initials: Y
authors.lastname: Fu
authors.forename: Wei
authors.initials: W
authors.lastname: Tian
authors.forename: Emily B
authors.initials: EB
authors.lastname: Pratt
authors.forename: Lisa B
authors.initials: LB
authors.lastname: Dirling
authors.forename: Show-Ling
authors.initials: SL
authors.lastname: Shyng
authors.forename: Charles K
authors.initials: CK
authors.lastname: Meshul
authors.forename: David M
authors.initials: DM
authors.lastname: Cohen
mesh_terms: D000818:Animals; D027682:Cation Transport Proteins / Q000235:genetics* / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects*; D005453:Fluorescence; D055785:Gene Knockdown Techniques; D005947:Glucose / Q000494:pharmacology*; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D009532:Nickel / Q000494:pharmacology; D015363:Quinolones / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D022142:Secretory Vesicles / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D014105:Tosyl Compounds / Q000378:metabolism; D015032:Zinc / Q000378:metabolism / Q000494:pharmacology; D000075703:Zinc Transporter 8
chemical_list: D027682:Cation Transport Proteins; D007328:Insulin; D015363:Quinolones; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C542071:Slc30a8 protein, rat; D014105:Tosyl Compounds; D000075703:Zinc Transporter 8; D009532:Nickel; D005947:Glucose; D015032:Zinc; C084731:zinquin