title: Monitoring C-Peptide Storage and Secretion in Islet β-Cells In Vitro and In Vivo.
abstract: Human proinsulin with C-peptide-bearing Superfolder Green Fluorescent Protein (CpepSfGFP) has been expressed in transgenic mice, driven by the Ins1 promoter. The protein, expressed exclusively in β-cells, is processed and stored as CpepSfGFP and human insulin comprising only ∼0.04% of total islet proinsulin plus insulin, exerting no metabolic impact. The kinetics of the release of insulin and CpepSfGFP from isolated islets appear identical. Upon a single acute stimulatory challenge in vitro, fractional release of insulin does not detectably deplete islet fluorescence. In vivo, fluorescence imaging of the pancreatic surface allows, for the first time, visual assessment of pancreatic islet insulin content, and we demonstrate that CpepSfGFP visibly declines upon diabetes progression in live lepR(db/db) mice. In anesthetized mice, after intragastric or intravenous saline delivery, pancreatic CpepSfGFP (insulin) content remains undiminished. Remarkably, however, within 20 min after acute intragastric or intravenous glucose delivery (with blood glucose concentrations reaching >15 mmol/L), a small subset of islets shows rapid dispossession of a major fraction of their stored CpepSfGFP (insulin) content, whereas most islets exhibit no demonstrable loss of CpepSfGFP (insulin). These studies strongly suggest that there are "first responder" islets to an in vivo glycemic challenge, which cannot be replicated by islets in vitro.
authors.affiliation: Division of Metabolism, Endocrinology & Diabetes, University of Michigan Medical Center, Ann Arbor, MI.
authors.forename: Shuaishuai
authors.initials: S
authors.lastname: Zhu
authors.affiliation: Division of Metabolism, Endocrinology & Diabetes, University of Michigan Medical Center, Ann Arbor, MI.
authors.forename: Dennis
authors.initials: D
authors.lastname: Larkin
authors.affiliation: Department of Chemistry, University of Michigan, Ann Arbor, MI.
authors.forename: Shusheng
authors.initials: S
authors.lastname: Lu
authors.affiliation: Department of Cell Biology, University of Virginia, Charlottesville, VA.
authors.forename: Candice
authors.initials: C
authors.lastname: Inouye
authors.affiliation: Division of Metabolism, Endocrinology & Diabetes, University of Michigan Medical Center, Ann Arbor, MI.
authors.forename: Leena
authors.initials: L
authors.lastname: Haataja
authors.affiliation: Division of Metabolism, Endocrinology & Diabetes, University of Michigan Medical Center, Ann Arbor, MI.
authors.forename: Arfah
authors.initials: A
authors.lastname: Anjum
authors.affiliation: Department of Chemistry, University of Michigan, Ann Arbor, MI.
authors.forename: Robert
authors.identifier: http://orcid.org/0000-0001-9417-0534
authors.initials: R
authors.lastname: Kennedy
authors.affiliation: Department of Cell Biology, University of Virginia, Charlottesville, VA.
authors.forename: David
authors.identifier: http://orcid.org/0000-0002-3075-1580
authors.initials: D
authors.lastname: Castle
authors.affiliation: Division of Metabolism, Endocrinology & Diabetes, University of Michigan Medical Center, Ann Arbor, MI parvan@umich.edu.
authors.forename: Peter
authors.initials: P
authors.lastname: Arvan
mesh_terms: D000818:Animals; D001786:Blood Glucose / Q000187:drug effects / Q000378:metabolism; D002096:C-Peptide / Q000235:genetics / Q000378:metabolism*; D003920:Diabetes Mellitus / Q000378:metabolism*; D018450:Disease Progression; D005947:Glucose / Q000494:pharmacology; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D066298:In Vitro Techniques; D050417:Insulin-Secreting Cells / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D046529:Microscopy, Electron, Transmission; D011384:Proinsulin / Q000187:drug effects / Q000378:metabolism*
chemical_list: D001786:Blood Glucose; D002096:C-Peptide; D049452:Green Fluorescent Proteins; D011384:Proinsulin; D005947:Glucose