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Transport of ascorbic acid and dehydroascorbic acid by pancreatic islet cells from neonatal rats
A Zhou1, J H Nielsen, O Farver
1Department of Medical Physiology C, University of Copenhagen, Denmark.
Insights
Pancreatic islet cells actively transport ascorbic acid via a saturable mechanism. This process is influenced by glucose and various inhibitors, highlighting its complexity in peptide hormone production.
Area of Science:
- Biochemistry
- Endocrinology
- Cell Biology
Background:
- Endocrine pancreatic tissue produces amidated peptides like pancreastatin and amylin.
- Peptidylglycine alpha-amidating mono-oxygenase (PAM) is crucial for peptide processing.
- Ascorbic acid, copper, and oxygen are essential for PAM activity.
Purpose of the Study:
- To investigate the mechanism of ascorbic acid uptake in pancreatic islet cells.
- To characterize the transport system for ascorbic acid and its oxidized form, dehydroascorbic acid.
- To explore the regulation of ascorbic acid uptake by glucose and other substances.
Main Methods:
- Incubation of rat pancreatic islet cells with 14C-labelled ascorbic acid.
- Assessing the effects of various inhibitors (ouabain, phloridzin, etc.) on ascorbic acid uptake.
- Investigating dehydroascorbic acid uptake and its conversion to ascorbic acid.
- Analyzing the composition of isolated secretory granules.
Main Results:
- Pancreatic islet cells accumulate ascorbic acid via Na(+)-dependent active transport (Km 17.6 microM).
- Transport is inhibited by ouabain, phloridzin, cytochalasin B, amiloride, and probenecid.
- Glucose exhibits time-dependent inhibition or stimulation of uptake.
- Dehydroascorbic acid uptake is linear and converted to ascorbic acid intracellularly.
- Tri-iodothyronine inhibits uptake of both forms; glucocorticoids inhibit ascorbic acid uptake.
Conclusions:
- Pancreatic islet cells possess a specific, saturable transport system for ascorbic acid.
- Ascorbic acid uptake is regulated by glucose and hormonal factors.
- The findings provide insights into the supply of ascorbic acid for peptide amidation in the pancreas.
Abstract:
Several amidated biologically active peptides such as pancreastatin, thyrotropin-releasing hormone, pancreatic polypeptide and amylin are produced in endocrine pancreatic tissue which contains the enzyme necessary for their final processing, i.e. peptidylglycine alpha-amidating mono-oxygenase (EC 1.14.17.3). The enzyme needs ascorbic acid for activity as well as copper and molecular oxygen. The present work shows that pancreatic islet cells prepared from overnight cultures of isolated islets from 5-7-day-old rats accumulate 14C-labelled ascorbic acid by a Na(+)-dependent active transport mechanism which involves a saturable process (estimated Km 17.6 microM). Transport was inhibited by ouabain, phloridzin, cytochalasin B, amiloride and probenecid. Glucose inhibited or stimulated uptake, depending on the length of incubation time of the cells. The uptake of dehydroascorbic acid was linearly dependent on concentration. Dehydroascorbic acid was converted to ascorbic acid by an unknown mechanism after uptake. The uptake of both ascorbic acid and dehydroascorbic acid was inhibited by tri-iodothyronine, and uptake of ascorbic acid, but not of dehydroascorbic acid, was inhibited by glucocorticoids. Isolated secretory granules contained a fairly low concentration of iron but a high concentration of copper.