Engineering a Glucose-responsive Human Insulin-secreting Cell Line from Islets of Langerhans Isolated from a Patient with Persistent Hyperinsulinemic Hypoglycemia of Infancy*
- Wendy M. MacFarlane‡,
- Joanna C. Chapman§,
- Ruth M. Shepherd§,
- Molly N. Hashmi§,
- Noritaka Kamimura§,
- Karen E. Cosgrove§,
- Rachel E. O'Brien§,
- Philippa D. Barnes§,
- Alan W. Hart‡,
- Hilary M. Docherty‡,
- Keith J. Lindley¶,
- Albert Aynsley-Green¶,
- Roger F. L. James‖,
- Kevin Docherty‡ and
- Mark J. Dunne§**
- From the ‡Department of Molecular and Cell Biology, University of Aberdeen, Institute of Medical Sciences, Aberdeen AB25 2ZD, the §Institute of Molecular Physiology and Department of Biomedical Science, Sheffield University, Western Bank, Sheffield S10 2TN, the ¶Institute of Child Health, University of London, 30 Guilford Street, London WC1N 1EH, and‖Department of Surgery, University of Leicester, Leicester Royal Infirmary, Leicester LE2 7LX, United Kingdom
Abstract
Persistent hyperinsulinemic hypoglycemia of infancy (PHHI) is a neonatal disease characterized by dysregulation of insulin secretion accompanied by profound hypoglycemia. We have discovered that islet cells, isolated from the pancreas of a PHHI patient, proliferate in culture while maintaining a beta cell-like phenotype. The PHHI-derived cell line (NES2Y) exhibits insulin secretory characteristics typical of islet cells derived from these patients, i.e. they have no KATP channel activity and as a consequence secrete insulin at constitutively high levels in the absence of glucose. In addition, they exhibit impaired expression of the homeodomain transcription factor PDX1, which is a key component of the signaling pathway linking nutrient metabolism to the regulation of insulin gene expression. To repair these defects NES2Y cells were triple-transfected with cDNAs encoding the two components of the KATP channel (SUR1 and Kir6.2) and PDX1. One selected clonal cell line (NISK9) had normal KATPchannel activity, and as a result of changes in intracellular Ca2+ homeostasis ([Ca2+]i) secreted insulin within the physiological range of glucose concentrations. This approach to engineering PHHI-derived islet cells may be of use in gene therapy for PHHI and in cell engineering techniques for administering insulin for the treatment of diabetes mellitus.
Footnotes
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↵* This work was supported by grants from the Wellcome Trust, Medical Research Council, and the British Diabetic Association.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵** To whom correspondence should be addressed: Institute of Molecular Physiology and Dept. of Biomedical Science, Sheffield University, Western Bank, Sheffield, S10 2TN, UK. Tel.: 114-222-4636; Fax: 114-276-5413; E-mail: m.j.dunne@sheffield.ac.uk.
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↵2 M. J. Dunne and A. Aynsley-Green, unpublished observations.
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↵3 W. M. MacFarlane and K. Docherty, unpublished observations.
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↵4 W. M. Macfarlane, K. Docherty, and M. J. Dunne, manuscript in preparation.
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↵5 W. M. MacFarlane and H. M. Docherty, unpublished observations.
- Abbreviations:
- PHHI
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persistent hyperinsulinemic hypoglycemia of infancy
- KATP channel
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ATP-sensitive potassium channel
- [Ca2+]i
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intracellular free calcium ion concentration
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- Received March 1, 1999.
- Revision received September 20, 1999.
- The American Society for Biochemistry and Molecular Biology, Inc.
