A stable analogue of glucose-dependent insulinotropic polypeptide, GIP(LysPAL(16)), enhances functional differentiation of mouse embryonic stem cells into cells expressing islet-specific genes and hormones

Lamin Marenah, Janie McCluskey, Yasser Abdel-Wahab, Finbarr O'Harte, Neville McClenaghan, Peter Flatt

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Abstract

Embryonic stem (ES) cells can be differentiated into insulin-producing cells by conditioning the culture media. However, the number of insulin-expressing cells and amount of insulin released is very low. Glucose-dependent insulinotropic polypeptide (GIP) enhances the growth and differentiation of pancreatic P-cells. This study examined the potential of the stable analogue GIP(LysPAL(16)) to enhance the differentiation of mouse ES cells into insulin-producing cells using a five-stage culturing strategy. Semi-quantitative PCR indicated mRNA expression of islet development markers (nestin, Pdx1, Nkx6.1, Oct4), mature pancreatic P-cell markers (insulin, glucagon, Glut2, Sur1, Kir6.1) and the GIP receptor gene GIP-R in undifferentiated (stage 1) cells, with increasing levels in differentiated stages 4 and 5. IAPP and somatostatin genes were only expressed in differentiated stages. lmmunohistochemical studies confirmed the presence of insulin, glucagon, somatostatin and IAPP in differentiated ES cells. After supplementation with GIP(LysPAL16), ES cells at stage 4 released insulin in response to secretagogues and glucose in a concentration-dependent manner, with 35-100% increases in insulin release. Cellular C-peptide content also increased by 45% at stages 4 and 5. We conclude that the stable GIP analogue enhanced differentiation of mouse ES cells towards a phenotype expressing specific P-cell genes and releasing insulin.
LanguageEnglish
Pages941-947
JournalBiological Chemistry
Volume387
Issue number7
DOIs
Publication statusPublished - Jul 2006

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Islets of Langerhans
Hormones
Insulin
Glucose
Peptides
Genes
Embryonic Stem Cells
Somatostatin
Glucagon
Mouse Embryonic Stem Cells
Nestin
C-Peptide
Culture Media
Cell Culture Techniques
Phenotype
Polymerase Chain Reaction
Messenger RNA
Growth

Cite this

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title = "A stable analogue of glucose-dependent insulinotropic polypeptide, GIP(LysPAL(16)), enhances functional differentiation of mouse embryonic stem cells into cells expressing islet-specific genes and hormones",
abstract = "Embryonic stem (ES) cells can be differentiated into insulin-producing cells by conditioning the culture media. However, the number of insulin-expressing cells and amount of insulin released is very low. Glucose-dependent insulinotropic polypeptide (GIP) enhances the growth and differentiation of pancreatic P-cells. This study examined the potential of the stable analogue GIP(LysPAL(16)) to enhance the differentiation of mouse ES cells into insulin-producing cells using a five-stage culturing strategy. Semi-quantitative PCR indicated mRNA expression of islet development markers (nestin, Pdx1, Nkx6.1, Oct4), mature pancreatic P-cell markers (insulin, glucagon, Glut2, Sur1, Kir6.1) and the GIP receptor gene GIP-R in undifferentiated (stage 1) cells, with increasing levels in differentiated stages 4 and 5. IAPP and somatostatin genes were only expressed in differentiated stages. lmmunohistochemical studies confirmed the presence of insulin, glucagon, somatostatin and IAPP in differentiated ES cells. After supplementation with GIP(LysPAL16), ES cells at stage 4 released insulin in response to secretagogues and glucose in a concentration-dependent manner, with 35-100{\%} increases in insulin release. Cellular C-peptide content also increased by 45{\%} at stages 4 and 5. We conclude that the stable GIP analogue enhanced differentiation of mouse ES cells towards a phenotype expressing specific P-cell genes and releasing insulin.",
author = "Lamin Marenah and Janie McCluskey and Yasser Abdel-Wahab and Finbarr O'Harte and Neville McClenaghan and Peter Flatt",
note = "4th General Meeting of the International-Proteolysis-Society/International Conference on Protease Inhibitors, Quebec City, CANADA, OCT 15-19, 2005",
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T1 - A stable analogue of glucose-dependent insulinotropic polypeptide, GIP(LysPAL(16)), enhances functional differentiation of mouse embryonic stem cells into cells expressing islet-specific genes and hormones

AU - Marenah, Lamin

AU - McCluskey, Janie

AU - Abdel-Wahab, Yasser

AU - O'Harte, Finbarr

AU - McClenaghan, Neville

AU - Flatt, Peter

N1 - 4th General Meeting of the International-Proteolysis-Society/International Conference on Protease Inhibitors, Quebec City, CANADA, OCT 15-19, 2005

PY - 2006/7

Y1 - 2006/7

N2 - Embryonic stem (ES) cells can be differentiated into insulin-producing cells by conditioning the culture media. However, the number of insulin-expressing cells and amount of insulin released is very low. Glucose-dependent insulinotropic polypeptide (GIP) enhances the growth and differentiation of pancreatic P-cells. This study examined the potential of the stable analogue GIP(LysPAL(16)) to enhance the differentiation of mouse ES cells into insulin-producing cells using a five-stage culturing strategy. Semi-quantitative PCR indicated mRNA expression of islet development markers (nestin, Pdx1, Nkx6.1, Oct4), mature pancreatic P-cell markers (insulin, glucagon, Glut2, Sur1, Kir6.1) and the GIP receptor gene GIP-R in undifferentiated (stage 1) cells, with increasing levels in differentiated stages 4 and 5. IAPP and somatostatin genes were only expressed in differentiated stages. lmmunohistochemical studies confirmed the presence of insulin, glucagon, somatostatin and IAPP in differentiated ES cells. After supplementation with GIP(LysPAL16), ES cells at stage 4 released insulin in response to secretagogues and glucose in a concentration-dependent manner, with 35-100% increases in insulin release. Cellular C-peptide content also increased by 45% at stages 4 and 5. We conclude that the stable GIP analogue enhanced differentiation of mouse ES cells towards a phenotype expressing specific P-cell genes and releasing insulin.

AB - Embryonic stem (ES) cells can be differentiated into insulin-producing cells by conditioning the culture media. However, the number of insulin-expressing cells and amount of insulin released is very low. Glucose-dependent insulinotropic polypeptide (GIP) enhances the growth and differentiation of pancreatic P-cells. This study examined the potential of the stable analogue GIP(LysPAL(16)) to enhance the differentiation of mouse ES cells into insulin-producing cells using a five-stage culturing strategy. Semi-quantitative PCR indicated mRNA expression of islet development markers (nestin, Pdx1, Nkx6.1, Oct4), mature pancreatic P-cell markers (insulin, glucagon, Glut2, Sur1, Kir6.1) and the GIP receptor gene GIP-R in undifferentiated (stage 1) cells, with increasing levels in differentiated stages 4 and 5. IAPP and somatostatin genes were only expressed in differentiated stages. lmmunohistochemical studies confirmed the presence of insulin, glucagon, somatostatin and IAPP in differentiated ES cells. After supplementation with GIP(LysPAL16), ES cells at stage 4 released insulin in response to secretagogues and glucose in a concentration-dependent manner, with 35-100% increases in insulin release. Cellular C-peptide content also increased by 45% at stages 4 and 5. We conclude that the stable GIP analogue enhanced differentiation of mouse ES cells towards a phenotype expressing specific P-cell genes and releasing insulin.

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T2 - Biological Chemistry

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SN - 1431-6730

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