Enhanced gene transfer to pancreatic islets using glucagon-like peptide-1

byungwan lee, M. H. Kim, H. Y. Chae, H. J. Hwang, D. Kang, S. H. Ihm

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Objective: The efficient transfer of genes into intact islets is difficult since islets exist as clusters of differentiated cells with little replication potential. Cell proliferation in response to growth factors is known to be accompanied by loosening of cell-to-cell contacts and increasing paracellular permeability. In this study, we investigated whether gene delivery into intact islet cells was facilitated by modulating β-cell proliferation. Methods: Isolated rat islets were pretreated with glucagon-like peptide (GLP)-1 or human growth hormone for 24 hours, or with 300 mg/dL of glucose for 48 hours before transduction with a suboptimal dose of recombinant adenoviral vector expressing green fluorescent protein (GFP) and β-galactosidase (multiplicity of infection of 25). Transduction efficiency was assessed by measuring β-galactosidase activity and GFP expression using enzyme-linked immunosorbent assay, flow cytometry, and fluorescence microscopy. The numbers of 7-aminoactinomycin D-positive dead cells and 5-ethynyl-2-deoxyuridine (EdU)-positive proliferating cells were also monitored using flow cytometry and fluorescence microscopy. Results: The transduction efficiency of rat islet cells by a suboptimal dose of viral vector was significantly improved by GLP-1 pretreatment, accompanied by enhanced cell viability and cell proliferation. An increased GFP expression in islet cells after GLP-1 pretreatment was observed among the increased numbers of EdU-positive proliferating cells. Conclusion: Pretreatment of rat islets with GLP-1 enhanced the transduction efficiency of an adenoviral vector, reducing viral dose burden while improving islet cell viability. From a therapeutic standpoint, genetic modification of pancreatic islets combined with GLP-1 pretreatment may be a promising option for ex vivo gene therapy prior to islet transplantation.

Original languageEnglish
Pages (from-to)591-596
Number of pages6
JournalTransplantation Proceedings
Volume45
Issue number2
DOIs
Publication statusPublished - 2013 Mar 1

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Glucagon-Like Peptide 1
Islets of Langerhans
Green Fluorescent Proteins
Galactosidases
Genes
Cell Proliferation
Fluorescence Microscopy
Cell Survival
Flow Cytometry
Islets of Langerhans Transplantation
Deoxyuridine
Human Growth Hormone
Viral Load
Genetic Therapy
Permeability
Intercellular Signaling Peptides and Proteins
Enzyme-Linked Immunosorbent Assay
Glucose
Infection

All Science Journal Classification (ASJC) codes

  • Surgery
  • Transplantation

Cite this

lee, byungwan ; Kim, M. H. ; Chae, H. Y. ; Hwang, H. J. ; Kang, D. ; Ihm, S. H. / Enhanced gene transfer to pancreatic islets using glucagon-like peptide-1. In: Transplantation Proceedings. 2013 ; Vol. 45, No. 2. pp. 591-596.
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abstract = "Objective: The efficient transfer of genes into intact islets is difficult since islets exist as clusters of differentiated cells with little replication potential. Cell proliferation in response to growth factors is known to be accompanied by loosening of cell-to-cell contacts and increasing paracellular permeability. In this study, we investigated whether gene delivery into intact islet cells was facilitated by modulating β-cell proliferation. Methods: Isolated rat islets were pretreated with glucagon-like peptide (GLP)-1 or human growth hormone for 24 hours, or with 300 mg/dL of glucose for 48 hours before transduction with a suboptimal dose of recombinant adenoviral vector expressing green fluorescent protein (GFP) and β-galactosidase (multiplicity of infection of 25). Transduction efficiency was assessed by measuring β-galactosidase activity and GFP expression using enzyme-linked immunosorbent assay, flow cytometry, and fluorescence microscopy. The numbers of 7-aminoactinomycin D-positive dead cells and 5-ethynyl-2-deoxyuridine (EdU)-positive proliferating cells were also monitored using flow cytometry and fluorescence microscopy. Results: The transduction efficiency of rat islet cells by a suboptimal dose of viral vector was significantly improved by GLP-1 pretreatment, accompanied by enhanced cell viability and cell proliferation. An increased GFP expression in islet cells after GLP-1 pretreatment was observed among the increased numbers of EdU-positive proliferating cells. Conclusion: Pretreatment of rat islets with GLP-1 enhanced the transduction efficiency of an adenoviral vector, reducing viral dose burden while improving islet cell viability. From a therapeutic standpoint, genetic modification of pancreatic islets combined with GLP-1 pretreatment may be a promising option for ex vivo gene therapy prior to islet transplantation.",
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Enhanced gene transfer to pancreatic islets using glucagon-like peptide-1. / lee, byungwan; Kim, M. H.; Chae, H. Y.; Hwang, H. J.; Kang, D.; Ihm, S. H.

In: Transplantation Proceedings, Vol. 45, No. 2, 01.03.2013, p. 591-596.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Enhanced gene transfer to pancreatic islets using glucagon-like peptide-1

AU - lee, byungwan

AU - Kim, M. H.

AU - Chae, H. Y.

AU - Hwang, H. J.

AU - Kang, D.

AU - Ihm, S. H.

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N2 - Objective: The efficient transfer of genes into intact islets is difficult since islets exist as clusters of differentiated cells with little replication potential. Cell proliferation in response to growth factors is known to be accompanied by loosening of cell-to-cell contacts and increasing paracellular permeability. In this study, we investigated whether gene delivery into intact islet cells was facilitated by modulating β-cell proliferation. Methods: Isolated rat islets were pretreated with glucagon-like peptide (GLP)-1 or human growth hormone for 24 hours, or with 300 mg/dL of glucose for 48 hours before transduction with a suboptimal dose of recombinant adenoviral vector expressing green fluorescent protein (GFP) and β-galactosidase (multiplicity of infection of 25). Transduction efficiency was assessed by measuring β-galactosidase activity and GFP expression using enzyme-linked immunosorbent assay, flow cytometry, and fluorescence microscopy. The numbers of 7-aminoactinomycin D-positive dead cells and 5-ethynyl-2-deoxyuridine (EdU)-positive proliferating cells were also monitored using flow cytometry and fluorescence microscopy. Results: The transduction efficiency of rat islet cells by a suboptimal dose of viral vector was significantly improved by GLP-1 pretreatment, accompanied by enhanced cell viability and cell proliferation. An increased GFP expression in islet cells after GLP-1 pretreatment was observed among the increased numbers of EdU-positive proliferating cells. Conclusion: Pretreatment of rat islets with GLP-1 enhanced the transduction efficiency of an adenoviral vector, reducing viral dose burden while improving islet cell viability. From a therapeutic standpoint, genetic modification of pancreatic islets combined with GLP-1 pretreatment may be a promising option for ex vivo gene therapy prior to islet transplantation.

AB - Objective: The efficient transfer of genes into intact islets is difficult since islets exist as clusters of differentiated cells with little replication potential. Cell proliferation in response to growth factors is known to be accompanied by loosening of cell-to-cell contacts and increasing paracellular permeability. In this study, we investigated whether gene delivery into intact islet cells was facilitated by modulating β-cell proliferation. Methods: Isolated rat islets were pretreated with glucagon-like peptide (GLP)-1 or human growth hormone for 24 hours, or with 300 mg/dL of glucose for 48 hours before transduction with a suboptimal dose of recombinant adenoviral vector expressing green fluorescent protein (GFP) and β-galactosidase (multiplicity of infection of 25). Transduction efficiency was assessed by measuring β-galactosidase activity and GFP expression using enzyme-linked immunosorbent assay, flow cytometry, and fluorescence microscopy. The numbers of 7-aminoactinomycin D-positive dead cells and 5-ethynyl-2-deoxyuridine (EdU)-positive proliferating cells were also monitored using flow cytometry and fluorescence microscopy. Results: The transduction efficiency of rat islet cells by a suboptimal dose of viral vector was significantly improved by GLP-1 pretreatment, accompanied by enhanced cell viability and cell proliferation. An increased GFP expression in islet cells after GLP-1 pretreatment was observed among the increased numbers of EdU-positive proliferating cells. Conclusion: Pretreatment of rat islets with GLP-1 enhanced the transduction efficiency of an adenoviral vector, reducing viral dose burden while improving islet cell viability. From a therapeutic standpoint, genetic modification of pancreatic islets combined with GLP-1 pretreatment may be a promising option for ex vivo gene therapy prior to islet transplantation.

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