| Caspase-3 gene silencing for inhibiting apoptosis in insulinoma cells and human islets. | |
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MedLine Citation:
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PMID: 18828606 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Although islet transplantation has great potential to treat type I diabetes, most islet grafts do not function due to the host immune rejection, nonspecific inflammatory response and poor revascularization. Since caspase-3 plays a crucial role in apoptosis of transplanted islet cells, we used chemically synthesized small interfering RNAs (siRNAs) to silence caspase-3 in insulinoma (INS-1E) cells and human islets, and then determined whether caspase-3 gene silencing can prevent these cells from cytokine-induced apoptosis. Transfection of INS-1E cells and islets with siRNAs reduced caspase-3 transcripts by 50-67% and 50%, respectively. Additionally, apoptosis in transfected insulinoma cells was markedly inhibited. Since gene silencing did not last beyond two days, we converted potent siRNA into shRNA and constructed replication deficient adenoviral (Adv) vectors encoding these shRNAs driven by a U6 or H1 promoter. Compared to chemically synthesized siRNA, Adv-caspase-3-shRNA efficiently transduced islets, showed relatively higher and prolonged levels of gene silencing beyond five days, with higher gene silencing with a U6 promoter, and protected islets from cytokine-induced apoptosis. Finally, return to normoglycemia was achieved at 1 day post-transplantation of Adv-caspase-3-shRNA transduced islets under the kidney capsules of streptozotocin induced nonobese diabetic-severe combined immunodeficiency (NOD-SCID) mice and maintained beyond two weeks. Blood glucose levels returned to > or = 325 mg/dL upon removal of the islet graft-bearing kidney at 32 days after transplantation, confirming that transplanted islets were functional. |
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Authors:
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Guofeng Cheng; Lin Zhu; Ram I Mahato |
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Publication Detail:
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Type: Journal Article; Research Support, N.I.H., Extramural |
Journal Detail:
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Title: Molecular pharmaceutics Volume: 5 ISSN: 1543-8384 ISO Abbreviation: Mol. Pharm. Publication Date: 2008 Nov-Dec |
Date Detail:
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Created Date: 2009-05-12 Completed Date: 2009-06-11 Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 101197791 Medline TA: Mol Pharm Country: United States |
Other Details:
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Languages: eng Pagination: 1093-102 Citation Subset: IM |
Affiliation:
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Department of Pharmaceutical Sciences, University of Tennessee Health Sciences Center, Memphis, Tennessee 38103, USA. cheng_guofeng@yahoo.com |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Adenoviridae
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genetics Animals Apoptosis / genetics* Blood Glucose / genetics Caspase 3 / genetics* Cell Line, Tumor Diabetes Mellitus, Experimental / genetics Gene Silencing* Genetic Vectors Humans In Situ Nick-End Labeling Insulinoma / genetics* Islets of Langerhans / cytology, metabolism* Islets of Langerhans Transplantation / methods Mice Mice, Inbred NOD Mice, SCID Promoter Regions, Genetic RNA, Small Interfering / genetics Transfection |
| Grant Support | |
ID/Acronym/Agency:
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1R01DK69968/DK/NIDDK NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Blood Glucose; 0/RNA, Small Interfering; EC 3.4.22.-/Caspase 3 |
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