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dc.contributor.authorGautam, Pratigyaen
dc.contributor.authorFoale, Robert Den
dc.contributor.authorRecino, Ashaen
dc.contributor.authorZhao, Jingen
dc.contributor.authorGan, Shu Uinen
dc.contributor.authorWallberg, Majaen
dc.contributor.authorCalne, Royen
dc.contributor.authorLever, Andrewen
dc.date.accessioned2016-10-20T11:35:32Z
dc.date.available2016-10-20T11:35:32Z
dc.date.issued2016-08-30en
dc.identifier.issn1099-498X
dc.identifier.urihttps://www.repository.cam.ac.uk/handle/1810/260839
dc.description.abstractBackground: The lack of an ideal cell type that can be easily acquired, modified to produce insulin, and re-implanted has been a limitation for ex vivo insulin gene therapy. Canine diabetes is currently treated with human insulin and is a good model for human diabetes. Mesenchymal stromal cells (MSCs) are a promising candidate cell type for gene therapy. Here we optimised insulin production using lentiviral transduced canine MSCs aiming to evaluate their ability for use as surrogate beta cells. Method: Canine MSCs were derived from bone marrow and validated by measuring expression of MSC lineage specific markers. Lentivirus vectors encoding the proinsulin gene (with or without a Kozak sequence) under the control of SFFV, CMV, EF1α and SV40 promotors were generated and used to transduce primary cMSCs and a hepatocyte cell line. The insulin producing capacity of transduced primary canine MSCs was assessed by measuring the concentration of C-peptide produced. Result: Primary canine MSC could be readily expanded in culture and efficiently transduced using lentiviral vectors encoding proinsulin. Increasing the multiplicity of infection from 3 to 20, led to an increase in C-peptide secretion (1700 pmol/l to 4000 pmol/l). The SFFV promoter conferred the strongest transcriptional ability. Conclusion: Our results suggest that optimised lentiviral transduction of the insulin gene into primary canine MSCs renders these cells capable of secreting insulin both short- and long-term, in sufficient quantities in vitro to support their potential use in insulin gene therapy.
dc.description.sponsorshipThe study was funded by the Lollipop Trust. Work in the laboratory is supported by the Biomedical Research Centre.
dc.languageEnglishen
dc.language.isoenen
dc.publisherWiley
dc.rightsAttribution-NonCommercial-NoDerivatives 4.0 International
dc.rightsAttribution-NonCommercial-NoDerivatives 4.0 Internationalen
dc.rightsAttribution-NonCommercial-NoDerivatives 4.0 Internationalen
dc.rightsAttribution-NonCommercial-NoDerivatives 4.0 Internationalen
dc.rights.urihttps://creativecommons.org/licenses/by-nc-nd/4.0/
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/en
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/en
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/en
dc.subjectinsulinen
dc.subjectgene therapyen
dc.subjectmesenchymal stromal cellsen
dc.subjectMSCsen
dc.subjectdiabetesen
dc.subjectlentivirusen
dc.titlePromoter Optimisation of Lentiviral Vectors for E fficient Insulin Gene Expression in Canine Mesenchymal Stromal Cells: Potential Surrogate Beta Cellsen
dc.typeArticle
dc.description.versionThis is the final version of the article. It first appeared from Wiley via https://doi.org/10.1002/jgm.2900en
prism.endingPage321
prism.publicationDate2016en
prism.publicationNameThe Journal of Gene Medicineen
prism.startingPage312
prism.volume18en
dc.identifier.doi10.17863/CAM.5982
dcterms.dateAccepted2016-08-25en
rioxxterms.versionofrecord10.1002/jgm.2900en
rioxxterms.versionAMen
rioxxterms.licenseref.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/en
rioxxterms.licenseref.startdate2016-08-30en
dc.identifier.eissn1521-2254
rioxxterms.typeJournal Article/Reviewen
pubs.funder-project-idMRC (MR/N022939/1)
pubs.funder-project-idMRC (G0800142)
pubs.funder-project-idBritish Heart Foundation (PG/14/16/30699)
pubs.funder-project-idNC3Rs (NC/M001083/1)


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Attribution-NonCommercial-NoDerivatives 4.0 International
Except where otherwise noted, this item's licence is described as Attribution-NonCommercial-NoDerivatives 4.0 International