Gene silencing in the endocrine pancreas mediated by short-interfering RNA

Sean P. Bradley, Cristiana Rastellini, Marco A. Da Costa, Timothy F. Kowalik, Aaron B. Bloomenthal, Melissa Brown, Luca Cicalese, Giacomo P. Basadonna, Marc E. Uknis

Research output: Contribution to journalArticle

38 Citations (Scopus)

Abstract

Objectives: RNA interference as mediated by short-interfering RNA (siRNA) offers a nonviral means to silence genes in tissue; however, few data exist about gene therapy using siRNA in pancreas tissue. To determine if siRNA treatment could silence an endogenous gene in pancreatic islets, we developed a murine model using the endocrine pancreas. Methods: The insulin 2 (Ins2) gene was targeted with siRNA, and quantitative RT-PCR, fluorescent microscopy, and FACS were used to measure transcript levels and siRNA cellular uptake and transfection efficiency. Isolated pancreatic islets were transfected with siRNA in vitro using a liposomal delivery method in a dose titration (50-400 nM) or pooled from BALB/c mice having received siRNA (100 μg) via hydrodynamic tail vein injection. Results: The Ins2 transcript level was significantly reduced by 55% in vitro with FACS data showing a transfection efficiency over 45% with the 400 nM concentration. In vivo delivery of siRNA to pancreatic islets revealed a 33% reduction in Ins2 mRNA levels, although siRNA was able to be detected in 19% of isolated islet cells. Conclusion: We have successfully used RNA interference to silence an endogenous tissue-specific gene (Ins2) in pancreatic islets when transfected in vitro or administered in vivo.

Original languageEnglish (US)
Pages (from-to)373-379
Number of pages7
JournalPancreas
Volume31
Issue number4
DOIs
StatePublished - Nov 2005
Externally publishedYes

Fingerprint

Gene Silencing
Islets of Langerhans
Small Interfering RNA
Insulin
RNA Interference
Genes
Transfection
Hydrodynamics
Genetic Therapy
Tail
Pancreas
Microscopy
Veins
Polymerase Chain Reaction
Messenger RNA
Injections

Keywords

  • Endocrine pancreas
  • Gene silencing
  • Islets
  • RNA interference
  • siRNA

ASJC Scopus subject areas

  • Gastroenterology
  • Endocrinology

Cite this

Bradley, S. P., Rastellini, C., Da Costa, M. A., Kowalik, T. F., Bloomenthal, A. B., Brown, M., ... Uknis, M. E. (2005). Gene silencing in the endocrine pancreas mediated by short-interfering RNA. Pancreas, 31(4), 373-379. https://doi.org/10.1097/01.mpa.0000179730.69081.64

Gene silencing in the endocrine pancreas mediated by short-interfering RNA. / Bradley, Sean P.; Rastellini, Cristiana; Da Costa, Marco A.; Kowalik, Timothy F.; Bloomenthal, Aaron B.; Brown, Melissa; Cicalese, Luca; Basadonna, Giacomo P.; Uknis, Marc E.

In: Pancreas, Vol. 31, No. 4, 11.2005, p. 373-379.

Research output: Contribution to journalArticle

Bradley, SP, Rastellini, C, Da Costa, MA, Kowalik, TF, Bloomenthal, AB, Brown, M, Cicalese, L, Basadonna, GP & Uknis, ME 2005, 'Gene silencing in the endocrine pancreas mediated by short-interfering RNA', Pancreas, vol. 31, no. 4, pp. 373-379. https://doi.org/10.1097/01.mpa.0000179730.69081.64
Bradley SP, Rastellini C, Da Costa MA, Kowalik TF, Bloomenthal AB, Brown M et al. Gene silencing in the endocrine pancreas mediated by short-interfering RNA. Pancreas. 2005 Nov;31(4):373-379. https://doi.org/10.1097/01.mpa.0000179730.69081.64
Bradley, Sean P. ; Rastellini, Cristiana ; Da Costa, Marco A. ; Kowalik, Timothy F. ; Bloomenthal, Aaron B. ; Brown, Melissa ; Cicalese, Luca ; Basadonna, Giacomo P. ; Uknis, Marc E. / Gene silencing in the endocrine pancreas mediated by short-interfering RNA. In: Pancreas. 2005 ; Vol. 31, No. 4. pp. 373-379.
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