Delivery of TAT/PTD-fused proteins/peptides to islets via pancreatic duct

Dagmar Klein, Valeska Mendoza, Antonello Pileggi, R. Damaris Molano, Florencia M. Barbé-Tuana, Luca Inverardi, Camillo Ricordi, Ricardo L. Pastori

Research output: Contribution to journalArticle

14 Scopus citations

Abstract

Delivering cytoprotective proteins/peptides into pancreata prior to islet isolation through protein transduction (PT) is a novel strategy to enhance the yield of viable transplantable islets. Previous work has shown that the protein transduction domain PTD-5 efficiently transduced islets via the pancreatic duct. TAT/PTD is a well-characterized PTD with the capability to cross even the hemato-encephalic barrier. In this study, we investigated the utilization of the 11-aa TAT protein transduction domain (TAT/PTD) to deliver peptides or proteins of different sizes ranging from 1.2 to 120 kDa, as the TAT/PTD and TAT/PTD-BH4 peptide, or the TAT/PTD-β-galactosidase fusion protein, into islets through the pancreatic duct. Using flow cytometry analysis we found that TAT/PTD derivatives transduced practically 100% of the islet cell population. Moreover, confocal laser scanning microscopy in live, nonfixed islets confirmed these results assessing transduction of TAT/PTD molecules into intact nondisaggregated islets. TAT-β-galactosidase peptide conjugated to FITC was not compartment selective, as both cytoplasmic and nucleic cellular compartments were positively stained. Furthermore, TAT-β-galactosidase peptide delivery was highly effective, as even cells located in the inner core region of the islets were transduced. Finally, transduced TAT-β-galactosidase fusion protein was biologically active after islet isolation and manipulation, and islet insulin secretion capability was not compromised by peptide transduction. These findings suggest that the transduction of chimeric TAT/PTD proteins can represent an efficient tool of molecular delivery independent of the size, to enhance or modify a specific phenotype at the nuclei or cytoplasmic level.

Original languageEnglish (US)
Pages (from-to)241-248
Number of pages8
JournalCell Transplantation
Volume14
Issue number5
DOIs
StatePublished - Jan 1 2005

    Fingerprint

Keywords

  • Islet
  • Pancreatic duct
  • Protein transduction
  • Transplantation

ASJC Scopus subject areas

  • Cell Biology
  • Transplantation

Cite this

Klein, D., Mendoza, V., Pileggi, A., Molano, R. D., Barbé-Tuana, F. M., Inverardi, L., Ricordi, C., & Pastori, R. L. (2005). Delivery of TAT/PTD-fused proteins/peptides to islets via pancreatic duct. Cell Transplantation, 14(5), 241-248. https://doi.org/10.3727/000000005783983016