Differential rAAV2 transduction efficiencies and insulin secretion profiles in pure and co-culture models of human enteroendocrine L-cells and enterocytes
✍ Scribed by Shiue-Cheng Tang; Athanassios Sambanis
- Book ID
- 102891313
- Publisher
- John Wiley and Sons
- Year
- 2004
- Tongue
- English
- Weight
- 620 KB
- Volume
- 6
- Category
- Article
- ISSN
- 1099-498X
- DOI
- 10.1002/jgm.587
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✦ Synopsis
Abstract
Background
Cell‐based therapies for treating insulin‐dependent diabetes (IDD) can provide a more physiologic regulation of blood glucose levels in a less invasive fashion than insulin injections. Previously, we developed an engineered human enteroendocrine L‐cell model for regulated insulin release via recombinant adeno‐associated virus serotype 2, or rAAV2, transduction. The aim of this study was to evaluate the efficiency and selectivity of rAAV2‐mediated insulin gene delivery to enteroendocrine L‐cells in co‐culture with a prevailing number of enterocytes, which are the predominant cell type in intestinal epithelium.
Methods
We tested rAAV2 transduction in pure and co‐culture models of human cell lines of enterocytes (Caco‐2 and T84 cell lines) and enteroendocrine L‐cells (NCI‐H716 cell line). Non‐viral, chemical‐mediated transfection was used as a control. Transduced and transfected co‐cultures were subjected to insulin secretion studies.
Results
In pure cultures, rAAV2 exhibited a low transduction efficiency towards both Caco‐2 and T84 enterocytes, as opposed to a strong reporter expression in permissive NCI‐H716 L‐cells. In co‐cultures of NCI‐H716 L‐cells and Caco‐2 or T84 enterocytes, rAAV2 exhibited differential transduction efficiency with a strong preference towards NCI‐H716 L‐cells. The rAAV2‐transduced co‐culture achieved regulated insulin release against stimulation, whereas the chemically transfected co‐culture failed to respond.
Conclusions
This study demonstrated that rAAV2‐mediated insulin gene transfer can differentiate human intestinal cell types in vitro, in particular enterocyte and enteroendocrine L‐cell lines. We consider the AAV2 vector a useful tool in developing enteroendocrine L‐cell‐specific insulin gene delivery for IDD treatment, in terms of AAV2 avoiding enterocytes and targeting selectively L‐cells. Copyright © 2004 John Wiley & Sons, Ltd.