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Updated: Jul 14, 2026
Isolated Pancreatic Islet Treatment and Apoptosis Measurement
Published on: May 2, 2025
Effect of iNOS and NF-kappaB gene silencing on beta-cell survival and function
Daniel De Paula1, M Vitória L B Bentley, Ram I Mahato
1Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Brazil.
Purpose:
Type I diabetes results from beta-cell death and dysfunction, induced by infiltration of immune cells and local production of inflammatory cytokines. Therefore, we investigated the effect of iNOS and NF-kappaB gene silencing on beta-cell survival and function.
Methods:
Rat insulinoma INS-1E cells were transfected with chemically synthesized siRNA after complex formation with Lipofectamine 2000. Cells were then treated with a cocktail of inflammatory cytokines (IL-1beta+ TNF-alpha+ IFN-alpha), and glucose stimulated-insulin response and viability were determined. iNOS and NF-kappaB gene expression was assessed at mRNA level by real time RT-PCR. The effect of gene silencing was also correlated with cytokine-induced NO production and apoptosis.
Results:
Transfection of INS-1E cells with siRNAs silenced iNOS and NF-kappaB gene expression and reduced NO production in a sequence-specific manner without causing significant loss of cell viability and function. However, the abrogation of NO production did not prevent INS-1E cells from cytokine-induced apoptosis, suggesting that this event may not be totally dependent on NO production.
Conclusion:
The gene silencing approach presented here is capable of attenuating the effects of inflammatory cytokines, such as iNOS expression and NO production and it will help to identify new target genes to improve islet transplantation.
