Inhibition of Inflammatory Cytokine-Induced Response in Human Islet Cells by Withaferin A

BackgroundAfter islet cell transplantation, a substantial mass of islets are lost owing to nonspecific inflammatory reactions. Cytokine exposure before or after transplantation can upregulate expression of proinflammatory genes via the nuclear factor-κB signaling pathway, eventually resulting in islet loss.ObjectiveTo test the effects of a naturally occurring nuclear factor-κB inhibitor, withaferin A, on regulation of inflammatory genes in human islets.MethodsHuman pancreatic islets were isolated using a modified Ricordi protocol. Purified islets were cultured for 2 days. The effect of withaferin A treatment on islet cell viability was examined using the fluorescein diacetate-propidium iodide dye exclusion test, and on function using a static glucose stimulation assay. Islet cells were treated with a cytokine mixture (50 U/mL of interleukin-1β, 1000 U/mL of tumor necrosis factor-α, and 1000 U/mL of interferon-γ) for 48 hours with or without withaferin A, 1 μg/mL. Treated islets were used for real-time polymerase chain reaction (PCR) array analysis for expression of inflammatory genes, and expression of other selected genes was analyzed using real-time PCR with single primers.ResultsGlucose stimulation and viability assays demonstrated that withaferin A was not toxic to islet cells. Of 84 inflammation-related genes examined using real-time PCR array analysis, 9 were significantly upregulated by cytokine treatment compared with the control group. However, addition of withaferin A to the culture significantly inhibited expression of all genes.ConclusionWithaferin A significantly inhibits the inflammatory response of islet cells with cytokine exposure.