Interferon-gamma induced cell death in a cultured human salivary gland cell line

Abstract

Increased levels of several cytokines, including interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha), have been demonstrated in the salivary gland microenvironment of patients with Sjögren's syndrome (SS). How these cytokines may be contributing to the pathogenesis of the disease is not well understood. This study examined the role of IFN-gamma +/- TNF-alpha on cellular death in a cultured human salivary gland cell line (HSG). Cells treated long-term with IFN-gamma +/- TNF-alpha demonstrate a profound antiproliferative effect with a decrease in cell number to below that initially plated. Treatment of HSG cells with TNF-alpha alone did not have any significant effects on growth but did increase the expression of the IFN-gamma receptor. Cells labelled with propidium iodide and anti-digoxigenin dUTP/dATP were examined by flow cytometry to determine the percentage of cells exhibiting low DNA content and DNA strand breaks. The percentage of cells exhibiting subdiploid DNA and DNA strand breaks increased with increased time of exposure to the cytokines. The maximum percentage of cells exhibiting DNA degradation at 12 days was 58% for cells treated with IFN-gamma + TNF-alpha, 31% for IFN-gamma treated cells, and < 5% for TNF-alpha-treated and untreated cells. The cells with subdiploid ( < 2n) DNA were subsequently demonstrated to represent two populations, both with evidence of increased DNA strand breaks but with differing light scatter characteristics. One population had features of cells undergoing necrosis, whereas the second population exhibited features of apoptosis. These findings were confirmed by transmission electron microscopy. Cells not exposed to cytokines did not exhibit significant evidence of either death process. We conclude that long-term exposure of a human salivary gland epithelial cell line to IFN-gamma +/- TNF-alpha leads to increased DNA degradation and subsequent cell death. This suggests a potential SS disease mechanism and implicates the role of the epithelial cell in this disease as an important area for future study.