Objective: Sjögren's syndrome (SS) is a chronic inflammatory disease characterized by lymphocytic infiltration and destruction of the salivary gland. Our preliminary data have demonstrated that apoptosis may play a role in the etiology of SS. We hypothesize that such autoantigens as α-fodrin we have shown result from apoptosis may be associated with the immune response that leads to the destruction of the salivary glands. The objective of this study is to induce apoptosis via two different pathways and evaluate the presence of cleaved autoantigen on the cell surface resulting from this stimulation. We predict that antigens resulting from apoptosis will be pathway-dependent and that activated caspase 3 and cleaved α-fodrin will be associated with stimulation by TNF-α but not Staurosporine which activates the mitochondrial pathway. Methods: Using our salivary gland acinar (NS-SV-AC) and ductal (HSG) cell lines, we stimulated apoptosis by treating them separately with TNF-α and Staurosporine (Sta). Apoptotic forms were detected and photographed using Scanning Electron Microscopy and were stained using immunofluorescence and an antibody specific for the cleaved α-fodrin autoantigen fragment. Results: our results show that HSG ductal cells and acinar cells produce unique characteristic forms of apoptotic bodies as a result of TNF-α stimulation. These characteristic morphologies are readily distinguished from those produced during apoptosis using Staurosporine stimulation. These unique forms of apoptotic bodies are positively stained using immunofluorescence and an antibody specific for the cleaved fragment of α-fodrin. In both cases of TNF-α stimulation the apoptotic blebs stay aggregated together while when Staurosporine is used, they become dispersed. Conclusion: these results show that these unique characteristic forms of apoptotic bodies are autoantigenic since they present cleaved α-fodrin on their surface. These findings provide further support for our hypothesis that apoptotic antigens may elicit autoantibody formation in vivo.

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