Pro- and Antiapoptotic Proteins Regulate Apoptosis but Do Not Protect Against Cytokine-Mediated Cytotoxicity in Rat Islets and β-Cell Lines

• DOI: 10.2337/db05-1000
• Publication Date: 2006-05-10T22:48:46Z
• Authors: J. Jason Collier; Patrick T. Fueger; Hans E. Hohmeier; Christopher B. Ne...
• Keywords: 0301 basic medicine; Cell Survival; Apoptosis; Transfection; Cell Line; Rats; 3. Good health; Interferon-gamma; Islets of Langerhans; 03 medical and health sciences; bcl-2 Homologous Antagonist-Killer Protein; Gene Expression Regulation; Animals; Cytokines; RNA, Small Interfering; Proto-Oncogene Proteins c-akt; Cells, Cultured; Interleukin-1; bcl-2-Associated X Protein

Type 1 diabetes results from islet β-cell death and dysfunction induced by an autoimmune mechanism. Proinflammatory cytokines such as interleukin-1β and γ-interferon are mediators of this β-cell cytotoxicity, but the mechanism by which damage occurs is not well understood. In the current study, we present multiple lines of evidence supporting the conclusion that cytokine-induced killing of rat β-cells occurs predominantly by a nonapoptotic mechanism, including the following: 1) A rat β-cell line selected for resistance to cytokine-induced cytotoxicity (833/15) is equally sensitive to killing by the apoptosis-inducing agents camptothecin and etoposide as a cytokine-sensitive cell line (832/13). 2) Overexpression of a constitutively active form of the antiapoptotic protein kinase Akt1 in 832/13 cells provides significant protection against cell killing induced by camptothecin and etoposide but no protection against cytokine-mediated damage. 3) Small interfering RNA–mediated suppression of the proapoptotic protein Bax enhances viability of 832/13 cells upon exposure to the known apoptosis-inducing drugs but not the inflammatory cytokines. 4) Exposure of primary rat islets or 832/13 cells to the inflammatory cytokines causes cell death as evidenced by the release of adenylate kinase activity into the cell medium, with no attendant increase in caspase 3 activation or annexin V staining. In contrast, camptothecin- and etoposide-induced killing is associated with robust increases in caspase 3 activation and annexin V staining. 5) Camptothecin increases cellular ATP levels, whereas inflammatory cytokines lower ATP levels in both β-cell lines and primary islets. We conclude that proinflammatory cytokines cause β-cell cytotoxicity primarily through a nonapoptotic mechanism linked to a decline in ATP levels.