Increased availability of fatty acids causes cell death and dysfunction in beta-cell lines, isolated islets, and animal models of diabetes. From the MIN6 beta-cell line, we selected two subpools that are resistant to palmitate-induced apoptosis. Protection was not universal because palmitate-resistant cells remained sensitive to cytokine- and streptozotocin-induced apoptosis. Palmitate oxidation and incorporation into cholesterol ester (but not triglycerides) were significantly higher in palmitate-resistant cells than in control cells. Consistent with these findings, transcript profiling revealed increased expression in palmitate-resistant cells of several beta-oxidation genes as well as a 2.8-fold upregulation of stearoyl-CoA desaturase 1 (SCD1). Correspondingly, the oleate-to-palmitate ratio of palmitate-resistant cells was double that of palmitate-pretreated control cells. At least some of this additional oleate in palmitate-resistant cells was incorporated into cholesterol ester stored in the form of large cytosolic lipid bodies. However, blocking cholesterol ester formation did not render palmitate-resistant cells sensitive to palmitate-induced apoptosis. On the other hand, an inhibitor of SCD1, 10,12-conjugated linoleic acid, dose dependently overcame the resistance of palmitate-resistant cells to lipoapoptosis. Our results suggest that desaturation per se is more important in protecting beta-cells from the cytotoxic effects of palmitate than is the nature of neutral lipid storage pool thus generated.