ABSTRACT
Recently neuronal insulin resistance was suggested playing a role in Alzheimer's disease. Streptozotocin (STZ) is commonly used to induce impairment in insulin metabolism. In our previous work on undifferentiated SH-SY5Y cells the compound exerted cytotoxicity without altering insulin sensitivity. Nevertheless, differentiation of the cells to a more mature neuron-like phenotype may considerably affect the significance of insulin signaling and its sensitivity to STZ. We aimed at studying the influence of STZ treatment on insulin signaling in SH-SY5Y cells differentiated by retinoic acid (RA). Cytotoxicity of STZ or low serum (LS) condition and protective effect of insulin were compared in RA differentiated SH-SY5Y cells. The effect of insulin and an incretin analogue, exendin-4 on insulin signaling was also examined by assessing glycogen synthase kinase-3 (GSK-3) phosphorylation. STZ was found less cytotoxic in the differentiated cells compared to our previous results in undifferentiated SH-SY5Y cells. The cytoprotective concentration of insulin was similar in the STZ and LS groups. However, the right-shifted concentration-response curve of insulin induced GSK-3 phosphorylation in STZ-treated differentiated cells is suggestive of the development of insulin resistance that was further confirmed by the insulin potentiating effect of exendin-4. Differentiation reduced the sensitivity of SH-SY5Y cells for the non-specific cytotoxicity of STZ and enhanced the relative significance of development of insulin resistance. The differentiated cells thus serve as a better model for studying the role of insulin signaling in neuronal survival. However, direct cytotoxicity of STZ also contributes to the cell death.
Subject(s)
Cell Differentiation/physiology , Insulin Resistance/physiology , Insulin/pharmacology , Streptozocin/toxicity , Cell Differentiation/drug effects , Cell Line, Tumor , Cell Survival/drug effects , Exenatide/pharmacology , Glycogen Synthase Kinase 3/chemistry , Glycogen Synthase Kinase 3/metabolism , Humans , Phosphorylation/drug effects , Phosphorylation/physiology , Signal Transduction/drug effects , Tretinoin/pharmacologyABSTRACT
Recently, it is suggested that brain insulin resistance may contribute to the development of Alzheimer's disease; therefore, there is a high interest in its investigation. Streptozotocin (STZ) is often used to induce dysregulation of glucose and insulin metabolism in animal and cell culture models. Alteration in insulin sensitivity however, has not yet been assessed in neuronal cells after STZ treatment. We aimed at studying the concentration dependence of the protective effect of insulin on STZ-induced damage using SH-SY5Y cell line. Cells were treated with STZ and cell viability was assessed by resazurin reduction and lactate dehydrogenase release assays. Low serum (LS) medium was used as control damage. The effect of various concentrations (30, 100, 300, 1000 nM) of insulin was studied on cell viability and glycogen synthase kinase-3 (GSK-3) phosphorylation, an indicator of insulin signaling. STZ induced dose- and time-dependent cytotoxicity, its 1 mM concentration exerted a low, gradually developing damage. The cytoprotective effect of insulin was demonstrated in both STZ and LS groups. Its maximal effect was lower in the STZ-treated cells; however, its effective concentration remained largely unaltered. Insulin-induced GSK-3 phosphorylation was similar in the STZ- and LS-treated cells suggesting unchanged insulin signaling. Our present results indicate that STZ does not induce significant impairment in insulin sensitivity in SH-SY5Y cells, thus in this cell line it is not a good tool for studying the role of insulin resistance in neurodegeneration and to examine protective agents acting by improving insulin signaling.
