Sulforaphane neuroprotective effects against methylglyoxal-induced toxicity in SH-SY5Y cells

Methylglyoxal (MG) is a reactive dicarbonyl compound precursor of advanced glycation end products (AGEs) and is found in high levels in blood or tissues of diabetic models. MG has been correlated with diabetic complications such as macrovascular disease, retinopathy, nephropathy, aging, neuropathy and neurodegenerative conditions such as Alzheimer's disease (1). Neurodegeneration involves several mechanisms including mitochondrial dysfunction, abnormal protein aggregation and inflammation (2). MG is efficiently metabolized by the cytosolic glyoxalase system that is composed of two enzymes glyoxalase (GLO) 1 and GLO2 and a catalytic amount of GSH (3). Isothiocyanates are metabolites derived from the hydrolysis of glucosinolates, a class of compounds present in plants of the Brassica family. In particular, sulforaphane (SF), an isothiocyanate produced from glucoraphanin by the enzyme myrosinase, is known for its chemopreventive, anti-inflammatory, cardioprotective and neuroprotective actions (4,5,6). Aim of this study was to investigate the effects of SF on the interrelated mechanisms of biological glycation and MG-induced death in neuroblastoma SH-SY5Y cells. In particular, we focused on the signaling pathways of apoptosis and the modulation of the GLO system. Cell viability was evaluated by MTT and LDH assays, apoptotic cell death by measuring caspase 3 activity, ROS production by the 2',7'-dichlorofluorescein-diacetate assay, GSH levels by the monochlorobimane assay, MAPK activation and GLO1 expression by immunoblotting, GLO1 activity spectrophotometrically. MG induced neuronal cell apoptosis via activation of caspase 3 and pro-apoptotic MAPK-signalling pathways (JNK and p38) and reduced the activation of the pro-survival kinase Akt. Moreover MG significantly increased intracellular ROS production and decreased GSH levels. SF treatment significantly counteracted cell death and apoptosis induced by MG as measured by MTT, LDH and caspase 3 activity assays. SF partially inhibited MG induced activation of p38 MAPK and activated the pro-survival kinase Akt, whereas it did not show any effect on JNK activation. SF treatment reduced MG induced oxidative stress decreasing intracellular ROS production and increasing GSH levels. Moreover, for the first time, we demonstrated that SF is able to significantly increase GLO1 protein expression and activity, eliciting a fundamental role in MG detoxification. These findings suggest that SF possesses cytoprotective ability in the prevention of MG induced glycation involved in neurodegenerative complications of diabetic patients.