Metallothionein-III (MT-III), a metal-binding protein, is associated with resistance to neuronal injury. However, the underlying mechanism for its effects remains unclear. We therefore examined whether MT-III can induce VEGF expression and promote neuroprotective effects in brain endothelial bEND.3 cells. MT-III significantly induced VEGF mRNA and protein expression in bEND.3 cells in a dose- and time-dependent manner. Furthermore, MT-III treatment increased the stability of hypoxia-inducible factor lot (HIF-1 alpha) and stimulated transcription of a reporter gene under control of the VEGF promoter. MT-III also increased the accumulation of HIF-1 alpha in nuclei and increased HIF-1 alpha-binding to the VEGF promoter. MT-III increased PI3K/Akt and ERK1/2 phosphorylation according to Western blot analysis. However, pretreatment with PD98059 and LY294002 (ERK1/2 and Akt inhibitors) inhibited MT-III-induced stimulation of HIF-1 alpha protein expression and VEGF production. These results suggest that MT-III upregulates VEGF production in brain endothelial cells by a HIF-1 alpha-dependent mechanism. (c) 2008 Elsevier Inc. All rights reserved.