Organotellurane-Promoted Mitochondrial Permeability Transition Concomitant with Membrane Lipid Protection against Oxidation Felipe S. Pessoto, †,‡ Priscila A. Faria, ‡ Rodrigo L. O. R. Cunha, § João V. Comasseto, | Tiago Rodrigues, ‡ and Iseli L. Nantes* ,‡ Departamento de Biologia Molecular, UniVersidade Federal de São Paulo - UNIFESP, Brazil, Centro Interdisciplinar de InVestigação Bioquímica - CIIB, UniVersidade de Mogi das Cruzes - UMC, Brazil, Departamento de Biofísica, UniVersidade Federal de São Paulo - UNIFESP, Brazil, and Instituto de Química, UniVersidade de São Paulo - USP, Brazil ReceiVed March 27, 2007 Organotelluranes exhibit potent antioxidant properties as well as the ability to react with protein thiol groups and, thereby, they are good models to study the mechanism of the mitochondrial permeability transition (MPT). We evaluated the effects of the concentration of organotelluranes, namely RT-03 and RT-04, on rat liver mitochondria. At the concentration range of 0.25–1.0 μM, organotelluranes did not cause any mitochondrial dysfunction. At the concentration range of 5–10 μM, RT-03 and RT-04 caused the Ca 2+ -dependent opening of the (MPT) pore, regulated by Cyclosporin A. At the concentration range of 15–30 μM the swelling was not inhibited by Cyclosporin A and in the absence of Ca 2+ , a significant decrease of respiratory control ratio was observed due to concomitant phosphorylation impairment and uncoupling, transmembrane potential disruption, depletion of mitochondrial reduced thiol groups, and alterations in the bilayer fluidity. Above 100 μM, the organotelluranes caused complete inhibition of respiratory chain. Over the whole studied concentration range, RT-03 and RT-04 did not induce mitochondrial oxidative stress assessed by using the reactive oxygen and nitrogen species indicator 2′,7′- dichlorodihydrofluorescein diacetate. Further, the organotelluranes also exhibited protective effect against t-butyl hydroperoxide-induced oxidative stress as well as against Fe 2+ /citrate-induced peroxidation of mitochondrial membranes and PCPECL liposomes. These results point out that MPT pore opening can involve damage exclusively to mitochondrial membrane proteins. The exclusive antioxidant activity observed at nanomolar range is also an interesting new finding described in this work. Introduction The mitochondrial permeability transition (MPT) is a well- known event that can precede necrotic and apoptotic cell death (1–3). The onset of the MPT is initiated by the opening of the mitochondrial permeability transition pore (MPTP). MPTP is a high conductance pore in the inner mitochondrial membrane that can be opened by many agents such as Ca 2+ , Pi, alkaline pH, and reactive oxygen species (ROS), and prevented by the imunosuppressive drug cyclosporine A (CsA), Mg 2+ , acidic pH, and phospholipase inhibitors such as dibucaine, trifluoperazine, and quinacrine. MPTP opening leads to nonselective diffusion of solutes with molecular mass of up to 1500 Da through the inner mitochondrial membrane and promotes mitochondrial depolarization, uncoupling of oxidative phosphorylation and large amplitude swelling (4). These events, in turn, lead to ATP depletion and cell death. Although several proteins such as ADP/ ATP translocator (ANT), cyclophilin D, and possibly the voltage-dependent anion channel (VDAC) and hexokinase have been assigned as components of MPTP (1, 5, 6), the molecular composition of the MPTP remains uncertain. The prevailing model of pore structure does not explain the fact that the MPTP can exist in a regulated and unregulated form. The most incisive evidence against the hypothesis that the MPTP can be formed exclusively by the assembly of specific proteins is the finding that small exogenous pore-forming amphipathic peptides like mitochondrial targeting peptides, alamethicin, and mastoparan also induce onset of a Ca 2+ -dependent and CsA-inhibitable MPT at low concentrations and a CsA-insensitive MPT at higher concentrations (7–9). Considering that many agents that induce MPT such as ROS, oxidants, and thiol reactive agents exhibit the common ability to attack and modify membrane proteins, Lemasters and co-workers (9) proposed a model for the MPTP formation in which the chemical attack of mitochondrial membrane proteins leads to the misfolding and aggregation of these proteins to form aqueous channels. Initially, chaperone- like proteins including cyclophilin D regulate the conductance of solutes by the MPTP to confer the properties of regulated PT pores. Binding of chaperones to the misfolded proteins * To whom correspondence should be addressed: Iseli L. Nantes, Centro Interdisciplinar de Investigação Bioquímica - CIIB, Universidade de Mogi das Cruzes - UMC, Av. Dr C-andido Xavier de Almeida e Souza, 200, Mogi das Cruzes - SP, Brazil, CEP 08780-911. E-Mail: ilnantes@umc.br. † Departamento de Biologia Molecular, Universidade Federal de São Paulo-UNIFESP. ‡ Universidade de Mogi das Cruzes - UMC. § Departamento de Biofísica, Universidade Federal de São Paulo- UNIFESP. | Instituto de Química, Universidade de São Paulo-USP. Chart 1. Molecular Structure of Organotelluranes RT-03 and RT-04 Chem. Res. Toxicol. 2007, 20, 1453–1461 1453 10.1021/tx700092r CCC: $37.00  2007 American Chemical Society Published on Web 09/27/2007