Mechanism of Estrogen–mediated Cytoprotection Against Oxidative Insult: Increased Sequestration of Mitochondrial Calcium

Abstract

Abstract: : Purpose: Multiple defense mechanisms assemble at a point of regulation of mitochondrial control to provide a mitochondrial defense state which puts forward antioxidative cytoprotection against acute oxidative stress. These diverse systems involve a complex convergence of genomic and non–genomic associations; including, but not limited to, 17ß–estradiol (17ß–E₂)–induced activation of signal transduction survival pathways and regulation of mitochondrial calcium homeostasis so as to prevent opening of the mitochondrial permeability transition pore. Herein we illustrate how 17ß–E₂ is cytoprotective against oxidative insult which triggers excessive sustained increase in intracellular calcium ([Ca²⁺]_i). Methods: HLE–B3 cells were grown on 25mm cover slips and the cells preincubated with 17ß–E₂ (1 µM) or vehicle for 24 hrs prior to the bolus addition of 300 µM H₂O₂ and intracellular calcium movement monitored by Fura–2 imaging (n=70 cells per treatment). Results: 17ß–E₂ pretreatment of HLE–B3 cells lessened the peroxide–induced sustained [Ca²⁺]_i in spite of the fact that the hormone initially potentiates the influx of Ca²⁺ induced by peroxide.17ß–E₂–induced attenuation of [Ca²⁺]_i is mediated through the increased uptake and accumulation of Ca²⁺ into mitochondria. Similar results with 17α–E₂ indicate that the regulation of mitochondrial Ca²⁺ transport is an estrogen receptor–independent function. Mitochondrial sequestration of Ca²⁺ is blocked by the administration of antimycin, which nullifies mitochondrial membrane potential, thus preventing mitochondrial Ca²⁺ transport. Conclusions: The 17ß–E₂–induced attenuation of peroxide–induced sustained [Ca²⁺]_i puts forward a novel mechanism which confers cellular protection against oxidative stress through a process of increased mitochondrial sequestration of cytosolic Ca²⁺. Estradiols exert their protective effects (against oxidative stress), at least in part, by stimulating mitochondrial Ca²⁺ transport and adjusting, to a higher–level set point, mitochondrial tolerance for Ca²⁺ load, thereby preventing apoptotic and/or necrotic forms of cell death that are associated with mitochondrial dysfunction otherwise initiated by disproportionate Ca²⁺ overload. These data establish a new mechanistic role for antioxidants and identifies novel approaches for targeting mitochondrial function to reduce oxidative stress.

Keywords: calcium • mitochondria • antioxidants