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In vivo study of the mitochondrial dysfunction during ischemia and the effect of oxidative stress on cell proliferation Liu, Ran

Abstract

Ion influx and water imbalance are major causes of injury during ischemia. Knowledge of the instantaneous subcellular structural and functional changes occurring in vivo, both during ischemia and immediately after the onset of reperfusion, have not been well characterized, mainly due to the extremely rapid progression of these events. To better understand the mechanisms underlying injury during ischemia in vivo, here, we examine mitochondrial function using the bilateral common carotid artery occlusion model of stroke. Mitochondrial membrane potential (Ym) was examined using two-photon fluorescence imaging of the dye Rhodamine123, as an indicator of mitochondrial function. We demonstrate that mitochondrial permeability transition pore-induced Ym collapse occurs during ischemia concurrently with plasma membrane potential depolarization, and repolarized rapidly during reperfusion. Furthermore, we show that inhibition of Ym collapse with cyclosporine A does not result in any detectable attenuation of dendritic structural damage–either during the stroke event or 2 hours afterwards. Thus, these data suggest that mitochondrial dysfunction is an early event during stroke and could contribute to delayed injury at later time points. Oxidative stress is another proposed mechanism of ischemic injury, given that anti-oxidant proteins play a vital role in brain cell survival. To study the chronic effect of elevated oxidative stress in vivo, we examined cell proliferation within neurogenic regions of the brain of adult mice with compromised anti-oxidant defences (Sut mice). Sut mice possess a natural truncation mutation in the gene Slc7a11 resulting in a malfunctional cystine/glutamate exchanger (xCT). Under normal conditions, xCT supply intracellular cyst(e)ine for the production of glutathione, a major cellular anti-oxidant. Using bromodeoxyuridine labelling as an indicator of newborn cells, we found that the rate of subventricular-zone (SVZ) cell proliferation when normalized to tissue area was comparable between Sut and control mice. However, the cell proliferation rate within the denate gyrus (DG) was elevated in Sut mice. These results demonstrate that xCT expression plays a role in regulating cellular proliferation in the DG, but not the SVZ of adult mice. Futhermore, our in vivo observations clearly indicate that in the absence of xCT ongoing cellular proliferation can still persist.

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