Investigation of the effects of resveratrol on in vitro endothelial cell injury induced by hypoxia and reoxygenation and of the possible role of nrf2

Introduction and aim of study: The ischemic diseases of the central nervous system witness several biochemical and functional changes in the endothelial cells as well as in the parenchyma. The aim of this investigation is to investigate whether resveratrol protects human cerebral microvascular endothelial cells (HCMVE) against oxidative injury exerted by an in vitro model of ischemia-reperfusion: ?oxygen- glucose deprivation (OGD) and reoxygenation + glucose supplementation (RGS)? and in case of positivity, to study further if this effect is mediated by Nrf2 (Nuclear transcription factor erythroid 2p45-related factor 2)Materials and methods: HCMVE cells were submitted to OGD (6 hours) + RGS (15, 60 min and 24 hr) and at the end of the incubation periods, the reactive oxygen species were assessed using the fluorescent probes, APF (3?- (p-aminophenyl) florescein) and HPF (3?- (p-hydroxyphenyl) florescein). The effects of resveratrol on the formation of reactive oxygen species were investigated by pre-incubation for one hour with different doses ofresveratrol (0.1, 1, 10, 50 ve 100 ? M). In order to assess the effect of resveratrol on Nrf2 activation, cells were incubated for different periods ( 1, 3, 6, 9, and 24 hours) with 10 ? M resveratrol.Results: The analysis with APF and HPF probes showed statistically significant increases in the 60-min RGE (p= 0.004 and 0.016, respectively), following the 6-hour OGD application to the HCMVE cells. At the 60th minute of RGS following 6-hour OGD, resveratrol significantly decreased (p: 0.004), the production or reactive oxygen species, at all doses applied. In addition, resveratrol increased, at 60 minutes, the Nrf2 expression- the Nrf2 band attenuated starting with the 3rd hour, while completely disappearing at the 9th hour.Conclusions: This study showed that resveratrol decreases the production of reactive oxygen species in the HCMVE cells submitted to OGD + RGS, under the condition studied. In addition, it has been shown for the first time that resveratrol increases the expression of Nrf2 protein, in the HCMVE cells, under these conditions.