Hyperoxia promotes astrocyte cell death after oxygen and glucose deprivation

Abstract

Astrocyte dysfunction and death accompany cerebral ischemia/reperfusion and possibly compromise neuronal survival. Animal studies indicate that neuronal death, neurologic injury, and oxidative molecular modifications are worse in animals exposed to hyperoxic compared to normoxic ventilation during reperfusion after global cerebral ischemia. It is unknown, however, whether ambient O 2 affects brain cell survival using in vitro ischemia paradigms where mechanisms of injury to specific cell types can be more thoroughly investigated. This study tested the hypothesis that compared with the supraphysiological level of 20% O 2 normally used in cell culture, lower, more physiological O 2 levels protect astrocytes from death following oxygen and glucose deprivation. Primary rat cortical astrocytes were cultured under either 7 or 20% O 2 , exposed to O 2 , and glucose deprivation for 4 h, and then exposed to normal medium under either 7 or 20% O 2 . Cell death and 3-nitrotyrosine and 8-hydroxy-2-deoxyguanosine immunoreactivities were assessed at different periods of reoxygenation. Astrocytes exposed to low levels of O 2 during reoxygenation undergo less death and exhibit lower levels of protein nitration and nucleic acid oxidation when compared with those under high levels of O 2 during reoxygenation. These results support the hypothesis that the 20% O 2 normally used in cell culture exacerbates astrocyte death and oxidative stress in an in vitro ischemia/reperfusion model compared to levels that more closely approximate those that exist in vivo.

Physiologic Oxygen Concentration Enhances the Stem-Like Properties of CD133+ Human Glioblastoma Cells In vitro

“…At this reduced O 2 level, the capacity of the CD133 + cells to differentiate along both the glial and neuronal pathways was also enhanced. A similar increase in neuronal differentiation has been reported for normal neural stem cells maintained at in vitro oxygen levels of 3% to 5% (19, 20, 34), as well as increased astrocytic death in primary rat cultures at 20% O 2 (36). Whether the enhancement of stem cell phenotype at 7% O 2 is a result of an increase in stem cell traits or a release of an inhibition of stem cell traits resulting from 20% O 2 remains to be determined.…”

Sulforaphane protects astrocytes against oxidative stress and delayed death caused by oxygen and glucose deprivation

“…Fluorescence immunocytochemistry was performed on cell cultures on coverslips as previously described (Danilov and Fiskum, 2008). After OGD and 4 or 48 hr REOX, the coverslips were rinsed twice with 0.05 M potassium phosphate buffered saline (KPBS), pH 7.4, and fixed with 4% PFA for 10 min, rinsed twice followed by incubation with 1% Sodium Borohydride in phosphate buffer for 20 min.…”

Primary culture of cellular subtypes from postnatal mouse for in vitro studies of oxygen glucose deprivation

“…In our relatively pure cultures of primary astrocytes, durations of OGD from 30 to 120 min did not result in significant apoptosis or loss of astrocytes. Other researchers utilized 4-24 h of OGD in experiments examining the effects of OGD on astrocytes (Papadopoulos et al, 1997;Giffard et al, 2004;Chu et al, 2008;Danilov and Fiskum, 2008;Danilov et al, 2009). However, these findings do not mean that astrocytes are unaffected by shorter durations of OGD.…”