Apoptosis is not an invariable component of in vitro models of cortical cerebral ischaemia

Abstract

Characterising the mechanisms of cell death following focal cerebral ischaemia has been hampered by a lack of an in vitro assay emulating both the apoptotic and necrotic features observed in vivo. The present study systematically characterised oxygen-glucose-deprivation (OGD) in primary rat cortical neurones to establish a reproducible model with components of both cell-death endpoints. OGD induced a time-dependent reduction in cell viability, with 80% cell death occurring 24 h after 3 h exposure to 0% O 2 and 0.5 mM glucose. Indicative of a necrotic component to OGDinduced cell death, N-methyl-D-aspartate (NMDA) receptor inhibition with MK-801 attenuated neuronal loss by 60%. The lack of protection by the caspase inhibitors DEVD-CHO and z-VAD-fmk suggested that under these conditions neurones did not die by an apoptotic mechanism. Moderating the severity of the insult by decreasing OGD exposure to 60 min did not reduce the amount of necrosis, but did induce a small degree of apoptosis (a slight reduction in cell death was observed in the presence of 10 µM DEVD-CHO). In separate experiments purported to enhance the apoptotic component, cells were gradually deprived of O 2 , exposed to 4% O 2 (as opposed to 0%) during the OGD period, or maintained in serum-containing media throughout. While NMDA receptor antagonism significantly reduced cortical cell death under all conditions, a caspase-inhibitor sensitive component of cell death was not uncovered. These studies suggest that OGD of cultured cortical cells models the excitotoxic, but not the apoptotic component of cell death observed in vivo.

VEGF-Induced BBB Permeability is Associated with an MMP-9 Activity Increase in Cerebral ischemia: Both Effects Decreased by ANG-1

“…In contrast, a coadministration of Ang-1 and VEGF led to a decrease of vascular leakage ( Figure 5A). These results were confirmed using an in vitro model of BBB placed in OGD to mimic ischemia (Jones et al, 2004;Pellegrini-Giampietro et al, 1999;Wiessner et al, 2000). Blood-brain barrier perme- MMP-9, Ang-1, and VEGF in ischemic BBB damage S Valable et al ability was evaluated by 14 C-sucrose transport through the endothelial monolayer.…”

“…To mimic ischemic conditions, oxygen glucose deprivation (OGD) was used on the in vitro BBB model (Jones et al, 2004;Pellegrini-Giampietro et al, 1999;Wiessner et al, 2000). For OGD studies, Ringer-HEPES solution was first equilibrated with nitrogen and glucose was removed from the medium.…”

Soy phytoestrogens are neuroprotective against stroke-like injury in vitro

“…Using embryonic cortical cells enriched for neurons we demonstrate that low micromolar concentrations of soy phytoestrogens can be directly neuroprotective against many of the insults associated with ischemic injury. However, the OGD model results in both apoptosis and necrosis (Jones et al, 2004) and although our results demonstrate inhibition of caspase pathways, it is likely that necrotic mechanisms are influenced as well.…”

“…Oxygen-glucose deprivation (OGD) for 2 hours leads to both apoptotic and necrotic cell death (Jones et al, 2004). Twenty-four hours of pretreatment with 10 nM E2, or 1 μM, but not 0.1 μM, genistein, daidzein, or equol significantly reduced LDH release 48 hours after OGD (Figure 3).…”

Stem cell transplantation therapy for multifaceted therapeutic benefits after stroke

“…Cell death induced by different OGD episodes may be mediated or dominated by distinctive mechanisms, e.g. necrotic and apoptotic pathways (Jones et al , 2004)(Agudo-López et al , 2010). OGD can be used with different cell types, including ES cell-derived neural precursors and glial cells, to study their mechanisms of survival and differentiation under an ischemia-like condition.…”