Fig. 7

Endogenous EPO produced by astrocytes under hypothermic conditions reduced the number of cleaved caspase-3-positive apoptotic neurons. Neurons were exposed to oxygen/glucose deprivation (OGD) for 2 h and reperfused in astrocyte-conditioned medium (ACM) from OGD-hypothermic astrocytes with either control or EPO-neutralizing IgG. Neurons were stained with the neuron cell marker MAP-2 (green) and the apoptosis marker cleaved caspase-3 (red). Nuclei were stained with 4,6-diamidino-2-phenylindole (DAPI; blue). a Representative immunofluorescence images of MAP-2 staining after 24 h of reperfusion. Apoptotic cells were identified by positive staining for cleaved caspase-3. Bar, 50 μm. b Quantification of neuron apoptosis after OGD-associated injury. Blockade of EPO signaling using an EPO-neutralizing antibody promoted neuron apoptosis with 24 h of reperfusion. The addition of EPO to non-conditioned culture medium at the same concentration as in hypothermic ACM (18 pg/mL) did not change the population of cleaved caspase-3-positive cells. Data are the mean ± SEM (n=6 fields in each group). ^*P < 0.05 compared with the hypothermic ACM/control IgG group