The Effect of Therapeutic Mild Hypothermia on Brain Microvascular Endothelial Cells During Ischemia-Reperfusion Injury.

Abstract

To determine the cerebral protective effects of mild hypothermia (MH) on cerebral microcirculation. We established ischemia-reperfusion (I/R) injury and MH treatment models with rat brain microvascular endothelial cells (RBMECs) in vitro and examined the apoptotic changes. The cultured RBMECs were randomly divided into the control group, I/R group, and MH group, which was further divided into two subgroups: intra-ischemia hypothermia (IIH) and post-ischemia hypothermia (PIH). Cell morphological changes were assessed using fluorescence microscopy. Apoptotic rates were obtained by flow cytometry. Expressions of caspase-3, Bax, and Bcl-2 were analyzed by Western blot. I/R injury in vitro induced apoptosis of RBMECs. The apoptotic rates in the control group, I/R group, and MH group were 0.13, 19.04, and 13.13%, respectively (P<0.01). Compared with the I/R group, the MH group showed a significant decrease in the number of apoptotic cells, mainly in stage I apoptotic cells (P<0.0083). The caspase-3 and Bax expressions were significantly enhanced (P<0.05) in RBMECs after I/R injury, while substantial decreases in Bcl-2 expression were noted (P<0.05). Following MH intervention, the increase in caspase-3 and Bax expression was suppressed (P<0.05), while Bcl-2 expression significantly increased. The apoptotic rates or protein expressions between the two subgroups were not different significantly (P>0.05). These results indicate that MH could inhibit RBMEC apoptosis by preventing pro-apoptotic cells and early apoptotic cells from progressing to intermediate and advanced stages. This may be due to the effect of MH on I/R-induced apoptotic gene expression changes.