Effect and mechanism of conditioned medium from bone marrow-derived mesenchymal stem cells on the survival of cortical neurons followed oxygen－glucose deprivation injury

Abstract:

Objective：To explore the effect and mechanism of conditioned medium from bone marrow-derived mesenchymal stem cells（BMSCs） on the survival of neurons after oxygen-glucose deprivation（OGD） injury. Methods：BMSCs and neurons were respectively cultured from 6 to 8 weeks（60 to 80 g） or pregnant 16 to 18 days Sprague-Dawley rats. After culturing for 5 days，primary cultured neurons were collected to build the model of OGD. Then experiments were randomly divided into four groups，including OGD group，OGD+CM group（the medium of neuron was replaced by culture medium of BMSCs after oxygen-glucose deprivation），OGD+CM+LY294002 group，OGD+LY294002 group（inhibitor LY294002 of PI3K/AKT signaling pathways were added to the above of two groups）. Morphology of neurons were observed by electron and light microscope after culturing for 12 h later in 4 groups. All the groups of neurons were cultured for 2 h later，and the protein levels of active Casepase-3 and pAKT（Ser473） were detected by Western bolt. After 48 h later，the survival rate of neurons was analyzed. Results：The neurite of neurons followed OGD injury were destructive and partially disappeared. Organelle of neurons，such as endoplasmic reticulum and mitochondrion，were swelled. In OGD+LY294002 group，the survival rate of neurons and expression level of pAKT（Ser473） were decreased（t=3.679，P=0.021；t=2.938，P=0.042），while the level of active Casepase-3 was increased（t=4.733，P=0.009） compared with those in OGD+CM group. The survival rate of neurons and expression level of pAKT（Ser473） were increased（t=6.630，P=0.003；t=3.288，P=0.030），while the level of active Casepase-3 was decreased（t=3.454，P=0.026） in OGD+CM group than in OGD+LY294002 group. In comparison with OGD+CM，the survival rate and level of pAKT（Ser473） were decreased（t=14.255，P=0.000；t=3.872，P=0.018） in OGD+CM+LY294002 group，while level of active Casepase-3 was increased（t=6.699，P=0.003）. Conclusion：BMSCs promote the survival of neurons after OGD damage，and its mechanisms may be related to the activation of PI3K/AKT protein signaling pathway.