Tanshinone IIA pretreatment attenuates oxygen-glucose deprivation induced hippocampal neurons damage

Objective: Tanshinone IIA (Tan IIA) has been used in treatment of ischemic stroke in China. This study was designed to investigate the underlying neuroprotective mechanism of Tan IIA pretreatment in primary cultured rat hippocampal neurons incurred oxygen-glucose deprivation (OGD). Method: Hippocampal neurons was prepared from E18 embryonic rats and identified by immunocytochemical staining. After OGD and pretreatment with Tan IIA, cell viability was measured using MTT assay. Lactate dehydrogenase (LDH) activity, Caspase-3 activity and adenosine triphosphate (ATP) level were detected using assay kits. Cell apoptosis analysis was carried out with flow cytometry. Mitochondrial membrane potential and intracellular calcium were monitored by fluorospectrophotometer. Ubiquinol-cytochrome c reductase, Rieske iron-sulfur polypeptide 1 (UQCRFS1) expression was mesured by RT-PCR and Western blotting. Results: Hippocampal neuron were incurred OGD for 6 h, the cell survival was decreased to (61.5 ± 4.1)% comparing to the control group (set 100%). Pretreatment with Tan IIA at 3, 10, 30 and 100 mM, the cell survivals were (70.5 ± 2.1)%, (72.8 ± 4.4)%, (78.8 ± 3.4)% and (83.5 ± 3.1)% respectively and higher than that of only OGD-ed group. Exposure of the cells to OGD, cell apoptosis was induced, intercellular Caspase-3 activity was elevated, ATP level was decreased and LDH activity was increased in the culture supernatants. Pretreatment with Tan IIA alleviated all the changes. Furthermore, OGD may induce mitochondrial membrane potential decreasing and intracellular calcium level elevating in the neuron, Tan IIA pretreatment also alleviated these effects. When the cells were exposed to OGD, both mRNA and protein level of UQCRFS1 were decreased, Tan IIA pretreatment attenuated the changes caused by OGD. Conclusion: Tan IIA pretreatment can reduce energy damage of the neurons induced by OGD, which is associated with attenuating the changes in mitochondrial membrane potential and UQCRFS1 expression.