Abstract
Salidroside has been shown to exert neuroprotective effects against hypoxia. However, its mitochondrial protective mechanisms still remain elusive. The present study aimed to explore the mitochondrial protection of salidroside on PC12 cells and the involved mechanisms. The hypoxic injury of PC12 cells was triggered by CoCl2 stimulus. The contents of LDH release, SOD, GSH-PX, Na+-K+-ATPase, ATP, NAD+ and NADH were determined by using commercial biochemical kits. Clark-type oxygen electrode and Seahorse XFe24 analyzer were employed to evaluate cell respiration and measure oxygen consumption rate (OCR), respectively. Mitochondrial swelling and mitochondrial membrane potential (MMP) were measured by using isolated mitochondria from the brain tissue of mice. The proteins expression of cleaved Caspase-3, HIF-1α, ISCU1/2, COX10 and PFKP were tested by immunofluorescence and Western blot. While the genes expression of Caspase-3, HIF-1α, ISCU1/2, COX10 and miR-210 were tested by quantitative real-time PCR (qRT-PCR) analysis. Salidroside alleviated CoCl2-induced oxidative stress in PC12 cells as evidenced by increased cell viability, decreased LDH release and elevated GSH-PX and SOD activities. Salidroside could inhibit apoptosis by suppressing the level of cleaved Caspase-3 and Caspase-3. The enhanced mitochondrial energy synthesis by salidroside treatment was evidenced by the increases of Na+-K+-ATPase activity, ATP content, NAD+/NADH ratio, cellular respiration and OCR. In addition, salidroside could reduce mitochondrial swelling and MMP dissipation in isolated mitochondria. The results of immunofluorescence, Western blot and qRT-PCR analyses further revealed that salidroside raised the level of HIF-1α, ISCU1/2, COX10, and miR-210. Collectively, salidroside can reverse CoCl2-simulated hypoxia injury in PC12 cells partly by mitochondrial protection via inhibiting oxidative stress event, anti-apoptosis and enhancing mitochondrial energy synthesis.