Protective Effect Of Pyrroloquinoline Quinine Against Oxygen And Glucose Deprivation Injury In Nuro2A Cells

Objective: Stroke is a life-threatening disease characterized by rapidly developing clinical signs of focal or global disturbance of cerebral function due to cerebral ischemia. The physiopathologic mechanisms of cerebral ischemia reperfusion injury were extremely complex. Multiple pathways were involved in the ischemic process that ultimately leads to cell death. In particular, the neuron apoptosis after ischemia/reperfu- sion is one of the major pathways that leads to the process of cell death .Therefore ,prot- ectting neurons from apoptosis may be beneficial to the therapy of ischemic disease. Pyrroloquinoline quinine(PQQ),which is an essential nutrient, has been demonstrated to act as a strong antioxidant , and thought to be have the function against cerebral ischemia by inhibting the oxidative stress. For thus ,we hypothesized that PQQ might have a neuroprotective effect via modulation of multiple pathways associated with apoptosis.To investigate the neuroprotective effects of PQQ against oxygen and glucose deprivation (OGD) in cultured rat neuroblastoma cells Neuro2A and the possible mechanisms involved.Methods: Cultured rat neuroblastoma cells Neuro2A were pretreated or not pretreated with increasingly concentrations of PQQ , exposed to 2 hours combined OGD in an anaerobic chamber followed by reoxygenation of 6 hours .Cellular morphology was observed by inverted phase contract microscope. Assessment of cell viability was quantitatively performed by the reduction of 3-(4,5-Dimethylthialzol-2-yl) -2,5-diphenyl-tetrazolium bromide (MTT) at reoxygenation 6 hours after OGD, and the percentage of apoptotic cells was tested by flow cytometry and Hoechst 33342 staining. The contents of intracellular ROS and GSH level were measured with fluorospectrophotometer .Results: 1. After OGD treating in cortical neurons for 2 hours and reoxygenation treating for 6 hours the swelling of neuronal body were significant ,while PQQ could lessen the swelling of neuronal body. 2. PQQ(0.4,0.8,1.6,3.2,6.4,12.8μM)group increased the cell viability following OGD/reoxygenation, and, the maximal neuroprotection was afforded by 12.8μM PQQ ,while the toxic dose of PQQ appeared at 25.6μM. 3. After OGD treating in Neuro2A cells for 2 hours and reoxygenation treating for 6 hours the rate of apoptotic neurons increased significantly. PQQ(6.4,12.8μM) could significantly inhibited OGD/reoxygenation-induced apoptosis of cultured Neuro2A cells (P<0.01). 4. The content of ROS was increased and the level of GSH was decreased at 6h after OGD, we observed that PQQ (6.4,12.8μM) decreeased the content of ROS and increased the GSH level, significantly.Conclusion: 1. PQQ exhibits remarkable protection against hypoxia/Reoxygenat- ion injury in Neuro2A cells. 2. The mechanism of PQQ neuroprotection may related to its ability of inhibiting the oxidative stress and subsequently suppressing the neurons apoptosis.