Pretreatment Administration Of Triptolide Ameliorates OGD Cerebral Ischemia In Vitro

BackgroundCerebrovascular accident(CVA),a medical term for stroke when blood flow through part of your brain and stopped by a blockage of blood clot or come up with a rupture of blood vessel.There are two main types:an ischemic stroke is caused by a blockage;a hemorrhagic stroke is caused by the rupture of a blood vessel.Both types of stroke deprive part of the blood and oxygen in take of the brain,which cause brain cells to die.Many in vitro brain endothelial cell culture models have been developed and studied over the past several decades to understand the pathophysiological mechanisms and role of the BBB in stroke[1].Triptolide,purified from Tripterygium wilfordii Hook.F.(Celastraceae)with a broad range of bioactivities and promising anti-inflammatory and anti-angiogenesis property has been used to treat autoimmune and inflammatory diseases in China for centuries.Triptolide has been reported to suppress the tumours’ growth and metastasis of many kinds including melanoma.Six years ago,a chemical biologist and his colleagues at Johns Hopkins university school of medicine found a chemical way to change its property by adding glucose to the tripterygium wilfordii and turned it into a soluble molecules。This structural change has also reduced the drug’s side effect of patients and since then.Triptolide(TP)was seen as a good application prospect for traditional Chinese medicine.This study investigated the neuroprotective mechanism of the drug for cerebral ischemia according to its protective prospects to nervous system.ObjectivesThe amelioration of inflammatory and apoptosis as a result of cerebral ischemia is of great interest to researchers and scientists alike.This study aimed at delving into the morphology and pathogenesis of PC 12 cells following artificial damage by setting up oxygen and glucose deprivation(OGD)condition so as to establish an in vitro model of cerebral ischemia.Triptolide(TP)was reported to have prominent therapeutic potential for many pathologic conditions especially in neuroprotection area,so the administration of TP was employed in the setting up of ischemia in vitro neuronal model with PC 12 cells under OGD environment.The drug take effect trough involving in to the inflammation and apoptosis signaling pathways.Meanwhile,side effects that hinder its applying to the clinical treatment should be reformulated and improved.MethodsEstablishment of cell model,set time as variable amount and cultured PC 12 in condition of oxygen and glucose deprivation environment in water-saturated incubator.First the cell were pretreated with TP for 24 hours and conducted OGD for 4 hours then the cell was recovered in 37 ℃ and 5%C02 for 24 hours.The suitable concentration of TP was screened subsequently along with the employment of MTT assay.The trypan blue staining and Hoechst 33342 staining were applied to analyze cell apoptosis and necrosis.The successful quantification of protein expression levels involved in the inflammatory autophagy and apoptosis pathways was measured by western blotting analysis.Additionally,the anti-inflammatory effect of TP was accessed through the expression of TNF-a and iNOS.ResultsTriptolide has the potentiality of serving as an anti-inflammatory agent that could ameliorate cerebral ischemia injury caused by OGD.But the mechanism remains unclear,this study gives a theory basis for the prevention and cure to ischemia stroke.