The Establishment Of High Altitude Cerebral Model In Rats And Intervention Of 5,6,7,8-tetrahydroxyflavone

Objective: 1.Establishment and evaluation of an experimental rat model of HACE.2.To investigate the intervention of 5,6,7,8-tetrahydroxyflavone on rats with HACE and its primary mechanism.Methods: 1.Male Wistar rats were randomly divided into 4 groups: Control group,HH group,HH+2? group,HH+12/2? group.In addition to the control group,the rest of the groups were placed in a hypobaric hypoxia cabin at an altitude of 6000 m for 72 h according to different conditions.The optimal model of HACE was screened by weight change of rats,water content of brain tissue,permeability of blood-brain barrier and HE staining.In addition,models were further evaluated by indicators of oxidative stress and energy metabolism,levels of inflammatory cytokines,expression of AQP4 and VEGF.2.The anti-hypoxia pharmacodynamics of 5,6,7,8-tetrahydroxyflavone at low,medium and high doses were studied by using normobaric hypoxia test and acute hypobaric hypoxia test in mice to determine the optimal dosage.Equivalent dose conversion between mouse and rat according to the optimal dosage.Male Wistar rats were randomly divided into 4 groups: Control group(Control),HACE group(HACE),acetazolamide(ACZ),5,6,7,8-tetrahydroxyflavone(THF).In addition to the control group,the HACE group,ACZ group and THF group were placed in a hypobaric hypoxia cabin at an altitude of 6000 m for 3 days after intragastric administration at a local altitude for 3 days.Observe the pathological changes of the brain and detect the brain water content,EB content,related biochemical indicators,gene and protein expression.3.Eight-arm maze test was used to explore the effect of 5,6,7,8-tetrahydroxyflavone on cognition in HACE rats.Male Wistar rats successfully trained were randomly divided into three groups: Control group,HACE group,5,6,7,8-tetrahydroxyflavone group(THF,350mg/kg).Repetitive rat acute decompression experiment.On the 7th day of the experiment,behavioral test was carried out with eight-arm maze,and WME,WMEF,RME,RMEF,TE and TT were recorded respectively.Each rat was measured twice and averaged.To evaluate the protective effect of 5,6,7,8-tetrahydroxyflavone on HACE rats from the aspect of behavior.Results 1.Compared with the control group,the weight loss of HH+12/2? group was more obvious,the brain water content and EB content were increased,and the pathological changes of brain tissue were observed by HE staining.Therefore,HH+12/2? group is the best model of high altitude brain edema.In addition,through the oxidative stress and energy metabolism indicators,the levels of inflammatory cytokines,the expression of AQP4 and VEGF to further evaluate these models.The levels of O2-·,H2O2,MDA,LD,LDH,pyruvate and PK in HH+12/2? group were all increased(P<0.05 or P<0.01).The activity of SOD,CAT,GSH-PX,GSH-ST,Na+-K+-ATPase,Ca2+-Mg2+-ATPase activity and GSH content were significantly decreased(P<0.05 or P<0.01).The serum levels of NF?B-p65,IL-6,IL-1? and TNF-? increased significantly(P<0.05 or P<0.01),and the levels of AQP4 and VEGF that were closely related to brain edema were significantly higher(P<0.01).These datas further illustrate that the HH+12/2? is the best model.2.Normobaric hypoxia test and acute hypobaric hypoxia test showed that high-dose group(500 mg/kg)had the best anti-hypoxia activity,so the latter part of the experiment selected high dose administration,the best dose of 350 mg/kg through the equivalent dose conversion between species of rats and mice.Acute decompression rats experiment showed that 5,6,7,8-tetrahydroxyflavone can significantly reduce brain water content and EB content in HACE rats and improve the pathological changes of brain tissue.In addition,5,6,7,8-tetrahydroxyflavone could reduce O2-·,H2O2,MDA,LD,LDH and PK(P<0.05 or P<0.01)and increase the levels of SOD,GSH-PX,GSH-ST,Na+-K+-ATPase,Ca2+-Mg2+-ATPase activity and GSH content(P<0.05 or P<0.01),thereby attenuating oxidative stress.By downregulating NF?B-p65 pathway,the secretion of inflammatory cytokines IL-6,IL-1? and TNF-? and the expression of AQP4(P<0.05 or P<0.01)were decreased.The expression of HIF-1?(P<0.01),a key factor in hypoxia signaling pathway,was down-regulated while the expression of downstream proteins VEGF and MMP-9 were down-regulated(P<0.05 or P<0.01).3.Compared with control group,the rats in HACE group had significantly increased both short-term memory error(WME)and long-term memory error(RME,RMEF)and prolonged TT and TE(P<0.01).After the administration of THF,the WME,WMEF,RME,RMEF,TT significantly decreased(P<0.05 or P<0.01).Conclusions 1.The optimal conditions for establishing experimental rat model of HACE were preliminarily defined as the temperature difference between day and night(12/2?)of 6000 m combined with cold conditions.2.5,6,7,8-tetrahydroxyflavone has a good anti-hypoxia effect,and high-dose(350 mg/kg)has best anti-hypoxic activity;In addition,5,6,7,8-tetrahydroxyflavone(350 mg/kg)has a good effect on the prevention of HACE.The mechanism of 5,6,7,8-tetrahydroxyflavone is to reduce oxidative capacity and improve the anti-oxidative capacity.At the same time,through the NF?B pathway,it reduces the inflammatory response,reduces the expression of AQP4,decreases the water content of the brain,maintains the permeability of the blood-brain barrier and improves the histopathology of the brain.In addition,5,6,7,8-tetrahydroxyflavone can down-regulate the expression of HIF-1?,a key factor in the hypoxia signaling pathway,and its downstream related proteins VEGF and MMP-9,reduce vascular permeability,protect the blood-brain barrier,and prevent HACE occurs.3.5,6,7,8-tetrahydroxyflavone can improve the learning and memory function induced by HACE.