| Objective::Hyperlipidemia(HLP) is due to confusion of synthesis and cleanup of lipoproteins in the blood.The plasma lipoprotein is abnormal and the blood lipid is heighten.It is belong to"phlegm"and"blood stasis"in the traditinal Chinese medicine.It is the most important risk factor of cardiovascular disease,such as atherosclerosis. Controling HLP is the most important way of control cardiovascular disease.Cerebrovascular disease is one of the diseases which threatening human health. It was the leading cause of death in China. Ischemic cerebrovascular disease is in 60%～80%. Cerebrovascular diseases are common disease in middle and old aged people, with high morbidity, mortality and deformity.In order to abate and reduce apoplexy and decrease poststroke syndrome,we use acupuncture in early hyperlipidemia stage.It is a new view-angle of controling cardiovascular disease. It is very important in epidemiologic studies.The rationship between hyperlipidemia and apoplexy, neural stem cells(NSC) and neurotrophic factors(NF) was not discussed in earlier study.Now we try to build hyperlipemia-cerebral ischemia model,and observe acupuncture's effect.We want to know if acupunctre can rise the excrete of neurotrophic factors, accelerate proliferation , migration and differentiation of neural stem cell, and then advance rehabilitation and regeneration of neurons.Methods:Feed the rats with high fat fodder and then use FeCl3 to induce thromboangitis obliterans of middle cerebral artery to establish the model of hyperlipidemia combined with cerebral ischemia. There are 8 groups in this study, normal control group,modle group of hyperlipemeria, treatment group of hyperlipemia, modle group of cerebral ischemia, treatment group of cerebral ischemia, modle group of hyperlipemia combined with focal cerebral ischemia, treatmentⅠ,Ⅱg roups of hyperlipemia combined with focal cerebral ischemia. Needling Fenglong(ST40),Sanyinjiao(SP6) in treatment group of hyperlipemia with electrical acupuncture for 10 days. Needling Bahui(DU20),Fenglong(DU26) in treatment group of cerebral ischemia for 7 days.There are two stage in the treatmentⅠ,Ⅱgroups.These two groups were fed with high fat fodder to establish the hyperlipidemia model.The treatment groupⅠwas given electroacupuncture in ST40,SP6 for 10 days,and then two groups were made to be the model of hyperlipidemia combined with cerebral ischemia with FeCl3.These two groups were treated then with electroacupuncture in ST40 and SP6, and acupuncture in DU20,DU26 for 7 days.Do not use acupuncture in normal control and model groups. Use biochemical method to observe the change of blood-lipid. Use neuroethology index observe the neural symptom. Use immunohistochemical method to observe the expression of HE stain in the Hippocampus area and the frontal lobe cortex, and the expression of nestin,PCNA,vimentin,Tuj-1 in the subventricular zone (SVZ) and the frontal lobe cortex. Use Enzyme-linked immenosorbent assay(ELISA)to detect the concentration of NSE,NGF,BDNF,bFGF of the right brain homogenate.Results:1 Building the hyperlipemia with cerebral ischemia rats model.Feed the rats high fat fodder for 6 weeks,and then use FeCl3 to induce thromboangitis obliterans of middle cerebral artery to establish the model of hyperlipidemia combined with cerebral ischemia.The result of preexperiment show:that contrast with the normal control group,the CHO,LDL-C,TG level of the hyperlipidemia 4 or 6 week group is higher, P<0.01.So we can build the hyperlipidemia model by 4 or 6 weeks.Because 6 weeks model is close to the human hyperlipidemia pathology, we decide use 6 weeks to make hyperlipidemia modle.In the experiment,the results show that the CHO,LDL-C,TG level of the groups fed with high fat fodder is higher than the normal group(P<0.01) in 42 days. HLD-C level is lower than the normal group(P<0.01).The hyperlipemia modle is made successfully.Positive neurobehavioral reaction can be found in one to two hours after ischemia in the 52th day. The neurobehavioral score of all ischemic model groups significantly higher than normal and high blood lipid groups and also higher than all acupuncture groups(p<0.01). HE dyeing result shows that after made cerebral model, the ischemia area is in the frontal lobe cortex.The position is fixed. The cells in ischemia area is messy. Lots of neurons become denaturation, and the manifestations of cerebral edema were observed too.The neurons in the Hippocampus area become denaturation, thanatosis and disorder in range,too.The hyperlipemia with cerebral ischemia rats model is made successfully. It is the first time to build the hyperlipemia with cerebral ischemia rats model.This model is easy to made and repeat , is close to the human hyperlipidemia pathology,too.The mortality rate is lower.2 the blood lipid , HE dyeing,NSC,NF of the normal ratsNormal rats CHO 51.92±9.46mg/dL, HDL-C 26.80±8.16 mg/dL,LDL-C 30.50±8.19 mg/dL,TG47.98±12.18 mg/dL。HE dyeing result shows that,the order of the neurons of the right brain of the rats is in range.We can see the neurons shape clearly.There is little expression of nestin,PCNA,vimentin,Tuj-1 in the subventricular zone of the normal rat's right brain.(the extend of back and lateral position of lateral ventrical:nestin:the positive cells number 63±9.40 ; area density0.62±0.10% ; optical density0.50±0.03%.PCNA:the positive cells number185±10.60;area density1.93±0.11%; optical density1.22±0.07%.vimentin:the positive cells number95±6.31;area density 0.61±0.04%; optical density0.34±0.02%.Tuj-1:the positive cells number56±7.69:area density 0.96±0.13%:optical density0.22±0.03%.lateral wall of lateral ventrical:nestin:the positive cells number41±5.61;area density 0.35±0.05%;optical density0.23±0.03%. PCNA:the positive cells number114±22.79;area density1.25±0.25%; optical density0.94±0.19%. vimentin: the positive cells number 48±11.85;area density 1.05±0.26%; optical density 0.58±0.14%. Tuj-1:the positive cells number35±4.69; area density 0.88±0.12%; optical density 0.45±0.06%.)There is little expression of nestin,PCNA,vimentin,Tuj-1 in the frontal lobe cortex.(nestin:the positive cells number14±3.47;area density0.11±0.03%; optical density0.09±0.02%. PCNA:the positive cells number58±6.52;area density0.64±0.07%; optical density0.39±0.04%.vimentin:the positive cells number56±6.46; area density 0.22±0.03%; optical density0.11±0.01%.Tuj-1:the positive cells number13±5.14; area density 0.08±0.03%;optical density0.05±0.02%.)The concentration of NSE(41.70±3.47 ng/dL),NGF(79.36±7.07 pg/ml),BDNF(5.66±0.86μg/L),bFGF (51.20±6.05 ng/L)assume basis expression。3 the effect of acupuncture on the hyperlipidemia rats3.1 the effect of acupuncture on the blood lipid of the hyperlipidemia rats Contrast with the hyperlipidemia model rats, the CHO,TG,LDL-C level of hyperlipemia rats is lower (P<0.01) and the HDL-C level of hyperlipemia rats is higer(P<0.01).Acupuncture can lower the level of CHO,TG,LDL-C,raise the level of HDL-C, and adjust the level of blood lipid.3.2 the effect of acupuncture on NSC of the hyperlipidemia rats The results show that There is little expression of nestin,PCNA,vimentin,Tuj-1 in hyperlipemia rat's brain. There is a little expression of nestin,PCNA,vimentin,Tuj-1 in the subventricular zone and frontal lobe cortex. There is no difference among the positive cells number, area density, optical density of the normal group, hyperlipemia model group,and the hyperlipemia treatment group(P>0.05). Acupuncture can do nothing on the expression of NSC of the hyperlipidemia rats.3.3 the effect of acupuncture on NSE of the hyperlipidemia rats The NSE level of the right brain has no difference among the normal group, hyperlipemia model group,and the hyperlipemia treatment group (P>0.05). Hyperlipemia can not do anything on the NSE level of the hyperlipidemia rat's right brain.Acupuncture can not do anything on the concentration of NSE of the hyperlipidemia rats.3.4 the effect of acupuncture on NF of the hyperlipidemia ratsThe NGF,BDNF,bFGF level of hyperlipemia model group is lower than that of the normal group (P<0.05/P<0.01).The BDNF level of is hyperlipemia treatment group is higher than the hyperlipemia model group.Acupuncture can raise the level of BDNF of the hyperlipidemia rats.4 the effect of acupuncture on the ischemia rats4.1 the effect of acupuncture on the neurobehavioral score and HE dyeingIn the 59th day, the neurobehavioral score of the ischemia model and treatment group is higher than that of the normal group (p<0.01).That of the ischemia treatment group is lower than the ischemia model group (p<0.01).The study shows that acupuncture therapy can reduce the neurobehavioral score and improve neural founction.HE dyeing result shows the edema of the ischemia treatment group is lighten.The cell nucleus is normal.The normal neuron is more than the model group. Edema and vacuole formation in vascular endothelium can be seen.So acupuncture method can lighten the denaturation and thanatosis in neurons.4.2 the effect of acupuncture on NSC of the ischemia ratsAfter builing ischemia modle, the expression of nestin,PCNA,vimentin,Tuj-1 in the subventricular zone (SVZ) and frontal lobe cortex of the right brain is enhance.There are lots of positive cells, and the back lateral extension has come to being.The the positive cells number ,area density,optical density of the ischemia model and treatment group is higher than that of the normal group(p<0.01). That of the treatment group is higher than the model group(p<0.01).The NSC proliferate and differentiate after ischemia, and some NSC differentiate to glialcells and neurons.Acupuncture can promote the NSC to proliferate and differentiate(p<0.01).Using acupuncture in hyperlipemia stage can accelerate proliferation of NSC, migration to the ischemia area and differentiation of neural stem cell to the neuron and neuroglia. 4.3 the effect of acupuncture on NSE of the ischemia ratsThe NSE level of the ischemia modle rat's right brain is higher than that of the normal group(p<0.01). That of the treatment group is lower than that of the model group(p<0.01). The result show that the brain damage appeared after the ischemia. Acupuncture can lower the NSE level,and alleviate the cerebral lesion.4.4 the effect of acupuncture on NF of the ischemia ratThe NGF,BDNF,bFGF level of the ischemia modle rat's right brain is higher than that of the normal group(p<0.01). The NGF,bFGF level of the treatment group is higher than that of the model group(p<0.05/0.01).The result show that the brain can secrete NGF,BDNF,bFGF after the ischemia. So the brain can protect himself by NF.Acupuncture can raise the NGF,bFGF level,and protect the brain. 5 the effect of acupuncture on the hyperlipemia with cerebral ischemia rats5.1 the effect of acupuncture on the blood lipid of the hyperlipemia with cerebral ischemia ratsThe CHO,LDL-C level of the hyperlipemia with cerebral ischemia model and treatmentⅠ,Ⅱgroup is higher than that of the normal group(p<0.01).The TG level of the hyperlipemia with cerebral ischemia model and treatmentⅡgoup is higher than that of the normal group(p<0.01).The CHO,TG,LDL-C level of the treatmentⅠgroup is lower than that of the hyperlipemia with cerebral ischemia model group(p<0.01/0.05). There is no difference in the CHO,TG,LDL-C level between the treatmentⅠa ndⅡgroup. There is no difference in the HDL-C level between every two groups.But the HDL-C level in 59th of treatmentⅠis higher than that of 42th day.Acupuncture can lower the CHO,TG,LDL-C and raise the HDL-C level of the hyperlipemia with cerebral ischemia rats.The effect of the treatmentⅠgroup is better than that of the treatmentⅡgroup.It can adjust the level of blood lipid better when we use acupuncture in the hyperlipemia stage.5.2 the effect of acupuncture on the neurobehavioral score and HE dyeing of the hyperlipemia with cerebral ischemia ratsIn the 59th day, the neurobehavioral score of the treatmentⅠa ndⅡgroup is lower than that of the model group. There is no difference between the treatmentⅠa ndⅡgroup (p>0.05). So acupuncture can treat cerebral ischemia rats.HE dyeing result shows the edema of the treatmentⅠandⅡgroup is lighten.The cell nucleus is normal.The normal neuron is more than the model group. The edema and vacuole formation in vascular endothelium can be seen. The effect of the treatmentⅠgroup is better than that of the treatmentⅡgroup.So acupuncture method can lighten the denaturation and thanatosis in neurons. It can lighten brain damage better when we use acupuncture in the hyperlipemia stage.5.3 the effect of acupuncture on the NSC of the hyperlipemia with cerebral ischemia ratsThe expression of nestin,PCNA,vimentin,Tuj-1 in the subventricular zone (SVZ) and frontal lobe cortex of the right hyperlipemia with cerebral ischemia rat's brain is lighten than that of the ischemia rats.The the positive cells number ,area density,optical density of the hyperlipemia with cerebral ischemia model and treatment group is higher than that of the normal group(p<0.01). That of the treatmentⅠ,Ⅱgroup is higher than the model group(p<0.01). That of the treatmentⅡgroup is lower than that of the treatmentⅠgroup(p<0.01).The result show that hyperlipemia can lighten the expression of nestin,PCNA,vimentin,Tuj-1 of the hyperlipemia with cerebral ischemia model. Hyperlipemia can reduce proliferation of NSC, migration to the ischemia area and differentiation of neural stem cell to the neuron and neuroglia.Acupuncture can enhance the expression of nestin,PCNA,vimentin,Tuj-1 in the subventricular zone and frontal lobe cortex of the right brain,The expression of nestin,PCNA,vimentin,Tuj-1 and NGF,BDNF,bFGFin treatmentⅠgroup is higer than that of treatmentⅡg roup.Using acupuncture in hyperlipemia stage can accelerate proliferation of NSC, migration to the ischemia area and differentiation of neural stem cell to the neuron and neuroglia.5.4 the effect of acupuncture on the NSE of the hyperlipemia with cerebral ischemia ratsThe NSE level of the hyperlipemia with cerebral ischemia model group is higher than that of the normal and ischemia group (P<0.05/0.01).The NSE level of the hyperlipemia with cerebral ischemia model and treatment group is higher than that of the normal group(p<0.01). The NSE level of the treatmentⅠ,Ⅱgroup is lower than that of the model group(P<0.05/0.01). The NSE level of the treatmentⅡgroup is higher than that of the treatmentⅠgroup.The result show that hyperlipemia with cerebral ischemia can enhance brain damage after the ischemia.Acupuncture can alleviate cerebral lesion. Using acupuncture in hyperlipemia stage can accelerate neuronal rehabilitation and regeneration.5.5 the effect of acupuncture on the NF of the hyperlipemia with cerebral ischemia rats The NGF,BDNF,bFGF level of the hyperlipemia with cerebral ischemia model group is lower than that of the normal and ischemia group (P<0.01). The NGF,bFGF level of the treatmentⅠ,Ⅱgroup is higher than that of the model group(p<0.01) The NGF,BDNF level of the treatmentⅡgroup is lower than that of the treatmentⅠgroup (p<0.01).The result show that hyperlipemia with cerebral ischemia can lighten brain protection after the ischemia. Acupuncture can enhance the level of NGF,BDNF,bFGF,and protect the brain.The effect of treatmentⅠgroup is better than that of treatmentⅡgroup.So using acupuncture in hyperlipemia stage can accelerate secreting NF,and protect the brain. Conclusion:1 The model of hyperlipemia with cerebral ischemia rats is successful.2 Hyperlipemia can not do anything on proliferation , migration and differentiation of neural stem cell, but maybe lower some of the NF level, for example the level of NGF,BDNF,bFGF.3 The NSC proliferate and differentiate after ischemia, and some NSC differentiate to glialcells and neurons.Ischemia can raise the NSE, NGF,BDNF,bFGF level of the ischemia brain..4 Hyperlipemia with cerebral ischemia can reduce proliferation , migration and differentiation of neural stem cell in the SVZ and frontal lobe cortex of the ischemia brain, enhance the level of NSE, lower the level of the NGF,BDNF,bFGF of the ischemia brain. The hyperlipemia with cerebral ischemia can enhance brain damage and lighten brain protection after the ischemia.5 Acupuncture in ST40,SP6 can lower CHO,TG,LDL-C level of the hyperlipemia rats,raise the HDL-C level,and regulate blood lipid. It can not do anything on proliferation , migration and differentiation of neural stem cell in the SVZ and frontal lobe cortex of the ischemia brain and NSE level of the ischemia brain. But it can raise the BDNF level of the ischemia brain.6 Acupuncture in DU20,DU26 can improve the state after brain ischemia, lower the NSE level of the ischemia brain,enhance proliferate and differentiate of NSC in the SVZ and frontal lobe cortex after ischemia enhance the tendency of NSC differentiating to glialcells and neurons, and raise the NGF,bFGF level of the ischemia rats.7 Acupuncture in ST40,SP6,DU20,DU26 can lower CHO,TG,LDL-C level ,raise the HDL-C level,and regulate blood lipid. It can improve the state after brain ischemia, enhance proliferate and differentiate of NSC in the SVZ and frontal lobe cortex after ischemia, enhance the tendency of NSC differentiating to glialcells and neurons, and raise the NGF,BDNF,bFGF level,and lower the NSE level of the ischemia brain.8 Using Acupuncture in hyperlipemia stage can regulate blood lipid ,lower the NSE level, raise the NGF,BDNF,bFGF level of the ischemia brain and accelerate proliferation, migration and differentiation of NSC.9 Acupuncture can accelerate neuronal rehabilitation and regeneration through lowering the concentration of NSE,raise the expression of NGF,BDNF,bFGF and nestin,PCNA,vimentin,Tuj-1.In conclusion,there is close relationship between the NF and NSC.In this study,the level of NF raising has close relationship with the proliferating and differentiating of NSC when ischemia and hyperlipemia with cerebral ischemia. So we guess that the NF raising can accelerate the NSC to proliferate and differentiate.Acupuncture can accelerate proliferation , migration and differentiation of neural stem cell, and then advance rehabilitation and regeneration of neurons,through accelerating secretion and synthesis of the neurotrophic factor.Using Acupuncture in hyperlipemia stage, can accelerate neuronal rehabilitation and regeneration through lowering the concentratio of NSE, raise the expression of NF and NSC of the ischemia brain.
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