| ObjectiveStroke is one of the main causes of human death and disability,and acute ischemic stroke(acute ischemic stroke,AIS)accounted for about 80%of the total stroke.Effectively promote nerve repair after cerebral ischemia is a hotspot of AIS.Used to think that the central nervous system neurons unsustainable.Neural stem cells,Neural stem cells,NSCs)found that brings new hope for nerve repair.Begin to understand musk wake up traditional Chinese medicine(TCM)is widely used in treatment of ischemic stroke,but the mechanism of promoting nerve regeneration is unclear.Use of cerebral ischemia/reperfusion model,neural stem cells in vitro culture of lateral ventricle area,use the techniques,such as molecular biology,stem cell observation "begin to understand wake up" method for the influence of the endogenous NSC proliferation differentiation,method of analysis " to understand wake up" intervention in the molecular mechanism of endogenous NSC proliferation differentiation,reveal "begin to understand wake up" method is the essence of traditional Chinese medicine,enrich the theory of TCM in the treatment of stroke.Methods1.Experimental animalsNinety SPF grade female SD rats with a body weight of 250-300g were divided into 6 groups according to the random number table,24 in the high,medium and low concentration groups,and then divided into 1,3,7 and 14d groups,and 6 in the sham surgery group,model group and blank control group.Normal saline was given to the model group,tho sham operation group and the blank control group,while lmg/kg,2mg/kg and 4mg/kg were given to the musk ketone high,medium and low dose groups respectively.The drug was given by 10ml/kg body weight,once a day.2.Effect of musk ketone on GFAP and neu-n protein expression in the lateral subventricular region of MCAO rats Copy MCAO rats model,high(16.8 mu L/100 g)in mu(8.2 L/100 g)low(4.2 mu L/100 g)dose of musk ketone suspension liquid continuous lavage Id,3d,7d,14d after death,separation zone under lateral ventricle(subventricular zone,the SVZ)brain,neuron specific nucleoprotein protein immunoblot method Neu-N,collagen fibre acidic protein GFAP protein expression,use Image Lab enhancement imaging and data analysis,The protein expression of rats in each group was compared and analyzed.3.Effect of musk ketone on the proliferation and differentiation of GFAP and neu-n cells in the lateral subventricular region of MCAO rats Duplicate rat MCAO model,high(16.8 muL/100 g)in mu(8.2L/100 g)low(4.2 muL/100 g)dose of musk ketone mixed suspension continuous lavage 1d,3d,7d,14d after death,brain tissue separation zone under the lateral ventricle and paraformaldehyde after fixation,embedding,and sectioning immunofluorescence staining,fluorescence microscope,analyzed each rat neurons markers Neu-N,astrocytes marker expression of GFAP positive cells count.Results1.Effect of musk ketone on GFAP and Neu-N protein expression in the lateral subventricular region of MCAO ratsProtein imprinting results suggest that in MCAO rats SVZ area found that GFAP,Neu-N protein of proliferation,compared with model group,high concentration group of GFAP,Neu-N protein expression differences statistically significant(P<0.01),shows different concentrations of muscone on MCAO rats,GFAP protein expression the decreasing trend along with the increasing of drug concentration,Neu-N protein expression along with the increasing of drug concentration increasing trend.GFAP and neu-n protein deposits were found in the lateral ventricle of MCAO rats.Musk ketone concentration dependently down-regulated GFAP expression in lateral ventricle of MCAO rats.Musk ketone concentration dependently up-regulated the expression of Neu-N protein in the lateral ventricle of MCAO rats.2.Effect of musk ketone on the proliferation and differentiation of GFAP and neu-n cells in the lateral subventricular region of MCAO ratsThe results showed that the astrocyte count was:High concentrations of musk ketone group(33.67 +5.86)/vision,concentration of musk ketone group(39.67 +5.13)/vision musk ketone group,low concentration(55.33+2.31)/vision,model group(50.67+2.08)/view,the control group(56.33+9.0)/vision,blank control group(58.33+1.53)/vision,compared with blank control group,model group,low concentration of musk ketone group rats star-shaped glial cell activation count decreased significantly,the difference statistically significant(P<0.01).Compared with the model group,the activation number of astrocytes in rats in the medium-high concentration thymol group was significantly reduced,and the difference was statistically significant(P<0.01).The number of neuron cells was:High concentrations of musk ketone group(110.67+37.2)/vision,concentration of musk ketone group(91.33+36.95)/vision musk ketone group,low concentration(69.33+20.79)/vision,model group(54.33+13.15)/view,the control group(24.33+8.62)/vision,blank control group(12.33+9.29)/vision,compared with blank control group,model group,low concentration of musk ketone group rats neuron cell count increased obviously,statistically significant difference(P<0.01).Compared with the model group,the astrocyte count in rats with high concentration of thymol increased significantly,and the difference was statistically significant(P<0.01).Conclusion1.Focal ischemia in rat brain tissue can induce the proliferation of autologous neural stem cells and migrate to the cortex and striatum in the ischemic area,and eventually differentiate into neurons(neu-n-labeled positive cells)and glial cells(GFAP labeled positive cells),but the number is small.2.Endogenous NSCs proliferated significantly and differentiated into neurons after musk ketone gavage therapy,while the number of astrocytes differentiated was less than that of the model control group.3.Musk ketone can promote the proliferation of endogenous NSCs in MCAO rats,promote their differentiation into neurons,and reduce the proportion of astrocyte differentiation;High concentration of musk ketone has a stronger ability to promote the proliferation and differentiation of NSC. |
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