The Effect And Mechanisms Of Ligustilide On Pathological Intimal And Medial Hyperplasia In Spontaneously Hypertensive Rats

Objective:（1）To research the effect of ligustilide on pathological intimal and medial hyperplasia of thoracic aortic atherogenesis in spontaneously hypertensive rats.（2）To explore the mechanisms of ligustilide inhibits pathological intimal and medial hyperplasia in spontaneously hypertensive rats:1)To explore the the effect of ligustilide on blood pressure and blood lipids in spontaneously hypertensive rats.2)To explore the the effect of ligustilide on the migration of vascular smooth muscle cells（VSMCs）in spontaneously hypertensive rats.3)To explore the the effect of ligustilide on the expression of c-Myc and alpha smooth muscle actin protein.Methods:（1）Ten weeks old male spontaneously hypertensive rats were raised up with high fat diet for 3 weeks after intraperitoneally injecting 600000/U of vitamin D₃ to set up the atherosclerosis modeling.（2）Endarterium of thoracic aortics were measured after intraperitoneally injecting ligustilide concentration of 0,10,20,40 mg/kg respectively for2 weeks in the atherosclerosis modeling,and rats were raised up with normal diet at the same time.（3）The tail artery blood pressure of spontaneously hypertensive rats were measured by using noninvasive arterial detector every Monday 9 am.（4）Two milliliter of blood samples were collected from inferior vena cava of spontaneously hypertensive rats to measure serum lipid and calcium levels after centrifuging.（5）Thoracic aortic endarteriums of spontaneously hypertensive rats were extracted to culture the vascular smooth muscle cells with tissue adherent method.（6）Theα-SMA antibody was used to evaluate vascular smooth muscle cells by immunofluorescence method.（7）The effect of ligustilide on the migration of vascular smooth muscle cells in spontaneously hypertensive rats was tested by scratch wound-healing assay and transwell migration assay.（8）The effect of ligustilide on the expression of c-Myc andα-SMA protein levels in vascular smooth muscle cells was tested by protein immunoblot method in spontaneously hypertensive rats.（9）SPSS 23.0 statistical software was used to analyze the experimental data,the datas were tested by normal test and homogeneity of variance firstly.Single factor analysis of variance would be applied and two groups can be compared with the LSD method（L）when homogeneity of variance,while nonparametric test would be applied.P-values less than0.05 were considered significant and are indicated by asterisks.Results:（1）The thoracic aortic atherosclerosis was observed in the atherosclerosis modeling of spontaneously hypertensive rats after 3 weeks feeding.（2）There were significant differences in the endarterium between the three experimental groups treated with different ligustilide concentration of 10,20,40 mg/kg and control group（P<0.001）,which indicated that ligustilide had a inhibitory effect on the pathological intimal and medial hyperplasia.（3）There were significant differences in the blood pressure between the three experimental groups treated with different ligustilide concentration of 10,20,40 mg/kg and control group（P<0.001）,which indicated that ligustilide could reduced systolic and diastolic blood pressure in spontaneously hypertensive rats.（4）There were significant differences in serum cholesterol and triglycerides between the experimental group treated with ligustilide concentration of 40 mg/kg and control group（P<0.001）,while such differences of low density lipoprotein,high density lipoprotein and serum calcium were not found（P>0.05）.（5）The vascular smooth muscle cells were confirmed by optical microscope and cell immunofluorescence test.（6）There were significant differences in VSMCs migration between the three experimental groups treated with different ligustilide concentration of 10,20,40 mg/kg and control group（P<0.001）,which indicated that ligustilide had a inhibitory effect on the migration of VSMCs.（7）Compared with control group,the expression ofα-SMA was increased and c-Myc was decreased significantly in three experimental groups treated with different ligustilide concentration of 10,20,40 mg/kg（P<0.05）.Conclusion:Ligustilide can prevent the pathological intimal and medial hyperplasia of thoracic aortic,inhibite the migration of VSMCs,decrease systolic and diastolic blood pressure and reduce total cholesterol and triglycerides in spontaneously hypertensive rats.