| Objective:The experiment established a model of cerebral hemorrhage with three-dimensional positioning and type IV collagenase in rats,and were given normal saline,normal saline,Yiqihuoxue method(ginseng,Panax notoginseng extract)and Huoxue Huayu method(Danhong injection)for each Group to intervene.First,observe the neurological deficits,intracranial hematoma volume,and cerebral edema in each group at 24 hours of cerebral hemorrhage,and determine whether the methods of nourishing Qi and activating blood and promoting blood circulation and removing blood stasis are suitable for the early treatment of cerebral hemorrhage,and whether the two method have the ability to prevent brain The role of hematoma in early bleeding and improvement of prognosis.First,observe the neurological deficit,intracranial hematoma volume and cerebral edema in each group at 24 h of cerebral hemorrhage,and determine whether the Yiqi Huoxue method is suitable for early treatment of cerebral hemorrhage,and whether this method has the effect of preventing early hematoma expansion and cerebral hemorrhage and Improve the prognosis.Secondly,to explore the possible mechanism of Yiqihuoxue method influencing the early hematoma expansion rate,improving nerve function and prognosis,and provide a basis for the future application in the treatment of cerebral hemorrhage.Methods:1.Experimental grouping: 158 male SD rats with a weight range of 300 ± 20 g were randomly divided into a sham operation group(32),a blank control group(42),Yiqi Huoxue Group(42),and Huoxue Huayu group(42);2.Modeling: The basal ganglia area is stereotactically positioned by a brain stereotaxic instrument(coordinates: 0.2mm before the anterior condyle point,3.5mm beside the right side,and 5.5mm in depth),and then the microinjector is controlled by a microinjector pump,and slowly toward the skull Intravenous injection of 1ul type IV collagenase(0.5U/ul);3.Model evaluation and administration: The model was evaluated according to the bederson evaluation method 2 hours after the operation,and the model was determined to be included in the study.After 3 hours,the rats were intervened with the corresponding drugs(sham operation group: Gastrointestinal saline was given by gavage,10 ml / Kg · d-1,blank control group: Gastrointestinal saline was given by gavage,10 ml / Kg · d-1,Yiqi Huoxue group: Ginseng and Sanqi concentrated extract Liquid gavage,10 ml / Kg · d-1,Huoxue Huayu group: Danhong injection was given intraperitoneally,2.0ml / Kg · d-1.In this test,the rats in each group were administered once a day.Dose);4.Blood coagulation function and blood biochemical test: After the evaluation of the model is completed,blood is drawn through the inferior vena cava to detect whether there is any difference in blood coagulation and biochemical function between the groups before drug intervention;5.Evaluation of neurological deficits: After 24 h of ICH,the neurological deficit scores were performed on rats in each group according to the modified neurological deficit score method to determine the status of neurological damage(the higher the score,the more severe the neurological deficit);6.Measurement of cerebral hematoma volume: Hematoma volume was measured by histological examination,paraffin-embedded sections of rat brain tissue were used,and hematoma volume was calculated by multifield method(hematoma volume = Π/6 × longitudinal diameter * transverse diameter × section thickness(mm)× number of slices);7.Detection of cerebral edema: After 24 h of ICH,the dry and wet weights of the brain tissues on the affected side and the healthy side were measured,and each group was calculated based on the brain tissue water content(%)=(wet weight-dry weight)/wet weight × 100% Rat brain tissue water content;8.Blood-brain barrier permeability detection: Evans blue spectrophotometric standard curve was prepared with the aid of a microplate reader,and the blood-brain barrier permeability impairment was determined by measuring the EB content in the sample brain tissue.Observe the ultrastructural characteristics of blood-brain barrier of rats in each group by transmission electron microscope;9.Hematoxylin-eosin staining(HE)of brain tissue: observe the morphology of the brain tissue and the condition of nerve cells after ICH24 h by HE staining,measure the number of degeneration cells,and calculate the degeneration cell index;10.Immunohistochemical detection: Detection of MMP-2 and claudin-5 around hematoma at ICH 24 h by immunohistochemistry;11.Statistical analysis: Statistical analysis was performed by spass 19.0 statistical software,and each detection index was described by mean plus or minus standard deviation(?),and single factor analysis of variance was used to compare the data difference between two pairs,P<0.05 as a sign of statistical difference.Results:1.Blood coagulation function and blood biochemical level: Before drug intervention,there was no significant difference in blood coagulation function and blood biochemical level of each group(P>0.05);2.Evaluation of neurological deficits: There were no significant neurological dysfunctions in the sham operation group,and the neurological deficits in each group of cerebral hemorrhage were significant compared with sham operation(P<0.05).After ICH24 h,the two groups receiving drug intervention were compared with the blank control group.The neurological scores of the former two were significantly lower than those of the blank control group(P<0.05).Compared with the two experimental groups,the improvement of neural function in the Yiqi Huoxue group was better than that in the Huoxue Huayu group(P <0.05);3.Hematoma measurement: Compared with the blank control group,the two experimental groups compared with the blank control group after ICH24 h.The amount of intracranial hematoma in the two experimental groups was significantly lower than that in the blank control group.Huayu group(P<0.05);4.Detection of cerebral edema: There was no significant difference in water content between the groups on the healthy side,and the comparison P value between the groups was greater than 0.05.The measurement of the brain water content on the affected side showed that the water content of the sham operation group compared with the other groups It is significantly lower than the other three groups(P<0.05).Compared with the blank control group,the first two groups can reduce the water content of brain tissue and reduce brain edema(P<0.05).The effect of cerebral edema was not significantly different(P>0.05);5.Blood-brain barrier permeability test: After 24 h of ICH,compared with cerebral hemorrhage groups,no EB exudation was observed in the sham operation group(P <0.05),and the EB exudation amount in the blank control group was significantly higher than that in the two experimental groups(P<0.05),there was no significant difference in EB content between Yiqi Huoxue group and Huoxue Huayu group(P>0.05).The ultrastructure of transmission electron microscope showed that the nucleus of the neurons in the sham-operated group was round and the chromatin was evenly distributed.There were complete mitochondria,rough endoplasmic reticulum and ribosome in the cytoplasm,and the structure of vascular endothelial cells was also clear and complete And there is a tight junction structure between endothelial cells,and the microvascular basement membrane is continuous.In the blank control group,neuronal cell apoptosis,cell volume shrinkage,nuclear shrinkage,cytoplasmic electron cloud density increase,microvascular basement membrane discontinuity and other damages,the above conditions in the two experimental groups were significantly improved,Yiqihuoxue group More obvious,but the gap is not big;6.HE staining and degeneration cell index measurement: In each group of cerebral hemorrhage,enlarged tissue space,edema,and disordered arrangement of nerve cells can be seen.In contrast,the Yiqi Huoxue group was lighter than the Huoxue Huayu group.In terms of degeneration cell index,the cerebral hemorrhage groups were significantly higher than the sham operation group(P<0.05),and the degeneration cell index of the Yiqi Huoxue group and Huoxue Huayu group were significantly lower than those of the blank control group(P<0.05).The degeneration cell index of Huoxue group was significantly lower than that of Huoxue Huayu group(P<0.05);7.Immunohistochemical detection: The expression of claudin-5 protein around the hematoma in the sham operation group was significantly higher than that in the experimental groups(P<0.05).The expression of claudin-5 protein around the hematoma in the blank control group and the blood circulation group was significantly lower Yiqi Huoxue group(P<0.05),while the blank control group had no significant difference in expression of claudin-5 protein compared with Huoxue Huayu group(P>0.05);The sham operation group only had a small amount of MMP-2 expression,and the expression of MMP-2 in each group of cerebral hemorrhage was significantly higher than that in the sham operation group(P<0.05).Compared with the blank control group,MMP-2 in the Yiqi Huoxue group and Huoxue group The expression levels of Huayu group were significantly lower(P<0.05),and the expression of MMP-2 protein in Yiqi Huoxue group was significantly lower than that in Huoxue Huayu group(P<0.05).Conclusion:1.Yiqi Huoxue Method and Huoxue Huayu Method can be applied to the early treatment of cerebral hemorrhage,both of which can reduce the incidence of hematoma enlargement and improve the neural function of rats with cerebral hemorrhage.2.The various research indicators in the experiment show that the treatment effect of Yiqi Huoxue method on cerebral hemorrhage is better than the method of promoting blood circulation and removing blood stasis(including improvement of nerve function,prevention of hematoma expansion,reduction of brain tissue degeneration cell index,etc.).3.Yiqihuoxue method(ginseng,Panax notoginseng extract)and Huoxuehuayu method(Danhong injection)may play a role in brain tissue and blood brain by reducing the expression of MMP-2 protein or promoting the expression of claudin-5 protein The protective effect of the barrier to reduce the incidence of early hematoma expansion and improve nerve function. |
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