| Objective:To study the effect of Annao pingchong Decoction on TRPV4,KCa3.1 protein expression and serum TNF-?,ICAM-1 concentration in rats with intracerebral hemorrhage,explore its therapeutic effect on cerebral edema and neuroprotective mechanism in rats with intracerebral hemorrhage.Methods:According to the random number table method,SD rats were divided into six groups:sham operation group,model group,Annaopingchong Decoction low-dose group,Annaopingchong Decoction high-dose group,nimodipine group,HC-067047 group(TRPV4 inhibitor),intracerebral hemorrhage model was established by autologous blood caudate nucleus injection.Each group was divided intofour subgroups :6h,24 h,3d and 7d.After the model was made,rats were intervened with corresponding drugs.The sham operation group and mod el group were given the same dose of distilled water.Rats in each experimental group were scored for neurological deficits at the corresponding time points,then the rats were decapitated and brain tissue was colledted to detect brain water content and HE staining,blood was drawn from the abdominal aorta to detect serum TNF-? ICAM-1 concentration by the enzyme-linked immunosorbent assay.Immunohistochemical method and Western Blot were used to detect the TRPV4 and KCa3.1 protein expression in the brain tiss ue around the hematoma on the third day.Result:1.Neurological deficit score:Compared with the sham operation group,the neurological deficit score of each group of intracerebral hemorrhage modeling was significantly increased at all differenttime points(P<0.05);At 24 h,3d and 7d after intracerebral hemorrhage,compared with the model group,the neurological deficit scores of each drug group were lower than the model group(P<0.05);On the 7d after intracerebral hemorrhage,compared with the HC-067047 group,the neurological deficit scores of the low-dose group of Annaopingchong Decoction,the high-dose group of Annaopingchong Decoction and the nimodipine group were significantly reduced(P<0.05);On the 7th day after intracerebral hemorrhage,compared with the low-dose group of Annaopingchong Decoction and the nimodipine group,the neurological deficit score of the high-dose group of Annaopingchong Decoction was significantly reduced(P<0.05).2.Moisture content of brain tissue : Compared with the sham operation group,the water content of brain tissue in each group of intracerebral hemorrhage modeling was significantly increased at all different time points(P<0.05);At 24 h,3d and 7d after intracerebral hemorrhage,compared with the model group,the water content of the brain tissue of each drug group was lower than that of the model group(P<0.05);On the 7th day after intracerebral hemorrhage,compared with the low-dose group of Annaopingchong Decoction,the nimodipine group and the HC-067047 group,the degree of cerebral edema in the high-dose group of Annaopingchong Decoction was significantly reduced(P<0.05).3.HE staining:The brain damage of the model group,the Annaopingchong Decoction group,the nimodipine group,and the HC-067047 group was basically the same 6h after cerebral hemorrhage;At 24 h,3d,and 7d after intracerebral hemorrhage,compared with the model group,the Annaopingchong Decoction group,nimodipine group,and HC-067047 group can effectively improve brain tissue injury.4.Enzyme-linked immunosorbent assay:(1)TNF-?:Compared with the sham operation group,the concentration of TNF-? in each group of intracerebral hemorrhage modeling was significantly increased at different time points(P<0.05);At 24 h,3d,and 7d after intracerebral hemorrhage,compared with the model group,the TNF-? concentration of each drug group was lower than that of the model group(P<0.05);On the 3rd and 7th day after intracerebral hemorrhage,compared with the low-dose group of Annaopingchong Decoction,the nimodipine group and the HC-067047 group,the concentration of TNF-? in the high-dose group of Annaopingchong Decoction were significantly reduced(P<0.05).(2)ICAM-1:Compared with the sham operation group,the concentration of ICAM-1 in each group of intracerebral hemorrhage modeling was significantly increased at different time points(P<0.05);At 24 h,3d and 7d after intracerebral hemorrhage,compared with the model group,the concentration of ICAM-1 in each drug group was lower than that of the model group(P<0.05);On the 3rd and 7th day after intracerebral hemorrhage,compared with the low-dose group of Annao pingchong Decoction,the nimodipine group and the HC-067047 group,the concentration of ICAM-1 in the high-dose group of Annaopingchong Decoction were significantly reduced(P<0.05).5.Immunohistochemical:The results of TRPV4 and KCa3.1 in the brain tissue around the hematoma on the 3rd day after modeling in rats showed that the expressions of TRPV4 and KCa3.1 in each group of the intracerebral hemorrhage model were significantly increased comp ared with the sham operation group(P<0.05);compared with the model group,the expressions of TRPV4 and KCa3.1 in each drug group were lower than those in the model group(P<0.05);compared with the low-dose group of Annaopingchong Decoction,the expressions o f TRPV4 and KCa3.1 in the high-dose group of Annaopingchong Decoction,nimodipine group,and HC-067047 group were significantly reduced(P<0.05).6.Western Blot:(1)The results of TRPV4 in the brain tissue around the hematoma on the 3rd day of each experimental group showed that compared with the sham operation group,the expression of TRPV4 protein in the model group was significantly increased.The protein expres sion was significantly reduced(P<0.05);compared with the model group,the TRPV4 protein expression of each drug intervention group was reduced(P<0.05);compared with the low-dose group of Annaopingchong Decoction,the expression of TRPV4 in the high-dose group of Annaopingchong Decoction,nimodipine group,and HC-067047 group was significa ntly reduced(P<0.05);Compared with the high-dose group of Annaopingchong Decoction,the expression of TRPV4 in the nimodipine group and the HC-067047 group was significantly reduced(P<0.05).(2)The results of KCa3.1 in the brain tissue around the hematoma on the 3rd day of each experimental group showed that the expression of KCa3.1 protein in the model group and the low-dose group of Annaopingchong Decoction was significantly up-regulated compared with the sham operation group,and the nimodipine group an d HC-The expression of KCa3.1 protein in group-067047 was significantly reduced(P<0.05);compared with the model group,the KCa3.1 protein expression of each drug intervention group was reduced(P<0.05);compared with the low-dose group of Annaopingchong Decoction,the expression of KCa3.1 in the high-dose group of Annaopingchong Decoction,nimodipine group,and HC-067047 group was significantly reduced(P<0.05);compared with the high-dose group of Annaopingchong Decoction,the expression of KCa3.1 in the nimodipine group and the HC-067047 group was significantly reduced(P<0.05).Conclusion:1.In intracerebral hemorrhage model rats,the expression of TRPV4 and KCa3.1 in the brain tissue around the hematoma increased,and the serum TNF-? and ICAM-1 concentrations increased.2.Annaopingchong Decoction can effectively improve the neurological deficit score of rats with intracerebral hemorrhage,reduce cerebral edema and reduce inflammatory factors,thereby exerting a therapeutic effect on cerebral hemorrhage,and the therapeutic effect of Annao pingchong Decoction on intracerebral hemorrhage rats has a certain dose-effect relationship.3.Possible mechanism:down-regulating the expression of TRPV4 and KCa3.1,reducing intracellular calcium overload,inhibiting the activation of microgli,reducing the concentration of serum TNF-? and ICAM-1,inhibiting the inflammatory response,thus reducing brain edema and playing a protective role in neurological function. |
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