| Intracerebral hemorrhage(ICH),caused by cerebral vascular rupture,is a major public health problem.Worldwide,the annual incidence is about 10-30 per 100,000,accounting for 10-15% of all stroke patients,with high mortality and disability rates.Studies have shown that the incidence of ICH in the elderly is higher and has not decreased in recent decades,but the incidence of cerebral hemorrhage in young adults has been increasing in recent years,and there are only a few studies on the long-term mortality and functional defects of ICH in young adults.There are gender differences in the incidence,pathophysiological changes,complications,treatment and prognosis after intracerebral hemorrhage.The effect of age and sex on cerebral hemorrhage was explained.We used collagenase to induce female and male ICH mice of different ages,as well as elderly,middle-aged and young ICH mice of different genders.We studied the differences of cerebral hemorrhage mice of different ages and genders in the above aspects by assessing the motor function damage,memory function damage,depression and anxiety degree after ICH.We found that the younger the age of the mice with cerebral hemorrhage,the less the motor and memory impairment,and the lower the degree of depression and anxiety.It was also found that depression and anxiety were the main differences between the elderly male and female cerebral hemorrhage mice,while there were differences between the middle-aged and young cerebral hemorrhage mice in sports,memory,depression and anxiety.To date,no surgery or medication has significantly improved long-term outcomes in ICH patients,especially in memory,depression,and anxiety.Chocolate,green tea and cocoa powder contain a significant proportion of antioxidant molecules,mainly flavonoids,especially flavanols,which are mostly in the form of epicatechin.Flavanol(-)epicatechin(EC)has been shown to be beneficial for vascular and cognitive function in humans and ischemic and hemorrhagic stroke in mice.Our previous studies showed that oral EC 3h after intracerebral hemorrhage could protect the brain of mice after intracerebral hemorrhage.We mainly examined the effect of EC on histopathology and function of middle-aged male and female cerebral hemorrhage mice.Oral administration of EC or saline every 24 hours for 7consecutive days at 6 or 12 hours after collagenase-induced intracerebral hemorrhage.We then assessed lesion size,brain edema,neurological dysfunction,white matter damage,neuronal survival,memory function,depression,and anxiety.It was found that EC significantly reduced cerebral edema and neurological deficits 6h after intracerebral hemorrhage,but EC failed to significantly reduce cerebral edema and neurological deficits 12 h after intracerebral hemorrhage.6h after treatment with EC,long-term neurological impairment was significantly improved.Memory,depression,anxiety and other behaviors were significantly improved.In addition,we further explore the mechanism of EC action on ICH,We found that EC can reduce nuclear factor-E2-related factor 2-antioxidant response element,Expression of Kelch-like Ech-associated protein 1(KEAP1)in the Nrf2-ARE signaling pathway and promotes Nrf2 nuclear aggregation.It can promote the expression of Superoxide dismutase 1(SOD1)and NADPH-Quinone oxidoreductase 1(NQO1)and reduce Heme oxygenase-1(HO-1);It can also reduce white matter injury and increase neuronal survival after ICH.To verify whether EC has a protective effect on brain injury after ICH through nr F2-ARE signaling pathway,we used Nrf2 gene knockout mice to study the effect of Nrf2 signaling pathway after EC treatment.In conclusion,our study suggests that EC protects brain damage after ICH through Nrf2-dependent and Nrf2-independent pathways.The potential use of EC in the treatment of ICH deserves further investigation.Part Ⅰ Comparison of neurological impairment in mice with intracerebral hemorrhage at different ages Objective: To compare the motor function,memory function,depression and anxiety of male and female cerebral hemorrhage mice of different ages.Methods: Classic collagenase stereotactic injection was used to make ICH mouse model.In this part,mice were randomly divided into six groups: Male aged group,Male middle-aged group,Male young group,Female aged group,Female middle-aged group and Female Young group.Six groups of mice after the success of the building,from day 1 to 42 days,each line Neurological deficit score(NDS),Forelimb placing test(FPT),Hindlimb placing test,(HPT),Corner turn test(CTT);Starting from the 43 rd day,the suspension test(TST),Y-maze test(YMT),Novel object recognition(NOR),Sucrose preference test(SPT),light-dark box(LDB),and Forced swimming test(FST)were performed respectively.By comparing the motor function,memory function,depression and anxiety degree of male and female cerebral hemorrhage mice in three different ages,the situation of cerebral hemorrhage mice in different ages was observed.Results: The results showed that,in terms of motor function,from day 1 to day 42,compared with the elderly group and the middle-aged group,the male and female cerebral hemorrhage mice in the youth group had less motor function damage,and the difference was statistically significant(P<0.05).In terms of memory function,compared with the elderly group and the middle-aged group,the memory function damage of male and female cerebral hemorrhage mice in the youth group was less,and the difference was statistically significant(P<0.05).In terms of depression degree,compared with the elderly group and the middle-aged group,male and female cerebral hemorrhage mice in the youth group had lower depression degree,and the difference was statistically significant(P<0.05).In terms of anxiety level,compared with the elderly group and the middle-aged group of male and female cerebral hemorrhage mice,the anxiety level in the youth group was lower,and the difference was statistically significant(P<0.05).Conclusion: Compared with male and female cerebral hemorrhage mice of different ages,the motor and memory function damage of cerebral hemorrhage mice in the youth group is less,and the degree of depression and anxiety is less.Part Ⅱ Comparison of neurological impairment in mice with intracerebral hemorrhage of different genders Objective: To compare the levels of motor function,memory function,depression and anxiety in elderly,middle-aged and young cerebral hemorrhage mice of different genders.Methods: Classic collagenase stereotactic injection was used to make ICH mouse model.In this part,mice were randomly divided into six groups: Male aged group,Female aged group,Male middle-aged group,Female middle-aged group,Male Young Male group and Female Young Female group.Six groups of mice after the success of the building,from day 1 to 42 days,each line Neurological deficit score(NDS),Forelimb placing test(FPT),Hindlimb placing test,(HPT),Corner turn test(CTT);Starting from the 43 rd day,the suspension test(TST),Y-maze test(YMT),Novel object recognition(NOR),Sucrose preference test(SPT),light-dark box(LDB),and Forced swimming test(FST)were performed respectively.By comparing the motor function,memory function,depression and anxiety degree of male and female cerebral hemorrhage mice in old age,middle age and young age,the situation of cerebral hemorrhage mice of different genders was observed.Results: The results showed that,in terms of motor function,in the elderly cerebral hemorrhage mice,from day 1 to day 42,compared with the male,the female group had slightly less motor function damage,but the difference was not statistically significant.In middle-aged and young mice with cerebral hemorrhage,from day 1 to day 42,the motor function damage of the female group was less than that of the male group,and the difference was statistically significant(P<0.05).In terms of memory function,in elderly cerebral hemorrhage mice,compared with the male group,the female group showed a slight decline in memory function,but the difference was not statistically significant.In middle-aged and young mice with cerebral hemorrhage,the memory function of the female group decreased less than that of the male group,and the difference was statistically significant(P<0.05).In terms of depression degree,the depression degree of the elderly,middle-aged and young cerebral hemorrhage mice was more severe than that of the male group,and the difference was statistically significant(P<0.05).In terms of anxiety level,compared with male mice,female mice in the elderly,middle-aged and young mice with cerebral hemorrhage had more severe depression,and the difference was statistically significant(P<0.05).Conclusion: There is no significant difference in long-term motor function between the aged male and female patients with cerebral hemorrhage.The motor function of middle-aged and young female cerebral hemorrhage mice is better than that of male cerebral hemorrhage mice.There is no significant difference in memory function between the aged male and female cerebral hemorrhage mice.The memory function of middle-aged and young female mice is better than that of male mice with cerebral hemorrhage.The depression and anxiety of the aged,middle aged and young male mice are less than that of the female mice.Part Ⅲ Long-term neuroprotective effect of EC on intracerebral hemorrhage mice Objective: To investigate the neuroprotective effects of EC on motor function and memory impairment,depression and anxiety,and the protective mechanism of EC in middle-aged male and female intracerebral hemorrhage mice.Methods: Classic collagenase stereotactic injection was used to make ICH mouse model.In this study,mice were randomly divided into six groups: Male Sham surgery group,Female Sham surgery group,middle-aged Male placebo group,middle-aged Male treatment group,middle-aged Female placebo group,and middle-aged Female treatment group.Four groups of mice after the success of the building,from day 1 to42 days,each line Neurological deficit score(NDS),Forelimb placing test(FPT),Hindlimb placing test,(HPT),Corner turn test(CTT);Starting from the 43 rd day,the suspension test(TST),Y-maze test(YMT),Novel object recognition(NOR),Sucrose preference test(SPT),light-dark box(LDB),and Forced swimming test(FST)were performed respectively.To observe the neuroprotective effect of EC on middle-aged male and female cerebral hemorrhage mice,the differences between the sham group,the Vehicle group and the EC treatment group were compared.The protein changes of Keap1,Nrf2,SOD1,NQO1,HO-1 in ICH mice brain tissues of the Vehicle group and the EC treatment group were detected by Western blot(WB).Nrf2 gene knockout mice were used to study the role of Nrf2 signaling pathway after EC treatment.Results: The results showed that in terms of motor function,in middle-aged cerebral hemorrhage mice,the motor function damage was less in the EC treatment group compared with the placebo group from day 1 to day 42,and the difference was statistically significant(P<0.05).In terms of memory function,compared with the placebo group,the motor function of middle-aged cerebral hemorrhage mice in the EC treatment group was less impaired,and the difference was statistically significant(P<0.05).In terms of depression,compared with the vehicle group,the motor function of middle-aged cerebral hemorrhage mice in the EC treatment group was less impaired,and the difference was statistically significant(P<0.05).In terms of anxiety level,compared with the vehicle group,the motor function of middle-aged cerebral hemorrhage mice in the EC treatment group was less impaired and the difference was statistically significant(P<0.05).Compared with vehicle group,EC decreased Keap1 expression in Nrf2-ARE signaling pathway and promoted Nrf2 nuclear aggregation,promoted SOD1 and NQO1 expression and decreased HO-1 expression.After ICH,white matter injury and neuronal survival were alleviated,and the differences were statistically significant(P<0.05).In most behavioral tests,EC did not improve neurological impairment in Nrf2 knockout mice.Conclusion: EC treatment can significantly improve short-term and long-term motor impairment,memory impairment,depression and anxiety in middle-aged mice with cerebral hemorrhage.EC protects against brain damage after ICH through Nrf2 dependent and independent pathways. |
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