Study On The Role Of Heterozygous Disruption Of Beclin 1 In Arsenic-induced Neurobehavioral Deficits And Gut Microbiota Changes

PART I STUDY ON THE ROLE OF HETEROZYGOUS DISRUPTION OF BECLIN 1 IN ARSENIC-INDUCED NEUROBEHAVIORAL DEFICITSObjective:To observe the effects of arsenite on neurobehavior and brain morphology though the animal model of arsenite exposure via drinking water with heterozygous disruption of beclin 1 mice.To explore the role of heterozygous disruption of beclin1 in the effect of arsenic on neurobehavior and brain morphology.Methods:A total of 44 healthy female wild-type mice(beclin 1^+/+)and heterozygous disruption of beclin 1 mice(beclin 1^+/-),aged 7-8weeks,weighted 20-24 g,were divided into 4 groups as follows:beclin1^+/++Vehicle group,beclin 1^+/-+Vehicle group,beclin 1^+/++Arsenite group and beclin 1^+/-+Arsenite group?n=11 each group?.The Vehicle groups and Arsenite groups were treated with distilled water and 50 mg/L arsenite solution respectively.During exposure,the change in body weight of mice was recorded,the general behavior and the water consumption of mice were observed.After 6 months exposure,the morphology and number of neurons in mice brain were observed by H&E staining and Nissl staining.Morris water maze was used to evaluate the spatial memory and learning ability of mice.Open filed experiments were used to evaluate the autonomous behavior of mice.Results:?1?There was no significant difference in the body weight,general behavior and water consumption of beclin 1^+/+mice and beclin1^+/-mice in four groups.?2?H&E staining showed that no significant damage was observed in cerebral cortex and hippocampus between beclin1^+/+mice and beclin 1^+/-mice in Vehicle groups.Compared with the Vehicle groups,the cortex and hippocampus of the beclin 1^+/++Arsenite group had varying degrees of damage.But no significant damage was observed in beclin 1^+/-+Arsenite group.?3?Nissl staining showed that the nissl bodies in cerebral cortex and hippocampus were no significant change between beclin 1^+/+mice and beclin 1^+/-mice in Vehicle groups.Compared with the Vehicle groups,the nissl bodies in cerebral cortex and hippocampus of beclin 1^+/++Arsenite group were vacuolated degeneration,blurred edges and lighter staining,but no significant change in beclin 1^+/-+Arsenite group.?4?Morris water maze showed that the escape latency,swimming distance,the number of platform crossings and the time spent in the target quadrant were no significant differences between beclin 1^+/+mice and beclin 1^+/-mice in Vehicle groups?P>0.05?.Compared with Vehicle groups,the beclin 1^+/++Arsenite group significantly reduced the number of platform crossings and the time spent in the target quadrant?P<0.05?,significantly increased the escape latency and swimming distance?P<0.05?;while the beclin 1^+/-+Arsenite group was no significant change?P>0.05?.beclin 1^+/-+Arsenite group was significant difference compared with the beclin 1^+/+Arsenite group in the number of platform crossings and the time spent in the target quadrant?P<0.05?.?5?Open filed showed that the total distance,distance move in central and central square duration were no significant difference between beclin 1^+/+mice and beclin 1^+/-mice in Vehicle groups?P>0.05?.Compared with Vehicle groups,beclin 1^+/++Arsenite group significantly reduced the distance move in central and central square duration?P<0.05?,while the beclin 1^+/-+Arsenite group was no significant change?P>0.05?.beclin 1^+/-+Arsenite group was significant difference compared with the beclin 1^+/++Arsenite group in the distance move in central and central square duration?P<0.05?.Conclusion:Chronic exposure to arsenic via drinking water can induce neuronal damage and neurobehavioral deficits in mice.Heterozygous disruption of beclin 1 mitigates neuronal damage and neurobehavioral deficits induced by arsenic exposure.PART II STUDY ON THE ROLE OF HETEROZYGOUS DISRUPTION OF BECLIN 1 IN ARSENIC-INDUCED GUT MICROBIOTA CHANGESObjective:The purpose of this study was to explore the effect of arsenite exposure on gut microbiota and compare the differences of gut microbiota between beclin 1^+/+mice and beclin 1^+/-mice.To elucidate the role of heterozygous disruption of beclin 1 in arsenite-induced changes in gut microbiota.Based on this,the effect of changes in gut microbiota on brain function in mice was explored.Methods:Aseptically collect mouse feces,extract DNA,subjecte to16S ribosomal RNA gene sequencing,and then conduct biological information analysis.RT-qPCR experiments were used to detect the mRNA expression levels of cerebral intestinal peptides?Sstr1,Sstr2,Vip,Tac1,Sst,Ghsr?and major neurotransmitters?Hrt3a,Hrt4?in mouse cortical tissue.Results:?1?Alpha diversity analysis showed that Sobs,Shannon,Simpson,ace and Chao indexes were no significant difference between beclin 1^+/+mice and beclin 1^+/-mice in Vehicle groups?P>0.05?.Compared with Vehicle groups,beclin 1^+/++Arsenite group and beclin1^+/-+Arsenite group significantly decreased on Sobs,Shannon,ace and Chao indexes?P<0.05?,but significantly increased on Simpson indexes?P<0.05?.beclin 1^+/-+Arsenite group was significantly higher than beclin1^+/++Arsenite group on Shannon indexes?P<0.05?,but significantly lower than beclin 1^+/++Arsenite group on Simpson indexes?P<0.05?.?2?Venn diagram showed that four groups shared the compositional overlap of 460 core microbiota and each group has unique microbiota.?3??diversity analysis showed that there was obvious difference in the gut microbial community compositions among four groups.?4?The bacterial taxonomic composition showed that,the abundance of Proteobacteria and Deferribacteres were significantly decreased in beclin 1^+/++Arsenite group?P<0.05?,and had no significant changes in beclin 1^+/-+Arsenite group?P>0.05?.?5?RT-qPCR showed that the mRNA expression levels of Sstr1,Sstr2,Vip,Tac1,Sst,Ghsr,Hrt3 and Hrt4 were no difference between beclin 1^+/+mice and beclin 1^+/-mice in Vehicle groups?P>0.05?.Compared with Vehicle groups,the mRNA expression levels of Sstr1,Sstr2,Vip,Ghsr and Hrt3a were significantly decreased in beclin 1^+/++Arsenite group?P<0.05?,but were no difference in beclin 1^+/-+Arsenite group?P>0.05?.The mRNA expression levels of Sstr1,Sstr2,Vip,Tac1,Hrt3a and Hrt4 were significant differences between beclin 1^+/++Arsenite group and beclin 1^+/-+Arsenite group?P<0.05?.Conclusion:Chronic exposure to arsenic via drinking water can induce gut microbiota and heterozygous disruption of beclin 1 mitigates arsenite-induced changes in gut microbiota.Changes in gut microbiota can be transmitted to the brain through the gut-brain axis and have an impact on brain function.