The Effects Of Butyrate On The Cognitive Function Of Mice With Circadian Rhythm Disorder By IL-17/IL-10

Objectives: 1.To establish a circadian rhythm disorder model.2.To explore the effect of circadian rhythm disorder on cognitive function of mice and the improvement of butyrate after intervention.3.To explore the effect of circadian rhythm disorder on the morphological changes of hippocampal neurons in mice and the effect of butyrate intervention on them..4.To investigate the changes of interleukin 17(IL-17)and IL-10 protein levels secreted by spleen helper cell 17(Th17)and regulatory T cells(Treg)in mice with circadian rhythm disorder and after the intervention of butyrate.5.To explore the changes of RORγt,Foxp3,IL-17,IL-10 m RNA transcription levels in hippocampus of mice with circadian rhythm disorder and after butyrate intervention.Methods: Male specific-pathogen-free(SPF)C57BL/6J mice aged 6-8 weeks were randomly divided into 4 groups with 6 mice in each group: normal control(light-dark,LD)group :12h light :12h darkness(12L:12D;L = 1000 lux;D=0lux)for 4weeks;Circadian rhythm disorders(light-light,LL)group: 24-hour constant light,the intensity of light maintain 1000 lux,for 4 weeks;Circadian rhythm disorders +butyrate(LL+ B)Group: 24-hour constant light with light intensity of 1000 lux.During constant light,butyrate(1.2 g/kg)is administered daily by instillation for 4weeks.Circadian rhythm disorders + PBS(LL+ PBS)group: 24-hour constant light,with light intensity of 1000 lux,in the process of continuous illumination,daily intragastric administration of PBS 1.2 g/kg,for 4 weeks.All experimental mice were allowed to adapt to the SPF environment for one week before the start of the experiment under the conditions of 12L:12D,L=1000lux,and D=0lux.Adequate water and food were ensured during the experiment.After exposed to constant light for 4 weeks,all mice were subjected to Morris water maze(MWM)place navigation training,followed by spatial probe test after 5 days of circadian disorder.After the test,the mice were sacrificed and the morphological changes of neurons in hippocampal tissues of mice were observed by HE staining.The contents of IL-17、IL-10 in spleen were determined by Elisa,RT-q PCR was used to determine the transcriptional level of IL-17、IL-10、RORγt、Foxp3 m RNA of in hippocampal tissues of each group.Results: 1.The body weight of mice in the LL group increased significantly compared with that in the LD group(P<0.05).The body weight of the mice significantly decreased after exogenous butyrate supplementation,and the difference was statistically significant(P<0.05).2.The results of behavioral test showed that the escape latency of LD and LL+B group decreased gradually with the prolongation of training time,while the escape latency of LL and LL + PBS group did not change significantly.Compare the incubation period of mice in each group on the fourth day,The LL group was significantly longer than that of LD group(P < 0.05),and the LL+B group was significantly shorter than that of LD group(P < 0.05).In the space exploration experiment,compared with LD group,the number of crossing platform in LL group decreased statistically(P < 0.05);after butyrate supplementation,the number of crossing platform increased statistically(P < 0.05).3.HE staining showed that under light microscope,the structure of neurons in LD group was complete,the arrangement was orderly,,and the layers were 3-5 layers,the cytoplasm was stained evenly,and the nucleolus were clearly visible.In LL group,The number of neurons decreased,the gap increased,the level was unclear,the cell boundary was unclear;after the butyrate intervention,the arrangement of neurons was relatively neat,the cell boundary was clear,and a small number of neurons were abnormal in morphology and structure.4.Compared with the LD group,the expression of IL-17 protein in the spleen of LL group increased significantly(P < 0.05).while IL-10 decreased significantly(P <0.05).After butyrate supplementation,the expression of IL-17 protein in the spleen of mice was decreased,and the difference was statistically significant(P < 0.05).And the expression of IL-10 increased,and the difference was statistically significant(P <0.05).5.Compared with the LD group,the expression level of RORγt、IL-17 m RNA in the hippocampal tissue of LL group was increased,and the difference had a statistical significance(P<0.05,P<0.05).The expression level of Foxp3、IL-10 m RNA was decreased,and the difference had a statistical significance(P<0.05,P<0.05).After butyrate supplementation,the expression level of RORγt、IL-17 m RNA in thehippocampal tissue decreased,and the difference had a statistical significance(P<0.05,P<0.05).The expression level of Foxp3、IL-10 m RNA increased,and the difference had a statistical significance(P<0.05,P<0.05).Conclusions: 1.The model of CRD was successfully established.After CRD,the mice gained weight,energy metabolism was unbalanced,and the mice lost weight after exogenous supplementation of butyrate.2.After CRD,the cognitive function of mice was impaired,and the cognitive function was improved after supplementation of butyrate.3..The CRD changed the morphology of hippocampal neurons in mice.After supplementing butyrate,he abnormal morphology of hippocampal neurons in mice decreased.4.CRD can induce IL-17 / IL-10 related inflammatory response in peripheral immune system of mice.Butyrate may reduce this inflammatory response.5.CRD promotes inflammatory damage to the central nervous system by inducing IL-17 / IL-10 in the hippocampus of mice.Supplementation with butyrate may balance IL-17 / IL-10 in the hippocampal tissue of mice.In summary,CRD may impair cognitive function by disrupting the balance of peripheral immune system and central IL-17/IL-10,and butyrate may improve cognitive function by correcting this imbalance.