Rhizoma Anemarrhenae-Cortex Phellodendri Improved Learning And Memory Ability In Diabetic Mice By Inhibiting The Activation Of NLRP3 Inflammasomes

Objective:In recent years,diabetic cognitive dysfunction has received widespread attention.In this experiment,by measuring the content of NLRP3 inflammasome-related protein in the brain,the mechanism of the effect of Rhizoma Anemarrhenae(RA)-Cortex Phellodendri(CP)on the improvement of learning and memory ability of diabetic mice was explored,and the changes of related chemical components before and after salt burning were measured,which provided an experimental basis for the material basis of RA-CP to improve diabetic cognitive impairment.Methods:First,the activation of NLRP3 inflammasome in the brain of diabetic mice was verified.Twenty-five healthy male ICR mice with SPF level were adaptively fed for 1 w,randomly divided into blank group(n=10)and model group(n=15)according to body weight,and the model group mice were fed 4 w with high sugar and high fat,combined with intraperitoneal injection of low-dose streptozocin(STZ)8Omg·kg-1 to replicate the diabetes model,and after 2 w,fasting blood glucose was determined,and the blood glucose value was ≥11.1mmol·L-1 was the mold standard,and the weight was measured after successful model replication,and the behavioral test was determined by Morris water maze experiment and hot Y maze experiment,Western Blot method to determine NLRP3,Caspase-1,IL-18 protein content,and ELISA method to determine IL-1β content.Secondly,the effect of RA-CP on the brain of NLRP3 inflammasome in diabetic mice.The 140 mice were reproduced according to the first part of the diabetes model,divided into blank group(n=20)and model replication group(n=120),and after successful model replication,they were divided into model group(n=25),ShengRA-ShengCP group(Sheng group,n=20),YanRA-YanCP group(Yanzhi group,n=20),sitagliptin group(n=20),biological group and salt burning group gavage 3.916g·kg-1 corresponding RA-CP liquid,sitagliptin group gavage 0.016g·kg-1 sitagliptin aqueous solution,body weight and fasting blood glucose were measured every 2 w during administration,OGTT and ITT experiments were performed at the 8th w of gavage administration,autonomous activity experiments were performed at the 2nd,4th,6th,and 8th w,new object recognition experiments were performed at the 4th and 8th w,and Morris water maze experiments and hot Y maze experiments were also performed at the 8th w.After the behavioral experiment,glucose metabolism-related indexes(GSP,C peptide,INS)were measured,and the content changes of HOMA-IR and lipid metabolism-related indexes(TC,TG,HDL-C,LDL-C)were calculated.Western Blot measured NLRP3,Caspase-1,IL-18,IL-1β protein content in brain tissue,and IL-1β content by ELISA method.HE staining observed morphological and structural changes in hippocampal neurons.Finally,HPLC was used to explore the changes of new mangiferin,mangiferin and berberine content before and after RA-CP salt burning,so as to determine the material basis of salt burning on the efficacy.Results:After 2 w of model replication,blood glucose and body weight of mice in the model group increased significantly compared with the blank group(P<0.01).The number of learning and errors in hot Y maze increased significantly(P<0.01).The evasion latency period was longer than that of the blank group,and there were significant differences in the 8th,9th.10th and 11th test periods(P<0.05,P<0.01).The contents of NLRP3,Caspase-1,IL-18 and IL-1β increased significantly(P<0.01),indicating that the NLRP3 inflammasome in the brain of diabetic mice was activated,and the learning and memory ability of diabetic mice decreased.Compared with the blank group,the weight and blood glucose of mice in the model group increased significantly(P<0.01)within 8 w;in OGTT and ITT experiments,the blood glucose value and area under the curve at each time point increased significantly(P<0.01),GSP,C peptide,INS,HOMA-IR,TC and TG were significantly increased(P<0.01),HDL-C was significantly reduced(P<0.05),and LDL-C was significantly increased(P>0.05).Compared with the model group,the drug group significantly reduced body weight and blood glucose(P<0.05,P<0.01)during administration,significantly decreased blood glucose value and area under the curve at different time points in OGTT and ITT experiments(P<0.05,P<0.01),GSP,INS.TC and TG content was significantly reduced(P<0.05,P<0.01),HDL-C was significantly increased(P<0.05,P<0.01),LDL-C decreased but there was no significant difference(P>0.05),C-peptide has different decreasing tendencies(P<0.05,P<0.01).Behavioral experiments showed that compared with the blank group,the number of autonomous activity and standing in the model group was significantly reduced(P<0.01),the recognition index of new objects at the 8th w was significantly decreased(P<0.05),the evasion latency period was prolonged,and the retention time and number of entries in the target quadrant were significantly reduced(P<0.05,P<0.01).The number of learning and errors increased significantly(P<0.01);Compared with the model group,the number of voluntary activities and standing in the drug group increased to different degrees(P<0.01),the new object recognition index at the 8th w increased significantly(P<0.05,P<0.01),the number of learning and errors decreased significantly(P<0.05,P<0.01),and the escape latency period decreased(P<0.05,P<0.01),the percentage of target quadrant entries increased significantly(P<0.05).It was found that compared with the blank group,the number of neuronal pyramidal cells in the CA1 region of mice in the model group was reduced,the cells in the CA3 and DG regions were slightly less,the arrangement was loose,the degree of looseness in the CA3 region was serious,and some cells were deeply stained and solidified.Compared with the model group,the arrangement of hippocampal neuronal cells in the drug group was more regular,and the looseness phenomenon was reduced.Western Blot and ELISA experiments on mice in each group showed that compared with the blank group,the NLRP3related protein content of the model group was significantly increased(P<0.01).Compared with the model group,the content of NLRP3-related protein in the drug group was significantly reduced(P<0.01).After salt burning,the content of new mangiferin decreased to 0.6299%,the content of mangiferin increased to 0.9755%,and the content of berberine decreased to 2.6691%after salt burning of CP.Conclusion:NLRP3 inflammasome in the brain tissue of diabetic mice is activated,which in turn leads to a decline in learning and memory ability.RA-CP can regulate the abnormal glycolipid metabolism of diabetic mice,improve their learning and memory ability by inhibiting the activation of NLRP3 inflammasome and reducing the expression of related inflammatory factors,and the increase in mangiferin content in the salt group is the material basis for its effect to be better than that of the raw group.