| Objective Wistar rats were used to establish Type 2 diabetic models with intermittent hyperglycemia,and compared with the control group and persistent hyperglycemia group.The effect of blood glucose fluctuation on rat cognitive,hippocampal oxidative stress indicators and expression of apoptosis-related proteins was revealed.We tried to reveal the possible mechanism of blood glucose fluctuations exacerbating cognitive impairment in type 2 diabetes,while using butylphthalide to provide intervention,providing a certain laboratory basis for the treatment and intervention of diabetes with cognitive impairment.Methods 1.Sixty Wistar rats(male)were randomly divided into a diabetes group(type 2 Diabetes Mellitus,T2DM)and a control group(Control,C).After the diabetes group was induced by high-sugar and high-fat diet combined with low-dose streptozotocin(STZ)intraperitoneal injection to establish a type 2 diabetic rat model,the diabetic rats were then randomly divided into the Constant high glucose group(CHG),Intermittent high glucose group(IHG),Constant high glucose group treated with L-3-n-butylphthalide(CHG + NBP)and Intermittent high glucose group treated with L-3-n-butylphthalide(IHG + NBP),in which the rat models of the IHG and IHG+ NBP groups were established by changing the life rhythm and eating habits of rats.2.Observing the general conditions of rats in each group,including mental state,reaction ability,skin of limbs,food intake,water intake,urine output and blood glucose level.3.Evaluating the success of the blood glucose fluctuation model: At the6 th week after the blood glucose fluctuation begins,and observeing the blood glucose situation at 8 o’clock-12 o’clock,15-18 o’clock every day in the rat,and use the two-factor repeated measurement variance model statistical analysis.Comparing between the two groups by LSD-t method showed that IHG,IHG + NBP weresignificantly different from other groups,and the blood glucose fluctuations were successfully modeled.4.Detection of memory ability: Morris water maze experiment was used to test the spatial memory learning ability of rats.5.Detecting glutathione peroxidase(GSH-Px),total superoxide dismutase(T-SOD),and malondialdehyde(MDA)in hippocampus of each group of rats.6.To detect the expression of Bcl-2,Bax,Caspase-3 proteins in hippocampus of rats.Results 1.Changes in growth and metabolism indicators of rats: Compared with group C,rats in the T2 DM group have slower movements,a weaker mental state,a dull coat,and a slow response to external stimuli,and they have obvious symptoms of polydipsia and polyuria.2.Metabolic changes in rats: Compared with group C,the water consumption and urine output of rats in the T2 DM group were significantly increased(P <0.05,P <0.01);however,there was no difference in water consumption and urine output among the T2 DM rats(P>0.05);the intake of rats in the CHG group was higher than that in the C and IHG groups(P<0.05),but there was no significant difference in the intake of the rats in the C group and the IHG group(P> 0.05).3.Blood glucose changes in rats: After 6 weeks of blood glucose fluctuations,blood glucose differences between groups were statistically significant(F= 67.388,P<0.05=,and differences between the four groups were significantly at each time point(P<0.05);the time trend of blood glucose daily was significantly(F=6.882,P<0.05=;The interaction between groups and time was not significantly(F=2.680,P>0.05).That is,there was a significantly difference in the changes in the measured values of the five groups of rats at 9 time points.4.Changes in learning and memory abilities of rats: The differences between the five groups of rats during the positioning and navigation test to avoid the incubation period were significantly(F=5.457,P<0.05).The temporal change trend was significantly(F=104.190,P<0.05),showing the overall trend of the 5 groups,all of them decreased.The interaction between group and time was significantly(F=2.247,P<0.05),that is to say,there was a statistical difference in the change in the escape latency measured by the positioning and sailing experiments of the 5 groups of rats at 4 days.In the space exploration experiment on the 5th day,the target quadrant activity time ratio of the 5 groups was compared,andthe difference was significantly(P<0.05).5.Detection of oxidative stress indicators:Compared with the C group,the MDA content in the T2 DM group was increased(P<0.05),and the SOD and GSH-Px levels were decreased(P<0.05).Compared with the CHG group,the IHG group was larger.Rat MDA content increased(P<0.05),SOD and GSH-Px content decreased(P<0.05).Further comparison,compared with the CHG group,the MDA content of the rats in the CHG + NBP group was reduced(P<0.05),and the GSH-Px and SOD contents were increased(P<0.05).The same was true between the IHG group and the IHG + NBP group.6.Detection of apoptosis indicators(Western Blot method): Compared with group C,the expression of Bcl-2protein in the T2 DM group was reduced,and the difference was statistically significant(P<0.05);compared with the CHG group,The expression of IHG group was significantly reduced(P<0.05).The contents of Bax and Caspase-3 in the hippocampus of the T2 DM group were significantly higher than those in the C group(P<0.05),and the contents of the IHG group were significantly higher than those in the CHG group(P<0.05).Further analysis showed that compared with the CHG group,the expression of Bcl-2 in the CHG + NBP group increased,but the difference was not significant(P>0.05);the protein content of Bax and Caspase-3decreased,and the difference was not significant(P>0.05)This was also true between the IHG group and the IHG + NBP group(P>0.05).Conclusion A blood glucose fluctuation model can be established by changing the eating habits and biological rhythms of rats.2.Volatile hyperglycemia can aggravate the occurrence of apoptosis and oxidative stress injury in hippocampus,which is likely to be an important mechanism that triggers cognitive impairment in T2 DM rats.3.Butylphthalide may have a certain protective effect on the cognitive function of type 2 diabetic rats with blood glucose fluctuation through the above mechanisms. |
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