| Objective:1.To explore the neuroprotective effect of GLP-1/GIP dual-receptor agonist DA3-CH and GLP-1 single-receptor liraglutide on cerebral ischemia-reperfusion injury in rats with diabetes(neurological function)Defect score,infarct size)2.To investigate the effects of DA3-CH and liraglutide on endoplasmic reticulum stress and apoptosis in a rat model of cerebral ischemia-reperfusion injury with diabetes,and to clarify its effects on cerebral ischemia-reperfusion in diabetic rats Neuroprotective mechanisms of injury(endoplasmic reticulum stress-related proteins,apoptosis-related proteins)3.To observe and compare the strength of neuroprotective effect of DA3-CH and liraglutide in a rat model of cerebral ischemia-reperfusion injury with diabetes,and the effect on endoplasmic reticulum stress and apoptosisMethods:1.Groups:72 SD male rats were randomly divided into 4 groups:① blank group(Sham group,n=18);model group(Saline group,n=18);DA3 treatment group(DA3 group,n=18));Liraglutide treatment group(Lir group,n=18)2.Intervention measures:No treatment was given to the Sham group,and they were raised in the same environment as the other groups.The other 3 groups used STZ to induce the diabetes model.After successful diabetic membrane formation,DA3-CH and liraglutide(10 mmol/kg,once/day)were injected intraperitoneally3.Determination method:14 days after intraperitoneal injection of drugs,blood glucose of rats was measured.Subsequently,the Saline group,DA3 group,and Lir group were used to prepare the cerebral ischemia-reperfusion model(MCAO).After 2 hours of ischemia and 22 hours of reperfusion,the modified nervous system severity score(mNSS score)was evaluated,and the brain was decapitated and taken.The area of cerebral infarction was measured by TTC staining,and endoplasmic reticulum stress-related proteins(GRP78,CHOP,Capasel2),and apoptosis-related proteins(NEUN,Bax,Bcl-2)were determined by immunohistochemistry and Western Blot.)Expression levelResults:1.Blood glucose level:the blood glucose level of the drug intervention group was lower than the model group(DA3:21.55 ± 1.66 vs 24.5 ± 2.53,P<0.01;Lir:20.90 ±2.30 vs 24.5 ± 2.53,P<0.01),but there was no significant difference in blood glucose levels between the DA3-CH group and the liraglutide group(21.55±1.66 vs 20.90 ±2.30,P>0.05)2.Infarct area:The infarct area in the drug intervention group is smaller than that in the model group(DA3:12.58 ±3.85 vs 39.57 ± 3.12,P<0.001;Lir:18.69 ±2.90 vs 39.57 ±3.12,P<0.001).The infarct size was smaller than the liraglutide group(12.58 ±3.85 vs 18.69 ± 2.90,P<0.01)3.Modified nervous system severity score(mNSS):The mNSS score of the drug intervention group was smaller than the model group(DA3:8.67 ± 1.63 vs 12.50 ± 1.76,P<0.001;Lir:10.50 ± 1.08 vs 12.50 ±1.76,P<0.01).The mNSS score of the DA3 group was lower than that of the liraglutide group(8.67±1.63 vs 10.50±1.08,P<0.05)4.Endoplasmic reticulum stress-related protein expression levels4.1.Expression level of endoplasmic reticulum stress marker GRP78:Immunohistochemical results showed that the number of GRP78 positive cells in the cerebral cortex of the drug intervention group was less than that in the model group(DA3:10.00 ± 2.68 vs 33.33 ± 1.89,P<0.001;Lir:15.67 ± 1.85 vs 33.33 ± 1.89,P<0.001);the number of GRP78 positive cells in the DA3-CH group was less than that in the liraglutide group(10.00 ± 2.68 vs 15.67 ± 1.85,P<0.001)Western blot results showed that the expression of GRP78 in the cerebral cortex of the drug intervention group was less than that of the model group(DA3:0.50 ± 0.11 vs 0.73 ± 0.14,P<0.001;Lir:0.58 ±0.13 vs 0.73 ± 0.14,P<0.001);The expression of GRP78 in the DA3-CH group was lower than that in the liraglutide group(0.50 ± 0.11 vs 0.58 ± 0.13,P<0.001)4.2 Expression levels of endoplasmic reticulum stress injury proteins CHOP and Capase12:The immunohistochemical results showed that the number of CHOP positive cells in the cerebral cortex of the drug intervention group was less than that of the model group(DA3:8.67 ± 1.37 vs 39.67 ± 2.25,P<0.001;Lir:12.33 ± 1.77 vs 39.67 ± 2.25,P<0.001),the number of CHOP-positive cells in the DA3-CH group was less than that in the liraglutide group(8.67 ± 1.37 vs 12.33 ± 1.77,P<0.01).The number of Capase12 positive cells in the cerebral cortex of the drug intervention group was less than that of the model group(DA3:12.67 ± 1.37 vs 30.00 ± 1.79,P<0.001;Lir:15.67 ± 2.73 vs 30.00±1.79,P<0.001),DA3-CH The number of Capase12 positive cells in the group was less than that in the liraglutide group(12.67 ± 1.37 vs 15.67 ± 2.73,P<0.01)Western blot results showed that the expression of CHOP in the cerebral cortex of the drug intervention group was less than that of the model group(DA3:0.49 ± 0.04 vs 0.84 ± 0.08,P<0.001;Lir:0.76 ± 0.03 vs 0.84 ± 0.08,P<0.001),The expression of CHOP in the DA3-CH group was less than that in the liraglutide group(0.49±0.04 vs 0.76 ± 0.03,P<0.001);the expression of Capase12 in the cerebral cortex of the drug intervention group was less than the model group(DA3:0.45 ± 0.08 vs.0.68 ± 0.12,P<0.001;Lir:0.47 ± 0.07 vs 0.68 ± 0.12,P<0.001),the expression of Capase12 in the DA3-CH group was less than that in the liraglutide group(0.45 ± 0.08 vs 0.47 ± 0.07,P<0.01)5.Expression levels of apoptosis-related proteins5.1 Expression level of neuron survival marker NEUNImmunohistochemical results showed that the number of NEUN positive cells in the cerebral cortex of the drug intervention group was greater than that of the model group(DA3:24.56 ± 2.25 vs.10.67 ± 1.86,P<0.001;Lir:18.43 ± 2.35 vs.10.67 ± 1.86,P<0.001);the number of NEUN positive cells in DA3-CH group was more than that in liraglutide group(24.56 ± 2.25 vs 18.43 ± 2.35,P<0.001)Western blot results showed that the expression of NEUN in the cerebral cortex of the drug intervention group was greater than that of the model group(DA3:0.66 ± 0.08 vs 0.46 ± 0.09,P<0.001;Lir:0.58 ± 0.09 vs 0.46 ± 0.09,P<0.001);The expression of NEUN in the DA3-CH group was greater than that in the liraglutide group(0.66 ± 0.08 vs 0.58 ± 0.09,P<0.001)5.2 Expression of pro-survival factor Bcl-2 and pro-apoptotic factor BaxWestern blot results showed that Bax expression in the cerebral cortex of the drug intervention group was less than that of the model group(DA3:0.33 ± 0.03 vs 0.69 ±0.17,P<0.001;Lir:0.45 ± 0.15 vs 0.69 ± 0.17,P<0.001),The expression of Bax in the DA3-CH group was lower than that in the liraglutide group(0.33 ± 0.03 vs 0.45 ± 0.15,P<0.001).The expression level of Bcl-2 in the cerebral cortex of the drug intervention group was greater than that of the model group(DA3:0.75 ± 0.07 vs 0.38 ± 0.04,P<0.001;Lir:0.58 ± 0.05 vs 0.38 ± 0.04,P<0.001),DA3-CH The expression of Bcl-2 in the group was greater than that in the liraglutide group(0.75 ± 0.07 vs 0.58 ± 0.05,P<0.001)Conclusion:1.DA3-CH and liraglutide have obvious neuroprotective effects in a rat model of cerebral ischemia-reperfusion injury with diabetes,which can reduce the infarct size and the neurological deficit score2.DA3-CH and liraglutide exert neuroprotective effects in a rat model of cerebral ischemia-reperfusion injury with diabetes by inhibiting endoplasmic reticulum stress and thereby reducing apoptosis3.DA-CH has stronger neuroprotective effect,inhibitory effect on endoplasmic reticulum stress and apoptosis in rat models of cerebral ischemia-reperfusion injury with diabetes than liraglutide. |
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