Study On The Relationship Between MIF, MCP-1 And Alzheimer’s Disease, Vascular Dementia

ObjectiveThis study aimed to evaluate evaluated the changes of MIF,MCP-1 expression in peripheral blood of patients with Alzheimer’s disease(AD),vascular dementia(VD)and the relationship with dementia severity.Based on this,make the vascular dementia model by repeated close and open the rat’s common carotid arterys in both sides and injecting sodium nitroprusside(SNP) intraperitoneal,using Morris water-mazed palace to test the rat’s ability of study and memory and prove the succession of making the model.The expression of MIF,MCP-1 around the focus of CA1 in hippocampus of the rat models of vascular dementia was evaluated.The aim of this work was to clarify the role and Significance ofMIF、MCP-1 in the AD and VD.MethodsPart 1:The change of MIF、MCP-1 in the peripheral blood of patients with AD and VD1.Clinical data:patients was admitted to hospital of Guangdong provincial people’s hospital from june 2006 to january 2008.64 AD patients including 34 males and 30 females with an average age of 53～90(75.84±8.11).37 VD patients, including 28 males and 9 females with an average age of 56～91(74.35±8.93),50 healthy including 29 males and 21females with an average age of 53～87 (72.71±9.99) was used as control group matched with AD,VD in age,risk factor and complication.People without disease of heart,liver,and kidney;without infection, tumor,and rheumatism.2.The patients were evaluated by mini-mental state examination(MMSE),GDS. HIS,ADL and depression,anxiety scale inspection.The plasma level of MIF,MCP-1 was evaluated by ELISA.Peripheral blood mononuclear cell(PBMC) was collected. Then,the PBMC expression of MIF,MCP-1 mRNA was measured by quantitative PCR.3.All the numerical data have been statistical analysis by SPSSll.0 for windows statistical package.Measurement data was said in(mean±standard deviation),independent samples comparison with single-factor analysis of variance; count information use the scores,compared groups with x~2 test,a single factor regression use Spearman correlation analysis,and more regression using multiple regression analysis,F introduction=3.84,F deleted=2.71.P＜0.05 as statistical significance.Part 2:The levels ofMIF,MCP-1 around the focus of CA1 in hippocampus at the rat models of vascular dementia.1.Animal:The 20-week-age Wistar rats were supplied by Laboratory Animal Center of Guangzhou University of Traditional Chinese Medicine and raised in Research Center of Medical Science of Guangdong Provincial People’s Hospital. There were half of females and males in the rats and the weight of them was 300±20g.2.The vascular dementia model was made by repeating closing and opening the rat’s common carotid arterys on both sides and injecting sodium nitroprusside(SNP) intraperitoneally.The succession of making the model was proved by testing the rat’s ability of study and memory through Morris water-mazed palace test.The rats were divided equally into controlled group,model group,sham-operated group and each group has 8 cases.All the animals were assessed by Morris water-mazed palace test before and after operation.After one week the rats were killed and the cerebral tissues arround hippocampus were excised.One half of the cerebral tissue was stored in liquid nitrogen for extraction of protein and mRNA,another half was fixed by 10% formaldehyde for geometry morphologic measuring by hematoxylin and eosin(HE) staining and immunohistochemistry staining.3.All the numerical data have been statistical analysis by SPSS11.0 for windows statistical package.Measurement data was said in(mean±standard deviation),the positioning navigation test use Repeated measurements of the multi-factor analysis of variance,the space exploration test use single-factor analysis of variance.The remaining statistically was same as the first chapterResultsPart 1 The change of MIF、MCP-1 in the peripheral blood of patients with AD and VD1.VD group＞AD group＞control group,AD and VD group were higher than the control group(P＜0.018),the difference is statistically significant.But between AD and VD group,there was no significant difference(P＞0.102).2.According mini-mental state examination(MMSE) scores(severe group≤10, moderate group 11～20,mild group 21～24) and Global Deterioration Scale(GDS) grade(severe group 6～7,moderate group 4～5,mild group 3) to evaluation the severity of patients,if when the two score are inconsistent we use the more serious levels.AD Patients were graded into 18 cases in severe group,33 cases in moderate group and 13 cases in mild group.VD Patients were graded into 16 cases in severe group,36 cases in moderate group and 5 cases in mild group.The levels of plasma and PBMC mRNA of MIF,MCP-1 with AD and VD: severe group were higher than moderate and mild group(P＜0.041);but,there were no difference between moderate and mild group(P＞0.100). 3.The multiple regression analysis:MIF protein concentration related on MMSE score,cholesterol,C-reactive protein and MIF mRNA expression;MCP-1 protein concentration related on MMSE score,systolic blood pressure and MCP-1 mRNA expression;MIF,MCP-1 protein have no obvious correlation with gender,history of hypertension,diabetes,history,history of high blood lipids,smoking history,history of alcohol abuse,ESR,diastolic blood pressure,fasting glucose,triglycerides, low-density lipoprotein,high-density lipid Protein,folic acid and vitamin B 12.4.The Correlation Analysis between the expression of MIF,MCP-1 in patients with AD and VD and MMSE score:in AD group MIF,MCP-1 not only the concentration in the plasma r=-0.415,r=-0.366)but also the PBMC mRNA expression level(r=-0.47,r=-0.325) was negatively correlated with MMSE score(P＜0.009). in VD group MIF,MCP-1 not only the concentration in the plasma(r=-0.516, r=-0.3496)but also the PBMC mRNA expression level(r=-0.578,r=-0.505) as well was negatively correlated with MMSE score(P<0.015).Part 2:The levels ofMIF,MCP-1 around the focus of CAl in hippocampus at the rat models of vascular dementia.1.Determination of memory:test the memory of of the rats by Morris water maze,and found that through training of swimming in the maze have learned of the rats,vascular dementia model rats compared to sham-operated group,control group rats,the learning,memory capacity significantly lower.In the positioning navigation test,the model group compared to the sham-operated group and the control group escape latency significantly longer(P=0.000);In the space exploration test,the model group compared with the sham group,control group,in the original platform quadrant significantly reduce the number of cross platform(P＜0.007),but,there was no significant difference between the sham and control groups in these two experiments(P＞0.359).2.Pathological changes:HE staining,200 times normal light microscope control group and sham-operated group rat at hippocampal CA1 pyramidal cells are close and regular,and large round cells,nucleolus clear.But vascular dementia model rats hippocampal CA1 pyramidal cell layer depletion almost completely,a large number of inflammatory cell can be seen in fracture level pyramidal cells,nerve cells in apparent loss,and smaller cell size,and taking on the phenomenon of Chenghu pyknosis.3.The immunohistochemical changes:vascular dementia model rats compared with sham-operated and the control group,MIF,MCP-1 expression increased.in hippocampus CA1.4.It was applied western blot and quantitative PCR to detect the level of MIF、MCP-1 protein and the expression of their mRNA around the hippocampus CA1 in rats.The MIF、MCP-1 protein was in increased gradually.The MIF、MCP-1 mRNA were higher than the control(P＜0.049),but,there was no significant difference between the sham and control groups(P＞0.681).Conclusions1.MIF,MCP-1 protein and mRNA level in AD and VD patients increased, suggesting that MIF,MCP-1 take part in the pathogenesis of AD and VD,and may involved in the pathogenesis of inflammatory Mechanism.2.The more severe the disease in patients with AD and VD,the higher plasma level ofMIF,MCP-1 and PBMC expression of MIF,MCP-1 mRNA.MIF,MCP-1 may serve as an indicator of the severity of AD and VD.So it was animportant value of the examination of them to dynamically supervise pathologic change at state of AD and VD.3.MIF/MCP-1 protein concentration correlate on the MMSE score,cholesterol, C-reactive protein,SBP,mRNA level of expression,but,there are no obvious correlation in gender,age,history of hypertension,diabetes,history,history of high blood lipids,smoking history,history of alcohol abuse,ESR,Ye Acid,vitamin B12.4.Inflammatory reactions involved in the pathogenesis of vascular dementia. MIF、MCP-1 positive cells were observed around the focus of CA1 in hippocampus at the rat models of vascular dementia.And as the essential inflammatory mediators, MIF,MCP-1 may be involved in the process of VD inflammatory response.5.The level ofMIF,MCP-1 protein and the expression ofMIF,MCP-1mRNA around the CA1 in hippocampus at the rat models of vascular dementia were increased gradually.They may involve in the mechanism of immunity damnification of vascular dementia.Applications their antagonists will provide a new way for the prevention and treatment of vascular dementia in future.