Association Of Risk Gene Methylation In Peripheral Leukocyte Of Alzheimer's Disease

?Objective?Study the methylation level of BIN1,CX3CR1,CR1,SLC19A2,TNFRSF1 B and PRCC in peripheral leukocytes in AD to explore the role of DNA methylation in AD.?Method?40 patients with mild cognitive impairment?MCI group?and 40 patients with Alzheimer's disease?AD group?were included,and we selected 29 healthy volunteers with matching gender,age and education level as the normal control group?NC group?in this study.Every study members accepted Mini-mental State Examination?MMSE?,Montreal Cognitive Assessment?MoCA?and Activity Of Daily Living Scale were used.?ADL?,Neuropsychiatric Inventory?NPI?,Hamilton Depression Scale?HAMD?and Hamilton Anxiety Scale?HAMA?testing to assessed the cognitive function.Extracted DNA of peripheral blood leukocytes from vein in elbow of the three groups above.High-throughput sequencing was performed on Illumina Hiseq platform to calculate and analyze the methylation level of BIN1?CX3CR1?CR1?SLC19A2?TNFRSF1B and PRCC.We use T test to analyze the difference of DNA methylation level between groups,and logistic regression analysis to conduct multivariate analysis of risk factors.The area under the ROC curve was calculated for CpG sites with statistical difference in methylation level of this locus to assess the ensitivity of screening the patients with MCI and AD.?Rsults?1.The methylation levels of CX3CR1 and BIN1 in peripheral blood leukocytes was relatively high?methylation level > 0.1?,while the levels of CR1,SLC19A2,TNFRSF1 B and PRCC were relatively low?methylation level < 0.1?.2.There was a statistical difference in the overall methylation level of CX3CR1 and PRCC between the cognitive impairment group and the NC group?p < 0.05?.3.The methylation level of specific CpG sites in the AD risk genes was compared among the three groups,and the MCI group was different from NC group at the following sites?p < 0.05?: SLC19A2?30,57,64?,TNFRSF1B?73,199?,PRCC?46,181,194?,using multiple Logistic regression model to analysis the cognitive function of the group cases,results show that the SLC19A2?30,57,64?,TNFRSF1B?73,199?,PRCC?46,194?sites of DNA methylation level contrast are differences between the two groups?p < 0.1?;There was a difference in DNA methylation level between AD group and NC group at the following CpG loci?p < 0.05?: CX3CR1?42,51,64,71,85,125?,CR1?106,108,112?,TNFRSF1B?73,90?,PRCC?125,181?.The results of multivariate analysis showed that CpG loci of CX3CR1?42,51,64,71,85,125?,CR1?106,108,112?,TNFRSF1B?73,90?,PRCC?125,181?was different between the two groups.3.The results of ROC curve analysis suggested that SLC19A2₆4 and TNFRSF1B₁99 CpG sites had strong diagnostic sensibility in the screening of MCI patients?SLC19A2₆4:AUC=0.671,p =0.017?;TNFRSF1B₇3: AUC = 0.648,p = 0.040);In the screening of AD patients,CX3CR1₁25,CX3CR1₄2,CX3CR1₅1,CX3CR1₆4,CX3CR1₇1,CX3CR1₈5,TNFRSF1B₇3,PRCC₁25,PRCC₁81 and other 9 CpG sites showed strong diagnostic sensibility?CX3CR1₁25: AUC=0.723,p =0.02;CX3CR1₄2: AUC=0.725,p =0.002;CX3CR1₅1: AUC=0.720,p =0.003;CX3CR1₆4: AUC=0.768,p < 10-3;CX3CR1₇1: AUC=0.758,p < 10-3;CX3CR1₈5: AUC=0.714,p =0.003;TNFRSF1B₇3: AUC=0.727,p =0.002;PRCC₁25: AUC=0.668,P=0.021?.?The number after each gene indicates the position of the methylation site contained in the gene on the amplicons??Conclusion?1.DNA methylation alteration may regulate the expression of AD risk genes and affect the pathological progress of AD.2.The methylation level of CX3CR1 in peripheral leukocyte is high,and that might do good job in the diagnosis of AD.3.DNA methylation in peripheral blood leukocyte may be a biomarker for the identification and diagnosis of AD,CX3CR1 methylation may be a candidate biomarker for the screening of AD patients.