帕金森病患者PINK1 LRRK2基因单核苷酸多态性分析

A research on polymorphisms of PINK1,LRRK2 in patients with Parkinson's disease

目的 探讨PINK1基因rs45530340位点及LRRK2基因rs1491942位点单核苷酸多态性（single nucleotide polymorphism,SNP）与淮海地区汉族人群晚发散发性帕金森病（Parkinson＇s disease,PD）的相关性.方法 收集152例晚发散发性PD患者和160例年龄和性别相匹配的健康对照者,入组者均来自淮海地区汉族人群.提取外周血全基因组DNA,应用聚合酶链式反应（polymerase chain reaction,PCR）技术扩增包含多态位点的目的 基因片段,通过琼脂糖凝胶电泳（AGE）检测PCR产物.对PCR产物分别用DNA限制性内切酶NlalV和SmlI进行酶切,用聚丙烯酰胺凝胶电泳（PAGE）、硝酸银染色检测酶切产物,采用聚合酶链式反应-限制性片段长度多态性（restriction fragment length polymorphism,RFLP）技术分析基因型,计算所有研究对象两个SNP位点的基因型频率和等位基因频率.结果 （1）PINK1基因rs45530340位点基因型和等位基因在晚发散发性PD组和正常对照组中的分布无统计学差异（基因型：χ2=1.572,P=0.456;等位基因：χ2=1.318,P=0.251）.（2） LRRK2基因rs1491942 位点基因型和等位基因在晚发散发性PD组与正常对照组中的分布差异有统计学意义（基因型：χ2=6.802,P=0.033;等位基因C：χ2=7.448,P=0.006,OR=1.571,95%CI=1.135~2.176）.结论 （1）PINK1基因rs45530340 多态位点可能不是淮海地区汉族人群晚发散发性PD患者的危险因素.（2）LRRK2基因rs1491942多态位点C等位基因可能是淮海地区汉族人群晚发散发性PD患者的危险因素.

Objective The present study is to investigate the association of rs45530340, rs1491942, polymorphism of PINK1 gene,and LRRK2 gene with late-onset sporadic Parkinson＇s disease （PD） in Han people of Huaihai. Methods Total of 152 patients with late-onset sporadic PD from the Department of Neurology,Afliated Hospital of Xuzhou Medical College,were recruited All the subjects are Han ancestry, and predominantly from Huaihai. A cohort of unrelated 160 healthy controls was recruited from Physical Examination Center of the same hospital. Venous blood was collected from peripheral vein of each par- ticipant. Genome DNAs were extracted from leucocytes using standard protocols and stored at-20℃. To obtain targeting genes fragments of PINK1 gnen polymorphism rs45530340 and LRRK2 gene polymorphism rs1491942, Polymerase Chain Reaction （PCR） was performed. All PCR products were validated with Agarose Gel Electrophoresis. Restricted fragments length poly- morphism （RFLP） was applied to identify genotypes of rs45530340,and rs1491942 using NlalV and SmlI which are the restric- tion enzymes of DNA for the digestion of PCR products. All enzyme digestion products were underwent the Polyacrylamide Gel Electrophoresis （PAGE）. And then, the rapid silver diammine approach was adopted for the staining of nucleic acids in polyac- rylamide gels. Results There was no statistical difference in the distribution of rs45530340 genotype and allele frequencies between late-onset sporadic PD group and control group （allele： X2 = 1. 318, P = 0. 251; genotype： X2 = 1. 572, P = 0. 456 ）. Difference were statistically significant in the distributions of rs 1491942 genotypes and alleles frequencies between late-onset sporadic PD group and control group （for C allele PD group （43.4% ）, control group （32.8%）,X2 = 7. 448, P = 0. 006, OR = 1. 571,95%CI=1.135-2.176; genotype：X2=6. 802,P=0. 033 ）.Conclusion LRRK2 genepolymorphismrs1491942 is a pos-sible risk factor for late-onset sporadic PD in Han population of Huaihai. PINK1 gene polymorphism rs45530340 may not be a risk factor for late-onset sporadic PD in Hun population of Huaihai.