原发性骨质疏松症患者的载脂蛋白E基因多态性

背景:近年来,国外学者发现载脂蛋白E基因与骨质疏松症及其并发髋部骨折存在相关性。国内外学者对载脂蛋白E基因采用聚合酶链反应-限制性片段长度多态性进行基因多态性与骨密度、骨质疏松之间关系进行了初步研究。目的:观察载脂蛋白E基因多态性,分析其与原发性骨质疏松症、骨质疏松性骨折的相关性。单位:南昌市第一医院骨科。设计:临床对照观察。对象:原发性骨质疏松患者60例,男31例,女29例,其中髋部骨折组30例均为新近骨折并有X射线片为诊断依据,单纯性骨质疏松组30例;健康对照组30例,男16例,女14例。方法:选择2002-01/12南昌市第一医院骨科门诊、住院原发性骨质疏松患者60例,骨折组30例,单纯性骨质疏松组30例,健康对照组30例。全部受检对象均禁食12h以上,于次日早晨采静脉血,从分离的白细胞中提取DNA;进行基因位点DNA扩增聚合酶链反应-单链构象多态性分析。将已明确的载脂蛋白E2/2,E3/3,E4/4纯合子个体的模板DNA(中山大学达安基因诊断中心提供)进行PCR-SSCP分析,示快泳带中E3居前,E2居中,E4置后。将样本模板DNA与之同时电泳,比较快泳带的迁移率即可确定样本模板的载脂蛋白E基因型。主要观察指标:载脂蛋白E基因型。结果:参与者全部进入结果分析。①60例原发性骨质疏松患者(骨质疏松性髋部骨折组30例)和30例健康对照组中PCR扩增产物SSCP分析检测到6种基因型:载脂蛋白E2/2,E3/2,3/3,E4/2,4/3,4/4。各组均出现载脂蛋白E基因2型、3型、4型等位基因。②载脂蛋白E基因4型等位基因频率3组比较差异有非常显著性意义(χ2=17.520,P<0.01),携带载脂蛋白E基因4型频率骨质疏松组及骨质疏松性骨折组明显高于正常人群(χ2=4.904,16.681,P<0.01);骨质疏松性骨折组高于骨质疏松组(χ2=4.658,P<0.01)。结论:载脂蛋白E4与原发性骨质疏松症、骨质疏松性骨折有密切相关性;载脂蛋白E4是原发性骨质疏松症,尤其是骨质疏松性骨折一个有用的标志物。

Apolipoprotein E gene polymorphisms in patients with primary osteoporosis

BACKGROUND: In recent years, foreign scholars have found that apolipoprotein E gene was associated with osteoporosis and its complicated hip fracture. National scholars adopted polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) to primarily study the correlation of apolipoprotein E gene polymorphism with bone mineral density (BMD) and osteoporosis.OBJECTIVE: To observe the polymorphism of apolipoprotein E gene, and analyze its correlation with primary osteoporosis and osteoporotic fracture.SETTING: Department of Orthopaedics, Nanchang First Hospital.DESIGN: A clinical controlled observation.PARTICIPANTS: Sixty patients with primary osteoporosis (31 males and 29 females), were enrolled, including 30 cases of hip fracture, who were newly fractured and diagnosed according to radiogram, and 30 cases of simple osteoporosis; Thirty healthy subjects (16 males and 14 females)were also involved.METHODS: Sixty outpatients and inpatients with primary osteoporosis were selected from the Department of Orthopaedics, Nanchang First Hospital from January to December 2002, and they were divided into hip fracture group (n=30) and simple osteoporosis (n=30); Another 30 healthy physical examinees were taken as the healthy control group. All the subjects were fasted for more than 12 hours, and the samples of venous blood were collected on the next morning. DNA was extracted from the separated leucocytes. Gene site DNA amplification was analyzed with polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). The individual DNA templates (provided by Daan Gene Diagnosis Center of Sun Yat-sen University) of identified apolipoprotein E2/2, E3/3 and E4/4 homozygotes were analyzed with PCR-SSCP, and the band showed that E3 was in the front, E2 in the middle, followed by E4. They were in electrophoresis with DNA templates of the samples at the same time, and the apolipoprotein E genotype of the template could be identified by the migration rate of the band.MAIN OUTCOME MEASURES: Apolipoprotein E genotypes were observed.RESULTS: All the enrolled subjects were involved in the analysis of results. ① Six genotypes were detected with PCR-SSCP in the 60 patients with primary osteoporosis and 30 healthy controls, including apolipoprotein E2/2, E3/2, 3/3, E4/2, 4/3, 4/4. ② There were very significant differences in the allele frequency of apolipoprotein E4 among the three groups (χ2=17.520, P ＜ 0.01). The allele frequencies of apolipoprotein E4 in the simple osteoporosis group and osteoporotic fracture group were obviously higher that that in the normal control group (χ2=4.904, 16.681, P ＜ 0.01),it was also markedly higher in the osteoporotic fracture group than in the simple osteoporosis group (χ2=4.658, P ＜ 0.01).CONCLUSION: Apolipoprotein E4 is closely associated with primary osteoporosis and osteoporotic fracture. Apolipoprotein E is a useful marker for primary osteoporosis, especially for osteoporotic fracture.