Lack of association between HSV-1 DNA in the brain, Alzheimer's disease and apolipoprotein E4.
Lack of association between HSV-1 DNA in the brain, Alzheimer's disease and apolipoprotein E4.
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大脑中的 HSV-1 DNA、阿尔茨海默病和载脂蛋白 E4 之间缺乏关联。
DOI：
10.1080/135502801300069773
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发表时间：
2001
影响因子：
3.2
通讯作者：
Fischer,SH
中科院分区：
文献类型：
--
作者：
Marques,AR;Straus,SE;Fahle,G;Weir,S;Csako,G;Fischer,SH
An association between herpes simplex type 1 (HSV-1), ApoE4 and AD has been reported (Itzhaki et al., 1997). HSV-1 DNA was detected by the polymerase chain reaction (PCR) in the brains of 28/44 controls and 34/46 AD patients. The odds ratio for the presence of ApoE4 allele was 16.8 in HSV-1 positive AD versus 1.67 for HSV-1 negative AD. These Ūndings stimulated our own investigation of the association of HSV-1 and AD. AD patients and controls brains were provided by Dr Juan Troncoso from the Johns Hopkins University Alzheimer’s Disease Research Center, supported by NIH grant AG 05146. The pathological diagnosis of AD followed the recommendations of the Consortium to Establish a Registry for Alzheimer’s Disease (Mirra et al., 1991). The mean age (and ranges) of AD and controls were 79 (56 ą 93) and 79 (59ą93), respectively. All patients and controls were Caucasian. Autopsies were performed a maximum of 19 h post-mortem, with a mean post mortem delay of 9.5 h for AD and 9.2 h for controls. Brain samples were stored at Ð 808C until DNA extraction was performed. DNA was extracted from an~ 50 mg fragment of brain specimen using the QIAamp Tissue Kit (QIAGEN Inc., Valencia, CA, USA). The HSV primer sequences which amplify a 288 bp product from the glycoprotein B genes of both HSV-1 and HSV-2, were 5Ē-(biotin)-TGC TTG AAG CGG TCG GCG CGC-3Ē and 5Ē-GGA CGA CCA CGA GAC CGA CAT GG-3Ē. To verify the elimination of Taq polymerase inhibitors from the processed specimens, an internal control (IC), ampliŪed by these same primers, was included in each ampliŪcation reaction tube. The IC was constructed so that its ampliŪed product is approximately 50 bp longer than the HSV amplicon. The PCR was performed in the presence of 10 mM Tris-HCl (pH 8.3), 50 mM KCl, 1.5 mM MgCl2, 200 lM deoxynucleoside triphosphates, 2.5 U of Taq polymerase, 0.5 lM of each HSV primer, approximately 250 copies of IC, 25 lg/ml isopsoralen-10, 2.5 mg/ml BSA, 10% glycerol, with 5 ll of the extracted sample added, in a total volume of 50 ll. The thermal cycler program included an initial denaturation period of 5 min at 948C, followed by a two-step cycling program,(948C for 30 s and 728C for 2 min for 38 cycles) and concluded with an extension period of 5 min at 728C.Detection of the HSV and IC ampliŪcation products was performed using the DELFIA timeresolved Ŋuorescence hybridization assay (Fahle et al., 1999). The HSV-1 speciŪc probe was 5ĒTGA AAC CGG CCA ACG CCG CGA-3Ē. The internal IC-probe binding sequence was 5Ē-GCG ATG CTG TCG GAA ACG-3Ē. The HSV PCR was performed in triplicates on the extracted DNA from the frontal, temporal and occipital lobes of 10 AD and 14 controls, while the HSV PCR was performed in sextuplicates from extracted samples from Ūve AD and one control. For positive and negative controls, brain samples from HSV-1 latently infected (n= 3) and noninfected (n= 30) guinea pigs were run in triplicates. All samples were run in a blinded fashion. ApoE genotyping was performed by a modiŪed restriction fragment length polymorphism method (Wu et al., 2000).
