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Genetic factors determining the severity of hepatitis in patients receiving hepatitis virus infection : Significance of osteopontin promoter SNPs

决定肝炎病毒感染患者肝炎严重程度的遗传因素：骨桥蛋白启动子SNP的意义

基本信息

批准号：
16590628
负责人：
MOCHIDA Satoshi
金额：
$ 2.24万
依托单位：
Saitama Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16590628/
关键词：
Osteopontin SNP Th1 immune reaction fulminant hepatitis Chronic hepatitis C Promoter assay SRY 性差

项目摘要

Osteopontin is an extracellular matrix with RGD sequence, which also acts as a cytokine crucial for the initiation of Th1 immune reaction. Recently, we identified 4 SNPs (nt-155, -433, -616, -1,748) in promoter region of osteopontin gene, and demonstrated that SNP at nt -433 is a marker to reflect hepatitis activity in patients with chronic hepatitis C. The frequency of T/T in this SNP was 12% in the patients with sustained normal ALT levels, and this ratio was smaller than in those with elevated ALT levels (42%). Thus, it seems that hepatitis activity may be regulated in the patients through osteopontin expression in the liver determined by SNP at nt -443. Thus, in the present project, the function of osteopontin promoter SNPs was evaluated in vitro. Three haplotypes existed in Japanese showing homozygotes both in SNPs at nt -155 and nt -443 ; Type 1 : deletion, T, type 2 : deletion, C and type 3 : G, T, respectively. DNA samples of each haplotype were amplified by PCR between nt 0 and -658 in promoter region of the gene. The resultant fragments were subjected to dual-luciferase reporter assay following transfection into HepG2cells. Promoter activity was greater in cDNA of type 1 than in that of type 2, while the activity was markedly reduced in cDNA of type 3 compared to those of types 1 and 2. These data suggest that promoter activities were greater in cDNA with allele C in SNP at nt -443 than in those with allele T, and the activities were attenuated in cDNA with allele G compared to that with deletion mutation in SNPs at nt -155. Considering that SRY, a translation factor coded on Y chromosome, can bind to the oligonucleotide around nt -155 of osteopomtin promoter, SNP at nt -155 as well as SNP at nt -443 may be responsible for determination of hepatitis severity in patients following hepatitis virus infection and the significance of SNP at nt -155 might be different depending on sex of the patients.

骨桥蛋白是一种含有RGD序列的细胞外基质，也是一种启动Th 1免疫反应的关键细胞因子。最近，我们在骨桥蛋白基因启动子区发现了4个SNP（nt-155，-433，-616，-1，748），证明nt-433 SNP是反映慢性丙型肝炎患者肝炎活动性的标志物。在ALT水平持续正常的患者中，该SNP中T/T的频率为12%，并且该比例小于ALT水平升高的患者（42%）。因此，通过nt-443处SNP确定的肝脏中骨桥蛋白表达，似乎可以调节患者的肝炎活动。因此，在本项目中，骨桥蛋白启动子SNP的功能在体外进行了评估。在日本人群中存在3种单倍型，在nt-155和nt-443处均为纯合子，分别为：1型：缺失，T，2型：缺失，C，3型：G，T。每个单倍型的DNA样品通过PCR在基因启动子区的nt 0和-658之间扩增。转染HepG 2细胞后进行双荧光素酶报告基因检测。启动子活性在1型的cDNA中大于在2型的cDNA中，而活性在3型的cDNA中与1型和2型的cDNA相比显著降低。这些数据表明，在nt-443处SNP中具有等位基因C的cDNA的启动子活性高于具有等位基因T的cDNA，而在nt-155处SNP中具有等位基因G的cDNA的启动子活性比具有缺失突变的cDNA的启动子活性减弱。考虑到Y染色体上编码的翻译因子SRY可与骨桥蛋白启动子nt-155附近的寡核苷酸结合，nt-155和nt-443的SNP可能是决定肝炎病毒感染后患者肝炎严重程度的重要因素，且nt-155 SNP的意义可能因患者性别而异。