Castro Elena, Oviedo-Rodríguez Vladimir, Angel-Chávez Luis I
Centro Universitario de Investigaciones Biomédicas, Universidad de Colima, Colima, México.
BMC Cardiovasc Disord. 2008 Feb 29;8:5. doi: 10.1186/1471-2261-8-5.
Recessive mutations in WRN gene eliminate WRN protein function (helicase) and cause Werner syndrome. One of the most important clinical features of Werner syndrome patients are the premature onset and accelerated atherosclerosis process. Studies carried out on polymorphic WRN locus have shown that the alleles 1367R and 1074L confer protection for cardiovascular disease. Given that the levels of plasminogen activator inhibitor type 1 (PAI-1) were found to be significantly increased in Werner syndrome patients, is quiet possible that PAI-1 expression could be under regulation of WRN helicase. Therefore the purpose of this work was to evaluate the role of WRN polymorphism in modulating the expression of PAI-1.
In order to accomplish our aim, an array of primary cultured fibroblasts from normal adult donors was genotyped for polymorphisms of both the WRN and PAI-1 loci. In addition, steady state levels of WRN and PAI-1 were measured by semi-quantitative RT-PCR assays in such cultures. To search for the potential relationship between the lack of WRN protein and PAI-1 expression, heterozygous cultures of fibroblasts (1367RC/1074LF; WRN genotype) were treated with a molecule of interference RNA against WRN messenger RNA (mRNA).
We found that, carriers of 1367R and 1074L alleles of WRN shown to have low amounts of PAI-1 in plasma (7.56 +/- 5.02), as compared with carriers of 1367C and 1074F alleles (16.09 +/- 6.03). Moreover, fibroblasts from carriers with these alleles had low expression levels of PAI-1 mRNA. The treatment of heterozygous primary fibroblast cultures (1367RC/1074LF; WRN genotype) with iRNA against WRN mRNA caused PAI-1 overexpression. Treatment with normal PAI-1 inducers (TGFbeta, TNFalpha, or insulin) in these cultures and from those with genotypes 1367CC/1074FF and 1367RR/1074FL resulted in a genotype-dependent PAI-1 expression level.
Our results suggest that polymorphisms in the WRN gene might have a significant role regulating PAI-1 levels in healthy individuals and "normal states" as well as acute or chronic stress, obesity, aging, acute inflammation, among others, where characteristic high levels of insulin, TNF alpha and TGFbeta, could favor PAI-1 high levels in carriers with polymorphic variants (C and F alleles), beyond the levels reached by carriers with other alleles (R and L alleles).
WRN基因中的隐性突变会消除WRN蛋白功能（解旋酶）并导致沃纳综合征。沃纳综合征患者最重要的临床特征之一是过早发病和动脉粥样硬化进程加速。对多态性WRN基因座的研究表明，1367R和1074L等位基因对心血管疾病具有保护作用。鉴于在沃纳综合征患者中发现纤溶酶原激活物抑制剂1型（PAI-1）水平显著升高，PAI-1的表达很可能受WRN解旋酶的调控。因此，本研究的目的是评估WRN基因多态性在调节PAI-1表达中的作用。
为实现我们的目标，对来自正常成年供体的一系列原代培养成纤维细胞进行了WRN和PAI-1基因座多态性的基因分型。此外，通过半定量RT-PCR分析测定了这些培养物中WRN和PAI-1的稳态水平。为探究WRN蛋白缺失与PAI-1表达之间的潜在关系，用针对WRN信使核糖核酸（mRNA）的干扰RNA分子处理成纤维细胞杂合培养物（1367RC/1074LF；WRN基因型）。
我们发现，与1367C和1074F等位基因携带者相比，WRN基因1367R和1074L等位基因的携带者血浆中PAI-1含量较低（7.56±5.02）。此外，这些等位基因携带者的成纤维细胞中PAI-1 mRNA表达水平较低。用针对WRN mRNA的干扰RNA处理杂合原代成纤维细胞培养物（1367RC/1074LF；WRN基因型）会导致PAI-1过表达。在这些培养物以及基因型为1367CC/1074FF和1367RR/1074FL的培养物中，用正常的PAI-1诱导剂（转化生长因子β、肿瘤坏死因子α或胰岛素）处理会导致PAI-1表达水平呈现基因型依赖性。
我们的结果表明，WRN基因多态性可能在健康个体以及“正常状态”下，以及在急性或慢性应激、肥胖、衰老、急性炎症等情况下，对调节PAI-1水平发挥重要作用。在这些情况下，胰岛素、肿瘤坏死因子α和转化生长因子β的特征性高水平可能会使多态性变体（C和F等位基因）携带者的PAI-1水平高于其他等位基因（R和L等位基因）携带者所达到的水平。
