Hellerbrand C, Bosserhoff A K, Seegers S, Lingner G, Wrede C, Lock G, Schölmerich J, Büttner R
Dept. of Internal Medicine I, University of Regensburg, Germany.
Scand J Gastroenterol. 2001 Nov;36(11):1211-6. doi: 10.1080/00365520152584860.
The diagnosis of hereditary hemochromatosis (HH) before the onset of iron-overload has been difficult in the past. However, a convincing candidate gene for HH: HFE has been described recently. The aims of this study were: 1) To determine the prevalence of the hemochromatosis associated mutations C282Y and H63D of the HFE gene in patients from Southern Germany with hemochromatosis phenotype; and 2) to test two new, time- and cost-saving methods: automated SSCP-based capillary electrophoresis (SSCP-CE) and a PCR-ELISA technique for the analysis of HFE mutations.
HFE genotype was studied in 36 unrelated HH patients and 126 controls from Southern Germany. In addition, family screening was performed in 76 relatives. The C282Y and H63D mutations were detected using SSCP-CE and restriction length polymorphism (RFLP). The C282Y mutation was additionally analysed by a PCR-ELISA.
Twenty-six (72%) HH patients were homozygous for mutation C282Y, and three compound heterozygous for C282Y and H63D. One patient was homozygous for H63D. By performing family screening, six additional patients with the +/+ C282Y mutation were identified. The results of the SSCP-CE and the PCR-ELISA analysis agreed completely with data obtained by RFLP.
SSCP-CE and PCR-ELISA analysis proved to be reliable methods for HFE genotyping and therefore represent cost- and time-effective alternative methods to the widely used restriction analysis allowing large populations to be screened for HH associated with HFE mutations. Surprisingly, only 72% of our HH patients had the C282Y +/+ genotype. This indicates that hemochromatosis in Southern Germany is genetically more heterogeneous than in other regions. A challenge for the future will be to define the genetic or environmental factors responsible for iron-overload in HH patients who do not show typical alterations of the HFE gene.
过去，在遗传性血色素沉着症（HH）出现铁过载之前进行诊断一直很困难。然而，最近发现了一个令人信服的HH候选基因：HFE。本研究的目的是：1）确定德国南部具有血色素沉着症表型的患者中HFE基因的血色素沉着症相关突变C282Y和H63D的患病率；2）测试两种新的、节省时间和成本的方法：基于自动SSCP的毛细管电泳（SSCP-CE）和用于分析HFE突变的PCR-ELISA技术。
对36名来自德国南部的无亲缘关系的HH患者和126名对照进行HFE基因分型研究。此外，对76名亲属进行了家系筛查。使用SSCP-CE和限制性片段长度多态性（RFLP）检测C282Y和H63D突变。C282Y突变还通过PCR-ELISA进行了分析。
26名（72%）HH患者为C282Y突变纯合子，3名患者为C282Y和H63D复合杂合子。1名患者为H63D纯合子。通过家系筛查，又发现了6名携带+/+ C282Y突变的患者。SSCP-CE和PCR-ELISA分析的结果与RFLP获得的数据完全一致。
SSCP-CE和PCR-ELISA分析被证明是HFE基因分型的可靠方法，因此是广泛使用的限制性分析的经济高效替代方法，可用于对大量人群进行与HFE突变相关的HH筛查。令人惊讶的是，我们的HH患者中只有72%具有C282Y +/+基因型。这表明德国南部的血色素沉着症在遗传上比其他地区更加异质。未来的一个挑战将是确定在未表现出HFE基因典型改变的HH患者中导致铁过载的遗传或环境因素。
