Department of Internal Medicine I, University Medical Center Ulm, Ulm, Germany.
PLoS One. 2012;7(3):e32669. doi: 10.1371/journal.pone.0032669. Epub 2012 Mar 7.
Keratins 8 and 18 (K8/K18) are intermediate filament proteins that protect the liver from various forms of injury. Exonic K8/K18 variants associate with adverse outcome in acute liver failure and with liver fibrosis progression in patients with chronic hepatitis C infection or primary biliary cirrhosis. Given the association of K8/K18 variants with end-stage liver disease and progression in several chronic liver disorders, we studied the importance of keratin variants in patients with hemochromatosis.
The entire K8/K18 exonic regions were analyzed in 162 hemochromatosis patients carrying homozygous C282Y HFE (hemochromatosis gene) mutations. 234 liver-healthy subjects were used as controls. Exonic regions were PCR-amplified and analyzed using denaturing high-performance liquid chromatography and DNA sequencing. Previously-generated transgenic mice overexpressing K8 G62C were studied for their susceptibility to iron overload. Susceptibility to iron toxicity of primary hepatocytes that express K8 wild-type and G62C was also assessed.
We identified amino-acid-altering keratin heterozygous variants in 10 of 162 hemochromatosis patients (6.2%) and non-coding heterozygous variants in 6 additional patients (3.7%). Two novel K8 variants (Q169E/R275W) were found. K8 R341H was the most common amino-acid altering variant (4 patients), and exclusively associated with an intronic KRT8 IVS7+10delC deletion. Intronic, but not amino-acid-altering variants associated with the development of liver fibrosis. In mice, or ex vivo, the K8 G62C variant did not affect iron-accumulation in response to iron-rich diet or the extent of iron-induced hepatocellular injury.
In patients with hemochromatosis, intronic but not exonic K8/K18 variants associate with liver fibrosis development.
角蛋白 8 和 18（K8/K18）是中间丝蛋白，可保护肝脏免受各种形式的损伤。外显子 K8/K18 变体与急性肝衰竭的不良预后以及慢性丙型肝炎感染或原发性胆汁性肝硬化患者的肝纤维化进展相关。鉴于 K8/K18 变体与几种慢性肝病的终末期肝病和进展相关，我们研究了角蛋白变体在血色病患者中的重要性。
在 162 名携带纯合 C282Y HFE（血色病基因）突变的血色病患者中分析了整个 K8/K18 外显子区域。234 名肝脏健康的受试者作为对照。使用变性高效液相色谱法和 DNA 测序分析外显子区域的 PCR 扩增产物。还研究了先前生成的过表达 K8 G62C 的转基因小鼠对铁过载的易感性。还评估了表达 K8 野生型和 G62C 的原代肝细胞对铁毒性的敏感性。
我们在 162 名血色病患者中的 10 名（6.2%）和另外 6 名患者（3.7%）中发现了改变氨基酸的角蛋白杂合变体和非编码杂合变体。发现了两种新的 K8 变体（Q169E/R275W）。K8 R341H 是最常见的氨基酸改变变体（4 名患者），且仅与内含子 KRT8 IVS7+10delC 缺失相关。内含子而非氨基酸改变的变体与肝纤维化的发展相关。在小鼠或离体中，K8 G62C 变体不会影响富含铁的饮食或铁诱导的肝细胞损伤程度下的铁蓄积。
在血色病患者中，内含子而非外显子 K8/K18 变体与肝纤维化的发展相关。
