Btadini Waed, Abou Hassan Ossama K, Saadeh Dana, Abbas Ossama, Ballout Farah, Kibbi Abdul-Ghani, Dbaibo Ghassan, Darwiche Nadine, Nemer Georges, Kurban Mazen
Department of Dermatology, American University of Beirut, Beirut, Lebanon.
Department of Biochemistry and Molecular Genetics, American University of Beirut, Beirut, Lebanon.
PLoS One. 2015 Feb 6;10(2):e0115530. doi: 10.1371/journal.pone.0115530. eCollection 2015.
Hailey-Hailey disease (HHD) is an inherited blistering dermatosis characterized by recurrent erosions and erythematous plaques that generally manifest in intertriginous areas. Genetically, HHD is an autosomal dominant disease, resulting from heterozygous mutations in ATP2C1, which encodes a Ca2+/Mn2+ATPase. In this study, we aimed at identifying and analyzing mutations in five patients from unrelated families diagnosed with HHD and study the underlying molecular pathogenesis.
To genetically study Lebanese families with HHD, and the underlying molecular pathogenesis of the disease.
We performed DNA sequencing for the coding sequence and exon-intron boundaries of ATP2C1. Heat shock experiments were done on several cell types. This was followed by real-time and western blotting for ATP2C1, caspase 3, and PARP proteins to examine any possible role of apoptosis in HHD. This was followed by TUNEL staining to confirm the western blotting results. We then performed heat shock experiments on neonatal rat primary cardiomyocytes.
Four mutations were detected, three of which were novel and one recurrent mutation in two families. In order for HHD to manifest, it requires both the genetic alteration and the environmental stress, therefore we performed heat shock experiments on fibroblasts (HH and normal) and HaCaT cells, mimicking the environmental factor seen in HHD. It was found that stress stimuli, represented here as temperature stress, leads to an increase in the mRNA and protein levels of ATP2C1 in heat-shocked cells as compared to non-heat shocked ones. However, the increase in ATP2C1 and heat shock protein hsp90 is significantly lower in HH fibroblasts in comparison to normal fibroblasts and HaCaT cells. We did not find a role for apoptosis in the pathogenesis of HHD. A similar approach (heat shock experiments) done on rat cardiomyocytes, led to a significant variation in ATP2C1 transcript and protein levels.
This is the first genetic report of HHD from Lebanon in which we identified three novel mutations in ATP2C1 and shed light on the molecular mechanisms and pathogenesis of HHD by linking stress signals like heat shock to the observed phenotypes. This link was also found in cultured cardiomyocytes suggesting thus a yet uncharacterized cardiac phenotype in HHD patients masked by its in-expressivity in normal health conditions.
黑利-黑利病（HHD）是一种遗传性水疱性皮肤病，其特征为反复出现糜烂和红斑，通常出现在皮肤褶皱部位。从遗传学角度来看，HHD是一种常染色体显性疾病，由编码Ca2+/Mn2+ATP酶的ATP2C1基因杂合突变引起。在本研究中，我们旨在鉴定和分析来自无关家族的5例诊断为HHD的患者的突变情况，并研究其潜在的分子发病机制。
对黎巴嫩患有HHD的家族进行遗传学研究，并研究该疾病的潜在分子发病机制。
我们对ATP2C1的编码序列和外显子-内含子边界进行了DNA测序。对几种细胞类型进行了热休克实验。随后，对ATP2C1、半胱天冬酶3和PARP蛋白进行实时定量和蛋白质印迹分析，以研究细胞凋亡在HHD中的可能作用。接着进行TUNEL染色以证实蛋白质印迹分析结果。然后我们对新生大鼠原代心肌细胞进行了热休克实验。
检测到4个突变，其中3个是新突变，1个是两个家族中的复发性突变。为了使HHD表现出来，既需要基因改变也需要环境应激，因此我们对成纤维细胞（HH患者和正常人）和HaCaT细胞进行了热休克实验，模拟HHD中所见的环境因素。结果发现，以温度应激为代表的应激刺激导致热休克细胞中ATP2C1的mRNA和蛋白质水平相较于未热休克细胞有所增加。然而，与正常成纤维细胞和HaCaT细胞相比，HH患者成纤维细胞中ATP2C1和热休克蛋白hsp90的增加显著较低。我们未发现细胞凋亡在HHD发病机制中的作用。在大鼠心肌细胞上进行的类似方法（热休克实验）导致ATP2C1转录本和蛋白质水平出现显著差异。
这是黎巴嫩关于HHD的首份遗传学报告，我们在其中鉴定出ATP2C1的3个新突变，并通过将热休克等应激信号与观察到的表型联系起来，阐明了HHD的分子机制和发病机制
