Epidermolysis Bullosa Simplex keratinocytes show disturbed mitochondrial positioning and activity.
Epidermolysis Bullosa Simplex keratinocytes show disturbed mitochondrial positioning and activity.
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单纯性大疱性表皮松解症角质形成细胞显示线粒体定位和活性受到干扰
DOI：
10.1016/j.jid.2019.10.023
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Magin TM
中科院分区：
文献类型：
--
作者：
Vetter A;Jahn K;Bouameur JE;Kiritsi D;Magin TM
角蛋白细胞骨架蛋白保护表皮免受各种应激，并参与基本细胞过程的调节，如生长、分化和代谢。角蛋白5和角蛋白14（K14）突变导致的后果强调了角蛋白功能的日益复杂性，这些突变通过促进角蛋白细胞骨架塌陷成细胞质蛋白聚集体而导致皮肤起泡疾病单纯性大疱性表皮病（EBS）（Kiritsi和Nyström，2018）。最初被认为是由于受损的角蛋白细胞骨架引起的角质形成细胞脆性病症，EBS和相应小鼠模型的后续分析揭示EBS表型要复杂得多，因为角蛋白参与炎症、瘙痒、粘附和细胞迁移，从而影响伤口愈合、皮肤稳态和能量代谢的多个方面（Coulombe和Lee，2012，Kumar等人，2016年）。除了最近描述的线粒体在表皮分化、伤口愈合和衰老中的作用之外（Baris等人，2011年，Feichtinger等人，2014年，Hamanaka等人，2013），在EBS和几种角蛋白缺陷型小鼠品系中描述了线粒体形状改变、内含物形成和线粒体大小变化（阿尔瓦拉多和库伦贝，2014，陈等人，1994年，施瓦茨和Leube，2016年，Uttam等人，1996年）的报告。我们先前在缺乏所有角蛋白的角质形成细胞中发现了改变的线粒体脂质组成和活性（Kumar等人，在正常人角质形成细胞（NHK）中，线粒体主要分布在细胞核周围，这是一个富含成束角蛋白的区域（阿尔瓦拉多和库伦贝，2014）。为了研究EBS中的线粒体组成，我们使用了来自三名诊断为严重全身性EBS的患者的HPV E6 E7永生化角质形成细胞，基于临床表现、免疫荧光结果和突变分析，揭示了常见的KRT 14突变p.R125C。与NHK相反，EBS角质形成细胞中的线粒体除了核周积累之外还分散在整个细胞质中（图lg-1）。在这两种细胞类型中，共聚焦显微镜未能提供角蛋白和线粒体密切相关的证据，而是表明线粒体散布在角蛋白网络中（图la-l）。由于核周线粒体的密度很高，无法对其大小和数量进行更详细的分析，因此我们根据线粒体的核周分布或扩大的细胞质分布对线粒体的细胞分布进行了分类。基于对每个实验100-200个细胞的分析，74%的NHK细胞显示出核周线粒体的优势，而在26%的NHK中，除了核周模式之外，线粒体分散到细胞质中。在相同的细胞培养条件下，84%的EBS角质形成细胞显示线粒体的细胞质分布，只有16%的细胞仅含有核周积累的线粒体（图1 m）。为了进一步证实这一点，我们在三个独立的实验中分析了10-20个代表性细胞的细胞核和单个外周线粒体之间的距离。由于其核周密度高，无法分析核周围的前3 μm。在EBS细胞中，距离细胞核3 - 20 μm的线粒体数量是NHK细胞的2倍（图1 n）。
Keratin cytoskeletal proteins protect the epidermis against various stresses and participate in the regulation of fundamental cellular processes, such as growth, differentiation, and metabolism. The increasing complexity of keratin functions is underscored by consequences resulting from mutations in keratin 5 and keratin 14 (K14), which cause the skin-blistering disease epidermolysis bullosa simplex (EBS) by promoting the collapse of the keratin cytoskeleton into cytoplasmic protein aggregates (Kiritsi and Nyström, 2018). Originally regarded as a keratinocyte fragility condition owing to a compromised keratin cytoskeleton, subsequent analysis of EBS and corresponding mouse models revealed that the EBS phenotype is much more complex, because of the involvement of keratins in inflammation, itch, adhesion, and cell migration, thereby affecting multiple aspects of wound healing, skin homeostasis, and energy metabolism (Coulombe and Lee, 2012, Kumar et al., 2016). In addition to the recently described roles of mitochondria in epidermal differentiation, wound healing, and aging (Baris et al., 2011, Feichtinger et al., 2014, Hamanaka et al., 2013), altered mitochondrial shape, formation of inclusions, and changes in mitochondrial size have been described in EBS and in several strains of keratin-deficient mice (Alvarado and Coulombe, 2014, Chan et al., 1994, Schwarz and Leube, 2016, Uttam et al., 1996). Our previous finding of altered mitochondrial lipid composition and activity in keratinocytes lacking all keratins (Kumar et al., 2015) prompted us to investigate the intracellular distribution and activity of mitochondria in EBS keratinocytes.In normal human keratinocytes (NHKs), mitochondria are distributed predominantly around the nucleus, a domain enriched in bundled keratins (Alvarado and Coulombe, 2014). To investigate the mitochondrial composition in EBS, we used HPV E6E7 immortalized keratinocytes from three patients with the diagnosis of severe generalized EBS, based on the clinical appearance, immunofluorescence findings, and mutation analysis disclosing the common KRT14 mutation p. R125C. In contrast to NHKs, mitochondria in EBS keratinocytes were scattered throughout the cytoplasm in addition to a perinuclear accumulation (Figure 1 g–l). In both cell types, confocal microscopy failed to provide evidence for a close association of keratins and mitochondria but rather suggested that mitochondria were interspersed in the keratin meshwork (Figure 1 a–l). As the very high density of perinuclear mitochondria did not permit a more detailed analysis of their size and numbers, we classified the cellular distribution of mitochondria according to their perinuclear or expanded cytoplasmic distribution. Based on the analysis of 100–200 cells per individual experiment, 74% of NHK cells showed a predominance of perinuclear mitochondria, whereas in 26% of NHKs, mitochondria were spread out to the cytoplasm, in addition to a perinuclear pattern. Under identical cell culture conditions, 84% of EBS keratinocytes showed a cytoplasmic distribution of mitochondria, with only 16% of cells harboring exclusively perinuclear accumulated mitochondria (Figure 1 m). To substantiate this further, we analyzed the distance between the nucleus and single peripheral mitochondria in 10–20 representative cells in three independent experiments. Owing to their high perinuclear density, the first 3 μm around the nucleus could not be analyzed. The number of mitochondria found in a distance between 3 and 20 μm away from the nucleus was 2-fold higher in EBS cells than in NHK cells (Figure 1 n).
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中间丝作为细胞空间的组织者：它们如何影响线粒体结构和功能
影响因子：
6
作者：
N. Schwarz;R. Leube
通讯作者：
R. Leube
角蛋白调节 β 细胞线粒体形态、运动和稳态
DOI：
--
发表时间：
2017
期刊：
The FASEB Journal
影响因子：
--
作者：
J. Silvander;Sofie M Kvarnström;Angeli Kumari;Anup Shrestha;C. Alam;D. Toivola
通讯作者：
D. Toivola
影响因子：
--
作者：
Kiritsi, Dimitra;Nystrom, Alexander
通讯作者：
Nystrom, Alexander
影响因子：
4.3
作者：
Sanchez-Caballero, Laura;Guerrero-Castillo, Sergio;Nijtmans, Leo
通讯作者：
Nijtmans, Leo
影响因子：
4.8
作者：
Mironov, SL;Ivannikov, MV;Johansson, M
通讯作者：
Johansson, M