Kimberly A. Arnold, Mary C. Moran, Huishan Shi, Ivonne M. J. J. van Vlijmen-Willems, Diana Rodijk-Olthuis, Jos P. H. Smits, Matthew G. Brewer
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引用次数: 0
摘要
跨膜蛋白 claudin-1 对于被称为紧密连接(TJ)的表皮屏障结构的形成至关重要,已被证明在多种疾病中起着重要作用。这些疾病包括新生儿鱼鳞病和硬化性胆管炎综合征、特应性皮炎和各种病毒感染。为了建立一个模型来研究claudin-1在不同疾病中的作用,我们使用CRISPR/Cas9生成了缺乏claudin-1(CLDN1 KO)的人类永生化角质形成细胞(KC)系。然后,我们测定了 Claudin-1 的表达缺失是否会影响表皮屏障的形成/功能以及 KC 的分化/分层。claudin-1的缺失导致单层培养和器官型培养的屏障功能明显降低。CLDN1 KO细胞表现出屏障蛋白filaggrin和分化标记细胞角蛋白-10基因转录本的减少。在 CLDN1 KO 的器官型培养物中还观察到明显的形态差异,包括分层减弱和颗粒层形成减少。我们还在 CLDN1 KO 器官培养物的基底层检测到增殖性 KC 的增加。这些结果进一步支持了claudin-1在表皮屏障中的作用,并表明该蛋白在表皮适当分层中的额外作用。
CLDN1 knock out keratinocytes as a model to investigate multiple skin disorders
The transmembrane protein claudin-1 is critical for formation of the epidermal barrier structure called tight junctions (TJ) and has been shown to be important in multiple disease states. These include neonatal ichthyosis and sclerosing cholangitis syndrome, atopic dermatitis and various viral infections. To develop a model to investigate the role of claudin-1 in different disease settings, we used CRISPR/Cas9 to generate human immortalized keratinocyte (KC) lines lacking claudin-1 (CLDN1 KO). We then determined whether loss of claudin-1 expression affects epidermal barrier formation/function and KC differentiation/stratification. The absence of claudin-1 resulted in significantly reduced barrier function in both monolayer and organotypic cultures. CLDN1 KO cells demonstrated decreases in gene transcripts encoding the barrier protein filaggrin and the differentiation marker cytokeratin-10. Marked morphological differences were also observed in CLDN1 KO organotypic cultures including diminished stratification and reduced formation of the stratum granulosum. We also detected increased proliferative KC in the basale layer of CLDN1 KO organotypic cultures. These results further support the role of claudin-1 in epidermal barrier and suggest an additional role of this protein in appropriate stratification of the epidermis.
期刊介绍:
Experimental Dermatology provides a vehicle for the rapid publication of innovative and definitive reports, letters to the editor and review articles covering all aspects of experimental dermatology. Preference is given to papers of immediate importance to other investigators, either by virtue of their new methodology, experimental data or new ideas. The essential criteria for publication are clarity, experimental soundness and novelty. Letters to the editor related to published reports may also be accepted, provided that they are short and scientifically relevant to the reports mentioned, in order to provide a continuing forum for discussion. Review articles represent a state-of-the-art overview and are invited by the editors.