Department of Dermatology, Faculty of Medicine, Kagawa University, Kagawa, Japan.
J Dermatol. 2010 Nov;37(11):956-64. doi: 10.1111/j.1346-8138.2010.00932.x. Epub 2010 Aug 16.
The epidermal cornified cell envelope is a complex protein-lipid composite that replaces the plasma membrane of corneocytes and is crucial for epidermal barrier function. Loricrin is a major constituent of the epidermal cornified cell envelope, contributing approximately 70% by mass. In order to explore novel function of wild-type (WT) loricrin other than the major component of the epidermal cornified cell envelope, we transiently expressed construct encoding human WT and mutant loricrin (730insG) in HaCaT keratinocytes. HaCaT cells transfected with WT or mutant loricrin were at differentiation level. WT loricrin in the transfected cells was seen diffusely in the cytoplasm and nuclei. Positive transferase deoxytidyl uridine end labeling staining was observed in the nuclei of WT loricrin-transfected HaCaT keratinocytes. Data from the DNA fragmentation assay showed that only WT loricrin induced DNA ladders compared with that of mutant loricrin. WT loricrin-transfected HaCaT keratinocytes were susceptible to programmed cell death (PCD). Activation of caspase-14 was also seen. In contrast, PCD or activation of caspase-14 did not occur in mutant loricrin-transfected HaCaT cells. These results suggest that the expression of WT loricrin facilitates induction of PCD in HaCaT keratinocytes.
表皮的角质细胞包膜是一种复杂的蛋白脂质复合结构，它替代了角质细胞的质膜，对于表皮屏障功能至关重要。兜甲蛋白是表皮角质细胞包膜的主要成分之一，其质量占比约为 70%。为了探索野生型（WT）兜甲蛋白除了作为表皮角质细胞包膜的主要成分之外的新功能，我们在 HaCaT 角质细胞中瞬时表达了编码人 WT 和突变型兜甲蛋白（730insG）的构建体。转染 WT 或突变型兜甲蛋白的 HaCaT 细胞处于分化水平。在转染细胞中，WT 兜甲蛋白弥散分布于细胞质和细胞核中。在 WT 兜甲蛋白转染的 HaCaT 角质细胞的细胞核中观察到阳性转移酶脱氧尿苷末端标记染色。DNA 片段化分析结果显示，只有 WT 兜甲蛋白诱导了 DNA 梯状条带，而突变型兜甲蛋白则没有。WT 兜甲蛋白转染的 HaCaT 角质细胞易发生程序性细胞死亡（PCD）。也观察到了半胱氨酸蛋白酶-14 的激活。相比之下，突变型兜甲蛋白转染的 HaCaT 细胞未发生 PCD 或半胱氨酸蛋白酶-14 的激活。这些结果表明，WT 兜甲蛋白的表达促进了 HaCaT 角质细胞中 PCD 的诱导。
