Skin Research Institute of Singapore, Singapore, Singapore.
Institute of Medical Biology, Singapore, Singapore.
Exp Dermatol. 2024 Aug;33(8):e15143. doi: 10.1111/exd.15143.
Immortalized keratinocytes can offer a low-cost experimental platform for human skin research, with increased cell yield compared to primary cultures. However, the usefulness of these surrogate cell models is highly dependent on their ability to retain the phenotypic attributes of the parent cells. Keratins K14 and K5 are the hallmarks of undifferentiated, mitotically active basal keratinocytes. We observed occasional progressive loss of K14 expression in growing keratinocyte cell lines, with persistent retention of K5 and an epithelial phenotype, and investigated possible reasons for this. We show that K14 repression occurs by DNA promoter methylation of KRT14 gene and is compounded by histone deacetylation and by the presence of EGF. In vivo, keratinocytes shut down K14 synthesis as they commit to terminal differentiation and move from the basal to spinous layer, but by laser-capture microdissection of human epidermis we could detect no evidence of increased selective KRT14 methylation in this normal process. Loss of K14 expression suggests that epidermal identity of cultured keratinocytes can be compromised in certain tissue culture situations, possibly due to the immortalization method and persistent EGF supplementation.
永生化角质形成细胞可为人类皮肤研究提供低成本的实验平台，与原代培养相比，其细胞产量增加。然而，这些替代细胞模型的有用性高度依赖于它们保持亲本细胞表型特征的能力。角蛋白 K14 和 K5 是未分化、有丝分裂活性的基底角质形成细胞的标志。我们观察到在不断生长的角质形成细胞系中，K14 表达偶尔会逐渐丧失，而 K5 持续保留并保持上皮表型，我们对此进行了研究。我们表明，K14 抑制是通过 KRT14 基因的 DNA 启动子甲基化发生的，并因组蛋白去乙酰化和 EGF 的存在而加剧。在体内，当角质形成细胞向终末分化并从基底层移到棘层时，它们会关闭 K14 的合成，但通过对人表皮进行激光捕获显微解剖，我们在这个正常过程中没有发现 KRT14 甲基化选择性增加的证据。K14 表达的丧失表明，在某些组织培养情况下，培养的角质形成细胞的表皮特性可能受到损害，这可能归因于永生化方法和持续的 EGF 补充。
