NFAT5 Controls the Integrity of Skin II. NFAT5 Controls the Integrity of Skin II

The skin protects the human body against dehydration and harmful challenges. Keratinocytes (KCs) are the most frequent epidermal cells, and it is anticipated that KC-mediated transport of Na+ ions creates a physiological barrier of high osmolality against the external environment. We studied in KCs the role of NFAT5, a transcription factor whose activity is controlled by osmotic stress. Cultured KCs from adult mice secrete more than 300 proteins, and upon NFAT5 ablation, the secretion of several matrix proteinases, including metalloproteinase-3 (Mmp3) and kallikrein-related peptidase 7 (Klk7), was markedly enhanced. An increase in Mmp3 and Klk7 RNA levels was also detected in transcriptomes of Nfat5-/- KCs, along with increases of numerous components of ‘Epidermal Differentiation Complex’ (EDC), as proline-rich Sprr and S100 proteins. NFAT5 and Mmp3 are co-expressed in basal KCs from fetal and adult skin but not in skin of newborn mice. This is correlated with a strong increase in Mmp3 and Klk7 expression in KCs of newborn mice and suggests, along with the fragile epidermis of adult Nfat5-/- mice, a suppressive effect of NFAT5 on the expression of matrix proteases in skin. Our data suggest that NFAT5 controls the expression of matrix proteases in skin and contributes to the many fold changes during embryonal skin development and skin integrity in adults. Overall design: Expression profiling by high throughput sequencing of RNA from murine keratinocytes cultivated in vitro for one or for three weeks. Keratinocytes were prepared from the tail of adult mice (4 independent experiments)

皮肤是人体抵御脱水与有害侵袭的天然屏障。角质形成细胞（Keratinocytes，KCs）是表皮中占比最高的细胞类型，据推测，KC介导的钠离子转运可构建高渗透压的生理屏障，以对抗外界环境。我们针对角质形成细胞中受渗透压应激调控的转录因子NFAT5的功能展开了研究。
从成年小鼠体内分离培养的角质形成细胞可分泌超过300种蛋白质；当NFAT5基因敲除后，包括基质金属蛋白酶3（matrix metalloproteinase-3，Mmp3）与激肽释放酶相关肽酶7（kallikrein-related peptidase 7，Klk7）在内的多种基质蛋白酶的分泌量显著上调。在Nfat5基因敲除的角质形成细胞的转录组中，除Mmp3与Klk7的RNA水平升高外，表皮分化复合体（Epidermal Differentiation Complex，EDC）的众多组分——包括富含脯氨酸的Sprr蛋白与S100蛋白家族成员——的表达量也出现上调。
NFAT5与Mmp3在胎鼠与成年小鼠的基底角质形成细胞中共表达，但在新生小鼠皮肤中未检测到共表达。这与新生小鼠角质形成细胞中Mmp3与Klk7的表达量显著升高的现象相符，结合成年Nfat5敲除小鼠表皮脆弱的表型，提示NFAT5可抑制皮肤中基质蛋白酶的表达。
本研究数据表明，NFAT5可调控皮肤内基质蛋白酶的表达，并参与胚胎皮肤发育过程及成年个体皮肤完整性维持中的诸多表达变化。
实验整体设计：通过高通量测序对体外培养1周或3周的小鼠角质形成细胞的RNA进行表达谱分析。角质形成细胞均取自成年小鼠尾部组织，共开展4次独立实验。