Abstract

Background

Psoriasis is a chronic, inflammatory skin disease. It was reported that lncRNA Non-coding RNA-activated by DNA damage (NORAD) has potential regulatory effects on skin diseases. Our previous studies found that lncRNA NORAD was highly expressed and its potential target miR-26a was down-regulated in psoriasis model mice. Here, we aimed to investigate the role of NORAD in the development of psoriasis.

Methods

IL-22/LPS (interleukin-22/lipopolysaccharide)-stimulated HaCaT (human immortalized keratinocytes) cell model and imiquimod-induced mouse model were established. Keratin 6 (K6), Keratin 16 (K16), Keratin 17 (K17), and Cell division cycle 6 (CDC6) levels were detected by western blot. Cell activity was detected by CCK-8, MTT, and EdU assays. Quantitative real-time PCR was performed to examine the levels of NORAD, miR-26a, CDC6, K6, K16, and K17. Haematoxylin-eosin staining was applied to observe the degree of skin thickening and hyperplasia. Fluorescence in situ hybridization detects the location of NORAD. RNA immunoprecipitation, RNA pull-down, and Luciferase test were performed to detect the interaction between NORAD and miR-26a.

Results

In IL-22/LPS-stimulated HaCaT cells, NORAD, CDC6, and keratinocyte proliferation-related proteins (K6, K16, and K17) were up-regulated and miR-26a was down-regulated. Cell survival and proliferation were also increased. However, the results were reversed after interference with NORAD. Also, in vitro experiments revealed that NORAD negatively regulated miR-26a. In IL-22/LPS-stimulated HaCaT cells and skin of imiquimod-induced mice, we found that lower NORAD resulted in an increase of miR-26a and a decrease of CDC6, further decreased levels of keratinocyte proliferation-related proteins (K6, K16, and K17).

中文翻译：

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LncRNA NORAD 通过与 miR-26a 结合调节角质形成细胞增殖参与银屑病

 抽象的

 背景

牛皮癣是一种慢性炎症性皮肤病。据报道，lncRNA——由DNA损伤激活的非编码RNA（NORAD）对皮肤病具有潜在的调节作用。我们前期的研究发现lncRNA NORAD在银屑病模型小鼠中高表达，其潜在靶标miR-26a下调。在这里，我们的目的是研究 NORAD 在银屑病发生中的作用。

 方法

建立IL-22/LPS（白细胞介素22/脂多糖）刺激的HaCaT（人永生化角质形成细胞）细胞模型和咪喹莫特诱导的小鼠模型。通过蛋白质印迹检测角蛋白 6 (K6)、角蛋白 16 (K16)、角蛋白 17 (K17) 和细胞分裂周期 6 (CDC6) 水平。通过 CCK-8、MTT 和 EdU 检测检测细胞活性。进行实时定量 PCR 检测 NORAD、miR-26a、CDC6、K6、K16 和 K17 的水平。苏木精-伊红染色观察皮肤增厚、增生程度。荧光原位杂交可检测 NORAD 的位置。进行RNA免疫沉淀、RNA Pull-down和荧光素酶测试来检测NORAD和miR-26a之间的相互作用。

 结果

在 IL-22/LPS 刺激的 HaCaT 细胞中，NORAD、CDC6 和角质形成细胞增殖相关蛋白（K6、K16 和 K17）上调，而 miR-26a 下调。细胞存活和增殖也增加。然而，在 NORAD 干扰后，结果发生了逆转。此外，体外实验表明 NORAD 负调控 miR-26a。在IL-22/LPS刺激的HaCaT细胞和咪喹莫特诱导的小鼠皮肤中，我们发现较低的NORAD导致miR-26a增加和CDC6减少，进一步降低角质形成细胞增殖相关蛋白（K6、K16）的水平。和 K17）。