Department of Pharmacy, Xijing Hospital, Air Force Medical University, Xi'an 710032, Shaanxi, PR China; Laboratory of Pharmacotherapy of Life-Style Related Diseases, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai 980-8578, Miyagi, Japan.
Department of Pharmacy, Xijing Hospital, Air Force Medical University, Xi'an 710032, Shaanxi, PR China.
Cell Signal. 2019 May;57:58-64. doi: 10.1016/j.cellsig.2019.01.005. Epub 2019 Jan 19.
Thymic stromal lymphopoietin (TSLP), a master switch of allergic inflammation, plays an important role in the pathogenesis of allergic diseases. Although many compounds upregulate TSLP expression in vivo or in vitro, most of them are pollutants or toxicants. In the previous study, for the first time, we found that a steroid alkaloid derivative 02F04, which has a unique skeletal structure compared with other TSLP-inducing chemicals, significantly induced TSLP production in mouse keratinocytes. However, it is not investigated thoroughly that how 02F04 produces TSLP and why. In this study, we did a detailed investigation on the inducible effect and underlying molecular mechanism of 02F04 on TSLP production. We found that the peak time of TSLP mRNA level induced by 02F04 at 48 h led to a slow and continuous TSLP production in PAM212 cells. Besides, 02F04-induced TSLP production was significantly suppressed by inhibitors of Rho-associated protein kinase (ROCK), guanine nucleotide-binding protein subunit alpha q/11 (Gq/11) and extracellular signal-regulated kinase 1/2 (ERK1/2) at not only protein but also mRNA levels, and by siRNA-mediated knockdown of Gq or G11. This suggested that ROCK, Gq/11 and ERK1/2 signaling pathways were involved in 02F04-induced TSLP production. Increase in the level of p-ERK1/2 induced by 02F04 was suppressed by both inhibitors of ROCK and Gq/11, indicating that ROCK and Gq/11 molecules were located at the upstream of ERK1/2 to regulate 02F04-induced TSLP production. Gq/11 was located at the upstream of ROCK because the specific Gq/11 inhibitor of YM-254890 significantly reduced 02F04-induced actin stress fiber formation. Taken together, 02F04 upregulates a slow and continuous TSLP production through a novel Gq/11-ROCK-ERK1/2 signaling pathway. The thorough understanding the effect and mechanism of 02F04 on TSLP production is expected to supply it as a novel TSLP-regulating compound and a potential new tool for investigating the role of TSLP in allergic disorders.
胸腺基质淋巴细胞生成素 (TSLP) 作为过敏炎症的主开关，在过敏疾病的发病机制中发挥着重要作用。虽然许多化合物能在体内或体外上调 TSLP 的表达，但其中大多数是污染物或有毒物质。在之前的研究中，我们首次发现，与其他诱导 TSLP 的化学物质相比，具有独特骨架结构的甾体生物碱衍生物 02F04 可显著诱导小鼠角质形成细胞产生 TSLP。然而，02F04 如何产生 TSLP 以及为什么会产生 TSLP 尚未得到深入研究。在这项研究中，我们详细研究了 02F04 对 TSLP 产生的诱导作用及其潜在的分子机制。我们发现，02F04 在 48 小时诱导 TSLP mRNA 水平达到峰值，导致 PAM212 细胞中 TSLP 的产生呈缓慢持续状态。此外，02F04 诱导的 TSLP 产生在蛋白质和 mRNA 水平上均被 Rho 相关蛋白激酶 (ROCK)、鸟嘌呤核苷酸结合蛋白亚单位 alpha q/11 (Gq/11) 和细胞外信号调节激酶 1/2 (ERK1/2) 的抑制剂显著抑制，通过 Gq 或 G11 的 siRNA 介导的敲低也可显著抑制。这表明 ROCK、Gq/11 和 ERK1/2 信号通路参与了 02F04 诱导的 TSLP 产生。02F04 诱导的 p-ERK1/2 水平增加被 ROCK 和 Gq/11 的抑制剂抑制，表明 ROCK 和 Gq/11 分子位于 ERK1/2 的上游，以调节 02F04 诱导的 TSLP 产生。Gq/11 位于 ROCK 的上游，因为特异性 Gq/11 抑制剂 YM-254890 可显著减少 02F04 诱导的肌动蛋白应激纤维形成。综上所述，02F04 通过一种新型的 Gq/11-ROCK-ERK1/2 信号通路上调缓慢而持续的 TSLP 产生。深入了解 02F04 对 TSLP 产生的作用和机制有望将其作为一种新型的 TSLP 调节化合物，并为研究 TSLP 在过敏疾病中的作用提供一种新的潜在工具。
