沉默PCSK9通过抑制REG3A-EXTL3-PI3K-AKT通路促进UVB诱导角质形成细胞凋亡并抑制NF-κB通路的机制研究

UVB can induce apoptosis and is usually used in treatment of skin diseases characterized by hyperplasia of keratinocytes. However, it often leads to adverse reactions such as erythema because of triggering inflammatory pathways such as NF-κB. We have found that suppressing proprotein convertase subtilisin/kexin 9 (PCSK9) can significantly decrease proliferation and induce apoptosis of human keratinocytes, also suppress the NF-κB pathway. Interestingly, combination of UVB and si-hPCSK9 treatments has significant interactive effect on apoptosis. In addition, REG3A, was significantly decreased by suppressing PCSK9, while it could inhibit apoptosis and trigger NF-κB pathway of keratinocytes by the EXTL3-PI3K-AKT pathway. Therefore, we speculate that silencing PCSK9 can promote apoptosis and suppress NF-κB pathway in UVB-induced keratinocytes via inhibiting the REG3A-EXTL3-PI3K-AKT pathway. We try to reveal the mechanism of which PCSK9 regulates UVB-induced apoptosis and NF-κB pathway via REG3A-EXTL3-PI3K-AKT pathway, at the molecular, cellular and animal levels, and we expect to provide a new target and original theoretical basis for enhancing the efficacy and reducing inflammatory adverse reactions of UVB in the treatment of skin diseases by combing with PCSK9 interference.

UVB可诱导细胞凋亡，常用于治疗以角质形成细胞过度增殖为特征的皮肤病，因可触发NF-κB等炎性反应通路，常导致红斑等不良反应。我们发现角质形成细胞沉默枯草溶菌素转化酶9型（PCSK9）后，可促进凋亡并抑制NF-κB通路；且沉默PCSK9与UVB照射联用可协同诱导凋亡。进一步发现沉默PCSK9可降低REG3A的表达，而REG3A可通过激活EXTL3-PI3K-AKT通路而抑制细胞凋亡并诱导NF-κB通路，且UVB照射亦可激活PI3K-AKT通路。因此，我们假设：沉默PCSK9可以通过抑制REG3A-EXTL3-PI3K-AKT通路而促进UVB诱导的角质形成细胞凋亡并抑制其诱导的NF-κB通路。我们将从分子、细胞、动物水平，系统揭示沉默PCSK9促进UVB诱导凋亡并抑制NF-κB通路的机制，为通过联合干扰PCSK9的手段来增强UVB治疗效果并降低不良反应提供全新的作用靶点和原创性的理论基础。

UVB可诱导细胞凋亡，常用于治疗以角质形成细胞过度增殖为特征的皮肤病，因可触发NF-κB等炎性反应通路，常导致红斑等不良反应。我们发现角质形成细胞沉默枯草溶菌素转化酶9型（PCSK9）后，可促进凋亡并抑制NF-κB通路；且沉默PCSK9与UVB照射联用可协同诱导凋亡。进一步发现沉默PCSK9可降低REG3A的表达，而REG3A可通过激活EXTL3-PI3K-AKT通路而抑制细胞凋亡并诱导NF-κB通路，且UVB照射亦可激活PI3K-AKT通路。因此，我们假设：沉默PCSK9可以通过抑制REG3A-EXTL3-PI3K-AKT通路而促进UVB诱导的角质形成细胞凋亡并抑制其诱导的NF-κB通路。经验证，沉默HaCat细胞的PCSK9，对EXTL3及AKT的表达水平无显著影响。因为EXTL3与AKT是REG3A-EXTL3-PI3K-AKT通路的关键成分，因此我们推断，单纯的沉默PCSK9对REG3A-EXTL3-PI3K-AKT通路无影响，及时停止该通路的进一步验证实验。接下来，进一步探索PCSK9对UVB光损伤的影响以及可能的机制。UVB照射后引起小鼠皮肤明显损伤，其皮损中PCSK9表达显著升高。局部通过RNA干扰或药物抑制剂抑制PCSK9的表达，可以减轻UVB诱导的皮肤损伤，皮损中PCNA表达降低。而系统使用PCSK9抑制剂同样可减少UVB诱导的皮肤损伤和PCNA表达。进一步研究机制，PCSK9局部敲除和系统抑制均可抑制UVB诱导的cGAS-STING通路的激活；抑制PCSK9可降低 UVB 诱导下HaCat细胞中dsDNA 的释放。PCSK9抑制降低了UVB引发的DNA损伤，导致巨噬细胞中角化细胞释放的dsDNA和STING通路激活减少，从而减轻炎症和皮肤损伤。本研究首次发现PCSK9通过调节cGAS-STING通路调控UVB光损伤的机制。以上发现，对于探索UVB光损伤的机制及进一步明确光损伤的治疗靶点，具有重要的意义。