Key Laboratory of Big Data Mining and Precision Drug Design of Guangdong Medical University, Key Laboratory of Computer-Aided Drug Design of Dongguan City, Key Laboratory for Research and Development of Natural Drugs of Guangdong Province, School of Pharmacy, Guangdong Medical University, Dongguan, 523808, China.
State Key Laboratory of Bioactive Molecules and Druggability Assessment, Jinan University, Guangzhou, 511443, China.
Theranostics. 2024 Oct 7;14(17):6516-6542. doi: 10.7150/thno.98400. eCollection 2024.
: Abdominal aortic aneurysm (AAA) is an inflammatory, fatal aortic disease that currently lacks any effective drugs. Cryptotanshinone (CTS) is a prominent and inexpensive bioactive substance derived from , a well-known medicinal herb for treating cardiovascular diseases through its potent anti-inflammatory properties. Nevertheless, the therapeutic effect of CTS on AAA formation remains unknown. : To investigate the therapeutic effect of CTS in AAA, variety of experimental approaches were employed, majorly including AAA mouse model establishment, real-time polymerase chain reaction (PCR), RNA sequencing, western blot, co-immunoprecipitation, scanning/transmission electron microscopy (SEM/TEM), enzyme-linked immunosorbent assay (ELISA), seahorse analysis, immunohistochemistry, and confocal imaging. : In this study, we demonstrated that CTS suppressed the formation of AAA in apolipoprotein E knock-out (ApoE) mice infused with Ang II. A combination of network pharmacology and whole transcriptome sequencing analysis indicated that activation of the Keap1-Nrf2 pathway and regulation of programmed cell death in vascular smooth muscle cells (VSMCs) are closely linked to the anti-AAA effect of CTS. Mechanistically, CTS promoted the transcription of Nrf2 target genes, particularly Hmox-1, which prevented the activation of NLRP3 and GSDMD-initiated pyroptosis in VSMCs, thereby mitigating VSMC inflammation and maintaining the VSMC contractile phenotype. Subsequently, by utilizing molecular docking, together with the cellular thermal shift assay (CETSA) and isothermal titration calorimetry (ITC), a particular binding site was established between CTS and Keap1 at Arg415. To confirm the binding site, site-directed mutagenesis was performed, which intriguingly showed that the Arg415 mutation eliminated the binding between CTS and the Keap1-Nrf2 protein and abrogated the antioxidant and anti-pyroptosis effects of CTS. Furthermore, VSMC-specific Nrf2 knockdown in mice dramatically reversed the protective action of CTS in AAA and the inhibitory effect of CTS on VSMC pyroptosis. : Naturally derived CTS exhibits promising efficacy as a treatment drug for AAA through its targeting of the Keap1-Nrf2-GSDMD-pyroptosis axis in VSMCs.
腹主动脉瘤（AAA）是一种炎症性、致命的主动脉疾病，目前尚无任何有效的药物治疗方法。隐丹参酮（CTS）是一种从丹参中提取的重要且廉价的生物活性物质，丹参是一种治疗心血管疾病的著名草药，具有很强的抗炎作用。然而，CTS 对 AAA 形成的治疗效果尚不清楚。
为了研究 CTS 在 AAA 中的治疗作用，采用了多种实验方法，主要包括 AAA 小鼠模型的建立、实时聚合酶链反应（PCR）、RNA 测序、western blot、共免疫沉淀、扫描/透射电子显微镜（SEM/TEM）、酶联免疫吸附测定（ELISA）、海瑟分析、免疫组织化学和共聚焦成像。
在这项研究中，我们证明了 CTS 抑制了 Ang II 输注的载脂蛋白 E 敲除（ApoE）小鼠 AAA 的形成。网络药理学和全转录组测序分析的结合表明，Keap1-Nrf2 通路的激活和血管平滑肌细胞（VSMCs）中程序性细胞死亡的调节与 CTS 的抗 AAA 作用密切相关。从机制上讲，CTS 促进了 Nrf2 靶基因的转录，特别是 Hmox-1，从而防止了 NLRP3 的激活和 GSDMD 引发的 VSMCs 细胞焦亡，从而减轻了 VSMC 炎症并维持了 VSMC 的收缩表型。随后，通过利用分子对接，以及细胞热转移测定（CETSA）和等温滴定量热法（ITC），在 CTS 和 Keap1 之间建立了一个特定的结合位点在 Arg415。为了确认结合位点，进行了定点突变，结果有趣地表明，Arg415 突变消除了 CTS 与 Keap1-Nrf2 蛋白之间的结合，并消除了 CTS 的抗氧化和抗细胞焦亡作用。此外，在小鼠中特异性敲低 VSMC 中的 Nrf2 可显著逆转 CTS 在 AAA 中的保护作用，以及 CTS 对 VSMC 细胞焦亡的抑制作用。
天然来源的 CTS 通过靶向 VSMCs 中的 Keap1-Nrf2-GSDMD 细胞焦亡轴，显示出作为 AAA 治疗药物的有希望的疗效。
