School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing, 211198, China.
Wuxi Huishan District People's Hospital, Wuxi, 214187, China; Affiliated Hushan Hospital of Xingling College, Nantong University, 226019, China.
Phytomedicine. 2024 Jul;129:155629. doi: 10.1016/j.phymed.2024.155629. Epub 2024 Apr 17.
Acute pancreatitis (AP) is an inflammatory disorder of the exocrine pancreas, especially hyperlipidemia acute pancreatitis (HLAP) is the third leading cause of acute pancreatitis which is more severe with a greater incidence of persistent multiorgan failure. HLAP inflicts injury upon the organelles within the acinar cell, particularly mitochondria, the endolysosomal-autophagy system, and is accompanied by senescence-associated secretory phenotype (SASP). RAD, only two consists of Rhizoma Alismatis and Atractylodes macrocephala Rhizoma, which is best known for its ability to anti-inflammatory and lipid-lowering. Nevertheless, the mechanism by which RAD alleviates HLAP remains obscure, necessitating further investigation.
The study aimed to assess the effects of the RAD on HLAP and to elucidate the underlying mechanism in vivo and in vitro, offering a potential medicine for clinical treatment for HLAP.
C57BL/6 mice with hyperlipidemia acute pancreatitis were induced by HFD and CER, then administrated with RAD. AR42J were stimulated by cerulein or conditioned medium and then cultured with RAD. Serums were analyzed to evaluate potential pancreas and liver damage. Furthermore, tissue samples were obtained for histological, and protein investigations by H&E, Oil red staining, and Western blot. In addition, western blot and immunofluorescent staining were utilized to estimate the effect of RAD on mitochondrial function, autophagy flux, and SASP.
In vivo, RAD considerably alleviated systemic inflammation while attenuating TC, TG, AMY, LPS, inflammatory cytokines, histopathology changes, oxidative damage, mitochondrial fission, and autophagy markers in HLAP mice. Impaired autophagy flux and mitochondrial dysfunction resulted in a significant enhancement of NLRP3 and IL-1β in the pancreas. RAD could reverse these changes. In vitro, RAD significantly restored mitochondrial membrane potential and oxidative phosphorylation levels. RAD decreased Beclin-1 and LC3-II expression and increased LAMP-1 and Parkin-Pink expression, which showed that RAD significantly ameliorated HLAP-induced damage to the mitochondria function by suppressing mitochondrial oxidative damage and enhancing autophagy flux and mitophagy to remove the damaged mitochondria. In addition, we found that RAD could up-regulate the expression of BAX, and Bad and down-regulate the expression of p16, and p21, indicating that RAD could promote damaged cell apoptosis and alleviate SASP.
This study revealed that RAD ameliorates mitochondrial function to alleviate SASP through enhancing autophagy flux, mitophagy, and apoptosis which provided a molecular basis for the advancement and development of protection strategies against HLAP.
急性胰腺炎（AP）是一种胰腺外分泌的炎症性疾病，尤其是高脂血症性急性胰腺炎（HLAP）是急性胰腺炎的第三大病因，其病情更严重，持续性多器官衰竭的发生率更高。HLAP 会损伤胰腺泡细胞内的细胞器，特别是线粒体、内溶酶体自噬系统，并伴有衰老相关分泌表型（SASP）。RAD 仅由泽泻和白术组成，以其抗炎和降血脂能力而闻名。然而，RAD 缓解 HLAP 的机制尚不清楚，需要进一步研究。
本研究旨在评估 RAD 对 HLAP 的影响，并在体内和体外阐明其潜在机制，为 HLAP 的临床治疗提供潜在药物。
通过 HFD 和 CER 诱导 C57BL/6 小鼠发生高脂血症性急性胰腺炎，然后用 RAD 治疗。用 Cerulein 或条件培养基刺激 AR42J 细胞，然后用 RAD 培养。分析血清以评估潜在的胰腺和肝脏损伤。此外，通过 H&E、油红染色和 Western blot 分析组织样本进行组织学和蛋白质研究。此外，还利用 Western blot 和免疫荧光染色来评估 RAD 对线粒体功能、自噬通量和 SASP 的影响。
体内研究结果表明，RAD 可显著减轻 HLAP 小鼠的全身炎症，同时降低 TC、TG、AMY、LPS、炎症细胞因子、组织病理学变化、氧化损伤、线粒体裂变和自噬标志物。自噬通量和线粒体功能受损导致胰腺中 NLRP3 和 IL-1β显著增加，RAD 可逆转这些变化。体外研究结果表明，RAD 可显著恢复线粒体膜电位和氧化磷酸化水平。RAD 降低 Beclin-1 和 LC3-II 的表达，增加 LAMP-1 和 Parkin-Pink 的表达，表明 RAD 通过抑制线粒体氧化损伤和增强自噬通量和线粒体自噬来清除受损的线粒体，显著改善 HLAP 诱导的线粒体功能损伤。此外，我们发现 RAD 可以上调 BAX 和 Bad 的表达，下调 p16 和 p21 的表达，表明 RAD 可以促进受损细胞凋亡，减轻 SASP。
本研究表明，RAD 通过增强自噬通量、线粒体自噬和细胞凋亡来改善线粒体功能，从而减轻 SASP，为 HLAP 的防治策略提供了分子基础。
