目的 探讨夏枯草来源外泌体装载奥利司他（Prunella vulgaris-derived exosomes loading orlistat，PVENs-Orl）对肝癌细胞脂代谢及恶性生物学行为的影响及其潜在机制。方法 采用超速离心法提取PVENs，经透射电镜（transmission electron microscopy，TEM）及纳米颗粒追踪分析（nanoparticle tracking analysis，NTA）表征后，构建PVENs-Orl载药系统。通过CCK-8、克隆、划痕及Transwell实验评估PVENs-Orl对HepG2细胞增殖、迁移与侵袭的抑制作用。检测游离脂肪酸（free fatty acid，FFA）摄取、脂肪酸合成酶（fatty acid synthase，FASN）活性、三酰甘油（triglyceride，TG）含量、脂质过氧化指标及线粒体膜电位与形态。采用qRT-PCR检测脂代谢与铁死亡相关基因[酰基辅酶A合成酶长链家族成员4（acyl-CoA synthetase long-chain family member 4，ACSL4）、谷胱甘肽过氧化物酶4（glutathione peroxidase 4，GPX4）、溶质载体家族7成员11（solute carrier family 7 member 11，SLC7A11）、B细胞淋巴瘤-2相互作用蛋白3（Bcl-2 interacting protein 3，BNIP3L）、溶血磷脂酰胆碱酰基转移酶3（lysophosphatidylcholine acyltransferase 3，LPCAT3）、转铁蛋白受体（transferrin receptor，TFRC）]的表达。结果 成功提取并表征了PVENs，构建的PVENs-Orl能被HepG2细胞有效内化。PVENs及PVENs-Orl呈剂量相关性抑制HepG2细胞的增殖、迁移与侵袭（P＜0.001）。在脂代谢表型上，PVENs-Orl能抑制FASN活性（P＜0.01），同时引起细胞内FFA与TG蓄积（P＜0.01）。PVENs-Orl显著提升细胞内活性氧（reactive oxygen species，ROS）和丙二醛（malondialdehyde，MDA）水平（P＜0.01、0.001），增加乳酸脱氢酶（lactate dehydrogenase，LDH）释放（P＜0.001）。PVENs-Orl显著诱导线粒体膜电位下降（P＜0.01），并引发线粒体功能障碍（P＜0.001）。分子机制上，PVENs-Orl上调脂质过氧化及铁死亡促进相关基因表达（P＜0.001），下调抗氧化及铁死亡抑制相关基因表达（P＜0.001）。结论 PVENs可有效递送奥利司他，通过抑制FASN活性破坏脂质稳态，诱导线粒体功能障碍和脂质过氧化，并协同调控铁死亡关键基因表达，从而抑制肝癌细胞生长，为基于天然外泌体的脂代谢靶向治疗提供了实验依据。

Objective To investigate the effect and underlying mechanism of Prunella vulgaris-derived exosomes loaded with orlistat (PVENs-Orl) on lipid metabolism and malignant biological behaviors of hepatoma cells. Methods PVENs were extracted by ultracentrifugation. After characterization by transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA), a PVENs-Orl drug delivery system was constructed. The inhibitory effects of PVENs-Orl on the proliferation, migration and invasion of HepG2 cells were evaluated through CCK-8, cloning, scratch and Transwell assays. The free fatty acids (FFA) uptake, fatty acid synthase (FASN) activity, triglyceride (TG) content, lipid peroxidation indicators and mitochondrial membrane potential and morphology were detected. The expressions of genes related to lipid metabolism and ferroptosis [acyl-CoA synthetase long-chain family member 4 (ACSL4), glutathione peroxidase 4 (GPX4), solute carrier family 7 member 11 (SLC7A11), B-cell lymphoma-2 interacting protein 3 (BNIP3L), lysophosphatidylcholine acyltransferase 3 (LPCAT3), transferrin receptor (TFRC)] was detected by qRT-PCR. Results The PVENs were successfully extracted and characterized, and the constructed PVENs-Orl could be effectively internalized by HepG2 cells. PVENs and PVENs-Orl exhibited dose-dependent inhibition of the proliferation, migration and invasion of HepG2 cells (P < 0.001). In terms of lipid metabolism phenotype, PVENs-Orl could inhibit FASN activity (P < 0.01), and simultaneously cause intracellular accumulation of FFA and TG (P < 0.01). PVENs-Orl significantly increased the levels of intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) (P < 0.01, 0.001), and increased the release of lactate dehydrogenase (LDH) (P < 0.001). PVENs-Orl significantly induced a decrease in mitochondrial membrane potential (P < 0.01) and triggered mitochondrial dysfunction (P < 0.001). At the molecular mechanism level, PVENs-Orl upregulated the expressions of genes related to lipid peroxidation and ferroptosis promotion (P < 0.001), and downregulated the expressions of genes related to antioxidant and ferroptosis inhibition (P < 0.001). Conclusion PVENs could effectively deliver orlistat, disrupt lipid homeostasis by inhibiting FASN activity, induce mitochondrial dysfunction and lipid peroxidation, and synergistically regulate the expressions of key genes involved in ferroptosis, thereby inhibiting the growth of liver cancer cells. This provides experimental evidence for targeted lipid metabolism therapy based on natural exosomes.

R285.5

国家自然科学基金面上项目（82575174）；国家自然科学基金面上项目（82074425）；湖南省重点研发计划项目（2023SK2057）；湖南省自然科学基金资助项目（2023JJ40403，2025JJ30038）；湖南省中医药管理局研究项目（B2023089）；湖南中医药大学研究生创新项目（2025CX188，2025CX199）