Frion-Herrera Yahima, Zanotto Ilaria, Colognesi Martina, Signor Anna, Cuffaro Doretta, Chilin Adriana, Macchia Marco, Marzaro Giovanni, Digiacomo Maria, Gabbia Daniela, De Martin Sara
Department of Medicine, University of Padua, Clinical Medicine 1 and Thrombotic and Haemorrhagic Disease Unit, and Hemophilia Center, Padova University Hospital, Padua, Italy.
Department of Pharmaceutical and Pharmacological Sciences, University of Padova, Padova, Italy.
Eur J Pharmacol. 2025 Sep 5;1002:177847. doi: 10.1016/j.ejphar.2025.177847. Epub 2025 Jun 12.
Metabolic dysfunction-associated steatohepatitis (MASH) is a chronic liver disease with increasing incidence. This study aimed at assessing the beneficial effects of oleocanthalic acid (OcA), extracted from extra virgin olive oil, in cellular models of MASH. OcA effects on lipid accumulation, oxidative stress, inflammation, and fibrosis were evaluated on 2D HepG2 cells treated with a palmitic and oleic acid mixture (1:2 0.1 mM) to induce steatosis, HepG2 co-cultured with THP-1-derived M1-like macrophages, multicellular spheroids (MCS) composed of HepG2 and TGFβ1-activated LX-2 cells co-cultured with macrophages, and spheroids of 3T3-L1-derived adipocytes. OcA effect was assessed by evaluating lipid droplets after Bodipy stain, reactive oxygen species (ROS), and mRNA expression by qPCR. The expression of the fibrogenic marker α-smooth muscle actin (Acta2) was assessed by ICC. OcA interaction with the pro-oxidant NADPH oxidases NOX1 and NOX4 was investigated via molecular docking simulations. OcA reduced the size and number of lipid droplets and ROS in HepG2 cells. In the MCS model, besides reducing lipid accumulation, OcA promoted a switch in macrophages towards an anti-inflammatory phenotype. OcA decreased Acta2 expression in activated LX-2 cells and prevented the LX-2 cell activation induced by the secretome of 3T3-L1-derived adipocyte spheroids. Molecular docking simulations suggested that OcA could act as a selective NOX1 inhibitor, binding to NOX1 in close contact with the FAD molecule. In conclusion, OcA demonstrated promising anti-steatotic, anti-inflammatory, and anti-fibrotic effects in different in vitro models of MASH, thus representing a potential therapeutic option for preventing lipid accumulation and fibrogenesis in MASH, with a NOX1-dependend mechanism of action.
代谢功能障碍相关脂肪性肝炎（MASH）是一种发病率不断上升的慢性肝病。本研究旨在评估从特级初榨橄榄油中提取的油橄榄酸（OcA）在MASH细胞模型中的有益作用。在用棕榈酸和油酸混合物（1:2, 0.1 mM）处理以诱导脂肪变性的二维HepG2细胞、与THP-1衍生的M1样巨噬细胞共培养的HepG2、由HepG2和TGFβ1激活的LX-2细胞与巨噬细胞共培养组成的多细胞球体（MCS）以及3T3-L1衍生的脂肪细胞球体中，评估了OcA对脂质积累、氧化应激、炎症和纤维化的影响。通过评估Bodipy染色后的脂滴、活性氧（ROS）以及通过qPCR检测mRNA表达来评估OcA的作用。通过免疫细胞化学（ICC）评估纤维化标志物α-平滑肌肌动蛋白（Acta2）的表达。通过分子对接模拟研究OcA与促氧化剂NADPH氧化酶NOX1和NOX4的相互作用。OcA减少了HepG2细胞中脂滴的大小和数量以及ROS。在MCS模型中，除了减少脂质积累外，OcA还促进巨噬细胞向抗炎表型转变。OcA降低了活化的LX-2细胞中Acta2的表达，并阻止了3T3-L1衍生的脂肪细胞球体分泌组诱导的LX-2细胞活化。分子对接模拟表明，OcA可以作为一种选择性NOX1抑制剂，与FAD分子紧密接触的NOX1结合。总之，OcA在不同的MASH体外模型中显示出有前景的抗脂肪变性、抗炎和抗纤维化作用，因此代表了一种预防MASH中脂质积累和纤维化的潜在治疗选择，其作用机制依赖于NOX1。
