Department of Experimental Medicine, Section of Biotechnology, Medical Histology and Molecular Biology, Università della Campania "Luigi Vanvitelli", Naples, Italy.
R&D - IBSA Farmaceutici Italia, Lodi, Italy.
Lipids Health Dis. 2018 Feb 5;17(1):24. doi: 10.1186/s12944-018-0663-2.
Steatosis is a chronic liver disease that depends on the accumulation of intracellular fatty acids. Currently, no drug treatment has been licensed for steatosis; thus, only nutritional guidelines are indicated to reduce its progression. The aim of this study is to combine different nutraceutical compounds in order to evaluate their synergistic effects on a steatosis in vitro model compared to their separate use. In particular, three different formulations based on silymarin, curcumin, vitamin E, docosahexaenoic acid (DHA), choline, and phosphatidylcholine were assayed.
Human hepatocellular carcinoma cells (HepG2 cell line) were treated with a mixture of fatty acids in order to induce an in vitro model of steatosic cells, and then the amount of intracellular fat was evaluated by Oil Red O staining. The peroxisome proliferator-activated receptors α and γ (PPARα and γ) expression, closely correlated to lipid metabolism, was evaluated. The efficiency of these receptors was evaluated through the study of LPL mRNA expression, a marker involved in the lipolysis mechanism. Superoxide dismutase (SOD-2) and malondialdehydes (MDA) in lipid peroxidation were assayed as specific biomarkers of oxidative stress. In addition, experiments were performed using human liver cells stressed to obtain a steatosis model. In particular, the content of the intracellular fat was assayed using Oil Red O staining, the activation of PPARα and γ was evaluated through western blotting analyses, and the LPL mRNA expression level was analyzed through qRT-PCR.
All formulations proved effective on lipid content reduction of about 35%. The oxidative stress damage was reduced by all the substances separately and even more efficiently by the same in formulation (i.e. Formulation 1 and Formulation 3, which reduced the SOD-2 expression and induced the PPARs activation). Lipid peroxidation, was reduced about 2 fold by foormulation2 and up to 5 fold by the others compared to the cells pretreated with HO.Formulation 1, was more effective on PPARγ expression (2.5 fold increase) respect to the other compounds on FA treated hepathocytes. Beside, LPL was activated also by Formulation 3 and resulted in a 5 to 9 fold-increase respect to FA treated control.
Our results proved that the formulations tested could be considered suitable support to face steatosis disease beside the mandatory dietetic regimen.
脂肪变性是一种慢性肝病，取决于细胞内脂肪酸的积累。目前，尚无药物治疗被批准用于脂肪变性；因此，仅指示营养指南以减少其进展。本研究的目的是结合不同的营养化合物，以评估它们在体外脂肪变性模型中的协同作用，与它们的单独使用相比。特别是，基于水飞蓟素、姜黄素、维生素 E、二十二碳六烯酸（DHA）、胆碱和磷脂酰胆碱的三种不同配方进行了测定。
用人肝癌细胞系（HepG2 细胞系）用脂肪酸混合物处理，以诱导体外脂肪变性细胞模型，然后通过油红 O 染色评估细胞内脂肪含量。评估与脂质代谢密切相关的过氧化物酶体增殖物激活受体α和γ（PPARα和γ）的表达。通过研究参与脂肪分解机制的脂蛋白脂肪酶（LPL）mRNA 表达来评估这些受体的效率。超氧化物歧化酶（SOD-2）和丙二醛（MDA）在脂质过氧化中作为氧化应激的特异性生物标志物进行测定。此外，还使用人肝细胞进行了实验，以获得脂肪变性模型。特别是，通过油红 O 染色测定细胞内脂肪含量，通过 Western blot 分析评估 PPARα和γ的激活，通过 qRT-PCR 分析 LPL mRNA 表达水平。
所有配方均有效降低约 35%的脂质含量。所有物质单独减少氧化应激损伤，甚至配方中更有效地减少（即配方 1 和配方 3，减少 SOD-2 表达并诱导 PPARs 激活）。与用 HO 预处理的细胞相比，配方 2 将脂质过氧化减少约 2 倍，其他配方减少 5 倍。配方 1 对 PPARγ表达的影响（增加 2.5 倍）优于其他化合物对 FA 处理的肝细胞。此外，Formulation 3 还激活了 LPL，并导致与 FA 处理对照相比增加了 5 到 9 倍。
我们的结果证明，测试的配方可被认为是除了强制性饮食治疗外，治疗脂肪变性疾病的合适支持。
