Liver Unit, Queen Elizabeth Hospital, University Hospitals Birmingham NHS Foundation Trust, Birmingham, United Kingdom.
National Institute for Health Research (NIHR) Birmingham Biomedical Research Centre, Institute of Immunology and Immunotherapy, College of Medical and Dental Sciences, University of Birmingham, Birmingham, United Kingdom.
PLoS One. 2018 Jul 25;13(7):e0201419. doi: 10.1371/journal.pone.0201419. eCollection 2018.
Pharmacological defatting of rat hepatocytes and hepatoma cell lines suggests that the same method could be used to ameliorate macrovesicular steatosis in moderate to severely fatty livers. However there is no data assessing the effects of those drugs on primary human liver cells. We aimed to determine the effectiveness of a pharmacological cocktail in reducing the in vitro lipid content of primary human hepatocytes (PHH). In addition we sought to determine the cytotoxicity of the cocktail towards non-parenchymal liver cells.
Steatosis was induced in PHH by supplementation with a combination of saturated and unsaturated free fatty acids. This was followed by addition of a defatting drug cocktail for up to 48 hours. The same experimental method was used with human intra-hepatic endothelial cells (HIEC) and human cholangiocytes. MTT assay was used to assess cell viability, triglyceride quantification and oil red O staining were used to determine intracellular lipids content whilst ketone bodies were measured in the supernatants following experimentation.
Incubation of fat loaded PHH with the drugs over 48 hours reduced the intracellular lipid area by 54%, from 12.85% to 5.99% (p = 0.002) (percentage of total oil red O area), and intracellular triglyceride by 35%, from 28.24 to 18.30 nmol/million of cells (p<0.001). Total supernatant ketone bodies increased 1.4-fold over 48 hours in the defatted PHH compared with vehicle controls (p = 0.002). Moreover incubation with the drugs for 48 hours increased the viability of PHH by 11%, cholangiocytes by 25% whilst having no cytotoxic effects on HIEC.
These data demonstrate that pharmacological intervention can significantly decrease intracellular lipid content of PHH, increase fatty acids β-oxidation whilst being non-toxic to PHH, HIEC or cholangiocytes.
药理学方法去除大鼠肝细胞和肝癌细胞系的脂肪表明，该方法也可用于改善中等至重度脂肪肝的大泡性脂肪变性。但是，目前尚无数据评估这些药物对原代人肝细胞的影响。我们旨在确定药理鸡尾酒在降低原代人肝细胞（PHH）体外脂质含量方面的有效性。此外，我们还试图确定鸡尾酒对非实质细胞的细胞毒性。
通过添加饱和和不饱和游离脂肪酸的组合诱导 PHH 脂肪变性。然后在 48 小时内添加去脂药物鸡尾酒。使用相同的实验方法研究人肝内内皮细胞（HIEC）和人胆管细胞。MTT 测定法用于评估细胞活力，甘油三酯定量和油红 O 染色用于确定细胞内脂质含量，实验后在上清液中测量酮体。
将药物孵育于脂肪负荷 PHH 中 48 小时可使细胞内脂质面积减少 54％，从 12.85％降至 5.99％（p = 0.002）（油红 O 总面积的百分比），细胞内甘油三酯减少 35％，从 28.24 降至 18.30 nmol/百万个细胞（p<0.001）。与载体对照相比，去脂 PHH 在 48 小时内的上清液中总酮体增加了 1.4 倍（p = 0.002）。此外，药物孵育 48 小时可使 PHH 的活力增加 11％，胆管细胞增加 25％，而对 HIEC 无细胞毒性。
这些数据表明，药理学干预可以显著降低 PHH 的细胞内脂质含量，增加脂肪酸β氧化，同时对 PHH、HIEC 或胆管细胞无毒。
