Departamento de Bioquímica y Biología Estructural, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México. Ciudad de México, México.
Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional. Unidad Monterrey, Apodaca, Nuevo León, México.
Arch Med Res. 2021 Oct;52(7):692-702. doi: 10.1016/j.arcmed.2021.04.005. Epub 2021 May 6.
An elevated level of plasma uric acid has been widely recognized as a risk factor for non-alcoholic fatty liver disease (NAFLD), where oxidative stress and inflammation play an important role in the pathophysiology of the disease. Although the complete molecular mechanisms involved remain unknown, while under physiological conditions uric acid presents antioxidant properties, hyperuricemia has been linked to oxidative stress, chronic low-grade inflammation, and insulin resistance, basic signs of NAFLD.
Employing in vivo experimentation, we aim to investigate whether a high-fat diet rich in cholesterol (HFD), modifies the metabolism of purines in close relationship to molecular events associated with the development of NAFLD. In vitro experiments employing HepG2 cells are also carried out to study the phenomenon of oxidative stress.
Adult male rabbits were fed for 8 weeks an HFD to induce NAFLD. At the beginning of the experiment and every 15 d until the completion of the study, plasma levels of lipids, lipoproteins, and uric acid were measured. Liver tissue was isolated, and histology performed followed by the biochemical determination of hypoxanthine, protein expression of xanthine oxidoreductase (XOR) by western blot analysis, and xanthine oxidase (XO) activity using an enzymatic kinetic assay. Furthermore, we employed in vitro experimentation studying HepG2 cells to measure the effect of hypoxanthine and HO upon the production of radical oxygen species (ROS), XO activity, and cell viability.
Hepatic tissue from rabbits fed the HFD diet showed signs of NAFLD associated with an increased ROS concentration and an altered purine metabolism characterized by the increase in hypoxanthine, together with an apparent equilibrium displacement of XOR towards the xanthine dehydrogenase (XDH) isoform of the enzyme. This protein shift visualized by a western blot analysis, associated with an increase in plasma uric acid and hepatocyte hypoxanthine could be understood as a compensatory series of events secondary to the establishment of oxidative stress associated with the chronic establishment of fatty liver disease.
血浆尿酸水平升高已被广泛认为是非酒精性脂肪性肝病（NAFLD）的一个危险因素，其中氧化应激和炎症在疾病的病理生理学中起着重要作用。虽然涉及的完整分子机制尚不清楚，但在生理条件下，尿酸具有抗氧化特性，而高尿酸血症与氧化应激、慢性低度炎症和胰岛素抵抗有关，这些都是 NAFLD 的基本特征。
通过体内实验，我们旨在研究富含胆固醇的高脂肪饮食（HFD）是否会改变嘌呤代谢，与与 NAFLD 发展相关的分子事件密切相关。还进行了体外实验，以研究 HepG2 细胞中的氧化应激现象。
成年雄性兔喂食 HFD 8 周以诱导 NAFLD。在实验开始时和实验完成前每 15 天，测量血脂、脂蛋白和尿酸的血浆水平。分离肝组织，进行组织学检查，然后通过生化测定黄嘌呤、黄嘌呤氧化还原酶（XOR）的蛋白质表达进行Western blot 分析以及黄嘌呤氧化酶（XO）活性的酶动力学测定。此外，我们还进行了体外实验，研究 HepG2 细胞中黄嘌呤和 HO 对活性氧（ROS）产生、XO 活性和细胞活力的影响。
喂食 HFD 饮食的兔肝组织显示出与 NAFLD 相关的特征，伴有 ROS 浓度升高和嘌呤代谢改变，表现为黄嘌呤增加，同时酶的 XOR 明显向黄嘌呤脱氢酶（XDH）同工酶发生平衡位移。Western blot 分析显示的这种蛋白质移位，以及血浆尿酸和肝细胞黄嘌呤的增加，可被理解为与慢性脂肪肝相关的氧化应激建立相关的一系列补偿性事件。
