BACKGROUND AND AIMS

The etiology of non-alcoholic fatty liver disease (NAFLD) is poorly understood, with males and certain populations exhibiting markedly increased susceptibility. Using a systems genetics approach, involving multi-omic analysis of ∼100 diverse inbred strains of mice, we recently identified several candidate genes driving NAFLD. We investigated the role of one of these, liver pyruvate kinase (L-PK or Pklr) in NAFLD using patient samples and mouse models.

METHODS

We examined L-PK expression in mice of both sexes and in a cohort of bariatric surgery patients. We used liver-specific loss- and gain-of-function strategies in independent animal models of diet-induced steatosis and fibrosis. Following treatment, we measured several metabolic phenotypes, including obesity, insulin resistance, dyslipidemia, liver steatosis and fibrosis. Liver tissues were used for gene expression and immunoblotting, and liver mitochondria bioenergetics was characterized.

RESULTS

In both mice and humans, L-PK expression is upregulated in males via testosterone and is strongly associated with NAFLD severity. In a steatosis model, L-PK silencing in male mice improved glucose tolerance, insulin sensitivity and lactate/pyruvate tolerance compared to controls. Further, these animals had reduced plasma cholesterol levels and intrahepatic triglyceride accumulation. Conversely, L-PK overexpression in male mice resulted in augmented disease phenotypes. In contrast, female mice overexpressing L-PK were unaffected. Mechanistically, L-PK altered mitochondrial pyruvate flux and its incorporation into citrate and this, in turn, increased liver triglycerides via upregulated de novo lipogenesis and increased PNPLA3 levels accompanied by mitochondrial dysfunction. Also, L-PK increased plasma cholesterol levels via increased PCSK9 levels. L-PK silencing, on the other hand, reduced de novo lipogenesis, PNPLA3 and PCSK9 levels and improved mitochondrial function. Finally, in fibrosis model, we demonstrate that L-PK silencing in male mice reduced both liver steatosis and fibrosis, accompanied by reduced de novo lipogenesis and improved mitochondrial function.

CONCLUSION

L-PK acts in a male-specific manner in the development of liver steatosis and fibrosis. Given that NAFLD/NASH exhibit sexual dimorphism, our results have important implications for the development of personalized therapeutics.

中文翻译：

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肝丙酮酸激酶以性别特异性方式促进小鼠和人类的 NAFLD/NASH。

背景和目的

非酒精性脂肪性肝病 (NAFLD) 的病因知之甚少，男性和某些人群的易感性显着增加。使用系统遗传学方法，包括对约 100 种不同的近交系小鼠进行多组学分析，我们最近确定了几个驱动 NAFLD 的候选基因。我们使用患者样本和小鼠模型研究了其中一种肝丙酮酸激酶（L-PK 或Pklr ）在 NAFLD 中的作用。

方法

我们检查了两性小鼠和减肥手术患者队列中的 L-PK 表达。我们在饮食诱导的脂肪变性和纤维化的独立动物模型中使用了肝脏特异性功能丧失和获得策略。治疗后，我们测量了几种代谢表型，包括肥胖、胰岛素抵抗、血脂异常、肝脂肪变性和纤维化。肝组织用于基因表达和免疫印迹，并对肝线粒体生物能量学进行了表征。

结果

在小鼠和人类中，L-PK 表达在雄性中通过睾酮上调，并且与 NAFLD 严重程度密切相关。在脂肪变性模型中，与对照组相比，雄性小鼠的 L-PK 沉默改善了葡萄糖耐量、胰岛素敏感性和乳酸/丙酮酸耐受性。此外，这些动物的血浆胆固醇水平和肝内甘油三酯积累降低。相反，雄性小鼠中的 L-PK 过度表达导致疾病表型增加。相反，过度表达 L-PK 的雌性小鼠不受影响。从机制上讲，L-PK 改变了线粒体丙酮酸通量及其与柠檬酸的结合，这反过来又通过从头上调增加了肝脏甘油三酯脂肪生成和增加的 PNPLA3 水平并伴有线粒体功能障碍。此外，L-PK通过增加 PCSK9 水平来增加血浆胆固醇水平。另一方面，L-PK 沉默降低了从头脂肪生成、PNPLA3 和 PCSK9 水平并改善了线粒体功能。最后，在纤维化模型中，我们证明雄性小鼠中的 L-PK 沉默可减少肝脏脂肪变性和纤维化，同时减少从头脂肪生成和改善线粒体功能。

结论

L-PK 在肝脏脂肪变性和纤维化的发展中以男性特异性方式起作用。鉴于 NAFLD/NASH 表现出性别二态性，我们的结果对个性化治疗的发展具有重要意义。