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Abstract：
BACKGROUND: Extracellular adenosine triphosphate (ATP) is an important signal molecule. In previous studies, intensive research had revealed the crucial roles of family with sequence similarity 3 member A (FAM3A) in controlling hepatic glucolipid metabolism, islet β cell function, adipocyte differentiation, blood pressure, and other biological and pathophysiological processes. Although mitochondrial protein FAM3A plays crucial roles in the regulation of glucolipid metabolism via stimulating ATP release to activate P2 receptor pathways, its mechanism in promoting ATP release in hepatocytes remains unrevealed.
METHODS: db/db, high-fat diet (HFD)-fed, and global pannexin 1 (PANX1) knockout mice, as well as liver sections of individuals, were used in this study. Adenoviruses and adeno-associated viruses were utilized for in vivo gene overexpression or inhibition. To evaluate the metabolic status in mice, oral glucose tolerance test (OGTT), pyruvate tolerance test (PTT), insulin tolerance test (ITT), and magnetic resonance imaging (MRI) were conducted. Protein-protein interactions were determined by coimmunoprecipitation with mass spectrometry (MS) assays.
RESULTS: In livers of individuals and mice with steatosis, the expression of ATP-permeable channel PANX1 was increased (P < 0.01). Hepatic PANX1 overexpression ameliorated the dysregulated glucolipid metabolism in obese mice. Mice with hepatic PANX1 knockdown or global PANX1 knockout exhibited disturbed glucolipid metabolism. Restoration of hepatic PANX1 rescued the metabolic disorders of PANX1-deficient mice (P < 0.05). Mechanistically, ATP release is mediated by the PANX1-activated protein kinase B-forkhead box protein O1 (Akt-FOXO1) pathway to inhibit gluconeogenesis via P2Y receptors in hepatocytes. PANX1-mediated ATP release also activated calmodulin (CaM) (P < 0.01), which interacted with c-Jun N-terminal kinase (JNK) to inhibit its activity, thereby deactivating the transcription factor activator protein-1 (AP1) and repressing fatty acid synthase (FAS) expression and lipid synthesis (P < 0.05). FAM3A stimulated the expression of PANX1 via heat shock factor 1 (HSF1) in hepatocytes (P < 0.05). Notably, FAM3A overexpression failed to promote ATP release, inhibit the expression of gluconeogenic and lipogenic genes, and suppress gluconeogenesis and lipid deposition in PANX1-deficient hepatocytes and livers.
CONCLUSIONS: PANX1-mediated release of ATP plays a crucial role in maintaining hepatic glucolipid homeostasis, and it confers FAM3A\'s suppressive effects on hepatic gluconeogenesis and lipogenesis.
METHODS: db/db, high-fat diet (HFD)-fed, and global pannexin 1 (PANX1) knockout mice, as well as liver sections of individuals, were used in this study. Adenoviruses and adeno-associated viruses were utilized for in vivo gene overexpression or inhibition. To evaluate the metabolic status in mice, oral glucose tolerance test (OGTT), pyruvate tolerance test (PTT), insulin tolerance test (ITT), and magnetic resonance imaging (MRI) were conducted. Protein-protein interactions were determined by coimmunoprecipitation with mass spectrometry (MS) assays.
RESULTS: In livers of individuals and mice with steatosis, the expression of ATP-permeable channel PANX1 was increased (P < 0.01). Hepatic PANX1 overexpression ameliorated the dysregulated glucolipid metabolism in obese mice. Mice with hepatic PANX1 knockdown or global PANX1 knockout exhibited disturbed glucolipid metabolism. Restoration of hepatic PANX1 rescued the metabolic disorders of PANX1-deficient mice (P < 0.05). Mechanistically, ATP release is mediated by the PANX1-activated protein kinase B-forkhead box protein O1 (Akt-FOXO1) pathway to inhibit gluconeogenesis via P2Y receptors in hepatocytes. PANX1-mediated ATP release also activated calmodulin (CaM) (P < 0.01), which interacted with c-Jun N-terminal kinase (JNK) to inhibit its activity, thereby deactivating the transcription factor activator protein-1 (AP1) and repressing fatty acid synthase (FAS) expression and lipid synthesis (P < 0.05). FAM3A stimulated the expression of PANX1 via heat shock factor 1 (HSF1) in hepatocytes (P < 0.05). Notably, FAM3A overexpression failed to promote ATP release, inhibit the expression of gluconeogenic and lipogenic genes, and suppress gluconeogenesis and lipid deposition in PANX1-deficient hepatocytes and livers.
CONCLUSIONS: PANX1-mediated release of ATP plays a crucial role in maintaining hepatic glucolipid homeostasis, and it confers FAM3A\'s suppressive effects on hepatic gluconeogenesis and lipogenesis.
摘要：
背景：细胞外三磷酸腺苷（ATP）是重要的信号分子。在以往的研究中,深入研究揭示了具有序列相似性的家族3成员A（FAM3A）在控制肝糖脂代谢中的关键作用，胰岛β细胞功能，脂肪细胞分化,血压,以及其他生物学和病理生理过程。尽管线粒体蛋白FAM3A通过刺激ATP释放激活P2受体通路在调节糖脂代谢中发挥关键作用,其促进肝细胞ATP释放的机制仍未揭示。
方法：db/db，高脂肪饮食（HFD）喂养，和全局pannexin1(PANX1)基因敲除小鼠，以及个体的肝脏切片，在这项研究中使用。腺病毒和腺相关病毒用于体内基因过表达或抑制。为了评估小鼠的代谢状态，口服葡萄糖耐量试验（OGTT），丙酮酸耐量试验(PTT),胰岛素耐量试验(ITT),和磁共振成像（MRI）。蛋白质-蛋白质相互作用通过免疫共沉淀与质谱(MS)测定来确定。
结果：在患有脂肪变性的个体和小鼠的肝脏中，ATP通透性通道PANX1的表达增加(P<0.01)。肝PANX1过表达改善肥胖小鼠糖脂代谢失调。肝PANX1敲除或全局PANX1敲除的小鼠表现出糖脂代谢紊乱。肝脏PANX1的恢复挽救了PANX1缺陷小鼠的代谢紊乱（P<0.05）。机械上,ATP释放由PANX1激活的蛋白激酶B叉头盒蛋白O1（Akt-FOXO1）途径介导，通过P2Y受体抑制肝细胞中的糖异生。PANX1介导的ATP释放也激活了钙调蛋白（CaM）（P<0.01），与c-Jun氨基末端激酶（JNK）相互作用以抑制其活性，从而使转录因子激活蛋白-1（AP1）失活，抑制脂肪酸合成酶（FAS）的表达和脂质合成（P<0.05）。FAM3A通过热休克因子1（HSF1）刺激肝细胞中PANX1的表达（P<0.05）。值得注意的是,FAM3A过表达未能促进ATP释放，抑制糖异生和脂肪生成基因的表达，并抑制PANX1缺乏的肝细胞和肝脏中的糖异生和脂质沉积。
结论：PANX1介导的ATP释放在维持肝糖脂稳态中起着至关重要的作用，它赋予FAM3A对肝糖异生和脂肪生成的抑制作用。
方法：db/db，高脂肪饮食（HFD）喂养，和全局pannexin1(PANX1)基因敲除小鼠，以及个体的肝脏切片，在这项研究中使用。腺病毒和腺相关病毒用于体内基因过表达或抑制。为了评估小鼠的代谢状态，口服葡萄糖耐量试验（OGTT），丙酮酸耐量试验(PTT),胰岛素耐量试验(ITT),和磁共振成像（MRI）。蛋白质-蛋白质相互作用通过免疫共沉淀与质谱(MS)测定来确定。
结果：在患有脂肪变性的个体和小鼠的肝脏中，ATP通透性通道PANX1的表达增加(P<0.01)。肝PANX1过表达改善肥胖小鼠糖脂代谢失调。肝PANX1敲除或全局PANX1敲除的小鼠表现出糖脂代谢紊乱。肝脏PANX1的恢复挽救了PANX1缺陷小鼠的代谢紊乱（P<0.05）。机械上,ATP释放由PANX1激活的蛋白激酶B叉头盒蛋白O1（Akt-FOXO1）途径介导，通过P2Y受体抑制肝细胞中的糖异生。PANX1介导的ATP释放也激活了钙调蛋白（CaM）（P<0.01），与c-Jun氨基末端激酶（JNK）相互作用以抑制其活性，从而使转录因子激活蛋白-1（AP1）失活，抑制脂肪酸合成酶（FAS）的表达和脂质合成（P<0.05）。FAM3A通过热休克因子1（HSF1）刺激肝细胞中PANX1的表达（P<0.05）。值得注意的是,FAM3A过表达未能促进ATP释放，抑制糖异生和脂肪生成基因的表达，并抑制PANX1缺乏的肝细胞和肝脏中的糖异生和脂质沉积。
结论：PANX1介导的ATP释放在维持肝糖脂稳态中起着至关重要的作用，它赋予FAM3A对肝糖异生和脂肪生成的抑制作用。
