Impaired bile acid synthesis due to CYP7A1 and CYP7B1 suppression in malnutrition

营养不良造成全球一半的儿童死亡。肝功能异常，包括胆汁减少 酸合成，在严重营养不良中很常见。在肠道，胆汁酸缺乏导致营养不良， 吸收不良和生长受损。在肝脏中，胆汁酸通过核受体发出信号，包括法尼醇- X-受体（FXR）调节从能量稳态到凝血的广泛过程。我们发现 营养不良时胆汁酸合成减少导致FXR活化减少， FXR靶基因，包括凝血因子。由此产生的营养不良引起的凝血功能障碍可能是致命的。 营养不良时胆汁酸合成受损的原因尚不清楚。我正在进行的K 08项目使用了一种独特的 早期营养不良的小鼠模型，揭示了过氧化物酶体增殖物激活受体表达的缺失， 过氧化物酶体是一个重要的β-氧化步骤所必需的， 成熟胆汁酸我们发表的数据和新的初步数据表明，抑制两个关键的细胞色素 过氧化物酶体β-氧化上游的P450（P450）酶也有助于降低胆汁酸合成， 营养不良，独立于PPARα。肝细胞从胆固醇合成胆汁酸有两个平行的途径 途径。我们的数据表明，胆汁酸合成的经典途径受到抑制， CYP 7A 1，速率决定酶。我们预测，CYP 7A 1活性下降，因为 营养不良的肝细胞缺乏血红素，血红素是CYP 7A 1的必需辅基。同样，我们有 证据表明胆汁酸合成的替代途径是通过降低 CYP 7 B1是该途径中的关键酶。我们预测CYP 7 B1的表达降低， 营养不良上调其有效的转录抑制因子，固醇调节元件结合蛋白-1 （SREBP-1）。因此，我们假设，除了PPARα依赖性过氧化物酶体丢失，营养不良损害 胆汁酸合成也通过抑制CYP 7A 1和CYP 7 B1来损害经典和替代 合成途径。具体目的是：1）表征营养不良中血红素代谢的改变， 通过定量标记胆固醇的流量，测量 血红素合成酶，以及用血红素处理肝细胞和小鼠;以及2）确定 SREBP-1在CYP 7 B1转录抑制中的作用及其对胆汁酸减少的相对贡献 通过定义招募到CYP 7 B1启动子的辅抑制因子并抑制营养不良中的合成 SREBP-1与siRNA和药理学拮抗剂的体外和体内研究。预期成果包括： 了解营养状况如何影响胆汁酸稳态。这种方法是创新的，因为它 将探索作为营养不良和受损的胆汁酸合成之间的一种新的联系的胆汁酸抑制。的 拟议的研究是重要的，因为有可能开发新的战略，以恢复肝脏合成 在严重营养不良和以获得性胆汁酸缺乏为特征的其他消化系统疾病中发挥作用。

Malnutrition contributes to half of all global child deaths. Liver function abnormalities, including decreased bile acid synthesis, are common in severe malnutrition. In the intestine, bile acid deficiency leads to nutrient malabsorption and impaired growth. In the liver, bile acids signal through nuclear receptors including farnesoid- X-receptor (FXR) to regulate a wide range of processes from energy homeostasis to coagulation. We discovered that decreased bile acid synthesis in malnutrition causes decreased FXR activation and decreased expression of FXR target genes, including coagulation factors. The resulting malnutrition-induced coagulopathy can be fatal. It is not known why bile acid synthesis is impaired in malnutrition. My ongoing K08 project uses a unique mouse model of early-life malnutrition, revealing loss of expression of peroxisome proliferator-activated receptor- α (PPARα) and subsequent loss of peroxisomes, which are required for an essential β-oxidation step to generate mature bile acids. Our published data and new preliminary data indicate that suppression of two key cytochrome P450 (CYP) enzymes upstream of peroxisomal β-oxidation also contributes to decreased bile acid synthesis in malnutrition, independently of PPARα. Hepatocytes synthesize bile acids from cholesterol in two parallel pathways. Our data suggest that the classic pathway of bile acid synthesis is suppressed through decreased activity of CYP7A1, the rate-determining enzyme. We predict that decreased CYP7A1 activity occurs because malnourished hepatocytes are depleted of heme, the essential prosthetic group for CYP7A1. Similarly, we have evidence that the alternative pathway of bile acid synthesis is suppressed through decreased expression of CYP7B1, the key enzyme in this pathway. We predict that decreased expression of CYP7B1 occurs because malnutrition upregulates its potent transcriptional repressor, the sterol regulatory element-binding protein-1 (SREBP-1). Thus, we hypothesize that, in addition to PPARα-dependent peroxisome loss, malnutrition impairs bile acid synthesis also by suppressing CYP7A1 and CYP7B1 to impair both the classic and the alternative pathways of synthesis. The Specific Aims are 1) to characterize altered heme metabolism in malnutrition and its role in decreased CYP7A1 activity by quantifying flux of labeled cholesterol, measuring expression levels of heme synthesizing enzymes, and treating hepatocytes and mice with heme; and 2) to determine the role of SREBP-1 in the transcriptional repression of CYP7B1 and its relative contribution to decreased bile acid synthesis in malnutrition by defining the corepressors recruited to the CYP7B1 promoter, and by inhibiting SREBP-1 with siRNA and pharmacologic antagonists in vitro and in vivo. Expected outcomes include a deeper understanding of how nutritional status influences bile acid homeostasis. This approach is innovative because it will explore CYP suppression as a novel link between malnutrition and impaired bile acid synthesis. The proposed research is significant because of the potential to develop new strategies to restore liver synthetic function in severe malnutrition and in other digestive diseases characterized by acquired bile acid deficiency.