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

营养不良占全球儿童死亡总数的一半。肝功能异常，包括胆汁减少 酸合成，是严重营养不良的常见症状。在肠道中，胆汁酸缺乏导致营养不良 吸收不良和生长受阻。在肝脏，胆汁酸通过核受体传递信号，包括法尼醇- X受体(FXR)调节从能量动态平衡到凝血的广泛过程。我们发现 营养不良患者胆汁酸合成减少导致FXR活性降低和表达减少 包括凝血因子在内的FXR靶基因。由此导致的营养不良引起的凝血障碍可能是致命的。 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K08项目使用了一个独特的 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的表达缺失- α(PPARα)和随后的过氧体丢失，这是产生必要的β氧化步骤所必需的 成熟的胆汁酸。我们公布的数据和新的初步数据表明，两个关键的细胞色素受到抑制 过氧化体β氧化上游的P450(CyP)酶也是导致胆汁酸合成减少的原因之一 营养不良，独立于PPARα。肝细胞从胆固醇合成胆汁酸的两个平行过程 小路。我们的数据表明，胆汁酸合成的经典途径是通过减少 速率决定酶CYP7A1的活性。我们预测，细胞色素P7A1活性下降是因为 营养不良的肝细胞会耗尽血红素，这是细胞色素P7A1的基本修复基团。同样，我们有 胆汁酸合成的替代途径通过减少胆汁酸的表达而被抑制的证据 细胞色素P7B1，这是该途径中的关键酶。我们预测，细胞色素P7B1的表达下降是因为 营养不良上调其强大的转录抑制因子--类固醇调节元件结合蛋白-1 (SREBP-1)。因此，我们假设，除了依赖于PPARα的过氧化物体丢失外，营养不良还会损害 胆汁酸的合成也通过抑制CYP7A1和CYP7B1来削弱经典和替代的胆汁酸 合成的途径。具体目标是：1)确定营养不良和血红素代谢改变的特征 它通过量化标记胆固醇的流量，测量细胞色素P7A1的表达水平来降低细胞色素P7A1的活性 血红素合成酶，并用血红素治疗肝细胞和小鼠；2)确定 SREBP-1在细胞色素P7B1转录抑制中的作用及其对胆汁酸下降的相对贡献 通过定义招募到CYP7B1启动子的辅抑制子和抑制营养不良的合成 SREBP-1与siRNA和药理拮抗剂的体外和体内实验。预期结果包括更深层次的 了解营养状况如何影响胆汁酸动态平衡。这种方法是创新的，因为它 将探索抑制CYP作为营养不良和胆汁酸合成受损之间的新联系。这个 拟议的研究具有重要意义，因为有可能开发新的策略来恢复肝脏合成 在严重营养不良和其他以获得性胆汁酸缺乏为特征的消化系统疾病中起作用。

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.