Xenobiotic Receptors

pparα激活的生物标志物：先前的一项研究发现过氧化物酶体增殖物激活受体（pparα）参与脂质稳态的控制。pparα在禁食期间被激活，它控制脂肪酸运输和代谢相关基因的表达。代谢组学，利用超高效色谱连接电喷雾电离四极杆飞行时间质谱（UPLCESI-QTOFMS）平台，发现pparα活化的生物标志物。经pparα激活剂wy - 14643处理的小鼠尿液中，生物标志物21-类固醇羧酸11β -羟基-3,20-二氧孕-4-烯-21-oic酸（HDOPA）和11β，20-二羟基-3-氧孕-4-烯-21-oic酸（DHOPA）含量非常丰富。确定了HDOPA和DHOPA产生的分子机制和代谢途径。pparα特异性的HDOPA和20 α - dhopa的时间依赖性增加与肾上腺皮质增生、高皮质血症和脾脏萎缩的发展平行，在肾上腺切除小鼠中这种情况减弱。wy - 14643激活pparα诱导肝脏FGF21，引起下丘脑神经肽Y和针刺相关蛋白mrna增加，刺激针刺相关蛋白/神经肽Y神经元，激活下丘脑-垂体-肾上腺（HPA）轴，导致肾上腺皮质增生和皮质酮产生增加，揭示了pparα和HPA轴在控制能量稳态和免疫调节中的联系。皮质酮在体内和体外均被证明是21-羧酸的前体。在PPARalpha激活下，皮质酮的经典还原代谢途径被抑制，而另一种氧化途径被发现，导致碳21的连续氧化产生HDOPA。后者被还原为最终产物20 α - dhopa。肝细胞色素P450、醛脱氢酶（ALDH3A2）和21-羟基类固醇脱氢酶（AKR1C18）参与了这一途径。PPARalpha的激活导致Aldh3a2和Akr1c18的诱导，这两种基因都是通过将启动子荧光素酶报告基因构建体引入小鼠肝脏中来证实的靶基因。本研究揭示了ppar α的下游代谢生物标志物及其上游分子机制。人类pparα活化的生物标志物：为了鉴定与脂肪酸β -氧化增加相关的潜在的内源性pparα活化的生物标志物，在使用贝特药物和pparα激活剂非诺贝特的人类中也进行了代谢组学研究。健康志愿者口服非诺贝特2周，并通过UPLCESI-QTOFMS对其尿液进行分析。通过机器学习算法随机森林识别的生物标志物包括在第14天前泛酸（大于5倍）和乙酰肉碱（大于20倍）的显著消耗，观察结果与ppar α的已知靶标一致，包括泛酸激酶和编码参与酰基肉碱运输和合成的蛋白质的基因。血清胆固醇（-12.7%）、甘油三酯（-25.6%）、尿酸（-34.7%）以及尿丙基肉碱（大于10倍）、异丁基肉碱（大于2.5倍）、(S)-(+)-2-甲基丁基肉碱（5倍）和异戊基肉碱（大于5倍）均在第14天降低。这些生物标志物作为pparα激活指标的特异性在Ppara-null小鼠中得到了证实。尿泛酸和酰基肉碱可能被证明是人类ppar诱导的脂肪酸β氧化的有用指标。这项研究说明了药物代谢组学方法在人类和近亲繁殖小鼠模型中了解药物对脂质代谢的影响的效用。Farnesoid X受体（FXR）激活的生物标志物：FXR是一种核受体，可调节胆汁酸合成、代谢和转运相关基因的表达，因此在维持胆汁酸稳态中起主要作用。采用UPLCESI-QTOFMS监测了野生型和FXR缺失型小鼠饲喂胆酸（一种FXR配体）尿液的代谢组学反应。多变量数据分析显示，野生型和无fxr小鼠在胆酸饮食中增加最多的离子是对甲酚（4-甲基酚）、皮质酮和胆酸的代谢物。上述代谢物的结构特性通过化学合成和尿液代谢物的保留时间（RT）和/或串联质量碎片模式与真实标准的比较得到证实。牛头-3 α，6,7 α,12 α -tetrol （3 α,6,7 α，12 α -四羟基-5 -胆甾醇-26-酰基牛磺酸）是CA饮食中fxr缺失小鼠中代谢增加最多的代谢物之一，可能是通过诱导Cyp3a11有效羟基化有毒胆汁酸的标志。石胆酸诱导的胆汁淤积模型显示，Cyp3a11的表达增强是fxr缺失小鼠解毒胆汁淤积胆汁酸的主要防御机制。这些结果将有助于鉴定胆汁淤积的生物标志物和确定胆汁淤积的适应性分子机制。人妊娠X受体（PXR）与炎症性肠病（IBD）的发病机制有关。利福昔明是一种人类PXR激活剂，目前正处于治疗IBD的临床试验中，并已证明对克罗恩病和活动性溃疡性结肠炎有效。本研究主要探讨利福昔明对IBD的保护和治疗作用及其作用机制。在葡聚糖硫酸钠（DSS）诱导的IBD模型和三硝基苯磺酸（TNBS）诱导的IBD模型中，利福昔明对PXR人源化（hPXR）、野生型和无PXR小鼠进行利福昔明处理，以确定PXR在IBD中的保护作用。进一步评价利福昔明对dss治疗的hPXR和pxr无效小鼠的治疗作用。结果表明，通过体重减轻、腹泻、直肠出血、结肠长度和组织学评估，预先给药利福昔明改善了dss治疗和tnbs治疗的hPXR小鼠结肠炎的临床特征。此外，在出现症状后给予利福昔明的小鼠中，观察到hPXR小鼠的存活率和结肠炎症状的恢复较高，而在pxr阴性小鼠中则没有。利福昔明处理后，hPXR小鼠NF-kappaB靶基因明显下调。体外NF-kappaB报告基因检测显示，利福昔明对表达hPXR的结肠来源细胞系的NF-kappaB活性有抑制作用。这些发现证明了利福昔明通过人PXR介导的NF-kappaB信号级联抑制对IBD的预防和治疗作用，从而提示人PXR可能是治疗IBD的有效靶点。

Biomarkers for PPARalpha activation: A previous study identified the peroxisome proliferator-activated receptor alpha (PPARalpha) is involved in the control of lipid homeostasis. PPARalpha is activated during fasting where is controls expression of genes involved if fatty acid transport and metabolism. Metabolomics, using the ultra-performance chromatography-linked electrospray ionization quadrupole time-of-flight mass spectrometry (UPLCESI-QTOFMS) platform, was employed to discover biomarkers fof PPARalpha activation. The biomarkers 21-steroid carboxylic acids 11beta-hydroxy-3,20-dioxopregn-4-en-21-oic acid (HDOPA) and 11beta,20-dihydroxy-3-oxo-pregn-4-en-21-oic acid (DHOPA) were found to be very abundant in urine of mice treated with the PPARalpha activator Wy-14,643. The molecular mechanism and the metabolic pathway of HDOPA and DHOPA production were determined. The PPARalpha-specific time-dependent increases in HDOPA and 20alpha-DHOPA paralleled the development of adrenal cortex hyperplasia, hypercortisolism, and spleen atrophy, which was attenuated in adrenalectomized mice. Wy-14,643 activation of PPARalpha induced hepatic FGF21, which caused increased neuropeptide Y and agouti-related protein mRNAs in the hypothalamus, stimulation of the agouti-related protein/neuropeptide Y neurons, and activation of the hypothalamic-pituitary-adrenal (HPA) axis, resulting in increased adrenal cortex hyperplasia and corticosterone production, revealing a link between PPARalpha and the HPA axis in controlling energy homeostasis and immune regulation. Corticosterone was demonstrated as the precursor of 21-carboxylic acids both in vivo and in vitro. Under PPARalpha activation, the classic reductive metabolic pathway of corticosterone was suppressed, whereas an alternative oxidative pathway was uncovered that leads to the sequential oxidation on carbon 21 resulting in HDOPA. The latter was then reduced to the end product 20alpha-DHOPA. Hepatic cytochromes P450, aldehyde dehydrogenase (ALDH3A2), and 21-hydroxysteroid dehydrogenase (AKR1C18) were found to be involved in this pathway. Activation of PPARalpha resulted in the induction of Aldh3a2 and Akr1c18, both of which were confirmed as target genes through introduction of promoter luciferase reporter constructs into mouse livers in vivo. This study revealed downstream metabolic biomarkers for PPARalpha and the corresponding upstream molecular mechanisms. Biomarkers for human PPARalpha activation: Metabolomics was also performed in humans administered the fibrate drug and PPARalpha activator fenofibrate in order to identify latent, endogenous biomarkers of PPARalpha activation associated with increased fatty acid beta-oxidation. Healthy human volunteers were given fenofibrate orally for 2 weeks and their urine was profiled by UPLCESI-QTOFMS. Biomarkers identified by the machine learning algorithm random forests included significant depletion by day 14 of both pantothenic acid ( greater than 5-fold) and acetylcarnitine ( greater than 20-fold), observations that are consistent with known targets of PPARalpha including pantothenate kinase and genes encoding proteins involved in the transport and synthesis of acylcarnitines. It was also concluded that serum cholesterol (-12.7%), triglycerides (-25.6%), uric acid (-34.7%), together with urinary propylcarnitine ( greater than 10-fold), isobutyrylcarnitine ( greater than 2.5-fold), (S)-(+)-2-methylbutyrylcarnitine (5-fold), and isovalerylcarnitine ( greater than 5-fold) were all reduced by day 14. Specificity of these biomarkers as indicators of PPARalpha activation was demonstrated using the Ppara-null mouse. Urinary pantothenic acid and acylcarnitines may prove useful indicators of PPARalpha-induced fatty acid beta-oxidation in humans. This study illustrates the utility of a pharmacometabolomic approach to understand drug effects on lipid metabolism in both human populations and in inbred mouse models. Biomarkers for Farnesoid X receptor (FXR) activation: FXR is a nuclear receptor that regulates the expression of genes involved in synthesis, metabolism, and transport of bile acids and thus plays a major role in maintaining bile acid homeostasis. Metabolomic responses monitored by UPLCESI-QTOFMS were investigated in urine of wild-type and Fxr-null mice fed cholic acid, an FXR ligand. Multivariate data analysis between wild-type and Fxr-null mice on a cholic acid diet revealed that the most increased ions were metabolites of p-cresol (4-methylphenol), corticosterone, and cholic acid in Fxr-null mice. The structural identities of the above metabolites were confirmed by chemical synthesis and by comparing retention time (RT) and/or tandem mass fragmentation patterns of the urinary metabolites with the authentic standards. Tauro-3alpha,6,7alpha,12alpha-tetrol (3alpha,6,7alpha,12alpha-tetrahydroxy-5beta-cholestan-26-oyltaurine), one of the most increased metabolites in Fxr-null mice on a CA diet, is a marker for efficient hydroxylation of toxic bile acids possibly through induction of Cyp3a11. A cholestatic model induced by lithocholic acid revealed that enhanced expression of Cyp3a11 is the major defense mechanism to detoxify cholestatic bile acids in Fxr-null mice. These results will be useful for identification of biomarkers for cholestasis and for determination of adaptive molecular mechanisms in cholestasis. Human pregnane X receptor (PXR) has been implicated in the pathogenesis of inflammatory bowel disease (IBD). Rifaximin, a human PXR activator, is in clinical trials for treatment of IBD and has demonstrated efficacy in Crohn's disease and active ulcerative colitis. In the current study, the protective and therapeutic role of rifaximin in IBD and its respective mechanism were investigated. PXR-humanized (hPXR), wild-type, and Pxr-null mice were treated with rifaximin in the dextran sulfate sodium (DSS)-induced and trinitrobenzene sulfonic acid (TNBS)-induced IBD models to determine the protective function of human PXR activation in IBD. The therapeutic role of rifaximin was further evaluated in DSS-treated hPXR and Pxr-null mice. Results demonstrated that pre-administration of rifaximin ameliorated the clinical hallmarks of colitis in DSS-treated and TNBS-treated hPXR mice as determined by body weight loss and assessment of diarrhea, rectal bleeding, colon length, and histology. Additionally, higher survival rates and recovery from colitis symptoms were observed in hPXR mice, and not in Pxr-null mice when rifaximin was administered after the onset of symptoms. NF-kappaB target genes were markedly down-regulated in hPXR mice by rifaximin treatment. In vitro NF-kappaB reporter assays demonstrated inhibition of NF-kappaB activity following rifaximin treatment in colon-derived cell lines expressing hPXR. These findings demonstrated the preventive and therapeutic role of rifaximin on IBD through human PXR-mediated inhibition of the NF-kappaB signaling cascade, thus suggesting that human PXR may be an effective target for the treatment of IBD.