Secondary bile acid[

石胆酸是一种有毒的次级胆汁酸[1]。

孕烷X受体（PXR）是孕烯醇酮16α-腈（PCN）等剧毒类固醇的分子靶标，可诱导细胞色素P450 3A（CYP3A）的表达，保护身体免受有害化学物质的侵害。在这项研究中，我们证明PXR被有毒的胆汁酸-石胆酸（LCA）及其3-酮代谢物激活。此外，我们发现PXR调节参与胆汁酸生物合成、运输和代谢的基因的表达，包括胆固醇7α-羟化酶（Cyp7a1）和Na+非依赖性有机阴离子转运蛋白2（Oatp2）。最后，我们证明PXR的激活可以防止LCA引起的严重肝损伤。基于这些数据，我们提出PXR作为LCA的生理传感器，协调调节基因表达以降低这种有毒胆汁酸的浓度。这些发现表明，PXR激动剂可能被证明可用于治疗人类胆汁淤积性肝病[1]。

Maintenance and Treatment of PXR−/− and Control Mouse Populations. [1]
Adult male wild-type (PXR+/+) and PXR-null (PXR−/−) mice were maintained on standard laboratory chow and were allowed food and water ad libitum. Mice were treated with PCN (0.4 mg/g), dexamethasone (0.1 mg/g), sodium phenobarbital (PB, 0.1 mg/g), or LCA (0.125 mg/g). All inducers were dissolved in corn oil and injected intraperitoneally either once (PCN, dexamethasone, and PB) or twice (LCA) a day for 4 days. Animals were killed 24 h following the final injection. For LCA and PCN cotreatment, PCN was administered by gavage (0.5 mg/g in corn oil) for 3 days. Subsequently, PCN treatment was continued for a further 4 days, during which time animals also received a daily i.p. injection of LCA (0.25 mg/kg in corn oil). Mice were exsanguinated 24 h following the final injection and serum alanine transaminase (ALT) and sorbitol dehydrogenase (SDH) levels determined by using standard techniques. Livers were fixed in neutral-buffered formalin solution Sigma, embedded in paraffin wax, and stained with hematoxylin and eosin. Differences between SDH and ALT levels in vehicle- and LCA-treated and PCN- and LCA-treated animals were determined by using a one-way ANOVA. Significant differences were determined by using Duncan's multiple range post hoc test.

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Quantitation of LCA in Urine. [1]
LCA levels in urine were quantitated by atmospheric pressure ionization–liquid chromatography mass spectrometry (API-LCMS). Briefly, an equal volume of mouse urine and a 5 μg/ml methanolic solution of 2,2,4,4-d4-cholic acid (D4-cholic acid) were combined. Samples were sonicated, centrifuged (3,000 × g for 10 min), and filtered through a 0.45-μm filter unit before injection onto the analytical column of an LCMS instrument. LCA and D4-cholic acid were detected as the molecular ions ([M-H−]) 375 and 311 m/z, respectively, in the negative selected ion monitoring mode of the instrumentation. LCA concentrations in the study samples were calculated by comparison to standard solutions of LCA containing D4-cholic acid as internal standard. The significance of differences between mean values was analyzed by using an unpaired Student's t test.

[1]. The nuclear receptor PXR is a lithocholic acid sensor that protects against liver toxicity. Proc Natl Acad Sci U S A. 2001;98(6):3369-3374.

Lithocholic acid-2,2,4,4-d4 is a deuterated compound and a lithocholic acid.

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It has been almost 30 years since Hans Selye first showed that PCN treatment blocks the hepatotoxicity and mortality caused by LCA treatment in rats. In this report, we have used PXR-null mice to demonstrate that the orphan nuclear receptor PXR mediates the hepatoprotective effects of PCN against LCA-induced toxicity. Moreover, we have shown that PXR is activated by LCA and its 3-keto metabolite and coordinately regulates genes involved in the biosynthesis, transport, and metabolism of LCA. Our results indicate that PXR plays a fundamental role in protecting the liver against pathophysiological levels of LCA. PXR thus joins FXR as nuclear receptors that are activated by bile acids. Although PXR and FXR are activated by distinct sets of bile acids, both receptors are activated by LCA. This raises the interesting possibility that PXR and FXR cooperate to remove LCA from the body when its concentrations reach pathophysiological levels.
Several previous reports suggest that our findings may have implications in the treatment of human cholestatic liver disease. Notably, urine from patients suffering from cholestasis contains elevated levels of 6-hydroxylated bile acids (including the LCA metabolite hyodeoxycholic acid), which are products of CYP3A4. These findings suggest that increased 6-hydroxylation is a relevant mechanism for reducing the levels of toxic bile acids in humans. Elevated levels of 6-hydroxylated bile acids are also observed in the urine of healthy subjects treated with the PXR ligand rifampicin. Interestingly, rifampicin has been used successfully in the treatment of pruritus associated with intrahepatic cholestasis and, in some instances, has been reported to induce remission of cholestasis. The molecular basis for these clinical effects has remained obscure. Based on our data, we suggest that the anticholestatic effects of rifampicin may be mediated through PXR, and that potent PXR ligands may be more efficacious in the treatment of cholestasis, a severe hepatic disease for which there is no known cure. [1]

C24H36D4O3

380.60

380.323

83701-16-0

Lithocholic acid;434-13-9

16217405

White to off-white solid powder

1.085 g/cm3

510.992ºC at 760 mmHg

184-186ºC(lit.)

276.926ºC

5.507

57.53

2

3

4

27

574

9

[2H]C1(C[C@@]2([C@H]3CC[C@]4([C@H]([C@@H]3CC[C@@H]2C([C@@H]1O)([2H])[2H])CC[C@@H]4[C@H](C)CCC(=O)O)C)C)[2H]

SMEROWZSTRWXGI-POXZWENPSA-N

InChI=1S/C24H40O3/c1-15(4-9-22(26)27)19-7-8-20-18-6-5-16-14-17(25)10-12-23(16,2)21(18)11-13-24(19,20)3/h15-21,25H,4-14H2,1-3H3,(H,26,27)/t15-,16-,17-,18+,19-,20+,21+,23+,24-/m1/s1/i10D2,14D2

(4R)-4-[(3R,5R,8R,9S,10S,13R,14S,17R)-2,2,4,4-tetradeuterio-3-hydroxy-10,13-dimethyl-3,5,6,7,8,9,11,12,14,15,16,17-dodecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanoic acid

83701-16-0; Lithocholic acid-2,2,4,4-d4; Lithocholic Acid-d4; (4R)-4-[(3R,5R,8R,9S,10S,13R,14S,17R)-2,2,4,4-tetradeuterio-3-hydroxy-10,13-dimethyl-3,5,6,7,8,9,11,12,14,15,16,17-dodecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanoic acid; Lithocholic-2,2,4,4-d4 acid; Lithocholic Acid D4; Cholan-24-oic-2,2,4,4-d4 acid, 3-hydroxy-, (3a,5ss)-; (3a,5ss)-3-Hydroxycholan-24-oic-2,2,4,4-d4 acid; Lithocholic acid-d4; Lithocholic Acid-2,2,4,4-d4;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : 50 mg/mL (131.37 mM)

配方 1 中的溶解度: ≥ 5 mg/mL (13.14 mM) (饱和度未知) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将100 μL 50.0mg/mL澄清的DMSO储备液加入到900μL 20％SBE-β-CD生理盐水中，混匀。
*20% SBE-β-CD 生理盐水溶液的制备（4°C，1 周）：将 2 g SBE-β-CD 溶解于 10 mL 生理盐水中，得到澄清溶液。

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配方 2 中的溶解度: ≥ 5 mg/mL (13.14 mM) (饱和度未知) in 10% DMSO + 90% Corn Oil (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 50.0 mg/mL 澄清 DMSO 储备液加入900 μL 玉米油中，混合均匀。

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
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10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
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