Cholesterol-desaturating agent; Secondary metabolite; microbial metabolite; secondary bile acid; endogenous metabolite

β-Murine cholic acid fluorescent probe diluted solution (100 μM; 48 h) in primary hepatocytes [1].
β - Muricholic acid (100 µ M; 48 h) inhibits lipid accumulation in primary mouse liver cells [1].

Beta-murine cholic acid (0.5% beta-murine cholic acid given for 8 weeks) inhibits diet-induced or experimental cholic acid in rats [2].

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Restraint stress enhanced hepatic and serum BA levels in NASH mice [1]
Next, hepatic and serum BA levels were measured to investigate whether the correlation between corticosterone and BA in serum results from stress challenge. Under the MCD diet, stress significantly increased both serum and hepatic BA levels [23.6–38.8 μM (1.64-fold) and 0.399–0.673 μmol/g liver (1.69-fold), respectively] (Fig. 2A). The stress trended to elevate hepatic BA levels under conditions of the MCS diet [the control and stress were 0.026 and 0.137 μmol/g liver (5.26-fold), respectively] (Fig. 2A). In addition, the hepatic BA composition was investigated. Hepatic tauro-beta-muricholic acid (TβMCA) and taurocholic acid (TCA) levels were significantly increased in MCD-induced NASH. However, these taurine-conjugated BAs were not changed after challenge with stress (Fig. 2B). Hepatic taurochenodeoxycholic acid (TCDCA) and taurodeoxycholic acid (TDCA) levels were not changed in MCD-induced NASH (Fig. 2B). Interestingly, nonconjugated BA and beta-muricholic acid (βMCA) were significantly elevated after challenge with stress [1.14–3.46 nmol/g liver (3.03-fold)], and cholic acid (CA) tended to be elevated [0.84–2.24 nmol/g liver (2.67-fold)] (Fig. 2B). In the tested BA-related genes, the MCD diet increased hepatic expression levels of cytochrome P450 family (CYP) 7A1 (CYP7A1), solute carrier family (SLC) 51 beta (SLC51B, also called organic solute transporter subunit beta), and ATP-binding cassette (ABC) C4 genes (3.56-, 11.73-, and 7.09-fold, respectively) (Fig. 2C). Under the MCD diet, stress enhanced SCL51B and ABCC4 expression (11.73–15.55-fold and 7.09–14.47-fold, respectively) (Fig. 2C). Interestingly, hepatic CYP7A1 expression was induced by stress under the condition of the MCS diet (4.03-fold). The results may suggest that stress directly stimulates hepatic Cyp7a1 induction. Hepatic expression of the other BA-related factors, namely, hepatic nuclear factor 4 alpha (HNF4A), liver rich homologue 1 (LRH1), farnesoid X receptor (FXR), small heterodimer partner (SHP), CYP7B1, CYP8B1, CYP27A1, bile acid-CoA:amino acid N-acyltransferase (BAAT), ABCB11, ABCC2, SLC51A, ABCC3, ABCC5, SLC10A1, and solute carrier organic anion 1a1 (SLCO1A1), was not changed in either the MCS or MCD group after stress (Fig. 2C). The highest hepatic CYP7A1 protein level was observed in the MCD-stress group (Fig. 2D). Interestingly, both serum BA and corticosterone levels were positively correlated with hepatic CYP7A1 protein levels but not hepatic CYP7A1 mRNA levels (Fig. 2E). These results suggested the existence of the corticosterone-CYP7A1 protein-BA cascade in NASH livers after stress challenge.

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Restraint stress decreased hepatic lipid levels in NASH mice via elevated hepatic βMCA levels [1]
Oil Red O staining was performed to investigate whether stress can influence hepatic lipid homeostasis in the development of NASH. Oil Red O staining showed smaller lipid droplet sizes in the livers of the MCD-stress group than in the livers of the MCD-non-stress group (Fig. 4A). In addition, triglyceride (TG), total cholesterol (TChol), and NEFA levels were measured in serum and liver after stress challenge. Stress significantly decreased both hepatic TG and NEFA levels [624–353 mg/g liver (0.57-fold) and 0.336–0.242 Eq/g liver (0.72-fold), respectively] in the MCD group (Fig. 4B) without affecting fatty acid-related gene expression in the liver (Fig. 4C). However, stress had no effect on hepatic TChol (Fig. 4B) and cholesterol-related gene expression levels in the liver (Fig. S1B). Interestingly, βMCA inhibited lipid accumulation in primary-cultured mouse hepatocytes after exposure to palmitic acid (PA)/oleic acid (OA), while CA did not (Fig. 5A, B). However, the expression levels of lipid-related genes were not altered in the hepatocytes after treatment with βMCA (Fig. 5C). On the other hand, the PA/OA-decreased CYP7A1 mRNA and protein levels tended to recover after treatment with βMCA (Fig. 5D, E). These results suggest that βMCA can suppress hepatic lipid accumulation in NASH mice.

Culture of primary hepatocytes [1]
Primary hepatocytes were prepared as previously reported. In corticosterone treatment, after starvation with FBS-negative Williams' Medium E for 2 h, the hepatocytes were exposed to 0.1% dimethyl sulfoxide as vehicle and 30–1000 nM corticosterone. Six hours later, the cells were collected and subjected to quantitative PCR and western blot analysis. For lipid accumulation, the hepatocytes were exposed to 0.2% isopropanol/1% ethanol/1% FBS solution as vehicle, 100 µM palmitic acid, or 100 µM palmitic acid and 100 µM bile acids (βMCA or CA). Forty-eight hours later, the cells were collected and subjected to quantitative PCR and western blot analysis. Another cell set was subjected to Oil Red O staining. Oil Red O staining was performed using 60% isopropanol solution with Oil Red O dye. Then, haematoxylin and eosin (HE) staining was carried out with Mayer's haematoxylin solution and pure eosin solution. For quantification of lipid accumulation, after Oil Red O staining, the dye was extracted from the stained cells with 40 mM HCl/isopropanol solution. The dye concentration was determined by measuring the absorbance at 500 nm and used as the relative lipid amount.

Animal/Disease Models: 6-8 weeks, male C57L/J mice (litholithic diet (2% cholesterol and 0.5% cholic acid))[2]
Doses: 0.5% beta-murine cholic acid
Route of Administration: Use 0.5% beta-murine cholic acid Results of 8 weeks of murine bile acid feeding diet: diminished gallstone prevalence to 20% by Dramatically reducing bile cholesterol secretion rate, saturation index and intestinal absorption, as well as inducing phase boundary movement and enlarged E-zone, preventing cholesterol Transition from liquid crystal phase to solid crystals and stones.

[1]. Stress can attenuate hepatic lipid accumulation via elevation of hepatic β-muricholic acid levels in mice with nonalcoholic steatohepatitis. Lab Invest. 2021 Feb;101(2):193-203.

[2]. Effect of beta-muricholic acid on the prevention and dissolution of cholesterol gallstones in C57L/J mice. J Lipid Res. 2002 Nov;43(11):1960-8.

β-Mouse cholic acid is a type of mouse cholic acid compound with β-configurations at both the 6- and 7-positions. It belongs to the mouse cholic acid class, 6β-hydroxysterol, and 7β-hydroxysterol. It is the conjugate acid of β-mouse cholic acid.
See also: β-mouse cholic acid (note moved to).

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It has been reported that in humans, non-alcoholic steatohepatitis (NASH) increases ethylene glycol-CA and TCA levels and decreases the ratio of secondary bile acids to primary bile acids. In this study, the MCD diet reduced the ratio of TCDCA and TDCA levels to TβMCA and TCA levels in the liver of mice. Since the hydrophilicity order of bile acids is: conjugate bile acids > non-conjugate bile acids, and βMCA > CA > CDCA > DCA, these results may indicate that hydrophilic bile acid levels are upregulated in NASH. Furthermore, in nonalcoholic steatohepatitis (NASH), stress increases hepatic β-methylcholine (βMCA) and bile acid (CA) levels, while total β-methylcholine (TβMCA) and tricarboxylic acid (TCA) levels are not elevated. Moreover, stress does not alter the expression of the Baat gene, which regulates bile acid (BA) taurine binding. Although gut microbiota unbinding responses need to be considered, these observations support the possibility that stress-induced increases in hepatic βMCA and CA levels are induced by the expression of the Cyp7a1 gene, the rate-limiting enzyme in bile acid synthesis. FXR is a nucleobiliary bile acid receptor that is strongly activated by CDCA and induces Shp expression to inhibit Cyp7a1 gene expression. Although βMCA can negatively regulate FXR activation, no changes in Shp expression levels were observed in NASH livers after stress. Furthermore, the expression levels of Hnf4a and Lrh1 genes, which upregulate Cyp7a1 gene expression, were also unchanged after stress. These results suggest that stress-induced Cyp7a1 gene expression is mediated by other factors. Stress may promote pathological changes in NASH by increasing unconjugated bile acid (BA) levels. [1] In this study, stress led to increased hepatic β-methylcholic acid (βMCA) levels and enhanced CYP7A1 expression under MCD-induced NASH conditions. Furthermore, βMCA treatment protected hepatocytes from palmitic acid (PA)/oleic acid (OA)-induced lipid accumulation. Therefore, this study suggests that βMCA produced by CYP7A1 helps to inhibit hepatocyte lipid accumulation, but the mechanism by which βMCA inhibits hepatic lipid accumulation is unclear. Since βMCA is a murine bile acid that cannot be synthesized by humans, these phenomena were not observed in patients with non-alcoholic fatty liver disease (NAFLD). However, the results may suggest that βMCA can be used to treat NAFLD. [1] In clinical studies, assessing the effects of stress on disease is difficult, mainly because there are currently no clear stress indicators. This study used serum glucocorticoid levels as a stress indicator. However, serum glucocorticoid levels can vary depending on a variety of factors, including stress. The results of this study may not be entirely consistent with clinical data, but suggest that stress may reduce liver lipid levels in patients with NAFLD. Future research is needed to establish stress indicators to further understand the molecular mechanisms by which stress affects the human body. [1] In summary, this study shows that stress affects liver bile acid and lipid homeostasis in NASH mice, suggesting that βMCA may be used to treat NASH. It is expected that the detailed molecular mechanisms will be elucidated in the future to develop NAFLD treatment drugs. [1] Stress affects the human body and is known to cause certain diseases. However, the effect of chronic restraint stress on the development of non-alcoholic fatty liver disease (NAFLD) is unclear. This study shows that chronic restraint stress can reduce liver lipid accumulation during the development of non-alcoholic steatohepatitis (NASH) by increasing liver β-mouse bile acid (βMCA) levels in mice. Serum cortisol and corticosterone levels (i.e., stress markers in humans and rodents) were correlated with serum bile acid levels in NAFLD patients and mice with methionine choline deficiency (MCD) diet-induced NASH, suggesting that stress is associated with bile acid (BA) homeostasis in NASH. In mouse models, hepatic β-MCA and bile acid (CA) levels increased after stress stimulation. Considering that short-term stress can increase CYP7A1 protein levels in the liver of normal mice, and corticosterone can increase CYP7A1 protein levels in primary mouse hepatocytes, it is hypothesized that enhanced Cyp7a1 expression is associated with increased hepatic β-MCA levels induced by chronic stress. Interestingly, chronic stress reduced hepatic lipid levels in MCD-induced NASH mice. Furthermore, β-MCA inhibited lipid accumulation in palmitic acid/oleic acid-treated primary mouse hepatocytes, while CA had no such effect. Additionally, Cyp7a1 expression appears to be associated with lipid accumulation in hepatocytes. In summary, chronic stress can alter hepatic lipid accumulation in NASH mice by inducing Cyp7a1 expression, thereby disrupting bile acid homeostasis. This study found a novel role for β-mouse cholic acid (β-MCA) in the liver, suggesting that β-MCA may be used to treat non-alcoholic fatty liver disease (NAFLD). [1]

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This study investigated whether β-mouse cholic acid (a naturally occurring trihydroxy hydrophilic bile acid in rodents) can act as a cholesterol desaturant to prevent cholesterol gallstones, and whether it can promote the dissolution of gallstones compared to ursodeoxycholic acid (UDCA). In the gallstone prevention study, male C57L mice susceptible to gallstones were fed a stone-inducing diet (2% cholesterol and 0.5% bile acid) for 8 weeks, with or without 0.5% UDCA or β-mouse cholic acid. In the study of gallstone dissolution, mice with existing gallstones were fed a normal diet with or without 0.5% β-mouse cholic acid or UDCA for 8 weeks. All mice fed the stone-inducing diet developed cholesterol stones. The addition of β-mouse cholic acid and ursodeoxycholic acid (UDCA) significantly reduced bile secretion rate, saturation index and intestinal cholesterol absorption, thereby reducing the incidence of gallstones by 20% and 50%, respectively. At the same time, β-mouse cholic acid and UDCA also induced phase boundary transition and expanded the E region, thereby preventing cholesterol from transforming from the liquid crystal phase into solid crystals and stones. Compared with the control group (10%), the complete dissolution rate of gallstones reached 100% and 60% after 8 weeks of administration of β-mouse cholic acid and UDCA, respectively. We conclude that β-mouse cholic acid is more effective than UDCA in the treatment or prevention of diet-induced or experimental cholesterol gallstones in mice. [2]

C24H40O5

408.5714

408.288

2393-59-1

5283853

White to light yellow solid powder

1.184g/cm3

565.7ºC at 760mmHg

310ºC

3.75E-15mmHg at 25°C

1.558

3.448

4

5

4

29

637

11

C[C@H](CCC(=O)O)[C@H]1CC[C@@H]2[C@@]1(CC[C@H]3[C@H]2[C@H]([C@H]([C@H]4[C@@]3(CC[C@H](C4)O)C)O)O)C

DKPMWHFRUGMUKF-CRKPLTDNSA-N

InChI=1S/C24H40O5/c1-13(4-7-19(26)27)15-5-6-16-20-17(9-11-23(15,16)2)24(3)10-8-14(25)12-18(24)21(28)22(20)29/h13-18,20-22,25,28-29H,4-12H2,1-3H3,(H,26,27)/t13-,14-,15-,16+,17+,18+,20+,21+,22-,23-,24-/m1/s1

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

beta-muricholic acid; 2393-59-1; b-Muricholic acid; beta-MCA; 5beta-Cholanic acid-3alpha,6beta,7beta-triol; (3a,5b,6b,7b)-3,6,7-trihydroxy-Cholan-24-oic acid; 3alpha,6beta,7beta-Trihydroxy-5beta-cholan-24-oic Acid; (4R)-4-[(3R,5R,6S,7R,8S,9S,10R,13R,14S,17R)-3,6,7-trihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanoic acid;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

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

DMSO : ~100 mg/mL (~244.76 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (6.12 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (6.12 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (6.12 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)