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.

Beta-muricholic acid is a member of the class of muricholic acids in which the hydroxy groups at positions 6 and 7 both have beta configuration. It is a member of muricholic acids, a 6beta-hydroxy steroid and a 7beta-hydroxy steroid. It is a conjugate acid of a beta-muricholate.
See also: beta-Muricholate (annotation moved to).

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In humans, NASH has been reported to elevate glycol-CA and TCA levels and reduce the ratio of secondary BAs/primary BAs. In the present study, the MCD diet decreased the ratio of TCDCA and TDCA levels/TβMCA and TCA levels in the mouse livers. As the hydrophilicity of BA is in the order conjugated BAs > unconjugated BAs and βMCA > CA > CDCA > DCA, these results may suggest that hydrophilic BA levels are upregulated in NASH. In addition, stress-increased hepatic βMCA and CA levels in NASH without elevated hepatic TβMCA and TCA levels. Furthermore, stress did not alter the expression of the Baat gene, which regulates the taurine conjugation of BA. Although the deconjugation reaction by gut flora needs to be considered, the observation supported the possibility that the stress-increased hepatic βMCA and CA levels result from the induction of Cyp7a1 gene expression, which is the rate-limiting enzyme of BA synthesis. FXR is a nuclear BA receptor that is strongly activated by CDCA and induces Shp expression to suppress Cyp7a1 gene expression. Although βMCA can negatively act on FXR activation, changes in SHP expression levels were not observed in NASH livers after stress challenge. In addition, the Hnf4a and Lrh1 gene expression levels, which upregulate Cyp7a1 gene expression, were not altered after the stress challenge. These results suggest that the stress induction of Cyp7a1 gene expression is mediated by other factors. Stress could contribute to the pathology changes in NASH through an increase in unconjugated BA levels. [1]
In this study, under conditions of MCD-induced NASH, stress-elevated hepatic βMCA levels with enhanced CYP7A1 expression. Furthermore, treatment with βMCA protected against PA/OA-induced lipid accumulation in hepatocytes. Thus, the present study provides a view that CYP7A1-produced βMCA contributes to the suppression of lipid accumulation in hepatocytes, but it is still unknown how βMCA suppresses hepatic lipid accumulation. Since βMCA is murine BA and is not synthesized in humans, these phenomena could not be observed in human patients with NAFLD. However, the results may suggest that βMCA is available for NAFLD therapy.[1]
In clinical research, it is difficult to assess the effect of stress on diseases mainly because no index defines stress. In this study, the serum glucocorticoid level was used as the stress index. However, serum glucocorticoid levels can change due to various factors as well as stress. The results in this study may not necessarily match the clinical data but provide the possibility that stress can decrease hepatic lipid levels in patients with NAFLD. In the future, a stress index should be required to further understand the molecular mechanism by which stress influences our bodies.[1]
In conclusion, the present study demonstrated that stress influences hepatic BA and lipid homeostasis in NASH mice, indicating the possibility that βMCA is available for NASH therapy. It is expected that detailed molecular mechanisms will be elucidated to develop a NAFLD therapeutic drug.[1]

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Stress can affect our body and is known to lead to some diseases. However, the influence on the development of nonalcohol fatty liver disease (NAFLD) remains unknown. This study demonstrated that chronic restraint stress attenuated hepatic lipid accumulation via elevation of hepatic β-muricholic acid (βMCA) levels in the development of nonalcoholic steatohepatitis (NASH) in mice. Serum cortisol and corticosterone levels, i.e., human and rodent stress markers, were correlated with serum bile acid levels in patients with NAFLD and methionine- and choline-deficient (MCD) diet-induced mice, respectively, suggesting that stress is related to bile acid (BA) homeostasis in NASH. In the mouse model, hepatic βMCA and cholic acid (CA) levels were increased after the stress challenge. Considering that a short stress enhanced hepatic CYP7A1 protein levels in normal mice and corticosterone increased CYP7A1 protein levels in primary mouse hepatocytes, the enhanced Cyp7a1 expression was postulated to be involved in the chronic stress-increased hepatic βMCA level. Interestingly, chronic stress decreased hepatic lipid levels in MCD-induced NASH mice. Furthermore, βMCA suppressed lipid accumulation in mouse primary hepatocytes exposed to palmitic acid/oleic acid, but CA did not. In addition, Cyp7a1 expression seemed to be related to lipid accumulation in hepatocytes. In conclusion, chronic stress can change hepatic lipid accumulation in NASH mice, disrupting BA homeostasis via induction of hepatic Cyp7a1 expression. This study discovered a new βMCA action in the liver, indicating the possibility that βMCA is available for NAFLD therapy.[1]

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This study investigated whether beta-muricholic acid, a natural trihydroxy hydrophilic bile acid of rodents, acts as a biliary cholesterol-desaturating agent to prevent cholesterol gallstones and if it facilitates the dissolution of gallstones compared with ursodeoxycholic acid (UDCA). For gallstone prevention study, gallstone-susceptible male C57L mice were fed 8 weeks with a lithogenic diet (2% cholesterol and 0.5% cholic acid) with or without 0.5% UDCA or beta-muricholic acid. For gallstone dissolution study, additional groups of mice that have formed gallstones were fed chow with or without 0.5% beta-muricholic acid or UDCA for 8 weeks. One hundred percent of mice fed the lithogenic diet formed cholesterol gallstones. Addition of beta-muricholic acid and UDCA decreased gallstone prevalence to 20% and 50% through significantly reducing biliary secretion rate, saturation index, and intestinal absorption of cholesterol, as well as inducing phase boundary shift and an enlarged Region E that prevented the transition of cholesterol from its liquid crystalline phase to solid crystals and stones. Eight weeks of beta-muricholic acid and UDCA administration produced complete gallstone dissolution rates of 100% and 60% compared with the chow (10%). We conclude that beta-muricholic acid is more effective than UDCA in treating or preventing 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.)