The side-chain cleavage of cholesterol by cytochrome 450 CYP11A1 (ie, classical pathway of steroidogenesis) is not implicated in the biosynthesis of MBG, as it is for endogenous ouabain. Previous studies in toads, known to produce bufadienolides, demonstrated that administration of labeled cholesterol and cholanic acid, but not of the product of cholesterol side-chain cleavage, progesterone, resulted in the incorporation of a radioactive label into bufadienolide molecules. Because the extra-hepatic synthesis of bile acids has recently been described, we hypothesized that in placenta, in which high levels of MBG are detected in preeclampsia, bufadienolides could be synthesized from bile acids. In human trophoblast JEG-3 cells, we examined the impact of post-transcriptional silencing of two genes, encoding sterol 27-hydrolase (CYP27A1), an initiating enzyme for synthesis of cholanic acid from cholesterol, and CYP11A1, an enzyme which controls side-cleavage of cholesterol into pregnenolone, on production of MBG and progesterone. Levels of steroids were measured in cell culture media. In JEG-3 cells, silencing of CYP11A1 resulted in 90% decrease of the protein amount (Western blot), in 80% decrease of mRNA (qPCR), and reduced production of progesterone by 77%, but did not affect production of MBG as compared to non-transfected and mock-transfected cells. Silencing of CYP27A1, accompanied by 75% reduction in CYP27A1 protein, and by 65% reduction in mRNA level, conversely, did not affect production of progesterone, but suppressed production of MBG by 80% vs. that in non-transfected and mock-transfected cells. Thus, in human trophoblast cells, bufadienolide cardiotonic steroid MBG is synthesized from cholesterol via bile acid pathway which represents a novel pathway of hormone biosynthesis.

Agency
National Institute of Health (NIH)
Institute
National Institute on Aging (NIA)
Type
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
1ZIAAG000868-04
Application #
8335945
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
4
Fiscal Year
2011
Total Cost
$217,651
Indirect Cost
Name
National Institute on Aging
Department
Type
DUNS #
City
State
Country
Zip Code
Fedorova, Olga V; Zernetkina, Valentina I; Shilova, Victoria Y et al. (2015) Synthesis of an Endogenous Steroidal Na Pump Inhibitor Marinobufagenin, Implicated in Human Cardiovascular Diseases, Is Initiated by CYP27A1 via Bile Acid Pathway. Circ Cardiovasc Genet 8:736-45
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Fedorova, Olga V; Emelianov, Igor V; Bagrov, Konstantin A et al. (2015) Marinobufagenin-induced vascular fibrosis is a likely target for mineralocorticoid antagonists. J Hypertens 33:1602-10
Grigorova, Yulia N; Juhasz, Ondrej; Zernetkina, Valentina et al. (2015) Aortic Fibrosis, Induced by High Salt Intake in the Absence of Hypertensive Response, Is Reduced by a Monoclonal Antibody to Marinobufagenin. Am J Hypertens :
Jablonski, Kristen L; Fedorova, Olga V; Racine, Matthew L et al. (2013) Dietary sodium restriction and association with urinary marinobufagenin, blood pressure, and aortic stiffness. Clin J Am Soc Nephrol 8:1952-9
Fedorova, Olga V; Kashkin, Vladimir A; Zakharova, Irina O et al. (2012) Age-associated increase in salt sensitivity is accompanied by a shift in the atrial natriuretic peptide modulation of the effect of marinobufagenin on renal and vascular sodium pump. J Hypertens 30:1817-26
Haller, Steven T; Kennedy, David J; Shidyak, Amjad et al. (2012) Monoclonal antibody against marinobufagenin reverses cardiac fibrosis in rats with chronic renal failure. Am J Hypertens 25:690-6
Nikitina, Elena R; Mikhailov, Anton V; Nikandrova, Ekaterina S et al. (2011) In preeclampsia endogenous cardiotonic steroids induce vascular fibrosis and impair relaxation of umbilical arteries. J Hypertens 29:769-76
Fedorova, Olga V; Shapiro, Joseph I; Bagrov, Alexei Y (2010) Endogenous cardiotonic steroids and salt-sensitive hypertension. Biochim Biophys Acta 1802:1230-6

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