Endogenous ouabain: upregulation of steroidogenic genes in hypertensive hypothalamus but not adrenal.
BACKGROUND
Mammalian tissues contain a presumed endogenous Na+, K(+)-ATPase inhibitor that binds reversibly to the Na+ pump with high affinity and specificity. The inhibitor has been linked to the pathogenesis of experimental volume-expanded and human essential hypertension. This compound has been isolated from mammalian hypothalamus and appears to be an isomer of the plant-derived cardiac glycoside ouabain, if not ouabain itself. The objective of this study was to test the hypothesis that a biosynthetic pathway exists in mammalian tissues to produce a steroid derivative closely related to plant cardiac glycosides.


RESULTS
Using bioinformatics and genomic techniques, Milan hypertensive rat tissues were studied because this strain has a 10-fold increase in hypothalamic ouabain-like compound that is linked to the pathogenesis of the hypertension. A putative steroid biosynthetic pathway was constructed and candidate genes encoding enzymes in this pathway were identified from sequence databases. Differential expression of selected genes in the pathway was studied by microarray analysis and quantitative polymerase chain reaction, with functional validation by gene silencing using small interfering RNAs. Marked upregulation of genes coding for P450 side chain cleavage and Delta5-3beta-hydroxysteroid dehydrogenase/Delta5-Delta4- isomerase enzymes in hypertensive hypothalamus but not adrenal was found, compared with normotensive Milan rats. Knockdown of the latter gene decreased production of ouabain-like factor from neural tissue.


CONCLUSIONS
Our findings support the possibility that a unique steroid biosynthetic circuit exists in Milan rat brain, functioning independently from adrenal, which could account for the overproduction of the hypothalamic ouabain-like compound in this species.
Murrell
Julie R
JR
Medical Services, Massachusetts General Hospital, Charlestown, MA 02129, USA.
Randall
Jeffrey D
JD
Rosoff
James
J
Zhao
Ji-liang
JL
Jensen
Roderick V
RV
Gullans
Steven R
SR
Haupert
Garner T
GT
D000311:Adrenal Glands / Q000378:metabolism*; D000818:Animals; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000235:genetics*; D006973:Hypertension / Q000378:metabolism*; D007031:Hypothalamus / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis; D010042:Ouabain / Q000378:metabolism*; D016716:PC12 Cells; D016133:Polymerase Chain Reaction; D011375:Progesterone Reductase / Q000235:genetics*; D034622:RNA Interference; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors*; D013251:Steroid Isomerases / Q000235:genetics*
C021425:3 beta-hydroxysteroid oxidoreductase-delta(5) 3-ketosteroid isomerase; D009097:Multienzyme Complexes; D012333:RNA, Messenger; D010042:Ouabain; D011375:Progesterone Reductase; D002786:Cholesterol Side-Chain Cleavage Enzyme; D013251:Steroid Isomerases; D000254:Sodium-Potassium-Exchanging ATPase