The organic solute transporters alpha and beta are induced by hypoxia in human hepatocytes.
OBJECTIVE
The organic solute transporters alpha and beta (OSTα-OSTβ) form a heterodimeric transporter located at the basolateral membrane of intestinal epithelial cells and hepatocytes. Liver injury caused by ischaemia-reperfusion, cancer, inflammation or cholestasis can induce a state of hypoxia in hepatocytes. Here, we studied the effect of hypoxia on the expression of OSTα-OSTβ.


METHODS
OSTα-OSTβ expression was measured in Huh7 cells and primary human hepatocytes (PHH) exposed to chenodeoxycholic acid (CDCA), hypoxia or both. OSTα-OSTβ promoter activity was analysed in luciferase reporter gene assays. Binding of hypoxia-inducible factor-1 alpha (HIF-1α) to the OSTα-OSTβ gene promoters was studied in electrophoretic mobility shift assays (EMSA).


RESULTS
Expression of OSTα and OSTβ increased in PHH under conditions of hypoxia. Exposure of Huh7 cells or PHH to CDCA (50 μM) enhanced the effect of hypoxia on OSTα mRNA levels. In luciferase assays and EMSA, the inducing effect of low oxygen could be assigned to HIF-1α, which binds to hypoxia responsive elements (HRE) in the OSTα and OSTβ gene promoters. Site-directed mutagenesis of either the predicted HRE or the bile acid responsive FXR binding site abolished inducibility of the OSTα promoter, indicating that both elements need to be intact for induction by hypoxia and CDCA. In a rat model of chronic renal failure, the known increase in hepatic OSTα expression was associated with an increase in HIF-1α protein levels.


CONCLUSIONS
OSTα-OSTβ expression is induced by hypoxia. FXR and HIF-1α bind in close proximity to the OSTα gene promoter and produce synergistic effects on OSTα expression.
Department of Clinical Pharmacology and Toxicology, University Hospital Zurich, Zurich, Switzerland.
Carlos A
CA
Schaffner
Jessica
J
Mwinyi
Zhibo
Z
Gai
Wolfgang E
WE
Thasler
Jyrki J
JJ
Eloranta
Gerd A
GA
Kullak-Ublick
D000818:Animals; D001665:Binding Sites; D015687:Cell Hypoxia; D002460:Cell Line; D002635:Chenodeoxycholic Acid / Q000494:pharmacology; D004195:Disease Models, Animal; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000378:metabolism; D007676:Kidney Failure, Chronic / Q000378:metabolism; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D017207:Rats, Sprague-Dawley; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism; D020218:Response Elements; D014162:Transfection; D015854:Up-Regulation; D000099150:Receptor, Farnesoid X-Activated
D002635:Chenodeoxycholic Acid; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D026901:Membrane Transport Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D000099150:Receptor, Farnesoid X-Activated; C497442:HIF1A protein, human; C544003:Slc51a protein, rat; C544004:Slc51b protein, rat; C476575:organic solute transporter alpha, human; C476577:SLC51B protein, human
bile acid transport; chronic renal failure; gene regulation; ligand; nephrectomy; nuclear receptor; organic anion transport; rat liver