Substrate specificity of the rat liver Na+-bile salt cotransporter in Xenopus laevis oocytes and in CHO cells

It has been proposed that the hepatocellular Na+-dependent bile salt uptake system exhibits a broad substrate specificity in intact hepatocytes. In contrast, recent expression studies in mammalian cell lines have suggested that the cloned rat liver Na+-taurocholate cotransporting polypeptide (Ntcp) may transport only taurocholate. To characterize its substrate specificity Ntcp was stably transfected into Chinese hamster ovary (CHO) cells. These cells exhibited saturable Na+-dependent uptake of [H-3]taurocholate [Michaelis constant (K-m) of similar to 34 mu M] that was strongly inhibited by all major bile salts, estrone S-sulfate, bumetanide, and cyclosporin A. Ntcp cRNA-injected Xenopus laevis oocytes and the transfected CHO cells exhibited saturable Na+-dependent uptake of [H-3]taurochenodeoxycholate (K-m of similar to 5 mu M), [H-3]tauroursodeoxycholate (K-m of similar to 14 mu M), and [C-14]glycocholate (K-m of similar to 27 mu M). After induction of gene expression by sodium butyrate, Na+-dependent transport of [H-3]estrone 3-sulfate (K-m of similar to 27 mu M) could also be detected in the transfected CHO cells. However there was no detectable Na+-dependent uptake of [H-3]bumetanide or [H-3]cyclosporin A. These results show that the cloned Ntcp can mediate Na+-dependent uptake of all physiological bile salts as well as of the steroid conjugate estrone 3-sulfate. Hence, Ntcp is a multispecific transporter with preference for bile salts and other anionic steroidal compounds.