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The Journal of Cell Biology, Vol 127, 401-410, Copyright © 1994 by The Rockefeller University Press

ARTICLES

Fluorescent choleretic and cholestatic bile salts take different paths across the hepatocyte: transcytosis of glycolithocholate leads to an extensive redistribution of annexin II

JC Wilton, GM Matthews, RD Burgoyne, CO Mills, JK Chipman and R Coleman
School of Biochemistry, University of Birmingham, Edgbaston, United Kingdom.

We have used fluorescent derivatives of the choleretic bile salts cholate and chenodeoxycholate, the cholestatic salt lithocholate, and the therapeutic agent ursodeoxycholate to visualize distinct routes of transport across the hepatocyte and delivery to the canalicular vacuole of isolated hepatocyte couplets. The cholate and chenodeoxycholate derivatives produced homogeneous intracellular fluorescence and were rapidly transported to the vacuole, while the lithocholate analogue accumulated more slowly in the canalicular vacuole and gave rise to punctate fluorescence within the cell. Fluorescent ursodeoxycholate showed punctate intracellular fluorescence against a high uniform background indicating use of both pathways. Inhibition of vesicular transport by treatment with colchicine and Brefeldin A had no effect on the uptake of any of the compounds used, but it dramatically impaired delivery of both the lithocholate and the ursodeoxycholate derivatives to the canalicular vacuole. We conclude that while the chenodeoxycholate and cholate analogues traverse the hepatocyte by a cytoplasmic route, lithocholate and ursodeoxycholate analogues are transported by vesicle-mediated transcytosis. Treatment of couplets with glycine derivatives of lithocholate and ursodeoxycholate, but not cholate or chenodeoxycholate, led to a marked relocalization of annexin II, which initially became concentrated at the basolateral membrane, then moved to a perinuclear distribution and finally to the apical membrane as the incubation progressed. This suggests that lithocholate and ursodeoxycholate treatment leads to a rapid induction of transcytosis and that annexin II exchange occurs upon membrane fusion at all stages of the hepatocyte transcytotic pathway. These results indicate that isolated hepatocyte couplets may provide an inducible model system for the study of vesicle-mediated transcytosis.
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