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

Abstract

Abstract. We have used fluorescent derivatives of the i 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.ILE formation and secretion are exclusive to the liver, and they are essential to the emulsification and digestion of fats in the small intestine and in the detoxication and excretion of potentially toxic endobiotic and xenobiotic substances. The primary force for bile formation is thought to be the hepatobiliary transport of osmotically active bile salts. The vast majority of bile salts (90%) are reabsorbed across the intestine and taken up by hepatocytes from the portal blood for reexcretion (Coleman, 1987), while the remainder are synthesized de novo in the hepatocytes. The uptake and reexcretion of bile salts involves three distinct processes: uptake across the sinusoidal (basolateral) mem-

Efficient In Vitro vectorial transport of a fluorescent conjugated bile acid analogue by polarized hepatic hybrid WIF-B and WIF-B9 cells

“…35 Moreover, there was no dissociation of this bile acid derivative in liver, 29 and the apparent sinusoid-to-canaliculus transport time of this compound was only 0.5 seconds. 36 In hepatocyte cultures, CGamF has been observed to be transferred through the cells and to accumulate in BC of isolated hepatocyte couplets from normal 20,29,33,37 and mutant TR Ϫ rats. 33 Its uptake into the cytoplasm of cultured hepatocyte doublets was partly inhibited by removal of sodium from the medium and by addition of other bile acids.…”

Transcytosis: Crossing Cellular Barriers

“…Unfortunately, the roles of annexin XIIIa and XIIIb in transcytosis were not tested in these studies. Some evidence for a role for annexin II in transcytosis comes from studies performed in isolated hepatocyte couplets (620). Upon induction of transcytosis by the addition of bile salts, annexin II immunofluorescence increased dramatically and was redistributed sequentially from beneath the basolateral PM to perinuclear structures and finally to the apical pole (620).…”

Safra oluşumunda rol oynayan transport proteinleri Transport proteins in bile production

“…Bir mikrotübül inhibitörü olan kolşisin, litokolat ve ursodeoksikolatın vezikül içinde birikimini azaltmaktadır, ancak kolat ve kenodeoksikolat üzerinde çok az bir etkisi bulunmaktadır (29,30).…”