Intestinal Bile Acid Transport: Biology, Physiology, and Pathophysiology

Bile acid homeostasis, which is a critical part of human health, involves hepatic biosynthesis and vectorial transport of bile acids in a number of organs (Fig. 1). Bile acids are synthesized from cholesterol via a process that is found exclusively in the liver and involves a number of cytosolic and mitochondrial enzymes (1). Inborn errors in this process lead to neonatal cholestatic liver disease (2–4). Similarly, genetic disruption of cholesterol 7α-hydroxylase, a rate-limiting enzyme in bile acid biosynthesis, leads to a fatal form of cholestasis, which can be reversed by primary bile acid administration (5). Up-regulation of bile acid biosynthesis, which can be induced by bile acid wasting, has been used as a means of treating hypercholesterolemia (6). Bile acids, after being synthesized in the liver, are secreted into bile by an adenosine triphosphate–dependent transporter referred to as the bile salt excretory pump. Once in the intestine, bile salts play a crucial role in activating digestive enzymes and solubilizing fats and fat-soluble vitamins. A small percentage of bile salts may be reabsorbed in the proximal intestine by either passive or carrier-mediated transport processes (7–9). Most bile salts are reclaimed in the distal ileum by a sodium-dependent apically located bile acid transporter (10,11), referred to as apical sodium-dependent bile acid transporter (ASBT). Inside the enterocyte, bile salts may be bound to intracellular binding proteins (eg, ileal lipid binding protein (12)). At the basolateral surface of the enterocyte, a truncated version of ASBT is involved in vectorial transfer of bile acids into the portal circulation. Completion of the enterohepatic circulation occurs at the basolateral surface of the hepatocyte by a transport process that is primarily mediated by a sodium-dependent bile acid transporter (13). The enterohepatic circulation of bile salts is modified by bile acid transport processes in the bile duct epithelium and renal tubules.FIG. 1.: Enterohepatic circulation of bile salts. Bile salts are synthesized in the liver and excreted into the bile ducts by an ATP-driven transporter (bile salt excretory pump = 2). Most bile salts are reabsorbed in the terminal ileum by a sodium-dependent transporter (apical sodium dependent bile acid transporter [ASBT] = 3). ASBT is also expressed on the apical surface of cholangiocytes and renal proximal tubules. An organic anion transporter (5) is expressed on the apical surface of jejunum and ileum and mediated sodium-independent absorption of bile salts. At the basolateral surface of a truncated version of tASBT (4) mediates transport of bile salts into the portal circulation. tASBT is also expressed in cholangiocytes and renal tubule cells. The enterohepatic circulation of bile salts is completed at the basolateral surface of hepatocytes by a sodium-dependent transport process (sodium-dependent taurocholate transporting polypeptide [ntcp] = 1).Intestinal bile acid transport plays a key role in the enterohepatic circulation of bile salts. Molecular analysis of this process has recently led to important advances in our understanding of the biology, physiology and pathophysiology of intestinal bile acid transport. Apical sodiumdependent bile acid transporter is the major carrier protein involved in intestinal reclamation of bile salts. Complete genetic disruption of ASBT activity leads to pathologic bile acid induced diarrhea, whereas partial inhibition of ASBT-mediated transport can be used to treat hypercholesterolemia and cholestasis. Apical sodiumdependent bile acid transporter is expressed on the apical surface of ileal enterocytes, renal tubule cells, and cholangiocytes. A truncated alternatively spliced form of ASBT is expressed on the basolateral surface of cholangiocytes. Apical sodium-dependent bile acid transporter undergoes a biphasic pattern of expression in the rat ileum during normal development. The bile acid responsiveness of the ASBT gene is species specific and also depends on the experimental method used to perturb bile acid homeostasis. ASBT expression is up-regulated by corticosteroids and down-regulated in animal models by ileal inflammation. Specific patterns of adaptation of ASBT expression have been observed in various models of intestinal resection. The molecular basis of the regulation of ASBT in various settings suggests that transcriptional and posttranscriptional mechanisms both are operational. This review will summarize our current understanding of the biology, physiology, and pathophysiology of the apical sodium-dependent bile acid transporter. PHYSIOLOGY AND MOLECULAR BASIS OF INTESTINAL BILE ACID TRANSPORT For more than a century it has been known that intestinal reclamation of bile salts occurs in the ileum (1). Various experimental techniques have been used to study intestinal bile acid transport, including surgical exclusion and functional analysis of isolated segments of intestine and transport assays using everted intestine, isolated enterocytes, and brush border membrane vesicles (2–4). Evidence indicates that the majority of bile acid reclamation occurs in the terminal ileum (5–7). The physiologic relevance of jejunal transport of bile salts to the enterohepatic circulation is not adequately characterized (8,9). Kinetic analysis of ileal bile acid transport showed that it was a sodium-dependent carrier-mediated process (4,10). Using an expression cloning strategy based on this finding, Dawson et al. (11) cloned the hamster ileal sodium-dependent bile acid transporter. Northern blotting analysis showed a similarly sized transcript in kidneys, in which apical sodium-dependent bile acid transport was previously observed (12). Subsequently, the ASBT complement DNA (cDNA) has been cloned in human, rat, mouse, and rabbit ileum (13–16). The substrate specificity of human ASBT expressed in COS cells includes conjugated and unconjugated bile salts but not sulfated bile acids or estrogen (14). Western blotting and indirect immunofluorescence analysis has showed that the ASBT gene product is a 48-kd protein localized to the apical surface of ileal enterocytes and proximal renal convoluted tubules (15). It is also expressed on the apical surface of large bile duct epithelial cells (16,17). A novel truncated 19-kDa form of ASBT has been identified in ileum, kidneys, and cholangiocytes (18). It is the consequence of exon-2 skipping that results in a frameshift change and truncation of the ASBT peptide from 348 to 154 amino acids. Alteration of the carboxy-terminal amino acid sequence may permit sorting to the basolateral membrane (19). This truncated protein seemss to be functional and is most likely involved in facilitating efflux of bile salts at the basolateral membrane. Two additional proteins have been implicated in the process of intestinal bile acid transport. OATP3 is a member of the organic anion transport protein family and may facilitate carrier-mediated bile acid transport in the proximal intestine (9). The ileal lipid binding protein (also known as the ileal bile acid binding protein or ILBP) is a cytosolic protein, which binds bile salts and is localized in the terminal ileum (20). It is presumed to be the ileal intracellular binder of bile salts; however, there is no direct evidence that this protein is physiologically relevant to the process of intestinal bile salt transport. ILBP is not expressed in cholangiocytes or renal tubule cells (B. Shneider, unpublished data, January 1997). Therefore, ILBP does not seem to be essential to the process of bile acid transport. The function of ASBT is known from transfection studies and analysis of human disease. Transfection of a number of different cell lines and Xenopus laevis oocytes shows that the ASBT protein functions as a sodium-dependent conjugated bile acid transporter with an apparent 2:1 Na+: bile acid coupled electrogenic stoichiometry (10,11,19,21). In the terminal ileum, it is clear that ASBT plays a key role in bile acid reclamation. Physiologic and molecular analysis of two children with congenital ileal bile acid malabsorption and intractable diarrhea showed that the children had a compound heterozygote defect in the human ASBT gene (22,23). Additional transporters for bile salts, including OATP, may exist in the proximal intestine; however they seem to be of reduced quantitative importance overall in the reclamation of intestinal bile salts (8,9,24). The physiologic role of ASBT in kidney and bile duct epithelia is not certain. Presumably, ASBT in the kidney serves to facilitate tubular reabsorption of bile salts and explains the relatively low concentration of bile acids in the urine of healthy persons. Expression of ASBT in cholangiocytes may subserve cholehepatic shunting of bile acids, although the physiologic relevance of this process is unknown. Interestingly, ASBT has recently been identified in the gallbladder of humans (25) and rabbits (B. Shneider, unpublished data, November 2000), and may play a role in modifying bile composition and in the pathogenesis of gallstones (26). Study of knockout mice will ultimately be instructive in advancing our understanding of the range of physiologic functions of ASBT. APICAL SODIUM-DEPENDENT BILE ACID TRANSPORTER IN HUMAN HEALTH AND DISEASE Analysis of ASBT function in humans can be accomplished using various experimental approaches. Measurement of fecal bile acids, which is the gold standard measurement, is a noninvasive but cumbersome means of assessing intestinal malabsorption of bile salts. Serum bile acid response to a test-meal stimulus is a relatively easy but potentially unreliable in of assessing intestinal bile acid transport function (27). Normal intestinal bile acid transport is suggested by a greater than twofold increase in serum bile salts 2 hours after a standard meal. This approach requires the presence of a gallbladder and relatively normal intestinal transit times. Kinetic analysis of orally or intravenously administered radiolabeled bile acids can be used to determine intestinal bile acid transport and bile acid pool size (28). Direct assessment of ileal sodium-dependent bile acid transport can be performed using either ileal mucosa or brush border membrane vesicles derived from endoscopically obtained ileal biopsy specimens (22,29). 75SeHCAT ([23-75Se]25-homocholyltaurine) is a radiolabeled bile acid analog that can be used to assess the enterohepatic circulation of bile salts. First-pass intestinal clearance of SeHCAT and total body retention of this compound are excellent markers of intestinal bile acid transport function (30,31). This compound has been widely used outside of the United States, but unfortunately has not been licensed for use within the United States. Indirect assessment of bile acid malabsorption can be inferred from measurement of serum 7α-hydroxy-4-cholesten-3-one, a marker of bile acid biosynthesis (32,33) Apical sodium-dependent bile acid transporter plays a key role in a number of physiologic and pathophysiologic processes and thus has a central role in human health and disease (Table 1). The immediately obvious examples are diseases that result from genetic defects in ASBT. As described previously, two children have been described who have severe congenital diarrhea, steatorrhea, and failure to thrive secondary to a compound heterozygote mutation in the ASBT gene (23). Fecal bile acid analysis and kinetic analysis of orally administered radiolabeled bile salts showed marked bile acid malabsorption and a contracted bile acid pool. Transport assays using ileal mucosal biopsy specimens showed markedly reduced uptake of bile salts as compared with biopsy specimens from children with ileostomies and no intrinsic ileal disease. Twenty years after the childrens' initial presentation, analysis of the human ASBT gene was performed in one of the affected children. Using single-stranded conformational polymorphism (SSCP) and direct sequence analysis, one allele was found to contain two separate missense mutations, both of which abrogated sodium-dependent bile acid transport activity. The other allele was found to contain a splice site mutation, which is presumed to lead to exon 3 skipping. The potentially truncated messenger (mRNA) protein, or both is presumed to be unstable, or however, this was not in the of a functional protein after exon 2 this is to (18). The of a ASBT mutation during the cloning of human ASBT from an ileal from the ileum of a with disease does not seem to be a with for ASBT however, large studies will be to this Apical sodium-dependent bile acid transporter knockout mice will be in the functional of of ASBT Interestingly, a recently described hepatocyte knockout has markedly reduced ASBT protein expression and evidence of bile acid malabsorption ASBT in human in intestinal bile acid transport function have been implicated in various other A number of have that primary bile acid malabsorption is involved in intractable diarrhea of bile acid malabsorption may in a number of with In it has been that ileal bile acid transport be with an and defects in ileal bile acid transport may also be involved in however, the pathophysiologic is Apical sodium-dependent bile acid transporter and protein reduced in ileal biopsy obtained from with In ASBT expression is affected by intestinal disease and results in Bile acid malabsorption in leads to bile diarrhea and of the bile salt pool. bile acid malabsorption may be a for the of Various animal including a rat ileal this however, of an in humans is with and disease also leads to bile acid malabsorption and diarrhea Bile acid malabsorption has been in human and may lead to It is not known this is a direct of on ASBT or an indirect mediated by intestinal or bile acid malabsorption is in in of relatively normal ileal has also been to bile acid malabsorption Bile acid malabsorption is a major in children with malabsorption with is one involved in the pathogenesis of bile acid however the of malabsorption does not with the of bile acid wasting, that there may be specific in ASBT function in A defect in bile acid transport was suggested by in analysis of taurocholate transport using ileal biopsy specimens from children with The role of in bile acid malabsorption is not bile acid malabsorption is in in which ileal is apical sodium-dependent bile acid transport activity and ASBT protein expression are in rabbit models of ileal OF APICAL SODIUM-DEPENDENT BILE ACID TRANSPORTER ASBT can lead to human there are in which either surgical or in ASBT function has had important The most of is the of hypercholesterolemia and in and by of ileal bile acid reclamation leads to fecal of bile salts and hepatic of bile acid biosynthesis from The result is a increase in cholesterol that this is from the on the of the of partial ileal for in which surgical in ileal bile acid reabsorption lead to a in cholesterol and disease (6). the basis of important a number of have been which with ASBT function studies in and rabbits have that increase fecal bile acid and serum cholesterol In was observed in rabbits that one of induced bile acid has also been used in the of of cholestasis. In children with disease may and in liver disease after either partial or partial ileal it will be of to ASBT or novel bile acid will be as as surgical this approach can be to other of is not of the of which to be in cholestatic liver is to with ileal reclamation of bile salts OF APICAL SODIUM-DEPENDENT BILE ACID TRANSPORTER In of the of ASBT in human health and there is in understanding of normal regulation may lead to novel and to of current In of regulation may permit to diseases in which ASBT expression is The normal regulation of the expression of ASBT is (Table 2). ASBT has a pattern of including terminal ileum, proximal renal convoluted large and gallbladder In the rat intestine, ASBT protein and function are to the terminal of the small intestine In ASBT expression is primarily to the terminal of the small intestine whereas it is localized to the terminal in the hamster small intestine of ASBT of the the expression in the intestine may lead to to expression in proximal This be in disease and in which the terminal ileum is In the response of ASBT to intestinal is and is dependent on the of intestinal ileal ASBT expression is to of the intestine that ASBT. ASBT expression to be down-regulated in surgical models of intestinal but not in more ileal The physiologic of this be of the ileum into the proximal intestine does not ASBT expression and leads to reabsorption of bile salts and cholesterol malabsorption jejunum is used in small it does not have the to terminal ileal gene Physiologic and molecular studies have that intestinal bile acid transport is up-regulated in the intestine during normal development. This has been observed in and In this process is with an of intestinal ASBT protein and expression The mechanisms the expression of ASBT are not and bile acid can of intestinal bile acid transport A to the mechanisms the regulation of ASBT is the that the expression of the gene product in the kidney is during this (15). have not analysis of the regulation of ASBT in the bile duct In the of ASBT expression at is in a of expression that was This pattern is of the expression of the ileal lipid binding protein in the (20). assessment of normal rat indicates that transcriptional and posttranscriptional are involved in the regulation of ASBT Measurement of ASBT by blotting and ASBT by assays has been performed in rat and there is more ASBT in the intestine than in the and there is a increase in ASBT in the intestine, there is a increase in two that ASBT and change during normal intestinal development. BILE ACID The bile acid responsiveness of the ASBT gene is an of have observed that the ASBT gene is or of bile acids in experimental of assessment of and species may for of the that have been of the bile acid responsiveness of intestinal bile acid transport performed in and using or bile acid transport of bile salts was with by Using this regulation was observed Kinetic analysis of brush border membrane vesicles from the ileum of to either bile duct or showed regulation In studies from our analysis of bile salts, or bile acid showed no change in sodium-dependent bile acid transport or in ASBT protein expression have also that bile duct in no change in ASBT studies have that administration of which can be in the proximal intestine, leads to an increase in ASBT and protein in A study of bile duct or in regulation of bile acid transport of the ileal lipid binding protein, OATP3 and ASBT The number of that have been performed in mice seem to regulation of ASBT Bile acid leads to of whereas of bile salts in cholesterol knockout mice or after leads to of transport. of this will be to different The importance of the bile acid responsiveness of ASBT is most in the of there is a response to ASBT expression or to bile acid wasting, the of this approach will be of that the of ASBT An of the physiologic of the bile acid responsiveness of ASBT can be from analysis of of the of cholesterol in the rat and rabbit in rabbits but not leads to marked In both cholesterol leads to absorption of cholesterol and of bile acids from In there is no change in ASBT the of ASBT is for of the bile acid there is no increase in the pool size and no that bile acid The result is of cholesterol to bile acids and in In in the ASBT is up-regulated and is not for bile acid and the bile acid pool This leads to of bile acid biosynthesis and the major of cholesterol bile acid the response of ASBT is a major in cholesterol homeostasis. advances in our understanding of the molecular mechanisms of bile acid responsiveness may to the the bile acid responsiveness of the ASBT gene Bile acids the expression of the ileal lipid binding protein gene via of a of the and the acid Bile of the cholesterol gene is more It involves of of the which in the liver which is a for cholesterol gene and are expressed in the small intestine In it does not that is expressed in the small intestine and this is in ASBT an is likely to be involved The of disruption of the gene in mice on expression of ASBT are not clear analysis of the of bile acids on ASBT including analysis of the of and will likely the bile acid responsiveness of the ASBT AND bile acid malabsorption has been in with disease Using a rabbit et al. showed that rabbit ASBT is down-regulated by inflammation. corticosteroids ASBT expression in this ASBT also to be in a with in both the rat and the rabbit ileum OF OF INTESTINAL regulation in the intestine has been by using and in of the analysis of in and protein and studies of The two have identified a of and in a number of intestinal including ileal bile acid binding protein, and The and are the in both a of to be involved in the of In transcriptional analysis of the human showed that it as a in cell lines and specificity to cell lines proteins described in the proximal of Two of hepatic and both of which are previously found to play a key role in hepatic gene regulation The other is a member of the family of proteins This protein to be involved in the regulation of various intestinal and is also involved in intestinal cell Similarly, the family of to be involved in the regulation of a of intestinal Various also seem to be involved in the regulation of the including and The mechanisms by which direct expression is not in have observed in intestinal gene of one compared with may be one used to direct the of intestinal gene including regulation and responsiveness analysis of the mechanisms of intestinal gene regulation has analysis of the of intestinal potentially more analysis of the of to direct cell and In as described previously, can that specific and may be involved in no cell or adequately normal studies permit in of have been used in the of a range of intestinal intestinal and liver acid binding ileal lipid binding protein, and A number of seem to have from although clear understanding of the molecular mechanisms of and expression is small proximal can expression in liver, and intestine acid binding In the intestinal specificity of the gene is by an that is more than from the gene patterns of expression of have been observed to be by relatively small of the acid and lipid binding proteins OF IN OF INTESTINAL has been to play a key role in the regulation of the expression of a range of this of gene regulation in the It that plays a role in the regulation of various a and by and by in in or in analysis of the role of in intestinal gene regulation is leads to marked in in cells The human small intestine peptide transporter is to a small via of substrate In cells, is after with the of have been performed using in A of of in hepatic studies have been accomplished using of the transcriptional and The mechanisms of of are mediated by and specific binding proteins with and is a and binding proteins increase the of by with direct to Specific can at specific which both to and to the of and which may be by The characterized examples of this process and which have Specific most that are and Two specific binding and to and may This of is for intestinal can be by the of a will have in the range of to whereas may as as to is by cell using of after has been of can be induced by A number of transcriptional have been used in this and and The major of is which can result in An approach is to and the of Serum by serum of the has been used for this approach the use of a has been described for the analysis of This may be the and may be the method for in analysis of for the analysis of have also recently been characterized and potentially permit into the physiologic relevance of in studies reabsorption of bile salts plays a crucial role in human health and disease. This process is primarily localized to the terminal ileum and is mediated by a 48-kd sodium-dependent bile acid = ASBT is also expressed in renal tubule cells, and the skipping leads to a truncated version of which to the basolateral surface and mediates efflux of bile salts. mutation of ASBT leads to congenital diarrhea secondary to bile acid inhibition of ASBT may be in the of hypercholesterolemia and cholestasis. normal in the rat ileum, ASBT undergoes a biphasic pattern of expression with a and at the of The bile acid responsiveness of the ASBT gene is not clear and may be dependent on both the experimental used and the species being studies of the transcriptional and posttranscriptional regulation of the ASBT gene and analysis of ASBT knockout mice will into the biology, physiology, and pathophysiology of intestinal bile acid transport.