epithelium, may become disrupted in inflammation. leaky tight junctions could allow commensal bacteria and toxins to enter portal circulation and reach the liver, where they can trigger inflammation and fibrosis. the intestinal dysbiosis theory hypothesizes that yet unidentified environmental triggers (e.g., diet, medication, inflammation) reduce microbiota diversity and/or alter the population of specific species. the resulting imbalance between primary and secondary bile acids may lead to psc via the gut-liver axis. the primary bile acids cholic acid (ca) and chenodeoxycholic acid (cdca) are synthesized in the liver and undergo conjugation before being released into the small intestine to aid digestion. in the distal ileum, 95% of these conjugated bas are actively reabsorbed via asbt but 5% enter the colon and are converted by gut microbes into deconjugated secondary bile acids, predominantly deoxycholic acid (dca) and lithocholic acid (lca). dca and lca are then reabsorbed into portal circulation and reach the liver, where they serve as signaling molecules that maintain bile acid homeostasis. specifically, dca and lca and potent agonists of farnesoid x receptor (fxr) and takeda g protein-coupled receptor 5 (tgr5), both of which mediate anti-inflammatory and cholangioprotective effects upon activation. on cholangiocytes, tgr5 activation induces cftr to secrete chloride into bile ducts, which then drives anion exchanger 2 to secrete bicarbonate into bile canaliculi. bicarbonate serves to protect the apical surface of cholangiocytes from damage by bile acids. on macrophages, activation of fxr and tgr5 inhibits nf-κb, thereby reducing production of proinflammatory cytokines. therefore, it is hypothesized that a reduction in secondary bile acid production, as a result of dysbiosis, could lead to bile duct damage via decreased activation of fxr and tgr5. indeed, lower levels of secondary bile acids were found in psc patients, but a causal relationship is yet to be confirmed. familial intrahepatic cholestasis main article: progressive familial intrahepatic cholestasis familial intrahepatic cholestasis (fih) is a group of disorders that lead to intrahepatic cholestasis in children. most often, fih occurs during the first year of life, with an incidence rate of 1/50,000 to 1/100,000. there are three different versions of fih, with each causing a different severity of jaundice. typically, children exhibit recurrent jaundice episodes, which eventually become permanent. diagnosis usually occurs by analyzing laboratory features, liver biopsy results, dna/rna sequences, and biliary lipid analysis. the definitive treatment for fih is liver transplant which usually results in a high recovery rate. each type of fih is a result of a different mutation. the three genes thought to be involved include apt8b1, which encodes for the fic1 protein. the abcb11 gene encodes for the bile salt export pump (bsep) protein, and the abcb4 gene encodes for the multidrug resistance 3 (mdr3) protein. bsep and mdr3 are respectively responsible for transporting bile salt and phospholipid, two major constituents of bile, across the apical membrane of hepatocytes. alagille syndrome main article: alagille syndrome alagille syndrome is an autosomal dominant disorder that impacts five systems, including the liver, heart, skeleton, face, and eyes. in the early part of life (within the first three months), patients with alagille syndrome exhibit conjugated hyperbilirubinemia, severe pruritus, and jaundice. bile duct obliteration usually worsens over time, causing cirrhosis of the liver and eventual failure. diagnosis usually occurs using the classic criteria by looking at changes associated with the five systems discussed earlier. like fih, the definitive treatment is a liver transplant. almost all patients with alagille syndrome have mutations of the genes involved in the notch signaling pathway. most have a mutation of the jag1 gene, while a small minority have a mutation of the notch2 gene. sepsis a variety of factors associated with sepsis may cause cholestasis. typically, patients have conjugated hyperbilirubinemia and alkaline phosphatase (alp) elevation but not to extreme levels. sepsis-induced cholestasis may occur due to increased serum lipopolysaccharide levels. lipopolysaccharides can inhibit and down-regulate bile salt transporters in hepatocytes, thereby leading to cholestasis. as such, in the case of sepsis, cholestasis occurs not as a result of impaired obstruction but rather the disruption of bile flow. ischemic liver injury resulting from sepsis can also cause cholestasis. importantly, jaundice is not indicative of cholestasis in all cases. widespread hemolysis resulting from sepsis may release bilirubin, thereby overwhelming bilirubin reabsorption and excretion mechanism. tpn-based cholestasis total parenteral nutrition (tpn) is given to patients with intestinal failure or a variety of other gastrointestinal problems. under normal settings, tpn causes a slight elevation of alp levels. however, this does not indicate cholestasis alone. in the case of tpn-induced cholestasis, there is an excessive elevation of alp, gamma-glutamyltransferase (ggt), and conjugated bilirubin. without appropriate intervention, symptoms can quickly exacerbate, leading to liver cirrhosis and failure. cholestasis arising from tpn has a diverse range of causes, including toxicity to tpn components, underlying disorders, or a lack of enteral nutrition. without enteral food consumption, gallbladder function is greatly inhibited, leading to gallstone formation, subsequent blockage, and eventually cholestasis. cholestasis resulting from tpn may also be a result of reduced bile flow from portal endotoxins. with tpn, there is a reduction in gastrointestinal motility, immunity, with an increase in permeability. these changes facilitate bacteria growth and increase the amount of circulating endotoxin. moreover, given that patients using tpn often have underlying health problems, drugs being used with known liver toxicity may also cause cholestasis. lipids in tpn may cause cholestasis and liver damage by overwhelming clearage mechanisms. intravenous glucose can also cause cholestasis as a result of increased fatty acid synthesis and decreased breakdown, which facilitates the accumulation of fats. intrahepatic cholestasis of pregnancy (obstetric cholestasis) main article: intrahepatic cholestasis of pregnancy intrahepatic cholestasis of pregnancy (icp) is an acute cause of cholestasis that manifests most commonly in the third trimester of pregnancy. it affects 0.5–1.5% of pregnancies in europe and the us and up to 28% in women of mapuche ethnicity in chile. icp is characterized by severe pruritus and elevated serum levels of bile acids as well as transaminases and alkaline phosphatase. these signs and symptoms resolve on their own shortly after delivery, though they may reappear in subsequent pregnancies for 45–70% of women. in the treatment of icp, current evidence suggests ursodeoxycholic acid (udca), a minor secondary bile acid in humans, is the most effective drug for reducing pruritus and improving liver function. the etiology of icp is multifactorial and likely involves hormonal, genetic, and environmental factors. several observations suggest estrogen plays a major role: icp begins in the third trimester, when estrogen levels are highest, resolves after estrogen levels return to normal post-delivery, and occurs with higher incidence in multiple pregnancies, where estrogen levels are more elevated than usual. although estrogen's exact pathomechanism in icp remains unclear, several explanations have been offered. estrogen may induce a decrease in the fluidity of the hepatic sinusoidal membrane, leading to a decrease in the activity of basolateral na+/k+-atpase. a weaker na+ gradient results in diminished sodium-dependent uptake of bile acids from venous blood into hepatocytes by the sodium/bile acid cotransporter. more recent evidence suggests that estrogen promotes cholestasis via its metabolite estradiol-17-β-d-glucuronide (e2). e2 secreted into the canaliculi by mrp2 was found to repress the transcription of bile salt export pump (bsep), the apical abc transporter responsible for exporting monoanionic conjugated bile acids from hepatocytes into bile canaliculi. e2 was also found to upregulate mir-148a, which represses expression of the pregnane x receptor (pxr). pxr is a nuclear receptor in hepatocytes that senses intracellular bile acid concentrations and regulates gene expression accordingly to increase bile efflux. genetic predisposition for icp is suggested by familial and regional clustering of cases. several studies have implicated heterozygous mutations of the genes abcb11 and abcb4 in icp, which respectively encode the canalicular transport proteins bsep and multidrug resistance protein 3 (mdr3). mdr3 is responsible for exporting phosphatidylcholine, the major lipid component of bile, into bile canaliculi where it forms micelles with bile salts to prevent the latter from damaging luminal epithelium. bile flow requires canalicular secretion of both bile salts and phosphatidylcholine. mdr3 mutations are an established predisposing factor, found in 16% of icp cases. more recently, studies have demonstrated involvement of bsep mutations in at least 5% of cases. the v444a polymorphism of abcb11 in particular may lead to icp by causing a reduction in hepatic bsep expression and consequently decreased bile salt export. other notable mutations identified in icp patients include ones in the farnesoid x receptor (fxr), a nuclear receptor in hepatocytes which activates transcription of mdr3 and bsep upon binding intracellular bile acids, thereby increasing canalicular bile efflux. mechanism bile is secreted by the liver to aid in the digestion of fats. bile formation begins in bile canaliculi that form between two adjacent surfaces of liver cells (hepatocytes) similar to the terminal branches of a tree. the canaliculi join each other to form larger and larger structures, sometimes referred to as the canals of hering, which themselves join to form small bile ductules that have an epithelial surface. the ductules join to form bile ducts that eventually form either the right main hepatic duct that drains the right lobe of the liver, or the left main hepatic duct draining the left lobe of the liver. the two ducts join to form the common hepatic duct, which in turn joins the cystic duct from the gall bladder, to give the common bile duct. this duct then enters the duodenum at the ampulla of vater. in cholestasis, bile accumulates in the hepatic parenchyma. one of the most common causes of extrahepatic, or obstructive cholestasis, is biliary obstruction. this is better known as choledocholithiasis where gallstones become stuck in the common bile duct. hepatocyte transporters implicated in drug-induced cholestasis. mechanisms of drug-induced cholestasis drugs may induce cholestasis by interfering with 1) hepatic transporters, 2) bile canaliculi dynamics, and/or 3) cell structure and protein localization. hepatic transporters are essential for maintaining enterohepatic bile flow and bile acid homeostasis. therefore, their direct inhibition by certain drugs may lead to cholestasis. relevant transporters implicated include bsep, mdr3, mrp2-4, and ntcp. cholestasis can result from competitive inhibition of bsep by several drugs, including cyclosporine a, rifampicin, nefazodone, glibenclamide, troglitazone, and bosentan. bsep is the main transporter in hepatocytes responsible for exporting bile salts across the apical membrane into bile canaliculi. therefore, inhibiting bsep should cause cytotoxic bile salts to accumulate in hepatocytes, leading to liver injury and impaired bile flow. indeed, there is a strong association between bsep inhibition and cholestasis in humans, and bsep inhibitors are shown to induce cholestasis in vitro. however, hepatocytes have safety mechanisms that can compensate for impaired canalicular bile efflux. in response to cholestasis, mrp3 and mrp4 on the basolateral membrane are upregulated to allow efflux of accumulated bile salts into portal blood. similarly, mrp2 can accommodate additional bile flow across the apical membrane in cholestatic conditions. these compensatory mechanisms explain why some bsep inhibitors do not cause cholestasis. on the contrary, contrast, drugs that inhibit both mrp3/4 and bsep (e.g., rifampicin, troglitazone, bosentan) pose greater risk for cholestasis mdr3 is another key canalicular efflux transporter that is the target of inhibition by certain drugs. mdr3 secretes phosphatidylcholine into bile canaliculi, where it form micelles with bile salts to dissolve cholesterol as well as protect hepatocyte and cholangiocytes from damage by bile salts. mdr3 inhibition leads to low phospholipid concentrations in bile that damages cholangiocytes and leads to cholestasis. antifungal azoles such itraconazole have been shown to inhibit both mdr3 and bsep, thus giving them higher cholestatic potential. other mdr3-inhibiting drugs are chlorpromazine, imipramine, haloperidol, ketoconazole, saquinavir, clotrimazole, ritonavir, and troglitazone. another target for inhibition, mrp2 is an apical efflux transporter that mainly exports bilirubin glucuronide and glutathione into bile. however, mrp2 is also the preferential route of export for certain sulfated conjugated bas (taurolithocholic acid and glycolithocholic acid), so its inhibition could contribute to cholestasis. on the hepatocyte basolateral membrane, na+-taurocholate cotransporting peptide (ntcp) is the major transporter of conjugated bile acids. enterohepatic bile flow requires the concerted activity of both ntcp and bsep, which form the major route by which bas enter and exit hepatocytes respectively. therefore, ntcp inhibitors, such as cyclosporine a, ketoconazole, propranolol, furosemide, rifamycin, saquinavir, and ritonavir, should theoretically cause cholestasis by decreasing hepatocyte ba uptake. however, no relationship was found between ntcp inhibition and dic risk, possibly because basolateral sodium-independent oatps can partially compensate for bile salt uptake. therefore, ntcp