Bile acid-induced necrosis in primary human hepatocytes and in patients with obstructive cholestasis

Abstract

Accumulation of bile acids is a major mediator of cholestatic liver injury. Recent studies indicate bile acid composition between humans and rodents is dramatically different, as humans have a higher percent of glycine conjugated bile acids and increased chenodeoxycholate content, which increases the hydrophobicity index of bile acids. This increase may lead to direct toxicity that kills hepatocytes, and promotes inflammation. To address this issue, this study assessed how pathophysiological concentrations of bile acids measured in cholestatic patients affected primary human hepatocytes. Individual bile acid levels were determined in serum and bile by UPLC/QTOFMS in patients with extrahepatic cholestasis with, or without, concurrent increases in serum transaminases. Bile acid levels increased in serum of patients with liver injury, while biliary levels decreased, implicating infarction of the biliary tracts. To assess bile acid-induced toxicity in man, primary human hepatocytes were treated with relevant concentrations, derived from patient data, of the model bile acid glycochenodeoxycholic acid (GCDC). Treatment with GCDC resulted in necrosis with no increase in apoptotic parameters. This was recapitulated by treatment with biliary bile acid concentrations, but not serum concentrations. Marked elevations in serum full-length cytokeratin-18, high mobility group box1 protein (HMGB1), and acetylated HMGB1 confirmed inflammatory necrosis in injured patients; only modest elevations in caspase-cleaved cytokeratin-18 were observed. These data suggest human hepatocytes are more resistant to human-relevant bile acids than rodent hepatocytes, and die through necrosis when exposed to bile acids. These mechanisms of cholestasis in humans are fundamentally different to mechanisms observed in rodent models.

Sweroside ameliorates α-naphthylisothiocyanate-induced cholestatic liver injury in mice by regulating bile acids and suppressing pro-inflammatory responses

“…However, the role of increased hepatic bile acids in the progress of ANITinduced liver injury is not known. A bile acid mixture significantly increased the mRNA expression of pro-inflammatory cytokines in vitro in our study, which is consistent with results from a previous report [5] . TCA exhibited the highest increase after ANIT treatment in our study, and this factor increases the expression of MIP-2 in rodents and IL-8 in humans [6,7] .…”

“…Numerous studies have suggested that the intrahepatic accumulation of bile acids may directly or indirectly cause further damage to the liver [3] . One of the hypotheses regarding the pathophysiology of cholestasis is that accumulated hepatic bile acids directly induce cell apoptosis, as supported by studies on rat hepatocytes and human hepatoma lines [4,5] . Increasing attention has been paid the physiological role of bile acids acting as pro-inflammatory signals, which may trigger hepatic CXC chemokine formation during the development of cholestatic liver injury [6,7] .…”

The role of bile salts in liver regeneration

“…Dose-dependent stimulation of liver regeneration was also observed in mice given alisol B 23-acetate, a plant triterpenoid with FXR agonistic activity [58]. Lastly, the synthetic FXR agonist Px20350 could overcome defective regeneration in aged mice [39]. In a clinical context, impaired regeneration of the (small and/or compromised) remnant liver can result in PLF.…”

“…Although low bile salt levels impair liver regeneration, an intrahepatic bile salt overload causes hepatotoxic effects [39]. While diet containing 0.2 % cholic acid is stimulatory, feeding of a 1.0 % cholic acid diet resulted in mortality in hepatectomized mice indicating that toxic bile salt levels had been reached [40].…”

Mitochondrial dysfunction as a mechanism of drug-induced hepatotoxicity: current understanding and future perspectives

“…Other biomarkers such as miR-122 [244–247], full-length cytokeratin-18 [247–250], high mobility group box 1 (HMGB1) protein [247–249, 251], and argininosuccinate synthetase (ASS) [252] are generally considered indicators of mainly necrotic cell death [253]. On the other hand, increased plasma levels of the caspase-cleaved form of cytokeratin-18 [247, 248, 250, 254, 255] and activated caspases detected as enzyme activity and/or the protein by western blotting [164, 247, 249, 254], are markers of apoptotic cell death. However, due to the sensitivity of the assay, cleaved cytokeratin-18 levels should always be compared to the full-length version in order to assess the relative importance of apoptosis [247, 248, 254].…”