Dietary oxysterols are believed to be associated with the progression of non-alcoholic fatty liver disease (NAFLD). However, the molecular basis of the association between dietary oxysterols and NAFLD is poorly understood. We hypothesized that hepatic Niemann-Pick C1-Like 1 (NPC1L1), a cholesterol re-absorber from bile to the liver, would regulate hepatic oxysterol levels and affects NAFLD progression.

Considering the species differences in hepatic NPC1L1 expression, we used liver-specific NPC1L1 transgenic (NPC1L1^Tg) mice as a human model and demonstrated that oxysterol-rich heated cholesterol exacerbated high-fat diet-induced steatosis, an early stage of NAFLD, in a hepatic NPC1L1-dependent manner. Analyses of hepatic and biliary oxysterol levels in NPC1L1^Tg mice and in vitro oxysterol uptake assays with NPC1L1-overexpressing cells revealed that NPC1L1 can uptake some, but not all, oxysterols and suppress their biliary excretion. Furthermore, in vitro and in vivo analyses revealed that 22(R)-hydroxycholesterol (22R-OHC) and 25-hydroxycholesterol (25-OHC), which are NPC1L1 substrates, were primarily involved in steatosis progression, via the activation of liver X receptor α and retinoid-related orphan receptor γ, respectively. Consistent with these results, examination of clinical specimens revealed that among the 14 major oxysterols analyzed, plasma concentrations of 22R-OHC and 25-OHC were significantly positively correlated with hepatic fat accumulation in humans.

Among the major dietary oxysterols, 22R-OHC and 25-OHC are particularly potent in promoting the progression of hepatic steatosis in a hepatic NPC1L1-dependent manner.