Chain length and saturation status of fatty acids significantly affect cellular injury and lipid metabolism of primary human hepatocytes

Epidemiological and animal studies suggest that (dietary) lipids differ in their potential to drive the development and progression of non-alcoholic fatty liver disease (NAFLD). However, systematic analyses of the effects of different fatty acids (FAs) on hepatocellular steatosis and injury are missing.

The aim of this study was to systematically examine the effects exerted by FAs with different chain lengths and saturation statuses in an in vitro model of hepatocellular steatosis.

Methods:

Primary human hepatocytes (PHH) were treated with the following FAs complexed to albumin: palmitate (C16:0), stearate (C18:0), palmitoleate (C16:1) and oleate (C18:1). Subsequently, primary murine hepatocytes (pmH) and the human hepatoma cell line HepG2 were similarly treated with the different FAs.

Results:

Saturated FAs caused significant cytotoxicity already at doses as low as 0.2 mM, while doses of unsaturated FA as high as 1.2 mM did not affect viability of PHH. Furthermore, C18 FAs were more cytotoxic compared to C16 FAs. In subsequent experiments with subtoxic doses, all FA species induced a similar dose-dependent cellular lipid uptake and intracellular lipid accumulation. However, induction of oxidative stress and pro-inflammatory gene expression by saturated FAs was significantly higher compared to unsaturated FAs, while no difference was observed due to chain length. In HepG2 cells, the toxic range was similar to that in PHH, while pmH tolerated significantly higher doses of all four analysed FAs. In both pmH and HepG2 cells, lipid uptake was similar to PHH. However, in pmH, palmitoleate (C16:1) induced a greater intracellular triacylglyceride accumulation compared to palmitate (C16:0), while there was no difference with regard to C18 FAs. As seen in PHH, saturated FAs caused more oxidative stress in HepG2 cells and pmH, but no difference was observed regarding chain length. Furthermore, pro-inflammatory gene expression was only moderately induced in HepG2 cells and pmH compared to PHH. Surprisingly, in HepG2 cells as well as in pmH, the unsaturated FAs oleate or palmitoleate were the strongest inducers.

Conclusion:

Our study reveals significant differences in the effects caused by different types of FA with regard to induction of toxicity, lipid metabolism and pro-inflammatory gene expression in primary human hepatocytes. Saturated species appeared the most harmful and these detailed findings could be added to existing dietary recommendations to prevent the development or progression of NAFLD. Results in murine hepatocytes or human hepatoma cells only incompletely reflected the data obtained with primary human hepatocytes, meaning that any potential clinical translation of (in vitro) data obtained with these cells requires additional validation.