Taraxasterol alleviates fatty acid-induced lipid deposition in calf hepatocytes by decreasing ROS production and endoplasmic reticulum stress.

Increased concentrations of free fatty acids (FFA) induce reactive oxygen species (ROS) generation and endoplasmic reticulum (ER) stress, thus, increasing the risk of fatty liver in dairy cows during the periparturient period. In non-ruminants, Taraxasterol (Tara; a pentacyclic triterpenoid found in medicinal plants) plays an important role in anti-inflammatory and antioxidant reactions. Whether Tara can alleviate or prevent fatty liver in ruminants is unknown. We addressed whether Tara supply could dampen lipid accumulation, ROS production, and ER stress caused by FFA in calf hepatocytes. Primary calf hepatocytes were isolated from 5 healthy calves (1 d old, female, 30-40 kg, fasting, rectal temperature 38.7-39.7 °C). In the first experiment, hepatocytes were incubated with various concentrations of Tara (2.5, 5 and 10 μg/mL) for 12 h prior to 1.2 mM FFA challenge. Results indicated that the level of ROS was lowest with 5 μg/mL Tara. Thus, to further characterize the molecular mechanisms whereby Tara protects from FFA-induced lipid deposition in calf hepatocytes, we performed incubations with 5 μg/mL Tara for 12 h prior to a 1.2 mM FFA challenge for an additional 12 h. Results indicated that 1.2 mM FFA challenge increased mitochondrial membrane potential (MMP), enhanced expression of proteins and mRNA associated with endoplasmic reticulum stress (PERK, IRE1, GRP78, ATF6 and CHOP) and fatty acid synthesis (FASN, ACC1 and SREBP-1c), and ultimately led to increased lipid droplet synthesis. In contrast, Tara treatment alleviated these negative effects after 1.2 mM FFA challenge. To determine whether Tara protects against FFA-induced lipid droplet synthesis by alleviating oxidative stress, hepatocytes were treated with 5 μg/mL Tara for 22 h prior to H2O2 (440 μM) challenge for 2 h. Compared with H2O2 treatment alone, results revealed marked decreases in ROS, MMP and protein abundance of ER stress (GRP78, ATF6 and CHOP) and lipid droplet synthesis in response to Tara prior to H2O2 challenge. Data suggested that the increase of mitochondrial ROS production contributes to lipid accumulation in calf hepatocytes. Collectively, our in vitro data indicate that Taraxasterol alleviates fatty acid-induced lipid deposition. Further research is warranted to ascertain that Taraxasterol can be helpful in the therapeutic management of early lactation cows to control or alleviate excessive hepatic lipid deposition.