Methods: HepG2 cells were treated with chenodeoxycholic acid (CDCA) (200 μmol/L) (Sigma-Aldrich, St. Louis, MO) with and without the addition of ω3PUFA (5 μM EPA combined with 5 μM DHA) (Sigma Aldrich). Concentrations of CDCA and ω3PUFA for these experiments were based on our previously published work. HepG2 cells were treated with 200 μM CDCA ± ω3PUFA for 0.5 hours, and PPARα, LXRα, FXR and RXR mRNA levels were evaluated by RT-PCR using an ABI 7500 Sequence Detector and SYBR Green PCR Master Mix according to the manufacturer's instructions. Levels of mRNA were normalized to 18s ribosomal RNA quantified simultaneously in a multiplex RTPCR reaction. Specific primers were designed for the genes of interest (PPARα, FXR, LXRα, RXR). See Table 1 for primer sequences. All samples were analyzed in triplicate.
Cultured HepG2 cells were treated with ω3PUFA (EPA 5μM +DHA 5μM) alone; 200μM  chenodeoxycholic acid alone; and then with the combinations ω3PUFA, and varying concentrations of PPARα antibody (3 – 10 μg/mL). Apoptosis was evaluated using the Apo-ONE Homogeneous Caspase-3/7 Assay purchased from Promega Corporation (Madison, WI) and performed according to the manufacturer’s instructions. HepG2 cells were treated for 12 hours followed by the addition of caspase-3/7 reagent and incubation for 4 hours in the dark on a rocking shaker at low speed. Results were read at fluorescein 485nm/535nm with a Victor 2, Perkin-Elmer Wallace 1420 multilabel counter (Shelton, CT) as previously described.
Generally, when treated with chenodeoxycholic acid (CDCA), it's important for all people to take care with their diet. Liver gallstones patients have upper abdominal fullness, discomfort under the right flank, anorexia, bitter mouth or mouth odor and other symptoms, the patient would like to eat some enjoyably especially like grapes, oranges, hawthorn, apple has such obviously sour fruit in order to eliminate the above-described bad feeling. However, in patients with liver gallstones after eating these fruits tend condition actually worsened, more abdominal fullness, discomfort turned down by the right flank pain, loss of appetite and even vomiting. patients with choledocholithiasis obstructive jaundice and fever phenomenon due to the inflammation of the bile duct caused appears.