CLA Modulates Ruminal Epithelial Inflammation

The immortalized ruminal epithelial cell line RECs was cultured in DMEM (Gibco, New York, USA) containing 2% FBS, 1% penicillin/streptomycin and 1% epithelial cell additive at 37 °C with 5% CO₂, as described previously [29 (link)]. When REC monolayers reached 70–80% confluency, the cells were treated without (control group, CON) or with 0.1 μg/mL LPS (from Escherichia coli O111:B4, Sigma-Aldrich, Shanghai, China) for 3 h [29 (link)] after pretreatment with 50 or 100 μM cis-9,trans-11- (c-9,t-11-CLA+LPS group, c-9,t-11-CLA+LPS) or trans-10,cis-12-CLA (t-10,c-12-CLA+LPS group, t-10,c-12-CLA+LPS) (purity ≥ 96.0%, Sigma-Aldrich, Shanghai, China) for 24 h or not (LPS group, LPS). Six treatments were contained and each treatment had 4 replicates.