Oral supplementation of clay to dairy cattle has been reported to reduce toxicity of aflatoxin (AF) in contaminated feed. The objective of this study was to determine the effects of 3 concentrations of dietary clay supplementation (EcoMix) after an AF challenge on hepatic gene expression of 7 different inflammation markers. Ten multiparous rumen-cannulated Holstein cows [BW (mean ± SD) = 669 ± 20 kg and 146 ± 69 DIM] were assigned to 1 of 5 treatments in a randomized replicated 5 × 5 Latin square design balanced to measure carryover effects. Periods (21 d) were divided into an adaptation phase (d 1 to 14) and a measurement phase (d 15 to 21). From d 15 to 17, cows received an AF challenge consisting of 100 μg of aflatoxin B₁ (AFB₁)/kg of dietary DMI. AFB₁ was fitted into 10-mL gelatin capsules (TORPAC, Fairfield, NJ) and administered into the rumen through the cannula based on the average DMI obtained on d 12 to 14. Treatments were POS, no clay plus an AF challenge; 3 different concentrations of clay (0.5, 1, or 2% of dietary DMI) plus an AF challenge; and control (C), no clay and no AF challenge. Statistical analysis was performed using the MIXED procedure of SAS. Contrasts included CONT1 (POS vs. C), CONT2 (POS vs. the average of 0.5, 1, or 2%), and tests of linear and quadratic treatment effects of clay inclusion. When comparing POS with C, the AF challenge caused a 2.27-fold downregulation of haptoglobin (HP; P = 0.04) and tended to have a 1.06-fold downregulation of signal transducer and activator of transcription 3 (STAT3; P = 0.10). However, when supplemented with clay, cows had a linear increase in expression of nuclear factor kappa B subunit (NFKB1; P = 0.02) and a trend for linear increase of tumor necrosis factor (TNF; P = 0.10). In conclusion, liver gene expression profiling suggested that an AF challenge downregulated inflammation and there was a restorative effect when clay was supplemented orally that seemed to counteract the immunosuppression of AF.