Objective:To determine whether the effect of TNF-α on caspase8/3 activity in mouse adrenal cortex Y1 cells is mediated by TNF-R1. Methods:The expression of TNF-R1 was down-regulated by short hairpin RNAs(shRNAs) in Y1 cells which expressed TNF-R1 natively. The effect of TNF-α on caspase8/3 activity in native Y1 cells and miRNA-Y1 cells was evaluated by spectrophotometer. Results:Interference efficiency of shRNAs on TNF-R1 expression in miRNA-Y1 cells was 71.25%. The activity of caspase8/3 increased significantly in Y1 cells(P < 0.05) and unchanged in miRNA-Y1 cells after treated with TNF-α. Conclusion:The effect of TNF-α on caspase8/3 activity is mainly mediated by TNF-R1.