Objective We investigated the role of miR-181b in regulating atherosclerotic plaque formation in atherosclerosis (AS) by targeting Mex3B expression. Methods Male Sprague-Dawley rats were randomly divided into a normal group, model group, control group, and experimental group. A mixture of miR-181b-inhibitor-NC and miR-181b- inhibitor was injected into the control group and the experimental group, while rats in the normal group and model group received the same amount of saline. At 24 hours after the injection, a 600, 000 U/ kg dose of vitamin D3 was intraperitoneally injected into the rats in the model group, control group, and experimental group, which then received a daily 100 g high-fat diet. The rats in the normal group always received the same amount of a basic diet. After 60 days of feeding, the rats underwent carotid ultrasound examination to measure the intima-media thickness of the posterior wall of the artery (IMT) and the plaque area (square, S). RT-PCR was used to detect miR-181b expression and the mRNA of Mex3B protein, and a Western blot was used to detect the expression of Mex3B protein. A luciferase reporter gene analysis was used to examine the regulatory relationship between miR-181b and Mex3B. Results Compared with the normal group, carotid intima hyperplasia was evident in the model group, and we observed significant increases in carotid IMT, plaque S, miR-181b and Mex3B mRNA expression, and Mex3B protein expression (P< 0. 05). Compared with the model group, carotid intima hyperplasia was evident in the experimental group, and we observed significant decreases in carotid IMT, plaque S, miR-181b and Mex3B mRNA expression, and Mex3B protein expression (P< 0. 05). Luciferase reporter gene analysis demonstrated that miR-181b targets Mex3B. Conclusions miR-181b is involved in the inflammatory stress response in atherosclerosis by targeting Mex3B expression. Inhibition of miR-181b expression may significantly inhibit inflammatory responses and reduce plaque formation in AS.