Aortic dissection (AD) is the circumferential or transversal tear of the aorta wall that allows blood to infiltrate the layers. MicroRNA (miR) analyses have demonstrated a correlation between miR-320 family and AD. The underlying mechanism is yet unclear. The matrix metalloproteinases (MMPs) are a group of proteolytic enzymes that could catalyze the degeneration of the extracellular matrix and the destruction of the vasculature. In this study, we investigated whether miR-320 presented a role in regulating the production of MMPs in aortic dissection. In a cohort of 30 CE patients and 30 healthy controls, the transcription and secretion of MMP-1, MMP-2, MMP-3, MMP-8, MMP-9, and MMP-12 by monocytes were investigated. The monocyte from AD patients presented significantly elevated capacity of MMP expression than those from healthy controls. In contrast, the monocyte/macrophage expression of miR-320 was significantly lower in AD patients than in controls. In both AD patients and healthy controls, LPS-activation of macrophages resulted in MMP upregulation and miR-320 downregulation, in which the MMP expression was significantly higher while the miR-320 expression was significantly lower in AD patients than in healthy controls. Transfection of miR-320 mimic did not affect MMP gene transcription but significantly reduced the protein production in some MMPs, demonstrated that miR-320 were involved in the post-transcriptional regulation of MMPs. Together, these results demonstrated that miR-320 could regulate the expression of MMPs by macrophages, through which miR-320 may interfere with AD development.