Abstract: Objective To investigate the role of interleukin-1β (IL-1β) in regulating the expressions of matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of matrix metalloproteinase-3 (TIMP-3) in cultured human endometrial cells of the mid-secretory phase. Methods Human endometrial cells of the mid-secretory phase cultured in the absence of steroid hormones for 24 h were stimulated with rhIL-1β at concentrations ranging from 50 to 1 000 U/ml for 48 h. The expressions of MMP-9 and TIMP-3 protein in the cells were detected by flow cytometry using immunofluorescent method. Results MMP-9 was expressed at the level of 1 491.38±68.95 in the control group, and at 1 592.40±47.57, 1 702.63±75.31, 1 994.49±52.98, and 2 347.58±45.87 in response to cell treatment with IL-1β at 50, 100, 500, and 1 000 U/ml, respectively, suggesting that IL-1β significantly increased MMP-9 expression in a dose-dependent manner (P<0.05). The expression level of TIMP-3 was 1 643.31±61.29 in the control group, and was 1 597.27±49.07, 1443.93±81.23, , 1 343.28±54.80, and 1 157.85±47.95 in 50, 100, 500, and 1 000 U/ml IL-1β groups, respectively, suggesting that IL-1β decreased TIMP-3 expression dose-dependently (P<0.05). Conclusions IL-1β significantly upregulated the secretion of MMP-9 but downregulated the secretion of TIMP-3 in human endometrial cells of the mid-secretory phase, resulting in extracellular matrix decomposition to facilitate the invasion by extravillous cytotrophoblasts.