Abstract: AIM:To detect the alteration of Toll-like receptor 4 (TLR-4) pathway expression in cardiac microvascular endothelial cells (CMECs) during hypoxia-reoxygenation (H-R) injury. METHODS: CMECs isolated from the hearts of adult rats were exposed to hypoxia (94% N2, 5% CO2, 1% O2) for 6 h followed by 2, 12 or 24 h reoxygenation (95% air, 5% CO2). Proliferation of CMECs was assessed by MTT assay, TLR-4 and NF-κB expressions were analyzed by Western blot and levels of IL-6 and TNF-α were detected by ELISA kits. RESULTS: The proliferation ability of CMECs was significantly inhibited by H-R injury (P<0.01) and H-R injury increased the TLR-4 expression 2 and 12 h after reoxygenation (P<0.05). The level of NF-κB increased 2 and 24 h after reoxygenation (P<0.05) and H-R injury enhanced IL-6 and TNF-α secretion compared with those in control groups (P<0.05). CONCLUSION: H-R injury increases the TLR-4 and NF-κB expressions in CMECs and enhances secretion of IL-6 and TNF-α. Activation of TLR-4/NF-κB pathway may participate in H-R injury of CMECs.