Toll-like receptor 4 signaling in dysfunction of cardiac microvascular endothelial cells under hypoxia/reoxygenation

This study was designed to detect the role of Toll-like receptor 4 (TLR4) signaling in the dysfunction of cardiac microvascular endothelial cells (CMECs) after hypoxia/reoxygenation (H/R).

The cell viability of CMECs was measured by MTT assay. The migration of CMECs was detected by cell scratch wound assay. The expressions of TLR4, nuclear factor-kappa B (NF-κB) and eNOS were analyzed by Western blot. Secretions of nitric oxide (NO) and tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) were determined by NO detection kit and ELISA.

Lipopolysaccharide (LPS) incubation increased the expressions of TLR4, NF-κB, IL-6 and TNF-α in CMECs (P < 0.05 vs. control). The CMECs after H/R injury had impaired cell viability (P < 0.01 vs. control) and migration ability (P < 0.05 vs. control). Moreover, the expressions of TLR4, NF-κB, IL-6 and TNF-α were elevated after H/R in CMECs (P < 0.01 vs. control), while NO and the eNOS expression were significantly decreased. In contrast, administration of the TLR4-neutralizing antibody MTS510 prior to H/R injury down-regulated the expressions of IL-6 and TNF-α and attenuated the dysfunction of CMECs.

TLR4 and its signaling components can be activated by LPS and H/R in CMECs. Blocking the TLR4 signal pathway before H/R injury attenuates CMEC dysfunction.