Humic Acid Suppresses the LPS-Induced Expression of Cell-Surface Adhesion Proteins through the Inhibition of NF-κB Activation

Abstract

Humic acid (HA), a potential toxin when penetrating the drinking well water of blackfoot disease-endemic areas in Taiwan, has been implicated as one of the etiological factors of this disease. In this study, we investigated the effects of HA on the expression of human vascular endothelial-leukocyte adhesion molecules and the activation of nuclear factor kappa B (NF-κB) in cultured human umbilical vein endothelial cells (HUVECs). The expression of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin was monitored by flow cytometry. Pretreatment of HUVECs with HA inhibited the lipopolysaccharide (LPS)-induced expression of these three adhesion molecules in a dose- and time-dependent manner. Since NF-κB can regulate the expression of these adhesion molecules, NF-κB activation was assessed by electrophoretic mobility shift assay (EMSA). Our results reveal that the activation of NF-κB by LPS is suppressed by HA in a dose- and time-dependent manner. Furthermore, HA reduces NF-κB binding to DNA slightly, but completely inhibits the degradation of IκBα at a concentration of 100 μg/ml. Thus, all our data demonstrate that HA can inhibit the LPS-induced expression of adhesion molecules through the inhibition of NF-κB activation. HA may also suppress the immune or inflammatory reaction of HUVECs responsible for endotoxin, which could be one possible explanation for the causes of the infection and inflammation observed for patients with blackfoot disease. Our results also suggest that immune or inflammatory disturbance occurs for patients with blackfoot disease and that NF-κB may be a critical molecule in the pathogenesis of this disease.