Up-regulation interleukin-6 and interleukin-8 by activated protein C in lipopolysaccharide-treated human umbilical vein endothelial cells

Objective： To investigate the effect of activated protein C （APC） on inflammatory responses in human umbilical vein endothelial cells （HUVEC） stimulated with lipopolysaccharide （LPS）. Methods： The second passage of collagenase digested HUVEC was divided into the following groups： serum free medium control group （SFM control）, phosphate buffer solution control group （PBS control）, LPS group with final concentration of 1 μg/ml （LPS group）, APC group with final concentration of 7 μg/ml, Pre-APC group （APC pretreatment for 30 min prior to LPS challenge）, and Post-APC group （APC administration 30 min after LPS challenge）. Supernatant was harvested at 0, 4, 8, 12 and 24 h after LPS challenge. Interleukin-6 （IL-6） and Interleukin-8 （IL-8） levels were analyzed with ELISA. Cells were harvested at 24 h after LPS challenge, and total RNA was extracted. Messenger RNA levels for IL-6 and IL-8 were semi-quantitatively determined by RT-PCR. Results： Compared with control group, IL-6 and IL-8 levels steadily increased 4 to 24 h after LPS stimulation. APC treatment could increase LPS-induced IL-6 and IL-8 production. The mRNA levels of IL-6 and IL-8 exhibited a similar change. Conclusion： APC can further increase the level of IL-6 and IL-8 induced by LPS. The effect of these elevated cytokines is still under investigation.