Implication for long-term hypothermia on degradation of interleukin-8 mRNA in endothelial cells stimulated with lipopolysaccharides.

Abstract

This experimental study investigated the effects of long-term hypothermia on the production of interleukin (IL)-8 protein and its mRNA expression in endothelial cells stimulated by lipopolysaccharides (LPS). Human umbilical vein endothelial cells were separated into a non-cooling group (N group: 37°C) and a cooling group (C group: 30°C). These groups were incubated with LPS (1 μg/mL) for 0, 2, 6, 24, 48, 72, and 96 hours. Production of the IL-8 protein secreted into the supernatant and mRNA expression in the cells were measured using enzyme-linked immunoabsorbent assay (ELISA) and real-time reverse transcription polymerase chain reaction (RT-PCR) analysis. To evaluate mRNA stability, both groups were incubated with actinomycin D at 6 hours after incubation with LPS for 24 hours. The degradation ratio was calculated by comparing the total expression of mRNA at 6 hours versus 0 hours. The protein levels in the C group were significantly lower than the N group between 6 and 96 hours. The mRNA expression in the C group was also significantly lower than in the N group up to 48 hours, but at 72 hours it was significantly higher than N group. IL-8 mRNA was less degraded in the C group compared to the N group. Under long-term hypothermia, IL-8 protein production was suppressed, while IL-8 mRNA was stabilized after LPS treatment. The potential of IL-8 to produce an inflammatory response in endothelial cells may persist even during long-term hypothermia.