Implication for Long-Term Hypothermia on Degradation of Interleukin-8 mRNA in Endothelial Cells Stimulated with Lipopolysaccharides
Implication for Long-Term Hypothermia on Degradation of Interleukin-8 mRNA in Endothelial Cells Stimulated with Lipopolysaccharides
复制标题
DOI：
10.1089/ther.2012.0010
复制
发表时间：
2012-06-01
影响因子：
1.2
通讯作者：
Tanjoh, Katsuhisa
中科院分区：
文献类型：
--
作者：
Sakurai, Atsushi;Kinoshita, Kosaku;Tanjoh, Katsuhisa
本实验研究了长期低温对内皮细胞产生IL-8蛋白及其mRNA表达的影响。将人脐静脉内皮细胞分为未降温组(N组：37℃)和降温组(C组：30℃)。各组分别与1mUg/mLLPS孵育0、2、6、24、48、72、96h。采用酶联免疫吸附试验(EL ISA)和实时定量逆转录聚合酶链式反应(RT-PCR)检测细胞培养上清液中IL-8蛋白的产生和细胞内IL-8基因的表达。为评价mRNA的稳定性，两组均在与内毒素孵育24小时后6小时加入放线菌素D孵育。通过比较6h和0h的总mRNA表达来计算降解率。C组在6~96h的蛋白水平明显低于N组。C组的mRNA表达在48h也明显低于N组，但在72h时显著高于N组。与N组相比，C组IL-8mRNA降解程度较轻。在长期低温下，IL-8蛋白的产生受到抑制，而IL-8mRNA的表达在内毒素处理后趋于稳定。即使在长期低温期间，IL-8在内皮细胞中产生炎症反应的潜力也可能持续存在。
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 degrees C) and a cooling group (C group: 30 degrees C). These groups were incubated with LPS (1 mu g/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.