Sanlorenzo Lauren, Zhao Bin, Spight Donn, Denenberg Alvin G, Page Kristen, Wong Hector R, Shanley Thomas P
Division of Critical Care Medicine, Children's Hospital Medical Center, Cincinnati, OH, USA.
Crit Care Med. 2004 Nov;32(11):2284-92. doi: 10.1097/01.ccm.0000145580.96994.c9.
Application of heat shock before an inflammatory stimulus often results in an attenuated response to that stimulus. As a result, it has become increasingly appreciated that heat shock may induce cross-tolerance to a variety of stimuli based on in vitro and in vivo models. Circulating peripheral blood monocytes are key mediators of cytokine release following endotoxin challenge. The mitogen-activated protein kinases play a key role in the transcriptional regulation of this response including expression of tumor necrosis factor. As such, counterregulatory phosphatases that target mitogen-activated protein kinase may play a role in this heat shock-mediated effect. We hypothesized that prior heat shock to monocytes would induce a phosphatase, MKP-1, that regulated mitogen-activated protein kinase activity and subsequently conferred cross-tolerance to lipopolysaccharide stimulation.
Experimental.
University research foundation laboratory.
THP-1 human monocyte cell line.
THP-1 cells were exposed to either heat shock (43 degrees C, 1 hr) or normothermia (37 degrees C, 1 hr) and allowed to recover before stimulation with endotoxin (lipopolysaccharide).
Induction of a heat shock response was determined by heat shock protein-70 expression. Tumor necrosis factor and interleukin-10 were measured by enzyme-linked immunosorbent assay to assess heat shock inhibition of lipopolysaccharide-induced gene expression. The effect of heat shock on lipopolysaccharide-mediated activation of the p38 and ERK kinases was examined by measuring phospho-specific isoforms of p38 and ERK1/2 and correlated to in vitro kinase activity. Confirmatory data were generated from experiments employing either pharmacologic inhibition or genetic deletion of MKP-1. Heat shock induced the nuclear localized phosphatase, MKP-1, that attenuated p38 and ERK kinase activity resulting in significantly diminished tumor necrosis factor expression in response to lipopolysaccharide.
The effect of heat shock on decreasing the tumor necrosis factor response to lipopolysaccharide is conferred by induction of MKP-1, which negatively regulates p38 and ERK kinases. Modulation of phosphatase activity may be a potential strategy for attenuating acute inflammatory responses.
在炎性刺激之前应用热休克通常会导致对该刺激的反应减弱。因此，基于体外和体内模型，热休克可能诱导对多种刺激的交叉耐受这一点已得到越来越多的认识。循环外周血单核细胞是内毒素攻击后细胞因子释放的关键介质。丝裂原活化蛋白激酶在这种反应的转录调控中起关键作用，包括肿瘤坏死因子的表达。因此，靶向丝裂原活化蛋白激酶的反调节磷酸酶可能在这种热休克介导的效应中起作用。我们假设，对单核细胞预先进行热休克会诱导一种磷酸酶MKP-1，它调节丝裂原活化蛋白激酶的活性，并随后赋予对脂多糖刺激的交叉耐受。
实验性研究。
大学研究基金会实验室。
THP-1人单核细胞系。
将THP-1细胞暴露于热休克（43℃，1小时）或正常体温（37℃，1小时），并在脂多糖刺激前恢复。
通过热休克蛋白-70表达来确定热休克反应的诱导情况。通过酶联免疫吸附测定法测量肿瘤坏死因子和白细胞介素-10，以评估热休克对脂多糖诱导的基因表达的抑制作用。通过测量p38和ERK1/2的磷酸化特异性异构体来检测热休克对脂多糖介导的p38和ERK激酶激活的影响，并与体外激酶活性相关联。采用MKP-1的药理学抑制或基因缺失的实验产生了确证数据。热休克诱导了核定位的磷酸酶MKP-1，它减弱了p38和ERK激酶的活性，导致对脂多糖反应时肿瘤坏死因子的表达显著减少。
热休克对降低肿瘤坏死因子对脂多糖反应的作用是由MKP-1的诱导介导的，MKP-1对p38和ERK激酶起负调节作用。调节磷酸酶活性可能是减轻急性炎症反应的一种潜在策略。
