Glutamine's protection against cellular injury is dependent on heat shock factor-1
Glutamine's protection against cellular injury is dependent on heat shock factor-1
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DOI：
10.1152/ajpcell.00635.2005
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发表时间：
2006-06-01
影响因子：
5.5
通讯作者：
Wischmeyer, PE
中科院分区：
文献类型：
--
作者：
Morrison, AL;Dinges, M;Wischmeyer, PE
谷氨酰胺（GLN）已被证明可以保护细胞、组织和整个生物体免受压力和伤害。热休克蛋白（HSP）的表达增强被认为是这种保护的原因。到目前为止，还没有明确的机制数据证实这种关系。本研究验证了gln通过热休克因子-1 （HSF-1）介导的HSP通路激活负责细胞保护的假设。所有实验均采用野生型HSF-1（+/+）和敲除型（-/-）小鼠成纤维细胞。在同时处理模型中，用浓度为0 ~ 16 mM的GLN处理细胞，并暴露于热应激损伤中。采用芬那嗪加四氮唑盐、热休克蛋白70、热休克蛋白25检测细胞活力，Western blot检测细胞核HSF-1表达，EMSA检测HSF-1/热休克元件（HSE）结合。GLN以剂量依赖性方式显著减弱热应激诱导的HSF-1（+/+）细胞死亡；然而，GLN在HSF-1（-/-）细胞中失去了生存益处。GLN导致热应激后HSP-70和HSP-25表达呈剂量依赖性增加。在HSF-1（-/-）细胞中未观察到可诱导的HSP表达。在热应激前，GLN增加了细胞核中未磷酸化的HSF-1。这伴随着热应激后gln介导的HSF-1/HSE结合和磷酸化HSF-1核含量的增加。这是首次证明热应激损伤后gln介导的细胞保护与HSF-1表达和激活热休克反应的细胞能力有关。此外，gln介导的抗损伤保护机制似乎涉及应激前核HSF-1含量的增加和HSF-1启动子结合和磷酸化的增加。
Glutamine (GLN) has been shown to protect cells, tissues, and whole organisms from stress and injury. Enhanced expression of heat shock protein (HSP) has been hypothesized to be responsible for this protection. To date, there are no clear mechanistic data confirming this relationship. This study tested the hypothesis that GLN-mediated activation of the HSP pathway via heat shock factor-1 (HSF-1) is responsible for cellular protection. Wild-type HSF-1 (HSF-1(+/+)) and knockout (HSF-1(-/-)) mouse fibroblasts were used in all experiments. Cells were treated with GLN concentrations ranging from 0 to 16 mM and exposed to heat stress injury in a concurrent treatment model. Cell viability was assayed with phenazine methosulfate plus tetrazolium salt, HSP-70, HSP-25, and nuclear HSF-1 expression via Western blot analysis, and HSF-1/heat shock element (HSE) binding via EMSA. GLN significantly attenuated heat-stress induced cell death in HSF-1(+/+) cells in a dose-dependent manner; however, the survival benefit of GLN was lost in HSF-1(-/-) cells. GLN led to a dose-dependent increase in HSP-70 and HSP-25 expression after heat stress. No inducible HSP expression was observed in HSF-1(-/-) cells. GLN increased unphosphorylated HSF-1 in the nucleus before heat stress. This was accompanied by a GLN-mediated increase in HSF-1/HSE binding and nuclear content of phosphorylated HSF-1 after heat stress. This is the first demonstration that GLN-mediated cellular protection after heat-stress injury is related to HSF-1 expression and cellular capacity to activate an HSP response. Furthermore, the mechanism of GLN-mediated protection against injury appears to involve an increase in nuclear HSF-1 content before stress and increased HSF-1 promoter binding and phosphorylation.