Rinaldo J E, Gorry M, Strieter R, Cowan H, Abdolrasulnia R, Shepherd V
Department of Veterans Affairs Medical Center, Nashville, Tennessee.
Am J Respir Cell Mol Biol. 1990 Sep;3(3):207-16. doi: 10.1165/ajrcmb/3.3.207.
Heat shock proteins (HSPs) have been remarkably conserved throughout evolution. It has been assumed that induction of HSPs remains a stereotypic response to injury, important for survival of eukaryotic cells during euthermic injury. However, there are few studies of this phenomenon in endothelial cells, and none in pulmonary endothelial cells. We studied the induction of synthesis of 70-kD proteins in bovine pulmonary artery endothelial cells (BPAECs) in response to heat shock and to euthermic injury induced by bacterial endotoxin. First, in response to heat, BPAECs showed rapid and reversible heat-induced synthesis of 70-kD proteins, readily detectable by one-dimensional SDS-PAGE of [35S]methionine-labeled BPAECs. Heat shock at 42 degrees C for 3 h or 43 degrees C for 2 h suppressed total protein synthesis by 30% (P less than 0.001) but an increased rate of synthesis of 70-kD protein continued, representing an increasing fraction of total protein synthesis. Heat-induced synthesis of 70-kD protein returned to baseline levels 8 h after heat shock. Northern analysis showed that mRNA for a protein homologous to a conserved amino acid sequence in the family of species-homologous 70-kD heat shock proteins (HSP 70) was induced by a 15-min incubation at 42 degrees C and remained detectably increased for 6 h. We next assessed whether euthermic injury by bacterial endotoxin (LPS) generated a similar response. LPS was cytotoxic by BPAECs as assessed morphologically, by release of 51Cr from prelabeled cells, and by a significant suppression of total protein synthesis (range, 35 to 70%; P less than 0.001). Despite cytotoxicity, LPS did not induce 70-kD protein at a level that could be detected by SDS-PAGE, and no increase in mRNA for HSP 70 was detected by Northern analysis. LPS-injured BPAECs remained "competent" to induce both 70-kD proteins and mRNA for HSP 70 in response to heat shock. We conclude that at least quantitatively, induction of HSP 70 by BPAECs is not a stereotypic response to injury but rather is at least relatively injury-specific. However, competence to induce HSP 70 appears to be extremely resilient: it is retained in dysfunctional BPAECs in the face of profound inhibition of global protein synthesis, suggesting an important homeostatic role.
热休克蛋白（HSPs）在整个进化过程中都得到了显著的保守。人们认为，热休克蛋白的诱导仍然是对损伤的一种刻板反应，这对于真核细胞在体温正常时受到损伤期间的存活很重要。然而，关于内皮细胞中这种现象的研究很少，而在肺内皮细胞中则没有相关研究。我们研究了牛肺动脉内皮细胞（BPAECs）中70-kD蛋白合成的诱导情况，以应对热休克以及细菌内毒素诱导的体温正常时的损伤。首先，在热刺激下，BPAECs显示出快速且可逆的热诱导70-kD蛋白合成，通过对[35S]甲硫氨酸标记的BPAECs进行一维SDS-PAGE很容易检测到。42℃热休克3小时或43℃热休克2小时可使总蛋白合成抑制30%（P<0.001），但70-kD蛋白的合成速率仍持续增加，占总蛋白合成的比例越来越大。热诱导的70-kD蛋白合成在热休克8小时后恢复到基线水平。Northern分析表明，与物种同源的70-kD热休克蛋白（HSP 70）家族中保守氨基酸序列同源的一种蛋白的mRNA，在42℃孵育15分钟后被诱导，并且在6小时内仍可检测到增加。接下来，我们评估细菌内毒素（LPS）引起的体温正常时的损伤是否产生类似的反应。从形态学、预标记细胞中51Cr的释放以及总蛋白合成的显著抑制（范围为35%至70%；P<0.001）评估，LPS对BPAECs具有细胞毒性。尽管具有细胞毒性，但LPS并未诱导出可通过SDS-PAGE检测到水平的70-kD蛋白，Northern分析也未检测到HSP 70的mRNA增加。LPS损伤的BPAECs在热休克时仍“有能力”诱导70-kD蛋白和HSP 70的mRNA。我们得出结论，至少在数量上，BPAECs对HSP 70的诱导不是对损伤的刻板反应，而是至少相对具有损伤特异性。然而，诱导HSP 70的能力似乎极具弹性：在整体蛋白合成受到严重抑制的情况下，功能失调的BPAECs仍保留这种能力，这表明其具有重要的稳态作用。
