Turman M A, Kahn D A, Rosenfeld S L, Apple C A, Bates C M
Department of Pediatrics, Ohio State University, Columbus 43205, USA.
Biochem Mol Med. 1997 Feb;60(1):49-58. doi: 10.1006/bmme.1996.2556.
In animal models of cardiac or cerebral ischemic preconditioning, induction of heat shock proteins (HSPs), especially HSP70, correlates with protection from subsequent injury. The extent of HSP70 induction after stress correlates inversely with initial HSP70 levels. Primate cells, unlike nonprimate cells, express high basal levels of HSP70; thus, primate cells may respond differently to preconditioning than nonprimate cells. We have demonstrated that exposing cultured human proximal tubular epithelial cells (PTEC) to 12 h of hypoxia followed by a 24-h recovery period (hypoxic preconditioning) induces resistance to subsequent hypoxic injury. Herein, we characterize the expression of HSP70, HSP90, and heat shock cognate-70 (HSC70) in PTEC under basal conditions and after hypoxic preconditioning. By Northern blot analysis, we demonstrate that hypoxic preconditioning of PTEC increases mRNA for HSP70 > HSP90 > HSC70. With reverse transcription and polymerase chain reaction, mRNA transcripts from three different HSP70 genes (HSP70 A, B, and C) were detected in unstressed PTEC. Transcripts from these genes were also detected in freshly isolated human renal cortex, indicating that all three genes are expressed in vivo. By Western blot analysis, we demonstrate that PTEC express high basal levels of HSP70, HSC70, and HSP90. Hypoxic preconditioning did not lead to a significant increase in protein content of any of these HSPs, despite increased mRNA levels. This suggests that HSP accumulation cannot account for the development of cytoresistance after hypoxic preconditioning in PTEC. However, high basal expression of HSP70 in human PTEC may contribute to their innate resistance for hypoxia.
在心脏或脑缺血预处理的动物模型中，热休克蛋白（HSPs）尤其是HSP70的诱导与对随后损伤的保护作用相关。应激后HSP70的诱导程度与初始HSP70水平呈负相关。与非灵长类细胞不同，灵长类细胞表达高水平的基础HSP70；因此，灵长类细胞对预处理的反应可能与非灵长类细胞不同。我们已经证明，将培养的人近端肾小管上皮细胞（PTEC）暴露于12小时的缺氧状态，随后进行24小时的恢复期（缺氧预处理），可诱导其对随后的缺氧损伤产生抗性。在此，我们描述了基础条件下和缺氧预处理后PTEC中HSP70、HSP90和热休克同源蛋白-70（HSC70）的表达情况。通过Northern印迹分析，我们证明PTEC的缺氧预处理使HSP70 > HSP90 > HSC70的mRNA增加。通过逆转录和聚合酶链反应，在未受应激的PTEC中检测到来自三个不同HSP70基因（HSP70 A、B和C）的mRNA转录本。在新鲜分离的人肾皮质中也检测到了这些基因的转录本，表明这三个基因在体内均有表达。通过Western印迹分析，我们证明PTEC表达高水平的基础HSP70、HSC70和HSP90。尽管mRNA水平增加，但缺氧预处理并未导致这些HSPs中任何一种的蛋白质含量显著增加。这表明HSP的积累不能解释PTEC缺氧预处理后细胞抗性的产生。然而，人PTEC中HSP70的高基础表达可能有助于其对缺氧的固有抗性。
