Joannidis M, Cantley L G, Spokes K, Stuart-Tilley A K, Alper S L, Epstein F H
Universitätsklinik für Innere Medizin, Innsbruck, Austria.
Kidney Int. 1997 Jul;52(1):130-9. doi: 10.1038/ki.1997.312.
The isolated perfused rat kidney provides a model of selective hypoxia to the medullary thick ascending limb. To investigate the relationship between immediate early gene expression and the extent of hypoxic damage, we determined expression of the immediate early genes (IEG) c-fos and egr-1 in isolated perfused kidneys during standard perfusion and after various measures shown previously to be protective. mRNA levels of c-fos and egr-1 were markedly increased in kidneys after 90 minutes of standard perfusion with Krebs-Henseleit buffer containing albumin. Gene expression was most prominent in the outer medulla followed by papilla and cortex, a pattern reflected by the immunohistochemical demonstration of a prominent accumulation of both egr-1 and c-fos polypetides mainly in the medullary thick ascending limb (mTAL). Protective measures known to minimize morphological damage to the mTAL, including hyperoncotic perfusion, perfusion with glycine, or perfusion with a mixture of amino acids, decreased mRNA levels of c-fos and egr-1 in the outer medulla (by 50% and 35%, respectively) and the papilla (by 60 and 30%, respectively). Renal cortex showed only minor changes. In contrast, prevention of tubular transport by perfusion with 1 mM ouabain increased mRNA levels of c-fos and egr-1 in the outer medulla by 100% and 60%, respectively. Ouabain also dramatically increased mRNA levels of both IEGs in two lines of cultured renal epithelial cells. Changes in the level and distribution of the protein products of these IEGs were not detectable in perfused kidneys by immunohistochemistry. Hypoxic injury of the kidney stimulates IEG expression even in the absence of reperfusion. Protection against hypoxic injury in the mTAL correlates with suppression of IEG mRNA levels when protection is provided by amino acids or hyperoncotic perfusion, but not when provided by inhibition of Na,K-ATPase, which stimulates IEG expression. We conclude that diminished IEG expression is not a necessary concomitant of protection against hypoxic injury.
离体灌注大鼠肾脏为髓袢升支粗段提供了一个选择性缺氧模型。为研究即刻早期基因表达与缺氧损伤程度之间的关系，我们测定了在标准灌注期间以及采用先前显示具有保护作用的各种措施后，离体灌注肾脏中即刻早期基因（IEG）c-fos和egr-1的表达。在用含白蛋白的Krebs-Henseleit缓冲液进行90分钟标准灌注后，肾脏中c-fos和egr-1的mRNA水平显著升高。基因表达在外髓最为显著，其次是乳头和皮质，这种模式通过免疫组织化学显示egr-1和c-fos多肽主要在髓袢升支粗段（mTAL）中大量积累得以体现。已知能使mTAL形态损伤最小化的保护措施，包括高渗灌注、甘氨酸灌注或氨基酸混合物灌注，可降低外髓（分别降低50%和35%）和乳头（分别降低60%和30%）中c-fos和egr-1的mRNA水平。肾皮质仅显示轻微变化。相比之下，用1 mM哇巴因灌注以阻止肾小管转运，可使外髓中c-fos和egr-1的mRNA水平分别升高100%和60%。哇巴因还显著增加了两株培养的肾上皮细胞系中两种IEG的mRNA水平。通过免疫组织化学在灌注肾脏中未检测到这些IEG蛋白质产物水平和分布的变化。即使在没有再灌注的情况下，肾脏的缺氧损伤也会刺激IEG表达。当通过氨基酸或高渗灌注提供保护时，mTAL中针对缺氧损伤的保护与IEG mRNA水平的抑制相关，但当通过抑制Na,K-ATP酶提供保护时则不相关，因为抑制Na,K-ATP酶会刺激IEG表达。我们得出结论，IEG表达的减少并非针对缺氧损伤保护的必然伴随现象。
