Jung K Y, Uchida S, Endou H
Department of Pharmacology, Faculty of Medicine, University of Tokyo, Japan.
Toxicol Appl Pharmacol. 1989 Sep 15;100(3):369-82. doi: 10.1016/0041-008x(89)90285-8.
To clarify the characteristics of cellular ATP synthesis in individual nephron segments for assessing nephrotoxicity of chemicals, cellular ATP content was measured by the luciferin/luciferase system under various conditions using intact nephron segments isolated from male Sprague-Dawley rats. Increasing the duration of collagenase treatment of kidney slices significantly lowered the cellular levels of ATP newly synthesized from 2 mM glutamine in PST at 37 degrees C over 30 min (p less than 0.01). The tubular incubation time significantly affected the cellular ATP content in the early and middle portions (S2) of the proximal tubule (p less than 0.05 and p less than 0.01, respectively) over 20 min and in the late proximal tubule over 10 min. Among numerous substrates tested, such as D-glucose, glutamine, pyruvate, DL-lactate, and beta-hydroxybutyrate, the substrate utilization for maintaining cellular ATP content was entirely variable according to each nephron segment. Pyruvate and glutamine were the best substrates in the proximal tubule. On the other hand, ATP production from glutamine was less than that from the other substrates in the distally located nephron segments: medullary and cortical thick ascending limbs of Henle's loop (MAL and CAL, respectively), distal tubule, cortical and medullary collecting tubules (CCT and MCT, respectively). In general, glucose, pyruvate, and lactate appear to be equivalent in maintaining ATP content in the distal segments of renal tubules. A monovalent cation ionophore, monensin, at 10 micrograms/ml decreased the cellular ATP content in MAL, CAL, and MCT significantly. Mercuric chloride (HgCl2) was used as a model compound to study nephrotoxicity by investigating its effects on cellular ATP metabolism in microdissected nephron segments. HgCl2 at 1 x 10(-6) M significantly decreased ATP content only in S2 (p less than 0.05), clearly demonstrating S2 to be the most sensitive segment within the nephron. These results indicate that measurement of cellular ATP content would be a useful method forecasting the intrarenal toxic site and potency of possible nephrotoxic chemical compounds.
为阐明单个肾单位节段中细胞ATP合成的特征以评估化学物质的肾毒性，使用从雄性Sprague-Dawley大鼠分离的完整肾单位节段，通过荧光素/荧光素酶系统在各种条件下测量细胞ATP含量。在37℃下，将肾切片的胶原酶处理时间延长，显著降低了PST中由2 mM谷氨酰胺在30分钟内新合成的ATP的细胞水平（p<0.01）。肾小管孵育时间在20分钟内对近端小管的早期和中部（S2）以及在10分钟以上对近端小管晚期的细胞ATP含量有显著影响（分别为p<0.05和p<0.01）。在测试的众多底物中，如D-葡萄糖、谷氨酰胺、丙酮酸、DL-乳酸和β-羟基丁酸，维持细胞ATP含量的底物利用情况因每个肾单位节段而异。丙酮酸和谷氨酰胺是近端小管中最好的底物。另一方面，在位于远端的肾单位节段：髓袢升支粗段和皮质升支粗段（分别为MAL和CAL）、远端小管、皮质和髓质集合小管（分别为CCT和MCT）中，谷氨酰胺产生的ATP少于其他底物。一般来说，葡萄糖、丙酮酸和乳酸在维持肾小管远端节段的ATP含量方面似乎相当。10微克/毫升的单价阳离子离子载体莫能菌素显著降低了MAL、CAL和MCT中的细胞ATP含量。氯化汞（HgCl2）用作模型化合物，通过研究其对显微切割的肾单位节段中细胞ATP代谢的影响来研究肾毒性。1×10−6 M的HgCl2仅显著降低了S2中的ATP含量（p<0.05），清楚地表明S2是肾单位中最敏感的节段。这些结果表明，测量细胞ATP含量将是预测肾内毒性部位和可能的肾毒性化合物效力的有用方法。
