Baricos W H, Shah S V
Biochem J. 1984 Oct 15;223(2):393-9. doi: 10.1042/bj2230393.
We have examined the activity and distribution of cathepsin D (EC 3.4.23.5), a major renal lysosomal endoproteinase, in the various anatomical and functional areas of normal rat kidney. Cathepsin D-like activities (delta A280/h per mg of protein) in normal rat tissues were: cortex, 0.78 +/- 0.05, n = 37; medulla, 0.62 +/- 0.03, n = 12; papilla, 0.63 +/- 0.04, n = 12; tubules, 0.74 +/- 0.04, n = 28; glomeruli, 0.59 +/- 0.03, n = 28; and liver, 0.41 +/- 0.02, n = 28. Enzyme activity was maximal at pH 3.0-3.5 and inhibited more than 90% by pepstatin (6.7 micrograms/ml), suggesting that the enzyme is cathepsin D. In subsequent experiments we measured cathepsin D-like activity in cortex, tubules and glomeruli isolated from rats with puromycin aminonucleoside (PAN)-induced nephrotic syndrome. Treated animals (15 mg of PAN/100g body wt., intraperitoneally) developed proteinuria beginning 4 days after injection and exceeding 900 mg/24h on day 9. In two separate experiments involving 52 animals we observed a significant increase in cathepsin D-like activity in cortex (+82.7%), tubules (+109.6%) and glomeruli (+54.7%) isolated from PAN-treated rats killed during marked proteinuria (day 9, mean total urinary protein excretion: 937 +/- 94 mg/24h). This increase was observed whether the activity was expressed per mg of DNA or per mg of protein. Increased cathepsin D-like activity was first observed in cortex and tubules coincident with the onset of proteinurea (day 4, mean total urinary protein excretion: 112 +/- 23 mg/24h). In contrast with the significant elevation of renal cathepsin D-like activity, the activity (nmol/h per mg of protein) of alpha-L-fucosidase (EC 3.2.1.51), a non-proteolytic enzyme, was markedly decreased in the identical samples used for the measurement of cathepsin D-like activity: cortex (-46.4%); tubules (-46.1%); and glomeruli (-38.5%). In addition to changes in renal enzyme activities, PAN-treated rats excreted large amounts of cathepsin D-like activity in their urine (beginning on day 3) compared with nearly undetectable cathepsin D-like activity in the urine from control rats. The significant increases in glomerular and tubular cathepsin D activity may reflect an important role for this enzyme in the pathophysiology associated with PAN-induced nephrotic syndrome.
我们研究了组织蛋白酶D（EC 3.4.23.5）的活性和分布，它是一种主要的肾脏溶酶体内切蛋白酶，存在于正常大鼠肾脏的各个解剖和功能区域。正常大鼠组织中类组织蛋白酶D的活性（每毫克蛋白质每分钟A280的变化量）分别为：皮质，0.78±0.05，n = 37；髓质，0.62±0.03，n = 12；乳头，0.63±0.04，n = 12；肾小管，0.74±0.04，n = 28；肾小球，0.59±0.03，n = 28；肝脏，0.41±0.02，n = 28。酶活性在pH 3.0 - 3.5时最高，并且被胃蛋白酶抑制剂（6.7微克/毫升）抑制超过90%，这表明该酶是组织蛋白酶D。在随后的实验中，我们测量了从嘌呤霉素氨基核苷（PAN）诱导的肾病综合征大鼠分离出的皮质、肾小管和肾小球中的类组织蛋白酶D活性。处理过的动物（15毫克PAN/100克体重，腹腔注射）在注射后4天开始出现蛋白尿，在第9天超过900毫克/24小时。在涉及52只动物的两个独立实验中，我们观察到在显著蛋白尿期（第9天，平均总尿蛋白排泄量：937±94毫克/24小时）处死的PAN处理大鼠分离出的皮质（+82.7%）、肾小管（+109.6%）和肾小球（+54.7%）中，类组织蛋白酶D活性显著增加。无论活性是以每毫克DNA还是每毫克蛋白质来表示，这种增加都能被观察到。类组织蛋白酶D活性的增加首先在皮质和肾小管中观察到，与蛋白尿的开始（第4天，平均总尿蛋白排泄量：112±23毫克/24小时）同时出现。与肾脏类组织蛋白酶D活性的显著升高形成对比的是，α-L-岩藻糖苷酶（EC 3.2.1.51），一种非蛋白水解酶，在用于测量类组织蛋白酶D活性的相同样本中的活性（每毫克蛋白质每小时nmol数）显著降低：皮质（-46.4%）；肾小管（-46.1%）；肾小球（-38.5%）。除了肾脏酶活性的变化外，与对照大鼠尿液中几乎检测不到的类组织蛋白酶D活性相比，PAN处理的大鼠在尿液中排泄大量的类组织蛋白酶D活性（从第3天开始）。肾小球和肾小管组织蛋白酶D活性的显著增加可能反映了该酶在与PAN诱导的肾病综合征相关的病理生理学中的重要作用。
