CARBAMYL-PHOSPHATE SYNTHETASE AND ORNITHINE TRANSCARBAMYLASE ACTIVITIES IN ENZYME-DEFICIENT HUMAN-LIVER MEASURED BY RADIOCHROMATOGRAPHY AND CORRELATED WITH OUTCOME
CARBAMYL-PHOSPHATE SYNTHETASE AND ORNITHINE TRANSCARBAMYLASE ACTIVITIES IN ENZYME-DEFICIENT HUMAN-LIVER MEASURED BY RADIOCHROMATOGRAPHY AND CORRELATED WITH OUTCOME
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DOI：
10.1203/00006450-198907000-00021
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发表时间：
1989-07-01
影响因子：
3.6
通讯作者：
BRUSILOW, SW
中科院分区：
文献类型：
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作者：
TUCHMAN, M;TSAI, MY;BRUSILOW, SW
建立了测定氨基甲酰磷酸合成酶I(CPS I)和鸟氨酸转氨甲基酶(OTC)活性的灵敏、特异的放射层析方法。对23例CPS I(5例)和OTC缺乏症(18例)引起的高氨血症患者的冰冻肝组织中这些酶的活性进行了测定。此外，还对1例OTC缺乏症流产胎儿和4例正常人的肝脏进行了研究。这些分析使用放射性鸟氨酸作为底物，然后用高效液相色谱法分离反应中形成的瓜氨酸，并用放射性流动监测仪或闪烁计数器对这两种氨基酸中的放射性进行定量。CPS I法和OTC法均与孵育时间和组织匀浆浓度呈线性关系。该方法的灵敏度可使5毫克肝组织的CPS I和OTC活性低至0.1微克/克/分，CPS I或OTC缺乏症的诊断可分别低至0.5和0.05毫克组织。4例正常肝脏不同切片的CPS I活性为3.01±-。0.16微克/分(平均值+-.扫描电子显微镜，n=19)，OTC活性为93.4+-。6.3(平均值+-扫描电子显微镜，n=19)。在5例CPS I缺乏症患者中有1例和19例OTC缺乏症患者中有14例肝组织中可检测到残留酶活性。正常肝组织OTC/CPS I活性比值为31.2±-。1.3(平均值+-扫描电子显微镜，n=19)，而在CPS I缺乏的肝脏中，这一比率在343~5000之间，在OTC缺乏的肝脏中，这一比率在~lt；0.02~1.55之间。OTC/CPS I比值被发现是评估酶缺乏的可靠参数，以补偿观察到的个体样本之间的酶活性差异。OTC/CPS I比值似乎也与酶缺乏患者的病情严重程度有关。残留酶活性最高(正常的1-3%)的OTC缺陷患者预后较好。这些灵敏的方法将有助于在只有非常少量的肝组织可用时建立CPS I或OTC缺乏症的诊断，并且他们的结果可以被解释，即使这些酶的活性因采集或储存伪影而发生继发性降低。残留酶活性水平可能有助于预测这些患者的预后。
Sensitive and specific radiochromatographic methods to measure enzymatic activities of carbamyl phosphate synthetase I (CPS I) and ornithine transcarbamylase (OTC) were developed. The activities of these enzymes were assayed in frozen liver tissue obtained from 23 individuals with hyperammonemia caused by CPS I (five patients) and OTC deficiency (18 patients). In addition, livers of one aborted fetus with OTC deficiency and four normal individuals were studied. The assays use radioactive ornithine as a substrate followed by separation of citrulline formed in the reactions by HPLC and quantitation of the radioactivity in both aminoacids by a radioactivity flow monitor or by a scintillation counter. Both CPS I and OTC assays were linear with respect to length of incubation time and concentration of tissue homogenate. The sensitivity of the methods allowed measurements of CPS I and OTC activities as low as 0.1 .mu.mol/g/min on 5 mg of liver tissue and the diagnosis of CPS I or OTC deficiency could be established on as low as 0.5 and 0.05 mg of tissue, respectively. CPS I activity in different sections of four normal livers was 3.01 .+-. 0.16 .mu.mol/g/min (mean .+-. SEM, n = 19) and OTC activity was 93.4 .+-. 6.3 (mean .+-. SEM, n = 19). Residual enzymatic activity could be detected and measured in the liver tissues of one of the five subjects with CPS I deficiency and in 14 of 19 subjects with OTC deficiency. OTC/CPS I activity ratio in normal liver tissue was 31.2 .+-. 1.3 (mean .+-. SEM, n = 19), whereas this ratio ranged from 343 to > 5000 in CPS I deficient livers and from < 0.02 to 1.55 in OTC deficient livers. The OTC/CPS I ratio was found to be a reliable parameter for assessing enzymatic deficiency compensating for the observed variability of enzymatic activity between individual samples. OTC/CPS I ratio also seemed to correlate with the severity of disease in enzyme deficient patients. OTC-deficient patients with the highest residual enzymatic activity (1-3% of normal) had a better outcome. These sensitive methods would help to establish the diagnosis of CPS I or OTC deficiency whenever only a very small amount of liver tissue is available and their results could be interpreted even if secondary reduction in the activities of these enzymes occurred due to collection or storage artifacts. Levels of residual enzymatic activities may help to predict the prognosis of these patients.