Annals of Clinical Biochemistry, 1978, 15, 313-315

Effect of anticonvulsant drugs on urinary excretion of y-glutamyl transferase in women M. J. LANDONl, D. BATES, MAUREEN KIRKLEY, AND G. VENABLES

From the MRC Reproduction and Growth Research Unit, Princess Mary Maternity Hospital, Newcastle-on-Tyne and the Department of Neurology, Royal Victoria Infirmary, Newcastle-on-Tyne

SUMMARY Excretion of y-glutamyl transferase was found to be significantly higher in a group of epileptic patients on long-term anticonvulsant therapy than in normal subjects. The losses, which may reflect renal enzyme induction, were unrelated to drug dose or duration of therapy and were significantly lower than those found in normal healthy pregnant women at term.

Gamma-glutamyl transferase (y-GT) is a microsomal enzyme which may be involved in the intercellular turnover of glutathione and certain amino acids in a cyclic series of enzymatic events known as the yglutamyl cycle (Orlowski and Meister, 1970a). Estimation of this enzyme in plasma is used as an index of hepatic function when high levels may indicate liver damage. Plasma levels may also be increased by drugs such as anticonvulsants, because of hepatic enzyme induction (Rosalki et al., 1971; Skillen and Pierides, 1976) but whether these compounds have a similar effect on the kidney is not known. The kidneys contain significantly more y-GT than the liver (Orlowski and Szewczuk, 1961; Orlowski and Meister, 1970b) and its concentration in urine samples is increasingly used in the assessment of renal disease, particularly tubular ischaemia and the nephrotoxic effects of antibiotics (Levy and Dubach, 1972; Ward, 1975; Beck et al., 1977). There is evidence that increased excretion of this enzyme occurs during normal healthy pregnancy (Noble et al., 1977), and in the absence of renal disease this may be due to enzyme induction. If this is so it is reasonable to suggest that anticonvulsant drugs might have a similar effect, and the present study was undertaken to investigate this possibility.

history of urinary tract infection, renal disease, or any current condition involving the urogenital tract including menstruation were excluded from the study. A total of 97 samples were collected in dry sterile universal containers and analysed within three hours of collection. Each sample was tested with Labstick (Ames Co Ltd) to exclude proteinuria, glycosuria, and haematuria and then centrifuged at 1000 g for 10 minutes. For assay, 100 iLl of the supernatant was added to 1 m1 substrate (Boehringer Corporation) in a semi-micro cuvette with a 10 nun light path. The rate of reaction at 25°C was recorded for five minutes at 405 nm using a Unicam SP 1800 spectrophotometer. Drug intake was assessed using the scoring system of Richens and Rowe (1970) with no addition made for sodium valproate as there is no evidence yet that this drug has any inducing potential for y-GT (Richens, personal communication). This system was devised to give an idea of enzymeinducing potential but it may not accurately reflect the inducing properties of anyone prescription because of the various combinations of drugs used and because of genetic and acquired differences in drug metabolism. The smoking and drinking habits of the patients were also recorded, although no attempt was made to check the accuracy of these data.

Patients and methods Results Epileptic patients attending a routine neurological outpatient clinic for review were asked to provide a single clean catch MSU specimen. Patients with any

The excretion of y-GT was expressed as mU/mmol creatinine to correct for differences in urine flow rate, and the results are detailed in the Table. In general, the distribution of results was positively 'Present address: Division of Perinatal Medicine, Clinical skewed, and there was extensive overlap in the Research Centre, Watford Road, Harrow, Middlesex HAl various groups studied (Figure). Women taking 3UJ 313

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M. J. Landon, D. Bates, Maureen Kirkley, and G. Venables

314

Table Excretion of y-GT in an epileptic population (mean ± standard deviation) Subject!

n

v-GT excretion (mU/mmol creatinine)

Male epileptics Female epileptics Female normal (non pregnant) Female normal (term pregnancy)

55 42 42" 54"

2550 2656 1968 3210

± 792 ± 643 ± 402 ± 912

Drug scoret 10'3 9'8

± ±

6-7 6·9

Duration of therapy (years) 5·2 4·2

± ±

6-0 4·2

Age (years) 35·6 33-6

± ±

16-0 12-6

"Data from Noble et al. (1977). tAller Richens and Rowe (1970).

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Epileptic male

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n042

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n042

Figure Excretion of y-GT in patients on long-term anticonvulsant therapy compared with normal pregnant women at term and normal non pregnant women: - - ± - - - - - - - - - (mean ± standard deviation) anticonvulsant drugs excreted significantly more y-GT than normal non-pregnant women (t = 5'9; P = 0'(01) but significantly less than normal pregnant women at term (t = 3'49; P = 0'(01). The excretion of y-GT in the epileptic population as a whole was unrelated to smoking, drinking, number of drugs prescribed, length of treatment or the drug score. There was a tendency for y-GT excretion to increase with age in female epileptics (r = 0'33; P = 0'05) but not in males.

in years (Ewen and Griffiths, 1973; Skillen and Pierides, 1976). Although it can be assumed that, in the patients studied, there would be a concomitant rise in plasma y-GT levels, this is unlikely to make any significant contribution to the urinary losses because the molecular weight of this enzyme is of the order of 84000 (Tate and Ross, 1977) and it is thought not to be filtered by the glomerulus. In general, there is no evidence of any relationship between plasma and u~ine levels of this enzyme (Lehmann et al., 1970; Thiel, 1973) and y-GT in the urine is thought to be entirely renal in origin. Whether enzyme induction in the kidney has any functional consequences is not clear. In the rat phenobarbital administration produced a rise in pamino hippurate clearance, reflecting an increased renal plasma flow, but no such changes were apparent in man (Ohnhaus et al., 1977). Other aspects of renal function have not been investigated. The finding that the effect of pregnancy on y-GT excretion exceeds that produced by anticonvulsants is of great interest. It may be that the intense hormonal activity during pregnancy is a more effective stimulator of enzyme synthesis, although, in general, there is no evidence that enzyme induction is doserelated. As with plasma levels of this enzyme, urinary excretion was not related to the amount of drug ingested. Alternatively, it may be that the different isoenzyme forms of y-GT (Rosalki and Rowe, 1973; Patel and O'Gorman, 1973; Azzopardi and Jayle, 1973) vary in their capacity for induction, and their individual response may depend on the nature of the applied stimulus. Whatever its basis it may offer a biochemical explanation for some of the striking physiological changes in renal function that are apparent during pregnancy. We thank Professor D. A. Shaw and Dr N. E. F. Cartlidge for allowing us to study patients under their care.

Discussion It seems unlikely that the kidney should be exempt

from the enzyme-inducing potential of certain drugs, and the present data show that fory-GT at least the kidney may be as susceptible in this respect as the liver. As with plasma y-GT, the effect appears to be unrelated to drug, drug dose, or duration of therapy

References Azzopardi, 0., and Jayle, M. F. (1973). Formes rnoleculaires multiples de Ia garnma-glutarnyl-transpeptidase. Clinica Chimica Acta, 43, 163-169. Beck, P. R., Thomson, R. B., and Chaudhuri, A. K. R.

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Effect of anticonvulsant drugs on urinary excretion of y-glutamyl transferase in women (1977). Aminoglycoside antibiotics and renal function: changes in urinary y-gIutamyl transferase excretion. Journal of Clinical Pathology, 30, 432-437. Ewen, L. M., and Griffiths, J. (1973). Gamma-glutamyl transpeptidase: elevated activities in certain neurological diseases. American Journal of Clinical Pathology, 59, 2-9. Lehmann, D., Prentice, M., and Rosalki, S. B. (1970). Plasma y-glutamyl transpeptidase activity following renal transplantation. Annals ofClinical Biochemistry, 7,148-151. Levy, A., and Dubach, U. C. (1972). Glutamyl transpeptidase-Aktivitlit im Urin bei urorenalen Krankheiten, Klinische Wochenschrift, 50, 438-44 I. Noble, M. C. B., Landon, M. J., and Davison, J. M. (1971). The excretion of y glutamyl transferase in pregnancy. British Journal of Obstetrics and Gynaecology, 84, 522-527. Ohnhaus, E. E., Martin, J., Kinser, J., and Colombo, J. P. (1971). Enzyme induction and renal function in Man. British Journal of Clinical Pharmacology, 4, 33-38. Orlowski, M., and Meister, A. (I 970a). The y-glytamyl cycle: a possible transport system for amino acids. Proceedings of the National Academy of Sciences of the United States of America, 67, 1248-1255. Orlowski, M., and Meister, A. (I 970b). In Methods in Enzymology, 17 part A, edited by S. P. Colwick and N. A. Kaplan, p. 883. Academic Press, New York. Orlowski, M., and Szewczuk, A. (1961). Colorimetric determination of gamma-glutamyl transpeptidase activity in human serum and tissues with synthetic substrates. Acta Biochimica Polonica, 8, 189-200.

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Patel, S., and O'Gorman, P. (1973). Demonstration of yglutamyl transpeptidase isoenzymes using Cellogel electrophoresis. Clinica Chimica Acta, 49, 11-17. Richens, A., and Rowe, D. J. F. (1970). Disturbance of calcium metabolism by anticonvulsant drugs. British Medical Journal, 4, 73-76. Rosalki, S. B. and Rowe, J. A. (1973). Gamma glutamyl transpeptidase activity of human seminal fluid. Lancet, I, 323-324. Rosalki, S. B., Tarlow, D., and Rau, D. (1971). Plasma gamma-glutamyl transpeptidase elevation in patients receiving enzyme inducing drugs. Lancet, 2, 376-371. Skillen, A. W., and Pierides, A. M. (1976). Serum gamma gIutamyl transpeptidase and alkaline phosphatase activities in epileptics receiving anticonvulsant therapy. Clinica Chimica Acta, 72, 245-251. Tate, S. S., and Ross, M. E. (1977). Human kidney yglutamyl transpeptidase, Journal of Biological Chemistry, 252., 6042-6045. Thiel, K. G. (1973). Glutamyl transpeptidase-Aktivitat im Urin bei Gesuden und Nierenkranken, Klinische Wochenschrift, 51, 339-345. Ward, J. P. (1975). Gamma-glutamyl transpeptidase: A sensitive indicator of renal ischaemic injury in experimental animals and renal homograft rejection in man. Annals ofthe Royal Col/ege of Surgeons of England, 57, 248-261. Accepted for publication 18 August 1978

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