Primary oxaluria type 2 (l -glyceric aciduria): A rare cause of nephrolithiasis in children

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The Journal of Pediatrics June 1991

Primary oxaluria type 2 (L-glyceric aciduria): A rare cause of nephrolithiasis in children L. E. S e a r g e a n t , PhD, G. W. d e G r o o t , a n d J. C. H a w o r t h , MD

MD, L. A, Dilling, ART, C. J. M a l l o r y , BSc,

From the Department of Pediatrics and Child Health and the Department of Biochemistry and Molecular Biology, University of Manitoba, and the Children's Hospital, Winnipeg, Manitoba, Canada

L-Glyceric aciduria (primary hyperoxaluria type 2; McKusick 26000) is a rare disorder characterized by excessive urinary excretion of t-glycerate and oxalate. Clinical symptoms are due to renal stone formation caused by the hyperoxaluria. Deficiency of D-glycerate dehydrogenase activity (EC 1.1.1.29) hasbeen demonstrated in leukocytes from five of the eight recorded patients.I, 2 In addition, a deficiency of D-glycerate dehydrogenase and glyoxylate reductase (EC 1.1.1.26/79) has been demonstrated in liver tissue in one patient. 3 D-Glycerate dehydrogenase also has glyoxylate reductase activity,4, 5 and it has been proposed that the deficiency of glyoxylate reductase activity is responsible for the hyperoxaluria in this disorder. 1, 6 We present data that confirm the combined deficiencies of D-glycerate dehydrogenase and glyoxylate reductase activities in this disorder and report on eight additional patients. They belong to three Saulteaux-Ojibway Canadian Indian families living in two isolated communities in northwestern Ontario. METHODS Deuterated L-glyeeric acid was prepared by treating L-2,3,3-d3-serine (Merck Sharp & Dohme, Montreal, Canada) with nitrous acid as described.7 The product was used without further purification. Urine samples were spiked with d3-glyeeric acid and extracted with ethylacetate and ether as described, 8 and the dried residues reacted with Nmethyl-N- (tert-butyldimethylsilyl)trifluoroacetamide (Pierce Chemical Co., Rockford, Ill.) for 20 minutes at 70 ~ C. Glyceric acid was identified and quantitated by Stable isotope dilution with a Hewlett-Packard model HP 5993C gas chromatography-mass spectrometry system equipped with a splitless capillary injection port in the electron-impact mode. The analytic column, a 30 X 0.25 mm (internal Submitted for publication Nov. 19, 1990; accepted Jan. 16, 1991. Reprint requests: L. E. Seargeant, PhD, Department of Clinical Chemistry, Health Sciences Centre, 820 Sherbrook St., Winnipeg, Manitoba R3A 1R9, Canada.

9/22/28095

diameter) model DB-1701 fused silica column ( J & W Scientific, Folsom, Calif.), was run from 40 ~ to 290 ~ C at a rate of 10 ~ C/min. Quantitative values were determined from standard curves with mass fragments of 391 and 394 used for the nondeuterated and deuterated compounds, respectively. Oxalate concentration was measured enzymatically with a commercially available kit from Sigma Chemical Co. (St. Louis, Mo.). 9 D-Glycerate dehydrogenase activity was measured in a liver biopsy sample obtained from patient 1 with informed parental consent at the time of surgery. The sample was homogenized and dialyzed overnight as described by Mistry et al. 3 D-Glycerate dehydrogenase activity was measured spectrophotometrically by monitoring the reduction of hydroxypyruvate in the presence of the reduced form of nicotinamide-adenine dinucleotide phosphate (NADPH). Glyoxylate reductase activity was measured with glyoxylate and N A D P H used as substrates. 3 The detection limit for both assays was 2 gmol/min/gm protein. RESULTS Clinical and laboratory findings in primary hyperoxaluria type 2 are summarized in the Table. Urinary excretion of glyceric acid was increased 20- to 500-fold relative to creatinine excretion. Urine glycerate was identified as the L-isomer on the basis of its reaction with L-lactate dehydrogenase after isolation by column chromatography] Urine oxalate excretion was increased 1.5- to 10-fold. D-Glyceratedehydrogenase activity and glyoxylate reductase activity were undetectable in liver homogenates from patient 1. D-Glycerate dehydrogenase activity and glyoxylate reductase activity in three control liver samples (obtained post mortem) ranged from 76 to 90 #mol/min/ gm protein and 114 to 141/zmol/min/gm protein, respectively. DISCUSSION Our findings are in agreement with those of the single previous report that primary hyperoxaluria type 2 is associated with deficiencies of D-glycerate dehydrogen~ise ac-

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Clinical and laboratory observations

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Table. Clinical and laboratory findings in primary hyperoxaluria type 2 Patient No.

Age at diagnosis

Gender

Clinical signs and findings

1

2 yr

M

2

8 yr

F

3

15 yr

M

4 5

20 yr 6 mo

M F

6 7 8

6 yr 6 mo 18 yr

M M M

History of dysuria (2 mo) and intermittent hematuria; bladder calculus (2 • 3 cm) removed surgically Sibling of patient 1; recurrent, refractory urinary tract infections since 12 mo of age; bladder calculus (11 gm) removed surgically at age 35 too; no further symptoms First cousin once removed of patients 1 and 2; 3 small renal calculi detected at 18 mo; no urinary tract symptoms present; calculi no longer evident after conservative treatment and did not recur; at age 14-15 yr had intermittent abdominal pain, dysuria, and microscopic hematuria but no renal calculi Sibling of patient 3; symptom free No apparent relation to patients 1-4 but lives in same community; symptom free Sibling of patient 5; symptom free Sibling of patients 5 and 6; symptom free No relation to other patients; urethral calculus at 7 yr; multiple renal and bladder calculi at 12 yr followed by recurrent urinary tract infections; ureterolithotomy at 14 yr; bilateral nephrolithiasis on radiographic examination at 18 yr; had cerebral palsy and mental retardation from birth, probably unrelated to hyperoxaluria

Reference range

Glyceric acid (umol/mmol Cr)

Oxalic acid (/~mol/mmol Cr)

6,908

739

2,196

522

1,436

280

822 6,747

115 381

1,758 15,202 671

342 266 700

<28

< 190* <75"~

Cr, Creatinine. *From 6 months to 7 years of age. rAfter 7 years of age.

tivity and glyoxylate reductase activity. Both of these enzyme activities appear to be attributable to a single enzyme. 46 Deficiency of D-glycerate dehydrogenase activity presumably causes accumulation of its substrate, hydroxypyruvate, which is then converted to L-glycerate by the action of L-lactate dehydrogenase. Deficiency of glyoxylate reductase presumably causes impaired conversion of glyoxylate to glycolate. Conversion of glyoxylate to oxalate by L-lactate dehydrogenase would explain the observed hyperoxaluria. Hyperoxaluria type 2 (L-glyceric aciduria) can be distinguished from primary hyperoxaluria type 1, which is caused by deficiency of the hepatic peroxisomal enzyme alanine glyoxylate aminotransferase, by the finding of greatly increased t-glyceric acid excretion in the urine with normal glycolate excretion. The main clinical manifestation of both types is calcium oxalate nephrolithiasis. In type 1 the age of onset may be variable, depending on the amount of residual enzyme activity: In the more severe form, with onset

in early childhood, there frequently is progressive renal insufficiency, and death often occurs before 20 years of age. 10 Seven of the eight patients previously reported with type 2 hyperoxaluria initially had renal calculi between 18 months and 24 years of age. l, 2 One patient seems to have had no symptoms and was identified only because his younger brother also had the disorder. 2 One patient required a nephrectomy, but the others apparently had a generally benign course after the initial surgical treatment of their nephrolithiasis. Four of our eight patients were free of symptoms. Three patients have not had recurrences of their calculi; only one has had recurrent stone formation since his original nephrolithiasis at 12 years of age. Thus hyperoxaluria type 2 may be a much milder disease with a better long-term prognosis for renal function than is the case in type 1. W e believe that infants and children having renal calculi should be investigated for metabolic disease, including L-glyceric aciduria. This can be done in any laboratory

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Clinical and laboratory observations

The Journal of Pediatrics June 1991

conducting organic acid analysis for the detection of inherited metabolic disorders. L-Glyceric aciduria may be missed if only oxaiate is measured in urine because oxalate excretion may not be great. All patients with primary hyperoxaluria should be carefully investigated to determine whether they have type 1 or type 2 disease. It is probable that more patients with type 2 would be identified if all patients with hyperoxaluria were so classified, therefore allowing a more complete description of the disease, improved data on the prognosis, the development of a treatment protocol, and more accurate genetic counseling.

3.

4. 5. 6.

7.

We thank Dr. Gina Rempel for the clinical details of patient 8.

8.

REFERENCES 1. Williams HE, Smith LH Jr. L-Glycericaciduria: a new genetic variant of primary hyperoxaluria. N Engl J Med 1968;278: 233-9. 2. Chalmers RA, Tracey BM, Mistry J, Griffiths KD, Green A, Winterborn MH. L-Glycericaciduria (primary hyperoxaluria

9. 10.

type 2) in siblingsin two unrelated families. J Inherited Metab Dis 1984;7(suppl2):133-4. Mistry J, Danpure CJ, Chalmers RA. Hepatic D-glyceratedehydrogenase and glyoxylate reductase deficiency in primary hyperoxaluria type 2. Biochem Soc Trans 1988;16:626-7. Willis JE, Sallach HJ. Evidence for a mammalian D-glycerate dehydrogenase. J Biol Chem 1962;237:910-5. Dawkins PD, Dickens F. The oxidation of o- and L-glycerate by rat liver. Biochem J 1965;94:353-67. Danpure CJ, Jennings PR, Mistry J, et al. Enzymological characterization of a feline analogue of primary hyperoxaluria type 2: a model for the human disease. J Inherited Metab Dis 1989;12:403-14. Williams HE, Smith LH. The identificationand determination of glyceric acid in human urine. J Lab Clin Med 1968;71: 495-500. Goodman SI, Markey SP. Diagnosis of organic acidemias by gas chromatography-mass spectrometry. New York: Alan R. Liss, 1981. Crider QE, Curran DF. Simplifiedmethod for enzymatic urine oxalate assay. Clin Biochem 1984;17:31-5. Hillman RE. Primary hyperoxalurias. In: Scriver CR, Beaudet AL, Sly WS, Valle D, eds. The metabolic basis of inherited disease. New York: McGraw-Hill, 1989:933-44.

Use of citrulline as a diagnostic marker in the prospective t r e a t m e n t of urea cycle disorders Mark L. B a t s h a w , MD, a n d G e r a r d T. Berry, MD From the Children's Seashore House, Children's Hospital of Philadelphia, and the Department of Pediatrics, University of Pennsylvania School of Medicine, Philadelphia

In children born with a complete deficiency of one of the enzymes in the urea cycle (other than arginase), hyperammonemic coma occurs within the first few days of life. Plasma citrulline has been proposed as being critical in differentiating the various complete inborn errors of urea synthesis.I Citrulline is the product of the first two enzymes in the urea cycle, carbamyl phosphate synthetase and ornithine transcarbamylase; thus children with deficiencies in these enzymes should have trace or absent citrulline levels. Citrulline is the substrate for the later three enzymes in the urea cycle and should be elevated in deficiencies of argininosuccinate synthetase (citrullinemia) and argininosucciSupported by U.S. Public Health Service grants R0t-NS-28033 and P30-HD-26979. Submitted for publication Nov. 9, 1990; accepted Jan. 11, 1990. Mark L. Batshaw, MD, Children's Seashore House, 3405 Civic Center Blvd., Philadelphia, PA 19104. 9/22/27912

nate lyase (argininosuccinic aciduria). This algorithm is important because it permits rapid diagnosis and intervention in these disorders. Plasma citrulline levels have also been useful in determining whether a newborn infant at genetic risk for a urea cycle disorder is, in fact, affected. For OTC and CPS deficiencies, the neonate is given therapy from birth, and citrulline levels are followed as protein is gradually added CPS OTC

Carbamyl phosphate synthetase Ornithine transearbamylase

to the diet. 2 It had been assumed that extremely low or undetectable plasma levels of citrulline established the diagnosis of a urea cycle disorder even in the absence of hyperammonemia or clinical disease. In this report we present the case of a neonate at risk for CPS deficiency who had undetectable or trace amounts of citrulline in plasma in the first days of life but subsequently