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Renal iminoglycinuria without intestinal malabsorption of glycine and imino acids
386
March, 1970 T h e ]ournal of P E D I A T R I C S
Renal iminoglycinuria without intestinal malabsorption of glycine and imino acids Renal iminoglyeinuria was detected in a 12-year-old girl affected by congenital achromatopsia with severe amblyopia. Both parents and other relatives have renal hyperglycinur~a. The resutts of family studies suggest that renal iminoglycinuria ~s observed in homozygotes [,or a mutation which in heterozygotes causes isolated hyperglycinuria. In this patient the intestinal absorption of glycine and imino acids seems to be normal. The association between the ocular disease and the renal defect is probably fortuitous.
Francesco Tancredi, M.D., "x" Giancarlo Guazzi, M.D., and
Salvatore Auricchio, M.D. NAPLES~ ITALY
T H E R E ~q A L absorption of glycine, proline, and hydroxyproline probably occurs via several transport systems. O n e of these is shared by all three compounds, but has the largest affinity for proline? -4 A genetic deficiency of this system has been found in some individuals affected by iminoglycinuria 5-1~: Renal iminoglycinuria is observed kn homozygotes for a mutation which in heterozygotes causes renal hyperglycinuria2 -1~ An associated defect of intestinal absorption of imino acids has been observed in only two patients. 6, z We have studied the renal and intestinal transport of glycine and imino acids in a girl with renal iminoglycinuria and low visual acuity, and also in some members of her family. T h e transport of imino acids and glyFrom the Departments of Child Health and Neurology, University of Naples. ~Address: Department of Child Health, UMvers~ty o] Naples, Via Annunzlata 34, Naples, Italy.
Vol. 76, No. 3, pp. 386-392
cine was found to be reduced in the kidney but not in the intestine.
CASE REPORT Patient. The proposita (Fig. 1, IV, IO), a 12year-old girl, was identified in a screening program for aminoaciduria conducted in schools for blind children. Gestation, delivery, growth, and development were normal; she walked at 12 months and menstruated at 12 years. At the age of 4 months photophobia and low visuaI acuity were noticed. Her height and weight were normal. Mental retardation was not found by a psychologist nor was it thought to exist by her teachers. Since the age of 4 years she had been in an institution for Mind children, where she had made good progress in her studies. On neurologic examination, the girl was found to be normal with the exception of the ocular findings: horizontal pendular nystagmus and marked photophobia; fundus--slight pallor of the optic disc with normal borders; visual acuity
Fig. 1. Pedigree of Family R.
V
III
II
[]
[]
[]
proband, ,mmog[ycinurla and achroma)opsla
mate, not s~udled
[]
female, nor s)Udled.
2
[ ] achromatopsia withoul- hypergl.ycmurla
[]
0
/
I
,r
10
~p
16
4
deceased
normal glycmne excrehon
hypergtycmuna wdh atbm~sm
hyperglycmuna
/
3
,r 13
r
C)
"-4
3 88
Tancredi, Guazzi, and Auricchio
--3/50; color vision--able to read the first card of the Ishihara test; totally unable to set up any line at the Farnsworth 100 Hue test; electroretinogram (ERG)--scotopic a and b waves present and within normal limits, photopic components undetectable for absence of response following the first in the flicker-ERG (frequency: 30 flash/see.; energy: 1.5 joule/flash). As a conclusion of these findings, the diagnosis of congenital total achromatopsia was made. The resuIts of urine analyses and blood examinations for hemoglobin, red and white blood cells, urea, electrolytes, alkaline phosphatase, proteins, cholesterol, glucose, bilirubin, Wassermann test, pH, p C Q , and standard bicarbonate were normal. Endogenous creatinine clearance values ranged between 82.4 and 94.0 ml. per minute per 1.73 MY Bone radiographs, intravenous pyelogram, electrocardiogram, and electroencephalogram were interpreted to be within normal limits. Family. In the 18-year-old sister (IV, 8) and the 16-year-old brother (IV, 9) photophobia and low visual acuity were observed at the age of 4 to 6 months. They were able to study in normal classes in an institution for blind children for 2 years. Afterwards, at the age of 8 to 9 years, mental deterioration was detected and they were unable to continue their studies. At the time of our examination their visual acuity was 3/50. Clinical and ERG examinations showed that both were affected by congenital total achromatopsia. The parents (III, 12 and 13) were in good health, without ocular defects or mental retardation. Color vision tests and the ERG of the mother were normal The parents were not consanguineous, but both of their families had lived for many generations in a small village (about 3,000 inhabitants) in southern Italy. There was no history of ocular diseases or mental retardation in the father's family. C. C. (K, 1), an l 1-year-old girl, had had very low visual acuity from the first year of iife and was mentally retarded. A. G. (IV, 6), a 20-year-old man, had albinism and low visual acuity. V. G. (II, 10) and V. F. (II, 12), now 68 and 66 years of age, respectively, had had very low visual acuity from the first months of life, without appreciable mental retardation. METHODS
Free u r i n a r y amino acids were ,separated by high-voltage electrophoresis ( W h a t m a n
The Journal o[ Pediatrics March 1970
p a p e r , 3 m m . ; formic a c i d : a c e t i c acid buffer, p H 2.1, 3,500 v, 130 mA. for 40 rain.) a n d detected with n i n h y d r i n 0.2 p e r cent in acetone containing collidine 5 per cent. T h e quantitative analysis of amino acids in urine a n d p l a s m a was p e r f o r m e d on ion-exchange resins a c c o r d i n g to S p a c k m a n a n d associates, 11 using a B e c k m a n - S p i n c o A m i n o A c i d Analyzer m o d e l 120 B. All urine samples were stored i m m e d i a t e l y after collection at - 2 0 ~ C. 'Plasma was imm e d i a t e l y separated f r o m samples of venous blood a n d deproteinized with 1 p e r cent picric a c i d ? 2 T h e endogenous renal clearance for a m i n o acids were estimated u n d e r fasting conditions in the patient, in her b r o t h e r (IV, 9), and in her mother. T h e urine was collected d u r i n g a p e r i o d of 3 hours; blood samples were o b t a i n e d at the beginning a n d at the end of this period. I n the p a t i e n t a n d in h e r m o t h e r the study was r e p e a t e d while they received a diet rich in gelatin. O n this diet a 24 h o u r clearance of a m i n o acids was also d e t e r m i n e d in the patient. C r e a t i n i n e was m e a s u r e d in blood serum a n d in urine *a to estimate the g l o m e r u l a r filtration rate. U r i n a r y excretion (UVaA = /,IV[ p e r m i n ute per 1.73 M.2), renal clearance (Can = milliliters p e r m i n u t e p e r 1.73 M.2), a n d t u b u l a r absorption in percentage of the filtered load ( R T % ) were calculated according to Scriver a n d Davies. 1~ F o r the study of intestinal absorption, the a m i n o acids in stools were d e t e r m i n e d d u r ing a 3 day b a l a n c e period, both on n o r m a l diet a n d on a diet rich in gelatin. Stools were i m m e d i a t e l y frozen after collection a n d then homogenized. T h e a m i n o acids were extracted from h o m o g e n a t e a n d analyzed as described by G o o d m a n a n d associates. 7 O r a l tolerance tests with glycine (200 rag. p e r kilogram) a n d proline (100 rag. p e r kilogram) were p e r f o r m e d in the p a t i e n t a n d in n o r m a l control subjects in the m o r n ing u n d e r fasting conditions. Blood samples were collected before a n d 1, 2, 3, a n d 6 hours after t h e load. U r i n e samples were collected in the m o r n i n g u n d e r fasting conditions from the
Volume 76 Number 3
Renal iminoglycinuria
3 89
small quantities of proline and hydroxyproline were detected by column chromatography. I n urine of normal control subjects of the same age, imino acids were never found. T h e 24 h o u r excretions of gtycine, proline, and hydroxyproline were abnormally high, whereas the blood levels were normal (Table I ) . No other abnormality in the amino acid pattern of blood and urine was detected. T h e specific defect of the renal tubular absorption of glycine and imino acids was confirmed by very high endogenous clearances
parents, the siblings, and 20 other relatives. T h e glycine concentration was expressed as micromoles per milligrams of creatinine s and as micromoles per milligrams of total nitrogen? ~ R E S U L T S
High-voltage electrophoresis of the patient's urine revealed iminoglycinuria in all samples collected when she was on the diet rich in gelatin. While she was given a normal diet, however, only hyperglycinuria was evident on two occasions by this method, but
Table I. Blood levels and urinary excretion of glycine (Gly), proline (Pro), and hydroxyproline ( O H - P r o ) in the patient
Date
GIy
Plasma (#M/IO0 rot.) I Pro I
Nov. 6, 1967" June 22, 1968t June 28, 1968" July 4, 1968" Normal values$
26.67 31.08 22.90' 31.62 < 40
8.60 10.35 10.54 11.40 ~ 16
*'On a normal diet. t O n a diet rich in gelatin. SThis laboratory and data from literature w
OH-Pro Trace Trace Trace 0 0~2.5w
Gly
Urine (l~M/24 hr.) I 1 Pro
5,985 9,441
OH-Pro
437 3,014
< 1,800
129 1,619
0-trace
(Scrlver and Davles14; Dickinson and associates2~
Table II. U r i n a l / e x c r e t i o n (UVAA),renal clearance (CAA), and tubular absorption ( R T % ) of glycine (Gly), proline (Pro), and hydroxyproline ( O H - P r o )
Subject Patient
UVAA (#M/min./1.73 M. e) Gly Pro I OH-Pro
CA.A
(ml./min./1.73 M. e) Gly t Pro
Gly
RT% I Pro
1" 2t 35
4.75 6.99 6.82
0.347 2.229 2.020'
0.102 1.192 0.987
17.82 21.94 20.86
3.37 15.81 12.24
78.6 73.6 75.O
95.9 81.0 84.1
1" 2t
3.86 4.24
0 0
0 0
12.00 13.09
0 0
89.5 87.0
100.0 100.0
0.62
0
0
2.96
0
96.6
100.0
0.33 0.68 0.54 1.03 0.33-1.54
0 0 0 0 0-0.0.38
0 0 0 0
1.10 3.08 2.17 4.60 1.2-8.6
0 0 0 0 0-0.3
99.2 95.1 98.0 96.2 92.9-99.0
100.0 I00.0 100.0 100.0 99.8-100
Mother Brother (IV, 9) 1*
Controls A.G.* B.F.* A.F.* A.F.t
Normal valuesw
*'Three hour's clearance, on a normal diet. tThree hour's clearance, on a diet rich in gelatin. :~Twenty-fom" hour's clearance, on a diet rich in gelatin. w (from Scriver and Davies14).
-
390
The Journal o/ Pediatrics March 1970
Tancredi, Guazzi, and Aurlcchlo
260,
>.-
,,~, I~ ",~,
-"=----~=
controls
= patient
d lzu] | 10(~
-'~---'=--
~\
--
controls
-- patient
140
,,,
"'" 40
20
20
" ..... i
23
,~
56
HOURS OF STUDY
"
~'
1~ 12 3/I. 56 HOURS OF STUDY
Fig. 2. Left: Blood glycine levels after a load of glyeine (200 mg. per kilogram) by mouth. Right: Blood proline levels after a load of proline (100 rag. per kilogram) by mouth. No difference between patient and normal control subjects was observed.
and the low tubular reabsorption of the filtered glycine and proline. The renal clearance and the tubular absorption of hydroxyproline were not determined because of the very low levels of this imino acid in blood. That the tubular absorption of hydroxyproline was reduced was indicated by its increased urinary excretion despite its normal concentration in the blood. On a diet rich in gelatin the urinary excretion of glycine, proline, and hydroxyproline increased, without appreciable variations of their blood levels. This phenomenon was more marked for proline and hydroxyproline than for glycine (Table I I ) . The intestinal absorption of glycine and imino acids was not impaired in our patient. The content of glycine, proline, and hydroxyproline in stoool was normal, both on a free diet and on a diet rich in gelatinl The blood glycine and proline levels after oral tolerance tests were also normal (Fig. 2). No other case of iminoglycinuria was found among the relatives of the patient. Hyperglycinuria was observed in both parents and in some other relatives (Tables I I and I I I ) . In the mother no iminoglycinuria
was observed while on a diet rich in gelatin (Table I I ) .
DISCUSSION Renal iminoglycinuria has been described in 15 individuals belonging to 9 families from different ethnic backgroundsr -1~ 1~ Many cases have been observed in Ashkenazi Jewsy, 0, 10 Most of these patients are in good health, but 4 subjects are mentally retarded 5-~ and one is deaf. 8 Our patient also has congenital total achromatopsia. This disease is inherited in an autosomal recessive manner; only a few exceptions have been reported of its transmission as a sex-linked recessive trait 1~, 17 or as a dominant trait. ~a In the latter situation the occurrence of pseudodominance was not ruled out. In the family of our patient, .the disease is probably transmitted as a dominant trait with incomplete expression (Fig. 1). The ocular disease does not seem to be associated with the renal defect. Both siblings of the proposita also have congenital total achromatopsia, but they have normal excretion of glycine and imino acids, The ocular disease is present only in the family
Volume 76 Number 3
R e n a l irninoglycinuria
39 1
Table I I I . Urinary excretion of glycine in the family members Subject Normal values R. D. (III, 13), fafher B. A. (III, 12), mother R. F. (IV, 8), sister R. Fi. (IV, 9), ,brother B. A. (II, 2) P. C. (H, 9) V. G. (II, 10) V. F. (H, 12) A. C. (III, 2) A. V. (1II, 3) A. E. (IIL 5) B. A. (III, 9) F. V. (III, 19) R. M. (III, 14) V.. ~R. (III, 16) C. G. (IV, 2) S. N. (IV, 1) A. G. (IV, 4) A. O. (IV, 5) A. G. (IV, 6) F. A. (IV, 11) F. F. (IV, 12) C. C. (V, 1) C. A. (V, 2)
I
#M/mg. ereatinine 1.120 +- 0.533~ 2.105 8.200 1.294 1.186 2.156 5.502 0.234 0.630 1.953 2.283 4.400 12.875 0.230 0.828 0.680 1.947 O.64O 0.550 0.335 5.546 1.583 1.680 1.614 0.066
I
#M/mg. nitrogen ~ 0.160~ 0.467 1.302 0.132 0.087 0.603 1.376 0.018 0.060 0.306 0.40.8 0.372 1.348 0.045 0.037 0.042 0.380 0.098 0.082 0.022 0.396 0.144 0.070 0.018 0.019
Heterozygote Hetemzygote Normal Normal IIeterozygote Heterozygote Normal Normal Heterozygote Heterozygote Heterozygote Hetemzygote Normal Normal Normal Heterozygote Normal Normal Normal tIeterozygote Normal Normal Normal Normal
e M e a n _+ S.D., f r o m Rosenberg and associates, s t F r o m Scriver. 1~
of the mother, not in association with iminoglycinuria or hyperglycinuria. Both parents of our patient have hyperglycinuria (Table I I I and Fig. 1). Furthermore, family studies show that the hyperglycinuria is inherited as a dominant trait (Fig. 1). Heterozygotes for this defect apparently have isolated hyperglycinuria (only a few exceptions are known ~' 10); homozygotes have iminoglycinuria. Some evidence exists of the presence of more than one renal transport system for glycine and imino acids? 5 One of these is common to all three compounds (system A ) ; two (or more) others are responsible for a selective absorption of glycine and imino acids (system B ) ? ~ In the homozygotes for iminoglycinuria the remaining tubular absorption of glycine and imino acids is probably due to system B, as shown by the inability of high blood levels of proline to decrease further the renal tubular absorption of glycine in these subjects? ~ In our patient the diet rich in gelatin
depressed the tubular absorption of the imino acids more than that of glycine. Both observations support the hypothesis of the existence of independent renal transport systems for glycine and imino acids (system B). In the heterozygotes, the remaining activity of system A appears to be sufficient, at normal Mood amino acid concentrations, to absorb all of the filtered proline and hydroxyproline but only part of the glycine. Studies of the fecal excretion of imino acids and of blood levels of proline and glycine after oral tolerance tests show in our patient, as in 3 others, 8' ~0 that the renal defect is not associated with" impairment of intestinal transport of glycine and imino acids. In one of these patients s uptake of glycine-2-C~4 by intestinal biopsies~was found to be normal. In two other patients 6' ~ the intestinal transport of glycine (in one of these) and of imino acids (in both) was impaired. If these results are confirmed, it should be possible to distinguish different forms of iminoglycinuria, depending on
392
Tancredi, Guazzi, and Auricchio
whether the renal transport defect occurs alone or together with a n intestinal transport defect, as has been observed in cystinuria. ~o SUMMARY
R e n a l iminoglycinuria was detected in a 12-year-old girl affected by congenital achromatopsia with severe amblyopia. Both parents a n d other relatives have renal hyperglycinuria. Family studies support the following conclusions: (1) R e n a l iminoglycinuria is observed in homozygotes for a m u t a t i o n which in heterozygotes causes isolated hyperglycinuria; (2) T h e association between ocular disease a n d the renal defect is probably fortuitous. I n this p a t i e n t the intestinal absorption of glycine a n d imino acids seems to be normal.
The Journal of Pediatrics March 1970
7.
8.
9. 10.
11.
12. 13,
We are greatly indebted to Dr. F. Ponte, Department of Ophthalmology, University of Palermo, Italy, for the ophthalmic and ERG studies and for assessment of the diagnosis of congenital a~hromatopsia. REFERENCES
1. Scriver, C, R., Effort, M. L., and Schafer, I. A.: Renal tubular transport of proline, hydroxyproline and glycine in health and in familial hyperprolinemia, J. Clin. Invest. 43" 374, 1964. 2. Scriver, C. R., and Wilson, O. H.: Possible locations for common gene product in membrane transport of iminoacids and glycine, Nature 202= 92, 1964. 3. Seriver, C. R., and Goldman, I-t.: Renal tubular transport of proline, hydroxyproline and glycine. II. Hydroxy-L-proline as substrate and as inhibitor in vivo, J. Clin. Invest. 45: 1357, 1966. 4. Wilson, O. H., and Scriver, C. R.: Specificity of transport of neutral and basic amino acids in rat kidney, Amer. J. Physiol. 213: 185, 1967. 5. Tada, K., Morikawa, T., Ando, T., Yoshida, T., and Minagawa, A.: Prolinuria: a new renal tubular defect in transport of proline and glycine, Tohoku J. Exp. Med. 87: 133, 1965. 6. Morikawa, T., Tada, K., Ando, T., Yosbida, T., Yokoyama, Y., and Arakawa, T.: Prolinurla: defect in intestinal absorption of
14.
15.
16.
17.
18.
19.
20.
iminoacids and glycine, Tohoku J. Exp. Med. 90: 105, 1966. Goodman, S. I., Mclntyre, C. A., Jr., and O'Brien, D.: Impaired intestinal transport of proline in patient with familial iminoaciduria, J. P~I)IAT. 71: 246, 1967. Rosenberg, L. E., Durant, J. L., and Elsas, L. J.: Familial imin0glycinuria: An inborn error of renal tubular transport, New Eng. J. Med. 978: 1407, 1968. Whelan, D. T., and Scriver, C. R.: Cystathioninuria and renal iminoglycinuria in a pedigree, New Eng. J. M. 278: 924, 1968. Scriver, C. R.: Renal tubular transport of proline, hydroxyproline and glycine. III. Genetic basis for more than one mode of transport in human kidney, J. Clin. Invest. 47: 823, 1968. Spackman, D. I-t., Stein, W. H., and Moore, S.: Automatic recording apparatus for use in the chromatography of amino acids, Anal. Chem. 30: 1190, 1958. Stein, W. H., and Moore, S.: The free amino acids of human blood plasma, J. Biol. Chem. 211: 915, 1954. Owen, S. A., Iggo, B., Scandrett, F. J., and Stewart, C. P.: The determination of creatinine in plasma or serum and in urine: A critical examination, Biochem. J. 58: 426, 1954. Scriver, C. R., and Davies, E.: Endogenous renal clearance rates of free aminoacids in pre-pubertal children, Pediatrics 36" 592, 1965. Scrlver, C. R., and Wilson, O. H.: Aminoacid transport: Evidence for genetic control of two types in human kidney, Science 155: 1428, 1967. Spivey, B. E., Pearlman, J. T., and Burian, I~. M.: Electroretinographie findings (including flicker) in carriers of congenital Xlinked achromatopsia, Doc. Ophthal. 18: 367, 1964. Krill, A. E., and Schneiderman, A.: Retinal function studies, including the electroretlnogram, in an atypical monochromat, In Clinical electroretinography, Suppl. Vision Res., New York, 1966, Oxford Press, p. 351. Franceschetti, A., Francois, J., and Babel, J.: In Les h6rddo-d6gdn6rescenees chorio-rgtiniennes, vol. II, Paris, 1963, Mas.son & Cie, p. 1252. Rosenberg, L. E., Downing, S., Durant, J. L., and Segal, S. : Cystlnuria : biochemical evidence for three genetically distinct .diseases, J. Clin. Invest. 45: 365, 1966. Dickinsori, S. C., Rosenblum, H., and Hamilton, P. B.: Ion-exchange chromatography of free amino acids in the plasma of the newborn infant, Pediatrics 36" 2, 1965.
March, 1970 T h e ]ournal of P E D I A T R I C S
Renal iminoglycinuria without intestinal malabsorption of glycine and imino acids Renal iminoglyeinuria was detected in a 12-year-old girl affected by congenital achromatopsia with severe amblyopia. Both parents and other relatives have renal hyperglycinur~a. The resutts of family studies suggest that renal iminoglycinuria ~s observed in homozygotes [,or a mutation which in heterozygotes causes isolated hyperglycinuria. In this patient the intestinal absorption of glycine and imino acids seems to be normal. The association between the ocular disease and the renal defect is probably fortuitous.
Francesco Tancredi, M.D., "x" Giancarlo Guazzi, M.D., and
Salvatore Auricchio, M.D. NAPLES~ ITALY
T H E R E ~q A L absorption of glycine, proline, and hydroxyproline probably occurs via several transport systems. O n e of these is shared by all three compounds, but has the largest affinity for proline? -4 A genetic deficiency of this system has been found in some individuals affected by iminoglycinuria 5-1~: Renal iminoglycinuria is observed kn homozygotes for a mutation which in heterozygotes causes renal hyperglycinuria2 -1~ An associated defect of intestinal absorption of imino acids has been observed in only two patients. 6, z We have studied the renal and intestinal transport of glycine and imino acids in a girl with renal iminoglycinuria and low visual acuity, and also in some members of her family. T h e transport of imino acids and glyFrom the Departments of Child Health and Neurology, University of Naples. ~Address: Department of Child Health, UMvers~ty o] Naples, Via Annunzlata 34, Naples, Italy.
Vol. 76, No. 3, pp. 386-392
cine was found to be reduced in the kidney but not in the intestine.
CASE REPORT Patient. The proposita (Fig. 1, IV, IO), a 12year-old girl, was identified in a screening program for aminoaciduria conducted in schools for blind children. Gestation, delivery, growth, and development were normal; she walked at 12 months and menstruated at 12 years. At the age of 4 months photophobia and low visuaI acuity were noticed. Her height and weight were normal. Mental retardation was not found by a psychologist nor was it thought to exist by her teachers. Since the age of 4 years she had been in an institution for Mind children, where she had made good progress in her studies. On neurologic examination, the girl was found to be normal with the exception of the ocular findings: horizontal pendular nystagmus and marked photophobia; fundus--slight pallor of the optic disc with normal borders; visual acuity
Fig. 1. Pedigree of Family R.
V
III
II
[]
[]
[]
proband, ,mmog[ycinurla and achroma)opsla
mate, not s~udled
[]
female, nor s)Udled.
2
[ ] achromatopsia withoul- hypergl.ycmurla
[]
0
/
I
,r
10
~p
16
4
deceased
normal glycmne excrehon
hypergtycmuna wdh atbm~sm
hyperglycmuna
/
3
,r 13
r
C)
"-4
3 88
Tancredi, Guazzi, and Auricchio
--3/50; color vision--able to read the first card of the Ishihara test; totally unable to set up any line at the Farnsworth 100 Hue test; electroretinogram (ERG)--scotopic a and b waves present and within normal limits, photopic components undetectable for absence of response following the first in the flicker-ERG (frequency: 30 flash/see.; energy: 1.5 joule/flash). As a conclusion of these findings, the diagnosis of congenital total achromatopsia was made. The resuIts of urine analyses and blood examinations for hemoglobin, red and white blood cells, urea, electrolytes, alkaline phosphatase, proteins, cholesterol, glucose, bilirubin, Wassermann test, pH, p C Q , and standard bicarbonate were normal. Endogenous creatinine clearance values ranged between 82.4 and 94.0 ml. per minute per 1.73 MY Bone radiographs, intravenous pyelogram, electrocardiogram, and electroencephalogram were interpreted to be within normal limits. Family. In the 18-year-old sister (IV, 8) and the 16-year-old brother (IV, 9) photophobia and low visual acuity were observed at the age of 4 to 6 months. They were able to study in normal classes in an institution for blind children for 2 years. Afterwards, at the age of 8 to 9 years, mental deterioration was detected and they were unable to continue their studies. At the time of our examination their visual acuity was 3/50. Clinical and ERG examinations showed that both were affected by congenital total achromatopsia. The parents (III, 12 and 13) were in good health, without ocular defects or mental retardation. Color vision tests and the ERG of the mother were normal The parents were not consanguineous, but both of their families had lived for many generations in a small village (about 3,000 inhabitants) in southern Italy. There was no history of ocular diseases or mental retardation in the father's family. C. C. (K, 1), an l 1-year-old girl, had had very low visual acuity from the first year of iife and was mentally retarded. A. G. (IV, 6), a 20-year-old man, had albinism and low visual acuity. V. G. (II, 10) and V. F. (II, 12), now 68 and 66 years of age, respectively, had had very low visual acuity from the first months of life, without appreciable mental retardation. METHODS
Free u r i n a r y amino acids were ,separated by high-voltage electrophoresis ( W h a t m a n
The Journal o[ Pediatrics March 1970
p a p e r , 3 m m . ; formic a c i d : a c e t i c acid buffer, p H 2.1, 3,500 v, 130 mA. for 40 rain.) a n d detected with n i n h y d r i n 0.2 p e r cent in acetone containing collidine 5 per cent. T h e quantitative analysis of amino acids in urine a n d p l a s m a was p e r f o r m e d on ion-exchange resins a c c o r d i n g to S p a c k m a n a n d associates, 11 using a B e c k m a n - S p i n c o A m i n o A c i d Analyzer m o d e l 120 B. All urine samples were stored i m m e d i a t e l y after collection at - 2 0 ~ C. 'Plasma was imm e d i a t e l y separated f r o m samples of venous blood a n d deproteinized with 1 p e r cent picric a c i d ? 2 T h e endogenous renal clearance for a m i n o acids were estimated u n d e r fasting conditions in the patient, in her b r o t h e r (IV, 9), and in her mother. T h e urine was collected d u r i n g a p e r i o d of 3 hours; blood samples were o b t a i n e d at the beginning a n d at the end of this period. I n the p a t i e n t a n d in h e r m o t h e r the study was r e p e a t e d while they received a diet rich in gelatin. O n this diet a 24 h o u r clearance of a m i n o acids was also d e t e r m i n e d in the patient. C r e a t i n i n e was m e a s u r e d in blood serum a n d in urine *a to estimate the g l o m e r u l a r filtration rate. U r i n a r y excretion (UVaA = /,IV[ p e r m i n ute per 1.73 M.2), renal clearance (Can = milliliters p e r m i n u t e p e r 1.73 M.2), a n d t u b u l a r absorption in percentage of the filtered load ( R T % ) were calculated according to Scriver a n d Davies. 1~ F o r the study of intestinal absorption, the a m i n o acids in stools were d e t e r m i n e d d u r ing a 3 day b a l a n c e period, both on n o r m a l diet a n d on a diet rich in gelatin. Stools were i m m e d i a t e l y frozen after collection a n d then homogenized. T h e a m i n o acids were extracted from h o m o g e n a t e a n d analyzed as described by G o o d m a n a n d associates. 7 O r a l tolerance tests with glycine (200 rag. p e r kilogram) a n d proline (100 rag. p e r kilogram) were p e r f o r m e d in the p a t i e n t a n d in n o r m a l control subjects in the m o r n ing u n d e r fasting conditions. Blood samples were collected before a n d 1, 2, 3, a n d 6 hours after t h e load. U r i n e samples were collected in the m o r n i n g u n d e r fasting conditions from the
Volume 76 Number 3
Renal iminoglycinuria
3 89
small quantities of proline and hydroxyproline were detected by column chromatography. I n urine of normal control subjects of the same age, imino acids were never found. T h e 24 h o u r excretions of gtycine, proline, and hydroxyproline were abnormally high, whereas the blood levels were normal (Table I ) . No other abnormality in the amino acid pattern of blood and urine was detected. T h e specific defect of the renal tubular absorption of glycine and imino acids was confirmed by very high endogenous clearances
parents, the siblings, and 20 other relatives. T h e glycine concentration was expressed as micromoles per milligrams of creatinine s and as micromoles per milligrams of total nitrogen? ~ R E S U L T S
High-voltage electrophoresis of the patient's urine revealed iminoglycinuria in all samples collected when she was on the diet rich in gelatin. While she was given a normal diet, however, only hyperglycinuria was evident on two occasions by this method, but
Table I. Blood levels and urinary excretion of glycine (Gly), proline (Pro), and hydroxyproline ( O H - P r o ) in the patient
Date
GIy
Plasma (#M/IO0 rot.) I Pro I
Nov. 6, 1967" June 22, 1968t June 28, 1968" July 4, 1968" Normal values$
26.67 31.08 22.90' 31.62 < 40
8.60 10.35 10.54 11.40 ~ 16
*'On a normal diet. t O n a diet rich in gelatin. SThis laboratory and data from literature w
OH-Pro Trace Trace Trace 0 0~2.5w
Gly
Urine (l~M/24 hr.) I 1 Pro
5,985 9,441
OH-Pro
437 3,014
< 1,800
129 1,619
0-trace
(Scrlver and Davles14; Dickinson and associates2~
Table II. U r i n a l / e x c r e t i o n (UVAA),renal clearance (CAA), and tubular absorption ( R T % ) of glycine (Gly), proline (Pro), and hydroxyproline ( O H - P r o )
Subject Patient
UVAA (#M/min./1.73 M. e) Gly Pro I OH-Pro
CA.A
(ml./min./1.73 M. e) Gly t Pro
Gly
RT% I Pro
1" 2t 35
4.75 6.99 6.82
0.347 2.229 2.020'
0.102 1.192 0.987
17.82 21.94 20.86
3.37 15.81 12.24
78.6 73.6 75.O
95.9 81.0 84.1
1" 2t
3.86 4.24
0 0
0 0
12.00 13.09
0 0
89.5 87.0
100.0 100.0
0.62
0
0
2.96
0
96.6
100.0
0.33 0.68 0.54 1.03 0.33-1.54
0 0 0 0 0-0.0.38
0 0 0 0
1.10 3.08 2.17 4.60 1.2-8.6
0 0 0 0 0-0.3
99.2 95.1 98.0 96.2 92.9-99.0
100.0 I00.0 100.0 100.0 99.8-100
Mother Brother (IV, 9) 1*
Controls A.G.* B.F.* A.F.* A.F.t
Normal valuesw
*'Three hour's clearance, on a normal diet. tThree hour's clearance, on a diet rich in gelatin. :~Twenty-fom" hour's clearance, on a diet rich in gelatin. w (from Scriver and Davies14).
-
390
The Journal o/ Pediatrics March 1970
Tancredi, Guazzi, and Aurlcchlo
260,
>.-
,,~, I~ ",~,
-"=----~=
controls
= patient
d lzu] | 10(~
-'~---'=--
~\
--
controls
-- patient
140
,,,
"'" 40
20
20
" ..... i
23
,~
56
HOURS OF STUDY
"
~'
1~ 12 3/I. 56 HOURS OF STUDY
Fig. 2. Left: Blood glycine levels after a load of glyeine (200 mg. per kilogram) by mouth. Right: Blood proline levels after a load of proline (100 rag. per kilogram) by mouth. No difference between patient and normal control subjects was observed.
and the low tubular reabsorption of the filtered glycine and proline. The renal clearance and the tubular absorption of hydroxyproline were not determined because of the very low levels of this imino acid in blood. That the tubular absorption of hydroxyproline was reduced was indicated by its increased urinary excretion despite its normal concentration in the blood. On a diet rich in gelatin the urinary excretion of glycine, proline, and hydroxyproline increased, without appreciable variations of their blood levels. This phenomenon was more marked for proline and hydroxyproline than for glycine (Table I I ) . The intestinal absorption of glycine and imino acids was not impaired in our patient. The content of glycine, proline, and hydroxyproline in stoool was normal, both on a free diet and on a diet rich in gelatinl The blood glycine and proline levels after oral tolerance tests were also normal (Fig. 2). No other case of iminoglycinuria was found among the relatives of the patient. Hyperglycinuria was observed in both parents and in some other relatives (Tables I I and I I I ) . In the mother no iminoglycinuria
was observed while on a diet rich in gelatin (Table I I ) .
DISCUSSION Renal iminoglycinuria has been described in 15 individuals belonging to 9 families from different ethnic backgroundsr -1~ 1~ Many cases have been observed in Ashkenazi Jewsy, 0, 10 Most of these patients are in good health, but 4 subjects are mentally retarded 5-~ and one is deaf. 8 Our patient also has congenital total achromatopsia. This disease is inherited in an autosomal recessive manner; only a few exceptions have been reported of its transmission as a sex-linked recessive trait 1~, 17 or as a dominant trait. ~a In the latter situation the occurrence of pseudodominance was not ruled out. In the family of our patient, .the disease is probably transmitted as a dominant trait with incomplete expression (Fig. 1). The ocular disease does not seem to be associated with the renal defect. Both siblings of the proposita also have congenital total achromatopsia, but they have normal excretion of glycine and imino acids, The ocular disease is present only in the family
Volume 76 Number 3
R e n a l irninoglycinuria
39 1
Table I I I . Urinary excretion of glycine in the family members Subject Normal values R. D. (III, 13), fafher B. A. (III, 12), mother R. F. (IV, 8), sister R. Fi. (IV, 9), ,brother B. A. (II, 2) P. C. (H, 9) V. G. (II, 10) V. F. (H, 12) A. C. (III, 2) A. V. (1II, 3) A. E. (IIL 5) B. A. (III, 9) F. V. (III, 19) R. M. (III, 14) V.. ~R. (III, 16) C. G. (IV, 2) S. N. (IV, 1) A. G. (IV, 4) A. O. (IV, 5) A. G. (IV, 6) F. A. (IV, 11) F. F. (IV, 12) C. C. (V, 1) C. A. (V, 2)
I
#M/mg. ereatinine 1.120 +- 0.533~ 2.105 8.200 1.294 1.186 2.156 5.502 0.234 0.630 1.953 2.283 4.400 12.875 0.230 0.828 0.680 1.947 O.64O 0.550 0.335 5.546 1.583 1.680 1.614 0.066
I
#M/mg. nitrogen ~ 0.160~ 0.467 1.302 0.132 0.087 0.603 1.376 0.018 0.060 0.306 0.40.8 0.372 1.348 0.045 0.037 0.042 0.380 0.098 0.082 0.022 0.396 0.144 0.070 0.018 0.019
Heterozygote Hetemzygote Normal Normal IIeterozygote Heterozygote Normal Normal Heterozygote Heterozygote Heterozygote Hetemzygote Normal Normal Normal Heterozygote Normal Normal Normal tIeterozygote Normal Normal Normal Normal
e M e a n _+ S.D., f r o m Rosenberg and associates, s t F r o m Scriver. 1~
of the mother, not in association with iminoglycinuria or hyperglycinuria. Both parents of our patient have hyperglycinuria (Table I I I and Fig. 1). Furthermore, family studies show that the hyperglycinuria is inherited as a dominant trait (Fig. 1). Heterozygotes for this defect apparently have isolated hyperglycinuria (only a few exceptions are known ~' 10); homozygotes have iminoglycinuria. Some evidence exists of the presence of more than one renal transport system for glycine and imino acids? 5 One of these is common to all three compounds (system A ) ; two (or more) others are responsible for a selective absorption of glycine and imino acids (system B ) ? ~ In the homozygotes for iminoglycinuria the remaining tubular absorption of glycine and imino acids is probably due to system B, as shown by the inability of high blood levels of proline to decrease further the renal tubular absorption of glycine in these subjects? ~ In our patient the diet rich in gelatin
depressed the tubular absorption of the imino acids more than that of glycine. Both observations support the hypothesis of the existence of independent renal transport systems for glycine and imino acids (system B). In the heterozygotes, the remaining activity of system A appears to be sufficient, at normal Mood amino acid concentrations, to absorb all of the filtered proline and hydroxyproline but only part of the glycine. Studies of the fecal excretion of imino acids and of blood levels of proline and glycine after oral tolerance tests show in our patient, as in 3 others, 8' ~0 that the renal defect is not associated with" impairment of intestinal transport of glycine and imino acids. In one of these patients s uptake of glycine-2-C~4 by intestinal biopsies~was found to be normal. In two other patients 6' ~ the intestinal transport of glycine (in one of these) and of imino acids (in both) was impaired. If these results are confirmed, it should be possible to distinguish different forms of iminoglycinuria, depending on
392
Tancredi, Guazzi, and Auricchio
whether the renal transport defect occurs alone or together with a n intestinal transport defect, as has been observed in cystinuria. ~o SUMMARY
R e n a l iminoglycinuria was detected in a 12-year-old girl affected by congenital achromatopsia with severe amblyopia. Both parents a n d other relatives have renal hyperglycinuria. Family studies support the following conclusions: (1) R e n a l iminoglycinuria is observed in homozygotes for a m u t a t i o n which in heterozygotes causes isolated hyperglycinuria; (2) T h e association between ocular disease a n d the renal defect is probably fortuitous. I n this p a t i e n t the intestinal absorption of glycine a n d imino acids seems to be normal.
The Journal of Pediatrics March 1970
7.
8.
9. 10.
11.
12. 13,
We are greatly indebted to Dr. F. Ponte, Department of Ophthalmology, University of Palermo, Italy, for the ophthalmic and ERG studies and for assessment of the diagnosis of congenital a~hromatopsia. REFERENCES
1. Scriver, C, R., Effort, M. L., and Schafer, I. A.: Renal tubular transport of proline, hydroxyproline and glycine in health and in familial hyperprolinemia, J. Clin. Invest. 43" 374, 1964. 2. Scriver, C. R., and Wilson, O. H.: Possible locations for common gene product in membrane transport of iminoacids and glycine, Nature 202= 92, 1964. 3. Seriver, C. R., and Goldman, I-t.: Renal tubular transport of proline, hydroxyproline and glycine. II. Hydroxy-L-proline as substrate and as inhibitor in vivo, J. Clin. Invest. 45: 1357, 1966. 4. Wilson, O. H., and Scriver, C. R.: Specificity of transport of neutral and basic amino acids in rat kidney, Amer. J. Physiol. 213: 185, 1967. 5. Tada, K., Morikawa, T., Ando, T., Yoshida, T., and Minagawa, A.: Prolinuria: a new renal tubular defect in transport of proline and glycine, Tohoku J. Exp. Med. 87: 133, 1965. 6. Morikawa, T., Tada, K., Ando, T., Yosbida, T., Yokoyama, Y., and Arakawa, T.: Prolinurla: defect in intestinal absorption of
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iminoacids and glycine, Tohoku J. Exp. Med. 90: 105, 1966. Goodman, S. I., Mclntyre, C. A., Jr., and O'Brien, D.: Impaired intestinal transport of proline in patient with familial iminoaciduria, J. P~I)IAT. 71: 246, 1967. Rosenberg, L. E., Durant, J. L., and Elsas, L. J.: Familial imin0glycinuria: An inborn error of renal tubular transport, New Eng. J. Med. 978: 1407, 1968. Whelan, D. T., and Scriver, C. R.: Cystathioninuria and renal iminoglycinuria in a pedigree, New Eng. J. M. 278: 924, 1968. Scriver, C. R.: Renal tubular transport of proline, hydroxyproline and glycine. III. Genetic basis for more than one mode of transport in human kidney, J. Clin. Invest. 47: 823, 1968. Spackman, D. I-t., Stein, W. H., and Moore, S.: Automatic recording apparatus for use in the chromatography of amino acids, Anal. Chem. 30: 1190, 1958. Stein, W. H., and Moore, S.: The free amino acids of human blood plasma, J. Biol. Chem. 211: 915, 1954. Owen, S. A., Iggo, B., Scandrett, F. J., and Stewart, C. P.: The determination of creatinine in plasma or serum and in urine: A critical examination, Biochem. J. 58: 426, 1954. Scriver, C. R., and Davies, E.: Endogenous renal clearance rates of free aminoacids in pre-pubertal children, Pediatrics 36" 592, 1965. Scrlver, C. R., and Wilson, O. H.: Aminoacid transport: Evidence for genetic control of two types in human kidney, Science 155: 1428, 1967. Spivey, B. E., Pearlman, J. T., and Burian, I~. M.: Electroretinographie findings (including flicker) in carriers of congenital Xlinked achromatopsia, Doc. Ophthal. 18: 367, 1964. Krill, A. E., and Schneiderman, A.: Retinal function studies, including the electroretlnogram, in an atypical monochromat, In Clinical electroretinography, Suppl. Vision Res., New York, 1966, Oxford Press, p. 351. Franceschetti, A., Francois, J., and Babel, J.: In Les h6rddo-d6gdn6rescenees chorio-rgtiniennes, vol. II, Paris, 1963, Mas.son & Cie, p. 1252. Rosenberg, L. E., Downing, S., Durant, J. L., and Segal, S. : Cystlnuria : biochemical evidence for three genetically distinct .diseases, J. Clin. Invest. 45: 365, 1966. Dickinsori, S. C., Rosenblum, H., and Hamilton, P. B.: Ion-exchange chromatography of free amino acids in the plasma of the newborn infant, Pediatrics 36" 2, 1965.