574
October 1977 TheJournalofPEDIATRICS
Free amino acids in liver of patients with homocystinuria due to cystathionine synthase deficiency: Effects of vitamin Patients with homocystinuria due to cystathionine synthase deficiency do not have free homocystine in the liver when it is present in high concentrations in the plasma and the urine. The liver of these patients is capable of maintaining normal concentrations of cystine at a time when the plasma cystine concentration is severely reduced There is an increase in the methionine concentration of the liver which is reduced to normal concentrations during pyridoxine therapy.
David K. Rassin, Ph.D,,* Riceardo C. Longhi, M.D., and Gerald E. Gaull, M.D., S t a t e n I s l a n d a n d N e w Y o r k , N. Y.
DEFICIENCY of cystathionine synthase 1 is the most common etiology of the clinical signs and symptoms associated with the excretion of homocystine in the urine. ~. "~In some patients treatment with massive doses of pyridoxine ameliorates the biochemical abnormalities in the plasma and urine, i.e. increased methionine, easily measurable homocystine, and decreased or absent cystine. '-~ The clinical results of pyridoxine therapy are not clear as yet, for some abnormalities may persist or progress, e.g. ectopia lentis 9 19 and ultrastructural abnormalities of the hepatocytes. 1~ ~ Although the mechanism for the biochemical effects of pyridoxine is not clear, a persistent increase in activity of hepatic synthase 1~ 12 has been demonstrated in some, but not all cases responsive to B6. Pyridoxal phosphate, the coenzyme form of the B6 vitamer pyridoxine, protects cystathionine synthase from thermal inactivation in vitro both in liver ~2.13 and in skin From the Department o f Human Development and Genetics, Institute for Basic Research in Mental Retardation, and Department of Pediatrics and Clinical Genetics Center, Mount Sinai School of Medicine of the City University o f New York. Supported by the New York State Department of Mental Hygiene, National Institutes o f Health Clinical Genetics Center Grant GM-19443 and N I H Grant 00071. *Reprint address: Department of Human Development and Genetics, Institute for Basic Research in Mental Retardation, 1050 Forest Hill Road. Staten Island, NY 10314.
Vol. 91, No. 4, pp. 574-577
fibroblasts.14.15 Evidence suggesting a direct relationship between pyridoxine administration and a change in the biochemical behavior of hepatic cystathionine synthase has been presented. 1~ The effects of pyridoxine treatment on the concentration of amino acids in the liver, however, are not known. Therefore, the concentrations of free amino acids were measured in liver of patients with homocystinuria due to cystathionine synthase deficiency before and during pyridoxine therapy and are compared with hepatic amino acid concentrations in patients without this enzymatic deficiency. METHODS Liver from research subjects was obtained by use of a Menghini needle; those used as controls were part of a biopsy specimen obtained for other clinical indications. The specimens were washed briefly with saline, blotted, weighed, and then homogenized with 10% trichloroacetic acid in a glass-giass homogenizer. Supernatant solution was analyzed on an automatic amino acid analyzer and quantified by an on-line computer as previously described? ~ Plasma samples were precipitated with 3% sulfosalicyclic acid, and urine samples were acidified with hydrochloric acid before analysis on a Technicon TSM amino acid analyzer. The three subjects studied were two clinically typical brothers (D.B. and R.B. Jr.), who have vitamin B6-responsive cystathionine synthase deficiency (see Table II), and their father (R.B. Sr.) an obligate
Volume 91 Number 4
heterozygote. The enzymatic data have been presented in detail elsewhere? 3 Written informed consent was obtained for all research procedures, and the experimental protocols and consent forms were reviewed by the hospital's Human Investigations Committee. RESULTS Concentrations of the sulfur containing amino acids in the plasma and their rates of excretion in the urine of the two affected sibs were typical for cystathionine synthase deficiency (Table I): methionine, homocystine, and the mixed disulfide of homocysteine and cysteine were high, and cystine was low. These abnormalities returned to normal on a regimen of 250 mg pyridoxine HC1 per day. The concentrations of amino acids in the plasma and their rates of excretion in the urine of the obligate heterozygote were essentially normal both prior to and during administration of pyridoxine. The concentrations of methionine and its metabolites in the liver of the patients reflects only partially the concentrations of these amino acids observed in the plasma and urine on the same day (Table II). Prior to treatment with pyridoxine, the concentration of free methionine in the liver was increased; however, homocystine was not observed, there was no decrease in concentration of cystine, and no mixed disulfide of cysteine and homocysteine was observed. During treatment with pyridoxine, the only significant change was a decrease to normal in the methionine concentration of the affected sibs. The concentrations of the free amino acids in the liver of the obligate heterozygote were within normal limits both before and during administration of pyridoxine. DISCUSSION The present results differ from those in one previous repoW 7 in that we could not identify homocystine in the liver, even though the specimens of liver were homogenized with TCA promptly after biopsy. Homocystine was not identified in tissues obtained at autopsy from two patients with homocystinuria due to cystathionine synthase deficiency, TM or of one patient with homocystinuria due to 5,10-methylenetetrahydrofolate reductase deficiency." We osberved a small unknown peak on the amino acid chromatogram at an elution time close to, but not that of, homocystine, but not enough material was available to allow identification of this compound. We occasionally see a compound in some tissue and urine specimens from patients without synthase deficiency, as well as from normals, which is eluted near the position of homocystine but is not homocystine. Insufficient details are reported by Tada and co-workers '7 to determine
Liver amino acids in homocystinuria
575
Table I. Plasma concentration and urinary excretion of sulfur amino acids at the time of liver biopsy Patient D.B.
Patient R.B. Jr.
Father R.B. Sr.
--Be I + B~
-Bo I+Bo
--B,~ ] + B,~ ]
Plasma (~moles %) Taurine 11.0 6.7 Cystine 1.4 10.0 Methionine 44.2 3.6 Homocystine 10.9 0 Mixed 0.8 0 disulfide Methionine 4.0 0 sulfoxide Urine (#moles excreted/hr) Taurine 38.3 41.0 Cystine 0.6 4.5 Methionine 6.5 1.0 Homocystine 11.0 0 Mixed 2.3 0.2 disulfide Methionine 2.4 0 sulfoxide
8.5 2.4 17.8 6.7 1.6
7.2 11.9 3.6 0 0
5.3 9.3 4.4 0 0
6.4 10.0 3.2 0 0
2.1
0
0
0
42.! 1.! 6.0 17.5 4.0
27.8 4.3 1.0 0 0.1
10.8 2.6 1.0 0 0
14.8 4.3 0.9 0 0
2.0
0
0
0
All samplesof urineand plasmawere taken on the day of the liver biopsy with the exceptionof the plasmaof Father R.B. St. -B~; this sample was taken fivedays before the biopsywas performed. whether homocystine was identified by chromatographic position alone. Spaeth and Barber ~~showed that the ratio of erythrocyte methionine to plasma methionine was increased but that this was not true for homocystine. It is possible that homocystine is present in the liver in the form of an enlarged pool of S-adenosylhomocysteine. The equilibrium of the enzymatic hydrolysis of S-adenosylhomocysteine to homocysteine and adenosine is strongly in the direction of synthesis of S-adenosylhomocysteine?1 It is possible, therefore, that homocystine is present in the tissues of the patients and heterozygotes as an enlarged pool of S-adenosylhomocysteine and that it is S-adenosylhomocysteine which is responsible for the ultrastructural abnormalities found in the liver of both the treated patient and the obligate heterozygote. 1..... Also, we did not rule out the presence of homocysteine in the form of a mixed disulfide with liver proteins. A compound with properties similar to homocysteine has been found to be bound to plasma proteins, and it has been suggested that homocysteine may bind to other body proteins. ~ In summary, in affected patients with deficiency of cystathionine synthase there is an increase in the methionine concentration of the liver which appears to be reduced to normal by pyridoxine therapy. Furthermore, the liver is capable of maintaining normal concentrations of cystine in the liver at a time when the plasma
576
Rassin, Longhi, and Gaull
The Journal of Pediatrics October 1977
Table II, Free a m i n o acids in the liver
Patient D.B. (~mol/gm)
Cystathionine synthase* Cystine Methionine Homocystine Mixed disulfide~ Aspartate Threonine Serine Glutamine and asparagine Glutamate Glycine Alanine Valine Isoleucine Leucine Tyrosine Phenylalanine Taurine
Patient R.B. Jr. (pmol/gm)
--B~,
+ B,~
-B,~
4.0 0.28 0.43 0 0 1.10 0.73 1.49 2.32 3.45 1.66 0.80 0.37 0.90 0.25 0.10 0.90 0.75
8.4 0.66 0.02 0 0 0.85 2.76 2.28 0.83 4.83 1.46 1.18 0.34 0.16 0.16 0.08 0.07 0.20
6.2 1.79 0.20 0 0 1.05 1.58 1.00 2.86 4.64 2.52 1.40 0.75 0.88 0.84 0.31 0.18 0.58
[
Father R.B. Sr. (pmol /gm )
+ B,~
--B,~
10.7 1.23 0.08 0 0 0.91 3.67 0.93 2.51 3.73 2.01 1.39 0.30 0.02 0.56 0.16 0.11 0.54
65.3 1.09 0.07 0 0 0.40 2.70 0.36 1.63 2.52 1.26 0.69 0.47 0.14 0.28 0.23 0.12 0.56
I
+ B~,
Controls (n = 3) Mean +_ SEM
88.4 0.41 0.04 0 0 0.68 4.23 0.53 2.82 3.97 1.75 1.92 0.37 0!16 0.33 0.12 0.03 0.75
100 0.30 _+ 0.05 0.07 +_ 0.003 0 0 1.52 _+ 0.44 1.05 _+ 0.25 0.93 +_ 0.21 2,98 _+ 0.77 4.76 +_ 1.15 1.78 +_ 0.21 2.56 +_ 1.03 0.34 +_ 0.07 0.12 +_ 0.02 0.33 +_ 0.09 0.18 • 0.06 0.15 +_ 0.03 1.38 _+ 0.46
*Cystathioninesynthase is expressed as a percent of the mean control specific activity of the enzyme (mean control specific activity [n = 5] was 225 nanomoles cystathionine formed/mg protein/hour). These data have been presented in detail elsewhere~~but are included for convenient reference. Patient R.B. Jr. is patient 2, Patient D.B. is patient 3 and R.B. Sr. is the father?:' tThis is the mixed disulfide of cysteine and homocysteine (Cys-S-S-Hcys). c o n c e n t r a t i o n of cystine is severely reduced. Finally, free homocystine is not present in the liver at a time that it is present in high c o n c e n t r a t i o n s in the p l a s m a a n d urine, but it is possible t h a t it is p r e s e n t in the form o f Sadenosylhomocysteine or as a mixed disulfide with cysteine residues of proteins. The authors are grateful to Drs. M. Pastrana and I. Krasna for obtaining the liver biopsies. The expert technical assistance of Nevenka Vukovic and Lucille Donadio is appreciated. REFERENCES
1. Mudd SH, Finkelstein JD, Irreverre F, and Laster L: Homocystinuria: An enzymatic defect, Science 143:1433~ 1964. 2. Carson NAJ, Dent CE, Field CMB, and Gaull GE: HomocystinOria: Clinical and pathological review of ten cases, J PEDIATR 66:565, 1965. 3. Schimke RN, McKusick VA, Huang T, and Pollack AD: Homocystinuria: Studies of 20 families with 38 affected members, JAMA 193:711, 1965. 4. Barber GW, and Spaeth GL: Pyridoxine therapy in homocystinuria, Lancet 1:337, 1967. 5. Hooft C, Carton D, and Samyn W: Pyridoxine treatment in homocystinuria, Lancet 1:1384, 1967. 6. Turner B: Pyridoxine treatment in homocystinuria, Lancet 2:1151, 1967: 7. Cusworth DC, and Dent CE: Homocystinuria, Br Med Bull 25:42, 1969.
8. Gaull GE, Rassin DK, and Sturman JA: Enzymatic and metabolic studies of homocystinuria, Neuropadiatrie 1:199, 1970. 9. Hagberg B, Hambraeus L, and Bensch K: A case of homocystinuria with a dystonic neurological syndrome, Neuropadiatrie 1:337, 1970. 10. Gaull GE; Sturman JA, and Schaffner F: Homocystinuria due to cystathionine synthase deficiency: Enzymatic and ultrastructural studies, J PEDIATR 84:381, 1974. 11. Gaull GE, and Schaffner F: Electron microscopic changes in hepatocytes of patients with homocystinuria, Pediatr Res 5:23, 1971. 12. Mudd SH, Edwards WA, Loeb PM, Brown MS, and Laster LJ: Homocystinuria due to cystathionine synthase deftciency: The effect of pyridoxine, J Clin Invest 49:1762, 1970. 13. Longhi RC, Fleisher LD, Tallan HH, and Gaull GE: Cystathionine /3-synthase deficiency: A qualitative abnormality of the enzyme modified by vitamin B6 therapy, Pediatr Res 11:100, 1977. 14. Kim YJ, and Rosenberg LE: On the mechanism of pyridoxine responsive homocystinuria. II. Properties of normal and mutant cystathionine fi-synthase from cultured fibroblasts, Proc Natl Acad Sci 71:4821, 1974. 15. Fleisher LD, Longhi RC, Tallan HH, and Gaull GE: Cystathionine /3-synthase deficiency: Differences in thermostability between normal and abnormal enzyme from cultured human cells, Pediatr Res (in press). 16. Sturman JA, and Gaull GE: Taurine in the brain and liver of the developing human and monkey, J Neurochem 25:831, 1975. 17. Tada K, Yoshida T, and Arakawa T: Free amino acid
Volume 91 Number 4
pattern in the liver from the patients with amino acid disorders: Postmortem diagnosis of inborn errors of amino acid metabolism, Tohoku J Exp Med 101:223, 1970. 18. Brenton DP, Cusworth DC, and Gaull GE: Homocystinuria. Biochemical studies of tissues including a comparison with cystathioninuria, Pediatrics 35:50, 1965. 19. Kanwar YS, Manaligod JR, and Wong PWK: Morphologic studies in a patient with homocystinuria due to 5,10methylentetrahydrofolate reductase deficiency, Pediatr Res 10:598, 1976.
Liver amino acids in homocystinuria
20.
21. 22.
577
Spaeth GL, and Barber GW: Homocystinuria in a mentally retarded child and her normal cousin, Trans Am Acad Ophthal Otolaryngol 912:69, 1965. de la Haba G, and Cantoni GL: Enzymatic synthesis of S-adenosylhomocysteine, J Biol Chem 234:603, 1959. Brenton DP, and Cusworth DC: Homocystinuria: Metabolism of ~S methionine, Clin Sci 31:197, 1966.
October 1977 TheJournalofPEDIATRICS
Free amino acids in liver of patients with homocystinuria due to cystathionine synthase deficiency: Effects of vitamin Patients with homocystinuria due to cystathionine synthase deficiency do not have free homocystine in the liver when it is present in high concentrations in the plasma and the urine. The liver of these patients is capable of maintaining normal concentrations of cystine at a time when the plasma cystine concentration is severely reduced There is an increase in the methionine concentration of the liver which is reduced to normal concentrations during pyridoxine therapy.
David K. Rassin, Ph.D,,* Riceardo C. Longhi, M.D., and Gerald E. Gaull, M.D., S t a t e n I s l a n d a n d N e w Y o r k , N. Y.
DEFICIENCY of cystathionine synthase 1 is the most common etiology of the clinical signs and symptoms associated with the excretion of homocystine in the urine. ~. "~In some patients treatment with massive doses of pyridoxine ameliorates the biochemical abnormalities in the plasma and urine, i.e. increased methionine, easily measurable homocystine, and decreased or absent cystine. '-~ The clinical results of pyridoxine therapy are not clear as yet, for some abnormalities may persist or progress, e.g. ectopia lentis 9 19 and ultrastructural abnormalities of the hepatocytes. 1~ ~ Although the mechanism for the biochemical effects of pyridoxine is not clear, a persistent increase in activity of hepatic synthase 1~ 12 has been demonstrated in some, but not all cases responsive to B6. Pyridoxal phosphate, the coenzyme form of the B6 vitamer pyridoxine, protects cystathionine synthase from thermal inactivation in vitro both in liver ~2.13 and in skin From the Department o f Human Development and Genetics, Institute for Basic Research in Mental Retardation, and Department of Pediatrics and Clinical Genetics Center, Mount Sinai School of Medicine of the City University o f New York. Supported by the New York State Department of Mental Hygiene, National Institutes o f Health Clinical Genetics Center Grant GM-19443 and N I H Grant 00071. *Reprint address: Department of Human Development and Genetics, Institute for Basic Research in Mental Retardation, 1050 Forest Hill Road. Staten Island, NY 10314.
Vol. 91, No. 4, pp. 574-577
fibroblasts.14.15 Evidence suggesting a direct relationship between pyridoxine administration and a change in the biochemical behavior of hepatic cystathionine synthase has been presented. 1~ The effects of pyridoxine treatment on the concentration of amino acids in the liver, however, are not known. Therefore, the concentrations of free amino acids were measured in liver of patients with homocystinuria due to cystathionine synthase deficiency before and during pyridoxine therapy and are compared with hepatic amino acid concentrations in patients without this enzymatic deficiency. METHODS Liver from research subjects was obtained by use of a Menghini needle; those used as controls were part of a biopsy specimen obtained for other clinical indications. The specimens were washed briefly with saline, blotted, weighed, and then homogenized with 10% trichloroacetic acid in a glass-giass homogenizer. Supernatant solution was analyzed on an automatic amino acid analyzer and quantified by an on-line computer as previously described? ~ Plasma samples were precipitated with 3% sulfosalicyclic acid, and urine samples were acidified with hydrochloric acid before analysis on a Technicon TSM amino acid analyzer. The three subjects studied were two clinically typical brothers (D.B. and R.B. Jr.), who have vitamin B6-responsive cystathionine synthase deficiency (see Table II), and their father (R.B. Sr.) an obligate
Volume 91 Number 4
heterozygote. The enzymatic data have been presented in detail elsewhere? 3 Written informed consent was obtained for all research procedures, and the experimental protocols and consent forms were reviewed by the hospital's Human Investigations Committee. RESULTS Concentrations of the sulfur containing amino acids in the plasma and their rates of excretion in the urine of the two affected sibs were typical for cystathionine synthase deficiency (Table I): methionine, homocystine, and the mixed disulfide of homocysteine and cysteine were high, and cystine was low. These abnormalities returned to normal on a regimen of 250 mg pyridoxine HC1 per day. The concentrations of amino acids in the plasma and their rates of excretion in the urine of the obligate heterozygote were essentially normal both prior to and during administration of pyridoxine. The concentrations of methionine and its metabolites in the liver of the patients reflects only partially the concentrations of these amino acids observed in the plasma and urine on the same day (Table II). Prior to treatment with pyridoxine, the concentration of free methionine in the liver was increased; however, homocystine was not observed, there was no decrease in concentration of cystine, and no mixed disulfide of cysteine and homocysteine was observed. During treatment with pyridoxine, the only significant change was a decrease to normal in the methionine concentration of the affected sibs. The concentrations of the free amino acids in the liver of the obligate heterozygote were within normal limits both before and during administration of pyridoxine. DISCUSSION The present results differ from those in one previous repoW 7 in that we could not identify homocystine in the liver, even though the specimens of liver were homogenized with TCA promptly after biopsy. Homocystine was not identified in tissues obtained at autopsy from two patients with homocystinuria due to cystathionine synthase deficiency, TM or of one patient with homocystinuria due to 5,10-methylenetetrahydrofolate reductase deficiency." We osberved a small unknown peak on the amino acid chromatogram at an elution time close to, but not that of, homocystine, but not enough material was available to allow identification of this compound. We occasionally see a compound in some tissue and urine specimens from patients without synthase deficiency, as well as from normals, which is eluted near the position of homocystine but is not homocystine. Insufficient details are reported by Tada and co-workers '7 to determine
Liver amino acids in homocystinuria
575
Table I. Plasma concentration and urinary excretion of sulfur amino acids at the time of liver biopsy Patient D.B.
Patient R.B. Jr.
Father R.B. Sr.
--Be I + B~
-Bo I+Bo
--B,~ ] + B,~ ]
Plasma (~moles %) Taurine 11.0 6.7 Cystine 1.4 10.0 Methionine 44.2 3.6 Homocystine 10.9 0 Mixed 0.8 0 disulfide Methionine 4.0 0 sulfoxide Urine (#moles excreted/hr) Taurine 38.3 41.0 Cystine 0.6 4.5 Methionine 6.5 1.0 Homocystine 11.0 0 Mixed 2.3 0.2 disulfide Methionine 2.4 0 sulfoxide
8.5 2.4 17.8 6.7 1.6
7.2 11.9 3.6 0 0
5.3 9.3 4.4 0 0
6.4 10.0 3.2 0 0
2.1
0
0
0
42.! 1.! 6.0 17.5 4.0
27.8 4.3 1.0 0 0.1
10.8 2.6 1.0 0 0
14.8 4.3 0.9 0 0
2.0
0
0
0
All samplesof urineand plasmawere taken on the day of the liver biopsy with the exceptionof the plasmaof Father R.B. St. -B~; this sample was taken fivedays before the biopsywas performed. whether homocystine was identified by chromatographic position alone. Spaeth and Barber ~~showed that the ratio of erythrocyte methionine to plasma methionine was increased but that this was not true for homocystine. It is possible that homocystine is present in the liver in the form of an enlarged pool of S-adenosylhomocysteine. The equilibrium of the enzymatic hydrolysis of S-adenosylhomocysteine to homocysteine and adenosine is strongly in the direction of synthesis of S-adenosylhomocysteine?1 It is possible, therefore, that homocystine is present in the tissues of the patients and heterozygotes as an enlarged pool of S-adenosylhomocysteine and that it is S-adenosylhomocysteine which is responsible for the ultrastructural abnormalities found in the liver of both the treated patient and the obligate heterozygote. 1..... Also, we did not rule out the presence of homocysteine in the form of a mixed disulfide with liver proteins. A compound with properties similar to homocysteine has been found to be bound to plasma proteins, and it has been suggested that homocysteine may bind to other body proteins. ~ In summary, in affected patients with deficiency of cystathionine synthase there is an increase in the methionine concentration of the liver which appears to be reduced to normal by pyridoxine therapy. Furthermore, the liver is capable of maintaining normal concentrations of cystine in the liver at a time when the plasma
576
Rassin, Longhi, and Gaull
The Journal of Pediatrics October 1977
Table II, Free a m i n o acids in the liver
Patient D.B. (~mol/gm)
Cystathionine synthase* Cystine Methionine Homocystine Mixed disulfide~ Aspartate Threonine Serine Glutamine and asparagine Glutamate Glycine Alanine Valine Isoleucine Leucine Tyrosine Phenylalanine Taurine
Patient R.B. Jr. (pmol/gm)
--B~,
+ B,~
-B,~
4.0 0.28 0.43 0 0 1.10 0.73 1.49 2.32 3.45 1.66 0.80 0.37 0.90 0.25 0.10 0.90 0.75
8.4 0.66 0.02 0 0 0.85 2.76 2.28 0.83 4.83 1.46 1.18 0.34 0.16 0.16 0.08 0.07 0.20
6.2 1.79 0.20 0 0 1.05 1.58 1.00 2.86 4.64 2.52 1.40 0.75 0.88 0.84 0.31 0.18 0.58
[
Father R.B. Sr. (pmol /gm )
+ B,~
--B,~
10.7 1.23 0.08 0 0 0.91 3.67 0.93 2.51 3.73 2.01 1.39 0.30 0.02 0.56 0.16 0.11 0.54
65.3 1.09 0.07 0 0 0.40 2.70 0.36 1.63 2.52 1.26 0.69 0.47 0.14 0.28 0.23 0.12 0.56
I
+ B~,
Controls (n = 3) Mean +_ SEM
88.4 0.41 0.04 0 0 0.68 4.23 0.53 2.82 3.97 1.75 1.92 0.37 0!16 0.33 0.12 0.03 0.75
100 0.30 _+ 0.05 0.07 +_ 0.003 0 0 1.52 _+ 0.44 1.05 _+ 0.25 0.93 +_ 0.21 2,98 _+ 0.77 4.76 +_ 1.15 1.78 +_ 0.21 2.56 +_ 1.03 0.34 +_ 0.07 0.12 +_ 0.02 0.33 +_ 0.09 0.18 • 0.06 0.15 +_ 0.03 1.38 _+ 0.46
*Cystathioninesynthase is expressed as a percent of the mean control specific activity of the enzyme (mean control specific activity [n = 5] was 225 nanomoles cystathionine formed/mg protein/hour). These data have been presented in detail elsewhere~~but are included for convenient reference. Patient R.B. Jr. is patient 2, Patient D.B. is patient 3 and R.B. Sr. is the father?:' tThis is the mixed disulfide of cysteine and homocysteine (Cys-S-S-Hcys). c o n c e n t r a t i o n of cystine is severely reduced. Finally, free homocystine is not present in the liver at a time that it is present in high c o n c e n t r a t i o n s in the p l a s m a a n d urine, but it is possible t h a t it is p r e s e n t in the form o f Sadenosylhomocysteine or as a mixed disulfide with cysteine residues of proteins. The authors are grateful to Drs. M. Pastrana and I. Krasna for obtaining the liver biopsies. The expert technical assistance of Nevenka Vukovic and Lucille Donadio is appreciated. REFERENCES
1. Mudd SH, Finkelstein JD, Irreverre F, and Laster L: Homocystinuria: An enzymatic defect, Science 143:1433~ 1964. 2. Carson NAJ, Dent CE, Field CMB, and Gaull GE: HomocystinOria: Clinical and pathological review of ten cases, J PEDIATR 66:565, 1965. 3. Schimke RN, McKusick VA, Huang T, and Pollack AD: Homocystinuria: Studies of 20 families with 38 affected members, JAMA 193:711, 1965. 4. Barber GW, and Spaeth GL: Pyridoxine therapy in homocystinuria, Lancet 1:337, 1967. 5. Hooft C, Carton D, and Samyn W: Pyridoxine treatment in homocystinuria, Lancet 1:1384, 1967. 6. Turner B: Pyridoxine treatment in homocystinuria, Lancet 2:1151, 1967: 7. Cusworth DC, and Dent CE: Homocystinuria, Br Med Bull 25:42, 1969.
8. Gaull GE, Rassin DK, and Sturman JA: Enzymatic and metabolic studies of homocystinuria, Neuropadiatrie 1:199, 1970. 9. Hagberg B, Hambraeus L, and Bensch K: A case of homocystinuria with a dystonic neurological syndrome, Neuropadiatrie 1:337, 1970. 10. Gaull GE; Sturman JA, and Schaffner F: Homocystinuria due to cystathionine synthase deficiency: Enzymatic and ultrastructural studies, J PEDIATR 84:381, 1974. 11. Gaull GE, and Schaffner F: Electron microscopic changes in hepatocytes of patients with homocystinuria, Pediatr Res 5:23, 1971. 12. Mudd SH, Edwards WA, Loeb PM, Brown MS, and Laster LJ: Homocystinuria due to cystathionine synthase deftciency: The effect of pyridoxine, J Clin Invest 49:1762, 1970. 13. Longhi RC, Fleisher LD, Tallan HH, and Gaull GE: Cystathionine /3-synthase deficiency: A qualitative abnormality of the enzyme modified by vitamin B6 therapy, Pediatr Res 11:100, 1977. 14. Kim YJ, and Rosenberg LE: On the mechanism of pyridoxine responsive homocystinuria. II. Properties of normal and mutant cystathionine fi-synthase from cultured fibroblasts, Proc Natl Acad Sci 71:4821, 1974. 15. Fleisher LD, Longhi RC, Tallan HH, and Gaull GE: Cystathionine /3-synthase deficiency: Differences in thermostability between normal and abnormal enzyme from cultured human cells, Pediatr Res (in press). 16. Sturman JA, and Gaull GE: Taurine in the brain and liver of the developing human and monkey, J Neurochem 25:831, 1975. 17. Tada K, Yoshida T, and Arakawa T: Free amino acid
Volume 91 Number 4
pattern in the liver from the patients with amino acid disorders: Postmortem diagnosis of inborn errors of amino acid metabolism, Tohoku J Exp Med 101:223, 1970. 18. Brenton DP, Cusworth DC, and Gaull GE: Homocystinuria. Biochemical studies of tissues including a comparison with cystathioninuria, Pediatrics 35:50, 1965. 19. Kanwar YS, Manaligod JR, and Wong PWK: Morphologic studies in a patient with homocystinuria due to 5,10methylentetrahydrofolate reductase deficiency, Pediatr Res 10:598, 1976.
Liver amino acids in homocystinuria
20.
21. 22.
577
Spaeth GL, and Barber GW: Homocystinuria in a mentally retarded child and her normal cousin, Trans Am Acad Ophthal Otolaryngol 912:69, 1965. de la Haba G, and Cantoni GL: Enzymatic synthesis of S-adenosylhomocysteine, J Biol Chem 234:603, 1959. Brenton DP, and Cusworth DC: Homocystinuria: Metabolism of ~S methionine, Clin Sci 31:197, 1966.
