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A " NEW " DISORDER OF ISOLEUCINE CATABOLISM
1289
On hypothesis B, however, the frequency would be less than in the population if the value of p was <0-5 and greater for p >05. However, as noted above, it is clear that, if there is a contribution from maternal/fetal incompatibility to toxaemia, there must be a series of loci at which antigenic determinants occur. In addition, there must be important environmental factors as well as multigenic modification of the expression of the major genes which determine the
antigens. Twin Types and Pre-eclamptic Toxaemia It is difficult to accumulate sufficient data to study relationships of toxaemia of pregnancy and consanguinity except in countries where inbreeding is common. Further, studies of frequency of toxaemia in specified relatives or relatives by marriage of a mother raise diagnostic problems, since the acceptance of toxaemia in relatives retrospectively has to be on hearsay evidence, or on records from different hos-
pitals, which can be misleading. However, twin studies are free of these disadvantages because only the diagnosis in the index patient is needed. On either single-gene hypothesis A or B the frequency of toxaemia should be greater in unlike-sex than in likesex twin pregnancies, because the unlike-sex twins will all be dizygous (Dz). The difference would be greater if it was possible to identify zygosity in the twins or to estimate the frequency of the zygosity. In monozygous (MZ) twins the frequency of toxaemia would be the same as for single pregnancies. Expressions for hypothesis A for MZ twins are as for single births pq2, and for DZ twins pq 2(1--l ). For hypothesis B, for MZ twins again as for single births pq2, and for DZ twins pq—iq). Table III shows data assembled from four sources: the frequency in unlike-sex twins is consistently higher than in like-sex twins, although the differences are of borderline significance. However, when estimates are made of the frequency of toxaemia in MZ and DZ twins the differsignificant. Although zygosity frequencies are only approximations, the large of differences are suggestive. There is significances no difference in the sex ratio of the offspring of toxxmic pregnancies, and it is difficult to think of another hypothesis than incompatibility which would explain the twin data. When 20,000-25,000 records are available the situation should be further clarified. Among other calculations which will be possible will be the regression of the frequency of toxaemia on the coefficient of inbreeding from consecutive births in the Dogum Evi Hospital, Ankara. ences are
estimated
so
-
Requests for reprints should be addressed to A. C. S. REFERENCES 1.
2. 3 4.
5. 6. 7. 8.
9.
Stevenson, A. C., Johnston, H. J., Stewart, M. I. P., Golding, D. R. Bull Wld Hlth Org. 1966, 34, suppl. Einen, M. A., Toppozada, H. K. Br. J. prev. soc. Med. 1966, 20, 176. Hellman, L. M., Pritchard, J. A. Williams’ Obstetrics; p. 685. New York, 1971. Penrose, L. S. Ann. Eugen. 1946, 13, 141. Platt, R., Stewart, A. E., Emery, E. W. Lancet, 1958, i, 552. Kalmus, H. Ann. Eugen. 1946, 13, 146. Adams, E. M., Finlayson, A. Lancet, 1961, ii, 1373. Humphries, J. A. Bull. Johns Hopk. Hosp. 1960, 107, 271. Chesley, L. C., Annitto, J. E., Cosgrove, R. A. Obstet. Gynec. 1968, 32, 303.
A " NEW " DISORDER OF ISOLEUCINE CATABOLISM
R. S. DAUM O. A. MAMER
P. H. LAMM C. R. SCRIVER deBelle Laboratory of Biochemical Genetics, McGill Uni-
versity-Montreal Children’s Hospital Research Institute; Mass-Spectrometry Unit, Royal Victoria Hospital; and Biology Department (Human Genetics Sector), Faculty of Science, McGill University A disorder of isoleucine catabolism was found in a child with intermittent metabolic acidosis. The block occurs at the stage of propionate synthesis, and two intermediates—&agr;methylacetoacetate and &agr;-methyl-&bgr;-hydroxybutyrate— accumulate. The condition is apparently inherited.
Summary
INTRODUCTION
THIS report introduces another, apparently hereditary, disorder of isoleucine catabolism. Earlier reports
have described several forms of impaired isoleucine oxidation at the first (transamination)and second (decarboxylation) 2-5 steps in its catabolism. The new disorder affects the sixth step, responsible for conversion of oc-methylacetoacetate to propionate. As with many of the other inborn errors of branchedchain aminoacid catabolism, the presenting clinical feature in our proband was recurrent severe metabolic acidosis. CASE-REPORT
The proband is the 6-year-old son of two unrelated Dutch parents. He was breast-fed through much of his infancy, and his first admission to hospital was at 17 months of age, when he became comatose following an upper respiratory infection accompanied by fever and anorexia. Severe metabolic acidosis was demonstrated which gradually disappeared as the intercurrent infection subsided and corrective intravenous therapy was given. Two further admissions at 34 and 48 months of age for a similar course of events are recorded. Organic aciduria was then diagnosed. Thereafter three mild transitory episodes of metabolic acidosis were effectively countered in the 5th year of life by dietary management, comprising low-protein intake (2 g. per kg. per day) and maintenance of an adequate calorie intake during the illness. The physical and mental development of the proband are normal. Two other siblings and both parents are healthy. METHODS
Organic acids were extracted from urine by the method of Mamer et al.and the trimethylsilyl derivatives were examined on an LKB-9000 coupled gas-chromatography/I L-isoleucine, L-leucine, and mass-spectrometry unit. L-valine were given separately by mouth, each for 6 consecutive doses, at meal-time (75 mg. per kg. per dose) over a 2-day period. Urine was collected in 8-hour periods before, during, and after the loading tests. Urine samples were obtained, at random and after isoleucine loading, from family members, and examined for organic acids. RESULTS
Urine obtained from the proband during an episode of metabolic acidosis contained large amounts of three unusual silylated derivatives. The excreted amounts of these substances were relatively small between
1290
organic acids in the patient’s were a-methyl-&bgr;-hydroxybutyric acid and fx-methylacetoacetate. The presence of either compound in more than trace amounts is abnormal, and in this patient suggests an impairment in their conSince interconversion of version to propionate. and a-methyl-&bgr;-hydroxybutyrcc-methylacetoacetate ate occurs by an oxidation/reduction step and is analogous to the interconversion of the normal ketone bodies, P-hydroxybutyrate and acetoacetate, this may explain the predominance of the butyrate derivative over the acetoacetate compound in our patient. Although a catabolic pathway for isoleucine has been describedthe authenticity of the final conversion of the tiglyl-CoA through a-methyl-p-hydroxybutyryl-CoA and a-methylacetoacetyl-CoA to propionyl-CoA has not been confirmed in man. That this is the pathway was assumed by analogy with the reaction in animal tissues,7 and because of the close parallel to &bgr;-oxidation of straight-chain fatty acids in The accumulation of a-methyl- &bgr;-hydroxyman. butyrate and a-methylacetoacetate in the patient now confirms that these two substances are formed during
The urine
major,
unusual
isoleucine oxidation. The informative trait which offered this insight appears to be another autosomalrecessive inborn-error of metabolism. Tracing of gas-liquid chromatogram of trimethyl-silyl (T.M.S.) derivatives from urine of proband after a dietary load of L-isoleucine. Peaks 1 and
2, minor normal metabolites; 3, a-methyl-04 and 5, (x-methylacetoacetate. Peaks 4 and 5
hydroxybutyrate ; T.M.s.
represent oc-M.A.A.
enol.
derivatives of the cis and trans forms of the an abnormal amount of the
Hatching indicates
substance.
attacks of acidosis, but could be greatly augmented if the dietary intake of L-isoleucine was specifically increased The (see accompanying figure). " unknown " compounds were shown by gas-liquid chromatography and mass spectrometry to be ocmethyl-p-hydroxybutyric acid and oc-methylacetoacetate, the latter appearing as two silylated derivatives. The concentration of isoleucine and other aminoacids, in plasma and urine, was normal at all times. The parents and one sib had elevated levels of oc-methyl-&bgr;-hydroxybutyric acid in their urine; excretion was further increased after L-isoleucine feeding. Normal subjects excrete only trace amounts of this substance. DISCUSSION
Our patient had a severe metabolic acidosis, which is the common mode of presentation for patients with a hereditary disorder of branched-chain aminoacid metabolism. Perhaps because the isoleucine content of human milk is lower than that of cow’s milk, and because no intercurrent infection apparently provoked a catabolic episode early in life, the first clinical episode of acidosis and coma was delayed until late infancy. When the underlying metabolic disease was identified, a diet restricted in protein content but adequate in calories could be prescribed, particularly during periods of intercurrent infectious illness. This regimen clearly ameliorated the frightening episodes of acidosis and mental disorientation.
We thank Dr. J. C. Crawhall for making available the facilities for mass spectrometry. This work was supported by grants from the Medical Research Council (MT 1085, MA 3719, ME 3720), the Department of National Health and Welfare (604-7-459), the National Genetics Foundation (New York), and the Quebec Network of Genetic Medicine. R. S. D. is a medical student at McGill University.
Requests for reprints may be addressed to C. R. S., Montreal Children’s Hospital, 2300 Tupper Street, Montreal 108, Quebec, Canada. REFERENCES 1.
2. 3. 4. 5.
6. 7.
Jeune, M., Collombel, C., Michel, M., David, M., Guibaud, P., Guerrier, G., Albert, J. Annls Pédiat. 1970, 17, 85. Dancis, J., Levitz, M., Westall, R. G. Pediatrics, Springfield, 1960, 25, 72. Dancis, J., Hutzler, J., Rokkones, T. New Engl. J. Med. 1967, 276, 84. Schulman, J. D., Lustberg, T. J., Kennedy, J. L., Museles, M., Seegmiller, J. E. Am. J. Med. 1970, 49, 118. Scriver, C. R., Mackenzie, S., Clow, C. L., Delvin, E. Lancet, 1971, i, 310. Mamer, O. A., Crawhall, J. C., Tjoa, C. S. Clinica chim. Acta, 1971, 32, 171. Robinson, W. G., Bachhawat, B. N., Coon, M. J. J. biol. Chem. 1956, 218, 391.
INSENSITIVITY OF RESPIRATORY CENTRE TO CARBON DIOXIDE IN THE ENGA PEOPLE OF NEW GUINEA
V. BERAL
D.
J. C. READ Department of Medicine, University of Sydney, Sydney, Australia The ventilatory response to a rising carbon-dioxide (CO2) tension has been determined in healthy New Guinea natives using a new apparatus which was developed for field-work. The response in New Guineans did not vary greatly between individuals, in contrast to the finding in healthy Caucasians. In all twelve New Guinean subjects the indices of response fell in the lowest range of
Sum ary
On hypothesis B, however, the frequency would be less than in the population if the value of p was <0-5 and greater for p >05. However, as noted above, it is clear that, if there is a contribution from maternal/fetal incompatibility to toxaemia, there must be a series of loci at which antigenic determinants occur. In addition, there must be important environmental factors as well as multigenic modification of the expression of the major genes which determine the
antigens. Twin Types and Pre-eclamptic Toxaemia It is difficult to accumulate sufficient data to study relationships of toxaemia of pregnancy and consanguinity except in countries where inbreeding is common. Further, studies of frequency of toxaemia in specified relatives or relatives by marriage of a mother raise diagnostic problems, since the acceptance of toxaemia in relatives retrospectively has to be on hearsay evidence, or on records from different hos-
pitals, which can be misleading. However, twin studies are free of these disadvantages because only the diagnosis in the index patient is needed. On either single-gene hypothesis A or B the frequency of toxaemia should be greater in unlike-sex than in likesex twin pregnancies, because the unlike-sex twins will all be dizygous (Dz). The difference would be greater if it was possible to identify zygosity in the twins or to estimate the frequency of the zygosity. In monozygous (MZ) twins the frequency of toxaemia would be the same as for single pregnancies. Expressions for hypothesis A for MZ twins are as for single births pq2, and for DZ twins pq 2(1--l ). For hypothesis B, for MZ twins again as for single births pq2, and for DZ twins pq—iq). Table III shows data assembled from four sources: the frequency in unlike-sex twins is consistently higher than in like-sex twins, although the differences are of borderline significance. However, when estimates are made of the frequency of toxaemia in MZ and DZ twins the differsignificant. Although zygosity frequencies are only approximations, the large of differences are suggestive. There is significances no difference in the sex ratio of the offspring of toxxmic pregnancies, and it is difficult to think of another hypothesis than incompatibility which would explain the twin data. When 20,000-25,000 records are available the situation should be further clarified. Among other calculations which will be possible will be the regression of the frequency of toxaemia on the coefficient of inbreeding from consecutive births in the Dogum Evi Hospital, Ankara. ences are
estimated
so
-
Requests for reprints should be addressed to A. C. S. REFERENCES 1.
2. 3 4.
5. 6. 7. 8.
9.
Stevenson, A. C., Johnston, H. J., Stewart, M. I. P., Golding, D. R. Bull Wld Hlth Org. 1966, 34, suppl. Einen, M. A., Toppozada, H. K. Br. J. prev. soc. Med. 1966, 20, 176. Hellman, L. M., Pritchard, J. A. Williams’ Obstetrics; p. 685. New York, 1971. Penrose, L. S. Ann. Eugen. 1946, 13, 141. Platt, R., Stewart, A. E., Emery, E. W. Lancet, 1958, i, 552. Kalmus, H. Ann. Eugen. 1946, 13, 146. Adams, E. M., Finlayson, A. Lancet, 1961, ii, 1373. Humphries, J. A. Bull. Johns Hopk. Hosp. 1960, 107, 271. Chesley, L. C., Annitto, J. E., Cosgrove, R. A. Obstet. Gynec. 1968, 32, 303.
A " NEW " DISORDER OF ISOLEUCINE CATABOLISM
R. S. DAUM O. A. MAMER
P. H. LAMM C. R. SCRIVER deBelle Laboratory of Biochemical Genetics, McGill Uni-
versity-Montreal Children’s Hospital Research Institute; Mass-Spectrometry Unit, Royal Victoria Hospital; and Biology Department (Human Genetics Sector), Faculty of Science, McGill University A disorder of isoleucine catabolism was found in a child with intermittent metabolic acidosis. The block occurs at the stage of propionate synthesis, and two intermediates—&agr;methylacetoacetate and &agr;-methyl-&bgr;-hydroxybutyrate— accumulate. The condition is apparently inherited.
Summary
INTRODUCTION
THIS report introduces another, apparently hereditary, disorder of isoleucine catabolism. Earlier reports
have described several forms of impaired isoleucine oxidation at the first (transamination)and second (decarboxylation) 2-5 steps in its catabolism. The new disorder affects the sixth step, responsible for conversion of oc-methylacetoacetate to propionate. As with many of the other inborn errors of branchedchain aminoacid catabolism, the presenting clinical feature in our proband was recurrent severe metabolic acidosis. CASE-REPORT
The proband is the 6-year-old son of two unrelated Dutch parents. He was breast-fed through much of his infancy, and his first admission to hospital was at 17 months of age, when he became comatose following an upper respiratory infection accompanied by fever and anorexia. Severe metabolic acidosis was demonstrated which gradually disappeared as the intercurrent infection subsided and corrective intravenous therapy was given. Two further admissions at 34 and 48 months of age for a similar course of events are recorded. Organic aciduria was then diagnosed. Thereafter three mild transitory episodes of metabolic acidosis were effectively countered in the 5th year of life by dietary management, comprising low-protein intake (2 g. per kg. per day) and maintenance of an adequate calorie intake during the illness. The physical and mental development of the proband are normal. Two other siblings and both parents are healthy. METHODS
Organic acids were extracted from urine by the method of Mamer et al.and the trimethylsilyl derivatives were examined on an LKB-9000 coupled gas-chromatography/I L-isoleucine, L-leucine, and mass-spectrometry unit. L-valine were given separately by mouth, each for 6 consecutive doses, at meal-time (75 mg. per kg. per dose) over a 2-day period. Urine was collected in 8-hour periods before, during, and after the loading tests. Urine samples were obtained, at random and after isoleucine loading, from family members, and examined for organic acids. RESULTS
Urine obtained from the proband during an episode of metabolic acidosis contained large amounts of three unusual silylated derivatives. The excreted amounts of these substances were relatively small between
1290
organic acids in the patient’s were a-methyl-&bgr;-hydroxybutyric acid and fx-methylacetoacetate. The presence of either compound in more than trace amounts is abnormal, and in this patient suggests an impairment in their conSince interconversion of version to propionate. and a-methyl-&bgr;-hydroxybutyrcc-methylacetoacetate ate occurs by an oxidation/reduction step and is analogous to the interconversion of the normal ketone bodies, P-hydroxybutyrate and acetoacetate, this may explain the predominance of the butyrate derivative over the acetoacetate compound in our patient. Although a catabolic pathway for isoleucine has been describedthe authenticity of the final conversion of the tiglyl-CoA through a-methyl-p-hydroxybutyryl-CoA and a-methylacetoacetyl-CoA to propionyl-CoA has not been confirmed in man. That this is the pathway was assumed by analogy with the reaction in animal tissues,7 and because of the close parallel to &bgr;-oxidation of straight-chain fatty acids in The accumulation of a-methyl- &bgr;-hydroxyman. butyrate and a-methylacetoacetate in the patient now confirms that these two substances are formed during
The urine
major,
unusual
isoleucine oxidation. The informative trait which offered this insight appears to be another autosomalrecessive inborn-error of metabolism. Tracing of gas-liquid chromatogram of trimethyl-silyl (T.M.S.) derivatives from urine of proband after a dietary load of L-isoleucine. Peaks 1 and
2, minor normal metabolites; 3, a-methyl-04 and 5, (x-methylacetoacetate. Peaks 4 and 5
hydroxybutyrate ; T.M.s.
represent oc-M.A.A.
enol.
derivatives of the cis and trans forms of the an abnormal amount of the
Hatching indicates
substance.
attacks of acidosis, but could be greatly augmented if the dietary intake of L-isoleucine was specifically increased The (see accompanying figure). " unknown " compounds were shown by gas-liquid chromatography and mass spectrometry to be ocmethyl-p-hydroxybutyric acid and oc-methylacetoacetate, the latter appearing as two silylated derivatives. The concentration of isoleucine and other aminoacids, in plasma and urine, was normal at all times. The parents and one sib had elevated levels of oc-methyl-&bgr;-hydroxybutyric acid in their urine; excretion was further increased after L-isoleucine feeding. Normal subjects excrete only trace amounts of this substance. DISCUSSION
Our patient had a severe metabolic acidosis, which is the common mode of presentation for patients with a hereditary disorder of branched-chain aminoacid metabolism. Perhaps because the isoleucine content of human milk is lower than that of cow’s milk, and because no intercurrent infection apparently provoked a catabolic episode early in life, the first clinical episode of acidosis and coma was delayed until late infancy. When the underlying metabolic disease was identified, a diet restricted in protein content but adequate in calories could be prescribed, particularly during periods of intercurrent infectious illness. This regimen clearly ameliorated the frightening episodes of acidosis and mental disorientation.
We thank Dr. J. C. Crawhall for making available the facilities for mass spectrometry. This work was supported by grants from the Medical Research Council (MT 1085, MA 3719, ME 3720), the Department of National Health and Welfare (604-7-459), the National Genetics Foundation (New York), and the Quebec Network of Genetic Medicine. R. S. D. is a medical student at McGill University.
Requests for reprints may be addressed to C. R. S., Montreal Children’s Hospital, 2300 Tupper Street, Montreal 108, Quebec, Canada. REFERENCES 1.
2. 3. 4. 5.
6. 7.
Jeune, M., Collombel, C., Michel, M., David, M., Guibaud, P., Guerrier, G., Albert, J. Annls Pédiat. 1970, 17, 85. Dancis, J., Levitz, M., Westall, R. G. Pediatrics, Springfield, 1960, 25, 72. Dancis, J., Hutzler, J., Rokkones, T. New Engl. J. Med. 1967, 276, 84. Schulman, J. D., Lustberg, T. J., Kennedy, J. L., Museles, M., Seegmiller, J. E. Am. J. Med. 1970, 49, 118. Scriver, C. R., Mackenzie, S., Clow, C. L., Delvin, E. Lancet, 1971, i, 310. Mamer, O. A., Crawhall, J. C., Tjoa, C. S. Clinica chim. Acta, 1971, 32, 171. Robinson, W. G., Bachhawat, B. N., Coon, M. J. J. biol. Chem. 1956, 218, 391.
INSENSITIVITY OF RESPIRATORY CENTRE TO CARBON DIOXIDE IN THE ENGA PEOPLE OF NEW GUINEA
V. BERAL
D.
J. C. READ Department of Medicine, University of Sydney, Sydney, Australia The ventilatory response to a rising carbon-dioxide (CO2) tension has been determined in healthy New Guinea natives using a new apparatus which was developed for field-work. The response in New Guineans did not vary greatly between individuals, in contrast to the finding in healthy Caucasians. In all twelve New Guinean subjects the indices of response fell in the lowest range of
Sum ary