- Email: [email protected]
IMMUNOLOGICAL ASPECTS OF EXPERIMENTAL PULMONARY SCLEROSIS DUE TO IONISING RADIATIONS
1397 these findings, since I believe that the position regarding the use of tannic acid should be put in proper perspective. Diagnostic Radiology Department, St. Bartholomew’s Hospital, London, E.C.1.
R. A. KEMP HARPER.
IMMUNOLOGICAL ASPECTS OF EXPERIMENTAL PULMONARY SCLEROSIS DUE TO IONISING RADIATIONS
SIR,-Both in man and in animals high doses of ionising radiations applied to the chest may give rise to pulmonary fibrosis.! The cause is still unknown, but some aspects of the lesions suggest the presence of immunological factors.2 Eight rabbits, weighing 2-2-5 kg., were irradiated with telecobalt-therapy " (f.p.d. 50 cm.) limited to the right hemithorax, 4900 rad over a 7-day period. 3 months later, six rabbits were exposed at the same site to a second cycle of 5000 rad, given over 5 days. The rabbits were bled 2-3 months after each cycle. Fragments of the left lung (non-irradiated) and the right (irradiated) lung from each rabbit were fixed in formalin 10% and acetone, and embedded in paraffin. Sections were examined microscopically and histochemically. Lung tissue from the same rabbits and from non-irradiated rabbits, and liver and kidney tissue from treated and untreated rabbits, were freeze-dried and embedded in paraffin (in vacuo); the sections, from which the paraffin was removed with petrol ether, were either fixed or not with acetone, and finally tested with the following: (1) fluorescein-isothiocyanate (F.I.T.C.)labelled y-globulins, isolated by salting from the serum of the same animals obtained before the first irradiation; (2) F.LT.C.labelled y-globulins, isolated as before from the serum of the same animals obtained 2 months after the last irradiation; (3) F.I.T.c.-labelled anti-rabbit globulins from sheep serum; and (4) human complement and F.I.T.c.-tagged anti-human complement from sheep serum. Further controls were applied using unlabelled y-globulins before the fluorescent serum and heat-inactivated complement. After the skin of the left flank was shaved, two rabbits treated with 2 cycles of radiations, and four untreated rabbits of the same weight, were injected under sterile conditions with (1) 1 ml. of suspension of finely homogenised irradiated rabbitlung in saline solution; (2) 1 ml. of suspension of untreated homogenised rabbit-lung; (3) 1 ml. of untreated homogenised rabbit-liver; and (4) 1 ml. of saline solution. The rabbits were
Fig. 3-Rabbit treated with 9900 rad. (Haematoxylin and eosin. x 80, reduced by half.) Cutaneous response 60 hours after injection with irradiated lung homogenate. Around the injected unabsorbed material, extensive inflammatory reaction of the dermis with perivascular infiltrates of lymphomonocytes and eosinophil cells, and lesions of the vessel walls. Fig. 4-Lung of rabbit 6 days after injection with immunopotent cells of irradiated rabbits. (Haematoxylin and eosin. x 200, reduced by half.) Lymphomonocytes and eosinophil cells infiltrate the alveolar septa
diffusely. under continuous observation for 5 days, and skin changes were photographed. The cutaneous injection sites were removed, and histologically examined. Two untreated rabbits, weighing 2-2-5 kg., after shaving of the abdomen were injected intraperitoneally with 2-5 ml. of a heavy suspension (in saline solution) of homogenised spleen from two irradiated rabbits. 24 hours later, 0-5 ml. of the same suspensions and solutions already mentioned were injected subcutaneously and the injection sites were examined. The rabbits were bled 6 days after the intraperitoneal injection, and the lungs were microscopically examined.
All rabbits showed thickened periodic-acid/Schiffpositive, alveolar septa infiltrated by lymphomonocytes and eosinophil cells, with pronounced reticular proliferation ; the elastic framework was often disrupted. The small-vessel walls were swollen by plasma-like material, and showed many altered or proliferating endothelial cells. In three of the six rabbits given 2 cycles of irradiation there large zones of septal and vascular fibrosis, pronounced changes in the alveolar cells, reticulin and collagen production, and sclero-calcined nodules (fig. 1). F.l.T.c.-tagged y-globulins, obtained from two rabbits long after the irradiation, were specifically fixed to the alveolar and septal cells of the treated or untreated lungs of the same or other rabbits, but were not fixed in sections of other organs. The tagged y-globulins, obtained from the same rabbits before the first irradiation, were not fixed in sections of the lung or other organs. In the irradiated rabbits, whose y-globulins were fixed in the pulmonary tissue, y-globulins were found to be increased after the irradiation; this did not happen in the other treated rabbits. The irradiated lungs of five rabbits showed large amounts of globulins and specific complement-fixation which were not present in the untreated rabbit-lungs (fig. 2). The subcutaneous injection of irradiated or non-irradiated lung suspensions in treated animals caused a strong (histologically confirmed) cutaneous response of the delayedsensitisation type (fig. 3); only a slight non-specific reaction was present in untreated rabbits. The liver suspension and the saline solution did not cause any specific reaction. The subcutaneous injection of irradiated or non-irradiated lung suspensions in the control rabbits which were injected intraperitoneally with immunopotent cells of irradiated rabbits caused a histologically confirmed cutaneous reaction of the delayed sensitisation type. Besides vascular lesions the lungs of these rabbits showed were
1. Jennings, F. L., Arden, A. Archs Path. 1961, 71, 437. 2. Mariuzzi, G. M., Toti, A., Mori, R. Riv. Patol. clin. sper. 1961, 2, 287.
Fig. 1-Rabbit lung treated with 9900 rad. (Griedly’s silver-reticulin stain. x 200, reduced by half.) Thickened and proliferated reticular framework of the septa with partial disappearance of alveolar structure. Note silver-positive elements in the alveoli.
Fig. 2-Rabbit lung treated with 9900 rad. Complement and F.I.T.C. tagged anti-complement. ( x 600, reduced by half.) Brilliant fluorescence of
septal and endoalveolar cells.
1398
large zones of interstitial in the irradiated animals.
pneumonitis (fig. 4) similar
to
those
Our
findings demonstrate that pulmonary lesions which on to sclerosis can be produced in rabbits exposed to high doses of ionising radiations over the chest. Seemingly, autosensitisation plays a part in the histogenesis of these lesions, with production of anti-lung humoral and tissue autoantibodies; it is also possible to passively transfer this state of sensitisation to nonirradiated animals by means of immunopotent cells. How ionising radiation causes an autoimmune reaction should be investigated, using germ-free animals. can
go
A. A. V. C. R.
Departments of Morbid Anatomy and Radiology, University of Bologna, Italy.
M. MANCINI CORINALDESI TISON RIMONDI FERRACINI.
CHANDRA-KHETARPAL SYNDROME
SIR,-I recently had the opportunity of examining a 5-year-old child with the typical features of the ChandraKhetarpal syndrome (laevocardia, situs inversus viscerum, bilateral bronchiectasis, and paranasal sinus abnormalities 1). Only one other case has been reported2 since the original description in 1963. The child was brought to me with the complaints of cough with mucopurulent expectoration and dyspnoea on exertion for the past 6-8 months. The child was stunted, with slight cyanosis, early clubbing, and signs of bilateral basal bronchiectasis. The liver was slightly enlarged and on the left side. The cardiac impulse was felt in the 4th left intercostal space. X-ray of chest and abdomen and bronchography confirmed the presence of situs inversus viscerum, with Isevocardia, and The right lung had two main bronchi, and the left had three-an exact mirror image of the normal. Electrocardiography, vectorcardiography, fluoroscopy, and cardiac catheterisation did not reveal any cardiac defect.
bronchiectasis.
PATRICK LEE.
Hong Kong.
CHEDIAK-HIGASHI SYNDROME WITH FAMILIAL IDIOPATHIC HYPERLIPÆMIA
SIR,-A previously undescribed association between Chediak-Higashi syndrome and familial idiopathic hyperlipxmia was observed in two brothers born to white first-cousins. CASE l.-A 6-month-old boy was born with horizontal nystagmus and silver-grey hair. After the 3rd month, he had several acute episodes of sore throat and suppurative tonsillitis. On admission, skin pallor, nystagmus, partial albinism, photophobia, silver-grey hair, hepatomegaly, and splenomegaly were observed. During the hospital stay, eruptive xanthelasmas of face, neck, and thorax, andmilky appearance of the bloodsamples were observed. Laboratory examination showed: pancytopenia, white blood-cells with abnormal granulations typical of the Chediak-Higashi syndrome (see accompanying figure), a high content of neutral fats in the blood (3-150 mg. per 100 ml.), and a normal cholesterol level. CASE 2.-A boy who had jaundice in the first 24 hours of life which lasted a month, had a negative Coombs test, and no abnormal granulations in his white blood-cells. Before his admission to hospital at 23 months, he had had frequent bouts of acute suppurative tonsillitis. He had hyperthermia, cervical lymph-node enlargement, jaundice, generalised oedema, and abdominal pain. Examination revealed an acromegalic head, silver-grey hair, hepatomegaly, and splenomegaly. Laboratory examination showed: white blood-cells and bone-marrow with abnormal granulations typical of the Chediak-Higashi syndrome, pathological liver-function tests, and milky ascitic fluid with normal cytology (on smear) and negative bacteriology. 1. Chandra, R. K., Khetarpal, S. K. Indian J. Pediat. 1963, 30, 78. 2.
Sharpe,
M. V.
Lancet, 1963, ii,
42.
Bone-marrow showing white blood-cell with abnormal typical of Chédiak-Higashi syndrome. (X 1650.)
2 years later
a
sister
was
granulations
born who had jaundice in the first were found on clinical and
days of life. No abnormalities haematological examination. 4
The white blood-cell and bone-marrow changes identify the Chediak-Higashi syndrome.l-4 The eruptive xanthelasmas, milky serum, and high neutral fats blood-content of the first case, and the milky ascitic fluid without evidence of inflammation or malignancy of the second, are suggestive of familial idiopathic hyperlipa:mia. Saraiva et awl.5 demonstrated histochemically that the white blood-cell granulations are Sudan-positive. The electronic microscopy studies of Bernard et al.revealed that these granulations are made up of phospholipids and an unknown amorphous substance. These findings suggest a change in fat metabolism, probably resulting from the absence or deficiency of an enzyme in the cytoplasm of the white blood-cells. Thannhauser and Stanley, studying familial idiopathic hyperlipxmia in experiments using 13’I-labelled fat, suggested that an enzymatic deficiency was responsible for the fat storage in the cells. flagbergdescribed in two brothers an entity (called familial malignant hyperlipa:mia), characterised by
by him hyperlipaemia, milky serum, hepatomegaly, splenomegaly, jaundice, pancytopenia, and fat granules in the histiocytes, and differing from our two cases in the absence of partial albinism and silvergrey hair, and the presence of fat in the cytoplasm of the white blood-cells. From the published studies and the observations we report here, it seems reasonable to postulate that the basic cause of the changes noted in Chediak-Higashi syndrome is an inborn error of metabolism which involves both lipids and proteins.
It is our impression that the findings in our two cases are due to the same Chediak-Higashi gene, but showing a wider manifestation; another hypothesis would be a different mutation in the same locus-i.e., an allele of this gene. It seems unlikely that the syndromes in two brothers were due to the simultaneous homozygosis of two rare
independent
genes.
Department of Pediatrics, School of Medicine, University of Paraná, Curitiba, Pr, Brazil.
Brackenridge Hospital, 1. 2. 3. 4. 5. 6.
7.
8.
IZRAIL CAT IZRAIL CAT LEIDE PAROLIN MARINONI DINARTE JOSÉ GIRALDI MÁRIO BRAZ DE ALMEIDA ANTONIO SACHELLI NETO HOMERO BRAGA.
ATYS QUADROS DA SILVA. Austin, Texas, U.S.A. Beguez César, A. Bol. Soc. cubana Pediat. 1943, 15, 900. Chédiak, M. M. Rev. Hémat. 1952, 7, 362. Higashi, O. Tohoku J. Exp. Med. 1954, 59, 315; Sato, A. ibid. 1954, 60, 22; Sato, A. ibid. 1955, 61, 201. Donohue, W. L., Bain, H. W. Pediatrics, 1957, 20, 416. Saraiva, L. G., Azevedo, M., Correa, J. M., Carvalho, G., Prospero, J. D. Blood, 1959, 14, 1112. Bernard, J., Bessis, M., Seligmann, M., Chassigneux, J., Chomé, J. Pr. méd. 1960, 68, 563. Thannhauser, S. J., Stanley, M. M. Trans. Ass. Am. Physns. 1949, 62, 245 (cited by Hsia, D. Y.-Y. Errores Innatos del Metabolismo. Mexico, 1961). Hagberg, B., Hultquist, G., Svennerholm, L., Voss, H. Am. J. Dis. Child. 1954, 107, 267.
R. A. KEMP HARPER.
IMMUNOLOGICAL ASPECTS OF EXPERIMENTAL PULMONARY SCLEROSIS DUE TO IONISING RADIATIONS
SIR,-Both in man and in animals high doses of ionising radiations applied to the chest may give rise to pulmonary fibrosis.! The cause is still unknown, but some aspects of the lesions suggest the presence of immunological factors.2 Eight rabbits, weighing 2-2-5 kg., were irradiated with telecobalt-therapy " (f.p.d. 50 cm.) limited to the right hemithorax, 4900 rad over a 7-day period. 3 months later, six rabbits were exposed at the same site to a second cycle of 5000 rad, given over 5 days. The rabbits were bled 2-3 months after each cycle. Fragments of the left lung (non-irradiated) and the right (irradiated) lung from each rabbit were fixed in formalin 10% and acetone, and embedded in paraffin. Sections were examined microscopically and histochemically. Lung tissue from the same rabbits and from non-irradiated rabbits, and liver and kidney tissue from treated and untreated rabbits, were freeze-dried and embedded in paraffin (in vacuo); the sections, from which the paraffin was removed with petrol ether, were either fixed or not with acetone, and finally tested with the following: (1) fluorescein-isothiocyanate (F.I.T.C.)labelled y-globulins, isolated by salting from the serum of the same animals obtained before the first irradiation; (2) F.LT.C.labelled y-globulins, isolated as before from the serum of the same animals obtained 2 months after the last irradiation; (3) F.I.T.c.-labelled anti-rabbit globulins from sheep serum; and (4) human complement and F.I.T.c.-tagged anti-human complement from sheep serum. Further controls were applied using unlabelled y-globulins before the fluorescent serum and heat-inactivated complement. After the skin of the left flank was shaved, two rabbits treated with 2 cycles of radiations, and four untreated rabbits of the same weight, were injected under sterile conditions with (1) 1 ml. of suspension of finely homogenised irradiated rabbitlung in saline solution; (2) 1 ml. of suspension of untreated homogenised rabbit-lung; (3) 1 ml. of untreated homogenised rabbit-liver; and (4) 1 ml. of saline solution. The rabbits were
Fig. 3-Rabbit treated with 9900 rad. (Haematoxylin and eosin. x 80, reduced by half.) Cutaneous response 60 hours after injection with irradiated lung homogenate. Around the injected unabsorbed material, extensive inflammatory reaction of the dermis with perivascular infiltrates of lymphomonocytes and eosinophil cells, and lesions of the vessel walls. Fig. 4-Lung of rabbit 6 days after injection with immunopotent cells of irradiated rabbits. (Haematoxylin and eosin. x 200, reduced by half.) Lymphomonocytes and eosinophil cells infiltrate the alveolar septa
diffusely. under continuous observation for 5 days, and skin changes were photographed. The cutaneous injection sites were removed, and histologically examined. Two untreated rabbits, weighing 2-2-5 kg., after shaving of the abdomen were injected intraperitoneally with 2-5 ml. of a heavy suspension (in saline solution) of homogenised spleen from two irradiated rabbits. 24 hours later, 0-5 ml. of the same suspensions and solutions already mentioned were injected subcutaneously and the injection sites were examined. The rabbits were bled 6 days after the intraperitoneal injection, and the lungs were microscopically examined.
All rabbits showed thickened periodic-acid/Schiffpositive, alveolar septa infiltrated by lymphomonocytes and eosinophil cells, with pronounced reticular proliferation ; the elastic framework was often disrupted. The small-vessel walls were swollen by plasma-like material, and showed many altered or proliferating endothelial cells. In three of the six rabbits given 2 cycles of irradiation there large zones of septal and vascular fibrosis, pronounced changes in the alveolar cells, reticulin and collagen production, and sclero-calcined nodules (fig. 1). F.l.T.c.-tagged y-globulins, obtained from two rabbits long after the irradiation, were specifically fixed to the alveolar and septal cells of the treated or untreated lungs of the same or other rabbits, but were not fixed in sections of other organs. The tagged y-globulins, obtained from the same rabbits before the first irradiation, were not fixed in sections of the lung or other organs. In the irradiated rabbits, whose y-globulins were fixed in the pulmonary tissue, y-globulins were found to be increased after the irradiation; this did not happen in the other treated rabbits. The irradiated lungs of five rabbits showed large amounts of globulins and specific complement-fixation which were not present in the untreated rabbit-lungs (fig. 2). The subcutaneous injection of irradiated or non-irradiated lung suspensions in treated animals caused a strong (histologically confirmed) cutaneous response of the delayedsensitisation type (fig. 3); only a slight non-specific reaction was present in untreated rabbits. The liver suspension and the saline solution did not cause any specific reaction. The subcutaneous injection of irradiated or non-irradiated lung suspensions in the control rabbits which were injected intraperitoneally with immunopotent cells of irradiated rabbits caused a histologically confirmed cutaneous reaction of the delayed sensitisation type. Besides vascular lesions the lungs of these rabbits showed were
1. Jennings, F. L., Arden, A. Archs Path. 1961, 71, 437. 2. Mariuzzi, G. M., Toti, A., Mori, R. Riv. Patol. clin. sper. 1961, 2, 287.
Fig. 1-Rabbit lung treated with 9900 rad. (Griedly’s silver-reticulin stain. x 200, reduced by half.) Thickened and proliferated reticular framework of the septa with partial disappearance of alveolar structure. Note silver-positive elements in the alveoli.
Fig. 2-Rabbit lung treated with 9900 rad. Complement and F.I.T.C. tagged anti-complement. ( x 600, reduced by half.) Brilliant fluorescence of
septal and endoalveolar cells.
1398
large zones of interstitial in the irradiated animals.
pneumonitis (fig. 4) similar
to
those
Our
findings demonstrate that pulmonary lesions which on to sclerosis can be produced in rabbits exposed to high doses of ionising radiations over the chest. Seemingly, autosensitisation plays a part in the histogenesis of these lesions, with production of anti-lung humoral and tissue autoantibodies; it is also possible to passively transfer this state of sensitisation to nonirradiated animals by means of immunopotent cells. How ionising radiation causes an autoimmune reaction should be investigated, using germ-free animals. can
go
A. A. V. C. R.
Departments of Morbid Anatomy and Radiology, University of Bologna, Italy.
M. MANCINI CORINALDESI TISON RIMONDI FERRACINI.
CHANDRA-KHETARPAL SYNDROME
SIR,-I recently had the opportunity of examining a 5-year-old child with the typical features of the ChandraKhetarpal syndrome (laevocardia, situs inversus viscerum, bilateral bronchiectasis, and paranasal sinus abnormalities 1). Only one other case has been reported2 since the original description in 1963. The child was brought to me with the complaints of cough with mucopurulent expectoration and dyspnoea on exertion for the past 6-8 months. The child was stunted, with slight cyanosis, early clubbing, and signs of bilateral basal bronchiectasis. The liver was slightly enlarged and on the left side. The cardiac impulse was felt in the 4th left intercostal space. X-ray of chest and abdomen and bronchography confirmed the presence of situs inversus viscerum, with Isevocardia, and The right lung had two main bronchi, and the left had three-an exact mirror image of the normal. Electrocardiography, vectorcardiography, fluoroscopy, and cardiac catheterisation did not reveal any cardiac defect.
bronchiectasis.
PATRICK LEE.
Hong Kong.
CHEDIAK-HIGASHI SYNDROME WITH FAMILIAL IDIOPATHIC HYPERLIPÆMIA
SIR,-A previously undescribed association between Chediak-Higashi syndrome and familial idiopathic hyperlipxmia was observed in two brothers born to white first-cousins. CASE l.-A 6-month-old boy was born with horizontal nystagmus and silver-grey hair. After the 3rd month, he had several acute episodes of sore throat and suppurative tonsillitis. On admission, skin pallor, nystagmus, partial albinism, photophobia, silver-grey hair, hepatomegaly, and splenomegaly were observed. During the hospital stay, eruptive xanthelasmas of face, neck, and thorax, andmilky appearance of the bloodsamples were observed. Laboratory examination showed: pancytopenia, white blood-cells with abnormal granulations typical of the Chediak-Higashi syndrome (see accompanying figure), a high content of neutral fats in the blood (3-150 mg. per 100 ml.), and a normal cholesterol level. CASE 2.-A boy who had jaundice in the first 24 hours of life which lasted a month, had a negative Coombs test, and no abnormal granulations in his white blood-cells. Before his admission to hospital at 23 months, he had had frequent bouts of acute suppurative tonsillitis. He had hyperthermia, cervical lymph-node enlargement, jaundice, generalised oedema, and abdominal pain. Examination revealed an acromegalic head, silver-grey hair, hepatomegaly, and splenomegaly. Laboratory examination showed: white blood-cells and bone-marrow with abnormal granulations typical of the Chediak-Higashi syndrome, pathological liver-function tests, and milky ascitic fluid with normal cytology (on smear) and negative bacteriology. 1. Chandra, R. K., Khetarpal, S. K. Indian J. Pediat. 1963, 30, 78. 2.
Sharpe,
M. V.
Lancet, 1963, ii,
42.
Bone-marrow showing white blood-cell with abnormal typical of Chédiak-Higashi syndrome. (X 1650.)
2 years later
a
sister
was
granulations
born who had jaundice in the first were found on clinical and
days of life. No abnormalities haematological examination. 4
The white blood-cell and bone-marrow changes identify the Chediak-Higashi syndrome.l-4 The eruptive xanthelasmas, milky serum, and high neutral fats blood-content of the first case, and the milky ascitic fluid without evidence of inflammation or malignancy of the second, are suggestive of familial idiopathic hyperlipa:mia. Saraiva et awl.5 demonstrated histochemically that the white blood-cell granulations are Sudan-positive. The electronic microscopy studies of Bernard et al.revealed that these granulations are made up of phospholipids and an unknown amorphous substance. These findings suggest a change in fat metabolism, probably resulting from the absence or deficiency of an enzyme in the cytoplasm of the white blood-cells. Thannhauser and Stanley, studying familial idiopathic hyperlipxmia in experiments using 13’I-labelled fat, suggested that an enzymatic deficiency was responsible for the fat storage in the cells. flagbergdescribed in two brothers an entity (called familial malignant hyperlipa:mia), characterised by
by him hyperlipaemia, milky serum, hepatomegaly, splenomegaly, jaundice, pancytopenia, and fat granules in the histiocytes, and differing from our two cases in the absence of partial albinism and silvergrey hair, and the presence of fat in the cytoplasm of the white blood-cells. From the published studies and the observations we report here, it seems reasonable to postulate that the basic cause of the changes noted in Chediak-Higashi syndrome is an inborn error of metabolism which involves both lipids and proteins.
It is our impression that the findings in our two cases are due to the same Chediak-Higashi gene, but showing a wider manifestation; another hypothesis would be a different mutation in the same locus-i.e., an allele of this gene. It seems unlikely that the syndromes in two brothers were due to the simultaneous homozygosis of two rare
independent
genes.
Department of Pediatrics, School of Medicine, University of Paraná, Curitiba, Pr, Brazil.
Brackenridge Hospital, 1. 2. 3. 4. 5. 6.
7.
8.
IZRAIL CAT IZRAIL CAT LEIDE PAROLIN MARINONI DINARTE JOSÉ GIRALDI MÁRIO BRAZ DE ALMEIDA ANTONIO SACHELLI NETO HOMERO BRAGA.
ATYS QUADROS DA SILVA. Austin, Texas, U.S.A. Beguez César, A. Bol. Soc. cubana Pediat. 1943, 15, 900. Chédiak, M. M. Rev. Hémat. 1952, 7, 362. Higashi, O. Tohoku J. Exp. Med. 1954, 59, 315; Sato, A. ibid. 1954, 60, 22; Sato, A. ibid. 1955, 61, 201. Donohue, W. L., Bain, H. W. Pediatrics, 1957, 20, 416. Saraiva, L. G., Azevedo, M., Correa, J. M., Carvalho, G., Prospero, J. D. Blood, 1959, 14, 1112. Bernard, J., Bessis, M., Seligmann, M., Chassigneux, J., Chomé, J. Pr. méd. 1960, 68, 563. Thannhauser, S. J., Stanley, M. M. Trans. Ass. Am. Physns. 1949, 62, 245 (cited by Hsia, D. Y.-Y. Errores Innatos del Metabolismo. Mexico, 1961). Hagberg, B., Hultquist, G., Svennerholm, L., Voss, H. Am. J. Dis. Child. 1954, 107, 267.