GENETIC ÆTIOLOGY OF LACTOSE INTOLERANCE

188

in her fourth and sixth pregwas perfectly normal. She did not have any circulating thyroid autoantibodies and her chromosomal karyotype is normal. When all methods of estimating thyroid autoantibodies are taken into account, the incidence of positive results in mongol propositi was significantly lower in the Winnipeg series of Fialkow (1966) than in the series of Burgio et al. (1966). This may be partly due to the fact that Fialkow et al. (1965) state " Families in which there was previous knowledge of thyroid diseases were excluded ". The incidence in our series of mongols is similar to that of Fialkow (1966) (table ix*). Some of this variability between series may well be ascribable to different techniques of testing and possibly also to the fact that our mongol propositi were all living at home. Sex may also be a factor as thyroid autoantibodies have a higher incidence in females than in males.

than

one

mongol, namely

nancies, and her fifth child

deficiency lactose is not hydrolysed into its component monosaccharides, glucose and galactose, but remains

within the bowel lumen where it is metabolised to lactic acid. The " osmotic purge " of lactose and the toxic, acidic intestinal contents together cause abdominal pain, distension, and diarrhoea. Lactase deficiency is found in three groups of patients. In hereditary alactasia (Holzel et al. 1959) the main features are diarrhoea in infancy, with malnutrition (since lactose supplies 40% of the calories of milk). There is no blood-sugar rise after an oral dose of lactose, and remission is obtained by substitution of sucrose for lactose in the diet. Lactose intolerance probably persists throughout life (Davidson et al. 1964, Dumphy et al. 1965). In older children with symptoms from birth, virtual absence of lactase activity in the intestinal mucosa has been demonstrated. This condition has been described in two pairs of sibs, and probably has a genetic basis (Holzel et al. 1959, Davidson et al. 1964). We thank the United Liverpool Hospital Research Committee and In primary adult hypolactasia the patient has had a the Nuffield Foundation for generous financial support; Prof. P. M. Sheppard and Prof. J. R. Anderson for their help and advice; Mr. normal childhood, and lactose intolerance develops in W. T. A. Donohoe and the staff of the Department of Medicine, adult life. When jejunal tissue is examined, lactase University of Liverpool, who separated sera and coded the tubes; activity is always below the normal range, and occasionally Dr. J. Walker and Dr. J. Webster, the Lancashire County Council is completely absent. medical officers of health, divisions 4 and 5; Dr. J. S. G. Burnet, medical officer of health, Preston County Borough; and Dr. J. W. Secondary hypolactasia may be present at any age. Maybury and Dr. J. B. Wright (partners of P. J. W.) for allowing us There is histological abnormality of the small intestine access to their patients. Requests for reprints (including all tables) should be addressed to in many intestinal and systemic diseases-including coeliac disease; intestinal infections, mesenteric ischaemia, D. A. P. E., Nuffield Unit of Medical Genetics, Department of Medicine, Crown Street, Liverpool 7. carcinomatosis, and dermatitis herpetiformis. In these REFERENCES conditions, there is an associated deficiency of all the disaccharidase enzymes, sometimes with lactose or Burgio, G. R., Severi, F., Rossoni, R., Vaccaro, R. (1965) Boll. Soc. Ital. Biol. Sper. 41, 1005. sucrose intolerance (Arthur et al. 1966). Occasionally, (1966) Lancet, i, 497. because of toxic or metabolic factors, there is a generalised Dallaire, L., Flynn, D. (1966) Int. Congr. hum. Genet. abstract no. 72, p. 23. Doniach, D., Roitt, I. M. (1957) J. clin. Endocr. Metab. 17, 1293. enzyme deficiency in a histologically normal mucosa Fialkow, P. J. (1966) Am. J. hum. Genet. 18, 93. 1106. (Ferguson, Watson, Maxwell et al. 1967). (1967) Lancet, i, Uchida, I. A., Hecht, F., Motulsky, A. G. (1965) ibid. ii, 868. The aetiology of primary adult hypolactasia has not yet Fulthorpe, A. J., Roitt, I. M., Doniach, D., Couchmann, K. (1961) J. clin. been established. We describe here a family in which Path. 14, 654. two of six sibs have lactase deficiency. Hill, O. W. (1961) Br. med. J. i, 1793. —

—

—

—

—

—

Mellon, J. P., Day, B. Y., Green, D. M. (1963) J. ment. Defic. Saxena, K. M., Pryles, C. V. (1965) J. Pœdiat. 67, 363.

Res.

7, 31.

.

GENETIC ÆTIOLOGY OF LACTOSE INTOLERANCE ANNE FERGUSON M.B., B.Sc. Glasg. LECTURER IN

MEDICINE,

UNIVERSITY OF GLASGOW

J. D. MAXWELL M.B., B.Sc. Glasg. MCINTYRE RESEARCH SCHOLAR, ROYAL

INFIRMARY, GLASGOW

From the University Department

of Medicine, Glasgow Royal Infirmary

A 28-year-old man presented with diarrhoea and was found to have lactase deficiency. His family (parents and five sibs) have been investigated by lactose-tolerance tests and intestinal biopsy. Primary hypolactasia was demonstrated in one sister, and a generalised disaccharidase deficiency, secondary to a collagen disease, was present in another sister. Primary hypolactasia is probably a hereditary disease, transmitted as an autosomal recessive. Summary

Introduction

LACTASE (-galactosidase) is one of the disaccharidases in the mucosal cells of the small intestine. In lactase

Material

The pedigree is shown in fig. 1. The parents are not related. Case A (father).-This 56-year-old man had had a myocardial infarction at age 50. On admission to hospital he was clinically myxoedematous and has been treated with thyroid extract since then. Case B (mother).-This 50year-old woman had become thyrotoxic at age 48, and 0 Normal lactase

activity

partial thyroidectomy was per- 4D Primary adult hypolactosie formed 1 year later. She was 0 symptom-free and clinically 0 Secondary hypolactasia Male 0 Female euthyroid during the gastrointestinal investigation. Fig. 1-Pedigree, with two cases of primary adult hypolactasia. Case C (daughter).-At age 27, this 32-year-old woman had been admitted to hospital with pyrexia, abdominal pain, and ascites. A clinical diagnosis of tuberculous peritonitis was made. She responded dramatically to antituberculous therapy. 3 years later (8 months after chemotherapy was stopped) she presented with pyrexia, polyarthritis and a high erythrocytesedimentation

There was no response to antituberculous with a provisional diagnosis of " collagen disease ", steroid therapy was commenced. There was a rapid remission and at the time of the gastrointestinal investigation she was taking 10 mg. prednisolone daily. She was moderately cushingoid and had a mild, hypochromic anxmia (Hb, 11.0 g. per 100 ml.). This patient had had several episodes of abdominal distension and diarrhoea since her second illness;

therapy and,

rate.

189 these

were

unrelated to her

joint

symptoms and

were

not

precipitated by any dietary factor. Case D (son).-In 1959, 3 years after marriage, this man (the propositus) and his wife were investigated for possible

INTESTINAL DISACCHARIDASE LEVELS AND MAXIMUM BLOOD-SUGAR RISE IN LACTOSE AND GLUCOSE/GALACTOSE TOLERANCE TESTS

causes of sterility. He was found to be a tall, intelligent, healthy man. A left-sided varicocele was present. At operation for removal of this, the underlying testis was noted to be small. Testicular biopsy specimen revealed normal interstitial cells, normal cells of Sertoli, but complete absence of germinal cells. Chromosome analysis was normal, and del Castillo’s syndrome was diagnosed (del Castillo et al. 1947). In April, 1966, he had the first of several episodes of abdominal distension, flatulence, colicky periumbilical pain, and watery diarrhoea.

Symptoms lasted 4 or 5 days, with remissions of several weeks. He did not relate these symptoms to any dietary factor. Within 9 months his weight had fallen from 200 to 180 lb. (90 to 80 kg.). On investigation in December, 1966, the following results were obtained: Hb 16-4 g. per 100 ml.; blood-film normal; erythrocyte-sedimentation rate 4 mm. in lst hour (Westergren); blood urea and electrolytes, liver-function tests, serum-calcium normal; fsecal fat, 3-0 g. per day; xylose excretion, after 5 g. dose 2-8 g. (normal); barium enema and sigmoidoscopy normal. A biopsy specimen of jejunum showed normal mucosal histology, and lactase deficiency (see below). He has had no further symptoms after 4 months on a milk-free diet. Case E (daughter).-At age 14 this woman (now aged 28) had a pleural effusion from which no organism was cultured. She had antituberculous therapy for 3 months. She married at age 19 and has had five normal pregnancies. All the children are healthy. She has taken ferrous sulphate intermittently for several years. For 2 months before the gastrointestinal investigation she had been aware of occasional colicky abdominal pain, but had no abdominal distension or diarrhcea. Case F (son).-A healthy, unmarried 25-year-old man. -

o---0

!00g. Lactose test 50g. Glucose + 50 g. galactose test

*

Unit of

activity, (imole substrate hydrolysed

per minute.

Case G (son).-This healthy 23-year-old man had recently married and had no children. Case H (son).-This prepubertal 11-year-old boy had had no serious illnesses. Methods Tolerance Tests A solution of 100 g. lactose in 500 ml. water was ingested within a period of 5 minutes after an overnight fast. Samples for blood-sugar estimation were taken at 0, 15, 30, 60, 90, and 120 minutes. The next morning the test was repeated using a mixture of 50 g. glucose and 50 g. galactose. In cases D and H, the glucose/galactose test was omitted. Blood-sugars were estimated by the method of Hoffman (1937) modified for use with the ’AutoAnalyzer ’ (Technicon methodology N26). Intestinal Biopsy In all cases, the Crosby capsule was used to obtain a specimen from the proximal jejunum, with 10 cm. of the ligament of Treiz. In case A a duodenal specimen was also obtained. The tissue was carefully removed from the capsule and halved. Before routine sectioning and staining, one piece was examined under the dissecting microscope and classified as normal (with finger or leaf villi), ridged, or flat. The other was weighed on a microtorsion balance, homogenised in ice-cold water, and brought to a final concentration of 10 mg. per ml. Lactase, sucrase, and maltase activities were estimated using the method of Dahlqvist (1964) with minor modifications as previously described (Paton et al. 1966). Results Tolerance Tests

Blood-sugar curves for lactose and glucose/galactose tolerance tests are shown in fig. 2. The table lists the maximum blood-sugar rise for each test. In case D, after ingestion of 100 g. of lactose, the blood-sugar rose by only 6 mg. per 100 ml.; he complained of abdominal pain and distension and had eleven loose stools within the next 4 hours. In case E, blood-sugar rose by 14 mg. per 100 ml. after lactose. Atter an equivalent dose of glucose and galactose the blood-sugar rose by 120 mg. per 100 ml. She had no symptoms during or after the tests. In the other cases, curves obtained after, lactose were similar to those after glucose/galactose. Jejunal Microscopy

Biopsy specimens from cases A, B, D, E, F, and G microscopically normal, and only finger and leaf villi were present. The mucosa from case C, examined under the dissecting microscope, was ridged with no normal villi present. This moderate abnormality is non-specific, and often found in patients with systemic or gastrointestinal disease (Ferguson, Watson, Maxwell et al. 1967). In case H, most villi were leaf-shaped and a few mucosal ridges were present. Anderson and Townley (1962) state that leaf villi are a normal finding in children. were

Fig. 2-Blood-sugar

curves in cases A-H, after 100 g. lactose 50 g. glucose and 50 g. galactose.

or

D3

190

Provisionally, this was classified from an 11-year-old boy. Intestinal Enzymes

normal

A reduction in effective lactase activity could be due the presence of an enzyme inhibitor. However, Dumphy et al. (1965) demonstrated that an extract of The results of disaccharidase assay are shown in the mucosa from a case of hypolactasia had no inhibitory table. The previously established lower limits of normal effect on the lactase activity of a normal specimen. It is for sucrase and maltase activities are as described else- possible that there is a genetic inhibitor of enzyme where (Paton et al. 1966). Lactase activity below 1-0 formation-an abnormal gene which causes a reduction unit per g. mucosa is considered to be definitely abnormal: in the rate of synthesis of lactase in adult life. A more probable explanation, which includes hereditary levels between 1-0 and 1-4 units are borderline. These levels are similar to those cited by other workers who use alactasia of infants, is as follows. Three genes (alleles) L, 11 and l2, can occupy the locus which controls lactase a similar method for lactase assay (Sheehy and Anderson production. The normal gene, L, is dominant; the McMichael et al. et al. In 1965, Dumphy 1965, 1965). others are recessive. Normally lactase is produced case C lactase, sucrase, and maltase activities were subnormal. These deficiencies are the result of the throughout life under the influence of L in homozygotes non-specific mucosal abnormality secondary to a collagen (LL) and heterozygotes (Ll1 or Ll2). The gene 11 disease. This woman has a secondary hypolactasia with controls lactase production during the period of physiological requirement, in infancy and childhood. The no convincing evidence of a specific lactose intolerance. In case D, lactase activity was completely absent, genotype 11 11 results in primary adult hypolactasia. It while sucrase and maltase levels were normal. This is not truly abnormal and has a harmful effect only in patient had profuse diarrhoea during the lactose-tolerance societies in which milk forms part of the adult diet. For this reason, the gene is widespread and homozygotes are test. Case E, there was definite hypolactasia with a level of 0-5 units per g. mucosa. This woman had a flat probably common. 12 is definitely abnormal. The lactose-tolerance curve, with a very high glucose/ genotype 12 12 results in hereditary alactasia of infants-a galactose curve. She had no symptoms suggestive of rare disease when compared with adult hypolactasia. lactose intolerance. It is postulated that the total The genotype /1 12 could result in the rare situation of intestinal lactase activity had not yet fallen to the threshold hypolactasia presenting in childhood, with normal milk tolerance in infancy (Auricchio et al. 1963). Secondary level, below which clinical lactose intolerance can occur. hypolactasia is an example of an environmental factor Discussion causing an effect identical to that brought about by gene Several possible aetiologies of primary adult hypolactasia action. have been proposed. Cuatrecasas et al. (1965) noted that This is as yet no agreement on the true incidence of patients with " flat " lactose-tolerance curves had a lower primary hypolactasia in a normal population. Dumphy milk intake (one quart daily) than controls (two quarts et al. (1965) when studying milk intolerance, found daily). These workers admitted that some patients in the hypolactasia in one of his twelve controls. In Sheehy first group had avoided milk, since the patients had and Anderson’s (1965) series of fifty American Servicenoticed that their symptoms were related to ingestion one man had lactase activity of 0-7 units per g. men, of milk. In our case D, the daily milk intake was three mucosa. Auricchio et al. (1963) found hypolactasia with " " pints, and in case E it was one pint. Disuse is a very lactose intolerance in three of eighteen patients underunlikely aetiology of hypolactasia. going surgery for duodenal ulcer. Of patients with " Loss of lactase activity may be always secondary to a functional" diarrhoea, at least 20% have hypolactasia systemic disease (as in case C) or a mucosal insult (such (McMichael et al. 1965, Ferguson and Maxwell 1967). as infestation or gastrointestinal infection). In case D, Unfortunately, hypolactasia cannot be inferred if there lactase deficiency could be caused by the same abnormality is a history of milk intolerance, since there is normal (perhaps genetic) as the testicular lesion of del Castillo lactase activity in half of these patients (Dumphy et al. syndrome. In case E, pulmonary tuberculosis 14 years 1965). Perhaps some of those with normal lactase earlier could have caused mucosal damage: an association activity have an abnormality of lactose transport from is unlikely, but cannot be disproved. intestinal lumen to the site of lactase activity, de la In most cases, hypolactasia in the adult is not a conFuente and Sols (1962) have demonstrated this abnormality tinuing manifestation of childhood lactose intolerance in yeasts. (Hammerlei et al. 1965). In many cases, including our Lactose-tolerance tests must be critically assessed, and case D, onset of symptoms is fairly sudden in the third, probably are of value only in excluding hypolactasia fourth, or fifth decade (Dumphy et al. 1965). (Newcomer and McGill 1966). Thus for an assessment The incidence of hypolactasia varies in different races of the true incidence of hypolactasia, it will be necessary (McMichael et al. 1966) and tribes (Cook and Kajubi to carry out intestinal biopsy and direct enzyme assay in 1966). This suggests a genetic aetiology. Our findings a large number of healthy adults. of hypolactasia in two of six sibs support this theory. We thank Dr. D. Murray who reported the small-bowel histology; Since the parents are normal, and a brother and sister Prof. E. M. McGirr and Dr. W. C. Watson for their interest and advice; and Dr. M. Ferguson-Smith who encouraged us to undertake are affected, the pattern of inheritance is that of an this study. autosomal recessive gene. Requests for reprints should be addressed to A. F., University What is the mode of action of this gene ? It seems Department of Medicine, Royal Infirmary, 86 Castle Street, likely that there is normal lactase activity at birth, and Glasgow C.4. REFERENCES a gradual diminution in enzyme activity until, in adult C. M., Townley, R. R. W. (1962) Ciba Fdn Study Grps, no. 14, Anderson, the falls a the level below to threshold life, (related p. 39. patient’s dietary habits) and symptoms develop. In case D, Arthur, A. B., Clayton, B. E., Cotton, D. G., Seakins, J. W. T., Platt, J. W. (1966) Lancet, i, 172. this threshold was reached in April, 1966. In case E, Auricchio, S., Rubino, A., Landolt, M., Semenza, G., Prader, A. (1963) been has discovered before the of onset hypolactasia ibid. ii, 324. symptoms. References continued at foot ofnext column as

a

mucosa

to

(

191

TRAUMATIC HÆMOGLOBINURIA CAUSED BY KARATE EXERCISES

J. A. STREETON M.B. Melb.

From the

Alfred Hospital, Melbourne, A

of

Victoria 3181, Australia

after karate

hæmoglobinuria Summary exercises involving considerable case

trauma to

the hands is described, the hæmoglobinuria being prevented by the use of a sponge-rubber cushion. After investigation, which included the performance of the exercises under laboratory supervision, the hæmoglobinuria was diagnosed as being a variant of march hæmoglobinuria. Introduction

HAEMOGLOBiNiJRiA is occasionally noted in healthy young adults after marching, running, or walking (Gilligan and Blumgart 1941, Dacie 1954, de Gruchy 1964, Buckle 1965), but not after other equally strenuous forms of exercise such as cycling or swimming (Gilligan and Blumgart 1941). A different type of exercise was found to be the cause ofhaemoglobinuria in a young man referred to this hospital.

The karate exercises consisted of two 15-minute periods of hand-strengthening exercises. During the first period he chopped the ulnar borders of his hands alternately against the seat of a wooden stool at the rate of 120-140 beats per minute as hard as possible. In the second period he punched his clenched fists against a piece of blanket spread on a concrete floor at the same rate. The first set of exercises were performed in a sitting position, the second set was performed in a squatting

position. The effect of exercise was striking. The first specimen of urine passed 1 hour after commencing exercise was bright red. Spectroscopic examination showed peaks at 541 m[.L and 577 m[L corresponding to those of oxyhasmoglobin (Harboe 1959) and a small peak at 530 mfL corresponding to methtmoglobin. Microscopic examination revealed no red blood-cells, but a considerable number of granular casts filled with iron-containing granules, presumably hxmosiderin. 2 hours after exercise, his urine still contained haemoglobin (see table). His creatinine clearance did not change over the period of the experiment. His plasma 1 hour after starting exercise also contained hxmoglobin (Harboe 1959) (see table), and this persisted for A SUMMARY OF CHANGES IN PLASMA AND URINARY HEMOGLOBIN LEVELS OVER THE TWO TEST PERIODS

Case-report 26-year-old slaughterman presented with a 3-month history of passing red urine on many occasions, but only for A

few hours in the late afternoon. He did not undertake any exercise either at home or at work, apart from a type of karate exercise which he performed in the afternoon after returning home from work. He had not believed there was any association between his karate exercises and his red urine. Otherwise, his past and present history was unremarkable. Physical examination did not reveal any abnormalities, apart from notable callosities on the ulnar borders of his hands and over his knuckles. On this occasion, his urine was normal. Likewise, his haemoglobin, blood-film, white-blood-cell count, reticulocyte-count, erythrocyte-sedimentation rate, blood Wassermann reaction, direct Coomb’s test, Ham’s test, DonathLandsteiner test, blood-urea, creatinine clearance, liver function, serum-protein electrophoresis, and serum-haptoglobin were normal. X-rays of his hands were normal. A specimen of red urine was examined spectroscopically and found to contain oxyhaemoglobin and methtmoglobin. There were no red blood-cells present. Since no obvious cause could be found for his hsemoglobinuria, an experiment was arranged to see whether his karate exercises were responsible. Before commencing the tests, the patient was asked to abstain from his exercises for 1 week, and then to come to the hospital and carry out his exercises under supervision. Blood and urine samples were taken before and for 6 hours after exercise. a

severe

del

Castillo, E. B., Trabucco, A., de la Balze, F. A. (1947) J. clin. Endocr. Metab. 7, 493. Cook, G. C., Kajubi, S. K. (1966) Lancet, i, 725. Cuatrecasas, P., Lockwood, D. H., Caldwell, J. R. (1965) ibid. i, 14. Dahlqvist, A. (1964) in Disorders due to Intestinal Defective Carbohydrate Digestion (edited by P. Durand); p. 8. Rome. Davidson, M., Sobel, E. H., Kugler, M. M., Prader, A. (1964) Gastroenterology, 46, 737. Dumphy, J. V., Littman, A., Hammond, J. B., Forstner, G., Dahlqvist, A., Crane, R. K. (1965) ibid. 49, 12. Ferguson, A., Maxwell, J. D. (1967) Unpublished. — Watson, W. C., Maxwell, J. D., Fell, G. (1967) Gut (in the press). de la Fuente, G., Sols, A. (1962) Biochim. biophys. Acta, 56, 49. Hammerlei, U. P., Kistler, H., Ammann, R., Marthaler, T., Semenza, G. Auricchio, S., Prader, A. (1965) Am. J. Med. 38, 7. Hoffman, W. S. (1937) J. biol. Chem. 120, 51. Holzel, A., Schwarz, V., Sutcliffe, K. W. (1959) Lancet, i, 1126. McMichael, H. B., Webb, J., Dawson, A. M. (1965) ibid. i, 717. — — —

(1966) Br. med. J. ii, 1037.

Newcomer, A. D., McGill, D. B. (1966) Gastroenterology, 50, 340. Paton, E., Murray, E. S., Watson, W. C. (1966) Br. med. J. i, 459. Sheehy, T. W., Anderson, P. R. (1965) Lancet, ii, 1.

4 hours. His plasma-haptoglobin (Owen et al. 1959) was 120 mg. per 100 ml. before exercise, but was less than 20 mg. per 100 ml. 4 hours later. The mechanical and osmotic fragility and autohsemolysis (Dacie 1954), which were normal

before exercise, were unaffected. No fragmented red bloodcells were seen in blood smears. The second part of the test was performed 1 month later. In this, the patient carried out a similar exercise session, but on this occasion a piece of sponge rubber 4 in. thick was used to cushion the blows. No haemoglobin appeared in either his urine or his plasma, and his serum-haptoglobin levels were unchanged from the pre-exercise level of 120 mg. per 100 ml.

Discussion The haemoglobinuria which occurs after marching, running, or walking has been termed " march hxmoglobinuria ". It has been shown to be due to mechanical damage to erythrocytes in the soles of the feet, and can be prevented by the use of foam-rubber insoles or by walking on soft surfaces (Davidson 1964, Buckle 1965). In the present case, hasmoglobinuria developed after exercise involving considerable trauma to the hands only, and was prevented by the use of sponge rubber. It must be assumed that there was release of haemoglobin from erythrocytes damaged intravascularly by blows to the hands (Gilligan et al. 1943, Davidson 1964, Buckle 1965). The factors which predispose to the development of march hxmoglobinuria are not known, but may be related to the amount of haptoglobin available for combination with haemoglobin (Allison and ap Rees 1957), to the degree of haemolysis which can rapidly deplete the available haptoglobin (Langley et al. 1962, Prager et al. 1963), and to the concentration of free haemoglobin in the plasma (Yuile et al. 1949, Laurell and Nyman 1957, Murray et al. 1961). The area of body surface exposed to trauma may also be an important factor, since it would seem that the smaller the exposed body-surface area and the greater the