Preliminary Observations on Glucose Tolerance and Serum Proteins M

Br J Ind Med: first published as 10.1136/oem.23.1.67 on 1 January 1966. Downloaded from

Brit. J. industr. Med., I966, 23, 67.

Chronic Manganism: Preliminary Observations on Glucose Tolerance and Serum Proteins M. HASSANEIN, H. A. GHALEB, E. A. HAROUN, M. R. HEGAZY, and M. A. H. KHAYYAL From the Department of Medicine and the Department of Pharmacology and Therapeutics, Faculty of Medicine, Cairo University

An intravenous glucose tolerance test was carried out in i i patients with chronic manganese poisoning. Prolonged reactionary hypoglycaemia was observed. The underlying mechanism is discussed. It may be due to a disturbance of the hypothalamo-pituitary-adrenal axis. In seven of these patients total serum proteins were estimated and were separated electro- phoretically. The albumin: globulin ratios were lower in patients than in controls. There were significant reductions in serum albumin concentrations and increases in concentrations of cxl and ,B globulins.

Since the early discovery of chronic manganese explore factors which may help to elucidate these poisoning among workers manufacturing chlorine mechanisms. We have studied the effects of chronic gas in France using manganese ore (Couper, I837), manganism on glucose tolerance and on liver many other reports from Europe, North Africa, function, as shown by the electrophoretic pattern of India, and America have appeared (see, e.g., Nazif, serum proteins. The latter study was suggested by copyright. 1936; Flinn, Neal, Reinhart, Dallavalle, Fulton, and the similarities with Parkinsonism, the relation Dooley, 1940; Fairhall, I945; Boyer and Rodier, between which and impairment of liver finction is 1954; Rodier, I955; and Pefialver, I955, I957). The well established in Wilson's disease. total number of published cases has not exceeded 6oo, but manganism has aroused great interest Materials and Methods because it is severely incapacitative. Subjects In this study II patients with well- http://oem.bmj.com/ The neurological lesions found clinically are established chronic manganism and 20 healthy volunteers characteristic enough to warrant their categorization of the same age group were compared. All subjects were as a distinct clinical syndrome with many similarities men of 32 to 65 years, and all were in hospital for seven to but also marked differences from Parkinsonism to I5 days before testing. The patients had been (Hassanein, I964). Pulmonary complications may exposed to manganese ore for an average of seven years. also be observed, as by Boyer and Rodier (I954) in They all came from the same native village (Khattare in Moroccan workers. Penialver (1955), however, upper Egypt) and were brought to Kasr El-Aini Cairo rarely observed them in Cuban workers, and University Hospital for investigation and treatment. on September 26, 2021 by guest. Protected Hassanein (I964) never encountered them in an Glucose Tolerance Test The subjects were kept extensive survey of manganism in the United Arab on a high carbohydrate diet for seven to I5 days before Republic. Pulmonary lesions have been produced the test and fasted overnight. experimentally (Lloyd Davies, I946). Glucose was injected intravenously to eliminate There is universal agreement about the patho- differences in absorption from the alimentary tract. This genesis of chronic manganism and about the nature method gives more consistent results than oral ad- of its neurological features; and much ministration (Amatuzio, Stutzman, Vanderbilt, and and progress Nesbitt, 1953). After withdrawal of the fasting venous work has been done on the physiological role, sample glucose (0o33 g./kg. ideal body weight* as a 25% absorption, distribution, transport, and fate of w/v solution) was given over a two-minute period. Blood manganese in the body (Cotzias, I958); but the samples were taken o05, I, I-5, 2, 3, and 4 hr. later into possible mechanism or mechanisms underlying the sterilized tubes containing 4 mg. anticoagulant preserva- neurological lesions are still matters of speculation. tive/ml. blood (preservative: sodium fluoride/thymol/ The present work represents two attempts to *If the height of the subject was h cm., the ideal body Received for publication July 9, I964. weight was taken as (h-ioo)kg. 67 Br J Ind Med: first published as 10.1136/oem.23.1.67 on 1 January 1966. Downloaded from

68 M. Hassanein, H. A. Ghaleb, E. A. Haroun, M. R. Hegazy, and M. A. H. Khayyal lithium oxalate, IO/I/3, w/w/w) as described by Hepler of the test, but the difference was not significant at (I960), and stored in an ice chest until the end of the 0o5 and I hour. From I-5 to 4 hours, however, it was series. Blood glucose estimations were carried out by significantly lower, i.e., there was an exaggerated, Somogyi's method (1945) using Nelson's (I944) arseno- molybdate colour reagent. Optical densities were prolonged hypoglycaemia. This is illustrated in measured with a Zeiss spectrophotometer at 500 m,e. Fig. I, which shows a typical case. Some patients The method is not affected by reducing substances in complained of drowsiness during the hypoglycaemic the blood other than glucose. phase. The significance of the differences between the mean values of blood glucose in the controls and in the patients Tests on Serum Proteins The mean con- was subjected to the t-test (Bum, Finney, and Goodwin, centrations of total serum proteins and of the 1950). separated fractions (albumin and l °C2, f, and y globulins) are shown in Table III. The albumin: Electrophoretic Analysis of Serum Proteins globulin ratios were lower in the patients than in Total serum proteins were estimated in patients and controls fasted overnight using Greenberg's technique the controls, because the albumin concentrations (I929). Paper electrophoresis of the proteins was carried were lower and the y-globulin concentrations out essentially as described by Kamel (1956). higher. The concentrations of the other globulins were also higher, but only significantly so in the Results case of a2 globulin. Glucose Tolerance Tests The results ob- Discussion tained on i I patients and seven controls are shown in Tables I and II. In the patients the glucose levels The mean fasting blood glucose levels in patients were lower than in the controls from the beginning with chronic manganism were lower than in normal

TABLE I BLOOD GLUCOSE LEVELS IN PATIENTS WITH CHRONIC MANGANESE POISONING Patient Age Weight Dose Blood Glucose Level (mg./Ioo ml.) (yrs) (kg.) of Glucose copyright. (g.) Fasting After Intravenous Administration of Glucose (hours) 0-5 I 1I5 2 3 4 I 62 63 22 85 IOI 59 80 7I 71 8I 2 55 47 20 85 II9 83 69 76 73 66 3 6o 57.5 22 93 "14 83 62 56 55 72 4 58 60 22 76 I64 96 60 59 82 8I 5 55 62-5 22 63 io8 84 58 6i 63 65 http://oem.bmj.com/ 6 60 57-5 22 83 II6 7' 68 74 58 79 7 35 58 22 77 200 I48 65 6i 6i 64 8 47 50 20 75 138 48 39 39 40 58 9 40 60 22 64 I68 "14 64 52 60 7I 10 40 63 22 IOO 14I 9' 89 85 82 80 II 54 58 22 8i io8 64 67 67 60 63 Mean 80-2 I34.3 85-6 65-6 63 7 64-I 70'9 Standard deviation (S.D.) ±II-I ±3I 4 ±27-7 ±iI2-6 12-7 ±I2-3 ±8 3 Standard error (S.E.) ±8-3 ±3-3 ±9-47 ±3.8 ±3-8 +3-7 ±2-5 on September 26, 2021 by guest. Protected Percentage rise or fall from initial level +67-5 +6-7 -I8-3 -20-5 -20@ I -II *6

TABLE II INTRAVENOUS GLUCOSE TOLERANCE TEST OF PATIENTS WITH CHRONIC MANGANESE POISONING AND CONTROLS Mean Values for Blood Glucose Level (mg./Ioo ml.) Fasting 0-5 hour I hour 1I5 hours 2 hours 3 hours 4 hours Patients with chronic manganism Mean 80-2 134'3 85.6 65-6 63 7 64 I 70°9 S.E. ±3-3 ±9-5 ±8-3 ±3-8 ±3-0 ±3-7 ±2-5 Normal controls Mean 89.4 143-3 97.7 8I.4 84-3 80-3 82-o S.E. ±2-8 ±II-3 ±I3-0 ±5-3 ± I7 ±9-7 ±2*I Significance of difference between means P = 0-05 n.s. n.s. P < 0-05 P < 0-05 P < O-OI P < O-OI of patients and controls t = 21I3 t = 0-6I t = 0-82 t = 2-44 t = 2.5I t = 31I1 t = 3-39 n.s. not significant Br J Ind Med: first published as 10.1136/oem.23.1.67 on 1 January 1966. Downloaded from

Chronic Manganism: Preliminary Observations on Glucose Tolerance and Serum Proteins 69 TABLE III SERUM PROTEIN CONCENTRATIONS IN PATIENTS WITH CHRONIC MANGANESE POISONING AND IN CONTROLS Mean Serum Protein Concentrations (g./IOo ml.) Total Albumin Globulins Proteins

aX2 v Patients with chronic manganism Mean 6.94 3-58 0-42 o064 o-82 I'47 S.D. ±0-28 ±0 25 ±o0IO ±O-IO ±O-OI ±0-24 S.E. ±0-11 ±0-09 +0-04 ±00-4 ±0-05 ±o-09 Controls Mean 7.77 4'91 0-35 O055 O077 I-I9 S.D. ±0-I4 ±0-09 ±0-01 ±0-02 ±0°07 S.E. ±0-03 ±0-02 ±0-003 ±0o005 ±0-03 ±0-02 Significance of difference between P < 0001 P < 000I O0I < P < 005 P < 005 n.s. p < O-OI means of patients and controls t = 4-30 t = I3-90 t = I-91 t = 2-I8 t = o-85 t = 3°03 n.s. not significant

160 its short duration in our experiments excludes the possibility of impairment of carbohydrate meta- 140 bolism by the liver. The prolonged reactionary hypoglycaemia seems rather to indicate an impair- 120 / ment of adrenal function, as suggested by Thorn, and 100 Koepf, Lewis, Olsen (I940). This might be due to a primary functional lesion in the adrenal E I \ cortex or to a secondary effect of a disturbance in UJ 80so the hypothalamo-pituitary-adrenal axis. The obser- copyright. 0 vation by Rodier (i955) that patients with chronic u v 60/ manganism showed a reduction in urinary I7-ketosteroids could favour either suggestion. A lesion in 40_ the adrenals has been demonstrated by Grunstein 0 and Popowa (I929) in animals subjected to pro- 0 longed feeding with manganese salts. X + _ Prolonged reactionary hypoglycaemia might, http://oem.bmj.com/ together with other factors, exert a profound

I:S 4 influence on the physiological function of the central HOURS nervous system. Thus a 'feedback' mechanism may FIG. I. Intravenous glucose tolerance curve in a 47-year-old operate whereby the initial disturbance causes patient suffering from chronic manganism. hypoglycaemia, which in turn increases the disturbance. The disturbance of the serum protein patterns individuals (controls) but lay within normal limits. and the reduction of the albumin: globulin ratios on September 26, 2021 by guest. Protected The glucose tolerance curve in the patients was, in patients with chronic manganese poisoning may however, significantly lower after the primary peak be a reflection of liver involvement. It cannot be than in the controls (Table II and Fig. i). stated at this stage whether this disturbance has any Hypoglycaemia from manganese ion has pre- relation to the intracellular manganese traffic in the viously been reported by Rubenstein, Levin, and liver cells. The liver is known to produce a high Elliott (I962) who succeeded in controlling a turnover of manganese, with most of the metal diabetic patient at a certain stage of his disease with being excreted via the bile (Greenberg, Copp, a manganese salt. Attacks of hypoglycaemia have and Cuthbertson, 1943; Maynard and Fink, I956). also been reported in cases of post-encephalitic Although Rodier (I955) found a reduction in the Parkinsonism by Meakins (I940). In view of the urinary 17-ketosteroids in patients suffering from similarities in the neurological lesions in the two chronic manganese poisoning, it is too early to state conditions (Naby and Hassanein, I965) the causes that the possible endocrine disturbances in may well be similar. these patients would be responsible for the observed The low peak of the glucose tolerance curve and changes in the serum proteins pattern.

6 Br J Ind Med: first published as 10.1136/oem.23.1.67 on 1 January 1966. Downloaded from

70 M. Hassanein, H. A. Ghaleb, E. A. Haroun, M. R. Hegazy, and M. A. H. Khayyal The authors wish to express their deep indebtedness Griinstein, A. M., and Popowa, N. (I929). Arch. Psychiat. to Professor S. El Ridi and his staff for their help. Nervenkr., 87, 742. Quoted from Von Oettingen, W. F. (I935). Physiol. Rev., I5, I92. Hassanein, M. (I964). Med.3J. Cairo University. REFERENCES Hepler, 0. E. (i96o). Manual of Clinical Laboratory Methods, 4th ed., p. 257. Charles C. Thomas, Springfield, Illinois, U.S.A. Amatuzio, D. S., Stutzman, F. L., Vanderbilt, M. J., Kamel, G. (I956). M.Sc. Thesis, Faculty of Sciences, and Nesbitt, S. (I953). J. Clin. Invest., 32, 428. University of Ciiro. Boyer, J., and Rodier, J. (1954). Rev. Neurol., 90, 13. Maynard, L. S., and Fink, S. (1956). J. clin. Invest., 35, 831. Burn, J. H., Finney, D. J., and Goodwin, L. G. (I950). Lloyd Davies, T. A. (1946). Brit. J. industr. Med., 3, III. Biological Standardization, 2nd ed., p. 26. Oxford Meakins, J. C. (1940). Ann. intern. Med., I3, I830. University Press, London. Naby, S. Abd el, and Hassanein, M. (I965). J. Neurol. Cotzias, G. C. (x958). Physiol. Rev., 38, 503. Neurosurg. Psychiat., 28, 282. Couper, J. (i837). J. Chim. mid. (Paris), 2 ser, 3, 233. Nazif, M. (1936). J. Egypt publ. Hlth Ass., I0, I. Fairhall, L. T. (1945). Physiol. Rev., 25, i82. Nelson, N. (i944). J. biol. Chem., 153, 375. Flinn, R. H., Neal, P. A., Reinhart, W. H., Dallavalle, J. M., Pefialver, R. (I955). Industr. med. Surg., 24, I. Fulton, W. B., and Dooley, A. E. (1940). Chronic (I957). A.M.A. Arch. iniustr. Hlth, I6, 64. manganese poisoning in an ore-crushing mill. Public Rodier, J. (I955). Brit. J3. industr. Med., 12, 21. Health Bull. No. 247, Washington, D.C., U.S. Gov. Rubenstein, A. H., Levin, N. W., and Elliott, G. A. (I962). Print. Office. Lancet, 2, 1348. Greenberg, D. M. (I929). J. biol. Chem., 82, 545. Somogyi, M. (I945). J. biol. Chem., I6o, 69. Copp, H. D., and Cuthbertson, E. M. (I943). Ibid., Thorn, G. W., Koepf, G. H., Lewis, R. A., and Olsen, E. F- 1479 749. (1940). J. clin. Invest., i9, 8I3. copyright. http://oem.bmj.com/ on September 26, 2021 by guest. Protected