L. Emmett Holt

Glycogen storage disease

Summary Two cases of the hepatic form of glycogen storage disease (von Gierkes disease) have been studied in detail. Observations on liver slices freshly removed at biopsy have shown an abnormal stability of the glycogen, but a satisfactory glycogenolytic response was obtained upon admixture with fresh normal liver, confirming previous work that the glycogenolytic enzyme system is defective in this condition. Liver slices treated with various enzyme preparations, including phosphorylase and amylase, gave results which were inconclusive as to the nature of the enzyme defect. It was demonstrated that liver functions other than glycogenolysis may be affected in this disease, defective prothrombin formation and defective methylation being observed. In one patient a pronounced lipemia was present at the age of 2 years which had almost completely disappeared at the age of 6 years, suggesting that the associated disturbance of fat metabolism, which has been observed repeatedly in infants with this disease, may be a self-limited one. An unidentified reducing substance was found, probably belonging to the pentose group, in the urine of one of our patients. Particular attention is called to two indirect results of the impaired glycogenolysis: (a) an abnormal conversion of carbohydrate to fat, demonstrated by respiratory quotient studies, which leads to obesity; (b) impaired nitrogen retention, brought about apparently by excessive gluconeogenesis from protein in the effort to sustain blood sugar, a condition which may interfere with normal growth. We have shown that this latter condition may be satisfactorily treated by the administration of an extra protein meal at night.

The intravenous administration of fat

Summary 1. It is possible to prepare fat emulsions that can safely be administered by vein. 2. Our experience confirms that of others, suggesting that these preparations are of value in parenteral feeding. 3. The possible limitations of their use are discussed.

Nitrogen Balance in Experimental Tryptophane Deficiency in Man.

The observations here reported comprise a part of a more comprehensive study undertaken jointly by the Departments of Medicine and Pediatrics of the Johns Hopkins University on experimental amino acid deficiencies in man. The purpose of this study was twofold. It was felt desirable to determine human amino acid requirements by direct observations on man, rather than by inferences based on animal experiments, since it is now known that even closely related animal species differ in their amino acid requirements. A second purpose was to discover whether deficiencies of particular amino acids produced characteristic pathological changes which could be recognized by clinical or laboratory technics. It seemed probable that, in certain parts of the world at least, specific amino acid deficiencies might be present, and that a familiarity with the picture, such as could be obtained in an experimental study, would be of aid in recognizing and treating them. The present report deals with the nitrogen balance of 4 normal adult males, who ingested a diet deficient in tryptophane for periods of 2 to 5 weeks. The experimental diet consisted of fats, starch, sugar and certain fruits and vegetables selected for their low protein content (tomatoes, mushrooms, carrots, asparagus, apples, oranges, etc.). Of the total nitrogen ingested only 10% was in the form of food proteins, the remaining 90% being supplied as a mixture of amino acids devoid of tryptophane (an acid hydrolysate of casein to which 1% cystine was added). The total nitrogen intake was maintained throughout the experiment at 0.1 g per kilo per day. The intake of non-nitrogenous food was identical for all subjects, providing approximately 800 calories of fat and 1500 calories of carbohydrate per day. The total caloric intake varied in the different subjects between 2400 and 2700 calories per day.

Nitrogen Balance in Experimental Lysine Deficiency in Man.

As part of a general study designed to determine the amino acids essential for man, the nitrogen balance was studied in 5 human volunteers who were given a diet deficient in lysine for periods of 2 to 5 weeks in length. The basic diet and the conditions of the experiment were similar to those outlined in the accompanying report 1 on tryptophane deficiency. The amino acid mixture deficient in lysine was prepared by deamination of casein, a process which has been shown to destroy the biological activity of lysine; 2 this was followed by acid hydrolysis, with the addition of 1.5% tryptophane (to replace that lost during hydrolysis) and of 1% cystine. Before the deficient diet was instituted the subjects were given a diet identical with the experimental diet except that an amino acid mixture containing lysine was supplied. This mixture consisted of an enzymatic hydrolysate of casein, enriched by 1% cystine. A control period on an amino acid mixture containing lysine followed the deficiency period. In subjects Es. and Ha. the amino acid mixture in the control period was identical with that of the experimental period except that 6% lysine was added. In subjects Ra., He. and Hs., the amino acid mixture in the control period was identical with that used in the foreperiod. The observations on nitrogen balance in 4 male subjects are shown in Fig. 1. It may be seen that each subject developed a negative nitrogen balance within a few days after the lysine-deficient diet was started, and that nitrogen equilibrium was promptly restored when lysine was again supplied. The failure to maintain nitrogen equilibrium in the absence of an adequate supply of this amino acid indicates clearly that lysine is a dietary essential for the human subject.

Biosynthesis of Nicotinamide in Man.

Summary (1) Four subjects subsisted for a period of 3 months on a diet of purified vitamin-free foods furnishing a daily intake of 1.5-2.0 mg nicotinamide. (2) There were no symptoms or signs of nicotinic acid deficiency observed during this period. (3) The urinary output expressed as N-methylnicotinamide chloride showed no reduction suggestive of depletion of nicotinamide stores, a finding which suggests synthesis of the vitamin by the intestinal bacteria. (4) An attempt to inhibit bacterial synthesis of nicotinamide in the intestinal tract was not reflected by a drop in the urinary excretion of N-methylnicotinamide (F2). (5) This suggests that nicotinamide synthesis in the intestinal tract was accomplished by bacteria which were not sensitive to sulfasuxidine in the doses used.

A Biochemical Lesion of Lysine Deficiency in Man.

In a previous report 1 it was shown by nitrogen balance studies that lysine is a dietary essential for man. Observations on the urine were made at that time 2 in lysine-deficient subjects, but with the exception of an increased sulfate excretion no very definite changes were encountered. The present report describes further experiments which have confirmed our previous finding of a negative nitrogen balance and have extended our studies of the urinary constituents in this deficiency. These studies have revealed a striking rise in the non-ketone organic acids of the urine in this deficiency. The lysine-deficient diets for the 3 subjects in the present study (No. 4 ♀, No. 5 ♂ and No. 6 ♂) were similar to those described in our previous report. The protein moiety was provided by an acid hydrolysate of deaminized casein, supplemented by 1.5% l-tryptophane and 1% l-cystine; this was fed in quantities of 0.1 g N/kg/day. The total caloric intake of the subjects varied from 2400-2700 cals./day. Since these subjects had been accustomed to a somewhat larger nitrogen intake we attempted to avoid the delay in reaching an initial nitrogen equilibrium by maintaining them on a protein-free diet for two days. This was followed by a 4-day control period in which nitrogen was supplied in the quantity mentioned above in the form of a lysine free amino acid mixture which had been supplemented by 6% l (+) lysine. It had been planned to submit the subjects to the lysine-deficient diet for a 24-day period; however, 5 days after this regimen had been instituted, all 3 subjects complained of nausea, dizziness and hypersensitivity to metallic sounds. So objective signs were, however, discovered by clinical examinations. A study of the curves in Fig. 1 reveals that a definite negative nitrogen balance had developed in all three subjects during this brief period.

Thiamin Excretion Tests in Children with Paralytic Poliomyelitis

Summary The results of thiamin excretion tests in 7 children having paralytic poliomyelitis did not differ in the main from those found in normal children. This finding indicates that the status of thiamin nutrition need not be a factor in determining whether infection by poliomyelitis virus shall result in the paralytic, or in one of the nonparalytic forms of the disease. Some evidence has also been obtained that nicotinic acid deficiency need not be a prerequisite for the development of the paralytic form of the disease. The role of other members of the vitamin B complex in relation to paralytic poliomyelitis remains to be investigated.

The Presence of an Unknown Factor in Serum Which Influences Calcification.

It has been shown by Shipley 1 and the observation has been confirmed by Shipley, Howland and Kramer 2 that calcification of slices of cartilage and bone from a rickety animal takes place in vitro within 48 hours when the slices are immersed in normal blood serum at 37° C. In the course of a series of experiments upon calcification from solutions of inorganic salts, we have repeatedly observed that calcification takes place only when the concentrations of blood electrolytes are considerably reduced below the concentrations in the blood serum. When a salt mixture is used approximating as closely as possible the inorganic composition of blood serum, 3 no calcification whatever occurs, even in preparations which have been incubated as long as 10 days. A sample experiment is given in the table. It would therefore seem necessary to postulate the existence in the blood serum of some factor which enables calcification to take place in spite of the high concentration of electrolytes. It seems altogether improbable that the serum proteins could be concerned in producing this effect, since proteins in general, owing to their ionic nature, would be expected to inhibit rather than to promote calcification. Moreover, the experiments of Shipley, Howland and Kramer have shown that egg albumen and gelatin actually do inhibit the process. Experiments to determine the nature of this unknown factor are now in progress.