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Home , Scurvy, Thiamine deficiency, Wernicke encephalopathy

The Effect of Deficiency on Erythrocyte Metabolism

Stanley J. Wolfe, … , Myron Brin, Charles S. Davidson

J Clin Invest. 1958;37(11):1476-1484. https://doi.org/10.1172/JCI103739.

Research Article

Find the latest version: https://jci.me/103739/pdf THE EFFECT OF ON HUMAN ERYTHROCYTE METABOLISM 1, 2 By STANLEY J. WOLFE,8 MYRON BRIN, AND CHARLES S. DAVIDSON (From the Thorndike Memorial Laboratory, and Second and Fourth [Harvard] Medical Serv- ices, Boston City Hospital, and the Department of Medicine, Harvard Medical School, Boston, Mass.) (Submitted for publication August 14, 1957; accepted June 26, 1958) When mature mammalian erythrocytes are in- rapid clearing of ophthalmoplegia ensues (8). cubated in a medium there is a low level It was felt, therefore, that patients with Wernicke's of oxidation, but when methylene blue is added exhibiting extraocular muscle to the reaction mixture oxygen consumption and constituted a clearly defined example of glucose utilization are greatly increased as a human thiamine deficiency, and erythrocytes of result of activation of the pentose phosphate path- nine patients manifesting this disorder were stud- way (1, 2). Non-nucleated erythrocytes possess ied and compared with those from well-nourished the enzyme potential to recycle 5-carbon frag- controls and with those from a group thought to ments (pentose) to glucose-6-phosphate but not be deficient of B1 but not exhibiting opthalmo- to oxidize glucose-6-phosphate to pentose and plegia. carbon dioxide. Methylene blue, by virtue of its ability to carry electrons directly to oxygen, ac- THEORETICAL CONSIDERATIONS celerates the mechanism for glucose oxidation, Provided the necessary substrate, enzymes and permitting the subsequent nonoxidative reactions cofactors are present, the carbon compounds of of the pentose phosphate pathway to follow. Thi- the pentose phosphate pathway continuously re- amine pyrophosphate is an essential for cycle with regeneration of glucose-6-phosphate a reaction in this pathway (3, 4) and erythrocytes (9). Oxidation of glucose-6-phosphate and sub- of thiamine-deprived rats, when incubated with sequent decarboxylation of 6-phospho-gluconate methylene blue, have clearly exhibited impairment produces carbon dioxide arising from the first car- of the thiamine-dependent reaction (5-7). bon and the residue is a pentose, the first carbon In this report, observations have been extended of which was the second carbon of the original glu- to human thiamine deficiency. Studies from this cose molecule: laboratory have demonstrated that the opthalmo- (1) C plegia of Wernicke's encephalopathy is not affected (2) C (2) C by bed rest, withdrawal and administration (3) C --- (1) CO2 + (3) C - of a purified diet containing ascorbic acid and all (4) C (4) C of the B other than thiamine, but that (5) C (5) C when thiamine alone is added to the purified diet, (6) C (6) C Hexose- Pentose- I This paper was presented in part at the annual meet- phosphate phosphate ing of the American Society for Clinical Investigation, Subsequent reactions are nonoxidative. Ribu- May 6, 1957, Atlantic City, N. J. 2 This work was supported in part by a contract from lose-5-phosphate, the pentose phosphate resulting the Office of The Surgeon General, Department of the from the oxidation of 6-phospho-gluconate, can Army; in part by a fellowship (HF-5688) from the be converted to xylulose-5-phosphate and ribose-5- National Institute, Service; in part by enzymes (epimerase, isomerase) by a grant from The Foundation, Inc., New phosphate York, N. Y.; and in part by a grant from Merck Sharp & that are widely prevalent in animal tissues (10). Dohme, Division of Merck & Co., Inc., Rahway, N. J. Pentose phosphate may then participate in the All grants were made to Harvard University. reaction where thiamine pyrophos- 8 Public Health Service Research Fellow of the Na- tional Heart Institute. phate acts as an essential cofactor (3, 4), and in 1476 THIAMINE DEFICIENCY AND ERYTHROCYTE METABOLISM 1477 which the active glycolaldehyde portion of xylu- sugar, sedoheptulose-7-phosphate. The second lose-5-phosphate condenses with ribulose-5-phos- carbon of the original glucose molecule is now the phate (11). The product is the seven carbon first carbon of the sedoheptulose: .1 (2 )C ' (2) C (2) C (3) C: (3) C transketolase (3) C (4) C + (4) C - + (2) C (4) - C (5) C (5) C thiamine (3) C + (5) C (6) C (6) C pyrophosphate (4) C (6) C (5) C (6) C triose-phosphate Pentose-phosphate Heptulose-phosphate In the transaldolase reaction (12) the first three new hexose molecule corresponds to the second carbon atoms of heptulose phosphate condense carbon of the original glucose and CO2 recovered with the triose phosphate residue regenerating from a second cyclic oxidation is derived from the hexose phosphate. The first carbon atom of this original second carbon:

(2) C{ (3) Cl (2) C transaldolase (3) C (3) C (3) C + (4) C - * (2) C ------(2) CO2 + (2) C (4) C (5) C (4) C (4) C (5) C (6) C (5) C (5) C (6) C (6) C (6) C Heptulose- TricDse- Hexos Pentose- phosphate phosp hate phosphLate phosphate In accord with these considerations it has been mally and the first carbon of the molecule would be found that when normal human erythrocytes are recovered as C-1-C1402 in the expected amount. incubated with methylene blue and glucose la- Impairment of the transketolase reaction and sub- beled with radioactive carbon (C14) in the first sequent lessened regeneration of new hexose phos- position, and, separately, with glucose labeled phate, the first carbon of which would represent with C14 in the second position, the recoveries of the original carbon-2, would result in diminished carbon-i carbon dioxide (C-1-Cl402) and C-2- C-2-C1402 recovery. As the reaction is altered C1402 each fall within relatively narrow limits after the decarboxylation of hexose, pentose would (Table I) (1). The first two carbon atoms of the be expected to accumulate. Confirmation of these glucose molecule are those principally involved in expected results has been achieved utilizing the the oxidative process conforming to the observa- rats tion that 95 per cent of radioactive carbon dioxide erythrocytes of thiamine-deprived (5-7). recovered when erythrocytes and methylene blue MATERIALS AND METHODS were incubated with uniformly labeled glucose could be accounted for by recoveries of radio- Patients. All patients with suspected Wernicke's en- cephalopathy and/or beriberi brought to the authors' at- active CO2 when glucose labeled only in the first tention after their admission to the Boston City Hospital and second positions was used as substrates (1). are included in this study. Theoretically, thiamine deficiency would inter- Wernicke's encephalopathy (Table II). Nine sub- jects (seven male, two female) were studied. In eight, fere with the cycle at the transketolase reaction. initial observations were made prior to thiamine ad- Oxidation of hexose phosphate would begin nor- ministration and one (J.W.) was first studied three 1478 STANLEY J. WOLFE, MYRON BRIN, AND CHARLES S. DAVIDSON

TABLE I C14 recovery, QO2 and pentose accumulation in erythrocytes of 20 control subjects

C-1 C-2 recovery recovery % counts % counts Pentose Patient Age Sex added added Q02 accumulation Diagnosis pl.13 hrs. pg./fask W. B. 20 F 41.6 19.3 111 132 Normal G. F. 41 M 41.2 21.3 125 120 Cirrhosis J. B. 54 M 33.9 15.7 104 113 Cirrhosis R. W. 19 F 37.6 18.0 123 121 Normal C. D. 46 M 34.1 16.6 104 138 Normal E. H. 36 F 39.5 13.1 106 158 Normal D. M. 26 F 31.0 14.9 103 149 Normal W. B. 55 M 32.6 12.7 97 109 Cardiac C. B. 22 F 34.6 16.5 114 121 Enteritis J. C. 46 M 40.1 16.6 100 144 Penicillin reaction M. D. 41 F 40.7 20.2 112 118 Cirrhosis J. M. 60 M 35.3 17.9 110 135 Cirrhosis A. R. 67 M 36.3 17.4 98 135 Gastritis M. W. 33 F 30.8 14.4 86 129 Normal J. R. 54 M 34.5 13.9 102 135 Bronchiogenic carcinoma L. B. 46 M 32.1 18.0 127 154 Pneumonia S. W. 33 M 34.5 15.9 102 140 Normal A. B. 36 F 31.5 16.6 102 133 Normal A. S. 46 F 35.0 18.9 126 Thyrotoxicosis J. S. 30 M 37.8 14.1 135 136 Normal Mean S.D. 35.7 d 3.5 16.6 4i 2.3 109 12 133 4 13

hours after thiamine injection. The diagnosis was es- to admission. Control subjects had had normal diets for tablished by exhibition of weakness or paralysis of both at least several weeks. lateral rectus muscles which completely cleared in from Initial treatment and obtaining of samples. Intrave- two hours to four days after thiamine treatment. Other nous glucose was administered until examination by one signs consistent with the Wernicke syndrome (13) in- of the authors established the diagnosis, at which time cluded in seven (one developing only after blood was obtained for study. Thiamine hydrochloride, return of lateral rectus function), peripheral neuritis in 100 mg., was then administered intravenously or intra- seven, and delirium in eight, and in all. muscularly and patients were subsequently fed according Four had smooth, red tongues. A diagnosis of portal to clinical indications. Blood samples were obtained cirrhosis of the alcoholic type was made on clinical periodically after therapy as outlined in Table II, and grounds in three patients. clinical status ascertained by repeated physical exami- Suspected thiamine deficiency (Table III). Eight sub- nation. In one case of Wernicke's encephalopathy, thi- jects (six male, two female) in this category, all alco- amine had been given 3 hours before blood was obtained. holics, were studied prior to thiamine administration. In Samples not immediately studied were refrigerated for five (E.B., J.B., A.K., F.C., C.A.) exhibiting ataxia, a maximum of 8 hours. We have observed no loss of peripheral neuritis and confusion, Wernicke's encepha- activity in either normal or deficient cells so stored for lopathy was suspected but eliminated as ophthalmoplegia 12 hours or less. was absent. One (G.T.) had clinical , as well as Methods. Heparinized blood was centrifuged at 2,500 nystagmus and peripheral neuritis. In two subjects with rpm for 20 minutes at room temperature. Plasma and , and peripheral neuritis, beriberi was buffy coat were removed and packed erythrocytes were initially suspected, but was later discarded when other diluted to approximately 50 per cent in phosphate saline organic heart disease was recognized. buffer, pH 7.4 (NaCl, 0.015 M; KCl, 0.004 M; MgCl,, Control group (Table I). Twenty individuals (11 0.005 M; Na2HPO4-NaHPO, 0.02 M). Hematocrit male, 9 female), 9 of whom were normal laboratory per- was determined for each sample. sonnel and 11 hospital convalescents, were studied. Standard Warburg technique was used. Into the outer Diet evaluation. All patients with Wernicke's en- chamber of 10 ml. Warburg flasks were placed 0.4 ml. cephalopathy and suspected thiamine deficiency were prepared erythrocytes, 0.1 ml. 0.05 per cent methylene chronic alcoholics and had been imbibing heavily before blue, 0.1 ml. 10 mg./ml. solution of C14-labeled glucose,4 admission. Many had been found in rooms or on the and 0.4 ml. buffer; and into the center well, 0.2 ml. 15 street in states of inebriation and physical collapse. Diet per cent KOH. After equilibration for five minutes the histories were unreliable but it was presumed that all C14-labeled glucose obtained from H. S. Isbell, Na- had failed to ingest solid for at least one week prior tional Bureau of Standards, Washington, D. C. THIAMINE DEFICIENCY AND ERYTHROCYTE METABOLISM 1479

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v 4 .R . 4 co16 . Cd :g ..ivCd ,:co 4 ~c/IE a (: 0 J)*c~ oj ;. vi Owl,) ;.0 .4 .o ;. 1480 STANLEY J. WOLFE, MYRON BRIN, AND CHARLES S. DAVIDSON

TABLE III C14 recovery, Q02 and pentose accumulation in eight patients thought to be thiamine-deficient but not exhibiting ophthalmoplegia C-1 C-2 recovery recovery % counts % counts Pentose Patient added added Q02 accumulation Clinical state pl.13 hrs. g./1fask E. B. 32.5 15.2 108 Ataxia, nystagmus, peripheral neuritis, cirrhosis 41 Female J. B. 37.1 16.7 101 162 Nystagmus, peripheral neuritis 41 Male G. T. 33.5 14.8 107 143 Nystagmus, peripheral neuritis, scurvy 62 Male A. K. 35.9 16.3 94 126 Confusion, ataxia, nystagmus, barbiturate intoxi- 58 cation Male F. C. 44.4 15.0 117 178 Nystagmus, ataxia, peripheral neuritis 54 Female C. A. 33.7 13.6 85 Seizures, nystagmus, peripheral neuritis 53 Male W. D. 36.5 14.7 107 144 Anasarca, CHF*, alcoholic 46 : acute bacterial endocarditis Male A. R. 36.6 16.1 130 122 Anasarca,-CHF, , alcoholic 40 Final Dx: chronic cor pulmonale Male Mean 4 S.D. 36.3 E-3.7 15.3 4 1.0 106 4 14 146 i 21

* Congestive heart failure. mixture was incubated at 38° C. for three hours under erage of four corrected manometer readings (two incu- air. The reaction was terminated by addition of 0.2 ml. bated with glucose-I-C' and two with glucose-2-C') 100 per cent trichloracetic acid and, after equilibration, further corrected to 50 per cent hematocrit. radioactive CO, absorbed in the center well was precipi- Erythrocytes obtained before treatment from seven tated as barium carbonate, plated and counted in a Robin- subjects with Wernicke's encephalopathy were incubated son flow counter. The contents of the outer chamber with thiamine hydrochloride (0.25 mg.) and co- were transferred and diluted with 10 per cent trichlora- carboxylase5 (0.1 mg.) to determine whether the meta- cetic acid. Filtrates were analyzed for pentose by the bolic defect could be corrected in vitro. Serum bilirubin orcinol method (14). concentrations were determined by the method of Ducci Duplicate samples were incubated separately with glu- and Watson (15). cose-i-C" and glucose-2-C", the specific activities of which had been determined by converting to osazones, RESULTS plating and counting. Specific activity of glucose-i-C" When erythrocytes of control subjects were in- ranged between 4,100 and 17,000 cpm per mg. while that cubated with labeled glucose and methylene blue, of glucose-2-Co was between 5,500 and 27,000 cpm per mg. The fraction recovered as C'O, of counts added ini- carbon-1 recovery ranged between 30.8 per cent tially is represented as C-i and C-2, respectively, and is and 41.6 per cent and carbon-2 between 12.7 per the arithmetical average of duplicate recoveries. cent and 21.3 per cent. Minimum Q02 was 86 We have found a linear relationship between three- per three hours and maximum pentose accu- hour oxygen consumption and final hematocrit of pre- pl. pared erythrocytes between 45 and 55 per cent, and fig- 6 Purchased from Nutritional Biochemical Corp., Cleve- ures for QO, were calculated from the arithmetical av- land, Ohio. THIAMINE DEFICIENCY AND ERYTHROCYTE METABOLISM 1481

TABLE IV C14 recoveries, QO and pentose accumulation of various groups studied *

C-1 recovery C-2 recovery % counts % counts Pentose Number of added added QO2 accumulation experiments hrs. jAg./flask pl./3 d Controls 35.7 =i 3.5 16.6 4 2.3 109 1 12 133 1 3t 20 Wernicke's encephalopathy, 36.7 4 3.6 10.2 4 2.1 98 4 15 172 b 18 8 pretreatment (p = <0.01) (p = <0.01) Wernicke's encephalopathy, 31.7 i 4.3 13.8 4 2.1 106 41: 11 148 E 211 8 maximum recovery after treatment Wernicke's encephalopathy, 35.5 4 2.5 12.1 4 1.9 107 i 7 173 :1: 35§ 7 pretreatment, thiamine added in vitro Wernicke's encephalopathy, 33.4 :1 3.8 11.9 d 1.9 105 4 8 178 1 38§ 7 pretreatment, cocarboxylase added in vitro "Suspected" thiamine 36.3 1 3.7 15.3 4 1.0 106 4 14 146 4 21§ 8 deficiency

* All figures are the mean plus or minus standard deviation. Represents 19 experiments. Represents 7 experiments. § Represents 6 experiments. mutation was 158 ,ug. per flask (Tables I and IV). minishing pentose accumulation parallel to in- There appeared to be no relationship between vari- creased carbon-2 recovery was observed. Patient ous individual measurements and age, sex or physi- W. B., suffering from severe disease and cal condition of the donor. hepatic coma, did not have prompt improvement of Erythrocytes of patients with Wernicke's en- ophthalmoplegia and his red cells obtained 18 cephalopathy exhibited evidence of impairment of hours after treatment did not demonstrate in- the transketolase reaction. In seven of eight ex- creased carbon-2 recovery. Four days later, re- periments utilizing red cells obtained prior to covery from hepatic coma and clearing eye signs therapy, carbon-2 recovery was less than the low- were associated with return toward normal car- est control value. Mean carbon-2 recovery for bon-2 recovery. In all other subjects ophthalmo- this group was 10.2 per cent, significantly less than plegia cleared rapidly. that of the control (16.6 per cent) (Table IV). Other signs attributed to thiamine deficiency In five of seven experiments pentose accumula- were slow in improving or failed to disappear. tion was increased and the group mean of 172 yg. Of nine patients with Wernicke's encephalopathy, per flask was significantly different from the con- one achieved complete psychiatric recovery, two trol (133 pg. per flask). The extent of increased exhibited persistent confusion and disorientation pentose accumulation did not correlate with that of at time of discharge, and six were transferred to depressed carbon-2 recovery. Q02 was slightly mental hospitals with diagnoses of Korsakoff's diminished and carbon-1 recovery was not in- psychosis. Nystagmus and peripheral neuritis fluenced. Erythrocytes of J. W., given thiamine also tended to persist. two hours before blood was obtained, demonstrated Erythrocytes of eight subjects suspected of depressed carbon-2 recovery. Observations made thiamine deficiency but not exhibiting opthalmo- from 8 hours to 11 days after therapy revealed a plegia had no impairment of the transketolase re- return toward normal metabolic activity in terms action as evidenced by carbon-2 recoveries, Q02 of increased carbon-2 recovery, increased Q02, and pentose accumulation in the normal range and reduced pentose accumulation in the majority. (Tables III and IV). A decrease in carbon-1 recovery was also noted In seven experiments where deficient erythro- (Table II and IV). In two instances (G. F., cytes of patients with Wernicke's encephalopathy R. S.), however, an increased rather than a di- were incubated in vitro with thiamine and cocar- 1482 STANLEY J. WOLFE, MYRON BRIN, AND CHARLES S. DAVIDSON boxylase, there was only slight increase in carbon-2 in vitro is probably dephosphorylated before enter- recovery and no change in pentose accumulation ing the erythrocyte (16, 17) and its action would (Table IV). be identical to in vitro thiamine. It has also been demonstrated that methylene blue, although stim- DISCUSSION ulating erythrocyte respiration, does not stimulate An impairment of the transketolase reaction best production and indeed explains the decreased recycling of carbon-1 of may inhibit it (19), and possibly accounts for the pentose, the carbon-2 of the original glucose mole- failure of complete recovery in thiamine treated cule, to hexose phosphate and its subsequent re- rat erythrocytes. covery as C1402 in methylene blue activated eryth- While evidence for reduced transketolase ac- rocytes of patients with Wernicke's encephalop- tivity in thiamine-deficient erythrocytes is con- athy. As the first oxidative steps of the pathway vincing from tracer studies, the supporting pen- were not influenced, recovery of C-1-C1402 was tose data, although suggestive, are less clear cut. not appreciably affected whereas that of C-2-C1402 Both glucose remaining from the incubating me- was significantly reduced. These findings are dium and erythrocyte purine nucleosides and nu- consistent with the thesis that dietary thiamine de- cleotides that enter the trichloracetic acid filtrate ficiency led to insufficient erythrocyte thiamine give the orcinol reaction and tend to raise the pyrophosphate as cotransketolase. apparent pentose concentratioin. This would tend Following parenteral administration of thiamine to be greater in the deficient cells since less glucose to the deficient patients, a slow return toward is consumed. Moreover, decreases in pentose ac- normal metabolic activity as evidenced by increas- cumulation did not strictly parallel increased car- ing carbon-2 recovery was observed. Insufficient bon-2 recovery in erythrocytes of treated patients. data are available to determine the time required Erythrocytes of patients known to have been im- for complete recovery. Experience with thiamine- bibing heavily, eating poorly and exhibiting physi- deficient rats indicates this to be slow and incom- cal signs often ascribed to thiamine deficiency, but plete (5, 6). Non-nucleated erythrocytes possess not suffering from ophthalmoplegia, exhibited no a limited capacity to phosphorylate thiamine com- metabolic defect. Moreover, carbon-2 recovery pared with nucleated cells (16). This suggests correlated well with resolution of eye signs in the that a new generation of erythrocytes must be patients with ophthalmoplegia, and the single pa- produced before normal activity can be anticipated. tient whose signs did not rapidly improve also Impaired phosphorlylation of thiamine has also exhibited persisting erythrocyte abnormality. It been demonstrated in patients with hepatic cir- is probable that the red cell phenomenon occurs rhosis (17) and the subject (W.B.) with the most only in the most deficient subjects, i.e., those with severe liver disease exhibited the poorest response ophthalmoplegia, but consideratin should be given to therapy in terms of both regression of eye signs to the possibility that the rapidly reversible aberra- and improvement in erythrocyte metabolism. De- tion interfering with normal cranial func- layed recovery of other coenzymatic functions of tion is related to defective glucose catabolism with thiamine following treatment of patients with red cell and nerve cell being involved simultane- Wernicke's syndrome has recently been reported ously. Available information indicates, however, (18). that mammalian brain glucose catabolism is pri- The failure of in vitro incubation of deficient hu- marily by glycolytic pathways (20-22). Since man cells with cocarboxylase and thiamine to re- nystagmus, ataxia and have sult in significantly increased carbon-2 recovery been clearly related to thiamine deficiency (8, 23- is in contrast to observations with rat erythrocytes 25), normal erythrocyte metabolism in patients in which definite albeit incomplete increased car- with these signs but not ophthalmoplegia may be bon-2 recovery was obtained (5-7). Species dif- due to differences in severity of deficiency required ferences can be invoked since rat erythrocytes are before one or another coenzymatic function of thi- known to have a greater capacity than human amine is impaired. Furthermore, as slow disap- erythrocytes to phosphorylate thiamine to the ac- pearance of these signs is not unusual, the patients tive pyrophosphate (19). Cocarboxylase added involved may have demonstrated stigmata of an THIAMINE DEFICIENCY AND ERYTHROCYTE METABOLISM11483 earlier episode of thiamine deficiency and may ACKNOWLEDGMENT adequacy have been in a state of relative The authors gratefully acknowledge the technical as- at the time they were studied. sistance of the Misses Lois M. Betten, Leonore M. De- Disturbances of carbohydrate intermediary me- Carli and Elsie I. Puro. tabolism recognized in thiamine-deficient states have provided a basis for biochemical evaluation of REFERENCES thiamine deficiency (26). Keto acid concentration 1. Brin, M., and Yonemoto, R. H. Stimulation of the in the blood is known to be increased in thiamine glucose oxidative pathway in human erythrocytes deficiency and has been used to indicate the pres- by methylene blue. J. biol. Chem. 1958, 230, 307. ence of deficiency (27-30). However, these 2. Huennekens, F. M., Liu, L., Myers, H. A. P., and changes have been observed in many diseases ap- Gabrio, B. W. Erythrocyte metabolism. III. Oxidation of glucose. J. biol. Chem. 1957, 227, parently unrelated to thiamine deficiency, for ex- 253. ample, multiple sclerosis (31), cirrhosis (32), 3. Horecker, B. L., and Smyrniotis, P. Z. The coen- and combined systems disease (33), both in the zyme function of thiamine pyrophosphate in pen- fasting state and in response to carbohydrate loads. tose phosphate metabolism. J. Amer. chem. Soc. Other investigations of thiamine adequacy have 1953, 75, 1009. blood con- 4. Racker, E., de la Haba, G., and Leder, I. G. Thia- included direct measurement of the mine pyrophosphate, a coenzyme of transketolase. centration of the vitamin which has not proved J. Amer. chem. Soc. 1953, 75, 1010. useful (34), and measurement of urinary thiamine 5. Brin, M., Shohet, S. S., and Davidson, C. S. Effect excretions which provide only an index of tissue of thiamin deficiency on mammalian erythrocyte thiamine stores (35). metabolism. Fed. Proc. 1956, 15, 224. stimulated eryth- 6. Brin, M., Shohet, S. S., and Davidson, C. S. The Application of methylene blue effect of thiamine deficiency on the glucose oxi- rocyte metabolism to clinical evaluation of thia- dative pathway of rat erythrocytes. J. biol. Chem. mine status in man awaits collection and compila- 1958, 230, 319. tion of considerably more data. Defects noted in 7. Wolfe, S. J. The effect of administration of thia- patients with unequivocal deficiency vary as to mine analogues (oxythiamine and pyrithiamine) on rat erythrocyte metabolism. J. biol. Chem. 1957, degree and are sometimes minimal in individual 229, 801. cases. However, the erythrocyte represents a 8. Phillips, G. B., Victor, M., Adams, R. D., and David- most available tissue for the study of the thiamine- son, C. S. A study of the nutritional defect in deficient state and may provide a basis for its Wernicke's syndrome. J. clin. Invest. 1952, 31, clinical measurement. 859. 9. Cohen, S. S. The oxidation of glucose and glucose-6- SUMMARY phosphate in Chemical Pathways of Metabolism, D. M. Greenberg, Ed. New York, Academic Press 1. Erythrocytes of patients with Wernicke's Inc., 1954, vol. 1, p. 187. encephalopathy, when incubated with methylene 10. Dickens, F., and Williamson, D. H. Transformation blue and C14-labeled glucose, exhibited failure of of pentose phosphates by enzymes of animal origin. carbon of pentose second carbon of Nature (Lond.) 1955, 176, 400. the first (the 11. Srere, P. A., Cooper, J. R., Klybas, V., and Racker, the original glucose) to recycle to hexose and be E. Xylulose-5-phosphate, a new intermediate in recovered as radioactive CO2, indicating impaired the pentose phosphate cycle. Arch. Biochem. 1955, transketolation of pentose to heptulose. 59, 535. 2. The defect in transketolation was partially 12. Horecker, B. L., and Smyrniotis, P. Z. Purification after treatment of the patient with thia- and properties of yeast transaldolase. 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