Inhibition of Amino Acid Incorporation into Protein ofYoshida Ascites Hepatoma Cells by Glyceraldehyde

GUIDO G. GuiDOTTi, ALBERTOFONNESU,AND ENRICO CIARANFI(Institutes of General Pathology, Universities oj Milano and Firenze, and C.N.R. Center

for Research in Cell Pathology, University of Milan, Milan, Italy)

SUMMARY

DL-GIyceraldehyde, a known inhibitor of glycolysis which does not appreciably affect respiration, has been found to depress the incorporation of labeled leucine into protein of normal and neoplastic tissues, in vitro. With Yoshida ascites hepatoma cells,the inhibition of amino acid incorporation into protein occurred both in aerobic andanaerobic conditions, the L-isomer being apparently more effective than the D-isomer, at comparable concentrations. In anaerobic conditions, DL-glyceraldehydeand L-glyceraldehyde markedly inhibited glycolysis as well as amino acid incorporationinto protein. In the same conditions, D-glyceraldehyde and a,/8-dihydroxybutyralde-hyde (3-methylglyceraldehyde) inhibited incorporation more effectively than glycolysis. The addition of glucose to the incubation medium partially counteracted theinhibitory effect of DL-,L-, and D-glyceraldehyde on the aerobic incorporation of aminoacid into protein and relieved, at least in part, previously established inhibition.Glyceraldehyde failed to show appreciable effects on the concentrative capacity of thecell for a-aminoisobutyric acid, a model amino acid which shares a common transportsystem with several naturally occurring amino acids.

These results are compatible with the hypothesis that glyceraldehyde inhibitsglycolysis and amino acid incorporation into protein by independent mechanisms.

During the course of studies on energy control of proteinsynthesis in neoplastic tissues, a number of metabolic inhibitors were tested. Among them, glyceraldehyde, whoseL-isomer is known to inhibit glycolysis (14, 16, 19), wasfound to depress the incorporation of labeled amino acidsinto protein of Yoshida ascites hepatoma cells (7).

This article is devoted to a description of the effects ofthe racemic and isomerie forms of glyceraldehyde on theincorporation of labeled leucine into protein of normal andneoplastic tissues under in vitro aerobic and anaerobic conditions. Evidence suggesting that glyceraldehyde inhibitsamino acid incorporation into protein by mechanismsdifferent from those involved in the inhibition of glycolysiswill be presented.

MATERIALS AND METHODS

Preparation of tissues.—Yoshida ascites hepatoma cells(AH-130) were harvested from tumor-adapted Wistaralbino rats, 7-10 days after intraperitoneal transplantation. After a twofold dilution with ice-cold saline, theascitic fluid was centrifuged for 5 minutes at 500 X g at2°C. The packed cells were resuspeiided in cold saline

and recentrifuged as described above. The procedure wasrepeated twice and required approximately 20 minutes.

Received for publication January 10, 1964.

Cell preparations containing a visible sediment of erythro-cytes were discarded. After the last washing the packedcells were resuspended either in phosphate or bicarbonateRinger solution to yield a 1:5 dilution. Samples of thefinal suspension were collected for tissue dry weight determinations. Normal tissues were removed from 12-hourfasted albino rats, Wistar strain, weighing 150-170 gm.Liver and kidney cortex were sliced and collected in coldRinger phosphate. Diaphragm was excised, trimmed inchilled Ringer phosphate, and divided into hemidia-phragms.

Incubation procedure.—Incubation was performed inconventional Warburg flasks at 38°C.,in an atmosphere of

air (aerobic conditions) or of 95 per cent N2 :5 per centC02 (anaerobic conditions). The basic incubation mediawere Krebs-Ringer phosphate (25) for aerobic and Krebs-Ringer bicarbonate (25) for anaerobic experiments. Inexperiments with ascites tumor cells the main compartment of the Warburg flask contained 2.3 ml. of medium,0.2 ml. of the inhibitor solution (always adjusted to anosmolarity of 0.30), and 0.3 ml. of the final cell suspension(8-12 mg. of dry tissue). The side arm of the flask contained 0.5 Mmolesof DL-leucine-l-Cu (specific activity, 3.8mc/mmole) or 0.8 ¿miólesof a-aminoisobutyric acid-l-C14(AIB) (specific activity, 1.5 mc/mmole) dissolved in 0.2 ml.of water. The content of the side arm was tipped into the

900

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GUIDOTHet al.—Inhibitory Effects of Glyceraldehyde 901

TABLE 1EFFECTSOF DL-, L-, ANDD-ÛLYCERALDEHYDEON RESPIRATIONANDAEROBICINCORPORATION

OF LEÃœCINE-1-C14INTOPROTEINOF YoSHIDAAsCITESHEPATOMACELLS

INHIBITORCONC.(mu)00.51.02.03.55.010.015.0OXYGENCONSUMPTION

(M!Oì/mgdrywt/hr)DL-Glyceral-

dehyde8.0

±0.4—7.2

±0.36.9±0.17.2±0.36.9±0.36.8±0.4—L-Glyceraldehyde7.1

±0.57.7±0.87.0±0.86.3±0.46.5±0.40.4±0.3——D-Glyceraldehyde7.2

±O.G——7.2

±0.5—6.9

±0.46.4±0.36.5±0.6INCORPORATION

INTOPROTEIN(counts,'min/mgprotein)DL-Glyceraldehydc1330

±121—913

±146868±63660±89448±102307±7—L-Glyceraldehyde1110

±175816

±135710±69590±68476±105391±90——D-Glyceraldehyde1100

±222——799

±115—743

db134478±48294±46

Ascites hepatoma cells (8-12 mg., dry weight) wore incubated at 38°C.for 60 minutes in Krebs-Ringer phosphate containing 0.5 Amólesof DL-leucine-1-C14(sp. act., 3.8 mc/mmole). Medium volume,3 ml. Gas phase: air. The results are mean values of three experiments ±S.E.

main compartment after a thermal equilibration period of10 minutes. When oxygen consumption was measured,0.2 ml. of 20 per cent KOH was present on a roll of filterpaper in the central well. The inhibitors used, DL-, L-,and D-glyceraldehyde and a,/3-dihydroxybutyraldehyde(3-methylglyceraldehyde), were added at different concentrations between 0.5 and 20 mM (final). When present, glucose was 15 IHM. The final osmolarity of the incubation medium was always adjusted to 0.30-0.31. Insome experiments, labeled leucine substituted for theinhibitor in the main compartment of the flask, and viceversa. Normal tissues were incubated in a final volumeof 3 ml.

Oxygen consumption and anaerobic glycolysis weremeasured by standard manometric Warburg procedures(25). Readings were taken every 10-15 minutes aftertipping. At the end of the incubation period, trichloro-acetic acid was added (1 per cent, final concentration) to

TABLE 2EFFECTSOF DL-GLYCERALDEHYDEONRESPIRATIONANDAEROBIC

INCORPORATIONOF LEUciNE-1-C14INTOPROTEINOF NORMALRAT TISSUES

TISSUELiver

KidneyDiaphragmOXYGEN

CONSUMPTIONOilOs/mg drywt/hr)Control6.

6 ±0.4(5)8.1 ±0.2(5)2.3±0.1(5)DL-Glyceral-

dehyde6.1

±0.4(5)8. 3 ±0.5(5)2.3±0.1(5)INCORPORATION

INTOPROTEIN(counts/min/mgprotein)Control585

±67(8)280±19(5)39 ±4(3)DL-Glyceral-

dehyde167

±25(8)83 ±7(5)15 ±2(3)

Tissue slices and hemidiaphragms (10-20 mg., dry weight)were incubated at 38°C.for 60 minutes in Krebs-Ringer phosphatecontaining 0.5 jumólesof DL-leucine-1-C14(s.a., 3.8 mc/mmole).Medium volume, 3 ml. Gas phase: air. The concentration ofDL-glyceraldehyde was 10 HIM (final). The results are meanvalues ±S.E. No. of experiments in parentheses.

TABLE 3EFFECTSOF DL-, L-, ANDD-GLYCERALDEHYDEONGLYCOLYSISANDINCORPORATIONOF

LEüciNE-1-C14INTOPROTEINOF YOSHIDAASCITESHEPATOMACELLSUNDERANAEROBICCONDITIONS

INHIBITORCONC.(mi.)00.51.02.03.55.010.015.0GLYCOLYSIS

(/il COs/mg drywt/hr)DL-Glyceral-

dehyde42.7

±0.240.8±1.132.4±1.425.9±2.416.0±1.18.4±1.34.0±2.1—L-Glyceral

dehyde44.6

±0.935.0±0.221.7±1.511.0±1.05.0±1.03.5±0.2——D-Glyceral-

dehyde41.3

±0.6————41.8

±1.734.6±6.225.5±0.8INCORPORATION

INTOPROTEIN(counts/min/mgprotein)DL-Glyceral-

dehyde1630

±1901650±1491710±1031210±306430±10197±4033±7—L-Glyceral

dehyde2270

±1662390±981660±127319±13788±1335±8——D-Glyceral-denyde1910

±289————1720

±1821080±136336±78

Ascites hepatoma cells (8-12 mg., dry weight) were incubated at 38°C.for 60 minutes in Krebs-Ringer bicarbonate containing 0.5 /umolesof DL-leucine-1-C14(s.a., 3.8 mc/mmole) and glucose, 15 HIM.Medium volume, 3 ml. Gas phase: 95 per cent N2:5 per cent CO2. The results are mean values ofthree experiments ±S.E.

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902 Cancer Research Vol. 24, June 1964

stop the reaction and to precipitate proteins. The sedi-mented proteins were then purified according to Rabino-vitz, Olson, and Greenberg (22), plated and countedbeneath a mica-window Geiger-Mueller counter. Thecounting error was less than 2 per cent. Departures fromthis routine procedure will be described in the accompanying charts.

In permeability experiments with AIB, incubation wasstopped by chilling the flasks in an ice-bath. The cellsuspension was centrifuged at 0°C.,and AIB extraction

was carried out as previously described (8). Samples ofextraction fluid were transferred to stainless steel planchéispainted with shellac to bind the residue, evaporated undera lamp, and assayed for radioactivity in a gas flow counterfitted with a mylar window. Intracellular radioactivitywas calculated by correcting for the material trapped inthe extracellular phase (8, 9).

Chemicals.—OL-Leucine-1-C14 and a-aminoisobutyricacid-l-C14 were obtained from the C.E.A., Gif-sur-Y vette,France, and from the Radiochemical Centre, Amersham,England. Unlabeled AIB was purchased from MannResearch Laboratories, New York. All these compoundswere shown to be chromatographically pure by a unidimensional descending system which minimizes amino acidbreakdown (11). DL- and D-Glyceraldehyde (chromatographically pure) were obtained from Mann Research Lab-

TABLE 4EFFECTS OF O^-DIHYDROXYBUTYRALDEHYDEON GLYCOLYSISAND

INCORPORATIONOF LsuciNE-l-C14 INTO PROTEIN OF

YOSHIDA ASCITES IÕEPATOMACELLS UNDERANAEROBICCONDITIONS

IMBIBITO»cone,(mil)0

10GLYCOLYSIS

(¡AC0t/mg drywt/hr)43.7

±1.8(5)33.8 ±0.6(5)INCORPORATION

INTO PROTEIN(counts/min/mgprotein)2760

=t 155(4)272 =fc115(4)

The experimental conditions were as in Table 3. The resultsare mean values ±S.E. No. of experiments in parentheses.

oratories, New York. L-Glyceraldehyde and a,/î-dihy-droxybutyraldehyde were kindly supplied by LaboratoriFarmitalia, Milan, Italy, and were shown to be chromatographically pure (4).

RESULTSThe results presented in Table 1 show that DL-,L-, and

D-glyceraldehyde strongly inhibited the aerobic incorporation of labeled leucine into protein of ascites hepatomacells at concentrations which had little effect on oxygenconsumption. D-Glyceraldehyde appeared to be less ef-

CLtrooî O

15 30 60 75 90 105 120

MINUTES OF INCUBATIONChart 1.—Effect of glucose on the inhibition of DL-leucine-1-C14

incorporation into protein of Yoshida ascites hepatoma cells byDL-glyceraldehyde. Cells were incubated in air at 38°C. Glucose(15 HIM,final concentration) was added after 45 minutes of incubation (arrow). Curves represent leucine incorporation into proteinin the absence (O) and in the presence of glyceraldehyde (5 HIM)without (D) and with H glucose added. Points are means oftriplicate determinations.

TABLE 5EFFECTS OF DL-, L-, AND D-GLYCERALDEHYDEON AEROBIC INCORPORATIONOF LEUciNE-1-C"

INTO PROTEIN OF YOSHIDA ASCITES HEPATOMACELLS IN THE ABSENCEAND IN THE PRESENCE OF GLUCOSE

INCORPORATIONINTOPROTEIN(counts/min/mg protein)

INHIBITORCONC.(mu)01.02.03.55.010.015.020.0DL-GLYCERALDEHYDENo

glucose12101030

(-15%)807(-33%)788(-35%)615(-49%)307

(-75%)——Glucose17202120(+23%)1940

(+13%)1520(-11%)1150(-33%)471

(-73%)——L-GlyceraldehydeXo

glucose1030696

(-33%)692(-33%)643(-38%)516(-50%)280(-73%)——Glucose12201460(+20%)1580(+28%)1380

(+13%)1230(+1%)629(-48%)——D-GlyceraldehydeNo

glucose856692(-19%)689(-19%)—649

(-24%)431(-50%)252(-71%)206(-76%)Glucose12601480

(+18%)1500(+19%)—1440

(+14%)823(-35%)512(-59%)356

(-72%)

The experimental conditions were as in Table 1. When present, glucose had a concentration of 15HIM(final). The results are mean values of two experiments. Per cent effect in parentheses.

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GuiDOTTi et al.—Inhibitory Effects of Glyceraldehyde 903

fective at comparable concentrations. With DL-glyceral-dehyde, similar results were obtained also with normaltissues such as rat liver, kidney, or diaphragm (Table 2).

A marked inhibition of glycolysis and of leucine-1-C14incorporation into protein of ascites hepatoma cells occurred in anaerobic conditions with DL-and L-glyceralde-hyde (Table 3). D-Glyceraldehyde, at higher concentrations, also inhibited incorporation, but it was less effectiveon anaerobic glycolysis.

With ascites hepatoma cells, a moderate impairment ofthe glycolytic reaction was also observed with dihydroxy-butyraldehyde at a concentration which markedly inhibited the incorporation of leucine into protein (Table 4).

In additional experiments, hepatoma ascites cells wereincubated aerobically (95 per cent Oì'.5per cent CCy and

anaerobically (95 per cent N2:5 per cent C02) in a highpotassium, phosphate-bicarbonate medium, as suggestedby Rabinovitz, Olson, and Greenberg (23). Under theseconditions, the inhibitory effect of DL-glyceraldehyde anddihydroxybutyraldehyde were essentially similar to thosepresented above.

When ascites tumor cells were incubated aerobically,glucose appeared to counteract the inhibition of aminoacid incorporation into protein by DL-, L-, and D-glyceral-

TABLE 6EFFECTS OF DL-GLYCERALDEHYDEANDa ^-DIHYDROXYBUTYRAL

DEHYDEON AIB-l-C14 UPTAKE BY YOSHIDA ASCITESHEPATOMACELLS UNDER AEROBIC CONDITIONS

80^

70n3MU»r

60>ic_•o»

50Mt->

40^^^w

30tO>S

20o10iiii

lX*s

/s'XoX

/X/X

^XxX•'/X

/Xo'X

XX/^'

/X/f

/'s^°X

s^^^^^/

^*°'*'^""^XX_^—,'/.p^«,'/

B-<T.Jfa^^f>

^AJgr^f^A>^l

1 t t 11)15 30 45 60 90 1205

i—iC

0)oí_o.4

9^C£rt"^3

'o*jCDO^0»—1

200.croo

0zU

MINUTES OF INCUBATION

Chart 2.—Kinetics oíinhibition of glycolysis and DL-leucine-1-C'4 incorporation into protein of Yoshida ascites hepatoma cellsby L- and D-glyceraldehyde, under anaerobic conditions. Inthese experiments the tracer substituted for the inhibitor in themain compartment of the Warburg flask. After 10 minutes ofthermal equilibration, L-glyceraldehyde (5 mM, final concentration) and D-glyceraldehyde (10 mM, final concentration) weretipped into the main compartment and the incubation was started.Glucose had a final concentration of 15 mM. Points are means oftriplicate determinations. Values of leucine incorporation havebeen corrected for incorporation occurring during preincubation.Legends: Glycolysis in the absence (O) and in the presence ofL-glyceraldehyde (A) or D-glyceraldehyde (a); incorporation ofleucine into protein in the absence • and in the presence ofL-glyceraldehyde (A) or D-glyceraldehyde (H).

ADDITIONSControlsDL-Glyceralde-hyde

(10HIM)o,/3-Dihydroxy-butyralde-hyde

(10 HIM)UPTAKE(¿irnoles

AIB/gm. intracellularwater)30

min.2.07

±0.14(2)2.21

±0.18(2)—60

min.2.67

±0.07(7)2.46±0.13(7)3.38

±0.02(3)DISTRIBUTIONRATIO*30

min.7.88.3—60min.10.09.212.7

Ascites hepatoma cells (8-12 mg., dry weight) were incubatedat 38°C.for 30 and 60 minutes in Krebs-Ringer phosphate containing 0.8 junóles of a-aminoisobutyric acid-l-Cu (s.a., 1.5mc/mmole). Medium volume, 3 ml. Gas phase: air. Theresults are mean values ± S.E. No. of experiments in parentheses.

* pmoles AIB/ml cell water.

AmólesAIB/ml medium

dehyde (Table 5). This effect was marked at low concentration of the inhibitors, where glucose actually stimulatedthe incorporation reaction, and decreased as the concentration of aldehyde increased.

The rate of aerobic incorporation of labeled leucine intoprotein of ascites tumor cells in the absence of inhibitorsproceeded as a linear function of time for at least 2 hours(Chart 1). In these conditions the addition of DL-glyceraldehyde markedly depressed incorporation but never abolished it completely. Similar results were obtained whenL- or D-glyceraldehyde was used. The addition of glucosepartially reversed the inhibitory effect of glyceraldehyde.

Under anaerobic conditions, glycolysis and incorporationof labeled leucine into protein of ascites tumor cells werecompletely blocked within 30-45 minutes after the additionof L-glyceraldehyde (Chart 2). A similar result, as far asleucine incorporation is concerned, was also obtained withD-glyceraldehyde. On the other hand, the o-isomer depressed the glycolytic reaction but never inhibited it completely.

Finally, DL-glyceraldehyde did not impair the concentra-tive capacity of ascites hepatoma cells for AIB-l-C14, apoorly-metabolized model amino acid, whereas a,/3-dihy-droxybutyraldehyde actually enhanced its accumulation inthe cell water (Table 6).

DISCUSSION

It has been known for a long time that glyceraldehydeinhibits the formation of lactic acid from glucose in avariety of normal and malignant tissues (1-3, 5, 10, 13,15, 20, 24). In contrast, glyceraldehyde, at comparableconcentrations, fails to show a significant inhibitory effecton cell respiration. More recently, Lardy, Wiebelhaus,and Mann (14) found that the inhibition of glycolysis byglyceraldehyde is dependent upon the condensation of theL-isomer with triósephosphate to form L-sorbose-1-phos-

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904 Cancer Research Vol. 24, June 1964

phate which, in turn, depresses hexokinase activity. Under the same experimental conditions, the D-isomer, whichupon condensation with trióse phosphate yields the naturally occurring ester D-fructose-1-phosphate, does notaffect hexokinase activity.

Our results indicate that glyceraldehyde is also a potentinhibitor of the reaction of amino acid incorporation intoprotein of normal and malignant tissues and establish theconditions under which this inhibition can be obtained.

The occurrence of the inhibition of amino acid incorporation into protein of normal and malignant tissues underaerobic conditions in the absence of glucose and the factthat this effect can be observed with both L-and D-glyceral-dehyde (Tables 1 and 2) seem to indicate that differentmechanisms are involved in the inhibition of sugar andprotein metabolism. The results of anaerobic experiments, showing that D-glyceraldehyde and «,/î-dihydroxy-butyraldehyde inhibit amino acid incorporation moreeffectively than glycolysis (Tables 3 and 4), lead to thesame conclusion. In these conditions, high concentrationsof L- and DL-glyceraldehyde inhibit glycolysis and incorporation to a similar extent (Table 3). This result,however, is not at variance with the above hypothesis,since, in anaerobiosis, glycolysis is the sole energy sourcefor the cell, and the depression of incorporation is the resultof an energetic failure.

If glyceraldehyde affects glucose and protein metabolismby different mechanisms, a competition between inhibitionof glycolysis and amino acid incorporation would be expected to occur under suitable conditions. That this isindeed the case is shown by the antagonistic effect of glucose on the inhibition of incorporation in aerobic conditions(Table 5), by the prompt restoration of incorporation uponaddition of glucose after the inhibition by glyceraldehydehad been established (Chart 1), and, under anaerobicconditions, by the failure of L- and DL-glyceraldehyde tocause significant inhibition of incorporation at low concentrations still capable of depressing glycolysis (Table 3).A tentative explanation of this competition could be asfollows. When glucose is present, an active glycolysistakes place in hepatoma ascites cells, generating considerable amounts of triósephosphates. Since glyceraldehyde isknown to condense with dihydroxyacetone phosphate andsince the equilibrium of this condensation reaction stronglyfavors hexose phosphate formation (18), intracellular freeglyceraldehyde will decrease, thus decreasing the amountavailable to inhibit ammo acid incorporation. At present,the specific mechanism by which glyceraldehyde depressesthe incorporation of amino acids into protein is uncertain.A depletion of energy supply does not appear to be thecause, since glyceraldehyde does not appreciably affectcell respiration (Tables 1 and 2) and fails to uncoupleoxidative phosphorylation1 under circumstances in whicha strong inhibition of incorporation occurs. Similarly, theresults of experiments with AIB, an amino acid analogwhich undergoes concentrative transfer similar to that ofseveral naturally occurring amino acids (21), argue againsta direct effect of glyceraldehyde on the mechanism bywhich the cell accumulates amino acids intracellularly(Table 6). Experiments in progress show that glyceralde-

1A. Perm, to be published.

hyde condenses, in vitro, with SH-group bearing moleculessuch as cysteine and that the addition of cysteine almostcompletely prevents the inhibitory effect of L-glyceralde-hyde on anaerobic glycolysis of hepatoma ascites cells.Accordingly, one might postulate that, in the intact cell, acondensation of glyceraldehyde with cysteine might causean imbalance of the intracellular amino acid pool, thus impairing protein synthesis. However, conclusive evidenceof such an occurrence is left for further exploration.

Two more findings in our study deserve consideration.First, results presented in Table 1 seem to indicate thatD-glyceraldehyde inhibits aerobic incorporation of aminoacid into protein less effectively than L-glyceraldehyde atcomparable concentrations. However, D-glyceraldehydeis likely to be metabolized at a faster rate than L-glyceraldehyde in neoplastic tissues (6, 12), a result which, in theintact cell, would lead to different intracellular concentrations of the two isomers. If this is the case, our results donot necessarily preclude the possibility that D- and L-glyceraldehyde inhibit the incorporation of amino acidsinto protein to the same extent. Secondly, results shownin Table 3 indicate that D-glyceraldehyde, at high concentrations, exhibits a definite inhibitory effect on anaerobicglycolysis of hepatoma ascites cells. Similar results werereported by Needham and Lehmann (19), with embryonictissues and by Mendel, Strelitz, and Mundell (17) withJensen sarcoma cells. In contrast, D-glyceraldehyde failedto show significant inhibition of glucose phosphorylation byparticle-free preparations and the inhibitory effect of DL-glyceraldehyde in these conditions has been entirely ascribed to the L-isomer (14). The explanation for thisdiscrepancy is uncertain. However, one might postulatethat, in the intact cell, the D-isomer condensing with dihydroxyacetone phosphate might decrease the amount oftriósephosphates flowing through the Embden-Meyerhofpathway.

ACKNOWLEDGMENTSThe authors are indebted to Prof. B. Camerino of the Labora-

tori Farmitalia, Milan, for the generous supply of L-glyceraldehyde and a,/3-dihydroxybutyraldehyde. Valuable technicalassistance by Miss A. Arnaboldi is also acknowledged.

REFERENCES1. ADLER,E.; CALVET,F.; ANDGÃœNTHER,G. Zur Kenntnis der

Glycerinaldehyd-Hemmung des glycolytischen Kohlenhydra-tabbaues. Z. physiol. Chem., 249:40-56, 1937.

2. ASHFORD,C. A. Glycolysis in Brain Tissue. Biochem. J., 28:2229-36, 1934.

3. BAKER,Z. Studies on the Inhibition of Glycolysis by Glyceraldehyde. Biochem. J., 32:332-41, 1938.

4. BORENFHEÃœND,E., ANDBISCHE,Z. A New Spray for Spottingof Sugars on Paper Chromatograms. Arch. Biochem. Biophys.,67:239-40, 1957.

5. BOYLANDE., ANDBOYLAND,M. E. Studies in Tissue Metabolism. XI. The Action of Tumour Slices and Extracts on Different Carbohydrates. Biochem. J., 32:321-31, 1938.

6. GIACCIO,E. I.; KELLER,D. L.; ANDBOXER,G. E. The Production of L-a-Glycerophosphate during Anaerobic Glycolysis inNormal and Malignant Tissues. Biochim. Biophys. Acta, 37:191-93, 1960.

7. CIARANFI,E., ANoFoNNESu,A. Sui Rapporti fra metabolismoenergetico e incorporazione di aminoacidi nelle proteine dellecellule di tumori-ascite. Atti Accad. Naz. Lincei, Rend.,Classe Sci. Fis. Mat. Nat., 32:835-44, 1962.

8. GUIDOTTI,G., ANDMELLi,M. L. Amino Acid Transport by the

on March 24, 2020. © 1964 American Association for Cancer Research.cancerres.aacrjournals.org Downloaded from

GuiDOTTi et al.—Inhibitory Effects of Glyceraldehyde 905

Hepatic Cell in Cloudy Swelling. Proc. Soc. Exp. Biol. Med.,110:719-22, 1962.

9. HELMREICH,E., ANDKIPNIS, D. M. Amino Acid Transport inLymph Node Cells. J. Biol. Chem., 237:2582-89, 1962.

10. HOLMES,B. E. Glucose and Hexosediphosphate Breakdown inTumour Tissue. Biochem. J., 31:1730-35, 1937.

11. HUOGINS, A. K., ANDMOSES, V. Breakdown of 2-14C-Glycineduring Paper Chromatography. Nature, 191:668-70, 1961.

12. LANDAU,B. R., ANDMERLEVEDE, W. Initial Reactions in theMetabolism of D- and L-Glyceraldehyde by Rat Liver. J. Biol.Chem., 238:861-67, 1963.

13. LARDY, H. A., ANDPHILLIPS, P. H. Inhibition of Sperm Gly-colysis and Reversibility of the Effects of Metabolic Inhibitors. J. Biol. Chem., 148:343-47, 1943.

14. LARDY, H. A.; WIEBELHAUS, V. D.; AND MANN, K. M. TheMechanism by Which Glyceraldehyde Inhibits Glycolysis. J.Biol. Chem., 187:325-37, 1950.

15. MENDEL, B. Krebszelle und Glycerinaldehyd. Klin. Wochschr.,8:169-70, 1929.

16. MENDEL, B.; STRELITZ, F.; AND MUNDELL, D. L-GlycericAldehyde and Tumor Metabolism. Science, 88:149-50, 1938.

17. . D-Glyceric Aldehyde and Tumour Glycolysis. Nature,141:288, 1938.

18. MEYERHOF, O.; LOHMANN,K.; AND SCHUSTER,P. Ãœber die

Aldolase, ein Kohlenstoff-verknüpfendes Ferment. II. Aldol-kondensation von Dioxyacetonphosphorsäure mit Glycerinaldehyd. Biochem. Z., 286:319-35, 1936.

19. NEEDHAM,J., ANDLEUMANN,H. Intermediary CarbohydrateMetabolism in Embryonic Life. VIII. Glyceraldehyde andGlucolysis. Biochem. J., 31:1913-25, 1937.

20. NEEDHAM,J., ANDNOWINSKI, W. W. Intermediary Carbohydrate Metabolism in Embryonic Life. I. General Aspects ofAnaerobic Glucolysis. Biochem. J., 31:1165-84, 1937.

21. NOALL, M. W.; RIGOS, T. R.; WALKER,L. M.; ANDCHRISTEN-sen, H. N. Endocrine Control of Amino Acid Transfer. Science,126:1002-5, 1957.

22. RABINOVITZ,M.; OLSON, M. E.; ANDGREENBERQ, D. M. Independent Antagonism of Amino Acid Incorporation intoProtein. J. Biol. Chem., 210:837-49, 1954.

23. -. Relation of Energy Processes to the Incorporation ofAmino Acids into Proteins of the Ehrlich Ascites Carcinoma.Ibid., 213:1-9, 1955.

24. SÃœLLMANN,H. O. Wirkung des Glycerinaldehyds auf die Gly-kolyse in der Linse. Klin. Wochschr., 16:1583-84, 1937.

25. UMBREIT, W. W.; BURRIS, R. H.; ANDSTADFFER,J. F. Mano-metric Techniques, 3d ed. Minneapolis: Burgess PublishingCo., 1959.

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1964;24:900-905. Cancer Res   Guido G. Guidotti, Alberto Fonnesu and Enrico Ciaranfi  Ascites Hepatoma Cells by GlyceraldehydeInhibition of Amino Acid Incorporation into Protein of Yoshida

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