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Blood pyruvate and plasma glucose levels during oral and intravenous glucose tolerance tests in obese and non-obese women
Blood Pyruvate and Plasma Glucose Levels During Oral
and Intravenous
Glucose
Tolerance
Obese and Non-obese ByJ.
W. H. DOAR, VICTOR WYNN
in
Women
AND
D. G. CRAMP
creased, the incremental area above the fasting base line was not significantly greater in the obese group. Elevated blood pyruvate levels were observed in certain obese subjects whose glucose tolerance was normal. The fasting blood pyruvate, and area under the oral/intravenous blood pyruvate curve, correlated better with the degree of obesity than with glucose tolerance. It was not possible to determine whether the elevated blood pyruvate levels in obese subjects were due to impaired pyruvate removal or increased formation. (Metabolism 17: No. 8, August, 690-701, 1968)
Studies of venous blood pyruvate and plasma glucose levels before and after the administration of oral/intravenous glucose are described in a group of 98 women, 27 of whom were obese. The fasting plasma glucose level and intravenous glucose tolerance were similar in obese and non-obese groups. Oral glucose tolerance, however, was impaired in the obese group. The mean blood pyruvate level was elevated in the obese group, both before and after oral/intravenous glucose administration. While the total area under the oral/ curve was inintravenous pyruvate
0
Tests
IS COMMONLY ASSOCIATED with impaired oral glucose tolerance,1-3 though intravenous glucose tolerance has usually been found to be norma1.2~4 Fasting plasma non-esterified fatty acid (NEFA) levels6 and plasma insulin levels5s6 are often increased in obese subjects. Elevated blood pyruvate levels have been noted in subjects with mild diabetes mellitus7J and during glucocorticoid therapy0 and oral contraceptive administration.10 All three situations are associated with impaired glucose tolerance and often with some degree of obesity. We have investigated the inter-relationships of glucose tolerance, blood pyruvate levels and degree of adiposity in a group of control women. Studies of blood pyruvate levels in obesity are few. Some workers2*11-13 have found no sign&cant difference between fasting blood pyruvate levels in obese and non-obese subjects. Others, I4915 however, have observed the mean fasting blood pyruvate level to be elevated in obesity. BESITY
From
the Alexander
Simpson
Laboratories
for
Metabolic
Research,
St. Mary’s
Hospital
Medical School, London, England. Supported in part by Grant Ph-43-67-1344 from the National lnstitutes of Health, Bethesda, Md. Received for publication October 30,1967. J. W. H. DOAR, M. B. CANTAB, M.R.C.P.: Lecturer, Metabolic Unit, St. Mary’s Hospital, London, England, VICTOR WYNN, M.D. MELB, M.R.C.P., F. C. PATH.: Reader in Human Metabolism, University of London. D. G. CRAMP, A.I.M.L.T.: Research Assistant, Metabolic Unit St. Mary’s Hospital, London, England. 690
691
BLOOD PYRUVATB METHODS
Ninety-eight control female subjects, 27 of whom were 120 per cent or more of their “ideal body weight” were studied. The sources of volunteers were identical to those reported in the previous study of oral and intravenous glucose tolerance.22 None were known to have any endocrine or metabolic disease and none were taking drugs known to affect carbohydrate or intermediary metabolism such as glucocorticoids, oral contraceptives, thyroid hormones, appetite suppressants, and diuretics. All received previous dietary preparation containing at least 200 Gm. carbohydrate for three days before each test and oral and intravenous glucose tolerance tests were performed using glucose loads of 1.0 Cm/Kg. and 0.5 Gm./Kg. body weight respectively by methods previously described.10 When both tests were performed the interval was not greater than two weeks. Venous blood was obtained without cuff pressure from an indwelling cannula at intervals for estimation of plasma glucose by an automated glucose oxidase method16 and blood pyruvate by an automated enzymatic fluorimetric technique.17
Analysis of Results The degree of obesity was assessed by expressing each subject’s weight as a percentage of her “ideal body weight” determined from Tables of the Life Extension Institute of New York City (Documenta Geigy, 1956). The total area under the OGTT glucose curve was used as an index of glucose tolerance,lO and the K value calculated from the intravenous 30-, 40-, 50-, and 60-minute glucose values by the method of least squares, as an index of intravenous glucose tolerance. Blood pyruvate changes following glucose administration were assessed by the total area under the curve and the incremental area above the baseline level. The total area was derived from the expression: A=
xr + Xn + xa f xs + ___
. . . . . . . . Xn-1
2
Where A is the total area under the curve, and x1, xa . . . . . . . . xn represent the individual values. The incremental area was obtained by subtracting xr(n-1) from the total area. The significance of differences of means was assessed by Student’s “t” test and correlation coefficients and partial correlation coefficients calculated by standard methods.37
RESULTS Subjects per
cent
were
classified
into
of “ideal
body
weight”);
3 (27
subjects,
>120
and group obese.
The
ages
age 27 years,
three
of the subjects
range
1753;
group
groups;
group per cent). were
subjects,
Group
similar
2, mean
group 1 (44
2 (27
3 were
in all groups
age 28 years,
subjects, 166-120
80-106 per
considered (group
range
17-44;
cent; to be
1, mean group
3, mean age 33 years, range 17-62).
Fasting Plasma Glucose and Blood Pyruvate Levels The mean fasting plasma glucose level in the obese group (82 4 9 mg./ 100 ml.* ) was not significantly greater than that of group 1 (79 + 7 mg./ 106 ml., P > 0.10) or group 2 (82 2 9 mg./lOO ml.). Figure 1 shows the frequency distribution of the mean fasting blood pyruvate level in these three groups. When subjects were tested on two or more occasions, only values from the first test were used. Three obese sub“Mean + SD.
692
DOAR,
Fig. L-Fasting venous blood pyruvate levels in group 1 ( < 100 per cent IBW), group 2 (100-120 per cent IBW) and group 3 (> 120 per cent IBW). Figures in parentheses refer to numbers of subjects.
WYNN
AND
GROUP (44)
I
GROUP
2
CRAhfP
00000000000000000 qqqq~a!qo-qnoawkaJ~ L~-~~~~-~~ VENOUS
BLOOD
PYRUVATE
MG./
100 MLS.
jects had extremely high fasting blood pyruvate levels and the mean level of group 3 (0.72 * 0.38 mg.1100 ml.) was significantly greater than those of group 1 (0.49 -t- 0.12 mg./lOO ml.) and group 2 (0.50 +- 0.10 mg./lOO ml., P < 0.001). Excluding the three markedly abnormal subjects from the obese group reduced the mean value of group 3 to 0.60 +- 0.16 mg./lOO ml., a value still significantly above those of groups 1 and 2 (P < 0.01, and P < 0.02, respectively). Of the three obese subjects with extremely high fasting blood pyruvate levels two had normal oral glucose tolerance curves with areas of 672 and 674 units and one just exceeded our upper limit of normal (800 units) with an area of 814 units. All three subjects, however, had a positive family history of diabetes mellitus. The mean fasting blood pyruvate level was slightly higher in those subjects with a family history of diabetes. This difference was most marked in the obese group, where the mean values were 0.82 mg./lOO ml. and 0.65 mg./ 100 ml. for subjects with and without a family history of diabetes. Oral Glucose Tolerance Tests were performed on 33 subjects in group 1, 21 subjects in group 2, and 24 subjects in group 3. Oral glucose tolerance, assessed as total area under the curve, was impaired in group 3 (736 -+ 134 units) as compared to group 1 (609 I+ 83 units, P < 0.001) and group 2 (630 f 51 units, P < 0.001). Seven obese subjects in group 3 and one subject in group 1 had OGTT glucose areas above 800 units, a value which we regard as indicative of chemical diabetes in women of this age group.10 The mean fasting blood pyruvate level of the 7 obese subjects with im-
BLOOD
693
FYRUVATE 1.4 _
levels
0.2 _
5 ci x
5 d lz
B d 2
6 0 :
5 d kY
6 d 2
0
I
I
I
I
I
I
30
60
90
\20
z I+
PRE I PRE 2
during
tween the mean values of groups 2 and 3. MINUTES
paired glucose tolerance (0.84 mg./lOO ml.) was greater than that of the 17 obese subjects with normal oral glucose tolerance (0.69 mg./ 100 ml.). The mean blood pyruvate levels during oral glucose tolerance tests are shown in Fig. 2. While the mean values for groups 1 and 2 did not differ significantly, those of group 3 were significantly elevated. The mean OGTT pyruvate areas and incremental areas are shown in Fig. 3. The mean pyruvate area of the obese group (4.27 2 1.25 units) was signikantly greater than that of group 1 (3.05 + 0.52 units) and group 2 (3.20 zk 0.76 units, P < 0.001). When 3 obese subjects with very high fasting pyruvate levels were excluded, the mean pyruvate area of group 3 was 4.00 units, a value significantly above those of groups 1 and 2 (P < 0.01). The mean OGTT pyruvate incremental area of the obese group (1.40 + 0.89 units) was not significantly greater than that of group 1 (1.07 i- 0.58 units) or group 2 (1.30 f 0.66 units). lntracenous
Glucose Tolerance
Tests were performed on 35 subjects in group 1, 22 subjects in group 2 and 24 subjects in group 3. The mean intravenous glucose ‘K’ value of the obese ;;
5.00
t
5
-
4.00
5 % 2
3.00
t y
2.00
Fig. 3.-Mean OGTT pyruvate levels during area and incremental area in groups 1,2 and 3. Bars represent 1 standard error of mean.
PYRUVATE
ii! ii \
too
GROUP
I AREA
2
3
2 INdREMENTAL
3 AREA
694
DOAR,
WYNN
AND
CRAMP
Fig. 4.-Mean blood py ruvate levels during IVGTT’s in group 1 (O), group 2 (A) and group 3 (0 ), Significance levels refer to the difference between mean values of groups 2 and 3.
PRE I
PRE 2
30
60
90
MINUTES
subjects (1.93 f 0.56) was not significantly different from that of group 1 (1.82 + 0.71) or group 2 (1.89 + 0.95). Two subjects in group 1 and one in group 2 had ‘K’ values of less than 0.95. The lowest ‘K’ value found in the obese group, however, was 1.17. The mean intravenous glucose area of the obese subjects (1613 -C 260) was greater than that of group 1 (1330 k 278) and group 2 ( 1498 t 269). The mean intravenous glucose incremental area of the obese subjects (872 I+ 234) was also greater than that of group 1 (624 + 236) and group 2 (753 + 255). Only the differences between groups 1 and 3 were significant (P < 0.001). The mean blood pyruvate levels following intravenous glucose administration to groups 1, 2 and 3 are shown in Fig. 4. The results are similar to those found during the oral glucose tolerance tests. The mean pyruvate levels of obese subjects were significantly greater than those of groups 1 and 2. The mean IVGTT pyruvate areas and incremental areas are shown in Fig. 5. The mean pyruvate area of the obese group (3.32 * 0.75 units) was significantly greater than that of group 1 (2.35 zk 0.55 units, P < 0.001) and group 2 (2.54 2 0.70 units, P < 0.001). When 2 obese subjects with very high fasting blood pyruvate levels were excluded, the mean pyruvate area of group 3 was 3.22 units, a value significantly above those of groups 1 and 2 (P < 0.001). The mean pyruvate incrementa area, however, of group 3
L
3 Q 4.00 Fig. 5.-MeanIVGTT pyruvate area and incremental area in groups 1, 2 and 3. Bars represent 1 standard error of mean.
IVGTT.
_
I
#
-I
3.00
_
2 $ 5 2.00 _ L o
PYRUVATE
-
I
f 1.00 _ a P a0 GROUP
I
2 AREA
3
I 2 INCREMENTAL
3 AREA
695
BLOOD PYRUVATJZ 2x, _ I.8 _
.
l .
I.6 _ 2 i
.
. .
l
l
.
1.4 _
2 a
I.2 _ I.0
_
a
08
_
t w I
0.6
_
;
04_
3
. .
.
l*
.
E
0.2
.
_
E ,’
.
. .
.
.
.
l
l
O_
z t
-a2
_
c’ t:
-0.4
_
.
+
> -:
-0.6
1 I1 0.2 0.4
I OS O.G.T.T.
0.6
I1 I.0
11 1.2
PYRUVATE
1.4
I1 I.6
I.6
1 2.0
INCREMENTAL
2.2
I1 2.4
2.6
I
I
2.6
31)
II 3.2
3.4
AREA (UNITS)
Fig. 6.-Correlation between OGTT pyrnvate incremental area and IVGTT pyruvate incremental area in 61 control women. Regresion line is given by IVGTT pyruvate incremental area = 0.37(OGTT pyruvate incremental area) + 0.53. r = 0.51(P
DOAII, WYNN
696
AND CRAMP
vate area (r = 0.44, P < 0.001) correlated significantly with degree of obesity. The correlation between oral glucose and pyruvate area (r = 0.27, P < 0.05) became non-significant when the effect of degree of obesity was removed by partial correlation (r = 0.13, P > 0.10). In 81 IVGTT’s intravenous pyruvate area correlated with degree of obesity (r = 0.54, P < 0.001). There was no correlation between the intravenous “K” value and any parameter of intravenous blood pyruvate levels. Surprisingly, significant negative correlations were observed between the fasting pyruvate levels and oral and intravenous pyruvate areas (r = -0.29, P < 0.01 and r = -0.37, P < 0.001, respectively). Inspection of the scattergrams, however, showed that these two correlations were largely due to the three obese subjects with extremely high fasting pyruvate levels who had very small or even negative pyruvate incremental areas. When these three subjects were excluded, there was no significant correlation between the fasting pyruvate level and oral/intravenous pyruvate incremental areas in the remaining subjects. DISCUSSION
Although oral glucose tolerance was markedly impaired in the obese group as a whole, the intravenous “K” values were not significantly different in the three groups studied. This suggests a dissociation between the results of oral and intravenous glucose tolerance tests in obesity as has been previously noted.2 The explanation for this dissociation is not clear. The contribution of tissues such as liver, fat and muscle, to the removal of oral or intravenous glucose loads is not known. It is probable that hepatic uptake is relatively more important in the former situation, Conn and Newburgh18 suggested that impaired oral glucose tolerance in obesity might be due to decreased hepatic glycogen formation. Butterfield et al. la~zohave postulated that glucose uptake by muscle is decreased in this condition. They found the rate of uptake of glucose by forearm tissues to be reduced in obese subjects during oral and intravenous glucose tolerance tests. It is difficult, however, to assess the relative contributions of muscle and adipose tissue to glucose removal in this type of study. The lower overall forearm glucose uptake rates observed in their obese subjects may have been due to the large mass of relatively inert adipose tissue removing comparatively little glucose. Since intravenous glucose tolerance does not appear to be impaired in obesity, further work is required to evaluate the uptake of glucose by various tissues in obese and non-obese subjects. Loading doses of glucose based on body weight are not ideal for the study of obesity because the volume of distribution of glucose forms a lesser proresulting in greater extracellular portion of body weight in obese subjects,‘lJ* fluid concentrations of glucose during glucose tolerance tests. These differences, however, are likely to exert only a small effect on conventional parameters of glucose tolerance since West et a1.23 found that in duplicate OGTTs doubling the glucose load from 0.8 g/Kg. to 1.6 g/Kg. resulted in the mean
BLOOD
PYRUVATE
697
60 minute and 120 minute blood sugar levels being about 20 mg./lOO ml. higher. Assuming similar increases for the other points, this would increase the OGTT area by 110 units. Wahlberg4 observed that doubling the glucose load from 25 Gm. to 50 Gm. in 52 subjects caused the mean intravenous glucose ‘K’ value to increase by 0.24. Since these extreme changes caused only small changes in oral and intravenous glucose tolerance, bias resulting from a glucose load based on body weight in the present series would be even smaller. Impaired oral glucose tolerance in obese subjects has been found by manyQ but not all investigators. 24.25 Wahlberg4 using a 25-Cm. glucose load found no significant difference between the mean intravenous “K” value of 56 obese subjects compared with that of 144 control subjects. Vajda et al.,26 however, using a glucose load of 0.5 Gm./Kg. body weight found impaired intravenous glucose tolerance in a group of 25 obese adolescents. They used the time required for the blood sugar to return to the fasting level as a criterion of glucose tolerance. For reasons outlined above, when glucose is administered in amounts proportional to body weight, an obese subject with a “K” value identical to that of a non-obese subject will show higher plasma glucose levels and in consequence the return of the plasma glucose level to normal will be delayed. This was confirmed by the obese subjects in the present study who, despite normal intravenous glucose tolerance as judged by ‘K” values, had a mean total glucose area and incremental area significantly above those of the controls. Morse et al.2 using an intravenous glucose dose proportional to estimated body surface area (a parameter more closely related to the apparent volume of distribution of an intravenous glucose load) found no significant difference between “K” values of obese and non-obese subjects. Significantly elevated levels of blood pyruvate were found both before and during oral and intravenous glucose tolerance tests in the group of 27 obese subjects. The total area under the pyruvate curve was significantly greater during both tests in the obese group. The incremental areas above the base line, however, were not signihcantly greater than those of the non-obese subjects. These results agree with those of Haagenson’” who noted elevated fasting blood pyruvate levels but a normal oral pyruvate incremental area in a group of obese subjects. There were several significant correlations between blood pyruvate and plasma glucose levels. The relation between fasting plasma glucose and blood pyruvate levels (r = 0.21, P < 0.05) was shown to be due to both variables correlating with the degree of obesity and contrasts with that found by Fry and RutterfieldY in a series of 28 diabetics (r = 0.69, P < 0.01). The two studies, however, are not comparable and the range of fasting blood sugar was much greater in the diabetic series. In 61 paired tests there were strong correlations between the fasting blood pyruvate levels in the two tests (r = 0.74, P < 0.001) and oral and intravenous pyruvate incremental areas (r = 0.51, P < 0.001). The latter finding suggests that the ratio of the proportion of the glucose load passing down the glycolytic pathway to pyruvate in the OGTT to that following a similar
698
DOAR, WYNN AND CRAMP
route in the IVGTT was similar for all subjects and, further, that the rate of pyruvate catabolism in a given subject was fairly constant over the period of time between the two tests. When the degree of obesity was taken into account, the correlation between OGTT glucose area and pyruvate area became non-significant (r = 0.13, P > 0.10). Elevated fasting blood pyruvate levels have been found by some workers in mild cases of diabetes meWus though the pyruvate rise following glucose has usually been normal .?,*? It is possible that abnormal pyruvate levels in mild diabetes may be more closely related to the degree of obesity than is the abnormality of glucose tolerance. In the present series, seven obese women had fasting blood pyruvate levels above 0.80 mg./lOO ml. and of these four had oral glucose tolerance areas within our normal range. Raised fasting plasma insulin levels have been found in obese nondiabetic subjects6 and the present findings suggest that an elevated fasting venous blood pyruvate level should be added to the list of metabolic abnormalities found in this condition. In 81 IVGTT’s there was no relation between the “K” value and any parameter of pyruvate metabolism. Although .theWmean.-pyruvate incremental area was greater in obese subjects during both oral and intravenous glucose tolerance tests, the values were not significantly above those of the nonobese groups. Both the oral pyruvate area (r = 0.37, P < 0.001) and intravenous pyruvate area (r = 0.44, P < 0.001) correlated better with the degree of obesity than with any parameter of glucose tolerance. Elevated fasting blood pyruvate levels in obese subjects may be due to increased pyruvate formation and/or decreased pyruvate removal. If pyruvate removal were impaired a positive correlation might have been expected between the fasting level and the intravenous pyruvate incremental area. This, however, was not the case. The three obese subjects with extremely high fasting pyruvate levels had small or negative intravenous pyruvate incremental areas. Their intravenous glucose ‘K’ values were 2.13, 2.39 and 1.76, respectively, indicating that glucose was removed at a normal rate and that impaired pyruvate removal in these subjects is therefore unlikely. Alternatively, increased rates of pyruvate formation may be responsible for elevated fasting blood pyruvate levels in obesity. In this situation, assuming the rate of pyruvate removal to be proportional to the blood pyruvate concentration, the presentation of a pyruvate load, derived from oral/intravenous glucose, should result in a relatively unchanged pyruvate incremental area. In the present series with a glucose load based on body weight, the increment of glucose concentration in the ECF would have been greater in the obese subjects,2* resulting in the formation of more pyruvate. This could account for the slightly greater mean intravenous pyruvate incremental area found in the obese group. Venous blood pyruvate levels to some extent reflect changes in metabolism of local tissues and are not necessarily indicative of events in the body as a whole. Relative anoxia of forearm tissues in obesity could impair pyruvate
699
BLOOD PYRUVATJI
removal but should also lead to increased lactatelpyruvate ratios. No difference was found, however, between venous lactatelpyruvate ratios in obese and non-obese subjects (unpublished observations). Further, similar arterio-venous lactate differences, oxygen uptakes and lactate production rates have been observed in forearm tissues of obese and non-obese subjects.2s The possibility remains that other tissues, such as liver, may be producing greater than normal amounts of pyruvate in obese subjects. The fraction of an oral/intravenous glucose load that passes down the glycolytic pathway to pyruvate is unknown. This proportion may vary with degree of obesity, and the above observations could equally be explained in terms of impaired pyruvate degradation in obese subjects together with a greater proportion of the glucose load being utilised along other routes such as conversion to glycogen, oxidation via the hexose monophosphate shunt, and conversion to cu-glyceromonophosphate in adipose tissue for re-esteriflcation of fatty acids. The hexose monophosphate shunt is more active in adipose tissue than musclez9 and it has been suggested that increased glycogen formation may occur in forearm tissues in obese subjects in the fasting state.2* Clearly, direct studies of pyruvate degradation rates in vivo will be necessary to determine whether pyruvate catabolism is impaired in obese subjects. Lower rates of C1402 excretion were found following carbonyl Cl4 pyruvate administration to a group of obese hyperglycaemic mice compared with a group of controls, suggesting that pyruvate oxidation was impaired in the former.30 The effects of different precursor pool sixes in this type of study, however, are diflicult to evaluate. Addition of fatty acids has been found to impair pyruvate oxidation by heart muscle and diaphragm in vitro31 and it is possible that the elevated plasma NEFA levels found in some cases of obesity5 may have a similar effect. Administration of certain hormones, including thyroxine,32 growth hormone,33 cortisoP and oestrogen-progestagen combinationslO has been found to cause elevated blood pyruvate levels. In general, however, blood concentrations of these hormones are not increased in obesity.34-36 ACKNOWLEDGMENT The authors would like to thank Mr. M. J. R. Healy for statistical advice. REFERENCES 1. John, H. J.: A summary of the findings in 1100 glucose tolerance estimations. Endocrinology 13:388, 1929. 2. Morse, W. I., Sidorov, J. J., Soeldner, J. S., and Dickson, Il. C.: Observations on carbohydrate metabolism in obesity, Metabolism 9:666, 1966. 3. Smith, M., and Levine, It.: Obesity and Diabetes. Med. Clin. N. Amer. 48: 1387,1964. 4. Wahlberg, F.: Intravenous glucose tolerance in myocardial infarction, angina,
pectoris and intermittent claudication. Acta Med. &and. Supp. 453, 1966. 5. Beck, P., Koumans, J. H., Winterling, C. A., Stein, M. F., Daughaday, W. H., and Kipnis, D. M.: Studies of insulin and growth hormone secretion in human obesity. J. Lab. Clin. Med. 64:654, 1964. 6. Kreisberg, Pi. A., Boshell, B. R., DiPlacido, J., and Roddam, R. F.: Insulin secretion in obesity. New Eng. J. Med. 276: 314,1967. 7. Smith, M. J. H., and Taylor, K. W.:
700 Blood pyruvate and a-ketoglutarate in norma1 and diabetic subjects. Brit. Med. J. 2: 1035, 1956 8. Fry, J. K., and Butterfield, W. J. H.: Carbohydrate metabolism in diabetes-a possible intracellular block. Lancet 2:66, 1962. 9. Hennes, A. R., Wajchenberg, B. L., Fajans, S. S., and Conn, J. W.: The effect of adrenal steroids on blood levels of pyruvic and alpha-ketoglutaric acids in normal subjects. Metabolism 6:339, 1957. 10. Wynn, V. W., and Doar, J. W. H.: Some effects of oral contraceptives on carbohydrate metabolism. Lancet 2:715, 1966. 11. Arendt, E. C., and Pattee, C. J.: Studies on obesity II. Blood pyruvate and lactate curves in obese and normal subjects after ingestion of glucose. J. Clin. Endocr. 16:375, 1956. 12. Miller, A. T., and Thomas, B. M.: Pyruvate metabolism in obesity. Amer. J. Clin. Nutr. 4:619, 1956. 13. Stucklikova, E., HruSkova, J., Tenorovi, M., Novotna, B., Komirkova, A., and Riedl, 0.: Some changes in intermediary metabolism of obese patients. Clin. Chim. Acta 6:571, 1961. 14. Nyfos, L., and Skouby, A. P.: Blood pyruvic acid in obesity. Acta Med. Stand. 161:403, 1957. 15. Haagenson, N. R.: Inorganic phosphate and pyruvate in obese patients during glucose assimilation. Acta Med. Stand. 167: 177, 1960. 16. Cramp, D. G.: New automated method for measuring glucose by glucose oxidase. J. Clin. Path. 20:910, 1967. 17. -: Automated enzymatic fluorimetric method for the determination of pyruvic and lactic acids in blood. J. Clin. Path. 21:171, 1968. 18. Conn, J. W., and Newburgh, L. H.: Hyperglycaemia due to impaired hepatic glycogenesis. Proc. Sot. Exp. Biol. Med. 36: 236, 1937. 19. Butterfield, W. J. H., Hanley, T., and Whichelow, M. J.: Peripheral metabolism of glucose and free fatty acids during oral glucose tolerance tests. Metabolism 14:851, 1965. 20. -, Abrams, M. E., St. John, D. J. B., and Whichelow, M. J.: The intravenous glucose tolerance test: Peripheral disposal of the glucose load in controls and dia-
DOAR,
WYNN
AND
CRAMP
betics. Metabolism 16:19, 1967. 21. Fran&son, J. R. M., Conard, V., and Bastenie, P. A.: Measurement of the free glucose diffusion space in man by the rapid intravenous glucose tolerance test. Acta Endoer. (Kobenhavn) 32:463, 1959. 22. Doar, J. W. H.: A comparison of oral and intravenous glucose tolerance tests in 50 control women (to be published). 23. West, K. M., Wulff, J. A., Reigel, D. G., and Fitzgerald, D. T.: Oral carbohydrate tolerance tests. Arch. Intern. Med. 113:641, 1964. 24. Unger, R. H.: The standard twohour oral glucose tolerance test in the diagnosis of diabetes mellitus in subjects without fasting hyperglycaemia. Ann. Intern. Med. 47:1138, 1957. 25. Tyner, J. D.: The prediabetic state: Its relation to obesity and to diabetic heredity. Amer. J. Med. Sci. 185:704, 1933. 26. Vajda, B., Heald, F. P., and Mayer, J.: Intravenous glucose tolerance in obese adolescents. Lancet 1:902, 1964. 27. Anderson, J.. and Marks, V.: Pyruvate in diabetes mellitus-concentrations in urine and blood. Lancet 1:1159, 1962. 28. Rabinowitz, D., and Zierler, K. L.: Forearm metabolism in obesity and its response to intra-arterial insulin, Characterisation of insulin-resistance and evidence for adaptive hyperinsulism. J. Clin. Invest. 41: 2173,1962. 29. Randle, P. J.: Endocrine control of metabolism. Ann. Rev. Fhysiol. 25:291, 1963. 39. Guggenheim, K., and Mayer, J.: Studies of pyruvate and acetate metabolism in the hereditary obesity-diabetes syndrome of mice. J. Biol. Chem. 198:259, 1952. 31. Randle, P. J., Garland, P. B, Hales, C. N., and Newsholme, E. A.: The glucose fatty-acid cycle. Its role in insulin sensivity and the metabolic disturbances of diabetes mellitus. Lancet 1:785, 1963. 32. Macho, L.: The influence of endocrine glands on carbohydrate metabolism. IV. The effect of thyroid hormones on pyruvic acid metabolism. Arch. Internat. Physiol. 70:507, 1962. 33. Henneman, D. H., and Henneman, P. H.: Effects of human growth hormone on levels of blood and urinary carbohydrate and fat metabolites in man. J. Clin. Invest. 39:1239, 1960.
BLOOD
701
I’YHU\‘ATE
34. Lessof.
hl.
H.,
and Greenwood, F. secretion in obese Rep. 115:65, 1966. 35. Schteingart, D. Characteristics of the
McHardy
Young,
S.,
C.: Growth hormone subjects. Guy Hosp. E., and Conn, J. W.: increased adrenocorti-
cal function observed in many obese patients. Ann. N. Y. Acad. Sci. 131:388, 1965.
36. Yalow, and Berson,
R. S., Glick. S. M.. Roth, J., S. A.: Plasma insulin and
growth hormone levels in obesity and diabetes. Ann. N. Y. Acad. Sci. 131:375, 1965. 37. Snedecor, G. W.: Statistical Methods, (ed. 5). Ames, Iowa, Iowa State University Press, 1956.
and Intravenous
Glucose
Tolerance
Obese and Non-obese ByJ.
W. H. DOAR, VICTOR WYNN
in
Women
AND
D. G. CRAMP
creased, the incremental area above the fasting base line was not significantly greater in the obese group. Elevated blood pyruvate levels were observed in certain obese subjects whose glucose tolerance was normal. The fasting blood pyruvate, and area under the oral/intravenous blood pyruvate curve, correlated better with the degree of obesity than with glucose tolerance. It was not possible to determine whether the elevated blood pyruvate levels in obese subjects were due to impaired pyruvate removal or increased formation. (Metabolism 17: No. 8, August, 690-701, 1968)
Studies of venous blood pyruvate and plasma glucose levels before and after the administration of oral/intravenous glucose are described in a group of 98 women, 27 of whom were obese. The fasting plasma glucose level and intravenous glucose tolerance were similar in obese and non-obese groups. Oral glucose tolerance, however, was impaired in the obese group. The mean blood pyruvate level was elevated in the obese group, both before and after oral/intravenous glucose administration. While the total area under the oral/ curve was inintravenous pyruvate
0
Tests
IS COMMONLY ASSOCIATED with impaired oral glucose tolerance,1-3 though intravenous glucose tolerance has usually been found to be norma1.2~4 Fasting plasma non-esterified fatty acid (NEFA) levels6 and plasma insulin levels5s6 are often increased in obese subjects. Elevated blood pyruvate levels have been noted in subjects with mild diabetes mellitus7J and during glucocorticoid therapy0 and oral contraceptive administration.10 All three situations are associated with impaired glucose tolerance and often with some degree of obesity. We have investigated the inter-relationships of glucose tolerance, blood pyruvate levels and degree of adiposity in a group of control women. Studies of blood pyruvate levels in obesity are few. Some workers2*11-13 have found no sign&cant difference between fasting blood pyruvate levels in obese and non-obese subjects. Others, I4915 however, have observed the mean fasting blood pyruvate level to be elevated in obesity. BESITY
From
the Alexander
Simpson
Laboratories
for
Metabolic
Research,
St. Mary’s
Hospital
Medical School, London, England. Supported in part by Grant Ph-43-67-1344 from the National lnstitutes of Health, Bethesda, Md. Received for publication October 30,1967. J. W. H. DOAR, M. B. CANTAB, M.R.C.P.: Lecturer, Metabolic Unit, St. Mary’s Hospital, London, England, VICTOR WYNN, M.D. MELB, M.R.C.P., F. C. PATH.: Reader in Human Metabolism, University of London. D. G. CRAMP, A.I.M.L.T.: Research Assistant, Metabolic Unit St. Mary’s Hospital, London, England. 690
691
BLOOD PYRUVATB METHODS
Ninety-eight control female subjects, 27 of whom were 120 per cent or more of their “ideal body weight” were studied. The sources of volunteers were identical to those reported in the previous study of oral and intravenous glucose tolerance.22 None were known to have any endocrine or metabolic disease and none were taking drugs known to affect carbohydrate or intermediary metabolism such as glucocorticoids, oral contraceptives, thyroid hormones, appetite suppressants, and diuretics. All received previous dietary preparation containing at least 200 Gm. carbohydrate for three days before each test and oral and intravenous glucose tolerance tests were performed using glucose loads of 1.0 Cm/Kg. and 0.5 Gm./Kg. body weight respectively by methods previously described.10 When both tests were performed the interval was not greater than two weeks. Venous blood was obtained without cuff pressure from an indwelling cannula at intervals for estimation of plasma glucose by an automated glucose oxidase method16 and blood pyruvate by an automated enzymatic fluorimetric technique.17
Analysis of Results The degree of obesity was assessed by expressing each subject’s weight as a percentage of her “ideal body weight” determined from Tables of the Life Extension Institute of New York City (Documenta Geigy, 1956). The total area under the OGTT glucose curve was used as an index of glucose tolerance,lO and the K value calculated from the intravenous 30-, 40-, 50-, and 60-minute glucose values by the method of least squares, as an index of intravenous glucose tolerance. Blood pyruvate changes following glucose administration were assessed by the total area under the curve and the incremental area above the baseline level. The total area was derived from the expression: A=
xr + Xn + xa f xs + ___
. . . . . . . . Xn-1
2
Where A is the total area under the curve, and x1, xa . . . . . . . . xn represent the individual values. The incremental area was obtained by subtracting xr(n-1) from the total area. The significance of differences of means was assessed by Student’s “t” test and correlation coefficients and partial correlation coefficients calculated by standard methods.37
RESULTS Subjects per
cent
were
classified
into
of “ideal
body
weight”);
3 (27
subjects,
>120
and group obese.
The
ages
age 27 years,
three
of the subjects
range
1753;
group
groups;
group per cent). were
subjects,
Group
similar
2, mean
group 1 (44
2 (27
3 were
in all groups
age 28 years,
subjects, 166-120
80-106 per
considered (group
range
17-44;
cent; to be
1, mean group
3, mean age 33 years, range 17-62).
Fasting Plasma Glucose and Blood Pyruvate Levels The mean fasting plasma glucose level in the obese group (82 4 9 mg./ 100 ml.* ) was not significantly greater than that of group 1 (79 + 7 mg./ 106 ml., P > 0.10) or group 2 (82 2 9 mg./lOO ml.). Figure 1 shows the frequency distribution of the mean fasting blood pyruvate level in these three groups. When subjects were tested on two or more occasions, only values from the first test were used. Three obese sub“Mean + SD.
692
DOAR,
Fig. L-Fasting venous blood pyruvate levels in group 1 ( < 100 per cent IBW), group 2 (100-120 per cent IBW) and group 3 (> 120 per cent IBW). Figures in parentheses refer to numbers of subjects.
WYNN
AND
GROUP (44)
I
GROUP
2
CRAhfP
00000000000000000 qqqq~a!qo-qnoawkaJ~ L~-~~~~-~~ VENOUS
BLOOD
PYRUVATE
MG./
100 MLS.
jects had extremely high fasting blood pyruvate levels and the mean level of group 3 (0.72 * 0.38 mg.1100 ml.) was significantly greater than those of group 1 (0.49 -t- 0.12 mg./lOO ml.) and group 2 (0.50 +- 0.10 mg./lOO ml., P < 0.001). Excluding the three markedly abnormal subjects from the obese group reduced the mean value of group 3 to 0.60 +- 0.16 mg./lOO ml., a value still significantly above those of groups 1 and 2 (P < 0.01, and P < 0.02, respectively). Of the three obese subjects with extremely high fasting blood pyruvate levels two had normal oral glucose tolerance curves with areas of 672 and 674 units and one just exceeded our upper limit of normal (800 units) with an area of 814 units. All three subjects, however, had a positive family history of diabetes mellitus. The mean fasting blood pyruvate level was slightly higher in those subjects with a family history of diabetes. This difference was most marked in the obese group, where the mean values were 0.82 mg./lOO ml. and 0.65 mg./ 100 ml. for subjects with and without a family history of diabetes. Oral Glucose Tolerance Tests were performed on 33 subjects in group 1, 21 subjects in group 2, and 24 subjects in group 3. Oral glucose tolerance, assessed as total area under the curve, was impaired in group 3 (736 -+ 134 units) as compared to group 1 (609 I+ 83 units, P < 0.001) and group 2 (630 f 51 units, P < 0.001). Seven obese subjects in group 3 and one subject in group 1 had OGTT glucose areas above 800 units, a value which we regard as indicative of chemical diabetes in women of this age group.10 The mean fasting blood pyruvate level of the 7 obese subjects with im-
BLOOD
693
FYRUVATE 1.4 _
levels
0.2 _
5 ci x
5 d lz
B d 2
6 0 :
5 d kY
6 d 2
0
I
I
I
I
I
I
30
60
90
\20
z I+
PRE I PRE 2
during
tween the mean values of groups 2 and 3. MINUTES
paired glucose tolerance (0.84 mg./lOO ml.) was greater than that of the 17 obese subjects with normal oral glucose tolerance (0.69 mg./ 100 ml.). The mean blood pyruvate levels during oral glucose tolerance tests are shown in Fig. 2. While the mean values for groups 1 and 2 did not differ significantly, those of group 3 were significantly elevated. The mean OGTT pyruvate areas and incremental areas are shown in Fig. 3. The mean pyruvate area of the obese group (4.27 2 1.25 units) was signikantly greater than that of group 1 (3.05 + 0.52 units) and group 2 (3.20 zk 0.76 units, P < 0.001). When 3 obese subjects with very high fasting pyruvate levels were excluded, the mean pyruvate area of group 3 was 4.00 units, a value significantly above those of groups 1 and 2 (P < 0.01). The mean OGTT pyruvate incremental area of the obese group (1.40 + 0.89 units) was not significantly greater than that of group 1 (1.07 i- 0.58 units) or group 2 (1.30 f 0.66 units). lntracenous
Glucose Tolerance
Tests were performed on 35 subjects in group 1, 22 subjects in group 2 and 24 subjects in group 3. The mean intravenous glucose ‘K’ value of the obese ;;
5.00
t
5
-
4.00
5 % 2
3.00
t y
2.00
Fig. 3.-Mean OGTT pyruvate levels during area and incremental area in groups 1,2 and 3. Bars represent 1 standard error of mean.
PYRUVATE
ii! ii \
too
GROUP
I AREA
2
3
2 INdREMENTAL
3 AREA
694
DOAR,
WYNN
AND
CRAMP
Fig. 4.-Mean blood py ruvate levels during IVGTT’s in group 1 (O), group 2 (A) and group 3 (0 ), Significance levels refer to the difference between mean values of groups 2 and 3.
PRE I
PRE 2
30
60
90
MINUTES
subjects (1.93 f 0.56) was not significantly different from that of group 1 (1.82 + 0.71) or group 2 (1.89 + 0.95). Two subjects in group 1 and one in group 2 had ‘K’ values of less than 0.95. The lowest ‘K’ value found in the obese group, however, was 1.17. The mean intravenous glucose area of the obese subjects (1613 -C 260) was greater than that of group 1 (1330 k 278) and group 2 ( 1498 t 269). The mean intravenous glucose incremental area of the obese subjects (872 I+ 234) was also greater than that of group 1 (624 + 236) and group 2 (753 + 255). Only the differences between groups 1 and 3 were significant (P < 0.001). The mean blood pyruvate levels following intravenous glucose administration to groups 1, 2 and 3 are shown in Fig. 4. The results are similar to those found during the oral glucose tolerance tests. The mean pyruvate levels of obese subjects were significantly greater than those of groups 1 and 2. The mean IVGTT pyruvate areas and incremental areas are shown in Fig. 5. The mean pyruvate area of the obese group (3.32 * 0.75 units) was significantly greater than that of group 1 (2.35 zk 0.55 units, P < 0.001) and group 2 (2.54 2 0.70 units, P < 0.001). When 2 obese subjects with very high fasting blood pyruvate levels were excluded, the mean pyruvate area of group 3 was 3.22 units, a value significantly above those of groups 1 and 2 (P < 0.001). The mean pyruvate incrementa area, however, of group 3
L
3 Q 4.00 Fig. 5.-MeanIVGTT pyruvate area and incremental area in groups 1, 2 and 3. Bars represent 1 standard error of mean.
IVGTT.
_
I
#
-I
3.00
_
2 $ 5 2.00 _ L o
PYRUVATE
-
I
f 1.00 _ a P a0 GROUP
I
2 AREA
3
I 2 INCREMENTAL
3 AREA
695
BLOOD PYRUVATJZ 2x, _ I.8 _
.
l .
I.6 _ 2 i
.
. .
l
l
.
1.4 _
2 a
I.2 _ I.0
_
a
08
_
t w I
0.6
_
;
04_
3
. .
.
l*
.
E
0.2
.
_
E ,’
.
. .
.
.
.
l
l
O_
z t
-a2
_
c’ t:
-0.4
_
.
+
> -:
-0.6
1 I1 0.2 0.4
I OS O.G.T.T.
0.6
I1 I.0
11 1.2
PYRUVATE
1.4
I1 I.6
I.6
1 2.0
INCREMENTAL
2.2
I1 2.4
2.6
I
I
2.6
31)
II 3.2
3.4
AREA (UNITS)
Fig. 6.-Correlation between OGTT pyrnvate incremental area and IVGTT pyruvate incremental area in 61 control women. Regresion line is given by IVGTT pyruvate incremental area = 0.37(OGTT pyruvate incremental area) + 0.53. r = 0.51(P
DOAII, WYNN
696
AND CRAMP
vate area (r = 0.44, P < 0.001) correlated significantly with degree of obesity. The correlation between oral glucose and pyruvate area (r = 0.27, P < 0.05) became non-significant when the effect of degree of obesity was removed by partial correlation (r = 0.13, P > 0.10). In 81 IVGTT’s intravenous pyruvate area correlated with degree of obesity (r = 0.54, P < 0.001). There was no correlation between the intravenous “K” value and any parameter of intravenous blood pyruvate levels. Surprisingly, significant negative correlations were observed between the fasting pyruvate levels and oral and intravenous pyruvate areas (r = -0.29, P < 0.01 and r = -0.37, P < 0.001, respectively). Inspection of the scattergrams, however, showed that these two correlations were largely due to the three obese subjects with extremely high fasting pyruvate levels who had very small or even negative pyruvate incremental areas. When these three subjects were excluded, there was no significant correlation between the fasting pyruvate level and oral/intravenous pyruvate incremental areas in the remaining subjects. DISCUSSION
Although oral glucose tolerance was markedly impaired in the obese group as a whole, the intravenous “K” values were not significantly different in the three groups studied. This suggests a dissociation between the results of oral and intravenous glucose tolerance tests in obesity as has been previously noted.2 The explanation for this dissociation is not clear. The contribution of tissues such as liver, fat and muscle, to the removal of oral or intravenous glucose loads is not known. It is probable that hepatic uptake is relatively more important in the former situation, Conn and Newburgh18 suggested that impaired oral glucose tolerance in obesity might be due to decreased hepatic glycogen formation. Butterfield et al. la~zohave postulated that glucose uptake by muscle is decreased in this condition. They found the rate of uptake of glucose by forearm tissues to be reduced in obese subjects during oral and intravenous glucose tolerance tests. It is difficult, however, to assess the relative contributions of muscle and adipose tissue to glucose removal in this type of study. The lower overall forearm glucose uptake rates observed in their obese subjects may have been due to the large mass of relatively inert adipose tissue removing comparatively little glucose. Since intravenous glucose tolerance does not appear to be impaired in obesity, further work is required to evaluate the uptake of glucose by various tissues in obese and non-obese subjects. Loading doses of glucose based on body weight are not ideal for the study of obesity because the volume of distribution of glucose forms a lesser proresulting in greater extracellular portion of body weight in obese subjects,‘lJ* fluid concentrations of glucose during glucose tolerance tests. These differences, however, are likely to exert only a small effect on conventional parameters of glucose tolerance since West et a1.23 found that in duplicate OGTTs doubling the glucose load from 0.8 g/Kg. to 1.6 g/Kg. resulted in the mean
BLOOD
PYRUVATE
697
60 minute and 120 minute blood sugar levels being about 20 mg./lOO ml. higher. Assuming similar increases for the other points, this would increase the OGTT area by 110 units. Wahlberg4 observed that doubling the glucose load from 25 Gm. to 50 Gm. in 52 subjects caused the mean intravenous glucose ‘K’ value to increase by 0.24. Since these extreme changes caused only small changes in oral and intravenous glucose tolerance, bias resulting from a glucose load based on body weight in the present series would be even smaller. Impaired oral glucose tolerance in obese subjects has been found by manyQ but not all investigators. 24.25 Wahlberg4 using a 25-Cm. glucose load found no significant difference between the mean intravenous “K” value of 56 obese subjects compared with that of 144 control subjects. Vajda et al.,26 however, using a glucose load of 0.5 Gm./Kg. body weight found impaired intravenous glucose tolerance in a group of 25 obese adolescents. They used the time required for the blood sugar to return to the fasting level as a criterion of glucose tolerance. For reasons outlined above, when glucose is administered in amounts proportional to body weight, an obese subject with a “K” value identical to that of a non-obese subject will show higher plasma glucose levels and in consequence the return of the plasma glucose level to normal will be delayed. This was confirmed by the obese subjects in the present study who, despite normal intravenous glucose tolerance as judged by ‘K” values, had a mean total glucose area and incremental area significantly above those of the controls. Morse et al.2 using an intravenous glucose dose proportional to estimated body surface area (a parameter more closely related to the apparent volume of distribution of an intravenous glucose load) found no significant difference between “K” values of obese and non-obese subjects. Significantly elevated levels of blood pyruvate were found both before and during oral and intravenous glucose tolerance tests in the group of 27 obese subjects. The total area under the pyruvate curve was significantly greater during both tests in the obese group. The incremental areas above the base line, however, were not signihcantly greater than those of the non-obese subjects. These results agree with those of Haagenson’” who noted elevated fasting blood pyruvate levels but a normal oral pyruvate incremental area in a group of obese subjects. There were several significant correlations between blood pyruvate and plasma glucose levels. The relation between fasting plasma glucose and blood pyruvate levels (r = 0.21, P < 0.05) was shown to be due to both variables correlating with the degree of obesity and contrasts with that found by Fry and RutterfieldY in a series of 28 diabetics (r = 0.69, P < 0.01). The two studies, however, are not comparable and the range of fasting blood sugar was much greater in the diabetic series. In 61 paired tests there were strong correlations between the fasting blood pyruvate levels in the two tests (r = 0.74, P < 0.001) and oral and intravenous pyruvate incremental areas (r = 0.51, P < 0.001). The latter finding suggests that the ratio of the proportion of the glucose load passing down the glycolytic pathway to pyruvate in the OGTT to that following a similar
698
DOAR, WYNN AND CRAMP
route in the IVGTT was similar for all subjects and, further, that the rate of pyruvate catabolism in a given subject was fairly constant over the period of time between the two tests. When the degree of obesity was taken into account, the correlation between OGTT glucose area and pyruvate area became non-significant (r = 0.13, P > 0.10). Elevated fasting blood pyruvate levels have been found by some workers in mild cases of diabetes meWus though the pyruvate rise following glucose has usually been normal .?,*? It is possible that abnormal pyruvate levels in mild diabetes may be more closely related to the degree of obesity than is the abnormality of glucose tolerance. In the present series, seven obese women had fasting blood pyruvate levels above 0.80 mg./lOO ml. and of these four had oral glucose tolerance areas within our normal range. Raised fasting plasma insulin levels have been found in obese nondiabetic subjects6 and the present findings suggest that an elevated fasting venous blood pyruvate level should be added to the list of metabolic abnormalities found in this condition. In 81 IVGTT’s there was no relation between the “K” value and any parameter of pyruvate metabolism. Although .theWmean.-pyruvate incremental area was greater in obese subjects during both oral and intravenous glucose tolerance tests, the values were not significantly above those of the nonobese groups. Both the oral pyruvate area (r = 0.37, P < 0.001) and intravenous pyruvate area (r = 0.44, P < 0.001) correlated better with the degree of obesity than with any parameter of glucose tolerance. Elevated fasting blood pyruvate levels in obese subjects may be due to increased pyruvate formation and/or decreased pyruvate removal. If pyruvate removal were impaired a positive correlation might have been expected between the fasting level and the intravenous pyruvate incremental area. This, however, was not the case. The three obese subjects with extremely high fasting pyruvate levels had small or negative intravenous pyruvate incremental areas. Their intravenous glucose ‘K’ values were 2.13, 2.39 and 1.76, respectively, indicating that glucose was removed at a normal rate and that impaired pyruvate removal in these subjects is therefore unlikely. Alternatively, increased rates of pyruvate formation may be responsible for elevated fasting blood pyruvate levels in obesity. In this situation, assuming the rate of pyruvate removal to be proportional to the blood pyruvate concentration, the presentation of a pyruvate load, derived from oral/intravenous glucose, should result in a relatively unchanged pyruvate incremental area. In the present series with a glucose load based on body weight, the increment of glucose concentration in the ECF would have been greater in the obese subjects,2* resulting in the formation of more pyruvate. This could account for the slightly greater mean intravenous pyruvate incremental area found in the obese group. Venous blood pyruvate levels to some extent reflect changes in metabolism of local tissues and are not necessarily indicative of events in the body as a whole. Relative anoxia of forearm tissues in obesity could impair pyruvate
699
BLOOD PYRUVATJI
removal but should also lead to increased lactatelpyruvate ratios. No difference was found, however, between venous lactatelpyruvate ratios in obese and non-obese subjects (unpublished observations). Further, similar arterio-venous lactate differences, oxygen uptakes and lactate production rates have been observed in forearm tissues of obese and non-obese subjects.2s The possibility remains that other tissues, such as liver, may be producing greater than normal amounts of pyruvate in obese subjects. The fraction of an oral/intravenous glucose load that passes down the glycolytic pathway to pyruvate is unknown. This proportion may vary with degree of obesity, and the above observations could equally be explained in terms of impaired pyruvate degradation in obese subjects together with a greater proportion of the glucose load being utilised along other routes such as conversion to glycogen, oxidation via the hexose monophosphate shunt, and conversion to cu-glyceromonophosphate in adipose tissue for re-esteriflcation of fatty acids. The hexose monophosphate shunt is more active in adipose tissue than musclez9 and it has been suggested that increased glycogen formation may occur in forearm tissues in obese subjects in the fasting state.2* Clearly, direct studies of pyruvate degradation rates in vivo will be necessary to determine whether pyruvate catabolism is impaired in obese subjects. Lower rates of C1402 excretion were found following carbonyl Cl4 pyruvate administration to a group of obese hyperglycaemic mice compared with a group of controls, suggesting that pyruvate oxidation was impaired in the former.30 The effects of different precursor pool sixes in this type of study, however, are diflicult to evaluate. Addition of fatty acids has been found to impair pyruvate oxidation by heart muscle and diaphragm in vitro31 and it is possible that the elevated plasma NEFA levels found in some cases of obesity5 may have a similar effect. Administration of certain hormones, including thyroxine,32 growth hormone,33 cortisoP and oestrogen-progestagen combinationslO has been found to cause elevated blood pyruvate levels. In general, however, blood concentrations of these hormones are not increased in obesity.34-36 ACKNOWLEDGMENT The authors would like to thank Mr. M. J. R. Healy for statistical advice. REFERENCES 1. John, H. J.: A summary of the findings in 1100 glucose tolerance estimations. Endocrinology 13:388, 1929. 2. Morse, W. I., Sidorov, J. J., Soeldner, J. S., and Dickson, Il. C.: Observations on carbohydrate metabolism in obesity, Metabolism 9:666, 1966. 3. Smith, M., and Levine, It.: Obesity and Diabetes. Med. Clin. N. Amer. 48: 1387,1964. 4. Wahlberg, F.: Intravenous glucose tolerance in myocardial infarction, angina,
pectoris and intermittent claudication. Acta Med. &and. Supp. 453, 1966. 5. Beck, P., Koumans, J. H., Winterling, C. A., Stein, M. F., Daughaday, W. H., and Kipnis, D. M.: Studies of insulin and growth hormone secretion in human obesity. J. Lab. Clin. Med. 64:654, 1964. 6. Kreisberg, Pi. A., Boshell, B. R., DiPlacido, J., and Roddam, R. F.: Insulin secretion in obesity. New Eng. J. Med. 276: 314,1967. 7. Smith, M. J. H., and Taylor, K. W.:
700 Blood pyruvate and a-ketoglutarate in norma1 and diabetic subjects. Brit. Med. J. 2: 1035, 1956 8. Fry, J. K., and Butterfield, W. J. H.: Carbohydrate metabolism in diabetes-a possible intracellular block. Lancet 2:66, 1962. 9. Hennes, A. R., Wajchenberg, B. L., Fajans, S. S., and Conn, J. W.: The effect of adrenal steroids on blood levels of pyruvic and alpha-ketoglutaric acids in normal subjects. Metabolism 6:339, 1957. 10. Wynn, V. W., and Doar, J. W. H.: Some effects of oral contraceptives on carbohydrate metabolism. Lancet 2:715, 1966. 11. Arendt, E. C., and Pattee, C. J.: Studies on obesity II. Blood pyruvate and lactate curves in obese and normal subjects after ingestion of glucose. J. Clin. Endocr. 16:375, 1956. 12. Miller, A. T., and Thomas, B. M.: Pyruvate metabolism in obesity. Amer. J. Clin. Nutr. 4:619, 1956. 13. Stucklikova, E., HruSkova, J., Tenorovi, M., Novotna, B., Komirkova, A., and Riedl, 0.: Some changes in intermediary metabolism of obese patients. Clin. Chim. Acta 6:571, 1961. 14. Nyfos, L., and Skouby, A. P.: Blood pyruvic acid in obesity. Acta Med. Stand. 161:403, 1957. 15. Haagenson, N. R.: Inorganic phosphate and pyruvate in obese patients during glucose assimilation. Acta Med. Stand. 167: 177, 1960. 16. Cramp, D. G.: New automated method for measuring glucose by glucose oxidase. J. Clin. Path. 20:910, 1967. 17. -: Automated enzymatic fluorimetric method for the determination of pyruvic and lactic acids in blood. J. Clin. Path. 21:171, 1968. 18. Conn, J. W., and Newburgh, L. H.: Hyperglycaemia due to impaired hepatic glycogenesis. Proc. Sot. Exp. Biol. Med. 36: 236, 1937. 19. Butterfield, W. J. H., Hanley, T., and Whichelow, M. J.: Peripheral metabolism of glucose and free fatty acids during oral glucose tolerance tests. Metabolism 14:851, 1965. 20. -, Abrams, M. E., St. John, D. J. B., and Whichelow, M. J.: The intravenous glucose tolerance test: Peripheral disposal of the glucose load in controls and dia-
DOAR,
WYNN
AND
CRAMP
betics. Metabolism 16:19, 1967. 21. Fran&son, J. R. M., Conard, V., and Bastenie, P. A.: Measurement of the free glucose diffusion space in man by the rapid intravenous glucose tolerance test. Acta Endoer. (Kobenhavn) 32:463, 1959. 22. Doar, J. W. H.: A comparison of oral and intravenous glucose tolerance tests in 50 control women (to be published). 23. West, K. M., Wulff, J. A., Reigel, D. G., and Fitzgerald, D. T.: Oral carbohydrate tolerance tests. Arch. Intern. Med. 113:641, 1964. 24. Unger, R. H.: The standard twohour oral glucose tolerance test in the diagnosis of diabetes mellitus in subjects without fasting hyperglycaemia. Ann. Intern. Med. 47:1138, 1957. 25. Tyner, J. D.: The prediabetic state: Its relation to obesity and to diabetic heredity. Amer. J. Med. Sci. 185:704, 1933. 26. Vajda, B., Heald, F. P., and Mayer, J.: Intravenous glucose tolerance in obese adolescents. Lancet 1:902, 1964. 27. Anderson, J.. and Marks, V.: Pyruvate in diabetes mellitus-concentrations in urine and blood. Lancet 1:1159, 1962. 28. Rabinowitz, D., and Zierler, K. L.: Forearm metabolism in obesity and its response to intra-arterial insulin, Characterisation of insulin-resistance and evidence for adaptive hyperinsulism. J. Clin. Invest. 41: 2173,1962. 29. Randle, P. J.: Endocrine control of metabolism. Ann. Rev. Fhysiol. 25:291, 1963. 39. Guggenheim, K., and Mayer, J.: Studies of pyruvate and acetate metabolism in the hereditary obesity-diabetes syndrome of mice. J. Biol. Chem. 198:259, 1952. 31. Randle, P. J., Garland, P. B, Hales, C. N., and Newsholme, E. A.: The glucose fatty-acid cycle. Its role in insulin sensivity and the metabolic disturbances of diabetes mellitus. Lancet 1:785, 1963. 32. Macho, L.: The influence of endocrine glands on carbohydrate metabolism. IV. The effect of thyroid hormones on pyruvic acid metabolism. Arch. Internat. Physiol. 70:507, 1962. 33. Henneman, D. H., and Henneman, P. H.: Effects of human growth hormone on levels of blood and urinary carbohydrate and fat metabolites in man. J. Clin. Invest. 39:1239, 1960.
BLOOD
701
I’YHU\‘ATE
34. Lessof.
hl.
H.,
and Greenwood, F. secretion in obese Rep. 115:65, 1966. 35. Schteingart, D. Characteristics of the
McHardy
Young,
S.,
C.: Growth hormone subjects. Guy Hosp. E., and Conn, J. W.: increased adrenocorti-
cal function observed in many obese patients. Ann. N. Y. Acad. Sci. 131:388, 1965.
36. Yalow, and Berson,
R. S., Glick. S. M.. Roth, J., S. A.: Plasma insulin and
growth hormone levels in obesity and diabetes. Ann. N. Y. Acad. Sci. 131:375, 1965. 37. Snedecor, G. W.: Statistical Methods, (ed. 5). Ames, Iowa, Iowa State University Press, 1956.