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Effects of oral chromium supplementation on the glucose tolerance of elderly human subjects
Effects
of Oral Chromium
Glucose Tolerance By ROBERT A.
Supplementation
of Elderly Human
on the
Subjects
LEVINE, DAVID H. P. STREETEN
AIYD RICHARD J, DOISY The possible role of Cr deficiency in the impairment of glucose tolerance in elderly subjects has been studied. Chromium supplements, 150 pg. daily, were given to 10 apparently healthy inmates of an old-age home, whose oral glucose tolerance tests (G.T.T,‘s) were abnormal. Mean GTT became normal in 4 subjects (“Cr responders”) but remained abnormal in the other 6 subjects (“Cr non-responders”) during Cr administration. The beneficial effect of Cr in the “Cr responders” did not result from an increased release of insulin, since
plasma ILA and immune-reactive insulin levels were normal and rose normally after the glucose load both before and during Cr supplementation. Serum Cr concentration was found to rise significantly after the glucose load in old and young subjects but to a lower level in the elderly “Cr non-responders,” suggesting that an adequate rise in serum Cr might be necessary for normality of the G.T.T. The data support the possibility that Cr deficiency may lead to the impairment of the GTT in some elderly subjects. (Metabolism 17: No. 2, February, 114-125, 1968)
N AN EARLIER COMMUNICATION’ it was shown that the abnormal glucose tolerance of elderly human subjects was due neither to retarded glucose absorption nor to a reduced rise in plasma insulin-like activity after the glucose load, but to some other (unknown) cause of delayed cellular uptake of glucose from the plasma. It is known that the chromium content of human tissues declines with advancing age.” Furthermore, rats ingesting chromium-deficient diets develop abnormal glucose disposal rates”.4 and diminished fat-pad sensitivity to exogenous insulin.” On normal diets, rat fat-pads show decreasing sensitivity to insulin with increasing ageSss7 In view of these facts, the present study was instituted to ascertain whether elderly subjects with abnormal glucose tolerance tests would show an improved glucose tolerance during oral chromium supplementation, as had been observed in laborator\, animals and in some patients with diabetes mellitus.x
I
From the Departments of Medicine and Biochemistry, State University of Nerc York, Upstate Medical Center, S:yracuse, New York. Supported bt~ Grants AM -5252 und AM 03025 from the National Institute of Arthritiq and Metabolic Diseases. Received for pz~blicution April 28, 1967. ROBERT A4. LEVINE, M.D.: Intern in Medicine, Mount Sinai Hospital, New York, N. f. This paper wus written during tenure of a medical student summer traineeship of the NIH. DAVID H. P. STREETEN, M.B.. D. PHIL, M.R.C.P.: Professor of Medicine, State Uniuersifq of New York, Upstate Medical Center, Sgraeuse, N. I’. RICHARD J. DOISY, PH.D.: Assoc~te Professor of Biochemistry, State Unioersity of Netc l‘ork. C7psfatc AlcrlicuI Ccntc~, Syracuse, N. Y.
ORAL CHROMIUM
115
SUPPLEMENTATION METHODS
Clinical Material Fifteen elderly subjects (9 male, 6 female) in the Onondaga County Home were selected from volunteers for the study. The subjects were at least 74 years of age, had been in the Home for six months or longer, and had no family history of diabetes. They were alert, had no known disease (except for inactive rheumatoid arthritis in one subject and well treated hypothyroidism in another) and were taking no medications that might have interfered with glucose tolerance tests (GTT). All had fasting plasma glucose concentrations below 120 mg./ 100 ml. Sixteen healthy young adults (12 male and 4 female physicians, medical students and laboratory technicians, aged 19-32 years) served as a control group. One of these young subjects was found to have one abnormal and two normal GTT’s during the study. All subjects were prepared for 3 days prior to testing, with a diet containing at least 300 Gm. of carbohydrate daily. With the subjects in a seated or reclining position after fasting overnight, venous blood was drawn through a siliconized needle into 3 evacuated tubes for determinations of plasma glucose con~ntration, serum insulin-like activity @LA), and serum Cr concentration. The tubes for glucose determination contained potassium oxalate (20 mg.) and sodium fluoride (25 mg.). The tubes for Cr determination were special Pb-free Vacutainer tubes which had been shown to contain no measurable traces of water-soluble Cr compounds. Sera for Cr analysis were transferred to tubes thoroughly rinsed with distilled water and previously found to be free of detectable amounts of Cr. The subjects then ingested 100 Gm. of glucose in aqueous solution and venipuncture was repeated at 30, 60, 90 and 120 minutes. Glucose concentrations were measured in all plasma samples, and serum ILA and chromium determinations were carried out on the samples obtained at 0 and 60 minutes. The specimens were centrifuged within 2 hours after completion of the glucose tolerance tests and stored at -40” C. Plasma glucose concentrations were measured by a standard ferrocyanide method using the AutoAnalyzer (Technicon). Serum ILA levels were determined by a rat epididymal fat pad assay. r In the latter part of the study, simultaneous assays were performed with and without the addition of guinea pig anti-insulin serum (AIS). Serum chromium determinations were kindly performed by Fredric J. Feldman at the Walter Reed Army Medical Center using atomic absorption spectrophotometry.sJ’)
The premise upon which this pilot study was based was that impaired glucose tolerance might be due to Cr deficiency in some or all elderly subjects, which might have resulted from inadequate intake or inadequate body stores of Cr. A given individual might be normal with respect to serum Cr concentration and yet have moderate Cr deficiency. The time taken to replete an individual with orally administered Cr supplements would depend on the degree of deficiency and on the dosage of the administered Cr. If Cr deficiency was responsible for the observed abnormality in GTT, it was anticipated that Cr administration would restore the GTT to normal. Glucose tolerance tests were performed at approximately monthly intervals, the first L? tests in each individual being done before treatment. Plasma glucose concentrations were considered to be “diabetic” in at least one of the “baseline” tests in 9 of the elderly subjects and to be abnormal but without all of the diabetic features in one subject. by the criteria of Fajans and Corm11 modified because plasma and not whole-blood concentrations were measured in the present study. Plasma and whole-blood glucose concentrations differ somcwhat inconsistently from one another because of the variably lowered concentrations of glucose in the formed elements of the blood and because of the normal range of fluct~lations in the hematocrit. Since there are no published Iimits of normality of the plasma glucose concentrations after a glucose load, new criteria of abnormality were used. These were based on the evidence of Dillon12 that, assuming an hematocrit of 45 per cent, the criteria of Fajarrs and Connil for diabetes, using whole-blood glucose concentrations would correspond with a peak plasma glucose concentration exceeding 185 mg./lOO ml., and with 90 and 120minute concentrations above 160 and 140 mg./lOO ml.. respectively. Further studies on the
116
LWIXE,
Fig.
l.-Plasma
concentrations performed subjects stration. glucose served 100 248 at
in
mg./lOO
10
elderlv
Limits
of plasma
concentrations
122-278
90
min.
DOISY
&mose
Cr adminiob-
116-200
ml. at 30 mg./lOO
min.,
AND
in 20 GTT’s
before
were
STHEETEN
mg./
min., 146ml. at 60 mg./lOO
and
ml.
131-280
ml. at 120
min.
,
0.5 Hours
1.0 After
Glucose
1 5
,
2.0
Lood
elderly subjects were restricted to the 10 individuals in whom the baseline GTT was “diabetic” or “abnormal” on at least one occasion. The objective of the study was to administer Cr tablets to each individual for 3 or 4 months or until the mean results of at least 2 GTT’s were within normal limits. This treatment period was to be followed or preceded by the administration of placebo tablets. In fact, because some of these elderly volunteers were not prepared to continue in the study for the length of time planned, only 4 of the 10 elderly subjects received both placebo and Cr tablets, and the remaining 6 received only the Cr tablets for approximately 4 months (5 subjects) or 1 month (1 subject). Ten of the young individuals were given Cr tablets, each for 14 months. The Cr tablets contained 256 ug. CrCb.GHaO (each with 50 ug. Cr) in an inert base and were given 3 times daily with meals, since the solubility of CrCL is known to be reduced at alkaline pHs, and an acid pH might be expected to enhance absorption. RESULTS
Plasma Glucose Concentration-Baseline
Tests
Among the 16 young adult subjects, baseline glucose tolerance tests were all normal except for one abnormal test in one subject. Nine of the 15 elderly subjects (60 per cent) had at least one “diabetic” baseline test and at least one of the listed criteria of abnormality was present in one or more GTT in 13 of the 15 elderly subjects. The 20 “baseline” tests in the 10 elderly subjects who received Cr supplements are depicted in Fig. 1. It is evident that the plasma glucose concentration at 120 minutes exceeded 140 mg./lOO ml. in 18 of the 20 GTT’s the two exceptions being in the same individual (F. W. ).
ORAL
Table
CHROMIUM
l.-Mean
117
SUPPLEMENTATION
2-hour Plasma Glucose Levels During in 10 EMerly Subjects Group
Subiect
1 2 3 4
cv AN CZ AM
Age
Wt. in Kg.
%sf
F
81
57.5
2
M M
83 76 74
Tolerance
Tests
a
Sex
M
Glucose
68.2 69.1 77.7
Cr
Glucose Level, mg./lOO ml.
244
3
-t
2 3
+
2 5
+
2 2
+
212 209 237 192 206 191
5
hIH
F
81
35.9
2 1
+
6
MC
M
84
64.3
2 1
-t
196 166 168 162 234 Avg.
194% 12
209*
10
Group b GF
M
79
53.2
2 2
+
170 +
II
M
78
53.0
2 3
Ech
F
88
46.8
2 3
+
FW
F
96
58.0
2 3
+
106 167 140 153 131 135 100 Avg.
156*8 P <
119* 10 .05
Tests During Cr and Placebo Administration During Cr administration the mean plasma glucose concentrations in the GTT’s remained “diabetic” in 6 elderly subjects (group a) but were “normal”, in all respects, in the remaining 4 elderly subjects (group b, Fig. 2 and Table 1). In contrast with the considerable overlap between the mean plasma glucose concentrations on and off Cr supplements in the subjects of group a, there was no such overlap, at 90 and 120 minutes, in group b. This difference between the mean GTT results during Cr supplementation in the subjects of groups (a) and (b) was also a consistent finding in each of the individual tests on the subjects of the 2 groups. Thus, the subjects of these groups appeared to have responded to Cr administration like two separate populations, without overlap of any plasma glucose concentrations at 90 and 120 minutes. Morever, closer inspection of Fig. 2 suggests that these 2 populations were recognizably distinct even in the tests before Cr supplementation. Thus, the mean plasma glucose concentrations before Cr treatment were almost invariably lower at 90 and 120 minutes in the subjects who subsequently became normal on Cr (group b) than in the subjects who remained “diabetic” on Cr (group a).
118
LEVINE,
1
300
STREETEN
AND
DOISY
(b)
270
240
J
, 0
0.5
y-control
/ -0
Limits
/ 5
Hours
2,O
After
,j
-Control 0
Glucose
0.5
b0
Llmlts
3.5
2.0
Load
Fig. S.-Mean plasma glucose concentrations in GTT’s performed before and during Cr supplementation in 6 elderly subjects whose GTT s were abnormal both
on and off Cr (group a) and 4 elderly subjects whose mean GTT’s became normal on Cr (group b), Limits of baseline tests in all 10 elderly subjects are enclosed within heavy lines. The results of the GTT’s in 2 elderly subjects of group (b) are shown in Fig. 3. It is evident that the GTT was improved within 25 days of starting the Cr supplements in one subject (E.C.), while the other (H.H.) showed a more gradual improvement. The placebo tablets had no evident effect on the mean GTT’s of the 2 elderly subjects whose abnormal GTT’s showed no response to Cr (group a). The GTT results in the 2 elderly subjects of group (b) who were treated with the placebo, and both of whom happened to receive placebo tablets after treatment with Cr, are shown in Fig. 4. The prompt and consistent improvement in the results during Cr administration is evident, especially in the 90- and lZO-minute plasma glucose concentrations. All or most of the improvement in the results was still evident in the tests done 27 and 25 days after the substitution of placebo for Cr tablets. However, during continued administration of the placebo, the 60-, 90- and 120-minute plasma glucose concentrations were always above all of the corresponding values during Cr administration, except in the test done at 248 days in subject G.F., which was very similar to those performed during Cr supplementation. Among the 10 young subjects given Cr supplements, this treatment did not affect the consistently normal baseline GTT’s observed (Table 2). The one individual who had a single diabetic baseline GTT was consistently normal while receiving Cr.
ORAL
CHROMIUM
On Chromium (days 12-129)
Baseline (days I-1 2) Ooy
of Study_,_ 210
119
SUPPLEMENTATION
45
r
&29_
,73,
H. H
1 E.O.
T
Fig. 3.-Plasma glucose concentrations in all GTTs performed on 2 elderlv subjects of group (b) (H.H and E.C.) before and during Cr administration. Days of each GTT are numbered from the dav of the first baseline test. Plasma glucose concentrations are shown before and at 30, 60, 90, and 120 min. after each glucose load.
P
60
1
OnChrormum (days
Serum CT Concentrations
6-l 16)
(Table 3)
no significant differences between the young and elderly groups of subjects. There was a rise in serum Cr concentration from 0 to 60 minutes after the glucose (Table 3) which was highly significant (P < .OOl) in both the young and the elderly groups of subjects when all of the values were analyzed by the “t” test for non-independent variables (n = 55 and 60, respectively. At 60 minutes after the glucose load, the serum Cr level was significantly lower in the elderly subjects of group (a) than in the young subjects (P < .05) during baseline tests. Mean
fasting
serum Cr levels showed
Serum ILA Concentrations In the fasting state, mean serum ILA showed no significant differences between young and elderly groups of subjects (Table 3). There was an increase in ILA 60 minutes after glucose administration, which was significant in the young and the elderly groups both before and during Cr supplementation (P < .05 to < .005). Serum ILA levels were determined both in the presence and in the absence of guinea pig anti-insulin serum, in samples of human serum obtained during 57 of the glucose tolerance tests performed in the present study. The mean results of these determinations are shown in Table 4 in which the corresponding results obtained by Samaan and Fraserls are shown for comparison. It is
120
LEVINE,
Baseline (day. t-26) Day ol Study
z z
u
I80
~ 1
On Chromium (days 27-1201
AND
DOISY
On Placebo ldoys 121-231)
~
:
&;
(
STREETEN
_ 231
z
-
z
z .?
150 -
z
120.
(3
h a SOi/
Day of Study
:;:’
-TV Baseline ldoyr l-14)
34’ On Chromium (days l5-S2)
77
T”5-s On Placebo (days 53-2461
Fig. 4.-Plasma glucose concentrations in all GTT’s performed on 2 elderly subjects of group (b) (F.W. and G.F.) before Cr, during Cr and during placebo administration. Days of each GTT are numbered from the day of the first baseline test. Plasma glucose concentrations are shown before and ;lt 30, 60. 90 kind 120 min. after each glucose load.
evident both from the present data and from the findings of Samaan and Fraser that the immunologically reactive ILA is the only component of the serum insulin-like activity which increased after a glucose load, from mean values of 26 and 43 pU./ml. to mean values of 183 and 188 @J./ml. respectively in the 2 studies. When the tests on subjects with known abnormalities of carbohydrate tolerance were eliminated from the data, the mean rise in ILA after glucose was 116 pU./ml. (39 tests), in good agreement with the report of Yalow and Berson”* that serum insulin concentration, measured by immunoassay, rose by 118 @J/ml. after the same load of glucose. Thus, glucose-induced changes in ILA levels measured by the fat-pad assay closeIy reflect changes in immunoreactive insulin concentrations in the serum. Dietary
Cr Irltakc
Table 5 shows the Cr content of the total diet offered to each of the elderly institutionalized subjects over a T-day period and of the diet consumed bv one Table 2.-PEQsnza Glucose Conc~ntru~jons ~~g./~OO ml.) During In Young Subjects On and OfF Chromium (Mean -e SEMI
GTT’s
121
SUPPLEMENTATION
of Oral Chromium Administration on Serum ILA and Cr Concentrations
&3.-E5eet
est.7
ILA, ,-~O;ts/ml. No. of Subiects Tests
( Meaxr& SEM )
0’
Cr. ng./ml.#ean
-(_ “EM:
seline nCr
16 10
31 25
197f12 199t30
319’29 292246
23.2 L+I 24.312
27.912 32.6f2
4.5*2 7.6*2
Jaseline On Cr
10 10
20 26
1792 I9 174+-14
312+61 379 f 68
21.7*3 24.6& 1
27.624 29.6*3
5.8-t3 4.5*2
Baseline
6
12
197k-29
343*96
19.0*3
21.8*2
2.1*1
On Cr
6
15
188*22
427+
28.253.k
2.853.k
Group (b) Abnormal Old
Baseline
4
8
152klS
265+51
26.0+6
36.2+10b
Abnormal Old
On Cr
4
I1
155k4
308+53
25.0”33
31.526
-___
..lormal Old
108
24.3i:
1
11.5~6* 6.723
A Values are not determined by difference of (66-O’) mean values but are mean values for these differences in each subject. * Included in this mean value is one 60’ determination which was unusually high and may partially account for the higher value in this gronp. t Serum Cr determinations 60 min. after glucose were obtained in only S of the 6 elderly subjects of group (a).
young subject for a week. The daily intake of Cr was quite variable in both instances. Analyses of the drinking water indicated that Cr levels were below the limits of detection ( 10 ng./ ml. ) . Grudges in Body Weight There was no consistent change in body weight during these studies, mean baseline weight being 61.7 and 53.0 Kg., mean weight during Cr administration being 62.0 and 52.8 Kg. in groups (a) and (b), respectively. Table
&-Serum
KLA in the Presence and Absence of Guinea Pig
Anti-in~lin Sera (AIS) During Oral G,T.T.‘s MEAN SERUM ILA, ,-units/ml. FASTING Authors
%Y
ILA + AIS
Present
190
149*
Samaan ancl Fraser’::
-
164
Immunoreactive Insulin
22’
60 MINUTES Immunoreactive Insulin
ILA + AIS
26
309
147
183 (57)-/-
43
-
165
188 (10)
Values of Immuno-reactive Insulin are mean differences between total ILA and ILA measured in presence of AIS. ‘The non-immunoreactive component of the ILA was essentially the same at 60 minutes as in the fasting state, as shown above. In 12 GTT’s ILA levels were also determined in the presence and absence of AIS at 0, 30, 60, 90, and 120 minutes. The respective mean values of non-~munoreactive ILA obtained were 157, 149, 149, 139, 146 iu-units/ml., further suggesting that this component of ILA is unchanged in normal subjects during the G.T.T.‘s. TNumber of tests indicated in parentheses.
122
LEVIA-Fa0 STl{
Table S.-Chromium
Analysis of Total Daily Food
Elderly Subjects
Young
ClgO,U&ll
_____
l&y
1”
Day
2”
1.15 47
3 4 5 6 7”
9 91 35 63 5
Mean
= 52 pg./dav ---z
b.1
.\‘111111,11 Silh (%I ..i:
1 ” :3 -1 3 6
4 li<1 4.;
I I .5 SJr!
7-7 -_-_--__
Ilean
Com~os~tiotz of ~1~erI~ S~~j~c~s’ Diet “Days 1, 3, and ‘7 were selected as representing high, medium Day 1. 115 pg. Criday
z
65 ,iLg./tlat
and Low intakes of (:r.
Breakfast: Orange juice, farina, toast, milk, coffee. Lunch: Cream of tomato soup, egg souffle, tossed salad, applesauce, tea. Dinner: Meat loaf with gravy, mashed potato, beets, cottage cheese, cookies, milk,
tea.
Day 2. 47 pg. Cr/day Breakfast: Prune juice, oatmeal, one egg, 2 slices toast, 1 cup milk, coffee. Lunch: Chicken noodle soup, roast beef hash, crushed pineapple, 1 cup tea.
I)inner: Baked chicken, dressing, gravy, mashed potato, 1 slice bEad, peas, fruit jelto, milk, tea. Day 7. 5pg. Criday Breakfast: Prune juice, farina, egg, toast, milk, coffee. Lunch: Clam chowder, tuna fish sandwich, white cake, tea. Dinner: Creamed cod fish, mashed potato, peas, apricots, 1 slice bread, milk. Tap water in both groups was found to contain less than 10 ng. per ml.
The present studies have confirmed previous evidence that ~~abnori~lal~ies~ of the oral glucose tolerance test are common in elderly subjects. Thus, 60 per cent of 15 individuals had at least one baseline curve which was “diabetic” by criteria modified for plasma glucose concentrations from the whole-blood glucose values proposed by Fajans and Corm. I1 The elderly subjects were considerably lighter (mean weight 58.5 Kg.) th an the group of 16 young adults (mean weight 77.8 Kg.) of whom 15 were found to have GWs consistently within the stated limits of normality. However, the weight discrepancy could not have accounted for the impaired tolerance in the elderly subjects since the in these elderly individuals closely glucose load (100 Gm. or 1.71 Gm./Kg.) approximated the load of 1.75 Gm./Kg. recommended by Fajans and Conn. Moreover, there was no difference between the mean change in weight during chromium administration in the elderly individuals of groups (a) and (b) (mean weight change of 0.3 and 0.2 Kg., respectively), all of whom continued to receive the same diet. Chromium administration was associated with disappearance of all abi~o~al features of the mean GTT in 4 of the 10 elderly individuals so treated, and with no change in the GTT of the remaining 6 subjects. When the plasma glucose concentrations of these 2 groups of elderly subjects were compared, it was found that there was complete separation, without any overlap what-
ORAL
CHROMIUM
SUPPLEMENTATION
123
ever, between the 21 observations in the individuals of group (b) who, for convenience, will be referred to as “Cr responders” and the 29 observations in the individuals of group (a) (“C r nonresponders”) at 90 and 120 minutes after the glucose load during Cr supplementation (Fig 4). Similarly, comparison of the baseline GTT’s (Fig. 3) showed that there was very little overlap between the findings before Cr administration in the “responders” and the “non-responders.” Cr supplementation, therefore, brought to light the existence of two apparently distinct populations among the elderly subjects studied. The first of these, “Cr responders,” had relatively mild abnormalities of the GTT which were not consistently present but which were corrected during Cr administration. The second group, “Cr non-responders,” had more marked and consistently abnormal GTT’s with no improvement in the GTT during Cr supplementation. Since diet was virtually identical in the two groups and was unchanged from the baseline period to the period of Cr administration, it is unlikely that dietary factors other than Cr could have influenced these results. The 4 elderly “responders” to Cr showed variations in the length of time required on Cr supplementation before the GTT’s improved. In 3 subjects (G. F., E. C. and F. W.), glucose tolerance was normal 20, 25 and 26 days, respectively, after beginning the Cr supplement. The fourth subject (H. H.) showed no improvement after 33 days, some improvement at 61 days, and marked improvement after 117 days on the Cr salt. This pattern might imply that the elderly individuals had varying degrees of Cr deficiency. It has been shown that the GTT remained normal for about a month after the substitution of placebo for Cr tablets in the 2 “Cr responders” given the Cr and placebo tablets in this sequence. This observation would suggest that the improvement observed during Cr supplementation resulted from replenishment of a previously deficient body store of Cr, which remained replenished for a month after stopping the Cr tablets. Total body stores of Cr in man have been estimated at 4-6 rngs2 Thus, even a 25 per cent deficiency (corresponding to the change in tissue Cr concentration at age 80, compared with that at age 20) might imply a 1 mg. deficit. Assuming that 1 per cent of exogenous Cr was absorbed2J5 and that all absorbed Cr was retained, it would take 666 days to replace a 1 mg. deficit, when daily Cr supplementation was limited, as in this study, to 150 pg. Cr, which is 2-3 times the usual daily intake. Therefore, it is possible that some of the “non-responders” might have been severely deficient of Cr and might have shown improvement in the GTT if Cr supplementation had been continued over a longer period of time. The observation that the “non-responders” failed to show a significantly reduced serum Cr level before and during Cr supplementation is somewhat against this possibility but does not rule it out, since serum Cr concentrations may not be a good reflection of total body Cr content. The significantly lowered serum Cr level after glucose administration in the “non-responders” suggests that the glucose-induced rise in serum Cr concentration may be of greater importance than the fasting Cr level in influencing the disposition of a glucose load.
The Cr content of the food offered to the elderly subjects was similar to that actually consumed during a total of 7 days by one of the young subjects and was within the range considered to be normal. This finding makes it unlikely that deficiency of Cr in the diet offered to these institutionalized elderly subjects was responsible for any depletion of body Cr content which they might have manifested. However, it was not possible to exclude the strong possibility that failure to eat all of the food offered might have reduced Cr intake to inadequate levels in some of the elderly subjects. The studies of Schwarz et al4 clearly showed that in Cr-deficient rats, glucose disposal rates were markedly enhanced by Cr administration. Also, in vitro studies by Mertz et al.;’ demonstrated that glucose uptake by adipose tissue taken from Cr-deficient animals was greatly augmented by the addition of trace amounts of Cr. The mechanism of action of Cr on the disposition of a glucose load and the source of the increased serum Cr concentration which we and other9 have observed after glucose ingestion, are unknown. The findings of the present study have no direct bearing on the possibility that Cr deficiency may play a role in the pathogenesis of diabetes mellitus, though they suggest that such a possibility is worthy of study. It is of interest that Ghnsmam~ and Mertz+‘” have shown that the glucose tolerance tests of certain maturity-onset diabetics were improved during the administration of oral Cr supplements. The possible importance of dietary Cr as a glucose tolerance factor in human subjects is also suggested by the recent observation of Hopkins” that the abnormal GTT’s in infants suffering from kwashiorkor were frequently restored to normal upon oral Cr supplementation. Thus, cvidence is accumulating that Cr may play an important role in human as well as in animal metabolism.
Grateful acknowledgment is made to Mr. Bredric J. Feldmxn for kindly performing the Cr determinations, to Dr. Peter H. Wright for the supply of guinea pig anti-insulin serum, and to Dr. Kenneth W. Wright, Miss Beatrice Latremore and the Nursing Staff of the Onondaga County Home for their generous cooperation in the performance of this study. The insulin used as a standard (Lot #795372) was furnished by Dr. Otto K. Behrens. Eli Lilly & Company, Indianapolis, Indiana. Dr. Walter hlertz kindly supplied the chromium tablets and much helpful advice. We are indebted to h$rs. Anne B. Holden and Mrs. Carol Dolich for technical assistance. Some of the data were processed in the Researoh Computer Center of the Upstate hledical Center.
1. Streeten, D. II. I?., Cerstein, M. M., hIarmor, 8. hf., and Doisy. R. 1.: Rcduced Chlcose Tolerance in elderly human subjects. Diabetes 14:579, 1965. -7. Schroeder, H. A., Balassa. J. J., and Tipton, I. H.: Abnormal Trace Metals in h&n. J. Chron. Dis. 15:941, 1962. 3. hlertz, W.. Roginski. E. E., and Schroe-
der. II. A.: Some aspects of glucose metabolism of chromium deficient rats raised in a strictly controlled environment. J. Nutr. 86:167, 1965. 4. Schwarz, K.. and Mertz, W.: Chromium (III) and the glucose tolerance factor. Arch. Biochem. 55392. 1959. 5. hlertz, W., Roginski, E. E., and Schwarz, K.: Effect of trivalent chromium com-
ORAL
6.
7.
8.
9.
10.
Il.
CHROhIIUhI
SUPPLEMENTATION
plexes on glucose uptake by epididyma1 fat tissue of rats. J. Biol. Chem. 236:318, 1961. Hagen, J. M., Ball, E. G., and Cooper, 0.: Studies on the metabolism of Adipose Tissue. J. Biol. Chem. 234:781, 1959. Doisy, R. J.: Plasma insulin assay and adipose tissue metabolism. Endocrinology 72273, 1963. Glinsmann, W. H., and Mertz, W.: Effect of trivalent chromimn on glucose tolerance. Metabolism 15:510, 1966. Feldman, F. J., and Purdy, W. C.: The atomic absorption spectroscopy of chromium. Anal. Chim. Acta 33:273, 1965. Schroeder, H. A.: Chromium deficiency in rats: A Syndrome simulating Diabetes Mellitus with retarded growth. J. Nutr. 88:439, 1966. Fajans. S. S.. and Conn, J. W.: The early recognition of diabetes mellitus. Ann. NY Acad. Sci. 82208, 1959.
125 12. Dillon, R. S.: Importance of the hematocrit in interpretation of blood sugar. Diabetes 14:672, 1965. 13. Samaan, H., and Fraser, R.: Typical and atypical serum insulin-like activity in untreated diabetes mellitus. Lancet _‘: 311, 1963. 14. Yalow, R. S., and Berson, S. A.: Immunoassay of endogenous plasma insulin in man. J. Clin. Invest. 39:1157, 1960. 15. Mertz, W., Roginski, E. E., and Reba: R. C.: Biological activity and fate of trace quantities of intravenous chromium (III) in the rat. Amer. J. Physiol. 209:489, 1965. 16. Glinsmann, W. H., Feldman, F. J., and Mertz, W.: Plasma chromium after glucose administration. Science 152: 1243,1966. 17. Hopkins, L. L., and Maja, A. S.: Normalization of impaired glucose utilization and hypoglycemia by chrominm III in malnourished infants. Abstr. Fed. Proc. 25303, 1966.
of Oral Chromium
Glucose Tolerance By ROBERT A.
Supplementation
of Elderly Human
on the
Subjects
LEVINE, DAVID H. P. STREETEN
AIYD RICHARD J, DOISY The possible role of Cr deficiency in the impairment of glucose tolerance in elderly subjects has been studied. Chromium supplements, 150 pg. daily, were given to 10 apparently healthy inmates of an old-age home, whose oral glucose tolerance tests (G.T.T,‘s) were abnormal. Mean GTT became normal in 4 subjects (“Cr responders”) but remained abnormal in the other 6 subjects (“Cr non-responders”) during Cr administration. The beneficial effect of Cr in the “Cr responders” did not result from an increased release of insulin, since
plasma ILA and immune-reactive insulin levels were normal and rose normally after the glucose load both before and during Cr supplementation. Serum Cr concentration was found to rise significantly after the glucose load in old and young subjects but to a lower level in the elderly “Cr non-responders,” suggesting that an adequate rise in serum Cr might be necessary for normality of the G.T.T. The data support the possibility that Cr deficiency may lead to the impairment of the GTT in some elderly subjects. (Metabolism 17: No. 2, February, 114-125, 1968)
N AN EARLIER COMMUNICATION’ it was shown that the abnormal glucose tolerance of elderly human subjects was due neither to retarded glucose absorption nor to a reduced rise in plasma insulin-like activity after the glucose load, but to some other (unknown) cause of delayed cellular uptake of glucose from the plasma. It is known that the chromium content of human tissues declines with advancing age.” Furthermore, rats ingesting chromium-deficient diets develop abnormal glucose disposal rates”.4 and diminished fat-pad sensitivity to exogenous insulin.” On normal diets, rat fat-pads show decreasing sensitivity to insulin with increasing ageSss7 In view of these facts, the present study was instituted to ascertain whether elderly subjects with abnormal glucose tolerance tests would show an improved glucose tolerance during oral chromium supplementation, as had been observed in laborator\, animals and in some patients with diabetes mellitus.x
I
From the Departments of Medicine and Biochemistry, State University of Nerc York, Upstate Medical Center, S:yracuse, New York. Supported bt~ Grants AM -5252 und AM 03025 from the National Institute of Arthritiq and Metabolic Diseases. Received for pz~blicution April 28, 1967. ROBERT A4. LEVINE, M.D.: Intern in Medicine, Mount Sinai Hospital, New York, N. f. This paper wus written during tenure of a medical student summer traineeship of the NIH. DAVID H. P. STREETEN, M.B.. D. PHIL, M.R.C.P.: Professor of Medicine, State Uniuersifq of New York, Upstate Medical Center, Sgraeuse, N. I’. RICHARD J. DOISY, PH.D.: Assoc~te Professor of Biochemistry, State Unioersity of Netc l‘ork. C7psfatc AlcrlicuI Ccntc~, Syracuse, N. Y.
ORAL CHROMIUM
115
SUPPLEMENTATION METHODS
Clinical Material Fifteen elderly subjects (9 male, 6 female) in the Onondaga County Home were selected from volunteers for the study. The subjects were at least 74 years of age, had been in the Home for six months or longer, and had no family history of diabetes. They were alert, had no known disease (except for inactive rheumatoid arthritis in one subject and well treated hypothyroidism in another) and were taking no medications that might have interfered with glucose tolerance tests (GTT). All had fasting plasma glucose concentrations below 120 mg./ 100 ml. Sixteen healthy young adults (12 male and 4 female physicians, medical students and laboratory technicians, aged 19-32 years) served as a control group. One of these young subjects was found to have one abnormal and two normal GTT’s during the study. All subjects were prepared for 3 days prior to testing, with a diet containing at least 300 Gm. of carbohydrate daily. With the subjects in a seated or reclining position after fasting overnight, venous blood was drawn through a siliconized needle into 3 evacuated tubes for determinations of plasma glucose con~ntration, serum insulin-like activity @LA), and serum Cr concentration. The tubes for glucose determination contained potassium oxalate (20 mg.) and sodium fluoride (25 mg.). The tubes for Cr determination were special Pb-free Vacutainer tubes which had been shown to contain no measurable traces of water-soluble Cr compounds. Sera for Cr analysis were transferred to tubes thoroughly rinsed with distilled water and previously found to be free of detectable amounts of Cr. The subjects then ingested 100 Gm. of glucose in aqueous solution and venipuncture was repeated at 30, 60, 90 and 120 minutes. Glucose concentrations were measured in all plasma samples, and serum ILA and chromium determinations were carried out on the samples obtained at 0 and 60 minutes. The specimens were centrifuged within 2 hours after completion of the glucose tolerance tests and stored at -40” C. Plasma glucose concentrations were measured by a standard ferrocyanide method using the AutoAnalyzer (Technicon). Serum ILA levels were determined by a rat epididymal fat pad assay. r In the latter part of the study, simultaneous assays were performed with and without the addition of guinea pig anti-insulin serum (AIS). Serum chromium determinations were kindly performed by Fredric J. Feldman at the Walter Reed Army Medical Center using atomic absorption spectrophotometry.sJ’)
The premise upon which this pilot study was based was that impaired glucose tolerance might be due to Cr deficiency in some or all elderly subjects, which might have resulted from inadequate intake or inadequate body stores of Cr. A given individual might be normal with respect to serum Cr concentration and yet have moderate Cr deficiency. The time taken to replete an individual with orally administered Cr supplements would depend on the degree of deficiency and on the dosage of the administered Cr. If Cr deficiency was responsible for the observed abnormality in GTT, it was anticipated that Cr administration would restore the GTT to normal. Glucose tolerance tests were performed at approximately monthly intervals, the first L? tests in each individual being done before treatment. Plasma glucose concentrations were considered to be “diabetic” in at least one of the “baseline” tests in 9 of the elderly subjects and to be abnormal but without all of the diabetic features in one subject. by the criteria of Fajans and Corm11 modified because plasma and not whole-blood concentrations were measured in the present study. Plasma and whole-blood glucose concentrations differ somcwhat inconsistently from one another because of the variably lowered concentrations of glucose in the formed elements of the blood and because of the normal range of fluct~lations in the hematocrit. Since there are no published Iimits of normality of the plasma glucose concentrations after a glucose load, new criteria of abnormality were used. These were based on the evidence of Dillon12 that, assuming an hematocrit of 45 per cent, the criteria of Fajarrs and Connil for diabetes, using whole-blood glucose concentrations would correspond with a peak plasma glucose concentration exceeding 185 mg./lOO ml., and with 90 and 120minute concentrations above 160 and 140 mg./lOO ml.. respectively. Further studies on the
116
LWIXE,
Fig.
l.-Plasma
concentrations performed subjects stration. glucose served 100 248 at
in
mg./lOO
10
elderlv
Limits
of plasma
concentrations
122-278
90
min.
DOISY
&mose
Cr adminiob-
116-200
ml. at 30 mg./lOO
min.,
AND
in 20 GTT’s
before
were
STHEETEN
mg./
min., 146ml. at 60 mg./lOO
and
ml.
131-280
ml. at 120
min.
,
0.5 Hours
1.0 After
Glucose
1 5
,
2.0
Lood
elderly subjects were restricted to the 10 individuals in whom the baseline GTT was “diabetic” or “abnormal” on at least one occasion. The objective of the study was to administer Cr tablets to each individual for 3 or 4 months or until the mean results of at least 2 GTT’s were within normal limits. This treatment period was to be followed or preceded by the administration of placebo tablets. In fact, because some of these elderly volunteers were not prepared to continue in the study for the length of time planned, only 4 of the 10 elderly subjects received both placebo and Cr tablets, and the remaining 6 received only the Cr tablets for approximately 4 months (5 subjects) or 1 month (1 subject). Ten of the young individuals were given Cr tablets, each for 14 months. The Cr tablets contained 256 ug. CrCb.GHaO (each with 50 ug. Cr) in an inert base and were given 3 times daily with meals, since the solubility of CrCL is known to be reduced at alkaline pHs, and an acid pH might be expected to enhance absorption. RESULTS
Plasma Glucose Concentration-Baseline
Tests
Among the 16 young adult subjects, baseline glucose tolerance tests were all normal except for one abnormal test in one subject. Nine of the 15 elderly subjects (60 per cent) had at least one “diabetic” baseline test and at least one of the listed criteria of abnormality was present in one or more GTT in 13 of the 15 elderly subjects. The 20 “baseline” tests in the 10 elderly subjects who received Cr supplements are depicted in Fig. 1. It is evident that the plasma glucose concentration at 120 minutes exceeded 140 mg./lOO ml. in 18 of the 20 GTT’s the two exceptions being in the same individual (F. W. ).
ORAL
Table
CHROMIUM
l.-Mean
117
SUPPLEMENTATION
2-hour Plasma Glucose Levels During in 10 EMerly Subjects Group
Subiect
1 2 3 4
cv AN CZ AM
Age
Wt. in Kg.
%sf
F
81
57.5
2
M M
83 76 74
Tolerance
Tests
a
Sex
M
Glucose
68.2 69.1 77.7
Cr
Glucose Level, mg./lOO ml.
244
3
-t
2 3
+
2 5
+
2 2
+
212 209 237 192 206 191
5
hIH
F
81
35.9
2 1
+
6
MC
M
84
64.3
2 1
-t
196 166 168 162 234 Avg.
194% 12
209*
10
Group b GF
M
79
53.2
2 2
+
170 +
II
M
78
53.0
2 3
Ech
F
88
46.8
2 3
+
FW
F
96
58.0
2 3
+
106 167 140 153 131 135 100 Avg.
156*8 P <
119* 10 .05
Tests During Cr and Placebo Administration During Cr administration the mean plasma glucose concentrations in the GTT’s remained “diabetic” in 6 elderly subjects (group a) but were “normal”, in all respects, in the remaining 4 elderly subjects (group b, Fig. 2 and Table 1). In contrast with the considerable overlap between the mean plasma glucose concentrations on and off Cr supplements in the subjects of group a, there was no such overlap, at 90 and 120 minutes, in group b. This difference between the mean GTT results during Cr supplementation in the subjects of groups (a) and (b) was also a consistent finding in each of the individual tests on the subjects of the 2 groups. Thus, the subjects of these groups appeared to have responded to Cr administration like two separate populations, without overlap of any plasma glucose concentrations at 90 and 120 minutes. Morever, closer inspection of Fig. 2 suggests that these 2 populations were recognizably distinct even in the tests before Cr supplementation. Thus, the mean plasma glucose concentrations before Cr treatment were almost invariably lower at 90 and 120 minutes in the subjects who subsequently became normal on Cr (group b) than in the subjects who remained “diabetic” on Cr (group a).
118
LEVINE,
1
300
STREETEN
AND
DOISY
(b)
270
240
J
, 0
0.5
y-control
/ -0
Limits
/ 5
Hours
2,O
After
,j
-Control 0
Glucose
0.5
b0
Llmlts
3.5
2.0
Load
Fig. S.-Mean plasma glucose concentrations in GTT’s performed before and during Cr supplementation in 6 elderly subjects whose GTT s were abnormal both
on and off Cr (group a) and 4 elderly subjects whose mean GTT’s became normal on Cr (group b), Limits of baseline tests in all 10 elderly subjects are enclosed within heavy lines. The results of the GTT’s in 2 elderly subjects of group (b) are shown in Fig. 3. It is evident that the GTT was improved within 25 days of starting the Cr supplements in one subject (E.C.), while the other (H.H.) showed a more gradual improvement. The placebo tablets had no evident effect on the mean GTT’s of the 2 elderly subjects whose abnormal GTT’s showed no response to Cr (group a). The GTT results in the 2 elderly subjects of group (b) who were treated with the placebo, and both of whom happened to receive placebo tablets after treatment with Cr, are shown in Fig. 4. The prompt and consistent improvement in the results during Cr administration is evident, especially in the 90- and lZO-minute plasma glucose concentrations. All or most of the improvement in the results was still evident in the tests done 27 and 25 days after the substitution of placebo for Cr tablets. However, during continued administration of the placebo, the 60-, 90- and 120-minute plasma glucose concentrations were always above all of the corresponding values during Cr administration, except in the test done at 248 days in subject G.F., which was very similar to those performed during Cr supplementation. Among the 10 young subjects given Cr supplements, this treatment did not affect the consistently normal baseline GTT’s observed (Table 2). The one individual who had a single diabetic baseline GTT was consistently normal while receiving Cr.
ORAL
CHROMIUM
On Chromium (days 12-129)
Baseline (days I-1 2) Ooy
of Study_,_ 210
119
SUPPLEMENTATION
45
r
&29_
,73,
H. H
1 E.O.
T
Fig. 3.-Plasma glucose concentrations in all GTTs performed on 2 elderlv subjects of group (b) (H.H and E.C.) before and during Cr administration. Days of each GTT are numbered from the dav of the first baseline test. Plasma glucose concentrations are shown before and at 30, 60, 90, and 120 min. after each glucose load.
P
60
1
OnChrormum (days
Serum CT Concentrations
6-l 16)
(Table 3)
no significant differences between the young and elderly groups of subjects. There was a rise in serum Cr concentration from 0 to 60 minutes after the glucose (Table 3) which was highly significant (P < .OOl) in both the young and the elderly groups of subjects when all of the values were analyzed by the “t” test for non-independent variables (n = 55 and 60, respectively. At 60 minutes after the glucose load, the serum Cr level was significantly lower in the elderly subjects of group (a) than in the young subjects (P < .05) during baseline tests. Mean
fasting
serum Cr levels showed
Serum ILA Concentrations In the fasting state, mean serum ILA showed no significant differences between young and elderly groups of subjects (Table 3). There was an increase in ILA 60 minutes after glucose administration, which was significant in the young and the elderly groups both before and during Cr supplementation (P < .05 to < .005). Serum ILA levels were determined both in the presence and in the absence of guinea pig anti-insulin serum, in samples of human serum obtained during 57 of the glucose tolerance tests performed in the present study. The mean results of these determinations are shown in Table 4 in which the corresponding results obtained by Samaan and Fraserls are shown for comparison. It is
120
LEVINE,
Baseline (day. t-26) Day ol Study
z z
u
I80
~ 1
On Chromium (days 27-1201
AND
DOISY
On Placebo ldoys 121-231)
~
:
&;
(
STREETEN
_ 231
z
-
z
z .?
150 -
z
120.
(3
h a SOi/
Day of Study
:;:’
-TV Baseline ldoyr l-14)
34’ On Chromium (days l5-S2)
77
T”5-s On Placebo (days 53-2461
Fig. 4.-Plasma glucose concentrations in all GTT’s performed on 2 elderly subjects of group (b) (F.W. and G.F.) before Cr, during Cr and during placebo administration. Days of each GTT are numbered from the day of the first baseline test. Plasma glucose concentrations are shown before and ;lt 30, 60. 90 kind 120 min. after each glucose load.
evident both from the present data and from the findings of Samaan and Fraser that the immunologically reactive ILA is the only component of the serum insulin-like activity which increased after a glucose load, from mean values of 26 and 43 pU./ml. to mean values of 183 and 188 @J./ml. respectively in the 2 studies. When the tests on subjects with known abnormalities of carbohydrate tolerance were eliminated from the data, the mean rise in ILA after glucose was 116 pU./ml. (39 tests), in good agreement with the report of Yalow and Berson”* that serum insulin concentration, measured by immunoassay, rose by 118 @J/ml. after the same load of glucose. Thus, glucose-induced changes in ILA levels measured by the fat-pad assay closeIy reflect changes in immunoreactive insulin concentrations in the serum. Dietary
Cr Irltakc
Table 5 shows the Cr content of the total diet offered to each of the elderly institutionalized subjects over a T-day period and of the diet consumed bv one Table 2.-PEQsnza Glucose Conc~ntru~jons ~~g./~OO ml.) During In Young Subjects On and OfF Chromium (Mean -e SEMI
GTT’s
121
SUPPLEMENTATION
of Oral Chromium Administration on Serum ILA and Cr Concentrations
&3.-E5eet
est.7
ILA, ,-~O;ts/ml. No. of Subiects Tests
( Meaxr& SEM )
0’
Cr. ng./ml.#ean
-(_ “EM:
seline nCr
16 10
31 25
197f12 199t30
319’29 292246
23.2 L+I 24.312
27.912 32.6f2
4.5*2 7.6*2
Jaseline On Cr
10 10
20 26
1792 I9 174+-14
312+61 379 f 68
21.7*3 24.6& 1
27.624 29.6*3
5.8-t3 4.5*2
Baseline
6
12
197k-29
343*96
19.0*3
21.8*2
2.1*1
On Cr
6
15
188*22
427+
28.253.k
2.853.k
Group (b) Abnormal Old
Baseline
4
8
152klS
265+51
26.0+6
36.2+10b
Abnormal Old
On Cr
4
I1
155k4
308+53
25.0”33
31.526
-___
..lormal Old
108
24.3i:
1
11.5~6* 6.723
A Values are not determined by difference of (66-O’) mean values but are mean values for these differences in each subject. * Included in this mean value is one 60’ determination which was unusually high and may partially account for the higher value in this gronp. t Serum Cr determinations 60 min. after glucose were obtained in only S of the 6 elderly subjects of group (a).
young subject for a week. The daily intake of Cr was quite variable in both instances. Analyses of the drinking water indicated that Cr levels were below the limits of detection ( 10 ng./ ml. ) . Grudges in Body Weight There was no consistent change in body weight during these studies, mean baseline weight being 61.7 and 53.0 Kg., mean weight during Cr administration being 62.0 and 52.8 Kg. in groups (a) and (b), respectively. Table
&-Serum
KLA in the Presence and Absence of Guinea Pig
Anti-in~lin Sera (AIS) During Oral G,T.T.‘s MEAN SERUM ILA, ,-units/ml. FASTING Authors
%Y
ILA + AIS
Present
190
149*
Samaan ancl Fraser’::
-
164
Immunoreactive Insulin
22’
60 MINUTES Immunoreactive Insulin
ILA + AIS
26
309
147
183 (57)-/-
43
-
165
188 (10)
Values of Immuno-reactive Insulin are mean differences between total ILA and ILA measured in presence of AIS. ‘The non-immunoreactive component of the ILA was essentially the same at 60 minutes as in the fasting state, as shown above. In 12 GTT’s ILA levels were also determined in the presence and absence of AIS at 0, 30, 60, 90, and 120 minutes. The respective mean values of non-~munoreactive ILA obtained were 157, 149, 149, 139, 146 iu-units/ml., further suggesting that this component of ILA is unchanged in normal subjects during the G.T.T.‘s. TNumber of tests indicated in parentheses.
122
LEVIA-Fa0 STl{
Table S.-Chromium
Analysis of Total Daily Food
Elderly Subjects
Young
ClgO,U&ll
_____
l&y
1”
Day
2”
1.15 47
3 4 5 6 7”
9 91 35 63 5
Mean
= 52 pg./dav ---z
b.1
.\‘111111,11 Silh (%I ..i:
1 ” :3 -1 3 6
4 li<1 4.;
I I .5 SJr!
7-7 -_-_--__
Ilean
Com~os~tiotz of ~1~erI~ S~~j~c~s’ Diet “Days 1, 3, and ‘7 were selected as representing high, medium Day 1. 115 pg. Criday
z
65 ,iLg./tlat
and Low intakes of (:r.
Breakfast: Orange juice, farina, toast, milk, coffee. Lunch: Cream of tomato soup, egg souffle, tossed salad, applesauce, tea. Dinner: Meat loaf with gravy, mashed potato, beets, cottage cheese, cookies, milk,
tea.
Day 2. 47 pg. Cr/day Breakfast: Prune juice, oatmeal, one egg, 2 slices toast, 1 cup milk, coffee. Lunch: Chicken noodle soup, roast beef hash, crushed pineapple, 1 cup tea.
I)inner: Baked chicken, dressing, gravy, mashed potato, 1 slice bEad, peas, fruit jelto, milk, tea. Day 7. 5pg. Criday Breakfast: Prune juice, farina, egg, toast, milk, coffee. Lunch: Clam chowder, tuna fish sandwich, white cake, tea. Dinner: Creamed cod fish, mashed potato, peas, apricots, 1 slice bread, milk. Tap water in both groups was found to contain less than 10 ng. per ml.
The present studies have confirmed previous evidence that ~~abnori~lal~ies~ of the oral glucose tolerance test are common in elderly subjects. Thus, 60 per cent of 15 individuals had at least one baseline curve which was “diabetic” by criteria modified for plasma glucose concentrations from the whole-blood glucose values proposed by Fajans and Corm. I1 The elderly subjects were considerably lighter (mean weight 58.5 Kg.) th an the group of 16 young adults (mean weight 77.8 Kg.) of whom 15 were found to have GWs consistently within the stated limits of normality. However, the weight discrepancy could not have accounted for the impaired tolerance in the elderly subjects since the in these elderly individuals closely glucose load (100 Gm. or 1.71 Gm./Kg.) approximated the load of 1.75 Gm./Kg. recommended by Fajans and Conn. Moreover, there was no difference between the mean change in weight during chromium administration in the elderly individuals of groups (a) and (b) (mean weight change of 0.3 and 0.2 Kg., respectively), all of whom continued to receive the same diet. Chromium administration was associated with disappearance of all abi~o~al features of the mean GTT in 4 of the 10 elderly individuals so treated, and with no change in the GTT of the remaining 6 subjects. When the plasma glucose concentrations of these 2 groups of elderly subjects were compared, it was found that there was complete separation, without any overlap what-
ORAL
CHROMIUM
SUPPLEMENTATION
123
ever, between the 21 observations in the individuals of group (b) who, for convenience, will be referred to as “Cr responders” and the 29 observations in the individuals of group (a) (“C r nonresponders”) at 90 and 120 minutes after the glucose load during Cr supplementation (Fig 4). Similarly, comparison of the baseline GTT’s (Fig. 3) showed that there was very little overlap between the findings before Cr administration in the “responders” and the “non-responders.” Cr supplementation, therefore, brought to light the existence of two apparently distinct populations among the elderly subjects studied. The first of these, “Cr responders,” had relatively mild abnormalities of the GTT which were not consistently present but which were corrected during Cr administration. The second group, “Cr non-responders,” had more marked and consistently abnormal GTT’s with no improvement in the GTT during Cr supplementation. Since diet was virtually identical in the two groups and was unchanged from the baseline period to the period of Cr administration, it is unlikely that dietary factors other than Cr could have influenced these results. The 4 elderly “responders” to Cr showed variations in the length of time required on Cr supplementation before the GTT’s improved. In 3 subjects (G. F., E. C. and F. W.), glucose tolerance was normal 20, 25 and 26 days, respectively, after beginning the Cr supplement. The fourth subject (H. H.) showed no improvement after 33 days, some improvement at 61 days, and marked improvement after 117 days on the Cr salt. This pattern might imply that the elderly individuals had varying degrees of Cr deficiency. It has been shown that the GTT remained normal for about a month after the substitution of placebo for Cr tablets in the 2 “Cr responders” given the Cr and placebo tablets in this sequence. This observation would suggest that the improvement observed during Cr supplementation resulted from replenishment of a previously deficient body store of Cr, which remained replenished for a month after stopping the Cr tablets. Total body stores of Cr in man have been estimated at 4-6 rngs2 Thus, even a 25 per cent deficiency (corresponding to the change in tissue Cr concentration at age 80, compared with that at age 20) might imply a 1 mg. deficit. Assuming that 1 per cent of exogenous Cr was absorbed2J5 and that all absorbed Cr was retained, it would take 666 days to replace a 1 mg. deficit, when daily Cr supplementation was limited, as in this study, to 150 pg. Cr, which is 2-3 times the usual daily intake. Therefore, it is possible that some of the “non-responders” might have been severely deficient of Cr and might have shown improvement in the GTT if Cr supplementation had been continued over a longer period of time. The observation that the “non-responders” failed to show a significantly reduced serum Cr level before and during Cr supplementation is somewhat against this possibility but does not rule it out, since serum Cr concentrations may not be a good reflection of total body Cr content. The significantly lowered serum Cr level after glucose administration in the “non-responders” suggests that the glucose-induced rise in serum Cr concentration may be of greater importance than the fasting Cr level in influencing the disposition of a glucose load.
The Cr content of the food offered to the elderly subjects was similar to that actually consumed during a total of 7 days by one of the young subjects and was within the range considered to be normal. This finding makes it unlikely that deficiency of Cr in the diet offered to these institutionalized elderly subjects was responsible for any depletion of body Cr content which they might have manifested. However, it was not possible to exclude the strong possibility that failure to eat all of the food offered might have reduced Cr intake to inadequate levels in some of the elderly subjects. The studies of Schwarz et al4 clearly showed that in Cr-deficient rats, glucose disposal rates were markedly enhanced by Cr administration. Also, in vitro studies by Mertz et al.;’ demonstrated that glucose uptake by adipose tissue taken from Cr-deficient animals was greatly augmented by the addition of trace amounts of Cr. The mechanism of action of Cr on the disposition of a glucose load and the source of the increased serum Cr concentration which we and other9 have observed after glucose ingestion, are unknown. The findings of the present study have no direct bearing on the possibility that Cr deficiency may play a role in the pathogenesis of diabetes mellitus, though they suggest that such a possibility is worthy of study. It is of interest that Ghnsmam~ and Mertz+‘” have shown that the glucose tolerance tests of certain maturity-onset diabetics were improved during the administration of oral Cr supplements. The possible importance of dietary Cr as a glucose tolerance factor in human subjects is also suggested by the recent observation of Hopkins” that the abnormal GTT’s in infants suffering from kwashiorkor were frequently restored to normal upon oral Cr supplementation. Thus, cvidence is accumulating that Cr may play an important role in human as well as in animal metabolism.
Grateful acknowledgment is made to Mr. Bredric J. Feldmxn for kindly performing the Cr determinations, to Dr. Peter H. Wright for the supply of guinea pig anti-insulin serum, and to Dr. Kenneth W. Wright, Miss Beatrice Latremore and the Nursing Staff of the Onondaga County Home for their generous cooperation in the performance of this study. The insulin used as a standard (Lot #795372) was furnished by Dr. Otto K. Behrens. Eli Lilly & Company, Indianapolis, Indiana. Dr. Walter hlertz kindly supplied the chromium tablets and much helpful advice. We are indebted to h$rs. Anne B. Holden and Mrs. Carol Dolich for technical assistance. Some of the data were processed in the Researoh Computer Center of the Upstate hledical Center.
1. Streeten, D. II. I?., Cerstein, M. M., hIarmor, 8. hf., and Doisy. R. 1.: Rcduced Chlcose Tolerance in elderly human subjects. Diabetes 14:579, 1965. -7. Schroeder, H. A., Balassa. J. J., and Tipton, I. H.: Abnormal Trace Metals in h&n. J. Chron. Dis. 15:941, 1962. 3. hlertz, W.. Roginski. E. E., and Schroe-
der. II. A.: Some aspects of glucose metabolism of chromium deficient rats raised in a strictly controlled environment. J. Nutr. 86:167, 1965. 4. Schwarz, K.. and Mertz, W.: Chromium (III) and the glucose tolerance factor. Arch. Biochem. 55392. 1959. 5. hlertz, W., Roginski, E. E., and Schwarz, K.: Effect of trivalent chromium com-
ORAL
6.
7.
8.
9.
10.
Il.
CHROhIIUhI
SUPPLEMENTATION
plexes on glucose uptake by epididyma1 fat tissue of rats. J. Biol. Chem. 236:318, 1961. Hagen, J. M., Ball, E. G., and Cooper, 0.: Studies on the metabolism of Adipose Tissue. J. Biol. Chem. 234:781, 1959. Doisy, R. J.: Plasma insulin assay and adipose tissue metabolism. Endocrinology 72273, 1963. Glinsmann, W. H., and Mertz, W.: Effect of trivalent chromimn on glucose tolerance. Metabolism 15:510, 1966. Feldman, F. J., and Purdy, W. C.: The atomic absorption spectroscopy of chromium. Anal. Chim. Acta 33:273, 1965. Schroeder, H. A.: Chromium deficiency in rats: A Syndrome simulating Diabetes Mellitus with retarded growth. J. Nutr. 88:439, 1966. Fajans. S. S.. and Conn, J. W.: The early recognition of diabetes mellitus. Ann. NY Acad. Sci. 82208, 1959.
125 12. Dillon, R. S.: Importance of the hematocrit in interpretation of blood sugar. Diabetes 14:672, 1965. 13. Samaan, H., and Fraser, R.: Typical and atypical serum insulin-like activity in untreated diabetes mellitus. Lancet _‘: 311, 1963. 14. Yalow, R. S., and Berson, S. A.: Immunoassay of endogenous plasma insulin in man. J. Clin. Invest. 39:1157, 1960. 15. Mertz, W., Roginski, E. E., and Reba: R. C.: Biological activity and fate of trace quantities of intravenous chromium (III) in the rat. Amer. J. Physiol. 209:489, 1965. 16. Glinsmann, W. H., Feldman, F. J., and Mertz, W.: Plasma chromium after glucose administration. Science 152: 1243,1966. 17. Hopkins, L. L., and Maja, A. S.: Normalization of impaired glucose utilization and hypoglycemia by chrominm III in malnourished infants. Abstr. Fed. Proc. 25303, 1966.