Serum insulin and growth hormone in lipoatrophic diabetes

Serum Insulin and Growth Hormone

in

Lipoatrophic Diabetes ByNAGUIBA. SAMAANANDJAMES W. CRAIG Hypersecretion of growth hormone has been suspected as a cause of the lipoatrophy and insulin resistance in lipoatrophic diabetes. Serum insulin and growth hormone studies were performed on three siblings with lipoatrophic diabetes to obtain information regarding the role of growth hormone in this syndrome; responses to the administration of glucose tolbutamide, and insulin were investigated. The subject with only mildly abnormal glucose tolerance had serum concentrations of immunoreactive insulin which were abnormally elevated after an oral glucose load, while the two patients with more severe diabetes had normal fasting insulin concentrations which failed to increase after glucose administration. After intravenous tolbutamide, there was an abnormally slow decline in the blood sugar in all three subjects as well as a subnormal rise in serum insulin in the patient with the

most severe diabetes. All patients showed resistance to insulin given intravenously but insulin antibodies were not detected in their sera. The fasting serum immunoreactive growth hormone concentration was normal (< 0.4 to 3.2 mpg./ ml.) in all cases but showed little or no rise in response to hypoglycemia. Fasting plasma nonesterified fatty acid concentrations varied from 529 to 1200 pEq./L. compared to 370~20 in normal subjects and decreased after glucose admlinistration. In these patients the serum insulin content appeared to decrease with increasing clinical severity or duration of diabetes. Increased levels of HGH were not required for maintenance of lipoatrophy or the severe insulin resistance. The relationship between failure to stimulate growth hormone secretion and the pathogenesis of lipoatrophic diabetes is unclear. (Metabolism 18: No. 6, June, 460-468, 1969)

L

AWRENCE’S ORIGINAL DESCRIPTION of lipoatrophic diabetes included several features: complete lipoatrophy; diabetes mellitus characterized by insulin resistance and the absence of ketosis; hepatosplenomegaly; hyperlipemia with cutaneous xanthomata; an increased basal metabolic rate with euthyroidism; and enlargement of the parotid glands and generalized From the Department of Medicine, Case Western Reserve University Sclaool of Medicine, t%veland, Ohio. These studies were supported by Grant T-46-G from the American Cancer Society, Inc., and were conducted in the Clinical Research Center of the University Hospitals of Cleveland which is supported by Grant FR-60 f rom the N.I.H. of the Department of Health, Education and Welfare. Received for publication January 6, 1969. NAGUIB A. SAMAAN, M.D. (ALEXANDRIA), M.R.C.P. (GREAT BRITAIN), PH.D. (LONDON): Formerly Research Associate, Case Western Reserve University School of Medicine, Cleueland, Ohio; presently Chief, Endocrinology Section, Department of Medicine, M.D. Anderson Hospital and Tumor Institute, University of Texas, Houston, Texas. JAMES W. CRAIG, M.D.: Assistant Professor of Medicine, Case Western Reserve University School of Medicine, Cleveland, Ohio. 460

METABOLISM, VOL. 18, No.

6

(JUNE)

1969

461

SERUM INSULIN AND GROWTH HORMONE

1ymphadenopathy.l In subsequent case reports, several other findings have been noted frequently, including prominence of muscles, an increased rate of growth in younger patients, genital hypertrophy, pseudo-acanthosis nigricans, and hirsutism. Parotid gland and lymph node enlargement have not been common features. The etiology of the syndrome is unknown, but growth hormone of anterior pituitary origin has been proposed as the factor responsible for fat mobilization and the insulin-resistant diabetes in these patients.2s3 In view of this possibility, the responses of serum growth hormone and of insulin to several stimuli have been studied in three siblings with lipoatrophic diabetes and are described in this article. MATERIALS

AND METHODS

The clinical features of the three subjects have been described information is presented in Table 1.

Table L-Clinical Patient Sex Age at time of testing (years) Lipoatrophy Duration of diabetes mellitus (years) Ketonuria Hyperlipemia Hepatomegaly Elevated basal metabolic rate Maximum daily insulin dose (units) Interval since last insulin administration Interval since last oral hypoglycemic agent (days)$

and Laboratory B.S. F

previously.4

Some of this

Data J.S. M

O.S. F

27 I)

27 1)

24 (I

15 t 0 0 *

10 0 0 0 0

13 0 0

2000

76 7 months

80 3 years

At least one month 5

9

0 *

12

0 = Absent * ‘= Present + = Observed only in association with starvation, infection, or emotional upset. t ‘= This is the minimum time, since it is the interval between the last dose of an oral hypoglycemic agent and the first of the series of tests which were performed on each subject. For the oral glucose tolerance tests, 50 Gm. of glucose was used. Tolbutamide tests were performed by injecting 1.081 Gm. of sodium tolbutamide (equivalent to 1.0 Gm. of tolbutamide) intravenously within a two-minute interval. The response to insulin-induced hypoglycemia was observed after injecting Regular Insulin intravenously in the amounts shown in Table 3. Blood glucose was measured on the AutoAnalyzer by a modification of the method of Hoffman5 and serum radioimmunoassayable insulin was determined by the double antibody technique of Morgan and Lazarow. 6 Insulin antibodies were measured by both the radioimmunoassay using 25 per cent sodium sulphate to precipitate the antigen antibody complex and the fat pad assay of Samaan and Fraser.’ Growth hormone was labeled by the method of Greenwood, et al.8 and assayed in serum by the radioimmunoassay technique of Schalch and Parkers as modified by Samaan et al.10 The plasma nonesterified fatty acids were measured by the method of Duncomb.

462

SAMAAN

Table 2.-Glucose

Minutes Fasting

30

60

178 100 2.1 491 371 20.8

241 132 2.2

NEFA pEq./L. Glucose mg. % Insulin pU./ml.

120 48 1.9 549 272 19.2

HGH mg./ml. NEFA fiEq./L. Glucose mg. % Insulin pU./ml. HGH mpg./ml. NEFA pEq./L.

<0.4 1200 525 4 <0.4 1019

955 546 5 <0.4 971

Glucose mg. % Insulin pU./ml. HGH mpg/ml.

O.S.

R.S.

Table 3.-Tolbutamide

R.S.

226 172 2.1 470 389 15 <0.4 575 621 8 <0.4 754

<0.4 860

120

150

186 104 3.1 418 334 16 <0.4 565 600 5

153 100 3.6 361 313 18

<0.4 545

Glucose mg. % Insulin pU./ml. HGH mpg./ml. Glucose mg. % Insulin &U./ml. HGH mpg/ml. Glucose mg. % Insulin pU./ml. HGH mpg/ml.

“15-minute f40-minute

92 8.8

co.4 307 7.2 <0.4 473 4 2.2

<0.4 688 654 4 <0.4

Tolerance Tests Minutes

Fasting

O.S.

90

443 17.8 <0.4 768 576 5

Subject

J.S.

CRAIG

Tolerance Tests

Subject -_

J.S.

AND

30

84 32 <0.4 315* 32* <0.4” 474 4 1.7

60

80 32 <0.4 297 21.6 <0.4 450f 4f 1.9t

90

70 36 co.4 278 21.6 <0.4 450 8 1.8

120

___ 150

66 20 0.6 258 4.8 <0.4 415 10 1.7

61 27 1.5 250 5 <0.4 400 12 1.9

value value

kSULTS

The data obtained from these studies are shown in Fig. (glucose tolerance tests), Fig. 2 and Table 3 ( tolbutamide and Table 4 (insulin response tests).

1 and Table 2 response tests)

Blood Glucose The fasting blood sugars ranged from slightly elevated in subject J.S. to 525 mg./lOO ml. in R.S.; the abnormality of glucose tolerance was likewise mildest in J.S. and most severe in R.S. As would be expected in diabetic patients, there was an abnormally slow fall in blood glucose following tolbutamide administration in all three subjects. Large doses of insulin were required to reduce the blood glucose concentration below 100 mg./lOO ml. in these patients. The dose of insulin used to produce hypoglycemia was the maximum amount which couId be administered without dangerous side effects. All patients developed signs and symptoms of hypoglycemia during the I.T.T. Patient OS. complained of retrosternal pain at 120 minutes.

SERUM

INSULIN

BLOOD

ma-

loo

AND

GLUCOSE

1 / 0

GROWTH

463

HORMONE

mc~m %

INSULIN

lo UNITS/ml

N E FA

@eq/litw

J. S.

NORMALS

I

I

60

120 DURING

ORAL

G.TT.

Fig. I-Blood glucose, serum insulin and NEFA levels of three patients with lipoatrophic diabetes during 50-Gm. oral glucose tolerance test compared with 15 normal subjects + SE. BLOOLI

SUGAR

INSULIN

mq %

p UNITS/ml

Fig. B.-Blood glucase and serum insulin in three patients with lipodiabetes atrophic during I.V. tolbutamide test compared with five normal subjects +- S.E. 0

30

60

So MINUTES

-+20

0 AFTER

30

60

SO

120

LV TOLBUTAMIDE

Serum Insulin Patient J.S., who was only moderately diabetic, showed abnormally high immunoassayable ins&n levels during the glucose toIerance test; in the more severely diabetic subjects, there was no significant increase in the serum insulin concentration in response to glucose. After intravenous tolbutamide, the increase in serum insulin concentration was greatest in J. S. and minimal and greatly delayed in R. S. Subject 0. S. showed no insulin response to oral glucose but a near normal rise in serum insulin after tolbutamide. Growth Hormone

(HGH)

Growth hormone was never detected in the serum of O.S. J.S. showed very

___

.._

HGH m,ug./ml.

0.1 Unit/Kg.

J.S.

Glucose mg. X HGH msg./ml. Glucose mg. X HGH mpg/ml. Glucose mg. % HGH mpg./ml. Glucose mg. % HGH mpg./ml. Glucose mg. %

Insulin Dose

80 Units 1 Unit/Kg. OS. 175 Units 3.6 Unit/Kg. R.S. 145 Units 3.5 Unit/Kg. 170 Units 4.1 Unit/Kg. Normals (15)

Subject

116 <0.4 320 <0.4 524 3.0 325 2.0 67 k2.3 2.5 k1.0

Fasting

62 <0.4 260 <0.4 462 2.1 302 <0.2 35 23.0 7.4 -Cl.5

30

Table

53 0.6 200 co.4 400 1.6 235 <0.2 45 22.3 23.3 22.2

60

4.-Insulin

64 2.2 125 <0.4 342 2.0 170 <0.2 55 10.9 22.1 k2.0

90

Tolerance

60 1.2 75 <0.4 287 <0.2 152 0.6 61 8.5 22.1 51.4

120

Minutes

Tests

64 <0.4 70 <0.4 223 0.4 110 1.7

150

75 f.04 184 .05 77 3.2

180

127 <0.2 63 1.2

210

97 0.3

270

-

75 4.0 63 1.6

360

SERUM INSULINANDGROWTHHORMONE

465

slight increases in the concentration of this hormone late in the glucose tolerance and tolbutamide response tests and when the blood sugar level was lowered by insulin. Small quantities of growth hormone were found in the serum of R.S., but the concentration did not rise significantly in response to the marked fall in the blood sugar level produced by large intravenous insulin doses. Non-esterified

Fatty Acicls (NEFA)

The fasting levels were either slightly or moderately higher than normal (37Or+20 pEq./L.). After glucose administration, the concentration declined at a rate similar to or faster than that observed in normal control subjects despite the impairment of glucose tolerance and the poor insulin response in subjects O.S. and R.S. DISGUSSION Speculations regarding the pathogenesis of lipoatrophic diabetes can be divided into those which involve a primary abnormality of adipose tissue and those which postulate that the adipose tissue changes are secondary. Pituitary growth hormone has been suggested as the agent responsible for both the adipose tissue changes and the insulin-resistant diabetes.2*3 However, no direct evidence of an excess of growth hormone has been found in patients with this syndrome. *,12-14One of the patients of Derot et all4 had a normal fasting serum level of growth hormone with a slight rise during an intravenous glucose tolerance test. Their second patient had slightly elevated levels which fluctuated during an oral glucose tolerance test; these authors had observed similar concentrations of growth hormone in other patients with poorly controlled diabetes mellitus who did not have lipoatrophy. Serum growth hormone levels were normal in the patient of Hamwi et. al.13 and showed only a slight rise at the sixth hour of a glucose tolerance test. The response of growth hormone to tolbutamide or to insulin-induced hypoglycemia has not been reported in such patients previously except for a mention by Oseidl” in an abstract that insulin did not produce any rise in the plasma levels of growth hormone in patients with generalized lipodystrophy. Our results confirm the absence of excessive levels of growth hormone in the serum of patients with lipoatrophic diabetes. In addition, they demonstrate a failure of the serum concentration of growth hormone to increase normally in response to a lowering of the blood sugar. Although the degree of hypoglycemia induced was not great, the blood sugar was lowered sufficiently (from 54 to 85%) that a rise in growth hormone concentration would have been expected. A rapidly falling blood glucose concentration has been observed to stimulate growth hormone secretion even in the absence of hypoglycemia. 16,17 Since our patients were studied after the establishment of lipoatrophy and diabetes mellitus, the participation of growth hormone in the development of the condition is not ruled out. However, an excessive amount of immunologically detectable growth hormone is apparently not responsible for the maintenance of lipoatrophy or insulin resistance. These findings do not, of course, exclude the involvement of some other lipotropic insulin antagonist or fat-mobilizing factor as reported by Louis et a1.16 and

466

SAMAAN

AND

CRAIG

Hamwi et al.13 The absence of clinically significant ketosis in these patients has been attributed to a lack of storage of mobilizable fat.l Low levels of growth hormone may also be associated with a decreased tendency to ket0sis.l” The reason for the failure of serum growth hormone to respond to the stimulus of blood sugar lowering is not clear. The presence of diabetes mellitus is, in itself, apparently not an adequate explanation for the failure of growth hormone response in these patients. During glucose tolerance tests in nonobese diabetic patients without lipoatrophy. Yalow et al.“O found a late rise in plasma growth hormone concentration which, however, tended to be less than in normal subjects; increases in plasma growth hormone concentration may also accompany insulin-induced decreases in the blood sugar concentration in diabetic patients.lr There has been no consistent evidence of insufficiency of other pituitary hormones in patients with lipoatrophic diabetes.2s21*22 All three of our patients’ thyroid function was normal and in OS. and R.S. the urinary excretion of 17-ketosteriods and 17-hydroxycorticoids was also normal. More specific tests of pituitary function have not been performed. The possibility exists that secretion of growth hormone might be suppressed by the presence of immunologically dissimilar growth hormone-like substance; the same substance might be involved in the pathogenesis of this syndrome. The majority of reports have indicated that the fasting levels of serum insulin are elevated in lipoatrophic diabetes when determined by either bioassayQ” or immunoassay;24~25 values at the upper limit of normal have also been found,13*14 and the fasting value was normal in the patient of Torikai et al.12 The increase in the concentration of serum insulin in response to glucose administration has varied from excessive14,25 to delayed or suboptimal.13s14 Samols found high fasting serum levels and an excessive insulin response to provocative tests in patients with lipoatrophy who had no abnormality of glucose tolerance.?” Samaan reported an abnormally high insulin-like activity in a mild diabetic patient suffering from the same disease.23 It is of interest that the patient (J.S.) in the present report who had the mildest diabetes of the shortest duration exhibited an excessive rise in serum insulin during the glucose tolerance test, while the other two patients failed to respond to glucose. Likewise, when tolbutamide was administered, J.S. had the greatest rise in serum insulin, while R.S. showed the smallest increase. Such observations suggest that the natural history of diabetes mellitus in patients with generalized lipoatrophy may include a phase of excessive insulin responsiveness followed by increasing insulin deficiency. Samaan23 and SamolsZ5 have speculated that the high levels of insulin in lipoatrophic patients are evidence of resistance to endogenous insulin. This insulin resistance has been attributed to a lipotropic insulin antagonistli or fat mobilizing factor, l3 to hyperlipemia,14 or to the absence of adipose tissue which is normally an important site of insulin action.l,” Field26 failed to demonstrate a circulating insulin antagonist in the serum of a patient with lipoatrophic diabetes and insulin resistance, and the plasma of the patient described by Schwartz et al.2 did not inhibit the effect of insulin on the rat hemidiaphragm. The patient studied by SamaarP and described by Tizard”? showed normal

SERUM

INSULIN

AND

GROWTH

467

HORMONE

values of insulin antagonists as measured by Vallence-Owen and absence of insulin antibodies. Insulin antibodies were not found in the serum of these patients described above. Subject O.S. showed no insulin response to oral glucose but a near normal rise in serum insulin after tolbutamide. This finding is consistent with the hypothesis that the mechanism of insulin release by tolbutamide is different from that of glucose. Chronic hyperglycemia may have blunted the glucose, mediated release mechanism or the process may have been impaired by structural or enzymatic alterations within the pancreatic beta cells.28* It is of interest that glucose produced a prompt decline in serum NEFA in our patients despite the abnormality of glucose tolerances and insulin responses. HamwiG patient showed a decrease in serum NEFA after glucose, and the patient of Schwartz et a1.2 had a normal fasting NEFA and a sluggish but definite fall after insulin administration. The observed abnormalities in serum growth hormone and insulin on the basis of lipoatrophic diabetes remains unexplained. ACKNOWLEDGMENTS The authors wish to thank Dr. Olof H. Pearson for his guidance laboratory facilities, and Mrs. Mary Young, Head Nurse, participation in the care of these patients.

Clinical

and for the use of his Research Center, for her

REFERENCES 1. Lawrence, R. D.: Lipodystrophy and hepatomegaly with diabetes, Iipaemia, and other metabolic disturbances. Lancet 1:724731, 773-775, 1946. 2. Schwartz, R., Schafer, I. A., and Generalized Iipoatrophy, Renold, A. E.: hepatic cirrhosis, distrubed carbohydrate metabolism and accelerated growth (Iipoatrophic diabetes). Amer. J. Med. 28: 973-985, 1960. 3. Hansen, A. E., McQuarrie, I., and Ziegler, M. R.: Lipohistiodiaresis. A syndrome of lipodystrophy universalis, accelerated growth, lipemia, hepatic cirrhosis and insulin-resistant diabetes without ketosis. Lancet 81:53%541, 1961. 4. Jolliff, J. W., and Craig, J. W.: Lipoatrophic diabetes and mental illness in three siblings. Diabetes 16:708-714, 1967. 5. Hoffman, W. S.: Rapid photoelectric method for determination of glucose in blood and urine. J. Biol. Chem. 120:51-55, 1937.

6. Morgan, C. R., and Lazarow, A.: Immunoassay of insulin: Two antibody system. Diabetes 12: 115-126, 1963. 7. Samaan, N. A., and Fraser, R.: Effect of circulating antibody to insulin on serum levels of insulin-like activity in rats, guinea pigs, and a diabetic patient. Brit. Med. J. 2:482-485, 1964. 8. Greenwood, F. C., Hunter, W. H., and of IrsrGlover, J. S.: The preparation labelled human growth hormone of high specific radioactivity. Biochem. J. 89: 11P 123, 1963. 9. Schalch, D. S., and Parker, M. L.: A sensitive double antibody immunoassay for human growth hormone in plasma. Nature (London) 203:1141-1142, 1964. 10. Samaan, N. A., Yen, S. S., Friesen, H., and Pearson, 0. H.: Serum placental lactogen levels during pregnancy and in trophoblastic disease. J. Clin. Endocr. 26: 1303-1308, 1966.

*Patient OS. was admitted again to the hospital and H.G.H. levels were measured in serum during a standard arginine infusion test. The maximum rise was 4.5 wg./ml. which was subnormal compared to 30.25 f s.e. 4.06 rryrg./ml. we found in 12 normal subjects of the same age range and sex. The I.T.T. test was also repeated using 200 units regular insulin intravenously. H.G.H. levels during the test were < 1 nyrg./ml. in spite of the fall of the blood sugar level from 225 mg./lOO ml. fasting to 45 mg./lOO ml. after 3% hours.

466 11. Duncomb: W. B.: The calorimetric microdetermination of non-esterifled fatty acids in plasma. Clin. Chim. Acta. 9:122125, 1964. 12. Malaisse, W. J., Malaisse, Lague F., Mayhew, D. A., and Wright, P. H.: Effect of sulfa&ureas upon insulin secretion by the rat in pancreas. In Proceedings of the Brook Lodge Symposium. Amsterdam, Excerpta Medica Foundation, 1967, pp. 49-60. 13. Torikai, T., Fuduchi, S., Sasaki, C., Ishigaki, J., Isawa, K., Suzuki, A., Maniki, T., Hashimoto, N., and Hashimoto, S.: Two sibling cases with lipoatrophic diabetes. Endoer. Jap. 12:197-208, 1965. 14. Hamwi, G. J., Kruger, F. A., Eymontt, M. J., Scarpellia, D. G., Gwinup, G., and Byron, R. : Lipoatrophic diabetes. Diabetes 15:262-268, 1966. 15. Derot, M., Rosselin, G., Assan, R., and Tchobroutsky, G.: Tau plasma-atiques de l’insuline et de l’hormone de croissance dans deux cas de diabete lipoatrophique. Societe Medical des Hopitaux de Paris. 117:601605, 1966. 16. Oseid, S.: Studies concerning carbohydrate and lipid metabolism in generalized lipodystrophy. Sixth Congress of International Diabetes Federation, Stockholm, 1967. 17. Louis, L. H., Corm, J. W., and Minick, M. C.: Lipoatrophic diabetes. Isolation and characterization of an insulin antagonist from urine. Metabolism 12:301-307, 1963. 18. Luft, R., Cerasi, E., Madison, L. L., Von Euler, U. S., Casa, L. D., and Roovete, A.: Effect of a small decrease in bloodglucose on plasma, growth hormone and urinary excretion of catecholamines in man. Lancet 291: 254-56, 1966. 19. Roth, J. Glick, S. M., Yalow, R. S., and Berson, S. A.: Plasma insulin and growth hormone levels in obesity and dia-

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betes. Ann. N.Y. Acad. Sci. 131:355-61, 1964. 20. Urgioti, E. J., Houssay, B. A., and Rietti, C. T.: Hypophyseal and adrenal factors essential for ketoacidosis of pancreatectomized dogs. Diabetes 12:301-307, 1963. 21. Yalow, R. S., Glick, S. M., Roth, J., and Berson, S. A.: Plasma inculin and growth hormone levels in obesity and diabetes. Ann, N.Y. Acad. Sci. 131:357, 1965. 22. Seip, M.: Lipodystrophy and gigantism with associated endocrine manifestations. Acta Paediat. 48:555574, 1959. 23. Boudin, G., de Gennes, J. L., Pepin, B., Barraine, R., and Saltiel, H.: Diabete lipo-atrophique avec menifestations neurologiques. Societe Medicale des Hopitaux de Paris 114:895-918, 1963. 24. Samaan, N. A.: The biological effects of insulin on the fat pad with and without antibody and the indication of two types of insulin in blood. Ph.D. Thesis, London University, 1964, p. 236. 25. Ruvalcaba, R. II., Samols, E., and Kelley, V. C.: Lipoatrophic diabetes. I. Studies concerning endocrine function and carbohydrate metabolism. Amer. J. Dis. Child. 109:279-286, 1965. 26. Samols, E. : Immunochemical aspects of insulin. In Leibel, B. S., and Wrenshall, G. A. (Eds. ) : On the Nature and Treatment of Diabetes. Amsterdam, Excerpta Medica Foundation, 1965. 27. Field, J. B.: Studies on the circulating insulin inhibitor found in some diabetic patients exhibiting chronic insulin resistance. J. Clin. Invest. 38:551-556, 1959. 28. Tizard, J. P. M.: Generalized lipodystrophy. Infectious mononucleosis. Mild infantile hemiplegia. Proc. Roy. Sot. Med. 47: 128-219, 1954.