Contents Detail:

THE LANCET, OCTOBER 21, 1989 p. 946-949

RESPONSES TO HUMAN AND PORCINE INSULIN IN HEALTHY SUBJECTS

R.J. HEINE, E.A.P. VAN DER HEYDEN, E.A.VAN DER VEEN

Departments of Internal Medicine and Endocrinology, Free University Hospital, PO Box 7057, 1007 MB Amsterdam, The Netherlands

SUMMARY

In a double-blind randomized study, eight healthy men received equimolar amounts of human or porcine insulin by infusion (50mU/kg per hour). Insulin potencies, in terms of the amount of glucose infused to maintain euglycemia, were almost identical.Hypoglycemia   (blood glucose concentration 2.0mmol/l or below[36mg/dl]) was then induced and the symptoms and hormonal counter-regulatory responses were recorded. The number of sympathoadrenergic  (but not neuroglycopenic) symptoms was significantly greater with the porcine than with human insulin,as was the integrated noradrenaline response. Glucagon, growth hormone, cortisol, and adrenaline responses were similar for the two insulins.

INTRODUCTION

Unawareness of hypoglycemia  ("hypoglycemia unawareness"), a hazard in insulin-treated diabetics is said to arise more frequently with human insulin than with animal insulins. (1-4)Consequently, there have been calls for the continued manufacture of porcine insulin. These have generated controversy because no consistent differences are reported between the counter-regulatory hormone responses to the different types of insulin. (5) Controlled trials comparing human and animal insulin have failed to demonstrate the frequency of hypoglycemia requiring treatment. (3,5-8) One study  (3) showed fewer sympathoadrenergic symptoms in patients transferred from porcine to human insulin. We have investigated the matter in healthy,rather than diabetic, volunteers to exclude the influences of autonomic neuropathy and variations in metabolic control on catecholamine responses.

SUBJECTS AND METHODS

Eight healthy male volunteers with a mean age of 23  (range 20-30years) and a mean body mass index of 22.9 kg/m^2  (range19.1-25.2) took part. They had never before received insulin injections and had no family history of diabetes. The Free University Hospital ethical committee gave permission for the experiments.

The insulins  (40U/ml) were provided by Nordisk. The semi-synthetic human, short-acting insulin was manufactured by chemical conversion of porcine to human insulin. The measured protein contents of the semi-synthetic human and highly purified porcine insulins were 1.41mg/ml  (batch 6843) and 1.43mg/dl  (batch 6929), respectively.

STUDY DESIGN

Each subject was studied on two occasions separated by one week.Subjects fasted from 2200 h the evening before and were admitted to the metabolic ward at 0800 h. After insertion of one cannula in a dorsal hand vein, for sampling of arterialised venous blood,and a second cannula in the antecubital fossa of the contralateral arm, for infusions, the subjects rested in a supine position of at least 1 hour. Samples were taken for basal values of glucose and hormone levels. (9) The subjects were randomized to receive human or porcine insulin and administration was double-blind. Then at time t=0 min, intravenous administration of the insulins, at an infusion rate of 50mU/kg per hour, for 150 min. was started. The insulins were diluted with "Haemaccel"   (Hoechst)to reduce insulin absorption by the infusion system. The "manual"glucose clamp technique was performed to maintain blood glucose concentration at 0.3mmol/l [5.4mg/dl] below fasting levels for120 min. (10) At t=120 min the glucose infusion was stopped while the insulin infusion continued, blood glucose concentrations were allowed to drop to 2.0mmol/l [36mg/dl]. Hypoglycemia was maintained until discontinuation of the insulin infusion at t=150min. Blood samples for blood glucose and hormone estimation were taken up to 210 min. Standard questions were used to record neuroglycopenic symptoms  (headache, dizziness, tiredness,inability to concentrate, anxiety, visual disturbances, nausea)and sympathoadrenergic symptoms  (sweating, trembling, warmth,palpitations, pallor) during the hypoglycemic episode. The heart rate was recorded at intervals until 210 min.

ANALYSIS

Blood glucose concentrations were measured by a glucose oxidase method  (Yellow Springs glucose analyzer, Yellow Springs, Ohio,USA). Plasma insulin was analyzed by radio immunoassay  (RIA) (Insik 5; Sorin Biomedica, Saluggia, Italy). Plasma glucagon was measured by RIA  (antiserum K 5563; Novo, Copenhagen, Denmark).Serum growth hormone   (HGHK-2; Sorin Biomedica, Saluggia, Italy)and serum cortisol   ("Cost-A-Count", DPC, Los Angeles, USA) were measured by RIA. Plasma noradrenaline and adrenaline were measured by high-performance liquid chromatography with electrochemical detection. (11) Results are reported as means  (95%confidence intervals) or as indicated. Wilcoxon matched-pairs signed-rank test was used for statistical evaluation.

RESULTS

Serum insulin levels were not significantly different during the human and porcine insulin infusions  (fig 1.1, fig 1.2, fig 1.3).The coefficients of variation of the blood glucose levels during the clamps were all less than 6%. Glucose requirements during the last 30 min  (t=90-120 min) of the euglycemic clamps, reflecting the potency of the insulins were not different -- 28   (95%confidence interval [CI], 11.5-44.5) mg/kg for human insulin compared to 29   (17.5-40.8) for porcine insulin. Blood glucose levels in the two groups were virtually identical throughout the experiments. Glucagon, cortisol, and growth hormone responses to hypoglycemia were almost identical for human and porcine insulin (fig 2.1, fig 2.2, fig 2.3). The significant differences between the insulins were  (for porcine insulin) a greater noradrenaline response, higher heart rate, and more sympathoadrenergic symptoms  (fig 3.1, fig 3.2, fig 3.3, and table below).

PLASMA NORADRENALINE AND ADRENALINE LEVELS, HEART RATE AND SYMPTOMS IN INSULIN PROVOKED HYPOGLYCEMIA
  Human  (95% CI) Porcine  (95% CI)
NORADRENALINE Human Porcine

Basal value  (nmol/l)

 0.79 (0.53-1.04) 0.89 (0.71-0.95)
Time to peak  (min) 33 (26-40) 30 (27-77)
Peak value  (nmol/l) 1.30 (0.99-1.61) 1.79 (1.46-2.12)*
** Response (nmol/l) 12.2 (4.0-20.5) 29.4 (20.9-37.9)*
ADRENALINE Human Porcine

Basal value  (nmol/l)

 0.31 (0.17-0.45) 0.28 (0.12-0.44)
Time to peak  (min) 33 (26-40) 35 (23-47)
Peak value  (nmol/l) 3.59 (2.23-4.95) 4.78 (3.37-6.19)
** Response (nmol/l) 98.4 (60.7-135.9 168.9 (102.9-234.9)

HEART RATE

Human

Porcine

Basal rate  (beat/min)

 61 (54-68) 62 (57-67)
Peak rate  (beat/min) 71 (64-78 77 (70-84)***

HYPOGLYCEMIC SYMPTOMS

Human

Porcine

Neuroglycopenic Symptoms (#)

 1.9 (1.0-2.8) 2.1 (0.9-3.3)
Sympathoadrenergic Symptoms (#) 2.9 (1.7-4.1) 3.6 (2.4-4.8)***
Porcine vs. human insulin 
*p <0.02
*** p <0.05
** Integrated  (area under curve) response between 120 and 210min.
DISCUSSION

We have found that the noradrenaline response to hypoglycemia in volunteers given human insulin is smaller than that in subjects receiving porcine insulin. Why has earlier work  (5,13-15)produced conflicting results? In most studies intravenous bolus administrations of insulin were used resulting in variable hypoglycemia and short-lived hyperinsulinemia. We used infusion of insulin over 2 hours, to allow physiological insulin levels and glucose requirements to stabilize before allowing the glucose levels to drop to 2mmol/l. Advantages of our protocol are the low variability of the glucose nadir and insulin levels. It also allowed comparison of insulin potency. (16) Variables such as degree of metabolic control, presence of autonomic neuropathy,duration of diabetes, presence of insulin antibodies, some drugs and alcohol, (17) affect hypoglycemia awareness. We therefore used healthy subjects.

The physicochemical characteristics of human and porcine insulin are different. (18) Porcine insulin is more lipophilic than human insulin and may have greater blood-brain barrier penetration.This may explain the differences in autonomic responses.

We thank Nordisk, Gentofte, Denmark for the insulins and J.J.P.Nauta for the statistical analysis.

Correspondence should be addressed to R.J.H.

 REFERENCES

1 Berger WG, Althaus BU, Reduced awareness of hypoglycemia after changing from porcine to human insulin in IDDM, Diabetes Care 1987;10:260-61

2 Teuscher A, Berger WG, Hypoglycemia unawareness in diabetics transferred from beef/pork to human insulin, Lancet 1987; ii:382-85

3 Berger WA, Keller U, Honegger B, Jaeggi E, Warnings of hypoglycemia during treatment with human and porcine insulin in diabetes mellitus, Lancet 1989; i:1041-44

4 Teuscher A, Egger M, Human insulin hypoglycaemia ( hypoglycemia , insulin shock, insulin reaction ) unawareness, Lancet 1989; i:1072

5 Berger M, Human insulin: much ado about hypoglycemia  (unawareness) Diabetologia 1987; 30:829-33

6 Sonnenberg GE, Chantelau E, Sundermann S, Hauff C, Berger M, Human and porcine regular insulins are equally effective in subcutaneous replacement therapy. Results of a double blind cross-over study in Type I diabetic patients with CSII, Diabetes 1982 31:600-02

7 Peacock I, Tattersall RB, Taylor A, Douglass CA, Reeves WG, Effects of new insulins on insulin and C-peptide antibodies, insulin dose and diabetic control, Lancet 1983 i:149-52

8 Home PD, Mann NP, Hutchinson AS, et al, A fifteen month double-blind cross-over study of the efficacy and antigenicity of human and pork insulins, Diabetic Med 1984 1:93-98

9 Auburnrad N, Rabin D, Diamond M, Lacy W, Use of heated superficial hand vein as alternative site for the measurement of amino acid concentrations and for the study of glucose and alanine kinetics, Metabolism 1981; 30:936-40

10 Ponchner, M, Heine RJ, Pernet A, et al. A comparison of the artificial pancreas   (glucose controlled insulin infusion system) and a manual technique for assessing insulin sensitivity during euglycemic clamping. Diabetologia 1984; 26: 420-25.

11 Popp-Snijders C, Geenen B, Van der Ileuden EAP. Serum noradrenaline is composed of plasma and platelet noradrenaline. Ann Class Biochem 1989; 26: 191-92.

12 Schluter KJ, Petersen K-G, Sontheimer J, Enzmann F, Kerp L. Different counterregulatory responses to human insulin  (recombinant DNA) and purified pork insulin. Diabetes Care 1982; 5 (suppl 2): 75-81.

13. Rotak C, Althoff P-H, Enzmann F, Schoffling K. Comparative studies on intermediary metabolism and hormonal counterregulation following human insulin  (recombinant DNA) and purified pork insulin in man. Diabetes Care 1982; 5 (suppl 2): 82-89.

14 Owens DR, Vorz JP, Tronier B, Keller U, Luzio S, Turko A. Hormonal counterregulatory responses to human semi-synthetic and recombinant DNA and porcine insulin induced hypoglycaemia ( hypoglycemia , insulin shock, insulin reaction ). Diabetes Res 1988; 8: 17-20.

15 Muller-Esch G, Hall P, Bekemuyts U, et al. Comparative study of hormonal counterregulation during GCIIS-guided insulin hypoglycaemia ( hypoglycemia , insulin shock, insulin reaction ) tests using human insulin (recombinant DNA) and pork insulin. Diabetes Res 1985; 2: 121-25.

16 Heine RJ, Ponchner M, Hanning I, et al. A comparison of the effects of semisynthetic human insulin and porcine insulin on transmembrance ion shifts and glucose metabolism during euglycaemic clamping. Acta Endocrinol 1984; 106:241-47.

17 Frier BM. hypoglycaemia ( hypoglycemia , insulin shock, insulin reaction ) and diabetes. Diabetic Med 1986; 3: 513-25.

18 Chance RP, Kroeff EP, Hoffman JA, Crank BH. Clinical, physical and biological properties of biosynthetic human insulin. Diabetes Care 1981; 4: 147-54.