Hepatic extraction and renal production of 3,3'-diiodothyronine and 3',5'-diiodothyronine in man.

The sequential deiodination of thyroxine (T4) gives rise to several iodothyronine analogs including 3,3'-diiodothyronine (3,3'-T2) and 3',5'-diiodothyronine (3',5'-T2). In vitro animal studies suggest that the liver and the kidneys are the main sites of both formation and degradation of 3,3'-T2 and 3',5'-T2. To determine the metabolism of 3,3'-T2 and 3',5'-T2 in human liver and kidneys plasma samples were obtained from (a) a brachial artery and a hepatic vein in 20 normal subjects, and from (b) a femoral artery and a renal vein in 11 normal subjects. Further, the hepatic plasma flow (a) and the renal plasma flow (b) were determined. Both plasma 3,3'-T2 and 3',5'-T2 levels were reduced in the hepatic venous blood as compared to arterial values (1.09 +/- 0.40 vs. 1.75 +/- 0.74 ng/dl (P < 0.02)) (mean +/- 1 SD). This resulted in a hepatic extraction of both, 3,3'-T2 and 3',5'-T2, which averaged 8.2 and 5.2 microgram/d, respectively. Plasma 3,3'-T2 as well as 3'5'-T2 levels were higher in the renal vein as compared to arterial values, 1.49 +/- 0.42 vs. 1.39 +/- 0.45 ng/dl (P < 0.05) and 2.35 +/- 0.83 vs. 2.09 +/- 0.81 ng/dl (P < 0.05), respectively. This positive venoarterial difference implies a net production of 3,3'-T2 and 3',5'-T2 in the kidneys of 1.2 and 3.0 microgram/d, respectively. It is concluded that the liver is an important site of 3,3'-T2 and 3',5'-T2 extraction in normal man. In contrast, the renal production of 3,3'-T2 as well as 3'5'-T2 exceeds the degradation and urinary excretion.

Kinetics of human connecting peptide in normal and diabetic subjects.

The metabolic clearance rate (MCR) of synthetic human connecting peptide (C-peptide) was measured with a single-dose injection technique in six normal and seven diabetic subjects and with a constant infusion technique in one normal subject. The MCR of C-peptide did not differ in normal subjects (4.4 ml/min per kg; range, 3.7-4.9) and in diabetic subjects (4.7 ml/min per kg; range, 3.7-5.8). Employment of both techniques in one subject gave similar MCR. The average half-life of C-peptide in plasma calculated from the last 1-h period of the single-dose injection studies was longer in the insulin-dependent diabetics (42.5 min; range, 39.4-48.5) than in the normal subjects (33.5 min; range, 24.9-45.3). These results indicate that the beta-cell secretory capacity of normal and insulin-dependent diabetic subjects can be compared by measuring the C-peptide concentration in peripheral venous plasma. The difference in the half-life of C-peptide in plasma between diabetics and normals suggests an altered kinetics of the disappearance of the peptide, while the overall metabolism, as expressed by the MCR, is similar.

Residual beta-cell function and its metabolic consequences.

Improvement in the sensitivity and specificity of the C-peptide immunoassay and studies of larger groups of patients have increased our knowledge of the importance of residual beta-cell function and its metabolic consequences in insulin-treated diabetic patients. During the first five to 10 years after the onset of diabetes mellitus residual beta-cell function is demonstrable in the majority of insulin-treated patients irrespective of the severity of the initial symptoms and only partly dependent on the patient's age at diagnosis. Residual beta-cell function facilitates good control. Stable patients have a higher C-peptide concentration in plasma than unstable ones, but unmeasurable C-peptide is not always associated with poor control. More data are needed before the full significance of an even minimal reserve of beta-cell function is elucidated. It also remains to be shown whether the reductive in beta-cell function in diabetic patients has a qualitative as well as quantitative component.

C-peptide response to glucagon. A test for the residual beta-cell function in diabetes mellitus.

Pancreatic beta-cell secretory activity was measured in 17 patients with insulin-dependent diabetes mellitus of less than 19 months' duration and in 10 nondiabetic subjects by means of the peripheral plasma C-peptide response to 1 mg. of glucagon I.V. The C-peptide response to a meal was also measured in the diabetic patients. Residual beta-cell function was present in all the diabetic patients as indicated by significant amounts of C-peptide in plasma. Significant increases in C-peptide were observed in 16 after glucagon stimulation and in 15 after the meal. Both absolute and relative increase in C-peptide were reduced in the diabetic patients. The increase in C-peptide was correlated to the fasting C-peptide concentration both after glucagon (r=0.86, p less than 0.001) and after the meal (r=0.66, p less than 0.01). The responses to the meal and to glucagon were correlated (r=0.77, p less than 0.005), indicating a high predictive value of the glucagon test as to how the beta-cells will respond during normal daily life.

Kinetics of human C-peptide in man.

A review of studies concerned with the metabolism of human C-peptide showed that the kidney is the major organ in C-peptide removal. The turnover rate of C-peptide was similar to that of proinsulin but considerably slower than that of insulin. An analysis of corresponding C-peptide and insulin concentrations in normal subjects after the administration of oral glucose or intravenous glucagon was used to define the relationships between the two peptides. These results show that different peripheral vein concentrations of insulin may correspond to identical C-peptide concentrations, depending on sampling conditions.

Prevalence of residual beta-cell function in insulin-dependent diabetics in relation to age at onset and duration of diabetes.

The influence of the age at onset as well as the duration of disease on the prevalence of residual beta-cell function was studied in insulin-dependent diabetic patients. Two hundred and sixty-seven patients presented at an early age (10-19.9 years) and 158 patients had a late onset (30-39.9 years of age). beta-cell function was evaluated by measuring serum C-peptide immunoreactivity. Fifty-six patients (21.0 per cent) in the early-onset group and 64 (40.5 per cent) in the late-onset group had residual beta-cell function. The prevalence of residual beta-cell function was almost 100 per cent during the first two years of disease and was lower thereafter in diabetics with early onset. About 15 per cent of all patients with a duration between 15 and 35 years had residual beta-cell secretory function.

Heterogeneity of circulating human C-peptide.

Values reported for serum C-peptide immunoreactivity in healthy individuals vary considerably. To assess factors that contribute to this finding, three human C-peptide assay systems were developed utilizing three distinct antisera that react differently with various C-peptide fragments. Preparations of natural pancreatic and synthetic human C-peptide standards were compared immunologically in these systems. The curves of the natural C-peptide and the synthetic preparations were not identical. The relative immunoreactivity of each standard varied, depending on the particular antiserum used. Serum C-peptide concentrations varied when measured in the different assay systems. Furthermore, the results of dilution and recovery tests and stability of the C-peptide during storage showed differences among the three assays. Gel filtration of serum indicated heterogeneity within the major C-peptide peak, and in addition, a smaller peak of lower molecular weight material was present in some samples. Although degradation of serum C-peptide may occur during storage, fragments of C-peptide may also be secreted or arise during in-vivo metabolism. Thus, the present studies indicate the need for careful standardization and checking of each particular assay system before its use in clinical studies.

Characterization of seven C-peptide antisera.

The plasma C-peptide immunoreactivity (CPR) in 10 normal subjects varied considerably when measured with different antisera in parallel assays. The CPR level correlated with the blank "CPR" value measured in plasma devoid of C-peptide and to a lesser degree with the sensitivity of the standard curves obtained with the individual antisera. Storage of plasma samples at different temperatures and for different lengths of time before the analyses were carried out resulted in further variation in the CPR results. This was caused by a time- and temperature-dependent fall in CPR, which was more pronounced with some antisera than with others. This sensitivity to storage of plasma did not correlate with the antigenic characteristics of the antisera as determined by their reactivity with 11 specific fragments of the C-peptide molecule. The contribution of human proinsulin to the CPR concentration relative in normal subjects was considered to be negligible even though the relative immunoreactivity of human proinsulin and C-peptide ranged from 11 to 143 per cent among these antisera. These results suggest that differences in C-peptide antisera are a major reason for the variation in the concentration of circulating CPR as measured in different C-peptide immunoassays.

Sulfonylureas. Why, which, and how?

Although controversies remain as to the usefulness of sulfonylureas, most evidence is in favor of their use in many if not patients with non-insulin-dependent diabetes mellitus. When used properly, sulfonylureas improve insulin secretion and action, and these effects may be maintained for years. If combined with hypocaloric dietary regulation, rapid- and short-acting sulfonylureas may help patients reach and maintain euglycemia without provoking chronic hyperinsulinemia or weight increase. There is no evidence that sulfonylurea treatment causes beta-cell exhaustion; instead, the antihyperglycemic effect helps improve beta-cell function. Sulfonylurea "failures" are often dietary failures or may be due to late introduction of these drugs, i.e., when beta-cell function is already attenuated. Desensitization of the insulinotropic effect of sulfonylureas may occur but might be avoided by discontinuous (less than 24 h/day) sulfonylurea exposure, i.e., once-daily administration of a short-acting sulfonylurea in a moderate dose. The most important adverse effect of sulfonylureas is long-lasting hypoglycemia, which may lead to permanent neurological damage and even death. This is mainly seen in elderly subjects who are exposed to some intercurrent event, e.g., acute energy deprivation or a drug interaction, i.e., aspirin. Long-acting sulfonylureas may be more likely to promote long-lasting hypoglycemia. The dose-response relationships of sulfonylureas have been poorly investigated, and sulfonylurea therapy should always be initiated and maintained at the lowest possible dose.

Correlations between fasting plasma C-peptide, glucagon-stimulated plasma C-peptide, and urinary C-peptide in insulin-treated diabetics.

This study correlated fasting plasma C-peptide (CP), plasma CP 6 min after stimulation with 1 mg glucagon i.v., and the mean of three 24-h urinary excretions of C-peptide (UCP)/creatinine in 132 insulin-treated diabetics. Patients were divided into three groups: group 1, stimulated CP less than 0.06 nM (n = 51); group 2, stimulated CP 0.06-0.60 nM (n = 48); and group 3, stimulated CP greater than 0.60 nM (n = 33). In all patients fasting CP was closely correlated to stimulated CP (r = .988, P less than .001), whereas the correlations between UCP and both fasting CP (r = .904, P less than .001) and stimulated CP r = .902, P less than .001) were slightly less pronounced. The associations between UCP and both fasting CP (r = .716, P less than .001) and stimulated CP (r = .731, P less than .001) were modest in group 2, and even more so in group 3 (r = .557, P less than .001 and r = .641, P less than .001, respectively). In conclusion, fasting CP is closely correlated to glucagon-stimulated CP in insulin-treated diabetics and can probably be used equally well in the assessment of beta-cell function. The associations between UCP and both fasting and glucagon-stimulated CP are less pronounced, especially in patients with well-preserved beta-cell function.

Influence of smoking on insulin requirement and metbolic status in diabetes mellitus.

This study was performed in order to examine the influence of tobacco smoking on carbohydrate and lipid metabolism and microangiopathy in diabetic patients with normal serum creatinine. Among 163 adult insulin-treated patients 114 smoked daily (smokers). Compared with nonsmokers, smokers had on the average a 15--20% higher insulin requirement (P < 0.001) and serum triglyceride concentration (P < 0.05), increasing to a 30% rise in heavy smokers (P < 0.01). The degree of retinopathy was equal in the two groups, as was the average creatinine clearance [99 +/- 2 (mean +/- 1 SEM) versus 101 +/- 4 ml/min in smokers compared with nonsmokers]. Smokers and nonsmokers were comparable regarding sex ratio, age at diabetic onset, duration of diabetes, residual beta-cell function, fasting hyperglycemia, and glycosuria. Evidently, tobacco smoking represents a strain on both carbohydrate and lipid metabolism in insulin-treated diabetes mellitus.

Interaction of ethanol and glipizide in humans.

The hypoglycemic effect of 2.5 mg glipizide and the potentiation of this effect by ethanol were studied in 10 normal-weight nondiabetic subjects. The reductions in blood glucose concentrations were similar in time of onset and extent (2 mM) whether glipizide was taken alone or in combination with ethanol. However, the return of blood glucose toward fasting level was delayed by ethanol. Beta-Cell secretory activity, evaluated from the concentrations of insulin and C-peptide, was unchanged by ethanol. The serum glipizide concentrations were reproducible within subjects, whereas there was a considerable interindividual variation. This heterogeneity in the rise in glipizide concentration was strongly correlated with blood glucose fall and insulin secretion. Thus, ethanol can prolong but does not augment the hypoglycemia induced by glipizide. The heterogeneity in glipizide concentration seems to be caused by an interindividual variation in kinetics.

Reproducibility of beta-cell function estimates in non-insulin-dependent diabetes mellitus.

We evaluated the reproducibility of different estimates of endogenous insulin secretion in 30 patients with non-insulin-dependent diabetes mellitus (NIDDM). Fasting blood glucose concentration was similar on the 2 days of study. The coefficients of variation of fasting plasma C-peptide, plasma C-peptide 6 min after the injection of 1 mg i.v. glucagon, and the increment in plasma C-peptide after glucagon were 16.0, 14.8 and 24.1%, respectively. The coefficients of variation of the corresponding plasma insulin values were 19.2, 24.8, and 34.8%, respectively. The coefficient of variation of 24-h urinary C-peptide excretion was 22.1%. Because fasting plasma C-peptide correlated closely with plasma C-peptide 6 min after glucagon (test 1: r = .70, P less than .01; test 2: r = .76, P less than .01), it seems that these two values can be used equally well as assessment of beta-cell function in NIDDM. In conclusion, fasting plasma insulin, fasting plasma C-peptide, and plasma C-peptide 6 min after glucagon stimulation showed a similar and acceptable degree of reproducibility. Plasma insulin 6 min after glucagon and increments in plasma insulin and C-peptide, as well as urinary C-peptide, seem to be less reproducible.

Acute actions of sulfonylurea drugs during long-term treatment of NIDDM.

The acute effect of sulfonylurea drugs during long-term treatment was evaluated in two separate studies. In the first study, the levels of plasma glucose, insulin, and C-peptide were measured after intake of 10 mg glyburide alone or together with a standardized mixed meal in 10 non-insulin-dependent diabetes mellitus (NIDDM) patients treated with 10-20 mg glyburide/day for greater than 2 yr. There was no acute effect of glyburide on the insulin and C-peptide responses to the meal or during continued fasting, indicating the absence of an acute insulinotropic action of glyburide during chronic treatment. The glucose increase after the meal was also unchanged, but a significant glucose reduction was found after glyburide intake during continued fasting, suggesting a sustained acute extrapancreatic (hepatic) effect. In the second study, the diurnal glycemic excursions in 8 NIDDM patients chronically and continuously (24 h/day) exposed to glipizide (2.5-7.5 mg 3 times/day) were similar when the drug was taken 30 min before each of the three main meals and when taken immediately before the meals. It is concluded that there is no acute insulinotropic action of sulfonylurea drugs during chronic continuous exposure, whereas an acute extrapancreatic action may prevail. During such exposure, there seems to be no clinical benefit in taking a sulfonylurea 30 min before rather than at the start of a meal.

Prevalence of fasting hyperglycemia and known non-insulin-dependent diabetes mellitus classified by plasma C-peptide: Fredericia survey of subjects 60-74 yr old.

A Danish population of 5699 individuals (60-74 yr old) was screened by fasting blood glucose (FBG) and interviewed about known diabetes. The distribution of FBG in individuals not known to have diabetes showed no sex difference or significant variation with age. Fasting hyperglycemia (FH), defined as FBG greater than or equal to mM in subjects without a history of diabetes, was found in 1.7% of men and women. Known diabetes (KD) had a prevalence of 3.9 and 5.0% in men and women, respectively. The prevalence rates of FH and KD increased significantly with age. In the two subgroups, plasma C-peptide was measured after overnight fasting and subsequently 6 min after an intravenous injection of glucagon. Based on the distribution of the C-peptide concentrations in non-insulin-treated KD subjects, lower limits for non-insulin-dependent diabetes mellitus (NIDDM) of 0.30 pmol/ml for fasting C-peptide and 0.60 pmol/ml for stimulated C-peptide were arbitrarily chosen. According to these cutoff points, only 38.5% of KD subjects treated with insulin had insulin-dependent diabetes mellitus, corresponding to 9.3% of all KD subjects. After exclusion of these patients, the prevalence of recognized NIDDM was 3.5% in men and 4.5% in women. All FH subjects except one had C-peptide values in the NIDDM interval. A close agreement between fasting and glucagon-stimulated C-peptide was seen. In epidemiological studies with an expected high prevalence of NIDDM, we propose to use fasting C-peptide for classification of patients with insulin-treated diabetes.

Decreased insulin removal contributes to hyperinsulinemia in obesity.

Peripheral plasma insulin and C-peptide concentrations during oral glucose tolerance tests were measured in 7 severely obese and 12 normal weight nondiabetic subjects. The insulin and C-peptide levels as well as incremental areas under the plasma curves were 2--5 times higher in the obese subjects (P less than 0.05). The C-peptide to insulin molar ratios as well as the relation between incremental areas under the plasma curves of the two peptides were used as relative measures of the hepatic insulin extraction. They were both reduced in the obese subjects, which suggests that decreased insulin removal may contribute to the hyperinsulinemia of obesity. Sixty minutes after the oral glucose load, all of the obese patients had higher peripheral venous insulin concentrations compared with those found in normal subjects with similar C-peptide levels. This suggests that the reduced insulin extraction is not entirely explained by increased beta-cell secretory activity, but is also a specific consequence of obesity.

Diminished immunoreactive gastric inhibitory polypeptide response to a meal in newly diagnosed type I (insulin-dependent) diabetics.

The release of immunoreactive gastric inhibitory polypeptide (IR-GIP) in response to a standard meal was examined in 10 normal subjects and 15 type I (insulin-dependent) diabetics 7 days (test I), 14 days (test II), and 3 months (test III) after time of diagnosis. During all three tests, the diabetics had significantly lower plasma IR-GIP concentrations than the controls from 15-90 min after the standard meal. The IR-GIP response in the diabetics measured as the integrated area under the response curve corresponded to 70% of that of normal subjects. beta-cell function evaluated from the C-peptide response to the meal increased significantly from test I to test III whereas the IR-GIP response was similar during all three tests. As GIP is known to potentiate glucose-induced insulin secretion and possibly the biosynthesis of insulin, the low IR-GIP responses in subjects with type I diabetes may significantly influence insulin levels and hyperglycemia.

Heterogeneity of circulating C-peptide.

Serum C-peptide levels vary when measured with different immunoassay systems. In order to assess the factors contributing to this finding, serum C-peptide was measured in two assays utilizing different antisera, but the same standards and labeled peptide. The antisera were characterized with synthetic C-peptide fragments and their reactivities towards some of these fragments differed. The results of dilution and recovery tests and stability of the C-peptide during storage showed differences between the two assays. Gel filtration experiments indicated heterogeneity within the major C-peptide peak, and, in addition, a smaller peak of lower molecular weight material was present in some sera. Although degradation of serum C-peptide may occur during storage or with freezing and thawing, fragments of C-peptide may also be secreted or arise during in vivo metabolism.

Glucose tolerance and B cell function in chronic alcoholism: its relation to hepatic histology and exocrine pancreatic function.

Glucose tolerance and B cell function were assessed in 30 consecutive chronic alcoholic patients without overt diabetes mellitus. Plasma glucose, insulin, and C peptide concentrations were measured during an oral glucose tolerance test. All patients underwent a liver biopsy and an exocrine pancreatic function test (Lundh test). Compared with the controls, the three groups of alcoholic patients (those with histologically normal livers, n = 12; those with steatosis, n = 10; and those with cirrhosis, n = 8) all had a two-fold increase in plasma concentrations of insulin as well as C peptide in the fasting state, despite normal fasting levels of glucose. After oral glucose all groups of patients had elevated plasma levels of glucose, insulin, and C peptide compared with the controls. The C peptide/insulin ratio was similar to that in the controls in all groups of alcoholics. Patients with decreased exocrine pancreatic function (n = 7) had a significantly lower insulin and C peptide response to glucose than the patients with normal exocrine pancreatic function. It is concluded that (1) chronic alcoholics even with histologically normal livers have endogenous insulin resistance, and (2) associated damage to the exocrine pancreas is more common than previously recognized and decompensation of B cell function could be demonstrated in patients with decreased exocrine pancreatic secretion.

ELISA for human proinsulin.

A micro enzyme-linked-immunosorbent-assay (ELISA) for monitoring circulating human proinsulin (hPI) was developed. A micro test plate was coated with guinea pig anti-insulin antibody. As labelling system peroxidase-labelled F(ab1)2-fragments of a guinea pig anti-human-C-peptide was used. The detection limit in buffer (95% level) was 0.6 pmol/l corresponding to 0.06 fmol/incubation well and to 1.2 pmol/l in serum, since samples were diluted 50%. Standard operating range was from 0-160 pmol/l. Interassay variation was 9% estimated from two human control materials (assayed within the range 6-9 pmol/l and 9-14 pmol/l, respectively). Insulin in samples did not interfere in concentrations below 400 pmol/l. Human C-peptide, porcine, and bovine proinsulins did not cross-react even at 10 000 pmol/l. In 38 healthy fasting subjects a reference range less than 1.2-13 pmol/l with a median of 4.1 pmol/l was found. Serum from total pancreatectomised patients showed values below the detection limit. The value from a patient with an insulinoma was 263 pmol/l. When stored at -20 degrees C human proinsulin appeared stable in serum or plasma for at least 9 mth. This ELISA, although among the most sensitive immunoassays for human proinsulin, is still not sensitive enough to measure the concentrations expected in samples from IDDM patients in the fasting state. In spite of this the method is useful in characterising beta-cell function in stimulated situations, as well as in the diagnosis of insulinoma.