Measurement of human proinsulin by an indirect two-site immunoradiometric assay

Summary An indirect two-site immunoradiometric assay is described for the measurement of human proinsulin in plasma. Polyethylene tubes coated with purified guinea-pig antibodies to insulin were used to extract proinsulin and insulin from plasma. Rabbit antibody to human C peptide was then added to react with the C-peptide moiety of the bound proinsulin. The uptake of this antibody was measured by the subsequent binding of¹²⁵I-sheep antibody to rabbit IgG. The binding of radioactivity to the tubes was a function of the proinsulin concentration in the sample. The sensitivity of the assay was 0.006 pmol/ml. Only 200 l of plasma was required in the assay and the¹²⁵I-labelled antibody was produced from readily available reagents. The polyethylene tubes remained stable for at least 5 months after coating. The mean fasting proinsulin level was 0.009 pmol/ml in sixteen normal subjects and 0.025 pmol/ml in twelve maturity onset diabetics. Oral glucose produced an 8 fold increase in proinsulin concentration but a decline in the plasma proinsulin/insulin molar ratio. Four patients with insulinoma had extremely elevated proinsulin levels and proinsulin/insulin ratios.     

Effects of streptozotocin in vitro on proinsulin biosynthesis,insulin release and ATP content of isolated rat islets of Langerhans

Summary Proinsulin synthesis, insulin release and intracellular ATP concentrations were measured in isolated rat islets of Langerhans under control conditions of in vitro incubation and after treatment with several concentrations of streptozotocin for different periods of time. It was found that streptozotocin inhibited proinsulin synthesis, as well as insulin release, in a time and concentration dependent manner. The characteristics of the inhibition of these two processes were similar in general terms, but one dissimilarity was noted, i.e. after 60 min exposure to a high concentration of streptozotocin, proinsulin synthesis was inhibited more than insulin release. ATP content was reduced by high concentrations of streptozotocin, but it was found that proinsulin synthesis and insulin release could be inhibited without any effect on ATP content by a low (0.22 mM) concentration of streptozotocin. The effect of streptozotocin on proinsulin synthesis was judged to be the result of a target specificity for the B-cell rather than a specific effect on proinsulin relative to total protein synthesis.This work was supported by the Swiss National Scientific Foundation (Grant No. 3106073), Berne, Switzerland, and by a grant-in-aid from Hoechst Pharmaceuticals, Frankfurt-Hoechst, GermanyPreliminary data from this work were presented at the European Symposium on Hypoglycemia, Rome, April 5–6, 1974     

Immunoradiometric assay of insulin,intact proinsulin and 32–33 split proinsulin and radioimmunoassay of insulin in diet-treated Type 2 (non-insulin-dependent) diabetic subjects

Summary Plasma insulin, intact proinsulin and 32–33 split proinsulin measured by specific immunoradiometric assays and insulin and C-peptide measured by radioimmunoassay were measured during a constant infusion of glucose test in ten diet-treated subjects with a history of Type 2 (non-insulin-dependent) diabetes (termed diabetic subjects), mean fasting plasma glucose 6.0 ± 1.0 mmol/l (mean ± SD), and 12 non-diabetic control subjects. Immunoreactive insulin concentrations measured by radioimmunoassay were 33 higher than insulin and 16 % higher than the sum of insulin and its precursors by immunoradiometric assay. The diabetic and non-diabetic subjects had similar fasting concentrations of insulin, intact proinsulin and 32–33 split proinsulin. The ratio of fasting intact proinsulin to total insulin was greater in the diabetic than the non-diabetic group 12.0 % (6.8–21.0 %, 1 SD range) and 6.3 % (4.0–9.8 %), respectively,p < 0.01), though the groups overlapped substantially. After glucose infusion, diabetic and non-diabetic subjects had similar intact proinsulin concentrations (geometric mean 4.9 and 5.2 pmol/l, respectively), but the diabetic group had impaired insulin secretion by immunoradiometric assay (geometric means 55 and 101 pmol/1,p < 0.05) or by radioimmunoassay C-peptide (geometric means 935 and 1410 pmol/1,p < 0.05), though not by radioimmunoassay insulin (87 and 144 pmol/1,p = 0.12), respectively. Individual immunoradiometric assay insulin responses to glucose expressed in terms of obesity were subnormal in nine of ten diabetic subjects. Radioimmunoassay insulin and C-peptide gave less complete discrimination ( subnormal responses in six of ten and eight of ten, respectively). Thus, raised proinsulin and proinsulin:total insulin ratio are not necessarily a feature of mild diet-treated Type 2 diabetic patients with subnormal insulin responses to glucose.     

Metformin restores prohormone processing enzymes and normalizes aberrations in secretion of proinsulin and insulin in palmitate-exposed human islets

Aim

To elucidate how proinsulin synthesis and insulin was affected by metformin under conditions of nutrient overstimulation.

Materials and Methods

Isolated human pancreatic islets from seven donors were cultured at 5.5 mmol/L glucose and 0.5 mmol/L palmitate for 12, 24 or 72 h. Metformin (25 μmol/L) was introduced after initial 12 h with palmitate. Proinsulin and insulin were measured. Expression of prohormone convertase 1/3 (PC1/3) and carboxypeptidase E (CPE), was determined by western blot. Adolescents with obesity, treated with metformin and with normal glucose tolerance (n = 5), prediabetes (n = 14), or type 2 diabetes (T2DM; n = 7) were included. Fasting proinsulin, insulin, glucose, 2-h glucose and glycated haemoglobin were measured. Proinsulin/insulin ratio (PI/I) was calculated.

Results

In human islets, palmitate treatment for 12 and 24 h increased proinsulin and insulin proportionally. After 72 h, proinsulin but not insulin continued to increase which was coupled with reduced expression of PC1/3 and CPE. Metformin normalized expression of PC1/3 and CPE, and proinsulin and insulin secretion. In adolescents with obesity, before treatment, fasting proinsulin and insulin concentrations were higher in subjects with T2DM than with normal glucose tolerance. PI/I was reduced after metformin treatment in subjects with T2DM as well as in subjects with prediabetes, coupled with reduced 2-h glucose and glycated haemoglobin.

Conclusions

Metformin normalized proinsulin and insulin secretion after prolonged nutrient-overstimulation, coupled with normalization of the converting enzymes, in isolated islets. In adolescents with obesity, metformin treatment was associated with improved PI/I, which was coupled with improved glycaemic control.     

Stimulation of proinsulin biosynthesis by K+ in rat pancreatic islets

Glucose-stimulated proinsulin biosynthesis is regulated mainly at the translational level. This study aims at investigating the possible role of the B-cell K⁺ content in such a process. In order to increase the islet cells K⁺ content, rat pancreatic islets exposed to a low D-glucose concentration (e.g., 2.5 mM) were incubated in the presence of 30 or 60 mM K⁺, as distinct from a control extracellular K⁺ concentration of 5 mM. Under these conditions, the K⁺ content of the islets, as judged from the net uptake of ⁸⁶Rb⁺ over 60 min incubation, was increased to a level comparable to that otherwise found in the presence of 16.7 mM D-glucose. In the presence of 2.5–4.0 mM D-glucose, the rise in K⁺ concentration from 5 to 30 and 60 mM caused a progressive increase in the incorporation of l-[4-³H]phenylalanine into both all islet peptides and (pro)insulin. A preferential stimulation of proinsulin biosynthesis was only observed in islets incubated at 60 mM K⁺ in the presence of 4.0 mM D-glucose. In relative terms, the K⁺-induced increase in biosynthetic variables was less pronounced, however, than that otherwise evoked by a rise in D-glucose concentration from 2.5 to 4.0 mM to 5.6 or 16.7 mM. These findings may suggest that the effect of D-glucose to increase the K⁺ content of islet cells represents one modality for coupling a rise in D-glucose concentration to stimulation of proinsulin biosynthesis.     

Stimulation of calcitonin secretion by calcium receptor activators: evaluation using a new, highly sensitive, homologous immunoradiometric assay for rat calcitonin

Current rat calcitonin immunoassays use human calcitonin antisera, and suffer from poor sensitivity, long incubation periods, nonspecific interferences, and unreliability. The homologous immunoradiometric assay (IRMA) for rat calcitonin described here overcomes these problems. Overnight incubation yields a detection limit of 0.4 pg/mL, a standard curve that is linear to >1800 pg/mL, and intra- and interassay coefficients of variation of <7%. Gel filtration chromatography of rat plasma and rat medullary thyroid carcinoma 44-2 cell media showed that the vast majority of immunoreactivity coeluted with calcitonin standard. In 44-2 cells, increasing extracellular Ca²⁺ concentration or incubation with the calcimimetic compound NPS R-467 markedly increased calcitonin secretion. Plasma calcitonin levels were elevated in rats anesthetized with ketamine/xylazine and in conscious rats with chronic renal insufficiency. Calcitonin levels decreased following EGTA-induced hypocalcemia and were undetectable after thyroparathyroidectomy. In normal conscious rats, plasma calcitonin levels averaged 3–5 pg/mL and increased up to 100-fold following calcium (Ca) infusion or NPS R-467 administration. The assay also quantified calcitonin in plasma of normal and Ca-injected mice. This assay has revealed that plasma calcitonin levels in normal rats are much lower than the detection limits of most existing assays, but can increase by 100-fold on activation of the C-cell Ca²⁺ receptor. The rat calcitonin assay described in this paper is available as a kit from Immutopics, Inc., 929 Calle Negocio, Suite A, San Clemente, CA 92673.     

High-performance liquid chromatography (HPLC): A rapid,flexible and sensitive method for separating islet proinsulin and insulin

Summary Evaluating islet function in vitro involves studying both insulin biosynthesis and release. For the former, it is necessary to resolve insulin from its precursor, proinsulin. This has been achieved in the past by various procedures, each of which suffers from major drawbacks in terms of resolution and the time involved. We show here that reversed phase high-performance liquid chromatography (HPLC) outperforms previous methods for separating proinsulin from insulin in is let extracts without any prepurification or concentration steps. This HPLC method is rapid (90 min for a complete cycle, including washing the column) and reproducible, while allowing for unambiguous separation and quantification of proinsulin and insulin.     

Hyperinsulinaemia in obesity is not accompanied by an increase in serum proinsulin/insulin ratio in groups of human subjects with and without glucose intolerance

Summary Serum proinsulin is disproportionately elevated compared to insulin in Type 2 (non-insulin-dependent) diabetes mellitus. We studied the effect of obesity on serum proinsulin with varying degrees of glucose intolerance. Serum proinsulin and insulin were measured during a 75 g oral glucose tolerance test in 73 obese and 74 non-obese subjects with normal, borderline or diabetic-type glucose tolerance. Proinsulin was assayed by a direct radioimmunoassay using proinsulin-specific antiserum. Fasting serum proinsulin and insulin and the summed values of proinsulin and insulin during oral glucose tolerance test were significantly, or tended to be, higher in obese subjects than in those without obesity in each category of glucose tolerance. However, the molar ratio of proinsulin to insulin was nearly the same between obese and non-obese groups with a similar degree of glucose tolerance. On the other hand, the proinsulin/insulin ratio increased progressively with the deterioration of glucose tolerance. We conclude that proinsulin secretion is disproportionately increased in the presence of glucose intolerance but not by obesity itself. Each Beta cell seems to function normally in obese subjects while glucose tolerance remains normal.This work was presented at the 6th International Congress on Obesity in October 1990, Kobe, Japan     

The balance between proinsulin biosynthesis and insulin secretion: where can imbalance lead?

Insulin is stored in pancreatic β-cells in β-granules. Whenever insulin is secreted in response to a nutrient secretagogue, there is a complementary increase in proinsulin biosynthesis to replenish intracellular insulin stores. This specific nutrient regulation of proinsulin biosynthesis is predominately regulated at the translational level. Recently, a highly conserved cis -element in the 5'-untranslated region (UTR) of preproinsulin mRNA, named ppIGE, has been identified that is required for specific translational regulation of proinsulin biosynthesis. This ppIGE is also found in the 5'-UTR of certain other translationally regulated β-granule protein mRNAs, including the proinsulin processing endopeptidases, PC1/3 and PC2. This provides a mechanism whereby proinsulin processing is adaptable to changes in proinsulin biosynthesis. However, relatively few β-granules undergo secretion, with most remaining in the storage pool for ∼5 days. Aged β-granules are retired by intracellular degradation mechanisms, either via crinophagy and/or autophagy, as another long-term means of maintaining β-granule stores at optimal levels. When a disconnection between insulin production and secretion arises, as may occur in type 2 diabetes, autophagy further increases to maintain β-granule numbers. However, if this increased autophagy becomes chronic, autophagia-mediated cell death occurs that could then contribute to β-cell loss in type 2 diabetes.     

Effect of pancreas transplantation and immunosuppression on proinsulin secretion

Insulin resistance and increased demand for insulin secretion occur after successful pancreas transplantation. To investigate the potential effects of immunosuppression and pancreas transplantation on fasting β-cell function, we studied fasting proinsulin and 32,33 split proinsulin secretion cross-sectionally and longitudinally in segmental pancreatic graft recipients (SPx, n = 18); in whole-pancreas graft recipients (WPx, n = 13); in nondiabetic kidney transplant recipients (Kx, n = 14) and in normal subjects (Ns, n = 14). Basal insulin secretion rates were significantly increased in SPx 15.8 (1.7), WPx 24.4 (4.5) and Kx 22.1 (2.1) vs Ns 9.7 (1.6) pmol min⁻¹ l⁻¹, p < 0.05, mean (SEM). Total proinsulin, intact proinsulin and 32,33 split proinsulin concentrations were significantly higher in all the transplanted groups than in normal subjects (p < 0.05), whereas the total proinsulin to C-peptide ratio and the 32,33 split proinsulin ratio were higher in SPx than in WPx, Kx and Ns ( < 0.05). In the longitudinal study, β-cell function in terms of proinsulin secretion remained stable for 1 year. In conclusion, fasting glucose homeostasis in pancreas-kidney transplant recipients is obtained at the expense of increased proinsulin secretion and increased insulin secretion rates, primarily induced by immunosuppression. In segmental pancreas graft recipients, increased fasting proinsulin and 32,33 split proinsulin relative to the number of β-cells transplanted indicate more stress on the residual β-cell and therefore higher secretory demand than in whole pancreas transplant recipients. © 1998 John Wiley & Sons, Ltd.     

Monomethylated-adenines potentiate glucose-induced insulin production and secretion via inhibition of phosphodiesterase activity in rat pancreatic islets

Monomethyladenines have effects on DNA repair, G-protein-coupled receptor antagonism and autophagy. In islet ß-cells, 3-methyladenine (3-MA) has been implicated in DNA-repair and autophagy, but its mechanism of action is unclear. Here, the effect of monomethylated adenines was examined in rat islets. 3-MA, N6-methyladenine (N6-MA) and 9-methyladenine (9-MA), but not 1- or 7-monomethylated adenines, specifically potentiated glucose-induced insulin secretion (3-4 fold; p ≤ 0.05) and proinsulin biosynthesis (～2-fold; p ≤ 0.05). Using 3-MA as a ‘model’ monomethyladenine, it was found that 3-MA augmented [cAMP]_i accumulation (2-3 fold; p ≤ 0.05) in islets within 5 minutes. The 3-, N6- and 9-MA also enhanced glucose-induced phosphorylation of the cAMP/protein kinase-A (PKA) substrate cAMP-response element binding protein (CREB). Treatment of islets with pertussis or cholera toxin indicated 3-MA mediated elevation of [cAMP]_i was not mediated via G-protein-coupled receptors. Also, 3-MA did not compete with 9-cyclopentyladenine (9-CPA) for adenylate cyclase inhibition, but did for the pan-inhibitor of phosphodiesterase (PDE), 3-isobutyl-1-methylxanthine (IBMX). Competitive inhibition experiments with PDE-isoform specific inhibitors suggested 3-MA to have a preference for PDE4 in islet ß-cells, but this was likely reflective of PDE4 being the most abundant PDE isoform in ß-cells. In vitro enzyme assays indicated that 3-, N6- and 9-MA were capable of inhibiting most PDE isoforms found in ß-cells. Thus, in addition to known inhibition of phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3′K)/m Target of Rapamycin (mTOR) signaling, 3-MA also acts as a pan-phosphodiesterase inhibitor in pancreatic ß-cells to elevate [cAMP]_i and then potentiate glucose-induced insulin secretion and production in parallel.     

Monomethylated-adenines potentiate glucose-induced insulin production and secretion via inhibition of phosphodiesterase activity in rat pancreatic islets

Monomethyladenines have effects on DNA repair, G-protein-coupled receptor antagonism and autophagy. In islet ß-cells, 3-methyladenine (3-MA) has been implicated in DNA-repair and autophagy, but its mechanism of action is unclear. Here, the effect of monomethylated adenines was examined in rat islets. 3-MA, N6-methyladenine (N6-MA) and 9-methyladenine (9-MA), but not 1- or 7-monomethylated adenines, specifically potentiated glucose-induced insulin secretion (3-4 fold; p ≤ 0.05) and proinsulin biosynthesis (∼2-fold; p ≤ 0.05). Using 3-MA as a ‘model’ monomethyladenine, it was found that 3-MA augmented [cAMP]_i accumulation (2-3 fold; p ≤ 0.05) in islets within 5 minutes. The 3-, N6- and 9-MA also enhanced glucose-induced phosphorylation of the cAMP/protein kinase-A (PKA) substrate cAMP-response element binding protein (CREB). Treatment of islets with pertussis or cholera toxin indicated 3-MA mediated elevation of [cAMP]_i was not mediated via G-protein-coupled receptors. Also, 3-MA did not compete with 9-cyclopentyladenine (9-CPA) for adenylate cyclase inhibition, but did for the pan-inhibitor of phosphodiesterase (PDE), 3-isobutyl-1-methylxanthine (IBMX). Competitive inhibition experiments with PDE-isoform specific inhibitors suggested 3-MA to have a preference for PDE4 in islet ß-cells, but this was likely reflective of PDE4 being the most abundant PDE isoform in ß-cells. In vitro enzyme assays indicated that 3-, N6- and 9-MA were capable of inhibiting most PDE isoforms found in ß-cells. Thus, in addition to known inhibition of phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3′K)/m Target of Rapamycin (mTOR) signaling, 3-MA also acts as a pan-phosphodiesterase inhibitor in pancreatic ß-cells to elevate [cAMP]_i and then potentiate glucose-induced insulin secretion and production in parallel.     

Evaluation of rat insulin messenger RNA in pancreatic and extrapancreatic tissues

Summary The purpose of these studies was to determine whether insulin detected immunochemically in extrapancreatic tissues of the adult rat is synthesized in situ by quantitating mRNA in these tissues. A blot hybridization assay was utilized with cloned ³²P-proinsulin cDNA. The lower limit of detection was estimated to be 3pg. Proinsulin mRNA concentration was found to be 1000–1500 g in isolated pancreatic islets and was easily detected in total pancreatic RNA at 10–15 pg/ g. Proinsulin mRNA was quantitated in rat insulinoma cells adapted to culture at levels 150 those in normal islets. Samples of RNA (20–50 g) enriched about 50-fold for mRNA sequences by repeated oligo-deoxythymidylate chromatography were assayed. No insulin mRNA was detected in 50 g samples of RNA from brain or in 20 g samples from subsections of brain or other extrapancreatic tissues. RNA samples were undegraded as assessed by ability to stimulate protein synthesis in a cell-free system. Proinsulin mRNA from pancreas was added to brain homogenates and recovered intact. Brain RNA samples with insulin mRNA levels 1:1000 that of pancreas would be predicted to have 50–75 pg proinsulin mRNA/50 g sample assayed if present. Because none was found, brain must have a concentration <1:6,000 that of pancreas. These findings suggest that immunoassayable insulin detected in extrapancreatic tissues of the adult rat is synthesized by the pancreas.     

Gastric bypass alters the dynamics and metabolic effects of insulin and proinsulin secretion.

AIMS: Hyperproinsulinaemia is associated with obesity and is a risk factor for Type 2 diabetes. We explored the dynamics of proinsulin and insulin and postprandial effects on glucose and lipids in subjects who had undergone gastric bypass (GBP) surgery compared with morbidly obese (MO) subjects and normal weight control subjects (NW). METHODS: Subjects free from diabetes were recruited: 10 previously MO subjects [body mass index (BMI) +/- sd, 34.8 +/- 6.2 kg/m2] who had undergone GBP surgery, 10 MO subjects (BMI 44 +/- 3.1 kg/m2) and 12 NW control subjects (BMI 23.2 +/- 2.4 kg/m2). After an overnight fast, a standard meal (2400 kJ) was ingested and glucose, proinsulin, insulin free fatty acids and triglycerides were determined up to 180 min. RESULTS: Fasting proinsulin was similar in the GBP group and NW control subjects, but threefold increased in MO subjects (P < 0.05). Postprandial AUC for glucose was similar in the three groups and AUC for proinsulin was high in MO, intermediate in the GBP group and lowest in NW control subjects (P for trend = 0.020). Postprandial proinsulin at 60 min was similar in the GBP group and MO subjects and twofold higher than in NW control subjects. Postprandial proinsulin at 180 min was normal in the GBP group, but fivefold increased in MO subjects (P = 0.008). Insulin increased rapidly at 30 min in the GBP group and was normal at 90 min, whereas insulin was still increased at 90-180 min in the MO subjects (P < 0.001). CONCLUSIONS: MO subjects, free from diabetes, have elevated proinsulin concentrations in the fasting as well as the postprandial phase. After GBP surgery markedly lower fasting and postprandial proinsulin concentrations were observed, although BMI was higher compared with NW control subjects.     

The serum levels of proinsulin and their relationship with IGFBP-1 in obese children

AIM: Serum proinsulin (PI) levels were investigated in obese children to determine whether PI is a sensitive indicator of insulin resistance, as previously shown in adults with type 2 diabetes mellitus (DM), and to evaluate their relationship with insulin-like growth factor-binding protein-1 (IGFBP-1) known as a predictor of the development of cardiovascular disease in diabetic adults. SUBJECTS AND METHODS: Forty-two obese children without DM (age, 12.1 +/- 1.5 year) and 42 age-matched control children were included in the study. The serum levels of PI, immunoreactive insulin (IRI), IGFBP-1 and free insulin-like growth factor-1 (IGF-1) were measured in the fasting state. RESULTS: The fasting levels of serum PI and IRI were significantly higher in obese children than in controls (PI, 10.5 +/- 6.8 vs. 5.6 +/- 2.0 pmol/l, p < 0.001; IRI, 72.0 +/- 41.8 vs. 32.7 +/- 19.5 pmol/l, p < 0.001). Serum IGFBP-1 levels were significantly lower in obese children than in controls (37.7 +/- 24.6 vs. 76.3 +/- 26.5 microg/l, p < 0.001). The ratio of PI to IRI (calculated as molar ratios) did not differ significantly between obese and control subjects (0.16 +/- 0.08 vs. 0.19 +/- 0.11, p = 0.08). For the whole group, serum PI levels correlated positively with IRI and inversely with IGFBP-1 (IRI, r = 0.67, p < 0.001; IGFBP-1, r = -0.49, p < 0.001). Serum IGFBP-1 levels correlated inversely with both BMI and IRI (BMI, r = -0.73, p < 0.001; IRI, r = -0.60, p < 0.001). Multiple regression analysis revealed that the best predictive parameters for IGFBP-1 were BMI and PI (R2 = 0.57, p < 0.001 and p < 0.05, respectively). CONCLUSION: These findings suggest that fasting serum PI levels may be a better predictor than fasting insulin levels for the future development of type 2 DM and cardiovascular disease in obese children, and PI, in addition to insulin, contributes to the suppression of hepatic IGFBP-1 production.     

Improved Beta Cell Function,with Reduction in Secretion of Intact and 32/33 Split Proinsulin,after Dietary Intervention in Subjects with Type 2 Diabetes Mellitus

Diet in Type 2 diabetes often reduces both hyperglycaemia and weight and both these factors may contribute to improvement in beta cell function. Sixty-nine subjects with newly diagnosed Type 2 diabetes had their glucose, insulin, intact and 32/33 split proinsulin measured at diagnosis, and after 16 (12–20) weeks of conventional diet. In the whole group following diet there was reduction in weight (p < 0.0001), Haemoglobin A₁ (p < 0.0001), and fasting glucose (p < 0.0001). There was a fall in fasting intact proinsulin (p < 0.03), 32/33 split proinsulin (p < 0.0001), and the percentage of fasting proinsulin to total insulin-like molecules (p < 0.0001). Subjects were separated according to weight loss (group 1 < 0.5 kg, group 2 0.5–4 kg, group 3 > 4 kg). Only subjects in group 3 (mean weight loss 6.2 kg) had a fall in fasting insulin (p < 0.04). The fasting glucose achieved following diet was positively correlated with the initial fasting glucose (r = 0.7) and negatively correlated with the degree of insulin deficiency at diagnosis (with the initial 30 min insulin r = ?0.62). The effect of diet on the final concentration of insulin, 32/33 split proinsulin and intact proinsulin was examined using multiple regression. The final insulin concentration was determined by the initial BMI and absolute weight loss, 32/33 split proinsulin concentration by the initial fasting glucose and absolute glucose fall, and intact proinsulin concentration by a combination of both weight loss and glucose reduction. We conclude that diet without weight loss increases the insulin secreted relative to the glucose concentration and reduces the concentration of proinsulin-like molecules. Weight loss reduces insulin resistance thereby lowering the insulin and intact proinsulin concentrations.     

The serum concentration of insulin, C-peptide, and proinsulin in patients with acute pancreatitis

Summary In 14 patients with acute pancreatitis during 16 episodes of the disease the concentrations of blood glucose, serum insulin (IRI), C-peptide (CP), and proinsulin (Pro) were determined in the fasting state on d 1, 2, 3, 5, and 10 after the attack. The peptides were measured using RIAs, and for determination fo CP two antibodies: Byk-Mallinckrodt’s and more specific M-1221 Novo antibodies were used. Apart from sporadic rises in the initial period of the disease, the blood glucose level did not change significantly and had a decreasing trend. On d 1 the mean serum IRI level was 0.17±0.04 (SD) nM, and it decreased on d 5 to 0.06±0.04 nM, rising again to 0.11±0.15 nM on d 10. The serum Pro concentration was on the same days: 11.1±12.6, 4.2±2.4 and 7.5±10.8 pM, whereas the serum CP values determined with M-1221 antibodies were 0.48±0.50, 0.34±0.19, and 0.52±0.25 nM, respectively. However, when serum CP was determined using Byk-Mallinckrodt kits, the concentration on d 1 was 1.90±1.12 nM and over the following days it decreased to 1.08±0.98 nM on d 5 and on d 10 it was 1.11±0.46 nM. In one patient (not included in the calculation of the mean values), in whom the second attack of acute pancreatitis had a fatal outcome, the serum levels of all three peptides were very high, with a particularly evident difference of CP-values, dependent on the antibodies used: 2.47 nM with M-1221 antibodies and 9.90 mM with Byk-Mallinckrodt kits on d 5. We hypothesize that the transient decrease of the serum peptides released into the blood in the process of insulin biosynthesis observed in the early period of acute pancreatitis is owing to their breakdown by the activated pancreatic proteases. Our observation suggests also that in acute pancreatitis in blood appeared an unidentified protein reacting with less specific antibodies to C-peptide.