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Honey is a food known for its medical properties. In this work, we have studied the impact of different types of honey on insulin signalling pathway. We found that honey extracts inhibit the enzyme PTP1B, one of the main negative regulators of insulin receptor signalling. HPLC-MS analysis allowed us to confirm the presence of several natural PTP1B inhibitors in the honey extracts analysed. Statistical analysis methods show a correlation between specific 1 H-NMR resonance frequencies/HPLC peaks and the inhibitory power of the samples. This finding will allow the prediction of the biological properties of honey samples applying relative simple analytical methods. Finally, we demonstrated that the treatment of HepG2 cells with honey extracts enhances the expression of insulin receptor, and stimulates glucose uptake. For the first time, our results demonstrate that bioactive components of honey could improve glycaemic control by both inhibiting PTP1B and stimulating the expression of insulin receptor in liver cells., (Copyright © 2019 Elsevier Masson SAS. All rights reserved.)

Insulin release in the perfused isolated rat pancreas was measured after stimulation with 16.5 mM glucose with and without somatostatin (cycle form, 100 ng/ml) in the medium. A complete blockage of the typical biphasic pattern of insulin release ocurred with somatostatin in the medium. Such blockage was abolished when cAMP (2.5 mM) and a 0.5 ml solution of glucagon (1 mg/ml) were continuously perfused for 20-minute periods and for 30-second periods correspondently. It did not take place when glibenclamide (HB-419) was perfused for a 20-minute period at a rate of 10 mug/ml. The results suggest that the adenylcyclase dependent mechanisms of glucose-induced insulin release are involved in the inhibition of the glucose-induced insulin secretion by somatostatin.

On gel filtration of acid-ethanol extracts from three pancreatic beta-cell adenomas 1.4% to 1.8% of total immunomeasurable insulin (IMI) eluted ahead of proinsulin. This high molecular IMI was resolved into three components. The presence of urea in the dilute acetic acid solutions of extracted tumor tissue did not influence the pattern of gel filtration. High molecular IMI dissolved in dilute acetic acid showed to be stable if immediately rechromatographed, but a partial dissociation to insulinlike and proinsulinlike components (ILC and PLC) was found if rechromatography was performed after 48 h of incubation. Mainly ILC and PLC were found on rechromatography provided high molecular IMI was dissolved and incubated briefly in 0.04M phosphate buffer, pH 7.4. It proved improbable that the proteolytic action of some protein being extracted with the hormones caused a splitting of high molecular IMI at pH 7.4. We conclude from our findings that the components of high molecular IMI are not precursors of proinsulin and insulin but are either self-associated products of the hormones or associations of insulin and proinsulin to other proteins extracted from insuloma tissue.

In agreement with data previously reported (Yalow and Berson 1973), after gel filtration of acid-ethanol extracts of islet cell adenomas (performed in 3 M acetic acid) 1.4% to 1.8% of total immunoreactive insulin (IRI) eluted ahead of proinsulin. The high-molecular IRI (HM-IRI) was, however, found to be heterogenous in size and consisted of at least three components, the major one having an estimated molecular weight of about 50,000. Under certain conditions, HM-IRI was partially dissociated into proinsulin- and insulin-like components. We conclude that HM-IRI does not represent a precursor of proinsulin and insulin, but probably a self-association product of the peptides or an association of insulin and proinsulin to other proteins extracted from tumor tissue.

The role of insulin secretion in the diurnal variation of glucose tolerance has been investigated. In ten healthy subjects, at 08.00 and at 18.00 after 10 hrs of fasting, a combination test of glucose and glucagon was performed. 1 mg glucagon was injected intravenously 40 min after the intravenous infusion of glucose (0.5 g/kg b.w.). Samples for blood sugar (BS) and serum immunoreactive insulin (IRI) were taken before and 2-5 min following the glucose and glucagon loads, and thereafter at 10 min intervals up to 85 min. In the afternoon test, the mean blood sugar values were higher, the differences in the 20-85 min values being statistically significant: the IRI values were statistically lower after glucose, while after glucagon, the increase of serum IRI was apparently similar in both morning and afternoon tests. However, the insulin/glucose ratio (I/G) was significantly lower at 18.00 at 55-85 men. Corresponding results were obtained in six additional healthy subjects when only glucagon (1 mg i.v.) was injected. In this case also, the mean insulin levels were lower in the afternoon after 5 min, while the BS values during the maximal insulin release (2-30 min) were comparable in both the morning and afternoon tests. In the 40-60 min interval, the BS levels were significantly higher in the afternoon. The existence of a diurnal variation in the blood sugar after intravenous glucose load, as well as after glucagon, seems to be correlated to a simultaneous diurnal variation in the insulin response, suggesting decreased pancreatic beta-cell activity in the afternoon.

The effect of glucagon on the inhibition of the insulin response to glucose induced by somatostatin was investigated in humans and in the isolated perfused rat pancreas. Both in vivo and in vitro somatostatin suppressed glucose-induced insulin release. This inhibitory effect of somatostatin was overcome by glucagon. Similar results have been observed in vitro by the infusion of theophylline or cyclic adenosine monophosphate.

Gel filtration of human serum extracts on Bio-Gel P-30 columns produced two peaks of material reactive with insulin antisera. The earlier eluting fraction appeared at the elution position of proinsulin (serum proinsulin-like component, PLC) while the second fraction corresponded in elution volume to insulin. In assays using porcine insulin-(131)I and an antiserum against porcine insulin, human pancreatic proinsulin was less reactive than human insulin. Serial dilutions of the serum PLC in the immunoassay showed immunological identity with the human proinsulin standard. Partial tryptic digestion of the serum PLC yielded products with increased immunological reactivity as estimated with insulin as the standard. With larger amounts of trypsin, all the serum PLC was converted to insulin-like components (desthreonine and desoctapeptide insulin). On the basis of these results we conclude that the earlier eluting fraction of human serum extracts is proinsulin. The fasting values of proinsulin in normal subjects ranged between 0.05 and 0.4 ng/ml, representing from 5 to 48% of the insulin concentration. In one subject the values of proinsulin were higher than those of insulin. After oral administration of 100 g of glucose, the proinsulin levels tended to rise similarly to insulin. Three obese patients with hyperinsulinemia had higher fasting levels of proinsulin and a greater increase after glucose than the normal subjects. As the high levels of proinsulin coexisted with raised insulin concentration in these obese subjects, the relative proportions of the two hormones were in the same range observed in the normal group. Thus hyperinsulinemia in these obese subjects was not accompanied by an increase in the fraction of serum proinsulin. When the values for serum proinsulin were expressed as percentage of the insulin levels, there was a decrease in the per cent proinsulin in the first hour of the glucose tolerance test. After the second hour, the per cent tended to rise towards the fasting levels.

An immunoassay for the measurement of human proinsulin and C-peptide is described. By a combination of this system and an insulin immunoassay, proinsulin, insulin, and C-peptide were identified in acid-ethanol extracts of serum after gel filtration. In keeping with the results of biosynthetic studies, insulin and C-peptide were found to be present in equimolar amounts. The absolute values of serum proinsulin determined independently in the two assays corresponded closely.