Demonstration of increased concentrations of circulating glycated insulin in human Type 2 diabetes using a novel and specific radioimmunoassay

Abstract

Aims/hypothesis. Glycation of insulin, resulting in impaired bioactivity, has been shown within pancreatic beta cells. We have used a novel and specific radioimmunoassay to detect glycated insulin in plasma of Type 2 diabetic subjects. Methods. Blood samples were collected from 102 Type 2 diabetic patients in three main categories: those with good glycaemic control with a HbA 1c less than 7%, moderate glycaemic control (HbA 1c 7-9%) and poor glycaemic control (HBA 1c greater than 9%). We used 75 age-and sex-matched non-diabetic subjects as controls. Samples were analysed for HbA 1c , glucose and plasma concentrations of glycated insulin and insulin.Results. Glycated insulin was readily detected in control and Type 2 diabetic subjects. The mean circulating concentration of glycated insulin in control subjects was 12.6±0.9 pmol/l (n=75). Glycated insulin in the good, moderate and poorly controlled diabetic groups was increased 2.4-fold (p<0.001, n=44), 2.2-fold (p<0.001, n=41) and 1.1-fold (n=17) corresponding to 29.8±5.4, 27.3±5.7 and 13.5±2.9 pmol/l, respectively. Conclusion/interpretation. Glycated insulin circulates at noticeably increased concentrations in Type 2 diabetic subjects. [Diabetologia (2003) [1]; the process of glycation in the beta cell is rapid and occurs in a time-and concentration-dependent manner [2]. Previous studies have shown that glycated insulin has a reduced ability to regulate plasma glucose homeostasis in vivo and to stimulate adipose tissue lipogenesis or glucose uptake and oxidation by isolated diaphragm and abdominal muscle in vitro [3,4,5]. Studies in healthy human volunteers using the hyperinsulinaemic-euglycaemic glucose clamp technique suggest that glycated insulin could contribute to insulin resistance in Type 2 diabetes mellitus [6].The site of glycation of human insulin has now been identified by electrospray tandem mass spectrometry as the N-terminal Phe 1 of the B-chain [7], enabling the development of a sensitive and specific radioimmunoassay to measure concentrations of glycated insulin and to identify glycated insulin in pancreatic islets using immunohistochemistry [8]. High performance liquid chromatography techniques used previously were neither sensitive nor reliable Formation of advanced glycation-end products (AGEs) plays an important role in long-term metabolic consequences of diabetes including ophthalmic, renal and atherosclerotic vascular complications. Glycation has been shown also to interfere with normal cellular functions including the activities of various enzymes such as Cu-Zn superoxide dismutase and several other peptide hormones. A growing body of evidence now exists to support the hypothesis that glycation of insulin in pancreatic beta cells occurs under conditions of hyperglycaemia. Glycated insulin has been identified in the pancreas of normal and diabetic animal models

Antibodies to post-translationally modified insulin in type 1 diabetes

“…Hyperglycaemia can unbalance the redox status later in the disease and contribute to a vicious cycle by providing additional neoepitopes to which the immune system is not tolerant. Indeed, insulin glycation has been detected both in the pancreas in vivo [29] and in the serum of individuals with diabetes [30]. Interestingly, we did not observe increased reactivity to glycated insulin in our study.…”

Role of Advanced Glycation End Products in Hypertension and Atherosclerosis: Therapeutic Implications

“…Furthermore, glycation of insulin and proinsulin has been found to occur in pancreatic b-cells contributing to bcell dysfunction in diabetic animal models [127]. Increased levels of circulating glycated insulin have been found in diabetic animal models and in humans with Type 2 diabetes, and this modified insulin showed reduced biological activity [128][129][130][131]. This potential of AGEs to exacerbate insulin resistance and abnormalities in glucose metabolism, will perpetuate excesses of methylglyoxal production and AGE formation, and the subsequent downstream detrimental effects on protein function.…”

Modulation of insulin action

“…Insulin itself has been shown to be glycated within beta cells, resulting in impaired biological activity [129]. In a recent report, by using a specific radioimmunoassay, it was shown that glycated insulin circulates at higher concentration in Type 2 diabetic patients [130].…”