Abstract

Hyperglycaemia stimulates a variety of biochemical abnormalities. The area of particular interest in this study is the influence of non-enzymatic glycation of proteins on iron homeostasis, and particularly on non-transferrin-bound iron (NTBI) and its possible relation to atherogenesis in both Type 2 diabetic and obese non diabetic subjects. The link between non-enzymatic glycation of proteins and iron homeostasis, and development of macrovascular disease may be mechanistically different in Type 2 diabetic and obese non diabetic subjects due to a difference in the protein glycation pattern. Because the following in vivo study required storage of samples for up to two years to complete the processing of all the samples, a storage study was carried out using different anticoagulants and addition of reduced glutathione (GSH) to samples to study the effects of storage, thawing and freezing of the samples on the level of malondialdehyde (MDA), a biomarker of lipid peroxidation. This storage study showed that EDTA attenuated the action of lipid oxidation compared with lithium heparin (LiH). A combination of GSH with either EDTA or LiH added more protection from lipid peroxidation in the first week of storage, but due to the thawing and freezing of the sample the action of GSH diminished through its autooxidation, meaning that addition of GSH to samples in the following in vivo study would be useless. An in vivo study was carried out on iron-related parameters in three subject groups: control (non-diabetic, non-obese), Type 2 diabetic and obese non diabetic. Glycated haemoglobin (HbA1C) was strongly correlated with NTBI in the diabetic group. Also the level of NTBI was significantly increased in Type 2 diabetic subjects compared with other groups while the level of total iron was significantly decreased. The study showed a strong positive correlation between NTBI and a biomarker of endothelium dysfunction (E-selectin) in all groups studied. Although it is not possible from the current data to know if there is a causal relationship between these two parameters, it remains a possibility that iron released from its binding sites could initiate oxidative damage to the endothelial cells and begin the process of atherogenesis. Positive correlation at the 90% confidence level between NTBI and a biomarker of inflammation, high sensitivity C-reactive protein, is another indicator in this study of a link between increases in NTBI, inflammation, endothelium dysfunction and atherosclerosis. This study also showed for first time that NTBI is present in higher levels in the plasma of obese subjects compared to controls despite the obese subjects having significantly lower total iron. An in vitro study found that glycation of transferrin half saturated with iron increased with increasing glucose concentration, leading to decreased capacity of transferrin to hold iron and increased release of free iron. Also co-incubation of transferrin half saturated with iron with low density lipoprotein (LDL) and glucose showed oxidation of LDL (measured as MDA). This may be explained by the effect of glycation, leading to release of free iron, which catalyses oxidation of LDL. In addition, glycation of LDL may enhance the oxidation of LDL catalysed by iron. Both studies indicate that the glycation of proteins has a major impact on iron homeostasis leading to release of non-enzymatic glycation and contributing to one of the most common complications of Type 2 diabetes, atherosclerosis

Keywords

Oxidative stress, Glycated proteins, Type 2 diabetes, Iron

Document Type

Thesis

Publication Date

2015

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