Brief technical note

A method for specifically measuring haemoglobin A_IC with a disposable commercial ion-exchange column

Diabetes mellitus is a vascular disease with many microvascular manifestations as retinopathy and macrovascular complications as coronary artery disease. A large number of diabetics with retinopathy may have unidentified coronary artery disease.

Evaluate whether diabetic retinopathy is a valid predictor of coronary artery disease.

Cross section study.

The study included 60 male patients chosen randomly from diabetic and ophthalmologic outpatient clinics. Twenty patients had type 2 diabetes mellitus with nonproliferative retinopathy, 20 patients with proliferative diabetic retinopathy and 20 patients with type 2 diabetes mellitus without retinopathy. All studied cases were not previously identified as coronary artery disease. Clinical examination stressed on conventional cardiovascular risk factors; duration, degree of control of hyperglycemia and erectile failure. Lipogram, urinary albumin excretion, resting, exercise ECG and echocardiography were carried out. All patients were subjected to fundus examination for evaluation of diabetic retinopathy and duplex ultrasound to assess carotid intimal medial thickness. Coronary angiography was undertaken for all cases.

There was no significant difference regarding age, BMI and blood pressure between the groups. However, duration of diabetes, erectile dysfunction and neuropathy revealed significant difference between different groups particularly the proliferative in relation to the nonproliferative diabetic group. Hypertriglyceridimia, urinary albumin excretion and serum cholesterol differ significantly between the studied groups. The ECG, echocardiography showed insignificant differences. Carotid intima media thickness showed an insignificant difference when comparing the three studied groups, however diabetic retinopathy group with coronary artery stenosis revealed a significant increase in carotid media thickness. Evidence of coronary artery stenosis was present in 80% of the proliferative diabetic retinopathy group and in 70% of the nonproliferative diabetic retinopathy group but was not present in nonretinopathic diabetic group. The stenotic lesions were involving one or two vessels. No one had left main disease.

Diabetic retinopathy is a good predictor of coronary artery disease that exceeds the conventional risk factors. Diabetics with retinopathy would benefit from early coronary angiography and diabetic retinocoronary clinics are warranted.

The development and progression of diabetic microvascular complications including nephropathy are related to the degree of glycemic control and oxidative stress and may be influenced by genetic factors. The aim of the present study was to investigate the association between haptoglobin (Hp) gene polymorphism and the occurrence of diabetic nephropathy in patients with type 2 diabetes mellitus and to find a possible link between Hp phenotypes and the inflammatory parameters; serum C-reactive protein (CRP), interleukin- 6 (IL-6), and Hp.

The study included 60 normotensive type 2 diabetic patients (> 5 years duration) categorized into three equal groups (normo-, micro-, and macroalbuminuric), according to urinary albumin excretion (UAE). In addition, 20 age- and sex-matched individuals were selected to serve as a control group. Serum CRP, IL-6, and Hp concentrations were measured and Hp phenotyping was conducted using polyacrylamide gel electrophoresis.

The frequency of Hp phenotype 1-1 (Hp 1-1) in diabetic patients with normoalbuminuria was 7/20 (35%) as compared with 1/20 (5%) in diabetics with macroalbuminuria (p = 0.02). However, the frequency of Hp 2-2 was greater in diabetics with macroalbuminuria (12/20, 60%) than in those with normoalbuminuria or controls (5/20, 25%; p = 0.03). Patients with diabetic nephropathy (micro- or macroalbuminuria) had higher levels of serum CRP, IL-6, and Hp than those without nephropathy (normoalbuminuria). Serum Hp levels in type 2 diabetics were higher in Hp phenotype 2-2 than in Hp 1-1; however, serum CRP and IL-6 levels did not differ significantly between Hp phenotype groups. Moreover, there were significant positive correlations between UAE and serum levels of CRP, IL-6, and Hp in diabetic patients.

Hp phenotype 2-2 is considered to be a major susceptibility gene for the development of nephropathy in type 2 diabetic patients. In addition, the significant association between inflammatory parameters and UAE indicates that inflammation may be a pathogenic mechanism of renal injury in type 2 diabetics. Moreover, serum IL-6 and Hp may be good prognostic factors for the development of nephropathy in the course of diabetes mellitus. Future research on the use of anti-inflammatory therapy may result in a new approach to the treatment and prevention of diabetic nephropathy.

This chapter discusses the steps involved in nonenzymatic glycosylation of proteins. Nonenzymatic glycosylation takes place in all proteins with a free-reacting lysine or valine in the presence of glucose. The formation of glycosylated plasma albumin, hemoglobin, and skin collagen provides a diagnostic index of short- to long-term time–concentration of glucose in vivo. A wide range of assay methods are available, with affinity chromatographic, isoelectric focusing, and spectrophotometric methods providing the best accuracy and versatility. Glycosylated hemoglobin assays indicate glucose pressure and are of diagnostic value in general diabetic control, while glycosylated plasma albumin determinations are preferable in acute episodes in the life of a diabetic, such as pregnancy, infection, stress, trauma, surgery. Glycosylated collagen estimations reflect tissue aging and are relevant in healing processes. Glycosylation alters the biologic activity of proteins, and these may relate to the manifold complications concomitant on the lifelong elevation of blood and tissue glucose in the diabetic.

The major utility of the glycosylated hemoglobin assay continues to be as a monitor for the effectiveness of diet/insulin or oral agent therapy in the normalization of serum glucose concentrations. During periods of poor glycemic control, as assessed by frequent positive urine glucose determinations, or by elevated fasting or postprandial blood glucose concentrations, or by episodes of ketoacidosis, the use of the glycosylated hemoglobin assay adds little information. Rather, it is most useful during periods of stable control, to assess the effectiveness of the combined therapy of diet, activity, and insulin or oral agents. As the glycosylated hemoglobin concentrations tend to reflect the metabolic control during the previous four to eight weeks, little is to be gained from more frequent use of this assay. An isolated elevated serum glucose concentration obtained in the office or hospital laboratory, associated with a near-normal serum glycosylated hemoglobin concentration need not demand drastic rearrangement of the patient's therapuetic regimen. The normal hemoglobin A_1c concentration reflects good blood glucose control over the past one to two months. The elevated blood glucose concentration may reflect a transient aberration secondary to stress, diet, or other factors. If, however, actual deterioration in the patient's metabolic control is occurring, this may not be reflected in the hemoglobin A_1c concentration for several weeks or months. Likewise, during a period of stable control, a normal office or lab glucose concentration associated with an elevated glycosylated hemoglobin concentration may indicate poor control, the normal glucose resulting from the peak effectiveness of either short- or intermediate-acting insulin. In this situation, more frequent glucose determinations would reveal elevation of blood glucose concentrations.

Whereas the glycosylated hemoglobin assay may help the physician and patient to assess glycemic control, this alone cannot lead to improved control. In this regard the use of home glucose monitoring with or without insulin infusion pumps has led to near-normalization of blood glucose concentrations in Type I and in pregnant diabetic patients [105, 175, 176]. The frequent determination of blood glucose concentrations on a daily basis can allow for optimum use of dietary and insulin therapy. Moreover, the reported variability in glycosylated hemoglobin after periods of poor control may reflect the time relation between blood glucose and hemoglobin A_1c concentrations, or the stability of the glycosylated hemoglobin, and may diminish the clinical utility of the glycosylated hemoglobin concentration in the individual patient. The clinical utility of the glycosylated albumin or protein assay in diabetes management remains to be established [211]. At this time the combined use of multiple daily home blood glucose determinations, coupled with several well-spaced hemoglobin A₁ determinations may be the most sensitive indicators of the state of metabolic control.

Thus, protein glycosylation provides an indication of glycemic regulation. Protein glycosylation may also contribute to the complications of diabetes mellitus, but such relationships, for the most part, remain to be established.