HbA1c determination from HemaSpot™ blood collection devices: comparison of home prepared dried blood spots with standard venous blood analysis.

AIM: To assess the clinical performance and patient acceptance of HemaSpot™ blood collection devices as an alternative blood collection method. METHODS: Adult men and women with any type of diabetes, routinely carrying out self-monitoring of blood glucose were recruited (n = 128). Participants provided a venous blood sample and prepared two HemaSpot dried blood spots, one at clinics and one at home. HbA1c analysis was by Tosoh G8 high-performance liquid chromatography. Participants also completed a questionnaire. RESULTS: Strong linear relationships been HbA1c levels in dried blood spots and venous blood were observed and a linear model was fitted to the data. Time between dried blood spot preparation and testing did not impact the model. Participants were accepting of the approach: 69.2% would use this system if available and 60.7% would be more likely to use this system than going to their general practitioner. CONCLUSIONS: The combination of a robust desiccating dried blood spot device, home sample preparation and return by post produces HbA1c data that support the use of a time-independent linear calibration of dried blood spot to venous blood HbA1c . A robust remote sample collection service would be valuable to people living with diabetes in urban areas who are working or house-bound as well as those living in remote or rural locations.

Preliminary study of hypoxia-related cardiovascular mediator-markers in patients with end-stage renal disease with and without diabetes and the effects of haemodialysis.

BACKGROUND: Evidence points to activation of pro-inflammatory and pro-thrombotic stimuli during the haemodialysis process in end-stage renal disease (ESRD) with potential to predispose to cardiovascular events. Diabetes is associated with a higher incidence of cardiovascular disease in haemodialysis patients. We tested the hypothesis that a range of mediators and markers that modulate cardiovascular risk are elevated in haemodialysis patients with diabetes compared to those without. METHODS: Men and women with diabetes (n = 6) and without diabetes (n = 6) aged 18-90 years receiving haemodialysis were recruited. Blood samples were collected and analysed pre- and post-haemodialysis sessions for (platelet-monocyte conjugates (PMC), oxidised LDL (Ox-LDL), endothelin 1 (ET-1) and vascular endothelial growth factor (VEGF-A). RESULTS: PMC levels significantly increased after haemodialysis in both groups (diabetes p = 0.047; non-diabetes p = 0.005). Baseline VEGF-A was significantly higher in people with diabetes (p = 0.009) and post-dialysis levels were significantly reduced in both groups (P = 0.002). Ox-LDL and CRP concentrations were not significantly different between groups nor affected in either group post-dialysis. Similarly, ET-1 concentrations were comparable in all patients at baseline, with no change post-dialysis in either group. CONCLUSIONS: In this pilot study, we have confirmed that circulating PMCs are increased following dialysis irrespective of diabetes status. This is likely to be a mechanistic process and offers a potential explanation for high rates of vascular events associated with haemodialysis. The higher VEGF-A concentrations between patients with and without diabetes is a previously unreported finding in diabetic ESRD. Further research is merited to establish whether VEGF-A is a marker or mediator (or both) of cardiovascular risk in haemodialysis.

A randomized crossover study to assess the effect of an oat-rich diet on glycaemic control, plasma lipids and postprandial glycaemia, inflammation and oxidative stress in Type 2 diabetes.

AIMS: In the UK, lifestyle intervention is first-line management in Type 2 diabetes. It is unclear what type of diet is most efficacious for improving glycaemic control. This study investigated the effects of an oat-enriched diet on glycaemic control, postprandial glycaemia, inflammation and oxidative stress compared with standard dietary advice. METHODS: In a randomized crossover design, 27 volunteers with Type 2 diabetes, managed on diet and lifestyle only, were observed for two consecutive 8-week periods following either the oat-enriched diet or re-enforced standard dietary advice. Volunteers attended at baseline (habitual intake) and 8 and 16 weeks. Measurements included basic clinical measurements and fasted and postprandial (3-h) glucose and insulin in response to a healthy test meal. Markers of inflammation and oxidative stress, including high-sensitivity C-reactive protein, interleukin 6, interleukin 18, tumour necrosis factor-alpha, adiponectin, thiobarbituric acid reactive substances, oxygen radical antioxidant capacity, oxidized LDL and urinary isoprostanes, were also measured at fasting and in the postprandial period. RESULTS: There were no diet-related effects on glycaemic control or glycaemic or insulinaemic responses to the test meal. Total cholesterol (5.1 ± 1.0 vs. 4.9 ± 0.8 mmol/l, P = 0.019) concentrations declined following the oat-enriched diet compared with standard dietary advice. There was a postprandial decline in adiponectin concentration (P = 0.009), but no effect of dietary intervention. None of the measures of oxidative stress or inflammation were altered by the oat-enriched diet compared with standard dietary advice. CONCLUSION: The oat-enriched diet had a modest impact on lipid lowering, but did not impact on oxidative stress or inflammation in these volunteers with Type 2 diabetes.

N-Acetylcysteine inhibits platelet-monocyte conjugation in patients with type 2 diabetes with depleted intraplatelet glutathione: a randomised controlled trial.

AIMS/HYPOTHESIS: The aim of this study was to determine whether oral dosing with N-acetylcysteine (NAC) increases intraplatelet levels of the antioxidant, glutathione (GSH), and reduces platelet-monocyte conjugation in blood from patients with type 2 diabetes. METHODS: In this placebo-controlled randomised crossover study, the effect of oral NAC dosing on platelet-monocyte conjugation and intraplatelet GSH was investigated in patients with type 2 diabetes (eligibility criteria: men or post-menopausal women with well-controlled diabetes (HbA(1c) < 10%), not on aspirin or statins). Patients (n = 14; age range 43-79 years, HbA(1c) = 6.9 ± 0.9% [52.3 ± 10.3 mmol/mol]) visited the Highland Clinical Research Facility, Inverness, UK on day 0 and day 7 for each arm of the study. Blood was sampled before and 2 h after oral administration of placebo or NAC (1,200 mg) on day 0 and day 7. Patients received placebo or NAC capsules for once-daily dosing on the intervening days. The order of administration of NAC and placebo was allocated by a central office and all patients and research staff involved in the study were blinded to the allocation until after the study was complete and the data fully analysed. The primary outcome for the study was platelet-monocyte conjugation. RESULTS: Oral NAC reduced platelet-monocyte conjugation (from 53.1 ± 4.5% to 42.5 ± 3.9%) at 2 h after administration and the effect was maintained after 7 days of dosing. Intraplatelet GSH was raised in individuals with depleted GSH and there was a negative correlation between baseline intraplatelet GSH and platelet-monocyte conjugation. There were no adverse events. CONCLUSIONS/INTERPRETATION: The NAC-induced normalisation of intraplatelet GSH, coupled with a reduction in platelet-monocyte conjugation, suggests that NAC might help to reduce atherothrombotic risk in type 2 diabetes. FUNDING: Chief Scientist Office (CZB/4/622), Scottish Funding Council, Highlands & Islands Enterprise and European Regional Development Fund. TRIAL REGISTRATION: isrctn.org ISRCTN89304265.

Diabetic fatty liver disease is associated with specific changes in blood-borne markers.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) can lead to cirrhosis and hepatocellular carcinoma and is strongly associated with obesity and insulin resistance. The aim of this study was to assess if plasma markers associated with NAFLD are increased in people with concomitant diabetes compared with those without. METHODS: A total of 68 participants were recruited from diabetes and liver clinics. Fatty liver disease was indicated by routine blood tests and ultrasonography. Forty-seven participants had type 2 diabetes; of them, 18 had no fatty liver disease as defined previously (DNoFLD) and 29 had fatty liver disease (DFLD); the remaining 21 had fatty liver disease but no diabetes (NonDFLD). Serum samples were analyzed for adiponectin (APN), alanine and aspartate aminotransferases and plasma for cholesterol, triglyceride, hyaluronic acid (HA), procollagen peptide III, alkaline phosphatase and fibrinogen. RESULTS: Hyaluronic acid and procollagen peptide III were significantly higher and adiponectin significantly lower in DFLD than NonDFLD and DNoFLD, the difference being particularly marked for hyaluronic acid and APN. There was no difference in these markers between NonDFLD and DNoFLD and no association between any plasma or serum marker and ultrasound grade of steatosis. CONCLUSION: We have identified markers of hepatic steatosis that appear to be specific for people with type 2 diabetes. A further longitudinal study is merited to assess the role of these markers in understanding the progression of hepatic steatosis and fibrosis in people with and without diabetes.

Blood rheology and albumin excretion in diabetic pregnancy.

Blood rheology is altered in diabetes and also in non-diabetic pregnant women. The cumulative effect of hyperfiltration, abnormal rheology of pregnancy, and diabetes could be one mechanism contributing to increased intraglomerular pressure and albuminuria in diabetic pregnancy. We conducted a prospective study of 22 Type 1 (insulin-dependent) diabetic patients and 22 non-diabetic women to determine if there was an association of altered blood rheology on glomerular function in diabetic pregnancy. Albumin excretion showed no increment with increasing gestation and was similar in diabetic and non-diabetic women throughout pregnancy (first trimester: 5.0 (3.0-14.0) vs 5.8 (3.7-10.7) mg l-1, p = 0.89; second trimester: 6.0 (5.0-12.0) vs 5.1 (3.6-10.4) mg l-1, p = 0.25; third trimester: 7.5 (3.5-16.0) vs 4.9 (2.9-7.3) mg l-1, p = 0.18). Red cell aggregation index increased in both groups between the first and third trimesters (diabetic patients: mean difference 2.0; Cl: 1.0-2.9, p = 0.003, and control patients: mean difference 2.3, Cl: 1.0-3.5, p = 0.002). Fibrinogen levels were significantly higher between the third and first trimesters in diabetic patients (mean difference 0.7, Cl: 0.2-1.3 g l-1, p = 0.006). Pregnancy, therefore, was associated with increased red cell aggregation, related in part to increased fibrinogen levels but the extent of change was similar in diabetic and nondiabetic women and appeared to have no adverse effect on glomerular function in pregnant insulin-dependent diabetic women.

A comparison of different methods of assessing free radical activity in type 2 diabetes and peripheral vascular disease.

Increased free radical activity in diabetes mellitus may contribute to the higher prevalence and mortality from macrovascular disease in diabetic patients. To investigate this, levels of plasma antioxidants (superoxide dismutase, caeruloplasmin, plasma, and lysate thiol), diene conjugates, lipid peroxides, and chemiluminescence were measured in diabetic and non-diabetic patients with peripheral vascular disease compared with healthy control subjects. Caeruloplasmin, diene conjugate ratio, and lipid peroxides were significantly increased in patients with vascular disease but there was no difference between diabetic and non-diabetic patients. Conjugated diene ratio correlated with caeruloplasmin (r = 0.40, p < 0.02) and inversely with superoxide dismutase level (r = 0.36, p < 0.05) but there was no significant correlation between other antioxidants and diene conjugates, lipid peroxides or chemiluminescence. The relationship between different indirect measurements of free radical activity is variable but there appears to be no additive effect of diabetes on the increased free radical activity associated with vascular disease.

Increased red cell aggregation in diabetes mellitus: association with cardiovascular risk factors.

Red cell aggregation may be higher in diabetic patients and may predispose to cardiovascular disease. Red cell aggregation was measured by a simple photometric method in 122 diabetic patients and 100 matched control subjects, to determine its relationship to cardiovascular risk factors. Red cell aggregation was significantly increased in both Type 1 (4.3 +/- 1.3 vs 3.4 +/- 1.2, p < 0.002) and Type 2 diabetic patients (5.5 +/- 1.5 vs 3.2 +/- 1.3, p < 0.0001). In all diabetic patients aggregation correlated with triglycerides, VLDL, and inversely with HDL and in Type 2 diabetic patients also with body mass index, hypertension, and inversely with duration of diabetes. On multiple regression analysis, triglycerides and body mass index showed an independent association with red cell aggregation and in Type 2 diabetic patients smoking was also associated with increased red cell aggregation. It is concluded that increased red cell aggregation may be one mechanism by which some cardiovascular risk factors could promote cardiovascular disease in diabetes.

Sarcoidosis: association with small bowel disease and folate deficiency.

A 30 year old woman with recurrent anaemia due to folate deficiency had evidence of sarcoid granuloma on small bowel biopsy but was presumed to have Crohn's disease. The diagnosis of small bowel sarcoidosis was not seriously considered until she developed systemic manifestations of sarcoidosis (cutaneous and pulmonary lesions) over the following 20 years. Sarcoidosis of the gastrointestinal tract, particularly the small bowel, is rare and this case is unusual because bowel pathology preceded more generalised lesions. As far as is known it is also the first case to be described presenting with malabsorption of folic acid.

Seasonal and climatic variation in cholesterol and vitamin C: effect of vitamin C supplementation.

Vitamin C (ascorbic acid) is an important anti-oxidant which may help to reduce free radical damage and atheroma formation in blood vessels. In a study in which a group of healthy volunteer subjects were followed up for 12 months and a group of patients with vascular disease taking Vitamin C supplements were followed for 23 months, we confirmed previous findings of seasonal variations in ascorbic acid and cholesterol and have shown an inverse relationship between leucocyte ascorbic acid and serum cholesterol levels. In healthy control subjects the increase in ascorbate and fall in cholesterol during the summer months was reversed when the weather changed to a more winter pattern, presumably due to dietary alterations. We found that ascorbic acid levels were lower in patients with peripheral vascular disease and that although normal ascorbic acid levels were achieved with Vitamin C supplementation, when supplements were stopped at the height of a normal summer, there was a fall in ascorbic acid and a rise in serum cholesterol to winter levels. Given these findings we suggest that patients with vascular disease should have Vitamin C supplements throughout the year.

Serum fructosamine/haemoglobin A1 ratio predicts the future changes in haemoglobin A1 in type 2 (non-insulin dependent) diabetic patients.

The aim of the study was to define clinical interpretation of the parallel measurements of serum fructosamine and HbA1 in diabetic patients. We studied 14 type 2 diabetic patients over a 16-wk period. The cross-sectional analysis showed no correlation between serum fructosamine and HbA1 concentrations during the period of changing glycaemic control. The correlations, however, became significant (P less than 0.05) at 12 (r = 0.60) and 16 (r = 0.87) weeks, i.e. after glycaemia had stabilised. Longitudinal analysis of individual patients' data over the 16-wk period showed a significant correlation between serum fructosamine and HbA1 (r = 0.55 to r = 0.94) which was present in 8 out of 14 patients. The changes in fructosamine concentration preceded those observed in HbA1. The ratio of fructosamine/HbA1 predicted the changes in HbA1 over the following month (r = 0.54, P less than 0.001). Thus, we demonstrated that the parallel measurement of fructosamine and HbA1 provides information on future trends in HbA1 concentration in diabetic patients.

Do rheological variables play a role in diabetic peripheral neuropathy?

Whole blood viscosity and its determinants were measured in diabetic patients with and without peripheral neuropathy to assess whether these variables could have a role in the microvascular aetiology of diabetic peripheral neuropathy. Although corrected whole blood viscosity at high and low shear rates (5.29 +/- 0.51 and 21.10 +/- 3.03 mPa s), plasma viscosity (1.41 +/- 0.13 mPa s), and red cell filtration ratio (0.49 +/- 0.04) in diabetic patients were significantly different from non-diabetic control subjects (high shear rate 4.83 +/- 0.54, low shear rate 17.36 +/- 2.78, plasma 1.29 +/- 0.09 mPa s, all p less than 0.001, and red cell filtration ratio 0.55 +/- 0.03, p less than 0.001) there were no significant differences between diabetic patients with neuropathy and those without. Blood rheology is altered to a similar extent in diabetic patients with and without neuropathy.

The relationship between colonisation, secretor status and in-vitro adhesion of Candida albicans to buccal epithelial cells from diabetics.

This study investigated whether oral candida infection in diabetics and adhesion of Candida albicans to buccal epithelial cells in vitro were related. Buccal cells from 50 patients with diabetes mellitus showed a significant increase in adhesion of C. albicans strain CDS 88 compared with those collected from 50 non-diabetic controls matched for age, sex and denture status. Oral candida carriage, candida infection and secretor status were also investigated in both groups. The frequency of carriage was increased, but not significantly, and there was a significantly higher incidence of candida infection in diabetic patients compared with controls. Diabetic patients who were non-secretors had a significantly increased frequency of oral candida carriage.

Evaluation of red cell deformability by a filtration method in type 1 and type 2 diabetes mellitus with and without vascular complications.

Previous studies of red cell deformability (RCD) in diabetic patients have produced conflicting results. We have therefore reassessed this problem with the Carri-Med Filtrometer, which measures RCD independently of the rate of clogging of the filter. The initial red cell filtration ratio relative to buffer (mean +/- SD) of 69 diabetic patients (28 type 1 and 41 type 2) was significantly impaired in both type 1 (0.470 +/- 0.047) and type 2 diabetic patients (0.488 +/- 0.065), when compared to 66 non diabetic control subjects (0.540 +/- 0.032; both p less than 0.001). A significant difference in RCD was noted between type 1 and type 2 diabetics (p less than 0.03), but no association with vascular complications was found. No significant correlations were noted between RCD and the duration of diabetes, age, HbA1, leucocyte count, or mean red cell volume. However RCD was inversely related to mean red cell haemoglobin concentration (r = -0.43, p less than 0.01). Diabetics had significantly lower mean red cell volume and significantly higher mean cell haemoglobin concentration than non-diabetics, but these changes could not explain the difference in RCD, which may be related to alterations in the red cell membrane.

Changes in phagocytic function with glycaemic control in diabetic patients.

Phagocytic function was assessed by serial whole blood chemiluminescence in poorly controlled type 2 (non-insulin dependent) diabetic patients during efforts to improve glycaemic control and compared with a group of well controlled type 1 (insulin dependent) diabetic patients. Chemiluminescence (corrected to a standard polymorphonuclear count) remained below normal (0.15-0.30 photons/second/cell) for most of the type 2 patients until 12 weeks when the value was significantly increased in patients showing improved glycaemic control (mean (range) 0.25 (0.01-0.43) photons/second/cell) compared with those showing no improvement (0.12(0.01-0.31) photons/second/cell). There was a significant inverse correlation of delta HbA1 with delta chemiluminescence. Although mean chemiluminescence for the type 1 diabetic patients was within the normal range, there was a wide scatter of values (0.19 (0.04-0.43) photons/second/cell) and there was no significant difference compared with the final value of type 2 patients with improved control. Glycaemic control is therefore a major determinant of phagocytic function in diabetic patients, but other factors must contribute, particularly in type 1 (insulin dependent) patients.

Association of hypertension with blood viscosity in diabetes.

Plasma and whole blood viscosity and its determinants were measured in 86 diabetic patients (29 hypertensive and 57 normotensive) and compared with 52 non-diabetic control subjects to assess whether hypertension has an additive and adverse effect on blood viscosity. Whole blood viscosity (corrected for haematocrit), at high and low shear rates (95 and 0.95 s-1), was significantly higher in both Type 1 (5.1 +/- 0.5 (+/- SD), 19.8 +/- 2.9) and Type 2 (5.2 +/- 0.3, 21.1 +/- 2.0) diabetic patients compared with control subjects (4.9 +/- 0.6, 17.4 +/- 2.6 mPa s, p less than 0.01). Corrected whole blood viscosity at high shear rate was significantly higher in hypertensive than in normotensive Type 2 diabetic patients (5.5 +/- 0.4 vs 5.2 +/- 0.3 mPa s, p less than 0.01). Plasma viscosity was significantly higher in diabetic patients compared with control subjects (1.4 +/- 0.1 vs 1.3 +/- 0.1 mPa s, p less than 0.01), but there was no difference between hypertensive and normotensive diabetic patients (1.4 +/- 0.1 vs 1.4 +/- 0.2 mPa s). Fibrinogen levels were similar in all the groups.

Long-term performance of the fructosamine assay.

We have investigated the long-term performance of the fructosamine assay based on secondary glycated protein standards and attempted to define the interpretation of varying degrees of increase in fructosamine concentration in comparison to haemoglobin A1 (HbA1) values both in insulin dependent (IDDM) and non-insulin dependent (NIDDM) diabetic patients. Between-batch imprecision of fructosamine over 5 months was (CV) 2.5% at 2.09 mmol/L, 2.8% at 3.52 mmol/L and 3.6% at 4.14 mmol/L. Variation of fructosamine concentration in vivo in stable diabetic patients monitored over 8-18 weeks was 2.3% to 7.1%. Fructosamine correlated with HbA1 both in IDDM (n = 110, r = 0.701, P less than 0.001) and NIDDM (n = 71, r = 0.764, P less than 0.001). Specificity and sensitivity of fructosamine for the prediction of degree of control assessed on the basis of HbA1 level (cut-off point for good vs. poor control, HbA1 = 10%) was determined. In NIDDM, specificity above 90% was achieved at a fructosamine concentration of 3.4 mmol/L with a corresponding sensitivity of 64.1%. 22.5% of patients were classified differently on the basis of fructosamine as compared to HbA1. In IDDM, specificity over 90% was achieved at 3.8% mmol/L fructosamine with a sensitivity of 35%. Discordancy rate between HbA1 and fructosamine based assessment of control was 31.8%. The assessment of diabetic control based on fructosamine may be different from that based on HbA1, particularly in IDDM. Fructosamine and HbA1 should be used as complementary rather than alternative tests.

Management of diabetes resistant to subcutaneous insulin with intravenous insulin via an implanted infusion pump.

Diabetes resistant to conventional subcutaneous insulin injection is a rare complication of insulin-dependent diabetes which poses a major management problem. We report three cases treated for a total of over seven patient years with fully implanted insulin infusion devices. Technical difficulties with the devices and their operation have been substantial but the patients are much improved and hospitalisation has been dramatically reduced. We suggest that implanted insulin pumps are a real treatment option for patients with this unusual syndrome.