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PubMed1986

The alkaline reducing activity of glycated serum proteins and its relevance to diabetes mellitus.

We investigated the ability of the fructosamine assay to detect glycation of serum proteins. We incubated both whole human serum and serum protein fractions in vitro with [14C]glucose, and analyzed for reducing activity and for uptake of 14C by protein. In all experiments, the reducing activity increased linearly with time for seven days and was correlated with 14C uptake (r = 0.94-0.98). Protein ketoamines were about fivefold more actively reducing than equimolar concentrations of deoxymorpholinofructose, the fructosamine standard, which explains why values for fructosamine in serum are higher than the expected concentration of protein ketoamines. We also used [14C, 2-3H]glucose to assess the contribution of the aldimine component to 14C uptake. Whole human serum and albumin incubated with [14C, 2-3H]glucose showed little uptake of 3H in relation to 14C. We conclude that glycated protein can be simply and reliably quantified by the fructosamine assay, and we discuss the relevance of this conclusion to the monitoring of diabetes.

Blood Proteins↗
PubMed1998

Long-term glycaemic control directly correlates with glomerular filtration rate in early Type 1 diabetes mellitus before the onset of microalbuminuria.

Hyperfiltration occurs early in diabetes mellitus and has been implicated in the development of microalbuminuria. Our aim was to re-examine the controversial relationship between glycaemic control and glomerular filtration (GFR) in normoalbuminuric, normotensive, non-obese patients with short duration Type 1 diabetes mellitus (DM). We studied 75 Type 1 DM patients, 35 male, aged 18-42 years, with a duration of diabetes of 4-8 years. GFR was determined by inulin clearance; hyperfiltration was defined as above 145 ml min(-1) 1.73 m(-2) (equivalent to 2 SD above mean for a control population). Analysis was by paired Student's t-testing and linear regression. GFR correlated significantly with HbA1c (r= 0.47, p < 0.0001) and fructosamine (r= 0.24, p = 0.035). Mean HbA1c and fructosamine in the 13 patients with hyperfiltration was significantly higher than in the rest of the group (HbA1c: 9.2% (95% C.I. 7.9-10.4%) vs 7.6 % (7.2-7.9), p= 0.002; fructosamine: 479 micromol l(-1) (450-507) vs 410 micromol l(-1) (388-432), p = 0.009. This significant difference persisted even when the two highest values of HbA1c or fructosamine were removed from analysis. Effective renal plasma flow, assessed by PAH clearance, also correlated in all patients with HbA1c (r=0.31, p=0.039). We conclude that poor glycaemic control directly correlates with hyperfiltration and renal hyperperfusion in early Type 1 DM.

Adult↗
PubMed2004

Circulating 1,5-anhydroglucitol levels in adult patients with diabetes reflect longitudinal changes of glycemia: a U.S. trial of the GlycoMark assay.

OBJECTIVE: 1,5-Anhydroglucitol (1,5AG) is a major circulating polyol arising primarily from ingestion and excreted competitively with glucose. Japanese studies have demonstrated reduced concentrations of 1,5AG in serum in hyperglycemic patients in comparison with euglycemic subjects and a gradual normalization of 1,5AG values for patients responding to antihyperglycemic therapies. In this first U.S. study, we assessed the ability of 1,5AG measurements to monitor glycemic control in a cohort of 77 patients with diabetes (22 with type 1 diabetes, 55 with type 2 diabetes) who presented with suboptimal glycemic control at baseline (defined as HbA(1c) >or=7%). RESEARCH DESIGN AND METHODS: Each patient received therapies consisting of combinations of diabetes education, nutritional counseling, and addition or dose adjustment of various insulins or oral antihyperglycemic medications. Therapy was targeted to reduce mean HbA(1c) by >or=1.0% over the monitoring period. 1,5AG, HbA(1c), fructosamine, and random glucose measurements were performed at baseline and at 2, 4, and 8 weeks after the initiation of therapy. RESULTS: 1,5AG, fructosamine, and glucose values progressed significantly toward euglycemia by week 2 of monitoring (Wilcoxon's signed-rank test, P < 0.05), with median changes of 93, -7, and -13% for 1,5AG, fructosamine, and glucose, respectively. In contrast, HbA(1c) values did not respond significantly to therapy until week 4. On an individual patient basis, 89.6% of patients displayed longitudinal changes of 1,5AG from baseline to week 8 in concordance with HbA(1c). 1,5AG was also highly correlated with HbA(1c) and fructosamine (Spearman rho = -0.6459 and -0.6751, respectively; both P < 0.0001). CONCLUSIONS: We conclude that 1,5AG responds sensitively and rapidly to changes in glycemia and monitors glycemic control in accordance with established markers.

Adult↗
PubMed2000

Reliability of history and physical examination findings for assessing control of glycemia in dogs with diabetes mellitus: 53 cases (1995-1998).

OBJECTIVE: To evaluate the reliability of history and physical examination findings for assessing control of glycemia in insulin-treated diabetic dogs. DESIGN: Retrospective study. ANIMALS: 53 insulin-treated dogs with diabetes mellitus. PROCEDURE: Medical records of insulin-treated diabetic dogs from June 1995 to June 1998 were reviewed, and information on owner perception of their dog's response to insulin treatment, physical examination findings, body weight, insulin dosage, and concentrations of food-withheld (i.e., fasting) blood glucose (FBG), mean blood glucose (MBG) during an 8-hour period, blood glycosylated hemoglobin (GHb), and serum fructosamine was obtained. Owner's perception of their dog's response to insulin treatment, physical examination findings, and changes in body weight were used to classify control of glycemia as good or poor for each dog. The FBG, MBG/8 h, blood GHb, and serum fructosamine concentrations were compared between well-controlled and poorly controlled insulin-treated diabetic dogs. RESULTS: Presence or absence of polyuria, polydipsia, polyphagia, lethargy, and weakness were most helpful in classifying control of glycemia. Mean FBG and MBG/8 h concentrations, blood GHb concentrations, and serum fructosamine concentrations were significantly decreased in 25 well-controlled diabetic dogs, compared with 28 poorly controlled diabetic dogs. Most well-controlled diabetic dogs had concentrations of FBG between 100 and 300 mg/dl, MBG/8 h < or = 250 mg/dl, blood GHb < or = 7.5%, and serum fructosamine < or = 525 mumol/L, whereas most poorly controlled diabetic dogs had results that were greater than these values. CONCLUSIONS AND CLINICAL RELEVANCE: Reliance on history, physical examination findings, and changes in body weight are effective for initially assessing control of glycemia in insulin-treated diabetic dogs.

Animals↗
PubMed1996

Physical activity and glycaemic control in children with diabetes mellitus.

The relationship between physical activity, glycaemic control, and skinfold thickness was studied in children with diabetes. Patients recorded details of activity in a home diary over a 6-day period, and provided eight serial capillary dried blood spots for glucose analysis during a 24-h period. Levels of activity were assessed using a semi-quantitative scoring scheme. No correlation was found between total activity score and glycaemic control. However, the activity score before 9 am showed significant correlation with mean blood glucose (p = 0.005) and fructosamine (p = 0.04). The time of rising in the morning showed significant correlation with blood glucose after lunch (p = 0.004) and with fructosamine (p = 0.04). A significant correlation was found between early morning activity and subscapular skinfold thickness. The association between activity and glycaemic control warrants further investigation, as it suggests that patients who engage in energetic activity early in the morning may achieve lower blood glucose and fructosamine levels than their less active peers.

Adolescent↗
PubMedDate not supplied

Scanning force microscopy reveals structural alterations in diabetic rat collagen fibrils: role of protein glycation.

BACKGROUND: The main functional property of collagen is to provide a supporting framework to almost all tissues: the effects of non-enzymatic glycation on this protein are deleterious and in diabetes mellitus contribute to the mechanism of late complications. The aim of this work is to provide evidence by scanning force microscopy of modifications in collagen structure caused by high glucose concentration, in vivo and in vitro, and to correlate the data with markers of non-enzymatic glycation. METHODS: Tendon fibrils were obtained from the tails of 8-month-old rats (BB/WOR/MOL¿BB) which developed diabetes spontaneously at least 12 weeks before they were killed, and from diabetes-resistant rats of the same strain (BB/WOR/MOL¿WB). A scanning force microscope (SFM; Nanoscope III) equipped with a Contact Mode Head was used for imaging. Band interval, diameter and depth of D-band gap were measured in non-diabetic and diabetic tail tendon fibrils and in fibrils incubated with glucose (0.5 M for 2 weeks). Fructosamine was determined in the tendon fibrils by a colorimetric method and pentosidine was evaluated in acid-hydrolyzed samples by coupled reverse phase-ionic exchange column HPLC. RESULTS: Incubated fibrils revealed modifications in radius (228+/-5 nm) and gap depth (3.65+/-0.10 nm) that closely reproduce diabetes-induced damage (236+/-3 and 3.20+/-0.04 nm respectively) and were significantly different from the pattern seen in non-diabetic fibrils (151+/-1 and 2.06+/-0.03 nm; p<0.001). Both fructosamine and pentosidine were higher in diabetic (3.82+/-1.43 nmol/mg and 2.23+/-0.24 pmol/mg collagen respectively) and in glucose-incubated fibrils (9.27+/-0.55 nmol/mg and 5.15+/-0.12 pmol/mg collagen respectively) vs non-diabetic tendons (1.29+/-0.08 nmol/mg and 0.88+/-0.11 pmol/mg collagen respectively; p<0.01); during the time course of incubation, an early increase in fructosamine was seen, whereas pentosidine increased later. The D-band parameter was similar in all three groups, indicating that axial organization is not modified by non-enzymatic glycation. CONCLUSION: This is the first description obtained with SFM of diabetes-induced ultrastructural changes in collagen fibrils. Moreover, the data presented are consistent with the concept that chronic exposure of collagen to glucose in vivo or in vitro leads to similar structural modifications in collagen fibrils, probably through crosslinks. The correlation between morphologic parameters and both markers of glycation provides strong evidence for a crucial role of this non-enzymatic modification.

Animals↗
PubMed1992

Insulin resistance and breast-cancer risk.

Life-style has a major influence on the incidence of breast cancer. To evaluate the effects of life-style related metabolic-endocrine factors on breast cancer risk we conducted a case-control study comparing 223 women aged 38 to 75 years presenting with operable (stage I or II) breast cancer and 441 women of the same age having no breast cancer, who participated in a population-based breast cancer screening program. Women reporting diabetes mellitus were excluded. Sera from 110 women of the same age group presenting with early stage melanoma, lymphoma or cervical cancer were used as a second 'other-cancer control group'. Serum levels of C-peptide were significantly higher in early breast cancer cases compared to controls. The same was found for the ratios C-peptide to glucose or C-peptide to fructosamine, indicating insulin resistance. Sex hormone binding globulin was inversely, triglycerides and available estradiol were positively related to C-peptide. Serum C-peptide levels were related to body mass index (BMI), and to waist/hip ratio (WHR), in particular in controls. However, the relative increase of C-peptide, C-peptide to glucose or C-peptide to fructosamine in cases was independent of BMI or WHR. The log relative risk was linearly related to the log C-peptide levels. Relative risk according to quintiles, and adjusted for age, family history, BMI and WHR, for women at the 80% level was 2.9 as compared with those at the 20% level for C-peptide. Elevated C-peptide or C-peptide to fructosamine values were not observed in the sera from women belonging to the 'other-cancer control group'. This study suggests that hyperinsulinemia with insulin resistance is a significant risk factor for breast cancer independent of general adiposity or body fat distribution.

Adult↗
PubMed1994

Effect of glycemic control on vitamin B12 metabolism in diabetes mellitus.

To determine the effect of glycemic control on vitamin B12 (B12) metabolism in diabetes mellitus, we studied B12 metabolism in 19 diabetic patients with poor glycemic control and 15 normal individuals. The diabetic patients had significantly higher total B12 binding capacity (3303 +/- 963 pg/ml), higher serum B12 levels (1173 +/- 503 pg/ml) and unsaturated B12 binding capacity (2131 +/- 902 pg/ml) when compared with the normal controls, but there was no difference in R-binder levels and the B12 binding ratio between the two groups. During a 2-week admission to establish glycemic control, the fructosamine levels in the diabetic patients decreased from 556 to 428 mumol/l and the total B12 binding capacity as well as unsaturated B12 binding capacity were significantly improved to the normal range (P < 0.01), but serum B12 levels, R-binder levels and the B12 binding ratio were not changed. There was a significant association between serum fructosamine levels and the total B12 binding capacity in poorly controlled diabetic patients and the decrease of fructosamine was correlated significantly with the change of total B12 binding capacity and serum B12 levels in diabetic patients. These results indicate the effects of glycemic control on B12 metabolism in diabetes mellitus.

Adult↗
PubMed1989

Early post-natal growth of large-for-dates babies of non-diabetic mothers is influenced by maternal glucose metabolism.

Growth of weight, length, head circumference and skinfold thickness (subscapular and triceps) from birth to 6 months in 53 large-for-dates (LFD) Chinese babies weighing greater than 4.0 kg at term and born to non-diabetic mothers was investigated and correlated with biochemical indices of maternal glucose tolerance at birth: glycosylated haemoglobin (HbA1), serum corrected fructosamine and the area under the oral glucose (50 g) tolerance (OGTT) curve. Growth in all physical dimensions, especially weight, showed a downward shift towards a reference mean. These changes in relative size were caused by slower growth velocities. None of the mothers had abnormally high concentrations of HbA1 or fructosamine nor an abnormal OGTT. However, weight velocities did show small but significant correlations with fructosamine (r = -0.42), and OGTT area units (r = 0.39) but not with HbA1. For some macrosomic babies born to apparently normal mothers, birth is seen to interrupt a process operating in prenatal life that accelerates growth. Covert abnormalities of maternal glucose homeostasis could explain this. Abnormal glucose tolerance during pregnancy might therefore be viewed as a continuum extending from (i) its maximum expression, the frankly diabetic state, through (ii) gestational diabetes to (iii) the mother who has no biochemically evident abnormality of glucose homeostasis but who has sufficient alteration to modify fetal growth. Post-natal growth of LFD babies is additional information which, when taken along with other markers of maternal glucose tolerance, might help to identify the mother at later perinatal risk.

Birth Weight↗
PubMed1995

Hyperglycemia and non-enzymatic glycation of serum and tissue proteins in chickens.

The objectives of these studies were to determine whether elevated plasma glucose concentrations in broiler breeder chickens (200-250 mg/dl) can result in the non-enzymatic attachment of glucose to serum proteins (fructosamine) and eventual cross-linking of tissue proteins (basement membrane thickness), and to investigate the effects of a factor that may influence this cross-linking process. In response to feeding the satiety factor calcium propionate (CaP, 1.7%), plasma glucose and fructosamine concentrations were increased (P < 0.05) from 1 to 9 weeks of age, whereas concentrations of plasma glucose and fructosamine in feed-restricted chicks were reduced for the first 7 weeks after hatch. In a second study, the age-related increase in kidney capillary basement membrane thickness was prevented (P < 0.05) by feeding the cross-linking inhibitor aminoguanidine (AG, 800 ppm) to 30-week-old broiler breeder hens for 34 weeks. The results from these studies suggest that concentrations of plasma glucose in chickens may, in fact, be exerting long-term detrimental effects on tissue proteins, which can be ameliorated by factors that limit the cross-linking reaction.

Administration, Oral↗
PubMed1996

Effect of acarbose on blood glucose profiles and plasma 1,5-anhydro-d-glucitol in type 2 diabetes poorly controlled by sulfonylurea therapy.

Sixteen patients with type 2 diabetes poorly controlled by glibenclamide (7.5-10.0 mg/day) were treated with acarbose (100 mg tds) for one week and the effect on the blood glucose profile, 24-hour urinary glucose excretion, plasma fructosamine, and plasma 1,5-anhydro-D-glucitol (1,5-AG) level was determined. The blood glucose profile was more stable and levels were lower during acarbose administration. In some patients, this improvement was maintained after discontinuing acarbose. The M-value, an indicator of blood glucose fluctuations, decreased significantly from 33.2 +/- 3.0 (mean +/- SEM) in the run-in period to 13.4 +/- 2.4 during acarbose therapy (P < 0.001), and rose again to 26.5 +/- 4.4 (P < 0.001) in the follow-up period. The 24-hour urinary glucose excretion and plasma fructosamine decreased similarly (P < 0.001 and P < 0.01, respectively) during and after acarbose therapy. Plasma 1,5-AG levels did not change significantly during acarbose therapy, but increased markedly afterwards (from 19.3 +/- 3.1 mumol 1(-1) to 25.0) +/- 3.1 mumol l-1, P < 0.001). Plasma 1,5-AG levels were significantly correlated with urinary glucose excretion one week earlier (r = 0.513, P < 0.006). These findings suggest that acarbose may improve glycemic control in type 2 diabetic patients poorly controlled by sulfonylurea therapy and that plasma 1,5-AG might be used as a marker of glycemic control cooperating with other markers such as fructosamine and urinary glucose determination for monitoring the short-term response to antidiabetic therapy.

Acarbose↗
PubMed2004

Non-enzymatic glycosylation of immunoglobulins in diabetic nephropathy.

BACKGROUND: Diabetic nephropathy is a relatively common microvascular complication in people suffering from diabetic mellitus. Chronic hyperglycemia leads to the accumulation of advanced glycosylation end products (AGEs) that covalently trap extravasated serum proteins such as immunoglobulins, albumin, and LDL through glucose derived cross-linking to the extra vascular matrix. METHODS: Serum fructosamine, glycosylated hemoglobin and percent glycosylation of IgG, IgA, IgM were measured in five different groups of human subjects: 50 normal individuals; 40 type 2 DM patients; 42 type 1 DM patients; 40 type 2 DM patients with nephropathy and 37 type 1 DM patients with nephropathy. RESULTS: Patients with long-term history of diabetes and chronic hyperglycemia as well as suffering from diabetic nephropathy showed an increased glycosylated hemoglobin level and serum fructosamine as compared to those with diabetes mellitus and to the normal individuals. Glycosylation of IgG, IgA and IgM showed an increase in both type 1 and type 2 DM patients with nephropathy as compared to the diabetic patients without any complication. A positive correlation has been observed between glycosylated IgG and glycosylated hemoglobin (R2=0.522, 0.5113, 0.7117, 0.673) in type 1 and type 2 DM without and with diabetic nephropathy, respectively, whereas correlation between glycosylated IgG and serum fructosamine was observed only in type 1 and type 2 DM without nephropathy (R2=0.7318, 0.5767). CONCLUSION: The present study suggests that glycosylation of IgG is an equivalent marker for advanced glycosylation as GHb and may have some role to play in the on onset of diabetic nephropathy by altering their immunoreactivity leading to microvascular complications.

Adolescent↗
PubMed2006

A comparison of glycemic effects of glimepiride, repaglinide, and insulin glargine in type 2 diabetes mellitus during Ramadan fasting.

Although diabetics may be exempted from Ramadan fasting, many patients still insist on this worship. Aim of the present study is to compare the effects of glimepiride, repaglinide, and insulin glargine in type 2 diabetics during Ramadan fasting on the glucose metabolism. Patients, who were willing to fast, were treated with glimepiride (n=21), repaglinide (n=18), and insulin glargine (n=10). Sixteen non-fasting control type 2 diabetics matched for age, sex, and body mass index were also included. Fasting blood glucose (FBG), post-prandial blood glucose (PBG), HbA1c, and fructosamine as well as lipid metabolism were evaluated in pre-Ramadan, post-Ramadan, and 1-month post-Ramadan time points. There was no significant change from pre-Ramadan in FBG, PBG, and HbA1c variables in fasting diabetics at post-Ramadan and 1-month post-Ramadan. However, PBG was found higher in non-fasting control diabetics at post-Ramadan and 1-month post-Ramadan (p<0.05 and p<0.001, respectively). In fructosamine levels, a significant increase was noted both in fasting group and non-fasting group at 1-month post-Ramadan (p<0.01 for all). However, no significant difference was found in the comparison of the changes in fructosamine levels between fasting group and non-fasting group. Risk of hypoglycemia did not significantly differ between fasting and non-fasting diabetics. There was no significant difference between three drug therapies regarding glucose metabolism and rate of hypoglycemia. No adverse effects on plasma lipids were noted in fasting diabetics. In this fasting sample of patients with type 2 diabetes, glimepiride, repaglinide, and insulin glargine did not produce significant changes in glucose and lipid parameters.

Adult↗
PubMed2006

[Ramadan fasting in type 2 diabetes mellitus].

This study was designed to assess the effects of fasting during Ramadan on weight, blood pressure, metabolic control and plasma lipoproteins in diabetic patients. This study was conducted in December 2000 (Ramadan 1421) when the length of fasting was 12 hours a day. It included 38 type 2 diabetic patients (20 males and 18 females). Mean patient age was 51.410.5 years and mean body mass index (BMI) 28.94.7kg/m2. Three patients were treated with diet and 35 with oral hypoglycemic agents. Clinical and biochemical parameters were evaluated during three periods: three weeks before Ramadan (T0), at the fourth week of Ramadan (T1) and three weeks after the end of Ramadan (T2). During the month of Ramadan, a decrease in weight (0.52kg) and no change in blood pressure were observed. No metabolic complication occurred in our patients. A significant effect of Ramadan fasting was observed on glycemic control and lipoprotein levels. In patients whose fructosamine level before Ramadan was higher than 340micromol/l, plasma fasting glucose and serum fructosamine increased during Ramadan (p<0.003) and returned to initial levels at the end of Ramadan T2; in these patients also, a decrease of HDL-cholesterol (p<0.01) associated with an increase of LDL-cholesterol (p<0.003) were observed at T1 and disappeared at T2. But, in patients whose fructosamine level at T0 was lower than 340micromol/l, no effect on glycemic control and no significant effect on serum lipoprotein levels were found during Ramadan month. Ramadan fasting in type 2 diabetic patients seems to cause slight effects on glycemia and lipoprotein levels when previous metabolic control is quite good; but fasting induces more deterioration when previous control is poor.

Adult↗
PubMed1998

Mood correlates with blood serotonin, but not with glucose measures in patients with recurrent suicidal behavior.

Depressive mood and impulsive-aggressive behavior are relevant to suicidal behavior and have been associated with decreased central serotonergic function. Impulsive behavior and aggression have also been related to abnormal glucose metabolism. To investigate these relationships further, blood serotonin (5-HT), blood glucose measures (glycated hemoglobin (HbA1C) and fructosamine) and self-reported depressive symptoms and anger were repeatedly measured, during a 1-year follow-up, in 106 patients who had recently attempted suicide for at least a second time. A major DSM-III-R Axis I diagnosis or use of antidepressants were reasons for exclusion. Intra-individual variations in blood 5-HT were inversely correlated with variations in depressive mood, self-depreciation, and anger, and positively with variations in fructosamine. Intra-individual variations of HbA1c or fructosamine were not correlated with mood. Mean blood 5-HT levels per patient were positively correlated with mean scores of depression and self-depreciation. The contrast between the intra-individual and the inter-individual correlations may reflect a difference between short- vs. long-term effects. The results do support an association between serotonergic function and mood, but not between glucose metabolism and mood, in recurrent suicide attempters.

Adolescent↗
PubMed1998

Oxidative damage, plasma antioxidant capacity, and glucemic control in elderly NIDDM patients.

A study of oxidative damage was made in elderly noninsulin-dependent diabetes mellitus (NIDDM) patients. A statistically significant increase in glucose and fructosamine was found in fasting NIDDM patients, as well as an increase in the oxidation induced by tert-butyl hydroperoxide. The Total Reactive Antioxidant Potential (TRAP) of the plasma was much reduced (p < .02) and the uricemia was unchanged. The erythrocytes of diabetic patients show greater basal oxidation products (p < .05), and the susceptibility of the diabetic erythrocytes to oxidation injury was also shown to increase in the oxidation induced by t-BOOH (p < .05). Linear regression studies showed that TRAP was associated directly with uric acid (p < .05) and inversely with fructosamine and with glucose (p < .03 and p < .05 respectively) in patients with NIDDM, but not in the controls. The levels of fructosamine were found to be related to the basal damage of the red blood cells (direct correlation, p < .001). This study suggest an useful approach to diabetic oxidative stress for clinical settings.

Adult↗
PubMed1998

Non-enzymatic glycosylation of alkaline phosphatase alters its biological properties.

Hyperglycaemia in poorly controlled diabetic patients induces non-enzymatic glycosylation (glycation) of proteins, altering their structure and physiological bioactivity. Alkaline phosphatase (ALP) is a membrane-bound exoenzyme which faces the extracellular compartment. We have investigated the glycation of intestinal alkaline phosphatase in vitro and the consequences of such molecular modifications on certain structural and functional characteristics. The effect of glycation on alkaline phosphatase specific activity was determined after incubation of the enzyme with different sugars for various periods of time. The formation of early reversible glycation products was determined by the measurement of fructosamine levels, while the appearance of advanced glycation end products was estimated by spectrofluorometric analysis. A decrease in the specific activity of ALP was associated both with an increase in fructosamine levels and with the appearance of AGE-characteristic fluorescence. Changes in these parameters were found to depend on the incubation time, and on the concentration and glycating capability of the sugar employed. Co-incubation with aminoguanidine slowed down the appearance of protein-linked fluorescence, and additionally curbed the decrease in enzymatic specific activity. A significant correlation between the levels of ALP-fructosamine and ALP-advanced glycation end product was observed. Patterns of protein bands fractionated by SDS-PAGE were essentially identical for the nonglycated controls and the glycated samples. The electrophoretic mobility of the band of alkaline phosphatase on cellulose acetate gels increased as a function of the incubation time and the glycosylating power of the carbohydrate used. The present study provides evidence for the in vitro glycation of alkaline phosphatase, and for the consecutive alteration of its activity and structure.

Alkaline Phosphatase↗
PubMed2003

Changes in glycation of fibrous type I collagen during long-term in vitro incubation with glucose.

The course of glycation of calf skin fibrous type I collagen was monitored in vitro under physiological conditions during an 8-week incubation period in order to take into account the long half-life of this protein. The formation of glycated compounds was measured by determining fructosamine, pentosidine, and carboxymethyllysine content. The incubation conditions were as physiological as possible in sterile saline phosphate buffer, except glucose concentration. With incubation medium containing 200 mmol glucose, fibrous collagen underwent solubilization; in addition an increase in fructosamine, pentosidine, and carboxymethyllysine content in both solubilized and remaining insoluble collagen was noticed. There was a spontaneous, restricted, and time-dependent native glycated state of collagen; high concentration glucose enhanced the formation of glycated compounds and induced changes in solubility and glycoxidated products. The production of pentosidine during incubation without glucose should be considered as an event resulting from the initial fructosamine. Whereas the production of carboxymethyllysine during long-term incubation with glucose provided indirect proof of an additional oxidative process after early glycated product formation. These experimental observations provide insight into the in vivo context of advanced glycation end product formation in chronic hyperglycemia and aging.

Animals↗