Erythrocyte membranes drawn from diabetic patients with poor metabolic control have increased protein glycosylation and decreased Ca2(+)-ATPase activity. A significant relationship was found between these two parameters. Similar results were obtained when protein glycosylation and Ca2(+)-ATPase activity were measured in membranes from normal erythrocytes preincubated with glucose. In this condition, both parameters showed a clear time and dose dependence. Incubation of erythrocyte membranes instead of intact erythrocytes with glucose and glucose-6-phosphate strongly suggests that only the glycosylation of the membrane inner-surface proteins can affect Ca2(+)-ATPase activity. The simultaneous presence of 10 mM glucose and 5 mM ATP in the incubation medium did not affect the degree of erythrocyte membrane protein glycosylation but significantly blocked the inhibitory effect of glucose on Ca2(+)-ATPase activity. However, 5 mM ATP only partially blocked the inhibitory effect of 100 mM glucose, suggesting a competitive mechanism of glucose and ATP for the enzyme active site. Our results show that glycosylation of erythrocyte membrane proteins significantly inhibits Ca2(+)-ATPase activity. This effect could contribute to the development of the capillary closure process observed in diabetic patients. Furthermore, it could represent an index of a general impairment of enzyme function arising in cells chronically exposed to high glucose levels.

The possible occurrence of increased non-enzymatic glycosylation of serum insulin-like growth factor binding protein-3 (IGFBP-3) in vivo and the changes that would simultaneously occur in serum levels of IGFBP-3 and insulin-like growth factor-1 (IGF-I) were investigated. We measured levels of IGF-I and IGFBP-3 and the degree of glycation of total serum protein and IGFBP-3, in serum samples obtained from patients with poorly controlled noninsulin-dependent diabetes (type 2) and from age-matched non-diabetic controls. Type 2 diabetic patients had significantly higher glycated serum protein (GlyP) levels. GlyP significantly correlated with age in the control (r = 0.315, P < 0.05) but not in the type 2 diabetes group. Control and diabetic subjects had comparable serum IGF-I levels and in both groups IGF-I levels tended to decrease with age (r = -0.567, P < 0.001 and r = -0.465, P < 0.05 for control and type 2 diabetic subjects, respectively). In the type 2 diabetes group, IGF-I levels showed a negative correlation with serum GlyP values (r = -0.476, P < 0.05). Type 2 diabetic and control patients had comparable serum IGFBP-3 levels, which were significantly higher in diabetic patients in the older age subgroups. A negative correlation was found between IGFBP-3 levels and age in the control (r = -0.705, P < 0.001) and in the type 2 diabetes groups (r = -0.463, P < 0.05). A significant negative correlation was found between IGFBP-3 levels and GlyP in control (r = -0.449, P < 0.002) but not in type 2 diabetic subjects. The mean glycated IGFBP-3 (GlyIGFBP-3) levels were higher in the oldest type 2 diabetic patients. In these patients, GlyIGFBP-3 was negatively associated with IGF-I levels (r = -0.447, P < 0.05). The IGF-I/IGFBP-3 molar ratio was significantly reduced in the 46-60-year-old type 2 diabetic group, whereas the IGF-I/IGFBP-3 ratio was positively and significantly correlated with GlyP levels only in the control group (r = 0.489, P < 0.01). Our results show that: a) increased non-enzymatic glycosylation of IGFBP-3 occurs in vivo; and b) this effect is accompanied by an increase in IGFBP-3 levels. These results suggest that the IGF-I/IGFBP-3 system is another target for the metabolic derangements of type 2 diabetes. Its alterations might play a role in diabetic complications.

Autoimmune diabetes is an organ specific and multifactorial disorder including insulin dependent diabetes mellitus (Type 1 Diabetes) and latent autoimmune diabetes in adults (LADA), which progresses to insulin dependency because of the beta cells destruction. Several polymorphisms in different genes have been associated with diabetes. The CTLA4 gene is considered a down regulator of T cell function, and the SUMO4 gene encodes a small ubiquitin-like modifier implicated in the intensity and duration of the immune response. We selected 62 LADA patients, 123 patients with Type 1 diabetes patients and 136 unrelated volunteers to study CTLA4 -318 C/T, 159 C/T, 3' STR and SUMO4 163 A/G polymorphisms by PCR. There was a statistical difference significant in the frequency of the allele 209pb for the 3'STR between LADA and Type 1 diabetes patients but not with respect the normal group, the frequencies were found to be 6.9%, 1.0% and 1.9%, respectively. However, no association with either of the polymorphisms has been found in the studied population. The knowledge of the several susceptibility loci in autoimmune diabetes will enhanced the prediction of individuals at high risk of developing the disease in order to establish the best treatment and the prevention of autoimmune diabetes.

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