A Coppelli

Reducing insulin resistance with metformin: the evidence today

Insulin resistance, defined as the inability of insulin to exert a normal biological action at the level of its target tissues, is one of the principal pathogenetic defects of type 2 diabetes. Metformin, the most widely-prescribed insulin-sensitizing agent in current clinical use, improves blood glucose control mainly by improving insulin-mediated suppression of hepatic glucose production, and by enhancing insulin-stimulated glucose disposal in skeletal muscle. Experimental studies show that metformin-mediated improvements in insulin sensitivity may be associated with several mechanisms, including increased insulin receptor tyrosine kinase activity, enhanced glycogen synthesis, and an increase in the recruitment and activity of GLUT4 glucose transporters. In adipose tissue, metformin promotes the re-esterification of free fatty acids and inhibits lipolysis, which may indirectly improve insulin sensitivity through reduced lipotoxicity. The improved glycaemia with metformin is not associated with increased circulating levels of insulin, and the risk of hypoglycaemia with metformin is minimal. The therapeutic profile of metformin supports its use for the control of blood glucose, in diabetic patients and for the prevention of diabetes in subjects with impaired glucose tolerance. Moreover, the improvement by metformin of cardiovascular risk factors associated with the dysmetabolic syndrome may account for the significant improvements in macrovascular outcomes observed in the UK Prospective Diabetes Study.

Effects of pancreas-kidney transplantation on diabetic retinopathy

The effects of pancreas transplantation (PTx) on diabetic retinopathy (DR) are still debated. We studied the course of DR in 48 patients (age: 40 ± 7 years; males/females 26/22, body mass index (BMI): 23.0 ± 2.4 kg/m2, duration of diabetes: 24 ± 8 years) bearing a successful PTx (combined with a kidney). Follow‐up ranged 6–60 months (median: 17 months). Before transplantation, according to the Eurodiab Study classification, 12 patients (25%) had nonproliferative retinopathy (NPDR; mild, moderate or severe), and 36 patients (75%) had laser‐treated and/or proliferative retinopathy (LT/PDR). During the follow‐up, in the NPDR group improvement/deterioration was defined as regression/progression to a lower/higher retinopathy grade; in the LT/PTD group, stabilization was defined as no new neo‐vessel formation or development of new lesions requiring laser‐treatment. In the NPDR group, five (41.7%) patients improved of one or more lesion grading, three (25%) patients showed no change, and four (33.3%) patients progressed of one grade. In the LT/PDR group, the post‐transplant data were: stabilization in 35 (97%) patients, and worsening in one (3%) patient. The number of improved/stabilized patients was significantly higher in the transplanted than in a control group of nontransplanted type 1 diabetic patients. In conclusion, despite a relatively short follow‐up period, successful PTx in our cohort of patients was associated with improvement and/or stabilization of DR in the majority of recipients.

MASSIVE ISOLATION, MORPHOLOGICAL AND FUNCTIONAL-CHARACTERIZATION, AND XENOTRANSPLANTATION OF BOVINE PANCREATIC-ISLETS

The limited availability of human donors makes the search for alternative islet sources mandatory for future developments in pancreatic islet transplantation. In this study, we report on the massive isolation of bovine islets of proven in vitro and in vivo viability. The islets were prepared by collagenase digestion, sequential filtrations, and density-gradient purification by modifying a technique previously developed in our laboratory for the porcine pancreas. The prepurification yield was 2,743 +/- 78 islet equivalents (IE)/g pancreas (mean +/- SE), with a postpurification recovery of 78.7 +/- 2.2%. Purity ranged from 80 to 90%. The histological and immunocytochemical studies demonstrated the identity and integrity of the islets with well-preserved insulin-, glucagon-, and somatostatin-containing cells. The morphological integrity of cultured bovine islets was demonstrated for up to 4 weeks from isolation. Insulin secretion from freshly isolated islets was similar at 3.3 mmol/l glucose (0.36 +/- 0.06 pmol.IE-1.min-1) and at 14 mmol/l glucose (0.42 +/- 0.00 pmol.IE-1.min-1), and it increased significantly (P < 0.01) at 25 mmol/l glucose (1.44 +/- 0.12 pmol.IE-1.min-1). Arginine, theophylline, and propionic acid increased insulin secretion from freshly isolated islets at 3.3 and 14 mmol/l glucose, but not at 25 mmol/l glucose. Islets cultured at 37 degrees C in CMRL 1066 culture medium for at least 10 days were shown to become responsive to a lower glucose concentration, as demonstrated by the significant increase of insulin release in response to 14 mmol/l glucose, when compared with basal secretion. This recovered responsivity to glucose was maintained after 4 weeks of culture.(ABSTRACT TRUNCATED AT 250 WORDS)

The biguanide compound metformin prevents desensitization of human pancreatic islets induced by high glucose

Pancreatic islet desensitization by high glucose concentrations is a temporary and reversible state of beta-cell refractoriness to glucose (and possibly other secretagogues), due to repeated or prolonged pre-exposure to increased glucose concentrations. We evaluated whether the oral antidiabetic agent metformin affects this phenomenon in isolated, human pancreatic islets, and whether the possible effects of the biguanide are influenced by the presence of a sulphonylurea, glyburide. Islets prepared from five human pancreases were incubated for 24 h in M199 culture medium containing either 5.5 or 22.2 mmol/l glucose, with or without a therapeutic concentration (2.4 microg/ml) of metformin. Then, the islets were challenged with either 3.3 mmol/l glucose, 16.7 mmol/l glucose, or 3.3 mmol/l glucose + 10 mmol/l arginine, and insulin release was measured. After incubation in the absence of metformin, the human islets exposed to 22.2 mmol/l glucose showed no significant increase in insulin release when challenged with 16.7 mmol/l glucose (confirming that hyperglycemia desensitizes pancreatic beta-cells). In the presence of metformin, the islets fully maintained the ability to significantly increase their insulin release in response to glucose, even when previously exposed to 22.2 mmol/l glucose. No major effect on arginine-induced insulin release was observed, whatever the culture conditions. The protective action of metformin was observed also when glyburide was present in the incubation medium, whereas the sulphonylurea alone did not affect insulin release from the islets previously exposed to high glucose concentrations. These in vitro results suggest that metformin can prevent the desensitization of human pancreatic islets induced by prolonged exposure to increased glucose concentrations.

Pancreas transplant alone determines early improvement of cardiovascular risk factors and cardiac function in type 1 diabetic patients

Background. The effects of pancreas transplant alone (PTA) on cardiovascular risk factors (CRF) and cardiac function in type 1 diabetes mellitus (T1DM) patients are still unsettled. Methods. We studied 13 T1DM patients who received PTA with portal drainage and 11 matched control patients. Parameters of glucose and lipid metabolism and several additional classic CRF were assessed before and up to 6 months posttransplant. Cardiac morphology and function were assessed by Doppler echocardiographic examination. Results. Insulin independence was promptly achieved and then maintained after PTA. Total and low-density lipoprotein cholesterol levels were significantly lower after transplantation, whereas high-density lipoprotein cholesterol and triglyceride concentrations did not change. Both systolic and diastolic blood pressure values and fibrinogen levels improved significantly. In addition, PTA determined a significant amelioration of several morphologic and functional cardiac indices. None of the measured parameters changed in the control patients. Conclusions. PTA with portal drainage induces an early improvement of CRF and ameliorates cardiac function in patients with T1DM.

Custom-Made Orthesis and Shoes in a Structured Follow-Up Program Reduces the Incidence of Neuropathic Ulcers in High-Risk Diabetic Foot Patients

Objective. The objective of this study was to assess the impact of a structured follow-up program on the incidence of diabetic foot ulceration (DFU) in high-risk diabetic patients. Research Design and Methods. A total of 1874 diabetic patients referred to the Diabetic Foot Unit of the University of Pisa were ranked based on the ulcerative risk score proposed by the International Consensus on Diabetic Foot. Out of 334 patients (17.8%) with a score ≥2, 298 accepted to participate in this prospective trial and were randomized into 2 groups: group A, which received standard treatment, and group B, in which the patients received, as a part of a structured prevention program, custom-made orthesis and shoes. Incidence of new DFUs was observed for no less than 1 year and in a subset of patients after 3 and 5 years, respectively. Incidence of new DFUs and recurrences were considered as primary endpoints to establish the effectiveness of the program; costs were also compared. Results. Among the patients enrolled in this follow-up analysis, 46% had neuropathy and deformities, 20% had previous ulceration, 25% had previous minor amputation, and 9% had neuro-osteoarthropathy. During the first 12-month follow-up, 11.5% of patients in group B developed a DFU compared with 38.6% in group A (P < .0001). In the extended follow-up, the cumulative incidence of ulcer in group B compared with group A was 17.6% versus 61% (P < .0001) after 3 years and 23.5% versus 72% (P < .0001) after 5 years, respectively. The net balance at the end of the follow-up was highly in favor of the prevention program, with a saving of more than €100 000 per year. Conclusions. The implementation of a structured follow-up with the use of orthesis and shoes can reduce the incidence of DFU in diabetic patients who are at high ulcerative risk and its related costs.

Characterization of peripheral benzodiazepine receptors in purified large mammal pancreatic islets

In this work, we evaluated the biochemical properties of peripheral benzodiazepine receptors (PBRs) in the porcine endocrine pancreas and their role in insulin release. Binding of [3H]1-(2-chlorophenyl-N-methyl-1-methyl-propyl)-3-isoquinolinecarboxa mide ([3H]PK-11195), a specific ligand of PBRs, to islet membranes was saturable and Scatchards analysis of saturation curve demonstrated the presence of a single population of binding sites, with a dissociation constant (Kd) value of 4.75 +/- 0.70 nM and a maximum amount of specifically bound ligand (Bmax) of 4505 +/- 502 fmol/mg of proteins. The pharmacological profile of PBRs was determined as the ability of PK-11195 and several benzodiazepine compounds to displace [3H]PK-11195 from these binding sites. The rank order of potency yielded the following affinity results: PK-11195 > 7-chloro-1,3-dihydro-1-methyl-5-(p-chlorophenyl)-2H-1,4-benzodiazepine-2 -on (Ro 5-4864) > diazepam > or = flunitrazepam >> flumazenil. Secretion studies demonstrated that PK-11195 (1 and 10 microM) and Ro 5-4864 (10 and 50 microM) significantly potentiated insulin secretion from freshly isolated porcine islets at 3.3 mM glucose. This potentiating effect was not observed at 16.7 mM glucose concentration nor by the addition of clonazepam. These results show the presence of PBRs in purified porcine pancreatic islets and suggest an implication of PBRs in the mechanisms of insulin release.

Effects of glibenclamide and metformin (alone or in combination) on insulin release from isolated human pancreatic islets

Abstract Isolated human pancreatic islets were prepared by collagenase digestion and density gradient purification, and the effects of glibenclamide (0.5 and 5.0 µmol/l) and metformin (20 and 200 µmol/l), alone or in combination, on insulin release were evaluated at varying glucose concentrations. At 3.3 mmol/l glucose level, the addition of 5.0 µmol/l glibenclamide or 5.0 µmol/l glibenclamide plus 200 µmol/l metformin caused a significant increase of insulin release, compared with glucose alone. At 16.7 mmol/l glucose concentration, a significant increase of insulin secretion, compared with glucose alone, was produced by the addition of either 5.0 µmol/l glibenclamide, 200 µmol/l metformin, or both 5.0 µmol/l glibenclamide and 200 µmol/l metformin. The effect of the combination of the two drugs was significantly higher than that with either drug used alone. Thus, glibenclamide was shown to have an insulinotropic effect on human islets at both low and high glucose concentrations, and metformin at high glucose concentrations. A possible synergistic effect of glibenclamide and metformin at high glucose concentrations is also suggested.

Long-term survival and function of isolated bovine pancreatic islets maintained in different culture media

Bovine islets are being evaluated for their potential in transplantation studies. We studied the recovery, morphology, and function of purified bovine islets cultured up to 4 weeks under varying conditions. Approximately 60% of the initial islet mass could be recovered after 4 weeks at 37°C in CMRL 1066 or M 199 culture medium, and the cultured islets were well preserved histologically and viable both in vitro and in vivo. On the other hand, culture with RPMI 1640 caused disaggregation of the islets within a few days, with altered in vitro viability. Thus, culturing purified bovine islets with appropriate media is a suitable procedure to maintain islet mass, morphology, and function in the long term.

Safety and Effectiveness of Therapeutic Magnetic Resonance in the Management of Postsurgical Lesion of the Diabetic Foot

To evaluate the safety and effectiveness of therapeutic magnetic resonance (TMR) in the management of the diabetic foot (DF), we treated a group of consecutive type 2 diabetic inpatients with wide postsurgical lesions (Group A: N = 10; age 67.7 ± 18.9 years, duration of diabetes 22.3 ± 6.6 years, 8.1 ± 1.1%, body mass index 29.4 ± 2.1 kg/m2), for 2 consecutive weeks, while admitted, with a low-intensity magnetic resonance equipment, in addition to standard treatment. Patients, compared with a matched control group with the same clinical characteristics (Group B), were then followed monthly for 6 months to evaluate healing rate (HR), healing time (HT), rate of granulation tissue (GT) at 3 months, and adverse events. HR was of 90% in Group A and 30% in Group B (P < .05); GT was 73.7 ± 13.2% in Group A versus 51.84 ± 18.77% in Group B (P < .05). HT in Group A was 84.46 ± 54.38 days versus 148.54 ± 78.96 days in Group B (P < .01). No difference in adverse events (5 in Group A and 6 in Group B) was observed throughout the study period. In this pilot study, the use of TMR at this dose and duration was safe. The results also permit the observation that TMR plus standard care offered a faster healing rate compared with standard care alone.