Peptide mapping and characterisation of glycation patterns of the glima 38 antigen recognised by autoantibodies in Type I diabetic patients.

AIMS/HYPOTHESIS: Glima 38 is an N-glycated neuroendocrine membrane protein of M(r) 38,000, which is recognised by autoantibodies in approximately 20% of patients with Type I (insulin-dependent) diabetes mellitus. The aim of this study was to characterise the carbohydrate moiety and generate peptide maps of glima 38. METHODS: Sera of high immunoreactivity to glima 38 were used to isolate 35-S methionine-labelled protein from betaTC-3 cells and a neuronal cell line GT1.7. Tunicamycin was used to inhibit N-glycation of glima 38 and define the core protein. The carbohydrate moiety was characterised for tunicamycin sensitivity, lectin binding and susceptibility to different endoglycosidases. The protein moiety was subjected to digestion by proteases to define peptide maps. RESULTS: The autoreactive epitopes in glima 38 recognised by Type I diabetic sera are conformational and independent of the carbohydrate moiety. Inhibition of N-glycation of glima 38 in vivo, shows a protein core of M(r) 22,000 in both pancreatic beta-(betaTC3) and neuronal (GT1.7) cell lines. The carbohydrate moieties in the two cell types are distinct but contain a similar amount of terminal sialic acid residues and at least five oligosaccharide chains Glima 38 binds Triticum vulgare and Ricinus communis I lectins. Endoproteinase treatment of the M(r) 22,000 core protein results in peptides of M(r) 4500 and M(r) 20,000 with Lys-C, and peptides of M(r) 4000 and M(r) 11,000-12,000 with Glu-C/V8 and Asp-N proteases. CONCLUSION/INTERPRETATION: The biochemical properties of glima 38 define it as a new autoantigen in Type I diabetes and provide a basis for its purification.

Hypoglycemia following inadvertent and factitious sulfonylurea overdosages.

OBJECTIVE: To recognize unreported sulfonylurea overdosages in hypoglycemic patients. CASES: We describe three patients with hypoglycemia due to inadvertent (in two patients) and factitious (in one patient) sulfonylurea overdosages. We review the world literature and summarize 43 previously published cases of inadvertently administered and 23 previously published cases of factitiously self-administered sulfonylurea overdosages with hypoglycemia. RESULTS: An inadvertently administered fulsonylurea overdosage usually occurred when a sulfonylurea was accidentally substituted for an intended medication with a similar generic or trade name. Features of the patients with a factitiously self-administered sulfonylurea overdosage included: 1) a history of the patient or patient's spouse having a medical job or sulfonylurea-treated diabetes mellitus; 2) an unusual affect or psychiatric history; 3) an abrupt onset of severe symptoms without previous milder symptoms; and 4) an absent hypoglycemic or hyperinsulinemic response to provocative testing. These features are not typical for an insulinoma. CONCLUSIONS: When a hypoglycemic patient denies antidiabetic medication use, we recommend sequentially performing: 1) a thorough pill inspection; 2) an interview for recently altered pill appearances; 3) a measurement of serum insulin and C-peptide levels during hypoglycemia; and 4) a blood or urine sulfonylurea screen. Discovery of an unreported sulfonylurea overdosage can eliminate the need to search for an insulinoma and prevent further overdosages from occurring.

Insulin therapy in insulin-dependent (type I) diabetes mellitus.

Intensive insulin therapy is suitable for individuals who want optimal glucose control and the least fluctuation in glucose levels. It permits a greater flexibility in life style and the opportunity to vary, within certain limits, meal and exercise schedules and meal content. This therapeutic approach is modeled on physiologic insulin secretion, but shortcomings in the subcutaneous route of delivery hamper the ability to control precisely the glucose levels. The therapy is most successful when the user is motivated to do the glucose monitoring and carbohydrate counting and educated to make rational decisions about how to adjust the insulin doses.

Biological activity of nasally administered insulin in normal subjects.

Nasally administered (IN) insulin has been advocated as a potentially useful alternative to subcutaneously administered regular insulin because of its more rapid onset and time to peak action and its shorter duration of action. This study further defines the pharmacodynamics of IN insulin by using a euglycemic clamp technique to determine the bioavailability of IN insulin as compared with intravenous (IV) insulin, and to ascertain whether multiple sequentially administered doses of IN insulin alter pharmacodynamics. Eight normal volunteers received 2 control doses of IV insulin (0.05 U/kg), and 3 high doses (0.7 U/kg) and 3 low doses (0.35 U/kg) of IN insulin with an absorption enhancer (tauro-24,25 dihydrofusidate) given sequentially over a 2 day period. A euglycemic clamp was performed with a Biostator (Ames) that infused dextrose to keep the subject's blood glucose at his fasting level. Analysis of dextrose infusion curves for the low and high doses of IN insulin revealed an onset of action of 9.4 +/- 0.4 and 10.5 +/- 0.3 minutes, time to peak action of 20.6 +/- 5.6 and 23.7 +/- 4.4 minutes and duration of action of 82.1 +/- 5.2 and 95 +/- 5.7 minutes respectively. Both the onset of action and time to peak action were slightly longer (P less than .05) for the high as compared with the low dose IN insulin, although this should not represent a clinically significant difference. The total dextrose requirement was 21.9 +/- 2.3 g for the low dose IN insulin and 34.1 +/- 3.3 g for the high dose IN insulin, the latter value being significantly greater (P less than .01) than the former.(ABSTRACT TRUNCATED AT 250 WORDS)

Selective unresponsiveness of pancreatic beta-cells to acute sulfonylurea stimulation during sulfonylurea therapy in NIDDM.

Patients with non-insulin-dependent diabetes mellitus (NIDDM) who have chronic hyperglycemia lose acute incremental insulin responses to glucose but are able to briskly respond to other beta-cell secretagogues. To investigate whether this is a defect specific for glucose or represents a more general phenomenon, we measured the insulin responses to acute intravenous tolbutamide in 10 obese patients with NIDDM both before and during sulfonylurea therapy with tolazamide. Comparable glycemia was achieved with oral dextrose 2 h before intravenous testing. To assess beta-cell responsiveness to a nonsulfonylurea secretagogue, 1 mg glucagon was administered intravenously during tolazamide therapy. In seven patients, the mean peak insulin increment 5 or 10 min after intravenous tolbutamide was 54 +/- 11 microU/ml when not receiving tolazamide (0.14 +/- 1.3 microU/ml) with tolazamide (P less than .001), even though serum insulin responded rapidly to intravenous glucagon. In four patients tested for reversibility of their refractoriness to intravenous tolbutamide during chronic tolazamide therapy, the mean peak insulin increment 1 wk after discontinuing tolazamide was 79 +/- 22 microU/ml. A relatively rapid development of refractoriness was documented in four patients who were tested only 12 h after beginning tolazamide therapy; the mean peak insulin increments 5-10 min after intravenous tolbutamide were undetectable (-0.5 microU/ml), yet responses to intravenous glucagon were evident. In these NIDDM patients, exposure of pancreatic beta-cells to sustained levels of sulfonylureas induces a reversible state of refractoriness to acute stimulation with sufonylureas but not to another secretagogue.(ABSTRACT TRUNCATED AT 250 WORDS)

High performance liquid chromatography used to distinguish the autoimmune hypoglycemia syndrome from factitious hypoglycemia.

Recurrent episodes of spontaneous hypoglycemia developed in a 30-yr-old woman who had received a brief course of insulin therapy 10 yr previously. She denied surreptitious insulin administration, and the autoimmune hypoglycemia syndrome was considered. Her insulin levels could not be reliably measured because of the presence of circulating antiinsulin antibodies, which interfere with standard RIA techniques. Reverse phase high performance liquid chromatographic analysis of serum obtained during a hypoglycemic episode revealed a mixture of beef and pork insulins but no human insulin, firmly establishing the diagnosis of factitious hypoglycemia. This case illustrates the value of reverse phase high performance liquid chromatography in characterizing patients in whom the autoimmune hypoglycemia syndrome is suspected.

Reduced solubility of short-acting soluble insulins when mixed with longer-acting insulins.

Using insulins from three manufacturers, we examined the recovery by radioimmunoassay of short-acting soluble insulin when mixed with long-acting insulin as a function of the ratio of the mixture and the time of pre-mixing. In ratios of 1:2, 1:3, and 1:5 (short- to long-acting insulin), all Novo, Nordisk, and Lilly short-acting insulins tested showed a significant loss of solubility when mixed with the respective company's long-acting insulin either for less than 75 s or for 20 min before centrifugation. In ratios of 1:1, Novo's Actrapid (regular) with Monotard (lente) and Lilly's regular with lente showed no significant loss of solubility when pre-mixed for less than 75 s, and the regular insulin also showed no significant loss when pre-mixed for 20 min. However, when Lilly's regular was mixed with either NPH or ultralente in a 1:1 ratio, a significant loss of solubility of the short-acting insulin occurred regardless of time [as was also found with Nordisk's Velosulin (regular) with insulatard (NPH)]. When Lilly regular was incubated with Lilly lente in ratios of 1:3 for less than 75 s, 20 min, 4 h, and 24 h before centrifugation, there was a progressive loss of solubility. In contrast, with the same ratios and times of pre-mixing, Lilly regular when mixed with Lilly NPH showed a rapid initial loss of solubility that plateaued by 20 min before centrifugation.(ABSTRACT TRUNCATED AT 250 WORDS)