Detection of IgE insulin antibody with radioallergosorbent test

Summary An in vitro method for detecting IgE insulin antibody using the principle of the radioallergosorbent test (RAST) is described. In six patients with insulin allergy, the RAST values were higher than in normal persons or insulin-treated diabetics without insulin-allergy. No differences were observed between normal persons and insulin-treated diabetics without insulin-allergy. Moreover, it was observed that in one patient treated with highlypurified insulin, there was a gradual decrease of RAST value parallel to the radioinsulin binding activity and clinical allergic symptoms. The RAST value of insulin is slightly inhibited by non-IgE antibodies and is, therefore, a semiquantitative value. However, the RAST is simple to perform and reproducible; It therefore is very useful in the detection of IgE insulin antibodies.     

Precipitin reactions between insulin,proinsulin, insulin chains and insulin antibody

Summary Insulin antibody was produced in guinea pigs and the precipitins tested by double diffusion in agarose gel. Pork, beef and monocomponent insulin produced precipitin lines. Proinsulin also produced a precipitin line with these antisera but no lines appeared with either the A-chain or the B-chain of insulin. There was good correlation between the precipitin titre and the radioimmunoassay titre.     

Protamine antibody production in diabetic subjects treated with NPH insulin

Treatment with neutral protamine Hagedorn (NPH) insulin predisposes individuals with diabetes to anaphylactoid reactions when given bolus protamine for heparin reversal during cardiovascular procedures. To prospectively examine production of protamine antibodies, 30 patients with non-insulin dependent diabetes were followed for 12 months from initiation of therapy with porcine NPH or Lente insulin. Twenty-one subjects were randomly assigned to NPH (protamine containing) and nine controls to Lente (protamine free) insulin. Protamine specific IgG antibody was produced by 6/21 (29%) of NPH-treated subjects and 0/9 controls. Among NPH treated subjects, there was no difference between protamine antibody producers and non-producers with regard to age, race, weight, or pre-treatment glycosylated hemoglobin. Both producer and non-producer groups received similar amounts of insulin and protamine and achieved similar glycemic control. Insulin antibodies were made by 4/6 (67%) of protamine antibody producers and by 6/15 (40%) of non-producers (NS). The authors conclude that one of three new diabetics who are treated with porcine NPH insulin will make IgG protamine antibodies. These antibodies do not affect insulin requirements, glycemic control, or insulin antibody production. Because of the frequency of protamine antibody production and the risk of anaphylaxis to bolus protamine administration in NPH treated diabetics, the authors suggest that NPH insulin-treated individuals should avoid heparin reversal by protamine.     

Cyclophosphamide inhibition of insulin antibody production,insulin resistance and experimental immunodiabetes

Summary Hyperglycemia, increase of insulin tolerance and striking changes in B- and A-cells of the islets of Langerhans occurred in guinea pigs producing precipitating insulin antibodies after immunization with insulin in adjuvant. Treatment with cyclophosphamide inhibited the production of insulin antibodies while the blood sugar and insulin tolerance remained on the level of nonimmunized control animals. Also the islets of Langerhans were less severely injured than those of animals not treated with cyclophosphamide.

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Zusammenfassung Hyperglykämie, erhöhte Insulintoleranz und auffällige Veränderungen der A- und B-Zellen der Langerhansschen Inseln wurden bei Meerschweinchen beobachtet, welche infolge Immunisierung mit Insulin in Adjuvans Insulin-präzipitierende Antikörper bilden. Cyclophosphamid-Behandlung verhindert die Bildung von Insulin-Antikörpern, während Blutzucker und Insulintoleranz auf dem gleichen Niveau bleiben wie bei nicht immunisierten Kontrolltieren. Auch die Langerhansschen Inseln sind weniger schwer geschädigt als bei nicht mit Cyclophosphamid behandelten immunisierten Tieren.

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Resumen En covayos productores de anticuerpos antinsulina precipitantes, tras inmunización con insulina y adyuvante, se ha podido observar: hiperglicemia, aumento de la tolerancia a la insulina y conspícuas modificaciones a cargo de las células A y B de las islas de Langerhans. El tratamiento con ciclofosfamida inhibía la producción de anticuerpos antinsulina, mientras la glucemia y la tolerancia insulínica permanecían en los mismos niveles que podían observarse en los animales de control no inmunizados. Las islas de Langerhans aparecían menos dañadas respecto a las de los animales no tratados con ciclofosfamida.

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Résumé Hyperglycémie, amélioration de la tolérance à l'insuline et modifications considérables à niveau des cellules B et A des ilots de Langerhans ont fait l'objet d'une observation, après immunisation par insuline et adjuvant, en cobayes produisant des anticorps anti-insuline précipitants. Le traitement par le cyclophosphamide inhibait la production d'anticorps anti-insuline, alors que la glycémie et la tolérance à l'insuline restaient aux mêmes niveaux qu'on pouvait vérifier chez les animaux de contrôle non immunisés. Même les îlots de Langerhans semblaient avoir subi des dommages moins importants par rapport à ceux des animaux qui n'avaient pas été traités par le cyclophosphamide.

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Riassunto Iperglicemia, aumento della tolleranza all'insulina e cospicue modificazioni a carico delle cellule B ed A delle isole di Langerhans sono stati osservati, dopo immunizzazione con insulina in adiuvante, in cavie produttrici di anticorpi anti-insulina precipitanti. Il trattamento con ciclofosfamide inibiva la produzione di anticorpi anti-insulina, mentre la glicemia e la tolleranza insulinica rimanevano agli stessi livelli riscontrabili negli animali di controllo non immunizzati. Anche le isole di Langerhans apparivano meno danneggiate rispetto a quelle degli animali non trattati con ciclofosfamide.

     

Cytokine profile and insulin antibody IgG subclasses in patients with recent onset Type 1 diabetes treated with oral insulin

Aims/hypothesis Tolerance to orally administered antigens may be generated through the induction of T helper cell type 2 and 3 (Th2/Th3) regulatory cells. We previously reported that treatment of recent onset Type 1 diabetes with oral insulin had no effect on residual beta cell function. The aim of this study was to evaluate whether this treatment produces a deviation in the immune response, with polarisation of the cytokine pattern and the induction of a Th2-like antibody response.Methods Mononuclear cells were collected from a total of 20 patients with Type 1 diabetes before and after 12 months of treatment with oral insulin (n=11) or placebo (n=9). Following stimulation of the cells with insulin or phytohaemagglutinin, levels of Th2 and Th3 cytokines (including TGF-, IFN-, IL-4 and IL-5) in the culture supernatants were assessed by ELISA. In addition, levels of total and specific insulin antibody IgG subclasses were measured by radioimmunoassay in serum samples drawn from 33 patients with Type 1 diabetes before and after 3, 6 and 12 months of therapy with oral insulin (n=18) or placebo (n=15).Results After 12 months of treatment, the release of TGF- was significantly higher in patients who received oral insulin compared with those who received placebo (p=0.025 and p=0.006 for lymphocytes challenged with insulin and phytohaemagglutinin respectively). The two groups had similar levels of IL-4 and IL-5 both at baseline and after 12 months of treatment. The release of IFN- was markedly reduced in patients treated with oral insulin compared with those who received placebo at the 12-month follow-up. Circulating levels of IgG1 and IgG3 subclasses directed against insulin were significantly lower in the oral insulin group than in the placebo group after 12 months of treatment (p=0.05 for IgG1 and p=0.014 for IgG3).Conclusions/interpretation The increased TGF- release observed in patients treated with oral insulin suggests that a regulatory response can be induced in vivo by this treatment. The lower levels of insulin antibody IgG1 and IgG3 subclasses present in patients exposed to oral insulin are consistent with a Th2 deviation of the immune response. The failure of oral insulin treatment to provide any measurable clinical benefit may be due to the timing of treatment initiation.     

Acute liver dysfunction complicated with uncontrollable glycemia due to insulin antibody: successful treatment with glucocorticoid and lispro insulin

Here, we report a case of acute liver dysfunction complicated with uncontrollable glycemia due to insulin antibody. The patient was admitted to our hospital due to diabetic ketoacidosis. He was administered insulin immediately, however, his fasting plasma glucose level remained unstable despite the insulin treatment. Blood biochemistry revealed severe liver dysfunction, although no markers including hepatitis virus or autoantibodies associated with autoimmune liver diseases were detected. The 125I-insulin binding rate was high (54%). The characteristics of insulin antibody in this patient were similar to the antibodies of IAS patients, therefore we administered oral glucocorticoid against insulin antibody. The reduction in the 125I-insulin binding rate and the binding capacity of the high affinity site of insulin antibodies were balanced after oral glucocorticoid therapy. In addition, preprandial subcutaneous regular insulin was switched to lispro insulin. Postprandial plasma glucose levels were relatively improved by lispro insulin. The etiology of acute liver dysfunction was unknown, however, we believe that the combination of oral glucocorticoid and lispro insulin was suitable and useful for preventing recurrent liver dysfunction in this patient.     

Anti-insulin receptor antibody and insulin resistance]

Type B insulin resistance syndrome is characterized by the presence of a circulating anti-insulin receptor antibody which inhibits insulin action. The syndrome is usually associated with autoimmune diseases, acanthosis nigricans and hyperinsulinemia. The antibody causes insulin resistance by inhibiting insulin binding to insulin receptors, decreasing insulin receptor number, i.e. down regulation of insulin receptors, and desensitizing insulin receptors which leads to decreased insulin signal transduction through insulin receptors. It also has the ability to elicit insulin action associated with or without increased autophosphorylation of insulin receptors. The antibody usually binds to the insulin receptor with decreased association rate and decreased dissociation rate. The antibody which elicits insulin action without activation of kinase activity of insulin receptors may induce conformational change of insulin receptors. These characteristic features of the anti-insulin receptor antibody may be informative in solving the mechanism of insulin action and may be a useful tool to study insulin receptor functions.     

Transplacental passage of insulin complexed to antibody.

The passage of plasma proteins across the placental barrier in humans is known to be highly selective. Thus, free maternal insulin has been reported not to cross the normal maternofetal barrier, although insulin-binding antibodies have been detected in newborn infants whose diabetic mothers received insulin therapy. In this report we demonstrate, with the use of a human antiserum that permits distinction between human and animal insulins, that insulin in the cord blood of each of two neonates of insulin-treated diabetic mothers was, in part, animal insulin. The higher the antibody titer of the mother the greater was the total insulin in the cord plasma and the greater was the fraction that was animal insulin. In case 1 cord plasma insulin was 0.7 unit/liter, of which 10% was animal insulin; in case 2 cord plasma insulin was 3.5 units/liter, of which 25% was animal insulin. The demonstration that antigen restricted from transplacental passage can be transferred while complexed to antibody raises the question whether such fetal exposure would induce partial or total immunologic unresponsiveness subsequently if the fetus were rechallenged with the same antigen.