Diabetologia

Mangili, R.; Viberti, G.; Vergani, D.

doi: 10.1007/BF00278745pmid: 2466725

125 31 31 9 9 R. Mangili G. C. Viberti D. Vergani Unit for Metabolic Medicine United Medical Schools of Guy's and St. Thomas's Hospitals Guy's Campus UK Department of Immunology King's College Hospital School of Medicine and Dentistry London UK Summary The presence of antibodies to glycosylated albumin was studied by means of a newly developed sandwich enzyme-linked immunosorbent assay in 29 long-standing Type 1 (insulin-dependent) diabetic patients with microvascular complications and in 20 normal subjects. Two types of antibody reactivity were detected. One directed against glucitol-albumin expressing G and M isotypes. The second, predominantly belonging to the IgG class, reacted with an epitope shared by non-glycosylated albumin and the ketoamine adduct of albumin glycosylation. Both types of antibodies, with affinity constant ranging from 10 4 to 10 7 (mol/l) −1 were found in normal and diabetic subjects, but higher litres were significantly more prevalent in the diabetic patients. These antibodies may represent the result of immune tolerance breakdown or, alternatively, be natural antibodies. Although their function remains to be established, their raised prevalence in Type 1 diabetes may be relevant to diabetic microvascular disease.

Ludvigsson, J.; Binder, C.; Mandrup-Poulsen, T.

doi: 10.1007/BF00278746pmid: 3069532

125 31 31 9 9 J. Ludvigsson C. Binder T. Mandrup-Poulsen Department of Paediatrics University Hospital Linköping Sweden Steno Memorial Hospital, Hagedorn Research Laboratory Gentofte Denmark Summary Blood was drawn from 74 children, 3–16 years old, at diagnosis of Type 1 (insulin-dependent) diabetes and before the first insulin injection. Insulin autoantibodies were detected with a polyethylen-glycol-method in 27/74 (36.4%) and with an immuno-electrophoretic method in 6/74 (8.1%). Islet cell cytoplasmic antibodies detected by indirect immuno-fluorescence were found in 49/74 patients (66.2%), who included as many as 23 of the 27 patients with insulin autoantibodies determined with the polyethylen-glycol-method ( p <0.01). The proportion of insulin autoantibody-positive patients who developed insulin antibodies during the first 9 months of insulin treatment was not significantly greater (51.8%) than that of insulin autoantibody-negative patients (44.6%), but patients with both islet cell antibodies and insulin autoantibodies at diagnosis produced more insulin antibodies during the first 9 months ( p <0.05). There was no difference in fasting or meal stimulated serum C-peptide after 3, 9 or 18 months as related to occurrence of insulin autoantibodies and/or islet cell antibodies. The correlation between insulin autoantibodies and islet cell antibodies indicates that both types of autoantibodies reflect the same immunological process, although the lack of correlation to C-peptide may indicate that they play a minor causal role. In addition, the results show that patients with an active autoimmune process evidently tend to produce more insulin antibodies during the first months of insulin treatment, but the islet cell antibodies and insulin autoantibodies-positive patients had at least as good residual B-cell function as patients without autoantibodies at diagnosis. If insulin antibodies produced as a response to exogenous insulin do have a negative effect on B-cell function our present results suggest that such mechanisms are of minor importance.

Flynn, M.; Edmonds, M.; Tooke, J.; Watkins, P.

doi: 10.1007/BF00278747pmid: 3234641

125 31 31 9 9 Dr. M. D. Flynn M. E. Edmonds J. E. Tooke P. J. Watkins Diabetic Department King's College Hospital Denmark Hill Camberwell Department of Physiology Charing Cross and Westminster Medical School London UK Medical Unit Torridge Ward Level 8 Royal Devon and Exeter Hospital Barrack Road EX2 5DW Exeter UK Summary The two major components of the microcirculation in the diabetic neuropathic foot have been examined in detail. Nutritive capillary blood flow was measured directly using the non-invasive technique of television microscopy, applied to the toe nailfold. Arteriovenous shunt flow was assessed using the technique of laser Doppler flowmetry, applied to the toe pulp. Fourteen diabetic patients with peripheral and autonomic neuropathy, 11 with no clinical evidence of neuropathy and 14 normal subjects were studied. Laser Doppler flowmetry (predominantly arteriovenous shunt flow) was increased more than three-fold ( p <0.01) in the diabetic patients with neuropathy compared to control subjects, (median 3.57, interquartile range 2.00–5.32 volts vs median 0.93, interquartile range 0.47–2.36 volts respectively). There was no evidence of skin capillary closure. The calculated capillary blood flow (erythrocyte flux) was significantly increased in the diabetic neuropathic patients compared to control subjects (median 76.4, interquartile range 34.4–109.8 picolitres/s vs median 23.2, range 8.0–44.8 picolitres/s, p <0.01). This study demonstrates that foot skin capillary blood flow is increased in diabetic patients with neuropathy. There is, therefore, no evidence to support the supposition that capillary ischaemia, either secondary to a “capillary steal phenomenon” or “advanced microangiopathy”, is a feature of diabetic neuropathy under resting conditions.

Takeda, H.; Kishikawa, H.; Shinohara, M.; Miyata, T.; Suzaki, K.; Fukushima, H.; Ichinose, K.; Shichiri, M.

doi: 10.1007/BF00278748pmid: 3069533

125 31 31 9 9 H. Takeda H. Kishikawa M. Shinohara T. Miyata K. Suzaki H. Fukushima K. Ichinose M. Shichiri Department of Metabolic Medicine Kumamoto University Medical School Kumamoto Japan Summary The role of epinephrine in platelet activation and the effect of an α 2 -adrenoceptor antagonist, midaglizole, during insulin-induced hypoglycaemia in Type 2 (noninsulin-dependent) diabetes mellitus were examined. The action of midaglizole as a platelet α 2 -antagonist was confirmed by in vitro studies using platelet-rich plasma and washed platelet suspension. Hypoglycaemia was induced by a bolus injection of short-acting insulin in 24 diabetic patients. They were divided into two groups, a control group ( n =12) and an α 2 -group ( n =12), and midaglizole was administered orally 60 min before insulin injection in the latter. Blood glucose and plasma C-peptide levels were significantly decreased ( p <0.005) by insulin injection in both groups. Counter-regulatory hormones, including epinephrine, and arginine vasopressin were similarly increased at the hypoglycaemic nadir compared with the levels at 0 min in both groups. Plasma β-thromboglobulin was increased at the hypoglycaemic nadir (165.5±12.6ng/ml) compared with the level at 0 min (121.0±11.5, p <0.005) in the control group, whereas no significant increase was demonstrated in the α 2 group. These results suggest that plasma epinephrine plays an important role in platelet activation during hypoglycaemia in Type 2 diabetes mellitus, and that the platelet activation is prevented by α 2 -adrenoceptor antagonist.

Wallensteen, M.; Dahlquist, G.; Persson, B.; Landin-Olsson, M.; Lernmark, Å.; Sundkvist, G.; Thalme, B.

doi: 10.1007/BF00278749pmid: 3069534

125 31 31 9 9 M. Wallensteen G. Dahlquist B. Persson M. Landin-Olsson Å. Lernmark G. Sundkvist B. Thalme Department of Paediatrics, Karolinska Institute St. Göran's children's Hospital Stockholm Sweden Department of Paediatrics, Karolinska Institute Sachs' Children's Hospital Stockholm Sweden Department of Paediatrics, Karolinska Institute Huddinge Hospital Stockholm Sweden Department of Internal Medicine, University of Lund Malmö General Hospital Malmö Sweden Summary The pattern of fall in B-cell function measured as plasma and 24 h urinary C-peptide excretion, as well as levels of islet cell antibodies, insulin antibodies and metabolic parameters, were followed for two years in 39 children aged 1–17 years prospectively from clinical onset of Type 1 (insulin-dependent) diabetes. At onset 32/36 patients had measurable plasma C-peptide (median 0.13 nmol/l). Maximum values of fasting and postprandial plasma C-peptide were reached at a median duration of three months. Thereafter both plasma and urinary C-peptide declined linearly. The median value of the rate of fall in postprandial plasma C-peptide was 0.019 nmol·1 −1 ·month −1 . Age at onset was positively correlated to the maximum value of postprandial plasma C-peptide in each patient (r s =0.57, p =0.0001) and throughout the observation time positively correlated to fasting and postprandial C-peptide and to the 24 h urinary C-peptide excretion (r s range 0.35–0.70, p =0.03–0.0001). The rate of fall of postprandial C-peptide was unrelated to age at onset and was strikingly parallel in different age groups. Islet cell antibodies were present in 87% of the patients at onset and decreased to 38% at 24 months. Islet cell antibody litres were not correlated to age at onset or to plasma or urinary C-peptide at any single observation. However, islet cell antibody negative patients had significantly higher ( p <0.05) postprandial plasma C-peptide values at 1, 9, and 12 months of duration, compared to islet cell antibody positive patients. Insulin antibodies and metabolic state at onset did not influence the C-peptide values. It is concluded that age at onset is the most important variable in predicting the duration and magnitude of endogenous insulin secretion during the first two years of Type 1 diabetes in children.

Wilkin, T.; Mirza, I.; Armitage, M.; Casey, C.; Scott-Morgan, L.

doi: 10.1007/BF00278750pmid: 3234642

125 31 31 9 9 T. Wilkin I. Mirza M. Armitage C. Casey L. Scott-Morgan Endocrine Section, Department of Medicine II General Hospital Southampton UK Summary Insulin autoantibodies, like islet cell antibodies, are found not only in the sera of newly diagnosed Type 1 (insulin-dependent) diabetic patients and their relatives, but also in patients with other autoimmunities who do not develop diabetes. Insulin autoantibodies are oligo/monoclonal and frequently binding-site restricted. As determinant selection is genetically determined, we questioned whether certain polymorphisms of insulin autoantibodies, identified by their binding site on the insulin molecule, could better discriminate for Type 1 diabetes, which is also HLA determined. First, we raised monoclonal antibodies to human insulin by classic fusion methods in order to determine the range of antibody polymorphism, and identified five distinct types by their binding profiles to a panel of insulin variants, using an enzyme-linked immunosorbent assay. Two of these polymorphisms, type A and type B, were subsequently found in insulin autoantibody positive human sera using the same panel of insulin variants, and successfully distinguished diabetes-related from diabetes-unrelated individuals. Thus, the type B polymorphism was responsible for binding in 60% of 41 insulin autoantibody positive individuals with polyautoimmune disease but no personal or family history of diabetes (diabetes unrelated), but in only 2% of a group which comprised 17 newly-diagnosed insulin autoantibody positive Type 1 diabetic patients, 19 insulin autoantibody positive discordant twins of Type 1 diabetes and six insulin autoantibody positive healthy siblings of Type 1 diabetic patients (diabetes related) ( p <0.01). Isolation of the type A polymorphism alone reduced the proportion of false negatives in the insulin autoantibody test for diabetes relatedness from 49% to 20% without diminishing its specificity. Thus, insulin autoantibody polymorphisms are more discriminating than the ‘nominal’ antibody, due possibly to linkage between immune response genes determining response to the type A epitope on the one hand, and susceptibility to Type 1 diabetes on the other.

Issa-Chergui, B.; Goldner-Sauvé, A.; Colle, E.; Prud'homme, G.; Lapchak, P.; Meide, P.; Seemayer, T.

doi: 10.1007/BF00278751pmid: 2853088

125 31 31 9 9 B. Issa-Chergui A. Goldner-Sauvé E. Colle G. J. Prud'homme P. H. Lapchak P. M. van der Meide T. A. Seemayer McGill University-Montreal Children's Hospital Research Institute Canada Department of Pathology McGill University Montreal Canada Primate Center TNO Rijswijk The Netherlands Summary A study of Class I and II major histocompatibility complex gene product expression by a rat insulinoma cell line (RINm5F) was performed using monoclonal antibodies and immunoperoxidase techniques. RINm5F cells were incubated with different concentrations of gamma interferon. RINm5F cells exhibit low levels of Class I molecules and are normally devoid of Class II gene products. Upon exposure to gamma interferon, RINm5F cells showed a dramatic increase in Class I expression. This expression was homogenous and could be detected on all cells after 18 h of incubation with as little as 1 unit/ml of interferon. In contrast, de novo Class II expression was not homogeneous and required 36 h of incubation with 10 units/ml of interferon. The number of RINm5F cells expressing Class II antigens was dose- and time-dependent. Interferon treatment did not affect the morphology of RINm5F cells as determined by ultrastructural analysis. Withdrawal of interferon from the culture medium for as long as 78 h diminished but did not abolish the expression of Class I and Class II molecules already induced. The ability of interferon to enhance expression of Class I gene products and induce de novo expression of Class II molecules on B-cell-derived RINm5F cells supports the hypothesis that aberrant expression of major histocompatibility complex gene products on pancreatic B cells may be an important factor in triggering the immune response in Type 1 (insulin dependent) diabetes mellitus.

Koltai, M.; Rösen, P.; Hadházy, P.; Ballagi-Pordány, Gy.; Köszeghy, A.; Pogátsa, G.

doi: 10.1007/BF00278752pmid: 2853089

125 31 31 9 9 M. Z. Koltai P. Rösen P. Hadházy Gy. Ballagi-Pordány A. Köszeghy G. Pogátsa National Institute of Cardiology Budapest Hungary Diabetes Research Institute Dusseldorf FRG Department of Pharmacodynamics Semmelweis Medical School Budapest Hungary Summary Before the onset of histologically detectable alterations of diabetic arteries, a considerable decrease of vasodilation ability develops. The role of an altered prostaglandin biosynthesis in this phenomenon was investigated in connection to the altered vascular adrenergic mechanisms. The effect of phenylephrine on prostacyclin production of isolated coronary arterial rings (100 μmol/l) as well as on conductivity of the coronary arterial bed (7.5-15-30-60 pmol·kg −1 ·min −1 ) were compared in 12 metabolically healthy and 12 alloxan-diabetic (560 μmol/kg) dogs. Furthermore, the effect of phentolamine (5 μmol/l) on the prostacyclin and thromboxane productions of the isolated vessels (coronary, femoral and basilar arteries) was investigated by radioimmunoassay. Although the basal prostacyclin amounts synthesized by healthy and diabetic coronary vessels were not different (5.1±1.6 and 4.9±1.4pg/mg vessel/30 min), similarly to femoral and basilar arteries, the diabetic arterial rings produced significantly ( p <0.05) more thromboxane than the control rings. The α-adrenergic blockade by phentolamine did not influence the prostacyclin production in the healthy arteries, but considerably ( p <0.05) increased it in the diabetic coronary arteries. Phentolamine normalised the thromboxane synthesis in the diabetic group ( p <0.01) and enhanced ( p <0.05) it in the metabolically healthy group. Phenylephrine was ineffective (98±6%) on the prostacyclin production in vitro versus the stimulated (150±22%) prostacyclin synthesis detected in the metabolically healthy group; and in vivo induced a more significant ( p <0.05) decrease in the coronary conductivity in diabetic than in control groups. These results refer to the supposition that altered adrenergic mechanisms are involved in the imbalamce of the vasoactive prostaglandins contributing to the high incidence of ischaemic heart disease in diabetes mellitus.

Vara, E.; Fernández-Martín, O.; García, C.; Tamarit-Rodríguez, J.

doi: 10.1007/BF00278753pmid: 3069535

125 31 31 9 9 E. Vara O. Fernández-Martín C. García J. Tamarit-Rodríguez Department of Biochemistry, Faculty of Medicine University Complutense Madrid Spain Summary Palmitate ability to modify D-(U- 14 C)glucose incorporation into different lipids (“de novo” synthesis), as well as sugar-stimulation of insulin release and 45 Ca 2+ -fluxes, was investigated in islets of fed and 48-h starved rats. The fatty-acid induced dose-dependent, correlative increments of insulin secretion, 45 Ca 2+ -influx and the “de novo” synthesis of each phospholipid fraction analysed at 20 mmol/l (but not 3 mmol/l) glucose. Omission of calcium reduced drastically ( p <0.001) insulin release and the “de novo” synthesis of neutral glycerolipids, leaving unaltered that of acidic phospholipids (phosphatidate and phosphoinositides). The increased synthesis of the latter is therefore not the consequence of stimulated secretion. It could initiate or contribute to maintain an increased turnover of islet phosphoinositides, thus generating some mediators of the calcium signalling system (inositol phosphates). Starvation led to a drastic reduction ( p <0.001) of both insulin secretion, “de novo” synthesis of each lipid fraction, and 45 Ca 2+ -influx in response to glucose and palmitate. The presence of a fatty-acid oxidation inhibitor (2-bromostearate or 2-tetradecylglycidate) prevented the effect of starvation on 45 Ca 2+ -influx, as it has been shown to do on insulin secretion and palmitate incorporation into islet lipids. It is finally suggested that palmitate might amplify the insulin secretory response of islets to glucose, through the stimulation of the “de novo” synthesis of phosphoinositides and the subsequent generation of inositol phosphates, which would contribute to accelerated calcium turnover.

Sodoyez-Goffaux, F.; Koch, M.; Dozio, N.; Brandenburg, D.; Sodoyez, J-Cl.

doi: 10.1007/BF00278754pmid: 3234643

125 31 31 9 9 F. Sodoyez-Goffaux M. Koch N. Dozio D. Brandenburg J-Cl. Sodoyez Department of Paediatrics University of Liege Belgium Department of Internal Medicine University of Liege Belgium German Research Institute, University of Aachen FRG Summary Human sera were tested for insulin antibodies by fluid and solid phase assays. Radioimmune titres determined with 125-I Tyr A14 insulin were not correlated with those obtained using insulin coated microplates and enzyme linked immunodetection ( n =60). Several reasons for this lack of correlation were found. Iodine substitution on the A14 residue of insulin may significantly alter the avidity of some insulin antibodies for their ligand; hence, disclosing a heretofore unsuspected pitfall for antibody determination by radio-immunoassay. Specificity for bovine insulin was easily demonstrable in fluid phase by comparing the binding of monoiodinated bovine, porcine and human insulin. By contrast, in solid phase assay, titres obtained with microplates coated with bovine or human insulin were almost equal, regardless of the serum specificity for bovine insulin. This lack of specificity of the solid phase assay is not due to denaturation or unavailability of the bovine specific epitope because: bovine specificity could be demonstrated by competitive assay, after preincubation of the serum with insulin of the different species; and, coating with crosslinked insulin dimers or oligomers instead of monomers did not unmask bovine specificity. It is concluded that radioimmune methods are best suited to study specificity but may be biased by the presence of the radioiodine label whereas solid phase assay detects low avidity antibodies with great efficiency but is less appropriate to study specificity.