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B Ziegler

Publications and source records attributed to B Ziegler.

At least 19 recordsLinked to original sources
PubMed1992

Monoclonal antibody-mediated cytotoxicity against rat beta cells detected in vitro does not cause beta-cell destruction in vivo.

Two monoclonal Beta-cell surface antibodies M10H6 und K14D10 were obtained by fusion of spleen cells of Balb/c mice with the myeloma cell line P(3)0. The monoclonal antibody M10H6 was induced by immunization with rat insulinoma cells finally boostered with disintegrated rat islets, whereas the K14D10 was generated after immunization with porcine proinsulin. Both monoclonals belong to the IgG2A isotype and were screened with insulin-producing rat insulinoma cells by an indirect immunofluorescence test as well as by a cellular enzyme linked immunosorbent assay. In addition to the cell surface binding on living Beta cells the monoclonals react with islets on cryostat sections of rat pancreas. The anti-islet cytotoxic potential of these monoclonals was measured by 51Chromium-release in the presence of complement or Fc-receptor bearing leucocytes using 51Chromium-labelled rat islet cells as target. Both antibody secreting hybridomas were propagated in syngeneic mice resulting in high levels of islet cell surface antibodies in ascites and sera from the recipient. High anti-islet cytotoxicity was mediated by ascites fluid, but no mouse developed hyperglycaemia. Furthermore, the repeated injections of the monoclonals into rats did not exert a diabetogenic action and failed to reduce the pancreatic insulin content although the attraction of the K14D10 to the pancreatic islets in vivo could be demonstrated. We conclude that islet cell surface antibody-mediated Beta-cell lysis in vitro may not be relevant to Beta-cell destruction in vivo.

Animals↗
PubMed1992

Characterization of two porcine proinsulin reactive monoclonal antibodies by immunostaining of beta-cells in pancreatic sections of different species.

Murine monoclonal antibodies against porcine proinsulin were generated by somatic cell hybridization. As detected by radioimmunoassay, 2 monoclonal antibodies KSPI14D4 and KSPI13G10 showed a strong binding to 125I-labelled porcine proinsulin but not to insulin. The species specificity of these 2 monoclonals was found to be different as shown by indirect immunofluorescence using sections of Bouin-fixed pancreata of different species. The KSPI14D4 recognized the proinsulin of pig, mouse, man, cattle, rat, dog, and cat but not that of guinea pig, whereas the KSPI13G10 bound to porcine proinsulin only. From these results it is concluded that KSPI14D4 effectively recognizes a wide-spread epitope located in one of the insulin-C-peptide junctions of the proinsulin molecule, whereas KSPI13G10 is directed to a species-specific epitope of the porcine connecting peptide.

Animals↗
PubMed1992

Detection of antibodies against both isoforms of glutamate decarboxylase in BB/OK rats by western blotting and immuno trapping enzyme activity assay.

The GABA-producing enzyme glutamate decarboxylase (GAD) is a prominent autoantigen in insulin-dependent diabetes mellitus (IDDM). Autoantibodies against GAD were found with a high prevalence in IDDM patients and in animal models for IDDM. The aim of this study was to detect autoantibodies against both isoforms of GAD in diabetic and non-diabetic but diabetes-prone BB/OK rats by Western blotting and to test their specificity to GAD by an immuno-trapping enzyme activity assay. Eighteen diabetic and 18 non-diabetic BB/OK rats (age 121 +/- 20 days) were investigated. In 10/18 (56%) of the diabetic and 13/18 (72%) of the non-diabetic BB/OK rats autoantibodies against at least one GAD-isoform were detected by Western blotting. In the immunotrapping enzyme activity assay, the mean value of the diabetic (1151 +/- 552 cpm, n = 11) and nondiabetic BB/OK rats (1978 +/- 1213 cpm, n = 10) was significantly (p < 0.01) increased compared to the LEW. 1A control rats (581 +/- 274 cpm, n = 12). 7/10 (70%) individual sera of the non-diabetic and 5/11 (45%) of the diabetic BB/OK rats were positive in this test. In conclusion, the prevalence of GAD autoantibodies in BB/OK rat is connected with the genetic susceptibility to IDDM but is not a predictor for the onset of the disease in BB/OK rats.

Animals↗
PubMed1992

[Cerebral multisystem atrophy in a patient with depressive hallucinatory syndrome. A case report].

In a female patient, aged 47 years at the beginning, the successive appearance of extrapyramidal signs preceded by depression, paranoid-hallucinatory psychosis, autonomic and cerebellar dysfunction was followed up over a period of 8 years. Autopsy revealed--in accordance with the clinical symptomatology--both olivo-ponto-cerebellar atrophy and a striatonigral degeneration. As a rule these changes are accompanied by dementia. The reported case is unusual in having first presented with severe depression and paranoid-hallucinatory symptoms.

Brain Stem↗
PubMed1991

Influence of target cell preparation on binding of monoclonal islet cell reactive antibodies (mc-ICRA) in cellular enzyme-linked immunosorbent assay (CELISA).

A rapid, effective and sensitive CELISA for the detection of monoclonal islet cell reactive antibodies (mc-ICRA) using the insulin-producing rat insulinoma cell line (RIN) is described. RIN cells are a suitable target for this monoclonal antibody assay as shown by a comparative study with normal rat islet cells. We tested the influence of the target cell preparation and obtained the best sensitivity and reliability with the CELISA using desiccated cells or desiccated cell homogenate with a cell number of 5 x 10(4) cells per well rather than an adsorbed cell homogenate. Furthermore, ethanol fixation of RIN cells resulted in a loss of antigenicity as shown particularly by the detection of islet cell surface antibodies. We also compared the binding of mc-ICRA in RIN-CELISA with data obtained by indirect immunofluorescence using viable RIN cells as targets. By permeabilization of the cell membrane by desiccation or sonication, more antibodies are detected in CELISA (surface and cytoplasmic antibodies), whereas in immunofluorescence on viable RIN cells, only surface reactive antibodies are detected.

Animals↗
PubMed1991

Characterisation of glutamine uptake in rat liver mitochondria.

Glutamine is taken up by rat liver mitochondria in an electroneutral manner with a Km of 3.3 mM and a Vmax of 33 nmol x min-1 x mg-1 at 10 degrees C. The uptake is driven by the mitochondrial pH/cytosolic pH difference in isolated mitochondria, as well as in the intact rat liver. The rate of uptake is stimulated at a more alkaline matrix pH due to a stimulation of mitochondrial glutaminase. Our data confirm the notion that glutamine metabolism is regulated by pH, not only at the site of its metabolism but also through regulation of its transport systems.

Animals↗
PubMed1991

Glucose tolerance behaviour before the onset of type I (insulin-dependent) diabetes in young people as a predictor of the further course of the disease: a retrospective analysis of 33 cases.

A study was made of glucose tolerance and insulin secretion in 33 persons who later developed insulin-dependent diabetes (aged 4-24 years) and observation continued further in the first years after manifestation. Patients who developed the typical labile type of diabetes were of normal weight and had either normal glucose tolerance tests before diagnosis or had impaired glucose tolerance (IGT) for a short interval of 2-16 months. Subjects with IGT over a significantly (p less than 0.01) longer period of 32.30 +/- 6.25 (normal body weight) or 94.71 +/- 20.62 (obese) months developed a milder form of diabetes with retarded insulin dependency in obese subjects. The severe and mild form of IDDM are distinct with respect to insulin requirement (0.75 +/- 0.03 or 0.28 +/- 0.04 U/kg b.w., P less than 0.01) and glucagon stimulated C-peptide (0.18 +/- 0.05 or 1.41 +/- 0.27, P less than 0.01) in the first 2.5-3.5 years after onset. The two forms were not different regarding HLA-DR antigens. Islet cell surface antibodies investigated in 15 probands at 27 occasions before diabetes onset had no prognostic value. The development of a mild form of IDDM may be expected in cases with pre-existing IGT for more than one year. The insulin secretion is of low predictive value under these conditions. The observation is of practical use and theoretical interest.

Adolescent↗
PubMed1991

Polymorphism of insulin antibodies in six patients with insulin-immune hypoglycaemic syndrome.

Insulin antibodies in six patients with immune hypoglycaemic syndrome were studied. The antibodies displayed a higher affinity for bovine insulin in two patients, were specific for human insulin in one patient and non-species specific in the other three patients. The predominant IgG subclass of the insulin antibodies was IgG4 in two patients, IgG3 in two and IgG1 in two. In one of these, the other three subclasses were also detectable. Insulin autoantibodies of four patients were homogeneous with regard to light chains (kappa), and those of the other two contained both kappa and gamma light chains. Analysis of insulin immune complex size by fast protein liquid chromatography was possible in three patients and demonstrated immune complexes with elution profile close to that of IgG, although not exactly superimposable to the one obtained with a mouse monoclonal insulin antibody. In two patients, avidity was too low to permit chromatography of the immune complexes, and, moreover, in these two cases insulin antibodies were of the IgG3 isotype and spontaneously formed aggregates independently of insulin binding. We conclude that insulin antibodies of the insulin immune syndrome are polymorphic but different from those generated by insulin therapy.

Adult↗
PubMed1991

[Production of monoclonal antibodies in serum-free medium in dialysis tubing].

A simple procedure for the production of pure monoclonal antibodies (mab) in dialysis tubing has been used. Hybridomas which produce pancreatic islet cell reactive monoclonal antibodies were grown in dialysis tubing containing serum free medium. The dialysis tubing was inserted into a flask with medium containing 7.5% foetal calf serum. The flask was placed on a roller and medium was changed every two days. The optimal time for harvesting the mab could be shown to be after 10 days of culture with a 4 fold increased immunoglobulin concentration in comparison to a conventional hybridoma culture. Immunoglobulin concentrations up to 110 mg/l and cell yields of 1.3 x 10(6)/ml have been obtained. The low concentration of contaminating low molecular weight proteins in the supernatant facilitated or saved purification of mab.

Animals↗
PubMed1991

CELISA for rapid screening of monoclonal islet cell surface antibodies using living rat insulinoma cells as target.

An improved rapid cell enzyme-linked immunosorbent assay (CELISA) is described which is suitable for the large scale screening of monoclonal antibodies to islet cell surface antigens. 5 x 10(4) insulin-producing rat insulinoma (RIN) cells were seeded per well in a 96-well flat-bottomed polystyrene plate coated one day before a 0.01% poly-D-lysine solution in PBS. After culture for 4 days in 200 microliters/well RPMI 1640 supplemented with 7.5% heat-inactivated fetal calf serum, the cell number per well was up to 2.1 x 10(5). These monolayer RIN cell cultures were used as a target for the detection of islet cell surface antibodies (ICSA) in the supernatants of hybridomas. The cells were used without fixation to avoid modification of sensitive surface antigens. Poly-D-lysine did not cause non-specific binding of immunoglobulins to the plastic wells as tested with irrelevant monoclonals. The specificity and sensitivity of the method is comparable to indirect immunofluorescence. All mc-ICSA primary screened by indirect immunofluorescence using viable RIN cell suspensions were positive in this CELISA. There was a correlation (r = 0.7; n = 44) between the antibody binding measured by CELISA and the indirect immunofluorescence technique. The advantage of this CELISA is that cell surface structures are well preserved in a viable cell monolayer used as target without chemical fixation. This assay procedure should be generally suitable for the initial screening of monoclonal antibodies to cell surface antigens of cells growing under culture conditions.

Animals↗
PubMed1991

Different multiple reactivity of monoclonal islet cell binding antibodies using indirect immunofluorescence technique on viable cells or cellular ELISA on desiccated cells as target.

Two commonly used methods for screening hybridoma supernatants secreting monoclonal islet cell reactive antibodies (mc-ICRA) were performed to investigate the specificity of the monoclonals established. For this, endothelial, neuroblastoma, murine subcutis and two myeloma cell lines were used as targets in comparison to the insulin-producing rat insulinoma cell line (RIN), either immobilized and permeabilized in cellular enzyme linked immunosorbent assay (CELISA) or in suspension of viable cells in the indirect immunofluorescence test. In addition, rat splenocytes were used for estimating multireactivity of mc-ICRA in ELISA. Using permeabilized target cells, we obtained a high multireactivity of the monoclonal antibodies (mab) tested, indicating a high incidence of molecular mimicry between cytoplasmic antigens of different cell lines. In contrast to CELISA, if only cell surface antigens of viable cells are accessible, detected by the immunofluorescence technique, the high multireactivity is not observed. For investigating the specificity of monoclonals, the complexity of target antigens used must be taken into consideration.

Animals↗
PubMed1991

[Scopolamine poisoning as a cause of acute paranoid hallucinatory psychoses].

In 1988 two patients intoxicated with scopolamine were treated in the department of psychiatry. Clinically an acute paranoid-hallucinatory psychosis was to be observed. Similar cases were reported from other parts of the country, partly proved by toxicological findings. In the cases seen by us scopolamine was applicated secretly in drinks and food with criminal intention. Diagnostic, therapeutic and historical aspects of intoxication with scopolamine are discussed.

Adult↗
PubMed1990

Development of cytotoxic islet cell antibodies in rats following damage of the pancreas by complete Freund's adjuvant combined with a nondiabetogenic dose of streptozotocin.

The possible relationship between destruction of pancreatic beta cells and islet cell surface antibodies (ICSA) was examined in a rat model using complete Freund's adjuvant (CFA), a lymphocyte activator, in combination with the beta cell toxin, streptozotocin (STZ). In addition to this treatment, the rat insulinoma cell line, RIN5AH, as a readily accessible source of insulin-producing cells, was utilized to potentiate the production of ICSA. Intraperitoneal injections of CFA to male Lewis rats, followed 24 h later by a single nondiabetogenic dose of STZ, produced a 47% (p less than 0.01) reduction in pancreatic insulin content associated with degranulation and necrosis of insulin-immunoreactive cells. Eight weeks after treatment, ICSA were detectable that mediated the complement-dependent lysis of neonatal rat islet cells. Injections of RIN5AH cells, following treatment with CFA/STZ, did neither increase the severity of histopathological changes in the exocrine pancreas nor the extent of beta cell necrosis, but gave rise to higher levels of cytotoxic ICSA. Immunization with RIN cells alone, although increasing ICSA levels above those of the other experimental groups, produced no major histopathological changes. These results indicate that ICSA are the consequence of beta cell damage, and they are not capable of promoting or initiating beta cell necrosis in this model.

Animals↗
PubMed1990

Survival of islet isografts despite cytotoxicity against pancreatic islets measured in vitro.

In this study the in vivo relevance of spleen cell anti-islet cytotoxicity measured in vitro was examined by transplantation of 1,200 syngeneic islets into the spleen of rats receiving 0.5 ml complete Freund's adjuvant (CFA) 24 h before 25 mg/kg body weight streptozotocin (STZ) was given. Control rats receiving CFA or STZ alone remained normoglycaemic whereas 12 out of 21 CFA/STZ-treated rats developed a severe hyperglycaemia after three combined treatments. After the first and second combined treatment splenocytes showed a significant cytotoxicity (p less than 0.01) against syngeneic islets measured by 51Cr-release. This cytotoxicity was not detectable after the third combined treatment. The CFA/STZ-induced diabetes with a residual pancreatic insulin content of only 5% was permanently reversed by intrasplenic islet isografts, but, surprisingly, syngeneic islets survived too, if transplanted at the time when an anti-islet cytotoxicity was measured in vitro. From our results we conclude that the polyclonal activation by complete Freund's adjuvant potentiates the beta cell-toxic effect of a low dose of streptozotocin and induced a transient splenocyte-mediated anti-islet cytotoxicity not recurrent after islet transplantation. Furthermore, our findings reveal a discrepancy between organ-specific immune reactions measured in vitro and those affecting the beta cells in vivo.

Animals↗
PubMed1990

Histopathological lesions in the pancreas of a rat model of diabetes induced with complete Freund's adjuvant and low-dose streptozotocin.

Neither injection of complete Freund's adjuvant (CFA) alone nor the administration of low doses of streptozotocin (STZ) to rats produced remarkable histopathological changes in the endocrine pancreas, but treatment with the combination of both resulted in necrosis of beta cells. When the combination of CFA/STZ was given two times, necrosis progressed, and the beta cell reserve was depleted to such an extend that persistent hyperglycemia ensued. These changes were associated with a significant reduction in the apparent islet size. A single injection of CFA induced pancreatitis and inflammatory lesions in the exocrine parenchyma with no insular involvement. Three injections caused extensive destruction of pancreatic acinar tissue but only moderate beta cell injury in the minority of islets. Apart from mild degranulation of beta cells, treatment with STZ did not produce histopathological changes in the pancreas. These results suggest that the acute inflammatory process induced by CFA may initially damage the beta cells, increasing thereby their susceptibility to the action of STZ.

Animals↗
PubMed1990

Autoimmune reactions in a patient with malignant insulinoma treated by multiple low dose streptozotocin.

A 66-year-old female patient with a malignant insulinoma was treated with streptozotocin (STZ; Zanosar) in 5 cycles every 4 weeks as 5 day courses with an intravenous dosage of 850 mg per day. Under this treatment hypoglycemic episodes decreased continuously in number as well as severity and - after a delay of 12 months after the last treatment - an overt diabetes mellitus appeared. Plasma insulin concentrations dropped immediately after starting of STZ therapy. On the other hand, islet cell surface antibodies and their complement-dependent cytotoxicity increased continuously, being at their highest 6 months after termination of STZ treatment. Thus, STZ is able to induce a specific immune response against islet cells with a progressive damage of malignant insulin producing cells.

Adenoma, Islet Cell↗
PubMed1990

The possibility that pancreatic beta cell destruction leading to type 1 diabetes is initiated by the release of cytokines by polyclonal activation of the immune system.

The cause of the destruction of the insulin-producing beta cells leading to type 1 diabetes is still unknown. Over the last few years it has become clear that autoimmune abnormalities, such as insulitis, autoantibodies against islet cell antigens and cellular cytotoxicity to beta cells are associated with the onset of type 1 diabetes. Nevertheless, it is still uncertain whether immune phenomena observed in human beings correspond to primary or secondary events in the development of type 1 diabetes. We do not know why the process of becoming diabetic is so lengthy in a risk proband with a genetic and immunological predisposition. Whatever the explanation of genetic association to the pathogenesis of type 1 diabetes may be, it seems sure that environmental factors may decisively influence the outcomes. In this article we summarize evidence implying that cytokines released during polyclonal activation of the immune system may initiate the beta cell destruction leading via autoimmune mechanisms of beta cell killing to insulin-dependent diabetes mellitus.

Animals↗