P. E. Beales

NOD mouse colonies around the world- recent facts and figures

Non-obese diabetic (NOD) mice are commonly used in autoimmune research. However, the diversity of these mice in developing autoimmune disease under different conditions prompted a group of researchers to compile a questionnaire on this subject. Here Paolo Pozzilli and colleagues comment on the results of this survey.

Troglitazone prevents insulin dependent diabetes in the non-obese diabetic mouse

Troglitazone has recently been introduced in the treatment of Type 2 diabetes. In addition to its anti-diabetic effects it acts as a perixosome proliferator activated receptor-gamma (PPAR-gamma) agonist and has anti-inflammatory properties by inhibiting macrophage tumour necrosis factor-alpha (TNF-alpha) secretion. It also inhibits the production of endothelial selectin (e-selectin). Troglitazone also reduces interleukin-1alpha induced nitric oxide production in pancreatic beta-cells, which may be relevant in preventing nitric oxide mediated damage to these cells in the Type 1 diabetes process. We tested troglitazone in the spontaneous model of autoimmune diabetes, the non-obese diabetic (NOD) mouse, to determine its effect on the disease process. When administered by gavage from weaning at a dose of 400 mg/kg body weight (n = 32), troglitazone reduced the incidence of diabetes by 16 weeks compared to controls (n = 32) in a pattern that was maintained up to the conclusion of the experiment at 31 weeks of age (p < 0.05). Insulitis was unaltered (index = 1.05 +/- 0.71 vs. 1.13 +/- 0.82, treated vs. controls, p = 0.78). The study was repeated using troglitazone in the diet of NOD mice (n = 24) to give a dose of approximately 200 mg/kg body weight in order to provide a more consistent level of troglitazone during the time course of the experiment. There was a reduction of diabetes incidence in this group but it did not reach significance. Insulin levels were reduced in gavage treated mice although such reduction did not reach significance (p < 0.07). We conclude that, in view of its effect on this model of autoimmune diabetes and because of its known function as an insulin sensitiser, troglitazone might be considered for potential use in those patients with Type 1 masquerading as Type 2 diabetes.

A wheat-based, diabetes-promoting diet induces a Th1-type cytokine bias in the gut of NOD mice

Dietary antigens are candidate environmental factors in the pathogenesis of type 1 diabetes. In the non-obese diabetic (NOD) mouse, an animal model of type 1 diabetes, cereal-based diets promote disease development, whereas the diets based on hydrolysed proteins or non-diabetogenic proteins are protective. The hypothesis that diabetogenic diets modulate the cytokine balance in the gut was tested. NOD mice were fed with NTP-2000 (mainly a wheat-based milk-free diet) or Prosobee (a semi-purified hypoallergenic diet based on soy protein isolate) or Prosobee plus casein (milk protein fraction). The mRNA levels of IFN-gamma, IL-10, TNF-alpha, TGF-beta, and inducible NO synthase in the small intestine and the Peyers patches were determined by semi-quantitative RT-PCR. Mice fed on the cereal-based NTP-2000 diet expressed higher levels of the Th1-type and pro-inflammatory markers IFN-gamma, TNF-alpha, and inducible NO synthase mRNA compared to the Prosobee-fed animals. The expression of the counterregulatory cytokines IL-10 and TGF-beta was unaffected. This resulted in a significant bias of the intestinal cytokine balance towards T helper cell type 1 after feeding NTP-2000. The cytokine mRNA levels in the gut-associated Peyers patches were not affected. Thus, modulation of gut immunoreactivity by diet may contribute to disease development in NOD mice.

Thiazolidinediones for the prevention of diabetes in the non‐obese diabetic (NOD) mouse: implications for human type 1 diabetes

Thiazolidinediones (TZDs) are a recently introduced generation of drugs acting as receptor agonists to reduce insulin resistance and used currently in combination with other hypoglycaemic agents for the treatment of type 2 diabetes. In addition, TZDs possess anti‐inflammatory properties that make them of interest for reducing the T‐cell inflammation occurring in the islets in type 1 diabetes.

Histological study of pancreatic beta-cell loss in relation to the insulitis process in the non-obese diabetic mouse

Non-obese diabetic (NOD) mice spontaneously develop an autoimmune diabetes with higher incidence in females than in males. In order to elucidate possible factors involved in the different incidence of diabetes between male and female mice, we studied the progression of pancreatic beta-cell loss in relation to mononuclear cell infiltration of the pancreas (insulitis). We examined the pancreas of 76 NOD mice (39 males and 37 females) of different ages. The beta-cell content was assessed by immunoperoxidase staining of sections with an anti-insulin serum and the severity of insulitis was determined by haematoxylin staining of the same sections. A semi-quantitative criterion was used to grade both parameters. The results showed that females have a faster loss of beta-cell mass, which progressively decreases with the increase of severity of insulitis. In males, a medium to severe degree of insulitis is required before initial loss of beta cells occurs. Under the age of 10 weeks there was a significantly lower content of beta cells in females than males (2.84±0.03 vs 2.67±0.07; P=0.02). Since we never observed a significant difference in the degree of mononuclear cell infiltration in age-matched males and females, these data support the hypothesis of weaker beta-cell resistance to immunological attack in female mice. Thus beta-cell sensitivity, in addition to immunological activity, is an important factor in the pathogenesis of insulin dependent diabetes.

Raised temperature reduces the incidence of diabetes in the NOD mouse

SummaryAn association between the incidence of childhood Type 1 (insulin-dependent) diabetes mellitus and the average yearly temperature in different countries has been reported, the incidence being higher in countries with a lower mean temperature. We have studied the effect of environmental temperature on the incidence of diabetes in an animal model of Type 1 diabetes, the non-obese diabetic (NOD) mouse. Female NOD mice were divided at weaning, with one group placed at a higher temperature (mean 23.7±1.7° C) and the other at a lower temperature (21.0±1.8° C). At 20 weeks of age 6 of 16 mice at lower temperature and 1 of 17 mice at higher temperature had developed diabetes (p < 0.02); at 30 weeks 10 of 16 and 5 of 17 mice had developed diabetes (p < 0.05). Non-diabetic animals in the low temperature group had a higher food intake than those in the high temperature group between 13–15 weeks of age (28.0±1.2 g/week vs 24.8± 0.7 g/week, P < 0.05). In a parallel experiment, histological examination showed that there were similar degrees of insulitis in the high and low temperature groups at seven weeks of age. We conclude that environmental temperature can affect the incidence of diabetes in the NOD mouse and that this may be related to alterations in food intake.

CD95 and CD95-ligand expression in endocrine pancreas of NOD, NOR and BALB/c mice

Summary The non-obese diabetic (NOD) mouse is widely used to study the pathogenesis of insulin-dependent diabetes mellitus. However, the mechanisms responsible for beta-cell destruction, in this model, are still poorly defined. The CD95/CD95L system among other effector systems has been implicated in beta-cell death. In this study we investigated in NOD, non-obese resistant (NOR) and Balb/c mice the expression of CD95 and CD95L in alpha and beta pancreatic cells by immunohistochemistry and immunofluorescence. We demonstrate that alpha cells in the islets of Langherans constitutively express CD95L forming a natural shield around beta cells. [Diabetologia (1997) 40: 1476–1479]

Labelling of lymphocytes with indium 111 oxine: effect on cell surface phenotype and antibody-dependent cellular cytotoxicity.

Indium 111 oxine is currently used to label peripheral lymphocytes in order to study the kinetics of these cells in vivo. Since the quantity of radioisotope for labelling is still a matter of controversy, we have investigated in vitro the effect of increasing the concentration of indium 111 oxine on the lymphocyte surface phenotype and the antibody-dependent cellular cytotoxicity (ADCC) using lymphocytes from normal subjects. The cell surface phenotype, as evaluated by 2 monoclonal antibodies, was not affected whereas ADCC, at any of the doses used, was significantly reduced compared to the baseline value. The implications of these results for the use of indium 111 oxine for the in vivo studies are discussed.

Tolbutamide reduces the incidence of diabetes mellitus, but not insulitis, in the non-obese-diabetic mouse.

SummaryThe functional state of beta cells may influence the rate of their destruction in Type 1 (insulin-dependent) diabetes mellitus. We examined the effect of diazoxide, which inhibits insulin secretion, or tolbutamide, which stimulates insulin secretion, upon the incidence of diabetes in the non-obese-diabetic (NOD) mouse. Female mice were treated from 3–30 weeks of age with diet containing diazoxide 250 mg·kg−1 or tolbutamide 125 mg·kg−1. The cumulative incidence of diabetes at 35 weeks was similar in the diazoxide (16 of 24) and control (18 of 24) groups, but reduced in the tolbutamide group (10 of 23,p<0.04 vs control group). In a second experiment, treatment was started from 9 weeks of age, by which time insulitis is already present. The cumulative incidence of diabetes at 35 weeks was 16 of 24 in controls, 15 of 24 on diazoxide and 11 of 24 on tolbutamide (p=NS vs control). A third experiment compared the effect of treatment from 3 weeks with control diet or diet containing tolbutamide 125 mg·kg−1 or 500 mg·kg−1. Diabetes was reduced by tolbutamide treatment, with a cumulative incidence of 25 of 31 in controls, 18 of 30 on tolbutamide 125 mg·kg−1 (p<0.04) and 14 of 32 on 500 mg·kg−1 (p<0.002), although the difference between the two treatment groups failed to reach statistical significance. A fourth experiment showed that treatment from 3–12 weeks with diazoxide 1000 mg·kg−1 increased the extent of insulitis compared with controls and animals treated with tolbutamide 500 mg·kg−1. Elucidation of the mechanisms by which tolbutamide reduces the incidence of diabetes in the NOD mouse has implications for human intervention trials.

Reduced cumulative incidence of diabetes but not insulitis following administration of chimeric human IL-15–murine IgG2b in NOD mice

It has been recently demonstrated that apoptosis is involved in beta‐cell destruction in the NOD mouse model of diabetes. The aim of the present study was to investigate whether IL‐15, a cytokine involved in the modulation of the apoptotic process, is capable of modifying the natural history of diabetes and/or insulitis in pre‐diabetic NOD mice. The rationale for the use of IL‐15‐IgG2b recombinant cytokine is related to its long half‐life (28 ± 4 h).