Prevention of the cytotoxic effect of IL-1 by human lysozyme on isolated rat islets.

Macrophages are present in the initial phase of the autoimmune process involved in the destruction of the endocrine pancreas in IDDM via the secretion of cytokines such as IL-1 beta. Macrophages also secrete lysozyme. Besides its action on the bacterial cell wall, lysozyme has an important physiological and immunological role. Human lysozyme is an in-situ modulator of the inflammatory reactions. We investigate the protective role of human lysozyme in vitro against the cytotoxic effect of IL-1 beta or of IL-1 beta combined with IFN-gamma on isolated rat islets. Precultured newborn rat islets were incubated with human or chicken lysozyme (50.000 U/ml) over 3 days. Human IL-1 beta (100 U/ml) or IL-1 beta (5 U/ml) + INF-gamma (100 U/ml) was added for the last 2 days and tritiated thymidine for the last 24 hrs. In another set of experiments, islets were exposed simultaneously to human lysozyme and IL-1 beta. Only pretreatment with human lysozyme abolished the lowering of the labelling index of the islet cell induced by IL-1 beta or by IL-1 beta and INF-gamma. Pycnotic nuclei were abundant in islets treated with IL-1 alone while they were not when islets were pretreated with human lysozyme. Chicken lysozyme had no protective effect in the same protocol. Human lysozyme was not protective when applied simultaneously with IL-1. Pretreatment of the islets by human lysozyme does not prevent the reduction of the insulin secretion induced by IL-1 beta. Human and chicken lysozyme differ further in their action when tested on fibroblasts proliferation. Only human lysozyme stimulates the latter. In conclusion, only human lysozyme seems to have a protective effect against the cytotoxicity of IL-1 in combination or not with IFN-gamma on islet cells in vitro. Moreover, to be protected, the islets have to be pretreated with lysozyme before the IL-1 application. Our in vitro results imply that natural aspecific immunity and its relation to the secretory function of the macrophage might be crucial for the prevention of the initial assault responsible for the onset of the immune process leading to insulin dependent diabetes.

[Importance of diets and their effect on fetal development: function and structure of the endocrine pancreas following protein deficiency during intrauterine life]. / Importance des régimes et leur influence sur le développement foetal: fonction et structure du pancréas endocrine suite à une carence en protéines au cours de la vie intra-utérine.

When an isocaloric low protein diet (8% versus 20%) is administrated to rats during gestation, the fetus or the neonate has a lower body weight and the structure and function of the endocrine pancreas is altered: islet cell proliferation, islet size and islet vascularisation are reduced when compared with a control group. If these fetal islets are cultured during 7 days in vitro, they secrete less insulin than normal islets in response to an AA challenge. When these newborns are fed with a low protein diet until adult age and analyzed at 70 days, the same alterations of the endocrine pancreas persist: the in vitro insulin secretion of the isolated islets in response to AA is dramatically depressed although their response to glucose is normal. However in vivo glucose and insulin levels in response to oral glucose challenge are abnormal. In addition, permanent functional alterations seem to persist when induced in utero. A normal diet (20% protein) given from birth to adulthood does not restore a normal insulin response in vivo to an oral glucose challenge.

Islet function in offspring of mothers on low-protein diet during gestation.

A low-protein diet (8 vs. 20%) administered during pregnancy affects the structure and function of the endocrine pancreas of the offspring. At 21.5 days of gestation, we reported a reduction of cell proliferation, islet size, islet vascularization, and pancreatic insulin content. In this study, we demonstrated an impairment of insulin secretion of these fetal islets when stimulated in vitro with amino acids such as arginine and leucine. If the offspring is kept on the same low-protein diet during suckling, weaning, and adulthood, fasting insulin levels remain low in the presence of normal blood glucose levels. Glucose tolerance at 70 days is impaired, with lower insulin response. In addition, permanent functional damage seems to be induced in utero by a low-protein diet, because a normal diet given from birth to adulthood does not restore normal insulin response after a glucose challenge. Our experimental results stress the impact of a balanced diet with qualitative and quantitative amino acid composition for the fetal endocrine pancreas to develop normally, without lasting functional and structural consequences in adulthood.

The development of the fetal rat intestine and its reaction to maternal diabetes. I. Morphometrical analysis in normal conditions.

In rodents, the fetal intestine develops rapidly during the last 5 days of gestation. The present investigation describes the events which occur in the duodenum, jejunum and ileum of fetal Wistar rat from day 16.5 to 21.5. The first villi and microvilli as well as endocrine cells already appear at 17.5 days in the duodenal mucosae. Goblet cells are detected at 18.5 days. The structure of the intestinal mucosa at 21.5 days is similar to that of adults. The evolution was quantified by morphometric analysis. The external and inner circumference, the length of the villi profile and the increased absorption area due to the villi profile were measured. We demonstrated that the total enlargement of the luminal surface area due to the villi and the microvilli in the duodenum of the fetus at 21.5 days is similar to that in the adult duodenum. This morphometric analysis could be used to detect possible disturbances in the development of the fetal intestine.

The development of the fetal rat intestine and its reaction to maternal diabetes. II. Effect of mild and severe maternal diabetes.

Diabetes during pregnancy induces specifically structural and functional changes in the fetal endocrine pancreas. Other organs are affected as well. In this study, the fetal intestinal tract which is in close connection with the endocrine pancreas was analysed during diabetic gestation. The disease was induced by two different doses of streptozotocin which led to a mild or severe diabetic state in the mother. In fetuses from mildly diabetic as well as from severely diabetic rats, the time sequence in the appearance of the differentiated cells was identical and similar to that of controls. However, morphometric analysis of the intestine of fetuses from severely diabetic rats revealed a decrease in each of the parameters measured which led to a general hypotrophy of the intestine. In the fetuses from mildly diabetic rats, the values of the morphometric parameters of the duodenal mucosa remained unchanged and comparable to those of the control group. The vascularisation of the duodenum is modified in these fetuses because the volume density of the blood vessels is significantly increased. In conclusion, both diabetic states of the mother induce various alterations in the fetal intestine, including the blood vessels. The nature of the structural changes observed in the intestine could lead to modifications in the function of the entero-pancreatic system in these fetuses.

Localization of high-affinity GABA uptake and GABA content in the rat duodenum during development.

The localization of high-affinity uptake sites for 3H gamma-aminobutyric acid (3H-GABA) was investigated in the rat duodenum during ontogenesis and also at the adult stage (from 15.5 days of fetal life up to 105 days post natum) by means of low- and high-resolution autoradiography. At all stages studied, specific endocrine cell types of the epithelium were labelled and an intense uptake was detected in the nervous tissue, especially in glial cells but also in scarce neurones. When the incubation medium was supplemented with beta-alanine (1 mM), a blocker of the glial uptake for GABA, the labelling persisted only in endocrine cells and in few neurones. The intensity and the frequency of the labelling decreased at later periods compared to the earlier developmental stages. The GABA content of the duodenum as measured by a new ion-exchange column chromatography-HPLC-coupled method was higher in the early postnatal period compared to later stages. These observations suggest that GABA, in addition to being a neurotransmitter, may play an important role during development of the duodenum.

Cell proliferation in pancreatic islets of rat fetuses and neonates from normal and diabetic mothers. An in vitro and in vivo study.

The influence of diabetic pregnancy on the fetal and newborn endocrine pancreas of rats was investigated in vitro and in vivo. A mild diabetic state was induced experimentally in the mother with 30 mg streptozotocin injected in the vein of the tail on the first day of gestation. The maternal blood sugar was 326 +/- 28 mg/dl at the end of the gestation. The in vitro experiment was performed on fetuses of 21.5 days. The endocrine pancreases were cultured during four or seven days and incubated the last 24 hours with tritiated thymidine. The healthy state of the islet cells after the two respective periods of culture was confirmed by an electron microscope study. After incubation with tritiated thymidine, a significantly higher percentage of labelled nuclei was observed in the islets of the diabetic group when compared with the controls. This was apparent after four days (diabetics: 54% - controls: 50%) and is obvious after seven days (diabetics: 28.8% - controls: 18%). For the in vivo experiment, two day old rats born at term from normal or diabetic mothers were injected (s.c.) with tritiated thymidine and killed two hours later. The proliferative capacity of the islet cells of pups born from diabetic mothers compared to the controls was higher when the percentage of labelled nuclei was calculated (respectively 5.4% versus 3.8%). An islet hypertrophy was also found in the diabetic group. In conclusion, our results combining in vitro techniques with in vivo observations demonstrate the higher proliferative rate of the fetal endocrine pancreas induced by a mildly diabetic feto-maternal environment.(ABSTRACT TRUNCATED AT 250 WORDS)

Localization of GABA high-affinity binding sites in the pancreas of neonatal rat.

The localization of gamma-aminobutyric acid (GABA) high-affinity binding sites was investigated in the exocrine and endocrine pancreas of neonatal rats by means of 3H-GABA autoradiography. GABA-binding was identified on Schwann cells and on the cells of the intralobular excretory ducts. In the endocrine part of the pancreas, no labelling was observed except in peripheral islet cells which, on the basis of their scarcity and distribution, could be somatostatin cells. Furthermore, peri-insular innervation showed considerable labelling.