Robert B. Elliott

Parenteral absorption of insulin from the lung in diabetic children.

Semi‐synthetic human insulin was delivered via a novel nebulizer to the respiratory tracts of six diabetic children. Blood glucose control obtained was at least as good as a control day when they received their usual dose of subcutaneous insulin.

PARTIAL PRESERVATION OF PANCREATIC β-CELL FUNCTION IN CHILDREN WITH DIABETES

Abstract 17 newly diagnosed insulin-dependent diabetic children received prednisone (0·25 mg/kg/day) for the first 12 months after diagnosis. The urinary C-peptide level of this corticosteroid treated group of diabetic children was significantly higher (p

Neonatal screening for cystic fibrosis, using immunoreactive trypsin assay in dried blood spots.

Our previous studies suggested that assay of immunoreactive trypsin (IRT) in dried blood spots might be a valuable neonatal screening test for cystic fibrosis (CF). We have developed a convenient, sensitive, human trypsin radioimmunoassay, which uses a 3-mm diameter disc of dried blood. The molecular species assayed in blood is trypsinogen. A retrospective study of 24 known cases of CF and appropriate controls confirmed that an elevated blood level of IRT is characteristic of all newborn CF infants, whether or not they have residual exocrine pancreatic function. IRT levels did, however, decline with time of sample storage. Guthrie cards from 5040 newborns were prospectively assayed: 2% of tests were reassayed because of elevated IRT. Thirty-three second samples (0.67% of the total) were requested, 32 were received, and 31 had normal IRT values. The baby with an elevated result had no clinical symptoms of CF at one month of age, normal stool trypsin activity, but 2 sweat tests gave grossly abnormal results. In contrast, a second infant in whom CF was diagnosed around the same time, had a similar 5-day blood spot IRT value, but severe clinical symptoms of CF and no stool trypsin activity. No false negatives are yet known. We conclude that blood spot IRT assay is a reliable and convenient neonatal screening test for CF.

A combined casein-free-nicotinamide diet prevents diabetes in the NOD mouse with minimum insulitis

We have previously shown that diabetes in the NOD mouse can be prevented if mice are placed from weaning on an infant formula diet in which the protein source is replaced with casein hydrolysate (Pregestimil) or soy protein (Prosobee), or if 1% nicotinamide is given in the drinking water. Nicotinamide somewhat suppresses insulitis but the hydrolysed casein formula does not. In this study, Prosobee was given concurrently with oral nicotinamide from weaning and their effects on the development of insulitis and diabetes measured. These effects were also assessed in mice given Prosobee alone from conception (day -20) or from weaning. Unlike the earlier experiments, a marked suppression of insulitis was observed when the diets and nicotinamide were given concurrently (mean insulitis scores +95% confidence intervals (back transformed): day 40 = 0.4% [0.03, 1.17] vs. 12.5% [2.52, 28.40] and at day 90 = 8.8% [3.65, 15.68] vs. 48.1% [33.89, 62.49], P = 0.0001). A similar suppression was observed on day 90 with Pregestimil combined with nicotinamide 7.3% [3.88, 11.70] vs. 43.8% [32.59, 55.35] (P = 0.0001). Qualitatively, introduction of Prosobee from conception appeared to elicit a greater degree of suppression of insulitis than when introduced from day 21. Insulitis lesions were examined immunohistochemically for CD4, CD8 and MAC-1 cells. The proportion of these cells was not different for any regime despite the great differences in total number of inflammatory cells in and around the islets of mice fed the combined diet. All the three dietary treatments (Prosobee from day -20, Prosobee from day 21, Prosobee+nicotinamide from day 21) resulted in substantial protection from diabetes in mice followed until 250 days. We conclude that the complete prevention of diabetes in the NOD mouse fed a casein-free diet together with nicotinamide is accompanied by marked inhibition of insulitis, which is not seen when either dietary agent is introduced alone. The somewhat greater suppression of insulitis in mice given the soy diet from conception compared to those fed from day 21 may indicate that even maternal diet during gestation may influence diabetes outcome in the offspring.

Low Dose Streptozotocin Causes Diabetes in Severe Combined Immunodeficient (Scid) Mice Without Immune Cell Infiltration of the Pancreatic Islets

Streptozotocin (stz) given in low doses (40 mg/kg body weight) on 5 consecutive days to susceptible strains of mice causes diabetes. Previous studies have shown that the induction of diabetes is associated with inflammatory infiltrates within the pancreatic islets. However, it is unclear whether stz causes limited beta cell destruction followed by insulitis or whether the diabetogen promotes immune cell influx into the pancreatic islets, followed by immune-mediated beta-cell destruction. It is also unclear whether stz given in sub-diabetogenic doses is capable of causing diabetes independent of cell-mediated processes. Here we have examined these possibilities in CB.17 Scid mice which lack functional T and B cells but have immunocompetent macrophages and NK cells. Low dose stz given to Scid mice caused diabetes in approximately 50% of mice of both sexes by 21 days (14/24 males; 10/18 females). Sections of pancreas were examined immunohistochemically for the presence of MAC-1 positive cells (macrophages and natural killer cells) in the exocrine, peri- and intra-islet regions at different time points following the administration of stz. There were no statistically significant differences in the number of immunoreactive cells in the three locations between tissues obtained from stz-injected mice (3, 7, 14 and 21 days after stz injection and at onset of diabetes) and buffer-injected Scid mice. Although diabetic Scid mice showed a reduced number of insulin immunoreactive cells and peri- and intra-islet distributed glucagon cells, no insulitis was seen histochemically. In parallel studies, normal Swiss male mice given stz at a similar dose developed diabetes (10/10) associated with insulitis which consisted predominantly of CD4, CD8 and MAC-1 cells. Balb/c mice given stz similarly, also developed diabetes (5/8) without showing insulitis, although a moderate increase in the number of macrophages were observed within several islets. These studies demonstrate that stz administered in multiple low doses to Scid mice can cause beta cell destruction and diabetes in the absence of immune cell infiltrate within the pancreatic islets.

CONTROLLED TRIAL OF AN ELECTROSTATIC PRECIPITATOR IN CHILDHOOD ASTHMA

A cross-over trial was conducted on 10 children with moderate to severe asthma, who had positive skin tests to Dermatophagoides pteronyssinus and nocturnal wheeze. An electrostatic precipitator was used during the night to remove airborne particles from the bedroom. During use of the precipitator peak expiratory flow rates were no better than in a control period.

Residual beta-cell function and islet cell antibodies in diabetic children.

Summary: There are no prospective studies evaluating residual β-cell function and islet cell antibody status during the first 2 years after the diagnosis of diabetes hi children. We report such a prospective study of 21 insulin-dependent diabetic children. The relationship of residual β-cell function during the 2 years after diagnosis, to age at diagnosis, islet cell antibody (ICA) status, histocompatability status, and exogenous insulin is reported.Twenty-one children were studied for at least 2 years after the diagnosis of diabetes. Blood was obtained at the time of diagnosis for the determination of ICA and histocompatability status. ICA status was determined 12 months after diagnosis, and again at 24 months if ICA had been detected at 12 months. Residual β-cell function was assessed serially by measuring 24-hr urinary C-peptide. The 24-hr C peptide excretion was expressed relative to 24-hr creatinine excretion. This ratio is termed urinary C peptide excretion (UCP). Islet cell antibodies were measured by indirect immunofluorescence on human O-group pancreas. Fluorescein-labeled anti-C3C was used to determine whether the islet cell antibodies were able to fix complement. A standard NIH micro-lymphocytotoxicity test was used to determine histocompatability status at the A, B, and C loci.Throughout the entire study period, the UCP of each diabetic was always below the mean of the group of nondiabetic children. For all diabetic children, the UCP decreased during the 24 months after diagnosis. Most childrens (14 of 21) UCP had faUen below 0.20 during the 12 months after diagnosis. Twelve of 21 diabetic children were ICA positive at diagnosis. Five had unaltered titers of ICA 12 months later and four had no ICA detectable at 12 months, whereas for three the titer had fallen by at least two dilutions. The magnitude of the UCP 12 months after diagnosis correlated significantly with age at the time of diagnosis (r = 0.557; P < 0.05). Similarly, there was also a significant correlation between the UCP 24 months after diagnosis and age at diagnosis (r = 0.446; P < 0.05). There was no significant difference between insulin dose, expressed as units/kg/day at 12 and 24 months after diagnosis. There was no significant relationship between ICA status and insulin dose at 12 or 24 months. There was no discernible relationship between the evolution of ICA or UCP with histocompatability antigens at the A, B, or C loci.Speculation: The positive correlation between urinary C peptide and age at diagnosis of diabetes suggests that pancreatic size and potential for subsequent growth might be factors influencing the impact of a diabetogenic insult. For insulin-dependent children, the impact of the diabetogenic process is usually greatest during the 12 months after diagnosis. Potential therapeutic efforts to limit islet cell damage need to be applied early, and it is possible they may not need to be continued much beyond 12 months after diagnosis.

Experience with neonatal screening for cystic fibrosis in New Zealand using measurement of immunoreactive trypsinogen

Abstract Neonatal cystic fibrosis (CF) screening has been performed in New Zealand for a total of 7 years. This study reports the experience with this procedure in New Zealand over a 4 year period and compares it with 2 years when diagnoses of CF were suggested by clinical features only. A total of 72 infants were confirmed as having CF during 4 years of screening. Twenty‐eight infants were found to have CF during 2 years in which screening was not performed. There were 29 false positive diagnoses during the screening years and six false negative diagnoses. Three of the false negative diagnoses occurred because of laboratory error, but three occurred because either the first or second measurement of immunoreactive trypsinogen (IRT) was normal. Faecal chymotrypsin was measured in samples from 434 infants at the time of the second IRT and assisted with the diagnosis for one infant which might otherwise have been missed. Only 42.5% of infants were asymptomatic at the time of the confirmatory sweat test. Significant morbidity and mortality was associated with meconium ileus which occurred in 24% of infants with CF. Improved ascertainment of cases of CF has occurred since screening began. Further follow‐up is required to determine other benefits of newborn screening.

Longitudinal study of islet cell antibodies and insulin autoantibodies and development of diabetes in non-obese diabetic (NOD) mice

We have previously shown the presence of circulating islet cell cytoplasmic antibodies (ICA) and insulin autoantibodies (IAA) in the NOD mouse before onset of insulin‐dependent diabetes mellitus (IDDM). Here we have determined the levels of the two autoantibodies in 28 female NOD mice longitudinally from approximately day 40 to day 250, to examine their ontogeny, association and predictive value for diabetes. All animals (11 diabetic, 17 non‐diabetic) showed varying levels of ICA at some stage, while IAA activity was found in 21 out of 28 mice. Expression of both the markers was seen in more than half of the animals by day 60, with higher levels and rates occurring subsequently in both diabetic and non‐diabetic groups. The expression of ICA did not always correlate with that of IAA. There was no apparent difference in the ontogeny of ICA and IAA between the two groups. During the study period the number of animals with ICA was similar in the two groups, while the number of those with IAA was higher in the diabetic animals. In this group declining and rising levels of ICA were seen just before clinical diabetes with frequent peaks of IAA. In the same animals, eight out of 11 mice showed co‐expression of high levels of both markers either intermittently or persistently prior to onset, whereas only one non‐diabetic animal showed this. We conclude that the ontogeny and serum level of ICA or IAA alone could not be used to predict the clinical onset of diabetes in these animals. However, co‐expression of high levels of both markers prior to onset may suggest a strong predisposition to clinical diabetes. This may have relevance to attempts to predict the onset of IDDM in humans who have one or both of these immunological markers.

First phase insulin release in the non-obese diabetic mouse: correlation with insulitis, beta cell number and autoantibodies

The ontogenic variation of beta cell function and its relationship with the degree of islet damage and levels of autoantibodies have been studied in the non-obese diabetic (NOD) mouse model. We conducted in vivo first phase insulin release (FPIR) in response to intravenous glucose and studied its correlation with the degree of insulitis, islet cell antibody (ICA) and insulin autoantibody (IAA) levels in female NOD mice cross-sectionally at days 40 (n = 19), 90 (n = 21), 150-160 (n = 21) and day 250 (n = 20). The mean +/- SEM FPIR values showed an age-related decline from day 40 (46.2 +/- 5.3 microU/ml) to day 150-160 (17.8 +/- 2.5 microU/ml) and then doubled at day 250 (34.5 +/- 5 microU/ml), while the mean +/- SEM insulitis scores increased progressively until day 150-160 (61.7 +/- 6.1%) and then declined slightly at day 250 to 50.2 +/- 6.2%. In female NOD mice with spontaneous diabetes (n = 4) and streptozotocin-induced diabetic Swiss mice (n = 5) FPIR was either absent or greatly attenuated. A statistically significant inverse correlation between FPIR and insulitis was found among NOD mice at days 90 (P = 0.02; r = -0.52) and 150-160 (P = 0.03; r = -0.48). However, no statistically significant correlation was observed at days 40 and 250. Morphometric techniques applied to day 150-160 pancreatic sections showed a statistically significant negative correlation between insulitis and beta cell number per unit area of islet tissue (P = 0.0001; r = - 0.75). At this age some islet beta cells showed different intensities of staining by immunofluorescence.(ABSTRACT TRUNCATED AT 250 WORDS)