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We investigated the effect of different doses of cyclosporin A (CyA) on glucose and insulin levels, as well as its residual effects on pancreatic islets ultrastructure after discontinuation of the drug. We studied four groups of Wistar rats. One control- (n = 5) and three experimental groups, n = 10 each, were treated with different doses of CyA i.m. for 14 days: group I, 5 mg/Kg; group II, 15 mg/Kg; and group III, 25 mg/Kg. Five animals of each group were sacrificed after 14 days, and the remaining five after 21 days to assess residual CyA effects. On the day of sacrifice, the rats underwent maltose absorption test, and glucose and insulin levels were measured. Pancreatic biopsies were obtained on day 21 to evaluate islets ultrastructure by electron microscopy. As a result, statistically significant, dose dependent (P < 0.05) increases in glucose and insulin levels were observed in CyA-treated groups. Groups II and III showed insulin levels significantly higher after fasting (P < 0.05) on day 14 comparing to the controls, while in groups I and II values returned to normal after CyA discontinuation. Group III showed persistently increased insulin levels on day 21. Pancreatic ultrastructural changes were observed only in group III. We can conclude that CyA effects on glucose and insulin levels were temporary and reversible at low doses. Ultrastructural changes in the pancreatic islets may occur with high doses of CyA.

Context: The development and evolution of different chronic diabetic complications may present variations among the different types and conditions of this disease., Objective: To evaluate the degree of microangiopathy in Type 1 diabetes mellitus (DM1) associated with autoimmune polyendocrinopathies (OSAD) or isolated DM1 (iDM1)., Patients: OSAD (n = 17) and iDM1 (n = 13) were over 15 years old at diagnosis of DM and were matched for diabetes duration (13.9 +/- 8.2 and 13.2 +/- 5.9 years respectively) and metabolic control (HbA1c: 6.4 +/- 1.9 and 6.8 +/- 1.4%)., Main Outcome Measures: Urinary albumin excretion (UAE; ELISA), the inversion of serum creatinine (1/C) level and indirect ophthalmoscopy., Results: Although the prevalence of hypertension was similar in both groups, the OSAD had inferior levels of UAE (7.4 +/- 2.5 vs. 17.3 +/- 9.2 micrograms/min; p < 0.05). Nephropathy was detected in 12% of the OSAD (none of them macroproteinuric) and in 39% of the iDM1. The UAE in the iDM1 correlated negatively with 1/C values (r = -0.7, p < 0.005), but the same did not occur in the OSAD (r = 0.2, ns). Among patients with retinopathy, the severe form was found in 29% of the OSAD and in 46% of the iDM1., Conclusions: OSAD was associated with a lower degree of microangiopathy, in spite of age at diagnosis, duration of diabetes and the metabolic control. In contrast with the iDM1, the increase in UAE of OSAD was not associated with reductions in GFR.

Adrenocortical autoantibodies (ACA), present in 60-80% of patients with idiopathic Addison's disease, are conventionally detected by indirect immunofluorescence (IIF) on frozen sections of adrenal glands. The large-scale use of IIF is limited in part by the need for a fluorescence microscope and the fact that histological sections cannot be stored for long periods of time. To circumvent these restrictions we developed a novel peroxidase-labelled protein A (PLPA) technique for the detection of ACA in patients with Addison's disease and compared the results with those obtained with the classical IIF assay. We studied serum samples from 90 healthy control subjects and 22 patients with Addison's disease, who had been clinically classified into two groups: idiopathic (N = 13) and granulomatous (N = 9). ACA-PLPA were detected in 10/22 (45%) patients: 9/13 (69%) with the idiopathic form and 1/9 (11%) with the granulomatous form, whereas ACA-IIF were detected in 11/22 patients (50%): 10/13 (77%) with the idiopathic form and 1/9 (11%) with the granulomatous form. Twelve of the 13 idiopathic addisonians (92%) were positive for either ACA-PLPA or ACA-IIF, but only 7 were positive by both methods. In contrast, none of 90 healthy subjects was found to be positive for ACA. Thus, our study shows that the PLPA-based technique is useful, has technical advantages over the IIF method (by not requiring the use of a fluorescence microscope and by permitting section storage for long periods of time). However, since it is only 60% concordant with the ACA-IIF method, it should be considered complementary instead of an alternative method to IIF for the detection of ACA in human sera.

We report a 32-year-old Black man, admitted to the ICU with coma and severe metabolic disturbances due to diabetic ketoacidosis. During the admission, rhabdomyolysis and acute renal failure (ARF) were diagnosed. After metabolic control and gradual decrease of creatine kinase levels, he presented a progressive improvement of renal function. We emphasize nontraumatic rhabdomyolysis as a poorly recognized pathogenetic factor for ARF in diabetic ketoacidosis and suggest that a better understanding of its mechanisms and an early application of protective measures is necessary.

Although rare, ketoacidosis may be induced by the occurrence of antibody mediated insulin resistance. Cases of 3 patients with ketoacidosis precipitated by immunologic insulin resistance (IIR) are reported. CASE REPORT--Three patients were admitted to the primary care unit of Hospital São Paulo in Diabetic Ketoacidosis. Demographic data of the patients (HML, DRJ and DIS) included: age (46.39 and 54 y.o.); sex (2F, 1M); diabetes mellitus (2 DM II and 1 pancreatic); duration of diabetes (6, 11 and 9 years) and BMI (17.5; 25.5 and 24.3 kg/m2. Admission laboratory data were: glucose (40, 38 and 22 mmol/L); pH (7.2; 6.9 and 7.2) and all had ketonuria. Insulin requirements for metabolic control were: HML: 1494U; DRJ: 1496U; DIS: 450U in a period of: 212, 206 and 72h. The plasmatic leves of Anti insulin antibodies (IA) measured by RIA (nU/mL) and ELISA (EI), where: HML: 7186, 3.26; DRJ: 7879, 3.42 and DIS: 8377, 2.88. HI was associated with marked decrease of both, insulin requirements and IA (HML: 3393, 1.39 after 10 months and DRJ: 4673, 2.34; DIS: 1510, after 18 months) at follow-up. DISCUSSION--The High Insulin requirements and time necessary to achieve the metabolic control guided us to the diagnosis of IIR. It was confirmed by high levels of AI and by the improvement in the metabolic control after the introduction of HI. CONCLUSION--The physician must be alert to severe IIR if there is no response after standard therapy to ketoacidosis. HI can be considered a valid alternative of treatment for IIR.

1. The literature suggests that the radioassay (RA) and ELISA detect different types of insulin antibodies (IA) (Wilkin et al., 1989. Diabetes, 38: 172-181). 2. In the present study we evaluated the relationship between these two antibodies and their involvement in the metabolic control of Type I diabetic (DMI) patients. 3. IA were measured by RA and ELISA in sera obtained from 34 patients (age: 9-16 years, median = 12.5 years; clinical duration of DMI: 0.1-11.0 years, median = 1.7 years) treated with different types of insulin [purified (bovine + porcine) N = 18, and monocomponent (porcine or human) N = 16] and submitted to various degrees of metabolic control as assessed by glycosylated serum protein (GSP) levels: range, 3.4-13.5%; median = 8.7%; normal value, 0.8-2.4%. 4. Insulin antibody levels measured by RA were: 3264 +/- 300 nU/ml (mean +/- SEM, normal value < 60 nU/ml) and by ELISA: 0.74 +/- 0.11 ELISA index (EI) (normal value, < 0.53). No correlation was found between IA levels measured by RA and ELISA, or between duration of the disease or insulin daily necessity and IA by either method. GSP was positively correlated with IA determined by ELISA (rS = 0.43, P < 0.01) but not with IA determined by RA. 5. The patients on purified bovine + porcine insulin had higher titers of IA by ELISA, compared to those of patients on monocomponent (0.96 +/- 0.15 vs 0.50 +/- 0.13 EI, P < 0.03, while IA levels measured by RA did not differ between groups. 6. These data show that RA or ELISA assays provide different serum titers of IA in insulin-treated diabetics and data obtained with ELISA correlated best with the metabolic control of Type I diabetic patients.

The authors evaluated the prevalence, magnitude, and contributing factors for osteopenia in insulin-dependent diabetes mellitus (IDDM). We measured bone mineral density (BMD) in the lumbar spine and femoral region in 90 patients aged 18-54 years with IDDM using dual-energy x-ray absorptiometry. The blood-glucose control, insulin dosage, duration of disease, and presence of chronic complications of diabetes were evaluated. Serum ionized calcium, magnesium, phosphorus, alkaline phosphatase (ALP), 25-hydroxycholecalciferol, immunoreactive parathyroid hormone (iPTH), and urinary calcium, phosphorus, and hydroxyproline were also analyzed. Thirty-one patients (34%) were classified as having a reduced BMD (less than 2 SD below the mean). The comparison between normal and low BMD patients showed that the osteopenics had a tendency to be younger (median, 28 years versus 32 years), showed a higher mean plasma glucose (15.5 +/- 5.0 mmol/L versus 12.9 +/- 3.8 mmol/L; p = 0.018), longer duration of disease (11.2 +/- 2.1 years versus 5.0 +/- 1.3 years; p = 0.004), and needed a higher insulin dosage (56 +/- 17 U/day versus 43 +/- 16 U/day; p < 0.001). There was a positive correlation between mean glucose levels, duration of disease, insulin dosage, and bone-mass decrease. A higher incidence of chronic complications, mainly retinopathy (58% versus 25%) and neuropathy (52% versus 22%) was found in the low BMD group. There was no alteration of serum calcium, phosphorus, iPTH, 25-hydroxycholecalciferol, and urinary calcium and phosphorus. The ALP levels were significantly higher in the osteopenic group, and magnesium and hydroxyproline levels were lower in the whole diabetic group, but these measurements did not correlate with BMD reduction.(ABSTRACT TRUNCATED AT 250 WORDS)

Case report on a child whose type I diabetes mellitus was diagnosed 23 months before the appearance of overt glucose intolerance. In this pre-IDDM stage of DMI were observed secondary enuresis, decreased growth speed, transient hyperglycemia and asymptomatic glycosuria. These alterations may represent the earliest clinical manifestation of impaired beta cell function. Immunologic markers (ICA and/or AAI) of DMI and abnormalities of the first-phase insulin secretion in response to intravenous glucose also may precede by several months the most common clinical picture of type I diabetes as they were detected in this child. If possible, markers and alterations should be tested in such patients and their young relatives with DMI in order to detect high risk individuals who may develop DMI. Such and accurate predictive ability should be a prerequisite to institution of appropriate therapy to preventing further beta cell destruction and severe metabolic decompensation, thus having the potential to reduce morbidity and mortality from new onset DMI.

1. The ability of glucose to suppress growth hormone (GH) secretion is well known and the glucose test is widely used for the diagnosis of acromegaly. However, when suspected acromegaly is associated with diabetes mellitus (DM) or impaired glucose tolerance (IGT) the interpretation of the GH response to the oral glucose tolerance test (OGTT) may be difficult. Recently, Hattori et al. (Journal of Clinical Endocrinology and Metabolism, 70: 771-778, 1990), using a highly sensitive (1.5 ng/l) polyclonal antibody-based immunoenzymometric assay, found no differences in the GH response to glucose load among control, IGT and DM patients. 2. We employed a less sensitive (100 ng/l) but monoclonal antibody-based immunoenzymometric assay to measure the serum GH levels of 19 normal subjects, 11 patients with DM and 11 patients with IGT to determine the effect of glucose intolerance on the GH response to the OGTT. 3. Complete suppression of GH (< 0.1 microgram/l) was achieved in 73% of the controls with a mean nadir of 0.17 +/- 0.16 microgram/l (range, < 0.1-0.6 microgram/l). GH was completely suppressed in 82% of the diabetics with a mean nadir of 0.58 +/- 1.21 micrograms/l (range, < 0.1-4.0 micrograms/l). However, complete suppression occurred in only 27% of the IGT patients with a nadir of 1.09 +/- 2.08 micrograms/l (range, < 0.1-7.0 micrograms/l), which was statistically higher than observed for controls and diabetics. 4. We conclude that plasma GH levels after glucose loading of IGT patients should be interpreted with caution because an abnormal response can be detected when some sensitive immunometric assays are employed.

The authors present a case of spontaneous ketoacidosis developed by one 32 years Krenak indian. The patient denied alcoholism and his mother had Type II diabetes for the last 3 years. The search to islets and insulin autoantibodies was negative in this patient. The basal C-peptide was found normal during follow-up. The patient received a short insulin therapy and now he shows good metabolic control (normal glycosylated hemoglobin) with oral hypoglycemic treatment. The environment influence, indian customs and the rarity of spontaneous ketoacidosis in these individuals were reviewed and discussed.

1. Insulin autoantibodies (IAA) of first-degree relatives of type I diabetic patients and recent-onset type I diabetics were measured by radioimmunoassay. A cut-off of 60 nU/ml was established on the basis of the values of normal control individuals. The intra-assay coefficient of variation was 9.2% for a moderately positive serum (1908 +/- 176 nU/ml (mean +/- SD), N = 7; range, 1708 to 2158 nU/ml). The interassay coefficient of variation was 23.8% for a negative (normal control) serum (28.1 +/- 6.7 nU/ml, N = 6; range, 22 to 39 nU/ml) and 14.5% in a highly positive serum (6185 +/- 899 nU/ml, N = 7; range, 5053 to 7009 nU/ml). 2. Insulin autoantibody levels (mean +/- SEM) were 19.3 +/- 2.8 nU/ml (range, -19 to 40 nU/ml) in 25 controls, 24.8 +/- 3.4 nU/ml (range, -17 to 59 nU/ml) in 41 type II diabetic patients, 18.5 +/- 2.4 nU/ml (range, -58 to 268 nU/ml) in 171 first-degree relatives of type I diabetic patients and 208.9 +/- 87.0 nU/ml (range, 10 to 1101 nU/ml) in 16 recent-onset type I diabetic patients. IAA levels were significantly higher in the last group compared with the other groups (P < 0.01). 3. None of the controls or type II diabetics exceeded the upper limit of normality. In contrast, 9 of 171 (5.3%) first-degree relatives and 9 of 16 (56.0%) recent-onset type I diabetic patients had IAA levels above the 60 nU/ml cut-off point.(ABSTRACT TRUNCATED AT 250 WORDS)

Human sera from 51 recent-onset insulin-dependent (type I) diabetic patients and 47 unrelated control subjects were screened for the possible presence of circulating factors reacting with several anti-pancreatic islet monoclonal antibodies (MoAb.ISL) in solid-phase radioimmunoassay methods (the original goal being the detection of anti-idiotypic islet cell antibodies and/or specific islet cell antigen-bearing immune complexes). MoAbs from the parental myeloma cell line and purified immunoglobulins (Igs) from different animal species were controls. Type I diabetic sera showed significantly increased binding to MoAb.ISL-coated wells compared with normal subjects (P less than .001). However, the same sera also tended to show a higher binding to the control (non-islet-related) MoAb. Sera from type I diabetic patients also reacted with horse, bovine, pig, rabbit, and goat IgG. Displacement of the binding has been obtained by F(ab')2 and/or Fc fragments of IgG. Evidence has been obtained regarding a similar reaction with human IgM. All the sera were negative when tested for rheumatoid factor by nephelometry. The circulating antibodies described have been proven to be different from islet cell autoantibodies. An anti-Ig antibody is thus present in the sera of recent-onset diabetic patients and represents an additional immunological phenomenon with possible physiopathological and clinical significance.

A major problem in standardization of the islet cell cytoplasmic antibody (ICA) assay is variation in sensitivity of the different human pancreas substrates used in individual laboratories. To circumvent this problem, we have developed an assay that utilizes Wistar-Furth rat pancreas as substrate, an anti-islet monoclonal antibody (A2B5) to identify islets and fluorescein-conjugated protein A to identify patient autoantibodies. Sera from 85 control subjects, 27 type I diabetics, and 17 subjects at high risk for developing type I diabetes were assayed in parallel with our standard ICA assay on human pancreas substrate and with Wistar-Furth rat pancreas as substrate. Two sera from control subjects (2 of 85) were ICA positive with rat pancreas compared to 1 of 85 with human pancreas substrate. Sera from 11 of 27 type I diabetics and 15 of 17 sera from high-risk subjects were ICA positive with either rat or human pancreas substrate. A correlation between the specific islet fluorescence readings on human and rat pancreas sections was found with sera from high-risk and control subjects. Furthermore, end-point titers of an ICA-positive serum were identical with both assays. Finally, incubation of an ICA-positive serum with glycolipids, extracted from either human or Wistar-Furth rat pancreas, blocked subsequent ICA binding. These findings suggest that Wistar-Furth rat pancreas expresses an identical or similar autoantigen to human pancreas.

The presence of other organ-specific autoimmune disorders in some patients with premature menopause has supported the concept of an autoimmune etiology. The authors analyzed the peripheral blood of 23 women with the diagnosis of premature menopause to detect the presence of monoclonal antibody-defined T-lymphocyte abnormalities and/or antiovarian antibodies. All subjects were less than 40 years of age with the duration of menopause ranging from less than 1 year to 11 years at the time of study. Thirty-five percent of the subjects had an elevated percentage of Ia+ (Dr-activated) T cells using monoclonal antibody L243. The percent T4 (helper) T8 (suppressor/cytotoxic) T cells and T4/T8 ratio were normal in the study group. Four subjects (approximately 17%) had elevated percentages of the age-related 3G5+ T cell subset. Two of the subjects with increased 3G5+ T cells also exhibited increased Ia+ T cells. Antiovarian steroid cell antibodies and antiadrenal cortical antibodies were present in approximately 9% of subjects. Anti-islet cell antibodies were not present. Thyroid antimicrosomal antibodies were present in 17% of subjects. Study subjects exhibited immunologic abnormalities that the authors hypothesize may play a role in the development of premature menopause in a larger percentage of patients than was previously suspected.

Transient hyperglycemia may represent the earliest manifestation of IDDM. In a series of 30 children referred for transient hyperglycemia, 8/30 (27%) developed IDDM within 10 months of their initial evaluation. Three children with detectable cytoplasmic islet cell antibodies (ICA) and/or positive insulin autoantibodies (CIAA) all developed IDDM, compared to 21% of ICA negative children and 13% who were CIAA negative. Of those with impaired glucose tolerance, 6/11 (55%) developed IDDM, as did 1/14 with normal OGTT. Of the children with a low "K rate" on IVGTT, 75% developed IDDM, compared to 1/13 with a normal "K rate". All four children (100%) whose first phase (1' + 3') insulin secretion never exceeded the first percentile developed IDDM within nine months, while no child with first phase insulin secretion above the first percentile (0/16) developed IDDM during 19 +/- 9 months (mean +/- SD) of follow-up. Thus, in our experience the oral glucose tolerance test is a less accurate predictor of impending IDDM; immunological abnormalities have the highest positive predictive value, while the first phase insulin secretion during an intravenous glucose tolerance test has the highest negative predictive value and the greatest overall accuracy of prediction.

We evaluated the expression of factor VIII and class II histocompatibility antigens on frozen sections of normal human and rat pancreases. The immunohistologic studies were performed with directly fluoresceinated anti-human factor VIII and monoclonal antibodies A2B5, 3G5 (anti-islet), L-243, I-2, OK1 (anti-human Ia-DR), Leu-10 (anti-human HLA-DQ), anti-human HLA-DP, and OX6 (anti-rat Ia). Islet endothelial cells of humans and Wistar, CD, and BB diabetes-prone rats could be distinguished from intra-acinar endothelial cells by markedly enhanced factor VIII immunoreactivity. Factor VIII-antibody staining of islet endothelial cells was specifically absorbed by prior incubation of anti-human factor VIII antibody with normal human plasma but not by incubation with factor VIII-deficient plasma. By double indirect immunofluorescence, normal human pancreatic ductal epithelium expressed Ia in five of six pancreases studied.

Murine monoclonal antibodies (MAbs) HISL-5, -9, and -14, generated after immunization of mice with human pancreatic islet cell preparations, recognize a differentiation antigen expressed by the pancreatic islet cells. These MAbs react strongly with all endocrine cell subtypes of human pancreatic islets, but minimally if at all with the exocrine acinar cells, vascular cells, and stromal connective tissue cells of the pancreas. The antigen is located on the cell surface (plasma membranes), as indicated by immunofluorescence staining of viable cell preparations. Besides the pancreatic islets, HISL-5, -9, and -14 antigenic determinants are also expressed by thyroid follicular cells, parathyroid chief cells, and anterior pituitary cells, other commonly involved targets in organ-specific autoimmune disorders. Preliminary biochemical findings indicated that the MAb-defined epitope(s) is trypsin sensitive and resistant to periodate oxidation and exposure to chloroform-methanol. Further biochemical studies, including single step MAb immunoaffinity chromatographic purification, indicate that the antigen recognized by the MAbs HISL-5, -9, and -14 is a 100 K glycoprotein.

A quantitative fluid-phase radioassay for autoantibodies reacting with insulin (competitive insulin autoantibody assay, CIAA) was developed. The assay's features include 1) use of a physiologic amount of 125I-labeled insulin, 2) parallel incubations with supraphysiologic cold insulin (competitive), and 3) an incubation time of 7 days and a single-step multiple-wash polyethylene glycol separation. Mean +/- SE CIAA levels in 50 controls were 8 +/- 1.4 nU/ml (range -16-33.3). In 36 cytoplasmic islet cell antibody (ICA)-positive nondiabetic first-degree relatives of type I (insulin-dependent) patients less than 30 yr of age, CIAA levels exceeded the normal range in 20 (55.6%) of 36 (mean 86.8 +/- 17.1 nU/ml). In 26 ICA-positive relatives greater than 30 yr of age, only 5 (19.2%) of 26 exceeded the normal range (mean 26.1 +/- 9.4 nU/ml); P less than .001 compared with younger ICA-positive relatives). Six ICA-negative HLA-identical siblings of type I diabetic patients had normal CIAA levels (mean 3.6 +/- 5.8 nU/ml), and only 2 of 13 ICA-negative identical twins discordant for diabetes (mean 15.4 +/- 6.6 nU/ml) exceeded the normal range. Nine (50%) of 18 ICA-positive schoolchildren exceeded the normal range (mean 105.3 +/- 36.7 nU/ml). Genetically susceptible subjects negative for CIAA (with only 3 exceptions) remained negative for CIAA on multiple determinations (3 conversions observed), and CIAA levels of positive subjects were relatively stable. Linear regression of the first CIAA level versus last (interval between sampling 1 mo to 10 yr) in genetically susceptible individuals showed a highly significant correlation (r = .95, P less than .001).(ABSTRACT TRUNCATED AT 250 WORDS)