Human fetal islet transplantation in type 1 diabetics: comparison of immunological effects between multiple implantation regimens.

Previous studies have suggested that the multiple transplants might be equally metabolically efficient to a single regimen for human adult islets. The aim of this study was to compare immunological and metabolic parameters after each of the two regimens of human fetal islets (HFI). Group A single transplants (n = 9) had 180 +/- 20 x 1000 HFI equivalents (IEQs) implanted via a single intramuscular injection. In group B multiple transplants (n = 8) islets were implanted by three consecutive injections of 60 +/- 10 x 1000 IEQs at 7-day intervals. We analyzed the immunological parameters of CD4/CD8 T lymphocyte ratios; islet cell antibodies (ICAs) and insulin antibodies (IAs). We estimated insulin secreting capacity (ISC) as the metabolic parameter. We observed that the CD4+/CD8+ T-cell ratio increased, peaking on day 90, in similar fashion in both groups: day -1: A = 1.18 +/- 0.03 versus B = 1.19 +/- 0.04; on day 90: A = 1.79 +/- 0.09, versus B = 1.75 +/- 0.08 (P = NS) immediately before the decrease in C-peptide levels. Thereafter the ratios rapidly decreased without statistical differences. The levels of ICAs did not change. The levels of IAs, which were increased before transplant, then decreased without statistical differences between the groups. The values of ISC increased after transplant and then decreased similar to the T-cell ratio. Our results demonstrated that regimens of multiple and single HFIs did not show differences in the kinetics of the immunological response presumably mediating graft destruction. The CD4/CD8 ratio increased as the C-peptide level decreased, peaking on day 90 at the time of a decrease in C-peptide. These results may be useful for clinical studies of HFIs for type 1 diabetic patients.

Human fetal islet transplantation in type 1 diabetic patients: comparison of metabolic effects between single and multiple implantation regimens.

Previous studies suggest that multiple transplantations might be equally efficient to a single regimen for human adult islets. The aim of this study was to compare metabolic parameters after each of the two regimens of human fetal islet (HFI) transplantation in type 1 diabetics. In group A (single transplant, n = 9), 180 +/- 20 x 1000 HFI equivalents (IEQs) were implanted by a single IM injection; in group B (multiple transplants, n = 8) islets were implanted as three consecutive injections (60 +/- 10 x 1000 IEQs) at 7-day intervals. We analyzed the metabolic parameters on days -1, 30, 60, 90, 120, 150, and 180 after the procedure. Among the metabolic parameters, we evaluated insulin secretion capacity-ISC (C peptide, RIA), metabolic control (HbA1c, chromatography), and insulin daily dose IDD. We found that C peptide levels increased, peaking on day 90 (A: 0.38 +/- 0.15; B: 0.34 +/- 0.19 nmol/L, P = NS) and then rapidly decreasing without differences, the HbA1c levels and IDD decreased in the same manner without differences between the groups. Our results demonstrate that multiple and single islet transplant regimens are equally efficient to temporarily restore a significant ISC with improvement of metabolic and clinical parameters. The results imply that the two regimens have an equal clinical value.

Relation between course of disease in type 1 diabetes and islet cell antibodies.

The aim of this study was to estimate the possibility of predicting the course of type 1 diabetes. We analyzed the importance of islet cell antibody levels and residual beta cell function in 46 newly diagnosed patients with diabetes. Islet cell antibodies (ICAs; Juvenile Diabetes Foundation [JDF] units) were determined at the time of diagnosis by the indirect immunofluorescent method. beta cell function was estimated by C peptide levels (nmol/L) before and after glucagon stimulation at the time of clinical remission. Of the 46 patients, 13 were ICA negative (group A). Among ICA-positive patients, ICAs were < 20 JDF units (group B) in 15, between 20 and 80 JDF in 9 (group C), and > 80 JDF in 9 (group D). In group A, 9 patients had clinical remission for 7.5 +/- 1.7 months. Their basal C peptide level was 0.26 +/- 0.05 nmol/L and it increased after stimulation to 44.5 +/- 2.5%. Ten patients in group B had remission for 6.2 +/- 1.5 months. Their basal C peptide levels (0.28 +/- 0.07 nmol/L) were similarly increased after stimulation (47.5 +/- 2.5%). In group C, all patients had remission and it was of the longest duration (14.7 +/- 1.5 months). They had the highest basal C peptide levels (0.45 +/- 0.12 nmol/L) with increases to 57.5 +/- 3.5%. Seven patients in group D with ICA levels > 80 JDF had a short remission (3.2 +/- 1.2 months) despite good basal C peptide levels (0.42 +/- 0.05 nmol/L) and excellent increases after stimulation (92.5%). Our results suggest that moderate levels of ICA are associated with good residual beta cell function and longer remission. Very high ICA levels (> 80 JDF) at the time of diagnosis despite better beta cell function are associated with short clinical remission. Therefore, high ICA levels could be a marker of strong autoimmune reaction and accelerated depletion of beta cell function.

[Response of pancreatic polypeptide to a protein rich meal in insulin non dependent diabetes melitus and autonomic neuropathy]. / Nivo pankreasnog polipeptida posle proteinskog test--obroka kod dijabeticara nezavislih od insulina i autonomnom neuropatijom.

Parasympathetic function and plasma hPP response to a protein rich meal were evaluated in 105 insulin non-dependent diabetic patients: 20 with autonomic neuropathy (group A), diagnosed by Clonidin test; 35 patients with neurophysiological evidence of polyneuropath (group B); 30 patients with autonomic neuropathy and polineuropathy (group C), and 20 patients without any sign of neuropathy (group D). Plasma hPP levels were determined by RIA using an anti-hPP antiserum, kindly provided by Prof. S. R. Bloom (Hammersmith Hospital, London). Blood was taken at 0. 45 and 60 minutes after the beginning of the meal. In groups A and C, the meal induced hPP increase was significantly lower than in group D (p 0.001). All group B patients had a marked increase in the peptide, similar to that in diabetics without neuropathy. These result ssuggest that diabetic autonomic neuropathy is associated with dysfunction of hPP secretion, and that the evaluation of hPP response to test meal may be a sensitive and simple method for the assessment of paraympathetic impairment in diabetes.

[Human fetal pancreatic islet transplantation in insulin-dependent diabetics: possibilities of early detection of transplant destruction]. / Tranplantacija ostrvaca humanog fetalnog pankreasa u insulin-zavisnih dijabeticara: mogucnost rane detekcije destrukcije transplantata.

The factors determining the outcome of human fetal islet transplantation in patients with insulin-dependent diabetes mellitus (IDDM) remain unclarified. In this study we analysed the ratio between immunoregulatory lymphocyte subpopulations in order to search for a possible marker of the immune destruction of transplanted islets. Human fetal islets were isolated by collagenase digestion, cultured for 14 days at 37 degrees C, 5% CO2, and implanted under fascia of m. rectus abdominis in 7 IDDM patients (5 pancreata per patient). After transplantation we evaluated simultaneously the level of metabolic control through HbA1c values determined by chromatography, the capacity of insulin secretion through the C-peptide levels (determined by radioimmunoassay) before and 6 minutes after 1 mg glucagon i.v. stimulation, and the ratio between CD4+ and CD8+ lymphocytes determined by immunofluorescence using monoclonal antibodies. We found that metabolic control after transplantation was improved together with the decrease of the insulin daily dose, and the improvement was simultaneous to the increase of both basal and glucagon-stimulated C-peptide levels. Four months after transplantation we detected a remarkable decrease in the secretion capacity, accompanied by the necessity for an increase in daily insulin dose to maintain optimal metabolic control. However, the loss of islet function was preceded by the increase in CD4+/CD8+ ratio, thus reflecting the presumable accumulation of CD4+ inducer T-lymphocytes. When the islet secretion capacity was destroyed, we found a decrease in CD4+/CD8+ ratio, reflecting the recruitment of CD8+ effector cells.(ABSTRACT TRUNCATED AT 250 WORDS)

[Use of cyclosporin A for remission induction in newly-detected insulin-dependent diabetes]. / Primena ciklosporina A u indukciji remisije u novootkrivenom insulini-zavisnom dijabetesu.

It has been postulated that some of the recent-onset insulin-dependent diabetics, after the initial use of insulin therapy, might develop the "honey moon period", i.e., a spontaneous remission of the disease, defined as the state of normal metabolic control maintained without insulin therapy. However, it has also been shown that spontaneous remission appears only in 5% of the patients treated with conventional insulin therapy and lasts, most frequently, not more than a few weeks. Different therapeutic regimens of immunosuppression and immunomodulation have been used worldwide in order to induce the remission, based on the findings that an autoimmune process underlies the pathogenesis of this type of diabetes. In this study, we have shown the results of the follow-up analysis of the effects of the treatment with cyclosporin A in 21 recent-onset insulin-dependent diabetics. In 15 of those patients insulin treatment was applied as bi-daily doses of monocomponent insulin preparations, and in 6 of them intensified insulin therapy with human insulin was used. In the first group, the remission was achieved in 46.66% and in the second group in 66.66%, which is a significantly higher incidence than in control groups treated only with insulin, without cyclosporin. Moreover, the duration of remission was longer in the patients treated with cyclosporin. The analysis of the residual beta cell secretory capacity has shown that C-peptide levels (taken as a marker for insulin secretion) were slightly higher in patients with the spontaneous remission than in those with the cyclosporin-induced remission both in basal conditions and after stimulation with 1 mg of glucagon. In the patients with cyclosporin A-induced remission we found an improved basal C-peptide secretion and, even more, we detected a significant improvement in beta cell response to the glucagon stimulation. The analysis of the first-phase insulin secretory response (the insulin response to rapidly injected glucose during the intravenous glucose tolerance test) which has been shown to be impaired very early during the development of diabetes, has demonstrated the lack of its recovery both in the spontaneous and in cyclosporin A-induced remissions. The analysis of the molar insulin/C-peptide ratio has detected the impairments of this ratio which remains decreased both in spontaneous and cyclosporin-induced remissions.

[Determinants of susceptibility to the induction of autoimmune diabetes in inbred strains of rats]. / Ispitivanje determinanti ostljivosti na indukciju autoimunog dijabetesa u cistih sojeva pacova.

Interleukin 2 (IL 2) proved to be important for the generation of autoreactive T cells in the pathogenesis of different autoimmune disorders, including diabetes. Also, we have previously shown that lymphoid cells obtained from inbred strains of rats Dark August (DA) and Albino Oxford (AO) exhibited a genetically determined difference in their ability to produce IL 2. In this study, in DA and AO rats, we analysed the susceptibility to the induction of autoimmune diabetes with multiple subdiabetogenic doses of alloxan. Rats were injected i.v. with 25 mg/kg b.w. of alloxan in 5 consecutive days. All DA rats developed moderate and persistent hyperglycaemia 15 days after the induction. In contrast, AO rats failed to develop the disease after the same regimen of induction. When the induction was made in (AO x DA)F1 rats, all of them developed the disease similar to the susceptible parental DA strain. However, the onset was delayed (the 40th day after the induction) and the glycaemia was constantly lower than in DA rats. Our results indicate that the difference in the response to the induction of autoimmune diabetes between susceptible DA strain, being high IL 2 producer, and resistant AO strain, being low IL 2 producer, is genetically determined. The results also suggest that this control could involve the gene determining the level of IL 2 production in DA and AO rats, thus signifying that the susceptibility to the induction of this form of autoimmune diabetes could be related to the level of IL 2 activity.

[The effect of somatostatin on specific binding of insulin to receptors in patients with non-insulin dependent diabetes]. / Efekat somatostatina na specificno vezivanje insulina za receptore u obolelih od insulin nazavisnog dijabetesa.

Tests were performed of the effects of the Somatostatin (SMS) upon the concentration of Insulin, Glucagon and STH, as well as of the effects of SMS upon the specific binding of the insulin to the receptors. The tests were carried out on eight insulin-independent diabetics, and five healthy volunteers. The tests were made with placebo, followed by 100 ug of the analogue SMS 201-995, known under the name of Sandostatin. Blood specimens for determining all parameters mentioned above were taken at 9 a.m., 3 p.m., 9 p.m., and 3 a.m. (09 h, 15 h, 21, and 03 h). The goal of the tests was to determine whether the SMS had any effect upon the glucoregulation, and at which level changes take place. In the group of healthy volunteers, a considerable decrease of insulin took place six hours after the administration of STH, and the decrease of the glucagon was especially marked, tending to increase again after six hours, while the specific binding of the insulin to the receptors, and the number of receptors were decreased six hours following the administration of the SMS when compared with the placebo, and without tending to reach the previous levels until 3 a.m. (03 h). In insulin-independent diabetics, the SMS leads to a considerable drop of concentrations of Insulin, Glucagon, STH and to a specific binding of the insulin.(ABSTRACT TRUNCATED AT 250 WORDS)

Detection of compounds in serum inhibiting insulin binding to erythrocyte receptors.

The erythrocytes were used as a model system to study insulin receptors in diabetic patients with highly increased insulin resistance. The specific binding of 125I-insulin to erythrocytes of these patients was markedly reduced. When erythrocytes prepared from non-diabetic subjects were exposed to serum of these patients, the decrease of insulin binding was reproduced. The binding of insulin showed a displacement curve that is parallel to the control values over the same range of hormone concentration, even in the case when control erythrocytes were preincubated with serum of diabetics with increased insulin resistance. These results indicate the presence of certain component in blood plasma of examined patients which alters insulin binding to receptors. The developed assay provides an efficient method for detection and identification of substances presented in the serum which can influence the binding of insulin to the specific sites as well as its biological effects.