Search PubMed⌕ Search

Biomedical subjects

W Besch

Publications and source records attributed to W Besch.

At least 37 records · Page 2Linked to original sources
PubMed1990

Humoral-mediated anti-islet cytotoxicity in diabetes-prone BB/OK rats--effect on beta-cell function and autologous islets.

The present study investigates whether sera from diabetic BB/OK rats impair beta-cell-specific functions of rat pancreatic islets and whether humoral-mediated cytotoxicity in BB rat sera also affects the autologous islets of normoglycaemic serum donors. Exposure of neonatal rat islets of Langerhans for 20 h to sera from diabetic BB rats and rabbit complement resulted in a decrease in insulin content and in insulin release upon glucose stimulation. However, the extent of islet alterations caused by sera from various BB rats was different as revealed by comparison of the ability of beta-cells to restore their functions during a 48 h recovery period. While the effect of some of the sera was completely reversible, in other cases the insulin content and/or the secretory capacity of the islets could not be fully restored indicating partially irreversible islet alterations due to pretreatment with cytotoxic BB rat serum. The potential effect of anti-islet cytotoxicity in BB rat sera against autologous islets of the serum donor indicated by enhanced 51Cr-release in the presence of rabbit complement was proven in 50% (6/12) of the investigated normoglycaemic BB/OK rats. These results suggest that humoral-mediated cytotoxicity may contribute to beta-cell alteration in this model of diabetes.

Animals↗
PubMed1990

Survival of islet isografts despite cytotoxicity against pancreatic islets measured in vitro.

In this study the in vivo relevance of spleen cell anti-islet cytotoxicity measured in vitro was examined by transplantation of 1,200 syngeneic islets into the spleen of rats receiving 0.5 ml complete Freund's adjuvant (CFA) 24 h before 25 mg/kg body weight streptozotocin (STZ) was given. Control rats receiving CFA or STZ alone remained normoglycaemic whereas 12 out of 21 CFA/STZ-treated rats developed a severe hyperglycaemia after three combined treatments. After the first and second combined treatment splenocytes showed a significant cytotoxicity (p less than 0.01) against syngeneic islets measured by 51Cr-release. This cytotoxicity was not detectable after the third combined treatment. The CFA/STZ-induced diabetes with a residual pancreatic insulin content of only 5% was permanently reversed by intrasplenic islet isografts, but, surprisingly, syngeneic islets survived too, if transplanted at the time when an anti-islet cytotoxicity was measured in vitro. From our results we conclude that the polyclonal activation by complete Freund's adjuvant potentiates the beta cell-toxic effect of a low dose of streptozotocin and induced a transient splenocyte-mediated anti-islet cytotoxicity not recurrent after islet transplantation. Furthermore, our findings reveal a discrepancy between organ-specific immune reactions measured in vitro and those affecting the beta cells in vivo.

Animals↗
PubMed1990

Radioiodination of peptide hormones and immunoglobulin preparations: comparison of the chloramine T and iodogen method.

Following optimization of the reaction conditions, e.g. concentration of oxidizing agents, reaction time, volume of reaction mixture, and pH, chloramine T and the new iodination reagent, Iodogen, were compared for their effectiveness in radioiodination of insulin, glucagon, human growth hormone (hGH), and rabbit anti-mouse IgG. The radioactive peptide hormones prepared were analyzed for the presence of aggregate and breakdown products by polyacrylamide gel electrophoresis (PAGE) at pH 8.9, the rabbit anti-mouse IgG was tested for the presence of low molecular weight damage products by gel filtration on Sephadex G-50. The results demonstrate that with respect to iodine incorporation, specific activity, and immunological reactivity either method can be used to prepare under carefully controlled conditions a wide range of tracers with high specific activity at minimal oxidation damage. These tracers are shown to be highly suitable in radioimmunoassays after previous purification by PAGE and gel filtration, respectively.

Chloramines↗
PubMed1990

Influence of polyethylene glycol (PEG) extraction on the C-peptide determination in sera from insulin-dependent diabetic patients with circulating insulin antibodies.

To investigate whether the unexpectedly high C-peptide levels in some insulin-dependent diabetic (IDDM) patients are due to co-determination of proinsulin bound to circulating insulin antibodies, 36 randomly selected sera from IDDM patients were assayed for C-peptide immunoreactivity (CPR) after polyethylene glycol (PEG) extraction, preceding incubation with proinsulin binding antibodies (LAB + PEG) or without pretreatment of the sera. Recovery of proinsulin was checked by addition of 1 nmol/l proinsulin to all sera. Recovery was found to be 101.5 +/- 4.0%. The mean values of concentrations were significantly lower (p less than 0.001) after treatment with PEG and IAB + PEG compared to the untreated sera. There was also a significant difference (p less than 0.05) between sera extracted with PEG alone or after IAB + PEG-treatment. However, no correlation (p greater than 0.1) was found to bound insulin (total minus free insulin) or to insulin binding capacity (IBC) of the sera. If an antiserum is not available with very low cross-reactivity with proinsulin to determine human C-peptide then sera should not be extracted with PEG alone but after additional incubation with a proinsulin binding antiserum. In spite of the extraction in some cases unexplicably high C-peptide levels may still be expected.

C-Peptide↗
PubMed1990

A sensitive sandwich enzyme immunoassay for measurement of insulin on microtiter plates.

A sensitive enzyme immunoassay for the measurement of insulin in human sera on microtiter plates was established. The assay is based on the sandwich technique with guinea pig anti-insulin IgG adsorbed at microtiter plate wells, human insulin as standard and the same anti-insulin IgG labeled with horseradish peroxidase. Standards used cover a range from 0 to 1200 pmol/l with a detection limit of 10 pmol/l. Coefficients of variation between 3-7% for intraassay precision and 5-11% for interassay precision were obtained over the concentration range of 80-1000 pmol/l. The correlation of EIA-data with those of a commercially available double antibody radioimmunoassay (r = 0.98) could be expressed by the equation: EIA = 0.97 RIA - 57 pmol/l. Normal fasting serum insulin concentrations in healthy subjects ranged from 11-165 pmol/l. In subjects with potentially diminished basal values concentrations of 10-79 pmol/l were determined. The insulin response in oral glucose tolerance tests of children was discussed, who had a constitutional tall stature or Turner's syndrome, respectively.

Adult↗
PubMed1990

Detection of antibodies to islet cell and splenic lymphocytes in diabetes-prone BB and adjuvant-streptozotocin treated Lewis rats by ELISA and immunoblot analysis.

An enzyme-linked immunosorbent assay (ELISA) has been developed to detect antibodies against surface components of rat islet and spleen lymphocytes. Live islet tumor RIN5 AH cells expressing characteristic ganglioside target antigens or rat spleen cells were immobilized onto wells of microtiter polystyrene plates precoated with poly-l-lysine and then incubated with test or normal rat sera. Cell surface-bound antibodies were quantitated after reaction with horseradish peroxidase-conjugated rabbit anti-rat Ig. With this assay, 46% (6/13) of sera from diabetes-prone BB rats and 100% (8/8) of sera from rats treated with complete Freund's adjuvant/streptozotocin (CFA/STZ) prior to immunization with RIN cells had islet cell surface antibodies: 54% (7/13) and 75% (6/8), respectively, were positive for lymphocyte antibodies (defined as the HRP anti-rat Ig binding exceeding the mean + 2SD of control group values). SDS polyacrylamide gel electrophoresis followed by immunoblotting analysis suggested that the islet cell antibodies in sera from the BB and CFA/STZ rats recognized RIN-cell components that were different in their molecular weights. These antigens were not detectable on spleen cells indicating that the ELISA described can be used to quantitate levels of islet cell specific antibodies which possibly reflect beta cell damage with progression to islet degeneration in the rat.

Animals↗
PubMed1989

Is the DNA synthesis of pancreatic islets influenced by the MHC and/or the genetic background?

Neonatal pancreatic rat islets were used to investigate the effect of MHC haplotype RT1u on islet replication measured by incorporation of 3H-thymidine and autoradiography. The special interest in RT1u resulted from the fact that BB rats, which develop an insulin-dependent diabetes, belong to the RT1u haplotype. Thus, the RT1u, when influencing the replication, may be involved in the development of the B-cell deficiency observed in diabetic rats. The incorporation of labeled thymidine into islets obtained from rat strains carrying the RT1u or RTIa haplotype revealed no differences. No matter whether the islets were isolated from Lewis, Wistar, or BB rats, no alteration of islet replication by RT1u islets was found. However, islets obtained from Wistar rats had a higher replicatory activity than islets from Lewis rats. This finding was confirmed by a decrease in DNA synthesis of BB rat islets, in which the Wistar background was substituted by the Lewis background. We conclude from our results that MHC has no influence on islet replication, whereas genetic background seems to be involved in the regulation of islet DNA synthesis.

Animals↗
PubMed1989

Early enhanced beta cell replication in normoglycaemic Wistar rats in response to a subdiabetogenic dose of streptozotocin.

The effect of a single subdiabetogenic dose of streptozotocin (30 mg/kg body weight) on pancreatic insulin content and relative beta cell volume has been studied in normoglycaemic Wistar rats treated with streptozotocin either 2, 3 or 14 days after STZ was given. A single intravenous injection of streptozotocin caused a significant reduction of pancreatic insulin content, islet and beta cell volume, accompanied by a significantly diminished islet insulin content. The glucose- and 3-isobutyl-1-methylxanthine-stimulated insulin secretion was significantly lower in islets obtained 2 or 3 days after streptozotocin injection compared with those of vehicle-treated controls. 14 days after streptozotocin injection, beta cell volume and pancreatic insulin content partially recovered. At this time islet insulin content and secretory responsiveness were enhanced compared to the early phase following streptozotocin administration. The incorporation of [3H] thymidine into islet DNA was significantly enhanced at day 2 or 3 after streptozotocin application, whereas at day 14 the DNA synthesis corresponded to values from control rats. Despite persisting normoglycaemia, the beta cell volume of streptozotocin-treated rats was only 52% vs. control values, thus indicating the unresponsiveness of residual beta cells to compensate spontaneously for the beta cell loss.

Animals↗
PubMed1989

Effect of serum from diabetes-prone BB/OK rats on neonatal rat pancreatic islets and islet cell suspensions.

Humoral-mediated cytolytic activity against neonatal rat pancreatic islet cells as well as islets of Langerhans was detected in sera of newly diagnosed diabetic BB/OK rats by measurement of enhanced 51Cr-release or insulin leakage. Beta-cell-specific functions of islet cell suspensions such as insulin secretion and (pro)insulin biosynthesis can be affected by BB rat serum in vitro.

Animals↗
PubMed1989

Spontaneous recovery of streptozotocin diabetes in mice.

With the aim of developing a model of experimental diabetes by which spontaneous recovery processes can be investigated, we used a lower (1) and a higher (2) dose of streptozotocin (SZ) to find out the lowest possible dose definitely inducing diabetes in mice through beta cell loss but preventing excessive damage to the endocrine pancreas which would exclude restoration processes. After application of SZ (1) to neonatal mice only male animals showed an overt diabetes in adult life. 70 percent of these mice had recovered 15 weeks after appearance of diabetes. Recovery was indicated by normalization of blood glucose, serum insulin, insulin secretion and biosynthesis of isolated pancreatic islets and a reenhancement of the pancreatic insulin content from lower than 10 to 30 percent of control values. After SZ (2) both sexes became hyperglycaemic, and the recovery rate was lower, but was increased by pregnancy in female mice. By means of this model it will be possible to investigate mechanisms and promoting factors of such restoration processes in more detail.

Animals↗
PubMed1989

Persistence of anti-islet ADCC after manifestation of type-1 (insulin-dependent) diabetes.

ADCC (antibody-dependent cellular cytotoxicity) against xenogenic islets in vitro has frequently been found with mononuclear blood cells and heat inactivated autologous serum from newly diagnosed Type-1 diabetics. Anti-islet ADCC, as measured by enhanced 51Cr-release of islets after a 6h-incubation, leads to functional alteration of islets such as a decrease in insulin content and in leucine incorporation. In a follow-up investigation over at least three years it was demonstrated that anti-islet ADCC in vitro disappears, if there is no more C-peptide secretion in vivo. Furthermore, anti-islet ADCC has also not been found in long-term Type-1 diabetics who had no C-peptide secretion but an acutely stimulated immune system due to infectious diseases. An acute immunocytolytic process against pancreatic beta cells in vivo seems to be the precondition for anti-islet ADCC in vitro.

Adolescent↗
PubMed1988

[Phase contrast microscopy demonstration of glomerular erythrocytes in urine: practicable in ambulatory practice?].

The use of phase-contrast microscopy in microhematuria, as proposed in 1979 by Birch and Fairley, renders morphological changes in red cells easily detectable and makes it possible to distinguish glomerular from non-glomerular bleeding. The aim of this study was to evaluate the practicability of this method as a routine laboratory test in ambulatory care. 60 patients with asymptomatic microhematuria (greater than or equal to 2 erythrocytes per high power field) were followed up over a one-year period. All patients were investigated by intravenous pyelography, ultrasound of urinary tract and three cytological examinations of the urine. The description of urine samples was done with phase-contrast microscopy by a first investigator at the beginning of the study and by a second after 12.8 months, blinded to clinical results and previous examinations. In 21 patients a definitive diagnosis was possible. In 18 patients the morphologic descriptions of the two investigators correlated with the clinical results. Only in two patients with established diagnosis there were differences between the urine description of the two investigators, and in one patient the interpretations of both investigators were wrong. These incorrect descriptions concerned patients with low-grade microhematuria. Thus, phase-contrast microscopy is a practicable method for the practitioner's use as a routine laboratory investigation. In low-grade microhematuria the method seems to be of minor value.

Ambulatory Care↗
PubMed1988

The effect of mixing regular- and intermediate-acting insulins on their absorption and biological activity.

Many insulin-dependent diabetics are now treated with different regimens of short- and intermediate-acting insulins. The new L-insulin S.N.C. (Berlin-Chemie) is thought to be appropriate for a combination with regular insulin. We investigated whether or not absorption kinetics and biological activity changes when L-insulin S.N.C. is mixed with regular insulin S.N.C. Eight non-obese healthy male subjects were connected to the BIOSTATOR. After an overnight fast 0.1 U of insulin S.N.C. and 0.3 U per kg b.w. of L-insulin S.N.C. or Monotard MC insulin (NOVO) were administered either in a mixture or separately. Injecting L-insulin S.N.C. and insulin S.N.C. in a mixture there was some, but not a statistically significant, delay in absorption of insulin. A mixture of insulin S.N.C. and Monotard provoked a marked delay of absorption during the first two hours. To examine biological activity we used the BIOSTATOR (glucose controlled dextrose infusion system). Glucose infusion rate necessary to keep blood glucose steady state concentration was taken as a measure of insulin action. Whereas mixing of insulin S.N.C. and L-insulin S.N.C. only slightly changed the profile of biological activity, a mixture of insulin S.N.C. and Monotard has a clear-cut higher potency during the late phase when compared with separately injected insulins. Therefore, insulin S.N.C. and Monotard should not be mixed. A mixture of insulin S.N.C. and L-insulin S.N.C. is also not recommended but can be done in exceptional cases.

Absorption↗
PubMed1988

The endocrine pancreas of BB/OK-rats before and at diagnosis of hyperglycaemia.

From 148 BB-rats 76 animals (51.4%) developed hyperglycaemia within 55th to 178th day of life. Already before diagnosis of diabetes a great variety in pancreatic insulin content and morphometrically determined relative beta-cell volume density were visible. Despite of this great heterogeneity two types in the course of beta-cell destruction could be observed. Verified by individual retrospective analysis one part of the rats is characterized by a more chronical course of diabetes development (pancreatic insulin content and beta-cell mass already more than 60 days before diagnosis of hyperglycaemia significantly reduced, occurrence of insulitis) whereas other animals show an acute form of beta-cell destruction (still about 20 days before diagnosis of diabetes pancreatic insulin content and relative beta-cell volume density in a normal range). Besides a drastical reduction of pancreatic insulin content at the time of diabetes diagnosis intense mononuclear infiltrations in the islets of Langerhans causing their destruction are demonstrable. As a result the residual beta-cell volume is significantly reduced. Rats with a plasma glucose higher than 13 mmol/l at diagnosis do not have any demonstrable beta-cells and an insulin content on the lowest detection limit of the method. In animals with a more mild hyperglycaemia (plasma glucose 8.3 to 13 mmol/l) there are rats with and the other ones without immunohistochemically detectable beta-cells. We conclude that prospective statements on the development of diabetes on the strength of analysis of residual beta-cells and pancreatic insulin content are not possible at present. We suppose that there exist different processes of beta-cell destruction which have to be investigated in further studies including immunological parameters.

Animals↗
PubMed1988

Diabetogenic effects of N-nitrosomethylurea in rats.

The influence of a single i.v. injection of 75 or 175 mg N-nitrosomethylurea (NMU)/kg into Wistar rats on plasma glucose, i.p. glucose tolerance, plasma insulin and glucagon, glucose stimulated insulin secretion and release as well as on the biosynthesis of insulin in isolated pancreatic islets have been investigated at set intervals between the 3rd day and the 6th week post injectionem. At the end of the experimental period all NMU-treated animals were alive but had significantly reduced body mass. All animals remained normoglycemic even after the glucose load. 3 days after drug application plasma insulin decreased only insignificantly in the 175 mg NMU-group, whereas plasma glucagon was significantly reduced in both drug-groups; after 3 and 6 weeks the reduction of plasma glucagon was still about 30 percent. In isolated islets the glucose stimulated insulin secretion, the percent stimulation of insulin release and insulin biosynthesis were significantly decreased.

Animals↗
PubMed1988

Intraindividual comparison of pharmacokinetics of insulin after intravenous, portal, subcutaneous and peritoneal administration.

To compare the kinetics of praehepatic and of posthepatic administered insulin, short term insulin deprived diabetic dogs were sequentially injected with 200 mU/kg of a monocomponent porcine insulin using either the intravenous, portal, subcutaneous or peritoneal route. After peritoneal insulin was applied, the peripheral plasma insulin levels increased immediately, their maxima were in the same range as after subcutaneous injection but the duration of elevation was shorter. There were portal-peripheral insulin-quotients greater than 1 after peritoneal and portal insulin administration but quotients less than 1 after subcutaneous and intravenous application. The time constant of insulin elimination was identical regardless of whether the praehepatic or the posthepatic route was used for application. The effectiveness of the administered insulin dose on blood glucose was found to be dependent on the posthepatic elevation of plasma insulin and its duration. The decrease in glycemia was initially identical in all tests but, on the whole, it was smaller after the two intravascular routes were used because of the shorter duration of elevated insulin levels. It is concluded that in an optimized management of insulin-dependent diabetes, the regime (doses and intervals or algorithms) must be adapted to the pharmacokinetic implications of the employed route of application.

Animals↗