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Biomedical subjects

W A Scherbaum

Publications and source records attributed to W A Scherbaum.

At least 109 records · Page 6Linked to original sources
PubMed1999

Prevalence of chronic complications, metabolic control and nutritional intake in type 1 diabetes: comparison between different European regions. EURODIAB Complications Study group.

This study compares the prevalence of chronic complications, the quality of metabolic control and the nutritional intake in people with type 1 diabetes in different European regions. The EURODIAB Complications Study included a sample of 3250 European patients with type 1 diabetes stratified for gender, age and diabetes duration. All examinations were performed using standardised, validated methods. HBA1c, LDL-cholesterol and fasting triglycerides were higher in the eastern European centres than in the southern or north-western European centres. Acute (severe ketoacidosis, severe hypoglycaemia) and chronic diabetes complications (retinopathy, nephropathy, neuropathy, cardiovascular disease) were all considerably more frequent in the eastern European centres. HbA1c was lower in the German centres than in the total EURODIAB cohort or in the north-western European centres, but severe hypoglycaemia and proliferative retinopathy were more common. Persons from the eastern European and the German centres consumed undesirably high amounts of cholesterol, total and saturated fat. Overall, improvements in the prevention, detection and management of diabetes complications in persons with type 1 diabetes are essential throughout Europe, particularly in eastern European regions. Since elevated LDL-cholesterol levels and hypertension were strikingly common in this relatively young cohort of European people with type 1 diabetes, generally more attention should be directed towards an adequate management of these cardiovascular risk factors.

Adult↗
PubMed1999

Detection of autoantibodies to the diabetes-associated antigen IA-2 by a sensitive enzyme-linked immunosorbent assay.

The tyrosine phosphatase like protein IA-2 is an important autoantigen in insulin-dependent diabetes mellitus (type 1 diabetes). Autoantibodies to IA-2 (IA-2A) are present in the serum of patients with type 1 diabetes even before the onset of the disease. Previously, we reported on a radioimmune assay to detect IA-2A, using E. coli-derived 125I-labelled IA-2 as antigen. Although this assay could be shown to be equivalent to the common reference method for IA-2A detection (radioligand assays using in vitro synthesised 35S-methionine labelled antigen), the disadvantages of both assays with respect to synthesis and handling of the radioactive antigen limit their use in routine laboratories. In this study, we have evaluated a non-radioactive enzyme-linked immunosorbent assay (ELISA) for the simple detection of IA-2A. We report on an ELISA where the biotinylated intracytoplasmic part of IA-2 (IA-2ic) is captured on streptavidin-coated plates. The sensitivity of the ELISA was similar to the validated radioligand assay, as it detected 47 of 69 (69%) patients with type 1 diabetes as compared to 46 of 68 (67 %) with the reference method for IA-2A detection (radioligand assays using in vitro synthesised 35S-methionine labelled antigen). Only 2 of 50 (4%) patients with autoimmune thyroid disease and 1 of 114 (1 %) healthy controls were detected in the ELISA, confirming specificity. There was a significant correlation between the ELISA and the radioligand assay (r = 0.64, p<0.001). We conclude that this ELISA is suitable to detect IA-2A in the serum of patients with type 1 diabetes with a similar sensitivity and specificity to the radioligand assay. This ELISA will allow rapid and simple measurement of IA-2A where the radioligand assay is inconvenient or not available.

Antibody Specificity↗
PubMed1999

Lymphocytes stimulate dehydroepiandrosterone production through direct cellular contact with adrenal zona reticularis cells: a novel mechanism of immune-endocrine interaction.

Adrenal androgen production was reduced by 80% in patients receiving T lymphocyte-suppressive medications compared to that in age-matched controls. In vitro, however, neither tacrolimus nor cyclosporin A reduced dehydroepiandrosterone (DHEA) release by adrenocortical cells. Therefore, we examined the potential role of lymphocytes in adrenal androgen production, using cocultures of human T lymphocytes and adrenocortical primary or transformed cells. Co-cultures led to a 4-fold elevation of DHEA levels (490.4 +/- 94.8% over basal), which was greater than the increase observed after the addition of maximal concentrations of ACTH (117.4 +/- 14.8%). Separation of cells by semipermeable membranes abolished this effect, and transfer of leukocyte-conditioned medium had little androgen-stimulating effect. These data suggested that the observed stimulation of androgen secretion required cell contact rather than soluble paracrine factor(s). Furthermore, we examined human adrenal glands for the presence of T lymphocytes and contact between these cells and steroid-secreting cells of the zona reticularis. Indeed, T lymphocytes expressing CD4 and CD8 antigens were present within human adrenal zona reticularis by immunohistochemical subtyping. Electron microscopic analyses demonstrated direct cell-cell contact between T lymphocytes and adrenocortical cells in situ. This study provides evidence for a novel mechanism of immune-endocrine interactions of direct T lymphocyte-adrenocortical cell contact-mediated stimulation of adrenal androgen secretion.

Adrenocorticotropic Hormone↗
PubMed1999

Direct binding of thyrotropin receptor autoantibody to in vitro translated thyrotropin receptor: a comparison to radioreceptor assay and thyroid stimulating bioassay.

Graves' disease is characterized by the presence of autoantibodies to the thyrotropin receptor (TSHR), which are pathogenic and responsible for disease activity. It is well recognized that the autoantibodies are heterogeneous and recognize a number of different conformational dependent epitopes on the TSHR. In this study, we have extended our previous observations to study the interaction of Graves' disease autoantibodies with TSHR ectodomain produced by in vitro transcription and translation reaction. The specific activity of the translated TSHR ectodomain has been increased by a log fold by adding an efficient ribosome binding Kozak sequence before the translation initiation codon as well as double labelling with 35S-methionine and 35S-cysteine during the translation reaction. Addition of canine pancreatic microsomes to the translation mix showed that the glycosylation of TSHR ectodomain did not occur efficiently for the nascent receptor protein. In order to determine the specificity and sensitivity of the improved assay with nonglycosylated TSHR ectodomain, we have studied 331 sera from Graves' disease patients and as controls 100 sera from patients with nonthyroid autoimmune disorders as well as sera from 200 normal control subjects with no family history of thyroid autoimmunity. With this large cohort of sera from Graves' disease and control individuals, 25% of Graves' disease sera immunoprecipitated the dual labeled, in vitro transcribed and translated TSHR ectodomain, exceeding the 98th percentile of the control sera. There was no correlation between the autoantibodies that immunoprecipitate the in vitro translated TSHR ectodomain and those that inhibit iodinated TSH binding in the radioreceptor assay and those with biological activity in a bioassay. The data are consistent with the finding that a proportion of Graves' disease autoantibodies can interact directly with TSHR ectodomain produced by in vitro transcription and translation. However, in contrast to the wide use of similar translation and immunoprecipitation assays to measure other autoantibodies for the diagnosis of autoimmune disorders, such as type 1 diabetes, the TSHR immunoprecipitation on its own is unsuitable for diagnosis of Graves' disease.

Animals↗
PubMed1998

[Progress in molecular medicine: "laser capture microdissection"].

BACKGROUND: With the unravelling of the human genome, we now face the challenge of defining the function and clinical relevance of single genes. To do this, we should be able to isolate normal and diseased cells from complex tissue structures to make them accessible to sensitive molecular analyses. Laser Capture Microdissection (LCM) was developed to meet this challenge. METHOD AND APPLICATION: LCM allows the precise dissection of cells with the help of a laser beam under direct visualization in the microscope, and the sterile transfer of these cells into a DNA or RNA isolation buffer. The technique is ideal for investigating cell-cell interactions, for performing mutation analyses, and for the production of high-quality cDNA libraries. Expression studies of known and unknown genes are currently employed successfully to define tissue- and simple cell-specific patterns which help elucidate the etiology and pathogenesis of colon, lung, breast, prostate, adrenal, ovary, and other organ tumors. The LCM system developed at the NIH is, therefore, an important part of the Cancer-Genome Anatomy Project (CGAP), which sequences and publishes the structures of genes that are expressed in human tumors. In combination with the modern cDNA arrays, it will thus be possible to analyze the expression of several thousands of genes in one step and to develop individual therapeutic strategies in the not too distant future. CONCLUSIONS: The LCM is a major advance in molecular medicine, enabling us to combine highly-sensitive gene analysis techniques with conventional histologic and morphologic methods. Applications range from research to diagnosis, and to monitoring disease progression.

Cell Separation↗
PubMed1998

A simplified procedure for the isolation of immunoglobulins from human serum using a novel type of thiophilic gel at low salt concentration.

By coupling 3-(2-mercaptoethyl)quinazoline-2,4(1H,3H)dione (MECH) to divinyl sulfone activated agarose, a novel thiophilic matrix was obtained which allows the binding of immunoglobulins from different sources. In contrast to other thiophilic gels, antibodies are bound at low ionic strength and can easily be desorbed in intact form by elution with dilute alkali. The potential of using the MECH-gel was demonstrated by the purification of antibodies from human and animal (goat, rabbit, mouse) sera. The functional integrity of the purified antibodies was established with cytoplasmic islet cell antibodies from the sera of patients with type I diabetes and autoantibodies against thyroid peroxidase from patients with Graves' and Hashimoto's disease.

Animals↗
PubMed1998

[B-mode ultrasound and modern Doppler ultrasound methods in diseases of the thyroid gland and parathyroid glands].

Color-coded duplex sonography is the main innovation in diagnostic ultrasound in recent years. It allows quantification of tissue vascularity and appreciation of vascular morphology. Due to the unique thyroid hypervascularity in Graves' disease this diagnosis can be made with color Doppler sonography alone. The decrease of vascularity during the course of disease is a relevant parameter throughout the follow-up. Hypervascularity is also observed within areas of inflammatory infiltration in thyroiditis, but the level is lower than in Graves' disease. Sonographic differentiation of benign from malignant thyroid nodules is not possible yet. Color-coded duplex sonography is not useful in the initial detection of parathyroid masses, but may be helpful in distinguishing parathyroid lesions from other cervical masses.

Diagnosis, Differential↗
PubMed1998

Relation of fibre intake to HbA1c and the prevalence of severe ketoacidosis and severe hypoglycaemia. EURODIAB IDDM Complications Study Group.

The effect of dietary fibre intake on glycaemic control is still controversial. This study analysed the intake of natural dietary fibre in patients with Type I diabetes mellitus enrolled in the EURODIAB IDDM Complications Study to determine any associations with HbA1c levels and with the prevalence of severe ketoacidosis or severe hypoglycaemia. Dietary intake was assessed by a 3-day dietary record. The relation between intake of fibre (total, soluble and insoluble) and HbA1c was examined in 2065 people with Type I diabetes. Associations with severe ketoacidosis (requiring admission to hospital) and severe hypoglycaemia (requiring the help of another person) were analysed in 2687 people with Type I diabetes. Total fibre intake (g/day) was inversely related to HbA1c (p = 0.02), independently of carbohydrate intake, total energy intake and other factors regarding lifestyle and diabetes management. Severe ketoacidosis risk fell significantly with higher fibre intake (p = 0.002), with an adjusted odds ratio of 0.48 (95 % confidence interval 0.27 to 0.84) in the highest quartile ( > or = 23.0 g fibre/day) compared with the lowest quartile ( < or = 13.7 g fibre/day). The occurrence of severe hypoglycaemia was not related to fibre intake. Beneficial effects of fibre on HbA1c and the risk of severe ketoacidosis were particularly pronounced in patients from southern European centres. This study shows that higher fibre intake is independently related to a reduction in HbA1c levels in European people with Type I diabetes. Furthermore, increased fibre intake may reduce the risk of severe ketoacidosis. These beneficial effects were already observed for fibre intake within the range commonly consumed by people with Type I diabetes.

Adolescent↗
PubMed1998

Immunological heterogeneity in type I diabetes: presence of distinct autoantibody patterns in patients with acute onset and slowly progressive disease.

Type I diabetes mellitus may represent a heterogeneous disorder with a distinct pathogenesis in patients with young and adult onset of the disease. To investigate whether serological markers directed to different autoantigens have the potential to distinguish acute onset from slowly progressive Type I diabetes we analysed antibodies to tyrosine phosphatases IA-2/ICA512 (IA-2A) and IA-2beta/phogrin (IA2betaA), antibodies to GAD65 (GADA) and cytoplasmic islet cell antibodies (ICA) in a non-selected group of diabetic patients clinically classified as having Type I or Type II diabetes at diagnosis. Both IA-2A and IA-2betaBA were found to be positively associated with onset before the age of 20 years and the presentation of classical features of Type I diabetes. In Type I diabetes 56 % (112/200) of patients were positive for IA-2A and 38 % (76/200) for IA-2betaA. In contrast, only 1 of 785 (0.1 %) patients with Type II diabetes had IA-2A and all of them were negative for IA-2betaA (p < 0.001). Among the patients with Type II diabetes 7.6% (n = 60) were ICA positive and 2.8% (n = 22) had GADA suggesting the presence of slowly progressive Type I diabetes. GADA were found in 8 of 60 (13.3 %) ICA positive subjects which was lower than the percentage detected in patients with acute onset of diabetes (115/157 73.2%) (p < 0.001). Blocking of double antibody positive sera showed that only 3 of 8 (37.5 %) patients with slowly progressive diabetes had ICA restricted to GAD or IA-2 whereas ICA were completely inhibited in 12 of 20 (60.0 %) patients with Type I diabetes. Among 193 patients with Type II diabetes available for follow-up, 35 % of ICA positives, 58 % of GADA positives and 60 % of those positive for both markers required insulin by 3 years. However, using strict criteria for the switch to insulin treatment the corresponding sensitivity of each marker was only low (9%, 10% and 5%). We show that clinical subtypes of Type I diabetes are associated with distinct humoral autoimmunity. IA-2A and GADA were associated with classical features of Type I diabetes whereas GADA and an uncharacterized ICA subspecificity indicate slowly progressive disease.

Acute Disease↗
PubMed1998

The role of amylin in the physiology of glycemic control.

Amylin is a 37-amino acid peptide hormone, discovered in 1987, which is co-located and co-secreted with insulin by the pancreatic beta-cells in response to nutrient stimuli. Like insulin, there is a deficiency of amylin in people with type 1 diabetes, while the changes in plasma amylin concentrations in people with impaired glucose tolerance and type 2 diabetes parallel those of insulin. It is well established that insulin regulates glycemic control by promoting glucose disposal. This paper reviews evidence from studies in animals and people with diabetes that amylin regulates the inflow of glucose to the circulation by delaying nutrient delivery and, thus, the appearance of meal-derived glucose, and also suppresses glucagon secretion in the postprandial period. It is suggested, therefore, that the actions of amylin complement those of insulin, and that the problems of glycemic control which continue to exist in people with diabetes, despite insulin replacement therapy, may be attributable to a deficiency in amylin. Preclinical and clinical studies with pramlintide, a synthetic analogue of human amylin, are also included in this brief review.

Amyloid↗
PubMed1998

Substitution therapy after surgery for autonomous adenomas.

A substitution therapy with L-thyroxine subsequent to surgery on the thyroid gland due to autonomous dysfunction depends in particular on the extent of resection. A specific postoperative therapy is not necessary in areas with sufficient iodine supply if the remaining part of the thyroid gland guarantees a euthyroid metabolic state. The radicalness of the intervention is dependent on the existence of an unifocal, multifocal or disseminated autonomy. Surgery due to disseminated autonomy always necessitates a substitutive therapy with thyroid hormones and in most cases surgery due to multifocal autonomies require the same treatment. The therapy is initiated with a dose of 1.5 microg L-thyroxine per kg body weight. Suppression of the TSH level is not necessary. Due to the greater risk of recurrence the L-thyroxine administration should be complemented in areas of iodine deficiency with approximately 100-200 mg iodide. After operations with functionally adequate thyroid remnants (8-10 ml), an exclusive prophylaxis with 200 mg iodide can be implemented. The result of surgery should be sonographically documented three months after the intervention.

Adenoma↗
PubMed1998

Application of a bioassay with CHO cells for the routine detection of stimulating and blocking autoantibodies to the TSH-receptor.

The importance of bioassays measuring stimulating and blocking autoantibodies to the TSH-receptor (TSH-R) by their effect on cAMP production in CHO cells transfected with the recombinant TSH-R is increasingly recognized. The standard technique for this bioassay is cumbersome, as it involves purification of serum IgG with polyethylene glycol (PEG) and resuspension in hypotonic buffer. We have therefore established a simpler approach for the detection of stimulating and blocking autoantibodies using JP09 CHO cells and unfractionated human serum. The cAMP concentration was measured by a highly sensitive commercial radioimmuno assay. Thyroid stimulating autoantibodies (TSAb) were present in 107 out of 126 patients with Graves' disease (85%) and in 4 out of 40 patients with Hashimoto's thyroiditis (10%). Specificity was confirmed by the fact that only 1 patient with insulin dependent diabetes mellitus (IDDM) out of 64 patients with different non-thyroid autoimmune disorders (46 with IDDM, 10 with stiff man syndrome and 8 with rheumatoid arthritis) and 2 out of 100 healthy controls (2%) were positive in this assay. In the subgroup of hyperthyroid Graves' disease patients 76 out of 83 (92%) had TSAb and the same number had TSH binding inhibiting immunoglobulin (TBII), as assessed by the commercial TRAK assay. Although both antibody types showed only a weak correlation (r = 0.30), a combination of TSAb and TBII detected 98% of all Graves' patients and 99% of the hyperthyroid subgroup. Thyroid blocking autoantibodies (TBAb) were measured in 4 out of 24 TSAb negative patients with Graves' disease (17%), who were hypothyroid and positive for TBII. A comparison of our bioassay with the standard bioassay using PEG precipitation showed a good correlation (r = 0.76,p < 0.001), demonstrating the feasibility of the simplified assay for the routine detection of TSAb and TBAb in Graves' disease.

Adult↗
PubMed1998

Recombinant IA-2 expressed in E. coli can be used for the routine detection of autoantibodies in Type-I diabetes.

We have investigated the possibility of measuring autoantibodies to IA-2 (IA-2A) using recombinant protein expressed in E. coli in a new radioassay. The intracellular part of IA-2 (IA-2ic) was expressed in E. coli as a biotinylated fusion protein and affinity-purified on a streptavidin column. The average yield of IA-2ic was about 1 mg purified protein from one litre of culture medium with E. coli. We could demonstrate the immunological activity of this material by blocking the autoantibody reactivity to in vitro synthesised IA-2ic. The IA-2ic fusion protein was then radiolabelled with 125I, purified by HPLC, and used in an immunoprecipitation assay for the detection of IA-2A. Sera from 46 of 68 (67%) patients with Type-I diabetes were positive by this radioassay, in contrst to only 2 of 50 (4%) patients with autoimmune thyroid disease and 1 of 114 (1 %) controls. There was a correlation between this radioassay and the previously established radioligand assay using synthesized 35S-methionine-labelled IA-2ic in vitro (r = 0.79, p < 0.001). We conclude that E. coli-derived IA-2 has the correct immunogenic conformation, and can be used for the detection of IA-2A with a similar sensitivity and specificity as the validated radioligand assay. This new assay can facilitate the measurement of IA-2A in routine laboratories where the radioligand assay is inconvenient or not available.

Adolescent↗
PubMed1998

mRNA levels of membrane-type 1 matrix metalloproteinase (MT1-MMP), MMP-2, and MMP-9 and of their inhibitors TIMP-2 and TIMP-3 in normal thyrocytes and thyroid carcinoma cell lines.

Thyroid cancer can degrade basement membranes and invade tissues. This depends on a cascade of matrix metalloproteinases involving membrane-type 1 matrix metalloproteinase (MT1-MMP), MMP-2, and MMP-9. We analyzed the expression and role of these MMPs and their specific inhibitors TIMP-2 and TIMP-3 in human highly purified thyroid epithelial, C 643, HTh 74, SW 1736, and 8505 C thyroid carcinoma and thyroid-derived fibroblast cell cultures. The effect of phorbol-myristate acetate (PMA), and of the inflammatory cytokines interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-alpha) on MMP and TIMP mRNA levels were monitored by semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) including an internal homologous competitor fragment. The highest MT1-MMP mRNA levels were found in thyroid-derived fibroblasts. The MT1-MMP mRNA expression was increased up to 10-fold by PMA, while all other growth factors tested had only negligible effects. The thyroid carcinoma cells themselves did not seem to play a crucial role in the production of MT1-MMP in thyroid tumors. Higher MMP-2 mRNA levels were found in all cell types investigated. The highest MMP-2 mRNA levels were determined in thyroid-derived fibroblasts and HTh 74 cells. We found a lack of MMP-2 response to IL-1, TNF-alpha, and phorbol esters. In unstimulated cells, MMP-9 mRNA was found near the detection limit or at low levels. In nearly all cell types, treatment with PMA, IL-1, and TNF-alpha caused an increase of the MMP-9 mRNA levels. The results of basal and stimulated MMP-2 and MMP-9 mRNA expression were confirmed at the protein level by gelatin zymography. TIMP-2 and TIMP-3 mRNAs were expressed at high levels. In contrast to the basal TIMP-3 mRNA levels, which varied over a great range, there were no striking differences the cell types from analyzing TIMP-2 mRNA. There were no or only slight stimulatory effects on TIMP-2 and TIMP-3 mRNA expression by IL-1, TNF-alpha, and PMA. Taken together, most enzymes of the MT-MMP/MMP class of proteases facilitating invasion of thyroid tumor cells seem to have been produced by fibroblasts, not by the tumor cells themselves. However, some dedifferentiated thyroid tumor cell lines may be capable of secreting some of these enzymes, as in the case of HTh 74 cells.

Cell Line↗
PubMed1998

Expression of Ob receptor in normal human adrenals: differential regulation of adrenocortical and adrenomedullary function by leptin.

The major effects of leptin, an adipostatic hormone produced in fat tissue, are exerted through the hypothalamic-pituitary-adrenal axis and the systemic sympathetic/adrenomedullary system at the level of the central nervous system. Here, we examined the direct effects of leptin on the adrenal gland, a peripheral end organ of both the hypothalamic-pituitary-adrenal axis and the sympathetic/adrenomedullary system. As cortical and chromaffin tissues are intermingled in the human adrenal, we employed the novel technique of laser capture microdissection to analyze these systems separately. Functional full-length leptin receptor messenger ribonucleic acid and all human isoforms Ob219.1-3 were demonstrated by RT-PCR in both cortical and medullary tissue. Immunohistochemical staining of leptin receptor protein, however, demonstrated a strong signal only in the adrenal cortex, whereas there was weak positive staining in the medulla. Corticotropin (ACTH)-induced adrenal aldosterone, cortisol, and dehydroepiandrosterone secretion was inhibited by leptin in a concentration-dependent manner, whereas this hormone had no significant effect on catecholamine release by primary cultures of human adrenal chromaffin cells. Leptin itself was not expressed in human adrenal tissue, excluding a local paracrine or autocrine function of this peptide. In conclusion, this is the first report identifying functional leptin receptor in human adrenal tissue and showing a differential action of leptin on human adrenocortical and chromaffin hormone production. This peripheral action of leptin on the adrenal gland provides an additional important link between the human stress response and body weight regulation.

Adrenal Cortex↗
PubMed1998

The effect of corticotropin-releasing hormone (CRH) on the adrenal medulla in hypophysectomized rats.

CRH occurs in the adrenal medulla of rats. We were interested to know whether CRH affects meduallary chromaffin cells in the absence of ACTH. We investigated the morphological changes of the adrenal medulla in Sprague Dawley rats with light and electron microscopy in normal rats, hypophysectomized rats, and hypophysectomized rats following injections of CRH (10 micrograms = 3 nmol for three days). Chromaffin cells were characterized by immunohistochemistry (anti-tyrosine hydroxylase). At light microscopy level chromaffin cells of hypophysectomized rats were reduced in number. On electron microscopy the number of granules and cell organelles were decreased. Following injections of CRH the medulla regained a more compact texture with cell organelles homogenously distributed, but with chromaffin granules still being reduced in number. Immunohistochemistry allowed the identification of chromaffin cells located within the adrenal cortex. In hypophysectomized rats these cells showed fewer signs of alterations compared to cells located within the medulla itself and had recovered better after treatment with CRH. In conclusion, CRH seems to exert a trophic effect on chromaffin cells in the absence of pituitary ACTH. This observation may provide further evidence for a close interaction of the two neuroendocrine stress systems.

Adrenal Medulla↗