On the occurrence of the diabetes-associated antigen GAD 65 in human sera.

BACKGROUND: Glutamic acid decarboxylase (GAD 65) is involved as an antigen in diabetes mellitus type 1 and is generally considered to be located intracellularly in pancreas ß-cells. In this study we demonstrate its appearance in 64 human sera samples representing a cross-section of a blood bank. METHOD: The proof of GAD 65 in sera was done using an enzyme-linked immunosorbent assay (ELISA) setup where it was detected by interaction with corresponding antibodies labeled with an enzyme. The enzyme catalyzes a substrate reaction, resulting in a change of color, that is used for the quantitative evaluation of the antigen-antibody interaction. RESULT: The measurements showed that GAD 65 exists in various amounts in the sera of blood donors, with an average concentration of 58.00 ng/ml. The correlation analysis of samples stored at -80° C and at room temperature demonstrates the stability of GAD 65 at room temperature. The correlation coefficient between the GAD 65 concentrations of samples stored at room temperature and of the same samples after 1 week shows that the molecule remains stable. CONCLUSION: Our results encourage us to propose antigen GAD 65, due to its frequency in human sera in different concentrations and its stability, as a biomarker for the early diagnosis of type 1 diabetes and related inflammations.

The diabetic antigen glutamic acid decarboxylase (GAD 65) in the human peripheral blood.

BACKGROUND: Glutamic acid decarboxylase (GAD 65) is a diabetes-associated antigen which is generally considered to be strictly intracellular. In order to better understand autoimmunity, this study demonstrates the appearance of GAD 65 in the peripheral human blood and presents implications for the diagnosis and therapy of some autoimmune diseases. METHODS: The GAD 65 molecules are detected by their interaction with monoclonal antibodies labeled with dyes in an experimental setup with fluorescence correlation spectroscopy (FCS). These interactions result in changes in Brownian motion measured as fluorescence fluctuations. Sera from 153 patients with diabetes mellitus type 1 and controls were investigated. To enable the representation of the molecule as a model for further discussions, we present structural visualizations of its hydrophobic properties, leading to possible interactions with the cell membrane lipids and epitope locations. RESULTS: The GAD65 antigen could be measured with a sensitivity of 2.65 microg/ml in 'clean systems' resulting from spiking experiments and human sera. The GAD 65 antigen could be identified in 8 patient sera: 4 children with diabetes mellitus type 1 and 4 adults initially taken as controls but who retrospectively showed signs of autoimmunity. CONCLUSION: We conclude that these findings are of significance for the concept of autoimmunity, i.e. in an initial step the immune system is primed by its accessibility to GAD 65. Our experimental results may also be important for the therapy of diabetes mellitus type 1 and other autoimmune diseases by the passive administration of GAD 65 antibodies.

[Sense and nonsense in antibiotic therapy for respiratory tract infections]. / Sinn und Unsinn der Antibiotikatherapie respiratorischer Infekte.

Whether an antibiotic successfully eradicates pathogens depends on the pathogens involved, on pharmacokinetics and bioavailability in the target tissue, and on the antimicrobial resistance of the pathogen. Other determinants are drug interactions, individual risk factors, age and compliance with respect to correct dosage and duration of therapy. In many cases, antimicrobial therapy is begun on an empirical basis, because the responsible pathogen can be identified in only half of all respiratory infections. The eradication of the pathogen has to be the first aim if treatment is to be curative and the development of resistance prevented. Long-term prevention of antimicrobial resistance will require a more critical prospective evaluation of the prescription of antibiotics. This paper considers rational and irrational measures in the antimicrobial therapy of respiratory infections.

Detection of tumour cells in the peripheral blood of patients with breast cancer. Development of a new sensitive and specific immunomolecular assay.

Malignant cells in the peripheral blood of patients with solid tumours are of considerable importance for the prognosis and therapeutic correlation. Their detection however is difficult due to lack of sensitivity, specificity and technical problems in standardisation. In this original article we show a new sensitive method overcoming the hitherto known difficulties by combining traditional antibody-techniques with a RT-PCR. Due to this method 2 tumour cells within 5 ml of peripheral blood can be detected in spiking experiments.

Cardiac myxomas: 24 years of experience in 49 patients.

OBJECTIVES: In this single-center study we reviewed our experience with a significant number of cardiac myxoma cases occurring over the past two decades. PATIENTS AND METHODS: Cardiac myxomas represented 86% of all surgically treated cardiac tumors at our center. Specifically, there were 49 consecutive patients, each with at least one myxoma. A detailed clinical, immunological, and echocardiographic long-term examination of 37 patients revealed one recurrent myxoma. RESULTS: Most myxomas originated from the left atrium (87.7%), but also much less frequently from the mitral valve (6.1%), from the right atrium (4.1%), and from the left and right atria (2.0%). The myxomas produced a prolapse into the left ventricle in 40.8% of the patients, mitral stenosis in 10.2%, and threatened left ventricular outflow tract obstruction in 2.0%. Multiple myxomas were found in 20.4% of the patients. Cardiac signs appeared in 93.9% of the patients. Preoperative embolic events had occurred in 26.5%. Immunologic alterations were present in 87.5%. For resection, a bilateral atriotomy was used. An additional aortotomy was needed to expose one mitral valve myxoma. Postoperatively, 81.1% of the patients remained without cardiac symptoms. The early mortality rate was 2.0% and the late mortality rate was 6.1%. Long-term prognosis was excellent with an actuarial survival rate of 0.74. Specific immunologic alterations were found in 71.4% of the patients. The actuarial freedom from reoperation of the myxoma was 0.96. The rate of reoperations was low with 2.0% after 24 years. CONCLUSIONS: Myxomas were usually detected and operated on in symptomatic patients. A high index of suspicion seems important for early diagnosis. Immunologic findings may play an additional role in confirming the diagnosis and the recurrence of a myxoma. Immediate surgical treatment was indicated because of the high risk of embolization or of sudden cardiac death. Also, a familial genesis must be excluded in myxoma patients.

[Soluble receptors of TNF-alpha in rheumatoid arthritis]. / Rastvorimye retseptory faktora nekroza opukholi pri revmatoidnom artrite.

The serum level of soluble TNF-alpha receptors with molecular mass 55 kDa (sTNF-a55R) was measured by enzyme immunoassay with commercial kits in 30 patients with rheumatoid arthritis (RA) and 38 healthy donors. High sTNF-a55R serum levels were registered in 90% of RA patients. These levels correlated with RA activity by DAS. Thus, assay for sTNF-a55R can be used for assessing RA activity.

Ultrasensitive detection and identification of fluorescent molecules by FCS: impact for immunobiology.

An experimental application of fluorescence correlation spectroscopy is presented for the detection and identification of fluorophores and auto-Abs in solution. The recording time is between 2 and 60 sec. Because the actual number of molecules in the unit volume (confocal detection volume of about 1 fl) is integer or zero, the fluorescence generated by the molecules is discontinuous when single-molecule sensitivity is achieved. We first show that the observable probability, N, to find a single fluorescent molecule in the very tiny space element of the unit volume is Poisson-distributed below a critical bulk concentration c*. The measured probability means we have traced, for example, 5 x 10(10) fluorophore molecules per ml of bulk solution. The probability is related to the average frequency, C, that the volume of detection contains a single fluorescent molecule and to the concentration, c, of the bulk solution. The analytical sensitivity of an assay is calculated from the average frequency C. In the Goodpasture experiment, we determined as analytical sensitivity a probability of 99.1% of identifying one single immune complex. Under these conditions, a single molecule event is proven. There exist no instrumental assumptions of our approach on which the experiment itself, the theoretical background, or the conclusion are based. Our results open up a broad field for analytics and diagnostics in solution, especially in immunology.

Unilateral high-altitude pulmonary edema (HAPE): a case report and discussion of pathophysiology.

High-altitude pulmonary edema (HAPE), a potentially life-threatening altitude adaptation disorder, is considered to be caused by an exaggerated increase in pulmonary blood pressure and a non-cardiogenic rise in pulmonary vascular permeability subsequent to alveolar hypoxia. A 40-year-old male mountaineer was affected by an advanced stage of HAPE at high altitude (Monte Rosa plateau, 4000 m). The symptoms abated immediately after the patient descended from the altitude. However, six hours after the symptoms had resolved, radiographic signs of pulmonary edema, confined to the right lung, were seen. This rarely described unilateral radiological pattern of HAPE resolved completely within two days. We suggest that aspiration events of nasal secretion, the right sleeping position at night and an elevated right diaphragm reduced the patient's compensatory hyperventilation capacity of the right lung. The resulting increased alveolar hypoxia in the right lung was responsible for unilateral edema. The pathophysiological mechanism underlying unilateral HAPE is discussed.

Detection and Identification of Single Molecules in Immunology and Immunohematology: Single (solution)-phase Single-molecule FCS.

In the Goodpasture experiment, we determined a probability of 99.1% of identifying one single immune complex. Under these conditions, a single molecule event is proven. There exist no instrumental assumptions of our approach on which the experiments themselves, the theoretical background or the conclusion are based on. Our results open up a broad field for analytics and diagnostics in solution, particularly in immunology and immunohematology.

[Function and repertoire of the immune system in body's defense]. / Funktion und Repertoire des Immunsystems bei körpereigener Abwehr.

This is a minireview on the organisation and activity of the human immune system with special reference to sport and--more precisely--stress by mountaineering. The activation of the immune system under physical exercise is shown and the immune depression after the sport documented. Hence the conclusion of increased susceptibility to diseases in the post activation phase--a sort of depression after alpine sport.

Tryptophan degradation and immune activation in Alzheimer's disease.

Alzheimer's disease (AD) is likely associated with systemic immune activation. During immune response, interferon-gamma stimulates indoleamine 2,3-dioxygenase (IDO) converting tryptophan to N-formylkynurenine followed by kynurenine in an ensuing step. Thus, IDO activity is estimated by the kynurenine per tryptophan quotient (Kyn/Trp). In 21 patients suffering from AD, in 20 controls of similar age, and in 49 blood donors we measured serum tryptophan and kynurenine concentrations by HPLC. Lower tryptophan concentrations were found in elderly control subjects compared to blood donors (62.1 vs. 73.0 microM, p < 0.005). Tryptophan concentrations tended to be still lower in AD patients (54.4 microM, p = 0.07) compared to elderly controls. Enhanced tryptophan degradation in patients was reflected by significantly increased Kyn/Trp (46.1 vs. 34.1 in elderly controls, p < 0.05). Correlations were found in patients between Kyn/Trp and concentrations of soluble immune markers in serum, i.e., neopterin, interleukin-2 receptor and tumor necrosis factor receptor (all p < 0.001). Increased Kyn/Trp was associated with reduced cognitive performance. Tryptophan degradation due to immune activation may exert impact on the pathogenesis of AD.

Time course of plasma soluble intercellular adhesion molecule-1 (sICAM-1) is related to severity of acute pancreatitis.

BACKGROUND/AIMS: In severe acute pancreatitis the release of cytokines indicates a key step from local to systemic inflammation. Increased plasma concentrations of circulating soluble intercellular adhesion molecule-1 (sICAM-1), a marker of leukocyte activation, were detected in necrotizing pancreatitis at the time of diagnosis, however, the exact role of sICAM-1 in the development of complications such as shock or organ dysfunction is unclear. Therefore, we investigated in what manner the time course of plasma sICAM-1 is associated with the development of severe pancreatitis and whether these results are of any predictive value for the further course of the disease. METHODOLOGY: In a medical intensive care unit we studied 29 consecutive patients admitted for acute pancreatitis. Plasma levels of sICAM-1 were measured serially over a period of 6 days and the time courses were assigned either to a group of patients with uncomplicated, mild disease or to patients who developed complications including multiple organ failure. RESULTS: In mild pancreatitis, decreasing and peak sICAM-1 concentrations were found in 88% of the patients with a mean maximal level of 574 +/- 59 ng/ml (SE) (upper limit of normal: 400 ng/ml) on day 1. Partial pancreatic necrosis was present in 24% and no deaths were observed. In severe pancreatitis an increase of sICAM-1 levels or an initial fall followed by an increase (relapsing response) was the predominant pattern (92%). Maximal values of 1453 +/- 136 ng/ml occurred on day 6, significantly different when compared to mild disease (p < 0.0001). Necrotizing pancreatitis was diagnosed in 75% and the mortality rate was 58%. The sensitivity in predicting severe pancreatitis using sICAM-1 plasma levels with an increasing or relapsing pattern was much higher (92%) when compared with serial C-reactive protein measurements (42%). CONCLUSIONS: In acute pancreatitis, increasing or relapsing plasma levels of sICAM-1 over 6 days after admission to hospital are associated with a high rate of pancreatic necrosis and a high mortality. Daily measurements of sICAM-1 would allow early recognition of patients prone to develop complications and follow a severe course.

Relation of serial measurements of plasma-soluble intercellular adhesion molecule-1 to severity of acute pancreatitis.

OBJECTIVE: Acute pancreatitis remains a clinical challenge because it is difficult to predict whether, in a given patient, the disease will be mild or will run a severe course with a possibly fatal outcome. The aim of this study was to investigate whether circulating soluble intercellular adhesion molecule-1 (sICAM-1) as a marker of leukocyte activation is related to the severity of the disease. METHODS: The study included 29 consecutive adults admitted with acute pancreatitis. Plasma sICAM-1 levels were measured serially over a period of 6 days, and values and time courses were correlated with clinical severity. RESULTS: Our patients fell into four groups on the basis of the following measurements: 1) Decreasing sICAM- levels with maximal values of 446 +/- 90 ng/ml (mean +/- SEM) slightly above the upper limit of normal were associated with uncomplicated mild disease in seven patients. 2) In nine patients with sICAM-1 concentrations reaching a peak of 743 +/- 121 ng/ml after 3 days, severe pancreatitis was present in 11% and pancreatic necrosis occurred in 33%. 3) A second increase of sICAM-1 (maximal level: 993 +/- 169 ng/ml) after an initial decrease (relapsing pattern, 7 patients) was associated with a severe course of disease in 71% including pancreatic necrosis in 43% and nosocomial pneumonia in 42%. 4) A rapid increase of sICAM-1 reaching highest maximal values of 1738 +/- 104 ng/ml (p < 0.0001) indicated fulminant pancreatic necrosis and a fatal outcome in six patients. CONCLUSIONS: Serial plasma sICAM-1 levels in patients with acute pancreatitis within the first 6 days after admission fall into four different groups of severity according to the shape of the curves. This suggests that the time course of elevated plasma sICAM-1 concentrations reflects the risk of developing necrosis and clinical complications in acute pancreatitis.

[Soluble adhesion molecules (P-selectin, ICAM-1 and ICAM-3) in rheumatoid arthritis]. / Rastvorimye molekuly adgezii (P-selektin, ICAM-1 i ICAM-3) pri revmatoidnom artrite.

AIM: Investigation of serum levels and clinical role of soluble intercellular molecules of adhesion (pICAM-1, PICAM-3 and pP-selectin) in rheumatoid arthritis (RA). MATERIALS AND METHODS: Enzyme immunoassay with Bender MedSystem kits (Austria) was employed to measure serum concentration of soluble intercellular molecules of adhesion in 36 RA patients. RESULTS: Elevated levels of serum pICAM-1, pICAM-3 and pP-selectin were registered in 74.2, 28.6 and 25.7% of RA patients, respectively. Content of pP-selectin more strongly correlated with activity and severity indices than that of pICAM-3 (p < 0.001). Content of pICAM-1 and clinical picture of RA were unrelated. CONCLUSION: Levels of pP-selectine can characterize RA activity.

[Frequency of immunologic disorders in acute schizophrenia]. / Zur Häufigkeit immunologischer Auffälligkeiten bei akuten Schizophrenien.

Immunological examinations in schizophrenic patients have shown that there are many alterations in both arms of the immune system, i.e. cellular and humoral activities. The results are quite heterogeneous, as not even all schizophrenics show these pathological changes. Immunological findings are assumed to be etiopathogenetically related to the disease process or to be an epiphenomenon. The present study supposes that immunological alterations as they can be found during the course of schizophrenia may be an indicator for somatic vulnerability or an epiphenomenon. 60 male inpatients, fulfilling DSM-IV criteria of schizophrenia where examined during their acute phases of psychosis and during their phases of clinical improvement, by means of a serological profile including cellular and humoral parameters. The control group consisted of 42 healthy male volunteers. It was the aim of this study to find out if there were (a) overall differences in the immune profiles between patients and control group and (b) differences between different categories of schizophrenic disorder. During the acute phase nearly half of the schizophrenic patients showed pathologic immunological parameters, whereas none of the controls did. During the phase of clinical improvement the number of patients with normal immunological findings predominated. Furthermore there was a difference between the Paranoid and the Disorganized Subtype, the latter showing more immunological abnormalities. The results of this study give further support to the hypothesis that immunological aberrations should not be seen as closely etiopathogenetically related to schizophrenic disorders, but rather as an epiphenomenon (e.g. as a stress marker) and/or as indicators for somatic vulnerability.

Increased serum neopterin concentrations in patients with Alzheimer's disease.

We measured serum neopterin concentrations in 24 patients with Alzheimer's disease (8 males, 16 females; age: 73.1+/-6.2 years; free of any infectious process) and fourteen controls of similar age (4 males, 10 females; age: 69.7+/-8.8 years). Compared to controls, significantly higher concentrations of neopterin (p< 0.01) were found in patients with Alzheimer's disease. Among patients, concentrations of neopterin were higher in those with lower mini-mental-state (p < 0.05), and an inverse correlation existed between mini-mental-state and neopterin concentrations. No such association existed with the duration of the disease. There were also significant correlations between neopterin and serum concentrations of immune activation markers such as soluble tumor necrosis factor (TNF) receptor and soluble interleukin-2 receptor (all p<0.01). Thus, increased concentrations of neopterin in serum of patients with Alzheimer's disease correlate with the severity of dementia. The data imply a chronic state of peripheral immune activation in Alzheimer's disease.