Synthesis, pharmacological and biophysical characterization, and membrane-interaction QSAR analysis of cationic amphiphilic model compounds.

Cationic amphiphilic drugs have a propensity to interact with biological interphases. This study was designed to gain more insight into the molecular properties of catamphiphilic drugs which govern this type of interaction. A series of phenylpropylamine model compounds were synthesized in which modifications were incorporated at the aromatic part of the molecule. The replacement of (45)Ca(2+) from phosphatidylserine monolayers served to monitor drug binding to the phospholipid. The influence on the phase-transition temperature of liposomes of dipalmitoylphosphatidic acid was measured to assess the perturbing action of the drugs on the structural organization of phospholipid assemblies. The antiarrhythmic activity of the compounds was determined in Langendorff preparations of guinea pig hearts to assess the membrane-stabilizing action. Quantitative structure-activity relationship (QSAR) models for these endpoints were developed using both intra- and intermolecular QSAR descriptors. Intermolecular membrane-interaction descriptors were derived from molecular dynamics simulations of the compounds in a model phospholipid monolayer. QSAR models were derived for all endpoints using partial least-squares regression (PLS) and a genetic algorithm tool, the genetic function approximation (GFA). Membrane-interaction descriptors appear to be of a particular importance in explaining the influence of the compounds on the phase-transition temperature of DPPA liposomes, while the other endpoints can be adequately modeled by intramolecular descriptors. The calcium-displacing activity at phosphatidylserine monolayers is governed by the electrostatic properties of the compounds. Measures of lipophilicity and molecular size are of particular importance for antiarrhythmic activity. Possible improvements to both the molecular modeling and the applied computational protocol of membrane-solute systems are identified and discussed.

Seroprevalence and disease association of antineutrophil cytoplasmic autoantibodies and antigens in HIV infection.

This prospective study was designed to determine the role of antineutrophil cytoplasmic autoantibodies (ANCA) in HIV-infected patients. Immunofluorescence tests (IFT) and enzyme-linked immunosorbent assays (ELISA) were applied to sera of 199 consecutive outpatients. In the IFT 20% were positive. An atypical ANCA pattern was demonstrated in 67% of these, 33% revealed a perinuclear staining (pANCA). Specific ELISA revealed proteinase 3 (n = 2), myeloperoxidase (n = 1), lysozyme (n = 2), lactoferrin (n = 1), cathepsin G (n = 1), and human leukocyte elastase (HLE, n = 6). The target antigen remained unidentified in 26 patients. Perinuclear ANCA-positive patients showed atypical antigens in eight of 13 cases; all six patients with anti-HLE revealed a pANCA pattern. The antigens of atypical ANCA-positive patients remained unidentified in 21 of 26 (81%) cases. No signs of vasculitis were present in the ANCA-positive patients. ANCA are frequently found in the sera of HIV-positive patients. They bind to a variety of antigens. No correlation was found between ANCA positivity and autoimmune or opportunistic diseases.

[Augmentation of the neuromuscular blocking effect of mivacurium during inhalation anesthesia with desflurane, sevoflorane and isoflurane in comparison with total intravenous anesthesia]. / Verstärkung der neuromuskulär-blockierenden Wirkung von Mivacurium während Inhalations-anästhesien mit Desfluran, Sevofluran und Isofluran im Vergleich zu totaler intravenöser Anästhesie.

To evaluate the enhancement of mivacurium-induced neuromuscular block by potent inhalational anaesthetic agents, dose-effect curves for mivacurium were determined in 84 patients scheduled for minor elective surgery during anaesthesia with 1.5 MAC (70% N2O) desflurane, sevoflurane and isoflurane and compared with those under total intravenous anaesthesia (TIVA). Acceleromyography (TOF-Guard) and train of four (TOF) stimulation of the ulnar nerve were used (2 Hz every 12 s). Mivacurium was administered in increments of 25 micrograms kg-1 until a depression of T1 > 95% was reached. ANOVA was used for statistical analysis (alpha = 0.05, beta = 0.2). The depression of T1 during potent inhalational anaesthesia was enhanced compared with TIVA. The ED50 and ED95 of mivacurium were 27 +/- 11 (SD) and 58 +/- 26 micrograms kg-1 for desflurane; 28 +/- 10 and 64 +/- 23 micrograms kg-1 for sevoflurane; and 27 +/- 13 and 55 +/- 27 micrograms kg-1 for isoflurane and were significantly lower than the 35 +/- 7 and 71 +/- 20 micrograms kg-1 for TIVA. The duration 25% revealed a significant difference between the volatile anaesthetic groups (10 +/- 2, 11 +/- 3, 11 +/- 3 min respectively) and the TIVA control group (8 +/- 3 min). The recovery index 25/75 and TOFO.80 were significantly prolonged by desflurane, sevoflurane and isoflurane compared with TIVA (RI25/75 9 +/- 4, 9 +/- 4, 10 +/- 5 respectively vs. 5 +/- 2 min; TOFO.80 27 +/- 10, 28 +/- 9, 29 +/- 9 respectively vs. 18 +/- 4 min). We conclude that the neuromuscular blocking effect of mivacurium is enhanced during anaesthesia with desflurane, isoflurane and sevoflurane compared with TIVA. The duration of action and the recovery time are prolonged. The dose of mivacurium used should be reduced if anaesthesia is maintained with volatile anaesthetics.

[The subspecies specificity of Babesia canis]. / Zur Artspezifität von Babesia canis.

The large Babesia species of dogs, Babesia canis, is transmitted by different ticks. Dermacentor reticulatus, Rhipicephalus sanguineus and haemaphysalis leachi are the known main vectors. Four B. canis isolates of different geographic origin were investigated for their transmission specificity and pathogenicity in infection trials. R. sanguineus ticks exclusively transmitted the Babesia isolate from Egypt. D. reticulatus was the vector for isolates from Hungary and France. Transmission of an South-African isolate was only possible by H. laechi. The B. canis isolates differed markedly in their pathogenicity. The South-African isolate was highly pathogenic and resulted in nearly complete mortality. Infections with the isolate from Hungary transmitted by D. reticulatus, also resulted in severe clinical disease which often ended fatal without treatment with an anti-Babesia drug. Clinical disease also resulted from infections with the French isolate while the isolate from Egypt was largely non-pathogenic. Challenge trials for investigations of cross-immunity demonstrated immunogenic differences between the individual isolates. The degree of immunogenicity appeared to be related to the pathogenicity of the Babesia isolates. Serological investigations revealed that antigen of the isolate from Hungary in ELISA reacted markedly higher than the other three antigens. Due to their vector specificity B. canis classification into three groups is possible: B. canis canis transmitted by D. reticulatus, B. canis vogeli transmitted by R. sanguineus and B. canis rossi transmitted by H. laechi.

Anti-neutrophil cytoplasmic antibodies in generalized autoimmune diseases.

Anti-neutrophil cytoplasmic antibodies (ANCA) are a system of autoantibodies which are strongly associated with the primary systemic vasculitides (PSV). cANCA, as detected by indirect immunofluorescence, are mostly reactive to proteinase 3 (PR3) and bear high sensitivity and specificity for Wegener's granulomatosis. pANCA have varied subspecificities and clinical associations. The most important subspecificity of pANCA is anti-myeloperoxidase (MPO), which is strongly associated with microscopic polyaniitis and pauci-immune crescentic glomerulonephritis. pANCA also occur at low to moderate frequency in other PSV, collagen vascular disease, inflammatory bowel disease and autoimmune liver disease. In systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA), pANCA of varied subspecificity have been found, including anti-MPO at low rate and low titer, while cANCA and anti-PR3 were generally absent. Consequently, anti-PR3 (PR3-ANCA) and anti-MPO (MPO-ANCA) at moderate and high titer are distinguishing features of the PSV and, in an appropriate clinical setting, argue strongly against the presence of SLE or RA. Since no significant clinical association has been found for other pANCA subspecificities, cANCA, PR3-ANCA and MPO-ANCA remain the critical elements of ANCA testing in the clinical diagnosis of generalized autoimmune diseases.

[The von Hippel-Lindau syndrome. Its differential diagnosis from cystic kidneys in adulthood]. / Das v. Hippel-Lindau-Syndrom. Differentialdiagnose von Zystennieren im Erwachsenenalter.

A 51-year-old patient with severe back pain had undergone resection of a benign cerebellar tumour when aged 15 years. In addition, polycystic kidney disease was diagnosed 24 years ago, bilateral phaeochromocytoma 2 years ago, and for 4 months before the present admission he had been on haemodialysis. The family history indicated autosomal dominant inheritance of the polycystic renal disease. His general condition was found to have deteriorated, he had pain on pressure over the upper thoracic and lower lumbar vertebrae, and the kidneys were enlarged on palpation. There were increased concentrations of calcium (3.01 mmol/l), parathormone (2.0 ng/l), carcinoembryonic antigen (13.5 micrograms/l) and TPA (69 U/l). Computed tomography demonstrated cystic and solid parts of much enlarged kidneys. Biopsy revealed a poorly differentiated clear-cell renal carcinoma. Further information concerning the previously removed brain tumour showed this to have been an haemangioblastoma of the cerebellar tonsils indicating the diagnosis of v. Hippel-Lindau disease. Nine other family members had been affected, but none had the full-blown picture of the disease. The patient died 3 weeks later from the rapidly advancing tumour. Autopsy showed the bilateral renal carcinoma, bilateral phaeochromocytoma and metastases to the sternum, femurs, vertebrae and liver.

Characterization and comparison of merozoite antigens of different Babesia canis isolates by serological and immunological investigations.

Merozoites of four Babesia canis isolates from Hungary, France, Africa, and Egypt were purified. Antigens were compared in an enzyme-linked immunosorbent assay (ELISA) and by immunoblotting. In the ELISA, antigen from the highly pathogenic isolate from Hungary showed the highest sensitivity for homologous and heterologous immune sera. This was confirmed by immunoblotting. Protein bands of the Hungarian isolate were strongly recognized by all B. canis immune sera, whereas the antigens from the other isolates showed only weak reactions with homologous and heterologous immune sera. Significant was a protein band of about 12 kDa appearing in all pathogenic isolates from Hungary, France, and South Africa but not in the apathogenic Egyptian isolate. This protein band may determine the virulence. For serological tests, the B. canis isolate from Hungary seems to be the one most suitable for detection of even mild infections.

[Polycythemia in the late metastasis of an erythropoietin-producing renal-cell carcinoma]. / Polyglobulie bei Spätmetastasierung eines Erythropoietin-produzierenden Nierenzellkarzinoms.

An 85-year-old woman had a right-sided renal cell carcinoma removed 20 years ago. At haemoglobin concentration. Two years ago she had a syncope, at which time the haemoglobin concentration was 16.9 g/dl. Ultrasound and computed tomography (CT) revealed an extensive retroperitoneal space-occupying lesion, which however was not investigated further, and no therapeutic consequences were drawn. An erythrocytosis (7.5 x 10(6)/microliters) and elevated haemoglobin concentration (> 20 g/dl) were found when she was examined after a fall in which she had sustained only minimal injury. The retroperitoneal mass had slightly increased in size. Histological examination of a CT-guided fine-needle biopsy revealed metastases of the hypernephroid carcinoma. The serum erythropoietin concentration was increased (42.4 U/l) and failed to increase even after repeated venesections, indicating erythropoietin production by the late metastases of the renal cell carcinoma. There was no evidence for any systemic haematological disease. Six months after the diagnosis of metastases the patient died at home, presumably of a cerebrovascular accident.

ANCA in systemic vasculitides, collagen vascular diseases, rheumatic disorders and inflammatory bowel diseases.

It was the aim of this study to reevaluate the diagnostic significance and clinical implication of ANCA after testing sera from 13,606 patients for the presence of ANCA. Our data confirm the high specificity (97%) and sensitivity (80%) of cANCA for Wegener's granulomatosis. pANCA were found in renal vasculitides (60%), collagen vascular diseases (SLE 20%, Sjögren's syndrome 26%, polymyositis 16%) and rheumatic disorders (Felty's syndrome 50%, rheumatoid arthritis 20%). A third fluorescence pattern in sera of patients with inflammatory bowel disease (ulcerative colitis 28/72, Crohn's disease 6/84), here called xANCA, was seen. Target antigens of granule proteins from PMN and monocytes (proteinase 3, myeloperoxidase, elastase, cathepsin G, lactoferrin, lysozyme) were identified.

Autoantibodies directed against lysozyme: a new target antigen for anti-neutrophil cytoplasmic antibodies (ANCA).

ANCA-positive sera from 1138 patients and ANCA-negative sera from 90 patients were screened for autoantibodies directed against lysozyme (LZ) by ELISA. Sera from 120 patients did react with LZ. 99 sera bound to LZ only, whereas 56 sera bound to further granule proteins, especially cathepsin G and lactoferrin. In the routine ANCA screening, most of the anti-LZ-positive sera showed a pANCA fluorescence. In total, 8% of 674 pANCA-positive sera did react with LZ. Clinically, anti-LZ antibodies were associated inflammatory rheumatologic, -renal and -bowel diseases.

[Elevated serum levels of ICAM-1 in Wegener's granulomatosis]. / Hohe Serumspiegel von ICAM-1 bei der aktiven generalisierten Wegenerschen Granulomatose.

Circulating soluble ICAM-1 (sICAM-1) isoforms are known to be elevated in inflammatory and autoimmune disorders (SLE, RA). A correlation between levels of circulating sICAM-1 and disease activity was described (5). In sera from 31 patients suffering from histologically confirmed Wegener's granulomatosis (WG) elevated levels of ICAM-1 were now detected in this primary systemic vasculitis. In generalized active disease circulating ICAM-1 levels were significantly higher when compared to remission and healthy control individuals. Sera from WG patients collected during intercurrent infections while on immunosuppressive drugs showed no elevation of circulating ICAM-1.

[Atropine versus glycopyrrolate in eye surgery. A comparison of rhythm disorders and heart frequency using Holter-ECG]. / Atropin versus Glycopyrrolat bei ophthalmochirurgischen Eingriffen. Vergleich der Rhythmusstörungen und der Herzfrequenz mit Holter-EKG.

Anticholinergics are indicated in ophthalmic surgery to inhibit oculocardiac reflexes (OCR). Although all anticholinergics exert an arrhythmic effect, comparative data on occurrence and severity of arrhythmias are not available. The aim of this investigation was to compare atropine (A) and glycopyrrolate (G) in adults undergoing ophthalmic surgery. One hundred patients (ASA class I and II, age range 20-60 yrs.) were given equipotent doses of either A (10 micrograms.kg-1) or G (5 micrograms.kg-1) in a randomized, double-blind fashion, before induction of anaesthesia. Continuous Holter monitoring was performed. The procedure was divided into the following phases: O (5 min before drug), I (5 min after drug), II (up to induction), III (induction until intubation), IV (intubation until operation), V (operation). Supraventricular, junctional, and ventricular arrhythmias were analyzed. Severe arrhythmia was judged to be greater than 5 events per min, or ventricular beats Lown classes III-V. The beat to beat analog signal was digitalized. The heart rate (HR) and the occurrence rate of OCR (greater than 20% decrease in heart rate, arrhythmias) was calculated by a computerized program. Analysed were: 1. the frequency of OCR, 2. the mean HR for each phase, 3. the frequency of all the various arrhythmias during the whole period, 4. as well as for each phase. The number of patients with severe arrhythmias 5. either for all the time or 6. for the individual phase were registered separately. There were no differences 1. in the frequency of OCR and 2. in mean HR during all phases between A and G.(ABSTRACT TRUNCATED AT 250 WORDS)

Sexual cycle of Babesia divergens confirmed by DNA measurements.

The DNA content of the developmental stages within the life cycle of Babesia divergens was measured by means of fluorescence microscopy using the DNA-specific bisbenzimide Hoechst 33258. By comparing the mean relative fluorescence intensities, the assumption of sexual reproduction in the gut of the tick vector (Ixodes ricinus) was confirmed. However, no proof of a pre- or postzygotic meiosis was found.

B cell activation by synthetic lipopeptide analogues of bacterial lipoprotein bypassing phosphatidylinositol metabolism and proteinkinase C translocation.

Synthetic analogues of bacterial lipoprotein induce proliferation of murine small resting B lymphocytes. We investigated the role of proteinkinase C (PKC) activation in lipopeptide-induced B cell stimulation. Using a standardized extraction procedure, there was no change in membrane bound and soluble PKC activity upon stimulation with lipopeptide. However, omitting Ca2+ chelators from the standard extraction medium resulted in a decrease of membrane bound PKC activity after stimulation. Lipopeptide failed to induce phosphoinositide degradation and the generation of the two second messengers cAMP and cGMP. To test whether guanosinetriphosphate-binding proteins are involved in lipopeptide-induced signal transfer we investigated the effect of LiCl, choleratoxin and pertussistoxin on B lymphocyte proliferation. LiCl and pertussistoxin did not inhibit cell activation, whereas choleratoxin reduced the proliferation rate at concentrations higher than 0.5 micrograms/ml. Similar results were observed when LPS was used as mitogen, whereas the anti-immunoglobulin-induced B cell activation was inhibited by all three compounds. Our results show, that B cell activation by bacterial lipopeptides bypasses phosphatidylinositol metabolism and PKC translocation.

The role of interleukin 2 (IL 2) in the proliferation of Theileria annulata-infected bovine lymphocytes.

The role of interleukin 2 (IL 2) in the proliferation of Theileria annulata-infected lymphoblastoid cells was studied. It was found that these cells neither require nor produce IL 2. This was based on the following findings: (a) The growth of T. annulata-infected cells was not enhanced by addition of IL 2 to the culture medium. (b) IL 2-dependent Con A-stimulated bovine peripheral blood lymphocytes (Con A-blasts) no longer required IL 2 after being infected with sporozoites of T. annulata. (c) The supernatants of T. annulata-infected cells did not support growth of IL 2-dependent Con A-blasts.