Adenovirus type 12 tumor antigen. II. Immunoprecipitation of protein kinase from infected and transformed cells by antisera to T antigen and some normal rat sera.

Protein kinase was found to be precipitated from adenovirus type 12 (Ad12)-infected KB cells and Ad12-transformed hamster cells by sera of tumor-bearing hamsters and rats: Immunoprecipitates obtained with T antigen reactive sera catalyzed transfer of ³²P from [γ-³²P]ATP to the γ-chain of IgG. Analogous products of control cells were without significant activity. Control hamster sera precipitated no protein kinase from infected and transformed cells. Some control rat sera (syngeneic with immune sera), however, were found to precipitate protein kinase from infected and transformed cells; particularly active in this respect were sera of female breeder rats. When partially purified, highly immunoreactive T antigen preparations from transformed cells were used as a source of enzymatic activity, protein kinase was detected only in precipitates obtained with immune sera.     

VEGF (vascular endothelial growth factor) concentration in serum and pleural fluid of patients with pleural malignancy and pleural tuberculosis

The purpose of this study was comparison of VEGF (vascular endothelial growth factor) levels in serum and pleural fluid and estimation of this test usefulness in diagnosis of pleural effusions. VEGF levels were measured by ELISA method in 68 patients (45 males and 23 females) aged 19-81 years. By Light's criteria in 16 cases transudate and in 52 cases exudate was recognized. By means of fluid cytology, pleural biopsy, microbiology or thoracoscopy in 10 cases pleural metastases from distant organs, in 15 cases coexisting pulmonary neoplasm, in 11 cases mesothelioma and in 16 cases tuberculosis were determined as a cause of fluid accumulation in pleural space. The mean VEGF levels were significantly higher (p < 0.001) in patients with exudates than in patients with transudates (3833 pg/ml and 325 pg/ml respectively). Based on likelihood ratios analysis, as a cut off value in differentiation of exudates and transudates a value 700 pg/ml was accepted. The sensitivity of this test was 75% and the specificity 93% and likelihood ratio (LR) 12.5. The mean VEGF level in exudates was seven times higher than mean VEGF level in serum (3833 pg/ml and 573 pg/ml respectively). Mean VEGF levels in malignant exudates (4615 pg/ml) were significantly higher than in tuberculous exudates (2073 pg/ml). As a cut off value in differentiation between malignant and tuberculous exudates a value of 4500 pg/ml was accepted. We conclude that our results suggests the local VEGF production in pleural cavity and the significant role of this cytokine in pleural exudates accumulations and also suggests the usefulness of VEGF estimation in pleural fluid in differentiation transudates from exudates and malignant from tuberculous pleural fluids.     

Seven novel and four recurrent point mutations in the factor VIII (F8C) gene

Haemophilia A is a X‐linked bleeding disorder, caused by deficiency in the activity of coagulation factor VIII due to mutations in the corresponding gene. The most common defect in patients is an inversion of the factor VIII gene that accounts for nearly 45% of individuals with severe hemophilia A. Point mutations and small deletions/insertions are responsible for the majority of cases with moderate to mild clinical course and for half of the severe hemophilia A occurrences. The majority of these mutations are “private”, because of the high mutation rate for this particular gene. We report on eleven pathological changes in the factor VIII sequence detected in male patients with haemophilia A or in female obligate carriers. Seven of these mutations are novel [E204N, E265X, M320T, F436C, S535C, N2129M and R2307P] and four have been previously identified [V162M, R527W, R1966X, and R2159C]. Genotype‐phenotype correlations and computer prediction analysis on the effect of missense mutations on the secondary structure of the factor VIII protein are performed and the relationships evaluated. © 2001 Wiley‐Liss, Inc.     

Changed visuomotor transformations during and after prolonged microgravity

A series of step-tracking experiments was conducted before, during, and after a 3-week space mission to assess the effects of prolonged microgravity on a non-postural motor-control task. In- and post-flight accuracy was affected only marginally. However, kinematic analyses revealed a considerable change in the underlying movement dynamics: too-small force and, thus, too-low velocity in the first part of the movements was mainly compensated by lengthening the deceleration phase of the primary movement, so that accuracy was regained at its end. The observed in-flight decrements in peak velocity and peak acceleration point to an underestimation of mass, in agreement with the re-interpretation hypothesis of Bock et. al. Post-flight no reversals of the in-flight changes (negative aftereffects) were found. Instead, there was a general slowing down, which could be due to post-flight physical exhaustion.     

A low-voltage activated,transient calcium current is responsible for the time-dependent depolarizing inward rectification of rat neocortical neurons in vitro

Intracellular recordings were obtained from rat neocortical neurons in vitro. The current-voltage-relationship of the neuronal membrane was investigated using current- and single-electrode-voltage-clamp techniques. Within the potential range up to 25 mV positive to the resting membrane potential (RMP: –75 to –80 mV) the steady state slope resistance increased with depolarization (i.e. steady state inward rectification in depolarizing direction). Replacement of extracellular NaCl with an equimolar amount of choline chloride resulted in the conversion of the steady state inward rectification to an outward rectification, suggesting the presence of a voltage-dependent, persistent sodium current which generated the steady state inward rectification of these neurons. Intracellularly injected outward current pulses with just subthreshold intensities elicited a transient depolarizing potential which invariably triggered the first action potential upon an increase in current strength. Single-electrode-voltage-clamp measurements reveled that this depolarizing potential was produced by a transient calcium current activated at membrane potentials 15–20 mV positive to the RMP and that this current was responsible for the time-dependent increase in the magnitude of the inward rectification in depolarizing direction in rat neocortical neurons. It may be that, together with the persistent sodium current, this calcium current regulates the excitability of these neurons via the adjustment of the action potential threshold.     

Shear stress induced membrane currents and calcium transients in human vascular endothelial cells

We have measured membrane currents induced by shear stress together with intracellular calcium signals in endothelial cells from human umbilical cord veins. In the presence of extracellular calcium (Ca²⁺]_o), shear stress induced an inward current at a holding potential of 0 mV which is accompanied by a rise in intracellular Ca²⁺ ([Ca²⁺]_i). In the absence of extracellular calcium shear stress was unable to evoke a calcium signal but still induced a membrane current. The voltage dependence of the shear stress induced current was obtained from difference currents evoked by linear voltage ramps before and during application of shear stress. Its reversal potential E_rev shifted from –2.3±0.8 mV (n=4) in a nominally Ca²⁺ free solution to +1.5±1.6 mV at 1.5 mM [Ca²⁺]_o (n=4) and to +21.9±4.4 mV (n=7) at 10 mM [Ca²⁺]_o. From our data we conclude that shear stress opens an ion channel that is 12.5±2.9 (n=7) times more permeable for calcium than for sodium or cesium.     

Role of H+ ions in volume and voltage of epithelial cell nuclei

Condensation of chromatin depends upon the ion composition in the cell nucleus. We tested in isolated nuclei of Madin-Darby canine kidney cells the influence of various ions on nuclear volume (i. e. DNA packing) and intranuclear voltage. After isolation, nuclei were superfused with cytosolic solutions in which Na⁺, K⁺, Ca²⁺ and H⁺ ions were varied. With video-imaging and microelectrode techniques nuclear volume and intranuclear potential were measured in response to the various ions. In control cytosolic solution, isolated nuclei exhibited an intranuclear electrical potential of –6.5±0.5 mV (relative to a reference electrode in the cytosolic solution) corresponding to a nuclear volume of 250±10 fl (n=104). Changing the Na⁺, K⁺ or free Ca²⁺ concentration in the superfusate in the physiological range resulted in minor changes of volume and intranuclear potential whereas pH altered both parameters dramatically. Nuclear swelling and intranuclear negative voltage increased with alkalinization and decreased when pH was reduced. An intact nuclear envelope was found to be no prerequisite for maintaining intranuclear negativity, indicating that the composition and functional state of nuclear chromatin rather than specific ion permeabilities of the nuclear envelope determine nuclear electrical potential. We present a model that explains nuclear volume and voltage on the basis of interaction between negatively charged DNA and positively charged histones of the nuclear chromatin.     

Basics of the virology of HIV-1 and its replication.

Human immunodeficiency virus is undoubtedly the causative agent of AIDS. The understanding of HIV-1 pathogenesis is essential to develop and maintain antiretroviral treatment and vaccination. Since the first isolation of HIV-1 in cell culture, thousands of publications dealing with HIV and/or AIDS per year were released. In this review we give a basic overview of the virology of HIV-1 including the functions of the different HIV-1 proteins required for effective viral replication. Moreover, we summarize the interactive processes between HIV-1 and its target cells. Finally, the HIV-1 specific immune response and the current status of antiretroviral therapy are briefly described in this review.     

Cushing's Syndrome Secondary to Olfactory Neuroblastoma

A case of olfactory neuroblastoma in a 36-year-old woman who presented with florid Cushing's syndrome is reported. A nasal polyp, which proved to be an olfactory neuroblastoma, was resected. The procedure was followed by complete remission from the endocrinologic abnormalities. Postoperatively, the patient was well for 5 years until recurrence of both Cushing's syndrome and the nasal polyp was noted. Following combined transnasal-transcranial resection of the tumor, which extended into the anterior cranial fossa, the patient again experienced complete remission of Cushing's syndrome. Immunohistochemistry showed the tumor to be positive for neuron-specific enolase, synaptophysin, chromogran-in, adrenocorticotropic hormone, (J-endorphin, and S-100 protein. Electron microscopy revealed neuritic processes containing microtubules and neurosecretory granules. This is the first reported case of Cushing's syndrome secondary to olfactory neuroblastoma.