Bortezomib inhibits human osteoclastogenesis.

In multiple myeloma, the overexpression of receptor activator of nuclear factor kappa B (NF-kappaB) ligand (RANKL) leads to the induction of NF-kappaB and activator protein-1 (AP-1)-related osteoclast activation and enhanced bone resorption. The purpose of this study was to examine the molecular and functional effects of proteasome inhibition in RANKL-induced osteoclastogenesis. Furthermore, we aimed to compare the outcome of proteasome versus selective NF-kappaB inhibition using bortezomib (PS-341) and I-kappaB kinase inhibitor PS-1145. Primary human osteoclasts were derived from CD14+ precursors in presence of RANKL and macrophage colony-stimulating factor (M-CSF). Both bortezomib and PS-1145 inhibited osteoclast differentiation in a dose- and time-dependent manner and furthermore, the bone resorption activity of osteoclasts. The mechanisms of action involved in early osteoclast differentiation were found to be related to the inhibition of p38 mitogen-activated protein kinase pathways, whereas the later phase of differentiation and activation occurred due to inhibition of p38, AP-1 and NF-kappaB activation. The AP-1 blockade contributed to significant reduction of osteoclastic vascular endothelial growth factor production. In conclusion, our data demonstrate that proteasomal inhibition should be considered as a novel therapeutic option of cancer-induced lytic bone disease.

Immunological memory stabilizing autoreactivity.

The etiopathologies of autoimmune diseases are complex. A broad variety of cell types and gene products are involved. However, clinical and experimental evidence suggests that the importance of an individual factor changes during the course of the disease. Factors and cell types that induce acute autoreactivity and initiate an autoimmune disease could be distinct from those that drive a chronic course of that disease. Autoreactive immunological memory, in particular B cell and plasma cell memory, contributes to chronicity through several mechanisms. Formation of autoreactive memory B cells leads to an increase in the numbers of autoreactive cells. In comparison to naive B cells, these memory B cells show a decreased threshold for activation. Additionally, a fraction of memory B cells express the chemokine receptor CXCR3, which supports their accumulation within chronically inflamed tissues. This may allow their escape from mechanisms for induction of peripheral tolerance. Within the inflamed tissue, inflammatory cytokines and autoantigens provide activation signals that promote plasma cell differentiation and survival. The autoantibodies produced locally by these plasma cells contribute to the severity of inflammation. Together, an autoreactive loop of autoantibody-induced inflammation is formed. Another integral part of immunological memory are long-lived plasma cells. These cells provide persistent humoral antibody memory. Though not all autoantibodies are produced by long-lived plasma cells, these cells have a special impact on immune pathology. Long-lived plasma cells are relatively resistant to existing therapies of immunosuppression and continuously secrete antibodies, without need for restimulation. Long-lived plasma cells provide titers of autoantibodies even during clinically quiescent phases and after immunosuppression. These persisting autoantibody titers, though often low and not causing acute clinical symptoms, are likely to maintain a low level of chronic inflammation and progressive tissue destruction, which reduces the threshold for another break of immunological tolerance.

Analysis and sorting of T cells according to cytokine expression.

Functional distinct populations of T helper cells can be defined according to the expression of cytokines. A remarkable diversity of cytokine expression has been demonstrated in single T cells even from clonal populations and up to now no stable surface markers have been described which on the single-cell level directly correlate with the secretion of a certain cytokine. Since cytokines are the major parameters of T cell effector function we have developed strategies which now allow to separate cells according to the specific cytokines they secrete. The "affinity matrix technology"--secreted molecules are relocated to the cell surface by an artificially created antibody matrix--allows to isolate cells according to a distinct secreted product. In addition to this universal, but laborious technology, we could demonstrate by high-sensitivity immunofluorescence that IFN-gamma and IL-10 but not IL-2 and IL-4 are specifically expressed in low copy number on the surface of cells secreting these cytokines. Surface IFN-gamma and IL-10 are the first unambiguous surface markers for pro-inflammatory IFN-gamma-secreting Th 0/1 cells and IL-10-producing anti-inflammatory Th2/3 cells. We have used purified cytokine-secreting T cells to study in individual T cells the sequential production of IL-2, IFN-gamma, and IL-10, the stability of IFN-gamma expression and the selective homing of IFN-gamma-producing cells into inflamed tissues.

Tumour oxygenation under normobaric and hyperbaric conditions.

Tumour oxygenation during exposure to normobaric and hyperbaric oxygen was assessed by means of a cryophotometric micromethod and a specially constructed pressure chamber. The tumours investigated were DS-carcinosarcomas implanted subcutaneously into the thigh of Sprague-Dawley rats. The animals were anaesthetised and put on a heating pad. Mean arterial blood pressure was monitored throughout the entire time of pressurisation. Cryobiopsies were removed from the tumours after exposure to O2 for 30 min at 1 bar (atmospheric pressure), 2, 3, or 4 bar. Oxyhaemoglobin saturation (HbO2) values of single red blood cells in tumour microvessels were determined photometrically in the frozen tissue samples as a quantitative measure for tumour oxygenation. Frequency distribution curves of HbO2 showed that there was a marked improvement of the O2 supply to tumour tissue in O2 atmospheres at 1 bar compared with exposure to air at 1 bar. Pressurisation in O2 atmospheres up to 2 and 3 bar did not substantially alter the distribution curve of the HbO2 values in comparison with the data sampled at 1 bar O2 exposure, yet led to a significant drop in the occurrence of HbO2 values below 5% saturation. Thus, pressurisation particularly raises those tissue O2 partial pressures (pO2) that are in a range where the radiosensitivity is critically influenced by the pO2. Pressurisation up to 4 bar caused a shift of the HbO2 distribution curve to values significantly higher than those from tumours exposed to O2 at 1 bar, with no values below 35% saturation. Using these data and a previously published model for computing tissue pO2 values it can be shown that radiobiological hypoxia is unlikely to occur in DS-carcinosarcomas exposed to an O2 atmosphere at 4 bar under the conditions chosen. Hyperbaric oxygenation is recommended as an efficient adjuvant of X irradiation, particularly in superficial tumours.

Impact of localized microwave hyperthermia on the oxygenation status of malignant tumors.

Upon heating at 40 degrees C for 30 minutes, the oxygenation of the tumor tissue significantly improved as compared with control conditions at 35 degrees C. In contradistinction to this, the tumor oxygenation significantly decreased directly after 43 degrees C- hyperthermia. A further temperature rise to 45 degrees C caused the oxygenation to drastically drop due to an almost complete cessation of nutritive blood flow. The changes in tumor oxygenation during hyperthermia seem to be predominatly mediated through changes in tumor blood flow, which showed the same directional changes.

Relationship between size and oxygenation status of malignant tumors.

Analyzing the oxygenation status of tumors by means of the oxyhemoglobin saturation of single red blood cells within microvessels, there is clear evidence that tissue hypoxia and anoxia are inherent features during advanced stages of malignant growth. This is a consequence of distinct deteriorations of the convective and diffusive O2 transport with increasing tumor size leading to an O2 depletion in peripheral tissue layers around nutritive blood vessels. These alterations are combined with inhomogeneities of the oxygen distribution, both spatial and temporal. With enlarging tumor mass, the heterogeneities of the O2 supply distinctly intensify. Although in vitro experiments utilizing multicell spheroids failed to show very low oxygen tensions in necrotic regions, it is still proposed that chronic oxygen depletion is a paramount factor for the induction of necrosis in solid tumors under "physiological" in situ conditions.

Heterogeneous oxygenation of rectal carcinomas in humans: a critical parameter for preoperative irradiation?

Tissue oxygenation was measured in 10 patients with differentiated adenocarcinoma in a very localized region in the middle part of the rectum (grade I - II, clinical stage II) by means of a cryophotometric micromethod. The results obtained clearly show that the oxygenation of differentiated rectal adenocarcinoma is distinctly lower than that of the normal rectal mucosa; tissue hypoxia or even anoxia are a common feature in those tumors; There exist considerable inter- individual differences among tumors of the same clinical staging and histological grading; substantial intra- individual heterogeneities in the oxygenation are evident within the same tumor and even within neighbouring microareas of the tissue. These findings imply that the commonly used classifications do not allow any conclusions concerning the oxygenation status, and probably the radiosensitivity of a tumor, respectively.

Effectiveness of respiratory hyperoxia, of normobaric and of hyperbaric oxygen atmospheres in improving tumor oxygenation.

Tumor oxygenation during respiratory hyperoxia is dependent on the tumor growth site, on the growth stage, and hence on the vascular pattern. Diffusion of O2 from the surrounding atmosphere contributes considerably to the oxygenation of subcutaneous tumors during normobaric exposure to a pure O2 atmosphere. However, hypoxia is still present in the tumor core under these conditions, and pressurization up to 4 bar is required to completely eradicate these hypoxic areas, thus enhancing the radiosensitivity of the tumors.

Intracapillary HbO2 saturation in tumor tissue of DS-carcinosarcoma during normoxia.

The measurements of the intravascular HbO2 saturation in tumor capillaries using the cryophotometric micromethod reveal that very low HbO2 saturation values predominate in malignant tumors. Under normoxic conditions only 8% of the measured values exceed 50% saturation. 53% of the intracapillary HbO2 saturation values are in the range of 0-10% HbO2 saturation. Great regional differences are seldom and can be found only in areas where a sufficient vascularization still exists. Taking into account the data of morphometric analysis of tumor vascularization and the parameters of respiratory gas exchange, the measured frequency distribution of HbO2 saturation values in tumor capillaries is simulated by means of different three-dimensional capillary structures; On the basis of these microcirculatory units, tissue pO2 values are computed in the intercapillary regions. As a result of these calculations it can be stated that the computed data derived from intracapillary HbO2 saturations are in sufficient agreement with pO2 values measured polarographically using gold- microelectrodes.

Impact of various thermal doses on the oxygenation and blood flow in malignant tumors upon localized hyperthermia.

Upon localized hyperthermia at modest thermal doses an increase in tumor blood flow can be observed in many tumors which is paralleled by an improvement of the oxygenation status of the tissue. At intermediate or high thermal doses a pronounced restriction of the tumor circulation becomes obvious leading to a deterioration of the tumor oxygenation. As a consequence, a further enhancement of the thermal response of tumors relative to normal tissues has to be expected at intermediate or high thermal doses.

Anxious and depressive symptoms in adolescents: epidemiological data of a large scale study in Dresden.

OBJECTIVES: The present large scale epidemiological study was designed to assess the prevalence of mental disorders in adolescents. METHODS: Two cross-sectional studies have been performed in adolescents in Dresden and the results of the examination of 627 high school and 485 secondary school students (mean age 15.5 years) are presented. Self rating procedures like the Beck Depression inventory (BDI) and clinical interviews have been used to enhance validity. RESULTS: The results underline the high prevalence rates of anxiety and depression in adolescents. Up to 30% of the 9th and 10th grades students suffer from mental disorders or are at risk for the development of mental disorders. CONCLUSIONS: Therefore primary prevention of mental disorders is desirable.

[Anxiety and depression in students. A study in Dresden middle schools and high schools]. / Angst und Depression bei Schülern. Eine Untersuchung an Dresdner Mittelschulen und Gymnasien.

OBJECTIVES: Recent investigations by the WHO have shown once again the high prevalence of mental disorders in European countries. Mainly in the field of mental disorders considerable deficits exist particularly with regard to primary prevention. METHODS: Although there is no doubt as to the importance of primary prevention within the scope of Public Health, there are deficits in this branch in Germany. This is unfortunate, considering that the proof of successful interventions is evidence for causal risk models. The current investigation is part of a study focusing upon the development, implementation and evaluation of a prevention program for anxiety and depression in adolescents and young adults. RESULTS: Presented here are the prevalence rates of anxiety disorders and depression ascertained in the first cross-sectional study carried out in secondary schools and high schools. A total of 627 high school students and 485 secondary school students in grades nine and ten in Dresden were examined.

Notch pathway inhibition controls myeloma bone disease in the murine MOPC315.BM model.

Despite evidence that deregulated Notch signalling is a master regulator of multiple myeloma (MM) pathogenesis, its contribution to myeloma bone disease remains to be resolved. Notch promotes survival of human MM cells and triggers human osteoclast activity in vitro. Here, we show that inhibition of Notch through the γ-secretase inhibitor XII (GSI XII) induces apoptosis of murine MOPC315.BM myeloma cells with high Notch activity. GSI XII impairs murine osteoclast differentiation of receptor activator of NF-κB ligand (RANKL)-stimulated RAW264.7 cells in vitro. In the murine MOPC315.BM myeloma model GSI XII has potent anti-MM activity and reduces osteolytic lesions as evidenced by diminished myeloma-specific monoclonal immunoglobulin (Ig)-A serum levels and quantitative assessment of bone structure changes via high-resolution microcomputed tomography scans. Thus, we suggest that Notch inhibition through GSI XII controls myeloma bone disease mainly by targeting Notch in MM cells and possibly in osteoclasts in their microenvironment. We conclude that Notch inhibition is a valid therapeutic strategy in MM.

Education research: cognitive performance is preserved in sleep-deprived neurology residents.

OBJECTIVE: To test the hypotheses that sleep deprivation in neurology residents is associated with performance deficits and that vigilance and cognitive performance is more compromised after overnight on-call duty compared to night shift. METHODS: Thirty-eight neurology residents of a university teaching hospital participated in a prospective single-blind comparison study. Residents were recruited according to their working schedule and divided into 3 groups: 24 hours overnight on-call duty, night shift, and regular day shift (controls). All participants underwent serial measurements of sleepiness and cognitive performance in the morning directly after or before their shift. Pupillary sleepiness test and Paced Auditory Serial Addition Test were applied. Perceived sleepiness was assessed by a questionnaire. RESULTS: Sleepiness was increased in residents after night shift and overnight call compared to controls while the type of night duty was not associated with the extent of sleepiness. Sleep-deprived residents did not show any performance deficits on the Paced Auditory Serial Addition Test. Cognitive performance was not associated with sleepiness measures. CONCLUSIONS: Night shift and overnight call duty have a similar impact on alertness in neurology residents. Sleep-deprived neurology residents may be able to overcome sleep loss-related performance difficulties for short periods.