Boswellic acids from frankincense inhibit lipopolysaccharide functionality through direct molecular interference

Lipophilic extracts of gum resins of Boswellia species (BSE) are used in folk medicine to treat various inflammatory disorders and infections. The molecular background of the beneficial pharmacological effects of such extracts is still unclear. Various boswellic acids (BAs) have been identified as abundant bioactive ingredients of BSE. Here we report the identification of defined BAs as direct inhibitors of lipopolysaccharide (LPS) functionality and LPS-induced cellular responses. In pull-down experiments, LPS could be precipitated using an immobilized BA, implying direct molecular interactions. Binding of BAs to LPS leads to an inhibition of LPS activity which was observed in vitro using a modified limulus amoebocyte lysate assay. Analysis of different BAs revealed clear structure-activity relationships with the classical β-BA as most potent derivative (IC(50)=1.8 μM). In RAW264.7 cells, LPS-induced expression of inducible nitric oxide synthase (iNOS, EC 1.14.13.39) was selectively inhibited by those BAs that interfered with LPS activity. In contrast, interferon-γ-induced iNOS induction was not affected by BAs. We conclude that structurally defined BAs are LPS inhibiting agents and we suggest that β-BA may contribute to the observed anti-inflammatory effects of BSE during infections by suppressing LPS activity.     

On the inhibition of 5-lipoxygenase product formation by tryptanthrin: mechanistic studies and efficacy in vivo

BACKGROUND AND PURPOSE

Leukotrienes (LTs) are pro-inflammatory mediators produced by 5-lipoxygenase (5-LO). Currently available 5-LO inhibitors either lack efficacy or are toxic and novel approaches are required to establish a successful anti-LT therapy. Here we provide a detailed evaluation of the effectiveness of the plant-derived alkaloid tryptanthrin as an inhibitor of LT biosynthesis.

EXPERIMENTAL APPROACH

We analysed LT formation and performed mechanistic studies in human neutrophils stimulated with pathophysiologically relevant stimuli (LPS and formyl peptide), as well as in cell-free assays (neutrophil homogenates or recombinant human 5-LO) and in human whole blood. The in vivo effectiveness of tryptanthrin was evaluated in the rat model of carrageenan-induced pleurisy.

KEY RESULTS

Tryptanthrin potently reduced LT-formation in human neutrophils (IC₅₀ = 0.6 µM). However, tryptanthrin is not a redox-active compound and did not directly interfere with 5-LO activity in cell-free assays. Similarly, tryptanthrin did not inhibit the release of arachidonic acid, the activation of MAPKs, or the increase in [Ca²⁺]_i, but it modified the subcellular localization of 5-LO. Moreover, tryptanthrin potently suppressed LT formation in human whole blood (IC₅₀ = 10 µM) and reduced LTB₄ levels in the rat pleurisy model after a single oral dose of 10 mg·kg⁻¹.

CONCLUSIONS AND IMPLICATIONS

Our data reveal that tryptanthrin is a potent natural inhibitor of cellular LT biosynthesis with proven efficacy in whole blood and is effective in vivo after oral administration. Its unique pharmacological profile supports further analysis to exploit its pharmacological potential.     

A closed system for islet isolation and purification using the COBE2991 cell processor may reduce the need of clean room facilities

During the isolation of human islets of Langerhans the digest has repeated direct contact with the ambient atmosphere. In order to fulfill GMP requirements in clinical applications, the entire cell preparation must be performed in clean room facilities. We hypothesized that the use of a closed system, which avoids the direct exposure of tissue to the atmosphere, would significantly ease the preparation procedure. To avoid the direct atmosphere exposure we tested a modification of the isolation and purification process by performing all islet preparation steps in a closed system. In this study we compared the isolation outcome of the traditional open preparation technique with the new closed system. Pancreata from 6-month-old hybrid pigs were procured in the local slaughterhouse. After digestion/filtration the digest was cooled, collected, and concentrated in centrifugation containers and purified thereafter in the COBE2991 by top loading (control). In the control group 502 +/- 253 IEQ per gram pancreas were purified. The total preparation time amounted to 12 h. In the closed system the digest was cooled and directly pumped into the COBE2991 for centrifugation followed by supernatant expelling. Bag filling, centrifugation, and expelling were repeated several times. Islets in pellet form were then purified by adding a gradient (bottom loading). Using this closed system 1098 +/- 489 IEQ per gram pancreas were purified with a total cell viability of 67 +/- 10% and a beta-cell viability of 41 +/- 13%. The total preparation time reduced to 6 h. After 24 h of cell culture the viability of beta-cells was still 56 +/- 10% and was only reduced after the addition of proapoptotic IL-1 and TNF-alpha to 40 +/- 4%, indicating that freshly isolated islets are not apoptotic. In conclusion, the closed system preparation is much faster, more effective, and less expensive than the traditional islet preparation. The closed system may be applicable for human islets preparations to restrict the need of clean room facilities for islet preparations to a minimum and may open the way for islet preparations without clean room demand.     

Dissociation between anterior and posterior cortical regions during self-specificity and familiarity: a combined fMRI-meta-analytic study

The familiarity to the subject of any potential stimuli presents one of the major difficulties for the investigation of the self; the separation of effects resulting from familiarity from self-effects being extremely problematic. The aim of this study was thus to investigate the neural distinction between self and familiarity by combining two sets of fMRI data with a meta-analysis. In the first fMRI experiment, regions responding to self/familiarity were investigated using the subject's own name and names of familiar others. These effects were confirmed and extended in a second fMRI experiment using the subject's own name and a stranger's name, as spoken by familiar and unfamiliar voices. Finally, a meta-analysis of self- and familiarity-related studies was conducted. Neural activity in the anterior brain regions, such as the anterior cingulate (ACC) and anterior insula (AI), was found to be specific for self-specific stimuli. In contrast, posterior brain regions, such as the posterior cingulate, were activated by familiar stimuli. Finally, the distinction between anterior and posterior regions for self and familiarity was confirmed by meta-analytic data. This study demonstrates a clear anterior-posterior cortical partition between self-specificity and familiarity.     

Auswirkungen der Leukozytendepletion auf die Lagerungsqualität von Vollblut

Background: According to the German Guidelines for Hemotherapy, preoperative autologous blood donations can be stored and transfused as whole blood. In contrast to the storage of blood components, however, storage of whole blood results in deterioration of the biological quality due to contamination with leukocytes and platelets. In this study we examined the influence of prestorage leukocyte depletion on the quality of filtered whole blood donations during storage. Material and Methods: Whole blood was donated by healthy volunteers (n=14, 500ml, 70?ml CPDA-1) and was filtered using an integrated whole blood filter system (Leukotrap A1, Fa. Pall, Dreieich, Germany). The leukocyte-depleted whole blood units were then stored for up to 49?days. Several hematological and biochemical parameters indicative of the quality of blood donations were determined during storage. Results: Our data indicate a quality of leukocyte-depleted whole blood donations comparable to buffy-coat-free red blood cell units. Conclusions: The Leukotrap A1 whole blood filter system is an interesting option for hospitals lacking the technical capacity to separate whole blood into components. The clinical suitability remains to be investigated by means of clinical studies.     

Change in perforin-positive peripheral blood lymphocyte (PBL) subpopulations following exercise

Perforin, one of the cytotoxic proteins of the immune system, plays a prominent role in protection against viral and bacterial infections. We investigated its expression in PBL and their CD3+, CD4+, CD8+ and CD16+ and/or CD56+ subpopulations in endurance athletes before and after a triathlon. Lymphocyte subpopulations were analysed by flow cytometry following separation of peripheral blood mononuclear cells and staining with antibodies against specific membrane antigens and intracellular perforin. The number of total lymphocytes decreased from 2.1 x 10(3)/microl before the triathlon to 1.0 x 10(3)/microl 1 h after the triathlon (P < 0.01). Interestingly, there was already a significant spontaneous decline in the percentage of CD3+/perforin+, and in CD8+/perforin+ cells, in the week proceeding the triathlon, when subjects were instructed to refrain from strenuous exercise training. The percentage of CD3+/perforin+, CD8+/perforin+ and CD16+ and/or CD56+/perforin+ cells in each lymphocyte subpopulation decreased 1 h after exercise even further from 14.3% to 5.8% (P < 0.05), 18.5% to 6.5% (P < 0.05) and 77.3% to 67.3%, respectively. However, at 18 h and 48 h after exercise the percentage of perforin-expressing CD3+, CD8+ and CD16+/56+ cells increased again towards baseline levels. Compared with normal controls, baseline perforin co-expression in CD3+ and CD8+ lymphocytes was significantly higher in trained athletes. From our data we conclude that trained athletes have an increased percentage of perforin+ PBL and that following exercise the percentage of perforin+ and therefore potentially cytotoxic lymphocytes transiently decreases in peripheral blood.     

Intrinsic neural network dynamics in catatonia

Catatonia is a transnosologic psychomotor syndrome with high prevalence in schizophrenia spectrum disorders (SSD). There is mounting neuroimaging evidence that catatonia is associated with aberrant frontoparietal, thalamic and cerebellar regions. Large‐scale brain network dynamics in catatonia have not been investigated so far. In this study, resting‐state fMRI data from 58 right‐handed SSD patients were considered. Catatonic symptoms were examined on the Northoff Catatonia Rating Scale (NCRS). Group spatial independent component analysis was carried out with a multiple analysis of covariance (MANCOVA) approach to estimate and test the underlying intrinsic components (ICs) in SSD patients with (NCRS total score ≥ 3; n = 30) and without (NCRS total score = 0; n = 28) catatonia. Functional network connectivity (FNC) during rest was calculated between pairs of ICs and transient changes in connectivity were estimated using sliding windowing and clustering (to capture both static and dynamic FNC). Catatonic patients showed increased static FNC in cerebellar networks along with decreased low frequency oscillations in basal ganglia (BG) networks. Catatonic patients had reduced state changes and dwelled more in a state characterized by high within‐network correlation of the sensorimotor, visual, and default‐mode network with respect to noncatatonic patients. Finally, in catatonic patients according to DSM‐IV‐TR (n = 44), there was a significant correlation between increased within FNC in cortico‐striatal state and NCRS motor scores. The data support a neuromechanistic model of catatonia that emphasizes a key role of disrupted sensorimotor network control during distinct functional states.