Spatial Cytoskeleton Organization Supports Targeted Intracellular Transport

The efficiency of intracellular cargo transport from specific sources to target locations is strongly dependent upon molecular motor-assisted motion along the cytoskeleton. Radial transport along microtubules and lateral transport along the filaments of the actin cortex underneath the cell membrane are characteristic for cells with a centrosome. The interplay between the specific cytoskeleton organization and the motor performance results in a spatially inhomogeneous intermittent search strategy. To analyze the efficiency of such intracellular search strategies, we formulate a random velocity model with intermittent arrest states. We evaluate efficiency in terms of mean first passage times for three different, frequently encountered intracellular transport tasks: 1) the narrow escape problem, which emerges during cargo transport to a synapse or other specific region of the cell membrane; 2) the reaction problem, which considers the binding time of two particles within the cell; and 3) the reaction-escape problem, which arises when cargo must be released at a synapse only after pairing with another particle. Our results indicate that cells are able to realize efficient search strategies for various intracellular transport tasks economically through a spatial cytoskeleton organization that involves only a narrow actin cortex rather than a cell body filled with randomly oriented actin filaments.     

Role of endogenous and exogenous antioxidants in the defence against functional damage and lipid peroxidation in rat liver mitochondria

Mitochondria are cellular organelles where the generation of reactive oxygen species may be high. They are, however, effectively protected by their high capacities of antioxidative systems, as enzymes and either water or lipid soluble low molecular weight antioxidants.These antioxidative defence systems can be effectively regenerated after or during an oxidative stress as long as the mitochondria are in an energized state. Energization of mitochondria mainly depends on the availability of suitable respiratory substrates which can provide hydrogen for the reduction of either the glutathione- or -tocopherol-system, since GSH is regenerated by glutathione reductase with the substrate NADPH and the -tocopheroxyl-radical likely by reduced coenzyme Q. It was shown that mitochondria do not undergo damages as long as they can keep a high energy state. The delicate balance between prooxidative/antioxidative activities can be shifted towards oxidation, if experimentally prooxidants were added. After exhaustion of the antioxidative defence systems damages of rnitochondrial functions become expressed followed by membrane injuries along with the oxidation and degradation of mitochondrial lipids and proteins leading finally to the total degradation of the mitoc hondria.Extramitochondrial antioxidants may assist the mitochondrial antioxidative defence systems in a complex way, whereby particularly ascorbic acid can act both as prooxidant and as antioxidant. (Mol Cell Biochem 174: 199–205, 1997)     

De-novo identification of specific exposure biomarkers of the alternative plasticizer di(2-ethylhexyl) terephthalate (DEHTP) after low oral dosage to male volunteers by HPLC-Q-Orbitrap-MS

Context: Human exposure biomonitoring relies on the availability of specific, sensitive biomarkers. For emerging chemicals, the identification (prediction, synthesis, verification) of such biomarkers is time and cost intensive.

Objective: This study aimed to further elucidate the urinary metabolic profile of the plasticizer di(2-ethylhexyl) terephthalate (DEHTP) in search of probably additional biomarkers of exposure.

Materials and methods: Urine samples of an oral low-dose volunteer study were analysed by HPLC-Q-Orbitrap-MS combined with a commercial data mining software. Metabolite identification was based on isotopic pattern, accurate masses of product ions and excretion profiles.

Results: Nine phase I metabolites of DEHTP were tentatively identified by HPLC-Q-Orbitrap-MS. Four previously described, side chain oxidized monoester metabolites were confirmed in all samples. In addition, five previously unknown downstream metabolites were tentatively identified.

Discussion and conclusion: The excretion profiles obtained by HPLC-Q-Orbitrap-MS were in good agreement with quantitative HPLC-QqQ-MS data. For the newly discovered metabolites, plausible excretion profiles, similar to the ones of the known metabolites, were obtained. The presented approach proved to be successful for metabolite screening in urine samples after low-dose exposure and will be applied in future human metabolism studies for a fast, reliable and cost effective identification of specific biomarkers of exposure.     

Increased Plasma Levels of Heparin-Binding Protein on Admission to Intensive Care Are Associated with Respiratory and Circulatory Failure

Purpose

Heparin-binding protein (HBP) is released by granulocytes and has been shown to increase vascular permeability in experimental investigations. Increased vascular permeability in the lungs can lead to fluid accumulation in alveoli and respiratory failure. A generalized increase in vascular permeability leads to loss of circulating blood volume and circulatory failure. We hypothesized that plasma concentrations of HBP on admission to the intensive care unit (ICU) would be associated with decreased oxygenation or circulatory failure.

Methods

This is a prospective, observational study in a mixed 8-bed ICU. We investigated concentrations of HBP in plasma at admission to the ICU from 278 patients. Simplified acute physiology score (SAPS) 3 was recorded on admission. Sequential organ failure assessment (SOFA) scores were recorded daily for three days.

Results

Median SAPS 3 was 58.8 (48–70) and 30-day mortality 64/278 (23%). There was an association between high plasma concentrations of HBP on admission with decreased oxygenation (p<0.001) as well as with circulatory failure (p<0.001), after 48–72 hours in the ICU. There was an association between concentrations of HBP on admission and 30-day mortality (p = 0.002). ROC curves showed areas under the curve of 0,62 for decreased oxygenation, 0,65 for circulatory failure and 0,64 for mortality.

Conclusions

A high concentration of HBP in plasma on admission to the ICU is associated with respiratory and circulatory failure later during the ICU care period. It is also associated with increased 30-day mortality. Despite being an interesting biomarker for the composite ICU population it´s predictive value at the individual patient level is low.     

Simultaneous quantification of cholesterol sulfate,androgen sulfates,and progestagen sulfates in human serum by LC-MS/MS

Steroids are primarily present in human fluids in their sulfated forms. Profiling of these compounds is important from both diagnostic and physiological points of view. Here, we present a novel method for the quantification of 11 intact steroid sulfates in human serum by LC-MS/MS. The compounds analyzed in our method, some of which are quantified for the first time in blood, include cholesterol sulfate, pregnenolone sulfate, 17-hydroxy-pregnenolone sulfate, 16-α-hydroxy-dehydroepiandrosterone sulfate, dehydroepiandrosterone sulfate, androstenediol sulfate, androsterone sulfate, epiandrosterone sulfate, testosterone sulfate, epitestosterone sulfate, and dihydrotestosterone sulfate. The assay was conceived to quantify sulfated steroids in a broad range of concentrations, requiring only 300 μl of serum. The method has been validated and its performance was studied at three quality controls, selected for each compound according to its physiological concentration. The assay showed good linearity (R² > 0.99) and recovery for all the compounds, with limits of quantification ranging between 1 and 80 ng/ml. Averaged intra-day and between-day precisions (coefficient of variation) and accuracies (relative errors) were below 10%. The method has been successfully applied to study the sulfated steroidome in diseases such as steroid sulfatase deficiency, proving its diagnostic value. This is, to our best knowledge, the most comprehensive method available for the quantification of sulfated steroids in human blood.