Regulatory potential of ethanol and retinoic acid on human monocyte functions

Retinoic acid (RA), a metabolic product of vitamin A, has been shown to affect a variety of immune functions, including monocytes. Monocyte functions and mediator production are also modulated by ethanol exposure. This study demonstrates that therapeutic doses of RA (0.1-10 microM) significantly increase transforming growth factor-beta (TGF beta) production both in THP-1, human myelomonocytic cells, and in human peripheral blood monocytes. We have previously reported TGF beta induction by ethanol in human M theta. Combination of RA stimulation with acute in vitro ethanol treatment, however, resulted in significantly lower M theta TGF beta production than TGF beta levels induced by RA alone (p \u3c 0.003). Down-regulation of M theta TGF beta production by ethanol was tested at the concentration range of 25-150 mM and occurred both at high and low RA concentrations (10-0.1 microM). In contrast to its inhibitory effect on RA-induced M theta TGF beta production, ethanol augmented TGF beta production induced by muramyl dipeptide (20 micrograms/ml), suggesting that ethanol can either up- or down-regulate M theta TGF beta production, depending on the costimulatory factors. RA also induced a moderate increase in M theta tumor necrosis factor-alpha (TNF alpha) production, which was down-regulated by ethanol both at the level of secreted and cell-associated TNF alpha. In addition to regulation of cytokine production, both RA and ethanol decreased expression of CD4 on THP-1 cells. The degree of inhibition of CD4 expression by RA was more significant than by ethanol, but RA-induced decrease in CD4 expression was not significantly affected by the combined stimulation with ethanol.(ABSTRACT TRUNCATED AT 250 WORDS

Plasma modification of microporous polymer membranes for application in biomimetic dissolution studies

Abstract Biorelevant dissolution is an indispensable tool utilized during formulation development and optimization for the prediction of in vivo bioavailability of pharmaceutical agents. Within that framework, membrane-permeation dissolution methodologies are widely used to model drug absorption. The current work evaluates polymer membrane surface modifications for production of biomimetic membranes to be employed in biorelevant dissolution studies. Biomimetic membranes exhibit hydrophilic and hydrophobic properties to simulate the intestinal membrane environment. Low temperature plasma treatment of microporous polyethersulfone (PES), nylon and polypropylene (PP) polymer membranes was applied to produce low energy surface layers with permanent hydrophobic and hydrophilic functionalities. Surface modifications on microporous polymer membranes were achieved by plasma treatments using tetrafluoromethane (CF4), perfluorohexane (C6F14), dichloromethane (DCM) and water (H2O). Surface characterization of treated membranes was evaluated using scanning electron microscopy energy dispersive x-ray spectroscopy (SEM-EDS), water contact angle (CA) and x-ray photoelectron spectroscopy (XPS) techniques. SEM-EDS analysis of polymer membranes treated with fluorinated and chlorinated solvents/gases depicts altered surface morphologies with enriched porosity. SEM-EDS and XPS analyses demonstrate the chemical modification at the surface of treated membranes is strongly influenced by the type of treatment gas or solvent. Results show fluorination as a more effective and less destructive treatment technique. XPS confirmed the presence of elemental fluorine functional groups at the surface of the PES and nylon membranes. Evaluating elemental changes (F/C ratio) from multiple techniques confirms fluorinated plasma treatments are localized to the surface of the membrane and do not significantly affect the bulk properties. In a supplemental study, a detailed comparison of the plasma treated polymer membranes and porcine intestines revealed the biomimetic nature of the modified membranes

Enantioseparation of hydroxyeicosatetraenoic acids by hydroxypropyl-gamma-cyclodextrin-modified micellar electrokinetic chromatography

Complete resolution of hydroxyeicosatetraenoic acid (HETE) enantiomers was achieved using hydroxypropyl-gamma-cyclodextrin (HP-gamma-CD)-modified MEKC. The optimum running conditions were determined to be utilizing a 30 mM phosphate-15 mM borate buffer (pH 9.0) containing 30 mM HP-gamma-CD and 75 mM SDS as the BGE, application of +30 kV as the effective voltage, and carrying out the experiment at 15 degrees C. The eluents were detected at 235 nm. The method was used successfully for the simultaneous separations of (S)- and (R)-enantiomers of regioisomeric 8-, 11-, 12-, and 15-HETEs. Subsequently, the optimized method was applied to evaluate the stereochemistry of 8- and 12-HETEs from the marine red algae, Gracilaria vermiculophylla and Gracilaria arcuata, respectively. The 8-HETE was found to be a mixture of 98% (R)-enantiomer and 2% (S)-enantiomer, while the 12-HETE was a mixture of 98% (S)-enantiomer and 2% (R)-enantiomer. The present study demonstrates that the HP-gamma-CD-modified MEKC method is simple and sensitive and provides unambiguous information on the configuration of natural and synthetic HETEs

Simultaneous LC-MS/MS analysis of eicosanoids and related metabolites in human serum, sputum and BALF

The differences among individual eicosanoids in eliciting different physiological and pathological responses are largely unknown because of the lack of valid and simple analytical methods for the quantification of individual eicosanoids and their metabolites in serum, sputum and bronchial alveolar lavage fluid (BALF). Therefore, a simple and sensitive LC-MS/MS method for the simultaneous quantification of 34 eicosanoids in human serum, sputum and BALF was developed and validated. This method is valid and sensitive with a limit of quantification ranging from 0.2 to 3 ng/mL for the various analytes, and has a large dynamic range (500 ng/mL) and a short run time (25 min). The intra- and inter-day accuracy and precision values met the acceptance criteria according to US Food and Drug Administration guidelines. Using this method, detailed eicosanoid profiles were quantified in serum, sputum and BALF from a pilot human study. In summary, a reliable and simple LC-MS/MS method to quantify major eicosanoids and their metabolites was developed and applied to quantify eicosanoids in human various fluids, demonstrating its suitability to assess eicosanoid biomarkers in human clinical trials