Comparison of three methods for the determination of serum alpha-1-antitrypsin in patients with pulmonary diseases

In patients with pulmonary diseases, serum alpha 1-antitrypsin (AAT) was measured by three methods: radial immunodiffusion (RID), trypsin inhibitory capacity assay (TIC) and by rate nephelometry with the immunosystem (NIA) in a total of 369 subjects (sarcoidosis, n = 35; asthma, n = 41; chronic obstructive bronchitis, n = 62; bronchogenic carcinoma, n = 93; pneumonia, n = 24; tuberculosis, n = 43; fibrosis, n = 22; healthy controls, n = 49). Considering all patients, AAT was found to be significantly elevated (p less than 0.01-0.001) in all methods (RID: 3.3 +/- 1.0 g/l; TIC: 2.7 +/- 0.4 g/l; NIA: 2.1 +/- 0.8 g/l) compared to healthy controls (RID: 2.1 +/- 0.3 g/l; TIC: 2.1 +/- 0.4 g/l; NIA: 1.2 +/- 0.3 g/l). The lowest mean values were found by means of the NIA method. The best correlation coefficient (R) was evaluated between the TIC and the NIA method (R = 0.96) in healthy controls, but the best correlated methods were the RID and the NIA (R = 0.93) in patients with pulmonary disease.     

Quantitative analysis of adenosine using liquid chromatography/atmospheric pressure chemical ionization-tandem mass spectrometry (LC/APCI-MS/MS)

Adenosine-secreting cellular brain implants constitute a promising therapeutic approach for the treatment of epilepsy. To engineer neural stem cells for therapeutic adenosine delivery, a reliable and fast analytical method is necessary to quantify cell-based adenosine release. Here we describe the development, optimization and validation of adenosine measurement using liquid chromatography–atmospheric pressure chemical ionization-tandem mass spectrometry (LC–APCI-MS/MS). LC–MS/MS in positive ion mode used selected reaction monitoring at m/z of 268.2/136.1 and 302.2/170.0 for adenosine and the internal standard, respectively. The bias was within 15% of the nominal value and evaluation of precision showed a relative standard deviation lower than 15% for all measured concentrations. The lower limit of quantification of adenosine was 15.6 ng/ml. Freeze and thaw stability and processed sample stability also fulfilled the acceptance criteria. Evaluation of the matrix effect showed that the method is not affected by relative matrix effects. The major advantages of this method are the absence of an extraction phase and the combination of the high selectivity and sensitivity characteristic for the LC–MS/MS technique, with a short run time of 4.5 min. These results demonstrate that this method is a useful tool to measure adenosine concentrations in culture medium released from stem cells in vitro.     

The ends of a large RNA molecule are necessarily close

We show on general theoretical grounds that the two ends of single-stranded (ss) RNA molecules (consisting of roughly equal proportions of A, C, G and U) are necessarily close together, largely independent of their length and sequence. This is demonstrated to be a direct consequence of two generic properties of the equilibrium secondary structures, namely that the average proportion of bases in pairs is ∼60% and that the average duplex length is ∼4. Based on mfold and Vienna computations on large numbers of ssRNAs of various lengths (1000–10 000 nt) and sequences (both random and biological), we find that the 5′–3′ distance—defined as the sum of H-bond and covalent (ss) links separating the ends of the RNA chain—is small, averaging 15–20 for each set of viral sequences tested. For random sequences this distance is ∼12, consistent with the theory. We discuss the relevance of these results to evolved sequence complementarity and specific protein binding effects that are known to be important for keeping the two ends of viral and messenger RNAs in close proximity. Finally we speculate on how our conclusions imply indistinguishability in size and shape of equilibrated forms of linear and covalently circularized ssRNA molecules.     

Cytokinin producing bacteria enhance plant growth in drying soil

Cytokinins can promote stomatal opening, stimulate shoot growth and decrease root growth. When soil is drying, natural cytokinin concentrations decrease in association with stomatal closure and a redirection of growth away from the shoots to the roots. We asked if decreased cytokinin concentrations mediate these adaptive responses by lessening water loss and promoting root growth thereby favouring exploration for soil water. Our approach was to follow the consequences for 12-d-old lettuce seedlings of inoculating the growing medium with cytokinin-producing bacteria under conditions of water sufficiency and deficit. Inoculation increased shoot cytokinins as assessed by immunoassay and mass spectrometry. Inoculation also promoted the accumulation of shoot mass and shortened roots while having a smaller effect on root mass. Inoculation did not raise stomatal conductance. The possible promoting effect of these cytokinins on stomatal conductance was seemingly hampered by increases in shoot ABA that inoculation also induced. Inoculation lowered root/shoot ratios by stimulating shoot growth. The effect was greater in non-droughted plants but remained sufficiently strong for shoot mass of inoculated droughted plants to exceed that of well-watered non-inoculated plants. We conclude that compensating for the loss of natural cytokinins in droughted plants interferes with the suppression of shoot growth and the enhancement of root elongation normally seen in droughted plants.     

Prevention of Candida albicans Biofilm Formation by Covalently Bound Dimethylaminoethylmethacrylate and Polyethylenimine

Candida albicans biofilms are a major cause of voice prosthesis deterioration in laryngectomized patients. The aim of this study was to produce a surface capable of inhibiting C. albicans biofilm formation. Dimethylaminoethylmethacrylate (DMAEMA) and polyethylenimine (PEI) moieties were covalently bound to the surface of polydimethylsiloxane (PDMS) or polymethylmethacrylate (PMMA) and subsequently quaternized. Physicochemical characterization of the grafted surfaces was carried out and their effect on C. albicans cell numbers was assessed using a modified Robbins device to grow the biofilms. Covalently bound quaternized polyDMAEMA (polyDMAEMAq) and PEI (PEIq) inhibited biofilm growth, with reductions up to 92%. Our approach may show promise for future application in medical devices such as catheters and prostheses.     

Identification of a series of substituted 2-piperazinyl-5-pyrimidylhydroxamic acids as potent histone deacetylase inhibitors

Pursuing our efforts in designing 5-pyrimidylhydroxamic acid anti-cancer agents, we have identified a new series of potent histone deacetylase (HDAC) inhibitors. These compounds exhibit enzymatic HDAC inhibiting properties with IC₅₀ values in the nanomolar range and inhibit tumor cell proliferation at similar levels. Good solubility, moderate bioavailability, and promising in vivo activity in xenograft model made this series of compounds interesting starting points to design new potent HDAC inhibitors.     

Sur2 mutations of Arabidopsis thaliana define a new locus involved in the control of auxin homeostasis

A new auxin homeostasis gene in Arabidopsis called SUR2 has been identified. This gene, mapped to the bottom of chromosome 4, is defined by two recessive nuclear mutants designated superroot2 (sur2), which display several abnormalities reminiscent of auxin effects. A number of these characteristics are similar to the phenotype of the previously described auxin-overproducing mutant superroot1 (sur1); however, several lines of evidences reveal that the SUR2 gene defines a new key point in the regulation of endogenous auxin concentrations. The phenotype of the sur1 sur2 double mutant is additive. Analysis by gas chromatography coupled to mass spectrometry indicated increased levels of free indole-3-acetic acid correlated with a decreased level of bound auxin in the sur2 mutant. These results suggest that SUR2 may be involved in the control of auxin conjugation.     

CALIB: a Bioconductor package for estimating absolute expression levels from two-color microarray data

In this article we describe a new Bioconductor package 'CALIB' for normalization of two-color microarray data. This approach is based on the measurements of external controls and estimates an absolute target level for each gene and condition pair, as opposed to working with log-ratios as a relative measure of expression. Moreover, this method makes no assumptions regarding the distribution of gene expression divergence. AVAILABILITY: http://bioconductor.org/packages/2.0/bioc Open Source.     

Pluronic and Tetronic Copolymers with Polyglycolyzed Oils as Self-Emulsifying Drug Delivery Systems

The potential of poly(ethylene oxide)-poly(propylene oxide) block copolymers Pluronic F127 (PF127) and Tetronic 304 (T304), 904 (T904) and 1307 (T1307) as components of solid self-(micro)emulsifying dosage forms, S(M)EDDS, was evaluated. The dependence of the self-associative properties of Tetronics on pH explained the low ability of the micelles to solubilize griseofulvin at acid pH (sevenfold increase) compared to at alkaline pH (12-fold). Blends of polyglycolyzed glycerides (Labrasol, Labrafac CC, and Labrafil M 1944CS) with each copolymer at two different weight ratios (80:20 and 60:40) were prepared, diluted in water, and characterized in terms of globule size, appearance and griseofulvin solubility. The blends with Labrasol led to microemulsions that are able to increase drug solubility up to 30-fold. SMEDD hard gelatine capsules filled with griseofulvin and Labrasol or Labrasol/copolymer 80:20 showed a remarkable increase in drug solubility and dissolution rate, particularly when T904, T1307 or PF127 was present in the blend. This effect was more remarkable when the volume of the dissolution medium was 200 ml (compared to 900 ml), which can be related to a higher stability of the microemulsion when there is a greater concentration of the copolymer and glyceride in the medium.