Recombinant human macrophage colony-stimulating factor augments pulmonary host defences against Aspergillus fumigatus

The in vivo and ex vivo effects of macrophage colony-stimulating factor (M-CSF) were studied in a profoundly neutropenic rabbit model in order to determine its potential to augment pulmonary host defence against Aspergillus. M-CSF (100-600 microg/kg/d) was administered prophylactically to neutropenic rabbits with pulmonary aspergillosis starting three days pre-inoculation and then throughout neutropenia. Rabbits receiving M-CSF had significantly increased survival (P=0.01) and decreased pulmonary injury, as measured by decreased pulmonary infarction (P=0.004), when compared with untreated controls. Microscopic studies demonstrated greater numbers of activated pulmonary alveolar macrophages (PAMs) in lung tissue of rabbits receiving M-CSF, in comparison to controls (P<0.001). PAMs harvested from rabbits treated with M-CSF had a significantly greater percent phagocytosis of Aspergillus fumigatus conidia than did PAMs from controls (P=0.04). These data indicate that prophylactic administration of M-CSF augments pulmonary host defence against A. fumigatus and suggest a potential role for this cytokine as adjunctive therapy in the treatment of pulmonary aspergillosis in the setting of profound neutropenia.     

Lead uptake and the effects of EDTA on lead-tissue concentrations in the desert species mesquite (Prosopis spp.)

Experimental results have shown that the desert plant species mesquite (Prosopis spp.) is capable of accumulating high levels of lead in the roots, translocating it to the aerial portion of the plant. One-week-old mesquite seedlings were treated for 7 d in a hydroponic culture using a modified Hoagland solution. Six treatments were used; three treatments contained only Pb [as Pb(NO3)2] at 25-, 50-, and 75-mg L(-1) levels and three treatments contained the same levels of Pb, but with equimolar concentrations of disodium ethylenediamine tetraacetic acid (EDTA). Our results showed that the plants exposed to 25-, 50-, and 75-mg Pb L(-1) treatments without EDTA concentrated in stems 524, 3726, and 1417 mg kg(-1), respectively. However, the plants treated with Pb-EDTA concentrated in stems 480-, 607-, and 1247-mg Pb kg(-1) for the 25-, 50-, and 75-mg Pb L(-1) treatments, respectively. Results for the roots followed a similar trend; without EDTA the Pb levels ranged from 16,055, 89,935, and 63,396 for the 25-, 50-, and 75-mg Pb L(-1) treatments, respectively, and with EDTA these levels were 9,562, 49,902, and 39,181 mg kg(-1) for the three treatments. However, the addition of EDTA increased lead movement to the leaves. The levels of Pb without EDTA were 20, 35, and 51 mg kg(-1) for the 25-, 50-, and 75-mg Pb L(-1) levels, respectively. Treatments with EDTA showed uptake levels of 105, 124, and 313 for the 25-, 50-, and 75-mg Pb L(-1) treatments. Further, the percent Pb in dry leaf tissues for all EDTA treatments were greater than 0.1%. However, only the 25-mg Pb L(-1) treatment was greater than 0.1%, compared to 0.04 and 0.08% for the 50- and 75-mg Pb L(-1) treatments, respectively. Preliminary transmission and scanning electron microscopy corroborate the presence of lead.     

Characterization of pathogenic and nonpathogenic strains of Xanthomonas axonopodis pv. manihotis by PCR-based DNA fingerprinting techniques

Strains of Xanthomonas axonopodis pv. manihotis (Xam) were characterized for pathogenicity and for DNA polymorphism using different PCR-based techniques. Using amplified restriction fragment length polymorphism (AFLP), strains were distinguished from each other and also from other Xanthomonas strains. Cluster analysis showed a high correlation between DNA polymorphism and pathogenicity. Four Xam strains were further analyzed using three PCR-based techniques, AFLP, AFLP-pthB and RAPD-pthB. Various primer combinations were used including primers specific to a Xam pathogenicity gene (pthB) along with RAPD or AFLP primers. The AFLP primer combinations EcoRI+T/MseI+A and EcoRI+T/MseI+T were the most efficient to discriminate among pathogenic and nonpathogenic Xam strains. Polymorphic bands were excised from the gel, amplified and cloned. Sequences analysis showed significant homology with bacterial pathogenicity island, genes involved in pathogenic fitness and regulators of virulence. Three cloned AFLP fragments were used as probes in DNA blot experiments and two of them showed significant polymorphism.     

Elimination of oxalate contamination in RNA isolation from<Emphasis Type="Italic">Rumex obtusifolius</Emphasis>

Many plant RNA isolation techniques aim to prevent contamination by means of secondary phenolics, carbohydrates, RNase, and other chemicals. However, when applied in our laboratory to the isolation of RNA fromRumex obtusifolius, these protocols failed to produce good quality RNA. A major problem was contamination of the RNA samples with the secondary metabolite oxalate. The relative quantities of guanidine isothiocyanate extraction buffer to plant tissue used in the protocol had significant effects on oxalate contamination. An increase in extraction buffer, from 1.5 mL in the original method to 15 mL per 200–300 mg of tissue in our protocol, removed the oxalate from the RNA. This RNA was of a good quality and was suitable for molecular biology applications.     

Interactions between nucleic acid bases. New parameters of potential functions and energy minima

The parameters of atom-atom potential functions suggested by one of the authors in 1979-1986 were slightly changed. The changes were performed to achieve a better agreement with experimental data of interaction energy values in global minima and hydrogen bond lengths. These changes resulted in better accord with experimental values of distances between the layers in DNA monomer crystals and between the base pairs in oligonucleotide duplexes. The refined potential functions were used to calculate the energy of interactions between nucleic acid bases in various mutual positions. The calculations revealed a few types of mutual base arrangements in minima of interaction energy for each pairwise base combination. A new type of minima was found, which correspond to a nearly perpendicular arrangement of base rings and the formation of the intermolecular hydrogen bond.     

Characterization of a Smad motif similar to Drosophila mad in the mouse Msx 1 promoter

Mouse Msx 1 gene, orthologous of the Drosophila msh, is involved in several developmental processes. BMP family members are major proteins in the regulation of Msx 1 expression. BMP signaling activates Smad 1/5/8 proteins, which associate to Smad 4 before translocating to the nucleus. Analysis of Msx 1 promoter revealed the presence of three elements similar to the consensus established for Mad, the Smad 1 Drosophila counterpart. Notably, such an element was identified in an enhancer important for Msx 1 regulation. Gel shift analysis demonstrated that proteins from 13.5 dpc embryo associate to this enhancer. Remarkably, supershift assays showed that Smad proteins are present in the complex. Purified Smad 1 and 4 also bind to this fragment. We demonstrate that functional binding sites in this enhancer are confined to the Mad motif and flanking region. Our data suggest that this Mad motif may be functional in response to BMP signaling.     

Genetic polymorphism in eight Chilean strains of the carotenogenic microalga Dunaliella salina Teodoresco (Chlorophyta)

Eight Chilean strains of Dunaliella salina obtained within a restricted geographic range, but exhibiting a high variability in their morphology, rate of growth and carotenogenic capacity, were analyzed by Random Amplified Polymorphic DNA (RAPD-PCR) Twenty of the 50 random primers (D, P, OPA and OPD series) that were tested amplified reproducible bands and were useful for comparative analysis of the strains. Of 107 polymorphic genetic markers, 49 were strain-specific. A great genetic variability was found among the strains in spite of their geographic proximity. In addition, phenetic analysis of the data showed close agreement between the morphophysiological attributes and the genetic diversity of the strains.     

Non-destructive quantitation of spermine in human prostate tissue samples using HRMAS 1H NMR spectroscopy at 9.4 T

We present the results of a study of human prostate specimens evaluated by high resolution magic angle spinning (1)H nuclear magnetic resonance (NMR) spectroscopy at 400 MHz (9.4 T) and by quantitative histopathology. We demonstrate that NMR and pathology data can be obtained from the same intact specimens, and report for the first time a linear correlation between the NMR measured concentration of spermine, a proposed endogenous inhibitor to prostate cancer growth, and the volume percentage of normal prostatic epithelial cells as quantified by histopathology. Our results show that NMR may serve as a critical tool for the investigation of the inhibitory mechanism of spermine in human subjects.