Sensitive fluorescence in situ hybridization signal detection in maize using directly labeled probes produced by high concentration DNA polymerase nick translation

The signal produced by fluorescence in situ hybridization (FISH) often is inconsistent among cells and sensitivity is low. Small DNA targets on the chromatin are difficult to detect. We report here an improved nick translation procedure for Texas red and Alexa Fluor 488 direct labeling of FISH probes. Brighter probes can be obtained by adding excess DNA polymerase I. Using such probes, a 30 kb yeast transgene, and the rp1, rp3 and zein multigene clusters were clearly detected.     

Similar hypotensive responses to resistance exercise with and without blood flow restriction

Low intensity resistance exercise (RE) with blood flow restriction (BFR) has gained attention in the literature due to the beneficial effects on functional and morphological variables, similar to those observed during traditional RE without BFR, while the effects of BFR on post-exercise hypotension remain unclear. The aim of the present study was to compare the blood pressure (BP) response of trained normotensive individuals to RE with and without BFR. In this cross-over randomized trial, eight male subjects (23.8 ± 4 years, 74 ± 3 kg, 174 ± 4 cm) completed two exercise protocols: traditional RE (3 x 10 repetitions at 70% one-repetition maximum [1-RM]) and low intensity RE (3 x 15 repetitions at 20% 1-RM) with BFR. Blood pressure measurements were performed after 15 min of seated rest (0), immediately after and 10 min, 20 min, 30 min, 40 min, 50 min and 60 min after the experimental sessions. Similar hypotensive effects for systolic BP (SBP) were observed for both protocols (P < 0.05) after exercise, with no differences between groups (P > 0.05) and no statistically significant difference for diastolic BP (P > 0.05). These results suggest that in normotensive trained individuals, both traditional RE and RE with BFR induce hypotension for SBP, which is important to prevent cardiovascular disturbances.     

Treatment with recombinant interferon-β reduces inflammation and slows cartilage destruction in the collagen-induced arthritis model of rheumatoid arthritis

We investigated the therapeutic potential and mechanism of action of IFN-β protein for the treatment of rheumatoid arthritis (RA). Collagen-induced arthritis was induced in DBA/1 mice. At the first clinical sign of disease, mice were given daily injections of recombinant mouse IFN-β or saline for 7 days. Disease progression was monitored by visual clinical scoring and measurement of paw swelling. Inflammation and joint destruction were assessed histologically 8 days after the onset of arthritis. Proteoglycan depletion was determined by safranin O staining. Expression of cytokines, receptor activator of NF-κB ligand, and c-Fos was evaluated immunohistochemically. The IL-1-induced expression of IL-6, IL-8, and granulocyte/macrophage-colony-stimulating factor (GM-CSF) was studied by ELISA in supernatant of RA and osteoarthritis fibroblast-like synoviocytes incubated with IFN-β. We also examined the effect of IFN-β on NF-κB activity. IFN-β, at 0.25 μg/injection and higher, significantly reduced disease severity in two experiments, each using 8–10 mice per treatment group. IFN-β-treated animals displayed significantly less cartilage and bone destruction than controls, paralleled by a decreased number of positive cells of two gene products required for osteoclastogenesis, receptor activator of NF-κB ligand and c-Fos. Tumor necrosis factor α and IL-6 expression were significantly reduced, while IL-10 production was increased after IFN-β treatment. IFN-β reduced expression of IL-6, IL-8, and GM-CSF in RA and osteoarthritis fibroblast-like synoviocytes, correlating with reduced NF-κB activity. The data support the view that IFN-β is a potential therapy for RA that might help to diminish both joint inflammation and destruction by cytokine modulation.     

GMP-140 (P-selectin) inhibits human neutrophil activation by lipopolysaccharide: analysis by proton magnetic resonance spectroscopy.

Proton magnetic resonance spectroscopy has been used to monitor the effect of GMP-140 on the stimulation of human peripheral blood neutrophils. Stimulation of neutrophils by lipopolysaccharide gives rise to a high resolution lipid spectrum from the intact cells. Fluid phase GMP-140, which prevents adhesion and development of inflammatory responses of neutrophils, was found to inhibit these changes in the lipid spectrum by up to 40%. Anti-GMP-140 Fab fragments reversed this effect while non-immune Fab fragments did not affect the observed inhibition by GMP-140.     

18S rRNA data indicate that Aschelminthes are polyphyletic in origin and consist of at least three distinct clades

The Aschelminthes is a collection of at least eight animal phyla, historically grouped together because the absence of a true body cavity was perceived as a pseudocoelom. Analyses of 18S rRNA sequences from six Aschelminth phyla (including four previously unpublished sequences) support polyphyly for the Aschelminthes. At least three distinct groups of Aschelminthes were detected: the Priapulida among the protostomes, the Rotifera-Acanthocephala as a sister group to the protostomes, and the Nematoda as a basal group to the triploblastic Eumetazoa.      

Epitope-specific changes in chondroitin sulfate/dermatan sulfate proteoglycans as markers in the lymphopoietic and granulopoietic compartments of developing bursae of Fabricius

The types and distributions of chondroitin sulfate proteoglycans within developing chick bursae of Fabricius were determined by indirect immunocytochemical analyses using mAb specific for chondroitin/dermatan sulfate epitopes. Analyses obtained from the use of well characterized mAb known to specifically identify chondroitin 4- and dermatan sulfates (antibody 2B6) and chondroitin 6-sulfate (antibody 3B3) were compared with those obtained from two additional mAb raised against chick chondroitin sulfates proteoglycans derived from hemopoietic tissue. The results indicate that chondroitin sulfate compositions of the adjacent lymphopoietic and granulopoietic compartments differ. Chondroitin 6-sulfate, notably absent from lymphopoietic regions, is a major chondroitin sulfate species in granulopoietic regions of day 13 bursae. Moreover, chondroitin 6-sulfate disappears from the granulopoietic compartment in a time course that corresponds to the decline in granulopoietic activity. Simultaneously, there is an apparent increase in chondroitin sulfates associated with developing medullary regions of lymphoid follicles. The content of chondroitin 4-/dermatan sulfates and, most significantly, of chondroitin/dermatan sulfates identified by antibodies raised against chick proteoglycans, increases within developing follicles. As a consequence, by day 18 of incubation, immunostained follicles become clearly demarcated from the connective tissue of the tunica propria. This study provides evidence that chondroitin sulfates are constituents of both lymphopoietic and granulopoietic microenvironments and that subtle changes occur within these proteoglycan structures during bursal development. These developmental changes in chondroitin sulfate compositions are consistent with these molecules playing a functional role in hemopoiesis.     

Interspecific and intraspecific comparisons of the period locus in the Drosophila willistoni sibling species

The period (per) locus has received much attention in molecular evolution studies because it is one of the best studied "behavioral genes" and because it offers insight into the evolution of repetitive sequences. We studied most of the coding region of per in Drosophila willistoni and confirmed previously observed patterns of conservation and divergence among distantly related species. Five regions are so highly diverged that they cannot be aligned, whereas a region encompassing the PAS domain is very conserved. Structural and nucleotide polymorphism patterns in the willistoni group are not the same as those observed in previously studied species. We sequenced the region homologous to the highly polymorphic threonine-glycine repeat of D. melanogaster in multiple strains of D. willistoni, as well as in other members of willistoni group, and found an unusual amount of conservation in this region. However, the next nonconserved region downstream in the sequence is quite variable and polymorphic for the number of repeated glycines. The glycine codon usage is significantly different in this glycine repeat as compared to other parts of the gene. We were able to plot the directionality of change in the glycine repeat region onto a phylogeny and find that the addition of glycines is the general trend with the diversification of the willistoni group.