Trapping Triatominae in silvatic habitats

Large-scale trials of a trapping system designed to collect silvatic Triatominae are reported. Live-baited adhesive traps were tested in various ecosystems and different triatomine habitats (arboreal and terrestrial). The trials were always successful, with a rate of positive habitats generally over 20% and reaching 48.4% for palm trees of the Amazon basin. Eleven species of Triatominae belonging to the three genera of public health importance (Triatoma, Rhodnius and Panstrongylus) were captured. This trapping system provides an effective way to detect the presence of triatomines in terrestrial and arboreal silvatic habitats and represents a promising tool for ecological studies. Various lines of research are contemplated to improve the performance of this trapping system.     

Solution structure of Pisum sativum defensin 1 by high resolution NMR: plant defensins, identical backbone with different mechanisms of action.

Pisum sativum defensin 1 (Psd1) is a 46 amino acid residue plant defensin isolated from seeds of pea. The three-dimensional structure in solution of Psd1 was determined by two-dimensional NMR data recorded at 600 MHz. Experimental restraints were used for structure calculation using CNS and torsion-angle molecular dynamics. The 20 lowest energy structures were selected and further subjected to minimization, giving a root-mean-square deviation of 0.78(+/- 0.22) A in the backbone and 1.91(+/-0.60) A for over all atoms of the molecule. The protein has a globular fold with a triple-stranded antiparalell beta-sheet and an alpha-helix (from residue Asn17 to Leu27). Psd1 presents the so called "cysteine stabilized alpha/beta motif" and presents identical three-dimensional topology in the backbone with other defensins and neurotoxins. Comparison of the electrostatic surface potential among proteins with high three-dimensional (selected using the softwares TOP and DALI) topology gave insights into the mode of action of Psd1. The surface topologies between proteins that present antifungal activity or sodium channel inhibiting activity are different. On the other hand the surface topology presents several common features with potassium channel inhibitors, suggesting that Psd1 presents this activity. Other common features with potassium channel inhibitors were found including the presence of a lysine residue essential for inhibitory activity. The identity of Psd1 in primary sequence is not enough to infer a mechanism of action, in contrast with the strategy proposed here.     

Static magnetic fields affect calcium fluxes and inhibit stress-induced apoptosis in human glioblastoma cells

BACKGROUND: Epidemiologic data revealed increased brain tumor incidence in workers exposed to magnetic fields (MFs), raising concerns about the possible link between MF exposure and cancer. However, MFs seem to be neither mutagenic nor tumorigenic. The mechanism of their tumorigenic effect has not been elucidated. METHODS: To evaluate the interference of MFs with physical (heat shock, HS) and chemical (etoposide, VP16) induced apoptoses, respectively, we exposed a human glioblastoma primary culture to 6 mT static MF. We investigated cytosolic Ca(2+) ([Ca(2+)](i)) fluxes and extent of apoptosis as key endpoints. The effect of MFs on HS- and VP16-induced apoptoses in primary glioblastoma cultures from four patients was also tested. RESULTS: Static MFs increased the [Ca(2+)](i) from a basal value of 124 +/- 4 nM to 233 +/- 43 nM (P < 0.05). MF exposure dramatically reduced the extent of HS- and VP16-induced apoptoses in all four glioblastoma primary cultures analyzed by 56% (range, 28-87%) and 44% (range, 38-48%), respectively. However, MF alone did not exert any apoptogenic activity. Differences were observed across the four cultures with regard to apoptotic induction by HS and VP16 and to MF apoptotic reduction, with an individual variability with regard to apoptotic sensitivity. CONCLUSION: The ability of static MFs to reduce the extent of damage-induced apoptosis in glioblastoma cells might allow the survival of damaged and possibly mutated cells.     

Genomewide linkage analysis of bipolar disorder by use of a high-density single-nucleotide-polymorphism (SNP) genotyping assay: a comparison with microsatellite marker assays and finding of significant linkage to chromosome 6q22

We performed a linkage analysis on 25 extended multiplex Portuguese families segregating for bipolar disorder, by use of a high-density single-nucleotide-polymorphism (SNP) genotyping assay, the GeneChip Human Mapping 10K Array (HMA10K). Of these families, 12 were used for a direct comparison of the HMA10K with the traditional 10-cM microsatellite marker set and the more dense 4-cM marker set. This comparative analysis indicated the presence of significant linkage peaks in the SNP assay in chromosomal regions characterized by poor coverage and low information content on the microsatellite assays. The HMA10K provided consistently high information and enhanced coverage throughout these regions. Across the entire genome, the HMA10K had an average information content of 0.842 with 0.21-Mb intermarker spacing. In the 12-family set, the HMA10K-based analysis detected two chromosomal regions with genomewide significant linkage on chromosomes 6q22 and 11p11; both regions had failed to meet this strict threshold with the microsatellite assays. The full 25-family collection further strengthened the findings on chromosome 6q22, achieving genomewide significance with a maximum nonparametric linkage (NPL) score of 4.20 and a maximum LOD score of 3.56 at position 125.8 Mb. In addition to this highly significant finding, several other regions of suggestive linkage have also been identified in the 25-family data set, including two regions on chromosome 2 (57 Mb, NPL = 2.98; 145 Mb, NPL = 3.09), as well as regions on chromosomes 4 (91 Mb, NPL = 2.97), 16 (20 Mb, NPL = 2.89), and 20 (60 Mb, NPL = 2.99). We conclude that at least some of the linkage peaks we have identified may have been largely undetected in previous whole-genome scans for bipolar disorder because of insufficient coverage or information content, particularly on chromosomes 6q22 and 11p11.     

Virus maturation targets the protein capsid to concerted disassembly and unfolding

Many animal viruses undergo post-assembly proteolytic cleavage that is required for infectivity. The role of maturation cleavage on Flock House virus was evaluated by comparing wild type (wt) and cleavage-defective mutant (D75N) Flock House virus virus-like particles. A concerted dissociation and unfolding of the mature wt particle was observed under treatment by urea, whereas the cleavage-defective mutant dissociated to folded subunits as determined by steady-state and dynamic fluorescence spectroscopy, circular dichroism, and nuclear magnetic resonance. The folded D75N alpha subunit could reassemble into capsids, whereas the yield of reassembly from unfolded cleaved wt subunits was very low. Overall, our results demonstrate that the maturation/cleavage process targets the particle for an "off pathway" disassembly, because dissociation is coupled to unfolding. The increased motions in the cleaved capsid, revealed by fluorescence and NMR, and the concerted nature of dissociation/unfolding may be crucial to make the mature particle infectious.     

Localization of a novel locus for autosomal recessive early-onset parkinsonism, PARK6, on human chromosome 1p35-p36

The cause of Parkinson disease (PD) is still unknown, but genetic factors have recently been implicated in the etiology of the disease. So far, four loci responsible for autosomal dominant PD have been identified. Autosomal recessive juvenile parkinsonism (ARJP) is a clinically and genetically distinct entity; typical PD features are associated with early onset, sustained response to levodopa, and early occurrence of levodopa-induced dyskinesias, which are often severe. To date, only one ARJP gene, Parkin, has been identified, and multiple mutations have been detected both in families with autosomal recessive parkinsonism and in sporadic cases. The Parkin-associated phenotype is broad, and some cases are indistinguishable from idiopathic PD. In > or = 50% of families with ARJP that have been analyzed, no mutations could be detected in the Parkin gene. We identified a large Sicilian family with four definitely affected members (the Marsala kindred). The phenotype was characterized by early-onset (range 32-48 years) parkinsonism, with slow progression and sustained response to levodopa. Linkage of the disease to the Parkin gene was excluded. A genomewide homozygosity screen was performed in the family. Linkage analysis and haplotype construction allowed identification of a single region of homozygosity shared by all the affected members, spanning 12.5 cM on the short arm of chromosome 1. This region contains a novel locus for autosomal recessive early-onset parkinsonism, PARK6. A maximum LOD score 4.01 at recombination fraction .00 was obtained for marker D1S199.     

In vitro proliferation and in vivo malignancy of cell lines simultaneously derived from a chemically-induced heterogeneous rat mammary tumor

Summary Identification of clones in primary tumors responsible for proliferation, invasion, and metastasis was carried out. Four different aneuploid established cell lines derived from a ductal infiltrating mammary rat tumor induced by 7,12-dimethylbenz[a]anthracene were studied for proliferative and growth features in vitro and for tumorigenic and metastatic potential in vivo in nude mice. Clones, named RM1, RM2, RM3, and RM4, were characterized by different proliferative activity. Clone RM1 showed the highest proliferative activity by both tritiated thymidine incorporation and S-phase flow cytometry, followed by clone RM4. Conversely, clones RM2 and RM3 showed a lower proliferation rate. Growth-promoting activity, tested on 3T3 Swiss cells, was high in all clones, although RM1 showed significantly lower growth factors—releasing activity. Nude mice tumorigenesis demonstrated a strong tumor induction of line RM1 (100% of the mice after 47±7 d) and a slightly lower tumor induction of line RM4 (70% of the mice after 69±9 d). Line RM3 showed tumor induction in 40% of the mice after 186±16 d. Lines RM2 showed no tumor induction. Metastasis occurred in mice treated with line RM1 only. Therefore, tumorigenesis and metastasis correlate with proliferation but not with the release of growth factors. In conclusion, flow cytometry monitoring of clones from heterogeneous primary tumors proved to be a suitable model for the study of in vivo malignancy and in vitro proliferation.     

Multicomponent structure of the thyrotropin receptor: relationship to Graves' disease

The thyrotropin receptor is proposed to contain both a glycoprotein and a ganglioside component. Monoclonal antibodies have been developed against soluble thyroid TSH receptor preparations and using Graves' lymphocytes. These show that initial recognition of thyrotropin requires the glycoprotein component, but that monoclonal antibodies to this component block thyrotropin function (blocking antibodies) rather than mimic thyrotropin. Monoclonal antibodies which stimulate thyroid activity in cultured cell systems (cAMP increase) or mouse bioassays, all interact with gangliosides. Using monoclonal antibodies to the glycoprotein component of the thyrotropin receptor, we show that two protein bands, molecular weights 18,000-23,000 and 50,000-55,000, are precipitated from detergent-solubilized preparations. Using a crosslinking procedure with 125I-labeled thyrotropin, we show that thyrotropin binding is related to the disappearance of the 18,000-23,000 molecular weight band on sodium dodecylsulfate gels and the appearance of a 30,000-33,000 molecular weight thyrotropin-membrane component complex. Higher molecular weight thyrotropin-membrane complexes of 75,000-80,000 and 250,000 are visualized when binding studies are performed at pH 7.4 in physiologic medium; larger amounts of the 30,000-33,000 complex are evident at pH 6.0 in a low salt medium. It is thus proposed that the glycoprotein component of the thyrotropin receptor is composed of two subunits with apparent molecular weights of 18,000-23,000 and 50,000-55,000; that the 18,000-23,000 subunit interacts with thyrotropin; and that different receptor subunits can exist depending on in vitro binding conditions. Using monoclonal-stimulating antibodies or natural autoimmune IgG preparations from patients' sera, we show that stimulating antibodies exhibit species-specific binding to human thyroid ganglioside preparations. Individual components or determinants of the thyrotropin receptor structure with specific autoimmune immunoglobulins.