Fractionation techniques in a hydro-organic environment. II. Acryloyl-morpholine polymers as a matrix for electrophoresis in hydro-organic solvents

The properties of gels prepared either from acryloyl-morpholine (ACM) or from its mixtures with acrylamide and crosslinked either with bisacrylylpiperazine or with methylenebisacrylamide have been described. ACM-containing gels are compatible with organic solvents. If polymerized in water and dried, they are able to reswell, e.g., in dimethyl sulfoxide or dimethylformamide. If polymerized in presence of dimethylformamide, they form perfectly clear gels, whose mechanical properties are by far superior than those of similar plain polyacrylamide formulations.     

Two genetic stocks of Steindachneridion melanodermatum living in sympatry in nature and genetic variability of wild parents and F₁ generation

Steindachneridion melanodermatum is a large Brazilian catfish, highly prized for sport fishing and for its meat. Specimens of this species, both caught in nature from Iguacu River and F(1) fish born in captivity, were analyzed with regard to patterns of RAPD molecular markers. Genetic similarity ranged from 0.57 to 0.95; two groups were determined for the wild specimens. The results suggest different genetic lineages in sympatry in nature. Heterozygosity and percentage of polymorphic loci were 0.31 and 79% and 0.23 and 62%, respectively, for the two populations of wild specimens and 0.26 and 66%, respectively, for those born in captivity.     

Banding pattern of A and B chromosomes of Prochilodus lineatus (Characiformes, Prochilodontidae), with comments on B chromosomes evolution

B chromosomes in Prochilodus lineatus, a migratory neotropical fish, were analyzed in a comparative study among populations from the Dourada lagoon (State of Paraná, Brazil) and from Mogi-Guaçu river (State of São Paulo, Brazil). The data on C-banding and fluorescent in situ hybridization with a satellite DNA probe (SATH1), indicate that the small metacentric B chromosome might correspond to an isochromosome. On the other hand, both populations presented a distinct set of B chromosomes, differentiated either by their number and by the presence of variant B types in the population from Mogi-Guaçu river. The present results indicate that the B chromosomes of P. lineatus should have an ancient origin, and have undergone a differential evolutionary pathway among distinct populations.     

Karyotypic variability in Iheringichthys labrosus (Teleostei, Pimelodidae) from the Tibagi River basin (Paraná State, Brazil)

Cytogenetic analyses were carried out in a populational sample of Iheringichthys labrosus from the Guaraúna River (Upper Tibagi River; Paraná State, Brazil) in order to provide a karyotypic comparison with another previously studied population from the Lower Tibagi River, characterized by the presence of 32m + 8sm + 6st + 10a (2n = 56, FN = 102) and occurrence of supernumerary chromosomes (80% of individuals). The 17 specimens of I. labrosus (6 females, 10 males and 1 of unknown sex) from the Upper Tibagi River showed 2n = 56 chromosomes, a karyotype formula of 14m + 32sm + 4st + 6a (FN = 106), without evidence of sex chromosome heteromorphism or supernumerary chromosomes. The heterochromatin was detected at telomeric and centromeric positions in several chromosomal pairs. The Ag-nucleolar organizer regions were heteromorphic and located at terminal position on short arms of the 16th chromosomal pair, suggesting a positive association with heterochromatic regions. The inter-populational karyotypic differentiation reported indicates distinct evolutionary pathways within I. labrosus in the Tibagi River basin.     

Fractionation techniques in a hydro-organic environment. I. Sulfolane as a solvent for hydrophobic proteins

Sulfolane (thiophene, tethrahydro-1,1-dioxide), at concentrations of 4 M or above, is an efficient solubilizing agent for water-insoluble proteins (e.g., zein or globin chains). In comparison with urea, it appears indefinitely stable in aqueous solutions and does not chemically modify proteins upon storage. Moreover, it favors protein structure, i.e., it increases their alpha-helix content, while urea decreases it. Sulfolane is compatible with electrophoretic techniques (it only slightly reduces polyacrylamide polymerization efficiency and it does not interfere with protein and peptide detection methods) and with chromatographic methods (it has negligible A280 nm). With hydrophilic proteins, sulfolane behaves as a mild denaturant and precipitates them at concentrations between 5 and 7 M.     

A polymorphism in the human serotonin 5-HT2A receptor gene may protect against systemic sclerosis by reducing platelet aggregation

Introduction

Platelet aggregation may contribute to the pathogenesis of systemic sclerosis: following activation, platelets release significant amounts of serotonin – which promotes vasoconstriction and fibrosis, and further enhances aggregation. The C+1354T polymorphism in the exonic region of the serotonin 2A receptor gene determining the His452Tyr substitution was associated with blunted intracellular responses after serotonin stimulation, and may have a role in susceptibility to scleroderma.

Methods

One hundred and fifteen consecutive systemic sclerosis patients and 140 well-matched healthy control individuals were genotyped by sequence-specific primer-PCR for the His⁴⁵²Tyr substitution of the serotonin 2A receptor gene, and associations were sought with scleroderma and its main clinical features. The functional relevance of the His⁴⁵²Tyr substitution was also assessed by evaluating the aggregation of platelet-rich plasma from His⁴⁵²/His⁴⁵² and His⁴⁵²/Tyr⁴⁵² healthy individuals after stimulation with adenosine diphosphate ± serotonin.

Results

The T allele of the C+1354T polymorphism was underrepresented in scleroderma patients compared with control individuals (5.2% versus 12.4%, P < 0.001, chi-square test and 1,000-fold permutation test) and its carriage reduced the risk for systemic sclerosis (odds ratio = 0.39, 95% confidence interval = 0.19 to 0.85, P < 0.01). Platelets from His⁴⁵²/Tyr⁴⁵² healthy subjects more weakly responded to serotonin stimulation compared with platelets from His⁴⁵²/His⁴⁵² individuals (3.2 ± 2.6-fold versus 9.6 ± 8.6-fold increase in aggregation, P = 0.017 by Kolmogorov–Smirnov test and P = 0.003 after correction for baseline adenosine diphosphate-induced aggregation values).

Conclusion

The His⁴⁵²Tyr substitution may influence susceptibility to systemic sclerosis by altering platelet aggregation in response to serotonin.     

Histopathological effects of cisplatin,doxorubicin and 5-flurouracil (5-FU) on the liver of male albino rats

Cisplatin, doxorubicin and fluorouracil (5-FU), drugs belonging to different chemical classes, have been extensively used for chemotherapy of various cancers. Despite extensive investigations into their hepatotoxicity, there is very limited information on their effects on the structure and ultra-structure of liver cells in vivo. Here, we demonstrate for the first time, the effects of these three anticancer drugs on rat liver toxicity using both light and electron microscopy. Light microscopic observations revealed that higher doses of cisplatin and doxorubicin caused massive hepatotoxicity compared to 5-FU treatment, including dissolution of hepatic cords, focal inflammation and necrotic tissues. Interestingly, low doses also exhibited abnormal changes, including periportal fibrosis, degeneration of hepatic cords and increased apoptosis. These changes were confirmed at ultrastructural level, including vesiculated rough endoplasmic reticulum and atrophied mitochondria with ill-differentiated cisternae, dense collection of macrophages and lymphocytes as well as fibrocytes with collagenous fibrils manifesting early sign of fibrosis, especially in response to cisplatin and doxorubicin -treatment. Our results provide in vivo evidence, at ultrastructural level, of direct hepatotoxicity caused by cisplatin, doxorubicin and 5-FU at both light and electron microscopi. These results can guide the design of appropriate treatment regimen to reduce the hepatotoxic effects of these anticancer drugs.     

The Ras guanine nucleotide exchange factor RasGRF1 promotes matrix metalloproteinase-3 production in rheumatoid arthritis synovial tissue

Introduction  

Fibroblast-like synoviocytes (FLS) from rheumatoid arthritis (RA) patients share many similarities with transformed cancer cells, including spontaneous production of matrix metalloproteinases (MMPs). Altered or chronic activation of proto-oncogenic Ras family GTPases is thought to contribute to inflammation and joint destruction in RA, and abrogation of Ras family signaling is therapeutic in animal models of RA. Recently, expression and post-translational modification of Ras guanine nucleotide releasing factor 1 (RasGRF1) was found to contribute to spontaneous MMP production in melanoma cancer cells. Here, we examine the potential relationship between RasGRF1 expression and MMP production in RA, reactive arthritis, and inflammatory osteoarthritis synovial tissue and FLS.     

Differentiation of repetitive DNA sites and sex chromosome systems reveal closely related group in Parodontidae (Actinopterygii: Characiformes)

Parodon and Apareiodon lack sufficiently consistent morphological traits to be considered a monophyletic group in Parodontidae. Species within this family are either sex-homomorphic or sex-heteromorphic (i.e., lacking a differentiated sex chromosome system, ZZ/ZW or ZZ/ZW(1)W(2)). In this study, a DNA fragment from the heterochromatin segment of the W chromosome of Apareiodon ibitiensis (named WAp) was microdissected and used for in situ mapping of nine Parodontidae species. The species were also characterized using a satellite DNA probe (pPh2004). The species were phylogenetically clustered according to 17 characters, which were examined by both classical and molecular cytogenetic techniques. Given the present results, the single ZZ/ZW sex chromosome system seems to have been derived from a paracentric inversion of a terminal WAp site onto the proximal regions of the short arms of a metacentric chromosome pair, followed by WAp site amplification. We reason that these events restrained recombination and favored differentiation of the W chromosome in some species. Moreover, co-hybridization experiments targeting the WAp and pPh2004 repetitive DNA sites of A. affinis suggest that the ZZ/ZW(1)W(2) sex chromosomes of this species may have arisen from a translocation between the proto-sex chromosome and an autosome. Our phylogenetic analysis corroborates the hypothesis of sex chromosome differentiation and establishes groups of closely related species. The phylogenetic reorganization in response to these new data supports the presence of internal monophyletic groups within Parodontidae.