Kinetic properties of the glucose 6-phosphatase of the liver from arthritic rats

According to previous reports, adjuvant-induced arthritic rats present reduced activities of the hepatic glucose 6-phosphatase. A kinetic study was done in order to characterize this phenomenon. Microsomes were isolated from livers of arthritic and control rats (Holtzman strain) and the glucose 6-phosphatase was measured at various temperatures (13-37 degrees C) and glucose 6-phosphate concentrations. Irrespective of the temperature, the enzyme from arthritic rats presented a reduction of both V(max) and K(M). Detergent treatment of liver microsomes from control rats increased the activity, but no increase was found when microsomes from arthritic rats were treated in the same way. The mannose 6-phosphatase activity of detergent-treated microsomes from arthritic rats was only 25% of the activity found with detergent-treated microsomes from control rats. Without detergent treatment, the mannose 6-phosphatase activities of both control and arthritic rats were minimal. The activation energy, derived from V(max), was not changed by arthritis. In vivo arthritic rats presented higher hepatic glucose 6-phosphate concentrations, a phenomenon that is consistent with a reduced activity of glucose 6-phosphatase. It was concluded that in arthritic rats, the hydrolase is probably reduced, without a similar change in the translocase activity.     

Structural features of an arabinogalactan gum exudates from Spondias dulsis (Anacardiaceae)

The tree Spondias dulcis, located in Venezuela, exudes a light-brown gum. The polysaccharide, isolated from the original gum, contains galactose, arabinose, mannose, rhamnose, glucuronic acid, and its 4-O-methyl derivative. Application of chemical methods, in combination with 1D and 2D NMR spectroscopy afforded interesting structural features of the gum polysaccharide. The unequivocal presence of rhamnose in the polymer structure was confirmed by chemical and spectral data [1H (1.03 ppm); 13C (16.92 ppm)]. Also confirmed was the existence of 3-O- and 6-O-substitutes galactose residues by the spectral data correlations observed in Heteronuclear Multiple Quantum Coherence (HMQC) and Heteronuclear Multiple Bond Correlation (HMBC). Also observed were unequivocal resonances for beta-D-glucuronic acid and its 4-O-methyl derivative, and the presence of 3-O-alpha-L-arabinofuranose and 3-O-beta-L-arabinopyranose residues.     

Cytoskeleton of human mononuclear cells as a possible peripheral marker for phenylalanine neurotoxicity in PKU

In this work we tested human mononuclear cells as a peripheral marker to study neurotoxicity of phenylalanine (Phe). Slices of cerebral cortex of rats or human mononuclear cells were incubated with different concentrations of Phe and/or Ala in the presence of ³²P-orthophosphate, the cytoskeletal fraction was extracted, and the radioactivity incorporated into intermediate filament proteins was measured. Our results show that 2 mM Phe as well as 1 mM Ala are effective in increasing the ³²P in vitro incorporation into IFs in both tissues. When cerebral cortex slices or mononuclear cells were incubated with different concentrations of Phe and/or Ala, the effects on the ³²P in vitro incorporation into IF proteins was compatible with an antagonistic mechanism of action of the two amino acids on the enzymes of the phosphorylating system. In addition, these blood cells may be a possible peripheral marker to study neurotoxicity of Phe in patients with PKU.     

Effects of Bucephalus sp. (Trematoda: Bucephalidae) on Perna perna mussels from a culture station in Ratones Grande Island,Brazil

This study reports the prevalence of Bucephalus sp. in Perna perna populations from a culture station of southern Brazil and its effect on the mussel reproductive tissue and immune system. The prevalence of Bucephalus sp. in P. perna (n = 1871) was considered low (3.1%) and did not seasonally vary. Histological sections of the mantle of infected mussels revealed a marked (80%) reduction of the reproductive tissue that was severe even in mussels exhibiting a moderate infection degree. The total (THC) and differential (DHC) hemocyte counts were lower in infected mussels (3.9 x 10(6) hem/ml; granular hemocytes = 33%) as compared with non-infected animals (5.5 x 10(6) hem/ml; granular hemocytes = 40%). The plasma protein concentration did not vary upon infection. Hemocyte infiltration was significantly higher only in mussels with a very heavy infection degree. The parasite sporocysts were never seen encapsulated by the host hemocytes. Our results indicate that Bucephalus sp. promotes a severe castration in its host and apparently evades the mussel immune system.     

Lipopolysaccharide-induced leukocyte lipid body formation in vivo: innate immunity elicited intracellular Loci involved in eicosanoid metabolism

Lipid bodies are rapidly inducible, specialized cytoplasmic domains for eicosanoid-forming enzyme localization, which we hypothesize to have specific roles in enhanced inflammatory mediator production during pathological conditions, including sepsis. However, little is known about the origins, composition, or functions of lipid bodies in vivo. We show that lipid body numbers were increased in leukocytes from septic patients in comparison with healthy subjects. Analogously, the intrathoracic administration of LPS into mice induced a dose- and time-dependent increase in lipid body numbers. Pretreatment with anti-CD14 or anti-CD11b/CD18 mAb drastically inhibited LPS-induced lipid body formation. Moreover, LPS failed to form lipid bodies in C3H/HeJ (TLR4 mutated) mice, demonstrating a requisite role for LPS receptors in lipid body formation. LPS-induced lipid body formation was also inhibited by the platelet-activating factor-receptor antagonists, suggesting a role for endogenous platelet-activating factor. The eicosanoid-forming enzymes, 5-lipoxygenase and cyclooxygenase-2, were immunolocalized within experimentally induced (LPS in mice) or naturally occurring (septic patients) lipid bodies. The proinflammatory cytokine involved in the pathogenesis of sepsis, TNF-alpha, was also shown to colocalize within lipid bodies. Prior stimulation of leukocytes to form lipid bodies enhanced the capacity of leukocytes to produce leukotriene B(4) and PGE(2). In conclusion, our studies indicate that lipid bodies formed after LPS stimulation and sepsis are sites for eicosanoid-forming enzymes and cytokine localization and may develop and function as structurally distinct, intracellular sites for paracrine eicosanoid synthesis during inflammatory conditions.     

AQR1 gene (ORF YNL065w) encodes a plasma membrane transporter of the major facilitator superfamily that confers resistance to short-chain monocarboxylic acids and quinidine in Saccharomyces cerevisiae

We report results on the functional analysis of Saccharomyces cerevisiae ORF YNL065w, predicted to code for a protein belonging to the poorly characterized major facilitator superfamily (MFS) of transporters that are involved in multidrug resistance (MDR). YNL065w is important for a moderate increase of yeast tolerance to ketoconazole and to the cationic dye crystal violet; it protects the cell against short-chain monocarboxylic acids (C(2)-C(6)), but not against highly liposoluble acids such as octanoic acid or the phenoxyacetic-acid herbicides 2,4-D and MCPA; it is also a determinant of resistance to the antiarrhytmic and antimalarial drug quinidine. The encoding ORF was, thus, denominated the AQR1 gene. Results obtained using an AQR1-lacZ fusion indicate that gene expression is very low and it is not stimulated under weak acid stress. The encoded putative transporter was localized in the plasma membrane by fluorescence microscopy observation of the overproduced Aqr1-GFP fusion protein distribution.     

Interleukin–10 promoter haplotypes are differently distributed in the Brazilian versus the Dutch population

The frequency of five different single nucleotide polymorphisms of the promoter interleukin-10 (IL-10) gene (-3575, -2849, 2763, -1082, -819) was compared between two healthy populations, one originating from the Netherlands and one from Rio de Janeiro, Brazil. A total of 321 Caucasian Dutch individuals and 293 Brazilians, grouped as Afro-Brazilians and Euro-Brazilians, were genotyped using PCR-RFLP. The frequencies of the genotypes in the Brazilian population were different (P<0.05) from the frequencies in the Dutch population in all but one (-2763) genotype. The comparison of genotype frequencies between Afro- and Euro-Brazilians did not demonstrate any differences. The haplotype combination of the most-distant three polymorphisms showed strong linkage disequilibrium. All eight possible combinations were observed in Brazilians, but only seven in Dutch Caucasians. The haplotype frequencies were also significantly different between Brazilians when compared with Dutch and also between Euro-Brazilians and Dutch. No differences were observed in haplotype frequencies between Afro-Brazilians and Euro-Brazilians. The -3575T/-2849G/-2763C is more frequent, while the AAA haplotype was much less represented in the Brazilian than in the Dutch population. The haplotype TAC, which was described in African-Americans, was observed only in Brazilians, almost exclusively among those of European origin. The results corroborate the data indicating that the Brazilian population exhibits a genetic admixture of Africans, Europeans, and Amerindians, and the data may serve as a background for clinical and immunological studies involving the IL-10 locus.     

Plant regeneration from callus cultures of <Emphasis Type="Italic">Maclura tinctoria</Emphasis>, an endangered woody species

Summary Some native species produce seeds with a low frequency of germination accompanied with a period of dormancy. These features make it difficult to produce new phenotypes through sexual propagation. Maclura tinctoria has been considered an endangered species due to extensive use of its wood and low frequency of seed germination. The objective of the present study is to establish an in vitro propagation system for this species. Organogenic friable callus formation from nodal segments has been obtained using woody plant medium (WPM) supplemented with 10.74 μM 1-naphthaleneacetic acid (NAA)+4.43 μM 6-benzylaminopurine (BA). Results indicate that the highest frequency of shoot formation is observed when WPM supplemented with 4.03 μM NAA+4.43 BA is used. For root formation, the use of WPM medium (pH adjusted to 7.0) supplemented with 23.62 μM indole-3-butyric acid (IBA) and 4.7gl⁻¹ activated charcoal is recommended. For acelimatization, subjecting rooted plantlets to 70%, 50%, and 30% mesh screen, each successively for a period of 7 d, has resulted in 97% plantlet survival.