Murine abortion is associated with enhanced interleukin-6 levels at the feto-maternal interface

CBA/JXDBA/2J murine abortion is known to be associated with increased local and peripheral Th1-cytokines levels. The role of the pro-inflammatory interleukin-6 (IL-6) in murine abortion remains unclear. In humans, IL-6 was reported to be elevated at the onset of spontaneous abortion. The aim of our study was to evaluate the levels of IL-6 during murine pregnancy in (1) the normal murine pregnancy combination CBA/JXBALB/c and in (2) the CBA/JXDBA/2J abortion prone mating combination. We measured IL-6 serum levels by ELISA and local (placental and decidual) IL-6 levels by flow cytometry and immunohistochemistry. The expression of the IL-6 receptor gp80 was further analyzed. We additionally evaluated the number of mast cells and macrophages at the feto-maternal interface as a putative IL-6 source in reproductive tissues. IL-6 and gp80 were expressed in decidual cells as well as in different trophoblast types. Flow cytometry analysis showed increased numbers of IL-6+ cells in abortion placentas and deciduas compared to control pregnant mice. We observed an elevated number of mast cells and macrophages at the feto-maternal interface from abortion mice in comparison to control mice. Interestingly, we found very high numbers of mast cells, macrophages and IL-6+ cells in resorption tissue compared to control tissues. Flow cytometry studies confirmed that macrophages are being an important source of IL-6 at the feto-maternal interface. The mRNA IL-6 levels were also enhanced in placenta and decidua from mice with high abortion rate compared to normal pregnant mice, as analyzed by RT-PCR. Our results suggest that IL-6 produced not only by immunocompetent cells such as macrophages and mast cells, but also by trophoblasts and decidua cells, is directly involved in the pathology of abortion.     

Post traumatic subcutaneous mycosis due to Fusarium solani

We report the case of a subcutaneous hyalohyphomycosis of a 24-year-old man, a rural worker with an ulcerative lesion in the right leg of approximately one year duration. It was caused by traumatic implantation of a yerba mate branch. The diagnosis was made by direct microscopic examination with 20% potassium hydroxide (KOH) and revealed several septate hyaline hyphae. It was confirmed by culture of several samples where Fusarium solani was isolated. The patient received local and systemic antifungal therapy. Therapeutic response could not be ascertained at any point in the disease.     

Structure of apolipophorin-III in discoidal lipoproteins. Interhelical distances in the lipid-bound state and conformational change upon binding to lipid

The structure of apolipophorin III in the lipid-bound state and the extent of the conformational change that takes place when the five-helix bundle apolipoprotein binds to a lipoprotein lipid surface were investigated by fluorescence resonance energy transfer in discoidal lipoproteins. Four intramolecular interhelical distances between helix pairs 1-4, 2-4, 3-4, and 5-4 were estimated by fluorescence resonance energy transfer in both the lipid-free and the lipid-bound states. Depending on the helices pairs, the intramolecular interhelical distances increased between 15 and > or = 20 A upon binding of the apolipoprotein to lipid, demonstrating for the first time that binding to lipid is accompanied by a major change in interhelical distances. Using discoidal lipoproteins made with a combination of apolipophorin III molecules containing donor and acceptor groups and apolipophorin III molecules containing neither donor nor acceptor groups, it was possible to obtain information about intermolecular interhelical distances between the helix 4 of one apolipoprotein and the helices 1, 2, 3, and 5 of a second apolipoprotein residing in the same discoidal lipoprotein. Altogether, the estimated intermolecular and intramolecular interhelical distances suggest a model in which the apolipoprotein arranges in pairs of antiparallel and fully extended polypeptide chains surrounding the periphery of the bilayer disc.     

Antimicrobial susceptibilities of strains of Nocardia brasiliensis isolated from soil of Tucumán

The activity of antimicrobial agents against soil isolates of N. brasiliensis was studied by determination of the minimum inhibitory concentrations (MICs) and disk diffusion technique, according to the National Committee for Clinical Laboratory Standards (NCCLS). The objectives were: (a) to study the patterns of sensitivity among regional strains of N. brasiliensis isolated from natural sources (soil) of different zones of Tucumán province; (b) to correlate these results with those previously obtained with regional strains of N. brasiliensis isolated from human mycetomas, as a contribution in the evaluation of the importance of the natural reservoir area of the potentially pathogen strains. The results obtained by both methods identified strains of N. brasiliensis from soils with similar patterns of susceptibility to the strains N. brasiliensis isolated from human mycetomas. This showed strains sensitive and resistant to antibiotics. The majority of the isolates of N. brasiliensis from soils showed higher susceptibilities to antibiotics than the strains isolated from human mycetomas. Among the antibiotics studied, cefotaxime, ceftriaxone and gentamicin were the most effective against all the regional strains tested, and these results are correlated with those obtained with regional strains that cause human mycetomas. This revised version was published online in June 2006 with corrections to the Cover Date.     

Synthesis and Phosphorylation of Maize Acidic Ribosomal Proteins : Implications in Translational Regulation

The objective of this research was to determine the role of acidic ribosomal protein (ARP) phosphorylation in translation. Ribosomes (Rbs) from germinated maize (Zea mays L.) axes had four ARP bands within 4.2 to 4.5 isoelectric points when analyzed by isoelectric focusing. Two of these bands disappeared after alkaline phosphatase hydrolysis. During germination a progressive change from nonphosphorylated (0 h) to phosphorylated ARP (24 h) forms was observed in the Rbs; a free cytoplasmic pool of nonphosphorylated ARPs was also identified by immunoblot and isoelectric focusing experiments. De novo ARP synthesis initiated very slowly early in germination, whereas ARP phosphorylation occurred rapidly within this period. ARP-phosphorylated versus ARP-nonphosphorylated Rbs were tested in an in vitro reticulocyte lysate translation system. Greater in vitro mRNA translation rates were demonstrated for the ARP-phosphorylated Rbs than for the non-ARP-phosphorylated ones. Rapamycin application to maize axes strongly inhibited S6 ribosomal protein phosphorylation, but did not interfere with the ARP phosphorylation reaction. We conclude that ARP phosphorylation does not depend on ARP synthesis or on ARP assembly into Rbs. Rather, this process seems to be part of a translational regulation mechanism.     

Effect of the auxin, 2-(2-methyl-4-chloro-phenoxy)propionic acid, on cell cycle regulation in maize embryonic tissues

During seed maturation, cells from embryonic tissues stop division at different phases of the cell cycle. In maize, neither these phases nor the effect of exogenous auxin on them are known. Disinfected whole maize ( Zea mays L. Mexican commercial hybrid H30) seeds or sectioned embryonic axes were incubated in Murashige and Skoog medium, with or without 2-(2-methyl-4-chlorophenoxy)propionic acid (MCPP), a synthetic auxin. For some in vitro experiments, radioactive [³H]-thymidine was also added. After the stated incubation period, meristems of mesocotyl, primary and seminal roots from embryonic axes were dissected, fixed, and analyzed under a microscope. The percentage of mitotic indices was recorded. In the labeling experiments, labeled and non-labeled percentage of mitotic figures (MI %) were determined. It was found that cell division is a programmed event in the meristematic tissues of maize embryonic axes. Populations of cells entering cell division were obseved during the germination process. The mesocotyl was the first tissue to divide, followed by seminal and primary roots.
Meristematic cells from dry embryos are arrested during the G₂ and G₁ phases of the cell cycle. MCPP has a differential effect, stimulating G₂ cells to enter cell division. It is concluded that MCPP might regulate the cell cycle at specific points.     

C4 gene polymorphism in primates: evolution,generation, and Chido and Rodgers antigenicity

Eleven new C4d genomic primate sequences of the fourth complement factor (C4) have been obtained. Seven of them belong to five species not yet explored for this gene: Pan paniscus (pygmy chimpanzee), Cercopithecus aethiops (green monkey), Macaca mulatta (rhesus monkey), Macaca fascicularis (cynomolgus), and Saguinus oedipus (cotton top tamarin). The New World monkeys (tamarins, four individuals) sequenced for C4 have a single C4d sequence only, which shows a B isotypic specificity and a Rodgers 3 (Rg3), Chido 1 (Ch1) antigenicity. Rg3 and Ch1 could thus be the oldest Rg/Ch specificity (at least 50 million years old) and Rg1, Rg2, Ch3, and Ch6 could be more recent human-specific antigens. Mechanisms of C4d polymorphism generation were analyzed by compiling all the presently available sequences. Examples of both point mutations and crossing-over events among C4d primate sequences could be detected. The problem of a possible trans-species inheritance of C4d polymorphism was addressed and two apparently contradicting dendrograms were obtained. One of them, constructed by using both exon and intron sequences, does not support trans-species evolution, but supports the proposed theory of extensive homogenization of the C4 genes occurring within each species, because alleles from each primate species cluster together. Another completely different dendrogram, obtained by using exon sequences only, suggests the existence of trans-species evolution for C4d polymorphism, because alleles belonging to different species cluster together in a way similar to that found for HLA class I or II alleles. However, orangutan sequences group together in both kinds of C4d sequence dendrograms and seem to have arisen from an ancestor different from that of chimpanzee, gorilla and man C4d sequences. Finally, further data have been obtained that support trans-species conservation of A-ness and B-ness and the existence of trans-specifically conserved allelic motifs, both in intronic and exonic sequences.     

The increase in cytosolic levels of protein kinase C precedes the growth factor requirements during the transition stem cells to differentiated progeny in the MCF-7 breast tumor cells.

The expression of regulatory genes within the context of a differentiation program can have profound long-term consequences in tissues with permanent renewing populations. The breast tumor cell line MCF-7 retains in culture some of the characteristics of a unidirectional differentiation pathway. We show that the cytosolic activity of the regulatory enzyme protein kinase C (PKC) precedes and continues the sequence of maturation in pre-differentiated subpopulations derived from a stem cell fraction. However, the activity declines in the most differentiated, post-mitotic fraction. These results indicate that PKC may be considered among the regulatory genes in MCF-7 cells that specify maturation of the stem cell progeny.     

Rapid chemiluminometric determination of gabapentin in pharmaceutical formulations exploiting pulsed‐flow analysis

In this study, a straightforward and automated pulsed flow‐based procedure was developed for the chemiluminometric determination of gabapentin [1‐(aminomethyl)cyclo‐hexaneacetic acid], a new generation antiepileptic drug, in different formulated dosage forms. The software‐controlled time‐based injection method capitalizes on the decrease of the background chemiluminescence (CL) readout of the luminol–hypochlorite reaction in the presence of gabapentin. In short, gabapentin works as a hypochlorite scavenger. The analytical procedure was implemented in a multi‐pumping flow network furnished with a suite of microdispensing solenoid‐actuated pumps. The diaphragm‐type micropumps might be configured to operate as fluid propellers, commutation units and metering injectors. A dynamic linear working range for gabapentin concentrations in the range 60–350 µmol/L was obtained, with an estimated detection limit of 40 µmol/L. The flow analyser handles about 41 injections/h and yields precise results (RSD < 2%). The miniaturized flow analyser thus has potential to be exploited for in‐line monitoring of drug manufacturing within the quality assurance framework of modern pharmaceutical companies. Copyright © 2008 John Wiley & Sons, Ltd.     

Infection with Usutu Virus Induces an Autophagic Response in Mammalian Cells

Usutu virus (USUV) is an African mosquito-borne flavivirus closely related to West Nile virus and Japanese encephalitis virus, which host range includes mainly mosquitoes and birds, although infections in humans have been also documented, thus warning about USUV as a potential health threat. Circulation of USUV in Africa was documented more than 50 years ago, but it was not until the last decade that it emerged in Europe causing episodes of avian mortality and some human severe cases. Since autophagy is a cellular pathway that can play important roles on different aspects of viral infections and pathogenesis, the possible implication of this pathway in USUV infection has been examined using Vero cells and two viral strains of different origin. USUV infection induced the unfolded protein response, revealed by the splicing of Xbp-1 mRNA. Infection with USUV also stimulated the autophagic process, which was demonstrated by an increase in the cytoplasmic aggregation of microtubule-associated protein 1 light chain 3 (LC3), a marker of autophagosome formation. In addition to this, an increase in the lipidated form of LC3, that is associated with autophagosome formation, was noticed following infection. Pharmacological modulation of the autophagic pathway with the inductor of autophagy rapamycin resulted in an increase in virus yield. On the other hand, treatment with 3-methyladenine or wortmannin, two distinct inhibitors of phosphatidylinositol 3-kinases involved in autophagy, resulted in a decrease in virus yield. These results indicate that USUV virus infection upregulates the cellular autophagic pathway and that drugs that target this pathway can modulate the infection of this virus, thus identifying a potential druggable pathway in USUV-infection.