Telepathology and continuous education: important tools for pathologists of developing countries

Education shows that active participation allows the best development of skills to acquire, and the results are better when the information is well documented. Now, with digital images and the Internet, in the case of the Static Telepathology (ST), it is easy to share macroscopic and microscopic photographs. The progress of the technologies enabled a form of Dynamic Telepathology (DT) named "virtual slides", with navigation tools, and can be moved around changing powers as desired, making any personal computer into a digital microscope. The use of these tools in continuous education leads to optimal development of knowledge. We reported the experience of a Latin-American Pathologist from La Rioja, a small Province of Argentina, and we also mentioned the electronic publications in Virtual Hispano-American Congresses of Pathology (VHACP) since 1997(18 reports in the case of ST) and in two Virtual Slide Congress (VSC). In the 1st (2005) and 2nd (2007) Internet VSCs two of our cases were digitized in Spain (case 1 and 3 respectively). In these Virtual Slides, the microscopic images can be moved remotely from any computer connected to the Internet, we should recognize that it will become a valuable continuing Medical Education tool in microscopy, probably related to the phrase "a picture is worth more than a thousand words", then we might add; "what about thousands of images?" Similarly, the autoevaluation test is very important. ST and DT, in support of Virtual Congresses allows learning, teaching and sharing of diseases in scientific presentations and the exchange of views in the forums, these are the optimum material for distance education. In addition we received CDs or DVDs and certificates as authors, recognized by European Institutions. The active participation and the autoevaluation test are the best tools for continuous medical education in telepathology, not only for pathologists in developing countries but for the entire world.     

Developmental changes of gut microflora and enzyme activity in rat pups exposed to fat-rich diet

The aim of this study was to investigate the effect of a high-fat (HF)/energy diet on the intestinal microbiota, the alkaline phosphatase (AP) activity, and related parameters of growth and obesity during the suckling and weaning periods in male Sprague-Dawley rats. From birth, nutrition in suckling pups was manipulated by feeding rat dams either HF or a standard diet, and then after weaning, by exposure of experimental pups to the HF, and control rats to normal diet. On days 15, 20, 40 the numbers of 2 microbial groups, i.e., Bacteroides/Prevotella (BAC) and the Lactobacillus/Enterococcus (LAB) in the jejunum, were determined by fluorescent in situ hybridization technique, and the AP activity was assayed histochemically. During all investigated periods HF pups gained body fat more rapidly than control animals, but from weaning they displayed significantly stunted growth resulting in final body weight loss. Obesity in HF rats was also accompanied by higher LAB and lower numbers of BAC and with permanently higher AP activity. Correlation of these data showed significant negative correlation between LAB, AP, and weight gain and energy efficiency, and significant positive correlation of BAC and AP activity with body fat. These data support the concept that postnatal nutritional experience represents an important factor affecting the ontogeny of intestinal microbial communities and intestinal function. These acquired changes could be a component of regulatory mechanisms involved in adverse and/or positive consequences of HF diet for adiposity, body weight, and energy-balance control in later life.     

COPI vesicle transport is a common requirement for tube expansion in Drosophila

Background

Tube expansion defects like stenoses and atresias cause devastating human diseases. Luminal expansion during organogenesis begins to be elucidated in several systems but we still lack a mechanistic view of the process in many organs. The Drosophila tracheal respiratory system provides an amenable model to study tube size regulation. In the trachea, COPII anterograde transport of luminal proteins is required for extracellular matrix assembly and the concurrent tube expansion.

Principal Findings

We identified and analyzed Drosophila COPI retrograde transport mutants with narrow tracheal tubes. γCOP mutants fail to efficiently secrete luminal components and assemble the luminal chitinous matrix during tracheal tube expansion. Likewise, tube extension is defective in salivary glands, where it also coincides with a failure in the luminal deposition and assembly of a distinct, transient intraluminal matrix. Drosophila γCOP colocalizes with cis-Golgi markers and in γCOP mutant embryos the ER and Golgi structures are severely disrupted. Analysis of γCOP and Sar1 double mutants suggests that bidirectional ER-Golgi traffic maintains the ER and Golgi compartments and is required for secretion and assembly of luminal matrixes during tube expansion.

Conclusions/Significance

Our results demonstrate the function of COPI components in organ morphogenesis and highlight the common role of apical secretion and assembly of transient organotypic matrices in tube expansion. Intraluminal matrices have been detected in the notochord of ascidians and zebrafish COPI mutants show defects in notochord expansion. Thus, the programmed deposition and growth of distinct luminal molds may provide distending forces during tube expansion in diverse organs.     

A mitochondrial kinase complex is essential to mediate an ERK1/2-dependent phosphorylation of a key regulatory protein in steroid biosynthesis

ERK1/2 is known to be involved in hormone-stimulated steroid synthesis, but its exact roles and the underlying mechanisms remain elusive. Both ERK1/2 phosphorylation and steroidogenesis may be triggered by cAMP/cAMP-dependent protein kinase (PKA)-dependent and-independent mechanisms; however, ERK1/2 activation by cAMP results in a maximal steroidogenic rate, whereas canonical activation by epidermal growth factor (EGF) does not. We demonstrate herein by Western blot analysis and confocal studies that temporal mitochondrial ERK1/2 activation is obligatory for PKA-mediated steroidogenesis in the Leydig-transformed MA-10 cell line. PKA activity leads to the phosphorylation of a constitutive mitochondrial MEK1/2 pool with a lower effect in cytosolic MEKs, while EGF allows predominant cytosolic MEK activation and nuclear pERK1/2 localization. These results would explain why PKA favors a more durable ERK1/2 activation in mitochondria than does EGF. By means of ex vivo experiments, we showed that mitochondrial maximal steroidogenesis occurred as a result of the mutual action of steroidogenic acute regulatory (StAR) protein -a key regulatory component in steroid biosynthesis-, active ERK1/2 and PKA. Our results indicate that there is an interaction between mitochondrial StAR and ERK1/2, involving a D domain with sequential basic-hydrophobic motifs similar to ERK substrates. As a result of this binding and only in the presence of cholesterol, ERK1/2 phosphorylates StAR at Ser(232). Directed mutagenesis of Ser(232) to a non-phosphorylable amino acid such as Ala (StAR S232A) inhibited in vitro StAR phosphorylation by active ERK1/2. Transient transfection of MA-10 cells with StAR S232A markedly reduced the yield of progesterone production. In summary, here we show that StAR is a novel substrate of ERK1/2, and that mitochondrial ERK1/2 is part of a multimeric protein kinase complex that regulates cholesterol transport. The role of MAPKs in mitochondrial function is underlined.     

BRED: A Simple and Powerful Tool for Constructing Mutant and Recombinant Bacteriophage Genomes

Advances in DNA sequencing technology have facilitated the determination of hundreds of complete genome sequences both for bacteria and their bacteriophages. Some of these bacteria have well-developed and facile genetic systems for constructing mutants to determine gene function, and recombineering is a particularly effective tool. However, generally applicable methods for constructing defined mutants of bacteriophages are poorly developed, in part because of the inability to use selectable markers such as drug resistance genes during viral lytic growth. Here we describe a method for simple and effective directed mutagenesis of bacteriophage genomes using Bacteriophage Recombineering of Electroporated DNA (BRED), in which a highly efficient recombineering system is utilized directly on electroporated phage DNA; no selection is required and mutants can be readily detected by PCR. We describe the use of BRED to construct unmarked gene deletions, in-frame internal deletions, base substitutions, precise gene replacements, and the addition of gene tags.     

Reduction of mineral nutrient availability accelerates flowering of Arabidopsis thaliana

The time of flowering is regulated by various environmental cues, and in some plant species, it is known to be affected by abiotic stresses. We investigated the effect of nutrient stress caused by an abrupt reduction of mineral nutrition on flowering of Arabidopsis thaliana. We used a hydroponic culture system that enabled us to precisely control nutrient levels. When plants were grown in full-strength nutrient solution for several weeks and then transferred to a diluted medium, the time from sowing to bud appearance was significantly shortened. This acceleration of flowering was more pronounced in short days than in long days, and stronger in the ecotype Landsberg erecta than in Columbia and San Feliu-2. The response was also affected by the age of plants at the beginning of nutrient stress and by the concentration of the diluted medium: earlier treatment and more diluted solutions strengthened the effect. Flowering was affected by nutrient stress, not by a change in the osmotic potential of the medium: addition of mannitol to a 1000-fold diluted solution had no effect on the promotion of flowering. When 3-week-old Landsberg erecta plants were exposed to 1000-fold diluted nutrient solution in an 8-h day length, flower bud appearance was strongly and reproducibly advanced by 10.8–12.8 d compared with control plants (which developed buds 41.1–46.2 d after sowing). This treatment can serve as an optimized protocol for future studies concerning physiological, molecular and ecological aspects of flower induction by nutrient stress in A. thaliana.     

Neoparamoeba perurans is a cosmopolitan aetiological agent of amoebic gill disease

Previously we described a new member of the Neoparamoeba genus, N. perurans, and showed that it is an agent of amoebic gill disease (AGD) of Atlantic salmon Salmo salar cultured in southeast Tasmania, Australia. Given the broad distribution of cases of AGD, we were interested in extending our studies to epizootics in farmed fish from other sites around the world. Oligonucleotide probes that hybridise with the 18S rRNA of N. perurans, N. branchiphila or N. pemaquidensis were used to examine archival samples of AGD in Tasmania as well as samples obtained from 4 host fish species cultured across 6 countries. In archival samples, N. perurans was the only detectable amoeba, confirming that it has been the predominant aetiological agent of AGD in Tasmania since epizootics were first reported. N. perurans was also the exclusive agent of AGD in 4 host species across 6 countries. Together, these data show that N. perurans is a cosmopolitan agent of AGD and, therefore, of significance to the global mariculture industry.     

Technical means for objectification of medical treatments in the area of the deep stabilisation spinal system

Information regarding two versions of an instrument called a muscle dynamometer, which enables detailed information about muscle activity in the deep stabilisation spinal system (DSSS), presented in this article. The MD01 (muscle dynamometer ver. 01) is a simple electromechanical instrument that allows measurement of muscle activity in two areas of the lumbar spine region. Measurements on patients have confirmed the usefulness of quantifying the initial state of a patient before rehabilitation as well as monitoring rehabilitation treatment; the MD01 is a suitable device for obtaining these measurements. However, a new and improved version of the MD01, the MD02, has been developed. The MD02 allows measurements in four different body regions and now has a PC interface, which allows achieving of patient information and data export for use with statistical software.