Inhibition of Melanosome Transfer from Melanocytes to Keratinocytes by Lectins and Neoglycoproteins in an In Vitro Model System

We propose that some of the critical molecules involved in the transfer of melanosomes from melanocytes to keratinocytes include plasma membrane lectins and their glycoconjugates. To investigate this mechanism, co‐cultures of human melanocytes and keratinocytes derived from neonatal foreskins were established. The process of melanosome transfer was assessed by two experimental procedures. The first involved labeling melanocyte cultures with the fluorochrome CFDA. Labeled melanocytes were subsequently co‐cultured with keratinocytes, and the transfer of fluorochrome assessed visually by confocal microscopy and quantitatively by flow cytometry. The second investigative approach involved co‐culturing melanocytes with keratinocytes, and processing the co‐cultures after 3 days for electron microscopy to quantitate the numbers of melanosomes in keratinocytes. Results from these experimental approaches indicate significant transfer of dye or melanosomes from melanocytes to keratinocytes that increased with time of co‐culturing. Using these model systems, we subsequently tested a battery of lectins and neoglycoproteins for their effect in melanosome transfer. Addition of these selected molecules to co‐cultures inhibited transfer of fluorochrome by approximately 15–44% as assessed by flow cytometry, and of melanosomes by 67–93% as assessed by electron microscopy. Therefore, our results suggest the roles of selected lectins and glycoproteins in melanosome transfer to keratinocytes in the skin.     

Tall gut formation in the rat embryo

Summary The formation of the tail portion of the primitive gut was investigated by light and electron microscopy in 10- and 11-day rat embryos. The observations permit the conclusion that the tail gut does not form as a posterior extension of the hindgut but originates from the tail bud mesenchyme by mechanism analogous to the secondary neurulation. It includes cell condensation, aquisition of apicobasal polarity and the radial, rosette-like arrangement around a central cavity. These cells bear the cytological characteristics of both the absorptive epithelial cells and the mesenchymal cells at their apical (adluminal) and abluminal ends respectively.     

Identification of tryptic peptides from large databases using multiplexed tandem mass spectrometry: simulations and experimental results

Multiplexed tandem mass spectrometry (MS/MS) has recently been demonstrated as a means to increase the throughput of peptide identification in liquid chromatography (LC) MS/MS experiments. In this approach, a set of parent species is dissociated simultaneously and measured in a single spectrum (in the same manner that a single parent ion is conventionally studied), providing a gain in sensitivity and throughput proportional to the number of species that can be simultaneously addressed. In the present work, simulations performed using the Caenorhabditis elegans predicted proteins database show that multiplexed MS/MS data allow the identification of tryptic peptides from mixtures of up to ten peptides from a single dataset with only three "y" or "b" fragments per peptide and a mass accuracy of 2.5 to 5 ppm. At this level of database and data complexity, 98% of the 500 peptides considered in the simulation were correctly identified. This compares favorably with the rates obtained for classical MS/MS at more modest mass measurement accuracy. LC multiplexed Fourier transform-ion cyclotron resonance MS/MS data obtained from a 66 kDa protein (bovine serum albumin) tryptic digest sample are presented to illustrate the approach, and confirm that peptides can be effectively identified from the C. elegans database to which the protein sequence had been appended.     

Discovery and SAR study of novel dihydroquinoline-containing glucocorticoid receptor agonists

We have recently reported the discovery of a novel class of glucocorticoid receptor (GR) antagonists, exemplified by 3, containing a 1,2-dihydroquinoline molecular scaffold. Further SAR studies of these antagonists uncovered chemical modifications conveying agonist functional activity to this series. These agonists exhibit good GR binding affinity and are selective against other nuclear hormone receptors.     

True bugs (Hemiptera,Heteroptera) as psyllid predators (Hemiptera,Psylloidea)

Data on natural enemies of psyllids are rare and can usually be found in papers about economically significant species. During an investigation of psyllid fauna in Serbia, natural enemies were investigated, too. True bugs were the most numerous among them. From 28 psyllid species, 21 species of true bugs from families Anthocoridae and Miridae were reared. Seven species of Anthocoridae were identified: Anthocoris amplicollis (Horváth, 1839), Anthocoris confusus Reuter, 1884, Anthocoris nemoralis (Fabricius, 1794), Anthocoris nemorum (Linnaeus, 1761), Orius majusculus Reuter, 1884, Orius minutus (Linnaeus, 1758) and Orius niger Wolff, 1811. The following 14 species of Miridae were identified: Atractotomus mali Meyer-Dür, 1843, Campylomma verbasci (Meyer-Dür, 1843), Deraeocoris flavilinea (A. Costa, 1862), Deraeocoris ruber (Linnaeus, 1758), Deraeocoris lutescens (Schilling, 1836), Heterocordylus genistae (Scopoli, 1763), Hypseloecus visci (Puton, 1888), Malacocoris chlorizans Panzer, 1794, Miris striatus (Linnaeus, 1758), Orthotylus marginalis Reuter, 1884, Psallus assimilis Stichel, 1956, Psallus quercus Kirschbaum, 1856, Psallus flavellus Stichel, 1933 and Pseudoloxops coccinea (Meyer-Dür, 1843). The aim of the research was to provide list of true bugs recorded as predators of psyllids in order to preserve their diversity and significance, especially on cultivated plants.     

Selective microRNA uridylation by Zcchc6 (TUT7) and Zcchc11 (TUT4)

Recent small RNA sequencing data has uncovered 3′ end modification of mature microRNAs (miRNAs). This non-templated nucleotide addition can impact miRNA gene regulatory networks through the control of miRNA stability or by interfering with the repression of target mRNAs. The miRNA modifying enzymes responsible for this regulation remain largely uncharacterized. Here we describe the ability for two related terminal uridyl transferases (TUTases), Zcchc6 (TUT7) and Zcchc11 (TUT4), to 3′ mono-uridylate a specific subset of miRNAs involved in cell differentiation and Homeobox (Hox) gene control. Zcchc6/11 selectively uridylates these miRNAs in vitro, and we biochemically define a bipartite sequence motif that is necessary and sufficient to confer Zcchc6/11 catalyzed uridylation. Depletion of these TUTases in cultured cells causes the selective loss of 3′ mono-uridylation of many of the same miRNAs. Upon TUTase-dependent loss of uridylation, we observe a concomitant increase in non-templated 3′ mono-adenylation. Furthermore, TUTase inhibition in Zebrafish embryos causes developmental defects and aberrant Hox expression. Our results uncover the molecular basis for selective miRNA mono-uridylation by Zcchc6/11, highlight the precise control of different 3′ miRNA modifications in cells and have implications for miRNA and Hox gene regulation during development.     

Microbiological surveillance of the surgical intensive care unit in Zagreb--a pivot for guideline-based therapy of severe sepsis

The aim of this retrospective study was to create guidelines for therapy of severe sepsis in surgical intensive care unit (ICU) for unknown causative agent based on antimicrobial susceptibility of causative bacteria. Seventy-four patients with severe sepsis from surgical ICU in 2003.-2005. were included in study. Their clinical and microbiological data were analyzed from the medical records. Antimicrobial susceptibility of the strains isolated from the blood-culture was tested by disk diffusion method according to CLSI (Clinical Laboratory Standard Institution). APACHE II score was used to predict the severity of illness. Statistical significance difference between results was tested by Mann-Whitney test and chi2 test. Important problem remained type of sepsis: mono-agent sepsis presented less therapeutic problem than sepsis caused with two or more agents (mixed sepsis). Methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa and Acinetobacter baumannii were predominant causative agents in both type of sepsis. There was remarkable increase of A. baumannii prevalence in 2005 compared to 2004 and to 2003. There was also decrease of MRSA prevalence in 2004 and 2005 compared to 2003. P. aeruginosa were the predominant causative agents in 2004. MRSA displayed good susceptibility to vancomycin and linezolide, whereas P. aeruginosa showed excellent susceptibility to ceftazidime and carbapenems. A. baumannii, third predominant causative agent, exhibited excellent susceptibility to ampicillin+ sulbactam and carbapenems. The recommended therapy is empirical and should cover all important pathogens.     

Morphological analyses allow to separate <Emphasis Type="Italic">Branchipus</Emphasis> species (Branchiopoda,Anostraca) from different geographic regions

Daphnia perform diel vertical migration (DVM), a predator-avoidance strategy to migrate towards deeper and colder layers in the water column in the morning and movement to the algae-rich surface layers in the evening. However, individuals performing DVM incur several trade-offs since they might suffer from resource limitation and a slower instantaneous birth rate in deeper depths. DVM patterns may be modified by abiotic factors such as temperature, food concentration, or pH and vary among different Daphnia species and genotypes. Furthermore, Daphnia host a variety of microparasites that might pose an additional factor influencing DVM behaviour. For infected individuals, migration into cooler temperature layers might slow down parasite growth. Moreover, parasites can increase opacity of their hosts. Non-migrating individuals might then be selectively purged from the upper layers by visually hunting predators. With these premises we asked, whether epidemics of the ichthyosporean parasite Caullerya mesnili affect or are affected by the DVM behaviour of Daphnia in Lake Greifensee, Switzerland by analysing the vertical distribution of Daphnia during day and night on two dates. Furthermore, we were interested whether a potential interaction depends on host genotype. We therefore studied the genotypic composition of the integrated population in regular sampling intervals over the course of one year and on a fine-grained vertical resolution during the Caullerya epidemic in late summer. Since Caullerya-infected Daphnia migrated equally well as uninfected ones, the findings of this study suggest that Caullerya epidemics neither affected nor were affected by the DVM behaviour of Daphnia. We observed clonal succession in the lake but could not link this succession to the Caullerya epidemic; all except one of the common multilocus genotypes were under-infected. In addition, outbreak and course of this Caullerya epidemic seemed to rely mainly on environmental cues. Because this first study only provides a snapshot of time, we hope that further studies will be done to verify our results.     

Phylogeography of the Vipera ursinii complex (Viperidae): mitochondrial markers reveal an east–west disjunction in the Palaearctic region

Aim The aim of this study was to elucidate the phylogeographical pattern of taxa composing the Vipera ursinii complex, for which the taxonomic status and the dating of splitting events have been the subject of much debate. The objectives were to delimit potential refugia and to date splitting events in order to suggest a scenario that explains the diversification of this species complex. Location Western Europe to Central Asia. Methods Sequences of the mitochondrial cytochrome b and NADH dehydrogenase subunit 4 (ND4) genes were analysed for 125 individuals from 46 locations throughout the distribution range of the complex. The phylogeographical structure was investigated using Bayesian and maximum likelihood methods. Molecular dating was performed using three calibration points to estimate the timing of diversification. Results Eighty‐nine haplotypes were observed from the concatenation of the two genes. Phylogenetic inferences supported two main groups, referred to in this study as the ‘ursinii clade’ and the ‘renardi clade’, within which several subclades were identified. Samples from Greece (Vipera ursinii graeca) represented the first split within the V. ursinii complex. In addition, three main periods of diversification were revealed, mainly during the Pleistocene (2.4–2.0 Ma, 1.4 Ma and 1.0–0.6 Ma). Main conclusions The present distribution of the V. ursinii complex seems to have been shaped by Quaternary climatic fluctuations, and the Balkan, Caucasus and Carpathian regions are identified in this study as probable refugia. Our results support a south–north pattern of colonization, in contrast to the north–south colonization previously proposed for this complex. The biogeographical history of the V. ursinii complex corroborates other biogeographical studies that have revealed an east–west disjunction (situated near the Black Sea) within a species complex distributed throughout the Palaearctic region.