Glycan-binding profile of DC-like cells

Modification of vaccine carriers by decoration with glycans can enhance binding to and even targeting of dendritic cells (DCs), thus augmenting vaccine efficacy. To find a specific glycan-“vector” it is necessary to know glycan-binding profile of DCs. This task is not trivial; the small number of circulating blood DCs available for isolation hinders screening and therefore advancement of the profiling. It would be more convenient to employ long-term cell cultures or even primary DCs from murine blood. We therefore examined whether THP-1 (human monocyte cell line) and DC2.4 (immature murine DC-like cell line) could serve as a model for human DCs. These cells were probed with a set of glycans previously identified as binding to circulating human CD14low/-CD16+CD83+ DCs. In addition, we tested a subpopulation of murine CD14low/-CD80+СD11c+CD16+ cells reported as relating to the human CD14low/-CD16+CD83+ cells. Manα1–3(Manα1–6)Manβ1–4GlcNAcβ1–4GlcNAcβ bound to both the cell lines and the murine CD14low/-CD80+СD11c+CD16+ cells. Primary cells, but not the cell cultures, were capable of binding GalNAcα1–3Galβ (Adi), the most potent ligand for binding to human circulating DCs. In conclusion, not one of the studied cell lines proved an adequate model for DCs processes involving lectin binding. Although the glycan-binding profile of BYRB-Rb (8.17)1Iem mouse DCs could prove useful for assessing human DCs, important glycan interactions were missing, a situation which was aggravated when employing cells from the BALB/c strain. Accordingly, one must treat results from murine work with caution when seeking vaccine targeting of human DCs, and certainly should avoid cell lines such as THP-1 and DC2.4 cells.     

An unusual presentation of a malignant jejunal tumor and a different management strategy

BACKGROUND: Malignant small bowel tumors are very rare and leiomyosarcoma accounts for less than 15% of the cases. Management of these tumors is challenging in view of nonspecific symptoms, unusual presentation and high incidence of metastasis. In this case report, an unusual presentation of jejunal sarcoma and management of liver metastasis with radiofrequency ablation (RFA) is discussed. CASE PRESENTATION: A 45-year-old male presented with anemia and features of small bowel obstruction. Operative findings revealed a mass lesion in jejunum with intussusception of proximal loop. Resection of bowel mass was performed. Histopathological findings were suggestive of leiomyosarcoma. After 3-years of follow-up, the patient developed recurrence in infracolic omentum and a liver metastasis. The omental mass was resected and liver lesion was managed with radiofrequency ablation. CONCLUSION: Jejunal leiomyosarcoma is a rare variety of malignant small bowel tumor and a clinical presentation with intussusception is unusual. We suggest that an aggressive management approach using a combination of surgery and a newer technique like RFA can be attempted in patients with limited metastatic spread to liver to prolong the long-term survival in a subset of patients.     

Species loss of stoneflies (Plecoptera) in the Czech Republic during the 20th century

1. Rapid expansion and intensification of anthropogenic activities in the 20th century has caused profound changes in freshwater assemblages. Unfortunately, knowledge of the extent and causes of species loss (SL) is limited due to the lack of reliable historical data. An unusual data set allows us to compare changes in the most sensitive of aquatic insect orders, the Plecoptera, at some 170 locations in the Czech Republic between two time periods, 1955–1960 and 2006–2010. Historical data (1890–1911) on assemblages of six lowland rivers allow us to infer even earlier changes. 2. Regional stonefly diversity decreased in the first half of the 20th century. Streams at lower altitudes lost a substantial number of species, which were never recovered. In the second half of the century, large‐scale anthropogenic pressure caused SL in all habitats, leading to a dissimilarity of contemporary and previous assemblages. The greatest changes were found at sites affected by organic pollution and a mixture of organic pollution and channelisation or impoundment. Colonisation of new habitats was observed in only three of the 80 species evaluated. 3. Species of moderate habitat specialisation and tolerance to organic pollution were most likely to be lost. Those with narrow specialisations in protected habitats were present in both historical and contemporary collections. 4. Contemporary assemblages are the consequence of more than a 100 years of anthropogenic impacts. In particular, streams at lower altitude and draining intensively exploited landscapes host a mere fragment of the original species complement. Most stonefly species are less frequently present than before, although their assemblages remain almost intact in near‐natural mountain streams. Our analyses demonstrate dramatic restriction of species ranges and, in some cases, apparent changes in altitudinal preference throughout the area.     

Endogenous fragment of hemoglobin, neokyotorphin, as cell growth factor

It is shown that neokyotorphin (the -globin fragment 137–141) stimulates proliferation of normal cells (murine embryonic fibroblasts, red bone marrow and spleen cells) and tumor cells (murine melanoma and transformed fibroblasts L929) in the absence or in the presence of fetal bovine serum. In contrast to serum deprivation conditions, the ability to potentiate L929 cell growth in the presence of fetal serum is strongly cell density dependent. The peptide also enhances the viability of L929 cells, murine embryonic fibroblasts and of the primary cultures of murine red bone marrow cells and splenocytes under serum-deprivation conditions for at least 72 h. The results of flow cytometry analysis suggest that the effect of neokyotorphin on survival of L929 cells in serum-free culture medium is due to maintenance of cell proliferation in the absence of growth factors. Along with cell cycle progression the peptide induces reversible reduction of L929 cell size.     

Moderate alcohol consumption is associated with better endothelial function: a cross sectional study

Background

Natural heterologous valved conduits with a diameter greater than 22 mm that can be used for right ventricular outflow tract reconstruction in adults are not commercially available. The purpose of this study was to measure by ultrasonography the maximum diameter of the distended jugular veins of horses and cattle, respectively, to identify a population of animals that would be suitable for post-mortem collection of jugular veins at sizes greater than 22 mm.

Methods

The study population included 60 Warmblood horses, 25 Freiberger horses, 20 Brown Swiss cows, and 20 Holstein cows (including 10 Holstein and 10 Red Holstein). The maximum cross-sectional diameter of the distended jugular veins was measured at a location half-way between the mandibular angle and the thoracic inlet. The thoracic circumference (heart girth length) was used as a surrogate of body size. The jugular vein diameters of the different populations were compared by analysis of variance and the association between heart girth length and jugular vein diameter was determined in each of the four study populations by linear regression analysis.

Results

There was considerable individual variation of jugular vein diameters within each of the four study populations. There was no statistically significant relationship between thoracic circumference and jugular vein diameter in any of the populations. The jugular vein diameters of Brown Swiss cows were significantly larger than those of any of the other populations. Warmblood horses had significantly larger jugular vein diameters compared to Freiberger horses.

Conclusion

The results of this study suggest that the production of bovine or equine xenografts with diameters of greater than 22 mm would be feasible. Differences between species and breeds need to be considered. However, prediction of the jugular vein diameter based on breed and heart girth length in an individual animal is inaccurate.     

Intragenic duplication and divergence in the spectrin superfamily of proteins

Many structural, signaling, and adhesion molecules contain tandemly repeated amino acid motifs. The alpha-actinin/spectrin/dystrophin superfamily of F-actin-crosslinking proteins contains an array of triple alpha-helical motifs (spectrin repeats). We present here the complete sequence of the novel beta-spectrin isoform beta(Heavy)- spectrin (beta H). The sequence of beta H supports the origin of alpha- and beta-spectrins from a common ancestor, and we present a novel model for the origin of the spectrins from a homodimeric actin-crosslinking precursor. The pattern of similarity between the spectrin repeat units indicates that they have evolved by a series of nested, nonuniform duplications. Furthermore, the spectrins and dystrophins clearly have common ancestry, yet the repeat unit is of a different length in each family. Together, these observations suggest a dynamic period of increase in repeat number accompanied by homogenization within each array by concerted evolution. However, today, there is greater similarity of homologous repeats between species than there is across repeats within species, suggesting that concerted evolution ceased some time before the arthropod/vertebrate split. We propose a two-phase model for the evolution of the spectrin repeat arrays in which an initial phase of concerted evolution is subsequently retarded as each new protein becomes constrained to a specific length and the repeats diverge at the DNA level. This evolutionary model has general applicability to the origins of the many other proteins that have tandemly repeated motifs.      

Comparisons of the molecular evolutionary process at rbcL and ndhF in the grass family (Poaceae)

We examine rate heterogeneity among evolutionary lineages of the grass family at two plasmid loci, ndhF and rbcL, and we introduce a method to determine whether patterns of rate heterogeneity are correlated between loci. We show both that rates of synonymous evolution are heterogeneous among grass lineages and that are heterogeneity is correlated between loci at synonymous sites. At nonsynonymous sites, the pattern of rate heterogeneity is not correlated between loci, primarily due to an aberrant pattern of rate heterogeneity at nonsynonymous sites of rbcL. We compare patterns of synonymous rate heterogeneity to predictors based on the generation time effect and the speciation rate hypotheses. Although there is some evidence for generation time effects, neither generation time effects nor speciation rates appear to be sufficient to explain patterns of rate heterogeneity in the grass plastid sequences.      

Expression patterns of germ line specific genes in mouse and human pluripotent stem cells are associated with regulation of ground and primed state of pluripotency

One of the main criteria of pluripotency is ability of cell lines to differentiate into the germ line. Pluripotent stem cell lines in ground state of pluripotency differ from the lines in primed state by their ability to give rise to the mature gametes. To understand molecular mechanisms involved in regulation of different states of pluripotency we investigated the expression patterns of germ line specific genes in different type pluripotent stem cells and mouse and human embryonic teratocarcinoma cells. We found that pluripotent stem cells in vitro, in blastocyst and gonocytes at stage E13.5 had similar expression patterns in contrast to the epiblast cells at stage E6.5. Quantitative real time PCR analysis showed that Vasa/Ddx4 expression in mouse and human embryonic stem cells was significantly lower than in blastocyst and gonocytes. Moreover, Vasa/Ddx4 and E-ras expression was significantly higher in mouse embryonic stem cells than in human embryonic stem cells. Our analysis of germ line specific gene expression in differentiating mouse embryonic stem and embryonic germ cells as well as in mouse embryonic teratocarcinoma cells maintained under conditions promoting cell reprogramming from primed to ground state of pluripotency (2i + LIF) revealed that only pluripotent stem cells are able to regulate the expression level of Oct4 and Vasa/Ddx4 and restore initial ground state, while in embryonic teratocarcinoma cells the expression level of these genes remained unchanged. We suggest that expression patterns of germ lines specific genes, in particular of Vasa/Ddx4, can underlie the regulation of ground and primed states of pluripotency.     

Antiproliferative action of valorphin in cell cultures.

The antiproliferative effects of the haemoglobin beta-chain fragment (33-39) (valorphin or VV-haemorphin-5) were studied in a panel of tumour cell lines and normal cells of different origin, using various methods of activity determination (trypan blue inclusion test, sulphorhodamine B staining, MTT staining, flow cytometry and clonogenic test). Valorphin suppressed the proliferation of tumour cells by 25%-95%, depending on the cell line. The maximal valorphin activity was detected in transformed cells of fibroblastic (L929) and epithelial (MCF-7) origin, transformed haematopoietic cells (K562, HL-60) being less sensitive. In normal cells, valorphin activity was several fold lower (10%-15%). A study of the dynamics of cell proliferation in L929 cells using a visual cell count and flow cytometry showed that valorphin induced reversible and relatively short (24 h) S-phase arrest of cell proliferation, accompanied by a reversible increase of cell size. The proliferation delay was followed by a comparatively long period of reversible resistance of the cells to the peptide (96 h) when the cells are dividing at normal rate. The same dynamics were demonstrated for A549, MCF-7 and primary murine breast carcinoma cells. On the basis of the data obtained, a pattern of regulation of cell growth by valorphin is suggested.     

Construction and Characterization of a Magnaporthe grisea Bacterial Artificial Chromosome Library

Diaz-Perez, S. V., Crouch, V. W., and Orbach, M. J. 1996. Construction and characterization of a Magnaporthe grisea bacterial artificial chromosome library. Fungal Genet. Biol. 20, 280-288. A bacterial artificial chromosome (BAC) library of Magnaporthe grisea containing 4128 clones with an average insert size of 66-kb has been constructed. This library represents seven genome equivalents of M. grisea and has been demonstrated to be representative of the genome by screening for the presence of several single-copy genes and DNA markers. The utility of the library for use in map-based cloning projects was shown by the spanning of a nine-cosmid, 207-kb DNA contig with only 3 BAC clones. In addition, using a lys1-3 auxotroph, we have shown that BAC clones at least 113 kb can be transformed into M. grisea to screen for complementation of mutations. Thus, BACs isolated in chromosome walks can be rapidly screened for the presence of the sought after gene. The ease of construction of BAC libraries and of isolation and manipulation of BAC clones makes the BAC system an ideal one for physical analyses of fungal genomes.