EBV persistence involves strict selection of latently infected B cells

EBV is found preferentially in IgD- B cells in the peripheral blood. This has led to the proposal that the recirculating memory B cell pool is the site of long-lived persistent infection. In this paper we have used CD27, a newly identified specific marker for memory B cells, to test this hypothesis. We show that EBV is tightly restricted in its expression. Less than 1 in 1000 of the infected cells in the peripheral blood are naive (IgD+, CD27-) and <1 in 250 are IgD+ memory cells. Furthermore, EBV was undetectable in the self-renewing peripheral CD5+ or B1 cells, a subset that has not been through a germinal center. No such restriction was observed in tonsillar B cells. Therefore, the virus has access to a range of B cell subsets in the lymph nodes but is tightly restricted to a specific long-lived compartment of B cells, the IgD-, CD27+, and CD5- memory B cells, in the periphery. We suggest that access to this compartment is essential to allow the growth-promoting latent genes to be switched off to create a site of persistent infection that is neither pathogenic nor a target for immunosurveillance.     

Oocyte growth and development in teleosts

Oocyte growth and development is an important issue in fish and fisheries biology. This paper reviews the information available on oocyte growth patterns and the rates and dynamics of oocyte growth in teleosts. In synchronous spawners, the weight of the gonad may represent as much as 40% of the overall body weight of the fish. In asynchronous spawners, the weight of the mature ovary is considerably less than in synchronous ovulators, but the ovary shows a more regular periodicity and may grow repeatedly many times during the breeding season. There is a huge variability in egg size in teleosts, with the largest known measuring up to 8 cm in diameter. Within the limits of variance set by genetic constraints, egg size may vary between populations of the same species. Oocytes in all teleosts undergo the same basic pattern of growth: oogenesis, primary oocyte growth, cortical alveolus stage, vitellogenesis, maturation and ovulation. The mechanisms that control oocyte growth are addressed in this review, albeit that the available information, as in all other vertebrates, is very limited. The main hormones that have been shown to affect ovarian growth are gonadotrophin, thyroid hormones, growth hormone, insulin and insulin-like growth factors. An overview of the determinants of fecundity, with particular reference to oocyte recruitment and atresia, is the focus of the second part of the paper. Genetics and nutrition have major effects on fecundity, and studies so far suggest that the determinants of fecundity usually operate during the early part of gametogenesis. The role of atresia in determining fecundity is less clear. The final part of this review highlights some areas of study that are priorities for research on ovarian development in fish.     

Vascular development: from precursor cells to branched arterial and venous networks

The adult vascular system is composed of an arterial, a venous and a lymphatic compartment. These different compartments respectively provide oxygen and nutrients to peripheral organs, remove carbon dioxide and waste products and maintain an immune barrier to defend the host against foreign organisms. Malfunctions of the vascular system represent a major cause of mortality and disease in developed countries. Understanding of the molecular mechanisms regulating vascular system development and maintenance is thus crucial for the design of therapies to cure vascular diseases. The molecules implicated in the control of physiological and pathological angiogenesis in the adult already function during embryonic development. Indeed, the survival of the embryo also critically depends on the establishment of a functional circulatory loop. Here we review our current knowledge about the emergence of endothelial precursor cells in the embryo, of their assembly into the primary vascular plexus and of the remodeling of this plexus into arteries and veins. We also focus on the molecular mechanisms controlling the development of arteries, veins and lymphatic vessels.     

Distribution of interstitial stem cells in Hydra

The distribution of interstitial stem cells along the Hydra body column was determined using a simplified cloning assay. The assay measures stem cells as clone-forming units (CFU) in aggregates of nitrogen mustard inactivated Hydra tissue. The concentration of stem cells in the gastric region was uniform at about 0.02 CFU/epithelial cell. In both the hypostome and basal disk the concentration was 20-fold lower. A decrease in the ratio of stem cells to committed nerve and nematocyte precursors was correlated with the decrease in stem cell concentration in both hypostome and basal disk. The ratio of stem cells to committed precursors is a sensitive indicator of the rate of self-renewal in the stem cell population. From the ratio it can be estimated that <10% of stem cells self-renew in the hypostome and basal disk compared to 60% in the gastric region. Thus, the results provide an explanation for the observed depletion of stem cells in these regions. The results also suggest that differentiation and self-renewal compete for the same stem cell population.     

Mouse oocytes derived from fetal germ cells are competent to support the development of embryos by in vitro fertilization

Figure 1B in the paper above appeared to be an enlargement of a portion of Figure 1A, while the legend indicated otherwise. This error had been made because the authors labeled the sections by date, but not by source, and because experimental and control sections were similar. The authors have now confirmed their overall results and have submitted new figures for Figure 1 and Figure 4A presented below. The original article to which this erratum refers, was published in Molecular Reproduction and Development 2006;73(10):1312–1317. Mol. Reprod. Dev. 75: 1688–1689, 2008. © 2008 Wiley‐Liss, Inc.     

The analysis of chromatin organisation allows selection of mouse antral oocytes competent for development to blastocyst

Mouse antral oocytes can be classified in two different types termed SN or NSN oocytes, depending on the presence or absence, respectively, of a ring of Hoechst 33342-positive chromatin surrounding the nucleolus. The aim of the present study was to test the developmental competence to blastocyst of the two types of oocytes. Here we show that following isolation, classification and culture of cumulus-free antral oocytes, 14.7% and 74.5% of NSN and SN oocytes, respectively, reached the metaphase II stage. When fertilised and further cultured none of the metaphase II NSN oocytes developed beyond the 2-cell stage whilst 47.4% of the metaphase II SN oocytes reached the 4-cell stage and 18.4% developed to blastocyst. The findings reported in this paper may contribute to improved procedures of female gamete selection for in vitro fertilisation of humans and farm animals. Furthermore, the selection of oocytes with better developmental potential may be of interest for studies on nuclear/cytoplasm interaction, particularly in nuclear-transfer experiments.     

Cell sorting during the regeneration of Hydra from reaggregated cells.

The role of cell sorting in the reorganization of Hydra cell reaggregates was studied. We quantitatively labeled ectodermal and endodermal cells by incubating whole animals in fluorescent beads or by injecting the beads into the gastric cavity. Beads were stably incorporated into the cells by phagocytosis. Our data show that dramatic cell sorting processes drive the formation of ectoderm and endoderm within the first 12 hr of reaggregation. After the ectoderm is established, no further rearrangement could be observed. We also tested the ability of cells to sort out with respect to their original position in Hydra by dissociating labeled apical and basal pieces of Hydra and measuring the clumping of labeled cells during reorganization. There was no increase in the clumping of cells during reorganization indicating that cell sorting is not involved in the formation of early activation centers. There was also no preferential incorporation of apically derived (presumptive head) tissue into tentacles that subsequently formed, indicating that after dissociation into single cells there is no predisposition of erstwhile presumptive head tissue to form heads.     

Germline stem cells and sex determination in Hydra

The sex of germline stem cells (GSCs) in Hydra is determined in a cell-autonomous manner. In gonochoristic species like Hydra magnipapillata or H. oligactis, where the sexes are separate, male polyps have sperm-restricted stem cells (SpSCs), while females have egg-restricted stem cells (EgSCs). These GSCs self-renew in a polyp, and are usually transmitted to a new bud from a parental polyp during asexual reproduction. But if these GSCs are lost during subsequent budding or regeneration events, new ones are generated from multipotent stem cells (MPSCs). MPSCs are the somatic stem cells in Hydra that ordinarily differentiate into nerve cells, nematocytes (stinging cells in cnidarians), and gland cells. By means of such a backup system, sexual reproduction is guaranteed for every polyp. Interestingly, Hydra polyps occasionally undergo sex-reversal. This implies that each polyp can produce either type of GSCs, i.e. Hydra are genetically hermaphroditic. Nevertheless a polyp possesses only one type of GSCs at a time. We propose a plausible model for sex-reversal in Hydra. We also discuss so-called germline specific genes, which are expressed in both GSCs and MPSCs, and some future plans to investigate Hydra GSCs.     

Migration of multipotent interstitial stem cells in Hydra

Stem cells in Hydra represent one of the phylogenetically most ancient stem cell systems and, therefore, provide information for reconstructing the early history of stem cell control mechanisms. Hydra's interstitial stem cells are multipotent and differentiate into both somatic cell types and germ line cells. Although it is well accepted that cells of the interstitial cell lineage are migratory, the in vivo migratory potential of multipotent interstitial stem cells has never been explored. Combining in vivo tracing of genetically labeled interstitial stem cells and tissue transplantation, we show that in contrast to precursor cells, multipotent interstitial stem cells are stationary. Only when exposed to tissue depleted of the interstitial cell lineage, interstitial stem cells start to migrate and to repopulate emptied stem cell niches. We conclude that multipotent interstitial stem cells in Hydra are static and that microenvironmental cues including signals derived from the interstitial cell lineage or from niche cells can trigger a shift in collective stem cell behavior to start migration.     

Isolation of biogenetically competent mitochondria from mammalian tissues and cultured cells

This article describes a quick basic method adapted for the purification of mammalian mitochondria from different sources. The organelles obtained using this protocol are suitable for the investigation of biogenetic activities such as enzyme activity, mtDNA, mtRNA, mitochondrial protein synthesis, and mitochondrial tRNA aminoacylation. In addition, these mitochondria are capable of efficient protein import and the investigation of mtDNA/protein interactions by DNA footprinting is also possible.     

Channel expression correlates with differentiation stage during the development of oligodendrocytes from their precursor cells in culture

Membrane currents in cultured murine oligodendrocytes and their precursors were characterized using the patch-clamp technique. Prior to recording, cells were identified by immunofluorescence using monoclonal antibodies characteristic of two types of precursor cells and two differentiation stages of oligodendrocytes. The most immature, A2B5 antigen-positive glial precursors, expressed four types of voltage-activated K+ currents and tetrodotoxin-sensitive Na+ currents. The more differentiated cells, O4 antigen-positive glial precursors, expressed similar K+ currents, but Na+ currents were recorded in only a minority of cells. In differentiated O1 and O10 antigen-positive oligodendrocytes the channels characteristic of precursor cells were no longer observed, but an inwardly rectifying K+ current was apparent. Thus, channel expression by cells of the oligodendrocyte lineage correlates with differentiation stage and is more complex in precursor cells than in oligodendrocytes.     

The development of cnidarian stinging cells: maturation and migration of stenoteles of Hydra vulgaris

The late stages of stenotele development and the migration and installation of freshly matured stenoteles in Hydra have been studied by kinetic and immunofluorescence investigations with rhodamine-labelled polyps. It was found that the high concentration of osmotic pressure-generating poly(-glutamic acid)s is synthesised exclusively within the lumen of the immature nematocyst. Assembly of the polymers, which is completed after approximately 0.5 days, is accompanied by a swelling of the capsule and ends when the cyst is mature. Active migration of the stenoteles to the tentacles begins only about 1.0 day later, and the time required for installation of a stenotele on the outer surface of the tissue amounts to about another 1.5 days. Furthermore, the results obtained suggest that the disintegration of the clusters of growing stenoteles, which begins 0.5 to 1.0 days before maturation, is a passive process; the ability of a nematocyte to migrate actively to the tentacles is acquired after maturation and might be controlled directly by regulating factors contained in the tissue.Abbreviations BrdU 5-bromo-2-deoxy-uridine - BSA bovine serum albumine - DIC differential-interference contrast - DTE 1,4-dithioerythritol - FITC fluoresceinisothiocyanate - MBS m-maleimidobenzoyl-n-hydroxysuccinimide ester - PBS phosphate-buffered saline - PEG poly(ethylene glycol) - p/GA poly(a/y-glutamic acid) - TRITC tetramethyl-rhodamine6-isothiocyanate - TROMI tetramethyl-rhodamine-5/6-maleimide     

The ultrastructure of the zymogen cells in Hydra viridis

Summary The zymogenic secretory cells of Hydra viridis are scattered between the digestive muscle cells of the gastric region. The mature zymogenic cells are located along the apical surface of the gastrodermal epithelium and contain numerous spherical secretory droplets. They appear to differentiate from stem cells located near the mesoglea. These stem cells resemble epidermal interstitial cells and are filled with free ribosomes. They differ from the interstitial cell in that they usually possess a small amount of granular endoplasmic reticulum. During the process of differentiation they elaborate a highly organized system of granular endoplasmic reticulum. This system becomes dispersed into vesicles as the secretory product is synthesized. There is no indication that the Golgi apparatus participates directly in the formation of the secretory droplets, and there is no indication of a membrane bounding the mature secretory droplet.The fate of the zymogenic cell following the discharge of its secretory product was not determined. It is possible that these cells revert back to a stage resembling the stem cell before resynthesizing a new supply of secretion. In this case the normal secretory process would be very similar to the events described in the dedifferentiation of the zymogenic cells during regeneration.This work was supported by Grant number GB-3262 from the National science Foundation.