Cutting edge: TGF-beta signaling is required for the in vivo expansion and immunosuppressive capacity of regulatory CD4+CD25+ T cells

Data regarding the role of TGF-beta for the in vivo function of regulatory CD4(+)CD25(+) T cells (Treg) are controversial. A transgenic mouse model with impaired TGF-beta signaling specifically in T cells was used to assess the role of endogenous TGF-beta for the in vivo function of CD4(+)CD25(+) Treg in a murine model of colitis induced by dextran sulfate. Transfer of wild-type, but not transgenic CD4(+)CD25(+) Treg was found to suppress colitis in wild-type mice. In addition, by transferring CFSE-labeled CD4(+)CD25(+) Treg we could demonstrate that endogenous TGF-beta promotes the expansion of CD4(+)CD25(+) Treg in vivo. Transgenic mice themselves developed reduced numbers of peripheral CD4(+)CD25(+) Treg and were more susceptible to the induction of colitis, which could be prevented by the transfer of wild-type Treg. These data indicate that TGF-beta signaling in CD4(+)CD25(+) Treg is required for their in vivo expansion and suppressive capacity.     

Preadolescents’ relationships with pet dogs: Relationship continuity and associations with adjustment

Research on human–animal interaction in children has been studied in isolation rather than integrated with core theories of children’s relationships. This study is one of the first to examine how children’s relationships with pet dogs are related to their human relationships (parent–child attachments, friendships) and to child adjustment, and to include observational assessment of children’s interactions with their pet dog. Children (9 to 11 years old, n = 99) completed questionnaires regarding relationships with pet dogs, parents, and friends. Half the children were observed interacting with their pet dog. Children and teachers reported children’s adjustment. Children who felt closer to their dogs were more securely attached to mothers and fathers and reported more positive qualities and less conflict with friends. Children with more secure attachments to mothers, and greater companionship with dogs, interacted more with their dogs. Parental attachment and friendship quality, but not the pet dog relationship, were related to child adjustment.     

Disappearance of a Novel Protein Component of the 26S Proteasome duringXenopusOocyte Maturation

We have prepared polyclonal antibodies againstXenopus20S proteasomes. The antibodies cross-react with several proteins that are common to 20S and 26S proteasomes and with at least two proteins that are unique to 26S proteasomes. The antibodies were used to analyze changes in the components of proteasomes during oocyte maturation and early development ofXenopus laevis.A novel protein with a molecular weight of 48 kDa, p48, was clearly detected in immature oocytes, but was found at very low levels in mature oocytes and ovulated eggs. p48 was reduced to low levels during oocyte maturation, after maturation-promoting factor was activated. The amount of p48 in eggs remained low during early embryonic development, but increased again after the midblastula transition. These results show that at least one component of 26S proteasomes changes during oocyte maturation and early development and suggest that alterations in proteasome function may be important for the regulation of developmental events, such as the rapid cell cycles, of the early embryo.     

Vascular endothelial growth factor inhibits programmed cell death of endothelial cells induced by clinorotation

The principal aim of this study was to investigate short- and long-term effects of clinorotation on human endothelial cells (EA hy 926 cell line) using a three-dimensional random positioning machine. Moreover, the impact of vascular endothelial growth factor (VEGF) was addressed. Immediately, within one hour and after four and twenty-four hours an increase of apoptotic cells was detected. VEGF significantly inhibited the amount of apoptotic endothelial cells (EC). VEGF reduced the amount of fas-positive EC. Moreover, after 24 hours, proliferating EC grew in form of three-dimensional multicellular spheroids and also as monolayers. The initially formed spheroids (maximum diameter 3 mm) remained stable up to the 15th day of clinorotation. Some spheroids revealed tubular structures. In addition, a clear increase of extracellular matrix proteins such as osteopontin and fibronectin was measured. The three-dimensional clinostat represents an important tool for cell biological experiments. VEGF significantly attenuated the changes of endothelial cells induced by simulated weightlessness in a cell protective manner.     

Phylogeography of island canary (Serinus canaria) populations

Island canaries (Serinus canaria) are characterised as a species living exclusively on North Atlantic islands, mainly on the Azores, Madeira and Canary Islands. Although they are very common in their habitats, their behaviour and breeding system has only recently been studied systematically. To advance the understanding of their ecology and to see if the rather isolated archipelagos are already promoting a genetic differentiation, we investigated their phylogeographic relationship as revealed by mtDNA sequences of the cytochrome b gene and investigated whether this measure corresponds to morphological characteristics within the islands. Genetic distances were very low throughout the distribution range of the species. Although the variation of genetic distances within the population of Pico (Azores) was larger than that on Madeira and Canary Islands, the genetic distances between island populations were very low throughout which prevented a clear phylogeographic differentiation. Moreover, morphological measurements did not reveal a consistent pattern to reliably separate the populations, although the measures of beak length and body weight revealed a clear island-specific differentiation. These data lead to the assumption that the colonisation of the Atlantic islands by the canaries occurred very recently, while there is no persisting gene flow between the populations.     

Knowledge‐guided laboratory evolution of protein thermolability

In rare but nevertheless important cases it is of practical interest to decrease the thermostability of an enzyme, that is, to increase thermolability in a controlled manner. In the present model study, this unconventional goal has been reached by applying directed evolution to the lipase from Pseudomonas aeruginosa (PAL). By utilizing the B‐factor iterative test (B‐FIT), previously developed to increase the thermostability of enzymes, it was possible to reduce the value from 71.6°C in the case of wild type (WT‐PAL) to 35.6°C (best mutant) without affecting the catalytic profile in terms of substrate acceptance or enantioselectivity at room temperature. Accordingly, saturation mutagenesis was performed at sites in PAL, which on the basis of its X‐ray structure, have the lowest B‐factors indicative of high rigidity. Focused mutations were introduced which can be expected to decrease rigidity, the ensuing increased flexibility leading to higher thermolability without changing the actual catalytic profile. Biotechnol. Bioeng. 2009;102: 1712–1717. © 2008 Wiley Periodicals, Inc.     

The placental analogue and the pattern of sexual reproduction in the cheilostome bryozoan Bicellariella ciliata (Gymnolaemata)

Background

Matrotrophy or extraembryonic nutrition ?C transfer of nutrients from mother to embryo during gestation ?C is well known and thoroughly studied among vertebrates, but still poorly understood in invertebrates. The current paper focuses on the anatomy and ultrastructure of the oogenesis and placentotrophy as well as formation of the brood chamber (ovicell) in the cheilostome bryozoan Bicellariella ciliata (Linnaeus, 1758). Our research aimed to combine these aspects of the sexual reproduction into an integral picture, highlighting the role of the primitive placenta-like system in the evolution of bryozoan reproductive patterns.

Results

Follicular and nutrimentary provisioning of the oocyte occur during oogenesis. Small macrolecithal oocytes are produced, and embryos are nourished in the ovicell via a simple placental analogue (embryophore). Every brooding episode is accompanied by the hypertrophy of the embryophore, which collapses after larval release. Nutrients are released and uptaken by exocytosis (embryophore) and endocytosis (embryo). Embryos lack specialized area for nutrient uptake, which occurs through the whole epidermal surface. The volume increase between the ripe oocyte and the larva is ca. 10-fold.

Conclusions

The ovicell is a complex organ (not a special polymorph as often thought) consisting of an ooecium (protective capsule) and an ooecial vesicle (plugging the entrance to the brooding cavity) that develop from the distal and the fertile zooid correspondingly. Combination of macrolecithal oogenesis and extraembryonic nutrition allows attributing B. ciliata to species with reproductive pattern IV. However, since its oocytes are small, this species represents a previously undescribed variant of this pattern, which appears to represent a transitional state from the insipient matrotrophy (with large macrolecithal eggs) to substantial one (with small microlecithal ones). Altogether, our results substantially added and corrected the data obtained by the previous authors, providing a new insight in our understanding of the evolution of matrotrophy in invertebrates.     

Cellular glycosylphosphatidylinositol-specific phospholipase D regulates urokinase receptor shedding and cell surface expression

The glycosylphosphatidylinositol (GPI) - anchored, multifunctional receptor for the serine proteinase, urokinase plasminogen activator (uPAR, CD87), regulates plasminogen activation and cell migration, adhesion, and proliferation. uPAR occurs in functionally distinct, membrane-anchored and soluble isoforms (s-uPAR) in vitro and in vivo. Recent evidence indicates that s-uPAR present in the circulation of cancer patients correlates with tumor malignancy and represents a valuable prognostic marker in certain types of cancer. We have therefore analyzed the mechanism of uPAR shedding in vitro. We present evidence that uPAR is actively released from ovarian cancer cells since the rate of receptor shedding did not correlate with uPAR expression. While s-uPAR was derived from the cell surface, it lacked the hydrophobic portion of the GPI moiety indicating anchor cleavage. We show that uPAR release is catalyzed by cellular GPI-specific phospholipase D (GPI-PLD), an enzyme cleaving the GPI anchor of the receptor. Thus, recombinant GPI-PLD expression increased receptor release up to fourfold. Conversely, a 40% reduction in GPI-PLD activity by GPI-PLD antisense mRNA expression inhibited uPAR release by more than 60%. We found that GPI-PLD also regulated uPAR expression, possibly by releasing a GPI-anchored growth factor. Our data suggest that cellular GPI-PLD might be involved in the generation of circulating prognostic markers in cancer and possibly regulate the function of GPI-anchored proteins by generating functionally distinct, soluble counterparts. J. Cell. Physiol. 180:225–235, 1999. © 1999 Wiley-Liss, Inc.