PAR-3 defines a central subdomain of the cortical actin cap in mouse eggs

AbstractThe evolutionarily conserved partitioning defective (PAR) protein PAR-3 is pivotal for establishing and maintaining cell polarity. During mammalian oocyte maturation, the radially symmetric oocyte is transformed into a highly polarized metaphase II (MII)-arrested egg. We therefore examined several aspects of PAR-3 expression during oocyte maturation. We cloned two novel PAR-3 transcripts from an oocyte library that likely encode proteins of Mr = 73 K and 133 K that are phosphorylated during maturation. PAR-3, which is found throughout the GV-intact oocyte, becomes asymmetrically localized during meiosis. Following germinal vesicle breakdown, PAR-3 surrounds the condensing chromosomes and associates with the meiotic spindles. Prior to emission of the first and second polar bodies, PAR-3 is located within a central subdomain of the polarized actin cap, which overlies the spindle. This cortical PAR-3 localization depends on intact microfilaments. These results suggest a role for PAR-3 in establishing asymmetry in the egg and in defining the future site of polar body emission

The “Soluble” Adenylyl Cyclase in Sperm Mediates Multiple Signaling Events Required for Fertilization

SummaryMammalian fertilization is dependent upon a series of bicarbonate-induced, cAMP-dependent processes sperm undergo as they “capacitate,” i.e., acquire the ability to fertilize eggs. Male mice lacking the bicarbonate- and calcium-responsive soluble adenylyl cyclase (sAC), the predominant source of cAMP in male germ cells, are infertile, as the sperm are immotile. Membrane-permeable cAMP analogs are reported to rescue the motility defect, but we now show that these “rescued” null sperm were not hyperactive, displayed flagellar angulation, and remained unable to fertilize eggs in vitro. These deficits uncover a requirement for sAC during spermatogenesis and/or epididymal maturation and reveal limitations inherent in studying sAC function using knockout mice. To circumvent this restriction, we identified a specific sAC inhibitor that allowed temporal control over sAC activity. This inhibitor revealed that capacitation is defined by separable events: induction of protein tyrosine phosphorylation and motility are sAC dependent while acrosomal exocytosis is not dependent on sAC

On the curvature of vortex moduli spaces

We use algebraic topology to investigate local curvature properties of the moduli spaces of gauged vortices on a closed Riemann surface. After computing the homotopy type of the universal cover of the moduli spaces (which are symmetric powers of the surface), we prove that, for genus g>1, the holomorphic bisectional curvature of the vortex metrics cannot always be nonnegative in the multivortex case, and this property extends to all Kaehler metrics on certain symmetric powers. Our result rules out an established and natural conjecture on the geometry of the moduli spaces.Comment: 25 pages; final version, to appear in Math.

Temporal changes in kin structure through a population cycle in a territorial bird, the red grouse Lagopus lagopus scoticus

Populations of red grouse (Lagopus lagopus scoticus) undergo regular multiannual cycles in abundance. The 'kinship hypothesis' posits that such cycles are caused by changes in kin structure among territorial males producing delayed density-dependent changes in aggressiveness, which in turn influence recruitment and regulate density. The kinship hypothesis makes several specific predictions about the levels of kinship, aggressiveness and recruitment through a population cycle: (i) kin structure will build up during the increase phase of a cycle, but break down prior to peak density; (ii) kin structure influences aggressiveness, such that there will be a negative relationship between kinship and aggressiveness over the years; (iii) as aggressiveness regulates recruitment and density, there will be a negative relationship between aggressiveness in one year and both recruitment and density in the next; (iv) as kin structure influences recruitment via an affect on aggressiveness, there will be a positive relationship between kinship in one year and recruitment the next. Here we test these predictions through the course of an 8-year cycle in a natural population of red grouse in northeast Scotland, using microsatellite DNA markers to resolve changing patterns of kin structure, and supra-orbital comb height of grouse as an index of aggressiveness. Both kin structure and aggressiveness were dynamic through the course of the cycle, and changing patterns were entirely consistent with the expectations of the kinship hypothesis. Results are discussed in relation to potential drivers of population regulation and implications of dynamic kin structure for population genetics.This work was funded by the Natural Environment Research Council, the European Union, University of Aberdeen and the Centre for Ecology & Hydrology.Peer reviewe

Ventilation Prior to Umbilical Cord Clamping Improves Cardiovascular Stability and Oxygenation in Preterm Lambs After Exposure to Intrauterine Inflammation

Background: Delaying umbilical cord clamping until after aeration of the lung (physiological-based cord clamping; PBCC) maintains cardiac output and oxygenation in preterm lambs at birth, however, its efficacy after intrauterine inflammation is not known. Given the high incidence of chorioamnionitis in preterm infants, we investigated whether PBCC conferred any benefits compared to immediate cord clamping (ICC) in preterm lambs exposed antenatally to 7 days of intrauterine inflammation.Methods: Ultrasound guided intraamniotic injection of 20 mg Lipopolysaccharide (from E. coli:055:B5) was administered to pregnant ewes at 0.8 gestation. Seven days later, ewes were anesthetized, preterm fetuses exteriorised via cesarean section, and instrumented for continuous measurement of pulmonary, systemic and cerebral pressures and flows, and systemic, and cerebral oxygenation. Lambs were then randomized to either PBCC, whereupon ventilation was initiated and maintained for 3 min prior to umbilical cord clamping, or ICC where the umbilical cord was cut and ventilation initiated 30 s later. Ventilation was maintained for 30 min.Results: ICC caused a rapid fall in systemic (by 25%) and cerebral (by 11%) oxygen saturation in ICC lambs, concurrent with a rapid increase in carotid arterial pressure and heart rate. The overshoot in carotid arterial pressure was sustained in ICC lambs for the first 20 min of the study. PBCC maintained cardiac output and prevented the fall in cerebral oxygen delivery at birth. PBCC lambs had lower respiratory compliance and higher respiratory requirements throughout the study.Conclusion: PBCC mitigated the adverse effects of ICC on oxygenation and cardiac output, and therefore could be more beneficial in preterm babies exposed to antenatal inflammation as it maintains cardiac output and oxygen delivery. The increased respiratory requirements require further investigation in this sub-group of preterm infants

Revival of the magnetar PSR J1622-4950: observations with MeerKAT, Parkes, XMM-Newton, Swift, Chandra, and NuSTAR

New radio (MeerKAT and Parkes) and X-ray (XMM-Newton, Swift, Chandra, and NuSTAR) observations of PSR J1622-4950 indicate that the magnetar, in a quiescent state since at least early 2015, reactivated between 2017 March 19 and April 5. The radio flux density, while variable, is approximately 100x larger than during its dormant state. The X-ray flux one month after reactivation was at least 800x larger than during quiescence, and has been decaying exponentially on a 111+/-19 day timescale. This high-flux state, together with a radio-derived rotational ephemeris, enabled for the first time the detection of X-ray pulsations for this magnetar. At 5%, the 0.3-6 keV pulsed fraction is comparable to the smallest observed for magnetars. The overall pulsar geometry inferred from polarized radio emission appears to be broadly consistent with that determined 6-8 years earlier. However, rotating vector model fits suggest that we are now seeing radio emission from a different location in the magnetosphere than previously. This indicates a novel way in which radio emission from magnetars can differ from that of ordinary pulsars. The torque on the neutron star is varying rapidly and unsteadily, as is common for magnetars following outburst, having changed by a factor of 7 within six months of reactivation.Comment: Published in ApJ (2018 April 5); 13 pages, 4 figure

Untersuchungen zum Einfluss von thrombozytären Wachstumsfaktoren auf den zellvermittelten Abbau eines nanopartikulären Knochenersatzstoffes auf Hydroxylapatitbasis : eine experimentelle Studie am Miniaturschwein

Ziel der vorliegenden tierexperimentellen Studie am Miniaturschwein war es, den Einfluss von plättchenreichem Plasma (PRP) auf den zellvermittelten Abbau eines nanopartikulären Hydroxylapatits (HA) in der Frühphase der Knochendefektheilung zu untersuchen. Hierzu wurden 26 männliche Miniaturschweine der Rasse Mini-Lewe in drei Versuchsgruppen eingeteilt und jeweils ein standardisierter Knochendefekt in der Intercondylarregion des rechten Femurs angelegt. Die Defekte wurden entweder mit dem Knochenersatzstoff (Gruppe I/PRP-,n = 11) oder dem Knochenersatzstoff kombiniert mit PRP (Gruppe II/PRP+, n = 11) befüllt. In einer Kontrollgruppe (n = 4) blieben die Defekte unbefüllt. Während der Implantationsoperation wurden bei sechs Tieren jeweils 250 ml Vollblut entnommen, aus dem anschließend durch fraktionierte Zentrifugation plättchenreiches Plasma gewonnen wurde. Die enthaltenen Thrombozyten wurden durch den Zusatz von Thrombin und Kalziumglukonat zur Degranulation angeregt, wodurch die enthaltenen Wachstumsfaktoren aus den alpha-Granula freigesetzt wurden. Zu diesen Wachstumsfaktoren gehören Platelet Derived Growth Factor AB und BB (PDGF AB, BB), Transforming Growth Factor ß 1 (TGF beta1), Vascular Endothelial Growth Factor (VEGF) und basic Fibroblast Growth Factor (bFGF). Die Konzentration der genannten Wachstumsfaktoren wurde mit Hilfe der ELISA-Technik bestimmt. Sie lagen zwischen Faktor 1,6 für TGF-beta1 und Faktor 24,4 für bFGF. 20 Tage post operationem fand die Explantation der operierten distalen Femura statt. Zur lichtmikroskopischen Untersuchung fanden die Knochen-Implantat-Proben Eingang in unterschiedliche Techniken der Einbettung (Paraffin-, Kunststoffeinbettungen), Präparation (Paraffinschnitte, Kunststoffschnitte und Schliffpräparationen), Färbung (Toluidinblau, Haematoxylin-Eosin, Safranin) und Histochemie (Enzym-, Immunhistochemie). Darüber hinaus wurden transmissionselektronenmikroskopische und computergestützte histomorphometrische Untersuchungen durchgeführt. Wie die Ergebnisse der Licht- und Transmissionselektronenmikroskopie aufgezeigt haben, erfolgt in den mit Knochenersatzmaterial behandelten Versuchsgruppen, unabhängig von der PRP-Applikation, die HA-Degradation hydrolytisch und Makrophagen-vermittelt. Die Makrophagen-Population wird durch Riesenzellen vom Langhans-Typ repräsentiert. Diese polarisierten Polykaryen adhärieren über ihre apikale Membrandomäne an den Implantatoberflächen. Das subplasmalemmale Zytoplasma ist immunhistochemisch durch Vimentin-Kondensationen gekennzeichnet. Nicht-adhärente, frei im Granulationsgewebe lokalisierte Polykaryen zeigen dagegen ein homogenes Vimentin-Verteilungsmuster im Zytoplasma. Der zelluläre Abbau des HA erfolgt mittels Phagozytose, indem die Polykaryen den "Fremdkörper" mit pseudopodienartigen Zytoplasmaausläufern umschließen und in ihr Zytoplasma inkorporieren. Diese Art der Degradation wird durch den post implantationem stattfindenden Zerfall des Knochenersatzmaterials in zahlreiche kleine Partikel unterstützt. Die hieraus resultierende Vergrößerung der Implantatoberfläche bietet einer Vielzahl von Zellen die Möglichkeit zur Haftung. Die festgestellten Expressionsmuster des CD44- Membranglykoproteins verweisen auf dessen funktionelle Rolle im Rahmen der Fusion mononukleärer Makrophagen zu multinukleären Riesenzellen. Die darüber hinaus beobachtete Umverteilung von CD44 von der apikalen zur basalen Membrandomäne bei Implantatassoziierten Polykaryen ist als transientes Geschehen im Zuge der Adhäsion zu interpretieren. Der hohe Aktivitätsstatus der adhärenten Polykaryen ist immunhistochemisch durch eine intensive Kathepsin K-Expression gekennzeichnet. Die vergleichende histomorphometrische Auswertung der mit HA aufgefüllten Defekte dokumentiert eine Verdopplung der Anzahl von Polykaryen in der Gruppe "Knochenersatzstoff mit PRP". Ein auf Basis der Messergebnisse durchgeführter Wilcoxon-Rangsummentest verweist auf den hochsignifikanten Einfluss (p < 0,01) des Faktors PRP auf die Ausdehnung Tartrat-resistenter saurer Phosphatase-positiver Areale in den Präparaten. Diese Effekte können sowohl auf den im PRP angereicherten Wachstumsfaktoren als auch auf dem homologen Charakter der PRP-Zubereitung beruhen. Die beobachteten Polykaryen – sogenannte "Fremdkörperriesenzellen" – sind auch immer Indikatoren einer stattfindenden Entzündungsreaktion. Die histomorphometrisch dargestellte, deutlich verstärkte Fremdkörperreaktion in Gruppe II/PRP+ kann auf die PRP-Applikation zurückgeführt werden. Im weiteren Heilungsverlauf kann dies zu einer Verzögerung der knöchernen Konsolidierung der Defekte führen.Aim of the current experimental study in Minipigs was to examine the effects of homologous platelet-rich plasma (PRP) on the cell-mediated degradation of a nanoparticulate hydroxyapatite (HA) during the early phase of bone defect healing. Twenty-six male "Lewe" minipigs were divided into three groups. Standardized bone defects were created in the intercondylar region of the right femur of each pig and were filled with HA (Group I/PRP-, n = 11) or HA + PRP (Group II/PRP+, n = 11). The defects of the control group (n = 4) were left empty. During the implantation procedure blood was drawn from six minipigs (250 ml each). PRP was isolated from these blood samples after several centrifugation steps. After the addition of thrombin and calcium gluconate growth factors were released from the alpha-granules of the thrombocytes which were enriched within the PRP. Some of these growth factors are Platelet Derived Growth Factor AB and BB (PDGF AB, BB), Transforming Growth Factor ß 1 (TGF beta1), Vascular Endothelial Growth Factor (VEGF) and basic Fibroblast Growth Factor (bFGF). The level of enrichment of these growth factors was controlled by the ELISA technique. Growth factor enrichment within the PRP ranged from 1.6 fold (TGF-beta1) to 24.4 fold (bFGF). After 20 days the treated distal femura were explanted. For light microscopical examination different tissue embedding methods (paraffine, plastic, resin), sectioning techniques (paraffine sections, plastic and resin sections, sawing and grinding sections), staining procedures (toluidine blue, hematoxylin eosin, safranin) and histochemical methods (enzyme- and immunohistochemistry) were performed. Additionally transmission electron microscopy and computer-assisted histomorphometry were used. The results of light microscopy and transmission electron microscopy showed that regardless of the addition of PRP, the HA is degraded by hydrolysis and macrophages. The population of macrophages consists of Langhans-type giant cells. The adhesion of the polarized polykaryons at the surfaces of the implant is mediated by the apical domain of the plasmamembranes. Vimentin condensations of the cytoplasm are attached to the apical plasmalemma. In contrast, non-adherent polykaryons of the granulation tissue reveal a homogeneous Vimentin distribution pattern within their cytoplasma. As shown ultrastructurally, the implant is degraded by means of phagocytosis. The implant particles are encircled by pseudopodia of the polykaryons and become incorporated into the cytoplasma. The degradation process is supported by disintegration of the bone substitute into numerous small particles after implantation. This disintegration causes enlargement of the implant surface and increases the probability of phagocyte adhesion. The pattern of CD44 expression points towards a functional role of the molecule during fusion of mononucleated macrophages into multinucleated giant cells. Implant-associated polykaryons show CD44 immunoreactivity only along the basal domains of the cytomembrane. This pattern can be interpreted as a temporal event during adhesion. Adherent polykaryons are further characterized by strong cathepsin K expression. The histomorphometric examination demonstrates twice as much foreign body giant cells in "Group II/PRP+" as in "Group I/PRP-". Based on these results, a Wilcoxon-signed-rank test was performed and a highly significant effect (p < 0.01) of PRP on the expansion of tartrate resistent acid phophatase (TRAP)-positive areas within bone defects could be demonstrated. This effect could be a result of the substution of PRP or of its homologous character. The polykaryons descibed in this work - so-called Foreign Body Giant Cells - are also indicators of inflammation. The enhanced cellular reaction observed in Group II/PRP+ must be interpreted as a strong foreign body reaction, triggered by the addition of PRP. It cannot be excluded that the strong inflammation reaction will lead to delayed bone formation in the course of healing

Physiological Stress Mediates the Honesty of Social Signals

Peer reviewedPublisher PD

DAF-12 Regulates a Connected Network of Genes to Ensure Robust Developmental Decisions

The nuclear receptor DAF-12 has roles in normal development, the decision to pursue dauer development in unfavorable conditions, and the modulation of adult aging. Despite the biologic importance of DAF-12, target genes for this receptor are largely unknown. To identify DAF-12 targets, we performed chromatin immunoprecipitation followed by hybridization to whole-genome tiling arrays. We identified 1,175 genomic regions to be bound in vivo by DAF-12, and these regions are enriched in known DAF-12 binding motifs and act as DAF-12 response elements in transfected cells and in transgenic worms. The DAF-12 target genes near these binding sites include an extensive network of interconnected heterochronic and microRNA genes. We also identify the genes encoding components of the miRISC, which is required for the control of target genes by microRNA, as a target of DAF-12 regulation. During reproductive development, many of these target genes are misregulated in daf-12(0) mutants, but this only infrequently results in developmental phenotypes. In contrast, we and others have found that null daf-12 mutations enhance the phenotypes of many miRISC and heterochronic target genes. We also find that environmental fluctuations significantly strengthen the weak heterochronic phenotypes of null daf-12 alleles. During diapause, DAF-12 represses the expression of many heterochronic and miRISC target genes, and prior work has demonstrated that dauer formation can suppress the heterochronic phenotypes of many of these target genes in post-dauer development. Together these data are consistent with daf-12 acting to ensure developmental robustness by committing the animal to adult or dauer developmental programs despite variable internal or external conditions