Sex-dependent effects of social status on the regulation of arginine-vasopressin (AVP) V1a, oxytocin (OT), and serotonin (5-HT) 1A receptor binding and aggression in Syrian hamsters (Mesocricetus auratus).

Dominance status in hamsters is driven by interactions between arginine-vasopressin V1a, oxytocin (OT), and serotonin 1A (5-HT1A) receptors. Activation of V1a and OT receptors in the anterior hypothalamus (AH) increases aggression in males, while decreasing aggression in females. In contrast, activation of 5-HT1A receptors in the AH decreases aggression in males and increases aggression in females. The mechanism underlying these differences is not known. The purpose of this study was to determine if dominance status and sex interact to regulate V1a, OT, and 5-HT1A receptor binding. Same-sex hamsters (N = 47) were paired 12 times across six days in five min sessions. Brains from paired and unpaired (non-social control) hamsters were collected immediately after the last interaction and processed for receptor binding using autoradiography. Differences in V1a, OT, and 5-HT1A receptor binding densities were observed in several brain regions as a function of social status and sex. For example, in the AH, there was an interaction between sex and social status, such that V1a binding in subordinate males was lower than in subordinate females and V1a receptor density in dominant males was higher than in dominant females. There was also an interaction in 5-HT1A receptor binding, such that social pairing increased 5-HT1A binding in the AH of males but decreased 5-HT1A binding in females compared with unpaired controls. These results indicate that dominance status and sex play important roles in shaping the binding profiles of key receptor subtypes across the neural circuitry that regulates social behavior.

The association between range usage and tibial quality in commercial free-range laying hens.

1. Bone tissue adapts continuously to metabolic calcium demands, as well as to external forces due to physical weight loading subject to hen movement. Limited calcium metabolism and, subsequently, its availability from the medullary bone, is a major factor contributing to reduced eggshell quality in hens in the late laying period (>60 weeks of age). 2. Increasing physical activity and biomechanical loading during hen rearing has been demonstrated to increase skeletal strength, enhancing bone mass as well as endocortical and periosteal bone metabolism. Presently, the consequences of range use during lay on bone quality characteristics in laying hens remain unknown. 3.The aims of this study were to characterise tibiotarsal bone indices and evaluate the impact of range access during lay on tibia bone quality in commercial free-range laying hens. 4. This exploratory study described and analysed the volumetric measurements, morphological mechanical and trabeculae indices of the tibiotarsal bone of 48 Lohmann Brown laying hens at 74 weeks of age. All bone parameters were obtained using micro-computed tomography and correlated with individual hen range use. 5. Range usage throughout lay was not associated with tibial trabecular architecture (bone volume and fraction, trabecular thickness, trabecular connectivity density and structural model index), or any other morphological characteristics (breaking strength, diaphyseal diameter, bone weight and bone mineral density) of the tibia (P > 0.05) when hens were 74 weeks of age. 6. The results demonstrated a large variation in individual bone characteristics and suggested that range usage was not associated with bone quality in commercial free-range laying hens used in this study. In conclusion, the bone health of free-range commercial laying hens may be positively impacted by other features, such as hen genetics, feed, the quality of pullet rearing, perch availability or other shed equipment, and the benefits of these variables exceed the benefit of range use.

Flock use of the range is associated with the use of different components of a multi-tier aviary system in commercial free-range laying hens.

1. The objective of this study was to investigate the association of using a multi-tier aviary system and access to range on flock uniformity in free-range laying hens, and to determine whether the extent of range use or flock uniformity can be predicted from the use of different levels of the aviary system.2. A total of 13,716 Lohmann Brown hens from five commercial free-range flocks housed in identical houses on the same farm were individually weighed at 16 weeks of age and allocated to five replicate areas within each house. Hen movement in the multi-tier aviary system and on the range was individually monitored using radio frequency identification (RFID). All hens had access to the range from 18 to 22 weeks of age and were exposed to the same management conditions.3. Whilst only one flock significantly changed its flock uniformity with time, they differed from each other in uniformity and body weight (P = 0.001).4. Hens spent most of their available time on the lower aviary tier (7.29 ± 0.029 h/hen/day) and on the upper aviary tier (4.29 ± 0.024 h/hen/day) while the least amount of time was spent on the range and in the nest boxes (0.93 ± 0.005 h/hen/day and 1.48 ± 0.007 h, respectively, P = 0.001).5. Range use was negatively correlated (r = -0.30) to the time spent on the upper aviary tier and positively correlated (r = 0.46) to the time spent on the lower aviary tier (P = 0.001). Bivariate analysis revealed that range and upper aviary resp. lower aviary tier usage had a significant curvilinear association.6. In conclusion, the study showed that range use was associated to the time hens spent on the different tiers of the aviary system. Flock uniformity varied between flocks but was not associated with either range and aviary system usage.

Uncharged Helical Modular Polypeptide Hydrogels for Cellular Scaffolds.

Grafted synthetic polypeptides hold appeal for extending the range of biophysical properties achievable in synthetic extracellular matrix (ECM) hydrogels. Here, N-carboxyanhydride polypeptide, poly(γ-propargyl-l-glutamate) (PPLG) macromers were generated by fully grafting the "clickable" side chains with mixtures of short polyethylene glycol (PEG) chains terminated with inert (-OH) or reactive (maleimide and/or norbornene) groups, then reacting a fraction of these groups with an RGD cell attachment motif. A panel of synthetic hydrogels was then created by cross-linking the PPLG macromers with a 4-arm PEG star molecule. Compared to well-established PEG-only hydrogels, gels containing PPLG exhibited dramatically less dependence on swelling as a function of cross-link density. Further, PPLG-containing gels, which retain an α-helical chain conformation, were more effective than standard PEG gels in fostering attachment of a human mesenchymal stem cell (hMSC) line for a given concentration of RGD in the gel. These favorable properties of PPLG-containing PEG hydrogels suggest they may find broad use in synthetic ECM.

Family Nurture Intervention in preterm infants alters frontal cortical functional connectivity assessed by EEG coherence.

AIM: To assess the impact of Family Nurture Intervention (FNI) on cortical function in preterm infants at term age. METHODS: Family Nurture Intervention is a NICU-based intervention designed to establish emotional connection between mothers and preterm infants. Infants born at 26-34 weeks postmenstrual age (PMA) were divided into two groups, standard care (SC, N = 49) and FNI (FNI, N = 56). Infants had EEG recordings of ~one hour duration with 124 lead nets between 37 and 44 weeks PMA. Coherence was measured between all pairs of electrodes in ten frequency bands. Data were summarised both within and between 12 regions during two sleep states (active, quiet). RESULTS: Coherence levels were negatively correlated with PMA age in both groups. As compared to SC infants, FNI infants showed significantly lower levels of EEG coherence (1-18 Hz) largely within and between frontal regions. CONCLUSION: Coherence in FNI infants was decreased in regions where we previously found robust increases in EEG power. As coherence decreases with age, results suggest that FNI may accelerate brain maturation particularly in frontal brain regions, which have been shown in research by others to be involved in regulation of attention, cognition and emotion regulation; domains deficient in preterm infants.

A 9-year retrospective evaluation of 102 pressure ulcer reconstructions.

OBJECTIVE: Several pressure ulcer (PU) risk factors including paralysis and age greater than 70 have been identified, while others such as nutrition are debated. The object of this study is to identify perioperative risk factors that may predict improved outcomes and reduced complications in primary and recurrent PU reconstructions. METHOD: A retrospective chart review of patients treated surgically for PUs from 2004 to 2013 at the University of Toledo Medical Center, Toledo, Ohio, US, was completed. Data collected included ulcer and medical history, as well as risk factors, complications and postoperative outcome. Data were statistically analysed for perioperative variances between primary and recurrent ulcers and closure status. RESULTS: A total of 49 patients with 102 reconstructions were reviewed. Spinal cord injured patients accounted for 90% receiving flap coverage of ulcers. Numerous differences between primary and recurrent ulcers were identified, including ulcer location, patient nutritional status, wound infection, postoperative course and recurrence. Multivariate analysis revealed a flap reconstruction prediction model using creatinine, haematocrit, haemoglobin, and prealbumin that is able to successfully predict closure outcome in 83.6% of cases. CONCLUSION: Many factors play a role in the development, course and treatment of PUs. It is vital to understand the role of patient risk factors in the development of PUs, to direct subsequent management and reconstruction, and to prevent future recurrences. DECLARATION OF INTEREST: The authors have no conflicts of interest to disclose.

Generalized platform for antibody detection using the antibody catalyzed water oxidation pathway.

Infectious diseases, such as influenza, present a prominent global problem including the constant threat of pandemics that initiate in avian or other species and then pass to humans. We report a new sensor that can be specifically functionalized to detect antibodies associated with a wide range of infectious diseases in multiple species. This biosensor is based on electrochemical detection of hydrogen peroxide generated through the intrinsic catalytic activity of all antibodies: the antibody catalyzed water oxidation pathway (ACWOP). Our platform includes a polymer brush-modified surface where specific antibodies bind to conjugated haptens with high affinity and specificity. Hydrogen peroxide provides an electrochemical signal that is mediated by Resorufin/Amplex Red. We characterize the biosensor platform, using model anti-DNP antibodies, with the ultimate goal of designing a versatile device that is inexpensive, portable, reliable, and fast. We demonstrate detection of antibodies at concentrations that fall well within clinically relevant levels.

Polymer Brushes as Functional, Patterned Surfaces for Nanobiotechnology.

Polymer brushes have many desirable characteristics such as the ability to tether molecules to a substrate or change the properties of a surface. Patterning of polymer films has been an area of great interest due to the broad range of applications including bio-related and medicinal research. Consequently, we have investigated patterning techniques for polymer brushes which allow for two different functionalities on the same surface. This method has been applied to a biosensor device which requires both polymer brushes and a photosensitizer to be polymerized on a patterned gold substrate. Additionally, the nature of patterned polymer brushes as removable thin films was explored. An etching process has enabled us to lift off very thin membranes for further characterization with the potential of using them as Janus membranes for biological applications.

Actin dynamics in vivo.

Actin dynamics in lamellipodia are driven by continuous cycles of actin polymerization, retrograde flow, and depolymerization. In the past year, advances have been made in identifying signaling pathways that regulate actin-filament uncapping and polymerization, in determining the role of myosin motor proteins in retrograde flow, and in evaluating the role of severing proteins in actin depolymerization. Both Listeria monocytogenes and Saccharomyces cerevisiae have emerged as powerful model organisms for studying actin dynamics in cells.

The yeast actin cytoskeleton.

Budding and fission yeast present significant advantages for studies of the actin cytoskeleton. The application of classical and molecular genetic techniques provides a facile route for the analysis of structure/function relationships, for the isolation of novel proteins involved in cytoskeletal function, and for deciphering the signals that regulate actin assembly in vivo. This review focuses on the budding yeast Saccharomyces cerevisiae and also identifies some recent advances from studies on the fission yeast Schizosaccharomyces pombe, for which studies on the actin cytoskeleton are still in their infancy.

Spatial control of actin polymerization during neutrophil chemotaxis.

Neutrophils respond to chemotactic stimuli by increasing the nucleation and polymerization of actin filaments, but the location and regulation of these processes are not well understood. Here, using a permeabilized-cell assay, we show that chemotactic stimuli cause neutrophils to organize many discrete sites of actin polymerization, the distribution of which is biased by external chemotactic gradients. Furthermore, the Arp2/3 complex, which can nucleate actin polymerization, dynamically redistributes to the region of living neutrophils that receives maximal chemotactic stimulation, and the least-extractable pool of the Arp2/3 complex co-localizes with sites of actin polymerization. Our observations indicate that chemoattractant-stimulated neutrophils may establish discrete foci of actin polymerization that are similar to those generated at the posterior surface of the intracellular bacterium Listeria monocytogenes. We propose that asymmetrical establishment and/or maintenance of sites of actin polymerization produces directional migration of neutrophils in response to chemotactic gradients.

Characterization and cloning of a novel glycoprotein expressed by stromal cells in T-dependent areas of peripheral lymphoid tissues.

A novel glycoprotein (gp) expressed by stromal cells of peripheral lymphoid tissue has been characterized immunohistochemically, biochemically, and at the molecular level. This molecule, gp38, was identified with a monoclonal antibody (mAb) (clone 8.1.1) previously shown to react with a subpopulation of thymic epithelium. This mAb generated a reticular labeling pattern in medullary and paracortical areas of lymph nodes and in splenic white pulp. At the ultrastructural level, labeling by the 8.1.1 mAb was restricted to fibroblastic reticular stromal cells. Serial sections of lymph node and spleen labeled with anti-CD3, anti-B220, and 8.1.1 mAbs clearly showed that the 8.1.1+ cells were associated with T cell-dependent areas. In severe combined immunodeficiency (SCID) or Nu/Nu mice, splenic white pulp also exhibited reticular labeling with the 8.1.1 mAb in the absence of detectable numbers of T cells, indicating that the appearance of 8.1.1-reactive stromal cells in discrete areas of peripheral lymphoid tissue was T cell independent. The cDNA encoding this stromal cell molecule was cloned by direct expression in COS cells and found to encode a 172 amino acid sequence with the typical features of a type I integral membrane protein. COS cells transfected with the gp38 clone direct the expression of an approximately 38-kD protein that reacts with the 8.1.1 mAb but not with isotype-matched controls. Comparison of the predicted amino acid sequence of 8.1.1 mAb but not with isotype-matched controls. Comparison of the predicted amino acid sequence of 8.1.1 with proteins in the National Biomedical Research Foundation (NBRF) data base showed that gp38 is very closely related to the early response protein OTS-8 obtained from a cDNA library of tumor promoting agent (TPA)-induced murine osteoblastic cell line, MC3T3-E1.

Ultrashort pulse compression and delivery in a hollow-core photonic crystal fiber at 540 nm wavelength.

We have fabricated a bandgap-guiding hollow-core photonic crystal fiber (PCF) capable of transmitting and compressing ultrashort pulses in the green spectral region around 532 nm. When propagating subpicosecond pulses through 1 m of this fiber, we have observed soliton-effect temporal compression by up to a factor of 3 to around 100 fs. This reduces the wavelength at which soliton effects have been observed in hollow-core PCF by over 200 nm. We have used the pulses delivered at the output of the fiber to machine micrometer-scale features in copper.

Recurrent massive pulmonary emboli in a critically ill patient with COVID-19.

We report the haematological management of a critically ill patient with coronavirus disease 2019 (COVID-19), with recurrent massive pulmonary emboli. A previous healthy 56-year-old man presented to the emergency department with severe hypoxaemic respiratory failure due to suspected COVID-19. He required invasive mechanical ventilation and transfer to the intensive care unit for increasing ventilatory requirements and cardiovascular instability. A computed tomography (CT) pulmonary angiogram demonstrated large bilateral pulmonary emboli with right heart strain, for which he received intravenous systemic thrombolysis followed by therapeutic weight-adjusted anticoagulation with low molecular weight heparin (dalteparin). Two weeks later, following an acute respiratory deterioration, a repeat CT pulmonary angiogram demonstrated a new saddle embolus with right heart strain requiring another regime of intravenous systemic thrombolysis. This occurred despite anti-Xa-guided therapeutic anticoagulation. The dose of therapeutic dalteparin was increased incrementally to an eventual dose of 12,500 units twice daily. A low threshold for radiological imaging should be considered in all COVID-19 patients with acute cardiorespiratory deterioration. Multidisciplinary team discussions highlighted aspects of balancing the risks of bleeding from anticoagulation vs. risk of death from pulmonary embolism. This report highlights the need for further research into the underlying mechanisms and optimal management of thrombotic complications in COVID-19.

The world according to Arp: regulation of actin nucleation by the Arp2/3 complex.

The coordination of cell shape change and locomotion requires that actin polymerization at the cell cortex be tightly controlled in response to both intracellular and extracellular cues. The Arp2/3 complex - an actin filament nucleating and organizing factor - appears to be a central player in the cellular control of actin assembly. Recently, a molecular pathway leading from key signalling molecules to actin filament nucleation by the Arp2/3 complex has been discovered. In this pathway, the GTPase Cdc42 acts in concert with WASP family proteins to activate the Arp2/3 complex. These findings have led to a more complete picture of the mechanism of actin filament generation and organization during cell motility.

Three regions within ActA promote Arp2/3 complex-mediated actin nucleation and Listeria monocytogenes motility.

The Listeria monocytogenes ActA protein induces actin-based motility by enhancing the actin nucleating activity of the host Arp2/3 complex. Using systematic truncation analysis, we identified a 136-residue NH(2)-terminal fragment that was fully active in stimulating nucleation in vitro. Further deletion analysis demonstrated that this fragment contains three regions, which are important for nucleation and share functional and/or limited sequence similarity with host WASP family proteins: an acidic stretch, an actin monomer-binding region, and a cofilin homology sequence. To determine the contribution of each region to actin-based motility, we compared the biochemical activities of ActA derivatives with the phenotypes of corresponding mutant bacteria in cells. The acidic stretch functions to increase the efficiency of actin nucleation, the rate and frequency of motility, and the effectiveness of cell-cell spread. The monomer-binding region is required for actin nucleation in vitro, but not for actin polymerization or motility in infected cells, suggesting that redundant mechanisms may exist to recruit monomer in host cytosol. The cofilin homology sequence is critical for stimulating actin nucleation with the Arp2/3 complex in vitro, and is essential for actin polymerization and motility in cells. These data demonstrate that each region contributes to actin-based motility, and that the cofilin homology sequence plays a principal role in activation of the Arp2/3 complex, and is an essential determinant of L. monocytogenes pathogenesis.

Temporal relationships of F-actin bundle formation, collagen and fibronectin matrix assembly, and fibronectin receptor expression to wound contraction.

Wound contraction can substantially reduce the amount of new tissue needed to reestablish organ integrity after tissue loss. Fibroblasts, rich in F-actin bundles, generate the force of wound contraction. Fibronectin-containing microfibrils link fibroblasts to each other and to collagen bundles and thereby provide transduction cables across the wound for contraction. The temporal relationships of F-actin bundle formation, collagen and fibronectin matrix assembly, and fibronectin receptor expression to wound contraction have not been determined. To establish these relationships, we used a cutaneous gaping wound model in outbred Yorkshire pigs. Granulation tissue filled approximately 80% of the wound space by day 5 after injury while wound contraction was first apparent at day 10. Neither actin bundles nor fibronectin receptors were observed in 5-d wound fibroblasts. Although fibronectin fibrils were assembled on the surfaces of 5-d fibroblasts, few fibrils coursed between cells. Day-7 fibroblasts stained strongly for nonmuscle-type F-actin bundles consistent with a contractile fibroblast phenotype. These cells expressed fibronectin receptors, were embedded in a fibronectin matrix that appeared to connect fibroblasts to the matrix and to each other, and were coaligned across the wound. Transmission EM confirmed the presence of microfilament bundles, cell-cell and cell-matrix linkages at day 7. Fibroblast coalignment, matrix interconnections, and actin bundles became more pronounced at days 10 and 14 coinciding with tissue contraction. These findings demonstrate that granulation tissue formation, F-actin bundle and fibronectin receptor expression in wound fibroblasts, and fibroblast-matrix linkage precede wound contraction.

The human Arp2/3 complex is composed of evolutionarily conserved subunits and is localized to cellular regions of dynamic actin filament assembly.

The Arp2/3 protein complex has been implicated in the control of actin polymerization in cells. The human complex consists of seven subunits which include the actin related proteins Arp2 and Arp3, and five others referred to as p41-Arc, p34-Arc, p21-Arc, p20-Arc, and p16-Arc (p omplex). We have determined the predicted amino acid sequence of all seven subunits. Each has homologues in diverse eukaryotes, implying that the structure and function of the complex has been conserved through evolution. Human Arp2 and Arp3 are very similar to family members from other species. p41-Arc is a new member of the Sop2 family of WD (tryptophan and aspartate) repeat-containing proteins and may be posttranslationally modified, suggesting that it may be involved in regulating the activity and/or localization of the complex. p34-Arc, p21-Arc, p20-Arc, and p16-Arc define novel protein families. We sought to evaluate the function of the Arp2/3 complex in cells by determining its intracellular distribution. Arp3, p34-Arc, and p21-Arc were localized to the lamellipodia of stationary and locomoting fibroblasts, as well to Listeria monocytogenes assembled actin tails. They were not detected in cellular bundles of actin filaments. Taken together with the ability of the Arp2/3 complex to induce actin polymerization, these observations suggest that the complex promotes actin assembly in lamellipodia and may participate in lamellipodial protrusion.

Pivotal role of VASP in Arp2/3 complex-mediated actin nucleation, actin branch-formation, and Listeria monocytogenes motility.

The Listeria monocytogenes ActA protein mediates actin-based motility by recruiting and stimulating the Arp2/3 complex. In vitro, the actin monomer-binding region of ActA is critical for stimulating Arp2/3-dependent actin nucleation; however, this region is dispensable for actin-based motility in cells. Here, we provide genetic and biochemical evidence that vasodilator-stimulated phosphoprotein (VASP) recruitment by ActA can bypass defects in actin monomer-binding. Furthermore, purified VASP enhances the actin-nucleating activity of wild-type ActA and the Arp2/3 complex while also reducing the frequency of actin branch formation. These data suggest that ActA stimulates the Arp2/3 complex by both VASP-dependent and -independent mechanisms that generate distinct populations of actin filaments in the comet tails of L. monocytogenes. The ability of VASP to contribute to actin filament nucleation and to regulate actin filament architecture highlights the central role of VASP in actin-based motility.