NANOG is required to establish the competence for germ-layer differentiation in the basal tetrapod axolotl.

Pluripotency defines the unlimited potential of individual cells of vertebrate embryos, from which all adult somatic cells and germ cells are derived. Understanding how the programming of pluripotency evolved has been obscured in part by a lack of data from lower vertebrates; in model systems such as frogs and zebrafish, the function of the pluripotency genes NANOG and POU5F1 have diverged. Here, we investigated how the axolotl ortholog of NANOG programs pluripotency during development. Axolotl NANOG is absolutely required for gastrulation and germ-layer commitment. We show that in axolotl primitive ectoderm (animal caps; ACs) NANOG and NODAL activity, as well as the epigenetic modifying enzyme DPY30, are required for the mass deposition of H3K4me3 in pluripotent chromatin. We also demonstrate that all 3 protein activities are required for ACs to establish the competency to differentiate toward mesoderm. Our results suggest the ancient function of NANOG may be establishing the competence for lineage differentiation in early cells. These observations provide insights into embryonic development in the tetrapod ancestor from which terrestrial vertebrates evolved.

Study on impact of robotic-assisted orthopaedic industrial noise (SIREN).

INTRODUCTION: The aim of this study was to evaluate noise exposure to the operating room staff consisting of the surgeon, assistant, anaesthetist and Mako Product Specialist (MPS) during Mako robotic-arm assisted total knee arthroplasty (TKA) and total hip arthroplasty (THA). We aimed to determine whether employees were exposed to noise at or above a lower exposure action value (LEAV) set out by the Noise at Work Regulations 2005, Health and Safety Executive (HSE), UK. MATERIALS AND METHODS: We prospectively recorded intra-operative noise levels in Mako robotic-arm assisted TKA and THA over a period of two months using the MicW i436 connected to an iOS device (Apple), using the Sound Level Meter App (iOS) by the National Institute for Occupation Safety and Health (NIOSH). Data obtained was then used to calculate "worst case" daily exposure value to assess if sound levels were compliant with UK guidelines. Comparison between operating room staff groups was performed with ANOVA testing. RESULTS: A total of 19 TKA and 11 THA operations were recorded. During TKA, for the primary surgeon and the assistant, the equivalent continuous sound pressure level (LAeq) was over 80 dB, exceeding the LEAV set out by the Noise at Work Regulations by HSE. During THA, the average LAeq and peak sound pressure levels did not exceed the LEAV. The calculated daily exposure for the primary surgeon in TKA was 82 dB. A Tukey post hoc test revealed that LAeq was statistically significantly lower in the anaesthetist and MPS (p < .001) compared to the primary surgeon and assistant in both TKA and THA. CONCLUSIONS: Operating room staff, particularly the primary surgeon and assistant are exposed to significant levels of noise during Mako robotic-arm assisted TKA and THA. Formal assessments should be performed to further assess the risk of noise induced hearing loss in robotic-arm assisted arthroplasty.

Thermodynamics, organisms and behaviour.

The physical origin of behaviour in biological organisms is distinct from those of non-living systems in one significant way: organisms exhibit intentionality or goal-directed behaviour. How may we understand and explain this important aspect in physical terms, grounded in laws of physics and chemistry? In this article, we discuss recent experimental and theoretical progress in this area and future prospects of this line of thought. The physical basis for our investigation is thermodynamics, though other branches of physics and chemistry have an important role. This article is part of the theme issue 'Thermodynamics 2.0: Bridging the natural and social sciences (Part 1)'.

Acoustic information about upper limb movement in voicing.

We show that the human voice has complex acoustic qualities that are directly coupled to peripheral musculoskeletal tensioning of the body, such as subtle wrist movements. In this study, human vocalizers produced a steady-state vocalization while rhythmically moving the wrist or the arm at different tempos. Although listeners could only hear and not see the vocalizer, they were able to completely synchronize their own rhythmic wrist or arm movement with the movement of the vocalizer which they perceived in the voice acoustics. This study corroborates recent evidence suggesting that the human voice is constrained by bodily tensioning affecting the respiratory-vocal system. The current results show that the human voice contains a bodily imprint that is directly informative for the interpersonal perception of another's dynamic physical states.

Development of a Gene-Activated Scaffold Incorporating Multifunctional Cell-Penetrating Peptides for pSDF-1α Delivery for Enhanced Angiogenesis in Tissue Engineering Applications.

Non-viral gene delivery has become a popular approach in tissue engineering, as it permits the transient delivery of a therapeutic gene, in order to stimulate tissue repair. However, the efficacy of non-viral delivery vectors remains an issue. Our lab has created gene-activated scaffolds by incorporating various non-viral delivery vectors, including the glycosaminoglycan-binding enhanced transduction (GET) peptide into collagen-based scaffolds with proven osteogenic potential. A modification to the GET peptide (FLR) by substitution of arginine residues with histidine (FLH) has been designed to enhance plasmid DNA (pDNA) delivery. In this study, we complexed pDNA with combinations of FLR and FLH peptides, termed GET* nanoparticles. We sought to enhance our gene-activated scaffold platform by incorporating GET* nanoparticles into collagen-nanohydroxyapatite scaffolds with proven osteogenic capacity. GET* N/P 8 was shown to be the most effective formulation for delivery to MSCs in 2D. Furthermore, GET* N/P 8 nanoparticles incorporated into collagen-nanohydroxyapatite (coll-nHA) scaffolds at a 1:1 ratio of collagen:nanohydroxyapatite was shown to be the optimal gene-activated scaffold. pDNA encoding stromal-derived factor 1α (pSDF-1α), an angiogenic chemokine which plays a role in BMP mediated differentiation of MSCs, was then delivered to MSCs using our optimised gene-activated scaffold platform, with the aim of significantly increasing angiogenesis as an important precursor to bone repair. The GET* N/P 8 coll-nHA scaffolds successfully delivered pSDF-1α to MSCs, resulting in a significant, sustained increase in SDF-1α protein production and an enhanced angiogenic effect, a key precursor in the early stages of bone repair.

Foraging Dynamics and Entropy Production in a Simulated Proto-Cell.

All organisms depend on a supply of energetic resources to power behavior and the irreversible entropy-producing processes that sustain them. Dissipative structure theory has often been a source of inspiration for better understanding the thermodynamics of biology, yet real organisms are inordinately more complex than most laboratory systems. Here we report on a simulated chemical dissipative structure that operates as a proto cell. The simulated swimmer moves through a 1D environment collecting resources that drive a nonlinear reaction network interior to the swimmer. The model minimally represents properties of a simple organism including rudimentary foraging and chemotaxis and an analog of a metabolism in the nonlinear reaction network. We evaluated how dynamical stability of the foraging dynamics (i.e., swimming and chemotaxis) relates to the rate of entropy production. Results suggested a relationship between dynamical steady states and entropy production that was tuned by the relative coordination of foraging and metabolic processes. Results include evidence in support of and contradicting one formulation of a maximum entropy production principle. We discuss the status of this principle and its relevance to biology.

Transcriptome-based screening of ion channels and transporters in a migratory chondroprogenitor cell line isolated from late-stage osteoarthritic cartilage.

Chondrogenic progenitor cells (CPCs) may be used as an alternative source of cells with potentially superior chondrogenic potential compared to mesenchymal stem cells (MSCs), and could be exploited for future regenerative therapies targeting articular cartilage in degenerative diseases such as osteoarthritis (OA). In this study, we hypothesised that CPCs derived from OA cartilage may be characterised by a distinct channelome. First, a global transcriptomic analysis using Affymetrix microarrays was performed. We studied the profiles of those ion channels and transporter families that may be relevant to chondroprogenitor cell physiology. Following validation of the microarray data with quantitative reverse transcription-polymerase chain reaction, we examined the role of calcium-dependent potassium channels in CPCs and observed functional large-conductance calcium-activated potassium (BK) channels involved in the maintenance of the chondroprogenitor phenotype. In line with our very recent results, we found that the KCNMA1 gene was upregulated in CPCs and observed currents that could be attributed to the BK channel. The BK channel inhibitor paxilline significantly inhibited proliferation, increased the expression of the osteogenic transcription factor RUNX2, enhanced the migration parameters, and completely abolished spontaneous Ca2+ events in CPCs. Through characterisation of their channelome we demonstrate that CPCs are a distinct cell population but are highly similar to MSCs in many respects. This study adds key mechanistic data to the in-depth characterisation of CPCs and their phenotype in the context of cartilage regeneration.

Mitochondrial dynamics and bioenergetics regulated by netrin-1 in oligodendrocytes.

Mitochondria are dynamic organelles that produce energy and molecular precursors that are essential for myelin synthesis. Unlike in neurons, mitochondria in oligodendrocytes increase intracellular movement in response to glutamatergic activation and are more susceptible to oxidative stress than in astrocytes or microglia. The signaling pathways that regulate these cell type-specific mitochondrial responses in oligodendrocytes are not understood. Here, we visualized mitochondria migrating through thin cytoplasmic channels crossing myelin basic protein-positive compacted membranes and localized within paranodal loop cytoplasm. We hypothesized that local extracellular enrichment of netrin-1 might regulate the recruitment and function of paranodal proteins and organelles, including mitochondria. We identified rapid recruitment of mitochondria and paranodal proteins, including neurofascin 155 (NF155) and the netrin receptor deleted in colorectal carcinoma (DCC), to sites of contact between oligodendrocytes and netrin-1-coated microbeads in vitro. We provide evidence that Src-family kinase activation and Rho-associated protein kinase (ROCK) inhibition downstream of netrin-1 induces mitochondrial elongation, hyperpolarization of the mitochondrial inner membrane, and increases glycolysis. Our findings identify a signaling mechanism in oligodendrocytes that is sufficient to locally recruit paranodal proteins and regulate the subcellular localization, morphology, and function of mitochondria.

Functional Interdependence in Coupled Dissipative Structures: Physical Foundations of Biological Coordination.

Coordination within and between organisms is one of the most complex abilities of living systems, requiring the concerted regulation of many physiological constituents, and this complexity can be particularly difficult to explain by appealing to physics. A valuable framework for understanding biological coordination is the coordinative structure, a self-organized assembly of physiological elements that collectively performs a specific function. Coordinative structures are characterized by three properties: (1) multiple coupled components, (2) soft-assembly, and (3) functional organization. Coordinative structures have been hypothesized to be specific instantiations of dissipative structures, non-equilibrium, self-organized, physical systems exhibiting complex pattern formation in structure and behaviors. We pursued this hypothesis by testing for these three properties of coordinative structures in an electrically-driven dissipative structure. Our system demonstrates dynamic reorganization in response to functional perturbation, a behavior of coordinative structures called reciprocal compensation. Reciprocal compensation is corroborated by a dynamical systems model of the underlying physics. This coordinated activity of the system appears to derive from the system's intrinsic end-directed behavior to maximize the rate of entropy production. The paper includes three primary components: (1) empirical data on emergent coordinated phenomena in a physical system, (2) computational simulations of this physical system, and (3) theoretical evaluation of the empirical and simulated results in the context of physics and the life sciences. This study reveals similarities between an electrically-driven dissipative structure that exhibits end-directed behavior and the goal-oriented behaviors of more complex living systems.

Discovery of peptide ligands targeting a specific ubiquitin-like domain-binding site in the deubiquitinase USP11.

Ubiquitin-specific proteases (USPs) reverse ubiquitination and regulate virtually all cellular processes. Defined noncatalytic domains in USP4 and USP15 are known to interact with E3 ligases and substrate recruitment factors. No such interactions have been reported for these domains in the paralog USP11, a key regulator of DNA double-strand break repair by homologous recombination. We hypothesized that USP11 domains adjacent to its protease domain harbor unique peptide-binding sites. Here, using a next-generation phage display (NGPD) strategy, combining phage display library screening with next-generation sequencing, we discovered unique USP11-interacting peptide motifs. Isothermal titration calorimetry disclosed that the highest affinity peptides (KD of ∼10 µm) exhibit exclusive selectivity for USP11 over USP4 and USP15 in vitro Furthermore, a crystal structure of a USP11-peptide complex revealed a previously unknown binding site in USP11's noncatalytic ubiquitin-like (UBL) region. This site interacted with a helical motif and is absent in USP4 and USP15. Reporter assays using USP11-WT versus a binding pocket-deficient double mutant disclosed that this binding site modulates USP11's function in homologous recombination-mediated DNA repair. The highest affinity USP11 peptide binder fused to a cellular delivery sequence induced significant nuclear localization and cell cycle arrest in S phase, affecting the viability of different mammalian cell lines. The USP11 peptide ligands and the paralog-specific functional site in USP11 identified here provide a framework for the development of new biochemical tools and therapeutic agents. We propose that an NGPD-based strategy for identifying interacting peptides may be applied also to other cellular targets.

Energy flows in gesture-speech physics: The respiratory-vocal system and its coupling with hand gestures.

Expressive moments in communicative hand gestures often align with emphatic stress in speech. It has recently been found that acoustic markers of emphatic stress arise naturally during steady-state phonation when upper-limb movements impart physical impulses on the body, most likely affecting acoustics via respiratory activity. In this confirmatory study, participants (N = 29) repeatedly uttered consonant-vowel (/pa/) mono-syllables while moving in particular phase relations with speech, or not moving the upper limbs. This study shows that respiration-related activity is affected by (especially high-impulse) gesturing when vocalizations occur near peaks in physical impulse. This study further shows that gesture-induced moments of bodily impulses increase the amplitude envelope of speech, while not similarly affecting the Fundamental Frequency (F0). Finally, tight relations between respiration-related activity and vocalization were observed, even in the absence of movement, but even more so when upper-limb movement is present. The current findings expand a developing line of research showing that speech is modulated by functional biomechanical linkages between hand gestures and the respiratory system. This identification of gesture-speech biomechanics promises to provide an alternative phylogenetic, ontogenetic, and mechanistic explanatory route of why communicative upper limb movements co-occur with speech in humans.

The quantification of gesture-speech synchrony: A tutorial and validation of multimodal data acquisition using device-based and video-based motion tracking.

There is increasing evidence that hand gestures and speech synchronize their activity on multiple dimensions and timescales. For example, gesture's kinematic peaks (e.g., maximum speed) are coupled with prosodic markers in speech. Such coupling operates on very short timescales at the level of syllables (200 ms), and therefore requires high-resolution measurement of gesture kinematics and speech acoustics. High-resolution speech analysis is common for gesture studies, given that field's classic ties with (psycho)linguistics. However, the field has lagged behind in the objective study of gesture kinematics (e.g., as compared to research on instrumental action). Often kinematic peaks in gesture are measured by eye, where a "moment of maximum effort" is determined by several raters. In the present article, we provide a tutorial on more efficient methods to quantify the temporal properties of gesture kinematics, in which we focus on common challenges and possible solutions that come with the complexities of studying multimodal language. We further introduce and compare, using an actual gesture dataset (392 gesture events), the performance of two video-based motion-tracking methods (deep learning vs. pixel change) against a high-performance wired motion-tracking system (Polhemus Liberty). We show that the videography methods perform well in the temporal estimation of kinematic peaks, and thus provide a cheap alternative to expensive motion-tracking systems. We hope that the present article incites gesture researchers to embark on the widespread objective study of gesture kinematics and their relation to speech.

Dissipative Structures, Organisms and Evolution.

Self-organization in nonequilibrium systems has been known for over 50 years. Under nonequilibrium conditions, the state of a system can become unstable and a transition to an organized structure can occur. Such structures include oscillating chemical reactions and spatiotemporal patterns in chemical and other systems. Because entropy and free-energy dissipating irreversible processes generate and maintain these structures, these have been called dissipative structures. Our recent research revealed that some of these structures exhibit organism-like behavior, reinforcing the earlier expectation that the study of dissipative structures will provide insights into the nature of organisms and their origin. In this article, we summarize our study of organism-like behavior in electrically and chemically driven systems. The highly complex behavior of these systems shows the time evolution to states of higher entropy production. Using these systems as an example, we present some concepts that give us an understanding of biological organisms and their evolution.

Predictors for recovery of macular function after surgery for primary macula-off rhegmatogenous retinal detachment.

PURPOSE: To determine predictors of best-corrected postoperative visual acuity (VA) in patients who underwent surgical intervention for macula-off rhegmatogenous retinal detachment. MATERIALS AND METHODS: Primary macula-off retinal detachments from the University of Colorado Primary Rhegmatogenous Retinal Detachment Database (2012-2017) were reviewed. The primary outcome measure was a postoperative VA of 20/40 or better at least 6 months after surgery. Patient demographics, medical history, duration of central vision loss before surgery, examination findings, operative technique, and postoperative findings were analyzed as possible predictors of postoperative visual recovery to 20/40 or better. Chi-square or Fisher's exact test was used to compare categorical variables, and Wilcoxon rank sum test was used for continuous variables. A multivariable logistic regression analysis was used to determine the adjusted odds ratios and 95% confidence intervals for variables that were significant in the univariable analyses. Statistical significance was set at p < 0.05. RESULTS: One hundred thirty-one patients met inclusion criteria. Eighty-one (61.8%) patients achieved a postoperative VA of 20/40 or better 6 or more months after surgery. Patients with a single retinal break were more likely than patients with more than one break to reach a postoperative VA of 20/40 or better (76.9% vs. 55.4%, p = 0.021). Patients with a better preoperative logMAR VA had better postoperative VA (p = 0.021). Duration of central vision loss prior to surgical repair was not related to final postoperative VA in this particular study. CONCLUSION: Postoperative recovery of visual acuity to 20/40 or better was significantly more common in patients with a single retinal break as well as in patients with better preoperative visual acuity. Duration of central vision loss prior to surgical repair was not significantly associated with postoperative VA.

Grasp Affordances in Bistable Perception of the Necker Cube.

Stimulus-response compatibility effects illustrate the mutual depen-dence of perception and action processes. Ellis and Tucker (2000) showed that object identification was facilitated when the response required a grip that was compatible with the stimulus. In the current study, we extend grip-compatibility effects to perception of the Necker cube. Participants reported the perceived orientation of a Necker cube by orienting a hand-held cube into a compatible or an incompatible position. Participants in the incompatible condition were quickly attracted to the FRB (front-side right bottom) percept, consistent with previous work. However, participants in the compatible condition showed an extended period of metastability, switching between the two perceptual states about equally. A second experiment replicated these results and showed that a control condition in which responses were made with a key press produced intermediate levels of metastability. These results are interpreted in terms of the dynamics of bistable perception.

Particle Flock Motion at Air-Water Interface Driven by Interfacial Free Energy Foraging.

From flocks of birds and sheep to colonies of bacteria, complex patterns and self-motion are found in all hierarchies of nature. Artificial nonliving systems provide useful insight, since living systems are complicated and may involve cognitive issues not found in nonliving matter. Herein, we report naturally flocking irregularly shaped benzoquinone (BQ) particles on the air-water interface that cross a gate. In this open system designed with absence of external control, the particle flock moves by Marangoni "surfing" driven by slow dissolution of weakly surface active BQ postulated to create inhomogeneous interfacial tension fields. The particle flocks move collectively through a gate placed in the air-water interface to the side that has higher interfacial tension. Position-sensitive surface tension measurements used for the first time in a multiparticle Marangoni motion system show unequivocally that flock motion and gate crossing proceed to areas of slightly higher interfacial tension. Flock crossing is accompanied by a low-high differential interfacial tension change from one side of the gate to the other, with the flock moving to the side with higher interfacial tension. Thus, the flocks move because they are foraging for interfacial free energy.

Highly efficient delivery of functional cargoes by the synergistic effect of GAG binding motifs and cell-penetrating peptides.

Protein transduction domains (PTDs) are powerful nongenetic tools that allow intracellular delivery of conjugated cargoes to modify cell behavior. Their use in biomedicine has been hampered by inefficient delivery to nuclear and cytoplasmic targets. Here we overcame this deficiency by developing a series of novel fusion proteins that couple a membrane-docking peptide to heparan sulfate glycosaminoglycans (GAGs) with a PTD. We showed that this GET (GAG-binding enhanced transduction) system could deliver enzymes (Cre, neomycin phosphotransferase), transcription factors (NANOG, MYOD), antibodies, native proteins (cytochrome C), magnetic nanoparticles (MNPs), and nucleic acids [plasmid (p)DNA, modified (mod)RNA, and small inhibitory RNA] at efficiencies of up to two orders of magnitude higher than previously reported in cell types considered hard to transduce, such as mouse embryonic stem cells (mESCs), human ESCs (hESCs), and induced pluripotent stem cells (hiPSCs). This technology represents an efficient strategy for controlling cell labeling and directing cell fate or behavior that has broad applicability for basic research, disease modeling, and clinical application.

Combined hydrogels that switch human pluripotent stem cells from self-renewal to differentiation.

The ability of materials to define the architecture and microenvironment experienced by cells provides new opportunities to direct the fate of human pluripotent stem cells (HPSCs) [Robinton DA, Daley GQ (2012) Nature 481(7381):295-305]. However, the conditions required for self-renewal vs. differentiation of HPSCs are different, and a single system that efficiently achieves both outcomes is not available [Giobbe GG, et al. (2012) Biotechnol Bioeng 109(12):3119-3132]. We have addressed this dual need by developing a hydrogel-based material that uses ionic de-cross-linking to remove a self-renewal permissive hydrogel (alginate) and switch to a differentiation-permissive microenvironment (collagen). Adjusting the timing of this switch can preferentially steer the HPSC differentiation to mimic lineage commitment during gastrulation to ectoderm (early switch) or mesoderm/endoderm (late switch). As an exemplar differentiated cell type, we showed that directing early lineage specification using this single system can promote cardiogenesis with increased gene expression in high-density cell populations. This work will facilitate regenerative medicine by allowing in situ HPSC expansion to be coupled with early lineage specification within defined tissue geometries.