搜索结果：Physical biology

Scalable suspension culture technologies are essential for the large-scale manufacturing of mesenchymal stem/stromal cells (MSCs). However, conventional microcarrier-based systems often fail to achieve scalable efficiency because cell-microcarrier dynamics vary across scales, necessitating complex, scale-dependent agitation protocols. To overcome this limitation, we designed a scale-up-oriented suspension culture system employing fluffy, fibrillated nanofiber scaffolds composed of chitosan and chitin. These nanofibers were readily suspended under continuous, gentle agitation and trapped cells on their surfaces, while spontaneously forming fluffy cell-scaffold aggregates through scaffold agglomeration. The low-adhesive chitosan nanofibers acted as physical spacers that prevented aggregate coalescence, thereby maintaining a microenvironment favorable for proliferation. By defining cell-scaffold aggregate size as the key scaling parameter-a biology-centric approach-, we successfully achieved scale-up from 30 mL to 5 L under continuous gentle agitation, yielding comparable specific growth rates of (3.66 ± 0.28) × 10-2 h-1 (30 mL), (3.27 ± 0.40) × 10-2 h-1 (1 L), and 3.50 × 10-2 h-1 (5 L), and reaching a total yield of 4.23 × 109 cells in the 5-L bioreactor (a single run). These findings demonstrate that fluffy nanofiber scaffolds enable a scale-up strategy that reproduces the cellular microenvironment in a manner that is less affected by scale-dependent physical forces. This concept provides a new framework for designing scalable culture environments applicable not only to cell therapy manufacturing but also to culture supernatant production and cell-based food materials.

During metaphase, chromosomes undergo oscillatory motion and exhibit distance-dependent coordinated movement with neighboring chromosomes within the spindle. However, the physical mechanism that gives rise to coordinated chromosome motion remains unresolved. Here, we combine quantitative live-cell imaging in PTK1 cells, targeted perturbations of spindle microtubules and chromatin condensation level, and minimal mechanical modeling to uncover the mechanical basis of chromosome coordination during metaphase. We show that chromosome oscillations are dampened by stabilizing microtubules or by reducing chromatin condensation, yet inter-chromosomal coordination measured by Pearson's correlation coefficient is preserved across all conditions. Consistently, simulations show that Pearson's correlation is insensitive to the mechanical parameters governing inter-chromosomal coupling. Together, these observations motivate the hypothesis that chromosome coordination is a mechanical signature of the physical spindle environment. To test this hypothesis, we developed a minimal mechanical model incorporating transient inter-chromosomal springs as a general representation of inter-chromosomal interactions, and show they are sufficient to generate correlated chromosome motion. To quantify coordination in a manner that reflects the mechanical properties of the surrounding spindle environment, we employed a microrheology-inspired analysis of time-lagged chromosome displacements applied to both experimental and simulated data, revealing that microtubules set the spatial range of coordination while chromatin condensation level modulates its strength. Together, our results support the hypothesis that coordinated chromosome motion is an emergent mechanical property of the mitotic spindle.

Spatial biology depends on the structural dynamics of the constituents of the cell. We propose that such dynamics during the cell cycle depend to a large extent on the origin of replication and its role in the trajectory of a replication hyperstructure. To explore this ori-Centric view, we adopt both historical and speculative approaches to the initiation of chromosome replication, focusing on Escherichia coli. We relate these approaches to the framework of concepts, results, and procedures within which science is structured; this framework can be considered a paradigm or a "thought style" that sometimes undergoes a major shift. We suggest that a paradigm shift is underway in the study of the cell cycle as shown by the increasing contribution of physical chemistry and, in particular, phase transitions.

Patients with Alzheimer's disease (AD) exhibit muscle decline and physical compromise. Probiotic supplements may mitigate muscle decline and physical impairment; however, empirical investigations remain limited. We hypothesized that probiotics improve muscle strength and physical performance by repairing intestinal leak in AD patients. We conducted a randomized, double-blind, monocenter trial of AD patients receiving either a placebo (n = 54, 68-84 years old) or a probiotic (Vivomixx 112 billion*, one capsule daily, n = 51, 72-84 years old) for four months. We measured handgrip strength (HGS), body composition, the Short Physical Performance Battery (SPPB), and plasma zonulin, a marker of intestinal permeability, in patients with AD at baseline and after 4 months. Four months of probiotic supplementation improved HGS, gait speed, and total SPPB scores, accompanied by reduced plasma zonulin (all p < 0.05). Patients with sarcopenia or reduced physical capacity (SPPB&#x2264;8) exhibited higher zonulin levels. Plasma zonulin was negatively associated with HGS, gait speed, and SPPB scores in univariate analyses (all p < 0.05). Multivariate models adjusting for age, cognition, body mass index, and nutritional status confirmed independent associations of zonulin with functional performance, particularly HGS and gait speed. Probiotics also reduced circulating markers of inflammation and oxidative stress. These results are derived from a male&#x2011;only cohort and may not be directly generalizable to female patients. Collectively, probiotics improve HGS and physical capacity by strengthening the intestinal barrier and reducing systemic inflammation and oxidative stress. Further studies should investigate the relative molecular and cellular mechanisms.

Weight gain in adulthood is a common potentially modifiable breast cancer risk factor. Intermittent 5:2 diets (two low-calorie days/week) have proven efficacy for weight loss among people with overweight or obesity and can promote sustained awareness and mindfulness of diet choice and help appetite control.This trial aims to test whether a less intensive 6:1 intermittent diet programme (one low-calorie day/week) is a feasible intervention to promote healthy eating and prevent weight gain in women at increased risk of breast cancer. Single arm prospective feasibility trial in 30 healthy weight women aged 18-40 years, at moderate or high risk of breast cancer (&#x2265;17% lifetime risk and/or &#x2265;3% 10&#x2009;year risk at 40 years), body mass index 20-25&#x2009;kg/m2. Participants will be entered to a 16-week 6:1 diet programme involving 1&#x2009;day consuming 1000 kcal and healthy eating for 6&#x2009;days a week. Participants will also be advised to meet physical activity recommendations for health (&#x2265;150&#x2009;min of moderate intensity physical activity/week and resistance exercise two times per week). The programme will be supported remotely by dietitian calls at baseline, week 1, 4, 8, 12 and 16. Participants will also be provided access to a trial-specific private monitored Facebook group which includes information and the opportunity for peer support.Co-primary outcomes are: (a) uptake to the trial, (b) retention rate, (c) adherence to the 6:1 diet and (d) participant feedback on acceptability of the programme. Secondary outcomes include characteristics of those recruited and completing the programme and a preliminary evaluation of benefits and harms. This includes changes in body weight and body composition (bioelectrical impedance), diet quality, physical activity, binge eating, sleep quality (Pittsburgh Sleep Quality Index), menstrual cycle length and potentially diet-related adverse events, that is, fatigue, constipation, dizziness, headache, indigestion. Exploratory outcomes include the impact of low-calorie days on dietary intake and physical activity across the week and any differences in adherence to the low calorie days across the menstrual cycle. This trial has been approved by South Central-Berkshire B Research Ethics committee (rec reference 24/SC/0321). Findings will be disseminated via peer-reviewed journals, national and international cancer prevention and obesity conferences and cancer prevention charitable bodies. ISRCTN14330469.

The Mediterranean lifestyle is increasingly recognized as a multidimensional determinant of health. However, cross-country comparisons using harmonized instruments remain limited. This study aimed to provide a comprehensive country-by-country comparison of Mediterranean lifestyle adherence and associated psychosocial and lifestyle correlates across 10 Mediterranean and neighboring countries participating in the MEDIET4ALL project. Cross-sectional data were collected from 4,010 participants (age: 37.2&#x202f;&#xb1;&#x202f;15.4&#x202f;years; 59.5% female) using the multinational MEDIET4ALL e-survey. Mediterranean lifestyle adherence was assessed using the MedLife Index and its three domains. Psychosocial status, sleep characteristics, physical activity, sedentary behaviour, social participation, and technology use were evaluated using validated instruments. Significant cross-country differences were observed in global MedLife adherence and across all domains (p&#x202f;<&#x202f;0.001, &#x3b7; 2&#x202f;=&#x202f;0.07-0.11), as well as in the distribution of adherence categories across countries (&#x3c7; 2&#x202f;=&#x202f;113.936, p&#x202f;<&#x202f;0.001). Spain consistently showed higher MedLife scores than several countries (z&#x202f;=&#x202f;3.42-8.12, adjusted p&#x202f;<&#x202f;0.001-0.02) and tended to display higher proportions of participants in the high-adherence category, whereas lower adherence was observed in multiple non-Mediterranean and North African contexts. Psychological distress differed significantly between countries (p&#x202f;<&#x202f;0.001), with several contexts showing elevated depression, anxiety, or stress levels (z&#x202f;&#x2248;&#x202f;3.57-14.29, adjusted p&#x202f;<&#x202f;0.001-0.05). Life satisfaction and social participation also varied substantially (194.86, p&#x202f;<&#x202f;0.001), with some European countries reporting lower social participation compared with Mediterranean and neighboring contexts (z&#x202f;=&#x202f;3.79-9.31, adjusted p&#x202f;<&#x202f;0.001-0.05). Sleep parameters and insomnia severity differed markedly across countries (H&#x202f;=&#x202f;66.64-198.63, p&#x202f;<&#x202f;0.001), with less favourable sleep profiles observed in several contexts (z&#x202f;&#x2248;&#x202f;3.28-12.82, adjusted p&#x202f;<&#x202f;0.001-0.05). Physical activity and sedentary behaviour showed pronounced variability (p&#x202f;<&#x202f;0.001), with Jordan reporting the lowest physical activity levels and Tunisia lower sedentary time. Mediterranean lifestyle adherence and its psychosocial and behavioural correlates vary substantially across countries, reflecting distinct constellations of sociocultural, socioeconomic, and lifestyle factors rather than dietary patterns alone. These findings highlight the importance of multidimensional, context-sensitive approaches to Mediterranean lifestyle promotion and provide a descriptive framework to inform tailored public-health strategies and future longitudinal and intervention research.

Human gingival fibroblasts (HGFs) are stromal cells that maintain periodontal tissue structure and extracellular matrix (ECM) dynamics. ECM stiffness serves as a physical cue that regulates HGF behavior and secretory profiles. This study investigated how substrate stiffness modulates the secretome of HGFs and observed the subsequent effects of this secretome on the osteogenic differentiation of human periodontal ligament cells (HPDLCs). HGFs isolated from healthy donors were cultured on polydimethylsiloxane substrates, representing soft or hard periodontal tissue under normal and lipopolysaccharide (LPS)-induced inflammatory conditions. The expression of cytokines and chemokines was analyzed using qRT-PCR and ELISA, with p38 MAPK inhibitors used to identify stiffness-associated signaling involvement. HPDLCs were treated with conditioned medium from HGFs (HGF-CM) under osteogenic induction, osteogenic marker expression was examined using qRT-PCR and immunofluorescence, with mineralization assessed by Alizarin red S staining. To establish mechanistic causality, functional blocking was conducted using a C-X-C motif chemokine receptor 4 (CXCR4) inhibitor. Hard substrates significantly increased the expression of anti-inflammatory cytokines and the C-X-C motif chemokine ligand 12 (CXCL12) in HGFs, whereas inhibition of p38 mitogen-activated protein kinase (MAPK) activity attenuated stiffness-associated CXCL12 expression. Under LPS-induced inflammatory conditions, hard substrates-maintained matrix metalloproteinase-9 suppression and tissue inhibitor of metalloproteinases 1 upregulation, although CXCL12 protein levels were decreased. Furthermore, HPDLCs treated with HGF-CM derived from hard substrates in osteogenic induction media exhibited elevated CXCR4 expression, increased osteogenic marker levels at days 14 and 21, and enhanced mineral deposition compared to those treated with HGF-CM from soft substrates. In addition, functional blocking with a CXCR4 inhibitor significantly reduced the expression of osteogenic markers (ALP, RUNX2, COL1A1, and OSX) and confirmed a subsequent decrease in matrix mineralization. Substrate stiffness modulated the paracrine behavior of HGFs, with CXCL12 serving as a representative example of a stiffness-responsive factor. These alterations in the HGF-derived secretome were associated with altered osteogenic and inflammatory responses in HPDLCs. These findings support the influence of the physical microenvironment on fibroblast-periodontal ligament cell interactions on anti-inflammatory response and periodontal tissue stabilization.

Intimate partner violence (IPV) remains a significant public health concern in New York City (NYC), with psychological and physical abuse highly prevalent. Hispanic and Latine adults may face elevated IPV risk due to gender norms, immigration-related stressors, and barriers to accessing services. However, differences in IPV patterns by nativity remain insufficiently understood. Data from the 2020 NYC Community Health Survey, a population-based survey of non-institutionalized adults, were analyzed. The analytic sample included 2,229 Hispanic and Latine respondents. Weighted descriptive statistics and stratified logistic regression models were used to estimate lifetime physical and psychological IPV prevalence and associated factors by nativity. U.S.-born Hispanic and Latine (USBH/L) adults reported higher prevalence of IPV than foreign-born Hispanic and Latine (FBH/L) respondents. However, nativity was not independently associated with IPV after adjustment. Risk factors varied across nativity groups. Among FBH/L adults, female gender and marital disruption were the strongest and most consistent predictors of IPV. Among USBH/L adults, binge drinking, disability, and mental health treatment showed stronger and more consistent associations. Having &#x2265;2 sexual partners was associated with higher odds of IPV across models. Older age (&#x2265;65 years) was consistently associated with lower IPV odds. IPV remains prevalent among Hispanic and Latine adults in NYC, with distinct patterns of associated risk factors by nativity. Prevention strategies should be culturally and linguistically responsive and address gender norms, substance use, and structural vulnerabilities, including disability and relationship instability. These findings highlight the importance of continued IPV surveillance to inform equitable, population-level interventions.

Philadelphia-negative myeloproliferative neoplasms (MPNs), encompassing polycythemia vera, essential thrombocythemia, and primary myelofibrosis, represent a group of chronic clonal hematopoietic disorders characterized by excessive proliferation of myeloid lineages and with a propensity for arterial and venous thrombosis. These disorders provide a unique human model of inflammation-driven, clone-mediated vascular disease with mechanistic insights highly relevant to cardiovascular medicine. The interplay of elevated hematocrit, leukocytosis, activated platelets, neutrophil extracellular traps, endothelial dysfunction, and chronic cytokine elevation creates a multifaceted prothrombotic state. Randomized evidence, such as the CYTO-PV (cytoreductive therapy in polycythemia vera) trial, demonstrates that correcting hematocrit meaningfully reduces major cardiovascular and thrombotic events. Discoveries like Janus Kinase 2 valine-to phenylalanine substitution on position 617-mutated clones, altered platelet biology, and NETosis inform evolving concepts in coronary artery disease, microvascular ischemia, and inflammatory thrombosis. This review synthesizes thrombosis biology in MPNs with actionable implications for cardiologists. Key lessons include the importance of clonal hematopoiesis as a cardiovascular risk factor, the role of inflammation and hematologic dysregulation in accelerating atherothrombosis, and the therapeutic relevance of antiplatelet optimization, hematocrit control, cytoreductive therapy, and anti-inflammatory interventions. The review also outlines diagnostic situations in cardiology where MPNs should be suspected, proposes opportunities for improved risk stratification, and identifies research priorities bridging the 2 specialties.

The interplay between cells and their surrounding microenvironment drives multiple cellular functions, including migration, proliferation, and cell fate transitions. The nucleus is a mechanosensitive organelle; however, the morphological and functional changes of the nucleus induced by a three-dimensional (3D) extracellular environment remain unclear. Here, we report that leukemia Jurkat cells selected after 3D growth conditions retain persistent nuclear changes even after being released from confinement. These altered cells showed aberrant nuclear wrinkling, visualized by the lamin B1 distribution and mediated by disrupted actin dynamics and protein kinase C (PKC)&#x3b2; signaling. Moreover, these cells presented changes in chromatin compaction, transcription, apoptosis, and in vivo dissemination. By combining biomechanical techniques and single-nucleus analysis, we have determined that these cells exhibit a distinct nuclear mechanical behavior and biophysical signature compared with control cells. Together, these findings demonstrate that 3D microenvironments alter leukemia cell biology by promoting persistent changes in chromatin organization, morphology, and mechanical response of the nucleus.

Laccases are the most common multicopper oxidase (EC 1.10.3.2); as a result, they have attracted great interest as natural cleaners for environmental&#x2002;bioremediation purposes. However, the fragile structure and low recyclability of laccases, along with their high price, are constraints in practical applications. Enzyme immobilization has been identified as one of the best&#x2002;strategies to circumvent these problems and the application of nanomaterials for enzyme immobilization has resulted in nanobiocatalysts (NBCs). Nanomaterials have great advantages for the immobilization of enzymes attributed to their special chemical and physical properties, including large specific surface area, tunable pore size, and strong solid mechanical stability. With better stability, efficacy and specificity, these nano-engineered systems have been very successful&#x2002;in the treatment of a variety of pollutants ranging from persistent organic pollutants and emerging contaminations to industrial waste. Herein, we give an updated review of the most recent advances in laccase-based NBCs, with a particular focus on&#x2002;new strategies for nanomaterial functionalization and laccase enzyme immobilization. Additionally, this review aims to gather information about laccase's sources, mechanisms of action, substrates, and mediators, providing a useful point of reference. This review also compiles numerous recent discoveries about the biology of laccase and its applications as a nanobiocatalyst and provides a concise synopsis that aids researchers in comprehending its possibilities. It also emphasizes the possible use of laccase-based NBCs for bioremediation, especially in dealing with emerging pollutants (EPs) like antibiotics, pharmaceutical residues, textile effluents, and other xenobiotics.

Understanding how nanoparticles move near liquid-solid interfaces is central to nanoscale transport in catalysis, biology, and soft materials. Here, we uncover the physical mechanisms governing anomalous surface diffusion of PEG-coated gold nanorods (AuNRs) near the silicon nitride (SiNx) membrane in liquid-phase transmission electron microscopy (LPTEM). By systematically tuning the ionic environment (H2O, 5 mM H2SO4, 1.5 mM NaCl, 5 mM PBS), we show how electrostatic screening and ion-specific surface interactions modulate the interaction landscape, altering the strength and abundance of binding sites that govern the confinement and mobility of nanoparticles. Statistical analyses and deep learning classification of particle trajectories reveal a tunable transition between fractional Brownian motion (FBM) in strongly interacting systems (H2O, H2SO4) and annealed transient time motion (ATTM) in screened environments (NaCl, PBS). These results establish electrostatic screening and specific ion effects as external controls that program near-surface transport, shifting the diffusion mechanism from FBM to ATTM and tuning the particle mobility. To further elucidate the interfacial dynamics, we introduce a passive nanorheology framework in LPTEM, modeling the near-surface environment of FBM-classified conditions as an effective viscoelastic medium. Leveraging translational and rotational trajectories as nanoscale rheological probes, we reconstruct frequency-dependent viscoelastic moduli to extract relaxation times and elastic-to-viscous crossover moduli that report on interaction strength at the SiNx interface. Together, these advances provide both control and diagnosis of interfacial mechanical response in LPTEM, positioning it as a quantitative tool for probing nanoscale transport in complex soft-matter and interfacial systems.

Mechanical forces are fundamental drivers of cutaneous tissue repair; however, the molecular mechanisms translating physical cues into fibroblast responses remain incompletely understood. This review examines the Piezo1-PI3K/Akt signaling axis as a putative convergent context-dependent mechanotransduction node, through which multiple mechanical inputs may be transduced into intracellular biochemical cascades. Direct evidence in dermal fibroblasts is strongest for the integrin-FAK arm; we synthesize findings across cell types to propose a multi-pathway framework encompassing four molecular bridges: CaM/CaMK-mediated activation, a synergistic integrin-FAK loop, calpain-mediated regulation, and a Panx1-ATP-P2Y2 paracrine pathway. Beyond signal relay, this axis may function as a spatiotemporal encoder coordinating fibroblast proliferation, migration, metabolic reprogramming, and myofibroblast differentiation. By contrasting physiological healing with dysregulated mechanotransduction in chronic wounds and hypertrophic scars, we highlight the therapeutic relevance of this axis and propose that its modulation may offer a strategy to restore mechanical homeostasis and improve clinical outcomes.

Inherited retinal disorders such as Norrie disease lack effective therapies, largely owing to the challenges of delivering genes to retinal tissues via intravitreal injection. The efficacy of gene delivery is constrained by physical impediments, including the inner limiting membrane and the viscoelastic characteristics of the vitreous humor. In this study, we present the development of vitreous humor-mimetic liposomes (VMLs), which have been shown to facilitate efficient retinal gene delivery by enhancing biocompatibility and tissue absorption. The VMLs were engineered by reproducing the lipidomic composition of the native vitreous, as identified through liquid chromatography-mass spectrometry analysis. The subretinal injection of VMLs was conducted to assess the efficiency of gene expression delivery by VMLs. Notably, VML #3 demonstrated a 1.3-fold enhanced GFP transfection compared to conventional nanoparticle, Lipofectamine 2000. Following intravitreal administration, VML #3 facilitated efficient delivery of pGFP-NDP (Norrie disease protein) plasmids and restored retinal vascularization in both Ndp-hemizygous and oxygen-induced retinopathy mouse models. The findings demonstrate the potential of tissue-inspired nanocarriers as effective platforms for enhancing intraocular delivery, suggesting a clinically applicable strategy for the treatment of inherited retinal diseases, including Norrie disease.

Spasticity is an individual, sensory-motor experience that is correlated with nonmotor symptoms of pain, fatigue, and dysesthesia. Patients and clinicians may have different understandings of spasticity. This discrepancy is complicated because spasticity is highly variable depending on the individual and the unique stressors in their environment. Listening to how patients explain their symptoms and what factors make their spasticity better or worse is key to effective symptom management. Good patient-clinician communication leads to applying the correct treatment of each symptom. The use of oral medications, physical therapy, botulinum toxins, intrathecal baclofen therapy, and adjunctive therapies are reported through patient experience.

The aging population is retaining more natural teeth, increasing the prevalence of exposed root surfaces. These surfaces are prone to abrasion, erosion, and biofilm accumulation, leading to tissue loss, hypersensitivity, and root caries. The objectives of this study were to: (1) evaluate the surface and mechanical properties of experimental resin coatings containing dimethylaminohexadecyl methacrylate (DMAHDM) and nanoparticles of amorphous calcium phosphate (NACP) following simulated toothbrushing, and (2) assess their antibacterial efficacy and protective effects compared with commercial controls. Experimental groups with urethane dimethacrylate (UDMA)/ triethylene glycol divinylbenzyl ether (TEGDVBE)+&#x202f;5% DMAHDM+&#x202f;10% (EX1) or 20% NACP (EX2), along with commercial controls (Seal&Protect and Vanish XT), were tested. Sound and artificially-demineralized dentin served as controls. Hardness, surface roughness, surface loss, and water contact angle were assessed before and after 10,000 brushing cycles. Additionally, Streptococcus mutans (S. mutans) biofilm colony-forming units (CFU) was measured, and surface morphology was evaluated using SEM. EX2 and Seal&Protect had similar hardness (0.12&#x202f;&#xb1;&#x202f;0.01)GPa (n&#x202f;=&#x202f;5, p&#x202f;>&#x202f;0.05), while the hardness values of EX1 and demineralized dentin were lower (0.09&#x202f;&#xb1;&#x202f;0.01)GPa and (0.07&#x202f;&#xb1;&#x202f;0.01)GPa (p&#x202f;<&#x202f;0.05). After 10,000 cycles of brushing, EX1, EX2, and Seal&Protect maintained smooth surfaces (Ra=0.4-0.5&#x202f;&#xb5;m; n&#x202f;=&#x202f;6, p&#x202f;>&#x202f;0.05), whereas Vanish and uncoated dentin became rougher (Ra=2.0&#x202f;&#xb1;&#x202f;0.5&#x202f;&#xb5;m; p&#x202f;<&#x202f;0.05). Surface loss after 10,000 cycles was minimal for experimental coatings (0.39-0.55&#x202f;&#xb5;m) and comparable to Seal&Protect (2.63&#x202f;&#xb5;m), while Vanish (8.42&#x202f;&#xb5;m) and dentin controls (19-27&#x202f;&#xb5;m) showed significantly greater loss (p&#x202f;<&#x202f;0.05). Both experimental coatings reduced S. mutans biofilm CFU counts by 7&#x202f;log, compared to all other groups (p&#x202f;<&#x202f;0.05), while retaining hydrophobicity. SEM showed intact coatings in the experimental groups and in Seal&Protect, but Vanish and uncoated samples exhibited cracking and dentin exposure. The DMAHDM-NACP UDMA/TEGDVBE resin coating demonstrated surface and mechanical properties comparable to Seal&Protect and superior surface stability compared to Vanish XT, while achieving potent antibacterial activity not observed in either commercial control. These findings indicate strong potential for this multifunctional resin coating to protect tooth root dentin via physical shielding and sustained antibacterial performance.

Rodents remain the predominant mammalian species used for biomedical research and must be humanely killed upon completion of the scientific work. Across the UK, cervical dislocation is reported as the most common method for humanely killing laboratory rodents. Cervical dislocation involves the separation of the vertebrae at the top of the spine and can be achieved manually or mechanically (e.g. using a tool). There is no standardised method for achieving consistent cervical dislocation in the desired location, and no dedicated tool specifically designed, validated and commercially available to achieve accurate and effective dislocation. Previous work has highlighted inaccuracy in method application and, as such, has the potential to risk animal welfare at killing. The aim of this work was to identify the techniques used by personnel across UK institutions for performing cervical dislocation using an online questionnaire. We found marked inconsistencies in technique and the use of tools to aid the application. Mice are predominantly killed via manual operation (i.e. without the aid of a tool), while rats are more often killed using mechanical aids. A wide range of improvised tools (i.e. not designed for killing) were reported, including pens, scissors, and cage scrapers. Further, there was little or no consensus regarding which physical actions are essential for a successful dislocation (i.e. a stretch and/or a twist), and a lack of reported institutional standard operating procedures. Further work is needed to establish validated methods and clear standards to ensure this common method is applied humanely and consistently.

F&#xf6;rster Resonance Energy Transfer (FRET) is a powerful technique for the detection and characterization of biomolecular interactions and conformational changes with subnanometer spatial resolution and a temporal resolution down to the timescale of fluorescence. While the technique is widely adopted in structural biology and biophysics, the evolution of single-molecule FRET has led to experimental setups with sophisticated optical layouts, multilaser excitation schemes, and time-resolved detection electronics. We here present an accessible alternative toward single-molecule FRET based on Brick-MIC, a recently introduced three-dimensional (3D)-printed microspectroscopy platform. The FRET-Brick uses continuous-wave excitation at 488&#x2009;nm with a minimal set of optomechanical components and photomultiplier detectors (PMTs). With this, we were able to significantly reduce the setup complexity retaining single-molecule sensitivity with dyes matching the sensitivity of PMTs. To maximize the photon output of Alexa488, ATTO488 (donors), Alexa555, ATTO542, and Cy3B (acceptors), we introduce ferrocene derivatives as photostabilizers that increase both dye brightness and remove dark-states. We benchmark the performance of the FRET-Brick with fluorophore-labeled oligonucleotide reference structures also in comparison to accessible volume simulations, and by detecting conformational changes in bacterial substrate-binding proteins. Our work demonstrates that qualitative and quantitative single-molecule FRTE (smFRET) measurements are possible with the minimalistic and cost-effective FRET-Brick.

Human &#x3b2; defensin type 3 (hBD-3) is recognized as one of the most intriguing antimicrobial peptides (AMPs) that holds the promise of solving drug resistance issues. hBD-3 can function (disruption of membrane integrity) in high salt environments, where most other AMPs fail. However, its functional mechanism at the molecular level remains elusive. To characterize its structure and dynamics during membrane crossing, long-time (a total of 57.0 &#x3bc;s) all-atom molecular dynamics simulations were conducted on hBD-3 monomers and dimers in both wild-type and analog (in which all three disulfide bonds are broken) forms that are embedded in four types of lipid membranes. Trajectory analysis was carried out using a statistical method&#x2500;conformational dynamics analysis to calculate contact matrices and then principal component analysis (PCA) and linear discriminant analysis (LDA), in order to discern structural changes upon various physical and chemical perturbations. The result shows that the major collective coordinate primarily distinguishes between the wild-type and analog forms of hBD-3. For the hBD-3 monomer, the analog undergoes significant structural loss due to the lack of stabilizing disulfide bonds; salt exerts a nearly consistent effect on the contact degrees of freedom of the protein, whereas changes in lipid membrane composition have an insignificant effect. For the hBD-3 dimer, no consistent relationship between structure and salt concentration is indicated, and variations in the chemical composition of model bacterial membranes have a limited effect on its dynamics. These results suggest that the wild-type and analog forms of hBD-3 may employ different mechanisms when crossing bacterial membranes. The effect of salt on hBD-3 dynamics can be mitigated by the high net charge density of the protein. Additionally, the hBD-3 dimer can distinguish between model Gram-positive and Gram-negative membranes, whereas the monomer cannot. Overall, these findings provide unique insights into the structure, dynamics, and membrane-disrupting mechanism of hBD-3.