Nuclear size-regulated emergence of topological packing order on growing human lung alveolospheres.

Within multicellular living systems, cells coordinate their positions with spatiotemporal accuracy to form various structures, setting the clock to control developmental processes and trigger maturation. These arrangements can be regulated by tissue topology, biochemical cues, as well as mechanical perturbations. However, the fundamental rules of how local cell packing order is regulated in forming three-dimensional (3D) multicellular architectures remain unclear. Furthermore, how cellular coordination evolves during developmental processes, and whether this cell patterning behavior is indicative of more complex biological functions, is largely unknown. Here, using human lung alveolospheres as a model system, by combining experiments and numerical simulations, we find that, surprisingly, cell packing behavior on alveolospheres resembles hard-disk packing but with increased randomness; the stiffer cell nuclei act as the hard disks surrounded by deformable cell bodies. Interestingly, we observe the emergence of topological packing order during alveolosphere growth, as a result of increasing nucleus-to-cell size ratio. Specifically, we find more hexagon-concentrated cellular packing with increasing bond orientational order, indicating a topological gas-to-liquid transition. Additionally, by osmotically changing the compactness of cells on alveolospheres, we observe that the variations in packing order align with the change of nucleus-to-cell size ratio. Together, our findings reveal the underlying rules of cell coordination and topological phases during human lung alveolosphere growth. These static packing characteristics are consistent with cell dynamics, together suggesting that better cellular packing stabilizes local cell neighborhoods and may regulate more complex biological functions such as organ development and cellular maturation.

Matrix confinement modulates 3D spheroid sorting and burst-like collective migration.

While it is known that cells with differential adhesion tend to segregate and preferentially sort, the physical forces governing sorting and invasion in heterogeneous tumors remain poorly understood. To investigate this, we tune matrix confinement, mimicking changes in the stiffness and confinement of the tumor microenvironment, to explore how physical confinement influences individual and collective cell migration in 3D spheroids. High levels of confinement lead to cell sorting while reducing matrix confinement triggers the collective fluidization of cell motion. Cell sorting, which depends on cell-cell adhesion, is crucial to this phenomenon. Burst-like migration does not occur for spheroids that have not undergone sorting, regardless of the degree of matrix confinement. Using computational Self-Propelled Voronoi modeling, we show that spheroid sorting and invasion into the matrix depend on the balance between cell-generated forces and matrix resistance. The findings support a model where matrix confinement modulates 3D spheroid sorting and unjamming in an adhesion-dependent manner, providing insights into the mechanisms of cell sorting and migration in the primary tumor and toward distant metastatic sites. STATEMENT OF SIGNIFICANCE: The mechanical properties of the tumor microenvironment significantly influence cancer cell migration within the primary tumor, yet how these properties affect intercellular interactions in heterogeneous tumors is not well understood. By utilizing calcium and calcium chelators, we dynamically alter collagen-alginate hydrogel stiffness and investigate tumor cell behavior within co-culture spheroids in response to varying degrees of matrix confinement. High confinement is found to trigger cell sorting while reducing confinement for sorted spheroids facilitates collective cell invasion. Notably, without prior sorting, spheroids do not exhibit burst-like migration, regardless of confinement levels. This work establishes that matrix confinement and intercellular adhesion regulate 3D spheroid dynamics, offering insights into cellular organization and migration within the primary tumor.

Effect of non-linear strain stiffening in eDAH and unjamming.

In cell clusters, the prominent factors at play encompass contractility-based enhanced tissue surface tension and cell unjamming transition. The former effect pertains to the boundary effect, while the latter constitutes a bulk effect. Both effects share outcomes of inducing significant elongation in cells. This elongation is so substantial that it surpasses the limits of linear elasticity, thereby giving rise to additional effects. To investigate these effects, we employ atomic force microscopy (AFM) to analyze how the mechanical properties of individual cells change under such considerable elongation. Our selection of cell lines includes MCF-10A, chosen for its pronounced demonstration of the extended differential adhesion hypothesis (eDAH), and MDA-MB-436, selected due to its manifestation of cell unjamming behavior. In the AFM analyses, we observe a common trend in both cases: as elongation increases, both cell lines exhibit strain stiffening. Notably, this effect is more prominent in MCF-10A compared to MDA-MB-436. Subsequently, we employ AFM on a dynamic range of 1-200 Hz to probe the mechanical characteristics of cell spheroids, focusing on both surface and bulk mechanics. Our findings align with the results from single cell investigations. Specifically, MCF-10A cells, characterized by strong contractile tissue tension, exhibit the greatest stiffness on their surface. Conversely, MDA-MB-436 cells, which experience significant elongation, showcase their highest stiffness within the bulk region. Consequently, the concept of single cell strain stiffening emerges as a crucial element in understanding the mechanics of multicellular spheroids (MCSs), even in the case of MDA-MB-436 cells, which are comparatively softer in nature.

Network physics of attractive colloidal gels: Resilience, rigidity, and phase diagram.

Colloidal gels exhibit solid-like behavior at vanishingly small fractions of solids, owing to ramified space-spanning networks that form due to particle-particle interactions. These networks give the gel its rigidity, and with stronger attractions the elasticity grows as well. The emergence of rigidity can be described through a mean field approach; nonetheless, fundamental understanding of how rigidity varies in gels of different attractions is lacking. Moreover, recovering an accurate gelation phase diagram based on the system's variables has been an extremely challenging task. Understanding the nature of colloidal clusters, and how rigidity emerges from their connections is key to controlling and designing gels with desirable properties. Here, we employ network analysis tools to interrogate and characterize the colloidal structures. We construct a particle-level network, having all the spatial coordinates of colloids with different attraction levels, and also identify polydisperse rigid fractal clusters using a Gaussian mixture model, to form a coarse-grained cluster network that distinctly shows main physical features of the colloidal gels. A simple mass-spring model then is used to recover quantitatively the elasticity of colloidal gels from these cluster networks. Interrogating the resilience of these gel networks shows that the elasticity of a gel (a dynamic property) is directly correlated to its cluster network's resilience (a static measure). Finally, we use the resilience investigations to devise [and experimentally validate] a fully resolved phase diagram for colloidal gelation, with a clear solid-liquid phase boundary using a single volume fraction of particles well beyond this phase boundary.

Matrix confinement modulates 3D spheroid sorting and burst-like collective migration.

While it is known that cells with differential adhesion tend to segregate and preferentially sort, the physical forces governing sorting and invasion in heterogeneous tumors remain poorly understood. To investigate this, we tune matrix confinement, mimicking changes in the stiffness and confinement of the tumor microenvironment, to explore how physical confinement influences individual and collective cell migration in 3D spheroids. High levels of confinement lead to cell sorting while reducing matrix confinement triggers the collective fluidization of cell motion. Cell sorting, which depends on cell-cell adhesion, is crucial to this phenomenon. Burst-like migration does not occur for spheroids that have not undergone sorting, regardless of the degree of matrix confinement. Using computational Self-Propelled Voronoi modeling, we show that spheroid sorting and invasion into the matrix depend on the balance between cell-generated forces and matrix resistance. The findings support a model where matrix confinement modulates 3D spheroid sorting and unjamming in an adhesion-dependent manner, providing insights into the mechanisms of cell sorting and migration in the primary tumor and toward distant metastatic sites.

Mechanical imbalance between normal and cancer cells drives epithelial defense against cancer.

Cell competition enables normal wildtype cells of epithelial tissue to eliminate mutant cells expressing activated oncoproteins such as HRasV12. However, the driving force behind this fundamental epithelial defense against cancer remains enigmatic. Here, we employ tissue stress microscopy and theoretical modeling and invent a new collective compressibility measurement technique called gel compression microscopy to unveil the mechanism governing cell competition. Stress microscopy reveals unique compressive stress experienced by the mutant cells, contrasting with predominantly tensile stress experienced by normal cells. A cell-based computer simulation then predicts that this compressive stress arises out of a mechanical imbalance between two competing populations due to a difference in their collective compressibility and rigidity. Gel compression microscopy empirically confirms the prediction and elucidates a three-fold higher compressibility of the mutant population than the normal population. Mechanistically, this difference stems from the reduced abundance and coupling of junctional E-cadherin molecules in the mutant cells, which weakens cell-cell adhesions and renders the mutant population more compressible. Taken together, our study elucidates both the physical principle and the underlying molecular mechanism driving cell competition in epithelial defense against cancer and opens new directions for mechanomedicine in cancer.

Constitutive model for the rheology of biological tissue.

The rheology of biological tissue is key to processes such as embryo development, wound healing, and cancer metastasis. Vertex models of confluent tissue monolayers have uncovered a spontaneous liquid-solid transition tuned by cell shape; and a shear-induced solidification transition of an initially liquidlike tissue. Alongside this jamming/unjamming behavior, biological tissue also displays an inherent viscoelasticity, with a slow time and rate-dependent mechanics. With this motivation, we combine simulations and continuum theory to examine the rheology of the vertex model in nonlinear shear across a full range of shear rates from quastistatic to fast, elucidating its nonlinear stress-strain curves after the inception of shear of finite rate, and its steady state flow curves of stress as a function of strain rate. We formulate a rheological constitutive model that couples cell shape to flow and captures both the tissue solid-liquid transition and its rich linear and nonlinear rheology.

Nature-inspired designs for disordered acoustic bandgap materials.

We introduce an amorphous mechanical metamaterial inspired by how cells pack in biological tissues. The spatial heterogeneity in the local stiffness of these materials has been recently shown to impact the mechanics of confluent biological tissues and cancer tumor invasion. Here we use this bio-inspired structure as a design template to construct mechanical metamaterials and show that this heterogeneity can give rise to amorphous cellular solids with large, tunable acoustic bandgaps. Unlike acoustic crystals with periodic structures, the bandgaps here are directionally isotropic and robust to defects due to their complete lack of positional order. Possible ways to manipulate bandgaps are explored with a combination of the tissue-level elastic modulus and local stiffness heterogeneity of cells. To further demonstrate the existence of bandgaps, we dynamically perturb the system with an external sinusoidal wave in the perpendicular and horizontal directions. The transmission coefficients are calculated and show valleys that coincide with the location of bandgaps. Experimentally this design should lead to the engineering of self-assembled rigid acoustic structures with full bandgaps that can be controlled via mechanical tuning and promote applications in a broad area from vibration isolations to mechanical waveguides.

Bridging the gap between collective motility and epithelial-mesenchymal transitions through the active finite voronoi model.

We introduce an active version of the recently proposed finite Voronoi model of epithelial tissue. The resultant Active Finite Voronoi (AFV) model enables the study of both confluent and non-confluent geometries and transitions between them, in the presence of active cells. Our study identifies six distinct phases, characterized by aggregation-segregation, dynamical jamming-unjamming, and epithelial-mesenchymal transitions (EMT), thereby extending the behavior beyond that observed in previously studied vertex-based models. The AFV model with rich phase diagram provides a cohesive framework that unifies the well-observed progression to collective motility via unjamming with the intricate dynamics enabled by EMT. This approach should prove useful for challenges in developmental biology systems as well as the complex context of cancer metastasis. The simulation code is also provided.

Discontinuous rigidity transition associated with shear jamming in granular simulations.

We investigate the rigidity transition associated with shear jamming in frictionless, as well as frictional, disk packings in the quasi-static regime and at low shear rates. For frictionless disks, the transition under quasi-static shear is discontinuous, with an instantaneous emergence of a system spanning rigid clusters at the jamming transition. For frictional systems, the transition appears continuous for finite shear rates, but becomes sharper for lower shear rates. In the quasi-static limit, it is discontinuous as in the frictionless case. Thus, our results show that the rigidity transition associated with shear jamming is discontinuous, as demonstrated in the past for isotropic jamming of frictionless particles, and therefore a unifying feature of the jamming transition in general.

Epithelial layer fluidization by curvature-induced unjamming.

The transition of an epithelial layer from a stationary, quiescent state to a highly migratory, dynamic state is required for wound healing, development, and regeneration. This transition, known as the unjamming transition (UJT), is responsible for epithelial fluidization and collective migration. Previous theoretical models have primarily focused on the UJT in flat epithelial layers, neglecting the effects of strong surface curvature that is characteristic of epithelial tissues in vivo. In this study, we investigate the role of surface curvature on tissue plasticity and cellular migration using a vertex model embedded on a spherical surface. Our findings reveal that increasing curvature promotes epithelial unjamming by reducing the energy barriers to cellular rearrangements. Higher curvature favors cell intercalation, mobility, and self-diffusivity, resulting in epithelial structures that are malleable and migratory when small, but become more rigid and stationary as they grow. As such, curvature-induced unjamming emerges as a novel mechanism for epithelial layer fluidization. Our quantitative model proposes the existence of a new, extended, phase diagram wherein local cell shape, cell propulsion, and tissue geometry combine to determine the epithelial migratory phenotype.

Compressive stress drives adhesion-dependent unjamming transitions in breast cancer cell migration.

Cellular unjamming is the collective fluidization of cell motion and has been linked to many biological processes, including development, wound repair, and tumor growth. In tumor growth, the uncontrolled proliferation of cancer cells in a confined space generates mechanical compressive stress. However, because multiple cellular and molecular mechanisms may be operating simultaneously, the role of compressive stress in unjamming transitions during cancer progression remains unknown. Here, we investigate which mechanism dominates in a dense, mechanically stressed monolayer. We find that long-term mechanical compression triggers cell arrest in benign epithelial cells and enhances cancer cell migration in transitions correlated with cell shape, leading us to examine the contributions of cell-cell adhesion and substrate traction in unjamming transitions. We show that cadherin-mediated cell-cell adhesion regulates differential cellular responses to compressive stress and is an important driver of unjamming in stressed monolayers. Importantly, compressive stress does not induce the epithelial-mesenchymal transition in unjammed cells. Furthermore, traction force microscopy reveals the attenuation of traction stresses in compressed cells within the bulk monolayer regardless of cell type and motility. As traction within the bulk monolayer decreases with compressive pressure, cancer cells at the leading edge of the cell layer exhibit sustained traction under compression. Together, strengthened intercellular adhesion and attenuation of traction forces within the bulk cell sheet under compression lead to fluidization of the cell layer and may impact collective cell motion in tumor development and breast cancer progression.

Shear-Driven Solidification and Nonlinear Elasticity in Epithelial Tissues.

Biological processes, from morphogenesis to tumor invasion, spontaneously generate shear stresses inside living tissue. The mechanisms that govern the transmission of mechanical forces in epithelia and the collective response of the tissue to bulk shear deformations remain, however, poorly understood. Using a minimal cell-based computational model, we investigate the constitutive relation of confluent tissues under simple shear deformation. We show that an initially undeformed fluidlike tissue acquires finite rigidity above a critical applied strain. This is akin to the shear-driven rigidity observed in other soft matter systems. Interestingly, shear-driven rigidity can be understood by a critical scaling analysis in the vicinity of the second order critical point that governs the liquid-solid transition of the undeformed system. We further show that a solidlike tissue responds linearly only to small strains and but then switches to a nonlinear response at larger stains, with substantial stiffening. Finally, we propose a mean-field formulation for cells under shear that offers a simple physical explanation of shear-driven rigidity and nonlinear response in a tissue.

Configurational fingerprints of multicellular living systems.

Cells cooperate as groups to achieve structure and function at the tissue level, during which specific material characteristics emerge. Analogous to phase transitions in classical physics, transformations in the material characteristics of multicellular assemblies are essential for a variety of vital processes including morphogenesis, wound healing, and cancer. In this work, we develop configurational fingerprints of particulate and multicellular assemblies and extract volumetric and shear order parameters based on this fingerprint to quantify the system disorder. Theoretically, these two parameters form a complete and unique pair of signatures for the structural disorder of a multicellular system. The evolution of these two order parameters offers a robust and experimentally accessible way to map the phase transitions in expanding cell monolayers and during embryogenesis and invasion of epithelial spheroids.

Weighting of toilet assessment scheme in China implementing analytic hierarchy process.

Advancing toilet technologies to address public health and sanitation issues are a concern of governments and organizations. This article mainly studies the assessment methods for the public toilets and some rural toilets considering from design to demolition to assist for the innovation of toilet technologies. The Analytic Hierarchy Process (AHP) and Life Cycle Assessment (LCA) methods were adopted to identify the assessment indicators and rank the weight. The outcome of Toilet Assessment Scheme (TAS), which includes a set of weightings and a classification system for the selected assessment indicators and sub-indicators. The weight calculation result showed that water resources, ecology, and indoor environmental quality are relatively high, which indicates that saving water, protecting the environment and optimizing the toilet environment should be given priority at the current stage. The individual questionnaire experts from the perspective of gender, profession, and generation, have different emphases on the evaluation scheme. This study can improve the comprehensiveness of toilet evaluation under the distinct background conditions, and will play a relevant role in the promotion of new toilet technology. The TAS can accelerate the toilet revolution in areas where toilets are scarce, and thus will improve the sanitary and health conditions of these populations.

LRRC75A-AS1 targets miR-199b-5p/PDCD4 axis to repress multiple myeloma.

BACKGROUND: Multiple functions of miR-199b-5p in diseases have been demonstrated by existing studies. However, never has the correlation between miR-199b-5p and multiple myeloma (MM) been established. METHODS: qRT-PCR analyzed RNA expression and western blot measured protein expression. Cell proliferation ability was tested via colony formation and EdU assays, and apoptosis was determined via TUNEL, flow cytometry and detection of apoptosis-related proteins. Position of LRRC75A antisense RNA 1 (LRRC75A-AS1) was recognized by FISH assay. RIP, RNA pull-down and luciferase reporter experiments explored the molecular interplay. RESULTS: GEO (Gene Expression Omnibus) data revealed miR-199b-5p upregulation in MM specimens, and qRT-PCR data verified miR-199b-5p upregulation in MM cells. Inhibiting miR-199b-5p markedly impeded MM cell proliferation and stimulated apoptosis. Moreover, we demonstrated the mechanism that miR-199b-5p was decoyed by LRRC75A-AS1 and miR-199b-5p targeted programmed cell death 4 (PDCD4) to repress its expression. Further, LRRC75A-AS1 was verified to repress proliferation and prompt apoptosis in a PDCD4-dependent way in MM cells. CONCLUSION: Our data displayed that miR-199b-5p was sequestered by LRRC75A-AS1 so that PDCD4 was released to repress MM, implying the targeting miR-199b-5p as a novel thought for improving MM therapy.

In primary airway epithelial cells, the unjamming transition is distinct from the epithelial-to-mesenchymal transition.

The epithelial-to-mesenchymal transition (EMT) and the unjamming transition (UJT) each comprises a gateway to cellular migration, plasticity and remodeling, but the extent to which these core programs are distinct, overlapping, or identical has remained undefined. Here, we triggered partial EMT (pEMT) or UJT in differentiated primary human bronchial epithelial cells. After triggering UJT, cell-cell junctions, apico-basal polarity, and barrier function remain intact, cells elongate and align into cooperative migratory packs, and mesenchymal markers of EMT remain unapparent. After triggering pEMT these and other metrics of UJT versus pEMT diverge. A computational model attributes effects of pEMT mainly to diminished junctional tension but attributes those of UJT mainly to augmented cellular propulsion. Through the actions of UJT and pEMT working independently, sequentially, or interactively, those tissues that are subject to development, injury, or disease become endowed with rich mechanisms for cellular migration, plasticity, self-repair, and regeneration.

[Fibular osteotomy and distal tibiofibular joint fusion for treatment of chronic valgus Pilon fracture malunion].

OBJECTIVE: To explore clinical effects of fibular osteotomy and distal tibiofibular joint fusion for chronic valgus Pilon fracture malunion. METHODS: From January 2014 to January 2017, 8 patients with chronic valgus Pilon fracture malunion were treated, including 7 males and 1 female, aged from 20 to 47 years old, 6 patients on the left side and 2 patients on the right side; according to Rüedi-Allgöwer classification, 1 case was typeⅠ, 3 cases were typeⅡand 4 cases were type Ⅲ; the time from injury to admission ranged from 7 to 21 months. All deformities were evaluated individually based on pre-operatively weight bearing X-ray and 3D CT scan, and 3D printing model was also used for preliminary surgery. Weight-bearing X-ray showed posterior subluxation of ankle joint in 5 cases. There were 5 cases of fibular fracture at primary injury, and 2 cases of fibular fracture malunion. Fibular osteotomy and distal tibiofibular syndesmosis fusion strategy was performed to reduce articular surface congruency and correct lower limb alignment. Postoperative complication, fracture healing time and reduction were regularly recorded. Clinical function was evaluated according to American Orthopedic Foot and Ankle Society (AOFAS) at 1 year after operation. RESULTS: All patients were followed up from 12 to 30 months. All incisions were primarily healed. No infection, neurovascular injuries or implant failure, lost of reduction occurred. Fracture healing time ranged from 13 to 19 weeks with good lower limb alignment. AOFAS score at 1 year after operation was 63 to 90 points, 1 patient got excellent result, 4 good and 3 fair. Seven patients returned to work at 6 to 14 months after opertaion. CONCLUSION: For chronic valgus Pilon fractures malunion, fibular osteotomy and distal tibiofibular syndesmosis fusion could effectively restore congruency and alignment, correct lower limb alignment, improve ankle joint function, reduce occurrence of complication, and receive short term clinical effects.

Irradiation Induces Epithelial Cell Unjamming.

The healthy and mature epithelial layer is ordinarily quiescent, non-migratory, solid-like, and jammed. However, in a variety of circumstances the layer transitions to a phase that is dynamic, migratory, fluid-like and unjammed. This has been demonstrated in the developing embryo, the developing avian airway, the epithelial layer reconstituted in vitro from asthmatic donors, wounding, and exposure to mechanical stress. Here we examine the extent to which ionizing radiation might similarly provoke epithelial layer unjamming. We exposed primary human bronchial epithelial (HBE) cells maintained in air-liquid interface (ALI) to sub-therapeutic doses (1 Gy) of ionizing radiation (IR). We first assessed: (1) DNA damage by measuring p-H2AX, (2) the integrity of the epithelial layer by measuring transepithelial electrical resistance (TEER), and (3) the extent of epithelial cell differentiation by detecting markers of differentiated airway epithelial cells. As expected, IR exposure induced DNA damage but, surprisingly, disrupted neither normal differentiation nor the integrity of the epithelial cell layer. We then measured cell shape and cellular migration to determine the extent of the unjamming transition (UJT). IR caused cell shape elongation and increased cellular motility, both of which are hallmarks of the UJT as previously confirmed. To understand the mechanism of IR-induced UJT, we inhibited TGF-ß receptor activity, and found that migratory responses were attenuated. Together, these observations show that IR can provoke epithelial layer unjamming in a TGF-ß receptor-dependent manner.

Unjamming and collective migration in MCF10A breast cancer cell lines.

Each cell comprising an intact, healthy, confluent epithelial layer ordinarily remains sedentary, firmly adherent to and caged by its neighbors, and thus defines an elemental constituent of a solid-like cellular collective [1,2]. After malignant transformation, however, the cellular collective can become fluid-like and migratory, as evidenced by collective motions that arise in characteristic swirls, strands, ducts, sheets, or clusters [3,4]. To transition from a solid-like to a fluid-like phase and thereafter to migrate collectively, it has been recently argued that cells comprising the disordered but confluent epithelial collective can undergo changes of cell shape so as to overcome geometric constraints attributable to the newly discovered phenomenon of cell jamming and the associated unjamming transition (UJT) [1,2,5-9]. Relevance of the jamming concept to carcinoma cells lines of graded degrees of invasive potential has never been investigated, however. Using classical in vitro cultures of six breast cancer model systems, here we investigate structural and dynamical signatures of cell jamming, and the relationship between them [1,2,10,11]. In order of roughly increasing invasive potential as previously reported, model systems examined included MCF10A, MCF10A.Vector; MCF10A.14-3-3ζ; MCF10.ErbB2, MCF10AT; and MCF10CA1a [12-15]. Migratory speed depended on the particular cell line. Unsurprisingly, for example, the MCF10CA1a cell line exhibited much faster migratory speed relative to the others. But unexpectedly, across different cell lines higher speeds were associated with enhanced size of cooperative cell packs in a manner reminiscent of a peloton [9]. Nevertheless, within each of the cell lines evaluated, cell shape and shape variability from cell-to-cell conformed with predicted structural signatures of cell layer unjamming [1]. Moreover, both structure and migratory dynamics were compatible with previous theoretical descriptions of the cell jamming mechanism [2,10,11,16,17]. As such, these findings demonstrate the richness of the cell jamming mechanism, which is now seen to apply across these cancer cell lines but remains poorly understood.