Pseudopod Tracking and Statistics During Cell Movement in Buffer and Chemotaxis.

Amoeboid cells such as the protist Dictyostelium, human neutrophils, and the fungus B.d. chytrid move by extending pseudopods. The trajectories of cell movement depend on the size, rhythm, and direction of long series of pseudopods. These pseudopod properties are regulated by internal factors such as memory of previous directions and by external factors such as gradients of chemoattractants or electric currents. Here a simple method is described that defines the X, Y time coordinates of a pseudopod at the start and the end of the extension phase. The connection between the start and end of an extending pseudopod defines a vector, which is the input of different levels of analysis that defines cell movement. The primary information of the vector is its spatial length (pseudopod size), temporal length (extension time), extension rate (size divided by time), and direction. The second layer of information describes the sequence of two (or more) pseudopods: the direction of the second pseudopod relative to the direction of the first pseudopod, the start of the second pseudopod relative to the extension phase of the first pseudopod (the second starts while the first is still extending or after the first has stopped), and the alternating right/left extension of pseudopods. The third layer of information is provided by specific and detailed statistical analysis of these data and addresses question such as: is pseudopod extension in buffer in random direction or has the system internal directional memory, and how do shallow external electrical or chemical gradients bias the intrinsic pseudopod extension. The method is described for Dictyostelium, but has been used successfully for fast-moving neutrophils, slow-moving stem cells, and the fungus B.d. chytrid.

Generation of a Virtual Cell using a Phase Field Approach to Model Amoeboid Crawling.

The process of chemotaxis of living cells is complex. Cells follow gradients of an external signal because the interior of the cells gets polarized. The description of the exterior and the interior of the cell together with its motion for the convenient realization of the computational modeling of the whole process is a complex technical problem. Here, we employ a phase field model to characterize the interior of the cell, permitting the integration of stochastic partial differential equations, responsible for the polarization in the interior of the cell, and simultaneously, the calculation of the shape deformations of the cell, including its locomotion. We detail the mathematical description of the process and the procedure to calculate numerically the phase field with a simple reaction-diffusion equation for a single concentration.

Confined migration: Microtubules control the cell rear.

Cell migration through complex 3D environments relies on the interplay between actin and microtubules. A new study shows that, when cells pass through narrow constrictions, CLASP-dependent microtubule stabilisation at the cell rear controls actomyosin contractility to enable nuclear translocation and preserve cell integrity.

MTF2 facilitates the advancement of osteosarcoma through mediating EZH2/SFRP1/Wnt signaling.

BACKGROUND: Osteosarcoma is a soft tissue neoplasm with elevated recurrence risk and highly metastatic potential. Metal response element binding transcriptional factor 2 (MTF2) has been revealed to exert multiple activities in human tissues. The present research was conducted to explore the functions and related response mechanism of MTF2 in osteosarcoma which have not been introduced yet. METHODS: Bioinformatics tools identified the differential MTF2 expression in osteosarcoma tissues. MTF2 expression in osteosarcoma cells was examined with Western blot. Cell Counting Kit-8 (CCK-8) assay, 5-Ethynyl-2'-deoxyuridine (EDU) staining, wound healing as well as transwell assays measured cell proliferation, migration and invasion, respectively. Flow cytometry assay detected the cellular apoptotic level. Western blot also measured the expressions of proteins associated with epithelial mesenchymal transition (EMT), apoptosis and enhancer of zeste homolog 2 (EZH2)/secreted frizzled-related protein 1 (SFRP1)/Wnt signaling. Co-immunoprecipitation (Co-IP) assay confirmed MTF2-EZH2 interaction. RESULTS: MTF2 expression was increased in osteosarcoma tissues and cells. MTF2 interference effectively inhibited the proliferation, migration and invasion of osteosarcoma cells and promoted the cellular apoptotic rate. MTF2 directly bound to EZH2 and MTF2 silence reduced EZH2 expression, activated SFRP1 expression and blocked Wnt signaling in osteosarcoma cells. EZH2 upregulation or SFRP1 antagonist WAY-316606 partly counteracted the impacts of MTF2 down-regulation on the SFRP1/Wnt signaling and the biological phenotypes of osteosarcoma cells. CONCLUSIONS: MTF2 might down-regulate SFRP1 to activate Wnt signaling and drive the progression of osteosarcoma via interaction with EZH2 protein.

CALU promotes lung adenocarcinoma progression by enhancing cell proliferation, migration and invasion.

BACKGROUND: Lung cancer is the second most common cancer with the highest mortality in the world. Calumenin as a molecular chaperone that not only binds various proteins within the endoplasmic reticulum but also plays crucial roles in diverse processes associated with tumor development. However, the regulatory mechanism of calumenin in lung adenocarcinoma remains elusive. Here, we studied the impact of calumenin on lung adenocarcinoma and explored possible mechanisms. METHODS: 5-ethynyl-2'-deoxyuridine assay, colony formation, transwell and wound healing assays were performed to explore the effects of calumenin on the proliferation and migration of lung adenocarcinoma cells. To gain insights into the underlying mechanisms through which calumenin knockdown inhibits the migration and proliferation of lung adenocarcinoma, we performed Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, Gene Set Enrichment Analysis and Ingenuity Pathway Analysis based on transcriptomics by comparing calumenin knockdown with normal A549 cells. RESULTS: The mRNA and protein levels of calumenin in lung adenocarcinoma are highly expressed and they are related to an unfavorable prognosis in this disease. Calumenin enhances the proliferation and migration of A549 and H1299 cells. Gene Set Enrichment Analysis revealed that knockdown of calumenin in A549 cells significantly inhibited MYC and V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog signaling pathways while activating interferon signals, inflammatory signals, and p53 pathways. Ingenuity pathway analysis provided additional insights, indicating that the interferon and inflammatory pathways were prominently activated upon calumenin knockdown in A549 cells. CONCLUSIONS: The anti-cancer mechanism of calumenin knockdown might be related to the inhibition of MYC and KRAS signals but the activation of interferon signals, inflammatory signals and p53 pathways.

Compressed intracellular motility via non-uniform temporal sampling in dynamic optical coherence tomography.

Significance: Optical coherence tomography has great utility for capturing dynamic processes, but such applications are particularly data-intensive. Samples such as biological tissues exhibit temporal features at varying time scales, which makes data reduction challenging. Aim: We propose a method for capturing short- and long-term correlations of a sample in a compressed way using non-uniform temporal sampling to reduce scan time and memory overhead. Approach: The proposed method separates the relative contributions of white noise, fluctuating features, and stationary features. The method is demonstrated on mammary epithelial cell spheroids in three-dimensional culture for capturing intracellular motility without loss of signal integrity. Results: Results show that the spatial patterns of motility are preserved and that hypothesis tests of spheroids treated with blebbistatin, a motor protein inhibitor, are unchanged with up to eightfold compression. Conclusions: The ability to measure short- and long-term correlations compressively will enable new applications in (3+1)D imaging and high-throughput screening.

[Formation, organization and function of invadosomes in cell motility and tumor invasion]. / Invadosomes - Entre mobilité et invasion, naviguer dans la dualité des fonctions cellulaires.

Invadosome is an umbrella term used to describe a family of cellular structures including podosomes and invadopodia. They serve as contact zones between the cell plasma membrane and extracellular matrix, contributing to matrix remodeling by locally enriched proteolytic enzymes. Invadosomes, which are actin-dependent, are implicated in cellular processes promoting adhesion, migration, and invasion. Invadosomes, which exist in various cell types, play crucial roles in physiological phenomena such as vascularization and bone resorption. Invadosomes are also implicated in pathological processes such as matrix tissue remodeling during metastatic tumor cell invasion. This review summarizes basic information and recent advances about mechanisms underlying podosome and invadopodia formation, their organization and function.

Title: Invadosomes - Entre mobilité et invasion, naviguer dans la dualité des fonctions cellulaires. Abstract: Le terme « invadosome ¼ désigne une famille de structures cellulaires, comprenant les podosomes et les invadopodes, qui constituent des zones de contact entre la membrane plasmique des cellules et la matrice extracellulaire. Ces structures contribuent au remodelage de la matrice grâce à un enrichissement local en enzymes protéolytiques qui dégradent ses constituants fibrillaires. Les invadosomes, présents dans des types cellulaires variés, contribuent à des processus physiologiques, tels que la vascularisation, ou pathologiques, comme l'invasion des tissus par les cellules métastatiques.

Novel hypoxia-induced HIF-1αactivation in asthma pathogenesis.

BACKGROUND: Asthma's complexity, marked by airway inflammation and remodeling, is influenced by hypoxic conditions. This study focuses on the role of Hypoxia-Inducible Factor-1 Alpha (HIF-1α) and P53 ubiquitination in asthma exacerbation. METHODS: High-throughput sequencing and bioinformatics were used to identify genes associated with asthma progression, with an emphasis on GO and KEGG pathway analyses. An asthma mouse model was developed, and airway smooth muscle cells (ASMCs) were isolated to create an in vitro hypoxia model. Cell viability, proliferation, migration, and apoptosis were assessed, along with ELISA and Hematoxylin and Eosin (H&E) staining. RESULTS: A notable increase in HIF-1α was observed in both in vivo and in vitro asthma models. HIF-1α upregulation enhanced ASMCs' viability, proliferation, and migration, while reducing apoptosis, primarily via the promotion of P53 ubiquitination through MDM2. In vivo studies showed increased inflammatory cell infiltration and airway structural changes, which were mitigated by the inhibitor IDF-11,774. CONCLUSION: The study highlights the critical role of the HIF-1α-MDM2-P53 axis in asthma, suggesting its potential as a target for therapeutic interventions. The findings indicate that modulating this pathway could offer new avenues for treating the complex respiratory disorder of asthma.

SRSF10 regulates migration of neural progenitor cells and granule cells and affects the formation of dentate gyrus during the development of mouse hippocampus.

Hippocampus is a critical component of the central nervous system. SRSF10 is expressed in central nervous system and plays important roles in maintaining normal brain functions. However, its role in hippocampus development is unknown. In this study, using SRSF10 conditional knock-out mice in neural progenitor cells (NPCs), we found that dysfunction of SRSF10 leads to developmental defects in the dentate gyrus of hippocampus, which manifests as the reduced length and wider suprapyramidal blade and infrapyramidal blade.Furthermore, we proved that loss of SRSF10 in NPCs caused inhibition of the differentiation activity and the abnormal migration of NPCs and granule cells, resulting in reduced granule cells and more ectopic granule cells dispersed in the molecular layer and hilus. Finally, we found that the abnormal migration may be caused by the radial glia scaffold and the reduced DISC1 expression in NPCs. Together, our results indicate that SRSF10 is required for the cell migration and formation of dentate gyrus during the development of hippocampus.

Fibronectin induces a transition from amoeboid to a fan morphology and modifies migration in Entamoeba histolytica.

Cell migration modes can vary, depending on a number of environmental and intracellular factors. The high motility of the pathogenic amoeba Entamoeba histolytica is a decisive factor in its ability to cross the human colonic barrier. We used quantitative live imaging techniques to study the migration of this parasite on fibronectin, a key tissue component. Entamoeba histolytica amoebae on fibronectin contain abundant podosome-like structures. By using a laminar flow chamber, we determined that the adhesion forces generated on fibronectin were twice those on non-coated glass. When migrating on fibronectin, elongated amoeboid cells converted into fan-shaped cells characterized by the presence of a dorsal column of F-actin and a broad cytoplasmic extension at the front. The fan shape depended on the Arp2/3 complex, and the amoebae moved laterally and more slowly. Intracellular measurements of physical variables related to fluid dynamics revealed that cytoplasmic pressure gradients were weaker within fan-shaped cells; hence, actomyosin motors might be less involved in driving the cell body forward. We also found that the Rho-associated coiled-coil containing protein kinase regulated podosome dynamics. We conclude that E. histolytica spontaneously changes its migration mode as a function of the substrate composition. This adaptive ability might favour E. histolytica's invasion of human colonic tissue. By combining microfluidic experiments, mechanical modelling, and image analysis, our work also introduces a computational pipeline for the study of cell migration.

An agent-based model for cell microenvironment simulation using FLAMEGPU2.

This work presents an advanced agent-based model developed within the FLAMEGPU2 framework, aimed at simulating the intricate dynamics of cell microenvironments. Our primary objective is to showcase FLAMEGPU2's potential in modelling critical features such as cell-cell and cell-ECM interactions, species diffusion, vascularisation, cell migration, and/or cell cycling. By doing so, we provide a versatile template that serves as a foundational platform for researchers to model specific biological mechanisms or processes. We highlight the utility of our approach as a microscale component within multiscale frameworks. Through four example applications, we demonstrate the model's versatility in capturing phenomena such as strain-stiffening behaviour of hydrogels, cell migration patterns within hydrogels, spheroid formation and fibre reorientation, and the simulation of diffusion processes within a vascularised and deformable domain. This work aims to bridge the gap between computational efficiency and biological fidelity, offering a scalable and flexible platform to advance our understanding of tissue biology and engineering.

Connexin 30 locally controls actin cytoskeleton and mechanical remodeling in motile astrocytes.

During brain maturation, astrocytes establish complex morphologies unveiling intense structural plasticity. Connexin 30 (Cx30), a gap-junction channel-forming protein expressed postnatally, dynamically regulates during development astrocyte morphological properties by controlling ramification and extension of fine processes. However, the underlying mechanisms remain unexplored. Here, we found in vitro that Cx30 interacts with the actin cytoskeleton in astrocytes and inhibits its structural reorganization and dynamics during cell migration. This translates into an alteration of local physical surface properties, as assessed by correlative imaging using stimulated emission depletion (STED) super resolution imaging and atomic force microscopy (AFM). Specifically, Cx30 impaired astrocyte cell surface topology and cortical stiffness in motile astrocytes. As Cx30 alters actin organization, dynamics, and membrane physical properties, we assessed whether it controls astrocyte migration. We found that Cx30 reduced persistence and directionality of migrating astrocytes. Altogether, these data reveal Cx30 as a brake for astrocyte structural and mechanical plasticity.

A geometrical theory of gliding motility based on cell shape and surface flow.

Gliding motility proceeds with little changes in cell shape and often results from actively driven surface flows of adhesins binding to the extracellular environment. It allows for fast movement over surfaces or through tissue, especially for the eukaryotic parasites from the phylum apicomplexa, which includes the causative agents of the widespread diseases malaria and toxoplasmosis. We have developed a fully three-dimensional active particle theory which connects the self-organized, actively driven surface flow over a fixed cell shape to the resulting global motility patterns. Our analytical solutions and numerical simulations show that straight motion without rotation is unstable for simple shapes and that straight cell shapes tend to lead to pure rotations. This suggests that the curved shapes of Plasmodium sporozoites and Toxoplasma tachyzoites are evolutionary adaptations to avoid rotations without translation. Gliding motility is also used by certain myxo- or flavobacteria, which predominantly move on flat external surfaces and with higher control of cell surface flow through internal tracks. We extend our theory for these cases. We again find a competition between rotation and translation and predict the effect of internal track geometry on overall forward speed. While specific mechanisms might vary across species, in general, our geometrical theory predicts and explains the rotational, circular, and helical trajectories which are commonly observed for microgliders. Our theory could also be used to design synthetic microgliders.

Simulation of Soluble and Bound VEGF-stimulated in vitro Capillary-like Network Formation on Deformed Substrate.

Capillary plexus cultivation is crucial in tissue engineering and regenerative medicine. Theoretical simulations have been conducted to supplement the expensive experimental works. However, the mechanisms connecting mechanical and chemical stimuli remained undefined, and the functions of the different VEGF forms in the culture environment were still unclear. In this paper, we developed a hybrid model for simulating short-term in vitro capillary incubations. We used the Cellular Potts model to predict individual cell migration, morphology change, and continuum mechanics to quantify biogel deformation and VEGF transport dynamics. By bridging the mechanical regulation and chemical stimulation in the model, the results showed good agreement between the predicted network topology and experiments, in which elongated cells connected, forming the network cords and round cells gathered, creating cobblestone-like aggregates. The results revealed that the capillary-like networks could develop in high integrity only when the mechanical and chemical couplings worked adequately, with the cell morphology and haptotaxis driven by the soluble and bound forms of VEGF, respectively, functioning simultaneously.

Derivation and simulation of a computational model of active cell populations: How overlap avoidance, deformability, cell-cell junctions and cytoskeletal forces affect alignment.

Collective alignment of cell populations is a commonly observed phenomena in biology. An important example are aligning fibroblasts in healthy or scar tissue. In this work we derive and simulate a mechanistic agent-based model of the collective behaviour of actively moving and interacting cells, with a focus on understanding collective alignment. The derivation strategy is based on energy minimisation. The model ingredients are motivated by data on the behaviour of different populations of aligning fibroblasts and include: Self-propulsion, overlap avoidance, deformability, cell-cell junctions and cytoskeletal forces. We find that there is an optimal ratio of self-propulsion speed and overlap avoidance that maximises collective alignment. Further we find that deformability aids alignment, and that cell-cell junctions by themselves hinder alignment. However, if cytoskeletal forces are transmitted via cell-cell junctions we observe strong collective alignment over large spatial scales.

Organogenesis: How active forces maintain integrity of migrating cells under pressure.

Cells experience dynamic internal and external forces during animal development. Two new studies reveal critical and unexpected roles for cytoskeletal regulators and nuclear positioning in maintaining the physical integrity of migrating leader cells during Caenorhabditis elegans organogenesis.

Follow the flow: Actin and membrane act as an integrated system to globally coordinate cell shape and movement.

Migratory cells are polarized with protrusive fronts and contractile rears. This spatial organization necessitates long-range coordination of the signals that organize protrusions and contractions. Cells leverage reciprocal interactions of short-range biochemical signals and long-range mechanical forces for this integration. The interface between the plasma membrane and actin cortex is where this communication occurs. Here, we review how the membrane and cortex form an integrated system for long-range coordination of cell polarity. We highlight the role of membrane-to-cortex-attachment proteins as regulators of tension transmission across the cell and discuss the interplay between actin-membrane and polarity signaling complexes. Rather than presenting an exhaustive list of recent findings, we focus on important gaps in our current understanding.

The role of placental kisspeptin in trophoblast invasion and migration: an assessment in Kiss1r knockout mice, BeWo cell lines and human term placenta.

Context There is mounting evidence implicating kisspeptin signalling in placental development and function. Aims This study aimed to elucidate kisspeptin's role in trophoblast invasion and migration using three experimental models. Methods First, we examined the mouse fetus and placenta in a kisspeptin receptor (Kiss1r) knockout (KO) model. Fetal/placental weights and gene expression (quantitative polymerase chain reaction) were assessed. Second, we determined kisspeptin effects on a human trophoblast (BeWo) cell line in vitro . Third, we examined KISS1 and KISS1R gene expression in human placenta from term and pre-term pregnancies. Key results No difference was found in fetal or placental weight between Kiss1r KO and wildtype mice. However, expression of the trophoblast invasion marker, Mmp2 mRNA, was greater in the placental labyrinth zone of Kiss1r KO mice. BeWo cell models of villus cytotrophoblast and syncytiotrophoblast cells exhibited kisspeptin protein expression, with greater expression in syncytiotrophoblast, consistent with KISS1 mRNA. Kisspeptin treatment inhibited the migratory potential of cytotrophoblast-like cells. Finally, while no difference was seen in KISS1 and KISS1R mRNA between term and pre-term placentas, we saw a difference in the relative expression of each gene pre-term. We also observed a positive correlation between KISS1 expression and maternal body mass index. Conclusions Our results indicate that kisspeptin may inhibit trophoblast invasion. Implications Further investigation is required to clarify specific regulatory mechanisms.

Real-Time In Vitro Migration Assay for Primary Murine CD8+ T Cells.

The adaptive immune response is reliant on a T cell's ability to migrate through blood, lymph, and tissue in response to pathogens and foreign bodies. T cell migration is a complex process that requires the coordination of many signal inputs from the environment and local immune cells, including chemokines, chemokine receptors, and adhesion molecules. Furthermore, T cell motility is influenced by dynamic surrounding environmental cues, which can alter activation state, transcriptional landscape, adhesion molecule expression, and more. In vivo, the complexity of these seemingly intertwined factors makes it difficult to distinguish individual signals that contribute to T cell migration. This protocol provides a string of methods from T cell isolation to computer-aided analysis to assess T cell migration in real-time under highly specific environmental conditions. These conditions may help elucidate mechanisms that regulate migration, improving our understanding of T cell kinetics and providing strong mechanistic evidence that is difficult to attain through animal experiments. A deeper understanding of the molecular interactions that impact cell migration is important to develop improved therapeutics.