Buckling forces and the wavy folds between pleural epithelial cells.

Cell shapes in tissues are affected by the biophysical interaction between cells. Tissue forces can influence specific cell features such as cell geometry and cell surface area. Here, we examined the 2-dimensional shape, size, and perimeter of pleural epithelial cells at various lung volumes. We demonstrated a 1.53-fold increase in 2-dimensional cell surface area and a 1.43-fold increase in cell perimeter at total lung capacity compared to residual lung volume. Consistent with previous results, close inspection of the pleura demonstrated wavy folds between pleural epithelial cells at all lung volumes. To investigate a potential explanation for the wavy folds, we developed a physical simulacrum suggested by D'Arcy Thompson in On Growth and Form. The simulacrum suggested that the wavy folds were the result of redundant cell membranes unable to contract. To test this hypothesis, we developed a numerical simulation to evaluate the impact of an increase in 2-dimensional cell surface area and cell perimeter on the shape of the cell-cell interface. Our simulation demonstrated that an increase in cell perimeter, rather than an increase in 2-dimensional cell surface area, had the most direct impact on the presence of wavy folds. We conclude that wavy folds between pleural epithelial cells reflects buckling forces arising from the excess cell perimeter necessary to accommodate visceral organ expansion.

SimuCell3D: three-dimensional simulation of tissue mechanics with cell polarization.

The three-dimensional (3D) organization of cells determines tissue function and integrity, and changes markedly in development and disease. Cell-based simulations have long been used to define the underlying mechanical principles. However, high computational costs have so far limited simulations to either simplified cell geometries or small tissue patches. Here, we present SimuCell3D, an efficient open-source program to simulate large tissues in three dimensions with subcellular resolution, growth, proliferation, extracellular matrix, fluid cavities, nuclei and non-uniform mechanical properties, as found in polarized epithelia. Spheroids, vesicles, sheets, tubes and other tissue geometries can readily be imported from microscopy images and simulated to infer biomechanical parameters. Doing so, we show that 3D cell shapes in layered and pseudostratified epithelia are largely governed by a competition between surface tension and intercellular adhesion. SimuCell3D enables the large-scale in silico study of 3D tissue organization in development and disease at a great level of detail.

Actomyosin fibers DApPLE epithelial apical junctions.

Epithelial cell morphology is essential for cellular homeostasis, but the mechanisms by which cell shape is established remain unclear. In this study, Marivin et al. (2022. J. Cell Biol.https://doi.org/10.1083/jcb.202111002) identify DAPLE as a linker between polarity complexes and the actomyosin network at apical junctions. By recruiting CD2P and activating Gαßγ-mediated RhoA signaling, DAPLE ensures proper cell shape and function.

Metastasising Fibroblasts Show an HDAC6-Dependent Increase in Migration Speed and Loss of Directionality Linked to Major Changes in the Vimentin Interactome.

Metastasising cells express the intermediate filament protein vimentin, which is used to diagnose invasive tumours in the clinic. We aimed to clarify how vimentin regulates the motility of metastasising fibroblasts. STED super-resolution microscopy, live-cell imaging and quantitative proteomics revealed that oncogene-expressing and metastasising fibroblasts show a less-elongated cell shape, reduced cell spreading, increased cell migration speed, reduced directionality, and stronger coupling between these migration parameters compared to normal control cells. In total, we identified and compared 555 proteins in the vimentin interactome. In metastasising cells, the levels of keratin 18 and Rab5C were increased, while those of actin and collagen were decreased. Inhibition of HDAC6 reversed the shape, spreading and migration phenotypes of metastasising cells back to normal. Inhibition of HDAC6 also decreased the levels of talin 1, tropomyosin, Rab GDI ß, collagen and emilin 1 in the vimentin interactome, and partially reversed the nanoscale vimentin organisation in oncogene-expressing cells. These findings describe the changes in the vimentin interactome and nanoscale distribution that accompany the defective cell shape, spreading and migration of metastasising cells. These results support the hypothesis that oncogenes can act through HDAC6 to regulate the vimentin binding of the cytoskeletal and cell-extracellular matrix adhesion components that contribute to the defective motility of metastasising cells.

Factor quinolinone inhibitors alter cell morphology and motility by destabilizing interphase microtubules.

Factor quinolinone inhibitors are promising anti-cancer compounds, initially characterized as specific inhibitors of the oncogenic transcription factor LSF (TFCP2). These compounds exert anti-proliferative activity at least in part by disrupting mitotic spindles. Herein, we report additional interphase consequences of the initial lead compound, FQI1, in two telomerase immortalized cell lines. Within minutes of FQI1 addition, the microtubule network is disrupted, resulting in a substantial, although not complete, depletion of microtubules as evidenced both by microtubule sedimentation assays and microscopy. Surprisingly, this microtubule breakdown is quickly followed by an increase in tubulin acetylation in the remaining microtubules. The sudden breakdown and partial depolymerization of the microtubule network precedes FQI1-induced morphological changes. These involve rapid reduction of cell spreading of interphase fetal hepatocytes and increase in circularity of retinal pigment epithelial cells. Microtubule depolymerization gives rise to FH-B cell compaction, as pretreatment with taxol prevents this morphological change. Finally, FQI1 decreases the rate and range of locomotion of interphase cells, supporting an impact of FQI1-induced microtubule breakdown on cell motility. Taken together, our results show that FQI1 interferes with microtubule-associated functions in interphase, specifically cell morphology and motility.

Symmetry at the 4-Cell Stage Is Associated with Embryo Aneuploidy.

The purpose of this study was to determine if morphometric parameters that can be measured quantitatively using a time-lapse embryo incubator are associated with aneuploidy. Embryos cultured in a time-lapse incubator and assessed with preimplantation genetic testing for aneuploidy (PGT-A) were analyzed retrospectively. Morphokinetic analysis included timing of cell divisions. Quantitative morphometric measurements included the distance between the second and first polar body, zona pellucida thickness at the pronuclear stage and at the 2-cell stage, and blastomere area at the 2- and 4-cell stages. Symmetry at the 2-cell stage was determined by percent difference between blastomeres; symmetry at the 4-cell stage was the percent difference between the smallest and largest blastomeres. Maternal age, blastocyst grade and day of biopsy were recorded. Euploid embryo characteristics were compared to aneuploid embryos. A receiver operating characteristic (ROC) curve was used to evaluate cell symmetry as a predictor of aneuploidy. Embryos (n = 182) from 21 patients (age 22-43; median = 34) were analyzed. Of the 182 embryos, 45% were euploid. Euploid and aneuploid embryos had similar morphokinetics and morphometry across many measures. As expected, age and blastocyst grade were associated with embryo ploidy. It was notable that, additionally, symmetry at the 4-cell stage (27% vs 31%, p = 0.01) was also associated with embryo ploidy. The optimized cutoff from the ROC curve to predict aneuploidy was determined to be 21%. Embryos with > 21% asymmetry at the 4-cell stage had high rates of aneuploidy while morphokinetic parameters were similar. In conclusion, this suggests that embryo selection models using time-lapse parameters would improve if they incorporate cleavage-stage morphometrics.

A PtdIns(3,4,5)P3 dispersal switch engages cell ratcheting at specific cell surfaces.

Force generation in epithelial tissues is often pulsatile, with actomyosin networks generating contractile forces before cyclically disassembling. This pulsed nature of cytoskeletal forces implies that there must be ratcheting mechanisms that drive processive transformations in cell shape. Previous work has shown that force generation is coordinated with endocytic remodeling; however, how ratcheting becomes engaged at specific cell surfaces remains unclear. Here, we report that PtdIns(3,4,5)P3 is a critical lipid-based cue for ratcheting engagement. The Sbf RabGEF binds to PIP3, and disruption of PIP3 reveals a dramatic switching behavior in which medial ratcheting is activated and epithelial cells begin globally constricting apical surfaces. PIP3 enrichments are developmentally regulated, with mesodermal cells having high apical PIP3 while germband cells have higher interfacial PIP3. Finally, we show that JAK/STAT signaling constitutes a second pathway that combinatorially regulates Sbf/Rab35 recruitment. Our results elucidate a complex lipid-dependent regulatory machinery that directs ratcheting engagement in epithelial tissues.

Tumor-initiating stem cell shapes its microenvironment into an immunosuppressive barrier and pro-tumorigenic niche.

Tumor-initiating stem cells (TSCs) are critical for drug resistance and immune escape. However, the mutual regulations between TSC and tumor microenvironment (TME) remain unclear. Using DNA-label retaining, single-cell RNA sequencing (scRNA-seq), and other approaches, we investigated intestinal adenoma in response to chemoradiotherapy (CRT), thus identifying therapy-resistant TSCs (TrTSCs). We find bidirectional crosstalk between TSCs and TME using CellPhoneDB analysis. An intriguing finding is that TSCs shape TME into a landscape that favors TSCs for immunosuppression and propagation. Using adenoma-organoid co-cultures, niche-cell depletion, and lineaging tracing, we characterize a functional role of cyclooxygenase-2 (Cox-2)-dependent signaling, predominantly occurring between tumor-associated monocytes and macrophages (TAMMs) and TrTSCs. We show that TAMMs promote TrTSC proliferation through prostaglandin E2 (PGE2)-PTGER4(EP4) signaling, which enhances ß-catenin activity via AKT phosphorylation. Thus, our study shows that the bidirectional crosstalk between TrTSC and TME results in a pro-tumorigenic and immunosuppressive contexture.

Viral Manipulation of a Mechanoresponsive Signaling Axis Disassembles Processing Bodies.

Processing bodies (PBs) are ribonucleoprotein granules important for cytokine mRNA decay that are targeted for disassembly by many viruses. Kaposi's sarcoma-associated herpesvirus is the etiological agent of the inflammatory endothelial cancer, Kaposi's sarcoma, and a PB-regulating virus. The virus encodes kaposin B (KapB), which induces actin stress fibers (SFs) and cell spindling as well as PB disassembly. We now show that KapB-mediated PB disassembly requires actin rearrangements, RhoA effectors, and the mechanoresponsive transcription activator, YAP. Moreover, ectopic expression of active YAP or exposure of ECs to mechanical forces caused PB disassembly in the absence of KapB. We propose that the viral protein KapB activates a mechanoresponsive signaling axis and links changes in cell shape and cytoskeletal structures to enhanced inflammatory molecule expression using PB disassembly. Our work implies that cytoskeletal changes in other pathologies may similarly impact the inflammatory environment.

Machine learning reveals mesenchymal breast carcinoma cell adaptation in response to matrix stiffness.

Epithelial-mesenchymal transition (EMT) and its reverse process, mesenchymal-epithelial transition (MET), are believed to play key roles in facilitating the metastatic cascade. Metastatic lesions often exhibit a similar epithelial-like state to that of the primary tumour, in particular, by forming carcinoma cell clusters via E-cadherin-mediated junctional complexes. However, the factors enabling mesenchymal-like micrometastatic cells to resume growth and reacquire an epithelial phenotype in the target organ microenvironment remain elusive. In this study, we developed a workflow using image-based cell profiling and machine learning to examine morphological, contextual and molecular states of individual breast carcinoma cells (MDA-MB-231). MDA-MB-231 heterogeneous response to the host organ microenvironment was modelled by substrates with controllable stiffness varying from 0.2kPa (soft tissues) to 64kPa (bone tissues). We identified 3 distinct morphological cell types (morphs) varying from compact round-shaped to flattened irregular-shaped cells with lamellipodia, predominantly populating 2-kPa and >16kPa substrates, respectively. These observations were accompanied by significant changes in E-cadherin and vimentin expression. Furthermore, we demonstrate that the bone-mimicking substrate (64kPa) induced multicellular cluster formation accompanied by E-cadherin cell surface localisation. MDA-MB-231 cells responded to different substrate stiffness by morphological adaptation, changes in proliferation rate and cytoskeleton markers, and cluster formation on bone-mimicking substrate. Our results suggest that the stiffest microenvironment can induce MET.

Effect of macrophages in semen on sperm quality.

BACKGROUND: This was a cross-sectional study in China which analyzed the levels of macrophages (Mφ) in semen and evaluated the influence of Mφ levels in semen on sperm quality. METHODS: The subjects involves 78 males, 25- to 35-year old. The samples were divided into a low group (Mφ < 6 × 105/ml) and a high group (Mφ > 6 × 105/ml). Evaluation included consideration of the influencing factors of male semen quality, macrophage concentration, sperm motility, morphology, membrane integrity DNA fragmentation index (DFI), anti-sperm antibodies (AsAb), IL-10, and IL-12 in semen. RESULTS: There was no difference in the physical or chemical indices of the semen, sperm concentration, AsAb, IL-10, or IL-12 between the two groups (P > 0.05). The percentage of sperm forward motility (PR%), the rate of normal sperm shape, and the integrity of cell membranes in the low group were higher than those in the high group (P < 0.05), while the percentage of sperm inactivity (IM%), the rate of sperm head deformity, the rate of deformity in the neck and middle segment, the sperm deformity index (SDI), the teratozoospermia index (TZI), and the sperm DFI in the low group were lower than those in the high group (P < 0.05). The concentration of Mφ in the semen was linearly correlated with sperm concentration, sperm PR%, IM%, sperm normal shape rate, head deformity rate, neck and middle deformity rate, SDI, TZI, sperm DFI, and sperm cell membrane integrity (P < 0.05), but there was no linear correlation with IL-10 or IL-12 (P > 0.05). CONCLUSIONS: The Mφ concentration in semen is not significantly correlated with semen volume or sperm concentration, but negatively correlated with sperm motility, morphology, cell membrane integrity, and DNA damage rate. There is no significant correlation between the macrophages and the concentration of IL-10 or IL-12.

MiR-182-5p, MiR-192-5p, and MiR-493-5p Constitute a Regulatory Network with CRISP3 in Seminal Plasma Fluid of Teratozoospermia Patients.

Numerous evidences suggested that microRNAs (miRs) could play an active and significant role during spermatogenesis. Cysteine-rich secretory protein (CRISP3) has a role in inflammatory response and is extremely over-expressed in adolescents with varicocele seminal plasma and modified semen analysis. Nowadays, the miRs expression's association with their target genes is well recognized. The aim of this study was evaluating the association of CRISP3 and four candidate miRs among teratozoospermia (TZ) infertile men. First, we have selected four miRs, miR-182-5p, miR-192-5p, miR-204-5p, and miR-493-5p bioinformatically. After that, RNA was extracted from semen samples of 21 TZ patients and 20 normozoospermia (Norm). Then, their expression levels were assessed using real-time polymerase chain reaction method. In the next step, we quantified the expression of two CRISP3 protein isoforms, targeted by these miRs, using western blotting. According to our results, up-regulation of miR-182-5p, miR-192-5p, and miR-493-5p was observed. MiR-182-5p, miR-192-5p, and miR-493-5p showed good AUC values which can be introduced as possible biomarkers of TZ. In addition, the expression level of the CRISP3 glycosylated (31 kDa) isoform was significantly lower in TZ patients than Norm ones. Notably, in TZ patients, there was a possibly positive correlation of glycosylated CRISP3 expression with normal sperm morphology. According to our results, CRISP3 protein can play a significant role in male infertility especially in maturation formation of spermatozoa. Also, deregulation of the studied miRs, miR-182-5p, miR-92-5p, and miR-493-5p, can suggest a regulatory network between these miRs and CRISP3 isoforms and suggest their regulatory roles in male infertility.

Seasonal changes in adenosine kinase in tanycytes of the Arctic ground squirrel (Urocitellus parryii).

Hibernation is a seasonal strategy to conserve energy, characterized by modified thermoregulation, an increase in sleep pressure and drastic metabolic changes. Glial cells such as astrocytes and tanycytes are the brain metabolic sensors, but it remains unknown whether they contribute to seasonal expression of hibernation. The onset of hibernation is controlled by an undefined endogenous circannual rhythm in which adenosine plays a role through the activation of the A1 adenosine receptor (A1AR). Seasonal changes in brain levels of adenosine may contribute to an increase in A1AR sensitivity leading to the onset of hibernation. The primary regulator of extracellular adenosine concentration is adenosine kinase, which is located in astrocytes. Using immunohistochemistry to localize and quantify adenosine kinase in Arctic ground squirrels' brain collected during different seasons, we report lower expression of adenosine kinase in the third ventricle tanycytes in winter compared to summer; a similar change was not seen in astrocytes. Moreover, for the first time, we describe adenosine kinase expression in tanycyte cell bodies in the hypothalamus and in the area postrema, both brain regions involved in energy homeostasis. Next we describe seasonal changes in tanycyte morphology in the hypothalamus. Although still speculative, our findings contribute to a model whereby adenosine kinase in tanycytes regulates seasonal changes in extracellular concentration of adenosine underling the seasonal expression of hibernation.

Human anterior pituitary's ACTH cells during the aging process: immunohistochemic and morphometric study.

Corticotrophs produce a hormone that stimulates the adrenal gland cortex to secrete glucocorticoids, which in turn have effects on carbohydrate and protein metabolism. Quantification, morphological characteristics, and distribution of corticotrophs in the anterior pituitary and changes in the number and shape of the cells during aging have been examined using immunohistochemical and morphometric methods. The material consisted of 14 anterior pituitaries taken from cadavers at routine autopsy. The tissue was processed by standard histological procedure and the obtained slices were stained by the monoclonal anti-ACTH antibody for corticotrophs identification. Digital images of stained histological sections were analyzed using the morphometric method with the Image J system. The volume density of ACTH positive cells was determined. The cases were classified into three age groups. One-way ANOVA showed that the volume density of the corticotrophs was significantly higher in the second and third group in relation to the first group. The difference in the volume densities of the corticotrophs between the genders was not significant. Morphometric and statistical analyses demonstrated a significant positive correlation between the corticotrophs volume densities and the age of the evaluated cases. Linear regression showed that age significantly predicts corticotrophs volume density. Corticotrophs significantly increase during the life span.

Microglial and peripheral immune priming is partially sexually dimorphic in adolescent mouse offspring exposed to maternal high-fat diet.

BACKGROUND: Maternal nutrition is critical for proper fetal development. While increased nutrient intake is essential during pregnancy, an excessive consumption of certain nutrients, like fat, can lead to long-lasting detrimental consequences on the offspring. Animal work investigating the consequences of maternal high-fat diet (mHFD) revealed in the offspring a maternal immune activation (MIA) phenotype associated with increased inflammatory signals. This inflammation was proposed as one of the mechanisms causing neuronal circuit dysfunction, notably in the hippocampus, by altering the brain-resident macrophages-microglia. However, the understanding of mechanisms linking inflammation and microglial activities to pathological brain development remains limited. We hypothesized that mHFD-induced inflammation could prime microglia by altering their specific gene expression signature, population density, and/or functions. METHODS: We used an integrative approach combining molecular (i.e., multiplex-ELISA, rt-qPCR) and cellular (i.e., histochemistry, electron microscopy) techniques to investigate the effects of mHFD (saturated and unsaturated fats) vs control diet on inflammatory priming, as well as microglial transcriptomic signature, density, distribution, morphology, and ultrastructure in mice. These analyses were performed on the mothers and/or their adolescent offspring at postnatal day 30. RESULTS: Our study revealed that mHFD results in MIA defined by increased circulating levels of interleukin (IL)-6 in the mothers. This phenotype was associated with an exacerbated inflammatory response to peripheral lipopolysaccharide in mHFD-exposed offspring of both sexes. Microglial morphology was also altered, and there were increased microglial interactions with astrocytes in the hippocampus CA1 of mHFD-exposed male offspring, as well as decreased microglia-associated extracellular space pockets in the same region of mHFD-exposed offspring of the two sexes. A decreased mRNA expression of the inflammatory-regulating cytokine Tgfb1 and microglial receptors Tmem119, Trem2, and Cx3cr1 was additionally measured in the hippocampus of mHFD-exposed offspring, especially in males. CONCLUSIONS: Here, we described how dietary habits during pregnancy and nurturing, particularly the consumption of an enriched fat diet, can influence peripheral immune priming in the offspring. We also found that microglia are affected in terms of gene expression signature, morphology, and interactions with the hippocampal parenchyma, in a partially sexually dimorphic manner, which may contribute to the adverse neurodevelopmental outcomes on the offspring.

Solid-Liquid Transition of Deformable and Overlapping Active Particles.

Experiments and theory have shown that cell monolayers and epithelial tissues exhibit solid-liquid and glass-liquid transitions. These transitions are biologically relevant to our understanding of embryonic development, wound healing, and cancer. Current models of confluent epithelia have focused on the role of cell shape, with less attention paid to cell extrusion, which is key for maintaining homeostasis in biological tissue. Here, we use a multiphase field model to study the solid-liquid transition in a confluent monolayer of deformable cells. Cell overlap is allowed and provides a way for modeling the precursor for extrusion. When cells overlap rather than deform, we find that the melting transition changes from continuous to first order like, and that there is an intermittent regime close to the transition, where solid and liquid states alternate over time. By studying the dynamics of five- and sevenfold disclinations in the hexagonal lattice formed by the cell centers, we observe that these correlate with spatial fluctuations in the cellular overlap, and that cell extrusion tends to initiate near fivefold disclinations.

Collective cell mechanics of epithelial shells with organoid-like morphologies.

The study of organoids, artificially grown cell aggregates with the functionality and small-scale anatomy of real organs, is one of the most active areas of research in biology and biophysics, yet the basic physical origins of their different morphologies remain poorly understood. Here, we propose a mechanistic theory of epithelial shells which resemble small-organoid morphologies. Using a 3D surface tension-based vertex model, we reproduce the characteristic shapes from branched and budded to invaginated structures. We find that the formation of branched morphologies relies strongly on junctional activity, enabling temporary aggregations of topological defects in cell packing. To elucidate our numerical results, we develop an effective elasticity theory, which allows one to estimate the apico-basal polarity from the tissue-scale modulation of cell height. Our work provides a generic interpretation of the observed epithelial shell morphologies, highlighting the role of physical factors such as differential surface tension, cell rearrangements, and tissue growth.

Phosphatidylinositol 4-kinase III beta regulates cell shape, migration, and focal adhesion number.

Cell shape is regulated by cell adhesion and cytoskeletal and membrane dynamics. Cell shape, adhesion, and motility have a complex relationship and understanding them is important in understanding developmental patterning and embryogenesis. Here we show that the lipid kinase phosphatidylinositol 4-kinase III beta (PI4KIIIß) regulates cell shape, migration, and focal adhesion (FA) number. PI4KIIIß generates phosphatidylinositol 4-phosphate (PI4P) from phosphatidylinositol and is highly expressed in a subset of human breast cancers. PI4KIIIß and the PI4P it generates regulate a variety of cellular functions, ranging from control of Golgi structure, fly fertility, and Akt signaling. Here, we show that loss of PI4KIIIß expression decreases cell migration and alters cell shape in NIH3T3 fibroblasts. The changes are accompanied by an increase in the number of FA in cells lacking PI4KIIIß. Furthermore, we find that PI4P-containing vesicles move to the migratory leading edge during migration and that some of these vesicles tether to and fuse with FA. Fusion is associated with FA disassembly. This suggests a novel regulatory role for PI4KIIIß and PI4P in cell adhesion and cell shape maintenance.

A device for investigation of natural cell mobility and deformability.

A microfluidic device made of polydimethylsiloxane was developed for continuous evaluation of natural migration mobility of many eukaryotic cells in relaxed and deformed state. The device was fabricated by standard photolithography and soft lithography techniques using the SU-8 3010 negative photoresist on a glass wafer as the master mold. The simple flow-free device exploits the chemotactic movement of cells through a set of mechanical barriers in the direction of concentration gradients of attractants. The barriers are formed by arrays of circular cross-section pillars with decreasing spacing 7, 5, and 3 µm. To pass through the obstacles, the cells are deformed and change their cytoskeletal architecture. The instantaneous migration velocities of cells are monitored in a time-lapse setup of the scanning confocal microscope. Thus, the cellular deformability and migratory activity can easily be evaluated. The functionality of the device was tested with model HeLa cells stably transfected with fluorescent Premo FUCCI Cell Cycle Sensor. The designed device has the potential to be implemented for testing the tendency of patients' tumors to metastasis.

Self-organization in brain tumors: How cell morphology and cell density influence glioma pattern formation.

Modeling cancer cells is essential to better understand the dynamic nature of brain tumors and glioma cells, including their invasion of normal brain. Our goal is to study how the morphology of the glioma cell influences the formation of patterns of collective behavior such as flocks (cells moving in the same direction) or streams (cells moving in opposite direction) referred to as oncostream. We have observed experimentally that the presence of oncostreams correlates with tumor progression. We propose an original agent-based model that considers each cell as an ellipsoid. We show that stretching cells from round to ellipsoid increases stream formation. A systematic numerical investigation of the model was implemented in [Formula: see text]. We deduce a phase diagram identifying key regimes for the dynamics (e.g. formation of flocks, streams, scattering). Moreover, we study the effect of cellular density and show that, in contrast to classical models of flocking, increasing cellular density reduces the formation of flocks. We observe similar patterns in [Formula: see text] with the noticeable difference that stream formation is more ubiquitous compared to flock formation.