Uniform Cell Research Articles

Interpenetrating Polymer Network of Hydroxyethyl Methacrylate and Chitosan as Cryogels for Tissue Engineering Applications

ABSTRACTTissue engineering is a vast expanding field with applications in areas such as tissue/organ transplantation, drug delivery, in vitro models, and so on. Biomaterials form an essential component in tissue engineering by acting as a template for cellular activity, therefore, novel tissue‐engineered biomaterials with innovative properties are in high demand. Hence, this work proposes an interpenetrating polymer network cryogel of chitosan and hydroxyethyl methacrylate (HEMA) as a tissue‐engineered biomaterial with uniform and high cell seeding throughout the cryogel matrix. The physical analysis of the cryogels demonstrated a highly macroporous structure exhibiting uniform pore size distribution and overall porosity through field emission‐scanning electron microscopy (FE‐SEM) analysis with pore sizes lying in the range of 50–200 μm and 150–400 μm in horizontal and transverse plane, respectively. The cryogels were also found to be degradable with an average percent degradation of 17.28 ± 1.47% in 4 weeks, and their mechanical properties revealed an average compressive strength of 0.05 MPa and an elastic modulus of 3 MPa. Further, biological characterization of the cryogel through direct contact test depicted an excellent biocompatibility with L929 mouse fibroblast and MC3T3‐E1 preosteoblasts with negligible presence of dead cells in and around the cryogel. Uniform and high cell seeding with an increasing proliferation trend of MC3T3‐E1 cells was observed on these cryogels through live‐dead staining and MTT assay at day 1, 3, and 7‐time point. Cell adherence studies via FE‐SEM analysis also depicted a similar trend with uniform MC3T3‐E1 cell distribution along with highly flattened morphology and extracellular matrix production. Therefore, based on their promising physico‐chemical and biological properties, HEMA‐Chitosan cryogels exhibit a strong potential application as tissue engineered biomaterials.

Humidity Resistant Biodegradable Starch Foams Reinforced with Polyvinyl Butyral (PVB) and Chitosan.

In this study, water-insoluble, moisture-resistant starch foams were prepared using an optimized one-step extrusion-foaming process in a ZSK-30 twin screw extruder. The extrusion parameters, including temperature, screw configuration, die diameter, water content, and feeding rates, were optimized to achieve foams with the lowest density and controlled expansion. A screw configuration made up of three kneading sections was found to be the most effective for better mixing and foaming. Polyvinyl butyral (PVB) acted as a plasticizer, resulting in foams with a density of 21 kg/m3 and an expansion ratio of 38.7, while chitosan served as a nucleating agent, reducing cell size and promoting a uniform cell size distribution. The addition of PVB and chitosan reduced the moisture sensitivity of the foams, rendering them hydrophobic and water-insoluble. The contact angle increased from 0° for control foams to 101.5° for foams containing 10% chitosan and 10% PVB. Confocal laser scanning microscopy (CLSM) confirmed the migration of chitosan to the foam surface, enhancing hydrophobicity. Aqueous biodegradation tests, conducted at 30 °C in accordance with ISO 14852 standards, demonstrated that despite enhanced moisture resistance, the foams remained readily biodegradable, achieving approximately 80% biodegradation within 80 days. These modified starch foams present a sustainable solution for packaging and insulation applications that demand long-term humidity resistance.

High-performance bio-based foam from agricultural waste luffa seed oil polyols

The present study aimed to develop eco-friendly polyurethane viscoelastic foam (PVF) with enhanced tear strength while preserving softness, comfort, and safety by utilizing bio-based polyols derived from luffa seed oil (LSO). Two types of bio-based polyols (PLSO-D and PLSO-M) were synthesized through the epoxidation process using diethylene glycol (DEG) and methanol (MeOH), respectively, for the production of LSO-based polyurethane viscoelastic foams (LSO-PVFs). The analyses revealed that these polyols exhibited high dispersion, appropriate hydroxyl values, and contained by-products such as oligomeric esters of fatty acids that facilitated physical and chemical crosslinking. LSO-PVFs demonstrated a uniform and fine cell structure with abundant open-cells, thick cell walls (55.5–76.6µm), and high apparent density (48.7–50.7kg/m3). Moreover, they exhibited enhanced tensile and tear strength (increasing up to 200% compared to petroleum-based PVFs), lower water absorption, excellent mildew resistance at 10% PLSO substitution. Additionally, these factors endowed LSO-PVFs with low indentation hardness and good bottom support performance, characterized by a compressive deflection coefficient of 2.30–3.54. This approach presents a sustainable alternative for foam production that offers comfort, durability while promoting the value-added utilization of agricultural waste.

Endodermal cells use contact inhibition of locomotion to achieve uniform cell dispersal during zebrafish gastrulation.

The endoderm is one of the three primary germ layers that ultimately gives rise to the gastrointestinal and respiratory epithelia and other tissues. In zebrafish and other vertebrates, endodermal cells are initially highly migratory with only transient interactions among one other, but later converge together to form an epithelial sheet. Here, we show that during their early, migratory phase, endodermal cells actively avoid each other through contact inhibition of locomotion (CIL), a characteristic response consisting of 1) actin depolymerization and membrane retraction at the site of contact, 2) preferential actin polymerization along a cell-free edge, and 3) reorientation of migration away from the other cell. We found that this response is dependent on the Rho GTPase RhoA. Expression of dominant-negative (DN) RhoA attenuated migration reorientation after cell-cell contact and increased the amount of time cells spent in contact with each other - behaviors consistent with a loss of CIL. Computational modeling predicted that CIL is required to achieve the efficient and uniform dispersal characteristic of endodermal cells. Consistent with our model, we found that loss of CIL via DN RhoA expression resulted in irregular clustering of cells within the endoderm. Finally, using a combination of pharmacological and genetic perturbations, we identify EphA2 as the cell surface receptor mediating endodermal CIL. Together, our results suggest that endodermal cells use EphA2- and RhoA-dependent CIL as a cell dispersal and spacing mechanism, demonstrating how tissue-scale patterns can emerge from local cell-cell interactions.

Septal wall synthesis is sufficient to change ameba-like cells into uniform oval-shaped cells in Escherichia coli L-forms

A cell wall is required to control cell shape and size to maintain growth and division. However, some bacterial species maintain their morphology and size without a cell wall, calling into question the importance of the cell wall to maintain shape and size. It has been very difficult to examine the dispensability of cell wall synthesis in rod-shaped bacteria such as Escherichia coli for maintenance of their shape and size because they lyse without cell walls under normal culture conditions. Here, we show that wall-less E. coli L-form cells, which have a heterogeneous cell morphology, can be converted to a mostly uniform oval shape solely by FtsZ-dependent division, even in the absence of cylindrical cell wall synthesis. This FtsZ-dependent control of cell shape and size in the absence of a cell wall requires at least either the Min or nucleoid occlusion systems for positioning FtsZ at mid cell division sites.

Spherical Shell Bioprinting to Produce Uniform Spheroids with Controlled Sizes.

Conventional cell spheroid production methods are largely manual, leading to variations in size and shape that compromise consistency and reliability for use in cell-based therapeutic applications. To enhance spheroid production, a spherical shell bioprinting system was implemented, enabling the high-throughput generation of uniform cell spheroids with precisely controlled sizes. The system encapsulates cells within thin alginate hydrogel shells formed through bioprinting and ion crosslinking reactions. Alginate-calcium ion crosslinking created alginate shells that contained gelatin-based bioinks with embedded cells, facilitating spontaneous cell aggregation within the shells and eliminating the need for plastic wells. By adjusting cell concentrations in the alginate-gelatin bioink, we achieved precise control over spheroid size, maintaining a sphericity above 0.94 and size deviations within ±10 µm. This method has been successfully applied to various cell types including cancer cells, fibroblasts, chondrocytes, and epithelial cells, demonstrating its versatility. This scalable approach enhances the reliability of cell therapy and drug screening, offering a robust platform for future biomedical applications.

Fabrication of Hydroxyl Modified Hollow Glass Microsphere Composite Isocyanate‐Based Polyimide Foam and Optimization Strategy Based on Different Bonding Mechanisms

ABSTRACTIn this study, the hydroxyl modified hollow glass microsphere (HM‐HGM) is added to different foaming slurries of isocyanate‐based polyimide foam (IBPIF) at varying ratios, and different bonding effects are formed to optimize the dispersion behavior. Then, the novel HGM composited IBPIF (IBPIF/HGM) is prepared. Hydroxyl groups on HM‐HGM establish hydrogen bonding effect with pyromellitic acid dimethyl ester and dimethyl formamide in the white slurry and react with isocyanate groups in the black slurry. The cell structure of IBPIF is altered to improve its sound absorption performance and mechanical behaviors. Compared with IBPIF/HGM‐0, the average cell size of IBPIF/HGM‐1 and BPIF/HGM‐5 decreases significantly. The sound absorption performance and mechanical behaviors of them are improved to some extent. Compared with samples in which the HM‐HGM is added alone to a single slurry, when the dosage ratio of HM‐HGM in black and white slurries is 1:1, IBPIF/HGM‐3 has more uniform cell structure. The change of IBPIF cell structure by the introduction of HM‐HGM and the unique structure of HM‐HGM can enhance the sound absorption performance and mechanical behaviors of IBPIF. The design idea of different bonding mechanisms significantly provides technical assistance to enhance the acoustic performance of polymeric foam materials.

Research on Melamine-Urea-Formaldehyde/Nano-Al2O3 Composite Foam Thermal Insulation Sealing Materials for Coal Mines.

In this study, melamine-urea-formaldehyde/nano-Al2O3 (MUF/nano-Al2O3) composite gel foams were produced by foaming with CO2 generated by CaCO3 and phosphoric acid. Nano-Al2O3 was introduced to the MUF matrix based on the optimum formulation obtained by the response surface methodology based on Box-Behnken design, and the effects of nano-Al2O3 on the cell structure, apparent density, compressive strength, pulverization ratio, thermal stability, and thermal conductivity were investigated. The results revealed that the introduction of nano-Al2O3 could improve the foaming and mechanical properties of MUF with smaller cell sizes, a narrower cell size distribution, decreased apparent density, higher compressive strength, and a decreased pulverization ratio. MUF/nano-Al2O3-1.5 with the most uniform cell size distribution had a 21% increased compressive strength, but its apparent density and pulverization ratio decreased by 10% and 13%, respectively. Although the char yield decreased with the introduction of nano-Al2O3, MUF/nano-Al2O3 composite foams still presented improved thermal stability under 300 °C, with a thermal conductivity of 0.068 W/(m·K) and a limiting oxygen index of more than 30%. Therefore, MUF/nano-Al2O3 composite gel foam could be employed in coal mines as thermal insulation sealing material due to its better cell structure and mechanical properties.

Safety and Effectiveness of Triple-Antenna Hepatic Microwave Ablation.

Microwave ablation is becoming a standard procedure for treating tumors based on heat generation, causing an elevation in the tissue temperature level from 50 to 60 °C, causing tissue death. Microwave ablation is associated with uniform cell killing within ablation zones, multiple-antenna capability, low complication rates, and long-term survival. Several reports have demonstrated that multiple-antenna microwave ablation is a promising strategy for safely, rapidly, and effectively treating large tumors. The key advantage of multi-antenna tumor microwave ablation is the creation of a large, well-defined ablation zone without excessively long treatment times or high power that can damage healthy tissue. The strategic positioning of multiple probes provides a fully ablated volume, even in regions where individual probe damage is incomplete. Accurate modeling of the complex thermal and electromagnetic behaviors of tissue is critical for optimizing microwave ablation because material parameters and tissue responses can change significantly during the procedure. In the case of multi-antenna microwave ablation, the calculation complexity increases significantly, requiring significant computational resources and time. This study aimed to evaluate the efficacy and safety of liver percutaneous microwave ablation using the simultaneous activation of three antennas for the treatment of lesions larger than 3 cm. Based on the known results from a single-probe setup, researchers can estimate and evaluate various spatial configurations of the three-probe array to identify the optimal arrangement. Due to the synergistic effects of the combined radiation from the three antennas, the resulting ablation zone can be significantly larger, leading to better outcomes in terms of treatment time and effectiveness. The obtained results revealed that volumetric damage and the amount of damaged healthy tissue are smaller for a three-antenna configuration than for microwave ablation using a single-antenna and two-antenna configurations.

Axonal spheroids are regulated by Schwann cells after peripheral nerve injury.

Axonal spheroids are hallmark features of neurodegeneration, forming along degenerating axons and contributing to disease progression. Despite their ubiquity across degenerative etiologies, the dynamics of spheroid disappearance, as well as their interactions with glial cells, remain poorly understood. Here, using an in vivo zebrafish model of peripheral nerve injury, we identified several patterns of spheroid disappearance that are regulated by Schwann cells. These results describe spheroid dynamics across their lifetimes, establish a role for the extra-axonal environment in altering spheroid outcomes, and identify a cellular mechanism whereby spheroid fates are altered.

Malignant Bone-Forming Neoplasm With NIPBL::BEND2 Fusion.

Conventional high-grade osteosarcomas are characterized by aggressive radiologic features, cytologic pleomorphism, and complex genomics. However, rare examples of osteosarcomas remain challenging due to unusual histology, such as sclerosing or osteoblastoma-like features, which may require molecular confirmation of their complex genetic alterations. We have encountered such a case in a 17-year-old man, who presented with a third metatarsal sclerotic bone lesion, found incidentally in the work-up of a foot trauma. The initial imaging revealed a lesion with sclerotic/blastic features proximally and lucent/lytic portion distally, findings interpreted consistent with osteoblastoma. The lesion was managed intra-lesionally with curettings and cryoablation; however, the microscopic findings were non-specific, showing a bland osteoblastic proliferation embedded in a densely sclerotic matrix. Subsequently, the patient developed two rapid recurrences; the first recurrence was treated similarly despite its associated soft tissue extension radiographically, and the histologic findings remained non-specific. The 2nd recurrence showed a large mass, with bone destruction and soft tissue extension and an open biopsy revealed features of osteosarcoma with lace-like osteoid deposition, albeit with uniform cytomorphology. The subsequent below knee amputation showed features compatible with high-grade osteosarcoma, including solid growth of uniform epithelioid cells, with vesicular nuclei and scant cytoplasm, set in a lace-like meshwork of osteoid matrix. There was significant mitotic activity and tumor necrosis. Tumor cells were positive for SATB2. Further molecular work-up was performed showing an unexpected NIPBL::BEND2 fusion, which has been previously reported in two cases of phosphaturic mesenchymal tumor (PMT). FGF23 (ISH) was performed and was negative. By DNA methylation profiling, unsupervised clustering and UMAP dimensionality reduction revealed grouping with high-grade osteosarcomas and not with the PMT group. The patient received chemotherapy post-amputation and is alive without evidence of disease, with 10-month follow-up. We report an aggressive, overtly malignant acral bone-forming tumor, harboring a NIPBL::BEND2 fusion. Further studies are needed to evaluate the recurrent potential of this fusion in osteosarcomas and its relationship with PMT.

On the design and fabrication of nanoliter-volume hanging drop networks

Hanging drop cultures provide a favorable environment for the gentle, gel-free formation of highly uniform three-dimensional cell cultures often used in drug screening applications. Initial cell numbers can be limited, as with primary cells provided by minimally invasive biopsies. Therefore, it can be beneficial to divide cells into miniaturized arrays of hanging drops to supply a larger number of samples. Here, we present a framework for the miniaturization of hanging drop networks to nanoliter volumes. The principles of a single hanging drop are described and used to construct the fundamental equations for a microfluidic system composed of multiple connected drops. Constitutive equations for the hanging drop as a nonlinear capacitive element are derived for application in the electronic-hydraulic analogy, forming the basis for more complex, time-dependent numerical modeling of hanging drop networks. This is supplemented by traditional computational fluid dynamics simulation to provide further information about flow conditions within the wells. A fabrication protocol is presented and demonstrated for creating transparent, microscale arrays of pinned hanging drops. A custom interface, pressure-based fluidic system, and environmental chamber have been developed to support the device. Finally, fluid flow on the chip is demonstrated to align with expected behavior based on the principles derived for hanging drop networks. Challenges with the system and potential areas for improvement are discussed. This paper expands on the limited body of hanging drop network literature and provides a framework for designing, fabricating, and operating these systems at the microscale.

Calcifying Nested Stromal-Epithelial Tumor of the Liver: A Rare Find

Abstract Introduction/Objective Calcifying Nested Stromal-Epithelial Tumor (CNSET) is a rare tumor of the liver with less than 40 cases described in the literature. It is a low-grade tumor characterized by a distinctive nested architecture surrounded by a cellular myofibroblastic stroma and psammomatous calcifications. CTNNB1 gene deletions have been identified in several cases. CNSET typically arises in the right lobe and there is a female predominance with a predilection for children, adolescents, and young adults. Methods/Case Report We report an interesting case of CNSET in a 13-year-old male with no pertinent medical history who presented with a one-month history of fatigue, paleness, and liver mass on physical examination. MRI abdomen showed a 9.8 cm heterogenous mass with a central scar involving the right lobe of the liver, radiologically favored to be fibrolamellar hepatocellular carcinoma. AFP levels were normal. The patient underwent a partial hepatectomy. On gross examination, the specimen showed a yellow, circumscribed, firm mass. H&E slides showed scattered round to oval nests of bland spindle and epithelioid cells arranged in short fascicles at the periphery of nests. Collagenous stroma surrounding the nests showed spindle cells with tapered nuclei and pinpoint nucleolus. Areas with calcification and osseous metaplasia were also present. Immunohistochemistry performed showed the tumor cells to be positive for pan-cytokeratin (weak), WT-1, and vimentin, and negative for Hepar-1, arginase-1, synaptophysin, chromogranin, S100, NUT-1, myogenin, and desmin. Beta-catenin showed nuclear and cytoplasmic staining pattern. INI-1 was retained within the tumor cells and Ki-67 showed a proliferative index of approximately 10-20%. On MSK-Impact (next- generation sequencing), a mutation in CTNNB1 was detected. Archer FusionPlex was negative for gene fusions within the tumor. Results (if a Case Study enter NA) NA Conclusion It is crucial to become familiar with this rare entity and to avoid mistaking it for hepatoblastoma. CNSET exhibits a biphasic morphology characterized by a nested architecture featuring bland and uniform epithelioid and spindle cells, surrounded by a cellular myofibroblastic stroma. Confirming the diagnosis can be aided by nuclear β- catenin and the expression of WT1.

CK20-negative Merkel Cell Carcinoma with Concomitant Squamous Cell Carcinoma In-Situ and Metastasis to the Parotid Gland

Abstract Introduction/Objective This report presents a patient with an aggressive CK20-negative neuroendocrine tumor of the dermis with metastasis to the parotid gland and a concomitant squamous cell carcinoma in-situ (SCCIS) and highlights diagnostic challenges of CK20-negative tumors. Methods/Case Report A centennial female with a history of non-melanoma skin cancer and leukemia presented to the dermatology clinic with a one-year history of an enlarging skin lesion on her right cheek and a subcutaneous mass on the right central lateral neck. On examination, the lesion appeared as a rough and raised red papule with a central crust. On CT, a large heterogeneously enhancing right parotid mass with surrounding satellite nodules, adjacent necrotic adenopathy, and an ulcerating subcutaneous mass in the adjacent right face. An incidental 3 mm right upper lobe nodule was identified. The pathology report noted two distinct malignant neoplasms within the specimen. Microscopy revealed ulceration with acanthosis and full thickness keratinocyte atypia, suggesting Bowen’s disease. On microscopic examination of the separate dermal tumor, there were sheets of uniform small cells with a high nuclear to cytoplasmic ratio and scant cytoplasm. IHC staining was positive for the neuroendocrine markers AE1/AE3, CAM5.2, synaptophysin, and CD56. CK20, CK7, and TTF-1 were negative. As some MCC may be negative for CK20, CK20 was performed twice. Differential diagnoses included SCCIS with CK20-negative MCC versus a metastatic neuroendocrine carcinoma of parotid gland to the skin. The patient transitioned to hospice care soon after the diagnosis. Results (if a Case Study enter NA) NA. Conclusion Our patient presents with an interesting case of a CK20-negative MCC with metastasis to the parotid gland. Since CK20 was stained negative twice, it complicated the diagnosis of MCC vs. small cell carcinoma. We suspected a more likely diagnosis of MCC metastasis to the parotid gland due to multiple reported cases (Day et al. 2016). Our patient also had an identifiable Bowen’s disease borderline to the neuroendocrine carcinoma. It is well known of the coexistence of MCC and squamous cell carcinoma in a cutaneous lesion (Suaiti et al. 2019). Thus, a metastatic MCC to the parotid gland is more likely in this case. As MCC is an aggressive cancer that metastasizes quickly, accurate diagnosis is crucial. With recent literature reporting the association of CK20-negativity with VN-MCC, and the presence of UV signature mutations in CK20- negative MCC, genomic screening would be a useful diagnostic tool.

Exploiting phenotypic heterogeneity to improve production of glutathione by yeast

BackgroundGene expression noise (variation in gene expression among individual cells of a genetically uniform cell population) can result in heterogenous metabolite production by industrial microorganisms, with cultures containing both low- and high-producing cells. The presence of low-producing individuals may be a factor limiting the potential for high yields. This study tested the hypothesis that low-producing variants in yeast cell populations can be continuously counter-selected, to increase net production of glutathione (GSH) as an exemplar product.ResultsA counter-selection system was engineered in Saccharomyces cerevisiae based on the known feedback inhibition of gamma-glutamylcysteine synthetase (GSH1) gene expression, which is rate limiting for GSH synthesis: the GSH1 ORF and the counter-selectable marker GAP1 were expressed under control of the TEF1 and GSH-regulated GSH1 promoters, respectively. An 18% increase in the mean cellular GSH level was achieved in cultures of the engineered strain supplemented with D-histidine to counter-select cells with high GAP1 expression (i.e. low GSH-producing cells). The phenotype was non-heritable and did not arise from a generic response to D-histidine, unlike that with certain other test-constructs prepared with alternative markers.ConclusionsThe results corroborate that the system developed here improves GSH production by targeting low-producing cells. This supports the potential for exploiting end-product/promoter interactions to enrich high-producing cells in phenotypically heterogeneous populations, in order to improve metabolite production by yeast.

A Scalable Platform for Generating Uniform Human-Induced Pluripotent Stem Cell (hiPSC)-Derived Kidney Organoids Using Micropatterning

A Scalable Platform for Generating Uniform Human-Induced Pluripotent Stem Cell (hiPSC)-Derived Kidney Organoids Using Micropatterning

Osteogenic effect of poly(lactic-co-glycolic acid) microcapsules with different molecular weights encapsulating bone morphogenetic protein 2.

This study aimed to explore the effects of bone morphogenetic protein 2 (BMP-2) encapsula-ted in poly(lactic-co-glycolic acid) (PLGA) microcapsules with different molecular weights on the osteogenic ability of osteoblasts. PLGA microcapsules with different molecular weights (12 000, 30 000) encapsulating BMP-2, were prepared using a dual-channel microinjection pump. The morphology and structure of the microcapsules were characterized by optical microscopy and scanning electron microscopy. The sustained-release performance of the microcapsules was characterized by phosphate buffered saline immersion method. The cell compatibility of the microcapsules was detected by the Calcein-AM/PI staining and CCK-8 method. The chemotactic effect of BMP-2-encapsulated microcapsules on MC3T3-E1 cells after 48 h of treatment was detected by the Transwell assay. The alkaline phosphatase activity assay and Alizarin Red S staining were used to characterize the effect of microcapsules on the osteogenic ability of MC3T3-E1 cells. Both types of microcapsules with different molecular weights exhibited smooth surfaces, as well as uniform and good cell compatibility. The chemotactic effect of the 12 000 microcapsules was outstanding. The 30 000 microcapsules had a longer sustained-release time, and the initial burst release was reduced by approximately 25% compared with the 12 000 microcapsules. In addition, 30 000 microcapsules performed better in long-term osteogenesis induction than 12 000 microcapsules. In this study, the release of BMP-2 is regulated by adjusting the molecular weight of PLGA, and the results indicate that 30 000 microcapsules can better induce the long-term osteogenic ability of MC3T3-E1 cells.

Mathematical model for constructing a matrix of coefficients thermal conductivity under low-temperature influence on the multilayer epidermis of biological tissue

Objective. The purpose of the study is to develop a mathematical model for constructing a matrix of thermal conductivity coefficients under low-temperature exposure to the multilayer epidermis of biological tissue. Method. The research is based on methods of thermo dynamic analysis, full-scale and computational modeling of processes under low-temperature influence. Result. A mathematical model of the matrix of thermal conductivity coefficients under low-temperature effects on biological tissue has been developed. Heat transfer in a multilayer medium under low-temperature influence on biological tissue is presented as a process with a discrete step in time and coordinate. Thermal properties are evenly distributed throughout the volume of layers of biological tissue. For exposure to cold, layers of the epidermis of biological tissue were used, divided into uniform identical cells, where the cell area of the biological tissue was taken to be equal to one. Conclusion. The results obtained in this work can be useful to specialists who deal with modeling problems with free boundaries.

Zygospore formation in Zygnematophyceae predates several land plant traits.

Recent research on a special type of sexual reproduction and zygospore formation in Zygnematophyceae, the sister group of land plants, is summarized. Within this group, gamete fusion occurs by conjugation. Zygospore development in Mougeotia, Spirogyra and Zygnema is highlighted, which has recently been studied using Raman spectroscopy, allowing chemical imaging and detection of changes in starch and lipid accumulation. Three-dimensional reconstructions after serial block-face scanning electron microscopy (SBF-SEM) or focused ion beam SEM (FIB-SEM) made it possible to visualize and quantify cell wall and organelle changes during zygospore development. The zygospore walls undergo strong modifications starting from uniform thin cell walls to a multilayered structure. The mature cell wall is composed of a cellulosic endospore and exospore and a central mesospore built up by aromatic compounds. In Spirogyra, the exospore and endospore consist of thick layers of helicoidally arranged cellulose fibrils, which are otherwise only known from stone cells of land plants. While starch is degraded during maturation, providing building blocks for cell wall formation, lipid droplets accumulate and fill large parts of the ripe zygospores, similar to spores and seeds of land plants. Overall, data show similarities between streptophyte algae and embryophytes, suggesting that the genetic toolkit for many land plant traits already existed in their shared algal ancestor. This article is part of the theme issue 'The evolution of plant metabolism'.

Arp2/3 complex activity enables nuclear YAP for naïve pluripotency of human embryonic stem cells.

Our understanding of the transitions of human embryonic stem cells (hESCs) between distinct stages of pluripotency relies predominantly on regulation by transcriptional and epigenetic programs with limited insight on the role of established morphological changes. We report remodeling of the actin cytoskeleton of hESCs as they transition from primed to naïve pluripotency which includes assembly of a ring of contractile actin filaments encapsulating colonies of naïve hESCs. Activity of the Arp2/3 complex is required for formation of the actin ring, to establish uniform cell mechanics within naïve colonies, to promote nuclear translocation of the Hippo pathway effectors YAP and TAZ, and for effective transition to naïve pluripotency. RNA-sequencing analysis confirms that Arp2/3 complex activity regulates Hippo signaling in hESCs, and impaired naïve pluripotency with inhibited Arp2/3 complex activity is rescued by expressing a constitutively active, nuclear-localized YAP-S127A. Moreover, expression of YAP-S127A partially restores the actin filament fence with Arp2/3 complex inhibition, suggesting that actin filament remodeling is both upstream and downstream of YAP activity. These new findings on the cell biology of hESCs reveal a mechanism for cytoskeletal dynamics coordinating cell mechanics to regulate gene expression and facilitate transitions between pluripotency states.