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1.
Phys Rev Lett ; 125(3): 038003, 2020 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-32745423

RESUMO

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.


Assuntos
Células Epiteliais/química , Células Epiteliais/citologia , Rim/química , Rim/citologia , Modelos Biológicos , Animais , Fenômenos Biofísicos , Movimento Celular/fisiologia , Forma Celular/fisiologia , Cães , Transição Epitelial-Mesenquimal , Células Madin Darby de Rim Canino , Transição de Fase
2.
Nat Commun ; 11(1): 3805, 2020 07 30.
Artigo em Inglês | MEDLINE | ID: mdl-32732886

RESUMO

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.


Assuntos
Forma Celular/fisiologia , Células Epiteliais/citologia , Organoides/citologia , Organoides/crescimento & desenvolvimento , Animais , Fenômenos Biomecânicos , Proliferação de Células/fisiologia , Simulação por Computador , Modelos Biológicos , Tensão Superficial
3.
Proc Natl Acad Sci U S A ; 117(29): 17399-17408, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32641513

RESUMO

The cytoskeleton plays a key role in establishing robust cell shape. In animals, it is well established that cell shape can also influence cytoskeletal organization. Cytoskeletal proteins are well conserved between animal and plant kingdoms; nevertheless, because plant cells exhibit major structural differences to animal cells, the question arises whether the plant cytoskeleton also responds to geometrical cues. Recent numerical simulations predicted that a geometry-based rule is sufficient to explain the microtubule (MT) organization observed in cells. Due to their high flexural rigidity and persistence length of the order of a few millimeters, MTs are rigid over cellular dimensions and are thus expected to align along their long axis if constrained in specific geometries. This hypothesis remains to be tested in cellulo Here, we explore the relative contribution of geometry to the final organization of actin and MT cytoskeletons in single plant cells of Arabidopsis thaliana We show that the cytoskeleton aligns with the long axis of the cells. We find that actin organization relies on MTs but not the opposite. We develop a model of self-organizing MTs in three dimensions, which predicts the importance of MT severing, which we confirm experimentally. This work is a first step toward assessing quantitatively how cellular geometry contributes to the control of cytoskeletal organization in living plant cells.


Assuntos
Fenômenos Fisiológicos Celulares , Forma Celular/fisiologia , Citoesqueleto/fisiologia , Células Vegetais/fisiologia , Citoesqueleto de Actina/efeitos dos fármacos , Citoesqueleto de Actina/metabolismo , Citoesqueleto de Actina/ultraestrutura , Actinas , Arabidopsis/metabolismo , Citocalasina D/farmacologia , Microtúbulos/metabolismo , Células Vegetais/efeitos dos fármacos , Células Vegetais/ultraestrutura , Protoplastos
4.
Int J Nanomedicine ; 15: 3903-3920, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32606657

RESUMO

Background: Researchers are trying to study the mechanism of neural stem cells (NSCs) differentiation to oligodendrocyte-like cells (OLCs) as well as to enhance the selective differentiation of NSCs to oligodendrocytes. However, the limitation in nerve tissue accessibility to isolate the NSCs as well as their differentiation toward oligodendrocytes is still challenging. Purpose: In the present study, a hybrid polycaprolactone (PCL)-gelatin nanofiber scaffold mimicking the native extracellular matrix and axon morphology to direct the differentiation of bone marrow-derived NSCs to OLCs was introduced. Materials and Methods: In order to achieve a sustained release of T3, this factor was encapsulated within chitosan nanoparticles and chitosan-loaded T3 was incorporated within PCL nanofibers. Polyaniline graphene (PAG) nanocomposite was incorporated within gelatin nanofibers to endow the scaffold with conductive properties, which resemble the conductive behavior of axons. Biodegradation, water contact angle measurements, and scanning electron microscopy (SEM) observations as well as conductivity tests were used to evaluate the properties of the prepared scaffold. The concentration of PAG and T3-loaded chitosan NPs in nanofibers were optimized by examining the proliferation of cultured bone marrow-derived mesenchymal stem cells (BMSCs) on the scaffolds. The differentiation of BMSCs-derived NSCs cultured on the fabricated scaffolds into OLCs was analyzed by evaluating the expression of oligodendrocyte markers using immunofluorescence (ICC), RT-PCR and flowcytometric assays. Results: Incorporating 2% PAG proved to have superior cell support and proliferation while guaranteeing electrical conductivity of 10.8 × 10-5 S/cm. Moreover, the scaffold containing 2% of T3-loaded chitosan NPs was considered to be the most biocompatible samples. Result of ICC, RT-PCR and flow cytometry showed high expression of O4, Olig2, platelet-derived growth factor receptor-alpha (PDGFR-α), O1, myelin/oligodendrocyte glycoprotein (MOG) and myelin basic protein (MBP) high expressed but low expression of glial fibrillary acidic protein (GFAP). Conclusion: Considering surface topography, biocompatibility, electrical conductivity and gene expression, the hybrid PCL/gelatin scaffold with the controlled release of T3 may be considered as a promising candidate to be used as an in vitro model to study patient-derived oligodendrocytes by isolating patient's BMSCs in pathological conditions such as diseases or injuries. Moreover, the resulted oligodendrocytes can be used as a desirable source for transplanting in patients.


Assuntos
Materiais Biomiméticos/farmacologia , Células da Medula Óssea/citologia , Diferenciação Celular , Nanofibras/química , Células-Tronco Neurais/citologia , Oligodendroglia/citologia , Tecidos Suporte/química , Compostos de Anilina/química , Animais , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Condutividade Elétrica , Gelatina/química , Grafite/química , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Nanofibras/ultraestrutura , Células-Tronco Neurais/metabolismo , Oligodendroglia/efeitos dos fármacos , Poliésteres/química , Ratos , Suínos , Tri-Iodotironina/farmacologia
5.
Int J Nanomedicine ; 15: 4471-4481, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32606689

RESUMO

Background: Ineffective integration has been recognized as one of the major causes of early orthopedic failure of titanium-based implants. One strategy to address this problem is to develop modified titanium surfaces that promote osteoblast differentiation. This study explored titanium surfaces modified with TiO2 nanotubes (TiO2 NTs) capable of localized drug delivery into bone and enhanced osteoblast cell differentiation. Materials and Methods: Briefly, TiO2 NTs were subjected to anodic oxidation and loaded with Metformin, a widely used diabetes drug. To create surfaces with sustainable drug-eluting characteristics, TiO2 NTs were spin coated with a thin layer of chitosan. The surfaces were characterized via scanning electron microscopy, atomic force microscopy, and contact angle measurements. The surfaces were then exposed to mesenchymal bone marrow stem cells (MSCs) to evaluate cell adhesion, growth, differentiation, and morphology on the modified surfaces. Results: A noticeable increase in drug release time (3 days vs 20 days) and a decrease in burst release characteristics (85% to 7%) was observed in coated samples as compared to uncoated samples, respectively. Chitosan-coated TiO2 NTs exhibited a considerable enhancement in cell adhesion, proliferation, and genetic expression of type I collagen, and alkaline phosphatase activity as compared to uncoated TiO2 NTs. Conclusion: TiO2 NT surfaces with a chitosan coating are capable of delivering Metformin to a bone site over a sustained period of time with the potential to enhance MSCs cell attachment, proliferation, and differentiation.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Quitosana/química , Liberação Controlada de Fármacos , Metformina/farmacologia , Nanotubos/química , Osteoblastos/citologia , Titânio/química , Fosfatase Alcalina/metabolismo , Animais , Adesão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Nanotubos/ultraestrutura , Osteoblastos/efeitos dos fármacos , Osteoblastos/ultraestrutura , Osteogênese/efeitos dos fármacos , Ratos Wistar , Molhabilidade
6.
Int J Nanomedicine ; 15: 4351-4362, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32606682

RESUMO

Purpose: The present study synthesized silver nanoparticles (AgNPs) using the aqueous extract of a traditional medicinal product consisting of an oleoresin (a combination of macromolecules of carbohydrates and proteins) exuded from the rhizome of the plant Ferula foetida (asafoetida gum) and evaluated its biological properties. Materials and Methods: The silver nanoparticles synthesized using asafoetida gum (As-AgNPs) were characterized using UV/Vis spectroscopy, fourier infrared (FTIR) spectroscopy, scanning electron microscopy (SEM) with energy dispersive X-ray analysis (EDX), X-ray diffraction (XRD) and transmission electron microscopy (TEM) and EADX. The cytotoxicity and antimicrobial activity As-AgNPs were evaluated against MCF-7 cell lines and selected microbial pathogens, respectively. Results: The synthesized silver nanoparticles were crystalline in nature with a spherical shape. The average particle size was 5.6-8.6 nm. The cytotoxicity of the synthesized As-AgNPs was evaluated against MCF-7 cell lines, and the As-AgNPs were found to be effective in inhibiting the multiplication of cancer cells. The As-AgNPs exhibited significant antimicrobial activity towards E. coli, K. pneumoniae and C. albicans. The MIC of the synthesized As-AgNPs was 7.80 µg/mL for E. coli ATCC 25922, Salmonella sp. WS50- and S. typhi; 15.60 µg/mL for S. typhimurium and S. aureus WS10, and 31.20 µg/mL for K. pneumoniae and S. aureus ATCC 43300-MRSA. In addition, MIC values of 15.60 µg/mL for C. albicans ATCC8436 and 31.20 µg/mL for C. krusei ATCC6258 were obtained. Conclusion: As asafoetida is a good traditional medicine, its involvement in the synthesis of AgNPs led the silver nanoparticles to exhibit good cytotoxic and antimicrobial effects.


Assuntos
Anti-Infecciosos/farmacologia , Ferula/química , Nanopartículas Metálicas/química , Gomas Vegetais/química , Prata/farmacologia , Antibacterianos/farmacologia , Antifúngicos/farmacologia , Candida albicans/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Humanos , Células MCF-7 , Nanopartículas Metálicas/ultraestrutura , Testes de Sensibilidade Microbiana , Tamanho da Partícula , Extratos Vegetais/química , Staphylococcus aureus/efeitos dos fármacos , Difração de Raios X
7.
PLoS One ; 15(7): e0236286, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32702004

RESUMO

Functional brown adipose tissue (BAT) was identified in adult humans only in 2007 with the use of fluorodeoxyglucose positron emission tomography imaging. Previous studies have demonstrated a negative correlation between obesity and BAT presence in humans. It is proposed that BAT possesses the capacity to increase metabolism and aid weight loss. In rodents it is well established that BAT is stimulated by the sympathetic nervous system with the interscapular BAT being innervated via branches of intercostal nerves. Whilst there is evidence to suggest that BAT possesses beta-3 adrenoceptors, no studies have identified the specific nerve branch that carries sympathetic innervation to BAT in humans. The aim of this study was to identify and trace the peripheral nerve or nerves that innervate human BAT in the supraclavicular region. The posterior triangle region of the neck of cadaveric specimens were dissected in order to identify any peripheral nerve branches piercing and/or terminating in supraclavicular BAT. A previously undescribed branch of the cervical plexus terminating in a supraclavicular adipose depot was identified in all specimens. This was typically an independent branch of the plexus, from the third cervical spinal nerve, but in one specimen was a branch of the supraclavicular nerve. Histological analysis revealed the supraclavicular adipose depot contained tyrosine hydroxylase immunoreactive structures, which likely represent sympathetic axons. This is the first study that identifies a nerve branch to supraclavicular BAT-like tissue. This finding opens new avenues for the investigation of neural regulation of fat metabolism in humans.


Assuntos
Tecido Adiposo/inervação , Clavícula/inervação , Adipócitos/citologia , Tecido Adiposo/anatomia & histologia , Idoso , Cadáver , Forma Celular , Clavícula/anatomia & histologia , Dissecação , Humanos , Tirosina 3-Mono-Oxigenase/metabolismo
8.
Int J Nanomedicine ; 15: 3695-3716, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32547023

RESUMO

Purpose: External and internal stimuli easily affect the retina. Studies have shown that cells infected with Toxoplasma gondii are resistant to multiple inducers of apoptosis. Nanoparticles (NPs) have been widely used in biomedical fields; however, little is known about cytotoxicity caused by NPs in the retina and the modulators that inhibit nanotoxicity. Materials and Methods: ARPE-19 cells from human retinal pigment epithelium were treated with silver nanoparticles (AgNPs) alone or in combination with T. gondii. Then, the cellular toxicity, apoptosis, cell cycle analysis, autophagy, ROS generation, NOX4 expression, and MAPK/mTOR signaling pathways were investigated. To confirm the AgNP-induced cytotoxicity in ARPE-19 cells and its modulatory effects caused by T. gondii infection, the major experiments carried out in ARPE-19 cells were performed again using human foreskin fibroblast (HFF) cells and bone marrow-derived macrophages (BMDMs) from NOX4-/ - mice. Results: AgNPs dose-dependently induced cytotoxicity and cell death in ARPE-19 cells. Apoptosis, sub-G1 phase cell accumulation, autophagy, JNK phosphorylation, and mitochondrial apoptotic features, such as caspase-3 and PARP cleavages, mitochondrial membrane potential depolarization, and cytochrome c release into the cytosol were observed in AgNP-treated cells. AgNP treatment also increased the Bax, Bik, and Bim protein levels as well as NOX4-dependent ROS generation. However, T. gondii-infected ARPE-19 cells inhibited AgNP-induced apoptosis, JNK phosphorylation, sub-G1 phase cell accumulation, autophagy, NOX4-mediated ROS production, and mitochondrial apoptosis. Furthermore, mitochondrial apoptosis was found in AgNP-treated HFF cells and BMDMs, and AgNP-induced mitochondrial apoptosis inhibition via NOX4-dependent ROS suppression in T. gondii pre-infected HFF cells and BMDMs was also confirmed. Conclusion: AgNPs induced mitochondrial apoptosis in human RPE cells combined with cell cycle dysregulation and autophagy; however, these effects were significantly inhibited by T. gondii pre-infection by suppression of NOX4-mediated ROS production, suggesting that T. gondii is a strong inhibitory modulator of nanotoxicity in in vitro models.


Assuntos
Apoptose/efeitos dos fármacos , Nanopartículas Metálicas/química , NADPH Oxidase 4/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Epitélio Pigmentado da Retina/parasitologia , Prata/farmacologia , Toxoplasmose/patologia , Animais , Autofagia/efeitos dos fármacos , Linhagem Celular , Forma Celular/efeitos dos fármacos , Modelos Animais de Doenças , Fibroblastos/efeitos dos fármacos , Fibroblastos/parasitologia , Fase G1/efeitos dos fármacos , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Modelos Biológicos , Fosforilação/efeitos dos fármacos
9.
PLoS One ; 15(6): e0234641, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32574164

RESUMO

Chondrocytes, comparable to many cells from the connective tissue, dedifferentiate and end up in a similar fibroblastoid cell type, marked by the loss of the specific expression pattern. Here, chondrocytes isolated from osteoarthritic (OA) patients were investigated. The OA chondrocytes used in this work were not affected by the loss of specific gene expression upon cell culture. The mRNA levels of known cartilage markers, such as SOX5, SOX6, SOX9, aggrecan and proteoglycan 4, remained unchanged. Since chondrocytes from OA and healthy tissue show different DNA methylation patterns, the underlying mechanisms of cartilage marker maintenance were investigated with a focus on the epigenetic modification by DNA methylation. The treatment of dedifferentiated chondrocytes with the DNA methyltransferase inhibitor 5-aza-2´-deoxycytidine (5-aza-dC) displayed no considerable impact on the maintenance of marker gene expression observed in the dedifferentiated state, while the chondrogenic differentiation capacity was compromised. On the other hand, the pre-cultivation with 5-aza-dC improved the osteogenesis and adipogenesis of OA chondrocytes. Contradictory to these effects, the DNA methylation levels were not reduced after treatment for four weeks with 1 µM 5-aza-dC. In conclusion, 5-aza-dC affects the differentiation capacity of OA chondrocytes, while the global DNA methylation level remains stable. Furthermore, dedifferentiated chondrocytes isolated from late-stage OA patients represent a reliable cell source for in vitro studies and disease models without the need for additional alterations.


Assuntos
Condrócitos/patologia , Decitabina/farmacologia , Osteoartrite/patologia , Adipogenia/efeitos dos fármacos , Adipogenia/genética , Biomarcadores/metabolismo , Cartilagem Articular/efeitos dos fármacos , Cartilagem Articular/metabolismo , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Forma Celular/efeitos dos fármacos , Forma Celular/genética , Células Cultivadas , Condrócitos/efeitos dos fármacos , Condrócitos/metabolismo , Colágeno Tipo II/metabolismo , DNA (Citosina-5-)-Metiltransferases/metabolismo , Metilação de DNA/efeitos dos fármacos , Metilação de DNA/genética , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Osteoartrite/genética , Osteogênese/efeitos dos fármacos , Osteogênese/genética
10.
PLoS Genet ; 16(6): e1008717, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32479493

RESUMO

Forces generated by the actomyosin cytoskeleton are key contributors to many morphogenetic processes. The actomyosin cytoskeleton organises in different types of networks depending on intracellular signals and on cell-cell and cell-extracellular matrix (ECM) interactions. However, actomyosin networks are not static and transitions between them have been proposed to drive morphogenesis. Still, little is known about the mechanisms that regulate the dynamics of actomyosin networks during morphogenesis. This work uses the Drosophila follicular epithelium, real-time imaging, laser ablation and quantitative analysis to study the role of integrins on the regulation of basal actomyosin networks organisation and dynamics and the potential contribution of this role to cell shape. We find that elimination of integrins from follicle cells impairs F-actin recruitment to basal medial actomyosin stress fibers. The available F-actin redistributes to the so-called whip-like structures, present at tricellular junctions, and into a new type of actin-rich protrusions that emanate from the basal cortex and project towards the medial region. These F-actin protrusions are dynamic and changes in total protrusion area correlate with periodic cycles of basal myosin accumulation and constriction pulses of the cell membrane. Finally, we find that follicle cells lacking integrin function show increased membrane tension and reduced basal surface. Furthermore, the actin-rich protrusions are responsible for these phenotypes as their elimination in integrin mutant follicle cells rescues both tension and basal surface defects. We thus propose that the role of integrins as regulators of stress fibers plays a key role on controlling epithelial cell shape, as integrin disruption promotes reorganisation into other types of actomyosin networks, in a manner that interferes with proper expansion of epithelial basal surfaces.


Assuntos
Actomiosina/metabolismo , Forma Celular , Proteínas de Drosophila/metabolismo , Células Epiteliais/metabolismo , Integrinas/metabolismo , Fibras de Estresse/metabolismo , Animais , Membrana Celular/metabolismo , Drosophila , Células Epiteliais/citologia , Fibras de Estresse/ultraestrutura
11.
Proc Natl Acad Sci U S A ; 117(27): 16027-16034, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32571946

RESUMO

Puzzle-shaped pavement cells provide a powerful model system to investigate the cellular and subcellular processes underlying complex cell-shape determination in plants. To better understand pavement cell-shape acquisition and the role of auxin in this process, we focused on the spirals of young stomatal lineage ground cells of Arabidopsis leaf epidermis. The predictability of lobe formation in these cells allowed us to demonstrate that the auxin response gradient forms within the cells of the spiral and fluctuates based on the particular stage of lobe development. We revealed that specific localization of auxin transporters at the different membranes of these young cells changes during the course of lobe formation, suggesting that these fluctuating auxin response gradients are orchestrated via auxin transport to control lobe formation and determine pavement cell shape.


Assuntos
Arabidopsis/metabolismo , Forma Celular/efeitos dos fármacos , Forma Celular/fisiologia , Ácidos Indolacéticos/metabolismo , Ácidos Indolacéticos/farmacologia , Proteínas de Arabidopsis , Transporte Biológico , Epiderme Vegetal/metabolismo , Folhas de Planta/metabolismo , Estômatos de Plantas/metabolismo
12.
J Vis Exp ; (158)2020 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-32364544

RESUMO

Macrophages are present in most vertebrate tissues and comprise widely dispersed and heterogeneous cell populations with different functions. They are key players in health and disease, acting as phagocytes during immune defense and mediating trophic, maintenance, and repair functions. Although it has been possible to study some of the molecular processes involved in human macrophage function, it has proved difficult to apply genetic engineering techniques to primary human macrophages. This has significantly hampered our ability to interrogate the complex genetic pathways involved in macrophage biology and to generate models for specific disease states. An off-the-shelf source of human macrophages that is amenable to the vast arsenal of genetic manipulation techniques would, therefore, provide a valuable tool in this field. We present an optimized protocol that allows for the generation of macrophages from human induced pluripotent stem cells (iPSCs) in vitro. These iPSC-derived macrophages (iPSC-DMs) express human macrophage cell surface markers, including CD45, 25F9, CD163, and CD169, and our live-cell imaging functional assay demonstrates that they exhibit robust phagocytic activity. Cultured iPSC-DMs can be activated to different macrophage states that display altered gene expression and phagocytic activity by the addition of LPS and IFNg, IL4, or IL10. Thus, this system provides a platform to generate human macrophages carrying genetic alterations that model specific human disease and a source of cells for drug screening or cell therapy to treat these diseases.


Assuntos
Técnicas de Cultura de Células/métodos , Células-Tronco Pluripotentes Induzidas/citologia , Macrófagos/citologia , Biomarcadores/metabolismo , Contagem de Células , Diferenciação Celular , Membrana Celular/metabolismo , Polaridade Celular , Forma Celular , Células Cultivadas , Corpos Embrioides/citologia , Humanos , Macrófagos/metabolismo , Fagocitose , Fenótipo
13.
PLoS Comput Biol ; 16(5): e1007611, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32379821

RESUMO

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.


Assuntos
Neoplasias Encefálicas/patologia , Biologia Computacional/métodos , Glioma/patologia , Contagem de Células/métodos , Movimento Celular/fisiologia , Forma Celular/fisiologia , Humanos , Modelos Biológicos , Modelos Teóricos , Simulação de Dinâmica Molecular
14.
Int J Nanomedicine ; 15: 2633-2646, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32368045

RESUMO

Objective: The aim of this study is to fabricate functional scaffolds to gene delivery bone morphogenetic protein-2 (BMP-2) plasmid for bone formation in bone tissue engineering. Methods: Dendriplexes (DPs) of generation 4 polyamidoamin (G4-PAMAM)/BMP-2 plasmid were prepared through microfluidic (MF) platform. The physiochemical properties and toxicity of DPs were evaluated by DLS, AFM, FESEM and MTT assay. In order to create a suitable environment for stem cell growth and differentiation, poly-l-lactic acid (PLLA) and poly-l-lactic acid/poly (ethylene oxide) (PLLA/PEO) scaffolds containing hydroxyapatite nanoparticles (HA) and DPs were fabricated by the electrospinning method. The osteogenic potency of the scaffolds on human adipose tissue-derived mesenchymal stem cells (hASCs) was investigated. Results: The results revealed that tuning the physical properties of DPs by adjusting flow parameters in microfluidic platform can easily improve the cell viability compared to conventional bulk mixing method. Also, the result showed that the presence of HA and DPs in PLLA/PEO scaffold enhanced alkaline phosphatase (ALP) activity and increased the amount of deposited Ca, as well as, related to osteogenesis gen markers. Conclusion: This study indicated that on using the MF platform in preparation of DPs and loading them along with HA in PLLA/PEO scaffold, the osteogenic differentiation of hASCs could be tuned.


Assuntos
Proteína Morfogenética Óssea 2/metabolismo , Osso e Ossos/fisiologia , Durapatita/química , Microfluídica , Nanofibras/química , Poliaminas/química , Engenharia Tecidual/métodos , Tecidos Suporte/química , Fosfatase Alcalina/metabolismo , Cálcio/metabolismo , Adesão Celular , Morte Celular , Diferenciação Celular , Proliferação de Células , Forma Celular , DNA/metabolismo , Dendrímeros/química , Humanos , Células-Tronco Mesenquimais/metabolismo , Nanopartículas/química , Tamanho da Partícula , Plasmídeos/metabolismo , Poliésteres/química , Resistência à Tração
15.
Adv Exp Med Biol ; 1246: 129-151, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32399829

RESUMO

Although the cytosolic Ca2+ signalling event in phagocytosis is well established, and the mechanism for generating such signals also understood, the target for the Ca2+ signal and how this relates to the phagocytic outcome is less clear. In this chapter, we present the evidence for a role of the Ca2+ activated protease, calpain, in phagocytosis. The abundant evidence for Ca2+ changes and calpain activation during cell shape changes is extended to include the specific cell shape change which accompanies phagocytosis. The discussion therefore includes a brief description of the domain structure of calpain and their functions. Also the mechanism by which calpain activation is limited at the cell periphery subdomains, and how this would allow phagocytic pseudopodia to form locally.


Assuntos
Cálcio/metabolismo , Calpaína/metabolismo , Fagocitose , Animais , Forma Celular , Citosol/metabolismo , Ativação Enzimática , Humanos , Pseudópodes/metabolismo
16.
PLoS One ; 15(5): e0232759, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32453737

RESUMO

SUMMARY: Reprogramming autologous adult cells to pluripotent cells allows for relatively safe cell replacement therapy. This can be achieved by nuclear transfer, cell fusion, or induced pluripotent stem cell technology However, the epigenetic memory of the cell is considered as a great challenge facing the complete reprograming of cells by these methods. Introducing oocyte-specific factors into differentiated cells may present a promising approach by mimicking cellular reprogramming during fertilization. METHODS: Human bone marrow mesenchymal stromal cells (hBM-MSCs) were cultured with different concentrations of human metaphase II (M II) oocyte extract (0.1, 1, 5, 10, 30 ng/µl). Reprogramming was assessed at various exposure times (1, 4, 7 days). Cells were tested for their proliferation rate, morphological changes, expression of pluripotency markers, expression of mesenchymal to epithelial transition markers, and mitochondrial rejuvenation. (mitochondrial localization, morphological changes, bioenergetics, transmembrane potential, and levels of reactive oxygen species, ROS). RESULTS: Treatment of human BM-MSCs with 10 ng/µl oocyte extract resulted in increased cell proliferation, which was associated with the upregulation of the pluripotency genes OCT-4, NANOG, and SOX-2 and a concomitant downregulation of mesenchymal-specific genes. MSCs exhibited small, immature round mitochondria with few swollen cristae localized proximal to the cell nucleus. This was accompanied by morphological cell changes, a metabolic shift towards oxidative phosphorylation, a high mitochondrial membrane potential, and increased ROS production. CONCLUSION: These data show that treatment with 10 ng/µl human MII-phase oocyte extract induced genetic and mitochondrial reprogramming of human BM-MSCs to a more embryonic phenotype.


Assuntos
Extratos Celulares/farmacologia , Reprogramação Celular/genética , Células-Tronco Mesenquimais/metabolismo , Mitocôndrias/metabolismo , Oócitos/metabolismo , Biomarcadores/metabolismo , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Células Cultivadas , Reprogramação Celular/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Potenciais da Membrana/efeitos dos fármacos , Células-Tronco Mesenquimais/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/ultraestrutura , Oócitos/efeitos dos fármacos , Consumo de Oxigênio/efeitos dos fármacos , Fatores de Tempo
17.
Int J Nanomedicine ; 15: 3281-3290, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32440124

RESUMO

Introduction: Cells exhibit high sensitivity and a diverse response to the nanotopography of the extracellular matrix, thereby endowing materials with instructive performances formerly reserved for growth factors. This finding leads to opportunities for improvement. However, the interplay between the topographical surface and cell behaviors remains incompletely understood. Methods: In the present study, we showed nanosurfaces with various dimensions of nanopits (200-750 nm) fabricated by self-assembling polystyrene (PS) nanospheres. Human adipose-derived stem cell behaviors, such as cell morphology, adhesion, cytoskeleton contractility, proliferation, and differentiation, were investigated on the prepared PS nanopit surface. Results: The osteogenic differentiation can be enhanced by nanopits with a diameter of 300-400 nm. Discussion: The present study provided exciting new avenues to investigate cellular responses to well-defined nanoscale topographic features, which could further guide bone tissue engineering and stem cell clinical research. The capability to control developing biomaterials mimicking nanotopographic surfaces promoted functional tissue engineering, such as artificial joint replacement, bone repair, and dental applications.


Assuntos
Tecido Adiposo/citologia , Diferenciação Celular , Nanoestruturas/química , Osteogênese , Poliestirenos/farmacologia , Células-Tronco/citologia , Fosfatase Alcalina/genética , Fosfatase Alcalina/metabolismo , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Nanoestruturas/ultraestrutura , Osteogênese/efeitos dos fármacos , Células-Tronco/efeitos dos fármacos , Células-Tronco/ultraestrutura
18.
Proc Natl Acad Sci U S A ; 117(24): 13541-13551, 2020 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-32467168

RESUMO

Within developing embryos, tissues flow and reorganize dramatically on timescales as short as minutes. This includes epithelial tissues, which often narrow and elongate in convergent extension movements due to anisotropies in external forces or in internal cell-generated forces. However, the mechanisms that allow or prevent tissue reorganization, especially in the presence of strongly anisotropic forces, remain unclear. We study this question in the converging and extending Drosophila germband epithelium, which displays planar-polarized myosin II and experiences anisotropic forces from neighboring tissues. We show that, in contrast to isotropic tissues, cell shape alone is not sufficient to predict the onset of rapid cell rearrangement. From theoretical considerations and vertex model simulations, we predict that in anisotropic tissues, two experimentally accessible metrics of cell patterns-the cell shape index and a cell alignment index-are required to determine whether an anisotropic tissue is in a solid-like or fluid-like state. We show that changes in cell shape and alignment over time in the Drosophila germband predict the onset of rapid cell rearrangement in both wild-type and snail twist mutant embryos, where our theoretical prediction is further improved when we also account for cell packing disorder. These findings suggest that convergent extension is associated with a transition to more fluid-like tissue behavior, which may help accommodate tissue-shape changes during rapid developmental events.


Assuntos
Forma Celular , Drosophila/crescimento & desenvolvimento , Animais , Anisotropia , Drosophila/citologia , Drosophila/genética , Drosophila/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Epitélio/metabolismo , Miosina Tipo II/genética , Miosina Tipo II/metabolismo
19.
Mol Pharmacol ; 97(6): 365-376, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32234808

RESUMO

Proteinase-activated receptors (PARs) are a four-member family of G-protein-coupled receptors that are activated via proteolysis. PAR4 is a member of this family that is cleaved and activated by serine proteinases such as thrombin, trypsin, and cathepsin-G. PAR4 is expressed in a variety of tissues and cell types, including platelets, vascular smooth muscle cells, and neuronal cells. In studying PAR4 signaling and trafficking, we observed dynamic changes in the cell membrane, with spherical membrane protrusions that resemble plasma membrane blebbing. Since nonapoptotic membrane blebbing is now recognized as an important regulator of cell migration, cancer cell invasion, and vesicular content release, we sought to elucidate the signaling pathway downstream of PAR4 activation that leads to such events. Using a combination of pharmacological inhibition and CRISPR/CRISPR-associated protein 9 (Cas9)-mediated gene editing approaches, we establish that PAR4-dependent membrane blebbing occurs independently of the Gα q/11- and Gα i-signaling pathways and is dependent on signaling via the ß-arrestin-1/2 and Ras homolog family member A (RhoA) signaling pathways. Together these studies provide further mechanistic insight into PAR4 regulation of cellular function. SIGNIFICANCE STATEMENT: We find that the thrombin receptor PAR4 triggers cell membrane blebbing in a RhoA-and ß-arrestin-dependent manner. In addition to identifying novel cellular responses mediated by PAR4, these data provide further evidence for biased signaling in PAR4 since membrane blebbing was dependent on some, but not all, signaling pathways activated by PAR4.


Assuntos
Membrana Celular/metabolismo , Membrana Celular/patologia , Receptores de Trombina/metabolismo , beta-Arrestinas/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismo , Animais , Sistemas CRISPR-Cas , Forma Celular , Técnicas de Inativação de Genes , Células HEK293 , Humanos , Músculo Liso Vascular/metabolismo , Ratos , Ratos Endogâmicos WKY , Receptores Acoplados a Proteínas-G/metabolismo , Receptores de Trombina/agonistas , Transdução de Sinais
20.
Arch Microbiol ; 202(7): 1677-1685, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32285166

RESUMO

Triterpenoid compounds are important bioactivity materials. Morchella is an abundant medicinal fungi found worldwide. In this study, we optimised the isolation and purification of triterpenoid compounds from Morchella mycelium fermentation. The results showed that the triterpenoid compounds yield was 35.22 mg/g, and we also identified two triterpenoid compounds using high-performance liquid chromatography. In addition, we evaluated the anti-tumour and antioxidant activity of the products, and the results showed that triterpenoid compounds from Morchella mycelium fermentation showed good bioactivity. The IC50 values of four cancer cell lines treated with the triterpenoid compounds for 48 h were 7.20, 14.96, 4.41, and 13.43 mg/mL, respectively. Morphological changes associated with the apoptosis of PC-3 cells were observed using confocal scanning laser microscopy after treatment with triterpenoid compounds for 48 and 72 h. The triterpenoid compounds also exhibited DPPH radical, hydroxyl, and ABTS-free radical scavenging activities in vitro. These results suggest that triterpenoid compounds from Morchella mycelium fermentation, which are found in functional foods and used in the field of pharmacology, might be excellent products for the treatment of cancer and age-related illnesses.


Assuntos
Ascomicetos/química , Micélio/química , Triterpenos/isolamento & purificação , Triterpenos/farmacologia , Antineoplásicos/farmacologia , Antioxidantes/farmacologia , Linhagem Celular Tumoral , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Fermentação , Humanos , Radical Hidroxila/química , Concentração Inibidora 50 , Células PC-3
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