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Estévez M, Batoni E, Cicuéndez M, Bonatti AF, Fernández-Marcelo T, De Maria C, González B, Izquierdo-Barba I, Vozzi G. Fabrication of 3D Biofunctional Magnetic Scaffolds by Combining Fused Deposition Modelling and Inkjet Printing of Superparamagnetic Iron Oxide Nanoparticles. Tissue Eng Regen Med 2025:10.1007/s13770-025-00711-2. [PMID: 40100619 DOI: 10.1007/s13770-025-00711-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2024] [Revised: 01/31/2025] [Accepted: 02/03/2025] [Indexed: 03/20/2025] Open
Abstract
BACKGROUND Recently, magnetic composite biomaterials have raised attention in bone tissue engineering as the application of dynamic magnetic fields proved to modulate the proliferation and differentiation of several cell types. METHODS This study presents a novel method to fabricate biofunctional magnetic scaffolds by the deposition of superparamagnetic iron oxide nanoparticles (SPIONs) through thermal Drop-On-Demand inkjet printing on three-dimensional (3D) printed scaffolds. Firstly, 3D scaffolds based on thermoplastic polymeric composed by poly-L-lactic acid/poly-caprolactone/poly(3-hydroxybutyrate-co-3-hydroxyvalerate) were fabricated by Fused Deposition Modelling. Then, in a second step, SPIONs were incorporated onto the surface of the scaffolds by inkjet printing following a designed 2D pattern. RESULTS A complete characterization of the resulting magnetic scaffolds was carried out attending to the surface SPIONs deposits, demonstrating the accuracy and versatility of the production technique, as well as the stability under physiological conditions and the magnetic properties. Biological evaluation with human bone marrow mesenchymal stems cells demonstrated biocompatibility of the scaffolds and increased osteogenic capability under the application of a magnetic field, due to the activation of mechanotransduction processes. CONCLUSION These results show that the developed 3D magnetic biofunctional scaffolds can be a very promising tool for advanced and personalised bone regeneration treatments.
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Affiliation(s)
- Manuel Estévez
- Departamento de Química en Ciencias Farmacéuticas, Facultad de Farmacia, Instituto de Investigación Sanitaria, Universidad Complutense de Madrid, Hospital 12 de Octubre i+12, Plaza Ramón y Cajal s/n, 28040, Madrid, Spain
| | - Elisa Batoni
- Department of Information Engineering, University of Pisa, Via Girolamo Caruso 16, 56122, Pisa, Italy
- Research Center "E. Piaggio", University of Pisa, Via Largo Lucio Lazzarino 1, 56122, Pisa, Italy
| | - Mónica Cicuéndez
- Departamento de Química en Ciencias Farmacéuticas, Facultad de Farmacia, Instituto de Investigación Sanitaria del Hospital Clínico San Carlos (IdISSC), Universidad Complutense de Madrid, 28040, Madrid, Spain
| | - Amedeo Franco Bonatti
- Research Center "E. Piaggio", University of Pisa, Via Largo Lucio Lazzarino 1, 56122, Pisa, Italy
| | - Tamara Fernández-Marcelo
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad Complutense de Madrid, Plaza Ramón y Cajal, s/n, 28040, Madrid, Spain
| | - Carmelo De Maria
- Department of Information Engineering, University of Pisa, Via Girolamo Caruso 16, 56122, Pisa, Italy.
- Research Center "E. Piaggio", University of Pisa, Via Largo Lucio Lazzarino 1, 56122, Pisa, Italy.
| | - Blanca González
- Departamento de Química en Ciencias Farmacéuticas, Facultad de Farmacia, Instituto de Investigación Sanitaria, Universidad Complutense de Madrid, Hospital 12 de Octubre i+12, Plaza Ramón y Cajal s/n, 28040, Madrid, Spain
- Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), 28029, Madrid, Spain
| | - Isabel Izquierdo-Barba
- Departamento de Química en Ciencias Farmacéuticas, Facultad de Farmacia, Instituto de Investigación Sanitaria, Universidad Complutense de Madrid, Hospital 12 de Octubre i+12, Plaza Ramón y Cajal s/n, 28040, Madrid, Spain.
- Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), 28029, Madrid, Spain.
| | - Giovanni Vozzi
- Department of Information Engineering, University of Pisa, Via Girolamo Caruso 16, 56122, Pisa, Italy
- Research Center "E. Piaggio", University of Pisa, Via Largo Lucio Lazzarino 1, 56122, Pisa, Italy
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2
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Albrecht FB, Schick AK, Klatt A, Schmidt FF, Nellinger S, Kluger PJ. Exploring Morphological and Molecular Properties of Different Adipose Cell Models: Monolayer, Spheroids, Gellan Gum-Based Hydrogels, and Explants. Macromol Biosci 2025; 25:e2400320. [PMID: 39450850 PMCID: PMC11904394 DOI: 10.1002/mabi.202400320] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/18/2024] [Revised: 10/04/2024] [Indexed: 10/26/2024]
Abstract
White adipose tissue (WAT) plays a crucial role in energy homeostasis and secretes numerous adipokines with far-reaching effects. WAT is linked to diseases such as diabetes, cardiovascular disease, and cancer. There is a high demand for suitable in vitro models to study diseases and tissue metabolism. Most of these models are covered by 2D-monolayer cultures. This study aims to evaluate the performance of different WAT models to better derive potential applications. The stability of adipocyte characteristics in spheroids and two 3D gellan gum hydrogels with ex situ lobules and 2D-monolayer culture is analyzed. First, the differentiation to achieve adipocyte-like characteristics is determined. Second, to evaluate the maintenance of differentiated ASC-based models, an adipocyte-based model, and explants over 3 weeks, viability, intracellular lipid content, perilipin A expression, adipokine, and gene expression are analyzed. Several advantages are supported using each of the models. Including, but not limited to, the strong differentiation in 2D-monolayers, the self-assembling within spheroids, the long-term stability of the stem cell-containing hydrogels, and the mature phenotype within adipocyte-containing hydrogels and the lobules. This study highlights the advantages of 3D models due to their more in vivo-like behavior and provides an overview of the different adipose cell models.
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Affiliation(s)
- Franziska B Albrecht
- Reutlingen Research Institute, Reutlingen University, Alteburgstraße 150, 72762, Reutlingen, Germany
- Faculty of Natural Science, University of Hohenheim, Schloss Hohenheim 1, 70599, Stuttgart, Germany
| | - Ann-Kathrin Schick
- Faculty of Science, Energy and Building Services, Esslingen University, Kanalstraße 33, 73728, Esslingen, Germany
| | - Annemarie Klatt
- Reutlingen Research Institute, Reutlingen University, Alteburgstraße 150, 72762, Reutlingen, Germany
| | - Freia F Schmidt
- Reutlingen Research Institute, Reutlingen University, Alteburgstraße 150, 72762, Reutlingen, Germany
| | - Svenja Nellinger
- Reutlingen Research Institute, Reutlingen University, Alteburgstraße 150, 72762, Reutlingen, Germany
| | - Petra J Kluger
- School of Life Sciences, Reutlingen University, Alteburgstraße 150, 72762, Reutlingen, Germany
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Burčík D, Macko J, Podrojková N, Demeterová J, Stano M, Oriňak A. Role of Cell Adhesion in Cancer Metastasis Formation: A Review. ACS OMEGA 2025; 10:5193-5213. [PMID: 39989825 PMCID: PMC11840620 DOI: 10.1021/acsomega.4c08140] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/09/2024] [Revised: 01/10/2025] [Accepted: 01/22/2025] [Indexed: 02/25/2025]
Abstract
Intercellular adhesion is accompanied by several physical quantities and actions. In this review, we tried to collect information about the influence of surface energy and its impact on cell-cell adhesion. It still undergoes development for cancer treatment. Data on receptor-ligand interactions that occur on circulating tumor cells (CTCs) are described, and adhesion receptors as therapeutic targets are collected. Additionally, the impact of surface roughness on the interactions between CTC cells and the surface was monitored. The effects of different cell adhesion molecules (CAMs) on cell adhesion, growth, and proliferation were investigated. This review offers general principles of cell adhesion, through the blockade of adhesion with blocking drugs and inhibitors like computational models that describe the process of adhesion. Some theoretical models based on the minimum of the total free energy of interaction between CAMs and selected organic molecules have been presented. The final aim was to find information on how modulation of the surface of CTCs (by medicals or physically) inhibits cancer metastases formation.
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Affiliation(s)
- Denis Burčík
- University of P. J. Safarik
in Kosice, Faculty of Sciences,
Institute of Chemistry, Department of Physical Chemistry, Moyzesova 11, 041 01 Kosice, Slovakia
| | - Ján Macko
- University of P. J. Safarik
in Kosice, Faculty of Sciences,
Institute of Chemistry, Department of Physical Chemistry, Moyzesova 11, 041 01 Kosice, Slovakia
| | - Natália Podrojková
- University of P. J. Safarik
in Kosice, Faculty of Sciences,
Institute of Chemistry, Department of Physical Chemistry, Moyzesova 11, 041 01 Kosice, Slovakia
| | - Jana Demeterová
- University of P. J. Safarik
in Kosice, Faculty of Sciences,
Institute of Chemistry, Department of Physical Chemistry, Moyzesova 11, 041 01 Kosice, Slovakia
| | - Michal Stano
- University of P. J. Safarik
in Kosice, Faculty of Sciences,
Institute of Chemistry, Department of Physical Chemistry, Moyzesova 11, 041 01 Kosice, Slovakia
| | - Andrej Oriňak
- University of P. J. Safarik
in Kosice, Faculty of Sciences,
Institute of Chemistry, Department of Physical Chemistry, Moyzesova 11, 041 01 Kosice, Slovakia
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Arriagada C, Lin E, Schonning M, Astrof S. Mesodermal fibronectin controls cell shape, polarity, and mechanotransduction in the second heart field during cardiac outflow tract development. Dev Cell 2025; 60:62-84.e7. [PMID: 39413783 PMCID: PMC11706711 DOI: 10.1016/j.devcel.2024.09.017] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2023] [Revised: 05/06/2024] [Accepted: 09/13/2024] [Indexed: 10/18/2024]
Abstract
Failure in the elongation of the cardiac outflow tract (OFT) results in congenital heart disease due to the misalignment of the great arteries with the left and right ventricles. The OFT lengthens via the accretion of progenitors from the second heart field (SHF). SHF cells are exquisitely regionalized and organized into an epithelial-like layer, forming the dorsal pericardial wall (DPW). Tissue tension, cell polarity, and proliferation within the DPW are important for the addition of SHF-derived cells to the heart and OFT elongation. However, the genes controlling these processes are not completely characterized. Using conditional mutagenesis in the mouse, we show that fibronectin (FN1) synthesized by the mesoderm coordinates multiple cellular behaviors in the anterior DPW. FN1 is enriched in the anterior DPW and plays a role in OFT elongation by maintaining a balance between pro- and anti-adhesive cell-extracellular matrix (ECM) interactions and controlling DPW cell shape, polarity, cohesion, proliferation, and mechanotransduction.
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Affiliation(s)
- Cecilia Arriagada
- Department of Cell Biology and Molecular Medicine, Cardiovascular Research Institute, Rutgers Biomedical and Health Sciences, 185 South Orange Ave., Newark, NJ 07103, USA
| | - Evan Lin
- Princeton Day School, Princeton, NJ, USA
| | - Michael Schonning
- Department of Cell Biology and Molecular Medicine, Cardiovascular Research Institute, Rutgers Biomedical and Health Sciences, 185 South Orange Ave., Newark, NJ 07103, USA
| | - Sophie Astrof
- Department of Cell Biology and Molecular Medicine, Cardiovascular Research Institute, Rutgers Biomedical and Health Sciences, 185 South Orange Ave., Newark, NJ 07103, USA.
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5
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Hudacova E, Abaffy P, Kaplan MM, Krausova M, Kubista M, Machon O. Single-cell transcriptomic resolution of osteogenesis during craniofacial morphogenesis. Bone 2025; 190:117297. [PMID: 39461490 DOI: 10.1016/j.bone.2024.117297] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/12/2024] [Revised: 10/07/2024] [Accepted: 10/15/2024] [Indexed: 10/29/2024]
Abstract
Craniofacial morphogenesis depends on complex cell fate decisions during the differentiation of post-migratory cranial neural crest cells. Molecular mechanisms of cell differentiation of mesenchymal cells to developing bones, cartilage, teeth, tongue, and other craniofacial tissues are still poorly understood. We performed single-cell transcriptomic analysis of craniofacial mesenchymal cells derived from cranial NCCs in mouse embryo. Using FACS sorting of Wnt1-Cre2 progeny, we carefully mapped the cell heterogeneity in the craniofacial region during the initial stages of cartilage and bone formation. Transcriptomic data and in vivo validations identified molecular determinants of major cell populations involved in the development of lower and upper jaw, teeth, tongue, dermis, or periocular mesenchyme. Single-cell transcriptomic analysis of Meis2-deficient mice revealed critical gene expression differences, including increased osteogenic and cell adhesion markers. This leads to affected mesenchymal cell differentiation and increased ossification, resulting in impaired bone, cartilage, and tongue formation.
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Affiliation(s)
- Erika Hudacova
- Department of Developmental Biology, Institute of Experimental Medicine, Czech Academy of Sciences, Videnska 1083, 14200 Prague, Czech Republic; Department of Cell Biology, Faculty of Science, Charles University, Vinicna 7, 12000 Prague, Czech Republic.
| | - Pavel Abaffy
- Laboratory of Gene Expression, Institute of Biotechnology, Czech Academy of Sciences, Prumyslova 595, 25200 Vestec, Czech Republic.
| | - Mehmet Mahsum Kaplan
- Department of Developmental Biology, Institute of Experimental Medicine, Czech Academy of Sciences, Videnska 1083, 14200 Prague, Czech Republic.
| | - Michaela Krausova
- Department of Developmental Biology, Institute of Experimental Medicine, Czech Academy of Sciences, Videnska 1083, 14200 Prague, Czech Republic
| | - Mikael Kubista
- Laboratory of Gene Expression, Institute of Biotechnology, Czech Academy of Sciences, Prumyslova 595, 25200 Vestec, Czech Republic.
| | - Ondrej Machon
- Department of Developmental Biology, Institute of Experimental Medicine, Czech Academy of Sciences, Videnska 1083, 14200 Prague, Czech Republic.
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Azizi L, Otani Y, Mykuliak VV, Goult BT, Hytönen VP, Turkki P. Talin-1 variants associated with spontaneous coronary artery dissection (SCAD) highlight how even subtle changes in multi-functional scaffold proteins can manifest in disease. Hum Mol Genet 2024; 33:1846-1857. [PMID: 39163585 PMCID: PMC11540920 DOI: 10.1093/hmg/ddae120] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/07/2024] [Revised: 07/08/2024] [Accepted: 08/09/2024] [Indexed: 08/22/2024] Open
Abstract
Variants of talin-1 (TLN1) have recently been linked with spontaneous coronary artery dissection (SCAD) a condition where a tear can form in the wall of a heart artery necessitating immediate medical care. One talin-1 variant, A2013T, has an extensive familial pedigree of SCAD, which led to the screening for, and identification of, further talin-1 variants in SCAD patients. Here we evaluated these variants with commonly used pathogenicity prediction tools and found it challenging to reliably classify SCAD-associated variants, even A2013T where the evidence of a causal role is strong. Using biochemical and cell biological methods, we show that SCAD-associated variants in talin-1, which would typically be classified as non-pathogenic, still cause a measurable impact on protein structure and cell behaviour, including cell movement and wound healing capacity. Together, this indicates that even subtle variants in central mechanosensitive adapter proteins, can give rise to significant health impacts at the individual level, suggesting the need for a possible re-evaluation of the scoring criteria for pathogenicity prediction for talin variants.
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Affiliation(s)
- Latifeh Azizi
- Faculty of Medicine and Health Technology, Tampere University, Arvo Ylpön katu, 33520 Tampere, Finland
| | - Yasumi Otani
- Department of Biochemistry, Cell & Systems Biology, Institute of Systems, Molecular & Integrative Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, United States
| | - Vasyl V Mykuliak
- Faculty of Medicine and Health Technology, Tampere University, Arvo Ylpön katu, 33520 Tampere, Finland
| | - Benjamin T Goult
- Department of Biochemistry, Cell & Systems Biology, Institute of Systems, Molecular & Integrative Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, United States
| | - Vesa P Hytönen
- Faculty of Medicine and Health Technology, Tampere University, Arvo Ylpön katu, 33520 Tampere, Finland
- Fimlab Laboratories, Biokatu 4, 33520 Tampere, Finland
| | - Paula Turkki
- Faculty of Medicine and Health Technology, Tampere University, Arvo Ylpön katu, 33520 Tampere, Finland
- Fimlab Laboratories, Biokatu 4, 33520 Tampere, Finland
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7
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Wu P, Sawaki S, Yamauchi K, Yokota K, Hakamada M, Mabuchi M. Long range juxtacrine signalling through cadherin for collective cell orientation. Acta Biomater 2024:S1742-7061(24)00627-5. [PMID: 39454932 DOI: 10.1016/j.actbio.2024.10.035] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2024] [Revised: 10/21/2024] [Accepted: 10/22/2024] [Indexed: 10/28/2024]
Abstract
Many life phenomena, such as development, morphogenesis, tissue remodelling, and wound healing, are often driven by orderly and directional migration of collective cells. However, when cells are randomly oriented or localized disorder exists in orderly oriented collective cells, cell migration cannot occur in an orderly manner although various motion modes such as global rotation and local swirling and/or various motion patterns such as radial pattern and chiral pattern often occur. Therefore, it is important to control cell orientation to ensure the orderly migration of collective cells. Here, we show that it is not force transmission, but juxtacrine signalling through cadherin that plays a critical role in the orientation of collective cells. Surprisingly, juxtacrine signalling for cell orientation reached cells on a plastic dish that were not directly subjected to mechanical stimulation, up to 7 mm away from the actuator. The present study suggests that even weak mechanical stimulation is transmitted in a long range without force transmission through juxtacrine signalling. The long range juxtacrine signalling might play an important role in various life phenomena. STATEMENT OF SIGNIFICANCE: Juxtacrine signalling is direct cell-cell contact-dependent signalling, which plays a crucial role in cell behaviors such as mechanosensing, mechanotransduction and collective cell behaviors, however, there is not enough understanding about juxtacrine signalling. The present study has demonstrated that juxtacrine signalling for collective cell orientation is transmitted over a long range through cadherin. To the best of our knowledge, this is the first report of long range juxtacrine signalling. This finding may lead to the elucidation of various life phenomena such as development, morphogenesis, tissue remodelling, and wound healing.
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Affiliation(s)
- Peizheng Wu
- Graduate School of Energy Science, Kyoto University, Yoshidahonmachi, Sakyo, Kyoto 606-8501, Japan.
| | - Shogo Sawaki
- Graduate School of Energy Science, Kyoto University, Yoshidahonmachi, Sakyo, Kyoto 606-8501, Japan
| | - Kei Yamauchi
- Graduate School of Energy Science, Kyoto University, Yoshidahonmachi, Sakyo, Kyoto 606-8501, Japan
| | - Kazuki Yokota
- Graduate School of Energy Science, Kyoto University, Yoshidahonmachi, Sakyo, Kyoto 606-8501, Japan
| | - Masataka Hakamada
- Graduate School of Energy Science, Kyoto University, Yoshidahonmachi, Sakyo, Kyoto 606-8501, Japan
| | - Mamoru Mabuchi
- Graduate School of Energy Science, Kyoto University, Yoshidahonmachi, Sakyo, Kyoto 606-8501, Japan
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Barcelona-Estaje E, Oliva MAG, Cunniffe F, Rodrigo-Navarro A, Genever P, Dalby MJ, Roca-Cusachs P, Cantini M, Salmeron-Sanchez M. N-cadherin crosstalk with integrin weakens the molecular clutch in response to surface viscosity. Nat Commun 2024; 15:8824. [PMID: 39394209 PMCID: PMC11479646 DOI: 10.1038/s41467-024-53107-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/10/2023] [Accepted: 09/30/2024] [Indexed: 10/13/2024] Open
Abstract
Mesenchymal stem cells (MSCs) interact with their surroundings via integrins, which link to the actin cytoskeleton and translate physical cues into biochemical signals through mechanotransduction. N-cadherins enable cell-cell communication and are also linked to the cytoskeleton. This crosstalk between integrins and cadherins modulates MSC mechanotransduction and fate. Here we show the role of this crosstalk in the mechanosensing of viscosity using supported lipid bilayers as substrates of varying viscosity. We functionalize these lipid bilayers with adhesion peptides for integrins (RGD) and N-cadherins (HAVDI), to demonstrate that integrins and cadherins compete for the actin cytoskeleton, leading to an altered MSC mechanosensing response. This response is characterised by a weaker integrin adhesion to the environment when cadherin ligation occurs. We model this competition via a modified molecular clutch model, which drives the integrin/cadherin crosstalk in response to surface viscosity, ultimately controlling MSC lineage commitment.
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Affiliation(s)
- Eva Barcelona-Estaje
- Centre for the Cellular Microenvironment, Advanced Research Centre, University of Glasgow, Glasgow, UK
| | - Mariana A G Oliva
- Centre for the Cellular Microenvironment, Advanced Research Centre, University of Glasgow, Glasgow, UK
| | - Finlay Cunniffe
- Centre for the Cellular Microenvironment, Advanced Research Centre, University of Glasgow, Glasgow, UK
| | | | - Paul Genever
- Department of Biology, University of York, York, UK
| | - Matthew J Dalby
- Centre for the Cellular Microenvironment, Advanced Research Centre, University of Glasgow, Glasgow, UK
| | - Pere Roca-Cusachs
- Institute for Bioengineering of Catalonia (IBEC), the Barcelona Institute of Technology (BIST), Barcelona, Spain.
- University of Barcelona, Barcelona, Spain.
| | - Marco Cantini
- Centre for the Cellular Microenvironment, Advanced Research Centre, University of Glasgow, Glasgow, UK.
| | - Manuel Salmeron-Sanchez
- Centre for the Cellular Microenvironment, Advanced Research Centre, University of Glasgow, Glasgow, UK.
- Institute for Bioengineering of Catalonia (IBEC), the Barcelona Institute of Technology (BIST), Barcelona, Spain.
- Institució Catalana de Recerca i Estudis Avançats (ICREA), Barcelona, Spain.
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Altmaier S, Le Harzic R, Stracke F, Speicher AM, Uhl D, Ehrlich J, Gerlach T, Schmidt K, Lemmer K, Lautenschläger F, Böse H, Neubauer JC, Zimmermann H, Meiser I. Cytoskeleton adaptation to stretchable surface relaxation improves adherent cryopreservation of human mesenchymal stem cells. Cryobiology 2024; 117:104958. [PMID: 39243925 DOI: 10.1016/j.cryobiol.2024.104958] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/04/2024] [Revised: 07/29/2024] [Accepted: 08/22/2024] [Indexed: 09/09/2024]
Abstract
Adherent cell systems are usually dissociated before being cryopreserved, as standard protocols are established for cells in suspension. The application of standard procedures to more complex systems, sensitive to dissociation, such as adherent monolayers, especially comprising mature cell types or tissues remains unsatisfactory. Uncontrolled cell detachment due to intracellular tensile stress, membrane ruptures and damages of adhesion proteins are common during freezing and thawing of cell monolayers. However, many therapeutically relevant cell systems grow adherently to develop their native morphology and functionality, but lose their integrity after dissociation. The hypothesis is that cells on stretchable substrates have a more adaptable cytoskeleton and membrane, reducing cryopreservation-induced stress. Our studies investigate the influence of stretchable surfaces on the cryopreservation of adherent cells to avoid harmful dissociation and expedite post-thawing cultivation of functional cells. A stretching apparatus for defined radial stretching, consisting of silicone vessels and films with specific surface textures for cell culture, was developed. Adherent human umbilical cord mesenchymal stem cells (hUC-MSCs) were cultivated on a stretched silicone film within the vessel, forming a monolayer that was compressed by relaxation, while remaining attached to the relaxed film. Compressed hUC-MSCs, which were cryopreserved adherently showed higher viability and less detachment after thawing compared to control cells without compression. Within three to seven days post-thawing, the hUC-MSCs recovered, and the monolayer reformed. These experiments support the hypothesis that cryopreservation success of adherent cell systems is enhanced by improved adaptability of the cytoskeleton and cell membrane, opening up new approaches in cryobiotechnology.
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Affiliation(s)
- Saskia Altmaier
- Department of Molecular and Cellular Biotechnology, Saarland University, 66123, Saarbrücken, Germany; Department of Cryosensor Technology, Fraunhofer Institute for Biomedical Engineering (IBMT), Joseph-von-Fraunhofer-Weg 1, 66820, Sulzbach, Germany
| | - Ronan Le Harzic
- Department of Cryosensor Technology, Fraunhofer Institute for Biomedical Engineering (IBMT), Joseph-von-Fraunhofer-Weg 1, 66820, Sulzbach, Germany
| | - Frank Stracke
- Department of Cryosensor Technology, Fraunhofer Institute for Biomedical Engineering (IBMT), Joseph-von-Fraunhofer-Weg 1, 66820, Sulzbach, Germany
| | - Anna Martina Speicher
- Department of Cryosensor Technology, Fraunhofer Institute for Biomedical Engineering (IBMT), Joseph-von-Fraunhofer-Weg 1, 66820, Sulzbach, Germany
| | - Detlev Uhl
- Center Smart Materials and Adaptive Systems (CeSMA), Fraunhofer Institute for Silicate Research ISC, Neunerplatz 2, 97082, Würzburg, Germany
| | - Johannes Ehrlich
- Center Smart Materials and Adaptive Systems (CeSMA), Fraunhofer Institute for Silicate Research ISC, Neunerplatz 2, 97082, Würzburg, Germany
| | - Thomas Gerlach
- Center Smart Materials and Adaptive Systems (CeSMA), Fraunhofer Institute for Silicate Research ISC, Neunerplatz 2, 97082, Würzburg, Germany
| | - Katharina Schmidt
- Department of Cryosensor Technology, Fraunhofer Institute for Biomedical Engineering (IBMT), Joseph-von-Fraunhofer-Weg 1, 66820, Sulzbach, Germany
| | - Katja Lemmer
- Department of Cryosensor Technology, Fraunhofer Institute for Biomedical Engineering (IBMT), Joseph-von-Fraunhofer-Weg 1, 66820, Sulzbach, Germany
| | | | - Holger Böse
- Center Smart Materials and Adaptive Systems (CeSMA), Fraunhofer Institute for Silicate Research ISC, Neunerplatz 2, 97082, Würzburg, Germany
| | - Julia C Neubauer
- Department of Cryosensor Technology, Fraunhofer Institute for Biomedical Engineering (IBMT), Joseph-von-Fraunhofer-Weg 1, 66820, Sulzbach, Germany
| | - Heiko Zimmermann
- Department of Molecular and Cellular Biotechnology, Saarland University, 66123, Saarbrücken, Germany; Department of Cryosensor Technology, Fraunhofer Institute for Biomedical Engineering (IBMT), Joseph-von-Fraunhofer-Weg 1, 66820, Sulzbach, Germany; Facultad de Ciencias del Mar, Universidad Católica del Norte, 1780000, Coquimbo, Chile
| | - Ina Meiser
- Department of Cryosensor Technology, Fraunhofer Institute for Biomedical Engineering (IBMT), Joseph-von-Fraunhofer-Weg 1, 66820, Sulzbach, Germany.
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10
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Habli Z, Zantout A, Al-Haj N, Saab R, El-Sabban M, Khraiche ML. Single-Cell Fluidic Force Spectroscopy Reveals Dynamic Mechanical Fingerprints of Malignancy in Breast Cancer. ACS APPLIED MATERIALS & INTERFACES 2024; 16:50147-50159. [PMID: 39105773 PMCID: PMC11440459 DOI: 10.1021/acsami.4c06335] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 08/07/2024]
Abstract
The interplay between cancer cell physical characteristics and metastatic potential highlights the significance of cancer cell mechanobiology. Using fluidic-based single-cell force spectroscopy (SCFS), quartz crystal microbalance with dissipation (QCM-D), and a model of cells with a spectrum of metastatic potential, we track the progression of biomechanics across the metastatic states by measuring cell-substrate and cell-to-cell adhesion forces, cell spring constant, cell height, and cell viscoelasticity. Compared to highly metastatic cells, cells in the lower spectrum of metastatic ability are found to be systematically stiffer, less viscoelastic, and larger. These mechanical transformations in cells within a cluster correlate with cells' metastatic potential but are significantly absent in single cells. Additionally, the response to chemotherapy is found to be highly dependent on cell viscoelastic properties in terms of both response time and magnitude. Shifts in cell softness and elasticity might serve as mechanoadaptive mechanisms during cancer cell metastasis, contributing to our understanding of metastasis and the effectiveness of potential therapeutic interventions.
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Affiliation(s)
- Zeina Habli
- Neural Engineering and Nanobiosensors Group, Biomedical Engineering Program, Maroun Semaan Faculty of Engineering and Architecture, American University of Beirut, Beirut 1107 2020, Lebanon
| | - Ahmad Zantout
- Neural Engineering and Nanobiosensors Group, Biomedical Engineering Program, Maroun Semaan Faculty of Engineering and Architecture, American University of Beirut, Beirut 1107 2020, Lebanon
| | - Nadine Al-Haj
- Department of Anatomy, Cell Biology, and Physiological Sciences, Faculty of Medicine, American University of Beirut, Beirut 1107 2020, Lebanon
| | - Raya Saab
- Department of Pediatrics, Stanford University School of Medicine, Palo Alto, California 94304, United States
| | - Marwan El-Sabban
- Department of Anatomy, Cell Biology, and Physiological Sciences, Faculty of Medicine, American University of Beirut, Beirut 1107 2020, Lebanon
| | - Massoud L Khraiche
- Neural Engineering and Nanobiosensors Group, Biomedical Engineering Program, Maroun Semaan Faculty of Engineering and Architecture, American University of Beirut, Beirut 1107 2020, Lebanon
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11
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Essebier P, Keyser M, Yordanov T, Hill B, Yu A, Noordstra I, Yap AS, Stehbens SJ, Lagendijk AK, Schimmel L, Gordon EJ. c-Src-induced vascular malformations require localised matrix degradation at focal adhesions. J Cell Sci 2024; 137:jcs262101. [PMID: 38881365 PMCID: PMC11267457 DOI: 10.1242/jcs.262101] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/07/2024] [Accepted: 06/06/2024] [Indexed: 06/18/2024] Open
Abstract
Endothelial cells lining the blood vessel wall communicate intricately with the surrounding extracellular matrix, translating mechanical cues into biochemical signals. Moreover, vessels require the capability to enzymatically degrade the matrix surrounding them, to facilitate vascular expansion. c-Src plays a key role in blood vessel growth, with its loss in the endothelium reducing vessel sprouting and focal adhesion signalling. Here, we show that constitutive activation of c-Src in endothelial cells results in rapid vascular expansion, operating independently of growth factor stimulation or fluid shear stress forces. This is driven by an increase in focal adhesion signalling and size, with enhancement of localised secretion of matrix metalloproteinases responsible for extracellular matrix remodelling. Inhibition of matrix metalloproteinase activity results in a robust rescue of the vascular expansion elicited by heightened c-Src activity. This supports the premise that moderating focal adhesion-related events and matrix degradation can counteract abnormal vascular expansion, with implications for pathologies driven by unusual vascular morphologies.
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Affiliation(s)
- Patricia Essebier
- Centre for Cell Biology of Chronic Disease, Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
| | - Mikaela Keyser
- Centre for Cell Biology of Chronic Disease, Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
| | - Teodor Yordanov
- Centre for Cell Biology of Chronic Disease, Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
| | - Brittany Hill
- Centre for Cell Biology of Chronic Disease, Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
| | - Alexander Yu
- Centre for Cell Biology of Chronic Disease, Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
| | - Ivar Noordstra
- Centre for Cell Biology of Chronic Disease, Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
| | - Alpha S. Yap
- Centre for Cell Biology of Chronic Disease, Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
| | - Samantha J. Stehbens
- Centre for Cell Biology of Chronic Disease, Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
- Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
| | - Anne K. Lagendijk
- Centre for Cell Biology of Chronic Disease, Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
| | - Lilian Schimmel
- Centre for Cell Biology of Chronic Disease, Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
| | - Emma J. Gordon
- Centre for Cell Biology of Chronic Disease, Institute for Molecular Bioscience, The University of Queensland, St. Lucia, Brisbane, Queensland, Australia4072
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12
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Wang C, Fan M, Heo SJ, Adams SM, Li T, Liu Y, Li Q, Loebel C, Alisafaei F, Burdick JA, Lu XL, Birk DE, Mauck RL, Han L. Structure-Mechanics Principles and Mechanobiology of Fibrocartilage Pericellular Matrix: A Pivotal Role of Type V Collagen. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.06.26.600498. [PMID: 38979323 PMCID: PMC11230444 DOI: 10.1101/2024.06.26.600498] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 07/10/2024]
Abstract
The pericellular matrix (PCM) is the immediate microniche surrounding resident cells in various tissue types, regulating matrix turnover, cell-matrix cross-talk and disease initiation. This study elucidated the structure-mechanical properties and mechanobiological functions of the PCM in fibrocartilage, a family of connective tissues that sustain complex tensile and compressive loads in vivo. Studying the murine meniscus as the model tissue, we showed that fibrocartilage PCM contains thinner, random collagen fibrillar networks that entrap proteoglycans, a structure distinct from the densely packed, highly aligned collagen fibers in the bulk extracellular matrix (ECM). In comparison to the ECM, the PCM has a lower modulus and greater isotropy, but similar relative viscoelastic properties. In Col5a1 +/- menisci, the reduction of collagen V, a minor collagen localized in the PCM, resulted in aberrant fibril thickening with increased heterogeneity. Consequently, the PCM exhibited a reduced modulus, loss of isotropy and faster viscoelastic relaxation. This disrupted PCM contributes to perturbed mechanotransduction of resident meniscal cells, as illustrated by reduced intracellular calcium signaling, as well as upregulated biosynthesis of lysyl oxidase and tenascin C. When cultured in vitro, Col5a1 +/- meniscal cells synthesized a weakened nascent PCM, which had inferior properties towards protecting resident cells against applied tensile stretch. These findings underscore the PCM as a distinctive microstructure that governs fibrocartilage mechanobiology, and highlight the pivotal role of collagen V in PCM function. Targeting the PCM or its molecular constituents holds promise for enhancing not only meniscus regeneration and osteoarthritis intervention, but also addressing diseases across various fibrocartilaginous tissues.
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Affiliation(s)
- Chao Wang
- School of Biomedical Engineering, Science and Health Systems, Drexel University, Philadelphia, PA 19104, United States
| | - Mingyue Fan
- School of Biomedical Engineering, Science and Health Systems, Drexel University, Philadelphia, PA 19104, United States
| | - Su-Jin Heo
- McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, United States
| | - Sheila M Adams
- Department of Molecular Pharmacology and Physiology, Morsani School of Medicine, University of South Florida, Tampa, FL 33612, United States
| | - Thomas Li
- School of Biomedical Engineering, Science and Health Systems, Drexel University, Philadelphia, PA 19104, United States
| | - Yuchen Liu
- School of Biomedical Engineering, Science and Health Systems, Drexel University, Philadelphia, PA 19104, United States
| | - Qing Li
- School of Biomedical Engineering, Science and Health Systems, Drexel University, Philadelphia, PA 19104, United States
| | - Claudia Loebel
- Department of Materials Science and Engineering, University of Michigan, Ann Arbor, MI 48109, United States
| | - Farid Alisafaei
- Department of Mechanical and Industrial Engineering, New Jersey Institute of Technology, Newark, NJ 07102, United States
| | - Jason A Burdick
- BioFrontiers Institute and Department of Chemical and Biological Engineering, University of Colorado, Boulder, CO 80309, United States
| | - X Lucas Lu
- Department of Mechanical Engineering, University of Delaware, Newark, DE 19716, United States
| | - David E Birk
- Department of Molecular Pharmacology and Physiology, Morsani School of Medicine, University of South Florida, Tampa, FL 33612, United States
| | - Robert L Mauck
- McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, United States
- Translational Musculoskeletal Research Center, Corporal Michael J. Crescenz Veterans Administration Medical Center, Philadelphia, PA 19104, United States
| | - Lin Han
- School of Biomedical Engineering, Science and Health Systems, Drexel University, Philadelphia, PA 19104, United States
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13
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Ferrai C, Schulte C. Mechanotransduction in stem cells. Eur J Cell Biol 2024; 103:151417. [PMID: 38729084 DOI: 10.1016/j.ejcb.2024.151417] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/27/2023] [Revised: 04/28/2024] [Accepted: 04/29/2024] [Indexed: 05/12/2024] Open
Abstract
Nowadays, it is an established concept that the capability to reach a specialised cell identity via differentiation, as in the case of multi- and pluripotent stem cells, is not only determined by biochemical factors, but that also physical aspects of the microenvironment play a key role; interpreted by the cell through a force-based signalling pathway called mechanotransduction. However, the intricate ties between the elements involved in mechanotransduction, such as the extracellular matrix, the glycocalyx, the cell membrane, Integrin adhesion complexes, Cadherin-mediated cell/cell adhesion, the cytoskeleton, and the nucleus, are still far from being understood in detail. Here we report what is currently known about these elements in general and their specific interplay in the context of multi- and pluripotent stem cells. We furthermore merge this overview to a more comprehensive picture, that aims to cover the whole mechanotransductive pathway from the cell/microenvironment interface to the regulation of the chromatin structure in the nucleus. Ultimately, with this review we outline the current picture of the interplay between mechanotransductive cues and epigenetic regulation and how these processes might contribute to stem cell dynamics and fate.
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Affiliation(s)
- Carmelo Ferrai
- Institute of Pathology, University Medical Centre Göttingen, Germany.
| | - Carsten Schulte
- Department of Biomedical and Clinical Sciences and Department of Physics "Aldo Pontremoli", University of Milan, Italy.
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14
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Zhang J, Kong X, Yang HJ, Zhang W, Chen M, Chen X. Ninjurin 2 Modulates Tumorigenesis, Inflammation, and Metabolism via Pyroptosis. THE AMERICAN JOURNAL OF PATHOLOGY 2024; 194:849-860. [PMID: 38325550 DOI: 10.1016/j.ajpath.2024.01.013] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/17/2023] [Revised: 01/08/2024] [Accepted: 01/19/2024] [Indexed: 02/09/2024]
Abstract
The nerve injury-induced protein 2 (NINJ2) belongs to a family of homophilic adhesion molecules and was initially found to be involved in nerve regeneration. However, the role of NINJ2 in other cellular processes is not well studied. The Ninj2-deficient mice generated in the current study had a short lifespan and were prone to spontaneous tumors, systemic inflammation, and metabolic defects. Comprehensive carbohydrate and lipid metabolic analyses were performed to better understand the metabolic traits that contribute to these phenotypes. Carbohydrate metabolic analyses showed that NINJ2 deficiency led to defects in monosaccharide metabolism along with accumulation of multiple disaccharides and sugar alcohols. Lipidomic analyses showed that Ninj2 deficiency altered patterns of several lipids, including triglycerides, phospholipids, and ceramides. To identify a cellular process that associated with these metabolic defects, the role of NINJ2 in pyroptosis, a programmed cell death that links cancer, inflammation, and metabolic disorders, was examined. Loss of NINJ2 promoted pyroptosis by activating the NOD-like receptor protein 3 (NLRP3) inflammasome. Taken together, these data reveal a critical role of NINJ2 in tumorigenesis, inflammatory response, and metabolism via pyroptosis.
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Affiliation(s)
- Jin Zhang
- Comparative Oncology Laboratory, University of California, Davis, Davis, California.
| | - Xiangmudong Kong
- Comparative Oncology Laboratory, University of California, Davis, Davis, California
| | - Hee Jung Yang
- Comparative Oncology Laboratory, University of California, Davis, Davis, California
| | - Weici Zhang
- Division of Rheumatology, Allergy and Clinical Immunology, University of California, Davis, Davis, California
| | - Mingyi Chen
- Department of Pathology, University of Texas Southwestern Medical Center, Dallas, Texas
| | - Xinbin Chen
- Comparative Oncology Laboratory, University of California, Davis, Davis, California.
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15
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Akinsuyi OS, Xhumari J, Ojeda A, Roesch LFW. Gut permeability among Astronauts during Space missions. LIFE SCIENCES IN SPACE RESEARCH 2024; 41:171-180. [PMID: 38670644 DOI: 10.1016/j.lssr.2024.03.003] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/11/2023] [Revised: 02/02/2024] [Accepted: 03/13/2024] [Indexed: 04/28/2024]
Abstract
The space environment poses substantial challenges to human physiology, including potential disruptions in gastrointestinal health. Gut permeability has only recently become widely acknowledged for its potential to cause adverse effects on a systemic level, rendering it a critical factor to investigate in the context of spaceflight. Here, we propose that astronauts experience the onset of leaky gut during space missions supported by transcriptomic and metagenomic analysis of human and murine samples. A genetic map contributing to intestinal permeability was constructed from a systematic review of current literature. This was referenced against our re-analysis of three independent transcriptomic datasets which revealed significant changes in gene expression patterns associated with the gut barrier. Specifically, in astronauts during flight, we observed a substantial reduction in the expression genes that are crucial for intestinal barrier function, goblet cell development, gut microbiota modulation, and immune responses. Among rodent spaceflight studies, differential expression of cytokines, chemokines, and genes which regulate mucin production and post-translational modifications suggest a similar dysfunction of intestinal permeability. Metagenomic analysis of feces from two murine studies revealed a notable reduction probiotic, short chain fatty acid-producing bacteria and an increase in the Gram-negative pathogens, including Citrobacter rodentium, Enterobacter cloacea, Klebsiella aerogenes, and Proteus hauseri which promote LPS circulation, a recipe for barrier disruption and systemic inflammatory activation. These findings emphasize the critical need to understand the underlying mechanisms and develop interventions to maintain gastrointestinal health in space.
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Affiliation(s)
- Oluwamayowa S Akinsuyi
- Institute of Food and Agricultural Sciences, Department of Microbiology and Cell Science, University of Florida, Gainesville 32611, FL, USA
| | - Jessica Xhumari
- Institute of Food and Agricultural Sciences, Department of Microbiology and Cell Science, University of Florida, Gainesville 32611, FL, USA
| | - Amanda Ojeda
- Institute of Food and Agricultural Sciences, Department of Microbiology and Cell Science, University of Florida, Gainesville 32611, FL, USA
| | - Luiz F W Roesch
- Institute of Food and Agricultural Sciences, Department of Microbiology and Cell Science, University of Florida, Gainesville 32611, FL, USA.
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16
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Ebrahim T, Ebrahim AS, Kandouz M. Diversity of Intercellular Communication Modes: A Cancer Biology Perspective. Cells 2024; 13:495. [PMID: 38534339 PMCID: PMC10969453 DOI: 10.3390/cells13060495] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/05/2024] [Revised: 02/27/2024] [Accepted: 03/10/2024] [Indexed: 03/28/2024] Open
Abstract
From the moment a cell is on the path to malignant transformation, its interaction with other cells from the microenvironment becomes altered. The flow of molecular information is at the heart of the cellular and systemic fate in tumors, and various processes participate in conveying key molecular information from or to certain cancer cells. For instance, the loss of tight junction molecules is part of the signal sent to cancer cells so that they are no longer bound to the primary tumors and are thus free to travel and metastasize. Upon the targeting of a single cell by a therapeutic drug, gap junctions are able to communicate death information to by-standing cells. The discovery of the importance of novel modes of cell-cell communication such as different types of extracellular vesicles or tunneling nanotubes is changing the way scientists look at these processes. However, are they all actively involved in different contexts at the same time or are they recruited to fulfill specific tasks? What does the multiplicity of modes mean for the overall progression of the disease? Here, we extend an open invitation to think about the overall significance of these questions, rather than engage in an elusive attempt at a systematic repertory of the mechanisms at play.
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Affiliation(s)
- Thanzeela Ebrahim
- Department of Pathology, Wayne State University School of Medicine, Detroit, MI 48202, USA
| | - Abdul Shukkur Ebrahim
- Department of Ophthalmology, Visual and Anatomical Sciences, Wayne State University School of Medicine, Detroit, MI 48202, USA
| | - Mustapha Kandouz
- Department of Pathology, Wayne State University School of Medicine, Detroit, MI 48202, USA
- Karmanos Cancer Institute, Wayne State University School of Medicine, Detroit, MI 48202, USA
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17
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Nguyen J, Gilbert PM. Decoding the forces that shape muscle stem cell function. Curr Top Dev Biol 2024; 158:279-306. [PMID: 38670710 DOI: 10.1016/bs.ctdb.2024.02.009] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 04/28/2024]
Abstract
Skeletal muscle is a force-producing organ composed of muscle tissues, connective tissues, blood vessels, and nerves, all working in synergy to enable movement and provide support to the body. While robust biomechanical descriptions of skeletal muscle force production at the body or tissue level exist, little is known about force application on microstructures within the muscles, such as cells. Among various cell types, skeletal muscle stem cells reside in the muscle tissue environment and play a crucial role in driving the self-repair process when muscle damage occurs. Early evidence indicates that the fate and function of skeletal muscle stem cells are controlled by both biophysical and biochemical factors in their microenvironments, but much remains to accomplish in quantitatively describing the biophysical muscle stem cell microenvironment. This book chapter aims to review current knowledge on the influence of biophysical stresses and landscape properties on muscle stem cells in heath, aging, and diseases.
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Affiliation(s)
- Jo Nguyen
- Institute of Biomedical Engineering, University of Toronto, Toronto, ON, Canada; Donnelly Centre, University of Toronto, Toronto, ON, Canada
| | - Penney M Gilbert
- Institute of Biomedical Engineering, University of Toronto, Toronto, ON, Canada; Donnelly Centre, University of Toronto, Toronto, ON, Canada; Department of Cell and Systems Biology, University of Toronto, Toronto, ON, Canada.
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18
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Hurst R, Brewer DS, Gihawi A, Wain J, Cooper CS. Cancer invasion and anaerobic bacteria: new insights into mechanisms. J Med Microbiol 2024; 73:001817. [PMID: 38535967 PMCID: PMC10995961 DOI: 10.1099/jmm.0.001817] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2023] [Accepted: 02/27/2024] [Indexed: 04/07/2024] Open
Abstract
There is growing evidence that altered microbiota abundance of a range of specific anaerobic bacteria are associated with cancer, including Peptoniphilus spp., Porphyromonas spp., Fusobacterium spp., Fenollaria spp., Prevotella spp., Sneathia spp., Veillonella spp. and Anaerococcus spp. linked to multiple cancer types. In this review we explore these pathogenic associations. The mechanisms by which bacteria are known or predicted to interact with human cells are reviewed and we present an overview of the interlinked mechanisms and hypotheses of how multiple intracellular anaerobic bacterial pathogens may act together to cause host cell and tissue microenvironment changes associated with carcinogenesis and cancer cell invasion. These include combined effects on changes in cell signalling, DNA damage, cellular metabolism and immune evasion. Strategies for early detection and eradication of anaerobic cancer-associated bacterial pathogens that may prevent cancer progression are proposed.
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Affiliation(s)
- Rachel Hurst
- Norwich Medical School, University of East Anglia, Norwich, Norfolk, NR4 7TJ, UK
| | - Daniel S. Brewer
- Norwich Medical School, University of East Anglia, Norwich, Norfolk, NR4 7TJ, UK
- Earlham Institute, Norwich Research Park Innovation Centre, Colney Lane, Norwich NR4 7UZ, UK
| | - Abraham Gihawi
- Norwich Medical School, University of East Anglia, Norwich, Norfolk, NR4 7TJ, UK
| | - John Wain
- Norwich Medical School, University of East Anglia, Norwich, Norfolk, NR4 7TJ, UK
- Quadram Institute Biosciences, Colney Lane, Norwich, Norfolk, NR4 7UQ, UK
| | - Colin S. Cooper
- Norwich Medical School, University of East Anglia, Norwich, Norfolk, NR4 7TJ, UK
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19
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Kim MH, Thanuthanakhun N, Kino-oka M. Stable and efficient generation of functional iPSC-derived neural progenitor cell rosettes through regulation of collective cell-cell behavior. Front Bioeng Biotechnol 2024; 11:1269108. [PMID: 38268936 PMCID: PMC10806250 DOI: 10.3389/fbioe.2023.1269108] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/31/2023] [Accepted: 12/21/2023] [Indexed: 01/26/2024] Open
Abstract
Although the potential of stem cells to differentiate into several cell types has shown promise in regenerative medicine, low differentiation efficiency and poor reproducibility significantly limit their practical application. We developed an effective and robust differentiation strategy for the efficient and robust generation of neural progenitor cell rosettes from induced pluripotent stem cells (iPSCs) incorporating botulinum hemagglutinin (HA). Treatment with HA suppressed the spontaneous differentiation of iPSCs cultured under undirected differentiation conditions, resulting in the preservation of their pluripotency. Moreover, treatment with HA during neural progenitor differentiation combined with dual SMAD inhibition generated a highly homogeneous population of PAX6-and SOX1-expressing neural progenitor cells with 8.4-fold higher yields of neural progenitor cells than untreated control cultures. These neural progenitor cells formed radially organized rosettes surrounding the central lumen. This differentiation method enhanced the generation of functional iPSC-derived neural progenitor cell rosettes throughout the culture vessel, suggesting that the regulation of collective cell-cell behavior using HA plays a morphogenetically important role in rosette formation and maturation. These findings show the significance of HA in the suppression of spontaneous differentiation through spatial homogeneity. The study proposes a novel methodology for the efficient derivation of functional iPSC-derived neural progenitor cell rosettes.
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Affiliation(s)
- Mee-Hae Kim
- Department of Biotechnology, Graduate School of Engineering, Osaka University, Osaka, Japan
| | | | - Masahiro Kino-oka
- Department of Biotechnology, Graduate School of Engineering, Osaka University, Osaka, Japan
- Research Base for Cell Manufacturability, Osaka University, Osaka, Japan
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20
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Shoyer TC, Gates EM, Cabe JI, Urs AN, Conway DE, Hoffman BD. Coupling during collective cell migration is controlled by a vinculin mechanochemical switch. Proc Natl Acad Sci U S A 2023; 120:e2316456120. [PMID: 38055737 PMCID: PMC10722971 DOI: 10.1073/pnas.2316456120] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/26/2023] [Accepted: 10/24/2023] [Indexed: 12/08/2023] Open
Abstract
The ability of cells to move in a mechanically coupled, coordinated manner, referred to as collective cell migration, is central to many developmental, physiological, and pathophysiological processes. Limited understanding of how mechanical forces and biochemical regulation interact to affect coupling has been a major obstacle to unravelling the underlying mechanisms. Focusing on the linker protein vinculin, we use a suite of Förster resonance energy transfer-based biosensors to probe its mechanical functions and biochemical regulation, revealing a switch that toggles vinculin between loadable and unloadable states. Perturbation of the switch causes covarying changes in cell speed and coordination, suggesting alteration of the friction within the system. Molecular scale modelling reveals that increasing levels of loadable vinculin increases friction, due to engagement of self-stabilizing catch bonds. Together, this work reveals a regulatory switch for controlling cell coupling and describes a paradigm for relating biochemical regulation, altered mechanical properties, and changes in cell behaviors.
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Affiliation(s)
- T. Curtis Shoyer
- Department of Biomedical Engineering, Duke University, Durham, NC27708
| | - Evan M. Gates
- Department of Biomedical Engineering, Duke University, Durham, NC27708
| | - Jolene I. Cabe
- Department of Biomedical Engineering, Virginia Commonwealth University, Richmond, VA23284
| | - Aarti N. Urs
- Department of Cell Biology, Duke University, Durham, NC27710
| | - Daniel E. Conway
- Department of Biomedical Engineering, The Ohio State University, Columbus, OH43210
| | - Brenton D. Hoffman
- Department of Biomedical Engineering, Duke University, Durham, NC27708
- Department of Cell Biology, Duke University, Durham, NC27710
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21
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Kenny FN, Marcotti S, De Freitas DB, Drudi EM, Leech V, Bell RE, Easton J, Díaz-de-la-Loza MDC, Fleck R, Allison L, Philippeos C, Manhart A, Shaw TJ, Stramer BM. Autocrine IL-6 drives cell and extracellular matrix anisotropy in scar fibroblasts. Matrix Biol 2023; 123:1-16. [PMID: 37660739 PMCID: PMC10878985 DOI: 10.1016/j.matbio.2023.08.004] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/15/2023] [Revised: 07/31/2023] [Accepted: 08/26/2023] [Indexed: 09/05/2023]
Abstract
Fibrosis is associated with dramatic changes in extracellular matrix (ECM) architecture of unknown etiology. Here we exploit keloid scars as a paradigm to understand fibrotic ECM organization. We reveal that keloid patient fibroblasts uniquely produce a globally aligned ECM network in 2-D culture as observed in scar tissue. ECM anisotropy develops after rapid initiation of a fibroblast supracellular actin network, suggesting that cell alignment initiates ECM patterning. Keloid fibroblasts produce elevated levels of IL-6, and autocrine IL-6 production is both necessary and sufficient to induce cell and ECM alignment, as evidenced by ligand stimulation of normal dermal fibroblasts and treatment of keloid fibroblasts with the function blocking IL-6 receptor monoclonal antibody, tocilizumab. Downstream of IL-6, supracellular organization of keloid fibroblasts is controlled by activation of cell-cell adhesion. Adhesion formation inhibits contact-induced cellular overlap leading to nematic organization of cells and an alignment of focal adhesions. Keloid fibroblasts placed on isotropic ECM align the pre-existing matrix, suggesting that focal adhesion alignment leads to active anisotropic remodeling. These results show that IL-6-induced fibroblast cooperativity can control the development of a nematic ECM, highlighting both IL-6 signaling and cell-cell adhesions as potential therapeutic targets to inhibit this common feature of fibrosis.
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Affiliation(s)
- Fiona N Kenny
- Randall Centre for Cell and Molecular Biophysics, King's College London, London, UK
| | - Stefania Marcotti
- Randall Centre for Cell and Molecular Biophysics, King's College London, London, UK
| | | | - Elena M Drudi
- Centre for Inflammation Biology & Cancer Immunology, Department of Inflammation Biology, School of Immunology & Microbial Sciences, King's College London, London, UK
| | - Vivienne Leech
- Department of Mathematics, University College London, UK
| | - Rachel E Bell
- Centre for Inflammation Biology & Cancer Immunology, Department of Inflammation Biology, School of Immunology & Microbial Sciences, King's College London, London, UK
| | - Jennifer Easton
- Centre for Inflammation Biology & Cancer Immunology, Department of Inflammation Biology, School of Immunology & Microbial Sciences, King's College London, London, UK
| | | | - Roland Fleck
- Centre for Ultrastructure Imaging, King's College London, UK
| | - Leanne Allison
- Centre for Ultrastructure Imaging, King's College London, UK
| | - Christina Philippeos
- Centre for Stem Cells and Regenerative Medicine, King's College London, London, UK
| | - Angelika Manhart
- Department of Mathematics, University College London, UK; Faculty of Mathematics, University of Vienna, Vienna, Austria
| | - Tanya J Shaw
- Centre for Inflammation Biology & Cancer Immunology, Department of Inflammation Biology, School of Immunology & Microbial Sciences, King's College London, London, UK.
| | - Brian M Stramer
- Randall Centre for Cell and Molecular Biophysics, King's College London, London, UK.
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22
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Son YJ, Keum C, Kim M, Jeong G, Jin S, Hwang HW, Kim H, Lee K, Jeon H, Kim H, Pahk KJ, Jang HW, Sun JY, Han HS, Lee KH, Ok MR, Kim YC, Jeong Y. Selective Cell-Cell Adhesion Regulation via Cyclic Mechanical Deformation Induced by Ultrafast Nanovibrations. ACS APPLIED MATERIALS & INTERFACES 2023. [PMID: 37751467 DOI: 10.1021/acsami.3c08941] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 09/28/2023]
Abstract
The adoption of dynamic mechanomodulation to regulate cellular behavior is an alternative to the use of chemical drugs, allowing spatiotemporal control. However, cell-selective targeting of mechanical stimuli is challenging due to the lack of strategies with which to convert macroscopic mechanical movements to different cellular responses. Here, we designed a nanoscale vibrating surface that controls cell behavior via selective repetitive cell deformation based on a poroelastic cell model. The vibrating indentations induce repetitive water redistribution in the cells with water redistribution rates faster than the vibrating rate; however, in the opposite case, cells perceive the vibrations as a one-time stimulus. The selective regulation of cell-cell adhesion through adjusting the frequency of nanovibration was demonstrated by suppression of cadherin expression in smooth muscle cells (fast water redistribution rate) with no change in vascular endothelial cells (slow water redistribution rate). This technique may provide a new strategy for cell-type-specific mechanical stimulation.
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Affiliation(s)
- Young Ju Son
- Center for Biomaterials, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
| | - Changjoon Keum
- Center for Advanced Biomolecular Recognition, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
| | - Minsoo Kim
- Center for Advanced Biomolecular Recognition, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
- Department of Chemistry, Hanyang University, Seoul 04763, Republic of Korea
| | - Goeen Jeong
- Center for Biomaterials, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
- Department of Materials Science and Engineering, Seoul National University, Seoul 08826, Republic of Korea
| | - Soyeong Jin
- Center for Advanced Biomolecular Recognition, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
- Department of Chemistry, Hanyang University, Seoul 04763, Republic of Korea
| | - Hae Won Hwang
- Center for Biomaterials, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
- Department of Materials Science and Engineering, Seoul National University, Seoul 08826, Republic of Korea
| | - Hyewon Kim
- Center for Biomaterials, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
| | - Kyungwoo Lee
- Center for Biomaterials, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
| | - Hojeong Jeon
- Center for Biomaterials, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
- KU-KIST Graduate School of Converging Science and Technology, Korea University, Seoul 02841, Republic of Korea
| | - Hojun Kim
- Center for Advanced Biomolecular Recognition, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
- Division of Bio-Medical Science and Technology, KIST School, Korea University of Science and Technology, Seoul 02792, Republic of Korea
| | - Ki Joo Pahk
- Department of Biomedical Engineering, Kyung Hee University, Yongin 17104, Republic of Korea
| | - Ho Won Jang
- Department of Materials Science and Engineering, Seoul National University, Seoul 08826, Republic of Korea
| | - Jeong-Yun Sun
- Department of Materials Science and Engineering, Seoul National University, Seoul 08826, Republic of Korea
| | - Hyung-Seop Han
- Center for Biomaterials, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
- Division of Bio-Medical Science and Technology, KIST School, Korea University of Science and Technology, Seoul 02792, Republic of Korea
| | - Kwan Hyi Lee
- Center for Advanced Biomolecular Recognition, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
- KU-KIST Graduate School of Converging Science and Technology, Korea University, Seoul 02841, Republic of Korea
- Division of Bio-Medical Science and Technology, KIST School, Korea University of Science and Technology, Seoul 02792, Republic of Korea
| | - Myoung-Ryul Ok
- Center for Biomaterials, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
- Division of Bio-Medical Science and Technology, KIST School, Korea University of Science and Technology, Seoul 02792, Republic of Korea
| | - Yu-Chan Kim
- Center for Biomaterials, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
- Division of Bio-Medical Science and Technology, KIST School, Korea University of Science and Technology, Seoul 02792, Republic of Korea
| | - Youngdo Jeong
- Center for Advanced Biomolecular Recognition, Biomedical Research Division, Korea Institute of Science and Technology (KIST), Seoul 02792, Republic of Korea
- Department of HY-KIST Bio-convergence, Hanyang University, Seoul 04763, Republic of Korea
- Division of Bio-Medical Science and Technology, KIST School, Korea University of Science and Technology, Seoul 02792, Republic of Korea
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23
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Turley TN, Theis JL, Evans JM, Fogarty ZC, Gulati R, Hayes SN, Tweet MS, Olson TM. Identification of Rare Genetic Variants in Familial Spontaneous Coronary Artery Dissection and Evidence for Shared Biological Pathways. J Cardiovasc Dev Dis 2023; 10:393. [PMID: 37754822 PMCID: PMC10532385 DOI: 10.3390/jcdd10090393] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2023] [Accepted: 09/08/2023] [Indexed: 09/28/2023] Open
Abstract
Rare familial spontaneous coronary artery dissection (SCAD) kindreds implicate genetic disease predisposition and provide a unique opportunity for candidate gene discovery. Whole-genome sequencing was performed in fifteen probands with non-syndromic SCAD who had a relative with SCAD, eight of whom had a second relative with extra-coronary arteriopathy. Co-segregating variants and associated genes were prioritized by quantitative variant, gene, and disease-level metrics. Curated public databases were queried for functional relationships among encoded proteins. Fifty-four heterozygous coding variants in thirteen families co-segregated with disease and fulfilled primary filters of rarity, gene variation constraint, and predicted-deleterious protein effect. Secondary filters yielded 11 prioritized candidate genes in 12 families, with high arterial tissue expression (n = 7), high-confidence protein-level interactions with genes associated with SCAD previously (n = 10), and/or previous associations with connective tissue disorders and aortopathies (n = 3) or other vascular phenotypes in mice or humans (n = 11). High-confidence associations were identified among 10 familial SCAD candidate-gene-encoded proteins. A collagen-encoding gene was identified in five families, two with distinct variants in COL4A2. Familial SCAD is genetically heterogeneous, yet perturbations of extracellular matrix, cytoskeletal, and cell-cell adhesion proteins implicate common disease-susceptibility pathways. Incomplete penetrance and variable expression suggest genetic or environmental modifiers.
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Affiliation(s)
- Tamiel N. Turley
- Molecular Pharmacology and Experimental Therapeutics Track, Mayo Clinic Graduate School of Biomedical Sciences, Mayo Clinic, Rochester, MN 55905, USA;
- Cardiovascular Genetics Research Laboratory, Mayo Clinic, Rochester, MN 55905, USA;
| | - Jeanne L. Theis
- Cardiovascular Genetics Research Laboratory, Mayo Clinic, Rochester, MN 55905, USA;
| | - Jared M. Evans
- Department of Quantitative Health Sciences, Division of Computational Biology, Mayo Clinic, Rochester, MN 55905, USA; (J.M.E.); (Z.C.F.)
| | - Zachary C. Fogarty
- Department of Quantitative Health Sciences, Division of Computational Biology, Mayo Clinic, Rochester, MN 55905, USA; (J.M.E.); (Z.C.F.)
| | - Rajiv Gulati
- Department of Cardiovascular Medicine, Mayo Clinic, Rochester, MN 55905, USA; (R.G.); (S.N.H.); (M.S.T.)
| | - Sharonne N. Hayes
- Department of Cardiovascular Medicine, Mayo Clinic, Rochester, MN 55905, USA; (R.G.); (S.N.H.); (M.S.T.)
| | - Marysia S. Tweet
- Department of Cardiovascular Medicine, Mayo Clinic, Rochester, MN 55905, USA; (R.G.); (S.N.H.); (M.S.T.)
| | - Timothy M. Olson
- Cardiovascular Genetics Research Laboratory, Mayo Clinic, Rochester, MN 55905, USA;
- Department of Cardiovascular Medicine, Mayo Clinic, Rochester, MN 55905, USA; (R.G.); (S.N.H.); (M.S.T.)
- Department of Pediatric and Adolescent Medicine, Division of Pediatric Cardiology, Mayo Clinic, Rochester, MN 55905, USA
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24
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Adebayo OE, Urcun S, Rolin G, Bordas SPA, Trucu D, Eftimie R. Mathematical investigation of normal and abnormal wound healing dynamics: local and non-local models. MATHEMATICAL BIOSCIENCES AND ENGINEERING : MBE 2023; 20:17446-17498. [PMID: 37920062 DOI: 10.3934/mbe.2023776] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/04/2023]
Abstract
The movement of cells during (normal and abnormal) wound healing is the result of biomechanical interactions that combine cell responses with growth factors as well as cell-cell and cell-matrix interactions (adhesion and remodelling). It is known that cells can communicate and interact locally and non-locally with other cells inside the tissues through mechanical forces that act locally and at a distance, as well as through long non-conventional cell protrusions. In this study, we consider a non-local partial differential equation model for the interactions between fibroblasts, macrophages and the extracellular matrix (ECM) via a growth factor (TGF-$ \beta $) in the context of wound healing. For the non-local interactions, we consider two types of kernels (i.e., a Gaussian kernel and a cone-shaped kernel), two types of cell-ECM adhesion functions (i.e., adhesion only to higher-density ECM vs. adhesion to higher-/lower-density ECM) and two types of cell proliferation terms (i.e., with and without decay due to overcrowding). We investigate numerically the dynamics of this non-local model, as well as the dynamics of the localised versions of this model (i.e., those obtained when the cell perception radius decreases to 0). The results suggest the following: (ⅰ) local models explain normal wound healing and non-local models could also explain abnormal wound healing (although the results are parameter-dependent); (ⅱ) the models can explain two types of wound healing, i.e., by primary intention, when the wound margins come together from the side, and by secondary intention when the wound heals from the bottom up.
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Affiliation(s)
- O E Adebayo
- Laboratoire de mathématiques de Besançon, UMR CNRS 6623, Université de Franche-Comté, Besançon 25000, France
| | - S Urcun
- Department of Engineering, Faculty of Science, Technology and Medicine, University of Luxembourg, Esch-sur-Alzette, Luxembourg
| | - G Rolin
- INSERM CIC-1431, CHU Besançon, Besançon 25000, France
- Université de Franche-Comté, EFS, INSERM, UMR RIGHT, F-25000 Besançon, France
| | - S P A Bordas
- Department of Engineering, Faculty of Science, Technology and Medicine, University of Luxembourg, Esch-sur-Alzette, Luxembourg
| | - D Trucu
- Division of Mathematics, University of Dundee, Dundee, DD1 4HN, United Kingdom
| | - R Eftimie
- Laboratoire de mathématiques de Besançon, UMR CNRS 6623, Université de Franche-Comté, Besançon 25000, France
- Division of Mathematics, University of Dundee, Dundee, DD1 4HN, United Kingdom
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25
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Tseng YT, Lai R, Oieni F, Standke A, Smyth G, Yang C, Chen M, St John J, Ekberg J. Liraglutide modulates adhesion molecules and enhances cell properties in three-dimensional cultures of olfactory ensheathing cells. Biomed Pharmacother 2023; 165:115084. [PMID: 37399717 DOI: 10.1016/j.biopha.2023.115084] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2023] [Revised: 06/15/2023] [Accepted: 06/26/2023] [Indexed: 07/05/2023] Open
Abstract
Cell transplantation using olfactory ensheathing cells (OECs) is a promising approach for nerve repair but there are numerous limitations with their delivery method. Three-dimensional (3D) cell culture systems potentially offer a powerful approach for cell production and delivery options. To further optimise the use of OECs, strategies to promote cell viability and maintain cell behaviours in 3D cultures become important. We previously demonstrated an anti-diabetic drug, liraglutide, could modulate OEC migration and re-model extracellular matrix in two-dimensional (2D) cultures. In the present study, we further investigated its beneficial effects in our 3D culture system using primary OECs. OECs treated with liraglutide at 100 nM showed improved cell viability and had modulated expression of N-cadherin and β1-integrin (two important cell adhesion molecules). When formed into 3D spheroids, the pre-treated OECs generated spheroids with an increased volume and a decreased cell density compared to control spheroids. OECs that subsequently migrated out of the liraglutide pre-treated spheroids had higher capacity for migration with increased duration and length, which was attributed to a reduction in the pauses during the migration. Moreover, OECs that migrated out from liraglutide spheroids had a more bipolar morphology consistent with higher migratory capacity. In summary, liraglutide improved the viability of OECs, modulated cell adhesion molecules, and resulted in stable 3D cell constructs which conferred enhanced migratory capacity on the OECs. Overall, liraglutide may potentially improve the therapeutic use of OECs for neural repair by enhancing the generation of stable 3D constructs and increasing the migratory behaviour of OECs.
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Affiliation(s)
- Yu-Ting Tseng
- Clem Jones Centre for Neurobiology and Stem Cell Research, Griffith University, Brisbane, QLD 4111, Australia; Menzies Health Institute Queensland, Griffith University, Southport, QLD 4222, Australia; Griffith Institute for Drug Discovery, Griffith University, Brisbane, QLD 4111, Australia
| | - Richard Lai
- Menzies Health Institute Queensland, Griffith University, Southport, QLD 4222, Australia
| | - Francesca Oieni
- Clem Jones Centre for Neurobiology and Stem Cell Research, Griffith University, Brisbane, QLD 4111, Australia; Menzies Health Institute Queensland, Griffith University, Southport, QLD 4222, Australia
| | - Andrea Standke
- Clem Jones Centre for Neurobiology and Stem Cell Research, Griffith University, Brisbane, QLD 4111, Australia; Menzies Health Institute Queensland, Griffith University, Southport, QLD 4222, Australia
| | - Graham Smyth
- Clem Jones Centre for Neurobiology and Stem Cell Research, Griffith University, Brisbane, QLD 4111, Australia; Menzies Health Institute Queensland, Griffith University, Southport, QLD 4222, Australia
| | - Chenying Yang
- Clem Jones Centre for Neurobiology and Stem Cell Research, Griffith University, Brisbane, QLD 4111, Australia; Menzies Health Institute Queensland, Griffith University, Southport, QLD 4222, Australia
| | - Mo Chen
- Clem Jones Centre for Neurobiology and Stem Cell Research, Griffith University, Brisbane, QLD 4111, Australia; Menzies Health Institute Queensland, Griffith University, Southport, QLD 4222, Australia; Griffith Institute for Drug Discovery, Griffith University, Brisbane, QLD 4111, Australia.
| | - James St John
- Clem Jones Centre for Neurobiology and Stem Cell Research, Griffith University, Brisbane, QLD 4111, Australia; Menzies Health Institute Queensland, Griffith University, Southport, QLD 4222, Australia; Griffith Institute for Drug Discovery, Griffith University, Brisbane, QLD 4111, Australia.
| | - Jenny Ekberg
- Clem Jones Centre for Neurobiology and Stem Cell Research, Griffith University, Brisbane, QLD 4111, Australia; Menzies Health Institute Queensland, Griffith University, Southport, QLD 4222, Australia; Griffith Institute for Drug Discovery, Griffith University, Brisbane, QLD 4111, Australia.
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26
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Alberici Delsin LE, Plutoni C, Clouvel A, Keil S, Marpeaux L, Elouassouli L, Khavari A, Ehrlicher AJ, Emery G. MAP4K4 regulates forces at cell-cell and cell-matrix adhesions to promote collective cell migration. Life Sci Alliance 2023; 6:e202302196. [PMID: 37369604 PMCID: PMC10300198 DOI: 10.26508/lsa.202302196] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/02/2023] [Revised: 06/12/2023] [Accepted: 06/16/2023] [Indexed: 06/29/2023] Open
Abstract
Collective cell migration is not only important for development and tissue homeostasis but can also promote cancer metastasis. To migrate collectively, cells need to coordinate cellular extensions and retractions, adhesion sites dynamics, and forces generation and transmission. Nevertheless, the regulatory mechanisms coordinating these processes remain elusive. Using A431 carcinoma cells, we identify the kinase MAP4K4 as a central regulator of collective migration. We show that MAP4K4 inactivation blocks the migration of clusters, whereas its overexpression decreases cluster cohesion. MAP4K4 regulates protrusion and retraction dynamics, remodels the actomyosin cytoskeleton, and controls the stability of both cell-cell and cell-substrate adhesion. MAP4K4 promotes focal adhesion disassembly through the phosphorylation of the actin and plasma membrane crosslinker moesin but disassembles adherens junctions through a moesin-independent mechanism. By analyzing traction and intercellular forces, we found that MAP4K4 loss of function leads to a tensional disequilibrium throughout the cell cluster, increasing the traction forces and the tension loading at the cell-cell adhesions. Together, our results indicate that MAP4K4 activity is a key regulator of biomechanical forces at adhesion sites, promoting collective migration.
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Affiliation(s)
- Lara Elis Alberici Delsin
- Vesicular Trafficking and Cell Signalling Research Unit, Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Montréal, Canada
| | - Cédric Plutoni
- Vesicular Trafficking and Cell Signalling Research Unit, Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Montréal, Canada
| | - Anna Clouvel
- Department of Bioengineering, McGill University, Montreal, Canada
| | - Sarah Keil
- Vesicular Trafficking and Cell Signalling Research Unit, Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Montréal, Canada
| | - Léa Marpeaux
- Vesicular Trafficking and Cell Signalling Research Unit, Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Montréal, Canada
| | - Lina Elouassouli
- Vesicular Trafficking and Cell Signalling Research Unit, Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Montréal, Canada
| | - Adele Khavari
- Department of Bioengineering, McGill University, Montreal, Canada
| | | | - Gregory Emery
- Vesicular Trafficking and Cell Signalling Research Unit, Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Montréal, Canada
- Department of Pathology and Cell Biology, Faculty of Medicine, Université de Montréal, Montréal, Canada
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27
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Tixi W, Maldonado M, Chang YT, Chiu A, Yeung W, Parveen N, Nelson MS, Hart R, Wang S, Hsu WJ, Fueger P, Kopp JL, Huising MO, Dhawan S, Shih HP. Coordination between ECM and cell-cell adhesion regulates the development of islet aggregation, architecture, and functional maturation. eLife 2023; 12:e90006. [PMID: 37610090 PMCID: PMC10482429 DOI: 10.7554/elife.90006] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2023] [Accepted: 07/12/2023] [Indexed: 08/24/2023] Open
Abstract
Pancreatic islets are three-dimensional cell aggregates consisting of unique cellular composition, cell-to-cell contacts, and interactions with blood vessels. Cell aggregation is essential for islet endocrine function; however, it remains unclear how developing islets establish aggregation. By combining genetic animal models, imaging tools, and gene expression profiling, we demonstrate that islet aggregation is regulated by extracellular matrix signaling and cell-cell adhesion. Islet endocrine cell-specific inactivation of extracellular matrix receptor integrin β1 disrupted blood vessel interactions but promoted cell-cell adhesion and the formation of larger islets. In contrast, ablation of cell-cell adhesion molecule α-catenin promoted blood vessel interactions yet compromised islet clustering. Simultaneous removal of integrin β1 and α-catenin disrupts islet aggregation and the endocrine cell maturation process, demonstrating that establishment of islet aggregates is essential for functional maturation. Our study provides new insights into understanding the fundamental self-organizing mechanism for islet aggregation, architecture, and functional maturation.
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Affiliation(s)
- Wilma Tixi
- Department of Translational Research and Cellular Therapeutics, Arthur Riggs Diabetes and Metabolism Research Institute, Beckman Research Institute, City of HopeDuarteUnited States
| | - Maricela Maldonado
- Department of Translational Research and Cellular Therapeutics, Arthur Riggs Diabetes and Metabolism Research Institute, Beckman Research Institute, City of HopeDuarteUnited States
- Department of Biomedical Engineering, College of Engineering, California State University, Long BeachLong BeachUnited States
| | - Ya-Ting Chang
- Department of Translational Research and Cellular Therapeutics, Arthur Riggs Diabetes and Metabolism Research Institute, Beckman Research Institute, City of HopeDuarteUnited States
| | - Amy Chiu
- Department of Translational Research and Cellular Therapeutics, Arthur Riggs Diabetes and Metabolism Research Institute, Beckman Research Institute, City of HopeDuarteUnited States
| | - Wilson Yeung
- Department of Translational Research and Cellular Therapeutics, Arthur Riggs Diabetes and Metabolism Research Institute, Beckman Research Institute, City of HopeDuarteUnited States
| | - Nazia Parveen
- Department of Translational Research and Cellular Therapeutics, Arthur Riggs Diabetes and Metabolism Research Institute, Beckman Research Institute, City of HopeDuarteUnited States
| | - Michael S Nelson
- Light Microscopy Core, Beckman Research Institute, City of HopeDuarteUnited States
| | - Ryan Hart
- Department of Neurobiology, Physiology and Behavior, University of California, DavisDavisUnited States
| | - Shihao Wang
- Department of Cellular and Physiological Sciences, Life Sciences Institute, University of British ColumbiaVancouverCanada
| | - Wu Jih Hsu
- Department of Cellular and Physiological Sciences, Life Sciences Institute, University of British ColumbiaVancouverCanada
| | - Patrick Fueger
- Department of Molecular & Cellular Endocrinology, Arthur Riggs Diabetes and Metabolism Research Institute, Beckman Research Institute, City of HopeDuarteUnited States
| | - Janel L Kopp
- Department of Cellular and Physiological Sciences, Life Sciences Institute, University of British ColumbiaVancouverCanada
| | - Mark O Huising
- Department of Neurobiology, Physiology and Behavior, University of California, DavisDavisUnited States
- Department of Physiology and Membrane Biology, School of Medicine, University of California, DavisDavisUnited States
| | - Sangeeta Dhawan
- Department of Translational Research and Cellular Therapeutics, Arthur Riggs Diabetes and Metabolism Research Institute, Beckman Research Institute, City of HopeDuarteUnited States
| | - Hung Ping Shih
- Department of Translational Research and Cellular Therapeutics, Arthur Riggs Diabetes and Metabolism Research Institute, Beckman Research Institute, City of HopeDuarteUnited States
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28
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Kulus J, Kranc W, Kulus M, Dzięgiel P, Bukowska D, Mozdziak P, Kempisty B, Antosik P. Expression of genes regulating cell division in porcine follicular granulosa cells. Cell Div 2023; 18:12. [PMID: 37550786 PMCID: PMC10408085 DOI: 10.1186/s13008-023-00094-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/19/2023] [Accepted: 07/27/2023] [Indexed: 08/09/2023] Open
Abstract
BACKGROUND Cell cycle regulation influences the proliferation of granulosa cells and affects many processes related to ovarian folliclular growth and ovulation. Abnormal regulation of the cell cycle can lead to many diseases within the ovary. The aim of this study was to describe the expression profile of genes within granulosa cells, which are related to the formation of the cytoskeleton, organization of cell organelles inside the cell, and regulation of cell division. Established in vitro primary cultures from porcine ovarian follicle granulosa cells were maintained for 48, 96, 144 h and evaluated via microarray expression analysis. RESULTS Analyzed genes were assigned to 12 gene ontology groups "actin cytoskeleton organization", "actin filament organization", "actin filament-based process", "cell-matrix adhesion", "cell-substrate adhesion", "chromosome segregation", "chromosome separation", "cytoskeleton organization", "DNA integrity checkpoint", "DNA replication initiation", "organelle fision", "organelle organization". Among the genes with significantly changed expression, those whose role in processes within the ovary are selected for consideration. Genes with increased expression include (ITGA11, CNN1, CCl2, TPM2, ACTN1, VCAM-1, COL3A1, GSN, FRMD6, PLK2). Genes with reduced expression inlcude (KIF14, TACC3, ESPL1, CDC45, TTK, CDC20, CDK1, FBXO5, NEK2-NIMA, CCNE2). For the results obtained by microarray expressions, quantitative validation by RT-qPCR was performed. CONCLUSIONS The results indicated expression profile of genes, which can be considered as new molecular markers of cellular processes involved in signaling, cell structure organization. The expression profile of selected genes brings new insight into regulation of physiological processes in porcine follicular granulosa cells during primary in vitro culture.
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Affiliation(s)
- Jakub Kulus
- Department of Diagnostics and Clinical Sciences, Institute of Veterinary Medicine, Nicolaus Copernicus University in Torun, Torun, Poland
| | - Wiesława Kranc
- Department of Anatomy, Poznan University of Medical Sciences, Poznan, Poland
| | - Magdalena Kulus
- Department of Veterinary Surgery, Institute of Veterinary Medicine, Nicolaus Copernicus University in Torun, Torun, Poland
| | - Piotr Dzięgiel
- Division of Histology and Embryology, Department of Human Morphology and Embryology, Wroclaw Medical University, Wroclaw, Poland
- Department of Physiotherapy, Wroclaw University School of Physical Education, Wroclaw, Poland
| | - Dorota Bukowska
- Department of Diagnostics and Clinical Sciences, Institute of Veterinary Medicine, Nicolaus Copernicus University in Torun, Torun, Poland
| | - Paul Mozdziak
- Physiology Graduate Faculty, College of Agriculture and Life Sciences, North Carolina State University, Raleigh, NC, 27695, USA
| | - Bartosz Kempisty
- Department of Veterinary Surgery, Institute of Veterinary Medicine, Nicolaus Copernicus University in Torun, Torun, Poland.
- Physiology Graduate Faculty, College of Agriculture and Life Sciences, North Carolina State University, Raleigh, NC, 27695, USA.
- Division of Anatomy, Department of Human Morphology and Embryology, Wroclaw Medical University, Wroclaw, Poland.
- Center of Assisted Reproduction, Department of Obstetrics and Gynecology, University Hospital and Masaryk University, Brno, Czech Republic.
| | - Paweł Antosik
- Department of Veterinary Surgery, Institute of Veterinary Medicine, Nicolaus Copernicus University in Torun, Torun, Poland
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29
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Dienemann S, Schmidt V, Fleischhammer T, Mueller JH, Lavrentieva A. Comparative analysis of hypoxic response of human microvascular and umbilical vein endothelial cells in 2D and 3D cell culture systems. J Cell Physiol 2023; 238:1111-1120. [PMID: 36947660 DOI: 10.1002/jcp.31002] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2022] [Revised: 02/24/2023] [Accepted: 03/07/2023] [Indexed: 03/24/2023]
Abstract
In vitro cultivation conditions play a crucial role in cell physiology and the cellular response to external stimuli. Oxygen concentrations represent an essential microenvironmental factor influencing cell physiology and behaviour both in vivo and in vitro. Therefore, new approaches are urgently needed to monitor and control oxygen concentrations in 2D and 3D cultures, as well as cell reactions to these concentrations. In this work, we modified two types of human endothelial cells-human microvascular (huMECs) and umbilical vein endothelial cells (huVECs) with genetically encoded hypoxia biosensors and monitored cell reactions in 2D to different oxygen concentrations. Moreover, we fabricated 3D cell spheroids of different cell numbers and sizes to reveal the onset of hypoxia in huVECs and huMECs. We could demonstrate a quantitative sensor response of two cell types to reduced oxygen supply in 2D and reveal different thresholds for hypoxic response. In 3D cell spheroids we could estimate critical construct sizes for the appearance of a hypoxic core. This work for the first time directly demonstrates different hypoxic signatures for huVECs and huMECs in 2D and 3D cell culture systems.
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Affiliation(s)
- Sandra Dienemann
- Institute of Technical Chemistry, Leibniz University of Hannover, Hannover, Germany
| | - Vanessa Schmidt
- Institute of Technical Chemistry, Leibniz University of Hannover, Hannover, Germany
| | - Tabea Fleischhammer
- Institute of Technical Chemistry, Leibniz University of Hannover, Hannover, Germany
| | - Julia H Mueller
- Institute of Technical Chemistry, Leibniz University of Hannover, Hannover, Germany
- Department of Cardiology and Angiology, Hannover Medical School, Hannover, Germany
| | - Antonina Lavrentieva
- Institute of Technical Chemistry, Leibniz University of Hannover, Hannover, Germany
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Karthika CL, Venugopal V, Sreelakshmi BJ, Krithika S, Thomas JM, Abraham M, Kartha CC, Rajavelu A, Sumi S. Oscillatory shear stress modulates Notch-mediated endothelial mesenchymal plasticity in cerebral arteriovenous malformations. Cell Mol Biol Lett 2023; 28:22. [PMID: 36934253 PMCID: PMC10024393 DOI: 10.1186/s11658-023-00436-x] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/06/2022] [Accepted: 03/06/2023] [Indexed: 03/20/2023] Open
Abstract
BACKGROUND Cerebral arteriovenous malformations (cAVM) are a significant cause of intracranial hemorrhagic stroke and brain damage. The arteriovenous junctions in AVM nidus are known to have hemodynamic disturbances such as altered shear stress, which could lead to endothelial dysfunction. The molecular mechanisms coupling shear stress and endothelial dysfunction in cAVMs are poorly understood. We speculated that disturbed blood flow in artery-vein junctions activates Notch receptors and promotes endothelial mesenchymal plasticity during cAVM formation. METHODS We investigated the expression profile of endothelial mesenchymal transition (EndMT) and cell adhesion markers, as well as activated Notch receptors, in 18 human cAVM samples and 15 control brain tissues, by quantitative real-time PCR (qRT-PCR) and immunohistochemical evaluation. Employing a combination of a microfluidic system, qRT-PCR, immunofluorescence, as well as invasion and inhibitor assays, the effects of various shear stress conditions on Notch-induced EndMT and invasive potential of human cerebral microvascular endothelial cells (hCMEC/d3) were analyzed. RESULTS We found evidence for EndMT and enhanced expression of activated Notch intracellular domain (NICD3 and NICD4) in human AVM nidus samples. The expression of transmembrane adhesion receptor integrin α9/β1 is significantly reduced in cAVM nidal vessels. Cell-cell adhesion proteins such as VE-cadherin and N-cadherin were differentially expressed in AVM nidus compared with control brain tissues. Using well-characterized hCMECs, we show that altered fluid shear stress steers Notch3 nuclear translocation and promotes SNAI1/2 expression and nuclear localization. Oscillatory flow downregulates integrin α9/β1 and VE-cadherin expression, while N-cadherin expression and endothelial cell invasiveness are augmented. Gamma-secretase inhibitor RO4929097, and to a lesser level DAPT, prevent the mesenchymal transition and invasiveness of cerebral microvascular endothelial cells exposed to oscillatory fluid flow. CONCLUSIONS Our study provides, for the first time, evidence for the role of oscillatory shear stress in mediating the EndMT process and dysregulated expression of cell adhesion molecules, especially multifunctional integrin α9/β1 in human cAVM nidus. Concomitantly, our findings indicate the potential use of small-molecular inhibitors such as RO4929097 in the less-invasive therapeutic management of cAVMs.
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Affiliation(s)
- C L Karthika
- Cardiovascular Diseases and Diabetes Biology, Rajiv Gandhi Centre for Biotechnology (RGCB), Thiruvananthapuram, Kerala, 695014, India
| | - Vani Venugopal
- Cardiovascular Diseases and Diabetes Biology, Rajiv Gandhi Centre for Biotechnology (RGCB), Thiruvananthapuram, Kerala, 695014, India
| | - B J Sreelakshmi
- Cardiovascular Diseases and Diabetes Biology, Rajiv Gandhi Centre for Biotechnology (RGCB), Thiruvananthapuram, Kerala, 695014, India
| | - S Krithika
- Cardiovascular Diseases and Diabetes Biology, Rajiv Gandhi Centre for Biotechnology (RGCB), Thiruvananthapuram, Kerala, 695014, India
| | - Jaya Mary Thomas
- Cardiovascular Diseases and Diabetes Biology, Rajiv Gandhi Centre for Biotechnology (RGCB), Thiruvananthapuram, Kerala, 695014, India
| | - Mathew Abraham
- Department of Neurosurgery, Sree Chitra Tirunal Institute for Medical Sciences and Technology, Thiruvananthapuram, Kerala, 695011, India
| | - C C Kartha
- Department of Neurology, Amrita Institute of Medical Sciences, Amrita Vishwa Vidyapeetham, Kochi, Kerala, 682041, India
| | - Arumugam Rajavelu
- Department of Biotechnology, Bhupat & Jyoti Mehta School of Biosciences, Indian Institute of Technology, Madras, Chennai, Tamil Nadu, 600036, India
| | - S Sumi
- Cardiovascular Diseases and Diabetes Biology, Rajiv Gandhi Centre for Biotechnology (RGCB), Thiruvananthapuram, Kerala, 695014, India.
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Li X, McLain C, Samuel MS, Olson MF, Radice GL. Actomyosin-mediated cellular tension promotes Yap nuclear translocation and myocardial proliferation through α5 integrin signaling. Development 2023; 150:dev201013. [PMID: 36621002 PMCID: PMC10110499 DOI: 10.1242/dev.201013] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/09/2022] [Accepted: 12/19/2022] [Indexed: 01/10/2023]
Abstract
The cardiomyocyte phenotypic switch from a proliferative to terminally differentiated state results in the loss of regenerative potential of the mammalian heart shortly after birth. Nonmuscle myosin IIB (NM IIB)-mediated actomyosin contractility regulates cardiomyocyte cytokinesis in the embryonic heart, and NM IIB levels decline after birth, suggesting a role for cellular tension in the regulation of cardiomyocyte cell cycle activity in the postnatal heart. To investigate the role of actomyosin contractility in cardiomyocyte cell cycle arrest, we conditionally activated ROCK2 kinase domain (ROCK2:ER) in the murine postnatal heart. Here, we show that α5/β1 integrin and fibronectin matrix increase in response to actomyosin-mediated tension. Moreover, activation of ROCK2:ER promotes nuclear translocation of Yap, a mechanosensitive transcriptional co-activator, and enhances cardiomyocyte proliferation. Finally, we show that reduction of myocardial α5 integrin rescues the myocardial proliferation phenotype in ROCK2:ER hearts. These data demonstrate that cardiomyocytes respond to increased intracellular tension by altering their intercellular contacts in favor of cell-matrix interactions, leading to Yap nuclear translocation, thus uncovering a function for nonmuscle myosin contractility in promoting cardiomyocyte proliferation in the postnatal heart.
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Affiliation(s)
- Xiaofei Li
- Cardiovascular Research Center, Lifespan Cardiovascular Institute, Rhode Island Hospital, Department of Medicine, Division of Cardiology, Alpert Medical School of Brown University, Providence, RI 02903, USA
| | - Callie McLain
- Cardiovascular Research Center, Lifespan Cardiovascular Institute, Rhode Island Hospital, Department of Medicine, Division of Cardiology, Alpert Medical School of Brown University, Providence, RI 02903, USA
| | - Michael S. Samuel
- Centre for Cancer Biology, an alliance between SA Pathology and the University of South Australia, Adelaide 5000, Australia
- Adelaide Medical School, Faculty of Health and Medical Sciences, University of Adelaide, Adelaide 5000, Australia
| | - Michael F. Olson
- Department of Chemistry and Biology, Toronto Metropolitan University, Toronto, Ontario, M5B 2K3 Canada
| | - Glenn L. Radice
- Cardiovascular Research Center, Lifespan Cardiovascular Institute, Rhode Island Hospital, Department of Medicine, Division of Cardiology, Alpert Medical School of Brown University, Providence, RI 02903, USA
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Valat A, Fourel L, Sales A, Machillot P, Bouin AP, Fournier C, Bosc L, Arboléas M, Bourrin-Reynard I, Wagoner Johnson AJ, Bruckert F, Albigès-Rizo C, Picart C. Interplay between integrins and cadherins to control bone differentiation upon BMP-2 stimulation. Front Cell Dev Biol 2023; 10:1027334. [PMID: 36684447 PMCID: PMC9846056 DOI: 10.3389/fcell.2022.1027334] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/24/2022] [Accepted: 12/05/2022] [Indexed: 01/06/2023] Open
Abstract
Introduction: Upon BMP-2 stimulation, the osteoblastic lineage commitment in C2C12 myoblasts is associated with a microenvironmental change that occurs over several days. How does BMP-2 operate a switch in adhesive machinery to adapt to the new microenvironment and to drive bone cell fate is not well understood. Here, we addressed this question for BMP-2 delivered either in solution or physically bound of a biomimetic film, to mimic its presentation to cells via the extracellular matrix (ECM). Methods: Biommetics films were prepared using a recently developed automated method that enable high content studies of cellular processes. Comparative gene expressions were done using RNA sequencing from the encyclopedia of the regulatory elements (ENCODE). Gene expressions of transcription factors, beta chain (1, 3, 5) integrins and cadherins (M, N, and Cad11) were studied using quantitative PCR. ECM proteins and adhesion receptor expressions were also quantified by Western blots and dot blots. Their spatial organization in and around cells was studied using immuno-stainings. The individual effect of each receptor on osteogenic transcription factors and alkaline phosphatase expression were studied using silencing RNA of each integrin and cadherin receptor. The organization of fibronectin was studied using immuno-staining and quantitative microscopic analysis. Results: Our findings highlight a switch of integrin and cadherin expression during muscle to bone transdifferentiation upon BMP-2 stimulation. This switch occurs no matter the presentation mode, for BMP-2 presented in solution or via the biomimetic film. While C2C12 muscle cells express M-cadherin and Laminin-specific integrins, the BMP-2-induced transdifferentiation into bone cells is associated with an increase in the expression of cadherin-11 and collagen-specific integrins. Biomimetic films presenting matrix-bound BMP-2 enable the revelation of specific roles of the adhesive receptors depending on the transcription factor. Discussion: While β3 integrin and cadherin-11 work in concert to control early pSMAD1,5,9 signaling, β1 integrin and Cadherin-11 control RunX2, ALP activity and fibronectin organization around the cells. In contrast, while β1 integrin is also important for osterix transcriptional activity, Cadherin-11 and β5 integrin act as negative osterix regulators. In addition, β5 integrin negatively regulates RunX2. Our results show that biomimetic films can be used to delinate the specific events associated with BMP-2-mediated muscle to bone transdifferentiation. Our study reveals how integrins and cadherins work together, while exerting distinct functions to drive osteogenic programming. Different sets of integrins and cadherins have complementary mechanical roles during the time window of this transdifferentiation.
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Affiliation(s)
- Anne Valat
- Grenoble Institute of Engineering, CNRS UMR 5628, LMGP, Grenoble, France
| | - Laure Fourel
- Grenoble Institute of Engineering, CNRS UMR 5628, LMGP, Grenoble, France
| | - Adria Sales
- U1292 Biosanté, INSERM, CEA, CNRS EMR 5000 Biomimetism and Regenerative Medicine, University Grenoble Alpes, Grenoble, France
| | - Paul Machillot
- Grenoble Institute of Engineering, CNRS UMR 5628, LMGP, Grenoble, France
- U1292 Biosanté, INSERM, CEA, CNRS EMR 5000 Biomimetism and Regenerative Medicine, University Grenoble Alpes, Grenoble, France
| | - Anne-Pascale Bouin
- U1209 Institut for Advanced Biosciences, CNRS 5309, University Grenoble Alpes, La Tronche, France
| | - Carole Fournier
- Grenoble Institute of Engineering, CNRS UMR 5628, LMGP, Grenoble, France
| | - Lauriane Bosc
- U1292 Biosanté, INSERM, CEA, CNRS EMR 5000 Biomimetism and Regenerative Medicine, University Grenoble Alpes, Grenoble, France
| | - Mélanie Arboléas
- Grenoble Institute of Engineering, CNRS UMR 5628, LMGP, Grenoble, France
| | - Ingrid Bourrin-Reynard
- U1209 Institut for Advanced Biosciences, CNRS 5309, University Grenoble Alpes, La Tronche, France
| | - Amy J. Wagoner Johnson
- Grenoble Institute of Engineering, CNRS UMR 5628, LMGP, Grenoble, France
- Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign, Urbana, IL, United States
- Carle Illinois College of Medicine, Urbana, IL, United States
- Carl R. Woese Institute for Genomic Biology, Urbana, IL, United States
| | - Franz Bruckert
- Grenoble Institute of Engineering, CNRS UMR 5628, LMGP, Grenoble, France
| | - Corinne Albigès-Rizo
- U1209 Institut for Advanced Biosciences, CNRS 5309, University Grenoble Alpes, La Tronche, France
| | - Catherine Picart
- Grenoble Institute of Engineering, CNRS UMR 5628, LMGP, Grenoble, France
- U1292 Biosanté, INSERM, CEA, CNRS EMR 5000 Biomimetism and Regenerative Medicine, University Grenoble Alpes, Grenoble, France
- Institut Universitaire de France (IUF), Paris, France
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Luís C, Soares R, Fernandes R, Botelho M. Cell-adhesion Molecules as Key Mechanisms of Tumor Invasion: The Case of Breast Cancer. Curr Mol Med 2023; 23:147-160. [PMID: 34365950 DOI: 10.2174/1566524021666210806155231] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/03/2020] [Revised: 03/10/2021] [Accepted: 03/23/2021] [Indexed: 12/16/2022]
Abstract
Cancer is a major health problem worldwide and the second leading cause of death following cardiovascular diseases. Breast cancer is the leading cause of mortality and morbidity among women and one of the most common malignant neoplasms prompt to metastatic disease. In the present review, the mechanisms of the major cell adhesion molecules involved in tumor invasion are discussed, focusing on the case of breast cancer. A non-systematic updated revision of the literature was performed in order to assemble information regarding the expression of the adhesion cell molecules associated with metastasis.
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Affiliation(s)
- Carla Luís
- Department of Biomedicine, Faculty of Medicine, University of Porto, Porto, Portugal
- Instituto de investigação e inovação em saúde, i3s, University of Porto, Porto, Portugal
- LABMI-PORTIC, Laboratory of Medical & Industrial Biotechnology, Porto Research, Technology and Innovation Center, Porto, Portugal
| | - Raquel Soares
- Department of Biomedicine, Faculty of Medicine, University of Porto, Porto, Portugal
- Instituto de investigação e inovação em saúde, i3s, University of Porto, Porto, Portugal
| | - Rúben Fernandes
- Departament of Biomedicine, Faculty of Medicine, University of Porto, Porto, Portugal
- LABMI-PORTIC, Laboratory of Medical & Industrial Biotechnology, Porto Research, Technology and Innovation Center, Porto, Portu
| | - Mónica Botelho
- Instituto de investigação e inovação em saúde, i3s, University of Porto, Porto, Portugal
- National Health Institute Ricardo Jorge, Porto, Portugal; Polytechnic Institute of Porto, Porto, Portugal
- LABMI-PORTIC, Laboratory of Medical & Industrial Biotechnology, Porto Research, Technology and Innovation Center, Porto, Portugal
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Fischer NG, Aparicio C. Junctional epithelium and hemidesmosomes: Tape and rivets for solving the "percutaneous device dilemma" in dental and other permanent implants. Bioact Mater 2022; 18:178-198. [PMID: 35387164 PMCID: PMC8961425 DOI: 10.1016/j.bioactmat.2022.03.019] [Citation(s) in RCA: 28] [Impact Index Per Article: 9.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2021] [Revised: 02/14/2022] [Accepted: 03/12/2022] [Indexed: 02/06/2023] Open
Abstract
The percutaneous device dilemma describes etiological factors, centered around the disrupted epithelial tissue surrounding non-remodelable devices, that contribute to rampant percutaneous device infection. Natural percutaneous organs, in particular their extracellular matrix mediating the "device"/epithelium interface, serve as exquisite examples to inspire longer lasting long-term percutaneous device design. For example, the tooth's imperviousness to infection is mediated by the epithelium directly surrounding it, the junctional epithelium (JE). The hallmark feature of JE is formation of hemidesmosomes, cell/matrix adhesive structures that attach surrounding oral gingiva to the tooth's enamel through a basement membrane. Here, the authors survey the multifaceted functions of the JE, emphasizing the role of the matrix, with a particular focus on hemidesmosomes and their five main components. The authors highlight the known (and unknown) effects dental implant - as a model percutaneous device - placement has on JE regeneration and synthesize this information for application to other percutaneous devices. The authors conclude with a summary of bioengineering strategies aimed at solving the percutaneous device dilemma and invigorating greater collaboration between clinicians, bioengineers, and matrix biologists.
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Affiliation(s)
- Nicholas G. Fischer
- MDRCBB-Minnesota Dental Research Center for Biomaterials and Biomechanics, University of Minnesota, 16-212 Moos Tower, 515 Delaware St. SE, Minneapolis, MN, 55455, USA
| | - Conrado Aparicio
- MDRCBB-Minnesota Dental Research Center for Biomaterials and Biomechanics, University of Minnesota, 16-212 Moos Tower, 515 Delaware St. SE, Minneapolis, MN, 55455, USA
- Division of Basic Research, Faculty of Odontology, UIC Barcelona – Universitat Internacional de Catalunya, C/. Josep Trueta s/n, 08195, Sant Cugat del Valles, Barcelona, Spain
- Institute for Bioengineering of Catalonia (IBEC), The Barcelona Institute of Science and Technology (BIST), C/. Baldiri Reixac 10-12, 08028, Barcelona, Spain
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Chau TCY, Keyser MS, Da Silva JA, Morris EK, Yordanov TE, Duscyz KP, Paterson S, Yap AS, Hogan BM, Lagendijk AK. Dynamically regulated focal adhesions coordinate endothelial cell remodelling in developing vasculature. Development 2022; 149:285926. [PMID: 36314606 DOI: 10.1242/dev.200454] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2021] [Accepted: 10/20/2022] [Indexed: 12/13/2022]
Abstract
The assembly of a mature vascular network involves coordinated endothelial cell (EC) shape changes, including the process of EC elongation. How EC elongation is dynamically regulated in vivo is not fully understood. Here, we have generated a zebrafish mutant that is deficient for the integrin adaptor protein Talin 1 (Tln1). Using a new focal adhesion (FA) marker line expressing endothelial Vinculinb-eGFP, we demonstrate that EC FAs function dynamically and are lost in our tln1 mutants, allowing us to uncouple the primary roles of FAs in EC morphogenesis from the secondary effects that occur due to systemic vessel failure or loss of blood flow. Tln1 loss led to compromised F-actin rearrangements, perturbed EC elongation and disrupted cell-cell junction linearisation in vessel remodelling. Finally, chemical induction of actin polymerisation restored actin dynamics and EC elongation during vascular morphogenesis. Together, we identify that FAs are essential for EC elongation and junction linearisation in flow-pressured vessels and that they influence actin polymerisation in cellular morphogenesis. These observations can explain the severely compromised vessel beds and vascular leakage observed in mutant models that lack integrin signalling. This article has an associated 'The people behind the papers' interview.
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Affiliation(s)
- Tevin C Y Chau
- Division of Cell and Developmental Biology, Institute for Molecular Bioscience, The University of Queensland, St Lucia, Queensland 4072, Australia
| | - Mikaela S Keyser
- Division of Cell and Developmental Biology, Institute for Molecular Bioscience, The University of Queensland, St Lucia, Queensland 4072, Australia
| | - Jason A Da Silva
- Division of Cell and Developmental Biology, Institute for Molecular Bioscience, The University of Queensland, St Lucia, Queensland 4072, Australia
| | - Elysse K Morris
- Division of Cell and Developmental Biology, Institute for Molecular Bioscience, The University of Queensland, St Lucia, Queensland 4072, Australia
| | - Teodor E Yordanov
- Division of Cell and Developmental Biology, Institute for Molecular Bioscience, The University of Queensland, St Lucia, Queensland 4072, Australia
| | - Kinga P Duscyz
- Division of Cell and Developmental Biology, Institute for Molecular Bioscience, The University of Queensland, St Lucia, Queensland 4072, Australia
| | - Scott Paterson
- Organogenesis and Cancer Program, Peter MacCallum Cancer Centre and The PeterMac Callum Department of Oncology, The University of Melbourne, Melbourne, Victoria 3000, Australia
| | - Alpha S Yap
- Division of Cell and Developmental Biology, Institute for Molecular Bioscience, The University of Queensland, St Lucia, Queensland 4072, Australia
| | - Benjamin M Hogan
- Organogenesis and Cancer Program, Peter MacCallum Cancer Centre and The PeterMac Callum Department of Oncology, The University of Melbourne, Melbourne, Victoria 3000, Australia.,Department of Anatomy and Physiology, The University of Melbourne, Parkville, Victoria 3010, Australia
| | - Anne Karine Lagendijk
- Division of Cell and Developmental Biology, Institute for Molecular Bioscience, The University of Queensland, St Lucia, Queensland 4072, Australia.,School of Biomedical Sciences, The University of Queensland, Brisbane, Queensland 4072, Australia
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36
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Zaman R, Islam RA, Chowdhury EH. Evolving therapeutic proteins to precisely kill cancer cells. J Control Release 2022; 351:779-804. [DOI: 10.1016/j.jconrel.2022.09.066] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2022] [Revised: 09/27/2022] [Accepted: 09/29/2022] [Indexed: 10/31/2022]
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Santerre K, Cortez Ghio S, Proulx S. TGF-β-Mediated Modulation of Cell-Cell Interactions in Postconfluent Maturing Corneal Endothelial Cells. Invest Ophthalmol Vis Sci 2022; 63:3. [PMID: 36194422 PMCID: PMC9547359 DOI: 10.1167/iovs.63.11.3] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022] Open
Abstract
Purpose Transforming growth factor-beta (TGF-β) is known to influence many cell functions. In the corneal endothelium, TGF-β1 exerts contextual effects, promoting endothelial–mesenchymal transition in proliferating cells and enhancing barrier integrity in early confluent maturing cells. Herein, we studied how TGF-β isoforms participate in the formation of corneal endothelial intercellular junctions. Methods Corneal endothelial cells (CECs) were cultured using a two-phase media approach. When CECs reached confluence, the proliferation medium was replaced with maturation medium, which was supplemented or not with TGF-β isoforms. The cell morphology (circularity index), intercellular junction protein expression, trans-endothelial electrical resistance (TEER), and permeability of 7-day postconfluent CECs were assessed. Gene transcription and signaling pathways that were activated following maturation in the presence of TGF-β2 were also studied. The beneficial effect of TGF-β2 on CEC maturation was evaluated using ex vivo corneas mounted on a corneal bioreactor. Results The results showed increases in circularity index, membrane localization of junction-related proteins, and TEER when TGF-β isoforms were individually added during the maturation phase, and TGF-β2 was the most effective isoform. Gene profiling revealed an increase in extracellular matrix-related gene expression. In ex vivo cell adhesion experiments, CECs that were matured in the presence of TGF-β2 had a higher circularity index and cell density and exhibited cell membrane-localized junction-related protein expression at earlier time points. Conclusions These results suggest that TGF-β2 can strengthen cell–cell and cell–substrate adhesion, which accelerates barrier integrity establishment and thus enhances CEC functionality.
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Affiliation(s)
- Kim Santerre
- Centre de recherche du Centre hospitalier universitaire (CHU) de Québec-Université Laval, axe médecine régénératrice, Hôpital du Saint-Sacrement, Québec, Québec, Canada.,Centre de recherche en organogénèse expérimentale de l'Université Laval/LOEX, Québec, Québec, Canada.,Département d'Ophtalmologie et d'oto-rhino-laryngologie-chirurgie cervico-faciale, Faculté de médecine, Université Laval, Québec, Québec, Canada
| | - Sergio Cortez Ghio
- Centre de recherche du Centre hospitalier universitaire (CHU) de Québec-Université Laval, axe médecine régénératrice, Hôpital du Saint-Sacrement, Québec, Québec, Canada.,Centre de recherche en organogénèse expérimentale de l'Université Laval/LOEX, Québec, Québec, Canada
| | - Stéphanie Proulx
- Centre de recherche du Centre hospitalier universitaire (CHU) de Québec-Université Laval, axe médecine régénératrice, Hôpital du Saint-Sacrement, Québec, Québec, Canada.,Centre de recherche en organogénèse expérimentale de l'Université Laval/LOEX, Québec, Québec, Canada.,Département d'Ophtalmologie et d'oto-rhino-laryngologie-chirurgie cervico-faciale, Faculté de médecine, Université Laval, Québec, Québec, Canada
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Cessna H, Baritaki S, Zaravinos A, Bonavida B. The Role of RKIP in the Regulation of EMT in the Tumor Microenvironment. Cancers (Basel) 2022; 14:cancers14194596. [PMID: 36230521 PMCID: PMC9559516 DOI: 10.3390/cancers14194596] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2022] [Revised: 09/21/2022] [Accepted: 09/21/2022] [Indexed: 12/03/2022] Open
Abstract
Simple Summary Raf kinase inhibitor protein (RKIP) expression in cancer cells is significantly reduced and promoting cancer cells growth and invasiveness. Overexpresssion of RKIP has been reported to mediate pleiotropic anti-cancer activities including the inhibition of survival signaling pathways, sensitization to cell death by cytotoxic drugs, inhibition of invasion, EMT and metastasis. The molecular mechanism by which RKIP inhibits EMT is not clear. In this review, we have examined how RKIP inhibits the selected EMT gene products (Snail, vimentin, N-cadherin, laminin alpha) and found that it involves signaling cross-talks between RKIP and each of the EMT gene products. These findings were validated by bioinformatic analyses demonstrating in various human cancers a negative correlation between the expression of RKIP and the expression of the EMT gene products. These findings suggest that targeting RKIP induction in cancer cells will result in multiple hits by inhibiting tumor growth, metastasis and reversal of chemo-immuno resistance. Abstract The Raf Kinase Inhibitor Protein (RKIP) is a unique gene product that directly inhibits the Raf/Mek/Erk and NF-kB pathways in cancer cells and resulting in the inhibition of cell proliferation, viability, EMT, and metastasis. Additionally, RKIP is involved in the regulation of cancer cell resistance to both chemotherapy and immunotherapy. The low expression of RKIP expression in many cancer types is responsible, in part, for the pathogenesis of cancer and its multiple properties. The inhibition of EMT and metastasis by RKIP led to its classification as a tumor suppressor. However, the mechanism by which RKIP mediates its inhibitory effects on EMT and metastases was not clear. We have proposed that one mechanism involves the negative regulation by RKIP of the expression of various gene products that mediate the mesenchymal phenotype as well as the positive regulation of gene products that mediate the epithelial phenotype via signaling cross talks between RKIP and each gene product. We examined several EMT mesenchymal gene products such as Snail, vimentin, N-cadherin, laminin and EPCAM and epithelial gene products such as E-cadherin and laminin. We have found that indeed these negative and positive correlations were detected in the signaling cross-talks. In addition, we have also examined bioinformatic data sets on different human cancers and the findings corroborated, in large part, the findings observed in the signaling cross-talks with few exceptions in some cancer types. The overall findings support the underlying mechanism by which the tumor suppressor RKIP regulates the expression of gene products involved in EMT and metastasis. Hence, the development of agent that can selectively induce RKIP expression in cancers with low expressions should result in the activation of the pleiotropic anti-cancer activities of RKIP and resulting in multiple effects including inhibition of tumor cell proliferation, EMT, metastasis and sensitization of resistant tumor cells to respond to both chemotherapeutics and immunotherapeutics.
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Affiliation(s)
- Hannah Cessna
- Department of Microbiology, Immunology & Molecular Genetics, David Geffen School of Medicine, Jonsson Comprehensive Cancer Center, University of California at Los Angeles, Los Angeles, CA 90095, USA
| | - Stavroula Baritaki
- Laboratory of Experimental Oncology, Division of Surgery, School of Medicine, University of Crete, 71003 Heraklion, Greece
| | - Apostolos Zaravinos
- Department of Life Sciences, School of Sciences, European University Cyprus, Nicosia 2404, Cyprus
- Basic and Translational Cancer Research Center (BTCRC), Cancer Genetics, Genomics and Systems Biology Laboratory, Nicosia 1516, Cyprus
| | - Benjamin Bonavida
- Department of Microbiology, Immunology & Molecular Genetics, David Geffen School of Medicine, Jonsson Comprehensive Cancer Center, University of California at Los Angeles, Los Angeles, CA 90095, USA
- Correspondence:
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Ouyang M, Zhu Y, Wang J, Zhang Q, Hu Y, Bu B, Guo J, Deng L. Mechanical communication-associated cell directional migration and branching connections mediated by calcium channels, integrin β1, and N-cadherin. Front Cell Dev Biol 2022; 10:942058. [PMID: 36051439 PMCID: PMC9424768 DOI: 10.3389/fcell.2022.942058] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/12/2022] [Accepted: 07/18/2022] [Indexed: 11/13/2022] Open
Abstract
Cell–cell mechanical communications at a large spatial scale (above hundreds of micrometers) have been increasingly recognized in recent decade, which shows importance in tissue-level assembly and morphodynamics. The involved mechanosensing mechanism and resulted physiological functions are still to be fully understood. Recent work showed that traction force sensation in the matrix induces cell communications for self-assembly. Here, based on the experimental model of cell directional migration on Matrigel hydrogel, containing 0.5 mg/ml type I collagen, we studied the mechano-responsive pathways for cell distant communications. Airway smooth muscle (ASM) cells assembled network structure on the hydrogel, whereas stayed isolated individually when cultured on glass without force transmission. Cell directional migration, or network assembly was significantly attenuated by inhibited actomyosin activity, or inhibition of inositol 1,4,5-trisphosphate receptor (IP3R) calcium channel or SERCA pump on endoplasmic reticulum (ER) membrane, or L-type calcium channel on the plasma membrane. Inhibition of integrin β1 with siRNA knockdown reduced cell directional migration and branching assembly, whereas inhibition of cell junctional N-cadherin with siRNA had little effect on distant attractions but blocked branching assembly. Our work demonstrated that the endoplasmic reticulum calcium channels and integrin are mechanosensing signals for cell mechanical communications regulated by actomyosin activity, while N-cadherin is responsible for traction force-induced cell stable connections in the assembly.
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Affiliation(s)
| | | | | | | | | | | | | | - Linhong Deng
- *Correspondence: Mingxing Ouyang, ; Linhong Deng,
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40
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Endothelial cell spreading on lipid bilayers with combined integrin and cadherin binding ligands. Bioorg Med Chem 2022; 68:116850. [PMID: 35714536 DOI: 10.1016/j.bmc.2022.116850] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2022] [Revised: 05/19/2022] [Accepted: 05/20/2022] [Indexed: 11/21/2022]
Abstract
Endothelial cells play a central role in the vascular system, where their function is tightly regulated by both cell-extracellular matrix (e.g., via integrins) and cell-cell interactions (e.g., via cadherins). In this study, we incorporated cholesterol-modified integrin and N-cadherin peptide binding ligands in fluid supported lipid bilayers. Human umbilical vein endothelial cell adhesion, spreading and vinculin localization in these cells were dependent on ligand density. One composition led to observe a higher extent of cell spreading, where cells exhibited extensive lamellipodia formation and a qualitatively more distinct N-cadherin localization at the cell periphery, which is indicative of N-cadherin clustering and a mimic of cell-cell contact formation. The results can be used to reconstitute the endothelial-pericyte interface on biomedical devices and materials.
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Vernaz ZJ, Lottero-Leconte RM, Alonso CAI, Rio S, Morales MF, Arroyo-Salvo C, Valiente CC, Lovaglio Diez M, Bogetti ME, Arenas G, Rey-Valzacchi G, Perez-Martinez S. Evaluation of sperm integrin α5β1 as a potential marker of fertility in humans. PLoS One 2022; 17:e0271729. [PMID: 35917320 PMCID: PMC9345343 DOI: 10.1371/journal.pone.0271729] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2021] [Accepted: 07/06/2022] [Indexed: 11/24/2022] Open
Abstract
Sperm selection for assisted reproduction techniques is generally based on basic parameters, while key aspects of sperm competence and its journey from the deposition site to the fertilization site are overlooked. Consequently, identifying molecular markers in spermatozoa that can efficiently predict the fertility of a semen sample could be of great interest, particularly in cases of idiopathic male infertility. When spermatozoa reach the female reproductive tract, it provides to them the cellular and molecular microenvironment needed to acquire fertilizing ability. In this sense, considering the role that integrin α5β1 of spermatozoa plays in reproduction-related events, we investigated the correlation between the subcellular localization of sperm integrin α5β1 and early embryo development outcome after in vitro fertilization (IVF) procedures in human. Twenty-four semen samples from normozoospermic men and metaphase II (MII) oocytes from healthy women aged under 38 years, from couples who underwent IVF cycles, were used in this work. Sperm α5β1 localization was evaluated by immunofluorescence assay using an antibody against integrin α5 subunit. Integrin α5β1 was mainly localized in the sperm acrosomal region (45.33±7.89%) or the equatorial segment (30.12±7.43%). The early embryo development rate (data obtained from the Fertility Center) correlated positively with the localization of α5β1 in the acrosomal region (number of usable embryos / inseminated oocytes: ρ = 0.75; p<0.01 and number of usable embryos/total number of two pronuclear zygotes: ρ = 0.80; p<0.01). However, this correlation was not significant when the equatorial segment mark was evaluated. In addition, human sperm released from co-culture with bovine oviductal epithelial cells (BOEC) showed a significant enrichment in the acrosomal localization pattern of α5β1 compared to those sperm that were not co-cultured with BOEC (85.20±5.35% vs 35.00±17.09%, respectively, p<0.05). In conclusion, the evaluation of sperm integrin α5β1 immunolocalization could be a useful tool to select sperm with fertilizing ability from human semen samples before IVF procedures.
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Affiliation(s)
- Zoilo José Vernaz
- Laboratorio de Biología de la Reproducción en Mamíferos, Centro de Estudios Farmacológicos y Botánicos (CEFYBO), Universidad de Buenos Aires/CONICET, Buenos Aires, Argentina
| | - Raquel María Lottero-Leconte
- Laboratorio de Biología de la Reproducción en Mamíferos, Centro de Estudios Farmacológicos y Botánicos (CEFYBO), Universidad de Buenos Aires/CONICET, Buenos Aires, Argentina
| | - Carlos Agustín Isidro Alonso
- Laboratorio de Biología de la Reproducción en Mamíferos, Centro de Estudios Farmacológicos y Botánicos (CEFYBO), Universidad de Buenos Aires/CONICET, Buenos Aires, Argentina
| | - Sofía Rio
- Laboratorio de Biología de la Reproducción en Mamíferos, Centro de Estudios Farmacológicos y Botánicos (CEFYBO), Universidad de Buenos Aires/CONICET, Buenos Aires, Argentina
| | | | - Camila Arroyo-Salvo
- Laboratorio de Biología de la Reproducción en Mamíferos, Centro de Estudios Farmacológicos y Botánicos (CEFYBO), Universidad de Buenos Aires/CONICET, Buenos Aires, Argentina
| | - Carla C. Valiente
- PROCREARTE- Red de Medicina Reproductiva y Molecular, Buenos Aires, Argentina
| | - María Lovaglio Diez
- PROCREARTE- Red de Medicina Reproductiva y Molecular, Buenos Aires, Argentina
| | - María Eugenia Bogetti
- Laboratorio de Biología de la Reproducción en Mamíferos, Centro de Estudios Farmacológicos y Botánicos (CEFYBO), Universidad de Buenos Aires/CONICET, Buenos Aires, Argentina
| | - Gabriela Arenas
- PROCREARTE- Red de Medicina Reproductiva y Molecular, Buenos Aires, Argentina
| | | | - Silvina Perez-Martinez
- Laboratorio de Biología de la Reproducción en Mamíferos, Centro de Estudios Farmacológicos y Botánicos (CEFYBO), Universidad de Buenos Aires/CONICET, Buenos Aires, Argentina
- * E-mail:
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Adherens junctions stimulate and spatially guide integrin activation and extracellular matrix deposition. Cell Rep 2022; 40:111091. [PMID: 35858563 DOI: 10.1016/j.celrep.2022.111091] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2021] [Revised: 05/04/2022] [Accepted: 06/22/2022] [Indexed: 11/03/2022] Open
Abstract
Cadherins and integrins are intrinsically linked through the actin cytoskeleton and are largely responsible for the mechanical integrity and organization of tissues. We show that cadherin clustering stimulates and spatially guides integrin activation. Adherens junction (AJ)-associated integrin activation depends on locally generated tension and does not require extracellular matrix ligands. It leads to the creation of primed integrin clusters, which spatially determine where focal adhesions will form if ligands are present and where ligands will be deposited. AJs that display integrin activation are targeted by microtubules facilitating their disassembly via caveolin-based endocytosis, showing that integrin activation impacts the stability of the core cadherin complex. Thus, the interplay between cadherins and integrins is more intimate than what was once believed and is rooted in the capacity of active integrins to be stabilized via AJ-generated tension. Altogether, our data establish a mechanism of cross-regulation between cadherins and integrins.
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Martinez Villegas K, Rasouli R, Tabrizian M. Enhancing metabolic activity and differentiation potential in adipose mesenchymal stem cells via high-resolution surface-acoustic-wave contactless patterning. MICROSYSTEMS & NANOENGINEERING 2022; 8:79. [PMID: 35846175 PMCID: PMC9276743 DOI: 10.1038/s41378-022-00415-w] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/16/2022] [Revised: 05/20/2022] [Accepted: 06/08/2022] [Indexed: 06/05/2023]
Abstract
Acoustofluidics has shown great potential for label-free bioparticle patterning with excellent biocompatibility. Acoustofluidic patterning enables the induction of cell-cell interactions, which play fundamental roles in organogenesis and tissue development. One of the current challenges in tissue engineering is not only the control of the spatial arrangement of cells but also the preservation of cell patterns over time. In this work, we developed a standing surface acoustic wave-based platform and demonstrated its capability for the well-controlled and rapid cell patterning of adipose-derived mesenchymal stem cells in a high-density homogenous collagen hydrogel. This biocompatible hydrogel is easily UV crosslinked and can be retrieved within 3 min. Acoustic waves successfully guided the cells toward pressure nodal lines, creating a contactless alignment of cells in <5 s in culture media and <1 min in the hydrogel. The acoustically patterned cells in the hydrogel did not show a decrease in cell viability (>90%) 48 h after acoustic induction. Moreover, 45.53% and 30.85% increases in metabolic activity were observed in growth and differentiation media, respectively, on Day 7. On Day 14, a 32.03% change in metabolic activity was observed using growth media, and no significant difference was observed using differentiation media. The alkaline phosphatase activity showed an increase of 80.89% and 24.90% on Days 7 and 14, respectively, for the acoustically patterned cells in the hydrogel. These results confirm the preservation of cellular viability and improved cellular functionality using the proposed high-resolution acoustic patterning technique and introduce unique opportunities for the application of stem cell regenerative patches for the emerging field of tissue engineering.
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Affiliation(s)
| | - Reza Rasouli
- Department of Biological and Biomedical Engineering, McGill University, Montreal, QC Canada
| | - Maryam Tabrizian
- Department of Biological and Biomedical Engineering, McGill University, Montreal, QC Canada
- Faculty of Dental Medicine and Oral Health Sciences, McGill University, Montreal, QC Canada
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44
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Islam S, Parker J, Dash BC, Hsia HC. Human iPSC-Vascular smooth muscle cell spheroids demonstrate size-dependent alterations in cellular viability and secretory function. J Biomed Mater Res A 2022; 110:1813-1823. [PMID: 35815599 DOI: 10.1002/jbm.a.37423] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2022] [Revised: 06/08/2022] [Accepted: 06/15/2022] [Indexed: 11/11/2022]
Abstract
Human-induced pluripotent stem cells (hiPSC) and their differentiated vascular cells have been revolutionizing the field of regenerative wound healing. These cells are shown to be rejuvenated with immense potentials in secreting paracrine factors. Recently, hiPSC-derived vascular smooth muscle cells (hiPSC-VSMC) have shown regenerative wound healing ability via their paracrine secretion. The quest to modulate the secretory function of these hiPSC-VSMC is an ongoing effort and involves the use of both biochemical and biophysical stimuli. This study explores the development and optimization of a reproducible, inexpensive protocol to form hiPSC-VSMC derived spheroids to investigate the implications of spheroid size on viability and paracrine secretion. Our data show the successful formation of different sizes of spheroids using various amount of hiPSC-VSMC. The hiPSC-VSMC spheroids formed with 10,000 cells strike an ideal balance between overall cell health and maximal paracrine secretion. The conditioned medium from these spheroids was found to be bioactive in enhancing human dermal fibroblast cell proliferation and migration. This research will inform future studies on the optimal spheroid size for regenerative wound healing applications.
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Affiliation(s)
- Sara Islam
- Section of Plastic Surgery, Department of Surgery, Yale School of Medicine, Yale University, New Haven, Connecticut, USA
| | - Jackson Parker
- Section of Plastic Surgery, Department of Surgery, Yale School of Medicine, Yale University, New Haven, Connecticut, USA
| | - Biraja C Dash
- Section of Plastic Surgery, Department of Surgery, Yale School of Medicine, Yale University, New Haven, Connecticut, USA
| | - Henry C Hsia
- Section of Plastic Surgery, Department of Surgery, Yale School of Medicine, Yale University, New Haven, Connecticut, USA.,Department of Biomedical Engineering, Yale University, New Haven, Connecticut, USA
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45
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Fu C, Wang J, Pallikkuth S, Ding Y, Chen J, Wren JD, Yang Y, Wong KK, Kameyama H, Jayaraman M, Munshi A, Tanaka T, Lidke KA, Zhang XA. EWI2 prevents EGFR from clustering and endocytosis to reduce tumor cell movement and proliferation. Cell Mol Life Sci 2022; 79:389. [PMID: 35773608 PMCID: PMC10428948 DOI: 10.1007/s00018-022-04417-9] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2022] [Revised: 05/27/2022] [Accepted: 06/06/2022] [Indexed: 12/01/2022]
Abstract
EWI2 is a transmembrane immunoglobulin superfamily (IgSF) protein that physically associates with tetraspanins and integrins. It inhibits cancer cells by influencing the interactions among membrane molecules including the tetraspanins and integrins. The present study revealed that, upon EWI2 silencing or ablation, the elevated movement and proliferation of cancer cells in vitro and increased cancer metastatic potential and malignancy in vivo are associated with (i) increases in clustering, endocytosis, and then activation of EGFR and (ii) enhancement of Erk MAP kinase signaling. These changes in signaling make cancer cells (i) undergo partial epithelial-to-mesenchymal (EMT) for more tumor progression and (ii) proliferate faster for better tumor formation. Inhibition of EGFR or Erk kinase can abrogate the cancer cell phenotypes resulting from EWI2 removal. Thus, to inhibit cancer cells, EWI2 prevents EGFR from clustering and endocytosis to restrain its activation and signaling.
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Affiliation(s)
- Chenying Fu
- University of Oklahoma Health Sciences Center, Oklahoma City, USA
| | - Jie Wang
- University of Oklahoma Health Sciences Center, Oklahoma City, USA
| | | | - Yingjun Ding
- University of Oklahoma Health Sciences Center, Oklahoma City, USA
| | - Junxiong Chen
- University of Oklahoma Health Sciences Center, Oklahoma City, USA
| | | | - Yuchao Yang
- University of Oklahoma Health Sciences Center, Oklahoma City, USA
| | | | | | | | - Anupama Munshi
- University of Oklahoma Health Sciences Center, Oklahoma City, USA
| | - Takemi Tanaka
- University of Oklahoma Health Sciences Center, Oklahoma City, USA
| | | | - Xin A Zhang
- University of Oklahoma Health Sciences Center, Oklahoma City, USA.
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Condensation of the Drosophila nerve cord is oscillatory and depends on coordinated mechanical interactions. Dev Cell 2022; 57:867-882.e5. [PMID: 35413236 DOI: 10.1016/j.devcel.2022.03.007] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2021] [Revised: 01/19/2022] [Accepted: 03/14/2022] [Indexed: 11/21/2022]
Abstract
During development, organs reach precise shapes and sizes. Organ morphology is not always obtained through growth; a classic counterexample is the condensation of the nervous system during Drosophila embryogenesis. The mechanics underlying such condensation remain poorly understood. Here, we characterize the condensation of the embryonic ventral nerve cord (VNC) at both subcellular and tissue scales. This analysis reveals that condensation is not a unidirectional continuous process but instead occurs through oscillatory contractions. The VNC mechanical properties spatially and temporally vary, and forces along its longitudinal axis are spatially heterogeneous. We demonstrate that the process of VNC condensation is dependent on the coordinated mechanical activities of neurons and glia. These outcomes are consistent with a viscoelastic model of condensation, which incorporates time delays and effective frictional interactions. In summary, we have defined the progressive mechanics driving VNC condensation, providing insights into how a highly viscous tissue can autonomously change shape and size.
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Zou H, Yang Z, Chan YS, Yeung SKA, Alam MK, Si T, Xu T, Yang M. Single cell analysis of mechanical properties and EMT-related gene expression profiles in cancer fingers. iScience 2022; 25:103917. [PMID: 35252814 PMCID: PMC8889141 DOI: 10.1016/j.isci.2022.103917] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2021] [Revised: 01/07/2022] [Accepted: 02/07/2022] [Indexed: 02/07/2023] Open
Abstract
Collective cell migration is associated with cancer metastasis. Cancer fingers are formed when groups of migrating cancer cells follow the leader cells in the front. Epithelial to mesenchymal transition (EMT) is a critical process of cancer metastasis. However, the role of EMT in cancer finger formation remains unclear. In this work, we investigated the EMT-associated mechanical properties and gene expression at single-cell levels in non-small lung cancer fingers. We found that leader cells were more elastic and less sticky than follower cells. Spatial EMT-related gene expression profiling in cancer fingers revealed cellular heterogeneity. Particularly, SNAIL and VIM were found to be two key genes that positively correlated with leader cell phenotypes and controlled cancer finger formation. Silencing either SNAIL or VIM, decreased cancer cell elasticity, cancer finger formation and migration, and increased adhesiveness. These findings indicated that SNAIL and VIM are two driver genes for cancer finger formation.
Spatial mapping of EMT genes and mechanical properties of cancer finger at single cell level Cancer cell elasticity and adhesiveness are two physical biomarkers for leader cells SNAIL and VIM drive finger cell formation and are potential targets for therapy
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Barhouse PS, Andrade MJ, Smith Q. Home Away From Home: Bioengineering Advancements to Mimic the Developmental and Adult Stem Cell Niche. FRONTIERS IN CHEMICAL ENGINEERING 2022. [DOI: 10.3389/fceng.2022.832754] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
Abstract
The inherent self-organizing capacity of pluripotent and adult stem cell populations has advanced our fundamental understanding of processes that drive human development, homeostasis, regeneration, and disease progression. Translating these principles into in vitro model systems has been achieved with the advent of organoid technology, driving innovation to harness patient-specific, cell-laden regenerative constructs that can be engineered to augment or replace diseased tissue. While developmental organization and regenerative adult stem cell niches are tightly regulated in vivo, in vitro analogs lack defined architecture and presentation of physicochemical cues, leading to the unhindered arrangement of mini-tissues that lack complete physiological mimicry. This review aims to highlight the recent integrative engineering approaches that elicit spatio-temporal control of the extracellular niche to direct the structural and functional maturation of pluripotent and adult stem cell derivatives. While the advances presented here leverage multi-pronged strategies ranging from synthetic biology to microfabrication technologies, the methods converge on recreating the biochemical and biophysical milieu of the native tissue to be modeled or regenerated.
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Integrin β1 orchestrates the abnormal cell-matrix attachment and invasive behaviour of E-cadherin dysfunctional cells. Gastric Cancer 2022; 25:124-137. [PMID: 34486077 PMCID: PMC8732838 DOI: 10.1007/s10120-021-01239-9] [Citation(s) in RCA: 13] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/03/2021] [Accepted: 08/19/2021] [Indexed: 02/07/2023]
Abstract
BACKGROUND Tumour progression relies on the ability of cancer cells to penetrate and invade neighbouring tissues. E-cadherin loss is associated with increased cell invasion in gastric carcinoma, and germline mutations of the E-cadherin gene are causative of hereditary diffuse gastric cancer. Although E-cadherin dysfunction impacts cell-cell adhesion, cell dissemination also requires an imbalance of adhesion to the extracellular matrix (ECM). METHODS To identify ECM components and receptors relevant for adhesion of E-cadherin dysfunctional cells, we implemented a novel ECM microarray platform coupled with molecular interaction networks. The functional role of putative candidates was determined by combining micropattern traction microscopy, protein modulation and in vivo approaches, as well as transcriptomic data of 262 gastric carcinoma samples, retrieved from the cancer genome atlas (TCGA). RESULTS Here, we show that E-cadherin mutations induce an abnormal interplay of cells with specific components of the ECM, which encompasses increased traction forces and Integrin β1 activation. Integrin β1 synergizes with E-cadherin dysfunction, promoting cell scattering and invasion. The significance of the E-cadherin-Integrin β1 crosstalk was validated in Drosophila models and found to be consistent with evidence from human gastric carcinomas, where increased tumour grade and poor survival are associated with low E-cadherin and high Integrin β1 levels. CONCLUSIONS Integrin β1 is a key mediator of invasion in carcinomas with E-cadherin impairment and should be regarded as a biomarker of poor prognosis in gastric cancer.
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Swamy H, Glading AJ. Contribution of protein-protein interactions to the endothelial barrier-stabilizing function of KRIT1. J Cell Sci 2021; 135:274104. [PMID: 34918736 PMCID: PMC8917353 DOI: 10.1242/jcs.258816] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2021] [Accepted: 12/08/2021] [Indexed: 11/20/2022] Open
Abstract
Krev-interaction trapped 1 (KRIT1) is an endothelial scaffold protein that promotes adherens junction (AJ) stability. The precise mechanism by which KRIT1 promotes barrier stabilization is unclear. We tested the ability of a panel of KRIT1 constructs containing mutations that inhibit Rap1 binding, ICAP1 binding, disrupt KRIT1's protein tyrosine binding domain (PTB), or direct KRIT1 to the plasma membrane, either alone or in combination, to restore barrier function in KRIT1-deficient endothelial cells. We found that ablating the 192NPAY195 motif or disrupting the PTB domain was sufficient to restore AJ protein localization and barrier function to control levels, irrespective of the junctional localization of KRIT1 or Rap1 binding. The ability of our KRIT1 constructs to rescue AJ/barrier function in KRIT1 depleted endothelial cells correlated with decreased 1 integrin activity and maintenance of cortical actin fibers. Together, our findings indicate that Rap1 binding, ICAP1 binding, and junctional localization are not required for the ability of KRIT1 to stabilize endothelial contacts, and suggest that the ability of KRIT1 to limit integrin activity may be involved in barrier stabilization.
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Affiliation(s)
- Harsha Swamy
- Department of Pharmacology and Physiology, University of Rochester, Rochester, NY, USA
| | - Angela J Glading
- Department of Pharmacology and Physiology, University of Rochester, Rochester, NY, USA
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