scholarly journals Biomechanical and biochemical assessment of YB-1 expression in melanoma cells

2021 ◽  
Author(s):  
Anna Magdalena Cykowska ◽  
Ulf Krister Hofmann ◽  
Aadhya Tiwari ◽  
Corinna Kosnopfel ◽  
Rosa Riester ◽  
...  
Keyword(s):  
Epithelial To Mesenchymal Transition ◽  
Proteome Analysis ◽  
Melanoma Cells ◽  
Cell Stiffness ◽  
Knock Out ◽  
Mesenchymal Transition ◽  
Force Microscopy ◽  
Atomic Force ◽  
Rearrangement Process ◽  
Actomyosin Cytoskeleton

Malignant melanoma is the most lethal form of skin cancer; its incidence has increased over the last five decades. Y-box binding protein 1 (YB-1) plays a prominent role in mediating metastatic behavior by promoting epithelial-to-mesenchymal transition (EMT) processes. Migratory melanoma cells exhibit two major phenotypes: elongated mesenchymal or rounded amoeboid. The actomyosin cytoskeleton is key in both phenotypes, but intermediate filaments also undergo a significant rearrangement process, switching from cytokeratin-rich to vimentin and nestin-rich network. In this study, we aimed to investigate to what extent YB-1 impacts the biomechanical (cell stiffness) and biochemical aspects of melanoma cells and their cytoskeleton. To this end, we subjected A375 YB-1 knock-out and parental cells to atomic force microscopy investigations (stiffness determination), immunolabelling, and proteome analysis. We found that YB-1 expressing cells were significantly stiffer compared to the corresponding YB-1 knock-out cell line. Proteomic analysis revealed that expression of YB-1 results in a strong co-expression of nestin, vimentin, fascin-1, and septin-9. In the YB-1 knock-out nestin was completely depleted, but zyxin was strongly upregulated. Collectively, our results showed that YB-1 knock-out acquires some characteristics of mesenchymal phenotype but lacks important markers of malignancy and invasiveness such as nestin or vimentin. We posit that there is an association of YB-1 expression with an amoeboid phenotype, which would explain the increased migratory capacity.

scholarly journals Bornyl cis-4-Hydroxycinnamate Suppresses Cell Metastasis of Melanoma through FAK/PI3K/Akt/mTOR and MAPK Signaling Pathways and Inhibition of the Epithelial-to-Mesenchymal Transition

2018 ◽  
Vol 19 (8) ◽  
pp. 2152 ◽  
Author(s):  
Tzu-Yen Yang ◽  
Mei-Li Wu ◽  
Chi-I Chang ◽  
Chih-I Liu ◽  
Te-Chih Cheng ◽  
...  
Keyword(s):  
Cell Migration ◽  
Signaling Pathways ◽  
Melanoma Cell ◽  
Epithelial To Mesenchymal Transition ◽  
Melanoma Cells ◽  
Migration And Invasion ◽  
Mesenchymal Transition ◽  
Cell Migration And Invasion ◽  
Cell Metastasis ◽  
A375 Melanoma

Bornyl cis-4-hydroxycinnamate, a bioactive compound isolated from Piper betle stems, has the potential for use as an anti-cancer agent. This study investigated the effects of bornyl cis-4-hydroxycinnamate on cell migration and invasion in melanoma cells. Cell migration and invasion were compared in A2058 and A375 melanoma cell lines treated with/without bornyl cis-4-hydroxycinnamate (1–6 µM). To examine whether bornyl cis-4-hydroxycinnamate has a potential anti-metastatic effect on melanoma cells, cell migration and invasion assays were performed using a Boyden chamber assay and a transwell chamber in A2058 and A375 cells. Gelatin zymography was employed to determine the enzyme activities of MMP-2 and MMP-9. Cell lysates were collected for Western blotting analysis of matrix metalloproteinase (MMP)-2, MMP-9 and tissue inhibitors of metalloproteinase-1/2 (TIMP-1/2), as well as key molecules in the mitogen-activated protein kinase (MAPK), focal adhesion kinase (FAK)/ phosphatidylinositide-3 kinases (PI3K)/Akt/ mammalian target of rapamycin (mTOR), growth factor receptor-bound protein 2 (GRB2) signaling pathways. Our results demonstrated that bornyl cis-4-hydroxycinnamate is a potentially useful agent that inhibits melanoma cell migration and invasion, and altered melanoma cell metastasis by reducing MMP-2 and MMP-9 expression through inhibition of the FAK/PI3K/Akt/mTOR, MAPK, and GRB2 signaling pathways. Moreover, bornyl cis-4-hydroxycinnamate inhibited the process of the epithelial-to-mesenchymal transition in A2058 and A375 melanoma cells. These findings suggested that bornyl cis-4-hydroxycinnamate has potential as a chemotherapeutic agent, and warrants further investigation for its use in the management of human melanoma.

scholarly journals Nanomechanical properties of enucleated cells: contribution of the nucleus to the passive cell mechanics

2020 ◽  
Vol 18 (1) ◽  
Author(s):  
Yuri M. Efremov ◽  
Svetlana L. Kotova ◽  
Anastasia A. Akovantseva ◽  
Peter S. Timashev
Keyword(s):  
Mechanical Properties ◽  
Cell Mechanics ◽  
Normal Cells ◽  
Force Microscopy ◽  
Nanomechanical Properties ◽  
Atomic Force ◽  
Fibrosarcoma Cells ◽  
Actomyosin Cytoskeleton ◽  
Small Deformations

Abstract Background The nucleus, besides its functions in the gene maintenance and regulation, plays a significant role in the cell mechanosensitivity and mechanotransduction. It is the largest cellular organelle that is often considered as the stiffest cell part as well. Interestingly, the previous studies have revealed that the nucleus might be dispensable for some of the cell properties, like polarization and 1D and 2D migration. Here, we studied how the nanomechanical properties of cells, as measured using nanomechanical mapping by atomic force microscopy (AFM), were affected by the removal of the nucleus. Methods The mass enucleation procedure was employed to obtain cytoplasts (enucleated cells) and nucleoplasts (nuclei surrounded by plasma membrane) of two cell lines, REF52 fibroblasts and HT1080 fibrosarcoma cells. High-resolution viscoelastic mapping by AFM was performed to compare the mechanical properties of normal cells, cytoplasts, and nucleoplast. The absence or presence of the nucleus was confirmed with fluorescence microscopy, and the actin cytoskeleton structure was assessed with confocal microscopy. Results Surprisingly, we did not find the softening of cytoplasts relative to normal cells, and even some degree of stiffening was discovered. Nucleoplasts, as well as the nuclei isolated from cells using a detergent, were substantially softer than both the cytoplasts and normal cells. Conclusions The cell can maintain its mechanical properties without the nucleus. Together, the obtained data indicate the dominating role of the actomyosin cytoskeleton over the nucleus in the cell mechanics at small deformations inflicted by AFM.

Betulinic acid suppresses NGAL-induced epithelial-to-mesenchymal transition in melanoma

2013 ◽  
Vol 394 (6) ◽  
pp. 773-781 ◽  
Author(s):  
Dorina Gheorgheosu ◽  
Michaela Jung ◽  
Bilge Ören ◽  
Tobias Schmid ◽  
Cristina Dehelean ◽  
...  
Keyword(s):  
Betulinic Acid ◽  
Cancer Metastasis ◽  
Epithelial To Mesenchymal Transition ◽  
Melanoma Cells ◽  
Antitumoral Activity ◽  
Cellular Phenotype ◽  
Mesenchymal Transition ◽  
Neutrophil Gelatinase Associated Lipocalin ◽  
Neutrophil Gelatinase ◽  
The Impact

Abstract Betulinic acid (BA) exhibits antitumoral activity by blocking proliferation, invasion, and angiogenesis. However, the impact of BA on epithelial-to-mesenchymal transition (EMT), a hallmark of cancer metastasis induced among others by neutrophil gelatinase-associated lipocalin (NGAL), remains unknown. The present study aimed at determining the effect of BA on NGAL-induced EMT. In A375 melanoma cells, BA downregulated mesenchymal markers, increased epithelial markers, and inhibited cytoskeletal reorganization. In addition, BA limited endogenous NGAL production and further suppressed EMT induced by exogenously added NGAL and the corresponding invasive cellular phenotype. In conclusion, BA interferes with EMT-associated changes, a mechanism to antagonize invasive melanoma cells.

Role for stress fiber contraction in surface tension development and stretch-activated channel regulation in C2C12 myoblasts

2008 ◽  
Vol 295 (1) ◽  
pp. C160-C172 ◽  
Author(s):  
Francesca Sbrana ◽  
Chiara Sassoli ◽  
Elisabetta Meacci ◽  
Daniele Nosi ◽  
Roberta Squecco ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Myosin Ii ◽  
Cell Stiffness ◽  
C2c12 Myoblasts ◽  
Tension Development ◽  
Force Microscopy ◽  
Atomic Force ◽  
Sphingosine 1 Phosphate ◽  
Membrane Stretching

Membrane-cytoskeleton interaction regulates transmembrane currents through stretch-activated channels (SACs); however, the mechanisms involved have not been tested in living cells. We combined atomic force microscopy, confocal immunofluorescence, and patch-clamp analysis to show that stress fibers (SFs) in C2C12 myoblasts behave as cables that, tensed by myosin II motor, activate SACs by modifying the topography and the viscoelastic (Young's modulus and hysteresis) and electrical passive (membrane capacitance, Cm) properties of the cell surface. Stimulation with sphingosine 1-phosphate to elicit SF formation, the inhibition of Rho-dependent SF formation by Y-27632 and of myosin II-driven SF contraction by blebbistatin, showed that not SF polymerization alone but the generation of tensional forces by SF contraction were involved in the stiffness response of the cell surface. Notably, this event was associated with a significant reduction in the amplitude of the cytoskeleton-mediated corrugations in the cell surface topography, suggesting a contribution of SF contraction to plasma membrane stretching. Moreover, Cm, used as an index of cell surface area, showed a linear inverse relationship with cell stiffness, indicating participation of the actin cytoskeleton in plasma membrane remodeling and the ability of SF formation to cause internalization of plasma membrane patches to reduce Cm and increase membrane tension. SF contraction also increased hysteresis. Together, these data provide the first experimental evidence for a crucial role of SF contraction in SAC activation. The related changes in cell viscosity may prevent SAC from abnormal activation.

scholarly journals Chemotherapy exposure increases leukemia cell stiffness

Blood ◽  
2006 ◽  
Vol 109 (8) ◽  
pp. 3505-3508 ◽  
Author(s):  
Wilbur A. Lam ◽  
Michael J. Rosenbluth ◽  
Daniel A. Fletcher
Keyword(s):  
Cell Death ◽  
Leukemia Cell ◽  
Vascular Complications ◽  
Cell Types ◽  
Cell Stiffness ◽  
Microfluidic Channels ◽  
Cell Deformability ◽  
Force Microscopy ◽  
Atomic Force ◽  
Acute Myeloid Leukemia Cells

Abstract Deformability of blood cells is known to influence vascular flow and contribute to vascular complications. Medications for hematologic diseases have the potential to modulate these complications if they alter blood cell deformability. Here we report the effect of chemotherapy on leukemia cell mechanical properties. Acute lymphoblastic and acute myeloid leukemia cells were incubated with standard induction chemotherapy, and individual cell stiffness was tracked with atomic force microscopy. When exposed to dexamethasone or daunorubicin, leukemia cell stiffness increased by nearly 2 orders of magnitude, which decreased their passage through microfluidic channels. This stiffness increase occurred before caspase activation and peaked after completion of cell death, and the rate of stiffness increase depended on chemotherapy type. Stiffening with cell death occurred for all cell types investigated and may be due to dynamic changes in the actin cytoskeleton. These observations suggest that chemotherapy itself may increase the risk of vascular complications in acute leukemia.

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