Biology subjects

Pabijan, J.

Publications and source records attributed to Pabijan, J..

6 recordsLinked to original sources

bioRxiv2024

Tubulin-Targeted Therapy in Melanoma Increases Cell Invasive Potential by Activation of Actomyosin Cytoskeleton - an in vitro study

One of the most dangerous aspects of cancers is their ability to metastasize, which is the leading cause of death. Hence, it holds significance to develop therapies targeting the eradication of cancer cells, in parallel, inhibiting metastases in cells surviving the applied therapy. Here, we focused on two melanoma cell lines - WM35 and WM266-4 - representing the less and more invasive melanomas. We investigated mechanisms of cellular processes regulating the activation of actomyosin being the effect of colchicine treatment. Additionally, we investigated biophysical aspects of supplement therapy using Rho-associated protein kinase (ROCK) inhibitor (Y-27632), and myosin II inhibitor ((-)-blebbistatin), focusing on the microtubules and actin filaments. We analyzed their effect on the proliferation, migration, and invasiveness of melanoma cells, supported by studies on cytoskeletal architecture using confocal fluorescence microscopy and nanomechanics using atomic force microscopy and novel method based on constriction channels. Our results showed that colchicine inhibits the migration of most melanoma cells, while for a small cell population, it paradoxically increases their migration and invasiveness. These changes are also accompanied by the formation of stress fibers compensating for the loss of microtubules. Simultaneous administration of selected agents led to the inhibition of this compensatory effect. Collectively, our results highlighted that colchicine led to actomyosin activation and increased cancer cell invasiveness. We emphasized a cellular pathway of Rho-ROCK-dependent actomyosin contraction to be responsible for the increased invasive potential of melanoma cells in tubulin-targeted therapy. HighlightsO_LIColchicine strongly increased (trans)migration in the subpopulation of melanoma cells. C_LIO_LIColchicine disrupted tubulin but compensated for the activation of the actomyosin network. C_LIO_LIRho-ROCK-targeted or myosin-targeted therapies vanished the side-effect of colchicine. C_LIO_LIIncreased actin polymerisation correlated with the transit time in the constriction channel. C_LIO_LIElastic modulus variations reflected observed alterations in the actomyosin network. C_LI

biophysics↗

bioRxiv2024

Migration, proliferation, and elasticity of bladder cancer cells on lectin-coated surfaces

The alterations in migration, proliferation, and mechanics of cells observed during cancer progression can potentially be linked to enhanced tumor invasiveness. These properties are frequently attributed to the ability to form distant metastasis; however, the direct mutual connection between these properties is not always proven. Here, we studied the migratory, proliferative, and mechanical phenotype of three bladder cancer cells originating from various stages of cancer progression, i.e., non-malignant cell cancer of ureter (HCV29 cells), bladder carcinoma (HT1376 cells) and transitional bladder carcinoma (T24 cells). The results were linked with the organization of actin filaments because of their major role in cell migration. The results classified cells into non-malignant, non-invasive, and invasive, revealing the significant impact of actin filaments in bladder cancer invasion. Based on the reported changes in cancer cell glycosylation, the potential applicability of the observed cancer-related changes to identify invasive cells was demonstrated for the lectin-coated surfaces, which is the potential surface modification for biosensors.

biophysics↗

bioRxiv2023

Multiscale Rheology of Aging Cancer Spheroids

Cancer spheroids offer a valuable experimental model that mimics the complexity and heterogeneity of solid tumors. Characterizing their mechanical response is crucial for understanding tumor development, progression, and drug response. Currently, whole live spheroids are analyzed primarily using image analysis, which is challenging, requires extended incubation times, and has limited imaging depth. Here, we present a new label-free approach for characterizing sub-superficial structures of bladder cancer spheroids and measuring their mechanical response at three distinct stages of cancer progression. We study the microrheological changes induced by aging at the cellular and cluster levels by conducting a multi-physics characterization and modeling approach. We find that spheroids exhibit viscoelastic behavior that can be described by fractional models. We show that spheroids are mechanically heterogeneous, with strong depth and time-dependent variations associated with evolving structural features. Our approach opens new possibilities to study 3D in vitro models, paving the way for the discovery of novel and more precise procedure in cancer diagnosis based on the use of mechanomarkers.

biophysics↗

bioRxiv2023

Reliable, standardized measurements for cell mechanical properties

Atomic force microscopy (AFM) has become indispensable for studying biological and medical samples. More than two decades of experiments have revealed that cancer cells are softer than healthy cells (for measured cells cultured on stiff substrates). The softness or, more precisely, the larger deformability of cancer cells, primarily independent of cancer types, could be used as a sensitive marker of pathological changes. The wide application of biomechanics in clinics would require designing instruments with specific calibration, data collection, and analysis procedures. For these reasons, such development is, at present, still very limited, hampering the clinical exploitation of mechanical measurements. Here, we propose a standardized operational protocol (SOP), developed within the EU ITN network Phys2BioMed, which allows the detection of the biomechanical properties of living cancer cells regardless of the nanoindentation instruments used (AFMs and other indenters) and the laboratory involved in the research. We standardized the cell cultures, AFM calibration, measurements, and data analysis. This effort resulted in a step-by-step SOP for cell cultures, instrument calibration, measurements, and data analysis, leading to the concordance of the results (Youngs modulus) measured among the six EU laboratories involved. Our results highlight the importance of the SOP in obtaining a reproducible mechanical characterization of cancer cells and paving the way toward exploiting biomechanics for diagnostic purposes in clinics.

biophysics↗

bioRxiv2022

Nanomechanical and microrheological properties of bladder cancer cells at cellular and spheroid levels

Cancer progression is associated with changes in cell mechanical and rheological properties that could be probed by atomic force microscopy (AFM). In this study, we applied AFM to measure elastic (by compressing the cells) and viscoelastic (by applying shear stress) properties of bladder cancers in relation to their culture morphology (in single cells, cell monolayers, and spheroids). Three different cell lines, HCV29 (non-malignant cell cancer of ureter), HT1376 (grade III bladder carcinoma), T24 (grade IV transitional cell carcinoma), were investigated. Nanoindentation measurements only differentiate between non-malignant and cancer cells, but it is difficult to distinguish between specific bladder cancers. By applying microrheological measurements, we confirm that non-malignant cells are more rigid than cancer cells but more importantly, it was possible to differentiate between two cancerous cell lines, regardless of the culture conditions. As each of them is characterized by a distinct actin filament network inside the cell, we showed that actin filaments are a key element in defining the rheological properties of spheroids originating from cells having thick actin bundles. Our results showed that HCV29 cells are more rigid than the studied cancer cells, indicating that normal cells are resistant to compressive and shear forces. Therefore, we conclude that cell mechanical and rheological properties may serve as a biophysical marker to distinguish normal and cancer cells of different malignancies.

biophysics↗

bioRxiv2022

Changes in nanomechanical properties of single neuroblastoma cells as a model for oxygen and glucose deprivation (OGD)

The biological processes underlying ischemic stroke, although complex, are better known than those related to biomechanical alterations of single cells. Mechanisms of biomechanical changes and their relations to the molecular processes are crucial for understanding the function and dysfunction of the brain. In our study, we applied atomic force microscopy (AFM) to quantify the alterations in biomechanical properties in neuroblastoma SH-SY5Y cells subjected to oxygen and glucose deprivation (OGD) and reoxygenation (RO). Obtained results reveal several characteristics. Cell viability remained at the same level, regardless of the OGD and RO conditions, but, in parallel, the metabolic activity of cells decreased with OGD duration. 24h RO did not recover the metabolic activity fully. Cells subjected to OGD appeared softer than control cells. Cell softening was strongly present in cells after 1h of OGD and, with longer OGD duration and in RO conditions, cells recovered their mechanical properties. Changes in the nanomechanical properties of cells were attributed to the remodelling of actin filaments, which was related to cofilin-based regulation and impaired metabolic activity of cells. The presented study shows the importance of nanomechanics in research on ischemic-related pathological processes such as stroke.

biophysics↗