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Dynamic cellular biomechanics in responses to chemotherapeutic drug in hypoxia probed by atomic force spectroscopy.

ABSTRACT: The changes in cellular structure play an important role in cancer cell development, progression, and metastasis. By exploiting single-cell, force spectroscopy methods, we probed biophysical and biomechanical kinetics (stiffness, morphology, roughness, adhesion) of brain, breast, prostate, and pancreatic cancer cells with standard chemotherapeutic drugs in normoxia and hypoxia over 12-24 hours. After exposure to the drugs, we found that brain, breast, and pancreatic cancer cells became approximately 55-75% less stiff, while prostate cancer cells became more stiff, due to either drug-induced disruption or reinforcement of cytoskeletal structure. However, the rate of the stiffness change decreased up to 2-folds in hypoxia, suggesting a correlation between cellular stiffness and drug resistance of cancer cells in hypoxic tumor microenvironment. Also, we observed significant changes in the cell body height, surface roughness, and cytoadhesion of cancer cells after exposure to drugs, which followed the trend of stiffness. Our results show that a degree of chemotherapeutic drug effects on biomechanical and biophysical properties of cancer cells is distinguishable in normoxia and hypoxia, which are correlated with alteration of cytoskeletal structure and integrity during drug-induced apoptotic process.

SUBMITTER: Alhalhooly L

PROVIDER: S-EPMC8202777 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Cell Adhesion on Dynamic Supramolecular Surfaces Probed by Fluid Force Microscopy-Based Single-Cell Force Spectroscopy.

Project description:Biomimetic and stimuli-responsive cell-material interfaces are actively being developed to study and control various cell-dynamics phenomena. Since cells naturally reside in the highly dynamic and complex environment of the extracellular matrix, attempts are being made to replicate these conditions in synthetic biomaterials. Supramolecular chemistry, dealing with noncovalent interactions, has recently provided possibilities to incorporate such dynamicity and responsiveness in various types of architectures. Using a cucurbit[8]uril-based host-guest system, we have successfully established a dynamic and electrochemically responsive interface for the display of the integrin-specific ligand, Arg-Gly-Asp (RGD), to promote cell adhesion. Due to the weak nature of the noncovalent forces by which the components at the interface are held together, we expected that cell adhesion would also be weaker in comparison to traditional interfaces where ligands are usually immobilized by covalent linkages. To assess the stability and limitations of our noncovalent interfaces, we performed single-cell force spectroscopy studies using fluid force microscopy. This technique enabled us to measure rupture forces of multiple cells that were allowed to adhere for several hours on individual substrates. We found that the rupture forces of cells adhered to both the noncovalent and covalent interfaces were nearly identical for up to several hours. We have analyzed and elucidated the reasons behind this result as a combination of factors including the weak rupture force between linear Arg-Gly-Asp and integrin, high surface density of the ligand, and increase in effective concentration of the supramolecular components under spread cells. These characteristics enable the construction of highly dynamic biointerfaces without compromising cell-adhesive properties.

| S-EPMC5406783 | biostudies-literature

Mechanics of spreading cells probed by atomic force microscopy.

Project description:Cellular adhesion and motility are fundamental processes in biological systems such as morphogenesis and tissue homeostasis. During these processes, cells heavily rely on the ability to deform and supply plasma membrane from pre-existing membrane reservoirs, allowing the cell to cope with substantial morphological changes. While morphological changes during single cell adhesion and spreading are well characterized, the accompanying alterations in cellular mechanics are scarcely addressed. Using the atomic force microscope, we measured changes in cortical and plasma membrane mechanics during the transition from early adhesion to a fully spread cell. During the initial adhesion step, we found that tremendous changes occur in cortical and membrane tension as well as in membrane area. Monitoring the spreading progress by means of force measurements over 2.5 h reveals that cortical and membrane tension become constant at the expense of excess membrane area. This was confirmed by fluorescence microscopy, which shows a rougher plasma membrane of cells in suspension compared with spread ones, allowing the cell to draw excess membrane from reservoirs such as invaginations or protrusions while attaching to the substrate and forming a first contact zone. Concretely, we found that cell spreading is initiated by a transient drop in tension, which is compensated by a decrease in excess area. Finally, all mechanical parameters become almost constant although morphological changes continue. Our study shows how a single cell responds to alterations in membrane tension by adjusting its overall membrane area. Interference with cytoskeletal integrity, membrane tension and excess surface area by administration of corresponding small molecular inhibitors leads to perturbations of the spreading process.

| S-EPMC3728925 | biostudies-literature

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| S-EPMC3637277 | biostudies-other

Subnanometre enzyme mechanics probed by single-molecule force spectroscopy.

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| S-EPMC4770092 | biostudies-literature

Single-molecule atomic force microscopy force spectroscopy study of A?-40 interactions.

Project description:Misfolding and aggregation of amyloid ?-40 (A?-40) peptide play key roles in the development of Alzheimer's disease (AD). However, very little is known about the molecular mechanisms underlying these molecular processes. We developed a novel experimental approach that can directly probe aggregation-prone states of proteins and their interactions. In this approach, the proteins are anchored to the surface of the atomic force microscopy substrate (mica) and the probe, and the interaction between anchored molecules is measured in the approach-retraction cycles. We used dynamic force spectroscopy (DFS) to measure the stability of transiently formed dimers. One of the major findings from DFS analysis of ?-synuclein (?-Syn) is that dimeric complexes formed by misfolded ?-Syn protein are very stable and dissociate over a range of seconds. This differs markedly from the dynamics of monomers, which occurs on a microsecond to nanosecond time scale. Here we applied the same approach to quantitatively characterize interactions of A?-40 peptides over a broad range of pH values. These studies showed that misfolded dimers are characterized by lifetimes in the range of seconds. This value depends on pH and varies between 2.7 s for pH 2.7 and 0.1 s for pH 7, indicating that the aggregation properties of A?-40 are modulated by the environmental conditions. The analysis of the contour lengths revealed the existence of various pathways for dimer dissociation, suggesting that dimers with different conformations are formed. These structural variations result in different aggregation pathways, leading to different types of oligomers and higher-order aggregates, including fibrils.

| S-EPMC3113552 | biostudies-literature

Minimizing pulling geometry errors in atomic force microscope single molecule force spectroscopy.

Project description:In atomic force microscopy-based single molecule force spectroscopy (AFM-SMFS), it is assumed that the pulling angle is negligible and that the force applied to the molecule is equivalent to the force measured by the instrument. Recent studies, however, have indicated that the pulling geometry errors can drastically alter the measured force-extension relationship of molecules. Here we describe a software-based alignment method that repositions the cantilever such that it is located directly above the molecule's substrate attachment site. By aligning the applied force with the measurement axis, the molecule is no longer undergoing combined loading, and the full force can be measured by the cantilever. Simulations and experimental results verify the ability of the alignment program to minimize pulling geometry errors in AFM-SMFS studies.

| S-EPMC2553119 | biostudies-literature

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| S-EPMC3397402 | biostudies-literature

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| S-EPMC3588056 | biostudies-literature

Interaction between amyloid-? peptide and heme probed by electrochemistry and atomic force microscopy.

Project description:Heme binds to amyloid ?-peptide (A?) in the brain of Alzheimer's disease (AD) patients, thus forming A?-heme complexes and leading the characteristic pathological features of AD. The interaction between heme and A? might have important biological relevance to AD etiology. In this work, the electrochemical performances of heme after incubation with A?1-42, A? fragments, and mutated A? were systematically investigated using cyclic voltammetry and differential pulse voltammetry. Our results indicated that His13 and His14 were possible binding sites, and A? bound two molecules of heme with a binding constant of K(a1) = 7.27 × 10(6) M(-1) (n(1) = 1.5) and K(a2) = 2.89 × 10(6) M(-1) (n(1) = 1.8). Detailed analysis with atomic force microscopy (AFM) of A?1-42 in the absence or presence of heme under the same incubation conditions showed that heme inhibited the formation of A? fibrils. According to results of the spectroscopic characterization, Arg5 was the key residue in making the heme-A?1-42 complex as a peroxidase.

| S-EPMC3629740 | biostudies-literature

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| S-EPMC5908543 | biostudies-literature

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