Microfluidic investigation of the effect of graphene oxide on mechanical properties of cell and actin cytoskeleton networks: experimental and theoretical approaches

Abstract

Biomechanical and morphological analysis of the cells is a novel approach for monitoring the environmental features, drugs, and toxic compounds’ effects on cells. Graphene oxide (GO) has a broad range of medical applications such as tissue engineering and drug delivery. However, the effects of GO nanosheets on biological systems have not been completely understood. In this study, we focused on the biophysical characteristics of cells and their changes resulting from the effect of GO nanosheets. The biophysical properties of the cell population were characterized as follows: cell stiffness was calculated by atomic force microscopy, cell motility and invasive properties were characterized in the microfluidic chip in which the cells are able to visualize cell migration at a single-cell level. Intracellular actin was stained to establish a quantitative picture of the intracellular cytoskeleton. In addition, to understand the molecular interaction of GO nanosheets and actin filaments, coarse-grained (CG) molecular dynamics (MD) simulations were carried out. Our results showed that GO nanosheets can reduce cell stiffness in MCF7 cells and MDA-MB-231 cell lines and highly inhibited cell migration (39.2%) in MCF-7 and (38.6%) in MDA-MB-231 cell lines through the GO nanosheets-mediated disruption of the intracellular cytoskeleton. In the presence of GO nanosheets, the cell migration of both cell lines, as well as the cell stiffness, significantly decreased. Moreover, after GO nanosheets treatment, the cell actin network dramatically changed. The experimental and theoretical approaches established a quantitative picture of changes in these networks. Our results showed the reduction of the order parameter in actin filaments was 23% in the MCF7 cell line and 20.4% in the MDA-MB-231 cell line. The theoretical studies also showed that the GO nanosheet–actin filaments have stable interaction during MD simulation. Moreover, the 2D free energy plot indicated the GO nanosheet can induce conformational changes in actin filaments. Our findings showed that the GO nanosheets can increase the distance of actin-actin subunits from 3.22 to 3.5 nm and in addition disrupt native contacts between two subunits which lead to separate actin subunits from each other in actin filaments. In this study, the biomechanical characteristics were used to explain the effect of GO nanosheets on cells which presents a novel view of how GO nanosheets can affect the biological properties of cells without cell death. These findings have the potential to be applied in different biomedical applications.

Graphene Enhances Actin Filament Assembly Kinetics and Modulates NIH-3T3 Fibroblast Cell Spreading

“…Changes in the microenvironment have also been also shown to affect the actin cytoskeleton. Recent studies have suggested that exposure to nanomaterials, in particular carbon-based nanomaterials (i.e., single-wall carbon nanotubes (SWCNT), graphene oxide, and graphene), can modulate actin polymerization [ 14 , 15 , 16 , 17 , 18 ].…”

“…In addition, a molecular dynamics (MD) simulation has shown that actin monomers adhere to graphene via weak interactions, including van der Waal forces, electrostatic interactions, and hydrogen bonding [ 34 ]. These interactions are not strong enough to dissociate two actin monomers compared to graphene oxide (GO) and reduced graphene oxide (rGO) [ 16 , 17 ], of which functional groups on graphene oxide can form hydrogen bonds with oxygen-containing residues in actin [ 34 ], thereby inducing actin disassembly [ 16 , 17 ]. While the effects of graphene on the actin cytoskeleton have been reported at the cellular level, how graphene modulates actin filament assembly is not well established.…”

Fingerprinting Metabolic Activity and Tissue Integrity of 3D Lung Cancer Spheroids under Gold Nanowire Treatment

“…Nevertheless, traditional twodimensional cell culture approaches are still being used as a route for the risk assessment of nanomaterials and chemical agents [16][17][18][19]. While 2D cell culture platforms will remain an attractive research strategy to investigate basic cellular behaviors [20,21], these simple two dimensional in vitro cell-based assays are not suited to address more complex biological questions where cellular responses, cell-to-cell, and cell-to-matrix interactions are governed by the tissue-specific architecture [17,[22][23][24].…”

“…This aspect is particularly important by nanomaterial-biology interactions where the uptake, transport, and distribution can be directly linked to the shape, composition, and size of the tissue structure [25]. Hence, the requirement for high-throughput and cost-effective preclinical cell-based assays for assessing the risk of nanoparticles and nanodrug screening studies needs to also involve functional 3D-tissue structures [1,2,21] that can bridge the gap between in vivo and in vitro models [6,26]. Here, three-dimensional spheroid models that are generated either by using commercial microplates, microfluidics systems [4] are ideally suited to address nanomaterial-biology interactions that require adequate cell-to-cell interactions and appropriate cell signaling events [3,27,28].…”

“…The current study sets out to provide a better understanding of how the uptake rate of AuNWs could be influenced by 3D multi-cellular spheroids. The use of nanomaterials as theranostic (therapy and diagnostic) tools promises advantages such as enhanced efficacy, better drug stability, biocompatibility or bioavailability, improved dosing, and reduced side effects [21,35,36]. Gold nanotoxicity has been studied by scientists due to its importance for biomedical applications, and it is well established and described for drug delivery platforms [37][38][39].…”