RESUMEN
Soluble signaling molecules and extracellular matrix (ECM) regulate cell dynamics in various biological processes. Wound healing assays are widely used to study cell dynamics in response to physiological stimuli. However, traditional scratch-based assays can damage the underlying ECM-coated substrates. Here, we use a rapid, non-destructive, label-free magnetic exclusion technique to form annular aggregates of bronchial epithelial cells on tissue-culture treated (TCT) and ECM-coated surfaces within 3 h. The cell-free areas enclosed by the annular aggregates are measured at different times to assess cell dynamics. The effects of various signaling molecules, including epidermal growth factor (EGF), oncostatin M, and interleukin 6, on cell-free area closures are investigated for each surface condition. Surface characterization techniques are used to measure the topography and wettability of the surfaces. Further, we demonstrate the formation of annular aggregates on human lung fibroblast-laden collagen hydrogel surfaces, which mimic the native tissue architecture. The cell-free area closures on hydrogels indicate that the substrate properties modulate EGF-mediated cell dynamics. The magnetic exclusion-based assay is a rapid and versatile alternative to traditional wound healing assays.
RESUMEN
This work describes a versatile and cost-effective cell culture method for micropatterning and growing adherent cells on porous membranes using pressure-sensitive double-sided adhesives. This technique also allows cell culture using conventional methods and their easy integration into microfluidic chip devices. Adhesives can be used to form different patterns of cultured cells, which can be used for cell proliferation and wound-healing models. To demonstrate the viability of our system, we evaluate the toxicity effect of five different adhesives on two distinct airway epithelial cell lines and show functional applications for cell patterning and microfluidic cell culture chip fabrication. We developed a sandwiched microfluidic device that enabled us to culture cells in a submerged condition and transformed it into a dynamic platform when required. The viability of cells and their inflammatory responses to IL-1ß stimulation were investigated. Our technique is applicable for conventional culturing of cells, widely available in biomedical research labs, while enabling the introduction of perfusion for an advanced dynamic cell culture model when needed.
Asunto(s)
Adhesivos , Microfluídica , Células Epiteliales , Dispositivos Laboratorio en un Chip , PulmónRESUMEN
Fibroblasts (mouse, NIH/3T3) are combined with MDA-MB-231 cells to accelerate the formation and improve the reproducibility of 3D cellular structures printed with magnetic assistance. Fibroblasts and MDA-MB-231 cells are cocultured to produce 12.5 : 87.5, 25 : 75, and 50 : 50 total population mixtures. These mixtures are suspended in a cell medium containing a paramagnetic salt, Gd-DTPA, which increases the magnetic susceptibility of the medium with respect to the cells. A 3D monotypic MDA-MB-231 cellular structure is printed within 24 hours with magnetic assistance, whereas it takes 48 hours to form a similar structure through gravitational settling alone. The maximum projected areas and circularities, and cellular ATP levels of the printed structures are measured for 336 hours. Increasing the relative amounts of the fibroblasts mixed with the MDA-MB-231 cells decreases the time taken to form the structures and improves their reproducibility. Structures produced through gravitational settling have larger maximum projected areas and cellular ATP, but are deemed less reproducible. The distribution of individual cell lines in the cocultured 3D cellular structures shows that printing with magnetic assistance yields 3D cellular structures that resemble in vivo tumors more closely than those formed through gravitational settling. The results validate our hypothesis that (1) fibroblasts act as a "glue" that supports the formation of 3D cellular structures, and (2) the structures are produced more rapidly and with greater reproducibility with magnetically assisted printing than through gravitational settling alone. Printing of 3D cellular structures with magnetic assistance has applications relevant to drug discovery, lab-on-chip devices, and tissue engineering.
RESUMEN
Controlled cell assembly is essential for fabricating in vitro 3D models that mimic the physiology of in vivo cellular architectures. Whereas tissue engineering techniques often rely on intrusive magnetic nanoparticles placed in cells and hydrogel encapsulation of cells to produce multilayered cellular constructs, we describe a high-throughput, label-free, and scaffold-free magnetic field-guided technique that assembles cells into a layered aggregate. An inhomogeneous magnetic field influences the diamagnetic cells suspended in a paramagnetic culture medium. Driven by the magnetic susceptibility difference and the field gradient, the cells are displaced toward the region of lowest field strength. Two cell lines are used to demonstrate the sequential assembly of layer-on-layer aggregates in microwells within 6 h. The effect of magnet size on the assembly dynamics is characterized and a microwell size criterion for the highest cell aggregation provided. Label-free magnetic-field-assisted assembly is relevant for on-demand scalable biofabrication of complex layered structures. Potential applications include drug discovery, developmental biology, lab-on-chip devices, and cancer research.
Asunto(s)
Campos Magnéticos , Ingeniería de Tejidos , Agregación Celular , HidrogelesRESUMEN
Acoustophoresis is rapidly gaining prominence in the field of cell manipulation. In recent years, researchers have extensively used this method for separating different types of cells from the bulk fluid. In this paper, we propose a novel acoustophoresis-based technique to capture infected or abnormal erythrocytes from blood plasma. A typical acoustic device consisting of a transducer assembly, microfluidic cavity, and a reflector is considered. Based on the concept of impedance matching, a pair of antibody-coated polystyrene layers is placed in the nodal regions of an acoustic field within the cavity. This technique allows bi-directional migration of the suspended cells to the biofunctionalized surfaces. Therefore, simultaneous capture of infected erythrocytes on both the layers is feasible. Finite element method is used to model the pressure field as well as the motion of erythrocytes under the influence of acoustic radiation, drag, and gravitational forces. A parametric analysis is done by varying the excitation frequency, driving voltage, and the thickness of the polystyrene layers. The resulting changes in the pressure amplitude and field pattern are investigated. The erythrocyte collection efficiency, rate of collection, and the cell distribution on the layer surfaces are also determined under different field conditions. The occurrence of transient cavitation in the blood plasma-filled cavity at the chosen frequency is taken into account by using its threshold pressure value as the limiting factor of pressure amplitude. The study provides an insight into the phenomenon and serves as a guideline to fabricate low-cost, multifunctional rapid diagnostic devices based on acoustophoretic separation.