RESUMEN
Diffusion and transport of small molecules within hydrogel networks are of high interest for biomedical and pharmaceutical research. Herein, using fluorescence correlation spectroscopy (FCS), we experimentally showed that the diffusion time in the hydrogel was directly related to the mechanical state (compression or swelling) and thus to the volume fraction of the gel. Following this observation, we developed cell-like barometers in the form of PAA microbeads, which when incorporated between cells and combined with a diffusion-based optical readout could serve as the first biosensors to measure the local pressure inside the growing biological tissues. To illustrate the potential of the present method, we used multicellular spheroids (MCS) as a tissue model, and it was observed that the growth-associated tissue stress was lower than 1 kPa, but significantly increased when an external compressive stress was applied.
RESUMEN
The surrounding microenvironment limits tumour expansion, imposing a compressive stress on the tumour, but little is known how pressure propagates inside the tumour. Here we present non-destructive cell-like microsensors to locally quantify mechanical stress distribution in three-dimensional tissue. Our sensors are polyacrylamide microbeads of well-defined elasticity, size and surface coating to enable internalization within the cellular environment. By isotropically compressing multicellular spheroids (MCS), which are spherical aggregates of cells mimicking a tumour, we show that the pressure is transmitted in a non-trivial manner inside the MCS, with a pressure rise towards the core. This observed pressure profile is explained by the anisotropic arrangement of cells and our results suggest that such anisotropy alone is sufficient to explain the pressure rise inside MCS composed of a single cell type. Furthermore, such pressure distribution suggests a direct link between increased mechanical stress and previously observed lack of proliferation within the spheroids core.
Asunto(s)
Microesferas , Presión , Esferoides Celulares/fisiología , Estrés Fisiológico/fisiología , Microambiente Tumoral/fisiología , Resinas Acrílicas/química , Animales , Línea Celular Tumoral , Proliferación Celular/fisiología , Elasticidad , Procesamiento de Imagen Asistido por Computador , Ratones , Microscopía ConfocalRESUMEN
Microfluidic bioreactors are expected to impact cell therapy and biopharmaceutical production due to their ability to control cellular microenvironments. This work presents a novel approach for continuous cell culture in a microfluidic system. Microcarriers (i.e., microbeads) are used as growth support for anchorage-dependent mammalian cells. This approach eases the manipulation of cells within the system and enables harmless extraction of cells. Moreover, the microbioreactor uses a perfusion function based on the biocompatible integration of a porous membrane to continuously feed the cells. The perfusion rate is optimized through simulations to provide a stable biochemical environment. Thermal management is also addressed to ensure a homogeneous bioreactor temperature. Eventually, incubator-free cell cultures of Drosophila S2 and PC3 cells are achieved over the course of a week using this bioreactor. In future applications, a more efficient alternative to harvesting cells from microcarriers is also anticipated as suggested by our positive results from the microcarrier digestion experiments.