An in vitro model of the tumor-lymphatic microenvironment with simultaneous transendothelial and luminal flows reveals mechanisms of flow enhanced invasion.

The most common cancers, including breast and skin, disseminate initially through the lymphatic system, yet the mechanisms by which tumor cells home towards, enter and interact with the lymphatic endothelium remain poorly understood. Transmural and luminal flows are important biophysical cues of the lymphatic microenvironment that can affect adhesion molecules, growth factors and chemokine expression as well as matrix remodeling, among others. Although microfluidic models are suitable for in vitro reconstruction of highly complex biological systems, the difficult assembly and operation of these systems often only allows a limited throughput. Here we present and characterize a novel flow chamber which recapitulates the lymphatic capillary microenvironment by coupling a standard Boyden chamber setup with a micro-channel and a controlled fluidic environment. The inclusion of luminal and transmural flow renders the model more biologically relevant, combining standard 3D culture techniques with advanced control of mechanical forces that are naturally present within the lymphatic microenvironment. The system can be monitored in real-time, allowing continuous quantification of different parameters of interest, such as cell intravasation and detachment from the endothelium, under varied biomechanical conditions. Moreover, the easy setup permits a medium-high throughput, thereby enabling downstream quantitative analyses. Using this model, we examined the kinetics of tumor cell (MDA-MB-231) invasion and transmigration dynamics across lymphatic endothelium under varying flow conditions. We found that luminal flow indirectly upregulates tumor cell transmigration rate via its effect on lymphatic endothelial cells. Moreover, we showed that the addition of transmural flow further increases intravasation, suggesting that distinct flow-mediated mechanisms regulate tumor cell invasion.

Mechanical characterization of porcine corneas.

An experimental program has been carried out in order to investigate the mechanical behavior of porcine corneas. We report the results of inflation tests on the whole cornea and uniaxial tests on excised corneal strips, performed on 51 fresh porcine eyes. Uniaxial tests have been performed on specimens cut from previously inflated corneas. The cornea behavior is characterized by means of elastic stiffness, measured on both average pressure-apex displacement and average uniaxial stress-strain curves; and by means of transversal contraction coefficient, peak stress, and failure stress measured on uniaxial stress-strain curves. Uniaxial tests performed on excised strips allowed to measure the anisotropy in the corneal stiffness and to compare the stiffness of the cornea with the one of the sclera. Viscous properties of the cornea have been obtained through uniaxial relaxation curves on excised corneal strips. The relevant geometrical parameters have been measured and, with the aid of the elastic thin shell theory, a stress-strain curve has been derived from the average inflation test data and compared with similar data available in the literature. The experimental system has been developed in view of future applications to the mechanical testing of both porcine and human corneas.