The focal mechanical properties of normal and diseased porcine aortic valve tissue measured by a novel microindentation device.

Cells sense and respond to the heterogeneous mechanical properties of their tissue microenvironment, with implications for the development of many diseases, including cancer, fibrosis, and aortic valve disease. Characterization of tissue mechanical heterogeneity on cellular length scales of tens of micrometers is thus important for understanding disease mechanobiology. In this study, we developed a low-cost bench-top microindentation system to readily map focal microscale soft tissue mechanical properties. The device was validated by comparison with atomic force microscopy nanoindentation of polyacrylamide gels. To demonstrate its utility, the device was used to measure the focal microscale elastic moduli of normal and diseased porcine aortic valve leaflet tissue. Consistent with previous studies, the fibrosa layer of intact leaflets was found to be 1.91-fold stiffer than the ventricularis layer, with both layers exhibiting significant heterogeneity in focal elastic moduli. For the first time, the microscale compressive moduli of focal proteoglycan-rich lesions in the fibrosa of early diseased porcine aortic valve leaflets were measured and found to be 2.44-fold softer than those of normal tissue. These data provide new insights into the tissue micromechanical environment in valvular disease and demonstrate the utility of the microindentation device for facile measurement of the focal mechanical properties of soft tissues.

Microdevice arrays with strain sensors for 3D mechanical stimulation and monitoring of engineered tissues.

Native and engineered tissue development are regulated by the integrative effects of multiple microenvironmental stimuli. Microfabricated bioreactor array platforms can efficiently dissect cue-response networks, and have recently integrated critical 2D and 3D mechanical stimulation for greater physiological relevance. However, a limitation of these approaches is that assessment of tissue functional properties is typically limited to end-point analyses. Here we report a new deformable membrane platform with integrated strain sensors that enables mechanical stretching or compression of 3D cell-hydrogel arrays and simultaneous measurement of hydrogel construct stiffness in situ. We tested the ability of the integrated strain sensors to measure the evolution of the stiffness of cell-hydrogel constructs for two cases. First, we demonstrated in situ stiffness monitoring of degradable poly (ethylene glycol)-norbornene (PEG-NB) hydrogels embedded with mesenchymal stromal cells (MSCs) and cultured with or without cyclic tensile stimulation for up to 15 days. Whereas statically-cultured hydrogels degraded and softened throughout the culture period, mechanically-stimulated gels initially softened and then recovered their stiffness corresponding to extensive cell network and collagen production. Second, we demonstrated in situ measurement of compressive stiffening of MSC-seeded PEG-NB gels cultured statically under osteogenic conditions, corresponding to increased mineralization and cellularization. This measurement technique can be generalized to other relevant bioreactor and organ-on-a-chip platforms to facilitate online, non-invasive, and high-throughput functional analysis, and to provide insights into the dynamics of engineered tissue development that are otherwise not available.

Polyacrylamide gel substrates that simulate the mechanical stiffness of normal and malignant neuronal tissues increase protoporphyin IX synthesis in glioma cells.

Protoporphyrin IX (PPIX) produced following the administration of exogenous 5d-aminolevulinic acid is clinically approved for photodynamic therapy and fluorescence-guided resection in various jurisdictions around the world. For both applications, quantification of PPIX forms the basis for accurate therapeutic dose calculation and identification of malignant tissues for resection. While it is well established that the PPIX synthesis and accumulation rates are subject to the cell's biochemical microenvironment, the effect of the physical microenvironment, such as matrix stiffness, has received little attention to date. Here we studied the proliferation rate and PPIX synthesis and accumulation in two glioma cell lines U373 and U118 cultured under five different substrate conditions, including the conventional tissue culture plastic and polyacrylamide gels that simulated tissue stiffness of normal brain (1 kPa) and glioblastoma tumors (12 kPa). We found that the proliferation rate increased with substrate stiffness for both cell lines, but not in a linear fashion. PPIX concentration was significantly higher in cells cultured on tissue-simulating gels than on the much stiffer tissue culture plastic for both cell lines. These findings, albeit preliminary, suggest that the physical microenvironment might be an important determinant of tumor aggressiveness and PPIX synthesis in glioma cells.

A metric for the stiffness of calcified aortic valves using a combined computational and experimental approach.

Calcific aortic valve disease is the most common heart valve disease. It is associated with a significant increase in cardiovascular morbidity and mortality and independently increases the cardiovascular risk. It is then important to develop parameters that can estimate the stiffness of the valve. Such parameters may contribute to early detection of the disease or track its progression and optimize the timing for therapy. In this study, we introduce a metric representing the stiffness of the native aortic calcified valve over a wide range of stenosis severities. Our approach is based on three-dimensional structural finite-element simulations and in vitro measurements. The proposed method is developed first in a pulse duplicator; its clinical applicability is then evaluated in three patients with severe aortic stenosis. Our results indicate that the value of the proposed metric varies considerably between healthy valves and valves with very severe aortic stenosis, from 0.001 to 7.38 MPa, respectively. The method introduced in this study could give useful information regarding the stiffness of the valve leaflets with potential application to the evaluation of aortic sclerosis and aortic stenosis.