Development of artificial tissues or organs is one of the actual tasks in regenerative medicine that requires observation and evaluation of intact volume microstructure of tissue engineering products at all stages of their formation, from native donor tissues and decellularized scaffolds to recipient cell migration in the matrix. Unfortunately in practice, methods of vital noninvasive imaging of volume microstructure in matrixes are absent. In this work, we propose a new approach based on high‐frequency acoustic microscopy for noninvasive evaluation and visualization of volume microstructure in tissue engineering products. The results present the ultrasound characterization of native rat diaphragms and lungs and their decellularized scaffolds. Verification of the method for visualization of tissue formation in the matrix volume was described in the model samples of diaphragm scaffolds with stepwise collagenization. Results demonstrate acoustic microscopic sensitivity to cell content concentration, variation in local density, and orientation of protein fibers in the volume, micron air inclusions, and other inhomogeneities of matrixes.

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We analyzed viability of mesenchymal stem cells seeded by static and dynamic methods to highly porous fibrous 3D poly-L-lactide scaffolds with similar physical and chemical properties, but different spatial organization modified with collagen. Standard collagen coating promoted protein adsorption on the scaffold surface and improved adhesive properties of 100 μ-thick scaffolds. Modification of 600-μ scaffolds with collagen under pressure increased proliferative activity of mesenchymal stem cells seeded under static and dynamic (delivery of 100,000 cells in 10 ml medium in a perfusion system at a rate of 1 ml/min) conditions by 47 and 648%, respectively (measured after 120-h culturing by MTT test). Dynamic conditions provide more uniform distribution of collagen on scaffold fibers and promote cell penetration into 3D poly-L-lactide scaffolds with thickness >600 μ.

The aim of this article is to verify the applicability of impulse acoustic microscopy as a tool for evaluating scaffolds in tissue engineering. Decellularized rat diaphragms and nonwoven polymer scaffolds made from cellulose diacetate, chlorinated polyvinyl chloride, and polysulfone were used as model objects. Optical microscopy, scanning electron microscopy, and histological research were used as reference methods in order to realize feasibility of the acoustic microscopy method in a regenerative medicine field. Direct comparison of different methods was carried out. The high frequency ultrasound microscopy is defined as a useful non‐invasive tool for studying and investigating the bulk morphology and bulk structure of opaque scaffolds, which allows to diagnose and to assume heterogeneous regions and defects inside the specimens. POLYM. ENG. SCI., 57:709–715, 2017. © 2017 Society of Plastics Engineers

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The fibrous porous structure of polymers can mimic the extracellular matrix of the native tissue, therefore such polymers have a good potential for use in regenerative medicine. Organs and tissues within the body exhibit different mechanical properties depending on their functionality, thus artificial scaffolds should have mechanical behaviors similar to the extracellular matrix in conditions like living organisms, primarily in aqueous media. Several methods have been investigated in aquatic environments, including noninvasive techniques based on ultrasonic focused beams for biological objectives. In this study we explored the tensile behavior of poly(L-lactide) nonwoven polymer scaffolds using high-frequency ultrasound microscopy combined with a horizontal testing machine, which provided a visualization of the reorganization and transformation of the dynamic volume microstructure. The mechanisms of unwinding, elongation, orientation, and deformation of polymer fibers under uniaxial tension were revealed. We observed an association between the lined plastic deformation from 100 to 400% and the formation of multiple necks in the fibers, which caused stress relaxation and significant rarefaction of the fibrous microstructure. It was shown that both peaks on the stress–strain curve corresponded to the microstructure of aligned fibers in terms of initial diameter and thinning fibers. We discuss the possible influence of these microstructure transformations on cell behavior.

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