High-Resolution Imaging for the Analysis and Reconstruction of 3D Microenvironments for Regenerative Medicine: An Application-Focused Review

Abstract

The rapid evolution of regenerative medicine and its associated scientific fields, such as tissue engineering, has provided great promise for multiple applications where replacement and regeneration of damaged or lost tissue is required. In order to evaluate and optimise the tissue engineering techniques, visualisation of the material of interest is crucial. This includes monitoring of the cellular behaviour, extracellular matrix composition, scaffold structure, and other crucial elements of biomaterials. Non-invasive visualisation of artificial tissues is important at all stages of development and clinical translation. A variety of preclinical and clinical imaging methods—including confocal multiphoton microscopy, optical coherence tomography, magnetic resonance imaging (MRI), and computed tomography (CT)—have been used for the evaluation of artificial tissues. This review attempts to present the imaging methods available to assess the composition and quality of 3D microenvironments, as well as their integration with human tissues once implanted in the human body. The review provides tissue-specific application examples to demonstrate the applicability of such methods on cardiovascular, musculoskeletal, and neural tissue engineering.

Modified Histopathological Protocol for Poly-ɛ-Caprolactone Scaffolds Preserving Their Trabecular, Honeycomb-like Structure

“…Despite the considerable developments in imaging techniques allowing for comprehensive in vivo and postmortem scaffold evaluation, and even the creation of 3D virtual models [ 29 , 30 ], little attention has been paid to the refinement and updating of histopathological techniques, which remain the basic tool for assessments of scaffold morphology, biocompatibility, and interactions with the surrounding cells. The standard techniques used today only allow for evaluations of selected representative fragments, where the scaffold is often cleared, melted, or swollen, and it is not possible to obtain the full cross-section view with the preserved scaffold’s honeycomb-like structure [ 31 , 32 ].…”

Characterization of Biocompatibility of Functional Bioinks for 3D Bioprinting

“…Laser confocal scanning microscopy (LCSM) can acquire multiple 2D images at different depths and reconstruct 3D images by the process known as optical sectioning in a sample, generating high-resolution 3D images ( Figure 3 ) [ 82 ]. This technique has been widely used in material science and life science, including tissue engineering and regenerative medicine [ 83 , 84 , 85 ]. With selective fluorescent labeling chosen by a researcher, the LCSM can provide precisely focused 3D images of subcellular structures such as nucleic acids, membranes, and mitochondria [ 59 , 86 ].…”

Imaging approaches for monitoring three‐dimensional cell and tissue culture systems

“…Scalability of the tool to reach particular spatial (moving from 2D to 3D) and temporal (real‐time acquisitions for living systems) resolution is a key factor in tool selection. This review focuses on laboratory‐grade methods, which are aimed at micro‐scale resolutions, however, some tools such as 2PEF/FLIM, SHG, THG, CARS, OCT, SRS and PAM can be scaled up to clinical‐grade methods for pathological, pre‐clinical or clinical applications [89, 241].…”

“…An additional factor to consider is moving towards high‐throughput automated image acquisition and analysis. High‐throughput research requires samples that can be imaged in high‐throughput and automated modalities, however, moving from 2D to 3D models is a challenge in high‐throughput applications due to the complexity of imaging as the scale increases [241]. Fluorescence modalities are well‐known for being incorporated into high‐throughput imaging systems.…”