Alginate Hydroxyapatite scaffold	PCL Skin scaffold

Micro-CT (Micro-Computed Tomography) is a modern preclinical imaging method allowing non-destructive visualizations and structure analysis yielding at a resolution of a few micrometers. Micro-CT has become a standard and essential tool for quantifying structure-function relationships, disease progression, and regeneration in preclinical models and has facilitated numerous scientific advancements in tissue engineering field over the past 30 years.

Micro-CT applications in tissue engineering

 Tissue engineering evolved from the field of biomaterials development and refers to the practice of combining scaffolds, cells, and biologically active molecules into functional tissues. The goal of tissue engineering is to assemble functional constructs that restore, maintain, or improve damaged tissues or whole organs. Artificial skin and cartilage are examples of engineered tissues. Micro-CT emerged as a commercially available tool in the middle of the ‘go-go’ years of tissue engineering (that is, the 1980s and 1990s), positioning it perfectly for widespread use as the problems targeted by tissue engineers necessitate non-destructive, 3D, quantitative imaging techniques.

Scaffold Characterization

 Scaffolds must balance mechanical properties with degradation kinetics and byproducts, sufficient porosity for cellular infiltration and seeding, and drug delivery characteristics, among other criteria. Thus, nondestructive quantification of microstructural characteristics such as porosity, surface-to-volume ratio, interconnectivity, and anisotropy is necessary for scaffold optimization, and micro-CT has the potential to provide comprehensive data on these parameters. Scaffold porosity and pore interconnectivity are key factors in biomaterial design to enable cell migration, proliferation, and extracellular matrix production and facilitate tissue in-growth and blood vessel invasion but come with trade-offs in other scaffold parameters, such as mechanical properties. Micro-CT has become a critical tool for quantitative and nondestructive assessment of internal scaffold microstructure to guide scaffold design and manufacture and enables non-destructive evaluation of both microstructural and mechanical behavior of multi-phase and fiber-reinforced scaffolds as well as longitudinal scaffold degradation.

Evaluate the ability of cell based tissue engineering bone constructs

 Micro-CT is also used to evaluate the ability of cell based tissue engineering bone constructs to form biologic mineralized matrix in vitro. The studies demonstrated that osteogenic differentiation of stem cells in vitro is dependent on substrate material and microstructural characteristics, cell source (for example, amniotic fluid- versus bone marrow-derived mesenchymal cells) [56], and dimensional (that is, 2D versus 3D)  and biomechanical culture conditions. Unlike other in vitro osteogenesis assays, micro-CT enables longitudinal quantification of the time course of mineralization in 3D without interfering with cell growth or mineral production, an important feature for comparison of various cell sources with different mineralization kinetics. In addition to scaffold microstructure, micro-CT enables assessment of tissue engineered bone formation in animal models (for example, high-density stem cell-mediated bone regeneration of calvarial defects).

Cell Distribution and Infiltration

Understanding the distribution and infiltration of cells within tissue constructs is crucial for successful tissue engineering. Micro-CT enables researchers to label cells or use contrast agents to visualize and quantify their distribution within scaffolds. This information helps in optimizing cell seeding techniques, ensuring uniform cell distribution, and evaluating the effectiveness of cell infiltration strategies. It also provides insights into cell behavior within the scaffold and their interaction with the surrounding microenvironment.

Bone scaffold implanted into rat skull

Vascularization Assessment

 The development of a functional vascular network is essential for the survival and integration of tissue-engineered constructs. Micro-CT allows for the visualization and quantification of blood vessels within tissue constructs. It helps researchers assess the formation, density, and connectivity of blood vessels, as well as their interaction with the surrounding tissue. This information is critical for evaluating the success of strategies aimed at promoting angiogenesis and ensuring proper vascularization of tissue-engineered constructs.

Tissue Integration and Host Response

 Micro-CT can provide insights into the integration of tissue-engineered constructs with the host tissue. By visualizing the interface between the implanted construct and the surrounding tissue, researchers can assess the quality of integration, identify potential complications, and monitor the host response. This information helps in evaluating the biocompatibility and functionality of tissue-engineered constructs in vivo.

Longitudinal Studies

 Microcomputed tomography enables longitudinal studies, allowing researchers to track the changes in tissue constructs over time. By acquiring multiple scans at different time points, scientists can monitor the development, growth, and degradation of tissue-engineered constructs. Longitudinal studies provide valuable information on the structural and functional changes that occur within the constructs, helping researchers understand their behavior and optimize their design.

Computational Modeling

 Micro-CT data can be used for computational modeling of tissue constructs. By converting the 3D images into digital representations, researchers can simulate and analyze the mechanical properties, fluid flow, and mass transport within the constructs. Computational modeling based on micro-CT data aids in optimizing scaffold design, predicting the behavior of tissue constructs, and guiding tissue engineering strategies.

Scaffold implanted into rat knee

Quality Control and Validation

 Micro-CT serves as a powerful tool for quality control and validation of tissue-engineered constructs with enabling non-destructive evaluation of scaffold properties, cell distribution, and tissue integration. Micro-CT scans can be compared against design specifications to ensure the accuracy and consistency of fabricated constructs. This imaging method also aids in identifying any structural defects, inconsistencies, or deviations from desired parameters.

In conclusion, Micro-CT has contributed to dramatic advances in tissue engineering, enabling fundamental studies in bone structure and function, quantitative evaluation of disease progression and treatment, development of new tissue engineering strategies, and contrast-enhanced soft tissue imaging. Both NDT and in vivo micro-CT systems are increasing in availability and application, and continued advancements and innovations promise to continue this trajectory into the future.

Below is a summary of some of the applications of micro-CT in the tissue engineering:

• Visualization and assessment of scaffolds’ internal microstructure

• Non-destructive quantification of microstructural scaffold characteristics such as porosity, surface-to-volume ratio, interconnectivity, and anisotropy

• Assessment of biomaterial properties to meet the biomechanical and biological requirements of complex tissues and organs

• Evaluation of effects of mechanical stimuli on tissue regeneration (e.g. evaluation of the effect of compressive loading or unloading on local bone formation and resorption)

• Longitudinal Quantification of scaffold integration and mineralization

• Non-destructive quantification of tissue engineering