Establishment of a microwell-array-based miniaturized thymic organoid model suitable for high-throughput applications

Area of Science:

• Immunology
• Developmental Biology
• Biotechnology

Background:

• T cell development is crucial and relies on thymic epithelial cells (TECs) within the thymic stroma.
• Existing in vitro thymus models lack the robustness and throughput for comprehensive analysis.

Purpose of the Study:

• To develop a novel, high-density microwell-array-based miniaturized thymus organoid (mTO) model.
• To assess the mTO's capacity to support T cell development and mimic native thymus organization.
• To evaluate the mTO's compatibility with live imaging and medium-/high-throughput applications.

Main Methods:

• Fabrication of miniaturized thymus organoids (mTOs) using high-density microwell arrays.
• Characterization of mTO cellular input, TEC phenotype, and complexity.
• Utilizing mTOs to investigate the role of fetal thymic mesenchyme in TEC differentiation.

Main Results:

• The mTO model successfully supports T cell commitment and development.
• mTOs exhibit key organizational characteristics and TEC complexity mirroring the native thymus.
• Fetal thymic mesenchyme plays a role in mature TEC sub-population differentiation beyond Foxn1 maintenance.

Conclusions:

• Miniaturized thymus organoids (mTOs) provide a robust in vitro model of the native thymus.
• The mTO platform is adaptable for medium-/high-throughput applications in thymus biology research.
• mTOs are validated for exploring thymus and organoid development, offering new avenues for discovery.