Innovative Platforms for Macrophage Behavior: Establishment of Stable Macrophage Reporters And Macrophage-Incorporated Airway Organoid

 Macrophages are an essential cellular component of innate immunity that perform immunosurveillance and maintain tissue homeostasis by detecting the presence of foreign pathogens and toxins, removing cell debris or dysfunctional cells, and releasing effector molecules such as cytokines, chemokines, and growth factors to the tissue microenvironment. Macrophage polarization is a highly dynamic process due to its sensory nature, and characterizing macrophage states has been a longstanding challenge. Some challenges have not been addressed in current phenotyping techniques, including one opportunity to measure cell behavior and failure to reflect the temporal determinates of cell fate. Given the macrophage's dynamic sensory nature, there is a need to measure macrophage cell fate equally dynamically. Here, we developed a responsive-element luciferase-tagged macrophage reporter cell line via lentiviral transduction to assess the temporal signals generated. Combined with bioluminescence temporal spectrometry that captures luciferase activities as the readouts, STAT6-RE THP-1 reporter can accurately report the dynamics of M2 polarization in single-dose administration of IL-4 and lung cancer spheroid model. These experiments allow for a better understanding of the association between molecular mechanisms and macrophage polarization, facilitating the development of therapeutic strategies for manipulating macrophages to attenuate inflammation and tumor progression and promote wound healing.

The lung is a vital respiratory organ for gas exchange. The airway epithelium is considered a dynamic structure that responds to environmental stimuli and closely coordinates with innate immune cells such as monocytes and macrophages to facilitate host immune defense against infection and maintain lung homeostasis. However, current in vitro models, including 2D air-liquid interface culture and 3D organoid, primarily focus on distal lungs and lack innate immune cellular components. In this thesis, a THP-1 cell incorporated apical-out airway organoid model is established to address these limitations. This novel model recapitulates the immunosuppressive microenvironment of lung airway epithelium and captures the development of monocytes to an airway macrophage-like phenotype. By incorporating the innate immune compartment, this model offers an advantage over traditional lung airway models for better investigating innate immunity in lung and respiratory diseases.