Effect of Genetic Knockout and Pharmacological Inhibition of Galectin-3 on Alveolar Type 2 Cell Differentiation in Organoids

Abstract

Chronic respiratory diseases present a large medical and economic burden on the industrialized world, only overshadowed by heart disease and cancer. These diseases result in punctuated lung injury and repair cycles with a sequalae of irreparably damaged tissue architecture. Lungs have multiple stem cell niches which repair damage after injury, one of which is in the alveoli. Within this niche, Alveolar type 2 cells (AT2) self-renew and differentiate into Alveolar type 1 (AT1) cells which are in contact with capillary beds allowing for gas exchange. After alveolar injury, AT2 cells undergo a transcriptionally distinct transitional intermediate cell state prior to AT1 cell differentiation. Currently, the only curative therapy for end-stage lung disease is transplantation, yet there is a paucity of donor lungs. Cell therapy approaches such as lung stem cell transplantation can ameliorate the donor shortage and provide therapeutic avenues beyond surgical interventions. Previous studies demonstrated these transplants can stimulate repair, yet AT2 to AT1 differentiation was halted. Transcriptional profiling revealed Galectin-3 was differentially expressed in the transplanted AT2 cells compared to native AT2 cells. Interestingly, Galectin-3 is specific to the transitional state seen in alveolar repair. However, little is known about the functional consequences of the development and resolution of this novel cell type. Therefore, I used galectin-3 genetic deletion and pharmacological inhibition in alveolar organoid models to determine its function. Pharmacological inhibition of galectin-3 increased expression AT1 cell marker genes in co-cultures and increased organoid size. These results suggest mesenchymal expression of galectin-3 inhibits AT1 differentiation, decreases AT2 proliferation and affects the transitional cell state. When Gal3 knockout AT2 cells were used in co-cultures with Gal3 inhibitor, AT1 marker gene expression further increased as did organoid forming efficiency. This thesis reveals the role of galectin-3 in lung organoid models and potentially demonstrates a novel method to increase AT1 differentiation in cell therapy models and lung repair.