Human gastric extracellular matrix hydrogels maintain organoid growth and reduce gene expression associated with stemness and inflammation

BACKGROUND: Human gastric organoids (HGOs) are 3-D cultures of primary gastric epithelial cells that serve as in vitro models for gastric epithelial physiology, development, and disease. The extracellular matrix material that organoids are maintained in provides crucial signals to the epithelial cells, regulating their function and growth. We here describe the application of human gastric ECM (hgECM) for HGO cultures, to provide a tissue- and species-specific growth matrix. Methods: We prepared hgECM from human gastric tissue by decellularization and then compared HGO growth, differentiation, and function in hgECM and in the traditionally used Matrigel. The composition, structure, and rheological properties of the hgECM were determined using tandem mass spectrometry, scanning electron microscopy, and rheometry. Transcriptional profiles of HGOs grown in hgECM and Matrigel were compared using single cell RNA sequencing (scRNAseq). Results: Proteome analysis showed that hgECM was rich in collagens, whereas Matrigel was predominantly composed of glycoproteins such as laminin. The structure of hgECM was fibrous, befitting its collagen composition, while Matrigel was porous. The viscosity and stiffness of hgECM were found to be lower than Matrigel. HgECM supported robust growth of HGOs. ScRNAseq showed that cell type composition of HGOs in each ECM was comparable. However, HGOs in hgECM had reduced expression of genes associated with stem cells, epithelial-mesenchymal transition, stress, and inflammation compared to HGO grown in Matrigel. Conversely, HGO culture in hgECM increased expression of gastric tissue-specific genes and of genes associated with growth and metabolism. Conclusions: HgECM hydrogels were highly compatible with HGO culture, and improved homeostatic functions of HGOs, including growth, differentiation, and reduced inflammation. HgECM may serve as a tissue- and species-specific growth matrix for HGO culture to improve the physiological relevance of organoid models. .