Project description:Cell and protein arrays have demonstrated remarkable utility in the high-throughput evaluation of biological responses; however, they lack the complexity of native tissue and organs and cannot replicate the environment of mature normal or diseased tissues. Here, we describe the development of tissue and organ extracellular matrix (ECM) arrays for screening biological outputs and systems analysis. Tissues and organs were chemically and mechanically processed and then formulated as particles without any enzymatic breakdown. The tissue ECM particles were spotted as two-dimensional tissue ECM arrays or incorporated with cells to generate three-dimensional cell-ECM microtissue arrays, and these methods were compatible with tissues prepared with varied processing techniques. The physical and biochemical properties, including the proteomic composition, of the tissue ECM arrays were characterized and compared with native tissues. The 2D arrays were validated through quantitative analysis of tissue-specific biological outputs of cell matrix production, cell adhesion and proliferation, and cell shape following culture with stem cells, cancer cells, and macrophages. Tissue-specific stem cell osteogenic differentiation on the arrays was comparable between 2D and 3D ECM environments, and revealed lung as an unexpected promoter of osteogenesis. Further gene ontology analysis of lung tissue ECM proteomics showed enrichment for families of proteins associated with skeletal development and osteogenesis. Stem cell biological outputs, along with cancer line adhesion and macrophage shape, were correlated with tissue proteomics, and network analysis identified several protein determinants of cell function. Our methodology enables broad screening of tissue and organ ECMs to connect tissue-specific composition with biological responses via systems analysis, providing a new resource for research and translation.
Project description:Due to the lack of a precise in vitro model that can mimic the nature microenvironment in osteosarcoma, the understanding of its resistance to chemical drugs remains limited. Here, we report a novel three-dimensional model of osteosarcoma constructed by seeding tumor cells (MG-63 and MNNG/HOS Cl #5) within in demineralized bone matrix scaffolds. Demineralized bone matrix scaffolds retain the original components of the natural bone matrix (hydroxyapatite and collagen type I), and possess good biocompatibility allowing osteosarcoma cells to proliferate and aggregate into clusters within the pores. Growing within the scaffold conferred elevated resistance to doxorubicin on MG-63 and MNNG/HOS Cl #5 cell lines as compared with two-dimensional cultures. Transcriptomic analysis showed an increased enrichment for drug resistance genes along with enhanced glutamine metabolism in osteosarcoma cells in demineralized bone matrix scaffolds. Inhibition of glutamine metabolism resulted a decrease in drug resistance of osteosarcoma, which could be restored by α-ketoglutarate supplementation. Overall, our study suggests that microenvironmental cues in demineralized bone matrix scaffolds can enhance osteosarcoma drug responses and that targeting glutamine metabolism may be a strategy for treating osteosarcoma drug resistance.
