BIOFABRICATION OF MICROPHYSIOLOGICAL 3D VASCULARIZED BONE MODELS FOR THE STUDY OF HUMAN PHYSIOLOGY AND DISEASE

Biofabrication is an emerging research field aimed at recapitulating in vitro the architecture and functionality of basic tissue units of the human body. A key aspect of biofabrication is the combination of 3D tissue models with microfabrication technologies (e.g. 3D printing, microfluidics) that allow to create spatially confined microenvironments (e.g. an interface between vasculature and a tissue-niche), apply electro-mechanical stimulations (e.g. compression, shear) and live track the dynamic interactions among different cell types. The design of biofabricated models allows to complement available pre-clinical assays and partially overcome their limitations, including the limited complexity of 2D cultures and the low-throughput of animal experiments. During the keynote I will present examples from the lab expertise in biofabrication describing different classes of multi-scale human 3D bone models for the study of: 1) basic mechanisms of breast cancer cell (BCC) dissemination to bone through blood vessels; 2) interactions between BCCs and an already established metastatic niche including immune cells (e.g. neutrophils, macrophages); 3) mechanisms of drug resistance mediated by the cross-talk between specific bone niches and BCCs. The common element across these models is the presence of perfusable microvascular networks that are generated by co-culturing endothelial cells (ECs) and mural cells through self-assembly. Importantly, vascular cells acquire an organ-specific signature when placed in the bone microenvironment, mimicking organotypic differences observed across vascular beds in vivo. We designed multi-scale bone models that recapitulated the bone microenvironment by including osteoblasts, osteoclasts, vascular cells and macrophages within fibrin or collagen matrices. We demonstrated that the bone microenvironment promoted the extravasation of circulating BCCs and the formation of micrometastases compared to control conditions (e.g. skeletal muscle microenvironment). Moreover, the presence of a bone microenvironment promoted the onset of drug resistance mechanisms when established metastases were exposed to doxorubicin or rapamycin. In particular, we showed that BCCs progressively skewed the polarization of resident macrophages towards an M2 pro-tumoral phenotype. We also observed that naive neutrophils perfused through vascularized bone models selectively killed BCCs. Finally, we designed a multi-analytical approach combining organs-on-a-chip and multi-omic analyses to identify the signature of ECs during aging, which might ultimately affect tissue functionality during physiology and disease. In particular, we observed an increase in vascular permeability during aging associated with compromised cell-cell junctions, potentially affecting the formation of metastases. Overall, we biofabricated multi-scale models of vascularized bone that allowed to mimic the dynamic, patho-physiological cross-talk among blood vessels, bone-resident cells and BCCs.