Bone-on-leaf-chip for the study of lung cancer bone metastasis.

Organ-on-a-chip systems provide invaluable preclinical insights into disease simulation, mechanism investigation and drug screening. By closely simulating the physiological conditions of human organs, these platforms enhance our understanding of complex biological processes. Here, we applied a leaf vein microfluidic chip as a controllable, endothelialized platform to investigate how hierarchical flow distribution and uniform shear influence tumor cell migration and behavior within a bone-mimetic microenvironment as a model demonstration. The hierarchical leaf vein architecture, resembling mammalian blood vessels, enables mechanistic studies of spatial distribution and migration under physiologically relevant conditions. Additionally, the system incorporates specialized chambers embedded with 3D hydrogel containing human umbilical vein endothelial cells (HUVECs) and bone stromal cells (HS-5) as the dormancy niche, and HUVECs and osteoclast precursor cells (THP-1) as the "vicious cycle" niche. These chambers serve as a demonstration of bone-mimetic units for examining specific microenvironmental responses. These bone microenvironments were modified by conditioned medium (CM) from primary tumor cells, facilitating their roles as the bone pre-metastatic niche. Cell morphology of lung cancer cells (A549) was observed throughout the dynamic culture process. Perfused medium and hydrogels were harvested to investigate the potential mechanisms. For the dormancy niche, the upregulation of angiogenin, MIP-3α, Wnt-5a, and TGF-β2 and the downregulation of CCL7 indicated that tumor-secreted factors may reactivate dormant tumor cells by activating angiogenesis, pro-inflammatory, and epithelial-mesenchymal transition related pathways. These changes in OPN and BMP-1 expression suggested potential involvement in bone microenvironment remodeling which were inferred from cytokine and gene expression profiles. For the "vicious cycle" niche, the upregulation of CCL5, CXCL5 and VCAM-1 may be associated with the recruitment of leukocytes and promotion of tumor invasion, based on cytokine profiling. These cytokines can serve as potential biomarkers for assessing disease progression or providing a basis for developing new targeted therapies. Taken together, the successful construction and application of this leaf vein chip establish a versatile, mechanistically tractable platform for future drug screening, pathological analysis, and microenvironment-targeted strategies relevant to bone metastasis.