An integrative alginate-based 3D in vitro model to explore epithelial-stromal cell dynamics in the breast tumor microenvironment.

The tumor microenvironment (TME) orchestrates cellular and extracellular matrix (ECM) interactions, playing a key role in tumorigenesis, tumor growth, and metastization. Investigating the interplay between stromal-epithelial cells within the TME is paramount for understanding cancer mechanisms but demands reliable biological models. 3D-models have emerged as powerful in vitro tools, but many fall short in replicating cell-cell/cell-matrix interactions. This study introduces a novel hybrid 3D-model of the breast TME, combining epithelial cells, cancer-associated fibroblasts (CAFs), and their ECM. To build the stromal compartment, porous 3D-printed alginate scaffolds were seeded with CAFs, which proliferated and produced ECM. The pores were infused with oxidized peptide-modified alginate hydrogel laden with MCF10A cells, forming the parenchymal compartment. The hybrid system supported epithelial morphogenesis into acini surrounded by fibroblasts and ECM, and could be readily solubilized to recover cells, their matrix, and sequestered soluble factors. Proteome profiling of the retrieved ECM showed upregulation of proteins associated with matrix assembly/remodeling, epithelial-to-mesenchymal transition (EMT), and cancer. The TME-like microenvironment induced a partial EMT in MCF10A cells, generating a hybrid population with epithelial and mesenchymal features, characteristic of aggressive phenotypes. Our model provided new insights into epithelial-stromal interactions within the TME, offering a valuable tool for cancer research in a physiologically-relevant 3D setting.

Metabolic Oscillations in Co-Cultures of Hepatocytes and Mesenchymal Stem Cells: Effects of Seeding Arrangement and Culture Mixing.

In vitro assembly of functional liver tissue is a prerequisite for the transplantation of tissue-engineered livers. There is an increasing demand for in vitro models that replicate complex events occurring in the liver. However, tissue engineering of implantable liver systems is currently limited by the difficulty of assembling three dimensional hepatocyte cultures of a useful size, while maintaining full cell viability. Recent reports have demonstrated that bone marrow mesenchymal stem cells (BM-MSCs) can provide a number of cues promoting hepatocyte growth and development. In this study, the effects of BM-MSCs co-culture on hepatocyte metabolism were evaluated as a function of scaffold seeding arrangement. BM-MSCs were co-cultured with hepatocytes in porous chitosan-heparin scaffolds using several seeding arrangements. The seeded scaffolds were subjected to orbital shaking to enhance mass transfer. BM-MSC-hepatocyte co-cultures exhibited higher rates of hepatocyte-specific functions, compared to hepatocyte-only cultures, regardless of the seeding arrangement. Cells formed smaller-compact spheroids in the heterotypic systems compared to mono-cultures of hepatocytes only. The spheroids exhibited reduction in size with time in all conditions except for the condition where BM-MSCs were seeded one day after seeding hepatocytes. In this condition, spheroids increased in size due to BM-MSC proliferation. Spheroid size reduction was hypothesized to be the result of cyclic shear stresses generated by the orbital shaking. Furthermore, results suggested that BM-MSC seeding onto preformed hepatocyte spheroids provide a degree of shear-protection and trophic stimuli. Overall, the results indicate that co-culturing hepatocytes with BM-MSCs enhanced their metabolic functions for the first week of culture. J. Cell. Biochem. 118: 3003-3015, 2017. © 2017 Wiley Periodicals, Inc.