In vitro three-dimensional (3D) tumour models mimic natural cancer tissue in vivo, bridging the gap between conventional 2D in vitro testing and animal models. Stromal and cancer tissues with extracellular matrix (ECM) can provide a tumour microenvironment (TME) with cell-to-cell and cell-to-ECM interactions. These interactions induce the exchange of biophysical factors, contributing to the progression, metastasis, and drug resistance of cancer. Here, we describe a 3D in vitro lung cancer model cultured in a microfluidic channel that is able to confirm the role and function of various stromal cells in tumourigenesis, thereby representing an in vivo-like TME. We founded that biophysical factors contribute to the role of fibroblast cells in tumour formation, especially, producing a nascent vessel-like tubular structure, resulting in the formation of vascularized tumour tissue. Fibroblast cells altered the gene expression of the cancer cells to enhance metastasis, survival, and angiogenesis. The device could be used for developing and screening anti-cancer drugs through the formation of the same multicellular tumour spheroids under TME interactions. We believe this microfluidic system provides interaction of TME for cancer research by culturing stromal tissue.

The purpose of this study was to demonstrate self‐organizing in vitro multicellular tumor spheroid (MCTS) formation in a microfluidic system and to observe the behavior of MCTSs under controlled microenvironment. The employed microfluidic system was designed for simple and effective formation of MCTSs by generating nutrient and oxygen gradients. The MCTSs were composed of cancer cells, vascular endothelial cells, and type I collagen matrix to mimic the in vivo tumor microenvironment (TME). Cell culture medium was perfused to the microfluidic device loaded with MCTSs by a passive fluidic pump at a constant flow rate. The dose response to an MMPs inhibitor was investigated to demonstrate the effects of biochemical substances. The result of long‐term stability of MCTSs revealed that continuous perfusion of cell culture medium is one of the major factors for the successful MCTS formation. A continuous flow of cell culture medium in the in vitro TME greatly affected both the proliferation of cancer cells in the micro‐wells and the sustainability of the endothelial cell‐layer integrity in the lumen of microfluidic channels. Addition of MMP inhibitor to the cell culture medium improved the stability of the collagen matrix by preventing the detachment and shrinkage of the collagen matrix surrounding the MCTSs. In summary, the present constant flow assisted microfluidic system is highly advantageous for long‐term observation of the MCTS generation, tumorous tissue formation process and drug responses. MCTS formation in a microfluidic system may serve as a potent tool for studying drug screening, tumorigenesis and metastasis.

Various methods for isolation of type I collagen using acids, bases, enzymes, and their combinations have been applied. However, a lack of standardization exists among type I collagens isolated by various approaches. Consequently, in this study, we assessed the influence of acetic acid residue on type I collagen isolated by pepsin-acetic acid treatment, the fabrication of collagen-based porous scaffolds, and the seeded cells on collagen scaffolds. Unlike the isolated collagen dialyzed by deionized water (DDW), collagen dialyzed by 0.5 M acetic acid (DAC) exhibited structural and thermal denaturation. Both DDW- and DAC-based porous scaffolds at all collagen concentrations (0.5, 1 and 2% w/v) showed the high degree of porosity (>98%), and their pore morphologies were comparable at the same concentrations. However, the DDW- and DAC-based collagen scaffolds displayed significant differences in their physical properties (weight, thickness, and volume) and swelling behaviors. In particular, the weight losses induced by mechanical stimulation reflected the high degradation of DAC-collagen scaffolds. In cell culture experiments using adipose-derived stem cells (ADSCs), the characteristics of mesenchymal stem cell (MSC) did not change in both DDW- and DAC-collagen scaffolds for 10 days, although cells proliferated less in the DAC-collagen scaffolds. Our results suggest that the elimination of acetic acid residue from isolated collagen is recommended to produce collagen scaffolds that provide a stable environment for cells and cell therapy-related applications.

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In the tumor microenvironment (TME), the extracellular matrix (ECM) provides a dynamic structure for cell adhesion and cancer cell motility, such as migration and invasion, as well as remodeling. Matrix metalloproteinases (MMPs) promote cancer cell motility, which contributes to inducing drug resistance and thereby acquiring aggressive features. The drug resistance-induced 3D in vitro tumor model can be an effective model for therapeutic strategies for anticancer drugs targeting aggressive cancer cells. Here, we describe highly drug-resistant multicellular tumoroids (MCTs)-ECM tumor grafts under a macroscale dense 3D in vitro model through a combination of numerous MCTs and a collagen matrix. MCTs-ECM tumor grafts promote the high activity of MMP2 and MMP9 compared to general MCTs and induced cancer cell motility. Then, after the administration of anticancer drugs, the tumor grafts show increased drug resistance, with both the sporadic distribution of necrotic cells and the reduction of apoptotic portions, by activating cancer cell motility. MCTs-ECM tumor graft could be useful as a macroscale tumor graft model for inducing drug resistance by activating cancer cell motility and evaluating the efficacy of anticancer drugs targeting cancer with aggressive features.