Exosome-mediated microRNA-497 delivery for anti-cancer therapy in a microfluidic 3D lung cancer model.

Non-small cell lung cancer (NSCLC) is one of the leading causes of death from cancer worldwide. The delivery and controlled regulation of miRNAs via exosomes is known as a potential therapeutic approach in the treatment of cancer. In this study, human cell-derived exosomes were used as delivery vehicles for miRNAs, and we investigated their anti-tumor and anti-angiogenic effects on NSCLCs that were cultured in 2D and 3D microfluidic devices. We demonstrated that exosomes that contained miRNA-497 (miR-497) effectively suppressed tumor growth and the expression of their associated genes, i.e., yes-associated protein 1 (YAP1), hepatoma-derived growth factor (HDGF), cyclin E1 (CCNE1), and vascular endothelial growth factor-A (VEGF-A), in A549 cells. Also, the level of VEGF-A-mediated angiogenic sprouting was decreased drastically in human umbilical vein endothelial cells (HUVECs) cultured in a microfluidic device. To mimic the in vivo-like tumor microenvironment of NSCLC, A549 cells were co-cultured with HUVECs in a single device, and miR-497-loaded exosomes were delivered to both types of cells. As a result, both the tube formation of endothelial cells and the migration of tumor decreased dramatically compared to the control. This indicated that miR-497 has synergistic inhibitory effects that target tumor growth and angiogenesis, so exosome-mediated miRNA therapeutics combined with the microfluidic technology could be a predictive, cost-efficient translational tool for the development of targeted cancer therapy.

Hydrogel-incorporating unit in a well: 3D cell culture for high-throughput analysis.

The microfluidic 3D cell culture system has been an attractive model because it mimics the tissue and disease model, thereby expanding our ability to control the local cellular microenvironment. However, these systems still have limited value as quantitative assay tools due to the difficulties associated with the manipulation and maintenance of microfluidic cells, and their lack of compatibility with the high-throughput screening (HTS) analysis system. In this study, we suggest a microchannel-free, 3D cell culture system that has a hydrogel-incorporating unit integrated with a multi-well plate (24- to 96-well plate), which can provide better reproducibility in biological experiments. This plate was devised considering the design constraints imposed by various cell biology applications as well as by high-throughput analysis where the physical dimensions of the micro-features in the hydrogel-incorporating units were altered. We also demonstrated that the developed plate is potentially applicable to a variety of quantitative biochemical assays for qRT-PCR, Western blotting, and microplate-reader-based assays, such as ELISA, viability assay, and high content-screening (HCS) as well as the co-culture for biological studies. Human neural progenitor cells (hNPCs) that produce pathogenic Aß species for modeling Alzheimer's disease (AD) were three-dimensionally cultured, and the efficacy of the inhibitors of Aß production was assessed by ELISA in order to demonstrate the performance of this plate.

On-Chip Lipid Extraction Using Superabsorbent Polymers for Mass Spectrometry.

Pretreatment of samples is one of the most important steps in analytical methods for efficient and accurate results. Typically, an extraction method used for lipid analysis with mass spectrometry is accompanied by complex liquid-liquid extraction. We have devised a simple, rapid, and efficient lipid extraction method using superabsorbent polymers (SAPs) and developed a high-throughput lipid extraction platform based on a microfluidic system. Since SAPs can rapidly absorb an aqueous solution from a raw sample and convert it into the gel, the lipid extraction process can be remarkably simplified. The hydrophobic lipid components were captured into the fibrous SAP gel and then solubilized and eluted directly into the organic solvent without significant interference by this polymer. The small-scale lipid extraction process minimizes the liquid handling and unnecessary centrifugation steps, thereby enabling the implementation of a SAP-integrated microfluidic lipid extraction platform. The SAP method successfully induced reproducible extraction and high recovery rates (95-100%) compared to the conventional Folch method in several lipid classes. We also demonstrated the feasibility of the SAP method for the analysis of lipids in complex biological samples, such as the brain and liver, as well as Escherichia coli. This small-scale SAP method and its microfluidic platform will open up new possibilities in high-throughput lipidomic research for diagnosing diseases because this new technique saves time, labor, and cost.