Three-dimensional Microphysiological Systems Research Articles

Comprehensive landscape of tumor angiogenesis via integrating RNA sequencing and three-dimensional microphysiological system.

e16058 Background: The prognosis of gastric cancer (GC) patients with peritoneal metastasis with malignant ascites is very poor. We developed an in vitro tumor angiogenesis platform “TumorAngioChip” to reproduce tumor angiogenesis in 45 gastric cancer patients and mapping their angiogenesis characteristics. This platform, which can predict the features of tumor angiogenesis and drug responsiveness in patient’s samples, opens up potential for a variety of combination therapies, including ramucirumab. Methods: We built a PDC library for the study based on samples of 45 GC patients receiving treatment at Samsung Medical Center. For all patient samples, whole exome and transcriptome sequencing was performed for subtype clustering. TumorAngioChip was fabricated by design prototyping through 3D printing and mass production through injection molding. Patient-derived tumor angiogenesis reconstructed on TumorAngioChip was analyzed using an algorithm for morphological image processing of 3D images obtained through confocal microscopy. Results: Of 45 GC patients, we investigated TumorAngioChip analysis and transcriptome sequencing to classify them as KDR-High (N = 24) and KDR-Low (N = 21) ( P = 4.481e-08, Wilcoxon signed-rank test). In the TumorAngioChip–patient mapping analysis, we clearly demonstrated that GC patients with high KDR level regardless to their GC TCGA subtype had high index of tumor-induced angiogenesis and cancer invasiveness measured by TumorAngioChip ( KDR-High vs. KDR-low group; mean sprouting length: 1.455 × 103 vs. 0.946 × 103, respectively, P < 0.0001; vessel density: 3.470 vs. 2.637, respectively, P < 0.0001; sprouts number: 219 vs. 111, respectively, P < 0.0001; vessel total length: 4.931 × 104 vs. 3.919 × 104, respectively, P = 0.0011). Conclusions: This study is the first attempt to elucidate cancer angiogenesis in large-scale GC patients via a microfluidic-based in vitro system. By integrating transcriptome sequencing and TumorAngioChip-based 3D morphological image processing, we demonstrated tumor angiogenesis mapping for GC patients and further revealed the benefits of ramucirumab in individual patients. The application of TumorAngioChip shows its potential as a novel preclinical drug screening platform to identify effective angiogenesis inhibitors reflecting TME for the first time.

Adipose-derived cells: building blocks of three-dimensional microphysiological systems.

ABSTRACTMicrophysiological systems (MPS) created with human-derived cells and biomaterial scaffolds offer a potential in vitro alternative to in vivo animal models. The adoption of three-dimensional MPS models has economic, ethical, regulatory, and scientific implications for the fields of regenerative medicine, metabolism/obesity, oncology, and pharmaceutical drug discovery. Key opinion leaders acknowledge that MPS tools are uniquely positioned to aid in the objective to reduce, refine, and eventually replace animal experimentation while improving the accuracy of the finding’s clinical translation. Adipose tissue has proven to be an accessible and available source of human-derived stromal vascular fraction (SVF) cells, a heterogeneous population available at point of care, and adipose-derived stromal/stem cells, a relatively homogeneous population requiring plastic adherence and culture expansion of the SVF cells. The adipose-derived stromal/stem cells or SVF cells, in combination with human tissue or synthetic biomaterial scaffolds, can be maintained for extended culture periods as three-dimensional MPS models under angiogenic, stromal, adipogenic, or osteogenic conditions. This review highlights recent literature relating to the versatile use of adipose-derived cells as fundamental components of three-dimensional MPS models for discovery research and development. In this context, it compares the merits and limitations of the adipose-derived stromal/stem cells relative to SVF cell models and considers the likely directions that this emerging field of scientific discovery will take in the near future.

A Microphysiological System for Studying Nonalcoholic Steatohepatitis.

Nonalcoholic steatohepatitis (NASH) is the most severe form of nonalcoholic fatty liver disease (NAFLD), which to date has no approved drug treatments. There is an urgent need for better understanding of the genetic and molecular pathways that underlie NAFLD/NASH, and currently available preclinical models, be they in vivo or in vitro, do not fully represent key aspects of the human disease state. We have developed a human in vitro co‐culture NASH model using primary human hepatocytes, Kupffer cells and hepatic stellate cells, which are cultured together as microtissues in a perfused three‐dimensional microphysiological system (MPS). The microtissues were cultured in medium containing free fatty acids for at least 2 weeks, to induce a NASH‐like phenotype. The co‐culture microtissues within the MPS display a NASH‐like phenotype, showing key features of the disease including hepatic fat accumulation, the production of an inflammatory milieu, and the expression of profibrotic markers. Addition of lipopolysaccharide resulted in a more pro‐inflammatory milieu. In the model, obeticholic acid ameliorated the NASH phenotype. Microtissues were formed from both wild‐type and patatin‐like phospholipase domain containing 3 (PNPLA3) I148M mutant hepatic stellate cells. Stellate cells carrying the mutation enhanced the overall disease state of the model and in particular produced a more pro‐inflammatory milieu. Conclusion: The MPS model displays a phenotype akin to advanced NAFLD or NASH and has utility as a tool for exploring mechanisms underlying the disease. Furthermore, we demonstrate that in co‐culture the PNPLA3 I148M mutation alone can cause hepatic stellate cells to enhance the overall NASH disease phenotype.

Human Induced Pluripotent Stem Cell-Derived Endothelial Cells for Three-Dimensional Microphysiological Systems.

Microphysiological systems (MPS), or "organ-on-a-chip" platforms, aim to recapitulate in vivo physiology using small-scale in vitro tissue models of human physiology. While significant efforts have been made to create vascularized tissues, most reports utilize primary endothelial cells that hinder reproducibility. In this study, we report the use of human induced pluripotent stem cell-derived endothelial cells (iPS-ECs) in developing three-dimensional (3D) microvascular networks. We established a CDH5-mCherry reporter iPS cell line, which expresses the vascular endothelial (VE)-cadherin fused to mCherry. The iPS-ECs demonstrate physiological functions characteristic of primary endothelial cells in a series of in vitro assays, including permeability, response to shear stress, and the expression of endothelial markers (CD31, von Willibrand factor, and endothelial nitric oxide synthase). The iPS-ECs form stable, perfusable microvessels over the course of 14 days when cultured within 3D microfluidic devices. We also demonstrate that inhibition of TGF-β signaling improves vascular network formation by the iPS-ECs. We conclude that iPS-ECs can be a source of endothelial cells in MPS providing opportunities for human disease modeling and improving the reproducibility of 3D vascular networks.

A strategy for integrating essential three-dimensional microphysiological systems of human organs for realistic anticancer drug screening.

Cancer is one of the leading causes of morbidity and mortality around the world. Despite some success, traditional anticancer drugs developed to reduce tumor growth face important limitations primarily due to undesirable bone marrow and cardiovascular toxicity. Many drugs fail in clinical development after showing promise in preclinical trials, suggesting that the available in vitro and animal models are poor predictors of drug efficacy and toxicity in humans. Thus, novel models that more accurately mimic the biology of human organs are necessary for high-throughput drug screening. Three-dimensional (3D) microphysiological systems can utilize induced pluripotent stem cell technology, tissue engineering, and microfabrication techniques to develop tissue models of human tumors, cardiac muscle, and bone marrow on the order of 1 mm(3) in size. A functional network of human capillaries and microvessels to overcome diffusion limitations in nutrient delivery and waste removal can also nourish the 3D microphysiological tissues. Importantly, the 3D microphysiological tissues are grown on optically clear platforms that offer non-invasive and non-destructive image acquisition with subcellular resolution in real time. Such systems offer a new paradigm for high-throughput drug screening and will significantly improve the efficiency of identifying new drugs for cancer treatment that minimize cardiac and bone marrow toxicity.