Understanding blood-brain barrier responses to inflammatory stimulation (such as lipopolysaccharide mimicking a systemic infection or a cytokine cocktail that could be the result of local or systemic inflammation) is essential to understanding the effect of inflammatory stimulation on the brain. It is through the filter of the blood-brain barrier that the brain responds to outside influences, and the blood-brain barrier is a critical point of failure in neuroinflammation. It is important to note that this interaction is not a static response, but one that evolves over time. While current models have provided invaluable information regarding the interaction between cytokine stimulation, the blood-brain barrier, and the brain, these approaches—whether in vivo or in vitro—have often been only snapshots of this complex web of interactions. We utilize new advances in microfluidics, organs-on-chips, and metabolomics to examine the complex relationship of inflammation and its effects on blood-brain barrier function ex vivo and the metabolic consequences of these responses and repair mechanisms. In this study, we pair a novel dual-chamber, organ-on-chip microfluidic device, the NeuroVascular Unit, with small-volume cytokine detection and mass spectrometry analysis to investigate how the blood-brain barrier responds to two different but overlapping drivers of neuroinflammation, lipopolysaccharide and a cytokine cocktail of IL-1β, TNF-α, and MCP1,2. In this study, we show that (1) during initial exposure to lipopolysaccharide, the blood-brain barrier is compromised as expected, with increased diffusion and reduced presence of tight junctions, but that over time, the barrier is capable of at least partial recovery; (2) a cytokine cocktail also contributes to a loss of barrier function; (3) from this time-dependent cytokine activation, metabolic signature profiles can be obtained for both the brain and vascular sides of the blood-brain barrier model; and (4) collectively, we can use metabolite analysis to identify critical pathways in inflammatory response. Taken together, these findings present new data that allow us to study the initial effects of inflammatory stimulation on blood-brain barrier disruption, cytokine activation, and metabolic pathway changes that drive the response and recovery of the barrier during continued inflammatory exposure.

The sophistication and success of recently reported microfabricated organs-on-chips and human organ constructs have made it possible to design scaled and interconnected organ systems that may significantly augment the current drug development pipeline and lead to advances in systems biology. Physiologically realistic live microHuman (μHu) and milliHuman (mHu) systems operating for weeks to months present exciting and important engineering challenges such as determining the appropriate size for each organ to ensure appropriate relative organ functional activity, achieving appropriate cell density, providing the requisite universal perfusion media, sensing the breadth of physiological responses, and maintaining stable control of the entire system, while maintaining fluid scaling that consists of ~5 mL for the mHu and ~5 μL for the μHu. We believe that successful mHu and μHu systems for drug development and systems biology will require low-volume microdevices that support chemical signaling, microfabricated pumps, valves and microformulators, automated optical microscopy, electrochemical sensors for rapid metabolic assessment, ion mobility-mass spectrometry for real-time molecular analysis, advanced bioinformatics, and machine learning algorithms for automated model inference and integrated electronic control. Toward this goal, we are building functional prototype components and are working toward top-down system integration.

We present a simple apparatus for improved surface modification of polydimethylsiloxane (PDMS) microfluidic devices. A single treatment chamber for plasma activation and chemical/physical vapor deposition steps minimizes the time-dependent degradation of surface activation that is inherent in multi-chamber techniques. Contamination and deposition irregularities are also minimized by conducting plasma activation and treatment phases in the same vacuum environment. An inductively coupled plasma driver allows for interchangeable treatment chambers. Atomic force microscopy confirms that silane deposition on PDMS gives much better surface quality than standard deposition methods, which yield a higher local roughness and pronounced irregularities in the surface.

The implementation of three-dimensional tissue engineering concurrently with stem cell technology holds great promise for in vitro research in pharmacology and toxicology and modeling cardiac diseases, particularly for rare genetic and pediatric diseases for which animal models, immortal cell lines, and biopsy samples are unavailable. It also allows for a rapid assessment of phenotype–genotype relationships and tissue response to pharmacological manipulation. Mutations in the TSC1 and TSC2 genes lead to dysfunctional mTOR signaling and cause tuberous sclerosis complex (TSC), a genetic disorder that affects multiple organ systems, principally the brain, heart, skin, and kidneys. Here we differentiated healthy (CC3) and tuberous sclerosis (TSP8-15) human induced pluripotent stem cells (hiPSCs) into cardiomyocytes to create engineered cardiac tissue constructs (ECTCs). We investigated and compared their mechano-elastic properties and gene expression and assessed the effects of rapamycin, a potent inhibitor of the mechanistic target of rapamycin (mTOR). The TSP8-15 ECTCs had increased chronotropy compared to healthy ECTCs. Rapamycin induced positive inotropic and chronotropic effects (i.e., increased contractility and beating frequency, respectively) in the CC3 ECTCs but did not cause significant changes in the TSP8-15 ECTCs. A differential gene expression analysis revealed 926 up- and 439 down-regulated genes in the TSP8-15 ECTCs compared to their healthy counterparts. The application of rapamycin initiated the differential expression of 101 and 31 genes in the CC3 and TSP8-15 ECTCs, respectively. A gene ontology analysis showed that in the CC3 ECTCs, the positive inotropic and chronotropic effects of rapamycin correlated with positively regulated biological processes, which were primarily related to the metabolism of lipids and fatty and amino acids, and with negatively regulated processes, which were predominantly associated with cell proliferation and muscle and tissue development. In conclusion, this study describes for the first time an in vitro TSC cardiac tissue model, illustrates the response of normal and TSC ECTCs to rapamycin, and provides new insights into the mechanisms of TSC.