The use of nanoparticles in medical applications is highly anticipated, and at the same time little is known about how these nanoparticles affect human tissues. Here we have simulated the oral uptake of 50 nm carboxylated polystyrene nanoparticles with a microscale, body-on-a-chip system (also referred to as multi-tissue microphysiological system or micro Cell Culture Analog). Using this system, we combined in vitro models of the human intestinal epithelium, represented by a co-culture of enterocytes (Caco-2) and mucin-producing (HT29-MTX) cells, and the liver, represented by HepG2/C3A cells, within one microfluidic device. The device also contained chambers that together represented all other organs of the human body. Measuring the transport of 50 nm carboxylated polystyrene nanoparticles across the Caco-2/HT29-MTX co-culture, we have found that this multi-cell layer presents an effective barrier to 90.5 ± 2.9% of the nanoparticles. Further, our simulation suggests that a larger fraction of the 9.5 ± 2.9% of nanoparticles that travelled across the Caco-2/HT29-MTX cell layer were not large nanoparticle aggregates, but primarily single nanoparticles and small aggregates. After crossing the GI tract epithelium, nanoparticles that were administered in high doses estimated in terms of possible daily human consumption (240 and 480 × 1011 nanoparticles/mL) induced the release of aspartate aminotransferase (AST), an intracellular enzyme of the liver that indicates liver cell injury. Using the GI ‘tract – liver – other tissue’ system allowed us to observe compounding effects and detect liver tissue injury at lower nanoparticle concentrations than expected from experiments with liver tissue only. Our results indicate that body-on-a-chip devices are highly relevant in vitro models for evaluating nanoparticle interactions with human tissues.

The use of engineered nanoparticles in food and pharmaceuticals is expected to increase, but the impact of chronic oral exposure to nanoparticles on human health remains unknown. Here, we show that chronic and acute oral exposure to polystyrene nanoparticles can influence iron uptake and iron transport in an in vitro model of the intestinal epithelium and an in vivo chicken intestinal loop model. Intestinal cells that are exposed to high doses of nanoparticles showed increased iron transport due to nanoparticle disruption of the cell membrane. Chickens acutely exposed to carboxylated particles (50 nm in diameter) had a lower iron absorption than unexposed or chronically exposed birds. Chronic exposure caused remodelling of the intestinal villi, which increased the surface area available for iron absorption. The agreement between the in vitro and in vivo results suggests that our in vitro intestinal epithelium model is potentially useful for toxicology studies.

The lining of the gastrointestinal (GI) tract is the largest surface exposed to the external environment in the human body. One of the main functions of the small intestine is absorption, and intestinal absorption is a route used by essential nutrients, chemicals, and pharmaceuticals to enter the systemic circulation. Understanding the effects of digestion on a drug or chemical, how compounds interact with and are absorbed through the small intestinal epithelium, and how these compounds affect the rest of the body is critical for toxicological evaluation. Our goal is to create physiologically realistic in vitro models of the human GI tract that provide rapid, inexpensive, and accurate predictions of the body's response to orally delivered drugs and chemicals. Our group has developed an in vitro microscale cell culture analog (microCCA) of the GI tract that includes digestion, a mucus layer, and physiologically realistic cell populations. The GI tract microCCA, coupled with a multi-chamber silicon microCCA representing the systemic circulation, is described and challenged with acetaminophen. Proof of concept experiments showed that acetaminophen passes through and is metabolized by the in vitro intestinal epithelium and is further metabolized by liver cells, resulting in liver cell toxicity in a dose-dependent manner. The microCCA response is also consistent with in vivo measurements in mice. The system should be broadly useful for studies on orally delivered drugs or ingestion of chemicals with potential toxicity.

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Objective Inflammatory activation of valve endothelium is an early phase of aortic valve disease pathogenesis, but subsequent mechanisms are poorly understood. Adult valve endothelial cells retain the developmental ability to undergo endothelial to mesenchymal transformation (EndMT), but a biological role has not been established. Here we test whether and how inflammatory cytokines (TNF-α and IL-6) regulate EndMT in embryonic and adult valve endothelium. Methods and Results Using in vitro 3D collagen gel culture assays with primary cells, we determined that IL-6 and TNF-α induce EndMT and cell invasion in dose dependent manners. Inflammatory-EndMT occurred through an Akt/NFκB-dependent pathway in both adult and embryonic stages. In embryonic valves, inflammatory-EndMT required canonical TGFβ signaling through Alk2/5 to drive EndMT. In adult valve endothelium, however, inflammatory-induced EndMT still occurred when Alk2/5 signaling was blocked. Inflammatory receptor gene expression was significantly upregulated in vivo during embryonic valve maturation. Endothelial-derived mesenchymal cells expressing activated NFκB were found distal to calcific lesions in diseased human aortic valves. Conclusions Inflammatory cytokine induced EndMT in valve endothelium is present in both embryonic and adult stages, acting through Akt/NFκB but differently utilizing TGFβ signaling. Molecular signatures of valve EndMT may be important diagnostic and therapeutic targets in early valve disease.

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Ingestion of titanium dioxide (TiO2) nanoparticles from products such as agricultural chemicals, processed food, and nutritional supplements is nearly unavoidable. The gastrointestinal tract serves as a critical interface between the body and the external environment, and is the site of essential nutrient absorption. The goal of this study was to examine the effects of ingesting the 30 nm TiO2 nanoparticles with an in vitro cell culture model of the small intestinal epithelium, and to determine how acute or chronic exposure to nano-TiO2 influences intestinal barrier function, reactive oxygen species generation, proinflammatory signaling, nutrient absorption (iron, zinc, fatty acids), and brush border membrane enzyme function (intestinal alkaline phosphatase). A Caco-2/HT29-MTX cell culture model was exposed to physiologically relevant doses of TiO2 nanoparticles for acute (four hours) or chronic (five days) time periods. Exposure to TiO2 nanoparticles significantly decreased intestinal barrier function following chronic exposure. Reactive oxygen species (ROS) generation, proinflammatory signaling, and intestinal alkaline phosphatase activity all showed increases in response to nano-TiO2. Iron, zinc, and fatty acid transport were significantly decreased following exposure to TiO2 nanoparticles. This is because nanoparticle exposure induced a decrease in absorptive microvilli in the intestinal epithelial cells. Nutrient transporter protein gene expression was also altered, suggesting that cells are working to regulate the transport mechanisms disturbed by nanoparticle ingestion. Overall, these results show that intestinal epithelial cells are affected at a functional level by physiologically relevant exposure to nanoparticles commonly ingested from food.

Understanding the role of mechanical forces on cell behavior is critical for tissue engineering, regenerative medicine, and disease initiation studies. Current hemodynamic bioreactors are largely limited to 2D substrates or the application of general flow conditions at a tissue level, which eliminates the investigation of some essential physiological and pathological responses. One example is the mesenchymal transformation of endothelial cells in response to shear stress. Endothelial to mesenchymal transformation (EndMT) is a valve morphogenic mechanism associated with aortic valve disease initiation. The aortic valve experiences oscillatory shear on the disease-susceptible fibrosa, and the role of hemodynamics on adult EndMT is unknown. The goal of this work was to develop and characterize a microfluidic bioreactor that applies physiologically relevant laminar or oscillatory shear stresses to endothelial cells and permits the quantitative analysis of 3D cell-extracellular matrix (ECM) interactions. In this study, porcine aortic valve endothelial cells were seeded onto 3D collagen I gels and exposed to different magnitudes of steady or oscillatory shear stress for 48 hours. Cells elongated and aligned perpendicular to laminar, but not oscillatory shear. Low steady shear stress (2 dyne/cm2) and oscillatory shear stress upregulated EndMT- (ACTA2, Snail, TGFB1) and inflammation- (ICAM1, NFKB1) related gene expression, EndMT-related (αSMA) protein expression, and matrix invasion when compared with static controls or cells exposed to high steady shear (10 and 20 dyne/cm2). Our system enables direct testing of the role of shear stress on endothelial cell mesenchymal transformation in a dynamic, 3D environment and shows that hemodynamics regulate EndMT in adult valve endothelial cells.