The ability to pattern three-dimensional microscale cultures opens new avenues for examining the effect of nonplanar mechanical environments on mammalian cells and tissues. Our lab has developed a method for generating suspended tissues with spatial control using open microfluidic principles called Suspended Tissue Open Microfluidic Patterning, or STOMP. STOMP utilizes spontaneous capillary flow and capillary pinning to pattern suspended, multi-region tissues. Using similar microfluidic principles as STOMP, we have developed a method to pattern large (cm-scale) models via semi-open microfluidic channels called Suspended Nonplanar and Planar, or SNaP, geometries. I design these devices with computer-aided design, fabricate components on stereolithography 3D printers, pattern devices with standard pipettes, and culture resulting tissues for short- and long-term time periods to model biological scenarios. With the broad statement that human tissue is generally nonplanar in mind, my research focuses on three different geometries of tissue, 1) a sinusoidal wave, 2) a transwell-like mogul, and 3) a multi-region dome, where each nonplanar geometry enables a different biomedical investigation. The sinusoidal wave construct allows us to ask if cells embedded in tissues with varying frequencies of undulation experience changes to cell morphology due to the topology of their environment; the transwell-like mogul enables investigation of cell proliferation of cells grown at or within an air-liquid interface; and the multi-region dome facilitates the study of tissue interfaces where a diseased region of cells meets a healthy region of cells, all within a single contiguous tissue. I am currently exploring these questions through multiple cultures where different device versions and/or multiple cell types are engaged to collect biological readouts which demonstrate SNaP as a translatable platform for the investigation of questions in biomechanics and regenerative medicine.

Less than 10% of drugs successfully transition from preclinical to clinical trials, principally due to the inability of currently used 2-dimensional models to simulate the 3-dimensional structure and function of human tissues. To develop 3D in vitro models of human vasculature for more efficacious screening of anti-atherosclerosis drugs, I created a device for constructing a perfusable tissue containing a lumen by leveraging open microfluidic patterning methods developed by our group: suspended tissue open microfluidic patterning (STOMP). The device can be used to pattern tissue with a hollow luminal structure lined with endothelial cells, which can be perfused via hollow posts the tissue is suspended between. Using surface tension-driven flow, a liquid hydrogel precursor solution flows through the open microfluidic channel and around the two hollow posts. After gelling, the tissue anchors to the post, contracts away from the sides of the microfluidic channel, and the STOMP device is removed. By adding a second STOMP device that can surround the first tissue another cell-laden hydrogel can be patterned around the first tissue, encapsulating it. To form a lumen in cardiac tissue, I will pattern the inner region with human umbilical vein endothelial cells (HUVECs) in an enzymatically degradable polyethylene glycol hydrogel, surrounded by human induced pluripotent stem cell-derived cardiomyocytes in fibrin hydrogel. Enzymatic degradation of the core region will form a cavity through which HUVECs will remodel the cavity walls, forming an endothelial lining. I will assess lining formation by adding fluorescent dextran to cell media being perfused through the device and measuring fluorescence through confocal microscopy in the surrounding region over time, allowing me to evaluate the permeability of the membrane to compare with physiological values. This model can then be used to screen treatments for atherosclerosis to study how drugs interact with cells in a 3D microenvironment. 

Environmental mechanical stress within a biological system is integral to proper cell fate, function and disease. These complex processes are affected by mechanotransduction, or the transfer of mechanical stimuli into biochemical signals. This occurs through the activation of mechanosensor proteins which transduce physical signals to the nucleus, leading to the activation of certain genes and cellular remodeling. Commonly used 2D cell culture techniques fail to replicate these forces, and are thus unable to activate mechanotransductive pathways seen in vivo. We have developed a method to apply physical stresses to 3D tissue models for investigating how these pathways impact functionality of the human physiological microenvironment. Our method was inspired by methodology created previously by our lab known as STOMP (suspended tissue open microfluidic patterning), which uses surface forces to pattern 3D hydrogel-based culture models. We use our method to create a cell-laden hydrogel suspended between two rows of disconnected rungs, referred to as tissue hooks. The hydrogel can then be transferred to a secondary device, where it is stretched to varying degrees, generating mechanical stress on the tissue. We use confocal fluorescent microscopy to observe cellular remodeling and use image analysis techniques and qPCR to quantify the activation of mechanotransduction pathways. While it is important to investigate how static mechanical stress on tissue impacts functionality, the human body is a dynamic environment. We created a system to dynamically stretch tissue cultures to further investigate cellular contractility within 3D tissue models. Instead of a static stretch, the cell-laden hydrogel is patterned to hooks with serpentine-style springs on the side. As the cell embedded hydrogel compacts, it pulls on the springs, allowing us to quantify the contractile forces. We plan to apply these models to study highly contractile tissue, such as skeletal muscle, and subsequent disease pathways shown in mechanotransduction.

The future of clinical research is expanding towards sampling that can be completed from the comfort of a participant's home. Blood samples allow for the collection of ribonucleic acid (RNA), which is relevant for gene sequencing that can track the progression of a disease. However, venous blood draws require trained phlebotomists at a healthcare facility, which may not be readily accessible in some areas. Dried blood spots are an existing remote sampling method, but rapid degradation of RNA and low blood volume can limit the scope of analyses that are possible. Previously, our lab developed homeRNA, which interfaces with the Tasso-SST (Tasso Inc.), a lancet-based device that draws blood from the upper arm. The addition of the engineered, spill-resistant container creates a channel through which participants can draw their own blood, stabilize the blood with RNAlater (Thermo Fisher Scientific), and ship the sample to a laboratory for analysis. The homeRNA+ project improves upon the original homeRNA by integrating a commercially available blood collection tube for better compatibility and doubling the maximum blood collection volume. Feedback from study participants over the United States across all age and race demographics generally find the blood collection process painless and the stabilization easy to perform. We expect samples to also have sufficient RNA integrity and yield for downstream analysis. The project serves a number of nationwide and global collaborators, including academic institutions like New York University and Boston University. I assist in receiving and processing biological samples from remote collection, ensuring proper handling by safely unpackaging, logging, and preserving returned samples in cold storage for future analysis. Additionally, I serve as a study coordinator by meeting with collaborators, manufacturing high volumes of kits in a timely manner, and managing inventories.

Microfluidics has enabled researchers to engineer environments with precisely controlled fluids in submillimeter scale, making it an essential tool in biomedical research. Moreover, the study of fluidic dynamics in both closed and open channels has been a major focus in the field of microfluidics. This study specifically examines capillary flow dynamics in open microfluidic systems where capillary flow refers to the spontaneous flow of liquids in narrow spaces without the assistance of external forces. The Lucas-Washburn-Rideal (LWR) law is commonly used to describe capillary flow dynamics in closed and open channels, microporous media, and threads that assumes a viscous regime in which capillary forces are counterbalanced by friction with the solid channel walls. However, in conditions beyond the viscous domain, inertial forces become significant, leading to an imbalance between wall friction and capillary force (e.g. the inertial regime at the onset of capillary motion). This study proposes a straightforward criterion for identifying inertial effects using the Lucas-Washburn-Rideal (LWR) law. This criterion is derived by analyzing the plot of a characteristic function, F, that is defined as the product of the cross-sectional area at the meniscus location, the travel distance, and the meniscus velocity. To validate this criterion, we present four different examples: open-channel devices with converging and diverging channel configurations, an open channel separated into two daughter channels in a symmetrical or asymmetrical configuration, and a two-phase capillary flow experiment in which pentanol pushes a plug of water in an open channel. These experiments successfully validate the proposed criterion for identifying inertial effects in capillary-driven flow within open channels. Furthermore, we also demonstrate that the Bosanquet equation can serve as an accurate model for open capillary flows in rectangular channels with progressively decreasing cross-sections. This study could impact the design of microfluidic systems that traditionally assume negligible inertial effects.

Microbial volatile organic compounds (mVOCs) are small molecules produced by microorganisms during biosynthetic pathways that easily diffuse through the air, interacting with other nearby organisms despite not physically touching. One method to measure this mVOC communication was the previous iteration of our co-culture device, a bottomless glass vial sealed onto a chip with two separated culture wells, where mVOCs could be released into the headspace. However, spores, a non-mVOC aerosol, also diffused through the headspace, and the device’s structure made it difficult to ensure a fully intact seal. We are developing a device that supports a co-culture that communicates through mVOCs only, uses an intact vial to improve encapsulation, and allows for solid-phase microextraction (SPME) coupled with gas chromatography-mass spectrometry. To test its efficacy, we use the sporulating fungus Aspergillus fumigatus and the bacteria Pseudomonas aeruginosa as model organisms, which are opportunistic pathogens often co-infecting cystic fibrosis patients. We assemble the device by inserting two layers of CNC milled polystyrene platforms containing wells into a vial. We add 1-octen-3-ol or isopentanol, mVOCs produced by A. fumigatus, to the first layer, and P. aeruginosa cultures to the second. The second layer contains polytetrafluoroethylene (PTFE) membranes that only mVOCs can diffuse through. We incubate the vials, plate the P. aeruginosa cultures onto agar, incubate, and observe their growth to assess mVOC communication. We anticipate higher concentrations of mVOCs to inhibit P. aeruginosa growth, demonstrating that the mVOCs interacted with microorganisms in the upper layer. In the future we will co-culture A. fumigatus and P. aeruginosa in the device to study their mVOC interaction, and explore using different biomarkers to determine their effects. This device could be used with other co-infecting pathogenic microorganisms to study their mechanisms and explore therapeutic possibilities.