Jonghyun Oh

Microfluidics-Assisted Fabrication of Gelatin-Silica Core–Shell Microgels for Injectable Tissue Constructs

Microfabrication technology provides a highly versatile platform for engineering hydrogels used in biomedical applications with high-resolution control and injectability. Herein, we present a strategy of microfluidics-assisted fabrication photo-cross-linkable gelatin microgels, coupled with providing protective silica hydrogel layer on the microgel surface to ultimately generate gelatin-silica core–shell microgels for applications as in vitro cell culture platform and injectable tissue constructs. A microfluidic device having flow-focusing channel geometry was utilized to generate droplets containing methacrylated gelatin (GelMA), followed by a photo-cross-linking step to synthesize GelMA microgels. The size of the microgels could easily be controlled by varying the ratio of flow rates of aqueous and oil phases. Then, the GelMA microgels were used as in vitro cell culture platform to grow cardiac side population cells on the microgel surface. The cells readily adhered on the microgel surface and proliferated over time while maintaining high viability (∼90%). The cells on the microgels were also able to migrate to their surrounding area. In addition, the microgels eventually degraded over time. These results demonstrate that cell-seeded GelMA microgels have a great potential as injectable tissue constructs. Furthermore, we demonstrated that coating the cells on GelMA microgels with biocompatible and biodegradable silica hydrogels via sol–gel method provided significant protection against oxidative stress which is often encountered during and after injection into host tissues, and detrimental to the cells. Overall, the microfluidic approach to generate cell-adhesive microgel core, coupled with silica hydrogels as a protective shell, will be highly useful as a cell culture platform to generate a wide range of injectable tissue constructs.

Siphon-driven microfluidic passive pump with a yarn flow resistance controller

Precise control of media delivery to cells in microfluidic systems in a simple and efficient manner is a challenge for a number of cell-based applications. Conventional syringe pumps can deliver culture media into microfluidic devices at precisely controlled flow rates, but they are bulky and require a power source. On the other hand, passive microflow-generating systems cannot maintain continuous, controllable and long-term delivery of media. We have developed an on-chip microflow control technology that combines flow rate control and passive, long-term delivery of media to microwell tissue culture chambers. Here, a passive flow is initiated using the siphon effect and a yarn flow resistor is used to regulate the flow rate in the microchannel. Using the yarn flow resistor, the medium flow rate into the microfluidic cell culture system is made adjustable to a few hundred microliters per hour. To evaluate the effects of controlled flow on microfluidic cell culture properties (feasibility test), we measured the cell alignment and cytoskeletal arrangement of endothelial cells cultured in a microwell array inside the microfluidic channel.

High‐throughput investigation of endothelial‐to‐mesenchymal transformation (EndMT) with combinatorial cellular microarrays

In the developing heart, a specific subset of endocardium undergoes an endothelial‐to‐mesenchymal transformation (EndMT) thus forming nascent valve leaflets. Extracellular matrix (ECM) proteins and growth factors (GFs) play important roles in regulating EndMT but the combinatorial effect of GFs with ECM proteins is less well understood. Here we use microscale engineering techniques to create single, binary, and tertiary component microenvironments to investigate the combinatorial effects of ECM proteins and GFs on the attachment and transformation of adult ovine mitral valve endothelial cells to a mesenchymal phenotype. With the combinatorial microenvironment microarrays, we utilized 60 different combinations of ECM proteins (Fibronectin, Collagen I, II, IV, Laminin) and GFs (TGF‐β1, bFGF, VEGF) and were able to identify new microenvironmental conditions capable of modulating EndMT in MVECs. Experimental results indicated that TGF‐β1 significantly upregulated the EndMT while either bFGF or VEGF downregulated EndMT process markedly. Also, ECM proteins could influence both the attachment of MVECs and the response of MVECs to GFs. In terms of attachment, fibronectin is significantly better for the adhesion of MVECs among the five tested proteins. Overall collagen IV and fibronectin appeared to play important roles in promoting EndMT process. Great consistency between macroscale and microarrayed experiments and present studies demonstrates that high‐throughput cellular microarrays are a promising approach to study the regulation of EndMT in valvular endothelium. Biotechnol. Bioeng. 2016;113: 1403–1412.

Interfacial Compression-Dependent Merging of Two Miscible Microdroplets in an Asymmetric Cross-Junction for In Situ Microgel Formation

Controlling the merging of different microdroplets in a microfluidics system could generate a multitude of complex droplets because of their inherent surface tension, but poses a significant challenge because of their high surface tension. Here, a novel microfluidic merging technique is introduced using an asymmetric cross-junction geometry which increases the interfacial compression between two microdroplets. Microdroplets of two viscous polymer solutions, oxidized dextran (ODX) and N-carboxyethyl chitosan (N-CEC), which can undergo a crosslinking reaction via Schiff base formation, are allowed to merge at the asymmetric cross-junction without the assistance of additional merging schemes. The N-CEC and ODX microdroplets being formed at their orifices contact at a more favorable position to overcome their interfacial tension through this asymmetric geometry, until the interfacial layer breaks and pushes the former (with higher viscosity) into the latter. On the other hand, a typical symmetric cross-junction geometry cannot induce merging, because of insufficient interfacial compression generated by direct collision between two droplets. The merged N-CEC and ODX droplets soon become completely homogeneous via diffusion, ultimately leading to in situ microgel formation. Changing the concentration of ODX further controls the crosslinking density of the microgels. In addition, the viability of cells encapsulated within the microgels is well maintained, demonstrating the biocompatibility of the entire process. Taken together, the microfluidic merging technique introduced here could be broadly applicable for engineering cell-encapsulated microgels for biomedical applications.