Edward Kang

Digitally tunable physicochemical coding of material composition and topography in continuous microfibres

Heterotypic functional materials with compositional and topographical properties that vary spatiotemporally on the micro- or nanoscale are common in nature. However, fabricating such complex materials in the laboratory remains challenging. Here we describe a method to continuously create microfibres with tunable morphological, structural and chemical features using a microfluidic system consisting of a digital, programmable flow control that mimics the silk-spinning process of spiders. With this method we fabricated hydrogel microfibres coded with varying chemical composition and topography along the fibre, including gas micro-bubbles as well as nanoporous spindle-knots and joints that enabled directional water collection. We also explored the potential use of the coded microfibres for tissue engineering applications by creating multifunctional microfibres with a spatially controlled co-culture of encapsulated cells.

Spheroid-based three-dimensional liver-on-a-chip to investigate hepatocyte–hepatic stellate cell interactions and flow effects

We have developed a three-dimensional (3D) liver-on-a-chip to investigate the interaction of hepatocytes and hepatic stellate cells (HSCs) in which primary 3D hepatocyte spheroids and HSCs are co-cultured without direct cell-cell contact. Here, we show that the 3D liver chip offers substantial advantages for the formation and harvesting of spheroids. The most important feature of this liver chip is that it enables continuous flow of medium to the cells through osmotic pumping, and thus requires only minimal handling and no external power source. We also demonstrate that flow assists the formation and long-term maintenance of spheroids. Additionally, we quantitatively and qualitatively investigated the paracrine effects of HSCs, demonstrating that HSCs assist in the maintenance of hepatocyte spheroids and play an important role in the formation of tight cell-cell contacts, thereby improving liver-specific function. Spheroids derived from co-cultures exhibited improved albumin and urea secretion rates compared to mono-cultured spheroids after 9 days. Immunostaining for cytochrome P450 revealed that the enzymatic activity of spheroids co-cultured for 8 days was greater than that of mono-cultured spheroids. These results indicate that this system has the potential for further development as a unique model for studying cellular interactions or as a tool that can be incorporated into other models aimed at creating hepatic structure and prolonging hepatocyte function in culture.

Microfluidic spinning of micro- and nano-scale fibers for tissue engineering

Microfluidic technologies have recently been shown to hold significant potential as novel tools for producing micro- and nano-scale structures for a variety of applications in tissue engineering and cell biology. Over the last decade, microfluidic spinning has emerged as an advanced method for fabricating fibers with diverse shapes and sizes without the use of complicated devices or facilities. In this critical review, we describe the current development of microfluidic-based spinning techniques for producing micro- and nano-scale fibers based on different solidification methods, platforms, geometries, or biomaterials. We also highlight the emerging applications of fibers as bottom-up scaffolds such as cell encapsulation or guidance for use in tissue engineering research and clinical practice.

Microfluidic Spinning of Flat Alginate Fibers with Grooves for Cell‐Aligning Scaffolds

Alginate microribbons with longitudinally grooved microstructures are continuously fabricated by means of a microfluidic system. The number and dimensions of the microgroovesare successfully controlled by regulation of the slit-shaped channel (yellow in figure). This method opens up the possibility of mass production of scaffolds for tissue engineering purposes, as it is proved that the grooved flat fibers can be used to align other types of cells in culture.

Novel PDMS cylindrical channels that generate coaxial flow, and application to fabrication of microfibers and particles

In this paper, we introduce a novel cylindrical channel that generates coaxial flow without using glass microcapillary or complicated silicon processing, and we demonstrate the fabrication of microparticles and microfibers using this channel. The simple fabrication process for cylindrical channels employs the deflection of free-standing thin PDMS membranes. Using this channel, alginate microparticles and microfibers were fabricated without clogging the downstream channel, and the dimensions of these particles and fibers could be successfully controlled by regulating the flow rate through the channels. We also developed a method to integrate the coaxial flow channel into rectangular microfluidic channel devices, which have a broad array of established applications. As proof of concept of this technology, we fabricated a microfluidic chip that incorporated 12 rectangular micromixers to generate a stepwise gradient across discrete output streams. These output streams simultaneously fed into 5 coaxial flow channels, each of which produced a microfiber of a different chemical composition. The fibers or particles generated by the proposed method may be used in biomedical and tissue engineering, as well as in drug delivery. We expect that our method will facilitate the construction of microfluidic factories within single PDMS devices.

Microfluidic wet spinning of chitosan-alginate microfibers and encapsulation of HepG2 cells in fibers

The successful encapsulation of human hepatocellular carcinoma (HepG2) cells would greatly assist a broad range of applications in tissue engineering. Due to the harsh conditions during standard chitosan fiber fabrication processes, encapsulation of HepG2 cells in chitosan fibers has been challenging. Here, we describe the successful wet-spinning of chitosan-alginate fibers using a coaxial flow microfluidic chip. We determined the optimal mixing conditions for generating chitosan-alginate fibers, including a 1:5 ratio of 2% (w∕w) water-soluble chitosan (WSC) solution to 2% (w∕w) alginate solution. Ratio including higher than 2% (w∕w) WSC solution increased aggregation throughout the mixture. By suspending cells in the WSC-alginate solution, we successfully fabricated HepG2 cell-laden fibers. The encapsulated HepG2 cells in the chitosan-alginate fibers were more viable than cells encapsulated in pure alginate fibers, suggesting that cross-linked chitosan provides a better environment for HepG2 cells than alginate alone. In addition, we found that the adhesion of HepG2 cells on the chitosan-alginate fiber is much better than that on the alginate fibers.

Micro/Nanometer-scale fiber with highly ordered structures by mimicking the spinning process of silkworm.

A new method for the microfluidic spinning of ultrathin fibers with highly ordered structures is proposed by mimicking the spinning mechanism of silkworms. The self-aggregation is driven by dipole-dipole attractions between polar polymers upon contact with a low-polarity solvent to form fibers with nanostrands. The induction of Kelvin-Helmholtz instabilities at the dehydrating interface between two miscible fluids generates multi-scale fibers in a single microchannel.

Development of a multi-layer microfluidic array chip to culture and replate uniform-sized embryoid bodies without manual cell retrieval†

We have developed a multi-layer, microfluidic array platform containing concave microwells and flat cell culture chambers to culture embryonic stem (ES) cells and regulate uniform-sized embryoid body (EB) formation. The main advantage of this platform was that EBs cultured within the concave microwells of a bottom layer were automatically replated into flat cell culture chambers of a top layer, following inversion of the multi-layer microfluidic array platform. This allowed EB formation and EB replating to be controlled simultaneously inside a single microfluidic device without pipette-based manual cell retrieval, a drawback of previous EB culture methods.

Large-Scale, Ultrapliable, and Free-Standing Nanomembranes

The creation and characterization of large-area ultrathin highly pliable free-standing PDMS membranes and their application to the study of cellular epithelia is described. The ultra-thin membranes permitted the straight forward calculation of cell monolayer moduli, derived from measured stress-strain curves. These measurements allowed the unprecedented detection of cellular-level injury in the epithelia caused by the rupture of cell-cell tight junctions in response to stretching.

An integrated microfluidic culture device to regulate endothelial cell differentiation from embryonic stem cells.

We developed an integrated microfluidic culture device to regulate embryonic stem (ES) cell fate. The integrated microfluidic culture device consists of an air control channel and a fluidic channel with 4×4 micropillar arrays. We hypothesized that the microscale posts within the micropillar arrays would enable the control of uniform cell docking and shear stress profiles. We demonstrated that ES cells cultured for 6 days in the integrated microfluidic culture device differentiated into endothelial cells. Therefore, our integrated microfluidic culture device is a potentially powerful tool for directing ES cell fate.