Three-Dimensional Cell Drawing Technique in Hydrogel Using Micro Injection System

Abstract

Fabrication of three-dimensional tissues using living cells is a promised approach for drug screening experiment and in vitro disease modeling. To study a physiological neuronal function, three-dimensional cell patterning and construction of neuronal cell network were required. In this study, we proposed a three-dimensional cell drawing methodology in hydrogel to construct the three-dimensional neuronal cell network. PC-12 cells, which were used as neuronal cell differentiation model, were dispensed into a collagen hydrogel using a micro injector with a three-dimensional position control. To maintain the three-dimensional position of cells, atelocollagen was kept at sol-gel transition state during cell dispensing. As the results, PC-12 cells were patterned in the atelocollagen gel to form square pattern with different depth. In the patterned cellular lines, PC-12 cells elongated neurites and form a continuous cellular network in the atelocollagen gel. It was suggested that our three-dimensional cell drawing technology has potentials to reconstruct three-dimensional neuronal networks for an investigation of physiological neuronal functions.

Multilayered Gel-Spotting Device for In Vitro Reconstruction of Hair Follicle-like Microstructure

“…Finally, the gel microconstructs were washed 2 times and covered with a cell culture medium. The collagen gel culture method has been described previously [ 11 , 13 , 14 ]. Each three-layered gel construct contained 1.6 × 10 4 mESCs and 1.6 × 10 4 mouse epidermal cells.…”

“…The microinjector consisted of a motorized stage (TSDM60-20X, Opto-Sigma, J and a 1 mL disposable syringe (SS-01T, TERUMO, Tokyo, Japan) or microdispenser Fixed Volume Microdispenser, Drummond Scientific, Broomall, PA, USA) to dispen gel solution (Figure 2b). The disposable syringe was held in a metal fixture with co solution inside to prevent gelation of the neutral collagen solution, as described i previous study [11]. A 27G needle (NN-2719S, TERUMO, Tokyo, Japan) was connec The microinjector consisted of a motorized stage (TSDM60-20X, Opto-Sigma, Japan) and a 1 mL disposable syringe (SS-01T, TERUMO, Tokyo, Japan) or microdispenser (5 µL Fixed Volume Microdispenser, Drummond Scientific, Broomall, PA, USA) to dispense the gel solution (Figure 2b).…”

“…A 27G needle (NN-2719S, TERUMO, Tokyo, Japan) was connec The microinjector consisted of a motorized stage (TSDM60-20X, Opto-Sigma, Japan) and a 1 mL disposable syringe (SS-01T, TERUMO, Tokyo, Japan) or microdispenser (5 µL Fixed Volume Microdispenser, Drummond Scientific, Broomall, PA, USA) to dispense the gel solution (Figure 2b). The disposable syringe was held in a metal fixture with coolant solution inside to prevent gelation of the neutral collagen solution, as described in our previous study [11]. A 27G needle (NN-2719S, TERUMO, Tokyo, Japan) was connected to the tip of the syringe to microspot the gel solutions.…”