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International Journal of Bioprinting Microfluidic spinning for neural models

by assembling HUVECs-loaded hollow microfiber the chip templates. BMF_3Dslice software was used to
microchips, an in vitro neural differentiation model, slice and import images to the computer connected to
capable of detecting axonal growth and related gene the 3D printer. The platform lifting distance was adjusted
expression in PC12 cells under the action of nerve growth according to the number of images and the required
factor (NGF), was constructed. This model is expected to accuracy, and the splicing mode was selected to print
provide a new platform for studying the development of the photosensitive resin chip template. Subsequently,
neurological diseases and evaluating drugs, and to offer the PDMS monomer and initiator were evenly mixed in
an avenue for personalized diagnosis. a 10:1 (v/v) solution and poured onto a resin template.
Bubbles were removed under vacuum, and the PDMS
2. Materials and methods prepolymer was cured at 80°C for 1 h. Subsequently, a
2.1. Materials PDMS layer with microchannels was obtained by peeling
NaA, GelMA (90%), calcium chloride dihydrate off the template. For the microfluidic spinning microchip,
(CaCl ·2H O), chitosan (η > 400 mPa·s), polystyrene irreversible sealing was required, that is, oxygen plasma
2
2
microspheres (PS microspheres; 5 µm, ex/em: 400/450 surface treatment was performed on two PDMS layers
nm), and polystyrene microspheres (PS microspheres; 5 with the same structure for alignment and sealing to
µm, ex/em: 488/518 nm) were purchased from Shanghai obtain the final microfluidic spinning microchip. The
Aladdin Biochemical Technology Co., Ltd. (Shanghai, microfiber assembly microchip did not require sealing
China). Pluronic® F-127 (PF-127), 2-hydroxy-4’-(2- and was sterilized as a backup. The dimensions of the
hydroxyethoxy)-2-methylphenylacetone (I2959, 98%), template and the microfluidic chip were measured using
acetic acid, isopropanol, dimethyl sulfoxide (DMSO), and an inverted fluorescence microscope (IX-73, Olympus,
4’, 6-diamino-2-phenylindole dihydrochloride (DAPI) Tokyo, Japan) or a stylus profilometer (AlphaStep D-300
were purchased from Sigma (Shanghai, China). The Stylus Profile, KLA, Milpitas, America).
3D printing resin, containing methacrylate monomers, 2.3. Preparation and characterization of composite
methyl oligomers, and photoinitiators, was purchased hollow microfibers
from Shenzhen Mofang Materials Technology Co., Ltd. NaA, GelMA, I2959, CaCl ·2H O, and PF-127 were all
Shenzhen, China. Sylgard 184 PDMS was purchased from dissolved in deionized water and sterilized for future use.
2
2
Dow Corning (Midland, America). Fetal bovine serum A 20% w/v PF-127 solution was used as the core flow, a
(FBS), horse serum, Roswell Park Memorial Institute mixture of 0.8% w/v NaA, 5% w/v GelMA, and 0.5% w/v
Medium 1640 (1640 medium), and double antibiotic photoinitiator I2959 was used as the sample flow, and
solution containing penicillin and streptomycin were a 4% (w/v) CaCl solution was used as the sheath flow.
purchased from Gibco Grand Island, America. Endothelial The core, sample, and sheath flows were injected into the
2
cell culture medium, supplemented with FBS, double microchannels of the microfluidic spinning microchip
antibiotic solution containing penicillin and streptomycin,
and endothelial cell growth additive, was purchased using a microinjection pump (Harvard, Holliston,
America). A UV device emitting rays at a wavelength of
from ScienCell, San Diego, America. CellTracker Green 365 nm was fixed above the microchip outlet position,
CMFDA, propidium iodide, Alexa Fluor® 488 phalloidin, and the microfibers were collected in a CaCl solution
and Alexa Fluor 568 Dextran (10 kDa) were purchased 2
from Life Invitrogen Carlsbad, America. Rabbit anti-von tank. To determine the composition of the composite
Willebrand factor (vWF), rabbit anti-β-Tubulin, goat microfibers, we characterized the composition of GelMA,
a mixture of CaA and GelMA, and the prepared CaA/
anti-rabbit IgG 568, blocking buffer, and antibody diluent
were all purchased from Abcam, Cambridge, England. GelMA microfibers using Fourier transform infrared
Nerve growth factor was purchased from R&D Systems (FTIR) spectroscopy (Thermo Fisher Scientific, Waltham,
(Minneapolis, America). America). To investigate the effect of flow rate on the
diameter of microfibers and facilitate the observation of
2.2. Fabrication and characterization of microchip the morphology of hollow microfibers, we mixed blue
According to the device manual, a 3D printing device fluorescent PS microspheres (5 µm, ex/em: 400/450 nm)
(nanoArch® SI40, BMF Material Technology Inc., and green fluorescent PS microspheres (5 µm, ex/em:
Shenzhen, China) was used to prepare microfluidic 488/518 nm) at a concentration of 0.1 mg/mL into the
chip templates. There exist two types of templates: sample solution and injected them into the corresponding
microfluidic spinning and microfiber assembly microchip microchannels. The flow velocity rates for core flow, sample
templates. We prepared two templates as described flow, and sheath flow were 5–100, 50–110, and 120–200
previously. 41,42 Solidworks software was used to design μL/min, respectively.

Volume 10 Issue 2 (2024) 266 doi: 10.36922/ijb.1797