Isolation and Culture of hiSMCs and hiNSCs

Primary human intestinal smooth muscle cells (hiSMCs) were obtained from
ScienCell and cultured in smooth muscle cell medium (SMCM, ScienCell). Cells
were cultured in T-75 cm² tissue culture flasks (Corning), maintained
at 37°C, 5% CO₂ humified atmosphere and harvested with 0.25%
trypsin-EDTA (Gibco) prior to seeding. Cells at passage number 4–5 were
used for the experiments. Human induced neural stem cells (hiNSCs) were
generated and expanded as previously described ^{[17 (link)]}. Briefly, expanded hiNSCs were lifted
off mouse embryonic fibroblasts (MEFs) using TrypLE Select and pelleted. The
cell pellet was resuspended in hiNSC expansion media consisting of KnockOut
Serum Replacement DMEM (Thermo Fisher), GlutaMax (Thermo Fisher), KnockOut SR
(Thermo Fisher), Antibiotic-Antimycotic (Thermo Fisher) and 2-mercapto (Thermo
Fisher), bFGF Basic (Thermo Fisher). The resuspended cell solution was vortexed
and passed through a 40μm filter to achieve single cell suspension. The
resulting single cells obtained from hiNSC colonies were then used for scaffold
seeding. The media were changed every other day.

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Chen Y., Guo C., Manousiouthakis E., Wang X., Cairns D.M., Roh T.T., Du C, & Kaplan D.L. (2020). Bi-layered Tubular Microfiber Scaffolds as Functional Templates for Engineering Human Intestinal Smooth Muscle Tissue. Advanced functional materials, 30(17), 2000543.

Publication 2020

T-75 cm2 tissue culture flasks trypsin-EDTA GlutaMax KnockOut SR Antibiotic-Antimycotic

Antibiotic Atmosphere Cells Cultured medium Edta Embryonic Fibroblasts Human Intestinal Mouse Neural stem cells Smooth muscle cell Tissue Trypsin

Corresponding Organization : Tufts University

Other organizations : Dalian Medical University

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Variable analysis

independent variables

Cell type (primary human intestinal smooth muscle cells (hiSMCs) and human induced neural stem cells (hiNSCs))

dependent variables

Not explicitly mentioned

control variables

Passage number (4-5) for hiSMCs
Culture conditions (37°C, 5% CO2, humified atmosphere) for both cell types
Culture media (smooth muscle cell medium (SMCM) for hiSMCs, hiNSC expansion media for hiNSCs)
Cell culture vessels (T-75 cm2 tissue culture flasks)

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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