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[Journal of Bioscience and Bioengineering] Compartmentalized microfluidic perfusion system to cultur


Abstract:

Microfluidic perfusion systems enable small-volume cell cultures under precisely controlled microenvironments, and are typically developed for cell-based high-throughput screening. However, most such systems are designed to manipulate dissociated single cells, not cell aggregates, and are thus unsuitable to induce differentiation in human induced pluripotent stem cells (hiPSCs), which is conventionally achieved by using cell aggregates to increase cell–cell interactions. We have now developed a compartmentalized microfluidic perfusion system with large flow channels to load, culture, and observe cell aggregates. Homogeneously sized cell aggregates to be loaded into the device were prepared by shredding flat hiPSC colonies into squares. These aggregates were then seeded into microchambers coated with fibronectin and bovine serum albumin (BSA) to establish adherent and floating cultures, respectively, both of which are frequently used to differentiate hiPSCs. However, the number of aggregates loaded in fibronectin-coated microchambers was much lower than in BSA-coated microchambers, suggesting that fibronectin traps cell aggregates before they reach the chambers. Accordingly, hiPSCs that reached the microchambers subsequently adhered. In contrast, BSA-coated microchambers did not allow cell aggregates to adhere, but were sufficiently deep to prevent cell aggregates from flowing out during perfusion of media. Immunostaining for markers of undifferentiated cells showed that cultures on both fibronectin- and BSA-coated microchambers were successfully established. Notably, we found that floating aggregates eventually adhered to surfaces coated with BSA upon differentiation, and that differentiation depends on the initial size of aggregates. Collectively, these results suggest that the microfluidic system is suitable for manipulating hiPSC aggregates in compartmentalized microchambers.

Yuki Kondo1, Koji Hattori2, §, Shota Tashiro1, Eri Nakatani1, Ryosuke Yoshimitsu1, Taku Satoh3, Shinji Sugiura3, Toshiyuki Kanamori3, Kiyoshi Ohnuma1, 4, , §, 1 Department of Bioengineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata 940-2188, Japan 2 Research Center for Stem Cell Engineering, National Institute of Advanced Industrial Science and Technology (AIST), Central 5th, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8565, Japan 3 Biotechnology Research Institute for Drug Discovery, National Institute of Advanced Industrial Science and Technology (AIST), Central 5th, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8565, Japan 4 Department of Science of Technology Innovation, Nagaoka University of Technology, 1603-1 Kamitomioka-cho, Nagaoka, Niigata 940-2188, Japan Received 29 December 2016, Accepted 24 March 2017, Available online 20 April 2017 Show less http://doi.org/10.1016/j.jbiosc.2017.03.014

Link: http://www.sciencedirect.com/science/article/pii/S138917231630706X

#04262017 #perfusion #cell #stemcell #Biologicalapplication

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