Browsing by Author "Yuan, Chen-Ching"

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Selection of alkaline phosphatase-positive induced pluripotent stem cells from human amniotic fluid-derived cells by feeder-free system
(2011-08-01)
Lu, Huai-En
;
Tsai, Ming-Song
;
Yang, Yao-Chen
;
Yuan, Chen-Ching
;
Wang, Tzu-Hao
Generation of induced pluripotent stem (iPS) cells from somatic cells has been successfully achieved by ectopic expression of four transcription factors, Oct4, Sox2, Klf4 and c-Myc, also known as the Yamanaka factors. In practice, initial iPS colonies are picked based on their embryonic stem (ES) cell-like morphology, but often may go on to fail subsequent assays, such as the alkaline phosphate (AP) assay. In this study, we co-expressed through lenti-viral delivery the Yamanaka factors in amniotic fluid-derived (AF) cells. ES-like colonies were picked onto a traditional feeder layer and a high percentage AF-iPS with partial to no AP activity was found. Interestingly, we obtained an overwhelming majority of fully stained AP positive (AP+) AF-iPS colonies when colonies were first seeded on a feeder-free culture system, and then transferred to a feeder layer for expansion. Furthermore, colonies with no AP activity were not detected. This screening step decreased the variation seen between morphology and AP assay. We observed the AF-iPS colonies grown on the feeder layer with 28%AP+ colonies, 45% AP partially positive (AP+/-) colonies and 27% AP negative (AP-) colonies, while colonies screened by the feeder-free system were 84%AP+ colonies, 16% AP+/- colonies and no AP- colonies. The feeder-free screened AP+ AF-iPS colonies were also positive for pluripotent markers, OCT4, SOX2, NANOG,TRA-1-60,TRA-1-81,SSEA-3 and SSEA-4 as well as having differentiation abilities into three germ layers in vitro and in vivo. In this study, we report a simplistic, one-step method for selection of AP+ AF-iPS cells via feeder-free screening. (C) 2011 Elsevier Inc. All rights reserved.
Submicron-grooved culture surface extends myotube length by forming parallel and elongated motif
(2013-08-01)
Yuan, Chen-Ching
;
Ma, Kuang-Jen
;
Li, Kuei-Chi
;
Chien, Hsi-Hsin
;
Lu, Huai-En
During skeletal muscle development, correct cellular orientation is vital to generate desired longitudinal contraction for functional muscle fibres. In this reported study, submicron-imprint lithography was used to generate submicron-grooved surfaces on polystyrene plates to induce striated myotubes in vitro. Mouse muscle myoblast cells cultured on a submicron-grooved surface migrated faster in a directionally uniform fashion; in comparison, cells cultured on a flat surface grew and migrated slower in indiscriminate directions. Subsequent maturation of the myoblast cells formed along the submicron-groove surface resulted in a tandem of parallel myotubes that were both longer and greater in circumference than in the case of the flat surface. In a functional test, the co-culture submicron-groove-grown myotubes with neurotransmitter secreting cells further demonstrated contraction abilities, suggesting submicron-groove-guided growth served to enhance myotube formation while retaining striated motifs and physiological functionality for muscle tissue engineering.