Large-Scale of Human Induced Pluripotent Stem Cell-Derived by Stirring-Type

Stem cells secrete paracrine factors including extracellular vesicles (EVs) which can mediate cellular communication and support the regeneration of injured tissues. Reduced oxygen (hypoxia) as a key regulator in development and regeneration may influence cellular communication via EVs. We asked whether hypoxic conditioning during human induced pluripotent stem cell (iPSC) culture effects their EV quantity, quality or EV-based angiogenic potential. We produced iPSC-EVs from large-scale culture-conditioned media at 1%, 5% and 18% air oxygen using tangential flow filtration (TFF), with or without subsequent concentration by ultracentrifugation (TUCF). EVs were quantified by tunable resistive pulse sensing (TRPS), characterized according to MISEV2018 guidelines, and analyzed for angiogenic potential. We observed superior EV recovery by TFF compared to TUCF. We confirmed hypoxia efficacy by HIF-1α stabilization and pimonidazole hypoxyprobe. EV quantity did not differ significantly at different oxygen conditions. Significantly elevated angiogenic potential was observed for iPSC-EVs derived from 1% oxygen culture by TFF or TUCF as compared to EVs obtained at higher oxygen or the corresponding EV-depleted soluble factor fractions. Data thus demonstrate that cell-culture oxygen conditions and mode of EV preparation affect iPSC-EV function. We conclude that selecting appropriate protocols will further improve production of particularly potent iPSC-EV-based therapeutics.

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Design of large-scale manufacturing of induced pluripotent stem cell derived cardiomyocytes

Chemical Engineering Research and Design ◽

10.1016/j.cherd.2013.08.021 ◽

2014 ◽

Vol 92 (6) ◽

pp. 1142-1152 ◽

Cited By ~ 16

Author(s):

Christopher Luke Darkins ◽

Carl-Fredrik Mandenius

Keyword(s):

Stem Cell ◽

Pluripotent Stem Cell ◽

Large Scale ◽

Induced Pluripotent Stem Cell ◽

Induced Pluripotent

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Error-Correcting Output Codes in Classification of Human Induced Pluripotent Stem Cell Colony Images

BioMed Research International ◽

10.1155/2016/3025057 ◽

2016 ◽

Vol 2016 ◽

pp. 1-13 ◽

Cited By ~ 11

Author(s):

Henry Joutsijoki ◽

Markus Haponen ◽

Jyrki Rasku ◽

Katriina Aalto-Setälä ◽

Martti Juhola

Keyword(s):

Stem Cell ◽

Pluripotent Stem Cell ◽

Large Scale ◽

Nearest Neighbor ◽

Distance Measure ◽

Classification Tree ◽

Induced Pluripotent Stem Cell ◽

Classification Problems ◽

Induced Pluripotent ◽

Error Correcting Output Codes

The purpose of this paper is to examine how well the human induced pluripotent stem cell (hiPSC) colony images can be classified using error-correcting output codes (ECOC). Our image dataset includes hiPSC colony images from three classes (bad, semigood, and good) which makes our classification task a multiclass problem. ECOC is a general framework to model multiclass classification problems. We focus on four different coding designs of ECOC and apply to each one of themk-Nearest Neighbor (k-NN) searching, naïve Bayes, classification tree, and discriminant analysis variants classifiers. We use Scaled Invariant Feature Transformation (SIFT) based features in classification. The best accuracy (62.4%) is obtained with ternary complete ECOC coding design andk-NN classifier (standardized Euclidean distance measure and inverse weighting). The best result is comparable with our earlier research. The quality identification of hiPSC colony images is an essential problem to be solved before hiPSCs can be used in practice in large-scale. ECOC methods examined are promising techniques for solving this challenging problem.

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Generation of human induced pluripotent stem cell-derived liver buds with chemically defined and animal origin-free media

Scientific Reports ◽

10.1038/s41598-020-73908-1 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Keisuke Sekine ◽

Shimpei Ogawa ◽

Syusaku Tsuzuki ◽

Tatsuya Kobayashi ◽

Kazuki Ikeda ◽

...

Keyword(s):

Stem Cell ◽

Mass Production ◽

Pluripotent Stem Cell ◽

Large Scale ◽

Induced Pluripotent Stem Cell ◽

Animal Origin ◽

Scale Production ◽

Clinical Applicability ◽

Induced Pluripotent

Abstract Advances in organoid technology have broadened the number of target diseases and conditions in which human induced pluripotent stem cell (iPSC)-based regenerative medicine can be applied; however, mass production of organoids and the development of chemically defined, animal origin-free (CD-AOF) media and supplements are unresolved issues that hamper the clinical applicability of these approaches. CD-AOF media and supplements ensure the quality and reproducibility of culture systems by lowering lot-to-lot variations and the risk of contamination with viruses or toxins. We previously generated liver organoids from iPSCs, namely iPSC-liver buds (iPSC-LBs), by mimicking the organogenic interactions among hepatocytes, endothelial cells (ECs), and mesenchymal cells (MCs) and recently reported the mass production of iPSC-LBs derived entirely from iPSCs (all iPSC-LBs), which should facilitate their large-scale production for the treatment of liver failure. However, in previous studies we used media originating from animals for differentiation except for the maintenance of undifferentiated iPSCs. Therefore, we developed a CD-AOF medium to generate all iPSC-LBs. We first developed a CD-AOF medium for hepatocytes, ECs, and stage-matched MCs, i.e., septum transversum mesenchyme (STM), in 2D cultures. We next generated all iPSC-LBs by incubating individual cell types in ultra-low attachment micro-dimple plates. The hepatic functions of all iPSC-LBs generated using the CD-AOF medium were equivalent to those of all iPSC-LBs generated using the conventional medium both in vitro and in vivo. Furthermore, we found that this CD-AOF medium could be used in several cell culture settings. Taken together, these results demonstrate the successful development of a CD-AOF medium suitable for all iPSC-LBs. The protocol developed in this study will facilitate the clinical applicability of all iPSC-LBs in the treatment of liver diseases.

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