Hydrodynamic characterization within a spinner flask and a rotary wall vessel for stem cell culture

Ghasemian, Masoud; Layton, Carys; Nampe, Daniel; Nieden, Nicole I. zur; Tsutsui, Hideaki; Princevac, Marko

doi:10.1016/j.bej.2020.107533

Cited by 21 publications

(28 citation statements)

References 32 publications

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“…Compared to a standard 125ml spinner flask or bioreactor, each well in a 96-well plate requires a maximum of 350 μ l of media, which is approximately a 3.5-fold reduction in total media consumption. The elimination of mechanical stirrers and tubing resolves unwanted fluidic shear stress gradients, decreases damaging cellular collisions, and reduces labor-intensive culture maintenance [41, 42, 43]. Moreover, by seeding each spheroid in its own well, the samples may be kept biologically independent, providing enhanced statistics and throughput for assays.…”

Section: Resultsmentioning

confidence: 99%

Microwell-Based Flow Culture Increases Viability and Restores Drug Response in Prostate Cancer Spheroids

Payne

Hashimoto

et al. 2022

Preprint

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3D cancer spheroids represent a highly promising model for study of cancer progression and therapeutic development. Wide-scale adoption of cancer spheroids, however, remains a challenge due to the lack of control over hypoxic gradients that may cloud the assessment of cell morphology and drug response. Here, we present a Microwell Flow Device (MFD) that generates in-well laminar flow around 3D tissues via repetitive tissue sedimentation. Using a prostate cancer cell line, we demonstrate the spheroids in the MFD exhibit improved cell growth, reduced necrotic core formation, enhanced cellular structure, and down-regulated expression of cell stress genes. The flow-cultured spheroids also exhibit an improved sensitivity to chemotherapy with greater transcriptional response. These results demonstrate how fluidic stimuli reveal the cellular phenotype previously impacted by severe hypoxia. Our platform advances 3D cellular models and enables study into hypoxia modulation, cancer metabolism, and drug screening within pathophysiologically relevant conditions.

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Section: Resultsmentioning

confidence: 99%

Microwell-Based Flow Culture Increases Viability and Restores Drug Response in Prostate Cancer Spheroids

Payne

Hashimoto

et al. 2022

Preprint

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“…Shear stress is a key factor in determining whether a bioreactor is suitable for the cultivation of shear-sensitive cells such as plant cells, mammalian cells, and stem cells (Ghasemian et al, 2020). Figure 10 shows the variation of shear stress with time in the cases of the initial and optimal Fig.…”

Section: Characteristics Of the Optimal Statementioning

confidence: 99%

Bayesian Optimization for Hydrodynamic Characterization of a Cylindrical Orbitally Shaken Bioreactor with a Bump at the Bottom

Wang

Isobe

Okano

et al. 2021

J. Chem. Eng. Japan

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An orbitally shaken bioreactor is the most popular system for cell cultivation in a suspension culture owing to its unique characteristics. However, conventional bioreactors struggle to meet the anticipated demand in practical applications due to their limitations. In this study, numerical simulations are conducted to evaluate the performance of a newly developed orbitally shaken bioreactor that is equipped with a vaulted "bump" at the bottom wall. The presence of this bump signi cantly improves the mass transfer and cell suspension ratio in the culture medium without increasing the shear stress; it also e ciently reduces the cell aggregation in the central part at the bottom wall. In addition, to enable a suitable cell culture environment for practical applications, the Bayesian algorithm is employed to optimize the control of three parameters, namely bump height (h b ), shaking velocity (ω), and shaking radius (R), of this reactor. For all the cases considered, the obtained data related to mass transfer, suspension ratio, and shear stress are analyzed. The simulation results show that the optimal bioreactor is preferable for cell cultivation compared with the initial state, owing to its high capability for mass transfer and suspending cells. It can be concluded that the methodology described in this paper is a feasible and reliable tool for performance prediction and process optimization in biotechnology.

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“…In the unique environment of space, free fall-induced weightlessness can modulate the intrinsic factors of tissue mechanical environment leading to reduced mechanical loads in tissue niche environments [ 8 ]. Modeling the effects of microgravity on Earth has been attempted with a multitude of strategies; however, many of these strategies in an effort to replicate free fall, generate non-physiologic fluid shear and rotational loading to the cells [ 9 ]. The interesting observation that traditional static cell culture plating results in pressure loading well below the physiologically relevant range may provide a unique path for cellular level modeling of the microgravity free fall environment.…”

Section: Introductionmentioning

confidence: 99%

“…The most studied physical stress relevant to the EB model is fluid shear stress [ 14 ], imparted by spinner flasks [ 15 ] or microfluidic channels [ 16 , 17 ], or rotating wall vessels [ 9 ] used in their culture. Generally, these studies have shown that fluidic shear stress promotes growth and maturation; however, this general observation has been refined as the study of mechanical regulation of EBs has expanded.…”

Section: Introductionmentioning

confidence: 99%

Differential Single Cell Responses of Embryonic Stem Cells Versus Embryoid Bodies to Gravity Mechanostimulation

Juran

Žvirblytė

Almeida

2022

Stem Cells and Development

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The forces generated by gravity have shaped life on Earth and impact gene expression and morphogenesis during early development. Conversely, disuse on Earth or during spaceflight, reduces normal mechanical loading of organisms, resulting in altered cell and tissue function. Although gravity mechanical loading in adult mammals is known to promote increased cell proliferation and differentiation, little is known about how distinct cell types respond to gravity mechanostimulation during early development. In this study we sought to understand, with single cell RNA-sequencing resolution, how a 60-min pulse of 50 g hypergravity (HG)/5 kPa hydrostatic pressure, influences transcriptomic regulation of developmental processes in the embryoid body (EB) model. Our study included both day-9 EBs and progenitor mouse embryonic stem cells (ESCs) with or without the HG pulse. Single cell t-distributed stochastic neighbor mapping shows limited transcriptome shifts in response to the HG pulse in either ESCs or EBs; this pulse however, induces greater positional shifts in EB mapping compared to ESCs, indicating the influence of mechanotransduction is more pronounced in later states of cell commitment within the developmental program. More specifically, HG resulted in upregulation of self-renewal and angiogenesis genes in ESCs, while in EBs, HG loading was associated with upregulation of Gene Ontology-pathways for multicellular development, mechanical signal transduction, and DNA damage repair. Cluster transcriptome analysis of the EBs show HG promotes maintenance of transitory cell phenotypes in early development; including EB cluster co-expression of markers for progenitor, post-implant epiblast, and primitive endoderm phenotypes with HG pulse but expression exclusivity in the non-pulsed clusters. Pseudotime analysis identified three branching cell types susceptible to HG induction of cell fate decisions. In totality, this study provides novel evidence that ESC maintenance and EB development can be regulated by gravity mechanostimulation and that stem cells committed to a differentiation program are more sensitive to gravity-induced changes to their transcriptome.

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Hydrodynamic characterization within a spinner flask and a rotary wall vessel for stem cell culture

Cited by 21 publications

References 32 publications

Microwell-Based Flow Culture Increases Viability and Restores Drug Response in Prostate Cancer Spheroids

Microwell-Based Flow Culture Increases Viability and Restores Drug Response in Prostate Cancer Spheroids

Bayesian Optimization for Hydrodynamic Characterization of a Cylindrical Orbitally Shaken Bioreactor with a Bump at the Bottom

Differential Single Cell Responses of Embryonic Stem Cells Versus Embryoid Bodies to Gravity Mechanostimulation

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