Effect of Culture Conditions on the Multinucleation of Human Adipose-Derived Stem Cells

Human adipose-derived stem cells (hADSCs) are types of mesenchymal stem cells (MSCs) that have been used as tissue engineering models for bone, cartilage, muscle, marrow stroma, tendon, fat and other connective tissues. Tissue regeneration materials composed of hADSCs have the potential to play an important role in reconstituting damaged tissue or diseased mesenchymal tissue. In this study, we assessed and investigated the osteogenesis of hADSCs in both two-dimensional (2D) and three-dimensional (3D) culture conditions. We confirmed that the hADSCs successfully differentiated into bone tissues by ARS staining and quantitative RT–PCR. To gain insight into the detailed biological difference between the two culture conditions, we profiled the overall gene expression by analyzing the whole transcriptome sequencing data using various bioinformatic methods. We profiled the overall gene expression through RNA-Seq and further analyzed this using various bioinformatic methods. During differential gene expression testing, significant differences in the gene expressions between hADSCs cultured in 2D and 3D conditions were observed. The genes related to skeletal development, bone development and bone remodeling processes were overexpressed in the 3D culture condition as compared to the 2D culture condition. In summary, our RNA-Seq-based study proves effective in providing new insights that contribute toward achieving a genome-wide understanding of gene regulation in mesenchymal stem cell osteogenic differentiation and bone tissue regeneration within the 3D culture system.

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Induction of Tendon-Specific Markers in Adipose-Derived Stem Cells in Serum-Free Culture Conditions

Tissue Engineering Part C Methods ◽

10.1089/ten.tec.2019.0080 ◽

2019 ◽

Vol 25 (7) ◽

pp. 389-400 ◽

Cited By ~ 3

Author(s):

Noelia D. Falcon ◽

Graham P. Riley ◽

Aram Saeed

Keyword(s):

Stem Cells ◽

Culture Conditions ◽

Adipose Derived Stem Cells ◽

Serum Free

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Characterisation and In Vivo Safety of Canine Adipose-Derived Stem Cells

Proceedings of the Latvian Academy of Sciences Section B Natural Exact and Applied Sciences ◽

10.2478/prolas-2018-0004 ◽

2018 ◽

Vol 72 (3) ◽

pp. 160-171

Author(s):

Uldis Bērziņš ◽

Ilze Matise-VanHoutana ◽

Ilze Pētersone ◽

Ilmārs Dūrītis ◽

Sergejs Ņikuļšins ◽

...

Keyword(s):

Stem Cells ◽

Intravenous Administration ◽

Adipogenic Differentiation ◽

Canine Model ◽

Culture Conditions ◽

Live Cells ◽

Autologous Serum ◽

Blast Transformation ◽

Adipose Derived Stem Cells

Abstract The study characterises canine adipose-derived stem cells (cASCs) in comparison to human ASCs (hASCs) and tests their safety in a canine model after intravenous administration. cASCs from two dogs were cultured under hypoxic conditions in a medium supplemented with autologous serum. They were plastic adherent, spindle-shaped cells that expressed CD73, CD90, and CD44 but lacked CD45, CD14, HLA-DR, and CD34. cASCs differentiated toward adipogenic, osteogenic, and chondrogenic lineages, although adipogenic differentiation capacity was low. Blast transformation reaction demonstrated that these cells significantly suppress T-cell proliferation, and this ability is dose-dependent. Intravenous administration of a cell freezing medium, therapeutic dose of cASCs (2 × 106 live cells/kg), and five times higher dose of cASCs showed no significant side effects in two dogs. Microscopic tissue lesions were limited to only mild, non-specific changes. There were no signs of malignancy. The results of the study indicate that cASCs are similar to hASCs and are safe for therapeutic applications in a canine model. The proposed methodology for ASC preparation on a non-routine basis, which includes individually optimised cell culture conditions and offers risk-adapted treatment, could be used for future personalised off-the-shelf therapies, for example, in myocardial infarction or stroke.

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Adipose-derived stem cells express higher levels of type VII collagen under specific culture conditions

Archives of Dermatological Research ◽

10.1007/s00403-017-1781-9 ◽

2017 ◽

Vol 309 (10) ◽

pp. 843-849

Author(s):

Yuichiro Maeda ◽

Toshio Hasegawa ◽

Akino Wada ◽

Tatsuo Fukai ◽

Hideo Iida ◽

...

Keyword(s):

Stem Cells ◽

Culture Conditions ◽

Adipose Derived Stem Cells ◽

Type Vii Collagen ◽

Specific Culture

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Characterization of human adipose-derived stem cells

Acta Cirurgica Brasileira ◽

10.1590/s0102-86502012000700007 ◽

2012 ◽

Vol 27 (7) ◽

pp. 471-476 ◽

Cited By ~ 19

Author(s):

Silvana Gaiba ◽

Lucimar Pereira de França ◽

Jerônimo Pereira de França ◽

Lydia Masako Ferreira

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Chondrogenic Differentiation ◽

Therapeutic Potential ◽

Culture Conditions ◽

In Vitro Cultivation ◽

Neural Cells ◽

Adipose Derived Stem Cells ◽

Differentiation Potential

PURPOSE: There is a growing scientific interest in the plasticity and therapeutic potential of adipose-derived stem cells (ASCs), which are multipotent and abundant in adipose tissue and can differentiate in vitro into multiple lineages, including adipocytes, chondrocytes, osteoblasts, neural cells, endothelial cells and cardiomyocytes. The aim of this study was to isolate, cultivate and identify ASCs. METHODS: Human adipose precursor cells were obtained from subcutaneous abdominal tissue. Recently dispersed cells were separated by density centrifugation gradient, cultured and then analyzed. RESULTS: Human ASCs were able to replicate in our culture conditions. The cells maintained their phenotypes throughout the studied period on different passages confirming they suitability for in vitro cultivation. We also induced their adipogenic, osteogenic and chondrogenic differentiation, verifying their mesenchymal stem cells potentiality in vitro. Flow cytometry results showed that these cells expressed CD73, CD90 and CD105, (mesenchymal stem-cells markers), contrasting with the lack of expression of CD16, CD34 and CD45 (hematopoietic cells markers). CONCLUSION: It was possible to isolate human adipose-derived stem cells by in vitro cultivation without adipogenic induction, maintaining their functional integrity and high proliferation levels. The cells demonstrated adipogenic, osteogenic and chondrogenic differentiation potential in vitro.

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Tissue Source and Cell Expansion Condition Influence Phenotypic Changes of Adipose-Derived Stem Cells

Stem Cells International ◽

10.1155/2017/7108458 ◽

2017 ◽

Vol 2017 ◽

pp. 1-15 ◽

Cited By ~ 7

Author(s):

Lauren H. Mangum ◽

Shanmugasundaram Natesan ◽

Randolph Stone ◽

Nicole L. Wrice ◽

David A. Larson ◽

...

Keyword(s):

Stem Cells ◽

Culture Conditions ◽

Adipose Derived Stem Cells ◽

Burned Skin ◽

Human Ascs ◽

Triglyceride Content ◽

Human Platelet Lysate ◽

Surface Staining ◽

Expansion Condition ◽

Subcutaneous Adipose

Stem cells derived from the subcutaneous adipose tissue of debrided burned skin represent an appealing source of adipose-derived stem cells (ASCs) for regenerative medicine. Traditional tissue culture uses fetal bovine serum (FBS), which complicates utilization of ASCs in human medicine. Human platelet lysate (hPL) is one potential xeno-free, alternative supplement for use in ASC culture. In this study, adipogenic and osteogenic differentiation in media supplemented with 10% FBS or 10% hPL was compared in human ASCs derived from abdominoplasty (HAP) or from adipose associated with debrided burned skin (BH). Most (95–99%) cells cultured in FBS were stained positive for CD73, CD90, CD105, and CD142. FBS supplementation was associated with increased triglyceride content and expression of adipogenic genes. Culture in hPL significantly decreased surface staining of CD105 by 31% and 48% and CD142 by 27% and 35% in HAP and BH, respectively (p<0.05). Culture of BH-ASCs in hPL also increased expression of markers of osteogenesis and increased ALP activity. These data indicate that application of ASCs for wound healing may be influenced by ASC source as well as culture conditions used to expand them. As such, these factors must be taken into consideration before ASCs are used for regenerative purposes.

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Selective Proliferation of Highly Functional Adipose-Derived Stem Cells in Microgravity Culture with Stirred Microspheres

Cells ◽

10.3390/cells10030560 ◽

2021 ◽

Vol 10 (3) ◽

pp. 560

Author(s):

Takanobu Mashiko ◽

Koji Kanayama ◽

Natsumi Saito ◽

Takako Shirado ◽

Rintaro Asahi ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Network Formation ◽

Embryonic Stem ◽

Adult Stem Cell ◽

Culture Conditions ◽

Therapeutic Effects ◽

Adipose Derived Stem Cells ◽

Mesenchymal Differentiation ◽

Microgravity Conditions

Therapeutic effects of adult stem-cell transplantations are limited by poor cell-retention in target organs, and a reduced potential for optimal cell differentiation compared to embryonic stem cells. However, contemporary studies have indicated heterogeneity within adult stem-cell pools, and a novel culturing technique may address these limitations by selecting those for cell proliferation which are highly functional. Here, we report the preservation of stemness in human adipose-derived stem cells (hASCs) by using microgravity conditions combined with microspheres in a stirred suspension. The cells were bound to microspheres (100−300 μm) and cultured using a wave-stirring shaker. One-week cultures using polystyrene and collagen microspheres increased the proportions of SSEA-3(+) hASCs 4.4- and 4.3-fold (2.7- and 2.9-fold increases in their numbers), respectively, compared to normal culture conditions. These cultured hASCs expressed higher levels of pluripotent markers (OCT4, SOX2, NANOG, MYC, and KLF), and had improved abilities for proliferation, colony formation, network formation, and multiple-mesenchymal differentiation. We believe that this novel culturing method may further enhance regenerative therapies using hASCs.

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