Three-Dimensional Spheroid Culture Increases Exosome Secretion from Mesenchymal Stem Cells

Kim, Mijin; Yun, Hee-Woong; Park, Do Young; Choi, Byung Hyune; Min, Byoung‐Hyun

doi:10.1007/s13770-018-0139-5

Cited by 112 publications

(98 citation statements)

References 38 publications

(47 reference statements)

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“…Changes in cell morphology, gene expression, etc. led to drastic difference of biological characteristics such as viability, differentiation, and cytokine secretion between MSCs cultured in spheroid and in monolayer [18,37,38]. Hence, the impact of spheroid formation on cell behaviors still needs further investigation.…”

mentioning

confidence: 99%

A Chemically Defined Serum-Free Culture System for Spontaneous Human Mesenchymal Stem Cell Spheroid Formation

Zhao

Xiao

et al. 2020

Stem Cells International

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Mesenchymal stem cells (MSCs) possess promising potential in tissue engineering and regenerative medicine. Previous studies demonstrated that spheroid formation of MSCs exhibited improved stemness maintenance and therapeutic potential compared with monolayer culture. To date, various spheroid culture systems have been developed but most of them required low adhesion conditions or special equipment. In this study, we demonstrated that inoculation of dissociated MSCs in TeSR-E8 medium could induce self-assemble spheroid formation in conventional tissue culture polystyrene dishes. Compared with monolayer culture, adipose-derived stem cell (ADSC) spheroids enhanced the proliferation and osteogenic capability of ADSCs compared with monolayer culture. When reseeded in normal serum-containing medium, the expression level of stemness biomarkers was even higher in spheroid-derived ADSCs than monolayer culture. Importantly, spheroid ADSCs could effectively promote the M2 polarization of macrophages both in vitro and in vivo. After transplantation into mouse, spheroid ADSCs improved the survival rate and significantly decreased serum levels of proinflammatory factors IL-1β and TNF-α following LPS challenge. In summary, we developed a 3D spheroid culture system through TeSR-E8 medium without the involvement of low adhesion conditions and special equipment, which provided a practical and convenient method for spheroid formation of MSCs with great potential for stem cell clinical therapy.

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mentioning

confidence: 99%

A Chemically Defined Serum-Free Culture System for Spontaneous Human Mesenchymal Stem Cell Spheroid Formation

Zhao

Xiao

et al. 2020

Stem Cells International

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“…In recent years, other researchers have used scaffolds, spheroid culture, or microcarrier-based 3D to culture MSCs [40][41][42]. Previously, Zhang et al [40] demonstrated that compared with 2D culture, human bone marrow-derived MSCs (hBM-MSCs) seeded in the 3D collagen scaffolds generated more exosomes with improved repair function in rats after traumatic brain injury.…”

Section: Discussionmentioning

confidence: 99%

Three-dimensional culture of MSCs produces exosomes with improved yield and enhanced therapeutic efficacy for cisplatin-induced acute kidney injury

et al. 2020

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Background: Exosomes derived from mesenchymal stem cells (MSC-exos) have been demonstrated with great potential in the treatment of multiple human diseases including acute kidney injury (AKI) by virtue of their intrinsic cargoes. However, there are major challenges of low yield and the lack of an established biomanufacturing platform to efficiently produce MSC-exos, thereby limiting their therapeutic application. Here, we aimed to establish a novel strategy to produce MSC-exos with a hollow fiber bioreactor-based three-dimensional (3D) culture system and evaluate the therapeutic efficacy of 3D-exosomes (3D-exos) on AKI. Methods: Mesenchymal stem cells (MSCs) were isolated from fresh human umbilical cord and cultured in twodimensional (2D) flasks. 2 × 10 8 MSCs were inoculated into the hollow fiber bioreactor for 3D culture. The culture supernatants were collected every 1 or 2 days for isolating exosomes. Exosomes from 2D (2D-exos) and 3D cultures were characterized by transmission electron microscopy, nanoparticle tracking analysis, and western blotting analysis of exosome markers. The yield of exosomes from 2 × 10 8 MSCs seeded in 2D and 3D culture system was compared, based on protein quantification. The therapeutic efficacy of 2D-exos and 3D-exos was investigated in a murine model of cisplatin-induced AKI in vivo and in vitro. Results: 3D culture did not significantly change the surface markers of MSCs, as well as the morphology, size, and exosomal markers of 3D-exos when compared to those of 2D-exos. Compared with conventional 2D culture, the 3D culture system increased total exosome production up to 19.4-fold. 3D-exos were more concentrated in the harvested supernatants (15.5-fold) than 2D-exos, which led to a higher exosome collection efficiency of 3D culture system. In vivo, both 2D-exos and 3D-exos significantly alleviated cisplatin-induced murine AKI evidenced by improved renal function, attenuated pathological changes of renal tubules, reduced inflammatory factors, and repressed T cell and macrophage infiltration. Impressively, 3D-exos were more effective than 2D-exos. Moreover, 3D-exos were taken up by tubular epithelial cells (TECs) with improved efficiency, thereby exhibiting superior antiinflammatory effect and improved viability of TECs in vitro.

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“…However, HS-27a showed a decrease in viable cells similarly to primary hMSCs and MS-5 are quiescent but do not shrink. Shrinking has already been reported for primary hMSCs [31,32,35,37,[67][68][69] and has been attributed to autophagy [32]. Reduced cell growth or arrest is known to trigger autophagy [70].…”

Section: Plos Onementioning

confidence: 90%

A comparative study of the capacity of mesenchymal stromal cell lines to form spheroids

et al. 2020

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Mesenchymal stem cells (MSC)-spheroid models favor maintenance of stemness, ex vivo expansion and transplantation efficacy. Spheroids may also be considered as useful surrogate models of the hematopoietic niche. However, accessibility to primary cells, from bone marrow (BM) or adipose tissues, may limit their experimental use and the lack of consistency in methods to form spheroids may affect data interpretation. In this study, we aimed to create a simple model by examining the ability of cell lines, from human (HS-27a and HS-5) and murine (MS-5) BM origins, to form spheroids, compared to primary human MSCs (hMSCs). Our protocol efficiently allowed the spheroid formation from all cell types within 24 hours. Whilst hMSC-spheroids began to shrink after 24 hours, the size of spheroids from cell lines remained constant during three weeks. The difference was partially explained by the balance between proliferation and cell death, which could be triggered by hypoxia and induced oxidative stress. Our results demonstrate that, like hMSCs, MSC cell lines make reproductible spheroids that are easily handled. Thus, this model could help in understanding mechanisms involved in MSC functions and may provide a simple model by which to study cell interactions in the BM niche.

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Three-Dimensional Spheroid Culture Increases Exosome Secretion from Mesenchymal Stem Cells

Cited by 112 publications

References 38 publications

A Chemically Defined Serum-Free Culture System for Spontaneous Human Mesenchymal Stem Cell Spheroid Formation

A Chemically Defined Serum-Free Culture System for Spontaneous Human Mesenchymal Stem Cell Spheroid Formation

Three-dimensional culture of MSCs produces exosomes with improved yield and enhanced therapeutic efficacy for cisplatin-induced acute kidney injury

A comparative study of the capacity of mesenchymal stromal cell lines to form spheroids

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