Dynamic Culture of Mesenchymal Stromal/Stem Cell Spheroids and Secretion of Paracrine Factors

Abstract

In recent years, conditioned medium (CM) obtained from the culture of mesenchymal stromal/stem cells (MSCs) has been shown to effectively promote tissue repair and modulate the immune response in vitro and in different animal models, with potential for application in regenerative medicine. Using CM offers multiple advantages over the implantation of MSCs themselves: 1) simpler storage, transport, and preservation requirements, 2) avoidance of the inherent risks of cell transplantation, and 3) potential application as a ready-to-go biologic product. For these reasons, a large amount of MSCs research has focused on the characterization of the obtained CM, including soluble trophic factors and vesicles, preconditioning strategies for enhancing paracrine secretion, such as hypoxia, a three-dimensional (3D) environment, and biochemical stimuli, and potential clinical applications. In vitro preconditioning strategies can increase the viability, proliferation, and paracrine properties of MSCs and therefore improve the therapeutic potential of the cells and their derived products. Specifically, dynamic cultivation conditions, such as fluid flow and 3D aggregate culture, substantially impact cellular behaviour. Increased levels of growth factors and cytokines were observed in 3D cultures of MSC grown on orbital or rotatory shaking platforms, in stirred systems, such as spinner flasks or stirred tank reactors, and in microgravity bioreactors. However, only a few studies have established dynamic culture conditions and protocols for 3D aggregate cultivation of MSCs as a scalable and reproducible strategy for CM production. This review summarizes significant advances into the upstream processing, mainly the dynamic generation and cultivation of MSC aggregates, for de CM manufacture and focuses on the standardization of the soluble factor production.

Mesenchymal stromal cell derived extracellular vesicles as a therapeutic tool: immune regulation, MSC priming, and applications to SLE

“…3D culture of MSCs englobes a great variety of techniques, including but not limited to microcarriers, diverse scaffolds, microgels, and spheroids, which can be then cultured in bioreactors or other types of vessels. 3D culture better mimics the natural cell conditions, and increases the levels of secreted angiogenic and immunomodulatory factors ( 178 ).…”

Advances in the application of extracellular vesicles derived from three-dimensional culture of stem cells

“…Bioreactor-based 3D culture could enable large-scale production of SCs [ 136 ]. Furthermore, 3D culture enhances the immunomodulatory and angiogenesis ability of MSCs, strengthens their paracrine activity, and promotes the secretion of cytokines and growth factors [ 137 , 138 ]. 3D culture can alter the characteristics of SCs, and EVs are secreted by them, similarly, some studies have confirmed that 3D culture will have an effect on EVs.…”

The Growing Importance of Three-Dimensional Models and Microphysiological Systems in the Assessment of Mycotoxin Toxicity

“…Dynamic culture in bioreactors brings several advantages over static methods for the generation of spheroids, including a homogeneous environment, better diffusion of oxygen and nutrients, and longer periods of cultivation [ 48 , 49 ]. The stirred-tank bioreactor (STR) allows an easy and large-scale production of spheroids using culture modalities that allow the control of several parameters, including oxygen and pH [ 48 , 50 ]. Their main disadvantage is related to the relatively high shear forces that could have a detrimental effect on the spheroids’ shape and structure.…”