ASC Spheroid Geometry and Culture Oxygenation Differentially Impact Induction of Preangiogenic Behaviors in Endothelial Cells

Abstract

Cell-based angiogenic therapies offer potential for the repair of ischemic injuries, while avoiding several of the limitations associated with material-based growth factor delivery strategies. Evidence supports that applying MSCs as spheroids rather than dispersed cells can improve retention and enhance therapeutic effect through increased secretion of angiogenic factors due to hypoxia. However, while spheroid culture appears to modulate MSC behavior, there has been little investigation of how major culture parameters that affect cellular oxygen tension, such as external oxygenation and culture size, impact the angiogenic potential of spheroids. We cultured equal numbers of adipose-derived stem cells (ASCs) as spheroids containing 10,000 (10k) or 60,000 (60k) cells each, in 20% and 2% oxygen. VEGF secretion varied among the sample groups, with 10k, 2% O2 spheroids exhibiting the highest production. Spheroid-conditioned media was applied to HUVEC monolayers, and proliferation was assessed. Spheroids of either size in 2% oxygen induced comparable proliferation compared to a 2 ng/ml VEGF control sample, while spheroids in 20% oxygen induced less proliferation. Spheroids were also applied in coculture with HUVEC monolayers, and induction of migration through a Transwell membrane was evaluated. Sixty thousand, 2% O2 spheroids induced similar levels of migration as VEGF controls, while 10k, 2% O2 spheroids induced significantly more. Ten thousand, 20% spheroids performed no better than VEGF-free controls. We conclude that the therapeutic ability of ASC spheroids to stimulate angiogenesis in endothelial cells is affected by both culture size and oxygenation parameters, suggesting that, while ASC spheroids offer potential in the treatment of injured and ischemic tissues, careful consideration of culture size in respect to in vivo local oxygen tension will be necessary for optimal results.

Wound Healing Potential of Chlorogenic Acid and Myricetin-3-O-β-Rhamnoside Isolated from Parrotia persica

“…Fibroblasts migrate toward the site of injury, proliferate, and establish an extracellular matrix to facilitate tissue reconstruction [ 5 ]. Endothelial cells are activated by signals expressed from keratinocytes and fibroblasts, leading to migration into the site to form new capillaries [ 6 , 7 , 8 , 9 , 10 ]. Wound contraction occurs over time in association with myofibroblasts and fibroblasts, which generate collagen and mature scars [ 11 ].…”

Systematic review on spheroids from adipose‐derived stem cells: Spontaneous or artefact state?

“…oxygen conditions compared to a 20% oxygen culture condition (Skiles et al, 2013), (Skiles et al, 2015). FABP4) in the spheroids grown in osteoblastic, chondrocytic and adipogenic media compared to the control group (Kapur et al, 2012).…”

Spheroid Mesenchymal Stem Cells and Mesenchymal Stem Cell-Derived Microvesicles: Two Potential Therapeutic Strategies

“…However, the ability of spheroid MSCs in stimulating angiogenesis and thus treating ischemic injuries can be affected by culture parameters such as culture size and external oxygenation. For example, MSC spheroids with a size of 10,000 cells cultured under 2% O 2 exhibited better production of vascular endothelial growth factor (VEGF) than spheroids containing 60,000 cells that were cultured under 20% O 2 [66]. Therefore, careful considerations should be given to both culture size and local oxygen tension to achieve optimal and consistent therapeutic results.…”