Department of Plastic and Reconstructive Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, China.
Stem Cell Res Ther. 2020 Aug 14;11(1):355. doi: 10.1186/s13287-020-01838-w.
Extensive passage of adipose-derived stem cells (ASCs) in vitro leads to loss of function. Endothelial colony-forming cells (ECFCs) can be isolated from adult peripheral blood. A 3D co-culture system may rescue in vitro ASC senescence.
A 3D co-culture model was successfully established using hyaluronic acid (HA) gel and a 10:1 ratio of late-passage ASCs and ECFCs. Cell density and culture conditions were optimized. Stem cell phenotype was characterized by flow cytometry. ELISA was used to measure the trophic effect of angiogenic growth factors and compare the effects of these factors between the 3-D co-culture and single-cell culture. Therapeutic potential of ASC/ECFC 3-D co-cultures was evaluated in a mouse chronic injury model.
Following incubation in a HA substrate 3D co-culture system, ASC morphology, phenotype, secretory profile, and differentiation capacity were restored. The ASC/ECFC co-culture increased the secretion of cytokines, such as hepatocyte growth factor, compared with single-cell 3D culture or monolayer culture. Mice radiation-ulcer wounds treated with ASC/ECFC 3-D co-cultures (spheroids) showed epithelialization and improved healing compared with wounds treated with ASCs or ECFCs alone. Further, transplanted ASC/ECFC spheroids exhibited superior angiogenic potential due to the ability of the ASCs to transdifferentiate into pericytes.
3D co-culture of ECFCs and ASCs in vitro restored native ASC properties by reversing cellular senescence and loss of trophic function. Transplant of ASC/ECFC 3D spheroids in vivo demonstrated pro-angiogenic capacity with improved therapeutic potential.
体外大量传代脂肪源性干细胞（ASCs）会导致功能丧失。内皮祖细胞（ECFCs）可从成人外周血中分离得到。三维共培养系统可能会挽救体外 ASC 衰老。
我们成功地使用透明质酸（HA）凝胶和晚期传代 ASC 与 ECFC 以 10:1 的比例建立了 3D 共培养模型。优化了细胞密度和培养条件。通过流式细胞术对干细胞表型进行了特征描述。ELISA 用于测量血管生成生长因子的营养作用，并比较 3D 共培养和单细胞培养之间这些因子的作用。在小鼠慢性损伤模型中评估了 ASC/ECFC 3D 共培养的治疗潜力。
在 HA 基质 3D 共培养系统孵育后，ASC 的形态、表型、分泌谱和分化能力得到恢复。与单细胞 3D 培养或单层培养相比，ASC/ECFC 共培养增加了细胞因子（如肝细胞生长因子）的分泌。与单独使用 ASCs 或 ECFCs 治疗相比，用 ASC/ECFC 3D 共培养（球体）治疗的小鼠辐射性溃疡伤口表现出上皮化和改善的愈合。此外，由于 ASC 能够转分化为周细胞，因此移植的 ASC/ECFC 球体表现出优越的血管生成潜能。
体外 ECFC 和 ASC 的 3D 共培养通过逆转细胞衰老和营养功能丧失恢复了 ASC 的固有特性。体内移植 ASC/ECFC 3D 球体显示出促血管生成能力，具有改善的治疗潜力。
