Data in support of effects of cell–cell contact and oxygen tension on chondrogenic differentiation of stem cells

While cell condensation has been thought to enhance chondrogenesis, no direct evidence so far confirms that cell-cell contact itself increases chondrogenic differentiation of stem cells, since the change of cell-cell contact is usually coupled with those of other cell geometry cues and soluble factors in cell culture. The present study semi-quantitatively examined the effect of cell-cell contact in a decoupled way. We fabricated two-dimensional micropatterns with cell-adhesive peptide arginine-glycine-aspartate (RGD) microdomains on a nonfouling poly(ethylene glycol) (PEG) hydrogel. Mesenchymal stem cells (MSCs) were well localized on the microdomains for a long time. Based on our micropattern design, single MSCs or cell clusters with given cell numbers (1, 2, 3, 6 and 15) and a similar spreading area per cell were achieved on the same substrate, thus the interference of soluble factor difference from cell autocrine and that of cell spreading area were ruled out. After 9-day chondrogenic induction, collagen II was stained to characterize the chondrogenic induction results; the mRNA expression levels of SOX9, collagen II, aggrecan, HIF-1α and collagen I were also detected. The statistics confirmed unambiguously that the extent of the chondrogenic differentiation increased with cell-cell contact, and even a linear relation between differentiation extent and contact extent was established within the examined range. The cell-cell contact effect worked under both hypoxia (5% O2) and normoxia (21% O2) conditions, and the hypoxia condition promoted the chondrogenic induction of MSCs on adhesive microdomains more efficiently than the normoxia condition under the same cell-cell contact extents.