Three-dimensional structural niches engineered via two-photon laser polymerization promote stem cell homing

A strategy to modulate the behavior of stem cells in culture is to mimic structural aspects of the native cell/extracellular matrix interaction. We applied femtosecond laser two-photon polymerization (2PP) to fabricate 3D microscaffolds, or "niches", using a hybrid organic–inorganic sol–gel called SZ2080. The niches, of sizes fitting in a cubic volume of 100 x 100 x 100 µm^3, were made by an external containment grid of horizontal parallel elements and by an internal 3D lattice. We developed two niche heights, 20 and 80-100 µm, and four lattice pore dimensions (10, 20, 30 µm, and graded). We used primary rat mesenchymal stem cells (MSC) to study cell viability, migration and proliferation in the niches, up to 6 culture days. MSCs preferentially stayed on/in the structures once they run into them through random migration from the surrounding flat surface, invaded those with a lattice pore dimension greater than 10 µm, and adhered to the internal lattice while the cell nuclei acquired a roundish morphology. In the niches, the highest MSC density was found in those areas where proliferation was observed, corresponding to the regions where the scaffold surface density available for cell adhesion was highest. The microgeometry inducing the highest cell density was the 20 µm high with graded pores, in which cell invasion was favored in the central region of large porosity and cell adhesion was favored in the lateral regions of high scaffold surface density. Cell density in the niches, 17±6 cells/(100x100 µm^2), did not significantly differ from that of the flat surface colonies. This implies that MSCs spontaneously homed and established colonies within the 3D niches. This study enlightens the crucial role played by the niche 3D geometry on MSC colonization in culture, with potential implications on the design of biomaterial scaffolds for synthetic niche engineering.