Micro-scale and meso-scale architectural cues cooperate and compete to direct aligned tissue formation

Abstract

Tissue and biomaterial microenvironments provide architectural cues that direct important cell behaviors including cell shape, alignment, migration, and resulting tissue formation. These architectural features may be presented to cells across multiple length scales, from nanometers to millimeters in size. In this study, we examined how architectural cues at two distinctly different length scales, “micro-scale” cues on the order of ~1–2 μm, and “meso-scale” cues several orders of magnitude larger (>100 μm), interact to direct aligned neo-tissue formation. Utilizing a micro-photopatterning (μPP) model system to precisely arrange cell-adhesive patterns, we examined the effects of substrate architecture at these length scales on human mesenchymal stem cell (hMSC) organization, gene expression, and fibrillar collagen deposition. Both micro- and meso-scale architectures directed cell alignment and resulting tissue organization, and when combined, meso cues could enhance or compete against micro-scale cues. As meso boundary aspect ratios were increased, meso-scale cues overrode micro-scale cues and controlled tissue alignment, with a characteristic critical width (~500 μm) similar to boundary dimensions that exist in vivo in highly aligned tissues. Meso-scale cues acted via both lateral confinement (in a cell-density-dependent manner) and by permitting end-to-end cell arrangements that yielded greater fibrillar collagen deposition. Despite large differences in fibrillar collagen content and organization between μPP architectural conditions, these changes did not correspond with changes in gene expression of key matrix or tendon-related genes. These findings highlight the complex interplay between geometric cues at multiple length scales and may have implications for tissue engineering strategies, where scaffold designs that incorporate cues at multiple length scales could improve neo-tissue organization and resulting functional outcomes.

Aligned multilayered electrospun scaffolds for rotator cuff tendon tissue engineering

“…However, initial cell seeding density in that study was 10-fold lower than the current study, and cells were not confluent on the surface of the scaffolds after 28 days of culture. Emerging evidence suggests that response to microarchitectural cues, cell proliferation, migration, symmetry of cell-cell contacts, and direction of cellular and matrix alignment are tightly coordinated [61–65]. In the initial period after seeding on electrospun fibers, MSCs demonstrate potent directionality in their migration response and in their cell-cell contacts on aligned compared to nonaligned electrospun fibers [66].…”

Aligned Nanofiber Topography Directs the Tenogenic Differentiation of Mesenchymal Stem Cells

“…Previous studies have investigated tendon gene expression from cells cultured on fibrous scaffolds where tendon gene expression was not significantly altered, although this was a comparison between aligned and random scaffolds [37]. In another study, Gilchrist et al compared aligned (parallel) and grid (perpendicular) architectural cues and observed an increase of gene expression of SCX and COL I from day 6 to day 12, with no significant differences in tendon gene expression between designs [22], similar to results observed in our scaffolds. Our findings suggest that alignment, rather than orientation, may be the predominant cue influencing tenogenic differentiation.…”

Micropatterning Strategies to Engineer Controlled Cell and Tissue Architecture in Vitro

“…Also, micro-gels that are guided by magnetic forces to assemble much like Tetris blocks have recently been introduced, in order to achieve higher-order multi-component tissues (3). Competing cues at different scales will likely also add complexity in the 3D situation, as has been observed in 2D (69). Overall, the need for architecturally controlled multilayered tissues represents a great challenge for tissue patterning and architectural engineering in the future.…”