Enhancing Tendon Regeneration: Investigating the Impact of Topography on the Secretome of Adipose-Derived Stem Cells

Abstract Background : Tendons are vital for maintaining integrity and movement, but current treatment options are insufficient for restoring them after injuries. Previous studies have shown that the secretome from mesenchymal stem cells (MSCs) promoted tendon regeneration. However, limited studies have explored the impact of the cellular microenvironment on the secretome of MSCs in tendon repair. This study aims to investigate how the topographic orientation impacts the secretome of human adipose-derived stem cells (ADSCs) and its effect on tendon repair and regeneration. Methods: Randomly oriented and aligned silk scaffolds were prepared by directional freeze-drying. Conditioned medium (CM) was generated from ADSCs cultured on the scaffolds with different topography (RCM: random scaffolds; ACM: aligned scaffolds). In vitro experiments were performed to access the effect of RCM and ACM on cell proliferation by live/dead staining, CCK-8 incubation, and Ki67 immunofluorescence. The effect on tenogenic differentiation of tendon stem/progenitor cells (TSPCs) and polarization of macrophages was confirmed by detecting the expression of related genes. Subsequently, RCM and ACM were injected into rats with patellar tendon defects. Tissue repair and immunomodulatory effects were evaluated through histological and immunohistochemical staining. Result: In vitro results showed that the ACM group had a more potent effect in promoting the proliferation of TSPCs as compared to RCM group. ACM group promoted tenogenic differentiation of TSPCs, as evidenced by higher expression of SCX, TNMD, and MKX in contrast to RCM. In addition, ACM group up-regulated the expression of M2-related anti-inflammatory genes including ARG-1 and IL-10, and down-regulated M1-related inflammatory genes including CCR7, iNOS, and IL-1β in RAW 264.7 cells, as compared to RCM group. The ACM group exhibited a greater formation of tendon-like tissues, as confirmed by histological evaluation, and a higher expression of tendon-related specific proteins, including SCX, TNMD, and COL I as shown by immunohistochemistry as compared to RCM group. The tissue sections of the ACM group showed a high expression of the M2 anti-inflammatory polarity-related protein ARG-1, and a low expression of the M1 pro-inflammatory polarity-related protein iNOS. These results were consistent with the in vitro findings. Conclusions : This study highlights the topographical dependency of ADSCs paracrine activities and demonstrates the potential of using oriented silk scaffolds to enhance the ADSCs secretome for tendon regeneration. These findings offer a promising, safer, and non-cell-based treatment option for tissue regeneration.