Anti-Inflammatory Effect of Adipose-Derived Mesenchymal Stem Cells for Flexor Tendon Healing

Despite advances in surgical techniques over the past three decades, tendon repairs remain prone to poor clinical outcomes. While previous attempts to improve flexor tendon healing have focused on the later stages of healing: i.e., proliferation and matrix synthesis), the early inflammatory phase of tendon healing remains poorly understood and its modulation during healing has not yet been studied. The overall goal of this work is therefore to determine whether flexor tendon healing can be enhanced through modulation of the inflammatory environment after surgical repair. To meet this objective, the inflammatory response after flexor tendon injury and repair was first characterized to help identify inflammation-related targets for future treatments. Temporal changes in immune cell population and gene expression of inflammation-, matrix degradation-, and extracellular matrix-related factors were examined. Of the observed changes, the most dramatic effect was the greater than 4000-fold up-regulation in the expression of the pro-inflammatory factor IL-1β. While some inflammation is essential for healing to occur, such high levels of pro-inflammatory cytokines may result in collateral tissue damage and impair tendon healing. To examine the effect of inflammatory cytokines on tendon fibroblasts, the native cell type that must eventually re-build the injured tendon, we established two in vitro models of inflammation: one induced by exogenous IL-1β and one induced by macrophages: the primary cellular source of IL-1β). We found that IL-1β and other pro-inflammatory cytokines secreted by macrophages induce further up-regulation of pro-inflammatory factors by tendon fibroblasts. Furthermore, exposure of tendon fibroblasts to an inflammatory environment led to up-regulation of factors related to matrix degradation and down-regulation of factors related to extracellular matrix formation. These changes in expression are likely detrimental to tendon healing in vivo. In an effort to reduce the negative effects of inflammatory cytokines on tendon fibroblasts, adipose-derived mesenchymal stem cells: ASCs) were incorporated into the two in vitro models and their ability to modulate inflammation was investigated. While ASCs were unable to counteract the effects of IL-1β directly, ASC co-culture with macrophages successfully suppressed the negative effects of macrophages on tendon fibroblasts. FACS analysis revealed that ASCs induced a phenotypic switch from a pro-inflammatory macrophage phenotype to an anti-inflammatory macrophage phenotype, thus resulting in exposure of tendon fibroblasts to fewer pro-inflammatory cytokines. In order to translate these in vitro findings in vivo, a surgically manageable scaffold capable of delivering and retaining ASCs at the repair site was developed. The fibrin/nanofiber scaffolds were well-tolerated in the in vivo flexor tendon repair setting and successful delivery of ASCs was achieved. Pilot data demonstrated promise with this approach, as demonstrated by down-regulation of IL-1β and MMP1 in the ASC treated animals. However, the pilot study was underpowered, and additional animals and assays are required to fully evaluate the effect of ASCs in vivo