Bioactive LbL-Assembled Multilayer Nano-films Up-regulate Tenogenesis and Angiogenesis Enabling Robust Healing of Degenerative Rotator Cuff Tendon In Vivo
Degenerative changes of rotator cuff tendon are often diagnosed in elder patients. Spontaneous healing of degenerative tendon is rather inefficient as a result of its lack of suitable microenvironment for tendon stem cell differentiation and vascularization. Herein, multilayer hyaluronic acid/chitosan (HA/CH) nano-films were assembled by layer-by-layer assembly method onto aligned poly(ε-caprolactone) (PCL) nanofibers towards degenerative tendon healing. Materials testing showed that the layer number of HA/CH nano-films could adjust hydrophilicity and wettability of the nanofiberous membranes. In vitro, the membrane with an optimal 8-layer (termed as 8LP) afforded aligned morphology of tendon stem/progenitor cells (TSPCs) and up-regulated mRNA expression of tenogenic markers (SCX, BGN). In a rabbit model with disorganized rotator cuff tendon, 8LP group up-regulated mRNA level of collagen I/III and tenascin (TNC) at 6-week, but not 12-week, post-surgery as compared to native PCL group. Next, vascular endothelial growth factor-loaded 8LP (termed as 8LP-V) was prepared. Compared to 8LP, 8LP-V produced higher level of angiogenesis in the tendon at 6- or 12-week post-surgery, thus supplying endogenous pre-tendon growth factors (TGF-β, IGF-1) to further enhance tenogenic transcriptional factors. As a result, 8LP-V yielded thicker collagen fibers and/or higher tendon stiffness as compared to 8LP and clinical pericardial patch groups. This study highlights rational design of LbL-assembled multilayer HA/CH films to up-regulate tenogenesis for robust healing of degenerative rotator cuff tendon.