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Functional fibrocartilage regeneration is a bottleneck during bone-tendon healing, and the currently available tissue-engineering strategies for fibrocartilage regeneration are insufficient because of a lack of appropriate scaffold that can load large seeding-cells and induce chondrogenesis of stem cells. The acellular fibrocartilage scaffold (AFS) contains active growth factors as well as tissue-specific epitopes for cell-matrix interactions, which make it a potential scaffold for tissue-engineered fibrocartilage. A limitation to this scaffold is that its low porosity inhibits cells loading and infiltration. Here, inspired by book appearance, we sectioned native fibrocartilage tissue (NFT) into book-shape to improve cells loading and infiltration, and then decellularized with four protocols: (1) 2% SDS for 6-h, (2) 2% SDS for 24-h, (3) 4 SDS for 6-h, (4) 4% SDS for 24-h, followed by nuclease digestion. The optimal protocol was screened with respect to microstructures, DNA residence, native ingredients reservation, and chondrogenic inducibility of the AFS. In vitro studies demonstrated that this screened scaffold is noncytotoxicity and low-immunogenicity, allows adipose-derived stromal cells (ASCs) attachment and proliferation, shows superior chondrogenic inducibility, and stimulates collagen or glycosaminoglycans secretion. The underlying mechanism for this chondrogenic inducibility may be related to hedgehog pathway activating. Additionally, a novel pattern for fabricating tissue-engineered fibrocartilage was developed to enlarge seeding-cells loading, namely, cell-sheets sandwiched by book-shaped scaffold. In-vivo studies indicate that this screened scaffold alone could induce endogenous cells to satisfactorily regenerate fibrocartilage at 16-week, as characterized by fibrocartilaginous extracellular matrix (ECM) deposition and good interface integration. Interleaving this book-shaped AFS with autologous ASCs-sheets significantly enhanced its ability to regenerate fibrocartilage. Cell tracking demonstrated that fibrochondrocytes, osteoblasts, and osteocytes in the healing interface at postoperative 8-week partly originated from the sandwiched ASCs-sheets. On that basis, we propose the use of this book-shaped AFS and cell sheet technique for fabricating tissue-engineered fibrocartilage to improve bone-tendon healing.

Surgical repair of rotator cuff tears presents a significant clinical challenge with high failure rates and inferior functional outcomes. Graft augmentation improves repair outcomes; however, currently available grafting materials have limitations. Although cell-seeded decellularized tendon slices may facilitate cell infiltration, promote tendon incorporation, and preserve original mechanical strength, the unique fibrocartilage zone is yet to be successfully reestablished. In this study, we investigated the biological and mechanical properties of an engineered tendon-fibrocartilage-bone composite (TFBC) with cyclic tension (3% strain; 0.2 Hz). Decellularized TFBCs seeded with bone marrow-derived mesenchymal stem cell (BMSCs) sheets and subjected to mechanical stimulation for up to 7 days were characterised by histology, immunohistochemistry, scanning electron microscopy, mechanical testing, and transcriptional regulation. The decellularized TFBC maintained native enthesis structure and properties. Mechanically stimulated TFBC-BMSC constructs displayed increased cell migration after 7 days of culture compared with static groups. The seeded cell sheet not only integrated well with tendon scaffold but also distributed homogeneously and aligned to the direction of stretch under dynamic culture. Developmental genes were regulated including scleraxis, which was significantly upregulated with mechanical stimulation. The Young's modulus of the cell-seeded constructs was significantly higher compared with the noncell-seeded controls. In conclusion, the results of this study reveal that the TFBC-BMSC composite provides an ideal multilayer construct for cell seeding and growth, with mechanical preconditioning further enhances cell penetration and differentiation. The BMSC cell sheet revitalised TFBC in conjunction with mechanical stimulation could serve as a novel and primed biological patch to improve rotator cuff repair., (Copyright © 2018 John Wiley & Sons, Ltd.)

At the tendon-to-bone insertion, there is a unique transitional structure: tendon, non-calcified fibrocartilage, calcified fibrocartilage, and bone. The reconstruction of this special graded structure after defects or damage is an important but challenging task in orthopedics. In particular, reconstruction of the fibrocartilage zone has yet to be successfully achieved. In this study, the development of a novel book-shape scaffold derived from the extracellular matrix of fibrocartilage was reported. Specifically, fibrocartilage from the pubic symphysis was obtained from rabbits and sliced into the shape of a book (dimensions: 10 mm × 3 mm × 1 mm) with 10 layers, each layer (akin to a page of a book) with a thickness of 100-μm. These fibrocartilage "book" scaffolds were decellularized using sequentially 3 freeze-thaw cycles, 0.1% Triton X-100 with 1.5 M KCl, 0.25% trypsin, and a nuclease. Histology and DNA quantification analysis confirmed substantial removal of cells from the fibrocartilage scaffolds. Furthermore, the quantities of DNA, collagen, and glycosaminoglycan in the fibrocartilage were markedly reduced following decellularization. Scanning electron microscopy confirmed that the intrinsic ultrastructure of the fibrocartilage tissue was well preserved. Therefore, the results of this study suggest that the novel "book" fibrocartilage scaffold could have potential applications in tissue engineering.

Meniscal tears in the avascular zone have a poor self-healing potential, however partial meniscectomy predisposes the knee for early osteoarthritis. Tissue engineering with mesenchymal stem cells and a hyaluronan collagen based scaffold is a promising approach to repair meniscal tears in the avascular zone. 4 mm longitudinal meniscal tears in the avascular zone of lateral menisci of New Zealand White Rabbits were performed. The defect was left empty, sutured with a 5-0 suture or filled with a hyaluronan/collagen composite matrix without cells, with platelet rich plasma or with autologous mesenchymal stem cells. Matrices with stem cells were in part precultured in chondrogenic medium for 14 days prior to the implantation. Menisci were harvested at 6 and 12 weeks. The developed repair tissue was analyzed macroscopically, histologically and biomechanically. Untreated defects, defects treated with suture alone, with cell-free or with platelet rich plasma seeded implants showed a muted fibrous healing response. The implantation of stem cell-matrix constructs initiated fibrocartilage-like repair tissue, with better integration and biomechanical properties in the precultured stem cell-matrix group. A hyaluronan-collagen based composite scaffold seeded with mesenchymal stem cells is more effective in the repair avascular meniscal tear with stable meniscus-like tissue and to restore the native meniscus., (Copyright © 2013 Wiley Periodicals, Inc., a Wiley Company.)