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Rapid and functional bone-tendon (B-T) healing remains a difficulty in clinical practice. Tissue engineering has emerged as a promising strategy to address this problem. However, the majority of tissue engineering scaffolds are loaded with stem cells to enhance the regenerability in B-T healing, which is complicated and inconvenient for clinical application. Accordingly, developing a cell-free scaffold with chemotactic function and chondrogenic inducibility may be an effective approach. In this study, a collagen affinity peptide derived from the A3 domain of von Willebrand factor (a hemostasis factor) was fused into the C-terminal of a stromal cell-derived factor-1α (SDF-1α) to synthesize a recombinant SDF-1α capable of binding collagen and chemotactic activity. The recombinant SDF-1α was then tethered on the collagen fibers of a book-shaped acellular fibrocartilage scaffold (BAFS), thus fabricating a novel scaffold (C-SDF-1α/BAFS) with chemotactic function and chondrogenic inducibility. In vitro tests determined that this scaffold was noncytotoxic and biomimetic, could attract stem cells migrating to the scaffold using sustainably released C-SDF-1α, and inducedthe interacting stem cells down the chondrogenic lineage. In vivo, the C-SDF-1α/BAFS significantly enhanced the B-T healing in a rabbit partial patellectomy model, as shown by the larger cartilaginous metaplasia region, better fibrocartilage regeneration, additional bone formation, and improved biomechanical properties. Therefore, the findings of the study demonstrate that the C-SDF-1α/BAFS could potentially be applied for B-T healing., (© 2020 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.)

Reducing rotator cuff failure after repair remains a challenge due to suboptimal tendon-to-bone healing. In this study we report a novel biomaterial with engineered tendon-fibrocartilage-bone composite (TFBC) and bone marrow-derived mesenchymal stem cell sheet (BMSCS); this construct was tested for augmentation of rotator cuff repair using a canine non-weight-bearing (NWB) model. A total of 42 mixed-breed dogs were randomly allocated to 3 groups (n = 14 each). Unilateral infraspinatus tendon underwent suture repair only (control); augmentation with engineered TFBC alone (TFBC), or augmentation with engineered TFBC and BMSCS (TFBC + BMSCS). Histomorphometric analysis and biomechanical testing were performed at 6 weeks after surgery. The TFBC + BMSCS augmented repairs demonstrated superior histological scores, greater new fibrocartilage formation and collagen fiber organization at the tendon-bone interface compared with the controls. The ultimate failure load and ultimate stress were 286.80 ± 45.02 N and 4.50 ± 1.11 MPa for TFBC + BMSCS group, 163.20 ± 61.21 N and 2.60 ± 0.97 MPa for control group (TFBC + BMSCS vs control, P = 1.12E-04 and 0.003, respectively), 206.10 ± 60.99 N and 3.20 ± 1.31 MPa for TFBC group (TFBC + BMSCS vs TFBC, P = 0.009 and 0.045, respectively). In conclusion, application of an engineered TFBC and BMSCS can enhance rotator cuff healing in terms of anatomic structure, collagen organization and biomechanical strength in a canine NWB model. Combined TFBC and BMSCS augmentation is a promising strategy for rotator cuff tears and has a high potential impact on clinical practice., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

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.)

The temporomandibular joint disk (TMJd) is an extremely dense and avascular fibrocartilaginous extracellular matrix (ECM) resulting in a limited regenerative capacity. The use of decellularized TMJd as a biocompatible scaffold to guide tissue regeneration is restricted by innate subcellular porosity of the ECM that hinders cellular infiltration and regenerative events. Incorporation of an artificial microporosity through laser micro-ablation (LMA) can alleviate these cell and diffusion based limitations. In this study, LMA was performed either before or after decellularization to assess to effect of surfactant treatment on porosity modification as well as the resultant mechanical and physical scaffold properties. Under convective flow or agitation schemes, pristine and laser ablated disks were decellularized using either low (0.1% w/v) or high (1% w/v) concentrations of sodium dodecyl sulfate (SDS). Results show that lower concentrations of SDS minimized collagen degradation and tissue swelling while retaining its capacity to solubilize cellular content. Regardless of processing scheme, laser ablated channels incorporated after SDS treatment were relatively smaller and more uniform than those incorporated before SDS treatment, indicating an altered laser interaction with surfactant treated tissues. Smaller channels correlated with less disruption of native biomechanical properties indicating surfactant pre-treatment is an important consideration when using LMA to produce artificial porosity in ex vivo derived tissues. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 1858-1868, 2018., (© 2017 Wiley Periodicals, Inc.)

Advances in cartilage tissue engineering have led to constructs with mechanical integrity and biochemical composition increasingly resembling that of native tissues. In particular, collagen cross-linking with lysyl oxidase has been used to significantly enhance the mechanical properties of engineered neotissues. In this study, development of collagen cross-links over time, and correlations with tensile properties, were examined in self-assembling neotissues. Additionally, quantitative MRI metrics were examined in relation to construct mechanical properties as well as pyridinoline cross-link content and other engineered tissue components. Scaffold-free meniscus fibrocartilage was cultured in the presence of exogenous lysyl oxidase, and assessed at multiple time points over 8weeks starting from the first week of culture. Engineered constructs demonstrated a 9.9-fold increase in pyridinoline content, reaching 77% of native tissue values, after 8weeks of culture. Additionally, engineered tissues reached 66% of the Young's modulus in the radial direction of native tissues. Further, collagen cross-links were found to correlate with tensile properties, contributing 67% of the tensile strength of engineered neocartilages. Finally, examination of quantitative MRI metrics revealed several correlations with mechanical and biochemical properties of engineered constructs. This study displays the importance of culture duration for collagen cross-link formation, and demonstrates the potential of quantitative MRI in investigating properties of engineered cartilages., Statement of Significance: This is the first study to demonstrate near-native cross-link content in an engineered tissue, and the first study to quantify pyridinoline cross-link development over time in a self-assembling tissue. Additionally, this work shows the relative contributions of collagen and pyridinoline to the tensile properties of collagenous tissue for the first time. Furthermore, this is the first investigation to identify a relationship between qMRI metrics and the pyridinoline cross-link content of an engineered collagenous tissue., (Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

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.

Regeneration of complex tissues, such as kidney, liver, and cartilage, continues to be a scientific and translational challenge. Survival of ex vivo cultured, transplanted cells in tissue grafts is among one of the key barriers. Meniscus is a complex tissue consisting of collagen fibers and proteoglycans with gradient phenotypes of fibrocartilage and functions to provide congruence of the knee joint, without which the patient is likely to develop arthritis. Endogenous stem/progenitor cells regenerated the knee meniscus upon spatially released human connective tissue growth factor (CTGF) and transforming growth factor-β3 (TGFβ3) from a three-dimensional (3D)-printed biomaterial, enabling functional knee recovery. Sequentially applied CTGF and TGFβ3 were necessary and sufficient to propel mesenchymal stem/progenitor cells, as a heterogeneous population or as single-cell progenies, into fibrochondrocytes that concurrently synthesized procollagens I and IIα. When released from microchannels of 3D-printed, human meniscus scaffolds, CTGF and TGFβ3 induced endogenous stem/progenitor cells to differentiate and synthesize zone-specific type I and II collagens. We then replaced sheep meniscus with anatomically correct, 3D-printed scaffolds that incorporated spatially delivered CTGF and TGFβ3. Endogenous cells regenerated the meniscus with zone-specific matrix phenotypes: primarily type I collagen in the outer zone, and type II collagen in the inner zone, reminiscent of the native meniscus. Spatiotemporally delivered CTGF and TGFβ3 also restored inhomogeneous mechanical properties in the regenerated sheep meniscus. Survival and directed differentiation of endogenous cells in a tissue defect may have implications in the regeneration of complex (heterogeneous) tissues and organs., (Copyright © 2014, American Association for the Advancement of Science.)

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.)