Bioactive Decellularized Tendon-Derived Stem Cell Sheet for Promoting Graft Healing After Anterior Cruciate Ligament Reconstruction

Abstract

Background: Stem cell sheets provide a scaffold-free option for the promotion of graft healing after anterior cruciate ligament reconstruction (ACLR). However, cell viability, stability, and potential uncontrolled actions create challenges for clinical translation. The decellularization of cell sheets may overcome these problems as studies have shown that the natural extracellular matrix of stem cells is bioactive and can promote tissue repair. Hypothesis: The decellularized tendon-derived stem cell (dTDSC) sheet can promote graft healing after ACLR. Study Design: Controlled laboratory study. Methods: An optimized decellularization protocol was developed to decellularize the TDSC sheets. A total of 64 Sprague-Dawley rats underwent ACLR with or without the dTDSC sheet wrapping the tendon graft (n = 32/group). At 2 and 6 weeks after surgery, graft healing was assessed by micro–computed tomography, histology, and biomechanical testing. The accumulation of iNOS+ and CD206+ cells and the expression of metalloproteinase 1 (MMP-1), MMP-13, and tissue inhibitor of metalloprotease 1 (TIMP-1) were assessed by immunohistochemistry. Results: The decellularization was successful, with the removal of 98.4% nucleic acid while preserving the collagenous proteins and bioactive factors. The expression of bone morphogenetic protein 2 (BMP-2) and VEGF in the dTDSC sheet was comparable with the TDSC sheet ( P > .05). Micro–computed tomography showed significantly more tunnel bone formation in the dTDSC sheet group. The dTDSC sheet group demonstrated better graft osteointegration and higher integrity of graft midsubstance with significantly higher ultimate failure load (16.58 ± 7.24 vs 8.93 ± 2.45 N; P = .002) and stiffness (11.97 ± 5.21 vs 6.73 ± 2.20 N/mm; P = .027). Significantly fewer iNOS+ cells but more CD206+ cells, as well as lower MMP-1 and MMP-13 but higher TIMP-1 expression, were detected at the tendon-bone interface and graft midsubstance in the dTDSC sheet group. Conclusion: An optimized decellularization protocol for producing bioactive dTDSC sheets was developed. Wrapping tendon graft with a dTDSC sheet promoted graft healing after ACLR, likely via enhancing bone formation and angiogenesis by BMP-2 and VEGF, modulating macrophage polarization and MMP/TIMP expression, and physically protecting the tendon graft. Clinical Relevance: dTDSC sheets alleviate the quality control and safety concerns of cell transplantation and can be used as a cell-free alternative for the promotion of graft healing in ACLR.

Cryopreserved Adipose-Derived Stem Cell Sheets: An Off-the-Shelf Scaffold for Augmenting Tendon-to-Bone Healing in a Rabbit Model of Chronic Rotator Cuff Tear

“…Third, it is unclear whether the improved repair efficacy of cell sheets is primarily the result of viable cells or the ECM within the sheets. Recently, Yao et al 28 evaluated the repair effect of decellularized tendon–derived stem cell sheets and found that they could promote graft healing after anterior cruciate ligament reconstruction. We could conclude from the current study that the ECM of the cell sheets promoted tendon-to-bone healing.…”

Global research trends and hotspots on tendon-derived stem cell: a bibliometric visualization study

“…Removing cellular immunogenicity from the cell sheet may be a Frontiers in Bioengineering and Biotechnology frontiersin.org better option, indicating that the cell sheet can be applied to largescale applications rather than personalized customization. In further research, Pauline Po Yee Lui et al (Shang et al, 2023) chose to decellularize the TDSCs-formed cell sheet, and found that the decellularized cell sheet also promoted graft healing, tunnel bone formation, and angiogenesis (Yao et al, 2023). Hence, the extracellular matrix in the cell sheet may play a pivotal role in promoting healing.…”

Advances Focusing on the Application of Decellularized Extracellular Matrix in Periodontal Regeneration

“…Moreover, different types of C-dECMs have been verified for their capacity to induce osteogenic differentiation of re-seeded BMSCs, including cell sheets derived from bone marrow mesenchymal stem cells (BMSCs), MC3T3 osteoblasts, and L929 fibroblasts [ 22 ]. In addition, C-dECM showed good effect in ligament regeneration: stem cell sheets-derived dECM provide an ectogenic scaffold-free option, enhancing bone formation and angiogenesis by BMP-2 and VEGF, modulating macrophage polarization and MMP/TIMP expression, and physically promoting ligament reconstruction [ 55 ]. In addition, many techniques have been used to construct C-dECM in vitro; for example, studies showed that the addition of a certain amount of ascorbic acid to the culture medium promoted ECM deposition and the formation of cell membranes [ 56 ].…”

“…The fabrication and in vitro analysis of dECM (solid dECM and soluble dECM) of various tissue origins are prerequisites for applications. The dECM can be generated and improved by a variety of methods, and complex periodontal regeneration modules based on the dECM were recently developed [ 20 , 48 , 55 , 87 ].…”