ABSTRACT: Hamstring tendon autograft is a routine graft for anterior cruciate ligament (ACL) reconstruction. However, ways of improving the healing between the tendon and bone is often overlooked in clinical practice. This issue can be addressed by using a biomimetic scaffold. Herein, a biomimetic nanofiber membrane of polycaprolactone/nanohydroxyapatite/collagen (PCL/nHAp/Col) is fabricated that mimics the composition of native bone tissue for promoting tendon-bone healing. This membrane has good cytocompatibility, allowing for osteoblast cell adhesion and growth and bone formation. As a result, MC3T3 cells reveal a higher mineralization level in PCL/nHAp/Col membrane compared with PCL membrane alone. Further in vivo studies in ACL reconstruction in a rabbit model shows that PCL/nHAp/Col-wrapped tendon may afford superior tissue integration to nonwrapped tendon in the interface between the tendon and host bone as well as improved mechanical strength. This study shows that PCL/nHAp/Col nanofiber membrane wrapping of autologous tendon is effective for improving tendon healing with host bone in ACL reconstruction.
Project description:Purpose:In order to accelerate the tendon-bone healing processes and achieve the efficient osteointegration between the tendon graft and bone tunnel, we aim to design bioactive electrospun nanofiber membranes combined with tendon stem/progenitor cells (TSPCs) to promote osteogenic regeneration of the tendon and bone interface. Methods:In this study, nanofiber membranes of polycaprolactone (PCL), PCL/collagen I (COL-1) hybrid nanofiber membranes, poly(dopamine) (PDA)-coated PCL nanofiber membranes and PDA-coated PCL/COL-1 hybrid nanofiber membranes were successfully fabricated by electrospinning. The biochemical characteristics and nanofibrous morphology of the membranes, as well as the characterization of rat TSPCs, were defined in vitro. After co-culture with different types of electrospun nanofiber membranes in vitro, cell proliferation, viability, adhesion and osteogenic differentiation of TSPCs were evaluated at different time points. Results:Among all the membranes, the performance of the PCL/COL-1 (volume ratio: 2:1 v/v) group was superior in terms of its ability to support the adhesion, proliferation, and osteogenic differentiation of TSPCs. No benefit was found in this study to include PDA coating on cell adhesion, proliferation and osteogenic differentiation of TSPCs. Conclusion:The PCL/COL-1 hybrid electrospun nanofiber membranes are biocompatible, biomimetic, easily fabricated, and are capable of supporting cell adhesion, proliferation, and osteogenic differentiation of TSPCs. These bioactive electrospun nanofiber membranes may act as a suitable functional biomimetic scaffold in tendon-bone tissue engineering applications to enhance tendon-bone healing abilities.
Project description:The incorporation of tendon graft into bone tunnel is one of the most challenging clinical issues in anterior cruciate ligament (ACL) reconstruction. As a biodegradable metal, Mg has appropriate mechanical strength and osteoinductive effects, thus may be a promising alternative to commercialized products used for graft fixation. Therefore, it was hypothesized that Mg based interference screws would promote tendon graft-bone junction healing when compared to Ti screws. Herein, we compared the effects of Mg and Ti screws on tendon graft healing in rabbits with ACL reconstruction via histological, HR-pQCT and mechanical analysis. The histological results indicated that Mg screws significantly improved the graft healing quality via promoting mineralization at the tendon graft enthesis. Besides, Mg screws significantly promoted bone formation in the peri-screw region at the early healing stage. Importantly, Mg screws exhibited excellent corrosion resistance and the degradation of Mg screws did not induce bone tunnel widening. In tensile testing, there were no significant differences in the load to failure, stress, stiffness and absorption energy between Mg and Ti groups due to the failure mode at the midsubstance. Our findings demonstrate that Mg screws can promote tendon graft healing after ACL reconstruction, implying a potential alternative to Ti screws for clinical applications.
Project description:Anterior cruciate ligament (ACL) ruptures are common and unfortunate injuries for many athletes. The standard therapy for ACL rupture is ACL reconstruction with either autograft, harvested from hamstring or patellar tendon, or allograft tendon from a tissue donor. Advances in tissue engineering have produced interventions to augment the healing process and may have applications when it comes to ACL reconstruction. In this Technical Note and accompanying video, we describe a simple technique to implant an amnion matrix graft with a tendon graft during ACL reconstruction. This procedure uses the proposed anti-inflammatory, scaffolding, and stem cell-producing effects of the amniotic membrane to biologically augment the healing process of an ACL reconstruction.
Project description:Despite the popularity of anterior cruciate ligament (ACL) reconstruction procedures, the ideal graft for reconstruction remains a matter of controversy. The ideal graft for ACL reconstruction should have histologic and biomechanical characteristics similar to those of the native ACL; should be quickly and fully incorporated within the bony tunnels; should maintain its viscoelastic properties for a long time; should have minimal donor-site morbidity; should be of sufficient length and diameter; should have minimal adverse effects on the extensor mechanism; should have no risk of rejection or disease transmission; and should be cost-effective and readily available. Synthetic grafts are not widely accepted because of their dangerous complications. The main sources of grafts for ACL reconstruction are allografts and autografts. Each type of graft has its own relative advantages and disadvantages. Allografts are not available in every country, besides being expensive, and there are many concerns regarding disease transmission. Autografts, particularly bone-patellar tendon-bone (BPTB), and hamstring tendon grafts have been the standard for ACL reconstruction. The main advantage of autogenous BPTB grafts is the direct bone-to-bone healing in the tunnel, whereas the main disadvantages of such grafts are related to donor-site morbidity, anterior knee pain, and extensor mechanism dysfunction. The popularity of autogenous hamstring tendon grafts for ACL reconstruction is increasing, but there are still concerns regarding the slow soft tissue-to-bone healing, with delayed healing and incorporation of the graft. We describe a technique for ACL reconstruction with autogenous hamstring-bone graft, aiming to produce a type of graft that combines the main advantages of BPTB and hamstring grafts, with avoidance of the main disadvantages of these 2 most commonly used graft types in ACL reconstruction.
Project description:Silk fibroin (SF) and fiber alignment were introduced into polycaprolactone (PCL)-based electrospun nanofibers as chemical and physical cues for tendon tissue engineering applications. The physicochemical properties of random PCL (RP) nanofibers, random PCL/SF (RPSF) nanofibers and aligned PCL/SF (APSF) nanofibers were characterized for fiber orientation and SF blending effects. An in vitro cell culture with rabbit dermal fibroblasts (RDFBs) on nanofibers indicated that SF promotes cell proliferation to a higher extent than fiber alignment. Cells aligned in the direction of fiber axes could be confirmed through scanning electron microscopy (SEM) observation and cytoskeleton staining. The quantitative real-time polymerase chain reaction (qRT-PCR) experiments indicated up-regulated gene expression of tendon marker proteins (type I collagen (Col I), fibronectin and biglycan) on APSF nanofibers and tendon reconstruction was confirmed from Col III gene expression. Animal experiments with Achilles tendon defect repairs in rabbits were carried out with RPSF and APSF scaffolds. The beneficial effects of fiber alignment were verified from histological and immunohistochemical staining, where cell migration and extracellular matrix protein deposition tend to stretch in a parallel direction along the axial direction of APSF nanofibers with enhanced Col I and tenascin C production. Biomechanical testing indicated the tensile stiffness and maximum load of cell-seeded APSF scaffolds were 60.2 and 81.3% of normal tendon values, respectively, which are significantly higher than cell-seeded RPSF or acellular APSF and RPSF scaffolds. These results suggest that APSF nanofiber scaffolds combined with RDFBs have the potential to repair the gap defects of Achilles tendons in vivo and to effectively restore the function and structure of tendons.
Project description:The exclusive autograft choice for medial collateral ligament (MCL) reconstruction that has been described until today is the semitendinosus tendon. However, this has some potential disadvantages in a knee with combined MCL-anterior cruciate ligament (ACL) injury, including weakening of the hamstring's anterior restraining action in an already ACL-injured knee and nonanatomic distal MCL graft insertion when leaving the semitendinosus insertion intact at the pes anserinus during reconstruction. Moreover, because some surgeons prefer to use the hamstring for autologous ACL reconstruction, the contralateral uninjured knee hamstring needs to be harvested as a graft source for the MCL reconstruction if autografts and not allografts are the surgeons' preference. We describe a technique for performing combined reconstruction of the MCL and ACL using ipsilateral quadriceps tendon-bone and bone-patellar tendon-bone autografts. This technique of MCL reconstruction spares the hamstring tendons and benefits from the advantage provided by bone-to-bone healing on the femur with distal and proximal MCL tibial fixation that closely reproduces the native MCL tibia insertion.
Project description:Runx2 is a powerful osteo-inductive factor and adipose-derived stem cells (ADSCs) are multipotent. However, it is unknown whether Runx2-overexpressing ADSCs (Runx2-ADSCs) could promote anterior cruciate ligament (ACL) reconstruction. We evaluated the effect of Runx2-ADSCs on ACL reconstruction in vitro and in vivo. mRNA expressions of osteocalcin (OCN), bone sialoprotein (BSP) and collagen I (COLI) increased over time in Runx2-ADSCs. Runx2 overexpression inhibited LPL and PPARγ mRNA expressions. Runx2 induced alkaline phosphatase activity markedly. In nude mice injected with Runx2-ADSCs, promoted bone formation was detected by X-rays 8 weeks after injection. The healing of tendon-to-bone in a rabbit model of ACL reconstruction treated with Runx2-ADSCs, fibrin glue only and an RNAi targeting Runx2, was evaluated with CT 3D reconstruction, histological analysis and biomechanical methods. CT showed a greater degree of new bone formation around the bone tunnel in the group treated with Runx2-ADSCs compared with the fibrin glue group and RNAi Runx2 group. Histology showed that treatment with Runx2-ADSCs led to a rapid and significant increase at the tendon-to-bone compared with the control groups. Biomechanical tests demonstrated higher tendon pullout strength in the Runx2-ADSCs group at early time points. The healing of the attachment in ACL reconstruction was enhanced by Runx2-ADSCs.
Project description:Many strategies, including various growth factors and gene transfer, have been used to augment healing after anterior cruciate ligament (ACL) reconstruction. The biological environment regulated by the growth factors during the stage of tendon-bone healing was considered important in controlling the integrating process. The purpose of this study was to evaluate the effects of bone marrow-derived mesenchymal stem cells (BMSCs) genetically modified with bone morphogenetic protein 2 (BMP2) and basic fibroblast growth factor (bFGF) on healing after ACL reconstruction. BMSCs were infected with an adenoviral vector encoding BMP2 (AdBMP2) or bFGF (AdbFGF). Then, the infected BMSCs were surgically implanted into the tendon-bone interface. At 12 weeks postoperatively, the formation of abundant cartilage-like cells, smaller tibial bone tunnel and significantly higher ultimate load and stiffness levels, through histological analysis, micro-computed tomography and biomechanical testing, were observed. In addition, the AdBMP2-plus-AdbFGF group had the smallest bone tunnel and the best mechanical properties among all the groups. The addition of BMP2 or bFGF by gene transfer resulted in better cellularity, new bone formation and higher mechanical property, which contributed to the healing process after ACL reconstruction. Furthermore, the co-application of these two genes was more powerful and efficient than either single gene therapy.
Project description:BACKGROUND:Tendon-bone interface healing and ligamentization of the graft in anterior cruciate ligament (ACL) reconstruction with autografts are important factors affecting treatment outcome. This study aimed to investigate the effectiveness of a cylindrical titanium-web (TW) in tendon-bone interface healing and graft maturation in ACL reconstruction. METHODS:Fourteen mature female CLAWN miniature swine underwent bilateral ACL reconstructions with patellar tendon (PT) autografts. In one limb, the TW/tendon complex was placed into the proximal side of the tibial tunnel. Only the graft was transplanted into the tunnel in the control limb. The proximal side of the graft was sutured into the stump of the native ACL and the distal end was stapled to the tibia. The animals were euthanized at 4 and 15?weeks postoperatively, for histological and biochemical analyses. RESULTS:Microscopic images in TW limbs showed that ingrowth of tendon-like tissue and mineralized bone tissue into the TW connected the bone and the tendon directly. In contrast, fibrous tissue intervened between the bone and tendon in the control limbs. The total amount of collagen cross-links (which defines the strength of collagen fibers) and the maturation of collagen cross-links in TW tendons were significantly higher (p?<?0.05) than those of control limbs. There was no significant difference in the ratio of dihydroxy-lysinonorleucine to hydroxy-lysinonorleucine (an indicator of tissue specific collagen maturation) between TW tendons and that of the native PT. CONCLUSIONS:TW promoted the maturation and formation of collagen cross-links in the grafted tendon while maintaining the cross-links pattern of native tendon collagen, and enabled direct binding of tendon to bone.
Project description:The basic biology of healing between a tendon graft and bone tunnel remains incompletely understood. Distinct variability in the morphological characteristics of the healing tendon-bone attachment site has been reported. We hypothesized that spatial and temporal differences in tendon-to-bone healing exist at different regions of a surgically created bone tunnel. Twenty-four male, Sprague-Dawley rats underwent anterior cruciate ligament (ACL) reconstruction in the left knee using a flexor digitorum longus tendon graft secured using suspensory periosteal fixation. Animals were sacrificed at 4, 7, 11, 14, 21, and 28 days after surgery and prepared for routine histology and immunohistochemical analysis of the healing enthesis at the intra-articular aperture (IAA), mid-tunnel, and extra-articular aperture (EAA). Six animals were used to measure mineral apposition rate (MAR) along the healing bone tunnel by double fluorochrome labeling at 14 and 28 days after surgery. The total area of calcified bone matrix was assessed with von Kossa staining and Goldner-Masson trichrome staining, respectively. The healing tendon-bone interface tissue exhibited a wide chondroid matrix at the IAA, in contrast to a narrow, fibrous matrix at the EAA. There were significantly more osteoclasts at the IAA compared to EAA throughout the study period, except 4 days after surgery (p < 0.05). Collagen continuity between the tendon graft and bone tunnel increased over time, with a more parallel orientation and increased collagen fiber continuity between tendon and bone at the EAA compared to the IAA. MAR was also significantly greater at the EAA at 4 weeks (p < 0.001). Significant differences in healing between the tendon graft and bone exist along the length of bone tunnel secured with suspensory fixation. The etiology of these differences is likely multifactorial in nature, including variable biological and biomechanical environments at different ends of the tunnel. Understanding these differences may ultimately allow surgeons to improve the quality of graft fixation and long-term outcomes after ACL reconstruction.