| Ligaments are fibrous connective tissue bands that connect bones and provide joint stability and protection of the joint surface,while also participating in joint motion control.Tendons are fibrous connective tissue that connects muscles to bones and transmits the force generated by muscles to maintain posture and movement of the body.After ligament and tendon injuries,due to poor blood supply and weak repair capability,it is easy to form scar tissue and incomplete healing,leading to long-term joint instability and functional impairment.Therefore,the treatment of ligament and tendon injuries has always been a difficult point in clinical orthopedics.Although the healing process of tendons is similar to that of ligaments,there are also some differences.Unlike ligaments,it is difficult for tendons to heal on their own after injury because of their poor blood supply and lack of cells and substances required for repair.Therefore,surgical repair of damaged tendons is usually necessary,such as by suture or transplantation to rebuild the damaged tendon.The process of tendon healing can be divided into three stages: the inflammatory response stage,the proliferative repair stage,and the remodeling stage.During the inflammatory response stage,the blood vessels surrounding the damaged tendon will dilate,and blood and cytokines will enter the damaged area,triggering an inflammatory response to promote the clearance of damaged tissue and cell repair.During the proliferative repair stage,stem cells and fibroblasts begin to proliferate in the damaged tendon area,forming new tissue.During the remodeling stage,the newly formed tissue gradually takes shape and adapts to the original function of the tendon,eventually forming a complete tendon.Similar to the healing of ligaments,the healing process of tendons also requires appropriate biomechanical stimulation and nutritional support to achieve optimal healing effects.In recent years,with the development of stem cell therapy,the use of stem cells to treat ligament and tendon injuries has become a highly anticipated method.Among them,mesenchymal stem cells(MSCs)from bone marrow have become one of the most promising types of cells due to their multipotent differentiation potential,easy accessibility and amplification,and immune regulation.In the process of mediating ligament repair by mesenchymal stem cells,cell migration,proliferation,differentiation,and collagen synthesis are the main cellular events.At the same time,mesenchymal stem cells can also secrete various growth factors,cytokines,and extracellular vesicles and other bioactive substances,participating in the regulation of extracellular matrix degradation and synthesis,cell apoptosis,and proliferation,promoting ligament and tendon healing.Therefore,we designed relevant experiments on the healing of anterior cruciate ligament and Achilles tendon injuries,exploring some methods to accelerate healing.ACL injury of the knee joint is a common type of ligament injury in clinical practice.Through a long understanding of the structure of the anterior cruciate ligament,we can use stem cells to assist in the repair of the anterior cruciate ligament.Similarly,for the Achilles tendon,stem cell therapy has also shown good therapeutic effects in clinical studies.In this review,we will discuss the current application of stem cell therapy in different types of tendon tissue repair,as well as its mechanism of action,in order to provide a reference for clinical treatment.Part One The Effect of Different Femoral Tunnel Positions on Postoperative Outcomes in Anterior Cruciate Ligament ReconstructionObjective: With the development of understanding and surgical techniques for anterior cruciate ligament(ACL)injury,different femoral tunnel positions have an impact on surgical outcomes.This study investigated the rehabilitation of patients using the IDEAL femoral tunnel positioning method based on the preservation of the stump and compared it with the low anatomical positioning method,aiming to explore the possible problems and solutions of this method and lay the foundation for further research.Methods: This study retrospectively analyzed 44 patients with ACL injury admitted from January 2019 to March 2020,who were divided into two groups according to the femoral tunnel positioning method: low anatomical positioning group(anatomical group;n=22)and IDEAL positioning group(IDEAL group;n=20),and compared the postoperative outcomes of the two groups.All patients underwent arthroscopic single-bundle reconstruction using autologous four-strand semitendinosus tendon grafts and followed a standardized rehabilitation program.Clinical examination,KT-1000 arthrometer measurement,IKDC subjective score,Lysholm score and Tegner score were performed before surgery and 24 months after surgery.Results: This study found that both groups of patients showed significant improvement in knee joint stability and function at the last follow-up,and there was no significant difference between the two groups in clinical scores(including KT-1000 arthrometer measurement,IKDC subjective score,Lysholm score and Tegner score),KT-1000 arthrometer measurement and ROM.The IDEAL group had slightly larger ROM than the low anatomical group(132.5°±5.3° vs 130.2°±6.6°),and the patients reported less difficulty in rehabilitation exercises.Some patients had bone tunnel enlargement phenomenon,but it did not affect postoperative recovery.No serious postoperative complications or ACL re-rupture were observed.Conclusions: This study compared the IDEAL femoral tunnel technique with preserved stump with the traditional low anatomical femoral positioning technique,both of which achieved good clinical outcomes.At the same time,it also discussed the common phenomenon of bone tunnel enlargement after ACL reconstruction,and intended to explore the use of basic experiments to promote tendon-bone healing to improve tunnel enlargement problems and the feasibility of acute ACL rupture suture repair,providing new ideas and methods for the treatment of ACL injury.Part Two The role of Runt-related transcription factor 1(RUNX1)in bone mesenchymal stem cell-mediated tendon-bone healing after anterior cruciate ligament reconstruction.Objective: Anterior cruciate ligament(ACL)injury can lead to functional disability and disability.ACL reconstruction often fails due to the failure of tendon-bone interface regeneration.This study aims to investigate the role of Runt-related transcription factor 1(RUNX1)in the tendon-bone healing of the reconstructed ACL using bone mesenchymal stem cells(BMSCs).Methods: Sprague Dawley rats were used as experimental animals,and BMSCs were isolated,cultured,and identified from femurs.To construct a RUNX1 upregulation vector,the c DNA sequence of RUNX1 was cloned into the lentiviral p LVX-Ac GFP1-N1.Then,the lentiviral vector and the helper vectors(p SPAX2 and p MD2.G)were co-transfected using Lipofectamine2000 according to the experimental protocol.The expression level of RUNX1 was detected by quantitative real-time polymerase chain reaction(q RT-PCR).The paraffin-embedded tendon-bone grafts were sectioned and deparaffinized in xylene and rehydrated in a gradient of ethanol.Hematoxylin and eosin(H&E)and Masson’s trichrome staining were performed to evaluate histopathological changes.The slices were imaged at 400 x magnification using a BX53 microscope.The bone and cartilage portions of the specimens were scanned using a Micro CT imaging system.Further,bone density(BMD)and bone volume fraction(BV/TV)were calculated.Results: Flow cytometry results showed that BMSCs were positive for the CD44 and CD90 antigens and negative for the CD34 and CD45 antigens.Then,we found that RUNX1-upregulated BMSCs could improve the biomechanical strength of the transplanted tendon after ACL reconstruction.Furthermore,based on histological observations,the upregulation of RUNX1 was associated with better recovery around the bone tunnel,a tighter tendon-bone interface,and more collagen fibers than the LV-NC-infected BMSCs group.Secondly,RUNX1-upregulated bone marrow mesenchymal stem cells promoted bone formation after ACL reconstruction,which was evidenced by the relief of severe loss and erosion of the cartilage and bone in the tibia and femur region,and an increase in the number of osteoblasts identified by upregulating alkaline phosphatase,osteocalcin,and osteopontin at the tendon-bone interface.Conclusions: The upregulation of the RUNX1 gene may contribute to accelerating the healing process between tendon and bone after anterior cruciate ligament(ACL)reconstruction using bone mesenchymal stem cells(BMSCs).RUNX1 is a transcription factor that plays a critical regulatory role in growth and differentiation processes.By increasing the expression level of RUNX1,the proliferation and differentiation of bone and tendon cells can be promoted,leading to faster ACL regeneration and better functional recovery.Part Three The role of leucine-rich repeat-containing 32(LRRC32)in rat tendon-derived stem cell-mediated Achilles tendon repair.Objective: Although many surgical or non-surgical treatments have been developed for the treatment of Achilles tendon injury,their prognosis is often unsatisfactory.Recently,a possible treatment approach using biological methods,such as tendon-derived stem cells(TDSCs)which are pluripotent,has been proposed.The objective of this study was to evaluate whether Leucine-rich repeat-containing 32(Lrrc32),which is rich in Leucine and contains 32 repetitive sequences,affects the tendon differentiation of TDSCs and thereby promotes Achilles tendon healing.Methods: Sprague-Dawley(SD,6-8 weeks old)rats were used as recipients or donors in this study.Six SD rats that did not undergo surgery were used as the source of TDSCs.The remaining 96 rats were divided into 4groups(24 rats per group): untreated control group,Achilles tendon injury and PBS application negative control group(NC group),Achilles tendon injury and TDSCs+LV-n C application group(TDSCs+LV-NC group),and Achilles tendon injury and TDSCs+LV-Lrrc32 application group(TDSCs+LV-Lrrc32group).TDSCs were infected with Lrrc32 overexpression recombinant lentivirus(LV-Lrrc32)and locally injected into the injured site of the rats.The euthanized rats were dissected,and the surrounding Achilles tendon tissue was collected.For histological evaluation,the isolated tendon tissue was dehydrated in an ethanol series,embedded in paraffin,and cut into sections with a thickness of 5 μm.The sections were stained with H&E to examine the general morphology of the healing tendon.Further observations were made through immunofluorescence(IF)staining and Western blot.Results: LV-Lrrc32,a recombinant slow virus infected TDSCs,was locally injected into the rat injury site.Four weeks after the surgery,the surrounding Achilles tendon tissue(about 0.5 cm)was analyzed.Pathological results showed that TDSCs overexpressing Lrrc32 significantly improved the morphological changes of the injured tendon.The expression of collagen-I in the extracellular matrix of the Lrrc32 overexpression group increased,the content of hydroxyproline increased,and the regenerated collagen fibers were more organized.In addition,four weeks after injecting TDSCs overexpressing Lrrc32,the expression of tendon-related genes such as tenomodulin(Tnmd),scleraxis(Scx),and decorin(Dcn)in the tendon healing area was upregulated.These results indicate that Lrrc32 promotes tenogenic differentiation of TDSCs in vivo.In addition,Lrrc32 overexpression also increased the expression of TGF-β1 and p-SMAD2/3,suggesting that the beneficial effects of Lrrc32 on tendon repair may be related to the expression of TGF-β1 and p-SMAD2/3.Conclusions: Our study found that TDSCs overexpressing Lrrc32 are more helpful in tendon healing than TDSCs expressing alone.Lrrc32 is a membrane protein related to cell adhesion and migration,and its expression increases significantly in the early stage of tendon injury.Through experiments,we have demonstrated that TDSCs overexpressing Lrrc32 have significant advantages in promoting tendon healing.These cells can enhance the growth and proliferation of tendon cells by enhancing the deposition of extracellular matrix and increasing cell adhesion ability,and can also inhibit inflammation in tendon fibrous tissue,thereby promoting tendon healing and repair.These findings provide a new strategy to enhance the role of TDSCs in tendon repair by changing their expression profile.TDSCs overexpressing Lrrc32 are more effective in promoting tendon healing. |
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