Ji-Ying Zhang

Circular RNA Related to the Chondrocyte ECM Regulates MMP13 Expression by Functioning as a MiR-136 ‘Sponge’ in Human Cartilage Degradation

Circular RNAs (circRNAs) are involved in the development of various diseases, but there is little knowledge of circRNAs in osteoarthritis (OA). The aim of study was to identify circRNA expression in articular cartilage and to explore the function of chondrocyte extracellular matrix (ECM)-related circRNAs (circRNA-CER) in cartilage. To identify circRNAs that are specifically expressed in cartilage, we compared the expression of circRNAs in OA cartilage with that in normal cartilage. Bioinformatics was employed to predict the interaction of circRNAs and mRNAs in cartilage. Loss-of-function and rescue experiments for circRNA-CER were performed in vitro. A total of 71 circRNAs were differentially expressed in OA and normal cartilage. CircRNA-CER expression increased with interleukin-1 and tumor necrosis factor levels in chondrocytes. Silencing of circRNA-CER using small interfering RNA suppressed MMP13 expression and increased ECM formation. CircRNA-CER could compete for miR-136 with MMP13. Our results demonstrated that circRNA-CER regulated MMP13 expression by functioning as a competing endogenous RNA (ceRNA) and participated in the process of chondrocyte ECM degradation. We propose that circRNA-CER could be used as a potential target in OA therapy.

A composite scaffold of MSC affinity peptide-modified demineralized bone matrix particles and chitosan hydrogel for cartilage regeneration

Articular cartilage injury is still a significant challenge because of the poor intrinsic healing potential of cartilage. Stem cell-based tissue engineering is a promising technique for cartilage repair. As cartilage defects are usually irregular in clinical settings, scaffolds with moldability that can fill any shape of cartilage defects and closely integrate with the host cartilage are desirable. In this study, we constructed a composite scaffold combining mesenchymal stem cells (MSCs) E7 affinity peptide-modified demineralized bone matrix (DBM) particles and chitosan (CS) hydrogel for cartilage engineering. This solid-supported composite scaffold exhibited appropriate porosity, which provided a 3D microenvironment that supports cell adhesion and proliferation. Cell proliferation and DNA content analysis indicated that the DBM-E7/CS scaffold promoted better rat bone marrow-derived MSCs (BMMSCs) survival than the CS or DBM/CS groups. Meanwhile, the DBM-E7/CS scaffold increased matrix production and improved chondrogenic differentiation ability of BMMSCs in vitro. Furthermore, after implantation in vivo for four weeks, compared to those in control groups, the regenerated issue in the DBM-E7/CS group exhibited translucent and superior cartilage-like structures, as indicated by gross observation, histological examination, and assessment of matrix staining. Overall, the functional composite scaffold of DBM-E7/CS is a promising option for repairing irregularly shaped cartilage defects.

Role of scaffold mean pore size in meniscus regeneration

UNLABELLED Recently, meniscus tissue engineering offers a promising management for meniscus regeneration. Although rarely reported, the microarchitectures of scaffolds can deeply influence the behaviors of endogenous or exogenous stem/progenitor cells and subsequent tissue formation in meniscus tissue engineering. Herein, a series of three-dimensional (3D) poly(ε-caprolactone) (PCL) scaffolds with three distinct mean pore sizes (i.e., 215, 320, and 515μm) were fabricated via fused deposition modeling. The scaffold with the mean pore size of 215μm significantly improved both the proliferation and extracellular matrix (ECM) production/deposition of mesenchymal stem cells compared to all other groups in vitro. Moreover, scaffolds with mean pore size of 215μm exhibited the greatest tensile and compressive moduli in all the acellular and cellular studies. In addition, the relatively better results of fibrocartilaginous tissue formation and chondroprotection were observed in the 215μm scaffold group after substituting the rabbit medial meniscectomy for 12weeks. Overall, the mean pore size of 3D-printed PCL scaffold could affect cell behavior, ECM production, biomechanics, and repair effect significantly. The PCL scaffold with mean pore size of 215μm presented superior results both in vitro and in vivo, which could be an alternative for meniscus tissue engineering. STATEMENT OF SIGNIFICANCE Meniscus tissue engineering provides a promising strategy for meniscus regeneration. In this regard, the microarchitectures (e.g., mean pore size) of scaffolds remarkably impact the behaviors of cells and subsequent tissue formation, which has been rarely reported. Herein, three three-dimensional poly(ε-caprolactone) scaffolds with different mean pore sizes (i.e., 215, 320, and 515μm) were fabricated via fused deposition modeling. The results suggested that the mean pore size significantly affected the behaviors of endogenous or exogenous stem/progenitor cells and subsequent tissue formation. This study furthers our understanding of the cell-scaffold interaction in meniscus tissue engineering, which provides unique insight into the design of meniscus scaffolds for future clinical application.

3D-Printed Poly(ε-caprolactone) Scaffold Augmented With Mesenchymal Stem Cells for Total Meniscal Substitution: A 12- and 24-Week Animal Study in a Rabbit Model

Background: Total meniscectomy leads to knee osteoarthritis in the long term. The poly(ε-caprolactone) (PCL) scaffold is a promising material for meniscal tissue regeneration, but cell-free scaffolds result in relatively poor tissue regeneration and lead to joint degeneration. Hypothesis: A novel, 3-dimensional (3D)–printed PCL scaffold augmented with mesenchymal stem cells (MSCs) would offer benefits in meniscal regeneration and cartilage protection. Study Design: Controlled laboratory study. Methods: PCL meniscal scaffolds were 3D printed and seeded with bone marrow–derived MSCs. Seventy-two New Zealand White rabbits were included and were divided into 4 groups: cell-seeded scaffold, cell-free scaffold, sham operation, and total meniscectomy alone. The regeneration of the implanted tissue and the degeneration of articular cartilage were assessed by gross and microscopic (histological and scanning electron microscope) analysis at 12 and 24 weeks postoperatively. The mechanical properties of implants were also evaluated (tensile and compressive testing). Results: Compared with the cell-free group, the cell-seeded scaffold showed notably better gross appearance, with a shiny white color and a smooth surface. Fibrochondrocytes with extracellular collagen type I, II, and III and proteoglycans were found in both seeded and cell-free scaffold implants at 12 and 24 weeks, while the results were significantly better for the cell-seeded group at week 24. Furthermore, the cell-seeded group presented notably lower cartilage degeneration in both femur and tibia compared with the cell-free or meniscectomy group. Both the tensile and compressive properties of the implants in the cell-seeded group were significantly increased compared with those of the cell-free group. Conclusion: Seeding MSCs in the PCL scaffold increased its fibrocartilaginous tissue regeneration and mechanical strength, providing a functional replacement to protect articular cartilage from damage after total meniscectomy. Clinical Relevance: The study suggests the potential of the novel 3D PCL scaffold augmented with MSCs as an alternative meniscal substitution, although this approach requires further improvement before being used in clinical practice.

Meniscus transplantation using treated xenogeneic meniscal tissue: viability and chondroprotection study in rabbits.

PURPOSE This was a preliminary study performed in vivo to evaluate the viability and the chondroprotective effects of irradiated deep-frozen xenogeneic meniscal tissue as a novel substitute for meniscus transplantation. METHODS Medial meniscectomies were performed on the right knees of 48 New Zealand white rabbits. The inner one-third of pig meniscus was harvested and then irradiated and deeply frozen. The treated xenogeneic meniscal tissues were then transplanted to 24 right knees (Xeno group), whereas 24 other knees received meniscus allograft transplantations (Allo group). The left knees of the Xeno group and Allo group received meniscectomies (Meni group) and sham operations (Sham group), respectively. The rabbits were killed at weeks 6, 12, and 24 postoperatively. The newly formed structure of the implanted tissue and cartilage of the medial compartment of each group was assessed by gross and semiquantitative histologic analysis. RESULTS After 24 weeks, the implanted xenogeneic meniscal tissue completely healed to the synovium and formed meniscus-like tissue. The chondrocyte-like cell infiltrated into the tissue with extracellular matrix including type II collagen and proteoglycans. The Xeno group showed significantly less cartilage degeneration than that of the Meni group in the medial tibial plateau at week 24 (P < .05). No significant difference was found between the Xeno group and the Allo group except for the meniscus-covered regions at week 24. From week 12 to week 24, almost no advanced cartilage degeneration was found in weight-bearing regions of the medial tibial plateau of the Xeno group. CONCLUSIONS The treated xenogeneic meniscal tissue healed to the synovium with tissue regeneration and slowed down articular cartilage degeneration in the short-term. The chondroprotection of xenograft transplantation was similar to that of allograft transplantation. CLINICAL RELEVANCE The treated xenogeneic meniscal tissue showed the potential for viability and slowed cartilage degeneration, but more studies are required for application in humans in the future.

Double-Bundle Anterior Cruciate Ligament Reconstruction Using Bone–Patellar Tendon–Bone Allograft Technique and 2- to 5-Year Follow-up

Background: Nonanatomic transtibial single-bundle anterior cruciate ligament reconstruction (SB-ACLR) with a bone–patellar tendon–bone (BPTB) allograft has been used for a long time and has shown the same satisfactory clinical results as an autograft; however, it has not been reported if a double-bundle ACLR (DB-ACLR) could be performed with a BPTB allograft and achieve even better results. Hypothesis: The DB-ACLR with a BPTB allograft is technically feasible and will be superior to the SB technique in restoring better anterior and rotating stability. Study Design: Cohort study; Level of evidence, 2. Methods: The study was performed with 56 patients, and 52 (25 in the DB group and 27 in the SB group) of them were followed up at 2 to 5 years. With an irradiated deep-frozen BPTB allograft, a standard single-incision arthroscopic technique was used, and the graft was fixed with bioabsorbable interference screws on both the femoral and tibial sides. Outcome assessment at final follow-up included International Knee Documentation Committee (IKDC), Tegner, and Lysholm scores; side-to-side difference by conventional KT-2000 arthrometer; total anteroposterior (AP) laxity by the back-pushing KT-2000 arthrometer; pivot shift (0, +, ++); range of motion (ROM); and isokinetic muscle strength evaluation. Results: Mean follow-up was 47.3 ± 11.5 and 58.2 ± 6.6 months for the DB group and SB group, respectively. A statistically significant difference in favor of the DB group was found with the total AP laxity at 30° (P < .05). The overall incidence of pivot shift in the DB group (4% ++) was significantly lower than that in the SB group (26%: 19% + and 7% ++; P = .029). No significant differences were found between the 2 groups in terms of IKDC score, Lysholm score, Tegner score, conventional KT-2000 arthrometer anterior laxity, ROM, and muscle strength. Conclusion: A DB-ACLR with a BPTB allograft is feasible and achieved more satisfactory results than the transtibial SB technique in terms of total AP stability and rotational stability in spite of no significant differences among other clinical parameters.

Runx2-Modified Adipose-Derived Stem Cells Promote Tendon Graft Integration in Anterior Cruciate Ligament Reconstruction

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.

Potential of Centrifugal Seeding Method in Improving Cells Distribution and Proliferation on Demineralized Cancellous Bone Scaffolds for Tissue-Engineered Meniscus

Tissue-engineered meniscus offers a possible solution to the regeneration and replacement problem of meniscectomy. However, the nonuniform distribution and declined proliferation of seeded cells on scaffolds hinder the application of tissue-engineered meniscus as a new generation of meniscus graft. This study systematically investigated the performances of different seeding techniques by using the demineralized cancellous bone (DCB) as the scaffold. Static seeding, injection seeding, centrifugal seeding, and vacuum seeding methods were used to seed the meniscal fibrochondrocytes (MFCs) and mesenchymal stem cells (MSCs) to scaffolds. Cell-binding efficiency, survival rate, distribution ability, and long-term proliferation effects on scaffolds were quantitatively evaluated. Cell adhesion was compared via cell-binding kinetics. Cell viability and morphology were assessed by using fluorescence staining. Combined with the reconstructed three-dimensional image, the distribution of seeded cells was investigated. The Cell Counting Kit-8 assay and DNA assay were employed to assess cell proliferation. Cell-binding kinetics and cell survival of the MFCs were improved via centrifugal seeding compared to injection or vacuum seeding methods. Seeded MFCs by centrifugation showed a more homogeneous distribution throughout the scaffold than cells seeded by other methods. Moreover, the penetration depth in the scaffold of seeded MFCs by centrifugation was 300-500 μm, much higher than the value of 100-300 μm by the surface static and injection seeding. The long-term proliferation of the MFCs in the centrifugal group was also significantly higher than that in the other groups. The results of the MSCs were similar to those of the MFCs. The centrifugal seeding method could significantly improve MFCs or MSCs distribution and proliferation on the DCB scaffolds, thus providing a simple, cost-effective, and effective cell-seeding protocol for tissue-engineered meniscus.

Thermogel-Coated Poly(ε-Caprolactone) Composite Scaffold for Enhanced Cartilage Tissue Engineering

A three-dimensional (3D) composite scaffold was prepared for enhanced cartilage tissue engineering, which was composed of a poly(ε-caprolactone) (PCL) backbone network and a poly(lactide-co-glycolide)-block-poly(ethylene glycol)-block-poly(lactide-co-glycolide) (PLGA–PEG–PLGA) thermogel surface. The composite scaffold not only possessed adequate mechanical strength similar to native osteochondral tissue as a benefit of the PCL backbone, but also maintained cell-friendly microenvironment of the hydrogel. The PCL network with homogeneously-controlled pore size and total pore interconnectivity was fabricated by fused deposition modeling (FDM), and was impregnated into the PLGA–PEG–PLGA solution at low temperature (e.g., 4 °C). The PCL/Gel composite scaffold was obtained after gelation induced by incubation at body temperature (i.e., 37 °C). The composite scaffold showed a greater number of cell retention and proliferation in comparison to the PCL platform. In addition, the composite scaffold promoted the encapsulated mesenchymal stromal cells (MSCs) to differentiate chondrogenically with a greater amount of cartilage-specific matrix production compared to the PCL scaffold or thermogel. Therefore, the 3D PCL/Gel composite scaffold may exhibit great potential for in vivo cartilage regeneration.

The Position of the Posterolateral Bundle Femoral Tunnel During Arthroscopic Double-Bundle Anterior Cruciate Ligament Reconstruction: A Cadaveric Study

PURPOSE The purpose was to find a simple guideline to help establish accurate positioning of the posterolateral bundle (PLB) femoral bone tunnel during double-bundle anterior cruciate ligament reconstruction by measuring the distance between the center of the PLB femoral footprint to the shallow and the deep articular cartilage borders of the lateral wall of the intercondylar notch. METHODS The femoral insertions of the anteromedial bundle and PLB of the anterior cruciate ligament were dissected in 22 male cadaveric knees, aged 25 to 45 years. By use of the intercondylar notch as the landmark, the distances between the center of the PLB femoral footprint and the shallow and the deep articular cartilage borders of the lateral wall of the intercondylar notch were measured with the knees flexed at 90°. The measured data (mean ± standard deviation) were evaluated and compared. RESULTS The center of the PLB was positioned 8.60 ± 1.52 mm and 8.65 ± 1.54 mm from the shallow and the deep cartilage borders of the lateral wall of the intercondylar notch, respectively (P = .95). The distance between the center of the PLB footprint to the low cartilage border of the lateral intercondylar wall was 5.05 ± 0.76 mm. CONCLUSIONS The findings suggest that the position of the center of the PLB femoral footprint is at the middle of the line joining the shallow and the deep borders of the femoral cartilage. CLINICAL RELEVANCE Surgeons can use our results as a guideline and use the PLB footprint remnant as a reference at the same time to locate the femoral PLB tunnel in a simple, easy, and repeatable way.