Hyaline-like Cartilage Research Articles

Collagen Type I and Type II Expression Evaluation on Cartilage Defect Regeneration Treated with Dwikora–Ferdiansyah–Lesmono–Purwati (DFLP) Scaffold Supplemented with Adipose–Derived Stem Cells (ASCs) or Secretome: An In-Vivo Study

Abstract Cell-based therapies such as Scaffold, stem cells, and secretome, are one of the alternatives to enhance the regeneration of hyaline-like cartilage in cases of cartilage defects. This study is an in-vivo experiment using animal models, in which we apply a composite of DFLP (Dwikora-Ferdiansyah-Lesmono-Purwati) Scaffold and Adipose-Derived Stem Cells (ASCs) or Secretome to an injury model on the distal femoral trochlea of New Zealand White Rabbits. The animals were divided into four groups: (1) control (K); (2) Scaffold only (S); (3) Scaffold + ASCs (SA); (4) Scaffold + Secretome (SS). Animals were terminated in the 12th week, and an immunohistochemistry (IHC) evaluation for Collagen type I and II were done. Statistical analysis shows that collagen type I IHC between groups shows no significant difference (p = 0.546). Collagen type II IHC shows significant difference between groups (p = 0,016). The findings in this study showed that Scaffold + ASCs group and Scaffold + Secretome have better collagen type II expression compared to the control group. DFLP Scaffold composite with ASCs or Secretome shows potential for cartilage regeneration therapy by increasing type II collagen expression as in hyaline-like cartilage which may be used for regenerative therapy for cartilage defects. Keywords : DFLP Scaffold; Adipose-Derived Stem Cells (ASCs); Secretome; Collagen Type I; Collagen Type IICorrespondence : ianperbowo@me.com

Osteochondral repair combining therapeutics implant with mesenchymal stem cells spheroids

Functional articular cartilage regeneration remains challenging, and it is essential to restore focal osteochondral defects and prevent secondary osteoarthritis. Combining autologous stem cells with therapeutic medical device, we developed a bi-compartmented implant that could promote both articular cartilage and subchondral bone regeneration. The first compartment based on therapeutic collagen associated with bone morphogenetic protein 2, provides structural support and promotes subchondral bone regeneration. The second compartment contains bone marrow-derived mesenchymal stem cell spheroids to support the regeneration of the articular cartilage. Six-month post-implantation, the regenerated articular cartilage surface was 3 times larger than that of untreated animals, and the regeneration of the osteochondral tissue occurred during the formation of hyaline-like cartilage. Our results demonstrate the positive impact of this combined advanced therapy medicinal product, meeting the needs of promising osteochondral regeneration in critical size articular defects in a large animal model combining not only therapeutic implant but also stem cells.

Decreased Expression of Heat Shock Protein 47 Is Associated with T-2 Toxin and Low Selenium-Induced Matrix Degradation in Cartilages of Kashin-Beck Disease.

Recent evidence suggests a role of type II collagen in Kashin-Beck disease (KBD) degeneration. We aimed to assess the abnormal expression of heat shock protein 47 (HSP47) which is associated with a decrease in type II collagen and an increase in cartilage degradation in KBD. Hand phalange cartilages were collected from KBD and healthy children. Rats were administered with T-2 toxin under the selenium (Se)-deficient diet. ATDC5 cells were seeded on bone matrix gelatin to construct engineered cartilaginous tissue. C28/I2 and ATDC5 cells and engineered tissue were exposed to different concentrations of T-2 toxin with or without Se. Cartilage degeneration was determined through histological evaluation. The distribution and expression of type II collagen and HSP47 were investigated through immunohistochemistry, western blotting, and real-time PCR. KBD cartilages showed increased chondronecrosis and extracellular matrix degradation in deep zone with decreased type II collagen and HSP47 expression. The low-Se + T-2 toxin animal group showed a significantly lower type II collagen expression along with decreased HSP47 expression. Decreased type II collagen and HSP47 in C28/I2 and ATDC5 cells induced by T-2 toxin showed a dose-dependent manner. Hyaline-like cartilage with zonal layers was developed in engineered cartilaginous tissues, with decreased type II collagen and HSP47 expression found in T-2 toxin-treated group. Se-supplementation partially antagonized the inhibitory effects of T-2 toxin in chondrocytes and cartilages. HSP47 plays a role in the degenerative changes of KBD and associated with T-2 toxin-induced decreased type II collagen expression, further promoting matrix degradation.

Macroscopic and histologic evaluation of cartilage regeneration treated using xenogenic biodegradable porous sponge cartilage scaffold composite supplemented with allogenic adipose derived mesenchymal stem cells (ASCs) and secretome: An in vivo experimental study.

Articular cartilage is an avascular, alymphatic, and anisotropic tissue, these characteristics cause significant healing problems to injuries to the cartilage tissue. To overcome this problem, various techniques have been developed and widely used, but the cost-effectiveness and resulting tissue regeneration have never achieved hyaline-like cartilage that has the best biomechanical properties. The idea of this experiment is to use a Biodegradable Porous Sponge Cartilage (BPSC) Scaffold to enhance the regeneration of hyaline-like cartilage combined with microfracture technique and Adipose Derived Stem Cells (ASCs) or secretome on an animal model. A model defect was made on the femoral trochlea of a New Zealand white rabbit. Four groups were made to compare different treatment methods for osteochondral defects. The groups were: (1) Control group; (2) Scaffold Group; (3) Scaffold + ASCs Group; (4) Scaffold + Secretome Group. After 12 weeks, we terminate the animal models, then a macroscopic evaluation using the International Cartilage Research Society (ICRS) scoring system and Oswestry Arthroscopy Score (OAS) was done, followed by sectioning the specimen for microscopic evaluation using the O'Driscoll scoring system. The mean score for all treatment group were better compared to the control group grossly and histologically. The best mean score for macroscopic and microscopic evaluation was the group given Scaffold + ASCs. The application of BPSC scaffold enhances cartilage regeneration in larger osteochondral defects. Furthermore, the addition of ASCs or secretome along with the scaffold implantation further enhances the cartilage regeneration, in which ASCs shows better results.

3D printing of fibre-reinforced cartilaginous templates for the regeneration of osteochondral defects

Successful osteochondral defect repair requires regenerating the subchondral bone whilst simultaneously promoting the development of an overlying layer of articular cartilage that is resistant to vascularization and endochondral ossification. During skeletal development articular cartilage also functions as a surface growth plate, which postnatally is replaced by a more spatially complex bone-cartilage interface. Motivated by this developmental process, the hypothesis of this study is that bi-phasic, fibre-reinforced cartilaginous templates can regenerate both the articular cartilage and subchondral bone within osteochondral defects created in caprine joints. To engineer mechanically competent implants, we first compared a range of 3D printed fibre networks (PCL, PLA and PLGA) for their capacity to mechanically reinforce alginate hydrogels whilst simultaneously supporting mesenchymal stem cell (MSC) chondrogenesis in vitro. These mechanically reinforced, MSC-laden alginate hydrogels were then used to engineer the endochondral bone forming phase of bi-phasic osteochondral constructs, with the overlying chondral phase consisting of cartilage tissue engineered using a co-culture of infrapatellar fat pad derived stem/stromal cells (FPSCs) and chondrocytes. Following chondrogenic priming and subcutaneous implantation in nude mice, these bi-phasic cartilaginous constructs were found to support the development of vascularised endochondral bone overlaid by phenotypically stable cartilage. These fibre-reinforced, bi-phasic cartilaginous templates were then evaluated in clinically relevant, large animal (caprine) model of osteochondral defect repair. Although the quality of repair was variable from animal-to-animal, in general more hyaline-like cartilage repair was observed after 6 months in animals treated with bi-phasic constructs compared to animals treated with commercial control scaffolds. This variability in the quality of repair points to the need for further improvements in the design of 3D bioprinted implants for joint regeneration. Statement of SignificanceSuccessful osteochondral defect repair requires regenerating the subchondral bone whilst simultaneously promoting the development of an overlying layer of articular cartilage. In this study, we hypothesised that bi-phasic, fibre-reinforced cartilaginous templates could be leveraged to regenerate both the articular cartilage and subchondral bone within osteochondral defects. To this end we used 3D printed fibre networks to mechanically reinforce engineered transient cartilage, which also contained an overlying layer of phenotypically stable cartilage engineered using a co-culture of chondrocytes and stem cells. When chondrogenically primed and implanted into caprine osteochondral defects, these fibre-reinforced bi-phasic cartilaginous grafts were shown to spatially direct tissue development during joint repair. Such developmentally inspired tissue engineering strategies, enabled by advances in biofabrication and 3D printing, could form the basis of new classes of regenerative implants in orthopaedic medicine.

Evaluation of autologous adipose-derived mesenchymal stem cell therapy in focal chondral defects of the knee: a pilot case series.

Aim: To evaluate the safety, pain, functional and structural improvements after autologous adipose-derived mesenchymal stem cell (ADMSC) therapy in combination with arthroscopic abrasion arthroplasty in focal chondral defects of the knee. Methods: Eight patients with a focal full thickness chondral defect of the knee underwent arthroscopic abrasion arthroplasty followed by postoperative intra-articular injections of autologous ADMSCs (50×106 ADMSCs at baseline and 6 months). Clinical outcome was assessed using numeric pain rating scale, Knee Injury and Osteoarthritis Outcome Score and the Western Ontario and McMaster Universities Osteoarthritis Index. Structural outcome was determined bymagnetic resonance imaging. Outcome was assessed over 24 months. Results: No serious adverse events occurred. Participants observed clinically significant improvement in pain and function. Magnetic resonance imaging analysis showed cartilage regeneration with T2 mapping values comparable to hyaline cartilage. Conclusion: Arthroscopic abrasion arthroplasty in combination with intra-articular ADMSC therapy results in reproducible pain, functional and structural improvements with regeneration of hyaline-like cartilage. Trial registration number: ACTRN12617000638336.

Reinforcing interpenetrating network hydrogels with 3D printed polymer networks to engineer cartilage mimetic composites

Engineering constructs that mimic the complex structure, composition and biomechanics of the articular cartilage represents a promising route to joint regeneration. Such tissue engineering strategies require the development of biomaterials that mimic the mechanical properties of articular cartilage whilst simultaneously providing an environment supportive of chondrogenesis. Here three-dimensional (3D) bioprinting is used to develop polycaprolactone (PCL) fibre networks to mechanically reinforce interpenetrating network (IPN) hydrogels consisting of alginate and gelatin methacryloyl (GelMA). Inspired by the significant tension-compression nonlinearity of the collagen network in articular cartilage, we printed reinforcing PCL networks with different ratios of tensile to compressive modulus. Synergistic increases in compressive modulus were observed when IPN hydrogels were reinforced with PCL networks that were relatively soft in compression and stiff in tension. The resulting composites possessed equilibrium and dynamic mechanical properties that matched or approached that of native articular cartilage. Finite Element (FE) modelling revealed that the reinforcement of IPN hydrogels with specific PCL networks limited radial expansion and increased the hydrostatic pressure generated within the IPN upon the application of compressive loading. Next, multiple-tool biofabrication techniques were used to 3D bioprint PCL reinforced IPN hydrogels laden with a co-culture of bone marrow-derived stromal cells (BMSCs) and chondrocytes (CCs). The bioprinted biomimetic composites were found to support robust chondrogenesis, with encapsulated cells producing hyaline-like cartilage that stained strongly for sGAG and type II collagen deposition, and negatively for type X collagen and calcium deposition. Taken together, these results demonstrate how 3D bioprinting can be used to engineer constructs that are both pro-chondrogenic and biomimetic of the mechanical properties of articular cartilage.

Enhanced chondrogenesis of human umbilical cord mesenchymal stem cells in a gelatin honeycomb scaffold.

Transplantation of chondrogenic stem cells is a promising strategy for cartilage repair, but requires improvements in cell sourcing, maintenance, and chondrogenic differentiation efficiency. We examined whether gelatin honeycomb scaffolds can enhance the proliferation, viability, and chondrogenic capability of human umbilical cord mesenchymal stem cells (HUCMSCs) compared to standard plate cultures. In addition, the survival and phenotypic stability of HUCMSC-derived chondrocytes in a scaffold were evaluated in mice over 6 weeks post-transplantation. Survival and proliferation rates of HUCMSCs were comparable between scaffold and plate culture. Scaffold culture in a chondrogenic differentiation medium induced more stable expression of the key hyaline cartilage genes COL2A1 and ACAN and the production of the respective proteins Type II collagen and aggrecan as well as glycosaminoglycan. These HUCMSC-differentiated chondrocytes also stably expressed cartilage genes and proteins in the scaffold 6 weeks after transplantation into nonobese diabetic/severe combined immunodeficient mice. These findings indicate that honeycomb-like gelatin scaffolds can promote the survival and chondrogenic differentiation of HUCMSCs to form hyaline-like cartilage.

The fiveyear outcome of a clinical feasibility study using a biphasic construct with minced autologous cartilage to repair osteochondral defects in the knee.

Autologous minced cartilage has been used to repair cartilage defects. We have developed a biphasic cylindrical osteochondral construct for such use in human knees, and report the fiveyear post-operative outcomes. Ten patients with symptomatic osteochondral lesion at femoral condyles were treated by replacing pathological tissue with the osteochondral composites, each consisted a DL-poly-lactide-co-glycolide chondral phase and a DL-poly-lactide-co-glycolide/β-tricalcium phosphate osseous phase. A flat chamber between the two phases served as a reservoir to house double-minced (mechanical pulverization and enzymatical dissociation) autologous cartilage graft. The osteochondral lesion was drill-fashioned a pit of identical dimensions as the construct. Graft-laden construct was press fit to the pit. Post-operative outcome was evaluated using Knee Injury and Osteoarthritis Outcome Score (KOOS) up to fiveyears. Regenerated tissue was sampled with arthroscopic needle biopsy for histology at oneyear, and imaged with magnetic resonance at one, three, and fiveyears to evaluate the neocartilage with MOCART chart. Subchondral bone integration was evaluated with computed tomography at three and fiveyears. Nine patients completed the five-year follow-up. Post-operative mean KOOS, except that of the "symptom" subscale, had been significantly higher than pre-operation from oneyear and maintained to fiveyears. The change of MOCRAT scores of the regenerated cartilage paralleled the change of KOOS. The osseous phase remained mineralized during the five-year period, yet did not fully integrate with the host bone. This novel construct for chondrocyte implantation yielded promising mid-term outcome. It repaired the osteochondral lesion with hyaline-like cartilage durable for at least fiveyears.

Single-Stage Autologous Chondrocyte-Based Treatment for the Repair of Knee Cartilage Lesions: Two-Year Follow-up of a Prospective Single-Arm Multicenter Study

Background: There is an unmet need for a single-stage cartilage repair treatment that is cost-effective and chondrocyte-based. Purpose: To evaluate the safety and preliminary efficacy of autologous freshly isolated primary chondrocytes and bone marrow mononucleated cells (MNCs) seeded into a PolyActive scaffold in patients with symptomatic cartilage lesions of the knee. Study Design: Case series; Level of evidence, 4. Methods: A total of 40 patients with symptomatic knee cartilage lesions were treated with freshly isolated autologous chondrocytes combined with bone marrow MNCs delivered in a biodegradable load-bearing scaffold. The treatment requires only 1 surgical intervention and is potentially a cost-effective alternative to autologous chondrocyte implantation. The primary chondrocytes and bone marrow MNCs were isolated, washed, counted, mixed, and seeded into a load-bearing scaffold in the operating room. Patients were followed up at 3, 6, 12, 18, and 24 months. Primary endpoints were treatment-related adverse events up to 3 months, adverse implant effects between 3 and 24 months, and the implant success rate at 3 months as measured by lesion filling. Results: Successful lesion filling (≥67% on magnetic resonance imaging) was found in 40 patients at 3 months and in 32 of the 32 patients analyzed at 24 months. Significant improvement over baseline was found for visual analog scale for pain from 3 months onward; Knee injury and Osteoarthritis Outcome Score (KOOS)-Pain and KOOS-Activities of Daily Living from 6 months onward; for KOOS-Symptoms and Stiffness, KOOS-Quality of Life and International Knee Documentation Committee from 12 months onward; and for KOOS-Sport and Recreation from 18 months onward. Hyaline-like repair tissue was found in 22 of 31 patients available for biopsy. Arthralgia and joint effusion were the most common adverse events. Scaffold delamination and adhesions led to removal of the implant in 2 patients. Conclusion: The treatment of knee cartilage lesions with autologous primary chondrocytes and bone marrow MNCs, both isolated and seeded into a load-bearing PolyActive scaffold within a single surgical intervention, is safe and clinically effective. Good lesion fill and sustained clinically important and statistically significant improvement in all patient-reported outcome scores were found throughout the 24-month study. Hyaline-like cartilage was observed on biopsy specimen in at least 22 of the 40 patients. Registration: NCT01041885 (ClinicalTrials.gov identifier)

Current solutions for the treatment of chronic articular cartilage defects in the knee.

Chondral and osteochondral defects in the knee are common and may lead to degenerative joint disease if treated inappropriately.Conventional treatments such as microfracture often result in fibrocartilage formation and are associated with inferior results. Additionally, microfracture is generally unsuitable for the treatment of defects larger than 2–4 cm2.The osteochondral autograft transfer system (OATS) has been shown to produce superior clinical outcomes to microfracture but is technically difficult and may be associated with donor-site morbidity. Osteochondral allograft use is limited by graft availability and failure of cartilage incorporation is an issue.Autologous chondrocyte implantation (ACI) has been shown to result in repair with hyaline-like cartilage but involves a two-stage procedure and is relatively expensive.Rehabilitation after ACI takes 12 months, which is inconvenient and not feasible for athletic patients.Newer methods to regenerate cartilage include autologous stem cell transplantation, which may be performed as a single-stage procedure, can have a shorter rehabilitation period and is less expensive than ACI. Longer-term studies of these methods are needed.Cite this article: EFORT Open Rev 2020;5:156-163. DOI: 10.1302/2058-5241.5.190031

Therapeutic effects and adaptive limits of an acellular technique by ultrapurified alginate (UPAL) gel implantation in canine osteochondral defect models

BackgroundThe aim of this study was to clarify the objective therapeutic effects of an acellular technique by ultrapurified alginate (UPAL) gel implantation in canine osteochondral defect models.MethodsTwo osteochondral defects (diameters: 3.0 and 5.0 mm) were created on each patellar groove in both knees of 10 dogs. Defects were divided into four groups (n = 10 each): Group 1, untreated 3.0-mm defect; Group 2, 3.0-mm defect with UPAL gel; Group 3, untreated 5.0-mm defect; and Group 4, 5.0-mm defect with UPAL gel. All surgical procedures were performed by individuals unfamiliar with the technique at an independent institution. Articular surfaces were evaluated grossly and histologically at 27 weeks after operation.ResultsUPAL gel-treated osteochondral defects showed significantly improved gross appearance in Group 4 and histological appearance in Groups 2 and 4. Reparative tissues in the 3.0-mm defect with UPAL gel were replaced by hyaline-like cartilage tissue. The 5.0-mm defects with UPAL gel were mostly covered with fibrocartilaginous tissue, whereas UPAL gel-untreated defects mostly remained uncovered by any tissue.ConclusionsAlthough an acellular technique using UPAL gel implantation significantly enhanced osteochondral repair in canines, reparative tissues of the large defect with alginate gel comprised of fibrocartilaginous tissue. This surgical technique is effective, especially for small cartilage injuries. Further improvements are required before clinical application in cases of severe osteochondral defects in humans.

Articular cartilage repair using autologous collagen-induced chondrogenesis (ACIC): a pragmatic and cost-effective enhancement of a traditional technique.

The autologous collagen-induced chondrogenesis technique is described, and the results of a 6-year follow-up clinical study using this technique are presented. 30 patients with International Cartilage Repair Society (ICRS) Grade III/IVa symptomatic chondral defects of the knee treated with enhanced microdrilling using atelocollagen were prospectively examined in this clinical series. The median age of the patients was 39.0years (range 19-61years). Patients were followed up to 72months. Clinical evaluation was performed using functional knee scores and radiologically. Both quantitative and qualitative assessments were performed. Statistically significant and clinically relevant improvement was observed in 2years and was sustained for the 6years of the study observation. At 6years, the mean Lysholm score was 79.7 (SD 6.8) compared to 52.6 (SD 10.7) pre-operatively (p < 0.05). The symptomatic Knee Injury and Osteoarthritis Outcome Score (KOOS) improved from 68.3 (SD 11.4) to 90.2 (SD 4.3) (p < 0.05). The subjective International Knee Documentation Committee (IKDC) also showed improvement from 39.1 (SD 4.1) to 81.6 (SD 7.8) (p < 0.05). The calculated T2* relaxation times were 26.0 (SD 4.2) seconds and 30.3 (SD 6.2) seconds for the repair tissue and native cartilage, respectively. The average magnetic resonance observation of cartilage repair tissue (MOCART) score was 78.5 (SD 9.6) for all lesions. The enhanced microdrilling using atelocollagen is an enhancement of the traditional microfracture method using an off-the-shelf product. When used to treat moderate to severe chondral lesions, this enhancement produces hyaline-like cartilage with a corresponding improvement in symptoms. IV.

Biologically Regulated Marrow Stimulation by Blocking TGF-β1 With Losartan Oral Administration Results in Hyaline-like Cartilage Repair: A Rabbit Osteochondral Defect Model

Background: Microfracture or bone marrow stimulation (BMS) is often the first choice for clinical treatment of cartilage injuries; however, fibrocartilage, not pure hyaline cartilage, has been reported because of the development of fibrosis in the repair tissue. Transforming growth factor β1 (TGF-β1), which can promote fibrosis, can be inhibited by losartan and potentially be used to reduce fibrocartilage. Hypothesis: Blocking TGF-β1 would improve cartilage healing in a rabbit knee BMS model via decreasing the amount of fibrocartilage and increasing hyaline-like cartilage formation. Study Design: Controlled laboratory study. Methods: An osteochondral defect was made in the patellar groove of 48 New Zealand White rabbits. The rabbits were divided into 3 groups: a defect group (defect only), a BMS group (osteochondral defect + BMS), and a BMS + losartan group (osteochondral defect + BMS + losartan). For the rabbits in the BMS + losartan group, losartan was administrated orally from the day after surgery through the day of euthanasia. Rabbits were sacrificed 6 or 12 weeks postoperatively. Macroscopic appearance, microcomputed tomography, histological assessment, and TGF-β1 signaling pathway were evaluated at 6 and 12 weeks postoperatively. Results: The macroscopic assessment of the repair revealed that the BMS + losartan group was superior to the other groups tested. Microcomputed tomography showed superior healing of the bony defect in the BMS + losartan group in comparison with the other groups. Histologically, fibrosis in the repair tissue of the BMS + losartan group was significantly reduced when compared with the other groups. Results obtained with the modified O’Driscoll International Cartilage Repair Society grading system yielded significantly superior scores in the BMS + losartan group as compared with both the defect group and the BMS group (F value: 15.8, P < .001, P = .012, respectively). TGF-β1 signaling and TGF-β-activated kinase 1 of the BMS + losartan group were significantly suppressed in the synovial tissues. Conclusion: By blocking TGF-β1 with losartan, the repair cartilage tissue after BMS was superior to the other groups and consisted primarily of hyaline cartilage. These results should be easily translated to the clinic because losartan is a Food and Drug Administration–approved drug and it can be combined with the BMS technique for optimal repair of chondral defects. Clinical Relevance: Biologically regulated marrow stimulation by blocking TGF-β1 (oral intake of losartan) provides superior repair via decreasing fibrocartilage formation and resulting in hyaline-like cartilage as compared with outcomes from BMS only.

Guided bone marrow stimulation for articular cartilage repair through a freeze-dried chitosan microparticle approach

A novel approach for stimulating articular cartilage repair was developed and evaluated in skeletally aged Arcott sheep with signs of early osteoarthritis. Freeze-dried (FD) chitosan formulations were optimized to produce ultraporous cylinders that slowly rehydrate and disperse into bioactive chitosan microparticles in coagulating blood plasma. FD-chitosan implants (80% Degree of Deacetylation, 85 kDa) were produced at 3 doses (initial concentrations of 5, 10, 20 mg/mL, pH 2.5). Full-thickness cartilage defects were created bilaterally in medial femoral condyles of 8–9 year-old sheep (N = 12), microdrilled with 11 holes, then in one knee per sheep, one implant cylinder was inserted into each bleeding drill hole. At 1 day (N = 2), 3 months (N = 5) and 9 months (N = 5) post-operative, repair tissues were analyzed macroscopically and by micro-computed tomography, histology, biochemistry, and mechanics. Chitosan microparticles were detected in day 1 subchondral blood clots and mostly cleared at 3 months. At 3 months, microdrill holes were 2-fold larger, filled with angiogenic granulation tissue, callus, and woven bone, with more chondroinduction in treated versus control drill holes (p = 0.021). At 9 months, biomaterial treatment enhanced bone plate repair and stimulated 68% cartilage resurfacing vs 53% for drill-only controls (p = 0.047). Both treated and control cartilage repair tissues had lower glycosaminoglycan content than intact cartilage and were thinner, stiffer, and more permeable. Upon indentation, hyaline-like repair cartilage showed poroelastic behavior. This study showed that FD-chitosan can be locally delivered to incorporate chitosan microparticles into subchondral bone blood clots and exert anabolic therapeutic effects on articular cartilage resurfacing in aged sheep knees.

Platelet-Rich Fibrin Facilitates One-Stage Cartilage Repair by Promoting Chondrocytes Viability, Migration, and Matrix Synthesis.

The main aim of this study is to develop a one-stage method to combine platelet-rich fibrin (PRF) and autologous cartilage autografts for porcine articular cartilage repair. The porcine chondrocytes were treated with different concentrations of PRF-conditioned media and were evaluated for their cell viability and extracellular glycosaminoglycan (GAG) synthesis during six day cultivation. The chemotactic effects of PRF on chondrocytes on undigested cartilage autografts were revealed in explant cultures. For the in vivo part, porcine chondral defects were created at the medial femoral condyles of which were (1) left untreated, (2) implanted with PRF combined with hand-diced cartilage grafts, or (3) implanted with PRF combined with device-diced cartilage grafts. After six months, gross grades, histological, and immunohistochemical analyses were compared. The results showed that PRF promotes the viability and GAG expression of the cultured chondrocytes. Additionally, the PRF-conditioned media induce significant cellular migration and outgrowth of chondrocytes from undigested cartilage grafts. In the in vivo study, gross grading and histological scores showed significantly better outcomes in the treatment groups as compared with controls. Moreover, both treatment groups showed significantly more type II collagen staining and minimal type I collagen staining as compared with controls, indicating more hyaline-like cartilage and less fibrous tissue. In conclusion, PRF enhances the viability, differentiation, and migration of chondrocytes, thus, showing an appealing capacity for cartilage repair. The data altogether provide evidences to confirm the feasibility of a one-stage, culture-free method of combining PRF and cartilage autografts for repairing articular cartilage defects. From translational standpoints, these advantages benefit clinical applications by simplifying and potentiating the efficacy of cartilage autograft transplants.

Human Amniotic Mesenchymal Stem Cell Sheets Encapsulating Cartilage Particles Facilitate Repair of Rabbit Osteochondral Defects

Background: Human amniotic mesenchymal stem cells (hAMSCs) are being widely applied in various fields. Therefore, hAMSCs represent a promising candidate to facilitate cartilage regeneration. Nonetheless, no studies have investigated the application of hAMSC sheets to repair cartilage defects in vivo. Purpose: To evaluate hAMSC sheets encapsulating cartilage particles to promote repair of rabbit osteochondral defects. Study Design: Controlled laboratory study. Methods: hAMSC sheets were constructed with passage 3 hAMSCs. The phenotypic and structural characteristics of hAMSC sheets were evaluated by flow cytometry and scanning electron microscopy, respectively. The potential for chondrogenic differentiation of hAMSC sheets was assessed by cartilage-specific marker staining, immunohistochemistry, and mRNA and protein expression (SOX9, COLII, and ACAN). Osteochondral defects (diameter, 3.5 mm; depth, 3 mm) were created in the left patellar grooves of 20 New Zealand White rabbits (female or male). The defects were treated with hAMSC sheet/cartilage particles (n = 5), cartilage particles (n = 5), hAMSC sheets (n = 5), or fibrin glue (n = 5). Macroscopic and histological evaluations of the regenerated tissue were conducted after 3 months. The survival time and differentiation of transplanted hAMSCs in the defect area were evaluated by immunofluorescence. Results: hAMSC sheets had a multilayered structure, with cells stacked layer by layer. Importantly, hAMSC sheets highly expressed phenotypic markers of mesenchymal stem cells. Cartilage-specific marker staining and immunohistochemistry were positive, and mRNA and protein expression was higher in the chondrogenically induced hAMSC sheet group than in the hAMSC sheet group (P < .05). hAMSC sheet/cartilage particles formed a large amount of hyaline-like cartilage in the defect area. In addition, macroscopic and histological scores were significantly higher than those in the other groups. Integration with surrounding normal cartilage and subchondral bone regeneration in the hAMSC sheet/cartilage particles group were better when compared with the other groups. A large number of human nuclear-specific antigen-positive cells were observed in the defect area of hAMSC sheet/cartilage particles and hAMSC sheet groups. Moreover, some positive cells expressed SOX9. Conclusion: hAMSC sheets encapsulating cartilage particles facilitate osteochondral defect repair. Clinical Relevance: Delivery of cells in the form of a cell sheet in conjunction with cartilage particles provides a novel approach for cell-based cartilage regeneration.

Scaffold channel size influences stem cell differentiation pathway in 3-D printed silica hybrid scaffolds for cartilage regeneration.

We report that 3-D printed scaffold channel size can direct bone marrow derived stem cell differentiation. Treatment of articular cartilage trauma injuries, such as microfracture surgery, have limited success because durability is limited as fibrocartilage forms. A scaffold-assisted approach, combining microfracture with biomaterials has potential if the scaffold can promote articular cartilage production and share load with cartilage. Here, we investigated human bone marrow derived stromal cell (hBMSC) differentiation in vitro in 3-D printed silica/poly(tetrahydrofuran)/poly(ε-caprolactone) hybrid scaffolds with specific channel sizes. Channel widths of ∼230 μm (210 ± 22 μm mean strut size, 42.4 ± 3.9% porosity) provoked hBMSC differentiation down a chondrogenic path, with collagen Type II matrix prevalent, indicative of hyaline cartilage. When pores were larger (∼500 μm, 229 ± 29 μm mean strut size, 63.8 ± 1.6% porosity) collagen Type I was dominant, indicating fibrocartilage. There was less matrix and voids in smaller channels (∼100 μm, 218 ± 28 μm mean strut size, 31.2 ± 2.9% porosity). Our findings suggest that a 200-250 μm pore channel width, in combination with the surface chemistry and stiffness of the scaffold, is optimal for cell-cell interactions to promote chondrogenic differentiation and enable the chondrocytes to maintain their phenotype.

Synthesis and biological evaluation of a bioinspired, tissue-adhesive gellan gum-based hydrogel designed for minimally invasive delivery and retention of chondrogenic cells.

A dopamine-modified, bioinspired gellan gum hydrogel (STM-148B) with improved physicochemical and biological characteristics, suitable for minimally invasive cell delivery and retention in the context of cartilage repair, is herein presented. STM-148B's putative game-changing design characteristics include a highly biocompatible, animal-free and chemically defined composition, reproducibility of manufacture and ease of formulation. STM-148B undergoes rapid ionic crossinking by physiologically relevant mono and divalent cations to form stable 3D hydrogels that possess excellent tissue adhesiveness, such that additional fixation aids are rendered superfluous. STM-148B hydrogels maintain viability of mammalian cells and further promote up-regulation of the expression of healthy chondrogenic extracellular matrix markers upon stimulation. STM-148B is currently undergoing pre-clinical safety and efficacy assessment as a medical device for cell delivery and retention focussing on regeneration of hyaline-like cartilage and may represent a valuable addition to the armamentarium of tissue-engineering therapies for treatment of focal cartilage lesions.

Full-Scale Osteochondral Regeneration by Sole Graft of Tissue-Engineered Hyaline Cartilage without Co-Engraftment of Subchondral Bone Substitute.

In this study, full-scale osteochondral defects are hypothesized, which penetrate the articular cartilage layer and invade into subchondral bones, and can be fixed by sole graft of tissue-engineered hyaline cartilage without co-engraftment of any subchondral bone substitute. It is hypothesized that given a finely regenerated articular cartilage shielding on top, the restoration of subchondral bones can be fulfilled via spontaneous self-remodeling in situ. Hence, the key challenge of osteochondral regeneration lies in restoration of the non-self-regenerative articular cartilage. Here, traumatic osteochondral lesions to be repaired in rabbit knee models are endeavored using novel tissue-engineered hyaline-like cartilage grafts that are produced by 3D cultured porcine chondrocytes in vitro. Comparative trials are conducted in animal models that are implanted with living hyaline cartilage grafts (LhCG) and decellularized LhCG (dLhCG). Sound osteochondral regeneration is gradually revealed from both LhCG and dLhCG-implanted samples 50-100 d after implantation. Quality regeneration in both zones of articular cartilage and subchondral bones are validated by the restored osteochondral composition, structure, phenotype, and mechanical property, which validate the hypothesis of this study.