Purpose: S100 calcium binding protein B (S100B) is expressed in human articular cartilage and has been proposed to reflect the chondrogenic potential and differentiation status of chondrocytes. On the other hand, S100B is higher expressed in osteoarthritic (OA) cartilage compared to normal cartilage and has been suggested to act as pro-inflammatory factor in OA. To gain more insight into the roles of S100B, this study investigated the expression of S100B in healthy and OA native cartilage tissue and chondrocytes in culture, either or not stimulated by growth factors or cytokines. Methods: Chondrocytes isolated from human healthy cartilage biopsies (4 donors) from non-weight bearing areas of the knee were expanded in monolayer culture. At passages 2, 3, 4, 5 and 10 spheroids were formed by seeding 200.000 chondrocytes/well of agarose-coated 96-wells plates. S100B gene expression was measured at the end of monolayer culture and after 2, 3 and 6 weeks in spheroid culture. In addition, sections of healthy articular cartilage and OA cartilage with Kellgren-Lawrence score (KL) II and III were stained for S100B protein expression by immunohistochemistry. This was also performed on sections of spheroids of passage 2 chondrocytes from healthy and KL II cartilage after 2 and 8 weeks of culture and after 8 weeks of culture in medium supplemented with bone morphogenetic protein 2 (BMP2), transforming growth factor beta (TGFβ) and interleukin 1 beta (IL-1β). Unless stated otherwise, the cell cultures were performed in cell culture medium supplemented with only 10 % pooled allogeneic serum at 37°C and 5% CO2. Results: Gene expression of S100B was higher in passage 2 chondrocytes compared to chondrocytes in passages 3, 4, 5 and 10, and expression increased over time in spheroid culture for passage 2 and 3 chondrocytes (Fig. 1). Positive staining for S100B was observed in the pericellular matrix of healthy cartilage and the staining was more intense in the pericellular matrix of KL II cartilage and decreased again in KL III cartilage (Fig. 2). S100B was present in spheroids of chondrocytes from healthy and KL II cartilage. However, the intensity of the staining decreased over time for spheroids of healthy chondrocytes, but not for spheroids of OA chondrocytes. In addition, the spheroids of healthy chondrocytes stained intense for aggrecan, while this was weak for spheroids of KL II chondrocytes (Fig. 3). The addition of the growth factors BMP2 and TGFβ to the culture medium increased S100B, while addition of IL-1 β decreased S100B (Fig. 4). Conclusions: The higher S100B gene expression by chondrocytes in monolayer culture at low passages and the increase upon culture in 3D spheroids support the association between S100B expression and the differentiation status of chondrocytes. The clear increase of S100B protein in mild (KL II) OA cartilage that was decreased in moderate (KL III) OA cartilage could mean that S100B is only increased the early phases of OA. Moreover, in the spheroids of the mild OA chondrocytes, high S100B levels were detected which were stable over time, while these levels decreased over time in spheroids from healthy chondrocytes. The spheroids of healthy chondrocytes already showed good aggrecan deposition after 2 weeks, whereas spheroids of mild OA chondrocytes showed very limited aggrecan deposition after 2 weeks, which was increased at 8 weeks. Therefore, it is likely that a high expression of chondrogenic genes was present for a longer time in the OA chondrocytes. This combined with higher S100B levels when the anabolic growth factors BMP2 and TGFβ were added to the spheroid cultures, suggests that there could be an association between S100B protein and an active cartilage tissue production, but not necessarily the differentiation status of the chondrocytes once normal cartilage tissue remodelling is reached. Addition of IL-1β decreased S100B levels, questioning its role as a pro-catabolic and pro-inflammatory factor.View Large Image Figure ViewerDownload Hi-res image Download (PPT)View Large Image Figure ViewerDownload Hi-res image Download (PPT)View Large Image Figure ViewerDownload Hi-res image Download (PPT)

Objective:Spherox (CO.DON AG) is an autologous chondrocyte implantation (ACI) product, consisting of spheroids of human autologous matrix-associated chondrocytes. The tendency of primary chondrocytes to dedifferentiate during cultivation and the high biologic variability caused by the autologous nature of the starting material makes it challenging to design a manufacturing process that performs consistently and delivers products that meet their intended function and the high quality criteria for cell-based ATMPs. The current study was submitted during the European authorization procedure, and addresses the requirement to justify the operational ranges of the manufacturing process using clinical data.Methods:In order to define the operational ranges, statistical correlation analyses were conducted between process parameters and clinical improvement data of 120 patients from Phase II and III treated with ACI (KOOS score, 1 year follow-up).Results:This approach identified cell culture time as a critical process parameter that negatively correlates with the product's efficacy. Subsequent analyses of the Phase III patients that were treated with chondrocyte spheroids that have been manufactured with shorter monolayer and spheroid cultivation times showed a higher average clinical improvement as well as a higher responder rate compared to the total group. In addition, retrospective analyses demonstrated superiority for the treatment with short-cultivated chondrocyte spheroids over micro-fracture treatment.Conclusion:These findings underscore the need to use clinical data to optimize the manufacturing process for autologous cell-based therapies. We expect that restricting the cultivation times during manufacturing minimizes the production of suboptimal batches, thus ensuring an efficacious product.

Purpose: The use of a fully autologous procedure for the manufacturing of cell-based therapies has a clear advantage for human clinical use. Moreover, also for in vitro research purposes it has advantages to culture cells without the addition of growth factors. Hyaline cartilage formation was observed after the fusion of precultured spheroids (self-aggregated pellets) from culture expanded chondrocytes, using culture medium with only autologous serum. This suggests that during the fusion of spheroids, a better chondrogenic environment is created, possibly by a change in growth factor expression by the chondrocytes themselves. Therefore, the aim of this study was to investigate the deposition of hyaline cartilage components by single and fused spheroids of passaged chondrocytes and to assess the expression of several chondrogenic growth factors by chondrocytes during monolayer expansion and subsequently during culture of single and fused spheroids. Methods: Chondrocytes isolated from human healthy cartilage biopsies (4 donors) from non-weight bearing areas of the knee were expanded in monolayer culture. At passage 2 spheroids were formed by seeding 200.000 chondrocytes/well of agarose-coated 96-wells plates. After 3.5 weeks of culturing the spheroids, 10 spheroids were transferred into one well allowing fusion. All cell cultures were performed in cell culture medium supplemented with 10 % autologous serum at 37°C and 5% CO2. No growth factors, cytokines, antibiotics or other supplements were added. Gene expression of the growth factors bone morphogenetic protein (BMP)2, BMP4, fibroblast growth factor 2 (FGF2), insulin like growth factor 1 (IGF1), transforming growth factor beta (TGFB) and cytokines interleukin 1 beta (IL1B) and tumor necrosis factor alpha (TNFA) were measured in passage 2 chondrocytes during monolayer culture and after 3 weeks in single spheroid cultures. Sections of single spheroids were stained for proteoglycan deposition after 6 weeks of culture. Sections of fused spheroids were stained after 10 weeks of culture by histology for proteoglycans and by immunohistochemistry for type II and I collagen, alkaline phosphatase (ALP), BMP2/4, FGF2, IGF1 and TGFβ. Results: FGF2 was detected in all chondrocyte monolayer cultures, but not in 3D spheroids. Similar results were observed for BMP4 in 3 out of 4 chondrocyte donors. A very weak BMP2 expression was found in spheroid culture of 1 chondrocyte donor. TGFB was expressed by all donors in both monolayer and spheroid culture. No expression of IGF1, IL1B and TNFA could be detected (Fig. 1). Proteoglycans were mainly present in the pericellular zones of single spheroids and more homogenously distributed in the extracellular matrix of fused spheroids. The fused spheroids showed intense type II and a weak type I collagen staining. No ALP or TGFβ could be detected. Some cells were weakly positive for FGF2, whereas most cells were positive for BMP2/4 and strongly positive for IGF1 (Fig. 2). Conclusions: The fusion of spheroids from passaged chondrocytes is an appropriate method to redifferentiate chondrocytes and stimulate hyaline cartilage deposition. The self-aggregation and culture of single spheroids already leads to some hyaline cartilage formation, but this is more pronounced after the fusion of spheroids. In these fused spheroids, chondrocytes express high levels of IGF1 and BMP2/4, which might create a good chondrogenic environment. This differs from the growth factor expression during expansion and single spheroid culture phases where the chondrocytes expressed mainly FGF2 and TGFB, and BMP4 and TGFB, respectively. Therefore, we propose that the clinical usage of the spheroids, which likely fuse upon implantation, limits subsequent formation of fibrocartilage. This is in line with the histological analyses of biopsies from patients treated with Spherox. Moreover, the cell culturing method is fully autologous and omits the use of allogeneic and xenogenic medium components, scaffolds and recombinant growth factors. This might have advantages to study in vitro cartilage formation. Current investigations often use addition of superphysiological concentrations of recombinant human TGFβ, but this is hard to translate towards in vivo use and care has to be taken in combinational treatments, as the effects of TGFβ are very strong and can overshadow effects of other treatments.View Large Image Figure ViewerDownload Hi-res image Download (PPT)

The objective of the investigations was the proof of the use of the neutral salt initiation as a construction material in the protecting silicate coating of concrete components, e.g. factory finished parts or reinforced concrete construction parts, by means of waterglass fused silica suspensions

Purpose: To evaluate the effect of 10% human serum (HS), 5% platelet-rich plasma (PRP), and 5% autologous conditioned plasma (ACP) on migration, proliferation, and extracellular matrix (ECM) synthesis of human meniscus cells. Methods: Cell migration and proliferation on stimulation with HS, PRP, and ACP were assessed by chemotaxis assays and measurement of genomic DNA content. Meniscus cells were cultivated in pellets stimulated with 10% HS, 5% PRP, or 5% ACP. Meniscal ECM formation was evaluated by histochemical staining of collagen type I, type II, and proteoglycans and by analysis of fibrochondrocyte marker gene expression. Results: Human meniscus cells were significantly attracted by all 3 blood-derived products (10% HS and 5% ACP: P = .0001, 5% PRP: P = .0002). Cell proliferation at day 9 was significantly increased on stimulation with 10% HS (P = .0001) and 5% PRP (P = .0002) compared with 5% ACP and controls. Meniscus cell pellet cultures showed the formation of a well-structured meniscal ECM with deposition of collagen type I, type II, and proteoglycans on stimulation with 10% HS, whereas 5% PRP or 5% ACP resulted in the formation of an inhomogeneous and more fibrous ECM. Stimulation with 10% HS and 5% ACP showed a significant induction of fibrochondrocyte marker genes such as aggrecan (HS: P = .0002, ACP: P = .0147), cartilage oligomeric matrix protein (HS: P = .0002, ACP: P = .0005), and biglycan (HS: P = .0002, ACP: P = .0003), whereas PRP showed no inducing effect. Conclusions: Among all tested blood-derived products, only stimulation with HS showed the formation of a meniscal ECM as well as positive cell proliferating and migrating effects in vitro. Regarding a potential biological repair of nonvascular meniscus lesions, our results may point toward the use of HS as a beneficial augment in regenerative meniscus repair approaches. Clinical Relevance: Our findings may suggest that HS might be a beneficial augment for meniscus repair.

Purpose: To evaluate the chondrogenic potential of platelet concentrates on human subchondral mesenchymal progenitor cells (MPCs) as assessed by histomorphometric analysis of proteoglycans and type II collagen. Furthermore, the migratory and proliferative effect of platelet concentrates were assessed. Methods: Platelet-rich plasma (PRP) was prepared using preparation kits (Autologous Conditioned Plasma [ACP] Kit [Arthrex, Naples, FL]; Regen ACR-C Kit [Regen Lab, Le Mont-Sur-Lausanne, Switzerland]; and Dr. PRP Kit [Rmedica, Seoul, Republic of Korea]) by apheresis (PRP-A) and by centrifugation (PRP-C). In contrast to clinical application, freeze-and-thaw cycles were subsequently performed to activate platelets and to prevent medium coagulation by residual fibrinogen in vitro. MPCs were harvested from the cortico-spongious bone of femoral heads. Chondrogenic differentiation of MPCs was induced in high-density pellet cultures and evaluated by histochemical staining of typical cartilage matrix components. Migration of MPCs was assessed using a chemotaxis assay, and proliferation activity was measured by DNA content. Results: MPCs cultured in the presence of 5% ACP, Regen, or Dr. PRP formed fibrous tissue, whereas MPCs stimulated with 5% PRP-A or PRP-C developed compact and dense cartilaginous tissue rich in type II collagen and proteoglycans. All platelet concentrates significantly (ACP, P = .00041; Regen, P = .00029; Dr. PRP, P = .00051; PRP-A, P < .0001; and PRP-C, P < .0001) stimulated migration of MPCs. All platelet concentrates but one (Dr. PRP, P = .63) showed a proliferative effect on MPCs, as shown by significant increases (ACP, P = .027; Regen, P = .0029; PRP-A, P = .00021; and PRP-C, P = .00069) in DNA content. Conclusions: Platelet concentrates obtained by different preparation methods exhibit different potentials to stimulate chondrogenic differentiation, migration, and proliferation of MPCs. Platelet concentrates obtained by commercially available preparation kits failed to induce chondrogenic differentiation of MPCs, whereas highly standardized PRP preparations did induce such differentiation. These findings suggest differing outcomes with PRP treatment in stem cellebased cartilage repair.

To analyze magnetic resonance imaging (MRI) at 3T and the clinical outcome in a short-term pilot study after treatment of retropatellar cartilage defects with microfracturing and subsequent covering with the cell-free chondrotissue® polyglycolic acid–hyaluronan implant.

Cell expansion in vitro is a prequisite to obtain a sufficient quantity of cells for cell-based cartilage repair of articular cartilage lesions. During this process verification of redifferentiation potential of highly expanded chondrocytes is required. Furthermore, cellular impurities of chondrocyte cultures have to be excluded. For this purpose, redifferentiation of expanded human chondrocytes in passage 3 or 5 was initiated in bioresorbable polyglycolic acid-fibrin (PGA-fibrin) scaffolds and selected potential markers were analysed during the process of cell expansion and redifferentiation. Chondrocyte expansion was accompanied by a decrease of collagen type II and COMP and an increase of collagen type I expression indicating cell dedifferentiation. Redifferentiation of chondrocytes in PGA-fibrin scaffolds was accompanied by an increase of collagen II/I ratio. Flow cytometric analyses revealed that in contrast to CD44 and CD49e, CD63 and CD166 showed significant changes in the number of positive cells during redifferentiation. CD14 and CD45 are not expressed by chondrocytes and are therefore possible candidates to detect specifically monocytes or haematopoetic cells in chondrocyte cultures. Characterization of surface antigen expression revealed two promising candidates (CD63 and CD166) to describe the process of redifferentiation, while CD14 and CD45 are suitable markers to exclude impurities by monocytes or haematopoetic cells.

Annulus fibrosus repair techniques for the intervertebral disc (IVD) address the unsolved problem of reherniation after IVD herniation and might facilitate the development of nucleus pulposus replacement techniques for IVD diseases. This study investigates the suitability of a bio-integrative annulus implant.Standardized box defects were applied to the annulus L3/4 and L4/5 of 16 sheep, followed by randomized insertion of the textile polyglycolic acid/polyvinylidene fluoride annulus implant in one of the defects. Explantation was conducted after 2, 6 and 12 weeks, followed by provocative pressure testing and histological analysis. At 2 weeks' follow-up, all specimens of the control defect group demonstrated uncontained herniated nucleus pulposus tissue in the annulus defects. For the treated specimens, the annulus implant consistently provided an effective barrier for herniating nucleus pulposus tissue, with no implant dislocation at all time-points. After 2 weeks, a homogeneous cell infiltration of the annulus implant was observed, leading to a progressive directional matrix build-up. Repair tissue thickness was significantly stronger with the annulus implant at all follow-ups (p < 0.01). No pronounced foreign body reaction and no difference in the amount of supra-annular scar tissue over the defect sites were observed. The implantation procedure inflicted annulus damage adjacent to the defect. At later time-points, however, no difference in comparison with the control defect group was evident. The investigated biointegrative annulus implant showed promising results with regard to biointegration, enhancement of repair tissue and function as a mechanical barrier in an ovine model. (c) 2013 The Authors. Journal of Tissue Engineering and Regenerative Medicine published by John Wiley & Sons, Ltd.

Synovial fibroblasts (SF) contribute to the pathogenesis of osteoarthritis (OA), but the effects of intra-articular cytokines on SF are not completely understood. The aim of this study was to characterize the interplay between tumor necrosis factor (TNF)α and the anti-inflammatory interleukin (IL)-10. Non-immortalized human SF and SF of the human cell line K4IM were stimulated with recombinant TNFα, IL-10, or TNFα + IL-10 (10 ng/ml each) for 24 h or transduced with an adenoviral vector overexpressing human IL-10 (hIL-10) and subsequently treated with 10 ng/ml TNFα for 24 h. Effects on the gene expression and protein synthesis of IL-6, IL-10, matrix metalloproteinases (MMP)-1, −3, type I collagen, β1-integrin, and CD44 were investigated via real-time detection polymerase chain reaction, immunofluorescence labeling, flow cytometry, and Western blotting. IL-10 release by transduced SF was confirmed with enzyme-linked immunosorbent assay. Both cell populations were activated by TNFα and by TNFα + IL-10, increasing their gene expression and protein synthesis of IL-6, IL-10, MMP-1, and MMP−3 and altering the synthesis of type I collagen, β1-integrin, and CD44. hIL-10 overexpression greatly elevated the gene expression and protein synthesis of IL-10. However, transduction did not significantly affect the gene expression of IL-6, MMP-1, and MMP−3 in SF. The increased expression of pro-inflammatory and catabolic mediators in TNFα-activated SF indicates their role in OA pathogenesis, suggesting they are a potential therapeutic target. Although the vigorousness of the responses of non-immortalized SF and K4IM clearly differ, the K4IM cell line seems to be a suitable model for non-immortalized human SF.

AIMS:To evaluate the impact of human plasma-derived fibronectin (FN) on human subchondral mesenchymal progenitor cells regarding cell migration, proliferation, and chondrogenic differentiation.MATERIALS & METHODS:Human subchondral mesenchymal progenitor cells were analyzed for their migration capacity upon treatment with human plasma-derived FN. Proliferation activity was evaluated by DNA content. For chondrogenesis, cells were cultured in high-density pellet cultures in the presence of FN, TGFβ3, and a combination thereof.RESULTS:Treatment of progenitors with FN significantly increased the number of migrating cells and elevated proliferative activity. Histological staining indicated formation of an extracellular matrix with type II collagen. Gene expression analysis gave no evidence for chondrogenic differentiation mediated by FN, but revealed a significant induction of type II collagen expression.CONCLUSION:FN has a potential to recruit human subchondral mesenchymal progenitor cells, possibly supporting proliferation and matrix assembly in cartilage repair procedures using bioactive implants after microfracture treatment.

The aim of our study was to analyse the clinical and histological outcome after the treatment of focal cartilage defects in non-degenerative and degenerative knees with bone marrow stimulation and subsequent covering with a cell-free resorbable polyglycolic acid–hyaluronan (PGA-HA) implant immersed with autologous platelet-rich plasma (PRP).

This study investigates the adhesion capacity of a polyglycolic acid- (PGA-) hyaluronan scaffold with a structural modification based on a planar polymer (PM) surface in a cadaver cartilage defect model. Two cadaver specimens were used to serially test multiple chondral matrices. In a cadaver hip model, cell free polymer-based cartilage implants with a planar bioinspired PM surface (PGA-PM-scaffolds) were implanted arthroscopically on 10 mm × 15 mm full-thickness femoral hip cartilage lesions. Unprocessed cartilage implants without a bioinspired PM surface were used as control group. The cartilage implants were fixed without and with the use of fibrin glue on femoral hip cartilage defects. After 50 movement cycles and removal of the distraction, a rearthroscopy was performed to assess the outline attachment and integrity of the scaffold. The fixation techniques without and with fibrin fixation showed marginal differences for outline attachment, area coverage, scaffold integrity, and endpoint fixation after 50 cycles. The PGA-PM-scaffolds with fibrin fixation achieved a higher score in terms of the attachment, integrity, and endpoint fixation than the PGA-scaffold on the cartilage defect. Relating to the outline attachment, area coverage, scaffold integrity, and endpoint fixation, the fixation with PGA-PM-scaffolds accomplished significantly better results compared to the PGA-scaffolds (P=0.03752, P=0.03078, P=0.00512, P=0.00512). PGA-PM-scaffolds demonstrate increased observed initial fixation strength in cadaver femoral head defects relative to PGA-scaffold, particularly when fibrin glue is used for fixation.

The aim of our study was to analyze the clinical outcome after repair of cartilage defects of the knee with subchondral drilling and resorbable polymer-based implants immersed with autologous platelet-rich plasma (PRP). Fifty-two patients with focal chondral defects were treated with subchondral drilling, followed by covering with a polyglycolic acid - hyaluronan (PGA-HA) implant (chondrotissue®) immersed with autologous PRP. At 5-year follow-up, patients' situation was assessed using the Knee Injury and Osteoarthritis Outcome Score (KOOS) and compared to the pre-operative situation. The KOOS showed clinically meaningful and significant (p < 0.05) improvement in all subcategories compared to baseline. Subgroup analysis showed that there were no differences in the clinical outcome regarding defect size and localization as well as degenerative condition of the knee. Cartilage repair was complete in 20 out of 21 patients at 4-year follow-up as shown by magnetic resonance observation of cartilage repair tissue (MOCART) scoring. Covering of focal cartilage defects with the PGA-HA implant and PRP after bone marrow stimulation leads to a lasting improvement of the patients' situation.

Disc degeneration alters disc height and mechanics of the spinal column and is associated with lower back pain. In preclinical studies gel-like materials or resorbable polymer-based implants are frequently used to rebuild the nucleus pulposus, aiming at tissue regeneration and restoration of tissue function. To compare the outcome of tissue repair, freeze-dried resorbable polyglycolic acid-hyaluronan (PGA/HA) implants without any bioactive components or bioactivated fibrin (fibrin-serum) was used in a degenerated disc disease model in New Zealand white rabbits. Animals with partial nucleotomy only served as controls. The T2-weighted/fat suppression sequence signal intensity in the nuclear region of operated discs as assessed by magnet resonance imaging was reduced in operated compared to healthy discs, indicating loss of water and did not change from week 1 to month 6 after surgery. Quantification of histological andimmunohistochemical staining indicated that the implantation of PGA/HA leads to significantly more repair tissue compared to nucleotomy only. Type II collagen content of the repair tissue formed after PGA/HA or fibrin-serum treatment is significantly increased compared to controls with nucleotomy only. The data indicate that intervertebral disc augmentation after nucleotomy has a positive effect on repair tissue formation and type II collagen deposition as shown in the rabbit model. (C) 2014 Elsevier Ltd. All rights reserved.

BACKGROUND:Three-dimensional (3D) culture in porous biomaterials as well as stimulation with growth factors are known to be supportive for intervertebral disc cell differentiation and tissue formation. Unless sophisticated releasing systems are used, however, effective concentrations of growth factors are maintained only for a very limited amount of time in in vivo applications. Therefore, we investigated, if an initial boost with transforming growth factor-beta 1 (TGF-beta 1) is capable to induce a lasting effect of superior cartilaginous differentiation in slightly and severely degenerated human annulus fibrosus (AF) cells.METHODS:Human AF tissue was harvested during surgical treatment of six adult patients with lumbar spinal diseases. Grading of disc degeneration was performed with magnet resonance imaging. AF cells were isolated and expanded in monolayer culture and rearranged three-dimensionally in a porous biomaterial consisting of stepwise absorbable poly-glycolic acid and poly-(lactic-co-glycolic) acid and a supportive fine net of non-absorbable polyvinylidene fluoride. An initial boost of TGF-beta 1 or TGF-beta 1 and hyaluronan was applied and compared with controls. Matrix formation was assessed at days 7 and 21 by (1) histological staining of the typical extracellular matrix molecules proteoglycan and type I and type II collagens and by (2) real-time gene expression analysis of aggrecan, decorin, biglycan, type I, II, III, and X collagens as well as of catabolic matrix metalloproteinases MMP-2 and MMP-13.RESULTS:An initial boost with TGF-beta 1 or TGF-beta 1 and hyaluronan did not enhance the expression of characteristic AF matrix molecules in our 3D culture system. AF cells showed high viability in the progressively degrading biomaterial. Stratification by grade of intervertebral disc degeneration showed that AF cells from both, slightly degenerated, or severely degenerated tissue are capable of significant up-regulations of characteristic matrix molecules in 3D culture. AF cells from severely degenerated tissue, however, displayed significantly lower up-regulations in some matrix molecules such as aggrecan.CONCLUSIONS:We failed to show a supportive effect of an initial boost with TGF-beta 1 in our 3D culture system. This underlines the need for further investigations on growth factor releasing systems.

In cartilage regeneration, bio-activated implants are used in stem and progenitor cell-based microfracture cartilage repair procedures. Our aim was to analyze the chondrogenic potential of freeze-dried resorbable polymer-based polyglycolic acid (PGA) scaffolds bio-activated with transforming growth factor-β3 (TGFB3) on human subchondral mesenchymal progenitor cells known from microfracture. Progenitor cells derived from femur heads were cultured in the presence of freeze-dried TGFB3 in high-density pellet culture and in freeze-dried TGFB3-PGA scaffolds for chondrogenic differentiation. Progenitor cell cultures in PGA scaffolds as well as pellet cultures with and without continuous application of TGFB3 served as controls. Release studies showed that freeze-dried TGFB3-PGA scaffolds facilitate a rapid, initial boost-like release of 71.5% of TGFB3 in the first 10 h. Gene expression analysis and histology showed induction of typical chondrogenic markers like type II collagen and formation of cartilaginous tissue in TGFB3-PGA scaffolds seeded with subchondral progenitor cells and in pellet cultures stimulated with freeze-dried TGFB3. Chondrogenic differentiation in freeze-dried TGFB3-PGA scaffolds was comparable to cultures receiving TGFB3 continuously, while non-stimulated controls did not show chondrogenesis during prolonged culture for 14 days. These results suggest that bio-activated, freeze-dried TGFB3-PGA scaffolds have chondrogenic potential and are a promising tool for stem cell-mediated cartilage regeneration.

The influence of gender on the biomechanical outcome after autologous chondrocyte implantation (ACI) including isokinetic muscle strength measurements has not been investigated. The present prospective study was performed to evaluate gender-specific differences in the biomechanical function 48 months after ACI.