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Chemokines are guiding cues for directional trafficking of mesenchymal stem cells (MSC) upon injury and local chemokine delivery at injury sites is an up-to-date strategy to potentiate and prolong recruitment of MSC. In this study we present the chemokine CCL25, also referred to as thymus-expressed chemokine, to mobilize human MSC along positive but not along negative gradients. We hence proceeded to design a biodegradable and injectable release device for CCL25 on the basis of poly(lactic-co-glycolic acid) (PLGA). The conducted studies had the objective to optimize PLGA microparticle fabrication by varying selected formulation parameters, such as polymer type, microparticle size and interior phase composition. We found that microparticles of D V,50 ∼75 µm and fabricated using end-capped polymers, BSA as carrier protein and vortex mixing to produce the primary emulsion yielded high chemokine loading and delayed CCL25 release. To determine bioactivity, we investigated CCL25 released during the microparticle erosion phase and showed that deacidification of the release medium was required to induce significant MSC mobilization. The designed PLGA microparticles represent an effective and convenient off-the-shelf delivery tool for the delayed release of CCL25. However, continuative in vivo proof-of-concept studies are required to demonstrate enhanced recruitment of MSC and/or therapeutical effects in response to CCL25 release microparticles. Statement of Significance With the discovery of chemokines, particularly CXCL12, as stimulators of stem cell migration, the development of devices that release CXCL12 has proceeded quickly in the last few years. In this manuscript we introduce CCL25 as chemokine to induce mobilization of human MSC. This study proceeds to demonstrate how selection of key formulation parameters of CCL25 loading into PLGA microparticles exerts considerable influence on CCL25 release. This is important for a broad range of efforts in in situ tissue engineering where the candidate chemokine and the delivery device need to be selected carefully. The use of such a cell-free CCL25 release device may provide a new therapeutic option in regenerative medicine. [ABSTRACT FROM AUTHOR]

Chemokines are potent stem cell homing and mobilization factors, and artificially increasing the concentrations of specific chemokines at injury sites is an up-to-date strategy to potentiate and prolong the recruitment of endogenous stem cells and to amplify in situ tissue regeneration. We briefly outline the latest progress in stem cell recruitment focusing on the interactions of mesenchymal stem cells (MSCs) with chemokines, complement cascade peptides, bioactive lipids, and glycosaminoglycans (GAGs). We present recent advances in state-of-the-art chemokine delivery devices suitable for various applications and critically evaluate the perspectives and challenges of the chemokine-guided in situ strategy for translation in regenerative medicine. [ABSTRACT FROM AUTHOR]

Abstract: Reprogramming is a new wave in cellular therapies to achieve the vital goals of regenerative medicine. Transdifferentiation, whereas the differentiated state of cells could be reprogrammed into other cell types, meaning cells are no more locked in their differentiated circle. Hence, cells of choice from abundant and easily available sources such as fibroblast and adipose tissue could be converted into cells of demand, to restore the diseased tissues. Before diverting this new approach into effective clinical use, transdifferentiation could not be simply overlooked, as it challenges the normal paradigms of biological laws, where mature cells transdifferentiate not only within same germ layers, but even across the lineage boundaries. How unipotent differentiated cells reprogram into another, and whether transdifferentiation proceeds via a direct cell-to-cell conversion or needs dedifferentiation. To address such questions, MSC were adipogenically differentiated followed by direct transdifferentiation, and subsequently examined by histology, immunohistochemistry, qPCR and single cell analysis. Direct cellular conversion of adipogenic lineage cells into osteogenic or chondrogenic resulted in mixed culture of both lineage cells (adipogenic and new acquiring osteogenic/chondrogenic phenotypes). On molecular level, such conversion was confirmed by significantly upregulated expression of PPARG, FABP4, SPP1 and RUNX2. Chondrogenic transdifferentiation was verified by significantly upregulated expression of PPARG, FABP4, SOX9 and COL2A1. Single cell analysis did not support the direct cell-to-cell conversion, rather described the involvement of dedifferentiation. Moreover, some differentiated single cells did not change their phenotype and were resistant to transdifferentiation, suggesting that differentiated cells behave differently during cellular conversion. An obvious characterization of differentiated cells could be helpful to understand the process of transdifferentiation. [Copyright &y& Elsevier]

Abstract: Extracellular matrix (ECM) is the non-cellular component of tissues, which not only provides biological shelter but also takes part in the cellular decisions for diverse functions. Every tissue has an ECM with unique composition and topology that governs the process of determination, differentiation, proliferation, migration and regeneration of cells. Little is known about the structural organization of matrix especially of MSC-derived adipogenic ECM. Here, we particularly focus on the composition and architecture of the fat ECM to understand the cellular behavior on functional bases. Thus, mesenchymal stem cells (MSC) were adipogenically differentiated, then, were transferred to adipogenic propagation medium, whereas they started the release of lipid droplets leaving bare network of ECM. Microarray analysis was performed, to indentify the molecular machinery of matrix. Adipogenesis was verified by Oil Red O staining of lipid droplets and by qPCR of adipogenic marker genes PPARG and FABP4. Antibody staining demonstrated the presence of collagen type I, II and IV filaments, while alkaline phosphatase activity verified the ossified nature of these filaments. In the adipogenic matrix, the hexagonal structures were abundant followed by octagonal structures, whereas they interwoven in a crisscross manner. Regarding molecular machinery of adipogenic ECM, the bioinformatics analysis revealed the upregulated expression of COL4A1, ITGA7, ITGA7, SDC2, ICAM3, ADAMTS9, TIMP4, GPC1, GPC4 and downregulated expression of COL14A1, ADAMTS5, TIMP2, TIMP3, BGN, LAMA3, ITGA2, ITGA4, ITGB1, ITGB8, CLDN11. Moreover, genes associated with integrins, glycoproteins, laminins, fibronectins, cadherins, selectins and linked signaling pathways were found. Knowledge of the interactive-language between cells and matrix could be beneficial for the artificial designing of biomaterials and bioscaffolds. [Copyright &y& Elsevier]

Purpose: Augmentation of the maxillary sinus with allogenic or alloplastic materials, as well as autologous bone grafts, has inherent disadvantages. Therefore, the aim of our study was to evaluate the long-term clinical repair effect of autologous periosteal bone grafts on atrophic maxillary bone. Patients and Methods: In the present retrospective cohort study, augmentation of the edentulous atrophic posterior maxilla was performed using autologous tissue-engineered periosteal bone grafts based on bioresorbable polymer scaffolds and, in a 1-step procedure, simultaneous insertion of dental implants. The clinical evaluation of 10 patients was performed by radiologic assessment of bone formation, with a follow-up of 5 years. Bone formation was further documented by measuring the bone height and by histologic examination. Results: Excellent clinical and radiologic results were achieved as early as 4 months after transplantation of the periosteal bone grafts. The bone height remained significantly (P < .05) greater (median 14.2 mm) than the preoperative atrophic bone (median 6.9 mm) during the 5-year observation period. Histologically, the bone biopsy specimens of 2 patients obtained after 6 months showed trabecular bone with osteocytes and active osteoblasts. No signs of bone resorption, formation of connective tissue, or necrosis were seen. Conclusion: Our results suggest that the transplantation of autologous periosteal bone grafts and implantation of dental implants in a 1-step procedure is a reliable procedure that leads to bone formation in the edentulous posterior maxilla, remaining stable in the long term for a period of at least 5 years. [Copyright &y& Elsevier]

Abstract: Growth, differentiation and migration factors facilitate the engineering of tissues but need to be administered with defined gradients over a prolonged period of time. In this study insulin as a growth factor for cartilage tissue engineering and a biodegradable PLGA delivery device were used. The aim was to investigate comparatively three different microencapsulation techniques, solid-in-oil-in-water (s/o/w), water-in-oil-in-water (w/o/w) and oil-in-oil-in-water (o/o/w), for the fabrication of insulin-loaded PLGA microspheres with regard to protein loading efficiency, release and degradation kinetics, biological activity of the released protein and phagocytosis of the microspheres. Insulin-loaded PLGA microspheres prepared by all three emulsification techniques had smooth and spherical surfaces with a negative zeta potential. The preparation technique did not affect particle degradation nor induce phagocytosis by human leukocytes. The delivery of structurally intact and biologically active insulin from the microspheres was shown using circular dichroism spectroscopy and a MCF7 cell-based proliferation assay. However, the insulin loading efficiency (w/o/w about 80%, s/o/w 60%, and o/o/w 25%) and the insulin release kinetics were influenced by the microencapsulation technique. The results demonstrate that the w/o/w microspheres are most appropriate, providing a high encapsulation efficiency and low initial burst release, and thus these were finally used for cartilage tissue engineering. Insulin released from w/o/w PLGA microspheres stimulated the formation of cartilage considerably in chondrocyte high density pellet cultures, as determined by increased secretion of proteoglycans and collagen type II. Our results should encourage further studies applying protein-loaded PLGA microspheres in combination with cell transplants or cell-free in situ tissue engineering implants to regenerate cartilage. [Copyright &y& Elsevier]

Abstract: Current tissue engineering strategies are focused on the restoration of pathologically altered tissue architecture by transplantation of cells in combination with supportive scaffolds and biomolecules. In recent years, considerable attention has been given to chitosan (CS)-based materials and their applications in the field of orthopedic tissue engineering. Interesting characteristics that render chitosan suitable for this purpose are a minimal foreign body reaction, an intrinsic antibacterial nature, and the ability to be molded in various geometries and forms such as porous structures, suitable for cell ingrowth and osteoconduction. Due to its favorable gelling properties chitosan can deliver morphogenic factors and pharmaceutical agents in a controlled fashion. Its cationic nature allows it to complex DNA molecules making it an ideal candidate for gene delivery strategies. The ability to manipulate and reconstitute tissue structure and function using this material has tremendous clinical implications and is likely to play a key role in cell and gene therapies in coming years. In this paper we will review the current applications and future directions of CS in articular cartilage, intervertebral disk and bone tissue engineering. [Copyright &y& Elsevier]

Summary: Objective: The aim of this study was to evaluate the growth characteristics of freshly isolated porcine chondrocytes in high-density pellet cultures and to preliminary investigate their use in an interactive in vitro model with synovial fibroblast cell lines to study rheumatoid arthritis (RA). Design: 1.8×106 chondrocytes/cm2 were seeded in 48-multiwell plates. Thickness, cell number and cell distribution in pellet cross sections were documented over a 22-day-long period. Alcian blue staining, type I and type II collagen staining, real-time reverse transcriptase polymerase polymerase chain reaction (RT-PCR) and high performance liquid chromatography (HPLC) were used to characterize cartilage extracellular matrix (ECM) formation, and cell proliferation was demonstrated by Ki67 staining. Furthermore, 2-week-old chondrocyte pellets were co-cultured for additional 2 weeks with two human synovial fibroblast cell lines derived from a normal donor (non-invasive cell line) and a RA patient (invasive–aggressive (IA) cell line), respectively. Results: Chondrocyte pellets from 11 individual preparations showed a significant increase in pellet thickness from 44±19μm (day 3) to 282±19μm (day 22). Calculation of chondrocyte distribution, cell number and pellet thickness indicated that pellet growth was due to ECM formation and not cell proliferation. This was also confirmed by low numbers of Ki67 positive chondrocytes and absence of cell clusters. HPLC, messenger RNA-analysis, histochemistry and antibody staining verified the expression of ECM components such as type II collagen, whereas type I collagen expression was very low. In contrast to the non-aggressive synovial fibroblast cell line, the IA synovial fibroblast cell line clearly showed cartilage invasion. Conclusion: Pellet formation of freshly isolated chondrocytes followed a reproducible developmental kinetics and showed typical immature hyaline cartilage properties. Such uniform cartilage pellets are very useful as a substrate for interactive cell culture models that simulate diseases like RA. [Copyright &y& Elsevier]

Several cell-based tissue-engineering therapies are emerging to regenerate damaged tissues. These strategies use autologous cells in combination with bioresorbable delivery materials. Major functions of a delivery scaffold are to provide initial mechanical stability, homogenous three-dimensional cell distribution, improved tissue differentiation, suitable handling and properties for delivery and fixation into patients. Delivery of cells can be achieved using injectable matrices, soft scaffolds, membranes, solid load-bearing scaffolds or immunoprotective macroencapsulation. Thus, to expand the clinical potential, next generation therapies will depend on smart delivery concepts that make use of the regenerative potential of stem cells, morphogenetic growth factors and biomimetic materials. [Copyright &y& Elsevier]

One of the milestones in tissue engineering has been the development of 3D scaffolds that guide cells to form functional tissue. Recently, mouldless manufacturing techniques, known as solid free-form fabrication (SFF), or rapid prototyping, have been successfully used to fabricate complex scaffolds. Similarly, to achieve simultaneous addition of cells during the scaffold fabrication, novel robotic assembly and automated 3D cell encapsulation techniques are being developed. As a result of these technologies, tissue-engineered constructs can be prepared that contain a controlled spatial distribution of cells and growth factors, as well as engineered gradients of scaffold materials with a predicted microstructure. Here, we review the application, advancement and future directions of SFF techniques in the design and creation of scaffolds for use in clinically driven tissue engineering. [Copyright &y& Elsevier]

Damaged or diseased articular cartilage frequently leads to progressive debilitation resulting in a marked decrease in the quality of life. Tissue engineering, a budding field in modern biomedical sciences, promises creation of viable substitutes for failing organs or tissues. It represents the amalgamation of rapid developments in cellular and molecular biology on the one hand and material, chemical and mechanical engineering on the other. Current tissue engineering approaches are mainly focused on the restoration of pathologically altered tissue structure based on the transplantation of cells in combination with supportive matrices and biomolecules. The ability to manipulate and reconstitute tissue structure and function in vitro has tremendous clinical implications and is likely to have a key role in cell and gene therapies in coming years. [Copyright &y& Elsevier]

Abstract: Tracking of transplanted stem cells is essential to monitor safety and efficiency of cell-based therapies. Magnetic resonance imaging (MRI) offers a very sensitive, repetitive and non-invasive in vivo detection of magnetically labeled cells but labeling with commercial superparamagnetic iron oxide nanoparticles (SPIONs) is still problematic because of low labeling efficiencies and the need of potentially toxic transfection agents. In this study, new experimental citrate-coated SPIONs and commercial Endorem and Resovist SPIONs were investigated comparatively in terms of in vitro labeling efficiency, effects on stem cell functionality and in vivo MRI visualization. Efficient labeling of human mesenchymal stem cells (MSCs) without transfection agents was only achieved with Citrate SPIONs. Magnetic labeling of human MSCs did not affect cell proliferation, presentation of typical cell surface marker antigens and differentiation into the adipogenic and osteogenic lineages. However, chondrogenic differentiation and chemotaxis were significantly impaired with increasing SPION incorporation. Transplanted SPION-labeled MSCs were visualized in vivo after intramuscular injection in rats by 7T-MRI and were retrieved ex vivo by Prussian Blue and immunohistochemical stainings. Though a careful titration of SPION incorporation, cellular function and MRI visualization is essential, Citrate SPIONs are very efficient intracellular magnetic labels for in vivo stem cell tracking by MRI. [Copyright &y& Elsevier]

Abstract: Although rabbits are commonly used as tendon repair model, interpretative tools are divergent and comprehensive scoring systems are lacking. Hence, the aim was to develop a multifaceted scoring system to characterize healing in a partial Achilles tendon defect model. A 3 mm diameter defect was created in the midsubstance of the medial M. gastrocnemius tendon, which remained untreated or was filled with a polyglycolic-acid (PGA) scaffold + fibrin and either left cell-free or seeded with Achilles tenocytes. After 6 and 12 weeks, tendon repair was assessed macroscopically and histologically using self-constructed scores. Macroscopical scoring revealed superior results in the tenocyte seeded PGA + fibrin group compared with the controls at both time points. Histology of all operated tendons after 6 weeks proved extracellular matrix (ECM) disorganization, hypercellularity and occurrence of irregular running elastic fibres with no significance between the groups. Some inflammation was associated with PGA implantation and increased sulphated proteoglycan deposition predominantly with the empty defects. After 12 weeks defect areas became hard to recognize and differences between groups, except for the increased sulphated proteoglycans content in the empty defects, were almost nullified. We describe a partial Achilles tendon defect model and versatile scoring tools applicable for characterizing biomaterial-supported tendon healing. [Copyright &y& Elsevier]

Abstract: The high-density micromass culture has been widely applied to study chondrocyte cell physiology and pathophysiological mechanisms. Since an integrated image has not been established so far, we analyzed the phenotypic alterations of human articular chondrocytes in this model on the broad molecular level. Freshly isolated chondrocytes were assembled as micromasses and maintained up to 6weeks in medium containing human serum. Formation of cartilaginous extracellular matrix (ECM) was evaluated by histological and immunohistochemical staining. At 0, 3 and 6weeks, chondrocyte micromasses were subjected to gene expression analysis using oligonucleotide microarrays and real-time RT-PCR. Micromasses developed a cartilaginous ECM rich in proteoglycans and type II collagen. On gene expression level, time-dependent expression patterns was observed. The induction of genes associated with cartilage-specific ECM (COL2A1 and COL11A1) and developmental signaling (GDF5, GDF10, ID1, ID4 and FGFR1 - 3) indicated redifferentiation within the first 3weeks. The repression of genes related to stress response (HSPA1A and HSPA4), apoptotic events (HYOU1, NFKBIA and TRAF1), and degradation (MMP1, MMP10 and MMP12) suggested a recovery of chondrocytes. Constant expression of other chondrogenic (ACAN, FN1 and MGP) and hypertrophic markers (COL10A1, ALPL, PTHR1 and PTHR2) indicated a pattern of phenotypic maintenance. Simultaneously, the expression of chondrogenic growth (BMP6, TGFA, FGF1 and FGF2) and transcription factors (SOX9, EGR1, HES1 and TGIF1), and other cartilage ECM-related genes (COMP and PRG4) was consistently repressed and expression of collagens related to dedifferentiation (COL1A1 and COL3A1) was steadily induced indicating a progressing loss of cartilage phenotype. Likewise, a steady increase of genes associated with proliferation (GAS6, SERPINF1, VEGFB and VEGFC) and apoptosis (DRAM, DPAK1, HSPB, GPX1, NGFRAP1 and TIA1) was observed. Sequence and interplay of identified expression patterns suggest that chondrocyte micromass cultures maintain a differentiated phenotype up to 3weeks in vitro and might be useful for studying chondrocyte biology, pathophysiology and differentiation. Cultivation longer than 6weeks leads to progressing dedifferentiation of chondrocytes that should be considered on long-term evaluations. [Copyright &y& Elsevier]

Abstract: This study was conducted to determine physiological and functional features of primary and immortalized canine chondrocytes. Chondrocytes were immortalized by introducing the catalytic component of human telomerase namely human telomerase reverse transcriptase (hTERT). Primary chondrocytes lost their characteristic phenotype and growth properties whereas the immortalized cells remained polygonal with rapid growth rate. The expression of chondrocyte-specific markers decreased many-fold whereas that of chondrocyte-non-specific gene increased in primary chondrocytes. The immortalized cells did not express chondrocyte-specific genes in monolayers. Both primary and immortalized cells were encapsulated in alginate microspheres to construct three-dimensional (3D) culture system. As the primary chondrocytes, also the telomerase-transfected cells adopted a chondrocyte-specific gene expression pattern in alginate culture. Thus, the expression of telomerase represents possibility to expand chondrocytes without limitation while maintaining the chondrocyte-specific phenotype in 3D cultures. Use of such cells provides a standardized tool for testing different tissue engineering applications in canine model. [Copyright &y& Elsevier]

Abstract: A variety of chemokines has been shown to recruit human bone marrow-derived mesenchymal stem cells (MSC) and may be potential candidates for chemokine-based tissue regeneration approaches. The aim of our study was to determine whether the chemokine CXCL7 stimulates migration of human bone marrow-derived MSC and to analyze the effect of CXCL7 on the recruitment of MSC on the broad molecular level. Chemotaxis assays documented that high doses of CXCL7 significantly recruited MSC. Gene expression profiling using oligonucleotide microarrays showed that MSC treated with CXCL7 differentially expressed genes related to cell migration, cell adhesion and extracellular matrix remodeling. Pathway analysis showed that CXCL7 induced the expression of all chemokines binding the interleukin (IL) receptors A and B, CXCR1 and CXCR2, as well as the IL6 signal transducer (gp130) and its ligands IL6 and leukemia inhibitory factor (LIF). Induction of differentially expressed chemokines CXCL1-3, CXCL5, and CXCL6 as well as LIF and gp130 in MSC by CXCL7 was verified by real-time polymerase chain reaction. Immunoassay of cell culture supernatants confirmed elevated levels of the interleukins 6 and 8 in MSC upon treatment with CXCL7. Chemotaxis assays showed that interleukin 6 did not recruit MSC. In conclusion, CXCL7 significantly stimulates the migration of human MSC in vitro. Pathway analysis suggests that recruitment of human MSC by CXCL7 is supported by the induction of ligands of the interleukin 8 receptors, synergistically activating the respective signaling pathways. [Copyright &y& Elsevier]

Abstract: For bone repair, transplantation of periosteal progenitor cells (PCs), which had been amplified within supportive scaffolds, is applied clinically. More innovative bone tissue engineering approaches focus on the in situ recruitment of stem and progenitor cells to defective sites and their subsequent use for guided tissue repair. Chemokines are known to induce the directed migration of bone marrow CD34− mesenchymal stem cells (MSCs). The aim of our study was to determine the chemokine receptor expression profile of human CD34− PCs and to demonstrate that these cells migrate upon stimulation with selected chemokines. PCs were isolated from periosteum of the mastoid bone and displayed a homogenous cell population presenting an MSC-related cell-surface antigen profile (ALCAM+, SH2+, SH3+, CD14−, CD34−, CD44+, CD45−, CD90+). The expression profile of chemokine receptors was determined by real-time PCR and immunohistochemistry. Both methods consistently demonstrated that PCs express receptors of all four chemokine subfamilies CC, CXC, CX3C, and C. Migration of PCs and a dose-dependent migratory effect of the chemokines CCL2 (MCP1), CCL25 (TECK), CXCL8 (IL8), CXCL12 (SDF1α), and CXCL13 (BCA1), but not CCL22 (MDC) were demonstrated using a 96-multiwell chemotaxis assay. In conclusion, for the first time, here we report that human PCs express chemokine receptors, present their profile, and demonstrate a dose-dependent migratory effect of distinct chemokines on these cells. These results are promising towards in situ bone repair therapies based on guiding PCs to bone defects, and encourage further in vivo studies. [Copyright &y& Elsevier]

Abstract: The aim of our study was the evaluation of a cell-free cartilage implant that allows the recruitment of mesenchymal stem and progenitor cells by chemo-attractants and subsequent guidance of the progenitors to form cartilage repair tissue after microfracture. Chemotactic activity of human serum on human mesenchymal progenitors was tested in 96-well chemotaxis assays and chondrogenic differentiation was assessed by gene expression profiling after stimulating progenitors with hyaluronan in high-density cultures. Autologous serum and hyaluronan were combined with polyglycolic acid (PGA) scaffolds and were implanted into full-thickness articular cartilage defects of the sheep pre-treated with microfracture. Defects treated with microfracture served as controls. Human serum was a potent chemo-attractant and efficiently recruited mesenchymal progenitors. Chondrogenic differentiation of progenitors upon stimulation with hyaluronan was shown by the induction of typical chondrogenic marker genes like type II collagen and aggrecan. Three months after implantation of the cell-free implant, histological analysis documented the formation of a cartilaginous repair tissue. Controls treated with microfracture showed no formation of repair tissue. The cell-free cartilage implant consisting of autologous serum, hyaluronan and PGA utilizes the migration and differentiation potential of mesenchymal progenitors for cartilage regeneration and is well suited for the treatment of cartilage defects after microfracture. [Copyright &y& Elsevier]

Abstract: Cartilage tissue engineering is applied clinically to cover and regenerate articular cartilage defects. In this study autologous human cartilage tissue engineering grafts based on bioresorbable polyglactin/polydioxanone scaffolds were analyzed on the broad molecular level. RNA from freshly isolated, primary and expanded adult articular chondrocytes and from three-dimensional cartilage grafts were used for gene expression profiling using oligonucleotide microarrays. The capacity of cartilage grafts to form cartilage matrix was evaluated after subcutaneous transplantation into nude mice. Gene expression profiling showed reproducibly the regulation of 905 genes and documented that chondrocytes undergo fundamental changes during cartilage tissue engineering regarding chondrocyte metabolism, growth, and differentiation. Three-dimensional assembly of expanded, dedifferentiated chondrocytes initiated the re-differentiation of cells that was accompanied by the reversal of the expression profile of multiple players of the transforming growth factor (TGF) signaling pathway including growth and differentiation factor-5 and inhibitor of differentiation-1 as well as by the induction of typical cartilage-related matrix genes such as type II collagen and cartilage oligomeric matrix protein. Cartilage grafts formed a cartilaginous matrix after transplantation into nude mice. Three-dimensional tissue culture of expanded articular chondrocytes initiates chondrocyte re-differentiation in vitro and leads to the maturation of cartilage grafts towards hyaline cartilage in vivo. [Copyright &y& Elsevier]

Abstract: The objective of our study was to evaluate the integration of autologous cartilage tissue engineering transplants based on resorbable polyglactin/polydioxanone scaffolds into full-thickness cartilage defects of horses. Cartilage biopsies were taken from the non-load-bearing area of the lateral talus of the left tibiotarsal joint of eight healthy Haflinger horses. Tissue engineering cartilage transplants were generated by three-dimensional arrangement of autologous chondrocytes in biocompatible and resorbable polymer scaffolds. Full-thickness cartilage defects of 8mm in diameter were created in the tubular bone condyle of the fetlock joint and cartilage grafts were fixed using an anchor system, while defects without grafting served as controls. After 6 and 12 months the repair tissue was evaluated histologically and showed formation of a cartilaginous tissue and good integration into the surrounding host tissue with firm bonding of the graft to the adjacent cartilage and the underlying subchondral bone. Biochemical analysis demonstrated that the content of glycosaminoglycans and hydroxyproline is comparable in repair tissue derived from treated and control defects. The use of three-dimensional autologous cartilage transplants based on resorbable polymer scaffolds ensures secure fixation, good integration of the graft into cartilage lesions, and is therefore suggested as a promising therapeutic option for the treatment of cartilage defects. [Copyright &y& Elsevier]

Tissue engineering of an elastic cartilage graft that meets the criterion for both structural and functional integration into host tissue, as well as allowing for a clinically tolerable immune response, is a challenging endeavour. Conventional scaffold technologies have limitations in their ability to design and fabricate complex-shaped matrix architectures of structural and mechanical equivalence to elastic cartilage found in the body. We attempted to investigate the potential of conventionally isolated and passaged chondrocytes (2D environment) when seeded and cultured in combination with a biomimetic hydrogel in a mechanically stable and biomimetic composite matrix to form elastic cartilage within ectopic implantation sites. In vitro cultured scaffold/hydrogel/chondrocytes constructs showed islets of cartilage and mineralized tissue formation within the cell-seeded specimens in both pig and rabbit models. Specimens with no cells seeded showed only vascularized fibrous tissue ingrowth. These studies demonstrated the potential of such scaffold/hydrogel/cell constructs to support chondrogenesis in vivo. However, it also showed that even mechanically stable scaffolds do not allow regeneration of a large mass of structural and functional cartilage within a matrix architecture seeded with 2D passaged chondrocytes in combination with a cell biomimetic carrier. Hence, future experiments will be designed to evaluate an initial 3D culture of chondrocytes, effect on cell phenotype and their subsequent culture within biomimetic 3D scaffold/cell constructs. [Copyright &y& Elsevier]

In this study we have used lectin histochemistry and scanning electron microscopy (SEM) to assess the growth and characterise the differentiation of human respiratory epithelial cells (REC) cultured on two biomaterial scaffolds. The first scaffold, based on a hyaluronic acid derivative, was observed to be non-adhesive for REC. This lack of adhesion was found to be unrelated to the presence of the hyaluronic acid binding domain on the surface of isolated REC. The other scaffold, consisting of equine collagen, was observed to encourage REC spreading and adhesion. Positive Ulex Europaeus agglutinin (UEA) lectin staining of this preparation indicated the presence of ciliated REC on the scaffold surface. However, the marked decrease in peanut agglutinin (PNA) positive staining, relative to that of control cultures and native tissue, indicates a dedifferentiation of the secretory cells of the REC monolayer. SEM analysis of REC cultured on the collagen scaffold confirmed the presence of ciliated cells thereby validating the UEA positive staining. The presence of both established and developing cilia was also verified. This study indicates that collagen biomaterials are appropriate for the tissue engineering of REC. Furthermore, that UEA and PNA staining is a useful tool in the characterisation of cells cultured on biomaterials, therefore helpful in identifying biomaterials that are suitable for specific tissue engineering purposes. [Copyright &y& Elsevier]