Cell-based Cartilage Regeneration Research Articles

Dual microparticles programmed delivery system regulating stem cell-based cartilage regeneration by cartilage-specific matrix hydrogels

Bone marrow mesenchymal stem cells (BMSCs)-based engineered cartilage usually faces the significant challenge of endochondral ossification tendency in an ectopic environment because of the inevitable vascular infiltration during chondrogenic differentiation and developmental stages. Additionally, current scaffold-free BMSC cartilage regeneration requires a long in vitro preinduction time for chondrogenic differentiation before in vivo transplantation, which limits its application in multiple cartilage defect repair. Therefore, ideal stem cell-based cartilage regeneration needs to meet time-dependent requirements of both early chondrogenic and late anti-angiogenic microenvironments in vivo. Here, we developed a cartilage-specific matrix hydrogel (CMH) with a dual microparticle-based programmed delivery system (dM-PDs) to dynamically regulate in vivo stem cell-based cartilage regeneration without in vitro preinduction. In this study, CMH scaffolds offer a three-dimensional matrix microenvironment for tissue regeneration, while dM-PDs has an early chondrogenic induction function to promote cartilage-specific differentiation and exerts a late anti-angiogenic effect to stabilize the cartilaginous phenotype. Stem cell-based cartilage regeneration was successfully achieved by sequentially dynamic regulation in vivo by dM-PDs, in which TGFβ3-loaded microparticles effectively activated the early-stage TGFβ/Smad signaling pathway and levatinib-loaded microparticles regulated the late-stage VEGF/TIMP signaling pathway. This study demonstrates a universal stem cell-based cartilage regeneration technique for clinical cartilage defects repair.

TNF-α activates RELA expression via TNFRSF1B to upregulate OPA1 expression and inhibit chondrogenic differentiation of human adipose stem cells

BackgroundTumor necrosis factor-alpha (TNF-α), one of the pro-inflammatory cytokines mediating the local inflammatory process in joints, inhibits cartilage formation and has a detrimental effect on stem cell-based cartilage regeneration for the treatment of osteoarthritis (OA). However, the mechanisms behind this inhibitory effect are still poorly understood. Mitochondrial morphological changes mediated by mitochondrial fusion and fission are highly plastic, are quite sensitive to environmental stimuli and play a crucial role in maintaining cell structure and function. In our study, chondrogenic differentiated human adipose stem cells (hADSCs) were exposed to TNF-α and the effect of TNF-α on the ability of hADSCs to chondrogenic differentiate and on mitochondrial fusion and fission was observed and analyzed. The aim was to investigate the role and mechanisms of mitochondrial fusion and fission regulation in the chondrogenic differentiation of hADSCs under normal conditions and under exposure to TNF-α.MethodsWe used flow cytometry to identify hADSCs immunophenotypes CD29, CD44, CD34, CD45, and HLA-DR. Alcian blue staining and Sirius red staining were used to observe the formation of proteoglycans and collagen during the chondrogenic differentiation of hADSCs, respectively. The mRNA and protein expression levels of the cartilage formation marker SOX9, type II collagen (COL2A1), and Aggrecan were measured by real-time fluorescent quantitative PCR (RT-qPCR) and western blot, respectively. The fluorescent probes MitoTracker® Red CMXRos and JC-1 were used to visualize mitochondria morphology and detect mitochondrial membrane electricity (MMP). Affymetrix PrimeView™ chips were used for gene expression profiling.ResultsThe results showed that the chondrogenic differentiation of hADSCs was inhibited in the presence of TNF-α that optic atrophy 1 (OPA1) expression was significantly upregulated and mitochondria were prolonged and interconnected during this process. Gene microarray and RT-qPCR data showed that the presence of TNF-α led to increased expression of TNFα receptor 2 (TNFRSF1B) and RELA during chondrogenic differentiation of hADSCs.ConclusionsTNF-α inhibits chondrogenic differentiation of human adipose stem cells by activating RELA expression through TNFRSF1B upregulating OPA1 expression thereby increasing mitochondrial fusion.

Evaluation of Autoregulated RUNX2 Suppression on Mesenchymal Stem Cell-based Cartilage Regeneration in Complex Pathological Environments

Evaluation of Autoregulated RUNX2 Suppression on Mesenchymal Stem Cell-based Cartilage Regeneration in Complex Pathological Environments

Inhibition of LDL receptor-related protein 3 suppresses chondrogenesis of stem cells, inhibits proliferation, and promotes apoptosis

Articular cartilage defects remain the most common and challenging joint disease. Cartilage lacks the self-healing capacity after injury due to its avascularity. Recently, stem cell-based therapy has been applied for cartilage regeneration. However, the critical target for stem cells during chondrogenesis remains unclear. We first reported that LDL receptor-related protein 3 (LRP3) expression was markedly increased during chondrogenesis in stem cells. Furthermore, LRP3 was an effective chondrogenic stimulator, as confirmed by knockdown and overexpression experiments and RNA sequencing. In addition, inhibition of LRP3 suppressed proliferation and induced apoptosis. Therefore, our study first defined a new chondrogenic stimulator, LRP3, with detailed clarification, which provided a novel target for stem cell-based cartilage regeneration.

The Hunt Is On! In Pursuit of the Ideal Stem Cell Population for Cartilage Regeneration.

Cartilage injury and degeneration are hallmarks of osteoarthritis (OA), the most common joint disease. OA is a major contributor to pain, loss of function, and reduced quality of life. Over the last decade, considerable research efforts have focused on cell-based therapies, including several stem cell-derived approaches to reverse the cartilage alterations associated with OA. Although several tissue sources for deriving cell-based therapies have been identified, none of the resident stem cell populations have adequately fulfilled the promise of curing OA. Indeed, many cell products do not contain true stem cells. As well, issues with aggressive marketing efforts, combined with a lack of evidence regarding efficacy, lead the several national regulatory bodies to discontinue the use of stem cell therapy for OA until more robust evidence becomes available. A review of the evidence is timely to address the status of cell-based cartilage regeneration. The promise of stem cell therapy is not new and has been used successfully to treat non-arthritic diseases, such as hematopoietic and muscle disorders. These fields of regenerative therapy have the advantage of a considerable foundation of knowledge in the area of stem cell repair mechanisms, the role of the stem cell niche, and niche-supporting cells. This foundation is lacking in the field of cartilage repair. So, where should we look for the ideal stem cell to regenerate cartilage? It has recently been discovered that cartilage itself may contain a population of SC-like progenitors. Other potential tissues include stem cell-rich dental pulp and the adolescent growth plate, the latter of which contains chondrocyte progenitors essential for producing the cartilage scaffold needed for bone growth. In this article, we review the progress on stem cell therapies for arthritic disorders, focusing on the various stem cell populations previously used for cartilage regeneration, successful cases of stem cell therapies in muscle and hemopoietic disorders, some of the reasons why these other fields have been successful (i.e., “lessons learned” to be applied to OA stem cell therapy), and finally, novel potential sources of stem cells for regenerating damaged cartilage in vivo.

A high-resolution route map reveals distinct stages of chondrocyte dedifferentiation for cartilage regeneration

Articular cartilage damage is a universal health problem. Despite recent progress, chondrocyte dedifferentiation has severely compromised the clinical outcomes of cell-based cartilage regeneration. Loss-of-function changes are frequently observed in chondrocyte expansion and other pathological conditions, but the characteristics and intermediate molecular mechanisms remain unclear. In this study, we demonstrate a time-lapse atlas of chondrocyte dedifferentiation to provide molecular details and informative biomarkers associated with clinical chondrocyte evaluation. We performed various assays, such as single-cell RNA sequencing (scRNA-seq), live-cell metabolic assays, and assays for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq), to develop a biphasic dedifferentiation model consisting of early and late dedifferentiation stages. Early-stage chondrocytes exhibited a glycolytic phenotype with increased expression of genes involved in metabolism and antioxidation, whereas late-stage chondrocytes exhibited ultrastructural changes involving mitochondrial damage and stress-associated chromatin remodeling. Using the chemical inhibitor BTB06584, we revealed that early and late dedifferentiated chondrocytes possessed distinct recovery potentials from functional phenotype loss. Notably, this two-stage transition was also validated in human chondrocytes. An image-based approach was established for clinical use to efficiently predict chondrocyte plasticity using stage-specific biomarkers. Overall, this study lays a foundation to improve the quality of chondrocytes in clinical use and provides deep insights into chondrocyte dedifferentiation.

Stem cell-based cartilage regeneration technology

Stem cell-based cartilage regeneration technology

Differentiation of adipose-derived stem cells to chondrocytes using electrospraying

An important challenge in the fabrication of tissue engineered constructs for regenerative medical applications is the development of processes capable of delivering cells and biomaterials to specific locations in a consistent manner. Electrospraying live cells has been introduced in recent years as a cell seeding method, but its effect on phenotype nor genotype has not been explored. A promising candidate for the cellular component of these constructs are human adipose-derived stem cells (hASCs), which are multipotent stem cells that can be differentiated into fat, bone, and cartilage cells. They can be easily and safely obtained from adipose tissue, regardless of the age and sex of the donor. Moreover, these cells can be maintained and expanded in culture for long periods of time without losing their differentiation capacity. In this study, hASCs directly incorporated into a polymer solution were electrosprayed, inducing differentiation into chondrocytes, without the addition of any exogenous factors. Multiple studies have demonstrated the effects of exposing hASCs to biomolecules—such as soluble growth factors, chemokines, and morphogens—to induce chondrogenesis. Transforming growth factors (e.g., TGF-β) and bone morphogenetic proteins are particularly known to play essential roles in the induction of chondrogenesis. Although growth factors have great therapeutic potential for cell-based cartilage regeneration, these growth factor-based therapies have presented several clinical complications, including high dose requirements, low half-life, protein instability, higher costs, and adverse effects in vivo. The present data suggests that electrospraying has great potential as hASCs-based therapy for cartilage regeneration.

The impact of photobiomodulation on the chondrogenic potential of adipose-derived stromal/stem cells.

Due to their capacity to differentiate into the chondrogenic lineage, adipose-derived stromal/stem cells (ASC) are a promising source of therapeutically relevant cells for cartilage tissue regeneration. Their differentiation potential, however, varies between patients. In our study, we aim to stimulate ASC towards a more reliable chondrogenic phenotype using photobiomodulation (PBM). LED devices of either blue (475nm), green (516nm) or red (635nm) light were used to treat human ASC from donors of varying chondrogenic potential. The treatment was applied either once during the 2D expansion phase or repeatedly during the 3D differentiation phase. Chondrogenic differentiation was assessed via pellet size, GAG/DNA content, histology and gene expression analysis. Reactions to PBM were found to be wavelength-dependent and more pronounced when the treatment was applied during expansion. Donors were assigned to responder categories according to their response to the treatment during expansion, whereby good responders were mainly donors with low intrinsic chondrogenic potential. Exposed to light, they revealed a particularly high relative increase in pellet size (more than twice the size of untreated controls after red light PBM), intense collagen type II immunostaining (low/absent in untreated controls) and activation of otherwise absent COL2A1 expression. Conversely, on a donor with high intrinsic chondrogenic potential, light had adverse effects. When applied with shorter wavelengths (blue, green), it led to reduced pellet size, GAG/DNA content and collagen type II immunostaining. However, when PBM was applied in 3D, the same donor was the only one to react with increased differentiation to all three wavelengths. We were able to demonstrate that PBM can be used to enhance or hamper chondrogenesis of ASC, and that success depends on treatment parameters and intrinsic cellular potential. The improvement of chondrogenesis in donors with low intrinsic potential highlights PBM as potent tool for cell-based cartilage regeneration. Its cost-effectiveness and ease of use make for an attractive treatment option to enhance the performance of ASC in cartilage tissue engineering.

Effects of Mechanical Compression on Chondrogenesis of Human Synovium-Derived Mesenchymal Stem Cells in Agarose Hydrogel.

Mechanical compression is a double-edged sword for cartilage remodeling, and the effect of mechanical compression on chondrogenic differentiation still remains elusive to date. Herein, we investigate the effect of mechanical dynamic compression on the chondrogenic differentiation of human synovium-derived mesenchymal stem cells (SMSCs). To this aim, SMSCs encapsulated in agarose hydrogels were cultured in chondrogenic-induced medium with or without dynamic compression. Dynamic compression was applied at either early time-point (day 1) or late time-point (day 21) during chondrogenic induction period. We found that dynamic compression initiated at early time-point downregulated the expression level of chondrocyte-specific markers as well as hypertrophy-specific markers compared with unloaded control. On the contrary, dynamic compression applied at late time-point not only enhanced the levels of cartilage matrix gene expression, but also suppressed the hypertrophic development of SMSCs compared with unloaded controls. Taken together, our findings suggest that dynamic mechanical compression loading not only promotes chondrogenic differentiation of SMSCs, but also plays a vital role in the maintenance of cartilage phenotype, and our findings also provide an experimental guide for stem cell-based cartilage repair and regeneration.

Nanoparticle-Mediated TGF-β Release from Microribbon-Based Hydrogels Accelerates Stem Cell-Based Cartilage Formation In Vivo.

Conventional nanoporous hydrogels often lead to slow cartilage deposition by MSCs in 3D due to physical constraints and requirement for degradation. Our group has recently reported macroporous gelatin microribbon (μRB) hydrogels, which substantially accelerate MSC-based cartilage formation in vitro compared to conventional gelatin hydrogels. To facilitate translating the use of μRB-based scaffolds for supporting stem cell-based cartilage regeneration in vivo, there remains a need to develop a customize-designed drug delivery system that can be incorporated into μRB-based scaffolds. Towards this goal, here we report polydopamine-coated mesoporous silica nanoparticles (MSNs) that can be stably incorporated within the macroporous μRB scaffolds,and allow tunable release of transforming growth factor (TGF)-β3. We hypothesize that increasing concentration of polydopamine coating on MSNs will slow down TGF- β3 release, and TGF-β3 release from polydopamine-coated MSNs can enhance MSC-based cartilage formation in vitro and in vivo. We demonstrate that TGF-β3 released from MSNs enhance MSC-based cartilage regeneration in vitro to levels comparable to freshly added TGF-β3 in the medium, as shown bybiochemical assays, mechanical testing, and histology. Furthermore, when implanted in vivo in a mouse subcutaneous model, onlythe group containing MSN-mediated TGF-β3 release supported continuous cartilage formation, whereas control group without MSN showed loss of cartilage matrix and undesirable endochondral ossification. The modular design of MSN-mediated drug delivery can be customized for delivering multiple drugs with individually optimized release kinetics, and may be applicable to enhance regeneration of other tissue types.

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.

Multi-Disciplinary Approaches for Cell-Based Cartilage Regeneration.

Articular cartilage does not regenerate in adults. A lot of time and resources have been dedicated to cartilage regeneration research. The current understanding suggests that multi-disciplinary approach including biologic, genetic, and mechanical stimulations may be needed for cell-based cartilage regeneration. This review summarizes contents of a workshop sponsored by International Combined Orthopaedic Societies during the 2019 annual meeting of the Orthopaedic Research Society held in Austin, Texas. Three approaches for cell-based cartilage regeneration were introduced, including cellular basis of chondrogenesis, gene-enhanced cartilage regeneration, and physical modulation to divert stem cells to chondrogenic cell fate. While the complicated nature of cartilage regeneration has not allowed us to achieve successful regeneration of hyaline articular cartilage so far, the utilization of multi-disciplinary approaches in various fields of biomedical engineering will provide means to achieve this goal faster. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:463-472, 2020.

An Injectable Bioorthogonal Dextran Hydrogel for Enhanced Chondrogenesis of Primary Stem Cells.

The purpose of this study is to corroborate the feasibility of an injectable dextran hydrogel, in situ formed through a catalyst-free bioorthogonal reaction, for cartilage regeneration with adult chondrocytes or adipose-derived stem cells (ADSCs). To this end, an injectable dextran (dextran-AN) hydrogel encapsulating the rabbit adult chondrocytes or rabbit ADSCs (rADSCs) or human ADSCs (hADSCs) was successfully prepared by simply incubating the cells with dibenzocyclooctyne-dextran conjugate and azide-dextran conjugate under physiological conditions through a copper-free bioorthogonal reaction without using additional reagents. The possibility of dextran-AN hydrogel/adult rabbit chondrocyte constructs for in vivo chondrogenesis was first assessed in a rabbit osteochondral defect model. Data indicated that the hydrogel was biocompatible against surrounding tissue and could promote adult chondrocyte survival and in vivo chondrogenesis within 8 weeks. Next, in vitro and in vivo chondrogenetic differentiation of rADSCs encapsulating in dextran-AN hydrogel was studied systemically. Compared to the agarose hydrogel as a positive control, the dextran-AN hydrogel elicited comparable ability to support in vitro chondrogenesis of rADSCs within 3 weeks but caused better cartilage regeneration within 6 weeks in a rabbit osteochondral defect model. Furthermore, miR29A downregulating of hADSCs was applied for in vivo chondrogenesis by subcutaneous in situ formation of dextran-AN hydrogel containing the hADSCs in a balb/c nude mouse model. Within 4-week regeneration, the hydrogel group illustrated upregulated mRNA expression level of chondrogenesis-related biomarkers. In addition, improved production of neocartilage extracellular matrices (type-II collagen and glycosaminoglycan) was found for miR29A-downregulatd hADSC group in comparison with miR29A-upregulated hADSC and untreated hADSC control groups. The results of this study show that bioorthogonal dextran-based hydrogel is capable of supporting in vitro and in vivo chondrogenesis of chondrocytes and human stem cells, thus highlighting high feasibility of bioorthogonal hydrogels for clinical translation in stem cell-based cartilage regeneration.

Impact of TGF-β family-related growth factors on chondrogenic differentiation of adipose-derived stem cells isolated from lipoaspirates and infrapatellar fat pads of osteoarthritic patients.

The success of cell-based approaches for the treatment of cartilage defects requires an optimal autologous cell source with chondrogenic differentiation ability that maintains its differentiated properties and stability following implantation. The objective of this study was to compare the chondrogenic capacity of mesenchymal stem cells (MSCs) isolated from lipoaspirates (ASCs) and infrapatellar fat pads (IFPSCs) of osteoarthritic patients and treated with transforming growth factor (TGF)-β family-related growth factors. Cells were cultured for 6 weeks in a 3D pellet culture system with the chimeric activin A/bone morphogenic protein (BMP)-2 ligand (AB235), the chimeric nodal/BMP-2 ligand (NB260) or BMP-2. To investigate the stability of the new cartilage, ASCs-treated pellets were transplanted subcutaneously into severe combined immunodeficiency (SCID) mice. Histological and immunohistochemical assessment confirmed that the growth factors induced cartilage differentiation in both isolated cell types. However, reverse transcription-quantitative PCR results showed that ASCs presented a higher chondrogenic potential than IFPSCs. In vivo results revealed that AB235-treated ASCs pellets were larger in size and could form stable cartilage-like tissue as compared to NB260-treated pellets, while BMP-2-treated pellets underwent calcification. The chondrogenic induction of ASCs by AB235 treatment was mediated by SMAD2/3 activation, as proved by immunofluorescence analysis. The results of this study indicated that the combination of ASCs and AB235 might lead to a cell-based cartilage regeneration treatment.

Link Protein N-Terminal Peptide as a Potential Stimulating Factor for Stem Cell-Based Cartilage Regeneration.

Background Link protein N-terminal peptide (LPP) in extracellular matrix (ECM) of cartilage could induce synthesis of proteoglycans and collagen type II in cartilaginous cells. Cartilage stem/progenitor cells (CSPCs), the endogenous stem cells in cartilage, are important in cartilage degeneration and regeneration. We hypothesized that LPP could be a stimulator for stem cell-based cartilage regeneration by affecting biological behaviors of CSPC. Methods CSPCs were isolated from rat knee cartilage. We evaluated the promoting effect of LPP on proliferation, migration, and chondrogenic differentiation of CSPCs. The chondrogenic differentiation-related genes and proteins were quantitated. Three-dimensional culture of CSPC was conducted in the presence of TGF-β3 or LPP, and the harvested pellets were analyzed to assess the function of LPP on cartilage regeneration. Results LPP stimulated the proliferation of CSPC and accelerated the site-directional migration. Higher expression of SOX9, collagen II, and aggrecan were demonstrated in CSPCs treated with LPP. The pellets treated with LPP showed more distinct characteristics of chondroid differentiation than those with TGF-β3. Conclusion LPP showed application prospect in cartilage regeneration medicine by stimulating proliferation, migration, and chondrogenic differentiation of cartilage stem/progenitor cells.

Pellet coculture of osteoarthritic chondrocytes and infrapatellar fat pad-derived mesenchymal stem cells with chitosan/hyaluronic acid nanoparticles promotes chondrogenic differentiation

BackgroundCell source plays a key role in cell-based cartilage repair and regeneration. Recent efforts in cell coculture have attempted to combine the advantages and negate the drawbacks of the constituent cell types. The aim of this study was to evaluate the chondrogenic outcome of articular chondrocytes (ACs) and infrapatellar fat pad (IPFP)-derived mesenchymal stem cells (MSCs) in direct coculture.MethodsACs and IPFP MSCs from the same patients with knee osteoarthritis (OA) were cocultured in monolayer and in pellets. The monocultures of each cell type were also used as controls. Morphological and histologic analysis, immunofluorescence staining, reverse transcription-polymerase chain reaction, and enzyme-linked immunosorbent assay were performed to characterize the chondrogenic differentiation of cocultures. Furthermore, the effects of chitosan/hyaluronic acid (CS/HA) nanoparticle exposure on the chondrogenesis of cocultures were examined.ResultsIn both monolayer and pellet coculture, the hypertrophy of MSCs and the inflammatory activities of ACs were inhibited, although the chondrogenic production in coculture was not promoted compared with that in monoculture. In addition, the exposure of CS/HA nanoparticles to pellet coculture improved the production of type II collagen and aggrecan.ConclusionsWe demonstrate for the first time that pellet coculture of ACs and IPFP MSCs with CS/HA nanoparticles could promote chondrogenic outcome while preventing the inflammatory status of ACs and the hypertrophic differentiation of MSCs. These findings suggest that the combination of ACs, IPFP MSCs, and CS/HA might be useful in cartilage repair in knee OA.

Synergistic effects on mesenchymal stem cell-based cartilage regeneration by chondrogenic preconditioning and mechanical stimulation

BackgroundMesenchymal stem cells (MSCs) hold promising translational potential in cartilage regeneration. However, the efficacy of MSC-based tissue engineering is not satisfactory in the treatment of cartilage defect because of the inevitable cellular functional changes during ex vivo cell expansion. How to maintain the chondrogenic capacity of MSCs to improve their therapeutic outcomes remains an outstanding question.MethodsBone marrow-derived MSCs were firstly primed in chondrogenic induction medium which was then replaced with normal growth medium to attain the manipulated cells (M-MSCs). Methacrylated hyaluronic acid (MeHA) was synthesized as a scaffold to encapsulate the cells. The MSC- or M-MSC-laden constructs were treated with dynamic compressive loading (DL) in a bioreactor or with free loading (FL) for 14 days. Afterwards, the constructs were implanted in nude mice or rat models of osteochondral defects to test their efficiency in cartilage regeneration or repair.ResultsData showed that the resulting M-MSCs exhibited superior chondrogenic differentiation potential and survivability compared with untreated MSCs. More importantly, we found that DL significantly promoted neocartilage formation in the MeHA hydrogel encapsulated with M-MSCs after 30 days of implantation in nude mice. Furthermore, the constructs laden with M-MSCs after DL for 14 days significantly enhanced cartilage healing in a rat model of osteochondral defect.ConclusionsFindings from this study highlight the importance of maintaining chondrogenic potential of MSCs by in-vitro chondrogenic preconditioning and a synergistic effect of mechanical stimulation in cartilage engineering, which may shed light on the stem cell-based tissue engineering for cartilage repair.

Featured Article: Invitro development of personalized cartilage microtissues uncovers an individualized differentiation capacity of human chondrocytes.

Personalized features in the treatment of knee injuries and articular replacement therapies play an important role in modern life with increasing demand. Therefore, cell-based therapeutic approaches for the regeneration of traumatic defects of cartilage tissue were developed. However, great variations in the quality of repair tissue or therapeutic outcome were observed. The aim of the study was to capture and visualize individual differentiation capacities of chondrocytes derived from different donors with regard to a possible personal regeneration capacity using a cell-based therapy. The redifferentiation potential of monolayer cultured cells was analyzed in a scaffold-free three-dimensional tissue model. Furthermore, stimulating options using cartilage maturation factors such as L-ascorbic acid and transforming growth factor beta 2 (TGF-β2) on this process were of special interest. Cells and tissues were analyzed via histological and immunohistochemical methods. Gene expression was measured by quantitative real-time polymerase chain reaction. In monolayer culture, cells from all donors showed an almost identical differentiation profile. In contrast, the differentiation state of cartilage-like three-dimensional microtissues revealed clear differences with respect to individual donors. Analyses at the protein and mRNA levels showed high variations regarding cartilage-typical matrix components (e.g. proteoglycans, collagen type II) and intracellular proteins (e.g. S100). Interestingly, only donor chondrocytes with a basic tendency to re-differentiate in a three-dimensional environment were able to increase this tissue-specific maturation when exposed to L-ascorbic acid and/or TGF-β2. Our approach revealed clear-cut possibilities for classification of individual donors into responders or non-responders. On the basis of these results an invitro platform could be designed to discriminate responders from non-responders. This invitro three-dimensional test system may be a suitable basis to establish a "personalized diagnostic tool" with the opportunity to assess the capacity of expanded chondrocytes to respond to an autologous cell-based therapy. Impact statement A challenge in cell-based cartilage regeneration therapies is the identification of a "personalized diagnostic tool" to predict the chondrogenic potency of cells from patients who are going to be treated with autologous cells. Comparing the phenotype of isolated chondrocytes from different donors invitro revealed an individual cartilage-specific differentiation capacity. These personalized features are not detectable invitro until the monolayer cells have the possibility to rearrange in 3D tissues. Cells from articular cartilage in monolayer culture may not be a suitable basis to discriminate responders from non-responders with respect to a personalized cell-based therapy to treat cartilage defects. A more physiological 3D (micro-)environment enable the cells to present their individual differentiation capacity. The here described microtissue model might be the basis for an invitro platform to predict the therapeutic outcome of autologous cell-based cartilage repair and/or a suitable tool to identify early biomarkers to classify the patients.

The role of laminins in cartilaginous tissues: from development to regeneration.

As a key molecule of the extracellular matrix, laminin provides a delicate microenvironment for cell functions. Recent findings suggest that laminins expressed by cartilage-forming cells (chondrocytes, progenitor cells and stem cells) could promote chondrogenesis. However, few papers outline the effect of laminins on providing a favorable matrix microenvironment for cartilage regeneration. In this review, we delineated the expression of laminins in hyaline cartilage, fibrocartilage and cartilage-like tissue (nucleus pulposus) throughout several developmental stages. We also examined the effect of laminins on the biological activities of chondrocytes, including adhesion, migration and survival. Furthermore, we scrutinized the potential influence of various laminin isoforms on cartilage-forming cells' proliferation and chondrogenic differentiation. With this information, we hope to facilitate the understanding of the spatial and temporal interactions between cartilage-forming cells and laminin microenvironment to eventually advance cell-based cartilage engineering and regeneration.