Association between expression of the Bone morphogenetic proteins 2 and 7 in the repair of circumscribed cartilage lesions with clinical outcome

<p>Abstract</p> <p>Background</p> <p>Although there is much known about the role of BMPs in cartilage metabolism reliable data about the <it>in vivo </it>regulation in natural and surgically induced cartilage repair are still missing.</p> <p>Methods</p> <p>Lavage fluids of knee joints of 47 patients were collected during surgical therapy. 5 patients had no cartilage lesion and served as a control group, the other 42 patients with circumscribed cartilage defects were treated by microfracturing (19) or by an Autologous Chondrocyte Implantation (23). The concentrations of BMP-2 and BMP-7 were determined by ELISA. The clinical status was evaluated using the IKDC Score prior to and 1 year following the operation.</p> <p>Results</p> <p>High level expression in the control group was found for BMP-2, concentrations of BMP-7 remained below detection levels. No statistical differences could be detected in concentrations of BMP-2 or BMP-7 in the lavage fluids of knees with cartilage lesions compared to the control group. Levels of BMP-7 did not change after surgical cartilage repair, whereas concentrations of BMP-2 statistically significant increased after the intervention (p < 0.001). The clinical outcome following cartilage regenerating surgery increased after 1 year by 29% (p < 0.001). The difference of the IKDC score after 1 year and prior to the operation was used to quantify the degree of improvement following surgery. This difference statistically significant correlated with initial BMP-2 (R = 0.554, p < 0.001) but not BMP-7 (R = 0.031, n.s.) levels in the knee joints.</p> <p>Conclusions</p> <p>BMP-2 seems to play an important role in surgically induced cartilage repair; synovial expression correlates with the clinical outcome.</p

Derivation of Chondrogenically-Committed Cells from Human Embryonic Cells for Cartilage Tissue Regeneration

Background: Heterogeneous and uncontrolled differentiation of human embryonic stem cells (hESCs) in embryoid bodies (EBs) limits the potential use of hESCs for cell-based therapies. More efficient strategies are needed for the commitment and differentiation of hESCs to produce a homogeneous population of specific cell types for tissue regeneration applications. Methodology/Principal Findings: We report here that significant chondrocytic commitment of feeder-free cultured human embryonic stem cells (FF-hESCs), as determined by gene expression and immunostaining analysis, was induced by coculture with primary chondrocytes. Furthermore, a dynamic expression profile of chondrocyte-specific genes was observed during monolayer expansion of the chondrogenically-committed cells. Chondrogenically-committed cells synergistically responded to transforming growth factor-b1 (TGF-b1) and b1-integrin activating antibody by increasing tissue mass in pellet culture. In addition, when encapsulated in hydrogels, these cells formed cartilage tissue both in vitro and in vivo. In contrast, the absence of chondrocyte co-culture did not result in an expandable cell population from FF-hESCs. Conclusions/Significance: The direct chondrocytic commitment of FF-hESCs can be induced by morphogenetic factor

Autologous chondrocyte implantation-derived synovial fluids display distinct responder and non-responder proteomic profiles

Hulme, Charlotte H. & Wilson, Emma L. - Equal contributorsBackground Autologous chondrocyte implantation (ACI) can be used in the treatment of focal cartilage injuries to prevent the onset of osteoarthritis (OA). However, we are yet to understand fully why some individuals do not respond well to this intervention. Identification of a reliable and accurate biomarker panel that can predict which patients are likely to respond well to ACI is needed in order to assign the patient to the most appropriate therapy. This study aimed to compare the baseline and mid-treatment proteomic profiles of synovial fluids (SFs) obtained from responders and non-responders to ACI. Methods SFs were derived from 14 ACI responders (mean Lysholm improvement of 33 (17–54)) and 13 non-responders (mean Lysholm decrease of 14 (4–46)) at the two stages of surgery (cartilage harvest and chondrocyte implantation). Label-free proteome profiling of dynamically compressed SFs was used to identify predictive markers of ACI success or failure and to investigate the biological pathways involved in the clinical response to ACI. Results Only 1 protein displayed a ≥2.0-fold differential abundance in the preclinical SF of ACI responders versus non-responders. However, there is a marked difference between these two groups with regard to their proteome shift in response to cartilage harvest, with 24 and 92 proteins showing ≥2.0-fold differential abundance between Stages I and II in responders and non-responders, respectively. Proteomic data has been uploaded to ProteomeXchange (identifier: PXD005220). We have validated two biologically relevant protein changes associated with this response, demonstrating that matrix metalloproteinase 1 was prominently elevated and S100 calcium binding protein A13 was reduced in response to cartilage harvest in non-responders. Conclusions The differential proteomic response to cartilage harvest noted in responders versus non-responders is completely novel. Our analyses suggest several pathways which appear to be altered in non-responders that are worthy of further investigation to elucidate the mechanisms of ACI failure. These protein changes highlight many putative biomarkers that may have potential for prediction of ACI treatment success

Two independent proteomic approaches provide a comprehensive analysis of the synovial fluid proteome response to Autologous Chondrocyte Implantation

Background: Autologous chondrocyte implantation (ACI) has a failure rate of approximately 20%, but it is yet to be fully understood why. Biomarkers are needed that can pre-operatively predict in which patients it is likely to fail, so that alternative or individualised therapies can be offered. We previously used label-free quantitation (LF) with a dynamic range compression proteomic approach to assess the synovial fluid (SF) of ACI responders and non-responders. However, we were able to identify only a few differentially abundant proteins at baseline. In the present study, we built upon these previous findings by assessing higher-abundance proteins within this SF, providing a more global proteomic analysis on the basis of which more of the biology underlying ACI success or failure can be understood. Methods: Isobaric tagging for relative and absolute quantitation (iTRAQ) proteomic analysis was used to assess SF from ACI responders (mean Lysholm improvement of 33; n = 14) and non-responders (mean Lysholm decrease of 14; n = 13) at the two stages of surgery (cartilage harvest and chondrocyte implantation). Differentially abundant proteins in iTRAQ and combined iTRAQ and LF datasets were investigated using pathway and network analyses. Results: iTRAQ proteomic analysis confirmed our previous finding that there is a marked proteomic shift in response to cartilage harvest (70 and 54 proteins demonstrating ≥ 2.0-fold change and p < 0.05 between stages I and II in responders and non-responders, respectively). Further, it highlighted 28 proteins that were differentially abundant between responders and non-responders to ACI, which were not found in the LF study, 16 of which were altered at baseline. The differential expression of two proteins (complement C1s subcomponent and matrix metalloproteinase 3) was confirmed biochemically. Combination of the iTRAQ and LF proteomic datasets generated in-depth SF proteome information that was used to generate interactome networks representing ACI success or failure. Functional pathways that are dysregulated in ACI non-responders were identified, including acute-phase response signalling. Conclusions: Several candidate biomarkers for baseline prediction of ACI outcome were identified. A holistic overview of the SF proteome in responders and non-responders to ACI  has been profiled, providing a better understanding of the biological pathways underlying clinical outcome, particularly the differential response to cartilage harvest in non-responders

The subchondral bone in articular cartilage repair: current problems in the surgical management

As the understanding of interactions between articular cartilage and subchondral bone continues to evolve, increased attention is being directed at treatment options for the entire osteochondral unit, rather than focusing on the articular surface only. It is becoming apparent that without support from an intact subchondral bed, any treatment of the surface chondral lesion is likely to fail. This article reviews issues affecting the entire osteochondral unit, such as subchondral changes after marrow-stimulation techniques and meniscectomy or large osteochondral defects created by prosthetic resurfacing techniques. Also discussed are surgical techniques designed to address these issues, including the use of osteochondral allografts, autologous bone grafting, next generation cell-based implants, as well as strategies after failed subchondral repair and problems specific to the ankle joint. Lastly, since this area remains in constant evolution, the requirements for prospective studies needed to evaluate these emerging technologies will be reviewed

Effect of transforming growth factor-β2 on biological regulation of multilayer primary chondrocyte culture

YesCytokines are extremely potent biomolecules that regulate cellular functions and play multiple roles in initiation and inhibition of disease. These highly specialised macromolecules are actively involved in control of cellular proliferation, apoptosis, cell migration and adhesion. This work, investigates the effect of transforming growth factor-beta2 (TGF-β2) on the biological regulation of chondrocyte and the repair of a created model wound on a multilayer culture system. Also the effect of this cytokine on cell length, proliferation, and cell adhesion has been investigated. Chondrocytes isolated from knee joint of rats and cultured at 4 layers. Each layer consisted of 2 × 105 cells/ml with and without TGF-β2. The expression of mRNA and protein levels of TGF-β receptors and Smad1, 3, 4, and 7 have been analysed by RT-PCR and western blot analysis. The effect of different supplementations in chondrocyte cell proliferation, cell length, adhesion, and wound repair was statistically analysed by One-way ANOVA test. Our results showed that the TGFβ2 regulates mRNA levels of its own receptors, and of Smad3 and Smad7. Also the TGF-β2 caused an increase in chondrocyte cell length, but decreased its proliferation rate and the wound healing process. TGF-β2 also decreased cell adhesion ability to the surface of the culture flask. Since, TGF-β2 increased the cell size, but showed negative effect on cell proliferation and adhesion of CHC, the effect of manipulated TGF-β2 with other growth factors and/or proteins needs to be investigated to finalize the utilization of this growth factor and design of scaffolding in treatment of different types of arthritis