Treatment Effects of Intra-Articular Allogenic Mesenchymal Stem Cell Secretome in an Equine Model of Joint Inflammation.

Background: Allogenic mesenchymal stem cell (MSC) secretome is a novel intra-articular therapeutic that has shown promise in in vitro and small animal models and warrants further investigation. Objectives: To investigate if intra-articular allogenic MSC-secretome has anti-inflammatory effects using an equine model of joint inflammation. Study Design: Randomized positively and negatively controlled experimental study. Method: In phase 1, joint inflammation was induced bilaterally in radiocarpal joints of eight horses by injecting 0.25 ng lipopolysaccharide (LPS). After 2 h, the secretome of INFy and TNFα stimulated allogeneic equine MSCs was injected in one randomly assigned joint, while the contralateral joint was injected with medium (negative control). Clinical parameters (composite welfare scores, joint effusion, joint circumference) were recorded, and synovial fluid samples were analyzed for biomarkers (total protein, WBCC; eicosanoid mediators, CCL2; TNFα; MMP; GAGs; C2C; CPII) at fixed post-injection hours (PIH 0, 8, 24, 72, and 168 h). The effects of time and treatment on clinical and synovial fluid parameters and the presence of time-treatment interactions were evaluated. For phase 2, allogeneic MSC-secretome vs. allogeneic equine MSCs (positive control) was tested using a similar methodology. Results: In phase 1, the joint circumference was significantly (p < 0.05) lower in the MSC-secretome treated group compared to the medium control group at PIH 24, and significantly higher peak synovial GAG values were noted at PIH 24 (p < 0.001). In phase 2, no significant differences were noted between the treatment effects of MSC-secretome and MSCs. Main Limitations: This study is a controlled experimental study and therefore cannot fully reflect natural joint disease. In phase 2, two therapeutics are directly compared and there is no negative control. Conclusions: In this model of joint inflammation, intra-articular MSC-secretome injection had some clinical anti-inflammatory effects. An effect on cartilage metabolism, evident as a rise in GAG levels was also noted, although it is unclear whether this could be considered a beneficial or detrimental effect. When directly comparing MSC-secretome to MSCs in this model results were comparable, indicating that MSC-secretome could be a viable off-the-shelf alternative to MSC treatment.

Focal articular surface replacement as primary treatment for focal chondral defects of the femoral condyles: A series of 157 cases.

BACKGROUND: Focal chondral defects (FCDs) of the femoral condyle are common. Treatment has heretofore primarily consisted of non-surgical and biological treatments. Focal articular surface replacement (FASR) is an emerging technique utilizing small implants to essentially fill the FCD. Here we report functional outcome and re-operation rates following FASR as a primary treatment for FCDs of the femoral condyles. METHODS: Retrospective analysis of a prospectively collected database including 327 FASR procedures was performed to identify patients who underwent FASR of the femoral condyle with a modular cementless metallic implant (HemiCAPTM) as a primary procedure. Knee Injury and Osteoarthritis Outcome Score (KOOS), Oxford Knee Score (OKS), SF-36 Health Status Survey (SF-36) and Visual Analog Scale (VAS) were collected before and 6 weeks, 6 months, and 4 years after surgery. Implant revision and re-operation rate were recorded. RESULTS: 157 patients were included with a mean follow-up of 9.4 ± 1.3 years (range 7.0 to 11.4 years). The average age was 40.2 ± 5.3 years, 85% involved the medial condyle, and the average defect size was 3.6 ± 0.5 cm2. Primary FASR resulted in functional improvement on the KOOS (+52%), OKS (+69%) and SF-36 (+50%) scores and a reduction in VAS scores (-70%) at 4-year follow-up. Revision rate was 0.64% and the re-operation rate was 11%. CONCLUSION: This retrospective case-series supports primary FASR with HemiCAPTM implants as an alternative to biological procedures to treat medium-sized FCDs (2.5-4 cm2) of the femoral condyle, although long-term follow-up is necessary to determine if the clinical outcome and low revision rate can be maintained.

[Swelling of the thigh after a scooter accident]. / Zwelling van het bovenbeen na een scooterongeval.

A 44-year-old man presented with a swelling of his right thigh three weeks after a fall of his scooter, in which he had experienced a sheering trauma on the tarmac. The painless swelling had increased gradually to a size of 20 x 10 cm. Ultrasonography showed a fluid filled separation of the subcutis and fascia. The diagnosis was a Morel-Lavallee lesion.

Focal articular surface replacement of knee lesions after failed cartilage repair using focal metallic implants: A series of 132 cases with 4-year follow-up.

BACKGROUND: Focal articular lesions of the knee can be treated using several different techniques with generally good results, but failures are difficult to manage. Focal articular surface replacement (FASR) using metal implants could be a promising technique that allows defect geometry matching, congruency restoration and defect propagation prevention. METHODS: 132 patients were included who underwent FASR between January 2009 and December 2013. Three different implants were used: 1. HemiCAP®; 2. UniCAP® and 3. HemiCAP® PF Classic for trochlear lesions. Primary outcome parameter was knee function assessed by Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) score with a 4-year follow-up, secondary outcomes included survivorship and complications. Statistical analyses were performed using GraphPad Prism. RESULTS: For all 132 surgeries combined (102 HemiCAP®, 11 UniCAP® and 19 HemiCAP® PF Classic implants), WOMAC scores significantly improved from 6 weeks onward until the end of the study (p < 0.001 for all time points). 4-year survival rate was 97.7%, and a re-operation rate of 12.1% was found. The HemiCAP® group revealed a slower WOMAC improvement in patients aged ≥40 years, combined with a trend towards lower final WOMAC scores and a higher re-operation rate in patients with a BMI ≥ 25. CONCLUSIONS: This report shows good to excellent clinical results of FASR as a salvage procedure after failed cartilage repair, with a low re-operation rate and a high survival of 97.7% at 4-year follow-up. Although longer follow-up is required, this could be a valuable treatment option in these challenging cases, without limiting future options for surgical interventions when deemed necessary.

Does resurfacing of asymptomatic full-thickness localized articular defects of the trochlea influence the outcome following unicompartmental knee arthroplasty of the medial compartment?: A retrospective cohort study with minimum seven-year follow-up.

BACKGROUND: Patellofemoral joint (PFJ) degeneration has been found not to affect outcome following medial unicompartmental knee arthroplasty (UKA). However, PFJ disease occasionally presents as isolated trochlear cartilage lesions, with little available evidence regarding treatment options or necessity. We evaluated the effectiveness of concomitant trochlear resurfacing in patients undergoing medial UKA with asymptomatic trochlear lesions. METHODS: We included 60 patients undergoing medial UKA with an associated full thickness lesion of the trochlea. A fixed bearing UKA implant (PKR™, Stryker, Warsaw, IND) was used in all cases. In 30 patients, trochlear lesions were resurfaced with a HemiCAP® PF Classic (HemiCAP® PFC) implant (Arthrosurface, Franklin, MA). Outcome measures included VAS-, KOOS-, WOMAC-, SF-36 scores and radiological assessment. RESULTS: Average length of follow-up was 97.4 months (range: 88-106 months). Both groups showed significant improvement for all clinical scores post-operatively compared to pre-operatively (p < 0.001 for all). The UKA group showed a better VAS score at all follow-up moments (p < 0.01 for all), but no differences were found between both groups at all time points for other outcome measures. None of the HemiCAP® PFC implants needed to be revised within the timeframe of the study. CONCLUSIONS: In this retrospective cohort study, we found a 100% survivorship of the HemiCAP® PFC implant at an average eight-year follow-up. However, no clinical benefits were found in performing trochlear resurfacing in conjunction with medial UKA for asymptomatic end-stage trochlear cartilage lesions. Therefore, these lesions can be safely ignored when performing a medial UKA.

Intra-articular Injections of Platelet-Rich Plasma Releasate Reduce Pain and Synovial Inflammation in a Mouse Model of Osteoarthritis.

BACKGROUND: Osteoarthritis (OA) is a degenerative joint disease leading to pain and disability for which no curative treatment exists. A promising biological treatment for OA is intra-articular administration of platelet-rich plasma (PRP). PRP injections in OA joints can relieve pain, although the exact working mechanism is unclear. PURPOSE: To examine the effects of PRP releasate (PRPr) on pain, cartilage damage, and synovial inflammation in a mouse OA model. STUDY DESIGN: Controlled laboratory study. METHODS: OA was induced unilaterally in the knees of male mice (n = 36) by 2 intra-articular injections of collagenase at days -7 and -5. At day 0, pain was measured by registering weight distribution on the hindlimbs, after which mice were randomly divided into 2 groups. Mice received 3 intra-articular injections of PRP or saline in the affected knee. Seven mice per group were euthanized at day 5 for assessment of early synovial inflammation and cartilage damage. Pain in the remaining mice was registered for a total of 3 weeks. These mice were euthanized at day 21 for assessment of cartilage damage and synovial inflammation on histological evaluation. Antibodies against iNOS, CD163, and CD206 were used to identify different subtypes of macrophages in the synovial membrane. RESULTS: Mice in the PRPr group increased the distribution of weight on the affected joint in 2 consecutive weeks after the start of the treatment ( P < .05), whereas mice in the saline group did not. At day 21, PRPr-injected knees had a thinner synovial membrane ( P < .05) and a trend toward less cartilage damage in the lateral joint compartment ( P = .053) than saline-injected knees. OA knees treated with saline showed less anti-inflammatory (CD206+ and CD163+) cells at day 5 than healthy knees, an observation that was not made in the PRPr-treated group. A higher level of pain at day 7 was associated with a thicker synovial membrane at day 21. The presence of CD206+ cells was negatively associated with synovial membrane thickness. CONCLUSION: In a murine OA model, multiple PRPr injections reduced pain and synovial thickness, possibly through modulation of macrophage subtypes. CLINICAL RELEVANCE: PRPr injections in early OA or shortly after joint trauma can reduce pain and synovial inflammation and may inhibit OA development in patients.

Pre-Treatment of Human Mesenchymal Stem Cells With Inflammatory Factors or Hypoxia Does Not Influence Migration to Osteoarthritic Cartilage and Synovium.

BACKGROUND: Mesenchymal stem cells (MSCs) are promising candidates as a cell-based therapy for osteoarthritis (OA), although current results are modest. Pre-treatment of MSCs before application might improve their therapeutic efficacy. HYPOTHESIS: Pre-treatment of MSCs with inflammatory factors or hypoxia will improve their migration and adhesion capacities toward OA-affected tissues. STUDY DESIGN: Controlled laboratory study. METHODS: We used real-time polymerase chain reaction to determine the effects of different fetal calf serum (FCS) batches, platelet lysate (PL), hypoxia, inflammatory factors, factors secreted by OA tissues, and OA synovial fluid (SF) on the expression of 12 genes encoding chemokine or adhesion receptors. Migration of MSCs toward factors secreted by OA tissues was studied in vitro, and attachment of injected MSCs was evaluated in vivo in healthy and OA knees of male Wistar rats. RESULTS: Different FCS batches, PL, or hypoxia did not influence the expression of the migration and adhesion receptor genes. Exposure to inflammatory factors altered the expression of CCR1, CCR4, CD44, PDGFRα, and PDGFRß. MSCs migrated toward factors secreted by OA tissues in vitro. Neither pre-treatment with inflammatory factors nor the presence of OA influenced MSC migration in vitro or adhesion in vivo. CONCLUSION: Factors secreted by OA tissues increase MSC migration in vitro. In vivo, no difference in MSC adhesion was found between OA and healthy knees. Pre-treatment with inflammatory factors influenced the expression of migration and adhesion receptors of MSCs but not their migration in vitro or adhesion in vivo. CLINICAL RELEVANCE: To improve the therapeutic capacity of intra-articular injection of MSCs, they need to remain intra-articular for a longer period of time. Pre-treatment of MSCs with hypoxia or inflammatory factors did not increase the migration or adhesion capacity of MSCs and will therefore not likely prolong their intra-articular longevity. Alternative approaches to prolong the intra-articular presence of MSCs should be developed to increase the therapeutic effect of MSCs in OA.

Mesenchymal stem cells reduce pain but not degenerative changes in a mono-iodoacetate rat model of osteoarthritis.

We studied the effects of intra-articularly injected bone marrow derived mesenchymal stem cells (MSCs), as well as freshly isolated bone marrow mononuclear cells (BMMNCs), on pain, cartilage damage, bone changes and inflammation in an in-vivo rat osteoarthritis (OA) model. OA was induced unilaterally by injection of mono-iodoacetate (MIA) and allowed to develop for 3 weeks. Then, animals were treated by intra-articular injection with MSCs, BMMNCs, or saline as a control. Four weeks later, pain was assessed with an incapitance tester, subchondral bone alterations were measured with µCT and cartilage quality and joint inflammation were assessed by histological analysis. Animals treated with MSCs distributed significantly more weight to the affected limb after treatment, which was not observed in the other groups. No statistically significant differences between treatment groups regarding cartilage damage, subchondral bone alterations and synovial inflammation were observed. Additional cell tracking experiments indicated adequate intra-articular cell injection and cell survival up to 2 weeks. In our OA model, injected MSCs were able to reduce MIA induced pain, as measured by an increased weight distribution to the affected limb. No statistically significant effects of the cellular therapies on structural damage and synovial inflammation were found.

Delayed gadolinium-enhanced MRI of cartilage (dGEMRIC) shows no change in cartilage structural composition after viscosupplementation in patients with early-stage knee osteoarthritis.

INTRODUCTION: Viscosupplementation with hyaluronic acid (HA) of osteoarthritic (OA) knee joints has a well-established positive effect on clinical symptoms. This effect, however, is only temporary and the working mechanism of HA injections is not clear. It was suggested that HA might have disease modifying properties because of its beneficial effect on cartilage sulphated glycosaminoglycan (sGAG) content. Delayed gadolinium-enhanced MRI of cartilage (dGEMRIC) is a highly reproducible, non-invasive surrogate measure for sGAG content and hence composition of cartilage. The aim of this study was to assess whether improvement in cartilage structural composition is detected using dGEMRIC 14 weeks after 3 weekly injections with HA in patients with early-stage knee OA. METHODS: In 20 early-stage knee OA patients (KLG I-II), 3D dGEMRIC at 3T was acquired before and 14 weeks after 3 weekly injections with HA. To evaluate patient symptoms, the knee injury and osteoarthritis outcome score (KOOS) and a numeric rating scale (NRS) for pain were recorded. To evaluate cartilage composition, six cartilage regions in the knee were analyzed on dGEMRIC. Outcomes of dGEMRIC, KOOS and NRS before and after HA were compared using paired t-testing. Since we performed multiple t-tests, we applied a Bonferroni-Holm correction to determine statistical significance for these analyses. RESULTS: All KOOS subscales ('pain', 'symptoms', 'daily activities', 'sports' and 'quality of life') and the NRS pain improved significantly 14 weeks after Viscosupplementation with HA. Outcomes of dGEMRIC did not change significantly after HA compared to baseline in any of the cartilage regions analyzed in the knee. CONCLUSIONS: Our results confirm previous findings reported in the literature, showing persisting improvement in symptomatic outcome measures in early-stage knee OA patients 14 weeks after Viscosupplementation. Outcomes of dGEMRIC, however, did not change after Viscosupplementation, indicating no change in cartilage structural composition as an explanation for the improvement of clinical symptoms.

Effect of Arthritic Synovial Fluids on the Expression of Immunomodulatory Factors by Mesenchymal Stem Cells: An Explorative in vitro Study.

BACKGROUND: In diseased joints, the catabolic environment results in progressive joint damage. Mesenchymal stem cells (MSCs) can have immunomodulatory effects by secreting anti-inflammatory factors. To exert these effects, MSCs need to be triggered by pro-inflammatory cytokines. To explore the potential of MSCs as a treatment for diseased joints, we studied the effect of synovial fluid (SF) from donors with different joint diseases and donors without joint pathology on the immunomodulatory capacities of human MSCs in vitro. We hypothesized that SF of diseased joints influences the immunomodulatory effects of MSCs. MATERIALS AND METHODS: MSCs were cultured in medium with SF of six osteoarthritis (OA) or six rheumatoid arthritis (RA) donors and three donors without joint pathology were used as control. Gene expressions of IL-6, HGF, TNFa, TGFb1, and indoleamine 2,3-dioxygenase (IDO) were analyzed. l-kynurenine concentration in conditioned medium (CM) by MSCs with SF was determined as a measure of IDO activity by MSCs. Furthermore, the effect of CM with SF on proliferation of activated lymphocytes was analyzed. RESULTS: Addition of SF significantly up-regulated the mRNA expression of IL-6 and IDO in MSCs. SF(OA) induced significantly higher expression of IDO than SF(control), although no difference in IDO activity of the MSCs could be shown with a l-kynurenine assay. Medium conditioned by MSCs with SF(OA or RA) suppressed activated lymphocyte proliferation in vitro more than medium conditioned by MSCs without SF or with SF(control). DISCUSSION: SF can influence the expression of genes involved in immunomodulation by MSCs and the effect on lymphocyte proliferation. We found indications for disease-specific differences between SFs but the variation between donors, even within one disease group was high. These data warrant further research to examine the potential application of MSC therapy in arthritic joints.

Platelet-rich plasma releasate inhibits inflammatory processes in osteoarthritic chondrocytes.

BACKGROUND: Platelet-rich plasma (PRP) has recently been postulated as a treatment for osteoarthritis (OA). Although anabolic effects of PRP on chondrocytes are well documented, no reports are known addressing effects on cartilage degeneration. Since OA is characterized by a catabolic and inflammatory joint environment, the authors investigated whether PRP was able to counteract the effects of such an environment on human osteoarthritic chondrocytes. HYPOTHESIS: Platelet-rich plasma inhibits inflammatory effects of interleukin-1 (IL-1) beta on human osteoarthritic chondrocytes. STUDY DESIGN: Controlled laboratory study. METHODS: Human osteoarthritic chondrocytes were cultured in the presence of IL-1 beta to mimic an osteoarthritic environment. Medium was supplemented with 0%, 1%, or 10% PRP releasate (PRPr, the active releasate of PRP). After 48 hours, gene expression of collagen type II alpha 1 (COL2A1), aggrecan (ACAN), a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)4, ADAMTS5, matrix metalloproteinase (MMP)13, and prostaglandin-endoperoxide synthase (PTGS)2 was analyzed. Additionally, glycosaminoglycan (GAG) content, nitric oxide (NO) production, and nuclear factor kappa B (NFκB) activation were studied. RESULTS: Platelet-rich plasma releasate diminished IL-1 beta-induced inhibition of COL2A1 and ACAN gene expression. The PRPr also reduced IL-1 beta-induced increase of ADAMTS4 and PTGS2 gene expression. ADAMTS5 gene expression and GAG content were not influenced by IL-1 beta or additional PRPr. Matrix metalloproteinase 13 gene expression and NO production were upregulated by IL-1 beta but not affected by added PRPr. Finally, PRPr reduced IL-1 beta-induced NFκB activation to control levels containing no IL-1 beta. CONCLUSION: Platelet-rich plasma releasate diminished multiple inflammatory IL-1 beta-mediated effects on human osteoarthritic chondrocytes, including inhibition of NFκB activation. CLINICAL RELEVANCE: Platelet-rich plasma releasate counteracts effects of an inflammatory environment on genes regulating matrix degradation and formation in human chondrocytes. Platelet-rich plasma releasate decreases NFκB activation, a major pathway involved in the pathogenesis of OA. These results encourage further study of PRP as a treatment for OA.

Clinically translatable cell tracking and quantification by MRI in cartilage repair using superparamagnetic iron oxides.

BACKGROUND: Articular cartilage has very limited intrinsic regenerative capacity, making cell-based therapy a tempting approach for cartilage repair. Cell tracking can be a major step towards unraveling and improving the repair process of these therapies. We studied superparamagnetic iron oxides (SPIO) for labeling human bone marrow-derived mesenchymal stem cells (hBMSCs) regarding effectivity, cell viability, long term metabolic cell activity, chondrogenic differentiation and hBMSC secretion profile. We additionally examined the capacity of synovial cells to endocytose SPIO from dead, labeled cells, together with the use of magnetic resonance imaging (MRI) for intra-articular visualization and quantification of SPIO labeled cells. METHODOLOGY/PRINICIPAL FINDINGS: Efficacy and various safety aspects of SPIO cell labeling were determined using appropriate assays. Synovial SPIO re-uptake was investigated in vitro by co-labeling cells with SPIO and green fluorescent protein (GFP). MRI experiments were performed on a clinical 3.0T MRI scanner. Two cell-based cartilage repair techniques were mimicked for evaluating MRI traceability of labeled cells: intra-articular cell injection and cell implantation in cartilage defects. Cells were applied ex vivo or in vitro in an intra-articular environment and immediately scanned. SPIO labeling was effective and did not impair any of the studied safety aspects, including hBMSC secretion profile. SPIO from dead, labeled cells could be taken up by synovial cells. Both injected and implanted SPIO-labeled cells could accurately be visualized by MRI in a clinically relevant sized joint model using clinically applied cell doses. Finally, we quantified the amount of labeled cells seeded in cartilage defects using MR-based relaxometry. CONCLUSIONS: SPIO labeling appears to be safe without influencing cell behavior. SPIO labeled cells can be visualized in an intra-articular environment and quantified when seeded in cartilage defects.