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This phase I clinical trial evaluated the safety and clinical efficacy of adipose‐derived stromal cells (ASCs) in osteoarthritis. Eighteen patients with severe knee osteoarthritis were treated with a single intra‐articular injection of autologous ASCs at low (2 × 106 cells), medium (10 × 106), or high (50 × 106) doses (n = 6 each). After 6 months, no serious adverse events were reported, and patients treated with low‐dose ASCs significantly improved in pain and function. Osteoarthritis (OA) is the most widespread musculoskeletal disorder in adults. It leads to cartilage damage associated with subchondral bone changes and synovial inflammation, causing pain and disability. The present study aimed at evaluating the safety of a dose‐escalation protocol of intra‐articular injected adipose‐derived stromal cells (ASCs) in patients with knee OA, as well as clinical efficacy as secondary endpoint. A bicentric, uncontrolled, open phase I clinical trial was conducted in France and Germany with regulatory agency approval for ASC expansion procedure in both countries. From April 2012 to December 2013, 18 consecutive patients with symptomatic and severe knee OA were treated with a single intra‐articular injection of autologous ASCs. The study design consisted of three consecutive cohorts (six patients each) with dose escalation: low dose (2 × 106 cells), medium dose (10 × 106), and high dose (50 × 106). The primary outcome parameter was safety evaluated by recording adverse events throughout the trial, and secondary parameters were pain and function subscales of the Western Ontario and McMaster Universities Arthritis Index. After 6 months of follow‐up, the procedure was found to be safe, and no serious adverse events were reported. Four patients experienced transient knee joint pain and swelling after local injection. Interestingly, patients treated with low‐dose ASCs experienced significant improvements in pain levels and function compared with baseline. Our data suggest that the intra‐articular injection of ASCs is a safe therapeutic alternative to treat severe knee OA patients. A placebo‐controlled double‐blind phase IIb study is being initiated to assess clinical and structural efficacy. Significance Although this phase I study included a limited number of patients without a placebo arm, it showed that local injection of autologous adipose‐derived stem cells was safe and well tolerated in patients with knee osteoarthritis. This study also provides encouraging preliminary evidence of efficacy. Larger and controlled long‐term studies are now mandatory to confirm whether this new strategy of cell therapy can improve pain and induce structural benefit in osteoarthritis.

Advanced Therapy Medicinal Products (ATMP) provide promising treatment options particularly for unmet clinical needs, such as progressive and chronic diseases where currently no satisfying treatment exists. Especially from the ATMP subclass of Tissue Engineered Products (TEPs), only a few have yet been translated from an academic setting to clinic and beyond. A reason for low numbers of TEPs in current clinical trials and one main key hurdle for TEPs is the cost and labor-intensive manufacturing process. Manual production steps require experienced personnel, are challenging to standardize and to scale up. Automated manufacturing has the potential to overcome these challenges, toward an increasing cost-effectiveness. One major obstacle for automation is the control and risk prevention of cross contaminations, especially when handling parallel production lines of different patient material. These critical steps necessitate validated effective and efficient cleaning procedures in an automated system. In this perspective, possible technologies, concepts and solutions to existing ATMP manufacturing hurdles are discussed on the example of a late clinical phase II trial TEP. In compliance to Good Manufacturing Practice (GMP) guidelines, we propose a dual arm robot based isolator approach. Our novel concept enables complete process automation for adherent cell culture, and the translation of all manual process steps with standard laboratory equipment. Moreover, we discuss novel solutions for automated cleaning, without the need for human intervention. Consequently, our automation concept offers the unique chance to scale up production while becoming more cost-effective, which will ultimately increase TEP availability to a broader number of patients.

Type-VI collagen is an integral part of the extracellular cartilage matrix. However, the exact amounts of type-VI collagen in normal and osteoarthritic human cartilage still are not known. In this study, we describe an inhibition enzyme-linked immunosorbent assay that was developed to quantitate type-VI collagen epitopes found in guanidinium chloride extracts from normal and osteoarthritic human cartilage. In 31 cartilage samples from various localizations of healthy adult human knees, type-VI collagen epitopes accounted for approximately 0.40% of the total collagen content. Interestingly, type-VI collagen epitopes increased about 4-fold in osteoarthritic cartilage. A statistically significant increase of type-VI collagen epitopes was found during early stages of the disease, with only a superficial roughening of the cartilage surface and a loss of proteoglycans. Thus, these findings indicate that type-VI collagen is a minor component of normal human articular cartilage and that the amount of type-VI collagen epitopes increases significantly during early stages of osteoarthritis.

Generation of advanced glycation end products (AGE) is an inevitable process in vivo and can be accelerated under pathologic conditions such as oxidative stress, e.g. in rheumatoid arthritis (RA). This process is mediated by the AGE-specific receptor (RAGE). In this study we analysed the presence of RAGE in RA and osteoarthritic (OA) synovial tissue using immunohistology. Frozen synovial tissue samples from 11 RA patients and 12 OA patients were treated with goat anti-RAGE immunoglobulin G (IgG) and rabbit antigoat IgG. Immunostaining was visualised with streptavidin horse radish peroxidase (chromogen amino-ethyl-carbazole). Cell differentiation was performed with antibodies against CD68, CD45RO, and CD20. In 9/11 RA and 8/12 OA synovial specimens, RAGE was detected in synovial lining, sublining, and stroma. In RA, many T cells (CD45RO(+)) and some macrophages (CD68(+)) showed positive immunostaining for RAGE, whereas B cells were mostly negative. We found no difference in staining patterns between the RA and OA samples. We detected RAGE in RA and OA synovial tissue. The presence of RAGE on macrophages, T cells, and some B cells suggests its role in the pathogenesis of inflammatory joint disease.

Cartilage defect treatment strategies are dependent on the lesion size and severity. Osteochondral explants models are a platform to test cartilage repair strategies ex vivo. Current models lack in mimicking the variety of clinically relevant defect scenarios. In this controlled laboratory study, an automated device (artificial tissue cutter, ARTcut®) was implemented to reproducible create cartilage defects with controlled depth. In a pilot study, the effect of cartilage defect depth and oxygen tension on cartilage repair was investigated. Osteochondral explants were isolated from porcine condyles. 4 mm chondral and full thickness defects were treated with either porcine chondrocytes (CHON) or co-culture of 20 % CHON and 80 % MSC (MIX) embedded in collagen hydrogel. Explants were cultured with tissue specific media (without TGF-β) under normoxia (20 % O2) and physiological hypoxia (2 % O2). After 28 days, immune-histological stainings (Collagen II and X, Aggrecan) were scored (modified Bern-score, 3 independent scorer) to quantitatively compare treatments outcome. ARTcut® represents a software-controlled device for creation of uniform cartilage defects. Comparing the scoring results of the MIX and the CHON treatment, a positive relation between oxygen tension and defect depth was observed. Low oxygen tension stimulated cartilaginous matrix deposition in MIX group in chondral defects and CHON treatment in full thickness defects. ARTcut® has proved a powerful tool to create cartilage defects and thus opens a wide range of novel applications of the osteochondral model, including the relation between oxygen tension and defect depth on cartilage repair.

Abstract Objective High failure rates of (trauma induced) knee injuries highlight the need to improve current treatment strategies aiming to decrease the number of secondary osteoarthritis developed by patients in later stage [1-3]. In this controlled laboratory study the stimulative effect of mesenchymal stromal cells (MSC) on chondrocyte (CHON) extracellular matrix production was investigated in an ex vivo cartilage defect model (chondral vs. full thickness defects) cultured under normoxic (20 % O2) and physioxic (2 % O2) conditions. Design Porcine CHON or co-culture of 20 % CHON and 80 % MSC (MIX) were embedded in collagen type I hydrogel, implanted into 4 mm diameter cartilage defects of osteochondral explants and cultured with tissue specific media without addition of TGF-β under normoxia and physioxia. Chondral defects were induced automatically, while full thickness defects were created with biopsy punch. After 28 days of culture, samples were histologically processed, and treatments outcome was evaluated using international cartilage repair society (ICRS)-II scoring. Results Under physioxic conditions, cartilage repair scoring results of the MIX treatment (chondral 8.67 ± 2.42, full thickness 5.67 ± 1.21) were close to those of CHON treatment (chondral 8.17 ± 0.75, full thickness 7.33 ± 1.21). Overall, scoring results were higher in physioxia compared to normoxia conditions in chondral defects, but less or no prevalent for full thickness defects. Conclusion Co-culture of CHON with MSC represents a promising approach to stimulate chondrogenic repair and tissue formation in our ex vivo model and reduces total amount of CHON needed for cell-based treatment. Competing Interest Statement The authors have declared no competing interest. Footnotes * Andrea Schwab University Hospital Wuerzburg, Department Tissue Engineering and Regenerative Medicine, AO Research Institute Davos, Clavadelerstrasse 8, 7270 Davos, Switzerland E-Mail: andrea.schwabataofoundation.org * Alexa Buß University Hospital Wuerzburg, Department Tissue Engineering and Regenerative Medicine, Roentgenring 11, 97070 Wuerzburg, Germany, Klinikum Würzburg Mitte gGmbH, Juliusspital, Juliuspromenade 19, 97070 Wuerzburg, Germany, E-Mail: alexa.bussatkwm-klinikum.de * Oliver Pullig University Hospital Wuerzburg, Department Tissue Engineering and Regenerative Medicine, Roentgenring 11, 97070 Wuerzburg, Germany, Fraunhofer Institute for Silicate Research ISC, Translational Center Regenerative Therapies, Roentgenring 11, 97070 Wuerzburg, Germany, E-Mail: oliver.pulligatisc.fraunhofer.de

Type‐VI collagen is an integral part of the extracellular cartilage matrix. However, the exact amounts of type‐VI collagen in normal and osteoarthritic human cartilage still are not known. In this study, we describe an inhibition enzyme‐linked immunosorbent assay that was developed to quantitate type‐VI collagen epitopes found in guanidinium chloride extracts from normal and osteoarthritic human cartilage. In 31 cartilage samples from various localizations of healthy adult human knees, type‐VI collagen epitopes accounted for approximately 0.40% of the total collagen content. Interestingly, type‐VI collagen epitopes increased about 4‐fold in osteoarthritic cartilage. A statistically significant increase of type‐VI collagen epitopes was found during early stages of the disease, with only a superficial roughening of the cartilage surface and a loss of proteoglycans. Thus, these findings indicate that type‐VI collagen is a minor component of normal human articular cartilage and that the amount of type‐VI collagen epitopes increases significantly during early stages of osteoarthritis.