Protective Properties of Conditioned Media From Adipose Stem Cells On Osteoarthritic Chondrocytes

Maria Isabel Guillén¹, Julia Platas¹, Vicente Mirabet², Miguel Angel Castejón³, Francisco Gomar⁴ and Maria Jose Alcaraz¹, ¹Pharmacology, University of Valencia, Burjasot, Valencia, Spain, ²Generalitat Valenciana, Valencia, Spain, ³De la Ribera University Hospital, Alzira, Spain, ⁴Surgery, University of Valencia and University Hospital, Valencia, Spain

Meeting: ACR/ARHP Annual Meeting 2012

Keywords: cartilage, chondrocytes, Inflammation, Mesenchymal stem cells and osteoarthritis

Session Information

Title: Biology and Pathology of Bone and Joint

Session Type: Abstract Submissions (ACR)

Background/Purpose: Adipose-derived mesenchymal stem cells (ASC) exhibit a high potential for cell therapy and they might also act as a cellular source for supplying soluble factors exerting anti-inflammatory or trophic effects on cells. Osteoarthritis (OA) involves the destruction of articular cartilage leading to disability. The purpose of this study was to investigate whether conditioned medium from adipose-derived mesenchymal stem cells (ASC-CM) improves the inflammatory and degradative response induced by interleukin-1β in OA chondrocytes and cartilage.

Methods: Adipose tissue from patients subjected to abdominal lipectomy surgery, was used for ASC isolation by collagenase treatment. Cells were incubated in DMEM/F12 containing 15% human serum. Cell phenotype was analyzed by flow cytometry with specific antibodies anti-CD105-PE, anti-CD90PerCP-eFluo 710, anti-CD34APC, and anti-CD45-PE (International Society of Cellular Therapy), and cellular viability with propidium iodide. The conditioned medium (ASC-CM) was collected after 48h of culture, centrifuged and stored at -80ºC in sterile conditions. Cartilage specimens were obtained from patients with diagnosis of advanced OA. Protocols were approved by the Institutional Ethical Committee. Chondrocytes were used in primary culture. Cartilage explants or isolated chondrocytes were stimulated with 10 ng/ml interleukin (IL)-1β for different times. Gene expression was analyzed by real-time PCR. Protein expression was investigated by ELISA. Nitric oxide (NO) production and matrix metalloproteinase (MMP) activity were measured as microplate fluorescence assays. Prostaglandin E₂ (PGE₂)was evaluated by RIA.

Results: IL-1β significantly increased the production of NO, PGE₂and MMP activity, and the expression of IL-1β, tumor necrosis factor-α (TNFα), vascular endothelial growth factor (VEGF) and a wide range of chemokines, after 24h, with respect to basal conditions. ASC-CM treatment of OA explants or chondrocytes reduced the production of NO which was dependent on inducible NO synthase down-regulation. In addition, PGE₂levels as well as MMP activity were significantly decreased. A lower expression of IL-1β, TNFα, VEGF and chemokines CCL-4, CCL-5, CCL-8, CCL-19, CCL-20, CXCL-1, CXCL-2, CXCL-3 and CXCL-8 was observed in chondrocytes treated with ASC-CM with respect to IL-1β controls. We have also evaluated the senescence marker β-galactosidase. In cells treated with IL-1β and ASC-CM for 7 days, we observed a significant reduction in senescence-associated β-galactosidase activity in comparison with chondrocytes treated with IL-1β.

Conclusion: The results of our study demonstrate the down-regulation of inflammatory and catabolic mediators by ASC-CM in OA explants and chondrocytes. Our data have revealed a protective role for ASC-CM in chondrocytes suggesting possible applications against cartilage degradation.

Disclosure:

M. I. Guillén,
None;

J. Platas,
None;

V. Mirabet,
None;

M. A. Castejón,
None;

F. Gomar,
None;

M. J. Alcaraz,
None.

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