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AB0069 Conditioned media from adipose-derived mesenchymal stem cells decreases senescence and enhances collagen ii expression in osteoarthritic chondrocytes
  1. M. I. Guillen1,
  2. J. Platas1,
  3. V. Mirabet2,
  4. M. A. Castejon3,
  5. F. Gomar4,
  6. M. J. Alcaraz1
  1. 1Pharmacology, University of Valencia, Burjasot
  2. 2Valencia Transfusion Center, Generalitat Valenciana, Valencia
  3. 3Traumatology, De la Ribera Hospital, Alzira
  4. 4Surgery, University of Valencia, Valencia, Spain

Abstract

Background Adipose-derived mesenchymal stem cells (ASC) might act as a cellular source of soluble factors exerting anti-inflammatory or trophic effects on cells. Osteoarthritis (OA) is characterized by the progressive loss of structure and functionality of articular cartilage.

Objectives In the present study we explored the effect of conditioned medium from adipose-derived mesenchymal stem cells (ASC-CM) on the metabolism of OA chondrocytes in primary culture.

Methods ADC were isolated from adipose tissue of patients subjected to abdominal lipectomy surgery, by collagenase treatment. Cells were incubated in DMEM/F12 + 15% human serum. Cell phenotype was analysed 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. ASC-CM was collected after 48h of culture, centrifuged and stored at -80°C in sterile conditions. Protocols were approved by the Institutional Ethical Committee. Chondrocytes were isolated from knee cartilage samples from patients with diagnosis of advanced OA. OA chondrocytes in primary culture were stimulated with 10 ng/ml interleukin (IL)-1β for 1 or 5 days. We have evaluated the senescence marker β-galactosidase by histochemical staining of cells. Collagen II production was analyzed by immunofluorescence, gene expression by real-time qPCR and protein expression by ELISA.

Results The percentage of senescent chondrocytes was increased by IL-1β treatment after 1 or 5 days whereas ASC-CM significantly counteracted this effect. When OA chondrocytes were treated with IL-1β and/or ASC-CM for 5 days, we observed a significant maintenance of collagen II expression in chondrocytes cultured with ASC-CM either in basal conditions or in the presence of IL-1β stimulation. Cytokine stimulation of chondrocytes augmented gene expression and protein release into the culture medium of matrix metalloproteinases (MMPs). Our data indicate that ASC-CM significantly decreased the production of MMP-13. In addition, AC-CM reduced gene expression and protein levels of the pro-inflammatory cytokine IL-6 and enhanced the production of the anti-inflammatory cytokine IL-10.

Conclusions We have shown that ASC-CM treatment of OA chondrocytes prevents the appearance of senescence features and the production of degradative mediators. These findings suggest a protective role for ASC-CM in OA chondrocytes.

Acknowledgements This work was funded by grants SAF2010-22048 (MINECO), RETICEF RD06/0013/2001 (MINECO, ISCIII, FEDER), and Prometeo2010-047 (Generalitat Valenciana).

Disclosure of Interest None Declared

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