Anti-Inflammatory Effect of Adipose-Derived Stromal Vascular Fraction on Osteoarthritic Temporomandibular Joint Synoviocytes

Abstract

Background: Osteoarthritis (OA) in the temporomandibular joint (TMJ) in the TMJ (TMJ-OA) is difficult to treat, and new alternative treatments are needed. Recently, adipose-derived stem cells (ASCs) have been introduced as a promising cell source because of their anti-inflammatory effects. However, the cost and availability of these cells limited broader applications of stem cell therapy. Thus, Thus, stromal vascular fraction (SVF) containing sufficient amount of ASCs at low cost can be an alternative. In this study, we aimed to demonstrate the use of uncultured, optimally isolated SVF for the treatment of TMJ-OA.

Methods: First, we optimized the method of isolation to harvest high-quality SVFs with a large yield of ASCs. Then, we analyzed the quantity of ASCs in the SVF and performed characterization of stem cell homology. Subsequently, to evaluate the anti-inflammatory effect of high-quality SVF, an in vitro study was performed to assess the expression patterns of inflammatory cytokines including prostaglandin E₂ (PGE₂), IL-6, and CXCL8/IL-8, COX2, TNF, IFN, CCL2/MCP-1 and CCL5/RANTES in co-culture with synoviocytes derived from the synovial fluid in the TMJ-OA patients.

Results: The SVF containing approximately 32% ASCs was isolated via the our optimized isolation method. The SVF significantly down-regulated certain inflammatory cytokines such as PGE2, CXCL8/IL-8 in TMJ-OA tissue-derived synoviocytes.

Conclusion: Although further study is needed, our study suggests that transplantation of adipose tissue-derived SVF cells might be a feasible and a novel therapeutic option for TMJ-OA in the future.

Keywords: Adipose-derived stem cells; Anti-inflammation; Osteoarthritis; SVF; Temporomandibular joint disorder.

Fig. 1

Isolation of the stromal vascular fraction enhanced the yield of adipose-derived stem cells. A Heterogeneous cell population of SVF cell pellet (left arrow). We isolated a large cell population from adipose tissue via the traditional isolation method. The microscopic view of heterogeneous cells on the hemocytometer shows that most of the cells were erythrocytes (right). B SVF isolated by means of Ficoll gradient centrifugation show absence of most erythrocytes (left, arrow) and a high purity of the cell population on the hemocytometer (right). C High-purity SVF contained numerous ASCs (15–20 µm) with rod-shaped morphology. Erythrocytes seemed smaller in size than ASCs, and adipocytes seemed larger in size than ASCs

Fig. 2

Flow cytometry histograms of human SVF. Red and black colored lines represent the specific antibodies indicated. Gray color indicates an isotype of each specific antibody. A Both CD34 and CD45 markers are highly expressed in the hematopoietic cell population. CD34 and CD45 cells have been described as representing about 14% and 7% of the SVF, respectively. B Both CD105 and CD90 markers are highly expressed on ASCs. CD105 and CD90 cells have been described as representing about 24% and 21% of the SVF, respectively

Fig. 3

Flow cytometry histograms of human ASCs. ASCs were selected by adherent capacity on a flask. Attached cells were harvested by trypsinization, and the expression levels were analyzed by MSC positive markers (CD73, CD105) and negative markers (CD34, CD45), respectively

Fig. 4

In vitro multi-lineage differentiation potential of ASCs was confirmed. A Adipogenesis was demonstrated by Oil Red O staining, B osteogenesis by Alizarin red S staining, and C chondrogenesis by Alcian blue staining. Cells were treated with either control (left panels) or differentiation media (right panels)

Fig. 5

Evaluation of co-culture conditions. A Schematic of the co-culture system is shown. B Morphology of synoviocytes. There was no difference in cell proliferation on day 2 and 7, and C tSVF(left), aSVF(middle), and ASCs (right) were evaluated on day 7. Original magnification: 40×

Fig. 6

Anti-inflammatory effects of SVF on synoviocytes secreted PGE2, CXCL8/IL-8 and IL-6. aSVF significantly down-regulates the expression of the main osteoarthritis inflammatory cytokines and chemokines. PGE2, CXCL8/IL-8 and IL-6 release is decreased by tSVF, aSVF, and ASCs co-culture. *p < 0.05, **p < 0.01 compared with each monoculture of synoviocytes

Fig. 7

Anti-inflammatory effects of SVF on synoviocytes secreted COX-2 and TNFα. aSVF, tSVF and ASCs does not significantly down-regulate the expression of COX-2 and TNFα