Enumeration and phenotypic characterization of synovial fluid multipotential mesenchymal progenitor cells in inflammatory and degenerative arthritis

Objectives

Synovial fluid (SF) contains rare clonogenic fibroblastic cells, but it is unknown whether these are true mesenchymal progenitor cells (MPCs) capable of trilineage differentiation or whether they participate in the pathogenesis of rheumatoid arthritis (RA). The present work evaluated SF for the presence of MPCs, to characterize these cells in relationship to bone marrow (BM) MPCs and to enumerate them in early and established RA, other inflammatory arthropathies and osteoarthritis (OA).

Methods

One hundred SF samples were evaluated (53 RA samples, 20 OA samples, 27 other samples). To investigate multipotentiality, cultures of SF-derived fibroblasts were subjected to chondrogenic, osteogenic and adipogenic differentiation assays, with BM-derived and skin-derived cultures used as positive and negative controls of differentiation. Trilineage differentiation of the individual clonogenic fibroblasts present was determined by direct measurements of produced Ca²⁺, sulphated glycosaminoglycan and lipid. Having established the MPC nature of individual SF clonogenic fibroblasts, fibroblast-colony forming unit assays were then used for enumeration of SF MPCs in early and late RA, OA and other arthropathies. The immunophenotype of SF MPCs before and after expansion was determined by multiparameter flow cytometry.

Results

Regardless of the nature of arthritis, polyclonal cultures of SF-derived fibroblastic cells possessed trilineage mesenchymal differentiation potentials. Individual clonogenic SF fibroblasts were also tripotential, confirming the MPC nature of SF cells. The number of MPCs in 1 ml SF was significantly higher in OA (median, 37) compared with RA (median, 2; P < 0.00001), corresponding to an average frequency of 2000 and 2 cells per 10⁶ of total cells, respectively. These differences were independent of SF volumes and inflammatory cell influx. No significant differences in MPC numbers were found between early and late RA (median, 3 and 2 cells/ml, respectively). Culture-expanded SF and BM MPCs had a similar phenotype (CD45^negD7-FIB^posCD13^posCD105^posCD55^posCD10^pos) but rare SF MPCs expressed low-affinity nerve growth factor receptor prior to expansion, a distinct marker of in vivo BM MPCs.

Conclusion

This work shows the presence of rare clonogenic tripotential MPCs in the SF, and is the first study to demonstrate that MPCs from two different anatomic sites have the same distinctive phenotype in vivo. The relative absence of SF MPCs in early and late RA does not support the concept of significant involvement in disease pathogenesis, but the relative abundance of these in OA provides the first direct evidence for MPC involvement in attempted joint repair in adults.