Multiple mesenchymal progenitor cell subtypes with distinct functional potential are present within the intimal layer of the hip synovium

Abstract

Background: The synovial membrane adjacent to the articular cartilage is home to synovial mesenchymal progenitor cell (sMPC) populations that have the ability to undergo chondrogenesis. While it has been hypothesized that multiple subtypes of stem and progenitor cells exist in vivo, there is little evidence supporting this hypothesis in human tissues. Furthermore, in most of the published literature on this topic, the cells are cultured before derivation of clonal populations. This gap in the literature makes it difficult to determine if there are distinct MPC subtypes in human synovial tissues, and if so, if these sMPCs express any markers in vivo/in situ that provide information in regards to the function of specific MPC subtypes (e.g. cells with increased chondrogenic capacity)? Therefore, the current study was undertaken to determine if any of the classical MPC cell surface markers provide insight into the differentiation capacity of sMPCs.

Methods: Clonal populations of sMPCs were derived from a cohort of patients with hip osteoarthritis (OA) and patients at high risk to develop OA using indexed cell sorting. Tri-differentiation potential and cell surface receptor expression of the resultant clones was determined.

Results: A number of clones with distinct differentiation potential were derived from this cohort, yet the most common cell surface marker profile on MPCs (in situ) that demonstrated chondrogenic potential was determined to be CD90⁺/CD44⁺/CD73⁺. A validation cohort was employed to isolate cells with only this cell surface profile. Isolating cells directly from human synovial tissue with these three markers alone, did not enrich for cells with chondrogenic capacity.

Conclusions: Therefore, additional markers are required to further discriminate the heterogeneous subtypes of MPCs and identify sMPCs with functional properties that are believed to be advantageous for clinical application.

Keywords: Clonal analysis; Heterogeneity; Hip; Osteoarthritis; Synovial progenitor cells.

Fig. 1

Experiment design flow chart

Fig. 2

In situ and in vitro cell surface marker data from 4 clones from the same patient. The in situ expression of each CD marker is represented by the vertical bar black. The in vitro expression of each marker in the clonal derived cell population is represented by the blue histogram. The isotype/negative control for each CD marker is represented by the red histogram

Fig. 3

Gene expression after adipogenic (a), chondrogenic (b) and osteogenic (c) differentiation. Results from 4 clonal cell lines from a single OA patient. The differentiated gene expression values are normalized to undifferentiated gene expression values from the same clone. *p < 0.05

Fig. 4

Histological analysis of differentiation. Oil Red O staining was employed to detect lipid accumulation after adipogenic differentiation (top row). All clones examined except clone # 3 demonstrated Oil Red O positive staining. Safranin O staining was employed to detect proteoglycan accumulation after chondrogenic differentiation (middle row). Only clone # 1 demonstrated positive Safranin O staining. Alizarin red staining was employed to detect calcium accumulation after osteogenic differentiation (bottom row). None of the clones examined demonstrated positive Alizarin red staining. A characterized and non-clonal MSC line was used as a positive control (right column) and demonstrated positive staining for Oil Red O, Safranin O and Alizarin Red. Scale bars equal 50 μm for Oil Red O and Alizarin Red stained images, and 200 μm for Safranin O stained images