The synovial fluid from patients with focal cartilage defects contains mesenchymal stem/stromal cells and macrophages with pro- and anti-inflammatory phenotypes

John Garcia,Charlotte Hulme,Claire Mennan,Sally Roberts,Yvonne M Bastiaansen-Jenniskens,Gerjo J.V.M Van Osch,Bernhard Tins,Peter Gallacher,Karina Wright

doi:10.1016/j.ocarto.2020.100039

Abstract

ObjectiveThe synovial fluid (SF) of patients with focal cartilage defects contains a population of poorly characterised cells that could have pathophysiological implications in early osteoarthritis and joint tissue repair. We have examined the cells within SF of such joints by determining their chondrogenic capacity following culture expansion and establishing the phenotypes of the macrophage subsets in non-cultured cells. DesignKnee SF cells were obtained from 21 patients receiving cell therapy to treat a focal cartilage defect. Cell surface immunoprofiling for stem cell and putative chondrogenic markers, and the expression analysis of key chondrogenic and hypertrophic genes were conducted on culture-expanded SF cells prior to chondrogenesis. Flow cytometry was also used to determine the macrophage subsets in freshly isolated SF cells. ResultsImmunoprofiling revealed positivity for the monocyte/macrophage marker (CD14), the haematopoietic/endothelial cell marker (CD34) and mesenchymal stem/stromal cell markers (CD73, CD90, CD105) on culture expanded cells. We found strong correlations between the presence of CD14 and the vascular cell adhesion marker, CD106 (r = 0.81, p = 0.003). Collagen type II expression after culture expansion positively correlated with GAG production (r = 0.73, p = 0.006), whereas CD90 (r = −0.6, p = 0.03) and CD105 (r = −0.55, p = 0.04) immunopositivity were inversely related to GAG production. Freshly isolated SF cells were positive for both pro- (CD86) and anti-inflammatory markers (CD163 and CD206). ConclusionsThe cellular content of the SF from patients with focal cartilage injuries is comprised of a heterogeneous population of reparative and inflammatory cells. Additional investigations are needed to understand the role played by these cells in the attempted repair and inflammatory process in diseased joints.

Highlights

A growing body of research has demonstrated that joint degeneration in osteoarthritis (OA) is due to mechanical attrition but is facilitated, at least in part, by an inflammatory environment [1,2]
Cultured cells were expanded by seeding at 5000 cells/cm2 and trypsinisation up to passage 3, after which multichromatic flow cytometry using fluorochrome-conjugated antibodies was used to determine the positivity of certain immune cell markers (CD14-PercPCy5.5, CD19-BV421, CD45-PE), the vascular cell adhesion marker (CD106-APC), haematopoietic/endothelial/adipose cell marker (CD34APC), the major histocompatibility complex class II marker (HLA-DRAPC), the mesenchymal stem/stromal cells (MSCs) markers (CD90-PE, CD73-BV421 and CD105-APC), and putative chondropotency markers (CD39-APC, CD44-PercP-Cy5.5 [11], CD49c-PE [11], CD151-PE [11], CD166-BV421 [12], and CD271-BV421 [13]), were examined as we described previously [14]
The positivity of CD14 could indicate the presence of monocytes/macrophagelike cells, possibly originating from the synovium, infrapatellar fat pad and/or subchondral bone in the event of deep cartilage defects

Summary

Introduction

A growing body of research has demonstrated that joint degeneration in osteoarthritis (OA) is due to mechanical attrition (through progressive wear or trauma) but is facilitated, at least in part, by an inflammatory environment [1,2]. The synovial fluid (SF), which contains both its own population of cells and molecules produced by other joint tissues, provides a means of evaluating the inflammatory status of the local environment Whilst molecules such as cytokines have been extensively explored in the SF as potential biomarkers of OA [4], few studies have investigated the cellular content of the SF, in early OA [5]. Rheumatoid, end-stage OA patients [6,7] These cells form part of the poorly understood innate immune system of the joint, that could hold important information on OA pathophysiology, as well as therapeutic targets at an early stage of disease progress and may contribute to disease pathophysiology. The changes that occur in early OA affect the articular cartilage (increased catabolic enzyme activity), subchondral bone (subchondral bone remodelling and osteophyte formation), meniscus (fibrillation of avascular region), synovium (hyperplasia and lymphocyte infiltrates), tendons and menisci (ruptures and tears) [8]

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Results

Conclusion