Comparative analysis of gene expression between articular cartilage-derived cells to assess suitability of fibronectin adhesion assay to enrich chondroprogenitors.

Abstract

Enhanced chondrogenesis and reduction in hypertrophy are essential pre-requisites for cell-based therapy in regenerative research for cartilage loss. Chondroprogenitors, isolated by fibronectin adhesion assay (FAA), have shown promising results in various preclinical studies due to their inherent characteristics. However, the need for monolayer culture and the effect of expansion on cell phenotype render differentiation between chondroprogenitors and chondrocytes (native cartilage cells) difficult. This is further complicated due to reported de-differentiation of chondrocytes in culture. Thus, the aim of our study was to harvest cells from articular cartilage and compare their gene expression to cells demonstrating adherence and non-adherence to fibronectin. Fresh-cells (FC) were isolated from human osteoarthritic knee joints(n = 3) and subjected to FAA. Cells unbound to fibronectin (20 min after plating) were termed as FAA-ve. Attached cells were further cultured for five population doublings and designated FAA+ve. RNA from all three cell groups was assessed for SOX-9, ACAN, COL2A1, COL1A1, RUNX2 and COL10A1. All three groups exhibited moderate to high expression of markers of chondrogenesis and marker of chondrocyte hypertrophy. FAA+ve group exhibited significantly lower levels of hypertrophy markers: RUNX2 (vs FC and FAA-ve, P = 0.018) and COL10A1(vs FAA-ve, P = 0.005). Our results demonstrated that fibronectin effectively isolated cells distinct from mature chondrocytes in terms of reduced hypertrophic tendency. This is noteworthy as cells isolated by FAA, retaining their inherent progenitor phenotype, with upregulation of chondrogenic markers may be used successfully for cartilage repair in future translational work.