Identification of a Prg4‐Expressing Articular Cartilage Progenitor Cell Population in Mice

Abstract

Objective We generated knock-in mice that express a tamoxifen-inducible Cre recombinase from the Prg4 locus (Prg4GFPCreERt2), and used these animals to fate-map the progeny of Prg4-positive articular cartilage cells at various ages. Methods We crossed Prg4GFPCreERt2 mice to Rosa26floxlacZ or Rosa26mTmG reporter strains, administered tamoxifen to the double heterozygous offspring at different ages, and assayed Cre-mediated recombination by histochemistry and/or fluorescence microscopy. Results In 1-month-old mice, the expression of the Prg4GFPCreERt2 allele mirrors expression of endogenous Prg4 and, when tamoxifen is given for 10 days, causes Cre-mediated recombination in ~70% of the superficial-most chondrocytes. Prg4GFPCreERt2 expressing cells are mostly confined to the top three cell layers of the articular cartilage in 1-month-old mice, but descendants of these cells are located in deeper regions of the articular cartilage in aged mice. At embryonic day 17.5, Prg4GFPCreERt2 expressing cells are largely restricted to the superficial-most cell layer of the forming joint, yet at approximately 1 year, progeny of these cells span the depth of the articular cartilage. Conclusions Our results indicate that Prg4-expressing cells located at the joint surface in the embryo serve as a progenitor population for all deeper layers of the mature articular cartilage. Also, our data reveal that Prg4GFPCreERt2 is expressed by superficial chondrocytes in young mice, but expands into deeper regions of the articular cartilage as the animals age. The Prg4GFPCreERt2 allele should be a useful tool for inducing efficient Cre-mediated recombination of floxed alleles at sites of Prg4 expression.

EGFR signaling is critical for maintaining the superficial layer of articular cartilage and preventing osteoarthritis initiation

“…Previous studies with chondrocyte-specific knockout of Mig-6 (10-12) also support our conclusion, as young mice exhibit hyperproliferation and hypercellularity in the top part of articular cartilage. A recent study identified Prg4-expressing cells located at the cartilage surface as chondrogenitors for deeper layers of the mature articular cartilage (19). We also observed a decrease in chondrocyte cellularity in an uncalcified area below the superficial layer, accompanied with an increase in the percentage of hypertrophic chondrocytes.…”

Induced superficial chondrocyte death reduces catabolic cartilage damage in murine posttraumatic osteoarthritis

“…We compared measures of cell number and the number of nuclei in the imaged cartilage volume (cell density) between an individual animal's left and right limbs and between the limbs of surface of articular cartilage (20 cell number or cell density, we measured the same specimens at weekly intervals ( Figure 1H and Supplemental Figure 4) and observed no systematic differences (Supplemental Figure 4 and Supplemental Table 2). …”

“…Mice that have a cDNA encoding a chimeric GFPCreERt2 protein knocked into the first coding exon of Prg4 can have Crerecombinase activity induced in most chondrocytes near the (10,20,30, and 40 μm) in control mice (C) and DTA-ablated mice (D) identify DAPI-stained chondrocyte nuclei (blue), nonrecombined membraneanchored dTomato-expressing chondrocytes (red), and Cre-recombined membrane-anchored eGFP-expressing chondrocytes (green). Identical thresholds were employed for all images.…”

Expression of p16INK4a is a biomarker of chondrocyte aging but does not cause osteoarthritis

“…Given our findings that interfering with p16 INK4a expression did not alter SASP production, investigating p16 INK4a loss in cell types with greater proliferative potential than chondrocytes may be of interest. For example, synovial fibroblasts and progenitor cells in the superficial zone of cartilage could be targeted for p16 INK4a loss with the Prg4 tm1(GFP/cre/ERT2)Abl allele (Kozhemyakina et al., 2015). Lineage tracing studies have suggested that subpopulations of synovial cells harbor regenerative potential (Decker et al., 2017) and a proliferative block in these cells through senescence may limit this capacity with aging.…”