Regulation of growth plate and articular chondrocyte differentiation : implications for longitudinal bone growth and articular cartilage formation

Abstract

Overall height and body proportions in humans are determined primarily by bone growth. Linear bone growth occurs at the growth plate, a thin layer of cartilage at the ends of long bones between the epiphysis and metaphysis. In the growth plate, resting/stem-like chondrocytes divide and give rise to proliferative chondrocytes, which, in turn, enlarge to become hypertrophic chondrocytes that ultimately undergo apoptotic cell death and are replaced by bone. Articular cartilage is an embryologically related but permanent tissue that lines the ends of long bones providing a lubricated surface for articulation and distributing loads to minimize stress on underlying subchondral bone. In both growth plate and articular cartilage, precise cell signaling mechanisms ensure normal bone growth and joint maintenance, respectively, by regulating cell differentiation, proliferation, and hypertrophy as well as matrix synthesis and turnover. A better understanding of these mechanisms has broad clinical implications for preventing, diagnosing, and treating skeletal diseases.

The aim of this thesis was to study the molecular mechanisms regulating growth plate and articular chondrocyte differentiation. In this regard, similarities and differences between these structurally similar yet functionally distinct skeletal tissues were also investigated.

We first explored gene expression related to the BMP signaling system in different layers of rat growth plate cartilage using manual microdissection, microarray, and real-time PCR (Paper 1). Our findings suggest a functional BMP signaling gradient across the growth plate where BMP antagonists are highly expressed in the resting and proliferative zones and BMP agonists are highly expressed in the hypertrophic zone. Gradients in BMP action may thus provide a key mechanism responsible for the spatial regulation of chondrogenesis in growth plate cartilage and thereby contribute to longitudinal bone growth.

Another important mechanism is the Ihh/PTHrP feedback system, which prevents premature hypertrophic differentiation in embryonic epiphyseal cartilage. However, less is known about its organization in the growth plate after birth when the area undergoes substantial remodeling. We therefore explored Ihh/PTHrP-related gene expression in postnatal rat growth plate and surveyed Ihh activity in the Gli1-lacZ mouse growth plate (Paper 2). We found that the embryonic Ihh/PTHrP feedback system is maintained postnatally except that the source of PTHrP has shifted to a more proximal location in the resting zone. This finding provides insight into the potential role of Ihh/PTHrP signaling in growth plate senescence and fusion.

Similar to the growth plate, articular cartilage is structurally organized into chondrocyte layers; however, its cellular differentiation program is not as well characterized. Thus, we explored the similarities and differences between articular and growth plate cartilage by comparing gene expression profiles of individual rat epiphyseal cartilage layers using bioinformatic approaches (Paper 3). Our findings revealed unexpected transcriptional similarities between the deeper zones of articular cartilage and the resting zone of growth plate cartilage as well as between articular cartilage superficial zone and growth plate cartilage hypertrophic zone, suggesting that in articular cartilage, superficial chondrocytes differentiate from chondrocytes in the deeper layers following a program that has some similarities to the hypertrophic differentiation program in growth plate cartilage.

Based on these findings, we hypothesized that microenvironment regulates chondrocyte differentiation into either articular or growth plate cartilage. We tested this hypothesis by transplanting growth plate cartilage to the articular surface in an EGFP rat model that enabled cell tracing (Paper 4). We found that hypertrophic differentiation appeared to be inhibited in growth plate cartilage transplanted to the articular surface. The transplanted cartilage also underwent structural remodeling into articular-like cartilage, which suggests that the synovial microenvironment inhibits hypertrophic differentiation and promotes articular cartilage formation.