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Objective. Chondrocytes of the epiphyseal growth zone are regulated by the Indian hedgehog (IHH)-parathyroid hormone-related protein (PTHrP) axis. In weight-bearing joints, this growth zone comes to be subdivided by the secondary ossification center into distinct articular and growth cartilage structures. The purpose of this study was to explore the cells of origin, localization, regulation of expression, and putative functions of IHH and PTHrP in articular cartilage in the mouse.Methods. We assessed IHH and PTHrP expression in an allelic PTHrP-LacZ-knockin mouse and several versions of PTHrP-null mice. Selected joints were unloaded surgically to examine load-induction of PTHrP and IHH.Results. The embryonic growth zone appears to serve as the source of PTHrP-expressing proliferative chondrocytes that populate both the forming articular cartilage and growth plate structures. In articular cartilage, these cells take the form of articular chondrocytes in the midzone. In PTHrP-knockout mice, mineralizing chondrocytes encroach upon developing articular cartilage but appear to be prevented from mineralizing the joint space by IHH-driven surface chondrocyte proliferation. In growing and adult mice, PTHrP expression in articular chondrocytes is loadinduced, and unloading is associated with rapid changes in PTHrP expression and articular chondrocyte differentiation.Conclusion. We conclude that the IHH-PTHrP axis participates in the maintenance of articular cartilage. Dysregulation of this system might contribute to the pathogenesis of arthritis.A growth zone at each end of a long bone drives linear growth. This zone is comprised of chondrocytes that move through an orderly differentiation program (round3flat3prehypertrophic3hypertrophic) that is regulated by the Indian hedgehog (IHH)-parathyroid hormone-related protein (PTHrP) feedback loop. IHH is produced by prehypertrophic chondrocytes and feeds back to control the proliferation of the round proliferative chondrocytes (RPCs) that lie early in the program.

The PTHrP gene is expressed in the periosteum and in tendon and ligament insertion sites in a PTHrPlacZ knockin reporter mouse. Here, we present a more detailed histological evaluation of PTHrP expression in these sites and study the effects of mechanical force on PTHrP expression in selected sites. We studied the periosteum and selected entheses by histological, histochemical, and in situ hybridization histochemical techniques, and tendons or ligaments were unloaded by tail suspension or surgical transection. In the periosteum, PTHrP is expressed in the fibrous layer and the type 1 PTH/PTHrP receptor (PTH1R) in the subjacent cambial layer. PTHrP has distinct temporospatial patterns of expression in the periosteum, one hot spot being the metaphyseal periosteum in growing animals. PTHrP is also strongly expressed in a number of fibrous insertion sites. In the tibia these include the insertions of the medial collateral ligament (MCL) and the semimembraneosus (SM). In young animals, the MCL and SM sites display a combination of underlying osteoblastic and osteoclastic activities that may be associated with the migration of these entheses during linear growth. Unloading the MCL and SM by tail suspension or surgical transection leads to a marked decrease in PTHrP/lacZ expression and a rapid disappearance of the subjacent osteoblastic population. We have not been able to identify PTHrP-lacZ in any internal bone cell population in the PTHrP-lacZ knockin mouse in either a CD-1 or C57Bl/6 genetic background.In conclusion, we have identified PTHrP expression in surface structures that connect skeletal elements to each other and to surrounding muscle but not in intrinsic internal bone cell populations. In these surface sites, mechanical force seems to be an important regulator of PTHrP expression. In selected sites and/or at specific times, PTHrP may influence the recruitment and/or activities of underlying bone cell populations.

The sites that receive ligament and tendon insertions (entheses) on the cortical surfaces of long bones are poorly understood, particularly as regards modeling and regulation. Entheses are classified as either fibrocartilagenous or fibrous based on their structures. Fibrous entheses typically insert into the metaphysis or diaphysis of a long bone, bear a periosteal component, and are modeled during long bone growth. This modeling forms a root system by which the insertions attach to the cortical surface. In the case of the medial collateral ligament, modeling drives actual migration of the ligament along the cortical surface in order to accommodate linear growth, whereas in other sites modeling may excavate a deep cortical root system (e.g., the teres major insertion) or a shallow root system with a large footprint (e.g., the latissimus dorsi insertion). We report here that conditionally deleting parathyroid hormone-related protein (PTHrP) in fibrous entheses via Scleraxis-Cre targeting causes modeling to fail in these three iterations of osteoclast-driven enthesis excavation or migration. These iterations appear to represent formes frusts of a common modeling strategy, presumably differing from each other as a consequence of differences in biomechanical control. In sites in which PTHrP is not induced, either physiologically or because of conditional deletion, modeling does not take place and fibrocartilage is induced. These findings represent the initial genetic evidence that PTHrP regulates periosteal/intramembranous bone cell activity on cortical bone surfaces and indicate that PTHrP serves as a load-induced modeling tool in fibrous insertion sites during linear growth.