The Rxr agonist SR 11237 causes disturbed skeletal morphogenesis in a rat model

Introduction: Retinoid X Receptor (RXR) is a type 2 nuclear receptor with important roles in cell death, development, metabolism and cell differentiation. RXR has the ability to form homodimers or heterodimers with other nuclear receptors (PPARs, VDR, LXL and others) in a permissive or non-permissive way. In mice, the non-permissive binding of RXR to the retinoic acid receptor (RAR) has been shown to be important in bone development (exposure to retinoic acid during embryonic development causes deletions and truncations in the forelimbs). However, although the impact of RXR itself is far-reaching, little is known about its relevance in long bone development. To determine its importance, the RXR agonist SR 11237 was used to identify the effects of RXR activation on endochondral ossification, a process through which long bones form by replacing a cartilaginous scaffold with bone. Purpose: To examine the effects of RXR specific activation on endochondral bone development in the rat. Methods: Sprague-Dawley rats were given an intraperitoneal injection (IP) of SR 11237 (pan-RXR specific agonist at a concentration of 25 mg per kg) or DMSO (vehicle) once a day from post-natal day 5 to 15. The animals were harvested on post-natal day 16 for Micro-computed Tomography (uCT) scanning (males only), histology and weight determination. For histology, the sections were stained with Safranin O / Fast green, picro-sirius red, Tartrate-resistant acid phosphatase (TRAP) and TUNEL. Further, p57 and SOX9 immunohistochemistry was completed on the sections. For uCT, the limbs were scanned at a resolution of 50 microns / voxel. Results: RXR activation by SR 11237 causes disturbed ossification and bone morphology in a rat model. uCT analyses and Safranin O staining of the rat long bones show premature growth plate closure and an infiltration of ossified tissue through the central epiphysis of the bone. In addition, the cells surrounding this invasion of ossified material are pre-hypertrophic in size and shape. This is consistent with the pattern of the p57 immunohistochemical staining. SOX9 (a proliferative marker) is also found in the cells surrounding the calcified tissue. In the RXR treated bones, the central epiphysis is highly positive for TRAP staining (an indicator for osteoclastic activity), and picro-sirius red staining reveals a large amount of collagen present. Further, TUNEL staining displays concentrated cell death at the osteo-chondral junction. Although this morphological disorganization is observed in all long bones, uCT also reveals a less calcified, pitted surface on the face of the scapula. The smaller bones of the hands and feet are also thinner and appear somewhat osteopenic. Males are more impaired than females, and the tibia is significantly more afflicted than the humerus and femur. The RXR treated animals of both sexes weigh 30% less than the controls. Conclusion: In a rat model, increased RXR signaling causes irregular ossification and premature closure of the growth plate. Consequently, this may result in long term effects on long bone and joint morphology, and lead to additional pathology (ex. Osteoarthritis) through joint mal-alignment or disruption of normal gait.