RANKL is a Marker and Mediator of Local and Systemic Bone Loss in Two Rat Models of Inflammatory Arthritis

Abstract

RANKL is an essential mediator of bone erosions, but the role of RANKL in systemic bone loss had not been studied in arthritis. RANKL protein was increased in rat joint extracts and serum at the earliest stages of arthritis. Osteoprotegerin (OPG) treatment reversed local and systemic bone loss, suggesting that RANKL is both a marker and mediator of bone loss in arthritis.Introduction: RANKL is well established as an essential mediator of bone erosions in inflammatory arthritis, but the role of RANKL in systemic bone loss in arthritis had not been studied. We hypothesized that serum RANKL could serve as both a mediator and as a novel biomarker for local and systemic bone loss in arthritis. We challenged this hypothesis in two established rat models of inflammatory arthritis. We sought to determine whether serum RANKL was elevated early in disease progression and whether RANKL suppression could prevent both local and systemic bone loss in these models. Materials and Methods: Detailed time-course studies were conducted in animals with collagen-induced (CIA) or adjuvant-induced (AIA) arthritis to evaluate the onset and progression of inflammation (paw swelling), bone erosions, osteoclast numbers, and RANKL protein levels in arthritic joints and in serum. Additional CIA and AIA rats (n ‫ס‬ 8/group) received placebo (PBS) or recombinant OPG (3 mg/kg three times weekly) for 10 days beginning 4 days after disease onset (first macroscopic evidence of hind paw erythema and edema) to assess the role of RANKL in local and systemic bone loss. Results: RANKL protein was significantly elevated in the joints and serum of CIA and AIA rats within 1-2 days of disease onset. Increased RANKL levels were associated with local (hind paw) and systemic (vertebral) osteopenia in both models. The RANKL inhibitor OPG prevented local and systemic osteopenia in both models of established disease. Conclusions: RANKL protein is significantly increased both locally and systemically during the earliest stages of inflammatory arthritis in rats, suggesting that serum RANKL might have prognostic value for bone erosions and systemic osteopenia in this condition. RANKL inhibition through OPG prevented local and systemic bone loss in these arthritis models, suggesting that RANKL inhibition is a promising new approach for treating bone loss in arthritis.

The Evolving Systemic and Local Biomarker Milieu at Different Stages of Disease Progression in Rat Adjuvant-Induced Arthritis

“…Bone erosions were significantly (p<0.05) increased in paw sections beginning 2 days after disease onset ( Fig. 1b; published previously in [42]) and peaked at 10 days after onset (Fig. 1b, c).…”

“…Joint swelling was examined by measuring average hind paw volume via water plethysmography [11] or diameter via precision calipers [42].…”

“…While peripheral blood provides an expedient source to assess systemic levels of these factors, joint tissue can also provide important information on local changes [40][41][42]. Data comparing local versus systemic levels of markers and mediators of RA progression are rare in the literature.…”

“…Data comparing local versus systemic levels of markers and mediators of RA progression are rare in the literature. However, some of the existing reports describe divergent levels when comparing for local and systemic compartments [42][43][44]. These reports highlight the likelihood that systemic markers and mediators of arthritis might not fully reflect the underlying local disease progression.…”

Prevention of glucocorticoid‐induced bone loss in mice by inhibition of RANKL

“…Total deoxypyridinoline (DPD) concentrations in urine were determined after acid hydrolysis by ELISA (Total DPD; Metra Biosystems, Mountain View, CA). Local protein content of RANKL and tartrate-resistant acid phosphatase 5b (TRAP-5b) was measured in bone extracts from humeri and L3 vertebrae that had been frozen in liquid nitrogen, pulverized, and extracted as previously described (24). Murine RANKL was measured using a commercially available species-specific Luminex antibody-immobilized microbead kit (Linco Research, St. Charles, MO), which showed ϳ5% cross-reactivity with recombinant hRANKL standards (data not shown).…”