In vitro expression of receptor activator of nuclear factor-kappaB ligand and osteoprotegerin in cultured equine articular cells.
OBJECTIVE
To determine concentrations of receptor activator of nuclear factor-kappaB ligand (RANKL) and osteoprotegerin (OPG) in equine chondrocytes and synoviocytes and to quantify changes in the OPG:RANKL ratio in response to exogenous factors.


METHODS
Samples of articular cartilage and synovium with grossly normal appearance obtained from metacarpophalangeal and metatarsophalangeal joints of 5 adult (1- to 8-year-old) horses.


METHODS
Cell cultures of chondrocytes and synoviocytes were incubated with human recombinant interleukin-1beta (hrIL-1beta; 10 ng/mL), lipopolysaccharide (LPS; 10 microg/mL), or dexamethasone (100nM) for 48 hours. Negative control cultures received no treatment. Cells and spent media were assayed for RANKL and OPG concentrations by use of western blot and immunocytochemical analyses. Spent media were also assayed for OPG concentration by use of an ELISA.


RESULTS
RANKL and OPG were expressed in equine chondrocytes and synoviocytes in vitro. Cell-associated RANKL and OPG concentrations were not impacted by exogenous factors. Soluble RANKL release into media was significantly increased by hrIL-1beta in chondrocyte but not in synoviocyte cultures. Soluble OPG release into media was significantly increased by hrIL-1beta and LPS in chondrocyte but not in synoviocyte cultures. The soluble OPG:RANKL ratio was significantly increased by LPS in chondrocyte cultures. Dexamethasone decreased OPG expression in synoviocytes.


CONCLUSIONS
RANKL and OPG proteins were expressed in equine articular cells. Release of these proteins may affect osteoclastogenesis within adjacent subchondral bone. Thus, RANKL and OPG may have use as biomarkers and treatment targets in horses with joint disease.
Department of Veterinary Clinical Medicine, College of Veterinary Medicine, University of Illinois, Urbana, IL 61801, USA. cbyron@ruffianequine.com
Christopher R
CR
Byron
Anne M
AM
Barger
Allison A
AA
Stewart
Holly C
HC
Pondenis
Timothy M
TM
Fan
D000818:Animals; D015153:Blotting, Western / Q000662:veterinary; D002358:Cartilage, Articular / Q000166:cytology / Q000378:metabolism*; D018929:Cell Culture Techniques; D019902:Chondrocytes; D004797:Enzyme-Linked Immunosorbent Assay / Q000662:veterinary; D006736:Horses / Q000378:metabolism*; D007150:Immunohistochemistry / Q000662:veterinary; D007596:Joints / Q000166:cytology / Q000378:metabolism*; D053244:Osteoprotegerin / Q000096:biosynthesis*; D053245:RANK Ligand / Q000096:biosynthesis*; D013583:Synovial Membrane / Q000166:cytology / Q000378:metabolism
D053244:Osteoprotegerin; D053245:RANK Ligand