The combined therapy with chondroitin sulfate plus glucosamine sulfate or chondroitin sulfate plus glucosamine hydrochloride does not improve joint damage in an experimental model of knee osteoarthritis in rabbits.

Osteoarthritis is the most common chronic joint disorder especially during aging. Although with controversies, glucosamine, both in its forms of sulfate and hydrochloride, and chondroitin sulfate are commonly employed to treat osteoarthritis. Due to the modest improve in the symptoms observed in patients treated with these drugs alone, a formulation combining both agents has been considered. The discrepant results achieved for pain control or structural improvement in osteoarthritis patients has been attributed to the quality of chemical formulations or different bias in clinical studies. The current study has been designed to test the effects of two different combined formulations with adequate pharmaceutical grade of these drugs in osteoarthritic joints, and to explore the underlying mechanisms modulated by both formulations in different osteoarthritis target tissues. Knee osteoarthritis was surgically induced in experimental rabbits. Some animals received the combined therapy (CT)1, (chondroitin sulfate 1200mg/day + glucosamine sulfate 1500mg/day), or the CT2 ((chondroitin sulfate 1200mg/day + glucosamine hydrochloride 1500mg/day). Neither CT1 nor CT2 significantly modified the cartilage damage or the synovial inflammation observed in osteoarthritic animals. Treatments were also unable to modify the presence of pro-inflammatory mediators, and the synthesis of metalloproteinases in the cartilage or in the synovium of osteoarthritic animals. Combined therapies did not modify the decrease in the subchondral bone mineral density observed in osteoarthritic rabbits. Therapies of chondroitin sulfate plus glucosamine sulfate or chondroitin sulfate plus glucosamine hydrochloride failed to improve structural damage or to ameliorate the inflammatory profile of joint tissues during experimental osteoarthritis.

The increase in O-linked N-acetylglucosamine protein modification stimulates chondrogenic differentiation both in vitro and in vivo.

Insulin is an inducer of chondrocyte hypertrophy and growth plate chondrogenesis, although the specific molecular mechanisms behind these effects are mostly unknown. Our aim was to investigate whether insulin-induced chondrocyte hypertrophy occurs through a modification in the amount of O-linked N-acetylglucosamine (O-GlcNAc)-modified proteins and in the expression of the key enzymes of this pathway, O-GlcNAc transferase and O-GlcNAcase (OGA). We also studied if O-GlcNAc accumulation per se, induced by an OGA inhibitor, was able to induce pre-hypertrophic chondrocyte differentiation both in vitro and in vivo. Insulin-induced differentiation of ATDC5 pre-chondrocytes occurred alongside a gradual increase in the accumulation of O-GlcNac-modified proteins (O-GlcNAcylated proteins), as well as an increase in the expression of O-GlcNAc transferase and OGA. In the absence of insulin, O-GlcNAc accumulation induced by thiamet-G, a specific OGA inhibitor, was able to increase the gene expression of differentiation markers, as well as the activity of MMP-2 and -9. Thiamet-G also activated pERK, p-JNK, and p-p38 and the O-GlcNAcylation of Akt. Thiamet-G administration to C57/bl mice induced a significant expansion in the growth plate height and in the hypertrophic zone height. Therefore, our results show that O-GlcNAc glycosylation has chondromodulating activity.

RANKL synthesized by articular chondrocytes contributes to juxta-articular bone loss in chronic arthritis.

INTRODUCTION: The receptor activator nuclear factor-kappaB ligand (RANKL) diffuses from articular cartilage to subchondral bone. However, the role of chondrocyte-synthesized RANKL in rheumatoid arthritis-associated juxta-articular bone loss has not yet been explored. This study aimed to determine whether RANKL produced by chondrocytes induces osteoclastogenesis and juxta-articular bone loss associated with chronic arthritis. METHODS: Chronic antigen-induced arthritis (AIA) was induced in New Zealand (NZ) rabbits. Osteoarthritis (OA) and control groups were simultaneously studied. Dual X-ray absorptiometry of subchondral knee bone was performed before sacrifice. Histological analysis and protein expression of RANKL and osteoprotegerin (OPG) were evaluated in joint tissues. Co-cultures of human OA articular chondrocytes with peripheral blood mononuclear cells (PBMCs) from healthy donors were stimulated with macrophage-colony stimulating factor (M-CSF) and prostaglandin E2 (PGE2), then further stained with tartrate-resistant acid phosphatase. RESULTS: Subchondral bone loss was confirmed in AIA rabbits when compared with controls. The expression of RANKL, OPG and RANKL/OPG ratio in cartilage were increased in AIA compared to control animals, although this pattern was not seen in synovium. Furthermore, RANKL expression and RANKL/OPG ratio were inversely related to subchondral bone mineral density. RANKL expression was observed throughout all cartilage zones of rabbits and was specially increased in the calcified cartilage of AIA animals. Co-cultures demonstrated that PGE2-stimulated human chondrocytes, which produce RANKL, also induce osteoclasts differentiation from PBMCs. CONCLUSIONS: Chondrocyte-synthesized RANKL may contribute to the development of juxta-articular osteoporosis associated with chronic arthritis, by enhancing osteoclastogenesis. These results point out a new mechanism of bone loss in patients with rheumatoid arthritis.

Nonsteroidal antiinflammatory drugs and prostaglandin E(2) modulate the synthesis of osteoprotegerin and RANKL in the cartilage of patients with severe knee osteoarthritis.

OBJECTIVE: Although the osteoprotegerin (OPG)/RANK/RANKL system is the main modulator of bone remodeling, it remains unclear whether it is regulated in cartilage during osteoarthritis (OA). The aim of this study was to examine whether nonsteroidal antiinflammatory drug (NSAID) treatment modulates the synthesis of OPG and RANKL in the cartilage of patients with OA, and to investigate whether prostaglandin E(2) (PGE(2)) modifies this system in human OA chondrocytes in culture. METHODS: A 3-month clinical trial was carried out in 20 patients with severe knee OA, all of whom were scheduled to undergo knee replacement surgery. Ten of these patients were treated with celecoxib, and the other 10 patients, who did not want to be treated, served as the control group. After surgery, cartilage was processed for molecular biology studies. We also used human OA chondrocytes to examine the effects of PGE(2) on OPG/RANKL synthesis, examining which surface receptors were affected by PGE(2). RESULTS: In patients with OA, celecoxib decreased RANKL synthesis in the cartilage, thereby increasing the OPG:RANKL ratio. In human OA chondrocytes in culture, PGE(2) elicited a dose- and time-dependent increase in the synthesis of RANKL, the extent of which was greater than that of OPG. Confocal microscopy revealed that PGE(2) induced RANKL transport to the cell membrane. Only EP2/EP4 agonists reproduced the effects of PGE(2) on OPG and RANKL induction. CONCLUSION: Long-term NSAID treatment inhibited the resorptive signal synthesized by chondrocytes. In vitro, PGE(2) regulated the expression and release of these key mediators of bone metabolism by articular chondrocytes. The role of OPG/RANK/RANKL in OA cartilage metabolism is still unknown, although the synthesis of these proteins would enable the cartilage to control the activity of subchondral bone cells.