Intermittent Hypoxic Therapy Inhibits Allogenic Bone-Graft Resorption by Inhibition of Osteoclastogenesis in a Mouse Model

Systemic Intermittent Hypoxic Therapy (IHT) relies on the adaptive response to hypoxic stress. We investigated allogenic bone-graft resorption in the lumbar spine in 48 mice. The mice were exposed to IHT for 1 week before surgery or 1 week after surgery and compared with controls after 1 and 4 weeks. Complete graft resorption was observed in 33–36% of the animals in the control group, but none in the preoperative IHT group. Increased bone-graft volume was demonstrated by micro-computed tomography in the preoperative IHT group after 1 week (p = 0.03) while a non-significant difference was observed after 4 weeks (p = 0.12). There were no significant differences in the postoperative IHT group. Increased concentration of immune cells was localized in the graft area, and more positive tartrate-resistant acid phosphatase (TRAP) staining was found in controls compared with IHT allogenic bone grafts. Systemic IHT resulted in a significant increase of the major osteoclast inhibitor osteoprotegerin as well as osteogenic and angiogenic regulators Tgfbr3, Fst3l, Wisp1, and Vegfd. Inflammatory cytokines and receptor activator of nuclear factor kappa-B ligand (RANKL) stimulators IL-6, IL-17a, IL-17f, and IL-23r increased after 1 and 4 weeks, and serum RANKL expression remained constant while Ccl3 and Ccl5 decreased. We conclude that the adaptive response to IHT activates numerous pathways leading to inhibition of osteoclastic activity and inhibition of allogenic bone-graft resorption.

P013 Serum osteoprotegerin and RANKL in patients with Juvenile Idiopathic Arthritis and their correlation with bone mineral density

Background Juvenile idiopathic arthritis (JIA) is the most common chronic rheumatic arthropathy of childhood and is associated with low bone mass, and may hasten the onset of osteoporosis later in life1. Bone loss occurs because of an imbalance between osteoclasts-activating factors like receptor activator of nuclear factor-κB ligand (RANKL) and its inhibitor osteoprotegerin (OPG) 2. Dual energy X-ray absorptiometry (DXA) is the preferred method for measuring bone mineral density (BMD) in children and to identify and follow individuals at risk for fracture 3. The objective is the Evaluation of serum levels of osteoprotegerin and RANKL and their correlation with BMD in JIA patients. Methods Forty JIA patients (according to the revised classification criteria of ILAR) and 40 healthy children individually matched for age, sex and race were included in this study. Children excluded from the study were those with primary and secondary causes of osteoporosis (such as chronic illness). All patients were assessed clinically by: age, sex, body mass index, type of JIA, disease duration and disease activity (by Juvenile Arthritis Disease Activity Score; JADAS 10). The functional disability was assessed by the Childhood Health Assessment Questionnaire (CHAQ). Blood samples were collected from JIA patients and healthy controls to determine serum levels of OPG and RANKL by ELISA. DXA scans were done using GE Healthcare Lunar DPX, Madison, Wisconsin. Bone mineral density of the L1-L4 lumbar spine and total body less head (TBLH) was evaluated in g/cm2 and expressed as Z score for age, sex according to the reference data given for this equipment. Results The study included 40 patients (25 females) with a mean age of 11.14 years and median disease duration of 2.5 years. As regard JIA type, 45% of patients were oligoarticular, 32.5% were polyarticular, and 22.5% were systemic JIA. Median JADAS 10 was 13.95. Patients (especially polyarticular JIA) had significantly higher serum RANKL levels and lower serum OPG and OPG/RANKL ratio when compared with controls (with p-value <0.001, 0.032 and <0.001 respectively). A diagnosis of low BMD (BMD Z-score ≤ -2) was given in 25% of patients (15% polyarticular and 10% systemic) by DXA of lumbar spine, and 20% (10% polyarticular and 10% systemic) by DXA of TBLH. On the other hand, no patient was given a diagnosis of osteoporosis (BMD Z-score ≤ -2 and a significant fracture history). Low BMD at lumbar spine and TBLH was negatively correlated with serum RANKL while positively correlated with OPG/RANKL ratio. Moreover, low BMD at lumbar spine was positively correlated with serum OPG level Conclusion High RANKL and low OPG levels appear to be associated with low bone mass in JIA patients. Patients with JIA (especially polyarticular and systemic subtype) are at increased risk of low bone mineral mass. Disclosure of Interests None declared

Serum RANKL levels in Chinese patients with ankylosing spondylitis: a meta-analysis

Objective We aimed to determine the association between serum receptor activator of nuclear factor-kappa B ligand (sRANKL) levels and ankylosing spondylitis (AS) in Chinese patients. Methods The PubMed, Cochrane Library, Embase, Chinese Biomedical Database, Web of Science, China National Knowledge Infrastructure, VIP, and Wan Fang databases were searched for studies conducted before October 1, 2020, without language restrictions. STATA version 12.0 and Revman version 5.3 were used to analyze the data. The standard mean differences (SMDs) and corresponding 95% confidence intervals (95% CIs) were calculated. Results Twelve clinical case–control studies, including 585 patients with AS and 423 healthy controls, were included. The combined SMD for sRANKL suggested that the sRANKL level was significantly higher in Chinese patients with AS than in healthy controls (SMD: 3.27, 95% CI 2.11–4.43, P < 0.00001). Serum RANKL-related factor osteoprotegerin (OPG) levels (SMD: 0.86, 95% CI 0.09–1.64, P < 0.03) were lower in the Chinese patients with AS than in healthy controls, and the RANKL/OPG ratio (SMD = 1.05, 95% CI 0.64–1.46, P < 0.00001) in Chinese patients with AS was approximately the same as that of healthy controls. Subgroup analysis indicated that patients from North and South China had higher sRANKL levels than controls; the sRANKL levels of patients from South China were higher in the subgroup with a Bath Ankylosing Spondylitis Functional Index (BASFI) of > 4 than those of patients in other subgroups. In terms of duration, patients with AS for > 8 years had higher sRANKL levels than health controls. Other subgroup analyses were conducted by region, language, source of control, age, and Bath Ankylosing Spondylitis Disease Activity Index (BASDAI). In these subgroups, the sRANKL levels were significantly higher in the patients with AS than in healthy controls. The BASFI and BASDAI were sources of heterogeneity. Conclusions The sRANKL levels are higher in Chinese patients with AS, especially among those from South China. sRANKL levels may be positively correlated with the pathogenesis of AS among Chinese patients.

Assessment of the prevalence of osteoporosis among children with hemophilia and its relation to serum 25(oh) vitamin D and serum rankl

Osteoporosis is one of the comorbidities that complicating hemophilia. Recurrent hemarthrosis, chronic arthropathy and immobilization all are factors that make patients with hemophilia prone to this complication. Till recently the major emphasis of the osteoporosis diagnosis is the dual-energy X-ray absorptiometry (DEXA) scan, however, the definition of osteoporosis in pediatrics not only low bone mineral density measured by DEXA scan but also required the presence of clinically significant fracture history. Investigation target is evaluating the osteoporosis prevalence between cases with hemophilia, assessing the risk factors increasing its incidence including diet habits, the severity of hemophilia, duration between the first diagnosis of the disease and enrollment in the study and chronic hepatitis C infection. Also, considering the role of vitamin D deficiency, and another potential indirect mechanism mediated through Receptor activator of NF-Kappa-B, the Receptor activator of NF-Kappa-B ligand (RANK-RANKL) pathway has been suggested in the pathogenesis of bone disease and osteoclastic activity but remains controversial.Thirty-nine hemophilia pediatric patients were recruited from hematology clinic, Cairo university hospital, history taking, and examination were done focusing on the musculoskeletal system and dietetic data. Twenty normal age-matching children enrolled as a control group. DEXA scan results showed normal bone mineral density in 28 patients (71.8%) and osteoporosis in 11 patients (28.2%). The median Z score of patients was -1.40 (-2.2 – -0.6). There was a statistically significant decrease of bone mineral density in patients with hemophilia comparing with a control group with P-value <0.001, also we observed significantly higher serum RANKL in the group of patients with low bone mineral density ensuring the relation between RANKL and osteoclastic activity.

Effect of Pistacia atlantica subsp. kurdica Gum in Experimental Periodontitis Induced in Wistar Rats by Utilization of Osteoclastogenic Bone Markers

The aim of this study was to assess the effect of local application of essential oil of Pistacia atlantica kurdica (EOK) gel in treatment of experimentally induced periodontitis in rats and its effect on osteoclastogenic bone markers. Twenty-four male Wistar rats of 250 to 350 g were used in this study and were allocated into four groups. Control negative (without induced periodontitis), control positive (induced experimental periodontitis left without treatment), treatment control (induced experimental periodontitis and treated with Chlorhexidine gel) and EOK treated group (induced experimental periodontitis treated with EOK gel). The animals were sacrificed after 30 days, and the mandibular central incisor and surrounding tissue were dissected from the mandible and further processed for preparing H&E slides. Inflammatory cells, osteoclast cells, and periodontal ligament (PDL) were examined and measured histologically. Finally, the mean concentrations of both markers, receptor activator of nuclear factor kappa-Β ligand (RANKL) and (Interleukin-1β) IL-1β, were analyzed by ELISA. A significant reduction of inflammatory reaction and osteoclast numbers with improvement of PDL and low mean concentrations of RANKL and IL-1β were seen in the EOK treated group in comparison to the control group and the chlorhexidine group as well. The extract showed a protective effect in the healing of periodontitis that had been induced in rats and decreased bone resorption by down regulation of serum RANKL and IL-1β markers.

Serum Receptor Activator of Nuclear Factor-κβ Ligand and Osteoprotegerin Levels and Ratio in Correlation with Bone Mineral Density

Osteoporosis is a disorder represented by manifestations of low bone mass, decreased bone tissue, and disrupted bonemicroarchitecture. The diagnosis of osteoporosis so far has been based on fracture manifestations after minimal trauma orby detecting low Bone Mineral Density (BMD). Measurement of Receptor Activator of Nuclear Factor-κβ Ligand (RANKL)and Osteoprotegerin (OPG) levels has opened the discourse of a more specific assessment of osteoblast and osteoclastregulation. The RANKL/OPG ratio can represent resorption and bone formation more significantly when correlated withBMD features. This study aimed to analyze the correlation between serum RANKL and OPG levels and ratio with BMD. A totalof 58 post-menopausal females from 13 elderly in Integrated Community Health Care Surabaya and Sidoarjo were enrolled.Data were collected by recording age, onset of menarche, onset of menopause, and Body Mass Index (BMI). Serum RANKLand OPG levels were evaluated using sandwich ELISA from Elabscience®. The RANKL/OPG ratio was obtained from the ratiobetween measured RANKL and OPG levels in serum. The proximal femur and lumbar spine BMDs were measured usingHologic® Discovery™ QDR™ Dual-Energy X-ray Absorptiometry (DEXA). Pearson's correlation test in this study showed nosignificant correlation between BMD and RANKL levels (lumbar: p=0.203; hip: p=0.283). The insignificant result was alsoshown in the correlation between BMD and OPG levels (lumbar: p=0.412; hip: p=0.617). A significant result between lumbarBMD and RANKL/OPG ratio was only found in the osteopenia subjects (p=0.001). The RANKL/OPG ratio had a significantcorrelation only with osteopenia-BMD in post-menopausal females. Therefore, it could be used as supporting data inosteoporosis screening.