Receptor Activator Of Nuclear factor-kappaB Ligand Concentrations Research Articles

The effect of vitamin D on the early clinical manifestations of menopausal syndrome and the production of cytokines, involved in bone remodeling

Introduction. The problem of early manifestations of menopausal syndrome is due to the high prevalence among postmenopausal women, a wide range of clinical manifestations, a sharp decrease in the quality of life, not always sufficient effectiveness of menopausal hormone therapy (MHT) in the presence of restrictions on its use.The aim. To evaluate the dynamics of clinical manifestations of menopausal syndrome and to determine the pathogenetic effects of vitamin D preparation when used in combination with menopausal hormone therapy in women of early postmenopausal age with menopausal syndrome.Materials and methods. 154 women were examined, of which 81 were characterized by the presence of clinical manifestations of menopausal syndrome. All women with menopausal syndrome received menopausal hormone therapy with estradiol and didrogesterone drug for 6 months, while 39 women additionally also took the vitamin D drug. Before and after therapy in patients with menopausal syndrome, the prevalence of symptoms of the Green scale was assessed. Concentrations of 25(OH) D, osteoprotegerin (OPG) and receptor activator of nuclear factor kappa-B ligand (RANKL) were determined in the blood serum of all women.Results and discussion. The use of MHT by women with menopausal syndrome leads to a decrease in the frequency of registration of a number of symptoms of the Green scale (p < 0.05). At the same time, a decrease in the frequency of registration of individual complaints is found only in the group of women who, along with MYT, received a vitamin D preparation (p < 0.05). Taking the vitamin D preparation is accompanied by an increase in its initially reduced serum concentrations to normal values (p < 0.001). The addition of cholecalciferol to the complex therapy of menopausal syndrome ensures the normalization of RANKL levels by reducing its production in the dynamics of treatment (p < 0.05) A decrease in the concentration of RANKL in response to the complex intake of a hormonal drug and cholecalciferol caused higher OPG/RANKL index values in the basic group by the end of therapy than in women of the comparison group (p < 0.01).Conclusions. The obtained effects of vitamin D preparation when prescribed in combination with menopausal hormone therapy (reduction of the frequency of symptoms of menopausal syndrome, reduction of initially elevated levels of RANKL) indicate the expediency of its use in the treatment of early manifestations of menopausal syndrome.

Effect of leukocyte-platelet-rich fibrin (L-PRF) on the rate of orthodontic tooth movement and expression of various biomarkers in gingival crevicular fluid.

To assess the outcome of leukocyte-platelet-rich fibrin (L-PRF) on the rate of maxillary canine retraction and its correlation with the levels of Receptor activator of nuclear factor kappa-B ligand (RANKL), osteoprotegerin (OPG), and RANKL:OPG in the gingival crevicular fluid (GCF) during comprehensive orthodontic treatment. Eighteen females who required all 1st premolars extraction for the correction of their class I bimaxillary protrusion malocclusions were included. The L-PRF plugs were placed in the experimental side 1st premolar extraction sockets. Canine retraction was performed by sliding mechanics. Canine retraction was assessed from the maxillary study models prepared just before the extraction (T0) and then at 1week (T1), 2weeks (T2), 4weeks (T3), and 8weeks (T4) after the 1st premolar extraction and placement of L-PRF plugs. The concentrations of RANKL and OPG in the GCF were evaluated at T0, T1, T2, T3, and T4. In experimental sides, the amount of canine retraction was statistically more during the T0-T1, T1-T2, and T2-T3 periods. The mean concentration of RANKL at T1, T2, and T3 was significantly more in the experimental sides. The mean concentration of OPG was significantly less in the experimental sides at T2, T3, and T4. The RANKL:OPG was significantly more in the experimental sides at T1, T2, T3, and T4. No significant correlation was found between amount of canine retraction and concentration of RANKL and OPG and RANKL to OPG ratio in GCF. The L-PRF accelerated the rate of maxillary canine retraction by 0.28mm over an 8-week period. The L-PRF favored the local osteoclastogenesis by enhancing the RANKL and suppressing the OPG concentrations. There was no significant correlation between the rate of maxillary canine retraction and expression of RANKL, OPG, and RANKL:OPG in GCF. The Clinical Trials Registry of India (Reg. No. CTRI/2020/10/028390, Date-13.10.2020).

Comparative study of the synovial levels of RANKL and OPG in rheumatoid arthritis, spondyloarthritis and osteoarthritis

IntroductionIn chronic arthropathies, there are several mechanisms of joint destruction. In recent years, studies have reported the implication of receptor activator of nuclear factor kappa-B ligand (RANKL) and osteoprotegerin (OPG) in the process of activation and differentiation of osteoclasts, a key cell in the development of bone erosion. The RANKL/OPG ratio is increased in the serum of patients with malignant diseases and lytic bone disease, as well as rheumatoid arthritis (RA). The objective of this study was to measure and compare the concentrations of OPG and RANKL in the synovial fluid (SF) of patients with rheumatoid arthritis, spondyloarthritis (SpA) and osteoarthritis (OA).MethodsThis was an observational and cross-sectional study with 83 patients, 33 with RA, 32 with SpA and 18 with OA, followed up regularly in the outpatient clinics of the Rheumatology Department of the Clinics Hospital of the Ribeirão Preto Medical School-USP. All patients were assessed for indications for arthrocentesis by the attending physicians at the time of SF collection and were evaluated for demographic variables and medication use. Disease activity was assessed in individuals with RA and SpA. The quantification of SF OPG and RANKL levels was performed by ELISA, and the correlations of the results with clinical, laboratory and radiological parameters were assessed.ResultsWe found no statistically significant difference in the RANKL and OPG levels among the groups. Patients with RA showed a positive correlation between the SF cell count and RANKL level (r = 0.59; p < 0.05) and the RANKL/OPG ratio (r = 0.55; p < 0.05). Patients with OA showed a strong correlation between C-reactive protein (CRP) and the RANKL/OPG ratio (r = 0.82; p < 0.05). There was no correlation between the OPG and RANKL levels and markers of inflammatory activity or the disease activity index in patients with RA or SpA.ConclusionWithin this patient cohort, the RANKL/OPG ratio was correlated with the SF cell count in patients with RA and with serum CRP in patients with OA, which may suggest a relationship with active inflammation and more destructive joint disease.

Differentiation of Functional Osteoclasts from Human Peripheral Blood CD14&lt;sup&gt;+&lt;/sup&gt; Monocytes

Osteoclasts (OCs) are bone-resorbing cells that play a pivotal role in skeletal development and adult bone remodeling. Several bone disorders are caused by increased differentiation and activation of OCs, so the inhibition of this pathobiology is a key therapeutic principle.Two key factors drive the differentiation of OCs from myeloid precursors: macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor kappa-B ligand (RANKL). Human circulating CD14+ monocytes have long been known to differentiate into OCs in vitro. However, the exposure time and the concentration of RANKL influence the differentiation efficiency. Indeed, protocols for the generation of human OCs in vitro have been described, but they often result in a poor and lengthy differentiation process. Herein, a robust and standardized protocol for generating functionally active mature human OCs in a timely manner is provided. CD14+ monocytes are enriched from human peripheral blood mononuclear cells (PBMCs) and primed with M-CSF to upregulate RANK. Subsequent exposure to RANKL generates OCs in a dose- and time-dependent manner. OCs are identified and quantified by staining with tartrate acid-resistant phosphatase (TRAP) and light microscopy analysis. Immunofluorescence staining of nuclei and F-actin is used to identify functionally active OCs. In addition, OSCAR+CD14- mature OCs are further enriched via flow cytometry cell sorting, and OC functionality quantified by mineral (or dentine/bone) resorption assays and actin ring formation. Finally, a known OC inhibitor, rotenone, is used on mature OCs, demonstrating that adenosine triphosphate (ATP) production is essential for actin ring integrity and OC function. In conclusion, a robust assay for differentiating high numbers of OCs is established in this work, which in combination with actin ring staining and an ATP assayprovides a useful in vitro model to evaluate OC function and to screen for novel therapeutic compounds that can modulate the differentiation process.

PS-BPB01-7: ANGIOTENSIN II INDUCES AORTIC RUPTURE AND DISSECTION IN OSTEOPROTEGERIN-DEFICIENT MICE

Background: The biological mechanism of action for osteoprotegerin (OPG), a soluble decoy receptor for the receptor activator of nuclear factor kappa-B ligand (RANKL) in the vascular structure, has not been elucidated. The study aim was to determine if OPG affects aortic structural integrity in angiotensin II-induced hypertension. Methods and Results: Mortality was higher (P &lt; 0.0001 by Log-rank test) in 8-week-old male homozygotes of OPG gene knockout (OPG -/- mice given subcutaneous administration of angiotensin II (1,000 ng/kg/min) for 28 days, with an incidence of 21% fatal aortic rupture (Chi-square, 8.024, P = 0.005), and 23% aortic dissection (Chi-square, 4.285, P = 0.038), than in age-matched wild type mice. The absence of OPG was associated with decreased medial and adventitial thickness and increased numbers of elastin breaks as well as with 2.5-fold increased periostin expression and soluble RANKL concentrations. PEGylated human recombinant OPG administration decreased all-causes mortality (-22%, P &lt; 0.001 by Log-rank test), the incidence of fatal aortic rupture (-13%, P = 0.08), and aortic dissection (-23%, P &lt; 0.001) with decreasing numbers of elastin breaks (P = 0.0539) at the supra-renal aorta, periostin expressions (P = 0.0011), and soluble RANKL concentrations (P = 0.0012) in angiotensin II-infused OPG-/- mice. Conclusions: These data suggest that OPG protects against aortic rupture and dissection in Ang II-induced hypertension by inhibiting RANKL activity and periostin expression.

Impact of Osteoprotegerin and RANKL on Non-ST-segment Elevation Myocardial Infarction

Objective: Osteoprotogerin and receptor activator of nuclear factor kappa-B ligand (RANKL) formed an alliance that joining between bone remaking and vascular reforming. The aim of this study is to quantify the blood concentrations of osteoprotogerin and of RANKL in diabetic patients with myocardial infarction and to evaluate their relationship with other perilous factors in reference to healthy subjects. Methods: A total of 76 diabetic patients with myocardial infarction of non-ST-segment raising aged 50 to 60 years and their matched 60 healthy control aged 45 to 55 years were randomly selected from Al-yarmook Hospital and evaluated for inclusion in this case–control study. Blood levels of osteoprotogerin and RANKL were assessed using high-sensitive enzyme-linked immunosorbent assay. Results: The outcomes of this study elucidated increment in the blood concentrations of osteoprotogerin and RANKL in those patients. Apparently, significant positive correlation directed between osteoprotogerin level with age and onset of diabetes. Otherwise, lipid profile, and inflammatory markers exhibited no significant correlation with osteoprotogerin level. Conclusion: Upraised blood levels of osteoprotogerin and RANKL in diabetic patients with non-ST-segment raising myocardial infarction may be of a consequence of diabetic complication on vascular cell.

ETS1 polymorphism rs73013527 in relation to serum RANKL levels among patients with RA.

We previously identified E26 transformation specific sequence 1 (ETS1) rs73013527 single nucleotide polymorphism associated with RA susceptibility and disease activity. In the present study, we aims to further investigate the association between ETS1 rs73013527 and receptor activator of nuclear factor kappa B ligand (RANKL), an index related to bone destruction and was reported to elevate in RA.We determined genotypes of ETS1 rs73013527, serum RANKL concentration, clinical characteristics (disease duration, disease activity score for 28 painful/swollen joints), and laboratory markers (rheumatoid factor, anti-citrullinated protein antibody, anti-keratin antibody, c-reactive protein, erythrocyte sedimentation rate) of 254 RA cases. Univariate and multivariate analysis were employed to explore the association between ETS1 rs73013527 and serum RANKL levels in RA patients.Univariate and multivariate analysis indicated no association of serum RANKL levels with patient age, gender, clinical characteristics, and laboratory markers. Univariate analysis, not multivariate analysis indicated genotype CT/TT of ETS1 rs73013527 was significantly associated with elevated RANKL levels in RA patients.ETS1 rs73013527 is in relation to serum RANKL levels among patients with RA. ETS1 probably might be an indirect factors involved in RANKL regulation in RA.

The inhibitory effect of melatonin on osteoclastogenesis of RAW 264.7 cells in low concentrations of RANKL and MCSF.

RAW 264.7 cells are one of the most recommended cell lines for investigating the activity and differentiation of osteoclasts. These cells differentiate into osteoclasts in the presence of two critical components: receptor activator of nuclear factor kappa B ligand (RANKL) and macrophage colony stimulating factor (MCSF). Melatonin (MEL) hormone has recently become one of the small molecules used in the field of bone regeneration and bone disease treatment, as it has the ability to inhibit the differentiation of osteoclasts directly by suppression of the NF-κB signaling pathway. The main aim of the current study is to determine sufficient RANKL/MCSF concentrations for differentiation of the cells to osteoclasts and to describe the repressive effect of MEL on the osteoclastogenesis of these cells. In this regard, it was found that 10 ng/mL of RANKL- and MCSF-containing medium is suitable for inducing osteoclastogenesis of the cells. In addition, melatonin at doses in the range of 100–1000 µM does not have a cytotoxic effect. Subsequently, results of tartrate resistant acid phosphatase (TRAP) activity, TRAP staining, and relative expressions of cathepsin K, nuclear factor of activated T cells one (NFATC1), and TRAP genes showed a suppressive effect of MEL —especially 800 µM— on RANKL-induced osteoclastogenesis of these cells.

Effects of isoflavones on bone turnover markers in peritoneal dialysis patients: a randomized controlled trial.

The aim of this study was to investigate the effects of soy isoflavones on serum markers of bone formation and resorption in peritoneal dialysis (PD) patients. In this randomized, double-blind, placebo-controlled trial, 40 PD patients were randomly assigned to either the soy isoflavone or the placebo group. The patients in the soy isoflavone group received 100mg soy isoflavones daily for 8weeks, whereas the placebo group received corresponding placebos. At baseline and the end of the 8th week, 7ml of blood was obtained from each patient after a 12- to 14-h fast and serum concentrations of bone formation markers (osteocalcin and bone alkaline phosphatase), bone resorption markers [N-telopeptide and receptor activator of nuclear factor kappa B ligand (RANKL)], and osteoprotegerin as an inhibitor of bone resorption were measured. Serum N-telopeptide concentration decreased significantly up to 27% in the soy isoflavone group at the end of week 8 compared to baseline (P = 0.003). Also, serum RANKL concentration reduced significantly up to 17% in the soy isoflavone group at the end of week 8 compared to baseline (P = 0.03). These bone resorption markers did not significantly change in the placebo group during the study. There were no significant differences between the two groups in mean changes of serum osteocalcin, bone alkaline phosphatase, and osteoprotegerin. This study indicates that daily administration of 100mg soy isoflavone supplement to PD patients reduces serum N-telopeptide and RANKL which are two bone resorption markers. CLINICALTRIALS.GOV: NCT03773029, 2018.

PS1102 HEMOPHILIA ASSOCIATED BONE PATHOLOGY IMPACTS QUALITY OF LIFE

Background:People with Hemophilia (PwH) have decreased skeletal health resulting in lower bone mineral density (BMD) and greater risk of fracture. We previously identified the cytokines c‐terminal telopeptide (CTX)‐1, receptor activator of nuclear factor kappa‐B ligand (RANKL), and osteopretegerin (OPG) as critical markers for bone metabolism in hemophilia.Aims:This study investigates the relationship between hemophilia‐associated bone pathology, factors including cytokines, factor VIII (FVIII) replacement therapy, and quality of life.Methods:Twelve participants with severe FVIII‐deficiency were enrolled in the single site IRB‐approved study. After a 72‐hour washout, plasma samples were obtained at 8 time points over the course of five days with FVIII infusions administered on Day 1 (50 units/Kg) and Day 3 (20 units/Kg). All participants had a physical examination and completed questionnaires on Day 1. The EQ‐5D‐3L was used to report participants’ self‐rated health. The Hemophilia Activities List (HAL) self‐reported physical activity. The International Society on Thrombosis and Haemostasis Bleeding Assessment Tool (ISTH‐BAT). BMD was measured as Z‐scores via Dual‐Energy X‐ray Absorptiometry (DEXA) Scanning. Joint assessment was performed using the Hemophilia Joint Health Score (HJHS). Plasma cytokine levels were measured using ELISA and magnetic bead multiplex kits.Results:Serum OPG concentrations increased throughout the study period (p = 7.26x10–5) while RANKL concentrations did not change (p = 0.19). The OPG:RANKL ratio exhibited a 6.2‐fold increase between Days 1 and 5. CTX‐1 concentrations were lower (p = 0.006) 30 minutes after the 50 U/kg FVIII infusion. CTX‐1 response for each participant was assessed using linear regression throughout the time course. Spine L1‐L4 Z‐score (SZ) and Hip Total Z‐score (HTZ) correlate with CTX‐1 response (p = 3.4x10–4 and 0.014, respectively). The mean age of participants was 25.2 ± 2.1 at time of first study visit. Participants had a mean SZ of −0.74 ± 0.34, HTZ of −0.17 ± 0.32, Hip Neck Z‐score (HNZ) of −0.16 ± 0.35, and HJHS of 20 ± 5. HTZ and HNZ decreased significantly with patient age (p = 0.027 and 0.032) while SZ and HJHS did not (p = 0.18 and 0.16). Consequently, age correction was applied to BMD comparisons. Statistical comparisons between HAL, EQ‐5D‐3L, and BHQ answers, BMD data, and HJHS are shown in Table 1.Summary/Conclusion:This prospective study demonstrates a relationship between FVIII deficiency and bone disease in PwH. OPG acts as a decoy ligand to RANKL, inhibiting osteoclastic bone resorption. The observed increase in OPG:RANKL ratio suggests FVIII has a direct impact on this pathway. Decreased CTX‐1 concentrations following factor infusion and correlations between CTX‐1 response and BMD further suggests that FVIII plays a role in moderating bone remodeling. Hemophilia‐associated bone and joint pathology is associated with decreased quality of life. Correlations between BMD and questionnaire responses demonstrate that poor bone and joint health is physically limiting, causes discomfort, and negatively impacts perceptions of personal well‐being. Furthermore, correlations between BMD and HJHS and ISTH‐BAT responses (Table 1) illustrate a relationship between bleeding management and skeletal health. PwH with lower BMD report more nose and gum bleeds, suggesting a link between FVIII replacement and skeletal health. Bleeds in the extremities were not correlated with decreased BMD or increased HJHS, but abdominal (stomach, iliopsoas) and other bleeds were. Analysis of these data is ongoing.image

Eimeria challenge adversely affected long bone attributes linked to increased resorption in 14-day-old broiler chickens

ABSTRACTThere is limited information on the effects of enteric pathogen on bone quality in rapidly growing broiler chicks. We examined tibia and femur attributes (length, diameter, relative weight of ash content [AC] to the BW, ash concentration [AP]) and serum bone-turnover markers including receptor activator of nuclear factor kappa-B ligand (RANKL) for resorption, alkaline phosphatase (ALP) for mineralization, and selected serum metabolites in 14-day-old broilers challenged with Eimeria. A total of 160 (80 males and 80 females) 1-day-old Ross × Ross 708 chicks were used. Based on BW, birds were placed within sex in cages (5 birds per cage) and fed chick starter diets to day 9 of age. On day 9, half of the cages were orally gavaged with 1 mL of Eimeria culture (100,000 oocysts of E. acervulina and 25,000 oocysts of E. maxima) and the other half (unchallenged control) received 1 mL 0.9% saline in distilled water. On day 14, 2 birds were randomly selected and necropsied for intestinal lesion score, blood, tibia, and femur samples. Data were analyzed in a 2 (challenged vs. unchallenged) × 2 (males vs. females) factorial arrangement. There was no interaction (P > 0.05) between Eimeria and sex on any measurement. Whereas there were no intestinal lesions in unchallenged birds, Eimeria resulted in lesion score (0 to 4) of 3.35, 2.59 and 0.11 in duodenum, jejunum and ileum, respectively. Eimeria challenge decreased (P < 0.05) tibia AC and AP by 10 and 8.2%, respectively but had no (P > 0.10) effect on femur attributes. Generally, males showed (P < 0.05) longer and wider bones with more AC compared with the female. Circulating serum RANKL concentration increased (P = 0.017) in response to Eimeria challenge and was negatively correlated with tibia AC (–0.731; P = 0.021). Our findings showed that Eimeria damage to the intestinal physiology had adverse effects on long bone attributes linked to increased resorption.

Abstract P6-09-01: In primary, non-metastatic breast cancer patients, increased serum levels of RANKL significantly correlate with tumor cell spread to the bone and the occurrence of bone metastasis whereas high levels of its soluble decoy receptor osteoprotegerin predict poor survival

Abstract Background: Receptor activator of nuclear factor kappa-B ligand (RANKL) is an essential protein for osteoclast regulation and its activity is controlled by its soluble decoy receptor osteoprotegerin (OPG). RANKL has been associated with benign as well as malignant bone disease and there is increasing evidence that RANKL may also directly affect breast cancer (BC) progression and metastasis to the bone. Here we assessed serum concentrations of RANKL and OPG in 509 patients with primary, non-metastatic BC and correlated the results with clinical parameters including the presence of disseminated tumor cells (DTCs) in the bone marrow (BM), survival and the risk of developing metastatic disease. Patients and Methods: Patients with first diagnosis of BC between Aug 2006 and Dec 2009 were included in our study. BM sampling was performed before surgery in an adjuvant setting and two BM aspirates were analyzed for DTCs using density centrifugation followed by immunocytochemistry applying the pan-cytokeratin antibody A45-B/B3. Blood was collected from each patient and sRANKL and OPG levels in the serum were measured by ELISA (Biomedica, Vienna, Austria). Results: Mean serum values for RANKL and OPG were 0.23±0.20 pmol/l and 4.24±1.70 pmol/l, respectively. RANKL levels were significantly lower in women above the age of 60 (p&amp;lt;0.0001) and RANKL/OPG ratios were higher in patients with lymph node involvement (p&amp;lt;0.05). High OPG levels were associated with a higher risk of death from BC (HR 1.94 95%CI 1.23-3.07; p=0.005) and multivariate analyses revealed OPG to be an independent prognostic marker for BC specific survival (p=0.035). DTCs were detected in 207/507 (41%) patients and RANKL levels were 33% higher in DTC-positive patients (p&amp;lt;0.0001). Interestingly, in DTC-negative patients, high RANKL levels were associated with a significantly better BC specific survival compared to low levels (HR 0.524; 95%CI 0.30-0.95; p=0.04). RANKL serum levels were significantly enhanced in patients that developed bone metastases (p=0.01) and patients in the highest quartile of RANKL had a significantly increased risk of developing bone metastases compared to those in the lowest RANKL quartile (HR 4.62, 95%CI 1.49-14.34, p=0.03). Conclusion: Increased OPG levels indicated poor survival and high RANKL significantly associated with an increased risk of developing bone metastases while indicating a positive prognostic marker in DTC-negative patients. These findings warrant further investigation as it may provide a rational for novel diagnostic or therapeutic approaches. Citation Format: Bittner A-K, Goebel A, Hoffmann O, Rauner M, Hofbauer LC, Kimmig R, Kasimir-Bauer S, Rachner TD. In primary, non-metastatic breast cancer patients, increased serum levels of RANKL significantly correlate with tumor cell spread to the bone and the occurrence of bone metastasis whereas high levels of its soluble decoy receptor osteoprotegerin predict poor survival [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P6-09-01.

Association between low bone mass and the serum RANKL and OPG in patients with nephrolithiasis

BackgroundNephrolithiasis is a risk factor for Osteopenia and osteoporosis. Receptor activator of nuclear factor kappaB ligand (RANKL) and osteoprotegerin (OPG) regulate bone remodeling and osteoclastogenesis. This study aimed to evaluate the relation between serum OPG, RANKL concentration, and bone mineral density (BMD) in patients with kidney stone disease.MethodsForty-four nephrolithiasis patients with either low bone mass or normal BMD (considered control group) were enrolled in this study. BMD was measured at lumbar spine (L1-L4) and femoral neck by dual-energy X-ray absorptiometry (DEXA). The serum OPG and RANKL were determined using the ELISA method.ResultsThe median levels of serum OPG were significantly higher in nephrolithiasis patients with low bone mass compared to the nephrolithiasis patients with normal BMD (3.9 pmol/l versus 3.1 pmol/l; P = 0.03), respectively. Negative correlation was detected between bone densities of femoral neck and OPG in patients with nephrolithiasis (r = −.0344, P = 0.02).ConclusionThe present study showed that high serum fasting OPG levels may be indicative of femoral neck BMD in patients with nephrolithiasis.

Cytokine levels in postmenopausal women depending on individual VDR, COL1A1, LRP5 gene polymorphisms

Objective: to study pro- and anti-infl ammatory cytokines serum levels at postmenopausal osteoporosis and their changes depending on separate polymorphic options of VDR, COL1A1 and LRP5 genes. Materials and methods: 180 postmenopausal women (60,0±0,77 years) were examined. Th e osteodensitometry was carried out by dual-energy x-ray absorptiometry method. Detection of 283 A>G (BsmI, rs1544410) polymorphisms of VDR gene, -1997 C>A (rs1107946) and 1546 (6252) G>T (Sp1 S>s, rs1800012) of COL1A1 gene, 3989 C>T (Ala1330Val, rs3736228) and 1999 G>A (Val667Met, rs4988321) of LRP5 gene was carried out by real-time PCR method. Enzyme-linked immunosorbent assay was used for defi nition of serum levels of interleukin (IL) -1β, -4, -6, -8, -10, -17A, tumor necrosis factor alpha (TNF-α), interferon gamma (INF-γ), osteoprotegerin (OPG), receptor activator of nuclear factor kappa-B ligand (RANKL). Results : it was established that women with postmenopausal osteoporosis had (P<0,05) increased serum IL-1-β, IL-8, IL-17A, TNF-α, RANKL concentration and decreased IL-4, IL-10 levels. Patients with GG genotype of rs1544410 polymorphism had also increased (P<0,05) IL-1-β, RANKL levels, OPG/RANKL index and decreased IL-10 values. Women with GT or TT genotypes of rs1800012 polymorphism of COL1A1 gene had (P<0,05) decreased IL-10, OPG values, OPG/RANKL index, but persons with CA or AA genotypes of rs1107946 polymorphism of COL1A1 gene had increased IL-4, IL-6, IL-8, IL-10, RANKL levels. IL-10 concentration decrease (P<0,05) was established for patients with GA genotype of rs4988321 polymorphism and CT or TT genotypes of rs3736228 polymorphism of LRP5 gene. Conclusion : the obtained data refl ect important pathogenetic aspects of postmenopausal osteoporosis and can be used for development of individualized treatment-and-prophylactic actions for women.

Effects of loxoprofen on the apical root resorption during orthodontic tooth movement in rats.

Studies have revealed that severe apical root resorption during tooth movement is caused by the noninfective inflammatory reaction of apical root tissues. We hypothesized that loxoprofen can suppress apical root resorption during tooth movement. Cyclic tensile force (CTF) of 10 kPa was applied to the human pulp cells for 48 hours by the Flexcell Strain Unit. Loxoprofen (10 and 100 μM) was added to the culture cells, and expression of cyclooxygenase (COX)-1, COX-2, interleukin (IL)-1β, receptor activator of nuclear factor kappa-B ligand (RANKL), tumor necrosis factor (TNF)-α, and macrophage colony-stimulating factor (M-CSF) were examined. To determine the effects of loxoprofen sodium on apical root reabsorption during tooth movement, the upper first molars of 7-week-old rats were subjected to mesial movement by 10g force for 30 days with or without the oral administration of loxoprofen. Gene expression and protein concentration of COX-1, COX-2, IL-1β, TNF-α, RANKL and M-CSF were significantly higher in the CTF group than in the control group. However, these levels were decreased by loxoprofen administration. After orthodontic tooth movement, the expression of IL-1β, TNF-α, RANKL and M-CSF decreased in the loxoprofen group than in the control group by immunohistochemical staining. In comparison to control group, less number of odontoclasts and a decrease in the amount of apical root resorption was observed in the loxoprofen group. Many osteoclasts became visible on the pressure side of the alveolar bone in the both groups, and the amount of tooth movement did not show a significant difference. These findings demonstrate that severe apical root resorption may be suppressed by loxoprofen administration, without a disturbance of tooth movement.

Receptor activator of nuclear factor kappa-B ligand (RANKL) but not sclerostin or gene polymorphisms is related to joint destruction in early rheumatoid arthritis

The aim of this study was to analyze relationships between receptor activator of nuclear factor kappa-B (RANKL), sclerostin and their gene polymorphisms with radiological progression in patients with early rheumatoid arthritis (RA). Patients with early RA (n = 407, symptomatic <1 year) (ARA criteria) examined radiologically at inclusion and after 24 months were consecutively included. Disease activity score and C-reactive protein were regularly recorded. Sclerostin, RANKL, and anti-CCP2 antibodies were analyzed in plasma at baseline using ELISAs. Data on gene polymorphism for sclerostin and RANKL were extracted from Immunochip analysis. Sex- and age-matched controls (n = 71) were identified from the Medical Biobank of Northern Sweden. The concentration of RANKL was significantly higher in patients compared with controls, median (IQR) 0.56 (0.9) nmol/L and 0.20 (0.25) nmol/L (p < 0.001), and in anti-CCP2-positive patients compared with sero-negative individuals. Sclerostin was significantly increased in female patients 0.59 (0.47–0.65) ng/mL compared with female controls 0.49 (0.4–0.65) ng/mL (p < 0.02). RANKL concentration was related to the Larsen score at baseline (p < 0.01), after 24 months (p < 0.001), and to radiological progression at 24 months (p < 0.001). Positivity of RANKL and anti-CCP2 yielded significant risk for progression with negativity for both as reference. No single nucleotide polymorphism encoding TNFSF11 or SOST was associated with increased concentrations of the factors. The concentration of RANKL was related to the Larsen score at baseline, at 24 months, and radiological progression at 24 months particularly in anti-CCP2-positive patients, while the concentration of sclerostin was unrelated to radiological findings.

Osteoprotegerin and osteoprotegerin/TRAIL ratio are associated with cardiovascular dysfunction and mortality among patients with renal failure

PurposeThe high prevalence of cardiovascular morbidity and mortality among patients with chronic kidney disease (CKD) is observed especially in those undergoing dialysis. Osteoprotegerin (OPG) and its ligands, receptor activator of nuclear factor kappa-B ligand (RANKL) and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) have been associated with cardiovascular complications. Our aim was to study their role as cardiovascular risk factors in stage 5 CKD patients. Patients and methodsOPG, RANKL and TRAIL concentrations were measured in 69 hemodialyzed CKD patients and 35 healthy volunteers. In CKD patients, cardiovascular dysfunction was assessed with aortic pulse wave velocity (AoPWV), carotid artery intima-media thickness (CCA-IMT), coronary artery calcium score (CACS) and N-terminal pro-B-type natriuretic peptide (NT-proBNP) serum concentrations. Cardiovascular and overall mortality data were collected during a 7-years follow-up. ResultsOPG plasma concentrations were higher in CKD patients comparing to controls. Total soluble RANKL was lower and OPG/RANKL ratio higher in patients. Soluble TRAIL concentrations did not differ between the groups and OPG/TRAIL ratio was higher in CKD patients. OPG and OPG/TRAIL positively predicted long-term mortality (all-cause and cardiovascular) in CKD patients. OPG positively correlated with AoPWV, CCA-IMT and NT-proBNP whereas OPG/TRAIL with AoPWV and NT-proBNP. Described relationships were independent of classical and non-classical cardiovascular risk factors, with exception of age. ConclusionsOur study confirmed the role of OPG as a biomarker of cardiovascular dysfunction and a predictor of mortality in stage 5 CKD. OPG/TRAIL ratio can be proposed as a predictor of cardiovascular dysfunction and mortality.

Influence of Hormone Replacement Therapy on Osteoprotegerin and Receptor Activator of Nuclear Factor Kappa-B Ligand Concentrations in Menopausal Women

The study was designed to compare the levels of the receptor activator of nuclear factor kappa-B ligand (RANKL) and osteoprotegerin (OPG) in the serum and saliva of menopausal women and verify whether there is a correlation between concentrations of these cytokines and bone mineral density (BMD) of the femur and mandible. Blood and saliva samples were collected from a group of 60 women during menopause--30 untreated (control group) and 30 treated with hormone replacement therapy (HRT) (research group). Densitometry was conducted on the femoral bone and mandible. BMD in the research group was significantly higher than in the control group. The OPG concentration was slightly higher in the research group. The concentration of RANKL was slightly higher in the control group. Differences in RANKL/OPG between groups were not high enough to be statistically significant. Analysis of the RANKL in saliva revealed a significantly higher concentration of this factor in the control group. Observed beneficial effects of HRT on bone tissue may be exerted through the RANKL/OPG ratio, but further research is required to clearly confirm this thesis.

The effect of ferric ion on the differentiation of osteoclast and bone resorption

To explore the effects of ferric ion on the differentiation from both RAW264.7 and bone marrow macrophages to osteoclast in vitro and bone resorption in vivo. In the presence of 50 ng/ml receptor activator of nuclear factor kappa-B ligand (RANKL), RAW264.7 was treated with ferric ammonium citrate (FAC). The formation of osteoclast was observed by staining of tartrate-resistant acid phosphatase (TRAP) and the TRAP positive cell counted. The expression levels of TRAP, cathepsin-K, nuclear factor of activated T cells c1 (NFATc1) and (receptor activator of NF-κB) RANK were measured by reverse transcription-polymerase chain reaction (RT-PCR).The control and iron overload groups were established by the intraperitoneal injection of normal saline and FAC. In vivo imaging system was employed to determine the bone density of femoral midportion and the fourth lumbar vertebra. After that, the bone marrow cells of femurs were used for osteoclast culture. The serum levels of ferritin, TRAP-5b, RANKL, osteoprotegerin (OPG) and C-terminal cross-linking telopeptide (CTX) were measured by enzyme-linked immunosorbent assay (ELISA). Ferric ion could stimulate the formation of TRAP positive cells in a dose-dependent manner. The expression levels of TRAP, cathepsin-K and NFATc1 in the FAC treated group were significantly higher those of the control group (P < 0.05) while the expression of RANK showed no statistical difference among these groups (P = 0.967). The bone marrow density of femoral midportion and the fourth lumbar vertebra of the iron-overload group decreased significantly versus the control group. The concentrations of ferritin, TRAP-5b, RANKL and CTX of the iron overload group were markedly higher than those of the control group (P < 0.05). Moreover, the concentration of ferritin showed a positive correlation with TRAP-5b and CTX respectively in the iron-overload group (r = 0.65, r = 0.76, P < 0.05). But no significant differences existed in the concentration of OPG for two groups (P > 0.05). Ferric ion may enhance the differentiation of osteoclast in vitro as well as bone resorption in vivo.

SRANKL/osteoprotegerin complex and biochemical markers in a cohort of male and female hemodialysis patients

The processes involved in bone remodeling are under the control of a multitude of systemic and local factors. Receptor activator of nuclear factor-kappaB ligand (RANKL)/osteoprotegerin (OPG) complex seems to be one of the major modulators of bone remodeling. In chronic renal failure, the cytokine systems involved in the regulation of bone turnover may be influenced, and are therefore likely to contribute to the pathogenesis of renal bone disease. The aim of the present study was the evaluation of RANKL/OPG complex in concert with other biochemical parameters in hemodialysis (HD) patients and the investigation of possible correlations between the serum levels of its components and several clinical parameters of these patients. We measured serum levels of intact PTH (iPTH), total serum RANKL (sRANKL), osteoprotegerin (OPG), alkaline phosphatase, osteocalcin (OC), and tartrate-resistant acid phosphatase (TRAP) in 104 HD patients and in 40 healthy controls. The average serum OPG level was significantly higher, whereas the average serum concentration of RANKL was nonsignificantly lower in patients on HD therapy than in age-matched healthy controls. Consequently, the mean sRANKL/OPG ratio was significantly lower in patients. Among HD patients, serum level of OPG increased significantly with aging and with a longer duration of hemodialysis. RANKL levels were inversely correlated with age nonsignificantly in the whole group of patients and significantly in the female subgroup (r=-0.322, p=0.035), whereas RANKL/OPG ratio declined significantly with age in the entire cohort of patients (r=-0.259, p=0.008). In addition, iPTH, OC, TRAP were significantly higher in female, whereas RANKL/OPG ratio was significantly higher in male than female patients. Lower values of sRANKL/OPG ratio in HD patients, as well as the age and duration of HD dependent increase of serum OPG and the age-dependent decrease of sRANKL concentration especially in women cannot be explained by the elimination of renal clearance only. Alterations in sRANKL/OPG ratio might reflect a compensatory mechanism to modulate bone remodeling in these patients.