Interactions of the Osteokines, Glucose/Insulin System and Vascular Risk Networks in Patients With Newly Diagnosed Type 2 Diabetes (VNDS 15).

BACKGROUND AND AIM: Bone as an endocrine organ regulates metabolic processes independently of mineral metabolism through the production/release of proteins collectively named 'osteokines'. Relevant connections were reported between the insulin/glucose system, calcification of the atherosclerotic plaque, and several osteokines. We aimed to test the hypothesis that the osteokine network could be involved in beta-cell function, insulin sensitivity, and vascular damage in a cohort of people with newly diagnosed type 2 diabetes (T2D). SUBJECTS AND METHODS: In 794 drug-naive, GADA-negative, newly-diagnosed T2D patients (mean ± SD age: 59 ± 9.8 years; BMI: 29.3 ± 5.3 kg/m2; HbA1c: 6.6 ± 1.3%) we assessed: plasma concentration of osteocalcin (OCN), osteopontin (OPN), RANKL, and its putative decoy receptor osteoprotegerin (OPG); insulin sensitivity (SI) by hyperinsulinemic euglycemic clamp; beta cell function (BCF), estimated by OGTT minimal modelling and expressed as derivative (DC) and proportional (PC) control. Echo-doppler of carotid and lower limb arteries were also performed in 708 and 701 subjects, respectively. RESULTS: OCN, RANKL and OPG were significantly associated with PC (p < 0.02); OCN was positively related to DC (p = 0.018). OPG was associated with lower IS (p < 0.001). Finally, the higher RANKL levels, the greater was the severity of atherosclerosis in common carotid artery (p < 0.001). Increased OPG and OPN concentrations were related to subclinical atherosclerosis in peripheral arteries of lower limbs (p = 0.023 and p = 0.047, respectively). CONCLUSION: These data suggest that, in patients with newly diagnosed T2D, the osteokine network crosstalks with the glucose/insulin system and may play a role in modulating the atherosclerotic process.

Effect of RANKL on Lower Depressive Symptoms In Hemodialysis Patients.

Depression and osteoporosis are common diseases in dialysis patients. In addition, patients with osteoporosis are more susceptible to depression. Contrary to previous anti-osteoporosis agents, denosumab and romosozumab could be used in dialysis patients and have similar action mechanisms for blocking RANKL. RANKL causes bone resorption after binding RANKL, but binding with OPG leads to suppress of bone resorption. In recent mice study, inhibition of RANKL with denosumab improved depressive-like phenotype. Besides, it was found that OPG was associated with depression. Therefore, this study aimed to investigate the association of depressive symptoms with RANKL and OPG in hemodialysis patients. We conducted a cross-sectional study with a total of 172 hemodialysis patients. The participants were measured for plasma RANKL, OPG, MMP-2, and MMP-9 levels. Logistic regression analysis was performed to evaluate the effect of RANKL and OPG on the presence of depressive symptoms. The depressive symptoms were observed in 90 (52.3%) subjects. RANKL tertile 3 had negative association with BDI score (ß - 4.527, 95% CI - 8.310 to - 0.743) in univariate analysis, and this association persisted even after multivariate adjustments (ß - 5.603, 95% CI - 9.715 to -1.491) in linear regression. In logistic regression between RANKL tertiles and depressive symptoms, RANKL tertile 3 had significantly lower unadjusted OR (0.40, 95% CI 0.19-0.86), and multivariate-adjusted OR (0.31, 95% CI 0.12-0.82) for depressive symptoms. OPG was not significantly associated with depressive symptoms. Higher plasma RANKL concentrations were significantly associated with lower depressive symptoms in HD patients.Trial registration WHO registry, No. KCT0003281, date: January 12, 2017.

Serum proinflammatory cytokines, receptor activator of nuclear factor kappa-Β ligand (RANKL), osteoprotegerin (OPG) and RANKL/OPG ratio in mild and severe COVID-19.

INTRODUCTION: Osteoporosis, a systemic skeletal disease, is characterized by a quantitative and qualitative, and progressive decrease in bone mass, which is related to inflammation. Since a cytokine storm is triggered in Coronavirus disease 2019 (COVID-19), this study aims to evaluate pro-inflammatory cytokines (TNF-α, IL-1ß), Receptor activator of nuclear factor-κB ligand (RANKL)/serum osteoprotegerin (OPG) ratio, and their relationship in mild and severe COVID-19. METHODS: This study was performed on 48 adult patients (18 mild, 18 severe COVID-19, and 12 healthy subjects as a control group). Serum OPG, RANKL, TNF-α, IL-1ß, 25-OH vitamin D, and ALKp were measured by ELISA and colorimetric assay. RESULTS: COVID-19 patients had a significant increase in RANKL, and RANKL/OPG in mild and severe form (p < 0.001) while OPG decreased significantly in severe form compared to healthy controls (p < 0.05). Inflammatory cytokines (TNF-α and IL-1ß) increased in both groups of patients whereas Alkaline phosphatase (ALKp) increased only in severe patients (p < 0.001). Both groups had 25-OH vitamin D deficiency in comparison to healthy ones (p < 0.001). Pearson's correlation coefficient was performed to determine the relationship between RANKL, OPG, ALKp, and 25-OH vitamin D with TNF-α and IL-1ß in mild and severe COVID-19, which was statistically significant. CONCLUSION: Serum RANKL/OPG ratio was elevated in COVID-19 individuals and is assumed to be a risk factor for BMD reduction and osteoporosis in these patients. Correlations between IL-1ß, TNF-α, ALKp, 25-OH vitamin D, OPG, RANKL, and RANKL/OPG ratio offered the potential role of these proinflammatory markers in the mechanism of osteoporosis in COVID-19 patients.

Analysis of inflammation and bone remodeling of atmospheric plasma therapy in experimental periodontitis.

BACKGROUND AND OBJECTIVE: The biological effects of atmospheric plasma (cold plasma) show its applicability for controlling the etiological factors that involve tissue repair. Thus, the study evaluated the effect of atmospheric plasma therapy in the control of tissue inflammation and bone remodeling in experimental periodontitis. METHODS: Fifty-six rats were subjected to ligation in the cervical region of the first maxillary molars (8 weeks). The animals were divided into two groups (n = 28): periodontitis without treatment group (P group), and periodontitis with atmospheric plasma treatment group (P + AP group). Tissue samples were collected at 2 and 4 weeks after treatment to analyze the inflammation and bone remodeling by biochemical, histomorphometric, and immunohistochemical analyses. RESULTS: Inflammatory infiltration in the gingival and periodontal ligament was lower in the P + AP group than in the P group (p < .05). The MPO and NAG levels were higher in the P + AP group compared to P group (p < .05). At 4 weeks, the TNF-α level was lower and the IL-10 level was higher in the P + AP group compared to P group (p < .05). In the P + AP group, the IL-1ß level increased in the second week and decreased in the fourth week (p < .05), the number of blood vessels was high in the gingival and periodontal ligament in the second and fourth week (p < .05); and the number of fibroblasts in the gingival tissue was low in the fourth week, and higher in the periodontal tissue in both period (p < .05). Regarding bone remodeling, the RANK and RANKL levels decreased in the P + AP group (p < .05). The OPG level did not differ between the P and P + AP groups (p > .05), but decreased from the second to the fourth experimental week in P + AP group (p < .05). CONCLUSIONS: The treatment of experimental periodontitis with atmospheric plasma for 4 weeks modulated the inflammatory response to favor the repair process and decreased the bone resorption biomarkers, indicating a better control of bone remodeling in periodontal disease.

The effect of interleukin-20 on periodontal tissue destruction in individuals with periodontitis.

BACKGROUND AND OBJECTIVE: Periodontitis is an inflammatory disease that destroys periodontal tissues. Interleukin-20 (IL-20), on the other hand, is known as a potent angiogenic, chemotactic, and pro-inflammatory cytokine associated with various chronic inflammatory disorders. IL-20 has a significant role in the regulation of osteoclastogenesis and osteoblastogenesis. This study aimed to evaluate the effect of IL-20 on periodontal destruction. METHODS: In this study, a total of 60 participants were included, 30 of whom were systemically and periodontally healthy (control group), and 30 were systemically healthy but had periodontitis (periodontitis group). Gingival crevicular fluid (GCF) and serum samples were collected from the participants for biochemical analysis. Enzyme-linked immunosorbent assay was used to determine the levels of IL-20, tumor necrosis factor (TNF)-α, IL1ß/IL-10, RANKL/osteoprotegerin (OPG), and matrix metalloproteinase-8 (MMP8). For statistical analysis, the independent t-test, Pearson correlation coefficients, and the Chi-square test were used. RESULTS: GCF IL-20, RANKL, RANKL/OPG, serum IL-20, RANKL, RANKL/OPG, MMP-8, TNF-α, IL-1B, and IL-1ß/IL-10 values were found to be statistically significantly higher in the periodontitis group than in the control group. GCF OPG and serum IL-10 values were found to be statistically significantly higher in the control group than in the periodontitis group. No statistically significant difference was observed between the groups in serum OPG values. A statistically significantly positive correlation was observed between serum IL-20 value and serum RANKL, RANKL/OPG, MMP-8, TNF-α, IL-1ß values, and periodontal clinical parameters. The ROC curves showed: AUC = 0.788 for GCF IL-20, and AUC = 1.000 for serum IL-20. CONCLUSION: According to the results of the study, IL-20 was found to be associated with periodontitis. The role of IL-20 in periodontal pathogenesis is related to osteoclastogenesis and collagen degradation. It is conceivable that IL-20 may increase bone destruction by both affecting the RANKL/OPG ratio and proinflammatory cytokines.

Associations of Bone Mineral Density with RANKL and Osteoprotegerin in Arab Postmenopausal Women: A Cross-Sectional Study.

Background and objective: There is limited information as to the association of several key bone markers with bone mineral density (BMD) in understudied ethnic groups. This study investigated the relationship between circulating levels of osteoprotegerin (OPG) and receptor activator of nuclear factor kappa-Β ligand (RANKL) with BMD in Arab postmenopausal women. Materials and methods: In this cross-sectional study, a total of 617 Saudi postmenopausal women from the Osteoporosis Registry of the Chair for Biomarkers of Chronic Diseases were included. Anthropometric data, BMD, and biochemical data were retrieved from the registry. Participants were stratified into three groups based on T-score; n = 169 with osteoporosis, n = 282 with osteopenia, and n = 166 normal. Analysis of bone markers including RANKL, OPG, osteocalcin, and N-terminal telopeptide (NTx) was completed using commercially available bioassays. Results: The results suggested that OPG was significantly and positively correlated with age in the osteoporosis group (r = 0.29, p < 0.05), while it was inversely correlated with BMD femoral neck left (r = −0.56, p < 0.001) and BMD femoral neck right (r = −0.37, p < 0.05) in the same group. Moreover, RANKL showed a significant inverse correlation with NTx in the osteopenia group (r = −0.37, p < 0.05). Furthermore, the RANKL/OPG ratio had a positive and significant correlation with BMI (r = 0.34, p < 0.05), BMD femoral neck left (r = 0.36, p < 0.05) and BMD femoral neck right (r = 0.35, p < 0.05) in the osteopenia group. By contrast, it showed a significant inverse correlation with waist to hip ratio in the osteoporosis group (r = −0.38, p < 0.05). Multiple regression analysis showed that OPG contributes to BMD variations in the osteopenia group (p = 0.03). Conclusions: In conclusion, changes in circulating levels of RANKL and OPG might be a protective mechanism contrary to the increased bone loss in postmenopausal women.

Administration of zoledronic acid alleviates osteoporosis in HIV patients by suppressing osteoclastogenesis via regulating RANKL expression.

BACKGROUND: Osteoporosis is a common phenomenon in HIV patients on tenofovir treatment, but its underlying mechanisms remain to be explored. METHODS: Quantitative real-time PCR was performed to analyze the expression of miR-302, miR-101, miR-145 and osteoclast-specific genes in the serum of HIV patients treated with tenofovir and ZOL. ELISA was used to evaluate the expression of RANKL, SMAD3 and PRKACB in the serum of these patients. Luciferase assay was carried out to explore the inhibitory effects of miR-302, miR-101 and miR-145 on the expression of PRKACB, RANKL and SMAD3, respectively. Western blot was used to examine the expression of genes involved in NF­κB and JNK signaling pathways. RESULTS: ZOL treatment significantly suppressed the expression of CTx and osteocalcin in HIV patients treated with tenofovir. The BMD loss of HIV patients treated with tenofovir was effectively hindered by ZOL treatment. Mechanistically, the expression of miR-302, miR-101, miR-145, RANKL, SMAD3 and PRKACB in the serum was remarkably activated by ZOL treatment. Luciferase assays showed that miR-302, miR-101 and miR-145 effectively suppressed the expression of PRKACB, RANKL and SMAD3, respectively, through binding to their 3' UTR. Furthermore, ZOL treatment notably restored the normal expression of osteoclast­specific genes while activating NF­κB and JNK signaling pathways. CONCLUSION: The findings of this study demonstrated that administration of ZOL suppressed the expression of RANKL via modulating signaling pathways of miR-101-3p/RANKL, miR-302/PRKACB/RANKL and miR-145/SMAD3/RANKL. Furthermore, down-regulated expression of RANKL by ZOL treatment alleviated osteoporosis in HIV-positive subjects treated with tenofovir.

Physical activity and Mediterranean diet as potential modulators of osteoprotegerin and soluble RANKL in gBRCA1/2 mutation carriers: results of the lifestyle intervention pilot study LIBRE-1.

PURPOSE: Emerging evidence suggests that the progesterone-mediated receptor activator of nuclear factor κB (RANK)/soluble RANK ligand (sRANKL)/osteoprotegerin (OPG) pathway plays an important role in mammary carcinogenesis and is hyperactivated in germline (g)BRCA1/2 mutation carriers. We analyzed the effects of a 3-month intensive lifestyle intervention within the LIBRE-1 study on the serum levels of OPG and sRANKL and hypothesized that the intervention program provides a beneficial impact on the biomarkers by increasing OPG and reducing sRANKL serum concentrations. METHODS: Serum levels of OPG and sRANKL of 49 gBRCA1/2 mutation carriers were quantified using enzyme-linked immunosorbent assays. We used previously collected blood samples from participants of the prospective LIBRE-1 study, who were randomized into an intervention group (IG), increasing physical activity and adherence to the Mediterranean diet (MedD) through supervised sessions from study entry to the first study visit after 3 months and a usual-care control group (CG). Differences in biomarker levels before and after the 3-month intervention were tested within and between study groups. RESULTS: The lifestyle intervention resulted in a significant increase in OPG for participants in both the IG (q = 0.022) and CG (q = 0.002). sRANKL decreased significantly in the IG (q = 0.0464) and seemed to decrease in the CG (q = 0.5584). An increase in the intake of Omega-3 polyunsaturated fatty acids was significantly associated with an increase in OPG (r = 0.579, q = 0.045). Baseline serum levels of sRANKL were a strong predictor for the change of sRANKL in the course of the intervention (ß-estimate = - 0.70; q = 0.0018). Baseline physical fitness (assessed as VO2peak) might predict the change of OPG in the course of the intervention program (ß-estimate = 0.133 pg/ml/ml/min/kg; p = 0.0319; q = 0.2871). CONCLUSION: Findings from this pilot study seem to confirm our hypothesis by showing an increase in OPG and decrease in sRANKL over a 3-month lifestyle intervention and suggest that increased physical activity and adherence to the MedD are potent modulators of the biomarkers OPG and potentially sRANKL.

Lumbar spine bone mineral density and trabecular bone score-adjusted FRAX, but not FRAX without bone mineral density, identify subclinical carotid atherosclerosis.

PURPOSE: Osteoporosis and atherosclerosis share common risk factors. Aim of this study was to test if FRAX (which is an algorithm that can identify subjects at risk of fracture), without or with BMD values, also adjusted for trabecular bone score (TBS) was able to identify subclinical atherosclerosis, evaluated by measurement of carotid intima media thickness (cIMT ≥ 0.9 mm) as compared to DXA values. METHODS: Ninety postmenopausal women underwent DXA measurement and cIMT evaluation. For each patient, the FRAX algorithm for major osteoporotic fracture (M) and for hip fracture (H) without BMD was computed, together with FRAX with BMD and TBS-adjusted FRAX. Serum levels of osteoprotegerin, sRANKL, and interleukin-6 were also measured. RESULTS: There were no differences in anthropometric parameters and cardiovascular risk factors between subjects with cIMT ≥ 0.9 mm (35% of subjects, group A) compared to those with cIMT < 0.9 mm (group B). The prevalence of osteoporosis and FRAX BMD, TBS-adjusted FRAX both for M and H were higher in group A compared to group B. The best ROC curves to identify subjects with a cIMT ≥ 0.9 mm were: lumbar spine T-score, with a threshold of - 2.5 SD (area under the curve, AUC 0.64; p = 0.02) with a sensibility of 50% and a specificity of 76%; TBS-adjusted FRAX H with a sensibility of 50% and a specificity of 72% (AUC 0.64; p = 0.01 with a threshold of 3%). Interleukin-6 positively correlated with FRAX BMD H and M. CONCLUSIONS: FRAX without BMD does not identify subclinical carotid atherosclerosis, while lumbar spine T-score and TBS-adjusted FRAX H similarly detected it with higher specificity for T-score.

Serum levels of osteoprotegerin, RANK-L & vitamin D in different stages of osteoarthritis of the knee.

Background & objectives: Osteoarthritis (OA) is the 11th leading cause of disability in the modern world, but till date, there have been no effective markers for monitoring the progression of OA. The three proteins RANK/RANK-Ligand and Osteoprotegerin (OPG) have been found to be the key regulators of bone metabolism. Interaction of RANK-Ligand with its receptor RANK triggers differentiation of osteoclasts leading to bone resorption. OPG on the other hand is protective as it is expressed by osteoblasts and bind RANKL with higher affinity preventing its interaction with RANK. The levels of these serum proteins are regulated by vitamin D and parathyroid hormones. Therefore, the present study, aimed to study the association of serum RANKL, OPG and vitamin D with disease severity in patients with knee OA. Methods: It was a cross-sectional study where 80 (43 women and 37 men) newly diagnosed subjects with OA knee were recruited. They were classified into four grades based on K-L grading and into two groups as early (grade 1+grade 2) and advanced (grade 3 + grade 4) based on the disease progression. Results: On comparing the biochemical parameters among the four grades decreasing vitamin D levels were seen with increasing severity of knee OA; an increasing trend of RANKL with increase in the severity of OA was seen; OPG was found to be elevated more in the early stages of OA. We also observed a strong association of RANKL/OPG ratio with disease severity. Interpretation & conclusions: Overall the results suggest that OPG may be considered as an early marker of the diseases.

Association of environmental cadmium exposure and bone remodeling in women over 50 years of age.

Chronic cadmium (Cd) toxicity is a significant health concern, and the mechanism of long-term low-dose Cd exposure on bone has not been fully elucidated yet. This study aimed to assess the association between long-term environmental Cd exposure and bone remodeling in women who aged over 50. A total of 278 non-smoking subjects from Cd-polluted group (n = 191) and non-Cd polluted group (n = 87) were investigated. Bone mineral density (BMD), the levels of three bone turnover markers (BTMs), including total procollagen type 1 amino-terminal propeptide (P1NP), collagen type 1 cross-linked C-telopeptide (ß-CTX), bone-specific alkaline phosphatase (BALP), together with serum soluble receptor activator of nuclear factor-κB ligand (sRANKL) and osteoprotegerin (OPG) were determined. Early markers of renal dysfunction were measured as well. Urinary Cd concentrations ranged from 0.41 to 87.31 µg/g creatinine, with a median of 4.91 µg/g creatinine. Age, BMD, T-score, and prevalence of osteoporosis showed no statistical differences among the quartiles of urinary Cd concentrations, while serum levels of P1NP, ß-CTX, and OPG were higher in the upper quartiles. Multivariate linear regression models indicated significantly positive associations of urinary Cd concentration with serum levels of P1NP, ß-CTX, BALP, sRANKL, and OPG. A ridge regression analysis with T-score and the three BTMs, sRANKL, and OPG, adjusted for age and body mass index (BMI), indicated that except for age and Cd exposure, ß-CTX was a predictor of T-score. These findings demonstrated that Cd may directly accelerate bone remodeling. Serum ß-CTX might be an appropriate biochemical marker for evaluating and monitoring Cd-related bone loss. Capsule: Cadmium (Cd) may directly accelerate bone remodeling and serum ß-CTX is a valuable biochemical marker for evaluating Cd-related bone loss.

Inflammatory markers in women with postpartum depressive symptoms.

Postpartum depression (PPD) is a devastating disorder affecting not only more than 10% of all women giving birth, but also the baby, the family, and the society. Compiling evidence suggests the involvement of the immune system in the pathophysiology of major depression; yet, the immune response in perinatal depression is not as well studied. The aim of this study was to investigate the alterations in peripheral levels of inflammatory biomarkers in 169 Swedish women with and without depressive symptoms according to the Edinburgh postnatal depression scale or the M.I.N.I neuropsychiatric interview at eight weeks postpartum. Among the 70 markers analyzed with multiplex proximity extension assay, five were significantly elevated in women with postpartum depressive symptoms in the adjusted LASSO logistic regression analysis: Tumor necrosis factor ligand superfamily member (TRANCE) (OR-per 1 SD increase = 1.20), Hepatocyte growth factor (HGF) (OR = 1.17) Interleukin (IL)-18 (OR = 1.06), Fibroblast growth factor 23 (FGF-23) (OR = 1.25), and C-X-C motif chemokine 1 (CXCL1) (OR 1.11). These results indicate that women with PPD have elevated levels of some inflammatory biomarkers. It is, therefore, plausible that PPD is associated with a compromised adaptability of the immune system.

Circulating Receptor Activator of Nuclear Factor kB Ligand and triglycerides are associated with progression of lower limb arterial calcification in type 2 diabetes: a prospective, observational cohort study.

BACKGROUND: Lower limb arterial calcification is a frequent, underestimated but serious complication of diabetes. The DIACART study is a prospective cohort study designed to evaluate the determinants of the progression of lower limb arterial calcification in 198 patients with type 2 diabetes. METHODS: Lower limb arterial calcification scores were determined by computed tomography at baseline and after a mean follow up of 31.20 ± 3.86 months. Serum RANKL (Receptor Activator of Nuclear factor kB Ligand) and bone remodeling, inflammatory and metabolic parameters were measured at baseline. The predictive effect of these markers on calcification progression was analyzed by a multivariate linear regression model. RESULTS: At baseline, mean ± SD and median lower limb arterial calcification scores were, 2364 ± 5613 and 527 respectively and at the end of the study, 3739 ± 6886 and 1355 respectively. Using multivariate analysis, the progression of lower limb arterial log calcification score was found to be associated with (ß coefficient [slope], 95% CI, p-value) baseline log(calcification score) (1.02, 1.00-1.04, p < 0.001), triglycerides (0.11, 0.03-0.20, p = 0.007), log(RANKL) (0.07, 0.02-0.11, p = 0.016), previous ischemic cardiomyopathy (0.36, 0.15-0.57, p = 0.001), statin use (0.39, 0.06-0.72, p = 0.023) and duration of follow up (0.04, 0.01-0.06, p = 0.004). CONCLUSION: In patients with type 2 diabetes, lower limb arterial calcification is frequent and can progress rapidly. Circulating RANKL and triglycerides are independently associated with this progression. These results open new therapeutic perspectives in peripheral diabetic calcifying arteriopathy. Trial registration NCT02431234.

LPS-induced inflammation disorders bone modeling and remodeling by inhibiting angiogenesis and disordering osteogenesis in chickens.

Inflammation plays a negative role in the growth and development of bone. However, the underlining mechanisms of inflammation caused abnormal bone development and even bone disease are still poorly understood, especially in chickens. In this study, we explored the influence of inflammation on bone formation in broilers for the first time by using lipopolysaccharide (LPS) to establish systemic inflammatory models in chickens with tibia as the research object. The measurements of production and tibial parameters showed an inefficient production performance and lower growth rate in LPS group. We also found a large amount of platelets, inflammatory cells in chickens' blood and higher levels of inflammatory factors in serum after LPS injection, meanwhile, increase in thrombus, chondrocyte nucleolysis, and osteoclasts and a reduction in blood vessels were observed in growth plate through histological observation. The qPCR analysis showed that the mRNA expression levels of NF-κB, TLR4, TF, TPO, and its receptor C-MPL enhanced, while VEGFA was inhibited in LPS group. In addition, in OPG/RANKL system, OPG was decreased while RANKL enhanced. It was also observed that the mRNA levels of MMP-9 and its inducing factor CD147 enhanced in LPS group. The western blot results were basically in consistent with mRNA test. Thus, we infer that inflammation can inhibit bone modeling and remodeling by affecting angiogenesis and osteogenesis, and result in negative effect on bone formation furtherly.

IL-3 Differentially Regulates Membrane and Soluble RANKL in Osteoblasts through Metalloproteases and the JAK2/STAT5 Pathway and Improves the RANKL/OPG Ratio in Adult Mice.

Bone remodeling comprises balanced activities between osteoclasts and osteoblasts, which is regulated by various factors, including hormones and cytokines. We previously reported that IL-3 inhibits osteoclast differentiation and pathological bone loss. IL-3 also enhances osteoblast differentiation and bone formation from mesenchymal stem cells. However, the role of IL-3 in regulation of osteoblast-osteoclast interactions and underlying mechanisms is not yet delineated. In this study, we investigated the role of IL-3 on the regulation of osteoblast-specific molecules, receptor activator of NF-κB ligand (RANKL), and osteoprotegerin (OPG) that modulate bone homeostasis. We found that IL-3 increases RANKL expression at both the transcriptional and translational levels, and it showed no effect on OPG expression in calvarial osteoblasts. The increased RANKL expression by IL-3 induces mononuclear osteoclasts; however, it does not induce multinuclear osteoclasts. Interestingly, IL-3 decreases soluble RANKL by reducing ectodomain shedding of membrane RANKL through downregulation of metalloproteases mainly a disintegrin and metalloproteinase (ADAM)10, ADAM17, ADAM19, and MMP3. Moreover, IL-3 increases membrane RANKL by activating the JAK2/STAT5 pathway. Furthermore, IL-3 enhances RANKL expression in mesenchymal stem cells of wild-type mice but not in STAT5a knockout mice. Interestingly, IL-3 restores RANKL expression in adult mice by enhancing bone-specific RANKL and decreasing serum RANKL. Furthermore, IL-3 increases the serum OPG level in adult mice. Thus, our results reveal, to our knowledge for the first time, that IL-3 differentially regulates two functional forms of RANKL through metalloproteases and the JAK2/STAT5 pathway, and it helps in restoring the decreased RANKL/OPG ratio in adult mice. Notably, our studies indicate the novel role of IL-3 in regulating bone homeostasis in important skeletal disorders.

Effects of Drynaria Total Flavonoid on the Microstructure of the Mandible in Ovariectomized Rats.

BACKGROUND The aim of this study is to investigate the effects of Drynaria total flavonoids (DTF) on mandible microarchitecture, serum estrogen (E2), osteoprotegerin (OPG), and receptor activator of nuclear factor kappa-B ligand (RANKL) levels in an ovariectomy-induced osteoporosis rat model. MATERIAL AND METHODS Thirty female Sprague-Dawley rats were divided into 5 groups (n=6 per group): sham surgery, ovariectomy (OVX), and low-dose, middle-dose, and high-dose DTF. Mandibular osteoporosis was induced by ovariectomy; an equal amount of ovary-sized fat tissue was removed from the sham group. The DTF-treated groups were given DTF gavage at different doses for 12 weeks; the sham and OVX groups were given saline. After the treatment phase, the effects of DTF on the microarchitecture of the mandible were evaluated by measuring bone density, maximum load, morphometric parameters, and histopathological alterations. Serum E2, OPG, and RANKL levels were measured. RESULTS The OVX group showed obvious osteoporosis in the mandible and decreased serum E2 levels and OPG/RANKL ratio. The low-dose group did not show significant improvement in mandibular microstructure. The middle-dose group showed significantly ameliorated osteoporosis. The high-dose group had further improvement in bone microstructures and increase of OPG/RANKL over the middle-dose group. Furthermore, ovariectomy significantly decreased serum E2, but DTF treatment failed to restore serum E2 levels. CONCLUSIONS Ovariectomy can cause significant bone loss in the rat mandible and a decrease in serum E2 and OPG/RANKL. DTF significantly improved the mandibular microstructure and restored OPG/RANKL balance, but it did not restore the decreased serum E2 concentration following ovariectomy.

Fibroblast growth factor 23 and 25(OH)D levels are related to abdominal obesity and cardiovascular risk in patients with polycystic ovarian syndrome.

Fibroblast growth factor 23 (FGF23) and Klotho are extensively studied in relation to bone metabolism and progression of chronic kidney disease. There is very limited information about their role in polycystic ovarian syndrome (PCOS). The aim of the present study was to investigate some bone markers in women with PCOS in relation to obesity and cardiovascular risk. In the study were included 80 patients, divided into three age-matched groups -Non-obese PCOS (n = 40); Obese PCOS (n = 20) and Obese control group (n = 20). Bone marker levels were measured by an enzyme-linked immunosorbent assay. Obese PCOS patients had higher levels of FGF23 and sRANKL, lower levels of 25(OH)D and higher prevalence of vitamin D deficiency compared to non-obese subjects. Patients with abdominal obesity (waist circumference >80 cm) independently of PCOS status had significantly higher levels of FGF23 (112.5 ± 86.5 vs. 73.4 ± 37.9 pg/ml; p = .023) and lower of 25(OH)D (35.8 ± 21.4 vs 47.8 ± 26.5 nmol/l; p = .034). Patients with PCOS at risk of cardiovascular diseases according to AE-PCOS consensus also had increased levels of FGF23 (111.6 ± 84.5 vs. 66.5 ± 35.1 pg/ml; p = .031) and decreased levels of 25(OH)D (31.9 ± 16.8 vs. 47.1 vs 28.4 nmol/l; p = .017) compared to those not at risk. There was no correlation between bone markers and blood glucose levels, insulin resistance or hormonal levels.

Sex Differences in Proatherogenic Cytokine Levels.

BACKGROUND: It has been shown that sex affects immunity, including cytokine production. Given that atherosclerosis is an inflammatory disease promoted by specific cytokines, such as interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α, we aimed at evaluating whether sex could affect the levels of these proatherogenic cytokines in a group of healthy adults. In this analysis, we also included other cytokines and peptides that have been implicated in atherosclerosis development and progression. METHODS: A total of 104 healthy adults were recruited; we measured circulating levels of IL-1ß, IL-6, TNF-α, angiotensins and angiotensin-converting enzyme-2 (ACE2), as well as osteoprotegerin and receptor activator of nuclear factor κB ligand (RANKL). RESULTS: IL-1ß, IL-6, and TNF-α were significantly higher in men as compared to women. They were all associated with testosterone and the testosterone/estradiol ratio. They remained significantly associated with sex (but not with hormones) after being tested for potential confounders. CONCLUSIONS: Sex seems to influence the levels of proatherogenic cytokines. This is consistent not only with sex differences in vulnerability to infections but also with the higher cardiovascular risk exhibited by the male gender as compared to the female gender. Nevertheless, this association is only partly explained by hormone levels.

The correlation between interleukin-34 and bone erosion under ultrasound in rheumatoid arthritis.

Background: Rheumatoid arthritis (RA) is a chronic inflammatory arthropathy characterized by excessive synovial hyperplasia and progressive joint destruction. Pro-inflammatory cytokines play major roles in the regulation of synovial inflammation. The contribution of interleukin-34 (IL-34) in RA pathogenesis has been strongly suggested in clinical studies.Aim: To investigate the correlation between plasma IL-34 and disease parameters in RA patients including disease activity score (DAS28), receptor activator of NF-[Formula: see text]B ligand (RANKL) concentration, synovitis and bone erosions under ultrasound.Methods: 60 RA patients and 20 healthy controls were from Huashan Hospital, patient's medical history, physical examination, laboratory examination and ultrasound data were collected and recorded, respectively. Blood samples of all participants were collected and the levels of IL-34 and RANKL were tested. The levels of IL-34 and RANKL in RA patients were compared with those of healthy controls. Furthermore, the correlation between IL-34, RANKL and disease parameters in RA patients was analyzed.Results: Both plasma levels of IL-34 and RANKL in RA patients were significantly higher than the healthy controls (p < .05). IL-34 was significantly related to disease activity scores (r = 0.43, p = .001); RANKL (r = 0.46, p = .0003) and bone erosions by ultrasound (r = 0.38, p = .002).Conclusions: The plasma IL-34 concentration in RA was significantly higher than the healthy controls and was significantly correlated with RANKL, as well as disease activity score and bone erosions by ultrasound. The IL-34 may be a new biological marker for disease activity and predictor for bone erosions in RA. Targeting IL-34 holds promise in the management of RA and, potentially, other osteoclasts driven diseases (erosive osteoarthritis and psoriatic arthritis for example).

Comparison of serum levels with bone content and gene expression indicate a contradictory effect of kidney transplantation on sclerostin.

In an attempt to clarify the mechanisms of post-transplant bone disease we investigated the bone content and gene expression of several bone-related proteins. After a successful kidney transplant, the content of sclerostin in bone biopsies was found to be increased as measured by immunohistochemistry, multiplex assay, and gene expression despite a concomitant decrease of sclerostin in the serum. The phosphorylation of beta-catenin was increased, confirming Wnt pathway inhibition, an effect accompanied by an increase of the receptor activator of nuclear factor kappa-Β ligand (RANKL) and a decrease of osteoprotegerin protein levels in both serum and bone. Thus, changes in circulating biomarkers after kidney transplantation cannot be easily extrapolated to concomitant changes occurring in the bone. Hence, overall treatment decisions post kidney transplant should not be based on serum biochemistry alone.