Effects of stepwise administration of osteoprotegerin and parathyroid hormone-related peptide DNA vectors on bone formation in ovariectomized rat model.

Osteoporosis is a metabolic bone disease that impairs bone mineral density, microarchitecture, and strength. It requires continuous management, and further research into new treatment options is necessary. Osteoprotegerin (OPG) inhibits bone resorption and osteoclast activity. The objective of this study was to investigate the effects of stepwise administration of OPG-encoded minicircles (mcOPG) and a bone formation regulator, parathyroid hormone-related peptide (PTHrP)-encoded minicircles (mcPTHrP) in osteoporosis. The combined treatment with mcOPG and mcPTHrP significantly increased osteogenic marker expression in osteoblast differentiation compared with the single treatment groups. A model of postmenopausal osteoporosis was established in 12-week-old female rats through ovariectomy (OVX). After 8 weeks of OVX, mcOPG (80 µg/kg) was administered via intravenous injection. After 16 weeks of OVX, mcPTHrP (80 µg/kg) was injected once a week for 3 weeks. The bone microstructure in the femur was evaluated 24 weeks after OVX using micro-CT. In a proof-of-concept study, stepwise treatment with mcOPG and mcPTHrP on an OVX rat model significantly improved bone microstructure compared to treatment with mcOPG or mcPTHrP alone. These results suggest that stepwise treatment with mcOPG and mcPTHrP may be a potential treatment for osteoporosis.

Correlates of Circulating Osteoprotegerin in Women with a Pathogenic or Likely Pathogenic Variant in the BRCA1 Gene.

BACKGROUND: Lower levels of osteoprotegerin (OPG), the decoy receptor for receptor activator of NFκB (RANK)-ligand, have been reported among women with a BRCA1 mutation, suggesting OPG may be marker of cancer risk. Whether various reproductive, hormonal, or lifestyle factors impact OPG levels in these women is unknown. METHODS: BRCA1 mutation carriers enrolled in a longitudinal study, no history of cancer, and a serum sample for OPG quantification, were included. Exposure information was collected through self-reported questionnaire at study enrollment and every 2 years thereafter. Serum OPG levels (pg/mL) were measured using an ELISA, and generalized linear models were used to assess the associations between reproductive, hormonal, and lifestyle exposures at the time of blood collection with serum OPG. Adjusted means were estimated using the fully adjusted model. RESULTS: A total of 701 women with a median age at blood collection of 39.0 years (18.0-82.0) were included. Older age (Spearman r = 0.24; P < 0.001) and current versus never smoking (98.82 vs. 86.24 pg/mL; Pcat < 0.001) were associated with significantly higher OPG, whereas ever versus never coffee consumption was associated with significantly lower OPG (85.92 vs. 94.05 pg/mL; Pcat = 0.03). There were no other significant associations for other exposures (P ≥ 0.06). The evaluated factors accounted for 7.5% of the variability in OPG. CONCLUSIONS: OPG is minimally influenced by hormonal and lifestyle factors among BRCA1 mutation carriers. IMPACT: These findings suggest that circulating OPG levels are not impacted by non-genetic factors in high-risk women.

Exploring the potential mechanism of Heng-Gu-Gu-Shang-Yu-He-Ji therapy for osteoporosis based on network pharmacology and transcriptomics.

ETHNOPHARMACOLOGICAL RELEVANCE: Heng-Gu-Gu-Shang-Yu-He-Ji (Osteoking, OK) is a well-known formula for fracture therapy. In clinic, OK is effective in treating fractures while alleviating osteoporosis (OP) symptoms. However, active components of OK and the associated molecular mechanisms remain not fully elucidated. AIM OF THE STUDY: This study aims to systematically evaluate the anti-osteoporosis efficacy of OK and for the first time combine network pharmacology with high-throughput whole gene transcriptome sequencing to study its underlying mechanism. MATERIALS AND METHODS: In this study, the osteoporosis model was established by the castration of both ovaries. The level of serum bone turnover factor was detected by enzyme-linked immunosorbent assay. Micro-CT and HE staining were used to observe the changes of bone histopathology, and nano-indentation technique was used to detect the biomechanical properties of rat bone. The main active Chemical components of OK were identified using UPLC-DAD. Efficacy verification and mechanism exploration were conducted by network pharmacology, molecular docking, whole gene transcriptomics and in vivo experiments. RESULTS: In our study, OK significantly improved bone microarchitecture and bone biomechanical parameters in OVX rats, reduced osteoclast indexes such as C-telopeptide of type I collage (CTX-I) and increased Osteoprotegerin (OPG)/Receptor activator of NF-κB ligand (RANKL) levels. Mechanistically, PI3K/AKT pathway was a common pathway for genome enrichment analysis (KEGG) of both network pharmacology and RNA-seq studies. G protein-ß-like protein (GßL), Ribosomal-protein S6 kinase homolog 2 (S6K2), and Phosphoinositide 3-kinase (PI3K) appeared differentially expression in the PI3K-AKT signaling pathway. These results were also confirmed by qRT-PCR and immunohistochemistry. CONCLUSIONS: OK may be used to treat osteoporosis, at least partly by activating PI3K/AKT/mTORC1 signaling pathway.

Downregulation of osteoprotegerin in colorectal cancer cells promotes liver metastasis via activating tumor-associated macrophage.

Osteoprotegerin (OPG) is a secreted cytokine that functions as a decoy receptor for receptor activator of nuclear factor kappa-B (RANK) ligand (RANKL). Anti-RANKL treatment for bone metastasis has been widely accepted for solid tumors. However, the mechanism of OPG-RANKL-RANK signaling in systemic colorectal cancer (CRC) metastasis remains unclear. In this study, we investigated the relevance and function of OPG expression in CRC liver metastasis. First, we performed in silico analysis using The Cancer Genome Atlas public database and found that lower OPG expression in CRC was associated with poor overall survival. Immunohistochemistry analyses using resected specimen from patients with CRC in our institute confirmed the result. Patient-matched primary CRC and liver metastases showed a significant downregulation of OPG expression in metastatic lesions. In CRC cell lines, OPG expression did not suppress cell proliferation and migration. However, OPG expression inhibited macrophage migration by suppressing the RANKL-RANK pathway. Moreover, in vivo mouse liver metastasis models showed that OPG expression in CRC cells suppressed liver metastases. In addition, treatment with an anti-RANKL neutralizing antibody also suppressed liver metastases. These results showed that downregulation of OPG expression in CRC cells promotes liver metastasis by activating tumor-associated macrophage, which can become a candidate for targeted therapy with anti-RANKL neutralizing antibody for CRC liver metastasis.

Cortical microarchitecture and remodeling-associated gene expression related to oral cancer prognosis.

The objective of this study was to assess the remodeling-associated gene expression in the mandible of patients diagnosed with oral squamous cell carcinoma (OSCC), investigating the cortical microarchitecture, and their influence on disease-free survival (DFS) and overall survival (OS) rates. A total of twenty-four patients who underwent mandibulectomy for OSCC treatment had two bone fragments harvested from the mandible for gene expression (RANK, RANKL, OPG, and SOST), and microarchitecture analysis, including bone volume, surface, mineral density, degree of anisotropy, and fractal dimension. The prognosis of the patients was assessed. The results revealed that RANK, RANKL, and SOST were predominantly downregulated, while OPG was completely downregulated. Tumors located adjacent to the posterior region of the mandible (p = 0.02), with a bone mineral density below 1.03 g/cm3 HA (p = 0.001), and a bone volume less than 86.47% (p = 0.03) were associated with poor outcomes. In conclusion, bone-remodeling-associated genes exhibited downregulation in the cortex of the mandible in OSCC patients. Additionally, the tumor's location within the mandible, bone volume, and cortical bone mineral density were identified as factors impacting DFS.

In vivo and in vitro action mechanism of treatment of glucocorticoid-induced osteoporosis by regulation of osteoprotegerin/receptor activator of nuclear factor-κB pathways by denshensu.

The research aimed to discuss the action mechanism of the treatment of glucocorticoid-induced osteoporosis (GIOP) by denshensu. In the research, 60 rats were purchased and divided into a control group, model group, estradiol group, and denshensu treatment group. Except for the control group, GIOP models were established for all other groups, and then the structural changes of osseous tissues as well as osteoprotegerin (OPG), expression of receptor activator of nuclear factor-κB ligands (RANKL) were detected. Besides, the changes in osteoclasts were observed by bone marrow-derived mononuclear phagocytes in vitro. The results showed that the micro-structure of bone trabeculae, bone mineral density (BMD), and bone metabolic markers of rats in the denshensu treatment group were enhanced significantly, while trabecular separation and structural model index were reduced (P<0.05). OPG messenger ribonucleic acid (mRNA) and protein levels in the hypothalamus and femur tissues were increased, while RANKL content was remarkably decreased (P<0.05). In addition, in vitro experiments revealed that denshensu inhibited the differentiation of positive osteoclasts, and osteoclast-related genes were reduced (P<0.05). To conclude, denshensu might inhibit the expressions of OPG and RANKL and further play a role in treating GIOP.

Association of rs2073618 polymorphism and osteoprotegerin levels with hypertension and cardiovascular risks in patients with type 2 diabetes mellitus.

There are reports of link of osteoprotegerin (OPG) gene polymorphism to type-2 diabetes (T2D) and hypertension (HTN). The objective of the study was to assess the allele frequency of OPG (rs2073618) gene polymorphism and its association with heart rate variability (HRV) and blood pressure variability profile as CVD risks in diabetes mellitus patients with hypertension undergoing treatment. T2D patients on treatment without hypertension (n = 172), with hypertension (n = 177) and 191 healthy volunteers were recruited for the study. Their blood pressure variability including baroreflex sensitivity (BRS), heart rate variability (HRV), OPG, insulin, lipid profile, receptor-activator for NFkB (RANK), receptor-activator for NFkB-Ligand (RANKL), and tumor necrosis factor-α (TNF-α) were estimated. Allele frequency of OPG (rs2073618) gene polymorphism was assessed from the DNA samples. BRS and HRV indices were decreased, and RANKL/OPG and TNF-α were increased in T2D and T2D + HTN groups, respectively compared to healthy control group. The reduction in BRS was contributed by increased inflammation and reduced SDNN of HRV in GG genotype in T2D + HTN. In GG + GC subgroup, it was additionally contributed by rise in RANKL/OPG level (ß - 0.219; p 0.008). Presence of mutant GG genotype contributed to the risk of hypertension among T2D patients (OR 3.004) as well as in general population (OR 2.79). It was concluded that CV risks are more in T2D patients with HTN expressing OPG rs2073618 gene polymorphism.

Osteoprotegerin genetic polymorphisms and their influence on therapeutic response to ibandronate in postmenopausal osteoporotic females.

OBJECTIVES: The present study investigated osteoprotegerin (OPG) genetic polymorphisms and their influence on the therapeutic response to ibandronate in postmenopausal osteoporotic females. METHODS: This case-control study included 135 postmenopausal females (89 osteoporotic females and 46 non-osteoporotic females). Each osteoporotic patient received a monthly 150 mg ibandronate tablet for six months, and blood samples were taken before and after treatment. Bone mineral density (BMD) was measured using DEXA Scan. Three SNPs (A163G, T245G, and G1181C) of the OPG gene were selected for analysis. RESULTS: Serum OPG levels were significantly lower in osteoporotic subjects than in the control group. The percentage changes in OPG levels in the osteoporotic group before and after treatment with ibandronate were significant (p < .001). After six months of therapy with ibandronate, the percentage changes in OPG levels with AA, TT, TC, GC, and GG genotypes were significant. Following six months of ibandronate treatment, the AA genotype of rs3134069, TT, TC genotypes of rs3102735, GG, and GC genotypes of rs2073618 SNP showed a significant increase in OPG levels. Age, BMI, and GC polymorphism (rs2073618 (G/C) G1181C) were inversely associated with low BMD. Adjusted odds ratios (OR) showed that BMI, GC, GG polymorphism (rs2073618 (G/C) G1181C) and TC polymorphism (rs3102735 (T/C) A163G) were inversely associated with low BMD. CONCLUSION: The inverse association of rs2073618 and rs3102735 with low BMD indicates the protective role of these SNPs in our population. More research is needed to replicate these results in another cohort and to determine the molecular processes by which such SNPs may influence BMD.

Identification and Association of Polymorphism rs2073618 of the Osteoprotegerin Gene in Type 2 Diabetics with and without Retinopathy.

OBJECTIVE: To identify and determine the association of SNP (rs2073618) of OPG gene in diabetics with and without retinopathy and in healthy controls. STUDY DESIGN: Descriptive study. Place and Duration of the Study: Department of Biochemistry and Molecular Biology, Army Medical College, Rawalpindi in collaboration with Chemical Pathology Laboratory, Pak Emirates Military Hospital, Rawalpindi and Armed Forces Institute of Ophthalmology, Rawalpindi, from June 2021 to May 2022. METHODOLOGY: Participants aged 25-70 years were inducted and divided into three equal groups. Group I consisted of diabetics with retinopathy (n = 50), group II was diabetics without retinopathy (n = 50), and group III was healthy individuals (n = 50). DNA was extracted and allele specific PCR technique was adopted using specifically designed primers. Results were analysed using the software Statistical Package for Social Sciences (SPSS) version 22.0 and online bio-informatics tool SNPstats. RESULTS: CC, CG, and GG genotypes were found to be present in 94%, 4%, and 2% in diabetics without retinopathy, 92%, 4%, and 4% in diabetics with retinopathy, respectively, and 100% presence of CC genotype only in healthy controls. C and G alleles were present in 96% and 4%, respectively, in diabetics without retinopathy, with 100% presence of only C allele in healthy subjects. The genotypic assessment using the models showed no significant association. CONCLUSION: SNP rs2073618 of OPG gene was identified in all study groups without any significant distribution or association with the development of diabetic retinopathy. The major genotype C/C was found in the majority of subjects in all groups. KEY WORDS: Allele specific PCR, Diabetic retinopathy, Single nucleotide polymorphism, Type 2 Diabetes mellitus.

The role of miR-433-3p in vascular calcification in type 2 diabetic patients: targeting WNT/ß-Catenin and RANKL/RANK/OPG signaling pathways.

BACKGROUND: Vascular calcification (VC) is a major predictor of cardiovascular diseases that represent the principal cause of mortality among type-2 diabetic patients. Accumulating data suggest the vital role of some microRNAs on vascular calcification as an epigenetic regulator. Thus, we assessed herein, the role of serum miR-433-3p in vascular calcification in type-2 diabetic patients. METHODS: Twenty healthy subjects (control group) and forty diabetic patients (20 without VC and 20 with VC) were involved in the study. miR-433-3p gene expression was measured. Runx2, Dickkopf-1 (DKK1), ß-catenin, Receptor activator of nuclear factor kappa-B ligand (RANKL), and osteoprotegerin (OPG) levels in serum were assessed by ELISA technique. RESULTS: Diabetes patients had significantly lower levels of miR-433-3p expression in comparison to the control group, with the lowest levels being found in diabetic patients with VC. Furthermore, Runx2, ß-catenin, and RANKL levels were significantly increased with concomitant lower DKK1 and OPG levels detected in the two diabetic groups especially those with VC. CONCLUSION: Collectively, the study documented that down-regulation of miR-433-3p may contribute to the development of VC through activating WNT/ß-Catenin and RANKL/RANK/OPG signaling pathways.

Bone destruction in chronic otitis media is not mediated by the RANKL pathway or estrogen receptor-alpha.

BACKGROUND: Chronic otitis media with or without cholesteatoma progresses with various degrees of bone resorption and remodeling. Estrogen mediates osteoprotective effects through the receptor activator of NF-κB ligand (RANKL) pathway, which is mainly mediated by estrogen receptor-alpha (ER-α). OBJECTIVES: The present study investigated the expression patterns of receptor activator of NF-κB (RANK), osteoprotegerin (OPG), RANKL, and ER-α in pathological tissue from patients with chronic otitis media to determine the roles of those factors in osteolytic mechanisms underlying the pathogenesis of chronic otitis media. METHODS: Normal and pathological specimens from 18 patients with chronic otitis media were examined. RESULTS: There were no significant differences in RANK, OPG, RANKL, or ER-α mRNA expression between normal and pathological specimens of epithelial tissue. CONCLUSIONS: Our findings suggested that RANK, OPG, RANKL, and ER-α are not associated with the bone destruction in chronic otitis media; other cytokines may directly activate the osteoclasts in chronic otitis media.

OPG/RANK/RANKL axis relation to cardiac iron-overload in children with transfusion-dependent thalassemia.

OPG/RANK/RANKL axis was reportedly involved in initiating various diseases, especially bone and cardiovascular diseases. This study aimed to assess the relationship between some OPG, RANK, and RANKL polymorphisms and alleles and iron-overload-induced cardiomyopathy in children with transfusion-dependent thalassemia (TDT). This study included 80 TDT children and 80 age and sex-matched controls. Real-time PCR was done for rs207318 polymorphism for the OPG gene and rs1805034, rs1245811, and rs75404003 polymorphisms for the RANK gene, and rs9594782 and rs2277438 polymorphisms for the RANKL gene. Cardiac T2* MRI and ejection fraction (EF) were done to assess the myocardial iron status and cardiac function. In this study, there were no significant differences in frequencies of the studied polymorphisms between cases and controls (p > 0.05 in all). In TDT children, OPG rs2073618 (G > C) had a significant relation to myocardial iron overload (p = 0.02). Its C allele had significantly more frequent normal EF than its G allele (p = 0.04). RANK rs75404403 (C > DEL) had a significant relation to cardiac dysfunction (p = 0.02). Moreover, the C allele of that gene had significantly more frequent affected EF than its DEL allele (p = 0.02). The A allele of RANKL rs2277438 (G > A) had significantly less frequent severe cardiac iron overload than the G allele (p = 0.04). In conclusion, the OPG/ RANK/RANKL genes may act as genetic markers for iron-induced cardiomyopathy in TDT children. Some of the studied genes' polymorphisms and alleles were significantly related to myocardial iron overload and cardiac dysfunction in TDT children.

Genetically predicted osteoprotegerin levels and the risk of cardiovascular diseases: A Mendelian randomization study.

PURPOSE: The relationship between circulating osteoprotegerin (OPG) levels and the risk of cardiovascular diseases (CVDs) has been the subject of conflicting results in previous observational and experimental studies. To assess the causal effect of genetically predicted OPG levels on the risk of a wide range of CVDs, we used the Mendelian randomization design. DESIGN: We initially extracted information of genetic variants on OPG levels and their corresponding effect values from the summary data based on the European ancestry genome-wide association study. Subsequently, we performed two-sample Mendelian randomization analyses to assess the causal effect of genetically predicted OPG levels on CVDs by using inverse variance weighting (IVW), MR-Egger, weighted median, and MR-PRESSO methods. We also conducted sensitivity analyzes as well as complementary analyses with a more relaxed threshold for the exposure genetic instrumental variable (P < 5 × 10-6) to test the robustness of our results. RESULTS: Our results indicated that genetically predicted OPG levels causally reduce the risk of atrial fibrillation (IVW OR = 0.84; 95% CI = 0.72-0.98; P = 0.0241), myocardial infarction(IVW OR = 0.89; 95% CI = 0.80-0.98; P = 0.0173) and coronary heart disease (IVW: OR = 0.90; 95% CI = 0.82-0.99; P = 0.0286). Further complementary analyses also confirmed the above results remain robust and we also identified a potential causal association of OPG levels with a reduced risk of hypertensive diseases(IVW OR = 0.94;95% CI = 0.88-1.00; P = 0.0394). CONCLUSION: This study provides compelling evidence for a causal relationship between genetically predicted OPG levels and risk reduction of coronary heart disease, myocardial infarction, and atrial fibrillation, indicating that OPG could potentially serve as a cardiovascular risk marker in clinical practice.

Reduced osteoprotegerin expression by osteocytes may contribute to rebound resorption after denosumab discontinuation.

Denosumab is an anti-RANKL Ab that potently suppresses bone resorption, increases bone mass, and reduces fracture risk. Discontinuation of denosumab causes rapid rebound bone resorption and bone loss, but the molecular mechanisms are unclear. We generated humanized RANKL mice and treated them with denosumab to examine the cellular and molecular conditions associated with rebound resorption. Denosumab potently suppressed both osteoclast and osteoblast numbers in cancellous bone in humanized RANKL mice. The decrease in osteoclast number was not associated with changes in osteoclast progenitors in bone marrow. Long-term, but not short-term, denosumab administration reduced osteoprotegerin (OPG) mRNA in bone. Localization of OPG expression revealed that OPG mRNA is produced by a subpopulation of osteocytes. Long-term denosumab administration reduced osteocyte OPG mRNA, suggesting that OPG expression declines as osteocytes age. Consistent with this, osteocyte expression of OPG was more prevalent near the surface of cortical bone in humans and mice. These results suggest that new osteocytes are an important source of OPG in remodeling bone and that suppression of remodeling reduces OPG abundance by reducing new osteocyte formation. The lack of new osteocytes and the OPG they produce may contribute to rebound resorption after denosumab discontinuation.

Establishment of a TNFRSF11B knock-out human induced pluripotent stem cell line (KSCBi002-B-2) via CRISPR/Cas9 system.

A TNFRSF11B (TNF Receptor Superfamily Member 11b) gene encodes a soluble decoy receptor, osteoprotegerin (OPG), which has a key role in repressing osteoclast differentiation. In this report, we generated a biallelic knock-out hiPSC line for the TNFRSF11B gene via CRISPR/Cas9. When TNFRSF11B Knock-out hiPSCs were differentiated into mesenchymal progenitor cells (MPCs), the expression level of OPG was significantly decreased compared to normal hiPSC-derived MPCs. This knock-out hiPSCs will provide a chance to study Paget disease of bone 5 (juvenile Paget disease).

Hypoxia caused by unilateral nasal obstruction decreases mandibular density in rats through inhibition of Cyp1a1 expression.

BACKGROUND: Oral breathing has an important impact on morphology and bone mineral density (BMD) in a mandible. This study aimed to investigate the hub genes and mechanism regulating the mandibular BMD decrease induced by nasal obstruction. METHODS: A unilateral nasal obstruction model was established in 1-week-old Wistar rats by electrocautery obstruction. BMD of the mandible was determined by micro-computed tomography. Transcriptome analysis was performed to identify differentially expressed genes (DEGs). Hub genes were identified by building protein-protein interaction network and verified by western blot. A hypoxic cell model was established in bone marrow mesenchymal stem cells (BMSCs) by using CoCl2. The expression of hypoxia-inducible factor-1α (HIF-1α), NF-kB ligand-receptor activator (RANKL), osteoprotegerin (OPG), and Cyp1a1 was detected by western blot. RESULTS: The mandibular BMD of rats in the unilateral nasal obstruction group was significantly decreased. A total of 38 DEGs were identified in nasal obstruction rats compared with normal rats. A ratio of RANKL/OPG in the mandible was elevated by nasal obstruction, while the Cyp1a1 was decreased. In vitro, the HIF-1α expression and RANKL/OPG ratio were upregulated by hypoxia while the Cyp1a1 expression was decreased. Pretreatment with Cyp1a1 activator, FICZ, could increase the expression of Cyp1a1 while attenuating the activation of HIF-1α and RANKL. CONCLUSION: Respiratory changes caused by nasal obstruction contribute to the decrease in Cyp1a1 expression in the mandible of juvenile rats, which is associated with disturbances in bone homeostasis controlled by the RANKL/OPG ratio.

Bone mineral density in egyptian children with juvenile idiopathic arthritis: possible correlation to serum RANKL / osteoprotegerin (OPG) ratio and OPG gene polymorphisms.

BACKGROUND: Children with juvenile idiopathic arthritis (JIA) are at higher risk of decreased bone mineral density (BMD) compared with healthy children due to genetic, disease and medication-related causes. This study aims to investigate the possible effects of osteoprotegerin (OPG) gene polymorphisms and serum levels of osteoprotegerin (OPG) and receptor activator of nuclear factor κB-ligand (RANKL) and RANKL/OPG ratio on BMD in children with JIA. METHODS: OPG gene rs2073617, rs3134069, serum RANKL, OPG and RANKL/OPG ratio were evaluated in 60 JIA children and 100 matched healthy controls. BMD was evaluated by lumbar dual energy X-ray absorptiometry (DEXA) according to which patients were classified in 2 groups (DEXA z-score above and below - 2). Composite disease activity was measured using the Juvenile Arthritis Disease Activity Score (JADAS) 27-joints. Articular damage was scored using the juvenile arthritis damage index (JADI). RESULTS: Patients aged 12.05 ± 3.2 years, included 38 females and 31% had BMD z-score below-2. Systemic-onset JIA was the most frequent phenotype (38%). Genotypes and alleles frequencies of the 2 studied polymorphisms did not differ between patients and controls (p > 0.05 for all) while serum RANKL and RANKL/OPG ratio were significantly higher in patients compared to controls (p = < 0.001 and 0.03 respectively). Patients with BMD < -2 had significantly greater frequencies of rs2073617 TT genotype and T allele (p < 0.001), higher serum RANKL, RANKL/OPG ratio (p = 0.01, 0.002), female predominance (p = 0.02), higher articular and extra-articular damage index (p = 0.008,0.009) and more frequent steroid usage (p = 0.02) compared to patients with BMD z-score >-2. Multivariate analysis showed rs2073617 TT genotype, RANKL/OPG ratio, long disease duration (above 36 months) and use of steroid to be associated with decreased BMD (p = 0.03,0.04,0.01,0.01 respectively) in JIA children. CONCLUSIONS: Egyptian children with JIA have decreased BMD. rs2073617 TT genotype and T allele, RANKL/OPG ratio are possible determinants of reduced BMD in JIA. Our results underline the importance of frequent monitoring of BMD in JIA children and trying to control disease activity to preserve long term bone health.

MiR-217 regulates autophagy through OPG/RANKL/RANK in giant cell tumors.

BACKGROUND: Increasing evidence suggests that microRNAs (miRNAs) play a crucial role in cancer development and progression. Our previous study showed remarkably lower levels of miR-217 in GCT cells and tissues, and miR-217 re-expression inhibited the occurrence and development of GCT in vitro; however, the associated mechanisms remain unknown. Thus, this study aimed to explore the mechanisms underlying the proliferation inhibitory effect of miR-217 in GCT cells. METHODS: The proliferative potential of the GCT cells was measured with an MTT assay and BrdU straining. Changes in GCT cell migration and invasion was assessed by a transwell assay. Finally, Western blot and RT-PCR assays were employed to evaluate OPG/RANKL/RANK signaling pathway-related protein expression. RESULTS: The excessive upregulation of miR-217 markedly suppressed GCT cell proliferation and tumorigenesis both in vitro and in vivo. miR-217 overexpression could inhibit the OPG/RANKL/RANK signaling pathway in vitro and in vivo. Furthermore, ALP activity was significantly decreased in GCT cells following miR-217 treatment. Importantly, miR-217 could inhibit autophagy-related protein expression and autophagosome/autolysosome formation in GCT cells and tissues. CONCLUSION: These results suggest that miR-217 upregulation could inhibit the occurrence and development of GCT by blocking autophagy. These findings offer an effective therapeutic target to improve the survival rates of patients with CGT in the future.

Prenatal Fumonisin Exposure Impairs Bone Development via Disturbances in the OC/Leptin and RANKL/RANK/OPG Systems in Weaned Rat Offspring.

The goal of the current study was to examine the effects of prenatal exposure to fumonisins (FBs) on bone properties and metabolism in weaned rat offspring divided into groups intoxicated with FBs at either 0 (the 0 FB group), 60 (the 60 FB group), or 90 mg/kg b.w. 0 (the 90 FB group). Female and male offspring exposed to FBs at a dose of 60 mg/kg b.w. had heavier femora. Mechanical bone parameters changed in a sex and FBs dose-dependent manner. Growth hormone and osteoprotegerin decreased in both sexes, regardless of FBs dose. In males osteocalcin decreased, while receptor activator for nuclear factor kappa-Β ligand increased regardless of FBs dose; while in females changes were dose dependent. Leptin decreased in both male FBs-intoxicated groups, bone alkaline phosphatase decreased only in the 60 FB group. Matrix metalloproteinase-8 protein expression increased in both female FBs-intoxicated groups and decreased in male 90 FB group. Osteoprotegerin and tissue inhibitor of metalloproteinases 2 protein expression decreased in males, regardless of FBs dose, while nuclear factor kappa-Β ligand expression increased only in the 90 FB group. The disturbances in bone metabolic processes seemed to result from imbalances in the RANKL/RANK/OPG and the OC/leptin systems.

Oncostatin M stimulates prostaglandin D2-induced osteoprotegerin and interleukin-6 synthesis in osteoblasts.

Oncostatin M produced by osteal macrophages plays a significant role in fracture healing. Osteoprotegerin (OPG) secreted by osteoblasts, binds to the receptor activator of nuclear factor-κB (RANK) ligand (RANKL) as a decoy receptor and prevents RANKL from binding to RANK, resulting in bone resorption suppression. Interleukin-6 (IL-6) is a pro-inflammatory cytokine and generally regulates bone resorption. However, accumulating evidence suggests that IL-6 plays pivotal roles in bone formation. We previously showed that prostaglandin D2 (PGD2) induces OPG synthesis by activating p38 mitogen-activated protein (MAP) kinase, stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK), and p44/p42 MAP kinase in osteoblast-like MC3T3-E1 cells. Furthermore, we demonstrated that PGD2 stimulates IL-6 synthesis by activating p38 MAP kinase and p44/p42 MAP kinase in MC3T3-E1 cells. In the present study, we investigated whether oncostatin M affects PGD2-stimulated OPG and IL-6 synthesis in MC3T3-E1 cells through MAP kinase activation. The osteoblast-like MC3T3-E1 cells and normal human osteoblasts were treated with oncostatin M and subsequently stimulated with PGD2. Consequently, oncostatin M significantly increased the PGD2-stimulated OPG and IL-6 release in both cells. Oncostatin M significantly enhanced mRNA expression levels of OPG and IL-6 induced by PGD2 similarly in both cells. Regarding the signaling mechanism, oncostatin M did not affect the phosphorylation of p38 MAP kinase, SAPK/JNK, and p44/p42 MAP kinase. Our results suggest that oncostatin M upregulates the PGD2-stimulated OPG and IL-6 synthesis in osteoblasts and therefore affects bone remodeling. However, OPG and IL-6 synthesis are not mediated through p38 MAP kinase, p44/p42 MAP kinase, or SAPK/JNK pathways.