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Periodontal disease is a common infectious disease that can lead to the loss of teeth. Hower how to effectively suppress the inflammation with medication is unclear. The aim of the present study was to investigate the anti-inflammatory effect of Oroxylin A in periodontitis and its potential role through heme oxygenase-1 (HO-1). Primary rat gingival fibroblasts (RGFs) were cultured using the tissue block method and identified by immunofluorescence. Following lipopolysaccharide (LPS) stimulation of RGFs, Oroxylin A was administered at 50, 100, 200 or 400 µg/ml. Reverse transcription-quantitative PCR was used to assess mRNA expression of cyclooxygenase (COX)-2, TNF-α, RANKL and osteoprotegerin (OPG). Western blotting was used to detect protein expression levels of COX −2, TNF-α, RANKL and OPG. Following HO-1 knockdown, the same treatment was performed. The expression of COX-2 in rat gingival tissue was observed by immunohistochemistry. One-way analysis of variance and Student's t test were used for statistical analysis. Oroxylin A downregulated mRNA expression of COX-2, TNF-α, RANKL and OPG in LPS-induced RGFs. With increase of Oroxylin A dose, the expression of HO-1 was gradually upregulated. When HO-1 was knocked down, Oroxylin A did not downregulate the expression of COX-2, TNF-α, RANKL and OPG in LPS-induced RGFs. Immunohistochemical results showed that expression of COX-2 was downregulated by Oroxylin A, and the expression of TNF-α, RANKL and OPG were also downregulated. Oroxylin A decreased expression of inflammatory cytokines in LPS-induced RGFs and had a good inhibitory effect on periodontitis in rats. [ABSTRACT FROM AUTHOR]

Dental epithelial-mesenchymal signaling is crucial for tooth development, but the detailed mechanism is not fully understood. Using microarray analysis, it was revealed that the expression of osteoprotegerin, an important factor regulating bone remodeling, significantly increased after removal of the dental epithelium. Immunohistochemical staining revealed that osteoprotegerin expression within the dental mesenchyme was quite low during the prenatal period, but significantly increased after birth. To investigate the influence of osteoprotegerin upon tooth development, first-molar tooth germs from embryonic day 14.5 (E14.5) Chinese Kunming mice were treated with different concentrations of osteoprotegerin. It was revealed that osteoprotegerin could inhibit the expression of odontogenic markers while promoting the expression of osteogenic markers, thereby disrupting tooth morphogenesis. These findings were further supported by in vitro and in vivo cultures. Finally, quantitative reverse transcription-polymerase chain reaction and immunofluorescence studies revealed that, after osteoprotegerin treatment, the activity of the wingless/integrated (Wnt)/β-catenin pathway increased, indicating that increased osteoprotegerin expression in prenatal tooth development could lead to uncontrolled upregulation of the Wnt/β-catenin pathway. [ABSTRACT FROM AUTHOR]

Previous studies have shown that Tougu Xiaotong capsule (TGXTC) has therapeutic effects on knee osteoarthritis (OA) through multiple targets. However, the mechanisms of action underlying its regulation of subchondral bone reconstruction remain unclear. In this study, we investigated the effects of TGXTC on subchondral bone remodeling. Eighteen six-month-old New Zealand white rabbits of average sex were randomly divided into the normal, model and TGXTC groups. The rabbit knee OA model was induced by a modified Hulth's method in the model and TGXTC groups, but not the normal group. Five weeks postoperatively, intragastric administration of TGXTC was performed for four weeks. After drug administration, the medial femoral condyle and tibia were prepared for observation of cartilage histology via optical microscopy and micro-computed tomography, the serum was collected for biochemical parameters assay and the subchondral bone isolated from the lateral femoral condyle was collected for detection of IL-1β and TNF-α mRNA and protein by reverse transcription-quantitative polymerase chain reaction and western blot analysis, respectively. The results showed that treatment with TGXTC significantly mitigated cartilage injury and subchondral bone damage, improved the parameter of subchondral trabecular bone, decreased alkaline phosphatase and tartrate-resistant acid phosphatase activity, and significantly reducing the osteoprotegerin/receptor activator of nuclear factor-κB ligand ratio, reduced the expression of IL-1β and TNF-α mRNA and protein. These results suggest that TGXTC could delay the pathological development of OA by regulating subchondral bone remodeling through regulation of bone formation and bone resorption and its relating inflammatory factors, and this may partly explain its clinical efficacy in the treatment of knee OA. [ABSTRACT FROM AUTHOR]

Osteoporosis and atherosclerosis are two chronic degenerative diseases that share several biochemical pathways and risk factors. Previous studies have associated osteoporosis with carotid atherosclerosis, cardiovascular mortality and stroke, but data on the relationship with peripheral artery disease are few and conflicting. The OPG/RANK/RANKL system and Wnt/beta catenin signaling seem to be deeply involved in the pathogenesis of bone alterations and atherosclerotic processes also affect arteries of the lower extremities. Hypovitaminosis D could also play a role in the relationship of these two diseases. New and larger studies are necessary to shed light on this association and to design new drugs able to act in both these chronic degenerative diseases. [ABSTRACT FROM AUTHOR]

Aseptic loosening and menopause‑induced osteoporosis are caused by an imbalance between bone formation and osteolysis. With an aging population, the probability of simultaneous occurrence of such conditions in an elderly individual is increasing. Strontium ranelate (SR) is an anti‑osteoporosis drug that promotes bone formation and inhibits osteolysis. The present study compared the effects of SR with those of the traditional anti‑osteoporosis drug alendronate (ALN) using an ovariectomized mouse model of osteolysis. The degree of firmness of the prosthesis and the surrounding tissue was examined, a micro‑CT scan of the prosthesis and the surrounding tissue was performed, and the levels of inflammatory and osteogenic and osteoclast factors were examined. It was observed that treatment with SR and ALN improved the bond between the prosthesis and the surrounding bone tissue by reducing the degree of osteolysis, thus improving the quality of bone around the prosthesis. SR increased the secretion of osteocalcin, runt‑related transcription factor 2 and osteoprotegerin (OPG). It additionally decreased the expression of the receptor activator of nuclear factor‑κB ligand (RANKL) and consequently increased the protein ratio OPG/RANKL, whereas ALN exhibited the opposite effect. Furthermore, SR and ALN suppressed tumor necrosis factor‑α and interleukin‑1β production, with SR exerting a more marked effect. The present results demonstrate that SR and ALN may stimulate bone formation and inhibit bone resorption in the ovariectomized mouse model of wear particle‑mediated osteolysis, with SR demonstrating better effects compared with ALN. [ABSTRACT FROM AUTHOR]

Dishevelled (Dvl)-2 represents one of the cytoplasmic proteins, which serves as a pivotal hub in signaling intermediates through a number of different signaling pathways associated with the Wnt family. The aim of the present study was to investigate the roles and mechanisms of Dvl-2 on synovial fibroblasts (SFBs) in osteoarthritis (OA). A Cell Counting kit-8 (CCK-8) assay was used to determine cell viability. An alkaline phosphatase (ALP) test kit was used to measure the activity of ALP. Western blot and reverse transcription-quantitative polymerase chain reaction analysis were used to evaluate the protein and mRNA expression, respectively. The results suggest that depletion of Dvl-2 significantly decreased the expression of osteoprotegerin (OPG) and ALP (P<0.05) and significantly increased the expression of receptor activator of nuclear factor-κB ligand (RANKL), ALP, osteonectin (ON), osteocalcin (OCN) and osterix (P<0.05). In addition, the depletion of Dvl-2 also significantly inhibited the expression of runt-related transcription factor 2 (Runx-2) and β-catenin in SFBs (P<0.05). The effect of Dvl-2 over-expression was opposite to the effect of Dvl-2 silencing. The inactivation of Wnt3a reversed the effect of Dvl-2 silencing. In conclusion, the results indicate that Dvl-2 regulated osteogenic differentiation of SFBs in OA. [ABSTRACT FROM AUTHOR]

Osteoprotegerin (OPG), additionally termed tumor necrosis factor receptor superfamily member 11B, is produced by vascular smooth muscle cells (VSMCs) and endothelial cells in the vasculature, and its release may be modulated by pro-inflammatory cytokines, including interleukin-1β and tumor necrosis factor-α. The present study investigated the effects of treatment with low-dose human recombinant OPG on abdominal aortic aneurysm (AAA) development in mice. Mice were treated with 1 µg human recombinant OPG four times (or vehicle) for 2 weeks prior to inducing AAA. A total of two different models for inducing AAA were used to investigate the hypothesis as to whether OPG is involved in key events of AAA development, using osmotic mini-pumps with angiotensin II in apolipoprotein-E (ApoE−/−) mice for 28 days or using periaortic application of CaCl2 on the aorta in C57Bl/6J mice for 14 days. OPG was continuously administered during the experimental period. Histological staining using Masson's trichrome, Verhoeff's van-Gieson and picro-sirius red, in addition to reverse transcription-quantitative polymerase chain reaction analysis of various markers, were used to analyze phenotypic alterations. Treatment with OPG had no inhibitory effect on AAA development in the angiotensin II model in ApoE−/− mice, which developed suprarenal aneurysms, although it increased vessel wall thickness of the aorta and total collagen in C57Bl/6J mice using the CaCl2 model that induced infrarenal dilation of the aorta. Treatment with OPG did not inhibit aneurysm development and key events, including inflammation, extracellular matrix or VSMC remodeling, in aortas from OPG-treated mice with periaortic treatment with CaCl2. The results indicated that mice treated with low levels of human recombinant OPG may have a more stable aneurysmal phenotype due to compensatory production of collagen and increased vessel wall thickness of the aorta, potentially protecting the aneurysm from rupture. Further studies investigating rupture models of AAA in addition to using higher levels of OPG are require to verify this speculation. Furthermore, treatment with low levels of OPG in patients with AAA may represent a novel therapeutic strategy for the treatment of AAA as well as attenuate the adverse effects associated with the administration of normal and high dosages of OPG. [ABSTRACT FROM AUTHOR]

The present study aimed to investigate the effects of strontium ranelate (SR), an anti‑osteoporotic drug, on osteolysis in an experimental mouse model of aseptic loosening. A total of 45 female C57BL/6J mice each received implantation of one titanium alloy pin into the tibia, followed by intraarticular injection of titanium particles. One week following surgery, mice were randomly divided into three groups: Control group (no additional treatment), SR625 group (treated with SR at a dose of 625 mg/kg/day), and SR1800 group (treated with SR at a dose of 1,800 mg/kg/day). SR was administered via oral gavage once every day for 12 weeks. Micro‑computed tomography scanning and hematoxylin/ eosin staining were used to assess osteolysis around the prosthesis. Immunohistochemistry and reverse transcription‑quantitative polymerase chain reaction analysis were used to measure the expression of receptor activator of nuclear factor‑κB ligand (RANKL) and osteoprotegerin (OPG). Compared with the control, the SR625 and SR1800 groups exhibited a significantly increased pulling force of the titanium alloy pin. Bone volume and the bone surface/volume ratio in the periprosthetic tissue were significantly increased in the SR‑treated groups. Significant differences were observed between the SR1800 group and control group with respect to trabecular thickness and trabecular number. Mechanistically, SR downregulated the expression of RANKL and upregulated the expression of OPG in the periprosthetic tissue. In addition, SR was observed to inhibit wear particle‑associated osteolysis in a dose‑dependent manner. In conclusion, the present data illustrated that SR inhibited titanium particle‑induced osteolysis in vivo. [ABSTRACT FROM AUTHOR]

Cynanchum wilfordii Hemsley has been used for the treatment of musculoskeletal diseases in traditional Republic of Korean medicine. The present study investigated the effects of C. wilfordii water extract (CW) on postmenopausal osteoporosis. Female mice were used and randomly assigned into a normal group and three ovariectomized (OVX) groups: OVX with vehicle (OVX + vehicle); OVX with 17β‑estradiol (E2; 10 μg/kg/day); and OVX with CW (1 mg/kg/day). Oral administration of CW or E2 intraperitoneal injection began 9 weeks after OVX and continued for 3 weeks. Following sacrifice, bone histology, bone mineral density (BMD) and bone mineral content (BMC) of the femur were observed. Serum osteocalcin concentration was analyzed. In addition, the expression levels of osteoprotegerin (OPG) and osterix were evaluated in human osteoblast‑like Saos‑2 cells. In the lateral and medial epicondyles of the CW‑administrated group, dense and well‑formed bone marrow cells with reduced bone marrow pores were observed. CW decreased the number of tartrate resistant acid phosphatase‑positive multinucleated osteoclasts. BMD and BMC were increased following increased serum osteocalcin levels by CW treatment. The expression levels of OPG and osterix were upregulated by CW treatment in vitro. The results suggested that C. wilfordii has an advantageous effect on osteoporosis and possesses the potential to be used in osteoporosis treatment. [ABSTRACT FROM AUTHOR]

Heat shock protein 90 (HSP90) is an ATP-dependent ubiquitous molecular chaperon which is important in cell homeostasis. The authors previously demonstrated that bone morphogenetic protein (BMP)-4 stimulates osteoprotegerin (OPG) production in osteoblast-like MC3T3-E1 cells, and that p70 S6 kinase positively regulates the OPG synthesis by BMP-4. The present study investigated the involvement of HSP90 in the BMP-4-stimulated OPG synthesis and the mechanism in MC3T3-E1 cells. HSP90 inhibitors, 17-allylamino-17demethoxy-geldanamycin (17-A AG), 17-dimethylamino-ethylamino-17-demethoxy-geldanamycin (17-DMAG) and geldanamycin significantly suppressed the BMP-4-stimulated OPG release. Geldanamycin markedly reduced the BMP-4-induced mRNA expression of OPG. 17-AAG and 17-DMAG significantly attenuated the phosphorylation of p70 S6 kinase induced by BMP-4 without affecting the BMP-4-induced phosphorylation of mothers against decapentaplegic homolog 1/5. The results suggest that HSP90 inhibitors suppress the BMP-4-stimulated OPG synthesis in osteoblasts, and that their suppressive effects are exerted through downregulating p70 S6 kinase. [ABSTRACT FROM AUTHOR]

(‑)‑Epigallocatechin gallate (EGCG) and chlorogenic acid (CGA), major flavonoids in green tea, and coffee, respectively, are recognized as possessing potential benefits in a multitude of human health conditions, including bone disorders. We have previously demonstrated that prostaglandin F2α (PGF2α), a potent bone remodeling mediator, stimulates the synthesis of osteoprotegerin (OPG) through the activation of p44/p42 mitogen‑activated protein kinase (MAPK), p38 MAPK and stress activated protein kinase/c‑Jun N‑terminal kinase (SAPK/JNK) in osteoblast‑like MC3T3‑E1 cells. In the present study, the effects of EGCG and CGA on PGF2α‑stimulated OPG synthesis in MC3T3‑E1 cells were investigated. EGCG significantly upregulated PGF2α‑stimulated OPG release, whereas CGA did not affect OPG release. The PGF2α‑induced expression level of OPG mRNA was enhanced by EGCG. Regarding the intracellular signaling underlying the effect of EGCG, EGCG failed to affect PGF2α‑stimulated phosphorylation of p44/p42 MAPK, p38 MAPK or SAPK/JNK. EGCG by itself markedly induced the phosphorylation of p44/p42 MAP kinase for up to 10 min and the status decreased subsequently, whereas EGCG did not significantly affect the phosphorylation status of p38 MAPK or SAPK/JNK within 60 min. These results indicated that EGCG, but not CGA amplifies the PGF2α‑stimulated OPG synthesis in osteoblasts. [ABSTRACT FROM AUTHOR]

Osteoprotegerin (OPG), a member of the tumour necrosis factor receptor (TNFR) superfamily of proteins known to be involved in a large number of biological systems, plays a pivotal role in bone remodelling. In addition to the roles of OPG in bone metabolism, it has been reported to be associated with a high cardiovascular risk in patients with metabolic syndrome. In most cases, the exact functions of OPG remain to be established; however, the widespread expression of OPG suggests that this molecule may have multiple biological activities, mainly in the cardiometabolic environment. The aim of this study was to evaluate the value of OPG as a predictive marker for cardiovascular and metabolic risk in osteoporotic patients. The study group comprised patients with osteoporosis, in order to evaluate the association between OPG serum levels and cardiovascular pathology. Our results revealed significant correlations between classical biochemical bone and metabolic parameters, such as osteocalcin and parathyroid hormone with lipid and glucose biomarkers, sustaining the crosstalk between calcium and bone parameters and cardiovascular risk. The OPG serum level proved to have a significant and independent predictive value for metabolic syndrome (MetS) as a cardiovascular risk standard in osteoporotic patients. The OPG serum levels were increased in patients with MetS as a protective response against the atherosclerotic lesions. The serum levels of 25‑hydroxy vitamin D had significant and independent predictive value for cardiovascular and metabolic risk in our subjects, sustaining the active role of vitamin D beyond the area of bone metabolism. [ABSTRACT FROM AUTHOR]

The aim of the present study was to examine the influence of technetium methylenediphosphonate (99Tc-MDP) on the proliferation and differentiation of human osteoblasts. Human iliac cancellous bone was isolated and cultured with either 99Tc-MDP, β fibroblast growth factor (as a positive control) or medium only (as a negative control). Proliferation was assessed by direct cell counting, CCK-8 assay and bromodeoxyuridine staining. The cell cycle and rate of apoptosis was assessed by propidium iodide staining and flow cytometry. Alkaline phosphatase (ALP) activity was assessed by the p-nitrophenyl phosphate method and mineralized nodules were stained with Alizarin Red. Expression of osteocalcin (OCN) and bone morphogenetic protein-2 (BMP-2) was assessed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and expression levels of osteoprotegerin (OPG) and receptor activator of NF-κB ligand (RANKL) were assessed by RT-qPCR and ELISA. Isolated human osteoblasts stained positively for ALP and developed mineralized nodules. Treatment with 10-5-10-10 M 99Tc-MDP enhanced proliferation and 48 h incubation with 10-8 M 99Tc-MDP increased the proportion of cells in S-phase, decreased the proportion in G0/G1 phase, and increased the cell proliferation index. The rate of apoptosis also increased, but the increase was not significant. Cells incubated with 10-6-10-9 M 99Tc-MDP for 3-9 days exhibited increased ALP activity and mineralized nodule development. 10-8 M 99Tc-MDP increased BMP-2 and OPG expression levels and OPG secretion, but OCN mRNA expression levels and RANKL secretion were not significantly altered at 72 h. 99Tc-MDP treatment induced osteoblast proliferation and differentiation without affecting apoptosis. These findings provide proof of concept for the future use of 99Tc-MDP in the treatment of bone-destructive diseases. [ABSTRACT FROM AUTHOR]

Mimosine, which is a natural plant amino acid present in the Leucaena genus, is able to induce hypoxia-inducible factors (HIFs). Previous evidence has indicated that HIF regulates angiogenesis-osteogenesis coupling in bone metabolism, and it has previously been reported that mimosine inhibits prostaglandin (PG)F2α-induced osteoprotegerin (OPG) synthesis without affecting interleukin-6 (IL-6) production in osteoblast-like MC3T3-E1 cells. In addition, PGE1 has been demonstrated to induce OPG synthesis via activation of p38 mitogen-activated protein (MAP) kinase and stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK) in these cells, and PGE1 stimulates IL-6 production via the activation of protein kinase A. In the present study, the effects of mimosine on the PGE1-stimulated synthesis of OPG and IL-6 were investigated in osteoblast-like MC3T3-E1 cells. The concentrations of OPG and IL-6 were measured using relevant ELISA kits. OPG mRNA was measured by semi-quantitative reverse transcription polymerase chain reaction. The phosphorylation of p38 MAP kinase and SAPK/JNK was analyzed by western blotting. Mimosine significantly reduced PGE1-induced release of OPG and OPG mRNA expression levels without affecting the release of IL-6. In addition, deferoxamine, which is also a normoxic HIF inducer, significantly inhibited PGE1-induced OPG release and OPG mRNA expression levels; however, it had little effect on IL-6 release. Furthermore, mimosine and deferoxamine failed to affect PGE1-stimulated phosphorylation of p38 MAP kinase or SAPK/JNK. These results strongly suggest that normoxic HIF inducers attenuate PGE1-stimulated OPG synthesis without affecting IL-6 production in osteoblasts. [ABSTRACT FROM AUTHOR]

Osteoprotegerin (OPG) is implicated in the pathogenesis of postmenopausal osteoporosis, and other metabolic bone diseases caused by estrogen deficiency. Previous studies have demonstrated that estrogen may stimulate OPG expression in osteoblast cells at the transcriptional level; however, whether estrogen can regulate OPG expression at a post-transcriptional level remains elusive. The present study aimed to investigate the role of microRNA (miRNA) in estrogen-mediated OPG production in human osteoblast-like MG-63 cells. The results from ELISA, western blotting and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) confirmed that estrogen may upregulate OPG expression. Mechanistic studies indicated that estrogen increased the activity of a luciferase reporter harboring the OPG 3'-untranslated region (3'-UTR). Bioinformatics analysis demonstrated that there is a potential targeting site in the OPG 3'-UTR for miRNA (miR)-145, which is associated with osteoblast differentiation. The results of an RT-qPCR suggested that estrogen suppressed miR-145 expression. In addition, dual-luciferase assay, RT-qPCR and western blot analysis indicated that miR-145 directly targets and negatively regulates OPG expression. Furthermore, transfection of cells with miR-145 mimics was able to partially inhibit the induction of OPG expression by estrogen, thus confirming the role of miR-145 in estrogen-mediated OPG induction. Taken together, the results of the present study demonstrated that estrogen may post-transcriptionally regulate OPG expression through suppression of miR-145 expression. [ABSTRACT FROM AUTHOR]

The present study aimed to investigate the effects of osthole on osteoclast formation and bone loss in a mouse model of 5/6 nephrectomy. The mice in control and osthole groups were treated 1 month following 5/6 nephrectomy with either a placebo or osthole, respectively. At 2 months post-nephrectomy, the L4 vertebrae were harvested. The bone mineral density (BMD) of cancellous bone was measured using micro-CT and tartrate-resistant acid phosphatase (TRAP) staining was performed to evaluate osteoclast formation. Immunohistochemistry staining and reverse transcription-quantitative polymerase chain reaction were performed to detect the expression of nuclear factor of activated T-cells, cytoplasmic-1 (NFATc-1), c-Fos, cathepsin K, Trap, matrix metalloproteinase 9 (Mmp9), osteoprotegerin (Opg) and receptor activator for nuclear factor-κB ligand (Rankl). Bone marrow cells were cultured with osthole, and osteoclast formation was shown by TRAP staining. Primary calvaria osteoblasts were cultured with osthole, and expression levels of Opg and Rankl were detected. Compared with the sham group, the BMD of mice in model group was significantly reduced. The numbers of osteoclasts and the expression levels of NFATc-1, c-Fos, cathepsin K and Mmp9 were significantly increased. Compared with the control group, the mice in the osthole group exhibited increased BMD of the L4 vertebrae, a reduction in osteoclast numbers and decreased expression levels of NFATc-1, c-Fos, cathepsin K and Mmp9. In vitro experiments also showed that osteoclast formation was decreased following treatment with osthole. Osteoprotegerin (Opg)/receptor activator for nuclear factor-κB ligand (Rankl) was upregulated by osthole treatment in the L4 vertebrae and in primary cultures of calvarial osteoblasts. Osthole inhibited osteoclast formation and partially reversed the bone loss induced by 5/6 nephrectomy in mice through the upregulation of OPG/RANKL. [ABSTRACT FROM AUTHOR]

Thiazolidinediones are traditional anti-diabetic therapeutic agents that have been associated with bone loss and increased fracture risk. However, the underlying mechanisms of this side effect require further elucidation. The present study aimed to investigate the effect of pioglitazone (PIO), a thiazolidinedione, on osteoblastogenesis, osteoclastogenesis and the osteoprotegerin (OPG) / receptor activator of nuclear factor-κB ligand (RANKL) / RANK system. The MC3T3-E1 murine pre-osteoblastic cell line was treated with PIO and processed for reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis of OPG, RANKL, peroxisome proliferator-activated receptor γ (PPARγ), Runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP) and osteocalcin (OCN), and western blotting analysis of OPG and RANKL. The culture medium was collected for ELISA analysis of OPG and RANKL. Murine bone marrow monocytes (BMMCs) were treated with PIO in the presence of RANKL and macrophage-colony stimulating factor and subjected to tartrate-resistant acid phosphatase (TRAP) staining and activity measurement, and RT-qPCR analysis of cathepsin K, TRAP and RANK. Co-culture of MC3T3-E1 and BMMCs was performed in the presence of PIO, and TRAP staining was also conducted. PIO inhibited the osteoblastic differentiation of MC3T3-E1 cells, and promoted the osteoclastic differentiation of BMMCs with or without co-culturing with MC3T3-E1 cells. ELISA analysis indicated increased RANKL and decreased OPG expression levels in the medium of MC3T3-E1 cells treated with PIO. PIO upregulated expression of RANKL and PPARγ and downregulated expression of OPG, RUNX2, ALP and OCN in MC3T3-E1 cells, while expression levels of RANK in BMMCs remained unchanged. These results suggest that PIO suppresses osteoblastogenesis and enhances osteoclastogenesis. In addition, PIO may also promote osteoclastogenesis by affecting the OPG-RANKL-RANK system. [ABSTRACT FROM AUTHOR]

This study was designed to investigate the anti-osteoporotic activity of polydatin and its possible underlying mechanism. Osteoporosis was induced in mice by ovariectomy (OVX) and the mice were divided into 5 groups: An OVX only group, polydatin groups (10, 20 and 40 mg/kg) and a sham group (n=10/group). After 12 weeks of treatment, body weight, uterine index and the dry weight of thigh-bones were recorded. In addition, the serum calcium, serum phosphorus, alkaline phosphatase (ALP) and osteoprotegerin (OPG) levels were also determined. Western blot analysis was then conducted to investigate the possible mechanism underlying the effect of polydatin via determining the expression of OPG, receptor activators of nuclear factor-ΚB ligand (RANKL) and β-catenin in the ST2 cell line. The results indicated that intraperitoneal injection of polydatin (10, 20 and 40 mg/kg/day) decreased body weight, and increased uterine index and dry weights of thigh-bones of ovariectomized mice (P<0.05), and polydatin also significantly increased the serum calcium, phosphorus, ALP and OPG of ovariectomized mice (P<0.05). Results of western blot analysis showed that polydatin upregulated the ratio of OPG/RANKL (P<0.05) and β-catenin protein in ST2 cells. In conclusion, the results demonstrated that polydatin exhibits anti-osteoporotic activity via regulating osteoprotegerin, RANKL and β-catenin. [ABSTRACT FROM AUTHOR]

Osteonecrosis, also termed aseptic necrosis, is the cellular death of bone components due to interruption of the blood supply. Glucocorticoid (GC) therapy is a common non-traumatic cause of osteonecrosis. However, the mechanism by which GCs induce osteonecrosis remains to be elucidated. The aim of the present study was to investigate the effects of GCs on osteoclast and osteoblast differentiation and function in a GC-induced osteonecrosis mouse model. BALB/c male mice (n=40; 4-weeks-old) were treated with dexamethasone and asparaginase for 8 weeks. The control group (n=20) was administered normal saline. The results demonstrated that the GC-treated group had a lower mean weight compared with the control group. Morphologically, 16/37 (43%) mice demonstrated significant osteonecrotic lesions in the GC-treated group. However, osteonecrotic lesions were not observed in the mice of the control group. Furthermore, immunohistochemistry demonstrated that the GC-treated group had a higher level of osteoprotegerin compared with the control group, without any change in the expression of receptor activator of nuclear factor-κB ligand. In addition, tartarate-resistant acid-phosphatase staining demonstrated significantly decreased osteoclasts in the areas of bone destruction in the GCs-treated group. Furthermore, the present study demonstrated that GCs increased expression levels of osterix and osteocalcin, and decreased expression of matrix metallopeptidase-9 to regulate the differentiation and function of osteoblasts and osteoclasts. The results of the present study suggested that GCs influence bone remolding resulting in decreased osteoclasts formation/differentiation. Therefore, regulating the differentiation and activity of the osteoclasts may be beneficial to the control and treatment of osteonecrosis. [ABSTRACT FROM AUTHOR]

Calcitonin-gene-related peptide (CGRP) is a neuropeptide, which is widely distributed throughout the central and peripheral nervous systems. Numerous mechanisms underlying the action of CGRP in osteoblast-associated cells have been suggested for bone growth and metabolism. The present study was designed to closely investigate the osteoblast- and osteoclast-associated mechanisms of the effect of CGRP administration on bone metabolism in primary osteoblasts. Primary osteoblasts were obtained from newborn rabbit calvaria and incubated with different concentrations of human CGRP (hCGRP), hCGRP and hCGRP (8-37), or without treatment as a control. Intracellular calcium (Ca2+) and cyclic adenosine monophosphate (cAMP) were detected following treatment, as well as the expression levels of osteoblast differentiation markers, including activating transcription factor-4 (ATF4) and osteocalcin (OC), and receptor activator of nuclear factor γB ligand (RANKL) and osteoprotegerin (OPG). The isolated primary osteoblasts were found to stain positively for ALP. hCGRP treatment had no significant effect on transient intracellular Ca2+ in the osteoblasts. Treatment of the osteoblasts with hCGRP led to elevations in the expression levels of cAMP, ATF4 and OPG, and downregulation in the expression of RANKL, in a dose-dependent manner. These effects were markedly reversed by the addition of hCGRP (8-37). The results of the present study demonstrated that CGRP administration not only stimulated osteoblast differentiation, as demonstrated by upregulated expression levels of ATF4 and OC in the hCGRP-treated osteoblasts, but also inhibited OPG/RANKL-regulated osteoclastogenesis. CGRP may act as a modulator of bone metabolism through osteoblast and osteoclast-associated mechanisms, which result in osteoblast formation with subsequent activation of bone formation. [ABSTRACT FROM AUTHOR]

Adenosine monophosphate-activated protein kinase (AMPK) is currently recognized to act as a key sensing enzyme in the regulation of cellular energy homeostasis. It has been previously demonstrated that prostaglandin E2 (PGE2) stimulates the synthesis of osteoprotegerin (OPG) through the activation of p38 mitogen-activated protein (MAP) kinase, p44/p42 MAP kinase and stress-activated protein kinase/ c-Jun N-terminal kinase (SAPK/JNK) in osteoblast-like MC3T3-E1 cells. In the present study, it was investigated whether AMPK is implicated in the PGE2-induced OPG synthesis in MC3T3-E1 cells. PGE2 was observed to induce the phosphorylation of AMPKα (Thr-172) and AMPKβ (Ser-108) in a time-dependent manner. PGE2 additionally induced the phosphorylation of acetyl-coenzyme A (CoA) carboxylase, a direct substrate of AMPK. Compound C, an inhibitor of AMPK, which attenuated the phosphorylation of acetyl-CoA carboxylase, significantly suppressed the PGE2-stimulated OPG release and the mRNA expression level. Compound C failed to affect the PGE2-stimulated phosphorylation of p38 MAP kinase or p44/p42 MAP kinase. On the contrary, the phosphorylation of SAPK/JNK was markedly attenuated by compound C. The results of the current study suggest that AMPK acts as a positive regulator in PGE2-stimulated OPG synthesis via SAPK/JNK signaling in osteoblasts. [ABSTRACT FROM AUTHOR]

Macrophages and osteoclasts release proinflammatory factors and promote osteoclastogenesis following the phagocytosis of wear particles. During this pathological process, receptor of nuclear factor κB ligand (RANKL) and tumor necrosis factor (TNF)-α are critical factors contributing to resorption and the inflammatory response. The present study aimed to construct recombination lentivirus vectors carrying TNF-α small interfering (si)RNA and osteoprotegerin (OPG) cDNA, and to examine the effects of Lenti-siTNFα-OPG on the wear particle-induced inflammatory response and osteoclastogenesis in a titanium (Ti) particle-induced-inflammatory response cell model. Lenti-siTNFα-OPG vectors were constructed and transnfected into RAW264.7 and MC3T3-E1 cells, respectively, prior to particle stimulation. The protein levels of TNF-α, OPG and RANKL were evaluated using western blot analysis and enzyme-linked immunosorbent assays, and the mRNA expression levels of the inflammatory factors, TNF-α, interleukin (IL)-1β and IL-6, as well as OPG and RANKL, were measured using reverse transcription-quantitative polymerase chain reaction analysis. The activity of alkaline phosphatase (ALP) was examined using an ALP kit. In the presence of the Lenti-siTNFα-OPG vector, the mRNA expression levels of the inflammatory factors and RANKL were downregulated, as were the protein levels of TNF-α. The mRNA expression and protein levels of OPG were upregulated, and ALP activity was increased. These findings suggested that Lenti-siTNFα-OPG transfection inhibited the wear particle-induced inflammatory response and osteoclastogenesis, which warrants further investigation for the prevention and/or treatment of wear particle-induced osteolysis. [ABSTRACT FROM AUTHOR]

Osteoporosis is characterized by the progressive loss of bone mass and the micro-architectural deterioration of bone tissue, leading to bone fragility and an increased risk of fracture. Gallium has demonstrated efficacy in the treatment of several diverse disorders that are characterized by accelerated bone loss. Osteoblasts orchestrate bone degradation by expressing the receptor activator of NF-κB ligand (RANKL), however they additionally protect the skeleton by secreting osteoprotegerin (OPG). Therefore, the relative concentration of RANKL and OPG in bone is a key determinant of bone mass and strength. The current study demonstrated that gallium nitrate (GaN) is able to counteract bone loss in an experimental model of established osteoporosis. Ovariectomized (OVX) rats exhibited significantly increased bone mineral density following GaN treatment for 4 and 8 weeks by 19.3 and 37.3%, respectively (P<0.05). The bone volume of the OVX + GaN group was increased by 40.9% (P<0.05) compared with the OVX group. In addition, the current study demonstrated that GaN stimulates the synthesis of OPG however has no effect on the expression of RANKL in osteoblasts, as demonstrated by RT-qPCR, western blotting and ELISA, resulting in an increase in the OPG/RANKL ratio and a reduction in osteoclast differentiation in vivo and in vitro. [ABSTRACT FROM AUTHOR]

The imbalance of subchondral bone remodeling is a common pathological feature in the progression of osteoarthritis. In the current study, using a rabbit model of knee osteoarthritis, the effects of the Tougu Xiaotong capsule (TGXTC) on the cartilage and subchondral bone were investigated. In addition, osteoprotegerin (OPG), an inducer of bone formation, and receptor activator of nuclear factor-κB ligand (RANKL), a regulator of bone resorption in the subchondral bone, were assessed, in order to further explore the protective role of TGXTC in subchondral bone remodeling. The rabbit model of knee osteoarthritis, which was induced by a modified version of Hulth's method, was treated with TGXTC or glucosamine hydrochloride for 4 or 8 weeks. Subsequently, the tibia and femur were harvested for observation of cartilage histology, and the subchondral bone was observed by scanning electron microscopy. The expression levels of OPG and RANKL at the gene and protein levels were determined by reverse transcription-quantitative polymerase chain reaction and western blotting. TGXTC and glucosamine hydrochloride were identified to mitigate cartilage injury, reduce trabecular number and thickness and accelerate trabecular separation. It was additionally observed that the level of OPG mRNA and protein expression was reduced, and the RANKL mRNA and protein expression level was increased, in addition to the observation of a lower OPG/RANKL ratio in the TGXTC and hydrochloride groups. Taken together, these results suggest that TGXTC may mitigate cartilage injury and subchondral sclerosis, thus delaying the pathological development of osteoarthritis. This is suggested to be mediated partly through the reduction of OPG expression and increase of RANKL expression, which reduces the OPG/RANKL ratio, suppressing excessive bone formation. [ABSTRACT FROM AUTHOR]

The present study aimed to investigate the effects of Wenyangbushen formula on the mRNA and protein expression levels of vascular endothelial growth factor (VEGF), osteoprotegerin (OPG), receptor activator of nuclear factor (NF)-κβ ligand (RANK), and RANK ligand (RANKL) in a rabbit model of steroid-induced avascular necrosis of the femoral head (SANFH). The present study also aimed to examine the potential mechanism underlying the effect of this formula on the treatment of SANFH. A total of 136 New Zealand rabbits were randomly divided into five groups: Normal group, model group, and three groups treated with the traditional Chinese medicine (TCM), Wenyangbushen decoction, at a low, moderate and high dose, respectively. The normal group and positive control group were intragastrically administered with saline. The TCM groups were treated with Wenyangbushen decoction at the indicated dosage. Following treatment for 8 weeks, the mRNA and protein expression levels of VEGF, OPG, RANK and RANKL in the femoral head tissues were determined using reverse transcription-quantitative polymerase chain reaction and western blot analyses, respectively. The data revealed that Wenyangbushen decoction effectively promoted the growth of bone cells, osteoblasts and chondrocytes, and prevented cell apoptosis in the SANFH. The mRNA and protein expression levels of OPG and VEGF were increased, while the levels of RANK and RANKL were reduced in the necrotic tissue of the model group, compared with those in the normal rabbits. Wenyangbushen treatment prevented these changes, manifested by an upregulation in the expression levels of VEGF and OPG, and downregulation in the expression levels of RANK and RANKL in a dose-dependent manner. It was concluded that treatment with Wenyangbushen formula alleviated necrosis of the femoral head induced by steroids. It was observed to promote bone cell, osteoblast and chondrocyte growth, as well as prevent cell apoptosis. In addition, it upregulated the expression levels of OPG and VEGF, and inhibited the expression levels of RANK and RANKL. These results suggest the potential use of Wenyangbushen formula as a possible approach for the effective treatment of SANFH. [ABSTRACT FROM AUTHOR]

The present study aimed to determine whether the mitogen-activated protein kinase (MAPK) signaling pathway is involved in the osteoprotegerin (OPG)-mediated inhibition of osteoclast differentiation and maturation. RAW264.7 cells were incubated with macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL) to stimulate osteoclastogenesis and treated with various concentrations of OPG, an inhibitor of osteoclast differentiation. The differentiation and activation of osteoclasts were monitored by tartrate-resistant acid phosphatase staining and bone resorption assays. The phosphorylation levels of p38-MAPK, c-Jun N-terminal kinase (JNK)-MAPK and extracellular signal-regulated kinase (ERK)-MAPK in the different treatment groups were determined by western blot analysis. The results confirmed that M-CSF + RANKL stimulated the differentiation and activation of osteoclasts as well as the phosphorylation of p38-MAPK, JNK-MAPK and ERK-MAPK in osteoclasts, which was attenuated by OPG treatment. These findings indicated that the MAPK signaling pathway is involved in the regulation of osteoclastogenesis and in the OPG-mediated inhibition of osteoclast differentiation and activation. [ABSTRACT FROM AUTHOR]

The aim of the present study was to investigate the alterations in the formation of cementocytes in response to orthodontic forces and to evaluate the contribution of these cells in the biological changes of tooth movement and associated root resorption. A total of 90 Sprague Dawley rats were randomly assigned to the control, high force, and low force groups. Intrusion forces of 10 and 50 g were applied on the rat molar to induce tooth intrusion. The tooth movement was observed from 0 to 14 days by micro-computed tomography, bone histometric analysis, tartrate-resistant acid phosphatase staining, as well as reverse transcription-quantitative PCR and immunofluorescence staining assays. The results suggested that under low force conditions, osteoclasts were distributed at a higher frequency on the bone side than on the root side. Under high force conditions, both sides suffered osteoclast infiltration. In the low force group, the cementocytes exhibited downregulated sclerostin (SOST) and osteoprotegerin (OPG) mRNA levels and a lower receptor activator of nuclear factor-κB ligand (RANKL)/OPG ratio over a certain period of time. The expression levels of these genes were lower compared with those of the osteocytes at each time-point. In the high force group, both cementocytes and osteocytes upregulated the SOST and RANKL/OPG ratio on days 7 and 14, while the cementocytes expressed higher levels of SOST mRNA than those noted in the osteocytes. These data suggested that cementocytes responded to the orthodontic force via modulation of the RANKL/OPG ratio and SOST expression. The biological response of cementocytes contributed to the mechanotransduction and homoeostasis of the roots under compression. Excessive forces may act as a negative factor of this regulatory role. These results expand our knowledge on the function of cementocytes. [ABSTRACT FROM AUTHOR]

In the present study, the effects of obesity on bone metabolism were investigated using a hyperphagic and obese rat model, the Otsuka Long-Evans Tokushima fatty (OLETF) rat, which exhibits normal glycemic control at 8 weeks of age. Body weight, food intake, fat mass, markers of bone resorption, the activities of tartrate-resistant acid phosphatase (TRAP) and cathepsin K, the number of osteoclasts in the proximal tibia, and the serum C-terminal crosslinking telopeptide level were higher in OLETF rats than those in control rats (Long-Evans Tokushima Otsuka; LETO). However, no differences in markers of bone formation, alkaline phosphatase activity, the number of osteoblasts in the proximal tibia or the serum osteocalcin level were observed. mRNA and protein levels of c-fms, receptor for activation of nuclear factor-κB (RANK), RANK ligand (RANKL), TRAP and cathepsin K were significantly increased in OLETF rats, although those levels of macrophage colony-stimulating factor (M-CSF) and osteoprotegerin (OPG) were similar to those in LETO rats. The level of serum tumor necrosis factor α (TNFα), and that of TNFα mRNA in bone, increased in association with the activation of NFκB. Furthermore, a frequency analysis and a colony formation assay respectively showed that the number of osteoclast precursors and the number of colony-forming cells induced by M-CSF each increased in OLETF rats compared with the control group. These results suggested that hyperphagia-induced obesity with normal glycemic control induces the upregulation of osteoclastogenesis that is associated with an increase in the expression of c-fms, RANK and RANKL, which is induced by TNFα, via the activation of NFκB. [ABSTRACT FROM AUTHOR]

Resveratrol is a naturally occurring polyphenol that possesses health-related properties, and is predominantly found in grapes and berries. Bone morphogenetic protein-4 (BMP-4) stimulates osteocalcin synthesis via p38 mitogen-activated protein (MAP) kinase in osteoblast-like MC3T3-E1 cells. The present study aimed to investigate the effects of resveratrol on BMP-4-induced osteoprotegerin (OPG) synthesis in MC3T3-E1 cells. Resveratrol alone had no effect on OPG expression levels, but significantly enhanced BMP-4-induced OPG release. In addition, resveratrol markedly amplified the mRNA expression levels of BMP-4-induced OPG. SB203580 is an inhibitor of p38 MAP kinase, which was shown to suppress BMP-4-stimulated OPG release. BMP-4-induced phosphorylation of p38 MAP kinase was also enhanced by resveratrol. Furthermore, SB203580 significantly reduced the resveratrol-induced amplification of BMP-4-stimulated OPG release. These results suggested that resveratrol was able to upregulate BMP-4-stimulated OPG synthesis via the amplification of p38 MAP kinase activity in osteoblasts. [ABSTRACT FROM AUTHOR]

The aim of the present study was to compare the osteoclast-inhibiting ability of recombinant osteoprotegerin (OPG) protein (rhOPG-Fc) and recombinant receptor activator of nuclear factor κB (rhRANK) in vitro and in vivo. Osteoclasts were cultured with either rhOPG-Fc or rhRANK for 9 days. The number of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells and resorption pits in bone slices were then counted. In the in vivo investigation, female mice were bilaterally ovariectomized (OVX) and intra-peritoneally injected with 3 mg/kg rhOPG-Fc or rhRANK for 12 weeks, respectively. Bone metabolism, bone mineral density and microstructure changes were then evaluated. The number of TRAP-positive cells and bone resorption pits decreased significantly following culture with either rhOPG-Fc or rhRANK, and this was more marked following culture with rhRANK compared with rhOPG-Fc. The levels of calcium and alkaline phosphatase in the serum were similar pre-OVX and after 12 weeks of treatment, while the levels of phosphorus in the serum were higher following treatment with rhRANK compared with rhOPG. The bone mineral density (BMD) of the whole body, femoral neck and L4 lumbar vertebral body in the mice treated with either rhOPG-Fc or rhRANK increased markedly. In addition, the mice treated with rhRANK exhibited significantly higher BMD in the femoral neck and lumbar vertebral body compared with those treated with rhOPG-Fc. Microcomputed tomography analysis demonstrated that the mice treated with rhRANK exhibited an increased bone volume and structure model index, and decreased trabecular spacing compared with those treated with rhOPG-Fc. rhRANK increased the inhibition of osteoclast differentiation and bone resorption, and rescued OVX-induced osteoporosis more effectively compared with rhOPG-Fc. [ABSTRACT FROM AUTHOR]

The receptor activator of nuclear factor-κB ligand (RANKL)/RANK/osteoprotegerin (OPG) system was identified in the late 1990s, ending the search for the specific factors expressed by osteoblasts and stromal cells in order to regulate osteoclastogenesis. The identification of the RANKL/RANK/OPG system was a breakthrough in bone biology; however, the system not only works as a dominant mediator in osteoclast activation, formation and survival, but also functions in other tissues, including the mammary glands, brain and lymph nodes. Evidence has indicated that the existence of the RANKL/RANK/OPG system in these tissues suggests that it may have specific functions beyond those in bone. Disorders of the RANKL/RANK/OPG system are associated with certain human diseases, including postmenopausal osteoporosis, rheumatoid arthritis (RA), bone tumors and certain bone metastatic tumors. Genetic studies have indicated that the RANKL/RANK/OPG system may be a key regulator in the formation of lymph nodes and in the autoimmune disease RA, which further suggests that the immune system may interact with the RANKL/RANK/OPG system. The present review aimed to provide an overview of the role of the RANKL/RANK/OPG system in osteoclastogenesis, bone disease and tissues beyond bone. [ABSTRACT FROM AUTHOR]

Macrophages, a type of immune cell, are the precursors of osteoclasts, and have important roles in bone remodeling and the immune system. In the present study, the RAW264.7 cell line was used as a macrophage model in order to study the macrophage changes during osteoclastogenesis. Receptor activator of nuclear factor ?B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) induce the formation of osteoclasts from several precursor cells. Observation of RAW264.7 macrophage osteoclastogenesis under the induction of RANKL and M-CSF revealed that except the few RAW264.7 macrophages that were differentiated into osteoclasts, almost all undifferentiated RAW264.7 macrophages underwent apoptosis. BRL-3A cells have no differentiation ability, and RANKL and M-CSF treatments did not induce BRL-3A cell apoptosis. When osteoprotegerin (OPG) was used to completely inhibit the differentiation of RAW264.7 macrophages to osteoclasts, apoptosis did not occur amongst the RAW264.7 macrophages despite the action of RANKL and M-CSF. Rac1, RhoA and RhoV are apoptosis-associated genes in the Rho guanosine triphosphate (GTP)ase family. Their expression levels were detected using quantitative polymerase chain reaction (qPCR). During the process of osteoclast differentiation, the mRNA expression of RhoV was significantly upregulated, while apoptosis occurred in a large proportion of macrophages. However, when macrophage apoptosis was inhibited by OPG, RhoV expression was significantly downregulated. Conversely, Rac1 and RhoA expression did not vary in correspondence with the apoptotic rate of the RAW264.7 macrophages. In conclusion, differentiation of RAW264.7 macrophages into osteoclasts resulted in their apoptosis. OPG inhibited RAW264.7 macrophage differentiation into osteoclasts, and thereby inhibited the apoptosis of RAW264.7 macrophages. RhoV mediated the apoptosis of RAW264.7 macrophages during osteoclast differentiation. [ABSTRACT FROM AUTHOR]

The involvement of osteoprotegerin (OPG) in bone metabolism has previously been established; however, whether OPG regulates chondrocytes directly and exerts precise cellular and molecular effects on chondrocytes remains to be determined. Thus, the present study aimed to investigate the direct effect of OPG on the viability, proliferation and functional consequences of chondrocytes. Primary chondrocytes were isolated from the knee of Sprague-Dawley rats. Passage 1 chondrocytes were identified by toluidine blue staining and used in the experiments. The cell proliferation induced by OPG at various concentrations was measured by a Cell Counting kit-8 (CCK-8) assay. Following pretreatment with mitogen-activated/extracellular signal-regulated kinase kinase (MEK) inhibitor U0126, extracellular signal-regulated kinase (ERK) inhibitor PD098059, and P38 mitogen-activated protein kinase (P38MAPK) inhibitor SB203580 for 30 min, chondrocytes were treated with OPG, and CCK-8 was performed. The cellular signals of MAPKs, including ERK, P38MAPK and c-Jun N-terminal protein kinase (JNK), were investigated by western blot analysis following treatment with OPG. The functional consequences following treatment with soluble OPG were analyzed by qPCR and western blot analysis. OPG increased chondrocyte proliferation with maximal effect at 10 ng/ml, and induced the phosphorylation of MEK and ERK but not P38MAPK or JNK. Suppression of ERK activity via PD098095 inhibited OPG-induced chondrocyte proliferation. Administration of OPG significantly downregulated ADAMTS-5 and upregulated tissue inhibitor of metalloproteinase (TIMP)-4 production, but had no effect on the expression of TIMP-1, -2 and -3, insulin-like growth factor I, transforming growth factor-β, basic fibroblast growth factor, bone morphogenetic protein-2, collagen II, aggrecan and ADAMTS-4. Suppression of ERK activity via PD098095 inhibited the alteration of ADAMTS-5 and TIMP-4 expression induced by OPG. OPG therefore regulated the proliferation of chondrocytes via MEK/ERK signaling, and directly affected chondrocytes by influencing the expression profile of ADAMTS-5 and TIMP-4. [ABSTRACT FROM AUTHOR]

Periodontitis is a common destructive inflammatory disease that leads to changes in the tooth-supporting tissues. Human periodontal ligament cells are essential in periodontal tissue regeneration. The traditional Chinese medicine icariin promoted bone formation, stimulated the osteogenic differentiation of preosteoblastic cells and inhibited osteoclast differentiation and bone resorption. Thus, in the present study, the effect of icariin on cell proliferation and the expression of osteoprotegerin (OPG), receptor activator of nuclear factor-κB ligand (RANKL), core binding factor α1 (Cbfa1) and osteocalcin (OC) was investigated in human periodontal ligament cells, by an MTT assay, qPCR and western blot analysis. The results demonstrated that icariin promoted cell proliferation in a dose- and time-dependent manner, upregulated OPG, Cbfa1 and OC expression, and downregulated RANKL production and the RANKL/OPG expression ratio. This suggested the potential value of icariin in treating alveolar bone resorption and promoting periodontal tissue regeneration, due to its ability to stimulate the proliferation and osteogenic differentiation of human periodontal ligament cells and inhibit osteoclast differentiation. [ABSTRACT FROM AUTHOR]