Effectof Yishen-Qingluo-Huoxue prescription on OPG/RANKL pathway in CFA rats and its mechanism / 中国药理学通报

To investigate the effect of Xinan Fang Yisheng-Qingluo-Huoxie prescription(YSF)on OPG/RANKL pathway in rats with rheumatoid arthritis induced by complete Freund's adjuvant(CFA)and its mechanism. Methods Thirty-two male SD rats were randomly divided into normal group, AIA group, QLY group and YSF group, with 8 rats in each group. AIA rats were induced except for the normal group. QLY and YSF were administered intragastrically on the first day after the model was successful for 30 days. The body weight and AI score of rats were observed. HE staining was used to observe the histomorphological changes of the ankle joint of rats. The expression of MMP3 and P-P65 was observed by immunohistochemistry. Serum levels of TNF-α, IL-1β, OPG and RANKL were detected by ELISA. The expression of OPG and RANKL proteins in synovium of rats was detected by Western blot and RT-PCR. Results Compared with AIA model, the body weight of the rats gradually returned to normal, and inflammation score decreased. HE staining showed that the bone was well preserved, no synovial infiltration was observed after YSF administration, and MMP3 and P-P65 showed low expression. The levels of TNF-α, IL-1β and RANKL in serum were significantly down-regulated, while the levels of OPG and OPG/RANKL were significantly up-regulated. In addition, Western blot and RT-PCR showed that OPG and OPG/RANKL were highly expressed in synovium of rats, while RANKL was on the contrary. Conclusions YSF may decrease inflammatory score and expression of TNF-α and IL-1β in serum by inhibiting inflammatory response. What's more, the increase of OPG/RANKL ratio can regulate the environment of joint bone destruction, thus alleviating RA.

Experimental study on the regulation of subchondral bone plate osteoprotegerin/receptor activator of nuclear factor kappa-B ligand system by berberine to retard the development of osteoarthritis in rabbits / 中国骨伤

OBJECTIVE@#To investigate the effect of intra-articular berberine injection on the structural remodeling of subchondral bone plate and osteoprotegerin/receptor activator of nuclear factor kappa-B ligand(OPG/RANKL) system expression in rabbits with osteoarthritis(OA).@*METHODS@#Forty 12-month-old male rabbits with an average of(2.73±0.18) kg of body weight, underwent left anterior cruciate ligament transection(ACLT), and were divided into berberine group and placebo groups after operation, 20 rabbits in each group. The berberine group received intra-articular injection of 100 μmol/L berberine 0.3 ml every week for 6 weeks. In placebo group, the same dose of 0.9% sodium chloride injection was injected into the left knee joint cavity every week for 6 weeks. Another 20 12-month-old male rabbits, weighing (2.68±0.18) kg, underwent sham operation on the left knee joint without intra-articular injection intervention (sham operation group). On the last day of the sixth week after operation, three groups of animals were sacrificed to obtain knee joint specimens. The femoral medial condyle samples were obtained for histological evaluation of cartilage and subchondral bone, Mankin scoring system was used to evaluate articular cartilage structure. Image-Pro Plus(IPP) software was used to evaluate subchondral bone plate bone volume(BV), bone volume/total volume(BV/TV), trabecular circumference(TC), mean trabecular thickness (Tb.Th). Real-time quantitative reverse transcription polymerization Enzyme chain reaction(reverse transcription-polymerase chain reaction, RT-PCR) was used to detect the mRNA expression levels of OPG and RANKL in subchondral bone tissue at 6 weeks after operation.@*RESULTS@#The cartilage structure evaluation showed that the surface of cartilage tissue in the sham operation group was smooth and flat, and the safranin coloration was full in the full thickness of the cartilage;the cartilage tissue in the berberine group showed uneven surface layer, and the staining of safranin O was mildly decreased;the surface layer fibrosis was seen in placebo group, Safranin O faded significantly. The Mankin score in the berberine group was lower than that in placebo group(P<0.01), but higher than that in sham operation group(P<0.01). The structural evaluation of subchondral bone plate showed that the trabecular bone in sham-operated group was densely arranged;after berberine intervention, the trabeculae were closely arranged;the subchondral bone trabeculae in placebo group were relatively sparse, and the distance between trabeculae was wider. Subchondral bone plate IPP software evaluation showed that BV, BV/TV, TC, Tb.Th in berberine group were higher than those in placebo group(P<0.01), BV, BV/TV, TC, Tb.Th in berberine group were higher than those in placebo group(P<0.01), while lower than the sham operation group (P<0.01). PCR test results showed that the expression of OPG mRNA in the berberine group was significantly higher than that in placebo group(P<0.01), and OPG mRNA in the berberine group was lower than that in sham operation group (P<0.01). There was no significant difference in mRNA expression of RANKL among three groups(P>0.05);the ratio of OPG/RANKL in berberine group was higher than that in placebo group(P<0.01), but lower than that in sham operation group(P<0.01).@*CONCLUSION@#Intra-articular injection of berberine can effectively inhibit the resorption of subchondral bone in the early stage of OA and delay the development of the disease. The specific mechanism may be that berberine maintains the balance of OPG/RANKL system by up-regulating the expression of OPG gene in subchondral bone.

The role of OPG/RANKL/RANK signaling pathway in the pathogenesis of giant-cell tumor of bone / 中国组织工程研究

BACKGROUND: Studies have shown that osteoprotegerin/receptor activator of nuclear factor κB ligand/nuclear factor κB receptor activator (OPG/RANKL/RANK) signaling pathway has a certain correlation with the pathogenesis of giant-cell tumor. Controlling the OPG/RANKL/RANK signaling pathway to affect the interaction between osteoblasts and osteoclasts can play a certain therapeutic role in giant-cell tumor of bone. OBJECTIVE: Ton introduce the relationship between the OPG/RANKL/RANK signaling pathway and the pathogenesis of giant-cell tumor of bone, and to summarize and discuss the new advances of the OPG/RANKL/RANK signaling pathway in the pathogenesis of giant-cell tumor of bone. METHODS: PubMed, Web of Science, and WanFang databases were searched for relevant articles published from 2001 to 2019 using the keywords of “OPG/RANKL/RANK, giant cell tumor of bone, pathogenesis, signal pathway, bone metabolism" in English and Chinese, respectively. A total of 53 articles were finally included for analysis and discussion after removal of old and repeated literatures. RESULTS AND CONCLUSION: OPG inhibits the proliferation and differentiation of osteoclasts, reduces the activity of mature osteoclasts, and blocks the binding of RANKL to RANK. RANKL binds to RANK on the surface of osteoclast progenitor cells to promote the differentiation and proliferation of osteoclast progenitor cells, thus accelerating osteoclast progression. After binding to RANKL receptor, RANKL activates signal factors such as nuclear factor-κB to promote the proliferation, differentiation and activation of osteoclasts, and to regulate the transcription and expression of related genes. Therefore, the OPG/RANKL/RANK is associated with the pathogenesis of giant-cell tumor.

Effects of Curcumin on Bone Metabolism Balance in Ovariectomized Osteoporosis Model Rats Based on OPG/ RANKL Signaling Pathway / 中国药房

OBJECTIVE：To investigate the effects of curcumin on bone metabolism balance in ovariectomized osteoporosis model rats ，and to investigate its potential mechanism. METHODS ：Totally female Wistar rats were randomly divided into blank group（group A ），model group （group B ），estradiol group [group C （positive control ），estradiol valerate 50 μg（/ kg·d）]， curcumin low-dose ，medium-dose and high-dose groups [group D-F ，55，110，165 μg（/ kg·d）]，with 15 rats in each group. Except for group A ，other rats were ovariectomized to establish osteoporosis model. After modeling ，group A and B were given normal saline intrgastrically ，and administration groups were given relevant medicine intrgastrically 30 mL/kg，once a day ，for consecutive 12 weeks. The contents of serum bone metabolism markers [BALP ，CBF-α1，CTX-Ⅰ，PINP and OC] were determined by ELISA. The bone mineral density （BMD）and trabecular structure indexes [relative bone volume fraction （BV/TV），trabecular number （Tb.N），trabecular thickness （Tb.Th），connectivity density （Conn. D ），trabecular separation （Tb.Sp）and structure model index （SMI）] were determined by micro CT imaging system. The mRNA and protein expression of OPG and RANKL in hypothalamus and femur were determined by RT-PCR and Western blotting assay. RESULTS ：Compared with group A ，the contents of serum bone metabolism markers ，BMD，BV/TV，Tb.N，Tb.Th，Conn.D，mRNA and protein expression of OPG were decreased significantly in group B ，while Tb.Sp ，SMI，mRNA and protein expression of RANKL were increased significantly @qq.com （P＜0.05）. Compared with group B ，the contents of serum bone metabolism markers ，BMD，BV/TV，Tb.N，Tb.Th，Conn.D，mRNA and protein expression of OPG were increased significantly in administration groups ；Tb.Sp，SMI，mRNA and protein expression of RANKL were decreased significantly ，in a dose-dependent manner among curcumin groups （P＜0.05 or P＜0.01）. CONCLUSIONS ：Curcumin can improve the level of bone metabolism，increase BMD ，improve the trabecular microstructure and inhibit bone absorption in ovariectomized osteoporosis model rats. Its mechanism may be related to the regulation of OPG/RANKL signaling pathway.

Effects of drynaria total flavonoids on mandible structure, serum E2 and OPG/RANKL balance in ovariectomized rats / 西安交通大学学报(医学版)

Objective: To explore the effects of drynaria total flavonoids on serum estrogen (E2) level, OPG/RANKL balance and bone microarchitecture of mandible in a rat model of bone loss induced by ovariectomy. Methods: We randomly divided 40 three-month-old female Sprague-Dawley rats into 5 groups (n=8 per group): sham-operation group (Sham), ovariectomized group (OVX), low-dose drynaria total flavonoids group (GB-L), high-dose drynaria total flavonoids group (GB-H) and estradiol valerate group (EV). The mandibular osteoporosis model was established by removing the rats' ovaries. After 12 weeks the rats were killed, and the serum estrogen levels were measured. OPG and RANKL were detected using ELISA method. Therapeutic effect of drynaria total flavonoids was observed by pathomorphology and histomorphometry of the mandibular molar sections. Results: Compared with those in the sham group, the mandible of rats in OVX group was loosely changed, the level of E2 decreased significantly (P<0.001), and OPG/RANKL decreased significantly (P<0.001). Compared with OVX group, the mandible microstructure of rats in EV group got a good repair, while the serum E2 level and OPG/RANKL increased significantly (both P<0.001). Compared with OVX group, the percentage of trabecular bone (%Tb.Ar) and OPG/RANKL increased in all the GB groups (P<0.001, P=0.029), (P<0.001, P=0.007), while GB-H group also had increased thickness of trabecular bone (Tb.Th) and serum OPG levels (P=0.019, P<0.001), reduced trabecular spacing (Tb.Sp)(P=0.001), but no effect on serum E2 level (P=0.432, P=0.459). Conclusion: Drynaria total flavonoids can inhibit the destruction of the mandibular trabecular structure, thus slowing down the mandibular resorption process. However, they cannot improve E2 concentration. The mechanism is realized by adjusting the balance of OPG/RANKL.

Research progress on molecular mechanism of Chinese materia medica affecting bone metabolism / 中草药

Dynamic balance of bone metabolism is one of the important factors to maintain normal osseous tissue function. When the balance is broken, it causes bone damage and even bone metabolic disease. However, the mechanism of bone metabolism is still unclear, the signal pathway is complex, and the treatment of diseases is still under study. The research on the mechanism of bone metabolism by Chinese materia medica has become a new direction in the treatment of bone metabolism diseases. At present, common mechanisms in bone metabolism include OPG/RANKL/RANK signal pathway, Wnt/β-catenin signal pathway, TGF-β/BMP/Smad signal pathway, and NF-κB signal pathway. In this paper, the research on the above pathways was reviewed, so as to provide a reference for further exploring the treatment of bone metabolism diseases with Chinese materia medica.

Research progress on the changes of subchondral bone in osteoarthritis and its molecular mechanisms / 复旦学报（医学版）

Osteoarthritis (OA) is one of the most common chronic osteoarthritic diseases,which can involve the whole joint.Subchondral bone is an important part of the joint and has a close relationship to the development of OA.The changes and mechanisms of subchondral bone in OA are complex and remain disputes.In this review,we will discuss the advances of the molecular mechanisms of subchondral bone in OA,which include the pathological changes and the roles of the OPG/RANKL/RANK system,transforming growth factor β (TGFβ),estrogen-estrogen receptors and lipid metabolism in OA.

Effects of Exercises on Bone Metabolism and OPG-RANKL-RANK System of Osteoclast Differentiation in Rats with Disuse Osteoporosis / 中国运动医学杂志

Objective To explore the effect of treadmill exercises on disuse osteoporosis and the media effect of OPG-RANKL-RANK system of osteoclast differentiation in the process.Methods Forty six-week-old male Sprague-Dawley rats were randomly divided into a normal control group,a disuse model group,a normal recovery group and an exercise recovery group,each of 10.The normal control group were sacrificed 4 weeks later without receiving any special treatment.The disuse model group were sacrificed after 4 weeks of tail suspension.The normal recovery group were sacrificed after 4 weeks of tail suspension and keeping quiet for 4 weeks.The exercise recovery group were sacrificed after 4 weeks of tail suspension and another 4 weeks of treadmill exercises.The indicators of bone mineral density (BMD),bone histomorphometry,bone tissue TRACP-5b staining,bone metabolism and cytokines related to the osteoclast differentiation OPG-RANKL-RANK system were tested immediately after the rats were sacrificed.Results The BMD,trabecular bone volume percentage (TBV),trabecular bone width (Tb.Wi),concentration of serum alkaline phosphatase (ALP) and the gene expression of bone marrow oculopneumoplethysmograph (OPG) of the disuse model goup were significantly lower than those of the normal control group (P＜0.01),while the trabecular bone spacing (Tb.Sp),average positive staining area percentage (APSAP) of tartrate-resistant acid phosphatase (TRACP)-5b,concentration of serum Ca2+ and TRACP-5b and the gene expression of bone marrow cytokines including RANKL,macrophage colony-stinulating factor (M-CSF),RANK,interleukin (IL)-6 and tumor necrosis factor (TNF)-α of the disuse model goup were significantly higher than those of the normal control group (P＜0.01).The BMD,TBV,Tb.Wi,concentration of serum ALP and the gene expression of bone marrow OPG of the exercise recovery group were significantly higher than those of the normal recovery group (P＜0.05 or P＜0.01).The Tb.Sp,APSAP,concentration of serum Ca2+ and TRACP-5b and the gene expression of RANKL,M-CSF,RANK,IL-6 and TNF-α of the exercise recovery group were significantly lower than those of the normal recovery group (P＜0.05 or P＜0.01).Conclusion The 4-week tail suspension can lead to disuse osteoporosis in rats.Treadmill exercise can promote the recovery of rats with the disuse osteoporosis.The occurrence of disuse osteoporosis and the effect of exercises on disuse osteoporosis were related with the expressing of OPG-RANKL-RANK system-related cytokines of osteoclast differentiation in bone marrow microenvironment.

The effect of Polygonatum Sibiricum Polysaccharide on the expression of OPG and RANKL in the rat model of ovariectomy-induced osteoporosis / 实用医学杂志

Objective To investigate PSP on bone microstructures,Ca,P,OPG and RANKL of osteoporotic rat model.Methods Thirty female rats randomly divided into five groups:Sham,OVX,H-,M-,L-PSP.Sham and OVX were irrigated stomachsaline;PSP solution was gavaged to other groups.After 8-week,bone microstructures of tibial metaphyseal,Ca,P,OPG and RANKL were measured.Results Body weight,Ca,P,RANKL,Tb.Sp of OVX were significantly increased compared to Sham,OPG,BV/TV,Tb.Th,Tb.N decreased.Body weight of H-,M-PSP,Ca and Tb.Sp of PSP,P and RANKL in H-PSP were decreased compared to OVX,OPG in H-,M-PSP,BV/TV,Tb.Th,Tb.N of PSP group increased.The differences were statistically significant (P ＜ 0.05).Conclusion PSP prevents osteoporosis by improving the microstructure of trabecular bone,reducing bone turnover,increasing OPG and reducing RANKL expression.

Effect of 1, 25-dihydroxy vitamin D3 combined with mechanical strain treatment on proliferation, differentiation, and expression of OPG and RANKL in osteoblastic MC3T3-E1 cells / 医用生物力学

Objective To investigate the effect of 1,25-(OH)2-vitamin D3 (VD3) or mechanical strain alone and their combined treatment on proliferation and differentiation of pre-osteoblast MC3T3-E1 cells in vitro, as well as gene and protein expression of osteoprotegerin (OPG) and receptor activator of nuclear factor-кB ligand (RANKL) in those cells. Methods MC3T3-E1 cells were treated with 10 nmol/L VD3, intermitted mechanical strain or with a combination of these two factors. Cell proliferation was assessed with flow cytometry, and alkaline phosphatase (ALP) activity was measured using a fluorometric detection kit. The mRNA expression of ALP, runt-related transcriptional factor 2 (Runx2), OPG, and RANKL genes was determined by real-time PCR. The proteins expression of Runx2, OPG, and RANKL was determined by Western blotting. ResultsVD3 inhibited the proliferation of MC3T3-E1 cells, but the mechanical strain had no effect on cell proliferation. Mechanical strain, VD3, and the combined treatment enhanced the ALP activity of MC3T3-E1 cells as well as the protein expression of Runx2. The effect of combined treatment was less pronounced than the effect of VD3 or mechanical strain alone. Mechanical strain promoted the gene and protein expression of osteoprotegerin (OPG) and increased the ratio of OPG/RANKL. However, the combination of VD3 and mechanical strain led to a decrease in ratio of OPG/RANKL. Conclusions Mechanical strain might be effective in inducing osteogenic differentiation and increasing bone formation. A joint stimulation with VD3 and strain can decrease proliferation and osteogenic differentiation and increase RANKL expression, which might affect bone remodeling. This study supplies some new data, which might be important in theoretical and clinical research of osteoporosis (OP) and other related bone diseases.

Influence of low frequency vibration on osteogenesis of bone marrow stromal cells and its mechanism / 中国康复理论与实践

@# Objective To explore the effect of low frequency vibration (LFV) on osteogenesis of bone marrow stromal cells (BMSCs) and OPG mRNA,RANKL mRNA expression.MethodsBMSCs from 10-week-old female SD rats were cultured and randomly divided into 2 groups:control group and vibration group.The BMSCs of vibration group was given LVF with 20 Hz,0.25 g,2 times/d,10 min/time for 7 days.The LFV was given from the eleventh day after BMSCs cultured.The osteogenesis and OPG mRNA,RANKL mRNA of all BMSCs in two group were tested at the end of LFV.ResultsCompared to the control groups,the BMSCs growth and proliferation improved in vibration group,but the alkaline phosphatase (ALP)activity and the number of mineral nodule did not changed significantly.The OPG mRNA elevated in vibration group,while the RANKL mRNA showed no change.ConclusionThe LFV may increase the growth and proliferation of BMSCs,which is attributed to the elevated OPG gene expression.The osteogenesis improved no significance,which may be associated with the too short vibration time.