Objective:To investigate whether acupotomy could inhibit subchondral bone remodeling in knee osteoarthritis(KOA)rabbits by regulating the activity of osteoblasts and osteoclasts.Methods:KOA rabbits were prepared by im...Objective:To investigate whether acupotomy could inhibit subchondral bone remodeling in knee osteoarthritis(KOA)rabbits by regulating the activity of osteoblasts and osteoclasts.Methods:KOA rabbits were prepared by immobilization for 6 and 9 weeks by Videman method.Nine groups of rabbits(control,6 weeks and 9 weeks model,6 weeks and 9 weeks acupotomy,6 weeks and 9 weeks electroacupuncture,and 6 weeks and 9 weeks drug groups)received acupotomy,electroacupuncture and risedronate sodium intervention,respectively,for 3 weeks.Results:Acupotomy can inhibit the activity of osteoclasts and osteoblasts in subchondral bone by reducing the proteins expression of cathepsin K(CK)and tartrate-resistant acid phosphatase(TRAP)and decreasing the proteins expression of osteocalcin(OCN)and alkaline phosphatase(ALP),to intercept the abnormal bone resorption and bone formation of subchondral bone in 6-week and 9-week immobilization-induced KOA rabbits.Conclusion:These findings indicated that acupotomy may be more advantageous than risedronate sodium intervention in modulating subchondral bone remodeling in KOA rabbits,especially in 9-week immobilization-induced KOA rabbits.展开更多
Objective:Myeloma bone disease(MBD)is the most common complication of multiple myeloma(MM).Our previous study showed that the serum levels of C3/C4 in MM patients were significantly positively correlated with the seve...Objective:Myeloma bone disease(MBD)is the most common complication of multiple myeloma(MM).Our previous study showed that the serum levels of C3/C4 in MM patients were significantly positively correlated with the severity of bone disease.However,the mechanism of C3 a/C4 a in osteoclasts MM patients remains unclear.Methods:The formation and function of osteoclasts were analyzed after adding C3 a/C4 a in vitro.RNA-seq analysis was used to screen the potential pathways affecting osteoclasts,and the results were verified by Western blot,q RT-PCR,and pathway inhibitors.Results:The osteoclast area per view induced by 1μg/m L(mean±SD:50.828±12.984%)and 10μg/m L(53.663±12.685%)of C3 a was significantly increased compared to the control group(0μg/m L)(34.635±8.916%)(P<0.001 and P<0.001,respectively).The relative m RNA expressions of genes,OSCAR/TRAP/RANKL/cathepsin K,induced by 1μg/m L(median:5.041,3.726,1.638,and 4.752,respectively)and 10μg/m L(median:5.140,3.702,2.250,and 5.172,respectively)of C3 a was significantly increased compared to the control group(median:3.137,2.004,0.573,and 2.257,respectively)(1μg/m L P=0.001,P=0.003,P<0.001,and P=0.008,respectively;10μg/m L:P<0.001,P=0.019,P<0.001,and P=0.002,respectively).The absorption areas of the osteoclast resorption pits per view induced by 1μg/m L(mean±SD:51.464±11.983%)and 10μg/m L(50.219±12.067%)of C3 a was also significantly increased(33.845±8.331%)(P<0.001 and P<0.001,respectively)compared to the control.There was no difference between the C4 a and control groups.RNA-seq analysis showed that C3 a promoted the proliferation of osteoclasts using the phosphoinositide 3-kinase(PI3 K)signaling pathway.The relative expressions of PIK3 CA/phosphoinositide dependent kinase-1(PDK1)/serum and glucocorticoid inducible protein kinases(SGK3)genes and PI3 K/PDK1/p-SGK3 protein in the C3 a group were significantly higher than in the control group.The activation role of C3 a in osteoclasts of MM patients was reduced by the SGK inhibitor(EMD638683).Conclusions:C3 a activated osteoclasts by regulating the PI3 K/PDK1/SGK3 pathways in MM patients,which was reduced using a SGK inhibitor.Overall,our results identified potential therapeutic targets and strategies for MBD patients。展开更多
The effects of vitamin D on osteoblast mineralization are well documented. Reports of the effects of vitamin D on osteoclasts, however, are conflicting, showing both inhibition and stimulation. Finding that resorbing ...The effects of vitamin D on osteoblast mineralization are well documented. Reports of the effects of vitamin D on osteoclasts, however, are conflicting, showing both inhibition and stimulation. Finding that resorbing osteoclasts in human bone express vitamin D receptor(VDR), we examined their response to different concentrations of 25-hydroxy vitamin D_3[25(OH)D_3](100 or 500 nmol·L^(-1)) and 1,25-dihydroxy vitamin D_3[1,25(OH)_2D_3](0.1 or 0.5 nmol·L^(-1)) metabolites in cell cultures. Specifically, CD14+ monocytes were cultured in charcoal-stripped serum in the presence of receptor activator of nuclear factor kappa-B ligand(RANKL) and macrophage colony-stimulating factor(M-CSF). Tartrate-resistant acid phosphatase(TRAP)histochemical staining assays and dentine resorption analysis were used to identify the size and number of osteoclast cells, number of nuclei per cell and resorption activity. The expression of VDR was detected in human bone tissue(ex vivo) by immunohistochemistry and in vitro cell cultures by western blotting.Quantitative reverse transcription-PCR(q RT-PCR) was used to determine the level of expression of vitamin D-related genes in response to vitamin D metabolites. VDR-related genes during osteoclastogenesis, shown by q RT-PCR, was stimulated in response to 500 nmol·L^(-1)of 25(OH)D_3and 0.1–0.5 nmol·L^(-1)of 1,25(OH)_2D_3,upregulating cytochrome P450 family 27 subfamily B member 1(CYP27B1) and cytochrome P450 family 24 subfamily A member 1(CYP24A1). Osteoclast fusion transcripts transmembrane 7 subfamily member 4(tm7sf4) and nuclear factor of activated T-cell cytoplasmic 1(nfatc1) where downregulated in response to vitamin D metabolites. Osteoclast number and resorption activity were also increased. Both 25(OH)D_3and1,25(OH)_2D_3reduced osteoclast size and number when co-treated with RANKL and M-CSF. The evidence for VDR expression in resorbing osteoclasts in vivo and low-dose effects of 1,25(OH)_2D_3on osteoclasts in vitro may therefore provide insight into the effects of clinical vitamin D treatments, further providing a counterpoint to the high-dose effects reported from in vitro experiments.展开更多
Bone marrow(BM) cavities are utilized for hematopoiesis and to maintain hematopoietic stem cells(HSCs). HSCs have the ability to self-renew as well as to differentiate into multiple different hematopoietic lineage cel...Bone marrow(BM) cavities are utilized for hematopoiesis and to maintain hematopoietic stem cells(HSCs). HSCs have the ability to self-renew as well as to differentiate into multiple different hematopoietic lineage cells. HSCs produce their daughter cells throughout the lifespan of individuals and thus, maintaining HSCs is crucial for individual life. BM cavities provide a specialized microenvironment termed "niche" to support HSCs. Niches are composed of various types of cells such as osteoblasts, endothelial cells and reticular cells. Osteoclasts are unique cells which resorb bones and are required for BM cavity formation. Loss of osteoclast function or differentiation results in inhibition of BM cavity formation, an osteopetrotic phenotype. Osteoclasts are also reportedly required for hematopoietic stem and progenitor cell(HSPC) mobilization to the periphery from BM cavities. Thus, lack of osteoclasts likely results in inhibition of HSC maintenance and HSPC mobilization. However, we found that osteoclasts are dispensable for hematopoietic stem cell maintenance and mobilization by using three independent osteoclast-less animal models. In this review, I will discuss the roles of osteoclasts in hematopoietic stem cell maintenance and mobilization.展开更多
Objective:To study the regulatory effect of feilongzhangxue on the levels of HMGB1-TLR4/RANKL-NF-κB signaling pathway related factors HMGB1,RANKL,rank,TRAF-6 and NF-κBp65 in osteoclasts,so as to explore the mechanis...Objective:To study the regulatory effect of feilongzhangxue on the levels of HMGB1-TLR4/RANKL-NF-κB signaling pathway related factors HMGB1,RANKL,rank,TRAF-6 and NF-κBp65 in osteoclasts,so as to explore the mechanism of feilongzhangxue intervention in RA;Methods:The osteoclasts with good activity were randomly divided into blank group,methotrexate control group and Zhuang medicine feilongzhang blood containing serum treatment group,which were divided into OC+blank group,OC+methotrexate control group,OC+Zhuang medicine feilongzhang blood containing serum group;The expression of HMGB1,RANKL,rank,TRAF-6 and NF-κBp65 mRNA was detected by RT-PCR;The protein expressions of HMGB1,RANKL,RANK,TRAF-6 and NF-κBp65 were detected by immunofluorescence.Results:PCR results showed that:Compared with the blank group,feilongzhangxue could effectively inhibit the expression levels of HMGB1,RANKL,rank,TRAF-6 and NF-κB p65 mRNA in OC cells,and the inhibitory effect was stronger than methotrexate.Immunofluorescence test results showed that:Compared with the blank group and methotrexate group,feilongzhangxue could effectively inhibit the protein expression of HMGB1,RANKL,rank,TRAF-6 and NF-κB p65 in OC cells.Conclusion:The effect of Zhuang medicine feilongzhangxue on hmgbl-tlr4/rankl-nf-κB signaling pathway of osteoclasts is through the regulation of related factors HMGB1,RANKL,rank,TRAF-6 and NF-κB p65,which may be the key mechanism of Zhuang medicine feilongzhangxue on rheumatoid arthritis.展开更多
Osteoporosis is caused by unbalanced bone reconstruction processes which leads to areduction in bone mineral density and changes in bone microstructure,resulting in increased fracture risk.Osteoclasts are the only bon...Osteoporosis is caused by unbalanced bone reconstruction processes which leads to areduction in bone mineral density and changes in bone microstructure,resulting in increased fracture risk.Osteoclasts are the only bone resorption cells in the body of human being,and are closely related to the pathogenesis of osteoporosis.In recent years,there have been more in-depth studies on osteoclasts and bone resorption.The pathogenesis of osteoporosis has always been an important research topic for many medical specialists.Cytokines,as important humoral factors that regulate bone metabolism,play an important role in osteoporosis,especially in the formation,differentiation,and apoptosis of osteoclasts.On the basis of referring to the related studies at home and abroad,this paper analyzes the action mechanism of cytokines such as interleukin-1,interleukin-6,nuclear factor KB ligand receptor activator(RANKL)and tumor necrosis factor(TNF)family members on osteoclasts,and clarifies the relationship between cytokines and osteoporosis,which provides a new angle for the prophylaxis and therapy of osteoporosis.展开更多
Prostaglandin E2(PGE2), a major cyclooxygenase-2(COX-2) product, is highly secreted by the osteoblast lineage in the subchondral bone tissue of osteoarthritis(OA) patients. However, NSAIDs, including COX-2 inhibitors,...Prostaglandin E2(PGE2), a major cyclooxygenase-2(COX-2) product, is highly secreted by the osteoblast lineage in the subchondral bone tissue of osteoarthritis(OA) patients. However, NSAIDs, including COX-2 inhibitors, have severe side effects during OA treatment. Therefore, the identification of novel drug targets of PGE2 signaling in OA progression is urgently needed. Osteoclasts play a critical role in subchondral bone homeostasis and OA-related pain. However, the mechanisms by which PGE2 regulates osteoclast function and subsequently subchondral bone homeostasis are largely unknown. Here, we show that PGE2 acts via EP4 receptors on osteoclasts during the progression of OA and OA-related pain. Our data show that while PGE2 mediates migration and osteoclastogenesis via its EP2 and EP4 receptors, tissue-specific knockout of only the EP4 receptor in osteoclasts(EP4 Lys M) reduced disease progression and osteophyte formation in a murine model of OA. Furthermore, OA-related pain was alleviated in the EP4 Lys M mice, with reduced Netrin-1 secretion and CGRP-positive sensory innervation of the subchondral bone. The expression of plateletderived growth factor-BB(PDGF-BB) was also lower in the EP4 Lys Mmice, which resulted in reduced type H blood vessel formation in subchondral bone. Importantly, we identified a novel potent EP4 antagonist, HL-43, which showed in vitro and in vivo effects consistent with those observed in the EP4 Lys Mmice. Finally, we showed that the Gαs/PI3 K/AKT/MAPK signaling pathway is downstream of EP4 activation via PGE2 in osteoclasts. Together, our data demonstrate that PGE2/EP4 signaling in osteoclasts mediates angiogenesis and sensory neuron innervation in subchondral bone, promoting OA progression and pain, and that inhibition of EP4 with HL-43 has therapeutic potential in OA.展开更多
Tissue-resident macrophages are highly specialized to their tissue-specific microenvironments,activated by various inflammatory signals and modulated by genetic and environmental factors.Osteoclasts and microglia are ...Tissue-resident macrophages are highly specialized to their tissue-specific microenvironments,activated by various inflammatory signals and modulated by genetic and environmental factors.Osteoclasts and microglia are distinct tissue-resident cells of the macrophage lineage in bone and brain that are responsible for pathological changes in osteoporosis and Alzheimer’s disease(AD),respectively.Osteoporosis is more frequently observed in individuals with AD compared to the prevalence in general population.Diagnosis of AD is often delayed until underlying pathophysiological changes progress and cause irreversible damages in structure and function of brain.As such earlier diagnosis and intervention of individuals at higher risk would be indispensable to modify clinical courses.Pleiotropy is the phenomenon that a genetic variant affects multiple traits and the genetic correlation between two traits could suggest a shared molecular mechanism.In this review,we discuss that the Pyk2-mediated actin polymerization pathway in osteoclasts and microglia in bone and brain,respectively,is the horizontal pleiotropic mediator of shared risk factors for osteoporosis and AD.展开更多
Elevated osteoclast(OC)activity is a major contributor to inflammatory bone loss(IBL)during chronic inflammatory diseases.However,the specific OC precursors(OCPs)responding to inflammatory cues and the underlying mech...Elevated osteoclast(OC)activity is a major contributor to inflammatory bone loss(IBL)during chronic inflammatory diseases.However,the specific OC precursors(OCPs)responding to inflammatory cues and the underlying mechanisms leading to IBL are poorly understood.We identified two distinct OCP subsets:Ly6C^(hi)CD11b^(hi) inflammatory OCPs(iOCPs)induced during chronic inflammation,and homeostatic Ly6C^(hi)CD11b^(lo)OCPs(hOCPs)which remained unchanged.Functional and proteomic characterization revealed that while iOCPs were rare and displayed low osteoclastogenic potential under normal conditions,they expanded during chronic inflammation and generated OCs with enhanced activity.In contrast,hOCPs were abundant and manifested high osteoclastogenic potential under normal conditions but generated OCs with low activity and were unresponsive to the inflammatory environment.Osteoclasts derived from iOCPs expressed higher levels of resorptive and metabolic proteins than those generated from hOCPs,highlighting that different osteoclast populations are formed by distinct precursors.We further identified the TNF-αand S100A8/A9 proteins as key regulators that control the iOCP response during chronic inflammation.Furthermore,we demonstrated that the response of iOCPs but not that of hOCPs was abrogated in tnf-α^(-/-)mice,in correlation with attenuated IBL.Our findings suggest a central role for iOCPs in IBL induction.iOCPs can serve as potential biomarkers for IBL detection and possibly as new therapeutic targets to combat IBL in a wide range of inflammatory conditions.展开更多
Biodegradation of tricalcium phosphate (TCP) ceramics was observed through mixed culture of osteoclasts and TCP discs in vitro in this study. Osteoclasts were isolated from newborn SD rat’s marrow of long bone and cu...Biodegradation of tricalcium phosphate (TCP) ceramics was observed through mixed culture of osteoclasts and TCP discs in vitro in this study. Osteoclasts were isolated from newborn SD rat’s marrow of long bone and cultured on TCP discs. The culture terminated at the 48th h and 96th h respectively. Under an inverted microscope, the osteoclasts imparted round or oval body with multinuclear and many thin processes. These cells were positively stained for tartrate-resistance acid phosphatase (TRAP). Scanning electron microscope showed that many resorption lacunae on TCP disc surface and their diameters were smaller than 20 μm. Osteoclasts were located in the lacunae. At the 96th h, the resorption lacunae become larger and osteoclasts showed degeneration. It is suggested that osteoclasts possess ability to re-absorb TCP ceramics under in vitro culturing condition.展开更多
Osteoclasts,the bone-resorbing cells,play a pivotal role in skeletal development and adult bone remodeling.They also participate in the pathogenesis of various bone disorders.Osteoclasts differentiate from cells of th...Osteoclasts,the bone-resorbing cells,play a pivotal role in skeletal development and adult bone remodeling.They also participate in the pathogenesis of various bone disorders.Osteoclasts differentiate from cells of the monocyte/macrophage lineage upon stimulation of two essential factors,the monocyte/ macrophage colony stimulating factor(M-CSF)and receptor activation of NF-кB ligand(RANKL).M-CSF binds to its receptor c-Fms to activate distinct signaling pathways to stimulate the proliferation and survival of osteoclast precursors and the mature cell.RANKL,however,is the primary osteoclast differentiation factor,and promotes osteoclast differentiation mainly through controlling gene expression by activating its receptor,RANK.Osteoclast function depends on polarization of the cell,induced by integrin αvβ3,to form the resorptive machinery characterized by the attachment to the bone matrix and the formation of the bone-apposed ruffled border.Recent studies have provided new insights into the mechanism of osteoclast differentiation and bone resorption.In particular,c-Fms and RANK signaling have been shown to regulate bone resorption by cross-talking with those activated by integrin αvβ3.This review discusses new advances in the understanding of the mechanisms of osteoclast differentiation and function.展开更多
The skeleton is a dynamic organ that is constantly remodeled. Proteins secreted from bone cells, namely osteoblasts, osteocytes,and osteoclasts exert regulation on osteoblastogenesis, osteclastogenesis, and angiogenes...The skeleton is a dynamic organ that is constantly remodeled. Proteins secreted from bone cells, namely osteoblasts, osteocytes,and osteoclasts exert regulation on osteoblastogenesis, osteclastogenesis, and angiogenesis in a paracrine manner. Osteoblasts secrete a range of different molecules including RANKL/OPG, M-CSF, SEMA3A, WNT5A, and WNT16 that regulate osteoclastogenesis. Osteoblasts also produce VEGFA that stimulates osteoblastogenesis and angiogenesis. Osteocytes produce sclerostin(SOST) that inhibits osteoblast differentiation and promotes osteoclast differentiation. Osteoclasts secrete factors including BMP6, CTHRC1, EFNB2, S1P, WNT10B, SEMA4D, and CT-1 that act on osteoblasts and osteocytes, and thereby influencea A osteogenesis. Osteoclast precursors produce the angiogenic factor PDGF-BB to promote the formation of Type H vessels, which then stimulate osteoblastogenesis. Besides, the evidences over the past decades show that at least three hormones or "osteokines"from bone cells have endocrine functions. FGF23 is produced by osteoblasts and osteocytes and can regulate phosphate metabolism. Osteocalcin(OCN) secreted by osteoblasts regulates systemic glucose and energy metabolism, reproduction, and cognition. Lipocalin-2(LCN2) is secreted by osteoblasts and can influence energy metabolism by suppressing appetite in the brain.We review the recent progresses in the paracrine and endocrine functions of the secretory proteins of osteoblasts, osteocytes, and osteoclasts, revealing connections of the skeleton with other tissues and providing added insights into the pathogenesis of degenerative diseases affecting multiple organs and the drug discovery process.展开更多
To dissect which subset of bone marrow monocyte is the major precursor of osteoclast,3-month-old rat bone marrow was obtained for single-cell RNA sequencing.A total of 6091 cells were acquired for detailed analysis,wi...To dissect which subset of bone marrow monocyte is the major precursor of osteoclast,3-month-old rat bone marrow was obtained for single-cell RNA sequencing.A total of 6091 cells were acquired for detailed analysis,with a median number of 1206 genes detected per cell and 17,959 genes detected in total.A total of 19 cell clusters were recognized,with the main lineages identified as B cells,Granulocytes,Monocytes,T cells,Erythrocytes and Macrophages.Monocytes were further divided into classical monocytes and non-classical monocytes.Compared with non-classical monocytes,classical monocytes highly expressed osteoclast differentiation related genes Mitf,Spi1,Fos and Csf1r.Additionally,biological processes of classical monocytes were related to osteoclast differentiation.qPCR revealed differentially expressed genes of classical monocytes played a role in osteoclast differentiation.In conclusion,classical monocytes were identified as the main precursors of osteoclasts in rats,and may contribute to osteoclast differentiation by regulating S100a4,S100a6,S100a10,Fn1,Vcan and Bcl2a1.The results of this study contribute to the understanding of the origin of osteoclasts and may provide potential biomarkers for early diagnosis of bone metabolic diseases,as well as molecular and cellular targets for clinical intervention in bone metabolic diseases.展开更多
The effect of lanthanum(Ⅲ) (La3+) on cytosolic free calcium ([Ca2+]i) in isolated rabbit mature osteoclasts was studied with the employment of fluo-3/AM as an intracellular calcium-sensitive fluorescent probe by usin...The effect of lanthanum(Ⅲ) (La3+) on cytosolic free calcium ([Ca2+]i) in isolated rabbit mature osteoclasts was studied with the employment of fluo-3/AM as an intracellular calcium-sensitive fluorescent probe by using a confocal laser scanning microscope. La3+ does not alter basal [Ca2+]i levels and cell spread area at the concentration of 1.00×10-8 mol·L-1. However, La3+ at higher concentrations ( 1.00×10-5 and 1.00×10-7 mol·L-1) decreases [Ca2+]i levels and cell spread area, and greater decreases are observed for the higher concentrations of La3+. Since [Ca2+]i affects cytoskeleton and the adhesion properties of osteoclasts, our results seem to suggest that La3+ inhibit bone resorption by decreasing [Ca2+]i in rabbit mature osteoclasts.展开更多
The effects of lanthanum （Ⅲ） on the bone resorbing activity of rabbit mature osteoclasts （OCs） in the presence of osteoblasts （OBs） were studied in vitro by measuring the number and area of absorption pits. La...The effects of lanthanum （Ⅲ） on the bone resorbing activity of rabbit mature osteoclasts （OCs） in the presence of osteoblasts （OBs） were studied in vitro by measuring the number and area of absorption pits. La（Ⅲ） at concentrations ranging from 1.00×10-5 to 1.00×10-8 mol·L-1 show no effect on mature OC number （P>0.05）. In the OC-OB co-culture systems without La（Ⅲ）, osteoblasts alone did not influence the pit number and area whether the two kinds of cells were in contact or not （P>0.05）. Under the OC-OB not-in-contact condition, the effect of La（Ⅲ） on the bone-resorbing activity of OCs was similar to that of La（Ⅲ） in the absence of OBs （P>0.05）. However, while OCs were in direct contact with OBs, the inhibitory effects of La（Ⅲ） on OCs′ bone-resorbing activity decreased at the concentrations of 1.00×10-5, 1.00×10-6 and 1.00×10-7 (mol·L-1), and the promotion effects increased at 1.00×10-8 (mol·L-1) （P<0.05）. The results suggest that direct cell-cell contact between OC and OB be essential for OBs to play their role in regulating the response of OCs to La（Ⅲ）.展开更多
Bone loss and deterioration of bone microarchitecture would increase the bone fragility and fracture risk,leading to the osteoporosis.More and more evidences proved that plant-derived polysaccharides could have a rema...Bone loss and deterioration of bone microarchitecture would increase the bone fragility and fracture risk,leading to the osteoporosis.More and more evidences proved that plant-derived polysaccharides could have a remarkable influence on osteoblasts and osteoclasts,exerting anti-osteoporosis effects.According to the previous research,the extract of Cibotium barumoz,Achyranthes bidentata,Curculigo orchioides,Epimedium brevicornum,Angelica sinensis,Polygonatum sibiricum,Dendrobium officinale,Morinda officinalis,Nelumbo mucifera,Diospyros kaki,Hordeum vulgare,Cistanche deserticola,Commiphora Myrrha and other plant-derived polysaccharides could benefit to the osteoblasts and osteoclasts.The essential mechanisms are mainly related to the activation or inhibition of many factors,including runt-related transcription factor 2(Runx2),B-catenin,osterix(Osx),activator protein-1(AP-1),osteocalcin(OCN/BGP),alkaline phosphatase(ALP),osteopontin(OPN),bone morphogenetic protein(BMP),phosphatidylinositol 3-kinase(PI3K)/C-Jun N-terminal kinase(JNK)/extracellular regulated protein kinase(ERK),osteoprotegerin(OPG),receptor activator of nuclear factor-kB(RANK),monocyte/macrophage colony-stimulating factor(M-CSF),tumor necrosis factor receptor-associated factor 6(TRAF-6),receptor activator of nuclear factor(NF)-KB ligand(RANKL),nuclear factor of activated T cells 1(NFATc1),c-Fos,matrix metallopeptidase-9(MMP-9),glycogen synthase kinase 3β(GSK3B)/B-catenin,nuclear factor E2-related factor 2(Nf2),as well as these related pathways,such as Wnt/p-catenin,BMP-2/SMAD1/5/8,PI3K/AKT,OPG/RANKL/RANK,NF-κB,MAPKs,etc.These plant-derived polysaccharides could improve the dynamic balance of bone formation and resorption through promoting the differentiation and maturation of osteoblast or inhibiting its formation.The reviewed plant-derived polysaccharides and their regulating mechanisms on the osteoclasts and osteoblasts provide the evidences for the development of osteoporosis therapeutics.展开更多
Gorham-Stout disease(GSD)is a sporadic chronic disease characterized by progressive bone dissolution,absorption,and disappearance along with lymphatic vessel infiltration in bone-marrow cavities.Although the osteolyti...Gorham-Stout disease(GSD)is a sporadic chronic disease characterized by progressive bone dissolution,absorption,and disappearance along with lymphatic vessel infiltration in bone-marrow cavities.Although the osteolytic mechanism of GSD has been widely studied,the cause of lymphatic hyperplasia in GSD is rarely investigated.In this study,by comparing the RNA expression profile of osteoclasts(OCs)with that of OC precursors(OCPs)by RNA sequencing,we identified a new factor,semaphorin 3A(Sema3A),which is an osteoprotective factor involved in the lymphatic expansion of GSD.Compared to OCPs,OCs enhanced the growth,migration,and tube formation of lymphatic endothelial cells(LECs),in which the expression of Sema3A is low compared to that in OCPs.In the presence of recombinant Sema3A,the growth,migration,and tube formation of LECs were inhibited,further confirming the inhibitory effect of Sema3A on LECs in vitro.Using an LEC-induced GSD mouse model,the effect of Sema3A was examined by injecting lentivirus-expressing Sema3A into the tibiae in vivo.We found that the overexpression of Sema3A in tibiae suppressed the expansion of LECs and alleviated bone loss,whereas the injection of lentivirus expressing Sema3A short hairpin RNA(shRNA)into the tibiae caused GSD-like phenotypes.Histological staining further demonstrated that OCs decreased and osteocalcin increased after Sema3A lentiviral treatment,compared with the control.Based on the above results,we propose that reduced Sema3A in OCs is one of the mechanisms contributing to the pathogeneses of GSD and that expressing Sema3A represents a new approach for the treatment of GSD.展开更多
Monocyte/macrophage lineage cells are highly plastic and can differentiate into various cells under different environmental stimuli. Bone-resorbing osteoclasts are derived from the monocyte/macrophage lineage in respo...Monocyte/macrophage lineage cells are highly plastic and can differentiate into various cells under different environmental stimuli. Bone-resorbing osteoclasts are derived from the monocyte/macrophage lineage in response to receptor activator of NF-κB ligand (RANKL). However, the epigenetic signature contributing to the fate commitment of monocyte/macrophage lineage differentiation into human osteoclasts is largely unknown. In this study, we identified RANKL-responsive human osteoclast-specific superenhancers (SEs) and SE-associated enhancer RNAs (SE-eRNAs) by integrating data obtained from ChIP-seq, ATAC-seq, nuclear RNA-seq and PRO-seq analyses. RANKL induced the formation of 200 SEs, which are large clusters of enhancers, while suppressing 148 SEs in macrophages. RANKL-responsive SEs were strongly correlated with genes in the osteoclastogenic program and were selectively increased in human osteoclasts but marginally presented in osteoblasts, CD4+ T cells, and CD34+ cells. In addition to the major transcription factors identified in osteoclasts, we found that BATF binding motifs were highly enriched in RANKL-responsive SEs. The depletion of BATF1/3 inhibited RANKL-induced osteoclast differentiation. Furthermore, we found increased chromatin accessibility in SE regions, where RNA polymerase II was significantly recruited to induce the extragenic transcription of SE-eRNAs, in human osteoclasts. Knocking down SE-eRNAs in the vicinity of the NFATc1 gene diminished the expression of NFATc1, a major regulator of osteoclasts, and osteoclast differentiation. Inhibiting BET proteins suppressed the formation of some RANKL-responsive SEs and NFATc1-associated SEs, and the expression of SE-eRNA:NFATc1. Moreover, SE-eRNA:NFATc1 was highly expressed in the synovial macrophages of rheumatoid arthritis patients exhibiting high-osteoclastogenic potential. Our genome-wide analysis revealed RANKL-inducible SEs and SE-eRNAs as osteoclast-specific signatures, which may contribute to the development of osteoclast-specific therapeutic interventions.展开更多
基金supported by the Beijing Municipal Natural Science Foundation(7192110)。
文摘Objective:To investigate whether acupotomy could inhibit subchondral bone remodeling in knee osteoarthritis(KOA)rabbits by regulating the activity of osteoblasts and osteoclasts.Methods:KOA rabbits were prepared by immobilization for 6 and 9 weeks by Videman method.Nine groups of rabbits(control,6 weeks and 9 weeks model,6 weeks and 9 weeks acupotomy,6 weeks and 9 weeks electroacupuncture,and 6 weeks and 9 weeks drug groups)received acupotomy,electroacupuncture and risedronate sodium intervention,respectively,for 3 weeks.Results:Acupotomy can inhibit the activity of osteoclasts and osteoblasts in subchondral bone by reducing the proteins expression of cathepsin K(CK)and tartrate-resistant acid phosphatase(TRAP)and decreasing the proteins expression of osteocalcin(OCN)and alkaline phosphatase(ALP),to intercept the abnormal bone resorption and bone formation of subchondral bone in 6-week and 9-week immobilization-induced KOA rabbits.Conclusion:These findings indicated that acupotomy may be more advantageous than risedronate sodium intervention in modulating subchondral bone remodeling in KOA rabbits,especially in 9-week immobilization-induced KOA rabbits.
基金supported by the National Natural Science Foundation of China(Grant Nos.81770110,81900131,and 82000219)the Anticancer Major Special Project of Tianjin(Grant No.12ZCDZSY18000)+4 种基金the Tianjin Municipal Natural Science Foundation(Grant Nos.18JCYBJC27200 and 18JCQNJC80400)the Tianjin Education Commission Research Project(Grant Nos.2018KJ043 and 2018KJ045)the Tianjin Health and Family Planning Commission(Grant No.15KG150)the Youth Incubation Fund of Tianjin Medical University General Hospital(Grant No.ZYYFY2019020)the Tianjin Science and Technology Planning Project(Grant No.20YFZCSY00060)。
文摘Objective:Myeloma bone disease(MBD)is the most common complication of multiple myeloma(MM).Our previous study showed that the serum levels of C3/C4 in MM patients were significantly positively correlated with the severity of bone disease.However,the mechanism of C3 a/C4 a in osteoclasts MM patients remains unclear.Methods:The formation and function of osteoclasts were analyzed after adding C3 a/C4 a in vitro.RNA-seq analysis was used to screen the potential pathways affecting osteoclasts,and the results were verified by Western blot,q RT-PCR,and pathway inhibitors.Results:The osteoclast area per view induced by 1μg/m L(mean±SD:50.828±12.984%)and 10μg/m L(53.663±12.685%)of C3 a was significantly increased compared to the control group(0μg/m L)(34.635±8.916%)(P<0.001 and P<0.001,respectively).The relative m RNA expressions of genes,OSCAR/TRAP/RANKL/cathepsin K,induced by 1μg/m L(median:5.041,3.726,1.638,and 4.752,respectively)and 10μg/m L(median:5.140,3.702,2.250,and 5.172,respectively)of C3 a was significantly increased compared to the control group(median:3.137,2.004,0.573,and 2.257,respectively)(1μg/m L P=0.001,P=0.003,P<0.001,and P=0.008,respectively;10μg/m L:P<0.001,P=0.019,P<0.001,and P=0.002,respectively).The absorption areas of the osteoclast resorption pits per view induced by 1μg/m L(mean±SD:51.464±11.983%)and 10μg/m L(50.219±12.067%)of C3 a was also significantly increased(33.845±8.331%)(P<0.001 and P<0.001,respectively)compared to the control.There was no difference between the C4 a and control groups.RNA-seq analysis showed that C3 a promoted the proliferation of osteoclasts using the phosphoinositide 3-kinase(PI3 K)signaling pathway.The relative expressions of PIK3 CA/phosphoinositide dependent kinase-1(PDK1)/serum and glucocorticoid inducible protein kinases(SGK3)genes and PI3 K/PDK1/p-SGK3 protein in the C3 a group were significantly higher than in the control group.The activation role of C3 a in osteoclasts of MM patients was reduced by the SGK inhibitor(EMD638683).Conclusions:C3 a activated osteoclasts by regulating the PI3 K/PDK1/SGK3 pathways in MM patients,which was reduced using a SGK inhibitor.Overall,our results identified potential therapeutic targets and strategies for MBD patients。
基金financial support from Orthopaedic Research UK (P 470)Arthritis Research UK (grant 20299 and Oxford EOTC)
文摘The effects of vitamin D on osteoblast mineralization are well documented. Reports of the effects of vitamin D on osteoclasts, however, are conflicting, showing both inhibition and stimulation. Finding that resorbing osteoclasts in human bone express vitamin D receptor(VDR), we examined their response to different concentrations of 25-hydroxy vitamin D_3[25(OH)D_3](100 or 500 nmol·L^(-1)) and 1,25-dihydroxy vitamin D_3[1,25(OH)_2D_3](0.1 or 0.5 nmol·L^(-1)) metabolites in cell cultures. Specifically, CD14+ monocytes were cultured in charcoal-stripped serum in the presence of receptor activator of nuclear factor kappa-B ligand(RANKL) and macrophage colony-stimulating factor(M-CSF). Tartrate-resistant acid phosphatase(TRAP)histochemical staining assays and dentine resorption analysis were used to identify the size and number of osteoclast cells, number of nuclei per cell and resorption activity. The expression of VDR was detected in human bone tissue(ex vivo) by immunohistochemistry and in vitro cell cultures by western blotting.Quantitative reverse transcription-PCR(q RT-PCR) was used to determine the level of expression of vitamin D-related genes in response to vitamin D metabolites. VDR-related genes during osteoclastogenesis, shown by q RT-PCR, was stimulated in response to 500 nmol·L^(-1)of 25(OH)D_3and 0.1–0.5 nmol·L^(-1)of 1,25(OH)_2D_3,upregulating cytochrome P450 family 27 subfamily B member 1(CYP27B1) and cytochrome P450 family 24 subfamily A member 1(CYP24A1). Osteoclast fusion transcripts transmembrane 7 subfamily member 4(tm7sf4) and nuclear factor of activated T-cell cytoplasmic 1(nfatc1) where downregulated in response to vitamin D metabolites. Osteoclast number and resorption activity were also increased. Both 25(OH)D_3and1,25(OH)_2D_3reduced osteoclast size and number when co-treated with RANKL and M-CSF. The evidence for VDR expression in resorbing osteoclasts in vivo and low-dose effects of 1,25(OH)_2D_3on osteoclasts in vitro may therefore provide insight into the effects of clinical vitamin D treatments, further providing a counterpoint to the high-dose effects reported from in vitro experiments.
文摘Bone marrow(BM) cavities are utilized for hematopoiesis and to maintain hematopoietic stem cells(HSCs). HSCs have the ability to self-renew as well as to differentiate into multiple different hematopoietic lineage cells. HSCs produce their daughter cells throughout the lifespan of individuals and thus, maintaining HSCs is crucial for individual life. BM cavities provide a specialized microenvironment termed "niche" to support HSCs. Niches are composed of various types of cells such as osteoblasts, endothelial cells and reticular cells. Osteoclasts are unique cells which resorb bones and are required for BM cavity formation. Loss of osteoclast function or differentiation results in inhibition of BM cavity formation, an osteopetrotic phenotype. Osteoclasts are also reportedly required for hematopoietic stem and progenitor cell(HSPC) mobilization to the periphery from BM cavities. Thus, lack of osteoclasts likely results in inhibition of HSC maintenance and HSPC mobilization. However, we found that osteoclasts are dispensable for hematopoietic stem cell maintenance and mobilization by using three independent osteoclast-less animal models. In this review, I will discuss the roles of osteoclasts in hematopoietic stem cell maintenance and mobilization.
基金Supported by the open foundation of Key Laboratory of Ethnomedicine(Minzu University of China),Ministry of Education,No.:KLEM-KF2018Z02Project for Improving Basic Capabilities of Middle-aged and Young Teachers in Guangxi Institutions of Higher Learning(No.:2020KY07016)+2 种基金Guangxi Health Commission Key Laboratory of Applied Fundamental Research of Zhuang Medicine(Guangxi University of Chinese Medicine)[Gui Wei Ke Jiao Fa(2020)No.17]Training Program under“139”Plan for Developing High-level Medical Talents in Guangxi[Gui Wei Ke Jiao Fa(2020)No.15]Traditional Chinese Medicine-Guangxi first-class discipline[No.Gui Jiao Ke Yan(2018)12]。
文摘Objective:To study the regulatory effect of feilongzhangxue on the levels of HMGB1-TLR4/RANKL-NF-κB signaling pathway related factors HMGB1,RANKL,rank,TRAF-6 and NF-κBp65 in osteoclasts,so as to explore the mechanism of feilongzhangxue intervention in RA;Methods:The osteoclasts with good activity were randomly divided into blank group,methotrexate control group and Zhuang medicine feilongzhang blood containing serum treatment group,which were divided into OC+blank group,OC+methotrexate control group,OC+Zhuang medicine feilongzhang blood containing serum group;The expression of HMGB1,RANKL,rank,TRAF-6 and NF-κBp65 mRNA was detected by RT-PCR;The protein expressions of HMGB1,RANKL,RANK,TRAF-6 and NF-κBp65 were detected by immunofluorescence.Results:PCR results showed that:Compared with the blank group,feilongzhangxue could effectively inhibit the expression levels of HMGB1,RANKL,rank,TRAF-6 and NF-κB p65 mRNA in OC cells,and the inhibitory effect was stronger than methotrexate.Immunofluorescence test results showed that:Compared with the blank group and methotrexate group,feilongzhangxue could effectively inhibit the protein expression of HMGB1,RANKL,rank,TRAF-6 and NF-κB p65 in OC cells.Conclusion:The effect of Zhuang medicine feilongzhangxue on hmgbl-tlr4/rankl-nf-κB signaling pathway of osteoclasts is through the regulation of related factors HMGB1,RANKL,rank,TRAF-6 and NF-κB p65,which may be the key mechanism of Zhuang medicine feilongzhangxue on rheumatoid arthritis.
文摘Osteoporosis is caused by unbalanced bone reconstruction processes which leads to areduction in bone mineral density and changes in bone microstructure,resulting in increased fracture risk.Osteoclasts are the only bone resorption cells in the body of human being,and are closely related to the pathogenesis of osteoporosis.In recent years,there have been more in-depth studies on osteoclasts and bone resorption.The pathogenesis of osteoporosis has always been an important research topic for many medical specialists.Cytokines,as important humoral factors that regulate bone metabolism,play an important role in osteoporosis,especially in the formation,differentiation,and apoptosis of osteoclasts.On the basis of referring to the related studies at home and abroad,this paper analyzes the action mechanism of cytokines such as interleukin-1,interleukin-6,nuclear factor KB ligand receptor activator(RANKL)and tumor necrosis factor(TNF)family members on osteoclasts,and clarifies the relationship between cytokines and osteoporosis,which provides a new angle for the prophylaxis and therapy of osteoporosis.
基金supported by grants from the National Key Research and Development Program of China (2020YFC2002800 to J.L. and 2018YFC1105102 to J.L.)the National Natural Science Foundation of China (91949127, 92168204 to J.L.)the Fundamental Research Funds for the Central Universities (22120210586)
文摘Prostaglandin E2(PGE2), a major cyclooxygenase-2(COX-2) product, is highly secreted by the osteoblast lineage in the subchondral bone tissue of osteoarthritis(OA) patients. However, NSAIDs, including COX-2 inhibitors, have severe side effects during OA treatment. Therefore, the identification of novel drug targets of PGE2 signaling in OA progression is urgently needed. Osteoclasts play a critical role in subchondral bone homeostasis and OA-related pain. However, the mechanisms by which PGE2 regulates osteoclast function and subsequently subchondral bone homeostasis are largely unknown. Here, we show that PGE2 acts via EP4 receptors on osteoclasts during the progression of OA and OA-related pain. Our data show that while PGE2 mediates migration and osteoclastogenesis via its EP2 and EP4 receptors, tissue-specific knockout of only the EP4 receptor in osteoclasts(EP4 Lys M) reduced disease progression and osteophyte formation in a murine model of OA. Furthermore, OA-related pain was alleviated in the EP4 Lys M mice, with reduced Netrin-1 secretion and CGRP-positive sensory innervation of the subchondral bone. The expression of plateletderived growth factor-BB(PDGF-BB) was also lower in the EP4 Lys Mmice, which resulted in reduced type H blood vessel formation in subchondral bone. Importantly, we identified a novel potent EP4 antagonist, HL-43, which showed in vitro and in vivo effects consistent with those observed in the EP4 Lys Mmice. Finally, we showed that the Gαs/PI3 K/AKT/MAPK signaling pathway is downstream of EP4 activation via PGE2 in osteoclasts. Together, our data demonstrate that PGE2/EP4 signaling in osteoclasts mediates angiogenesis and sensory neuron innervation in subchondral bone, promoting OA progression and pain, and that inhibition of EP4 with HL-43 has therapeutic potential in OA.
基金supported by the grants from the National Institution of Health(R01MH107205,R24OD024622,and U01TR002623)supported by Grantin-Aids for Scientific Research from the Japan Society for the Promotion of Science(19K10044).
文摘Tissue-resident macrophages are highly specialized to their tissue-specific microenvironments,activated by various inflammatory signals and modulated by genetic and environmental factors.Osteoclasts and microglia are distinct tissue-resident cells of the macrophage lineage in bone and brain that are responsible for pathological changes in osteoporosis and Alzheimer’s disease(AD),respectively.Osteoporosis is more frequently observed in individuals with AD compared to the prevalence in general population.Diagnosis of AD is often delayed until underlying pathophysiological changes progress and cause irreversible damages in structure and function of brain.As such earlier diagnosis and intervention of individuals at higher risk would be indispensable to modify clinical courses.Pleiotropy is the phenomenon that a genetic variant affects multiple traits and the genetic correlation between two traits could suggest a shared molecular mechanism.In this review,we discuss that the Pyk2-mediated actin polymerization pathway in osteoclasts and microglia in bone and brain,respectively,is the horizontal pleiotropic mediator of shared risk factors for osteoporosis and AD.
基金the support of the Society of Research Associates of the Lautenberg Center and the Harold B.Abramson Chair in Immunologythe grant support provided by the Israel Science Foundation(ISF)+6 种基金the Israeli Ministry of Healththe German-Israeli Project Cooperation of the German Research Foundation(DFG)the Postdoctoral Fellowships program of the German Cancer Research Center(DKFZ)the Israel Cancer Research Fund(ICRF)The Israel Ministry of Science and Technology,the Gross Foundationthe Bruce and Baila Waldholtz fundsthe Joseph and Matilda Melnick Funds。
文摘Elevated osteoclast(OC)activity is a major contributor to inflammatory bone loss(IBL)during chronic inflammatory diseases.However,the specific OC precursors(OCPs)responding to inflammatory cues and the underlying mechanisms leading to IBL are poorly understood.We identified two distinct OCP subsets:Ly6C^(hi)CD11b^(hi) inflammatory OCPs(iOCPs)induced during chronic inflammation,and homeostatic Ly6C^(hi)CD11b^(lo)OCPs(hOCPs)which remained unchanged.Functional and proteomic characterization revealed that while iOCPs were rare and displayed low osteoclastogenic potential under normal conditions,they expanded during chronic inflammation and generated OCs with enhanced activity.In contrast,hOCPs were abundant and manifested high osteoclastogenic potential under normal conditions but generated OCs with low activity and were unresponsive to the inflammatory environment.Osteoclasts derived from iOCPs expressed higher levels of resorptive and metabolic proteins than those generated from hOCPs,highlighting that different osteoclast populations are formed by distinct precursors.We further identified the TNF-αand S100A8/A9 proteins as key regulators that control the iOCP response during chronic inflammation.Furthermore,we demonstrated that the response of iOCPs but not that of hOCPs was abrogated in tnf-α^(-/-)mice,in correlation with attenuated IBL.Our findings suggest a central role for iOCPs in IBL induction.iOCPs can serve as potential biomarkers for IBL detection and possibly as new therapeutic targets to combat IBL in a wide range of inflammatory conditions.
基金This project was supported by grant from the National Natural Science Foundation of China (No.5949320O)
文摘Biodegradation of tricalcium phosphate (TCP) ceramics was observed through mixed culture of osteoclasts and TCP discs in vitro in this study. Osteoclasts were isolated from newborn SD rat’s marrow of long bone and cultured on TCP discs. The culture terminated at the 48th h and 96th h respectively. Under an inverted microscope, the osteoclasts imparted round or oval body with multinuclear and many thin processes. These cells were positively stained for tartrate-resistance acid phosphatase (TRAP). Scanning electron microscope showed that many resorption lacunae on TCP disc surface and their diameters were smaller than 20 μm. Osteoclasts were located in the lacunae. At the 96th h, the resorption lacunae become larger and osteoclasts showed degeneration. It is suggested that osteoclasts possess ability to re-absorb TCP ceramics under in vitro culturing condition.
文摘Osteoclasts,the bone-resorbing cells,play a pivotal role in skeletal development and adult bone remodeling.They also participate in the pathogenesis of various bone disorders.Osteoclasts differentiate from cells of the monocyte/macrophage lineage upon stimulation of two essential factors,the monocyte/ macrophage colony stimulating factor(M-CSF)and receptor activation of NF-кB ligand(RANKL).M-CSF binds to its receptor c-Fms to activate distinct signaling pathways to stimulate the proliferation and survival of osteoclast precursors and the mature cell.RANKL,however,is the primary osteoclast differentiation factor,and promotes osteoclast differentiation mainly through controlling gene expression by activating its receptor,RANK.Osteoclast function depends on polarization of the cell,induced by integrin αvβ3,to form the resorptive machinery characterized by the attachment to the bone matrix and the formation of the bone-apposed ruffled border.Recent studies have provided new insights into the mechanism of osteoclast differentiation and bone resorption.In particular,c-Fms and RANK signaling have been shown to regulate bone resorption by cross-talking with those activated by integrin αvβ3.This review discusses new advances in the understanding of the mechanisms of osteoclast differentiation and function.
基金supported in part by grants from 973 Program from the Chinese Ministry of Science and Technology (MOST) (2014CB964704 and 2015CB964503)the Strategic Priority Research Program of the Chinese Academy of Sciences (Grant No. XDB19000000)the National Natural Science Foundation of China (NSFC) (31371463, 81672119, and 81725010)
文摘The skeleton is a dynamic organ that is constantly remodeled. Proteins secreted from bone cells, namely osteoblasts, osteocytes,and osteoclasts exert regulation on osteoblastogenesis, osteclastogenesis, and angiogenesis in a paracrine manner. Osteoblasts secrete a range of different molecules including RANKL/OPG, M-CSF, SEMA3A, WNT5A, and WNT16 that regulate osteoclastogenesis. Osteoblasts also produce VEGFA that stimulates osteoblastogenesis and angiogenesis. Osteocytes produce sclerostin(SOST) that inhibits osteoblast differentiation and promotes osteoclast differentiation. Osteoclasts secrete factors including BMP6, CTHRC1, EFNB2, S1P, WNT10B, SEMA4D, and CT-1 that act on osteoblasts and osteocytes, and thereby influencea A osteogenesis. Osteoclast precursors produce the angiogenic factor PDGF-BB to promote the formation of Type H vessels, which then stimulate osteoblastogenesis. Besides, the evidences over the past decades show that at least three hormones or "osteokines"from bone cells have endocrine functions. FGF23 is produced by osteoblasts and osteocytes and can regulate phosphate metabolism. Osteocalcin(OCN) secreted by osteoblasts regulates systemic glucose and energy metabolism, reproduction, and cognition. Lipocalin-2(LCN2) is secreted by osteoblasts and can influence energy metabolism by suppressing appetite in the brain.We review the recent progresses in the paracrine and endocrine functions of the secretory proteins of osteoblasts, osteocytes, and osteoclasts, revealing connections of the skeleton with other tissues and providing added insights into the pathogenesis of degenerative diseases affecting multiple organs and the drug discovery process.
基金supported by the National Natural Science Foundation of China(Nos.11572209,11872260)National Natural Science Foundation of China(Key Program,No.11932014).
文摘To dissect which subset of bone marrow monocyte is the major precursor of osteoclast,3-month-old rat bone marrow was obtained for single-cell RNA sequencing.A total of 6091 cells were acquired for detailed analysis,with a median number of 1206 genes detected per cell and 17,959 genes detected in total.A total of 19 cell clusters were recognized,with the main lineages identified as B cells,Granulocytes,Monocytes,T cells,Erythrocytes and Macrophages.Monocytes were further divided into classical monocytes and non-classical monocytes.Compared with non-classical monocytes,classical monocytes highly expressed osteoclast differentiation related genes Mitf,Spi1,Fos and Csf1r.Additionally,biological processes of classical monocytes were related to osteoclast differentiation.qPCR revealed differentially expressed genes of classical monocytes played a role in osteoclast differentiation.In conclusion,classical monocytes were identified as the main precursors of osteoclasts in rats,and may contribute to osteoclast differentiation by regulating S100a4,S100a6,S100a10,Fn1,Vcan and Bcl2a1.The results of this study contribute to the understanding of the origin of osteoclasts and may provide potential biomarkers for early diagnosis of bone metabolic diseases,as well as molecular and cellular targets for clinical intervention in bone metabolic diseases.
文摘The effect of lanthanum(Ⅲ) (La3+) on cytosolic free calcium ([Ca2+]i) in isolated rabbit mature osteoclasts was studied with the employment of fluo-3/AM as an intracellular calcium-sensitive fluorescent probe by using a confocal laser scanning microscope. La3+ does not alter basal [Ca2+]i levels and cell spread area at the concentration of 1.00×10-8 mol·L-1. However, La3+ at higher concentrations ( 1.00×10-5 and 1.00×10-7 mol·L-1) decreases [Ca2+]i levels and cell spread area, and greater decreases are observed for the higher concentrations of La3+. Since [Ca2+]i affects cytoskeleton and the adhesion properties of osteoclasts, our results seem to suggest that La3+ inhibit bone resorption by decreasing [Ca2+]i in rabbit mature osteoclasts.
基金Project supported bythe National Natural Science Foundation of China (20031010 ,20271005)
文摘The effects of lanthanum （Ⅲ） on the bone resorbing activity of rabbit mature osteoclasts （OCs） in the presence of osteoblasts （OBs） were studied in vitro by measuring the number and area of absorption pits. La（Ⅲ） at concentrations ranging from 1.00×10-5 to 1.00×10-8 mol·L-1 show no effect on mature OC number （P>0.05）. In the OC-OB co-culture systems without La（Ⅲ）, osteoblasts alone did not influence the pit number and area whether the two kinds of cells were in contact or not （P>0.05）. Under the OC-OB not-in-contact condition, the effect of La（Ⅲ） on the bone-resorbing activity of OCs was similar to that of La（Ⅲ） in the absence of OBs （P>0.05）. However, while OCs were in direct contact with OBs, the inhibitory effects of La（Ⅲ） on OCs′ bone-resorbing activity decreased at the concentrations of 1.00×10-5, 1.00×10-6 and 1.00×10-7 (mol·L-1), and the promotion effects increased at 1.00×10-8 (mol·L-1) （P<0.05）. The results suggest that direct cell-cell contact between OC and OB be essential for OBs to play their role in regulating the response of OCs to La（Ⅲ）.
基金supported by the College students’innovation Project of Henan University(20221020006).
文摘Bone loss and deterioration of bone microarchitecture would increase the bone fragility and fracture risk,leading to the osteoporosis.More and more evidences proved that plant-derived polysaccharides could have a remarkable influence on osteoblasts and osteoclasts,exerting anti-osteoporosis effects.According to the previous research,the extract of Cibotium barumoz,Achyranthes bidentata,Curculigo orchioides,Epimedium brevicornum,Angelica sinensis,Polygonatum sibiricum,Dendrobium officinale,Morinda officinalis,Nelumbo mucifera,Diospyros kaki,Hordeum vulgare,Cistanche deserticola,Commiphora Myrrha and other plant-derived polysaccharides could benefit to the osteoblasts and osteoclasts.The essential mechanisms are mainly related to the activation or inhibition of many factors,including runt-related transcription factor 2(Runx2),B-catenin,osterix(Osx),activator protein-1(AP-1),osteocalcin(OCN/BGP),alkaline phosphatase(ALP),osteopontin(OPN),bone morphogenetic protein(BMP),phosphatidylinositol 3-kinase(PI3K)/C-Jun N-terminal kinase(JNK)/extracellular regulated protein kinase(ERK),osteoprotegerin(OPG),receptor activator of nuclear factor-kB(RANK),monocyte/macrophage colony-stimulating factor(M-CSF),tumor necrosis factor receptor-associated factor 6(TRAF-6),receptor activator of nuclear factor(NF)-KB ligand(RANKL),nuclear factor of activated T cells 1(NFATc1),c-Fos,matrix metallopeptidase-9(MMP-9),glycogen synthase kinase 3β(GSK3B)/B-catenin,nuclear factor E2-related factor 2(Nf2),as well as these related pathways,such as Wnt/p-catenin,BMP-2/SMAD1/5/8,PI3K/AKT,OPG/RANKL/RANK,NF-κB,MAPKs,etc.These plant-derived polysaccharides could improve the dynamic balance of bone formation and resorption through promoting the differentiation and maturation of osteoblast or inhibiting its formation.The reviewed plant-derived polysaccharides and their regulating mechanisms on the osteoclasts and osteoblasts provide the evidences for the development of osteoporosis therapeutics.
基金supported by the National Natural Science Foundation of China(No.81974321)the Discipline Innovation and Talent Introduction Program to Universities from Ministry of Education of China(Project 111)+3 种基金the Tackling Project for Science and Technology of Xinxiang City(No.GG2019003)the Natural Science Foundation of Henan Province of China(No.212300410173)the State Administration of Traditional Chinese Medicine Young Qi Huang Scholar,and the Innovation Team Project of Scientific Research of Traditional Chinese Medicine of Shanghai Health Committee(No.2022CX001)China.
文摘Gorham-Stout disease(GSD)is a sporadic chronic disease characterized by progressive bone dissolution,absorption,and disappearance along with lymphatic vessel infiltration in bone-marrow cavities.Although the osteolytic mechanism of GSD has been widely studied,the cause of lymphatic hyperplasia in GSD is rarely investigated.In this study,by comparing the RNA expression profile of osteoclasts(OCs)with that of OC precursors(OCPs)by RNA sequencing,we identified a new factor,semaphorin 3A(Sema3A),which is an osteoprotective factor involved in the lymphatic expansion of GSD.Compared to OCPs,OCs enhanced the growth,migration,and tube formation of lymphatic endothelial cells(LECs),in which the expression of Sema3A is low compared to that in OCPs.In the presence of recombinant Sema3A,the growth,migration,and tube formation of LECs were inhibited,further confirming the inhibitory effect of Sema3A on LECs in vitro.Using an LEC-induced GSD mouse model,the effect of Sema3A was examined by injecting lentivirus-expressing Sema3A into the tibiae in vivo.We found that the overexpression of Sema3A in tibiae suppressed the expansion of LECs and alleviated bone loss,whereas the injection of lentivirus expressing Sema3A short hairpin RNA(shRNA)into the tibiae caused GSD-like phenotypes.Histological staining further demonstrated that OCs decreased and osteocalcin increased after Sema3A lentiviral treatment,compared with the control.Based on the above results,we propose that reduced Sema3A in OCs is one of the mechanisms contributing to the pathogeneses of GSD and that expressing Sema3A represents a new approach for the treatment of GSD.
基金supported by National Research Foundation of Korea(NRF)grants funded by the Korean government(MSIP,No.2020R1A2C1006101 and No.2020M3A9B603885111 to SP)by the Tow Foundation(to K-HP-M).Figure 1a was generated by BioRender.
文摘Monocyte/macrophage lineage cells are highly plastic and can differentiate into various cells under different environmental stimuli. Bone-resorbing osteoclasts are derived from the monocyte/macrophage lineage in response to receptor activator of NF-κB ligand (RANKL). However, the epigenetic signature contributing to the fate commitment of monocyte/macrophage lineage differentiation into human osteoclasts is largely unknown. In this study, we identified RANKL-responsive human osteoclast-specific superenhancers (SEs) and SE-associated enhancer RNAs (SE-eRNAs) by integrating data obtained from ChIP-seq, ATAC-seq, nuclear RNA-seq and PRO-seq analyses. RANKL induced the formation of 200 SEs, which are large clusters of enhancers, while suppressing 148 SEs in macrophages. RANKL-responsive SEs were strongly correlated with genes in the osteoclastogenic program and were selectively increased in human osteoclasts but marginally presented in osteoblasts, CD4+ T cells, and CD34+ cells. In addition to the major transcription factors identified in osteoclasts, we found that BATF binding motifs were highly enriched in RANKL-responsive SEs. The depletion of BATF1/3 inhibited RANKL-induced osteoclast differentiation. Furthermore, we found increased chromatin accessibility in SE regions, where RNA polymerase II was significantly recruited to induce the extragenic transcription of SE-eRNAs, in human osteoclasts. Knocking down SE-eRNAs in the vicinity of the NFATc1 gene diminished the expression of NFATc1, a major regulator of osteoclasts, and osteoclast differentiation. Inhibiting BET proteins suppressed the formation of some RANKL-responsive SEs and NFATc1-associated SEs, and the expression of SE-eRNA:NFATc1. Moreover, SE-eRNA:NFATc1 was highly expressed in the synovial macrophages of rheumatoid arthritis patients exhibiting high-osteoclastogenic potential. Our genome-wide analysis revealed RANKL-inducible SEs and SE-eRNAs as osteoclast-specific signatures, which may contribute to the development of osteoclast-specific therapeutic interventions.
