Aqueous Extract of Salvia miltiorrhiza Bunge-Radix Puerariae Herb Pair Attenuates Osteoporosis in Ovariectomized Rats Through Suppressing Osteoclast Differentiation

Traditional herb pair Salvia miltiorrhiza Bunge-Radix Puerariae (DG) owns various biological activities including anti-inflammatory and anti-oxidative stress. Oxidative stress is one high-risk factor for osteoporosis, then effect of DG on osteoporosis and underlying mechanisms was explored both in vivo and in vitro. Firstly, the predication from network pharmacology hinted that DG has the potential for ameliorating osteoporosis. Consistent with predication, DG significantly restored bone loss and deficiency of type II collagen, decreased TRAP and Cathepsin K positive areas in femur. Meanwhile it improved important characteristics of microarchitectural deterioration of tissue, reduced the numbers of NFATc1-positive osteoclast in the vertebra as well as decreased the serum osteoclast-specific cytokine RANKL and OPG release in OVX rats exhibiting its protective effect against osteoporosis. In vitro, DG noticeably decreased osteoclastic-special marker protein expressions of RANK, c-Fos and NFATc1. Furthermore, autophagy pathway p62/LC3B, ROS production and NF-κB were all activated by RANKL stimulation and blocked by DG pretreatment. Moreover, autophagy inhibitors, ROS scavenger, Ca2+ chelator and NF-κB inhibitor remarkably suppressed c-Fos and NFATc1 expressions. Taken together, DG may ameliorate osteoporosis by regulating osteoclast differentiation mediated by autophagy and oxidative stress. This study provided a mechanistic basis for DG treating osteoporosis and offered a safe dose for DG in preventing and improving bone diseases.

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Effects of Isoliquiritigenin on Bone Metabolism and Uterus in Ovariectomized Rats

Current Developments in Nutrition ◽

10.1093/cdn/nzaa045_102 ◽

2020 ◽

Vol 4 (Supplement_2) ◽

pp. 469-469

Author(s):

Lara Sattgast ◽

Carmen Wong ◽

Daniel Doerge ◽

William Helferich ◽

Urszula Iwaniec ◽

...

Keyword(s):

Gene Expression ◽

Bone Resorption ◽

Epithelial Cell ◽

Osteoclast Differentiation ◽

Ovariectomized Rats ◽

Tartrate Resistant Acid Phosphatase ◽

Uterine Weight ◽

Uterine Epithelial Cell ◽

Ovx Rats

Abstract Objectives Isoliquiritigenin (ILQ) is a phenolic compound found in licorice and is a popular dietary supplement. ILQ exhibits model-specific antioxidant, anti-inflammatory, anti-tumor, and estrogenic activities. Limited data suggest the potential of ILQ to prevent or treat osteoporosis. Therefore, this study evaluated the effects of short-duration treatment with ILQ on bone and uterine tissue in estrogen-deplete ovariectomized (ovx) rats. The uterus was important to evaluate because ILQ stimulates proliferation of MCF7 breast cancer cells through an estrogen receptor-dependent mechanism. Methods Six-week-old rats (ovx'd at 4 weeks of age) were fed diets containing 0, 100, 250 or 750 ppm ILQ (n = 5/treatment) for 1 week and sacrificed. Gene expression in femur and uterus, blood markers of global bone turnover, body composition, and uterine weight and epithelial cell height were determined. In addition, the effect of ILQ on in vitro differentiation of osteoclasts derived from bone marrow was assessed. Results Treatment resulted in a dose-dependent increase in serum ILQ with levels reaching 2.4 ± 0.2 mM in rats receiving the highest dose. ILQ did not alter serum levels of osteocalcin, a global marker of bone formation, or osteocalcin gene expression in femur. Additionally, there was little or no effect of ILQ on genes related to osteoblast differentiation or activity in femur. These largely null findings contrast with a reduction in serum CTX, a global marker of bone resorption, at all dose levels of ILQ. At the gene level, ILQ resulted in lower mRNA for genes related to osteoclast differentiation and function in femur, including Acp5 (tartrate resistant acid phosphatase), Timp2 and Mmp2, and suppressed osteoclast differentiation in vitro. ILQ had no effect on the ovx-induced increase in body weight. Ovx resulted in lower uterine weight. Treatment with ILQ at 750 ppm resulted in development of severe uterine epithelial cell hyperplasia in two of five animals. Conclusions ILQ supplementation led to reduced biochemical and gene expression markers of bone resorption in vivo and reduced osteoclast differentiation in vitro without increasing estrogen-dependent gene expression. However, the potential benefits must be weighed against potential detrimental off-target effects, including uterine hypertrophy. Funding Sources NIH [P50AT006268].

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Action of Thioglycosides of 1,2,4-Triazoles and Imidazoles on the Oxidative Stress and Glycosidases in Mice with Molecular Docking

Letters in Drug Design & Discovery ◽

10.2174/1573413715666181212150955 ◽

2019 ◽

Vol 16 (6) ◽

pp. 696-710

Author(s):

Mahmoud Balbaa ◽

Doaa Awad ◽

Ahmad Abd Elaal ◽

Shimaa Mahsoub ◽

Mayssaa Moharram ◽

...

Keyword(s):

Oxidative Stress ◽

Molecular Docking ◽

Active Sites ◽

Biological Activities ◽

Imidazole Ring ◽

Quantitative Structure Activity Relationship ◽

Binding Interaction ◽

Qsar Study

Background: ,2,3-Triazoles and imidazoles are important five-membered heterocyclic scaffolds due to their extensive biological activities. These products have been an area of growing interest to many researchers around the world because of their enormous pharmaceutical scope. Methods: The in vivo and in vitro enzyme inhibition of some thioglycosides encompassing 1,2,4- triazole N1, N2, and N3 and/or imidazole moieties N4, N5, and N6. The effect on the antioxidant enzymes (superoxide dismutase, glutathione S-transferase, glutathione peroxidase and catalase) was investigated as well as their effect on α-glucosidase and β-glucuronidase. Molecular docking studies were carried out to investigate the mode of the binding interaction of the compounds with α- glucosidase and β -glucuronidase. In addition, quantitative structure-activity relationship (QSAR) investigation was applied to find out the correlation between toxicity and physicochemical properties. Results: The decrease of the antioxidant status was revealed by the in vivo effect of the tested compounds. Furthermore, the in vivo and in vitro inhibitory effects of the tested compounds were clearly pronounced on α-glucosidase, but not β-glucuronidase. The IC50 and Ki values revealed that the thioglycoside - based 1,2,4-triazole N3 possesses a high inhibitory action. In addition, the in vitro studies demonstrated that the whole tested 1,2,4-triazole are potent inhibitors with a Ki magnitude of 10-6 and exhibited a competitive type inhibition. On the other hand, the thioglycosides - based imidazole ring showed an antioxidant activity and exerted a slight in vivo stimulation of α-glucosidase and β- glucuronidase. Molecular docking proved that the compounds exhibited binding affinity with the active sites of α -glucosidase and β-glucuronidase (docking score ranged from -2.320 to -4.370 kcal/mol). Furthermore, QSAR study revealed that the HBD and RB were found to have an overall significant correlation with the toxicity. Conclusion: These data suggest that the inhibition of α-glucosidase is accompanied by an oxidative stress action.

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Effect of Shuangdan Mingmu capsule, a Chinese herbal formula, on oxidative stress-induced apoptosis of pericytes through PARP/GAPDH pathway

BMC Complementary Medicine and Therapies ◽

10.1186/s12906-021-03238-w ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Fujiao Nie ◽

Jiazhao Yan ◽

Yanjun Ling ◽

Zhengrong Liu ◽

Chaojun Fu ◽

...

Keyword(s):

Oxidative Stress ◽

Nitric Oxide ◽

Oxidative Damage ◽

Damage Model ◽

B Cell Lymphoma ◽

Cell Counting ◽

Network Pharmacology ◽

Diabetic Patients ◽

Induced Apoptosis

Abstract Background Diabetic retinopathy (DR) has become a worldwide concern because of the rising prevalence rate of diabetes mellitus (DM). Despite much energy has been committed to DR research, it remains a difficulty for diabetic patients all over the world. Since apoptosis of retinal microvascular pericytes (RMPs) is the early characteristic of DR, this study aimed to reveal the mechanism of Shuangdan Mingmu (SDMM) capsule, a Chinese patent medicine, on oxidative stress-induced apoptosis of pericytes implicated with poly (ADP-ribose) polymerase (PARP) / glyceraldehyde 3-phosphate dehydrogenase (GAPDH) pathway. Methods Network pharmacology approach was performed to predict biofunction of components of SDMM capsule dissolved in plasma on DR. Both PARP1 and GAPDH were found involved in the hub network of protein-protein interaction (PPI) of potential targets and were found to take part in many bioprocesses, including responding to the regulation of reactive oxygen species (ROS) metabolic process, apoptotic signaling pathway, and response to oxygen levels through enrichment analysis. Therefore, in vitro research was carried out to validate the prediction. Human RMPs cultured with media containing 0.5 mM hydrogen oxide (H2O2) for 4 h was performed as an oxidative-damage model. Different concentrations of SDMM capsule, PARP1 inhibitor, PARP1 activation, and GAPDH inhibitor were used to intervene the oxidative-damage model with N-Acetyl-L-cysteine (NAC) as a contrast. Flow cytometry was performed to determine the apoptosis rate of cells and the expression of ROS. Cell counting kit 8 (CCK8) was used to determine the activity of pericytes. Moreover, nitric oxide (NO) concentration of cells supernatant and expression of endothelial nitric oxide synthase (eNOS), superoxide dismutase (SOD), B cell lymphoma 2 (BCL2), vascular endothelial growth factor (VEGF), endothelin 1 (ET1), PARP1, and GAPDH were tested through RT-qPCR, western blot (WB), or immunocytochemistry (ICC). Results Overproduction of ROS, high apoptotic rate, and attenuated activity of pericytes were observed after cells were incubated with media containing 0.5 mM H2O2. Moreover, downregulation of SOD, NO, BCL2, and GAPDH, and upregulation of VEGFA, ET1, and PARP1 were discovered after cells were exposed to 0.5 mM H2O2 in this study, which could be improved by PARP1 inhibitor and SDMM capsule in a dose-dependent way, whereas worsened by PARP1 activation and GAPDH inhibitor. Conclusions SDMM capsule may attenuate oxidative stress-induced apoptosis of pericytes through downregulating PARP expression and upregulating GAPDH expression.

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Capsaicin, a TRPV1 Ligand, Suppresses Bone Resorption by Inhibiting the Prostaglandin E Production of Osteoblasts, and Attenuates the Inflammatory Bone Loss Induced by Lipopolysaccharide

ISRN Pharmacology ◽

10.5402/2012/439860 ◽

2012 ◽

Vol 2012 ◽

pp. 1-6 ◽

Cited By ~ 11

Author(s):

Megumi Kobayashi ◽

Kenta Watanabe ◽

Satoshi Yokoyama ◽

Chiho Matsumoto ◽

Michiko Hirata ◽

...

Keyword(s):

Bone Resorption ◽

Bone Loss ◽

Transient Receptor Potential ◽

Interleukin 1 ◽

Osteoclast Differentiation ◽

Bone Diseases ◽

Prostaglandin E ◽

Transient Receptor Potential Vanilloid ◽

Cox 2

Capsaicin, a transient receptor potential vanilloid type 1 (TRPV1) ligand, regulates nerve-related pain-sensitive signals, inflammation, and cancer growth. Capsaicin suppresses interleukin-1-induced osteoclast differentiation, but its roles in bone tissues and bone diseases are not known. This study examined the effects of capsaicin on inflammatory bone resorption and prostaglandin E (PGE) production induced by lipopolysaccharide (LPS) in vitro and on bone mass in LPS-treated mice in vivo. Capsaicin suppressed osteoclast formation, bone resorption, and PGE production induced by LPS in vitro. Capsaicin suppressed the expression of cyclooxygenase-2 (COX-2) and membrane-bound PGE synthase-1 (mPGES-1) mRNAs and PGE production induced by LPS in osteoblasts. Capsaicin may suppress PGE production by inhibiting the expression of COX-2 and mPGES-1 in osteoblasts and LPS-induced bone resorption by TRPV1 signals because osteoblasts express TRPV1. LPS treatment markedly induced bone loss in the femur in mice, and capsaicin significantly restored the inflammatory bone loss induced by LPS in mice. TRPV1 ligands like capsaicin may therefore be potentially useful as clinical drugs targeting bone diseases associated with inflammatory bone resorption.

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Network Pharmacology-Based Strategy for the Investigation of the Anti-Osteoporosis Effects and Underlying Mechanism of Zhuangguguanjie Formulation

Frontiers in Pharmacology ◽

10.3389/fphar.2021.727808 ◽

2021 ◽

Vol 12 ◽

Author(s):

Wang Gong ◽

Xingren Chen ◽

Tianshu Shi ◽

Xiaoyan Shao ◽

Xueying An ◽

...

Keyword(s):

Bone Resorption ◽

Bone Formation ◽

Bone Mass ◽

Signaling Pathway ◽

Osteoclast Differentiation ◽

Enrichment Analysis ◽

Underlying Mechanism ◽

Network Pharmacology ◽

Biological Processes

As the society is aging, the increasing prevalence of osteoporosis has generated huge social and economic impact, while the drug therapy for osteoporosis is limited due to multiple targets involved in this disease. Zhuangguguanjie formulation (ZG) is extensively used in the clinical treatment of bone and joint diseases, but the underlying mechanism has not been fully described. This study aimed to examine the therapeutic effect and potential mechanism of ZG on postmenopausal osteoporosis. The ovariectomized (OVX) mice were treated with normal saline or ZG for 4 weeks after ovariectomy following a series of analyses. The bone mass density (BMD) and trabecular parameters were examined by micro-CT. Bone remodeling was evaluated by the bone histomorphometry analysis and ELISA assay of bone turnover biomarkers in serum. The possible drug–disease common targets were analyzed by network pharmacology. To predict the potential biological processes and related pathways, GO/KEGG enrichment analysis was performed. The effects of ZG on the differentiation phenotype of osteoclasts and osteoblasts and the predicted pathway were verified in vitro. The results showed that ZG significantly improved the bone mass and micro-trabecular architecture in OVX mice compared with untreated OVX mice. ZG could promote bone formation and inhibit bone resorption to ameliorate ovariectomy-induced osteoporosis as evidenced by increased number of osteoblast (N.Ob/Tb.Pm) and decreased number of osteoclast (N.Oc/Tb.Pm) in treated group compared with untreated OVX mice. After identifying potential drug–disease common targets by network pharmacology, GO enrichment analysis predicted that ZG might affect various biological processes including osteoblastic differentiation and osteoclast differentiation. The KEGG enrichment analysis suggested that PI3K/Akt and mTOR signaling pathways could be the possible pathways. Furthermore, the experiments in vitro validated our findings. ZG significantly down-regulated the expression of osteoclast differentiation markers, reduced osteoclastic resorption, and inhibited the phosphorylation of PI3K/Akt, while ZG obviously up-regulated the expression of osteogenic biomarkers, promoted the formation of calcium nodules, and hampered the phosphorylation of 70S6K1/mTOR, which can be reversed by the corresponding pathway activator. Thus, our study suggested that ZG could inhibit the PI3K/Akt signaling pathway to reduce osteoclastic bone resorption as well as hamper the mTORC1/S6K1 signaling pathway to promote osteoblastic bone formation.

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Valsartan Reduces Left Ventricular Hypertrophy in Ovariectomized Spontaneous Hypertensive Rats

American Journal of Hypertension ◽

10.1093/ajh/hpz186 ◽

2020 ◽

Vol 33 (4) ◽

pp. 371-371

Author(s):

Hong Ding ◽

Ning-ying Li ◽

Xiang Zhang ◽

Pan-pan Zhang ◽

Jing Yu

Keyword(s):

Oxidative Stress ◽

Wall Thickness ◽

Interventricular Septum ◽

Posterior Wall ◽

Left Ventricular ◽

Ovariectomized Rats ◽

Control Group ◽

Hypertensive Rats ◽

Sham Control ◽

Ovx Rats

Abstract Objective To investigate the effects of valsartan on left ventricular mass, function, and oxidative stress in ovariectomized spontaneous hypertensive rats (SHR). Methods Twelve-week-old female SHRs were randomly divided into ovariectomy (OVX) control (n = 12), OVX + valsartan (n = 12), sham control (Sham, n = 13), and Sham + valsartan (n = 14) groups. Valsartan (30 mg/kg/day) or double-distilled water was given by oral gavage. After 12 weeks of valsartan or water treatment, left ventricular wall thickness and function, superoxide dismutase (SOD), glutathione peroxidase (GSH), and 8-hydroxydeoxyguanosine (8-OHdG) were assessed. Results There was a significant interaction between ovariectomy and valsartan on interventricular end-diastolic septum thickness (IVSTd), end-systolic interventricular septum thickness (IVSTs), left ventricular end-diastolic posterior wall thickness (LVPWTd), and left ventricular end diastolic diameter (LVEDD) (P < 0.05). Valsartan treatment in OVX rats decreased IVSTd, IVSTs, LVPWTd, and LVPWTs compared to OVX control (P < 0.05). Compared with Sham + control group, LVESP and ±dP/dt of LV were decreased while LVEDP was increased in OVX + control group (all P < 0.05). After valsartan treatment, LVESP and ±dP/dt of LV were increased and LVEDP was decreased in ovariectomized rats (all P < 0.05). Ovariectomy decreased GSH and SOD levels and increased 8-OHdG levels, which were reversed by valsartan treatment (all P < 0.05). Conclusion Valsartan treatment decreases oxidative stress, reduces LV hypertrophy, and improves cardiac function in overiectomized SHR.

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Baru Pulp (Dipteryx alata Vogel): Fruit from the Brazilian Savanna Protects against Oxidative Stress and Increases the Life Expectancy of Caenorhabditis elegans via SOD-3 and DAF-16

Biomolecules ◽

10.3390/biom10081106 ◽

2020 ◽

Vol 10 (8) ◽

pp. 1106

Author(s):

Natasha Rios Leite ◽

Laura Costa Alves de Araújo ◽

Paola dos Santos da Rocha ◽

Danielle Araujo Agarrayua ◽

Daiana Silva Ávila ◽

...

Keyword(s):

Oxidative Stress ◽

Caenorhabditis Elegans ◽

Life Expectancy ◽

Biological Activities ◽

Radical Scavenging ◽

Brazilian Savanna ◽

Beneficial Effects ◽

Dipteryx Alata

Fruits are sources of bioactive compounds that are responsible for several biological activities. Therefore, this study aimed to identify the chemical composition of the pulp of the Brazilian Savanna fruit Dipteryx alata; evaluate its toxic effects, influence on the life expectancy of the nematode Caenorhabditis elegans, and its antioxidant activities in vitro and in vivo; and describe the mechanisms involved. The chemical compounds identified include phenols, terpenes, fatty acid derivatives, vitamins, and a carboxylic acid. The in vitro antioxidant activity was demonstrated by radical scavenging methods. in vivo, the D. alata fruit pulp was not toxic and promoted resistance to oxidative stress in nematodes exposed to a chemical oxidizing agent. Furthermore, it promoted an increased life expectancy in wild-type nematodes and increased the expression of superoxide dismutase and the nuclear translocation of DAF-16. These results suggest that the beneficial effects identified are related to these two genes, which are involved in the regulation of metabolic activities, the control of oxidative stress, and the lifespan of C. elegans. These beneficial effects, which may be related to its chemical constituents, demonstrate its potential use as a functional and/or nutraceutical food.

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Fucoidan Prevents RANKL-Stimulated Osteoclastogenesis and LPS-Induced Inflammatory Bone Loss via Regulation of Akt/GSK3β/PTEN/NFATc1 Signaling Pathway and Calcineurin Activity

Marine Drugs ◽

10.3390/md17060345 ◽

2019 ◽

Vol 17 (6) ◽

pp. 345 ◽

Cited By ~ 5

Author(s):

Sheng-Hua Lu ◽

Yi-Jan Hsia ◽

Kuang-Chung Shih ◽

Tz-Chong Chou

Keyword(s):

Bone Loss ◽

Molecular Mechanisms ◽

Bone Erosion ◽

Osteoclast Differentiation ◽

Bone Diseases ◽

Nuclear Factor ◽

Activated T Cells ◽

Calcineurin Activity

Excessive osteoclast differentiation and/or function plays a pivotal role in the pathogenesis of bone diseases such as osteoporosis and rheumatoid arthritis. Here, we examined whether fucoidan, a sulfated polysaccharide present in brown algae, attenuates receptor activator of nuclear factor-κB ligand (RANKL)-stimulated osteoclastogenesis in vitro and lipopolysaccharide (LPS)-induced bone resorption in vivo, and investigated the molecular mechanisms involved. Our results indicated that fucoidan significantly inhibited osteoclast differentiation in RANKL-stimulated macrophages and the bone resorbing activity of osteoclasts. The effects of fucoidan may be mediated by regulation of Akt/GSK3β/PTEN signaling and suppression of the increase in intracellular Ca2+ level and calcineurin activity, thereby inhibiting the translocation of nuclear factor-activated T cells c1 (NFATc1) into the nucleus. However, fucoidan-mediated NFATc1 inactivation was greatly reversed by kenpaullone, a GSK3β inhibitor. In addition, using microcomputer tomography (micro-CT) scanning and bone histomorphometry, we found that fucoidan treatment markedly prevented LPS-induced bone erosion in mice. Collectively, we demonstrated that fucoidan was capable of inhibiting osteoclast differentiation and inflammatory bone loss, which may be modulated by regulation of Akt/GSK3β/PTEN/NFATc1 and Ca2+/calcineurin signaling cascades. These findings suggest that fucoidan may be a potential agent for the treatment of osteoclast-related bone diseases.

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Inhibition of Oxidative Neurotoxicity and Scopolamine-Induced Memory Impairment by γ-Mangostin: In Vitro and In Vivo Evidence

Oxidative Medicine and Cellular Longevity ◽

10.1155/2019/3640753 ◽

2019 ◽

Vol 2019 ◽

pp. 1-14 ◽

Cited By ~ 4

Author(s):

Youngmun Lee ◽

Sunyoung Kim ◽

Yeonsoo Oh ◽

Young-Mi Kim ◽

Young-Won Chin ◽

...

Keyword(s):

Oxidative Stress ◽

Memory Impairment ◽

Neuronal Damage ◽

Biological Activities ◽

Neuronal Cell Death ◽

Neuronal Cell ◽

Ros Generation ◽

Garcinia Mangostana

Among a series of xanthones identified from mangosteen, the fruit of Garcinia mangostana L. (Guttifereae), α- and γ-mangostins are known to be major constituents exhibiting diverse biological activities. However, the effects of γ-mangostin on oxidative neurotoxicity and impaired memory are yet to be elucidated. In the present study, the protective effect of γ-mangostin on oxidative stress-induced neuronal cell death and its underlying action mechanism(s) were investigated and compared to that of α-mangostin using primary cultured rat cortical cells. In addition, the effect of orally administered γ-mangostin on scopolamine-induced memory impairment was evaluated in mice. We found that γ-mangostin exhibited prominent protection against H2O2- or xanthine/xanthine oxidase-induced oxidative neuronal death and inhibited reactive oxygen species (ROS) generation triggered by these oxidative insults. In contrast, α-mangostin had no effects on the oxidative neuronal damage or associated ROS production. We also found that γ-mangostin, not α-mangostin, significantly inhibited H2O2-induced DNA fragmentation and activation of caspases 3 and 9, demonstrating its antiapoptotic action. In addition, only γ-mangostin was found to effectively inhibit lipid peroxidation and DPPH radical formation, while both mangostins inhibited β-secretase activity. Furthermore, we observed that the oral administration of γ-mangostin at dosages of 10 and 30 mg/kg markedly improved scopolamine-induced memory impairment in mice. Collectively, these results provide both in vitro and in vivo evidences for the neuroprotective and memory enhancing effects of γ-mangostin. Multiple mechanisms underlying this neuroprotective action were suggested in this study. Based on our findings, γ-mangostin could serve as a potentially preferable candidate over α-mangostin in combatting oxidative stress-associated neurodegenerative diseases including Alzheimer’s disease.

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Lipopolysaccharide- TLR-4 Axis regulates Osteoclastogenesis independent of RANKL/RANK signaling

BMC Immunology ◽

10.1186/s12865-021-00409-9 ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Mohammed S. AlQranei ◽

Linda T. Senbanjo ◽

Hanan Aljohani ◽

Therwa Hamza ◽

Meenakshi A. Chellaiah

Keyword(s):

Tumor Necrosis Factor ◽

Tumor Necrosis ◽

Osteoclast Differentiation ◽

Bone Diseases ◽

Raw 264.7 Cells ◽

Gram Negative ◽

Tnf Α ◽

Necrosis Factor ◽

In Cells

Abstract Background Lipopolysaccharide (LPS) is an endotoxin and a vital component of gram-negative bacteria’s outer membrane. During gram-negative bacterial sepsis, LPS regulates osteoclast differentiation and activity, in addition to increasing inflammation. This study aimed to investigate how LPS regulates osteoclast differentiation of RAW 264.7 cells in vitro. Results Herein, we revealed that RAW cells failed to differentiate into mature osteoclasts in vitro in the presence of LPS. However, differentiation occurred in cells primed with receptor activator of nuclear factor-kappa-Β ligand (RANKL) for 24 h and then treated with LPS for 48 h (henceforth, denoted as LPS-treated cells). In cells treated with either RANKL or LPS, an increase in membrane levels of toll-like receptor 4 (TLR4) receptor was observed. Mechanistically, an inhibitor of TLR4 (TAK-242) reduced the number of osteoclasts as well as the secretion of tumor necrosis factor (TNF)-α in LPS-treated cells. RANKL-induced RAW cells secreted a very basal level TNF-α. TAK-242 did not affect RANKL-induced osteoclastogenesis. Increased osteoclast differentiation in LPS-treated osteoclasts was not associated with the RANKL/RANK/OPG axis but connected with the LPS/TLR4/TNF-α tumor necrosis factor receptor (TNFR)-2 axis. We postulate that this is because TAK-242 and a TNF-α antibody suppress osteoclast differentiation. Furthermore, an antibody against TNF-α reduced membrane levels of TNFR-2. Secreted TNF-α appears to function as an autocrine/ paracrine factor in the induction of osteoclastogenesis independent of RANKL. Conclusion TNF-α secreted via LPS/TLR4 signaling regulates osteoclastogenesis in macrophages primed with RANKL and then treated with LPS. Our findings suggest that TLR4/TNF-α might be a potential target to suppress bone loss associated with inflammatory bone diseases, including periodontitis, rheumatoid arthritis, and osteoporosis.

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