Search

Four new ruthenium (II) polypyridine complexes bearing 18β-glycyrtinic acid derivatives, [Ru(bpy)2L](PF6)2 (Ru1), [Ru(dmb)2L](PF6)2 (Ru2), [Ru(dtb)2L](PF6)2 (Ru3) and [Ru(phen)2L](PF6)2 (Ru4) (bpy = 2,2-bipyridine, dmb = 4,4'-dimethyl-2,2'-bipyridine, dtb = 4,4'-di-tert-butyl-2,2'-bipyridine, phen = 1,10- phenanthroline and L is the GA modified new ligand) were designed and synthesized. Their antimicrobial activities against Staphylococcus aureus (S. aureus) were evaluated and all complexes showed obvious inhibitory effect, especially the minimum inhibitory concentration (MIC) value of Ru2 was 3.9 µg·mL-1. Meanwhile, Ru2 was found that could significantly inhibit the formation of biofilms. The membrane-compromising action mode was suggested to be their potential antibactericidal mechanism. In hemolysis experiments, Ru2 hardly showed cytotoxicity to mammalian erythrocytes. Furthermore, the synergism between Ru2 and common antibiotics, such as ampicillin, chloramphenicol, tetracyclinesand and ofloxacin, against S. aureus was also detected using the checkerboard method. Finally, a mouse skin infection model was established to evaluate the antibacterial activity of Ru2 in vivo, and the results showed that Ru2 could effectively promote wound healing in mice infecting with S. aureus. Moreover, the results of histopathological research were consistent with the results of hemolysis test, indicating that the Ru2 complex was almost non-toxic. Thus, it was demonstrated that the polypyridine ruthenium complexes modified with glycyrtinic acid (GA) is a promising strategy for developing interesting antibacterial agents.

New effective antimicrobial agents with novel modes of action are urgently needed due to the continued emergence of drug-resistant bacteria. Here, three ruthenium complexes functionalized with benzothiophene: [Ru(phen)2(BTPIP)](ClO4)2 (Ru(II)-1), [Ru(dmp)2(BTPIP)](ClO4)2 (Ru(II)-2) and [Ru(dmb)2(BTPIP)](ClO4)2 (Ru(II)-3) (dmb = 4,4'-dimethyl-2,2'-bipyridine, phen = 1,10-phenanthroline, dmp = 2,9-dimethyl-1,10-phenanthroline) have been synthesized and their antimicrobial activities in vitro were assessed. Minimum inhibitory concentration (MIC) assays indicated that the three Ru(II)-1, Ru(II)-2 and Ru(II)-3 complexes all showed antibacterial activity against Staphylococcus aureus and Pseudomonas aeruginosa. The most active Ru(II)-3 complex was further tested against biofilms. Furthermore, it was also tested whether complex Ru(II)-3 could serve as an antibacterial adjuvant. Interestingly, the checkerboard data showed that Ru(II)-3 selectively exhibited synergism with aminoglycoside antibiotics. More importantly, the observed synergetic effect might be attributed to the inhibition of the regulatory function of SaCcpA. Finally, in vivo bacterial infection treatment studies through a murine skin infection model and skin irritation test were also conducted. All in all, these results confirmed that ruthenium complexes functionalized with benzothiophene have good antimicrobial activity against Staphylococcus aureus.

In chronic infectious diseases caused by gram-negative bacteria, such as osteomyelitis, septic arthritis, and periodontitis, osteoclastic activity is enhanced with elevated inflammation, which disturbs the bone homeostasis and results in osteolysis. Lipopolysaccharide (LPS), as a bacteria product, plays an important role in this process. Recent evidence shows that an antimalarial drug artesunate attenuates LPS-induced osteolysis independent of RANKL. In this study we evaluated the effects of artesunate on LPS-induced osteoclastogenesis in vitro and femur osteolysis in vivo, and explored the mechanisms underlying the effects of artesunate on LPS-induced osteoclast differentiation independent of RANKL. In preosteoclastic RAW264.7 cells, we found that artesunate (1.56−12.5 μM) dose dependently inhibited LPS-induced osteoclast formation accompanied by suppressing LPS-stimulated osteoclast-related gene expression (Fra-2, TRAP, Cathepsin K, β3-integrin, DC-STAMP, and Atp6v0d2). We showed that artesunate (3.125−12.5 µM) inhibited LPS-stimulated nuclear factor of activated T cells c1 (NFATc1) but not NF-κB transcriptional activity; artesunate (6.25, 12.5 μM) significantly inhibited LPS-stimulated NFATc1 protein expression. Furthermore, artesunate treatment markedly suppressed LPS-induced Ca 2+ influx, and decreased the expression of PP2B-Aα (calcineurin) and pPLCγ1 in the cells. In addition, artesunate treatment significantly decreased the expression of upstream signals TLR4 and TRAF6 during LPS-induced osteoclastogenesis. Administration of artesunate (10 mg/kg, ip) for 8 days effectively inhibited serum TNF-α levels and ameliorated LPS (5 mg/kg, ip)-induced inflammatory bone loss in vivo. Taken together, artesunate attenuates LPS-induced inflammatory osteoclastogenesis by inhibiting the expression of TLR4/TRAF6 and the downstream PLCγ1-Ca 2+ -NFATc1 signaling pathway. Artesunate is a valuable choice to treat bone loss induced by gram-negative bacteria infection or inflammation in RANKL-independent pathway.

Sinomenine (SIN) is an anti-inflammatory and antiarthritic alkaloid derived from Sinomenium acutum, and the product Zhengqing Fengtongning produced from SIN has been marketed in China for treating rheumatoid arthritis (RA). Interestingly, we recently found that SIN could significantly ameliorate bone destruction induced by breast cancer cells in mice. Micro-CT examination showed that bone loss of the trabecular bones in tumor-bearing mice was markedly decreased by i.p. treatment of SIN at 150 mg/kg body weight. A mechanistic study demonstrated that SIN could suppress osteoclast formation and bone absorption induced by both MDA-MB-231 cells and MDA-MB-231 cell-conditioned medium (MDA-MB-231 CM) in preosteoclastic RAW264.7 cells. The MDA-MB-231 CM-induced osteoclast-related genes TRAP and OSCAR were obviously downregulated by SIN. In addition, mRNA expression of c-Fos and NFATc1 and nuclear translocation of c-Fos and NFATc1 protein were inhibited by SIN during MDA-MB-231 CM-induced osteoclastogenesis, while NF-κB signaling was not impacted by SIN. More interestingly, SIN was demonstrated to decrease hIL-8 mRNA expression in cultured MDA-MB-231 cells and to inhibit hIL-8 protein expression in MDA-MB-231 cells cocultured with preosteoclastic RAW264.7 cells while simultaneously downregulating CXCR1, the ligand of IL-8 related to bone destruction, during MDA-MB-231 CM-induced osteoclastogenesis. Previously, IL-8/CXCR1 was reported to be associated with the pathogenesis and progression of RA, and SIN was observed to markedly ameliorate bone erosion of RA patients. Our current findings may extend the utilization of SIN to preventing osteoclastogenesis and bone destruction in breast cancer patients and may enable IL-8/CXCR1 to serve as new targets for both anticancer and antiarthritic drug discovery.

Background Osteolytic diseases share symptoms such as bone loss, fracture and pain, which are caused by over-activated osteoclasts. Targeting osteoclast differentiation has emerged as a therapeutic strategy clinically. Dendrobine is an alkaloid isolated from Chinese herb Dendrobium nobile, with knowing effects of analgesia and anti-inflammation. The roles of dendrobine on osteoclasts and osteolysis remain unclear. Purpose Herein, the possible roles of dendrobine in osteoclastogenesis, inflammatory osteolysis and the underlying mechanism were explored. Methods Bone marrow-derived macrophages (BMMs) and RAW264.7 cells were employed to evaluate the roles of dendrobine on osteoclastogenesis, bone absorption and the underlying mechanism in vitro. LPS injection was used to cause inflammatory osteolysis in vivo. Results Dendrobine repressed osteoclastogenesis, bone resorption induced by receptor activator of nuclear factor kappa B ligand (RANKL) in vitro. Mechanistically, dendrobine inhibited RANKL-upregulated intracellular (ROS), p-p38, c-Fos expression and nuclear factor of activated T cells (NFATc1) nuclear translocation. Osteoclastic genes were reduced, and among them matrix metalloproteinase 9 (MMP9) mRNA was dramatically blocked by dendrobine. Moreover, it substantially suppressed MMP9 protein expression during osteoclastogenesis in vitro. Accordingly, oral 20 mg/kg/day dendrobine was capable of preventing LPS-induced osteolysis with decreased osteoclasts in vivo. Conclusion Taken together, dendrobine suppresses osteoclastogenesis through restraining ROS, p38-c-Fos and NFATc1-MMP9 in vitro, thus attenuates inflammatory osteolysis in vivo. This finding supports the discover of dendrobine as a novel osteoclast inhibitor for impeding bone erosion in the future.

Currently, dendritic cell-specific transmembrane protein (DC-STAMP), a multipass transmembrane protein, is considered as the master regulator of cell–cell fusion, which underlies the formation of functional multinucleated osteoclasts. Thus, DC-STAMP has become a promising target for osteoclast-associated osteolytic diseases. In this study, we investigated the effects of oridonin (ORI), a natural tetracyclic diterpenoid compound isolated from the traditional Chinese herb Rabdosia rubescens, on osteoclastogenesis in vivo and ex vivo. ICR mice were injected with LPS (5 mg/kg, ip, on day 0 and day 4) to induce inflammatory bone destruction. Administration of ORI (2, 10 mg·kg(−1)·d(−1), ig, for 8 days) dose dependently ameliorated inflammatory bone destruction and dramatically decreased DC-STAMP protein expression in BMMs isolated from LPS-treated mice. Treatment of preosteoclast RAW264.7 cells with ORI (0.78–3.125 μM) dose dependently inhibited both mRNA and protein levels of DC-STAMP, and suppressed the following activation of NFATc1 during osteoclastogenesis. Knockdown of DC-STAMP in RAW264.7 cells abolished the inhibitory effects of ORI on RANKL-induced NFATc1 activity and osteoclast formation. In conclusion, we show for the first time that ORI effectively attenuates inflammation-induced bone loss by suppressing DC-STAMP expression, suggesting that ORI is a potential agent against inflammatory bone diseases.

IKK-β is indispensable for inflammatory osteolysis, the functional residues of IKK-β are therapeutic drug targets for developing inhibitors to treat multiple diseases now. Thus it remains appealing to find the new residues of IKK-β to influence osteoclasts for alleviating bone loss diseases such as rheumatoid arthritis (RA). By employing IKK-β cysteine 46-A transgenic (IKK-βC46A) mice, we found that mutation of cysteine 46 to alanine in IKK-β exacerbated inflammatory bone destruction in vivo, and increased osteoclast differentiation and bone resorption ex vivo and in vitro. Consistent with these, IKK-β kinase activity as well as c-Fos, NFATc1 were up-regulated in bone marrow macrophages (BMMs) from IKK-βC46A mice during RANKL-induced osteoclastogenesis. Of interesting, we further identified and demonstrated that the expressions of mPGES-1 and caveolin-1 were heightened in BMMs of IKK-βC46A mice compared to those in WT mice in RANKL-induced osteoclastogenesis. Together, it revealed that mutating cysteine 46 in IKK-β could increase caveolin-1 and mPGES-1 expression to facilitate osteoclast differentiation and osteolysis. Cysteine 46 can serve as a novel target in IKK-β for designing inhibitors to treat osteolysis.

Background Osteoporosis is a threat to aged people who have excessive osteoclast activation and bone resorption, subsequently causing fracture and even disability. Inhibiting osteoclast differentiation and absorptive functions has become an efficient approach to treat osteoporosis, but osteoclast-targeting inhibitors available clinically remain rare. Kirenol (Kir), a bioactive diterpenoid derived from an antirheumatic Chinese herbal medicine Herba Siegesbeckiae, can treat collagen-induced arthritis in vivo and promote osteoblast differentiation in vitro, while the effects of Kir on osteoclasts are still unclear. Purpose We explore the role of Kir on RANKL-induced osteoclastogenesis in vitro and bone loss in vivo. Methods The in vitro effects of Kir on osteoclast differentiation, bone resorption and the underlying mechanisms were evaluated with bone marrow-derived macrophages (BMMs). In vivo experiments were performed using an ovariectomy (OVX)-induced osteoporosis model. Results We found that Kir remarkably inhibited osteoclast generation and bone resorption in vitro. Mechanistically, Kir significantly inhibited F-actinring formation and repressed RANKL-induced NF-κB p65 activation and p-p38, p-ERK and c-Fos expression. Moreover, Kir inhibited both the expression and nuclear translocation of NFATc1. Ca2+ oscillation and caveolin-1 (Cav-1) were also reduced by Kir during osteoclastogenesis in vitro. Consistent with these findings, 2–10 mg/kg Kir attenuated OVX-induced osteoporosis in vivo as evidenced by decreased osteoclast numbers and downregulated Cav-1 and NFATc1 expression. Conclusions Kir suppresses osteoclastogenesis and the Cav-1/NFATc1 signaling pathway both in vitro and in vivo and protects against OVX-induced osteoporosis. Our findings reveal Kir as a potential safe oral treatment for osteoporosis.

Background and purpose Osteoclasts are unique cells to absorb bone. Targeting osteoclast differentiation is a therapeutic strategy for osteolytic diseases. Natural marine products have already become important sources of new drugs. The naturally occurring nitrobenzoyl sesquiterpenoids first identified from marine fungi in 1998 are bioactive compounds with a special structure, but their pharmacological functions are largely unknown. Here, we investigated six marine fungus-derived nitrobenzoyl sesquiterpenoids on osteoclastogenesis and elucidated the mechanisms. Experimental approach Compounds were first tested by RANKL-induced NF-κB luciferase activity and osteoclastic TRAP assay, followed by molecular docking to characterize the structure-activity relationship. The effects and mechanisms of the most potent nitrobenzoyl sesquiterpenoid on RANKL-induced osteoclastogenesis and bone resorption were further evaluated in vitro. Micro-CT and histology analysis were used to assess the prevention of bone destruction by nitrobenzoyl sesquiterpenoids in vivo. Key results Nitrobenzoyl sesquiterpenoid 4, with a nitrobenzoyl moiety at C-14 and a hydroxyl group at C-9, was the most active compound on NF-κB activity and osteoclastogenesis. Consequently, nitrobenzoyl sesquiterpenoid 4 exhibited suppression of RANKL-induced osteoclastogenesis and bone resorption from 0.5 μM. It blocked RANKL-induced IκBa phosphorylation, NF-κB p65 and RelB nuclear translocation, NFATc1 activation, reduced DC-STAMP but not c-Fos expression during osteoclastogenesis in vitro. Nitrobenzoyl sesquiterpenoid 4 also ameliorated LPS-induced osteolysis in vivo. Conclusion and implications These results highlighted nitrobenzoyl sesquiterpenoid 4 as a novel inhibitor of osteoclast differentiation. This marine-derived sesquiterpenoid is a promising lead compound for the treatment of osteolytic diseases.

Dysregulated host immune homeostasis in sepsis is life-threatening even after a successfully treated bacterial infection. Lipopolysaccharide (LPS) is an endotoxin that is a major contributor to the aberrant immune responses and endotoxic shock in gram-negative bacterial sepsis. However, the current knowledge of the role of B cells in endotoxic shock is limited. Here, we report that CD1d expression in B cells and the percentage of CD5+CD1dhi regulatory B (Breg) cells decreased in a mouse model of endotoxic shock. Interestingly, IL-10 but not FasL expression in CD5+CD1dhi Breg cells in response to endotoxin was dramatically reduced in severe septic shock mice, and the regulatory function of CD5+CD1dhi Breg cells in vitro to control the Th1 response was also diminished. Adoptive transfer of CD5+CD1dhi Breg cells from healthy WT mice but not IL-10 deficient mice downregulated the IFN-γ secretion in CD4+ T cells and conferred protection against severe endotoxic shock in vivo. Our findings demonstrate the change and notable therapeutic potential of IL-10-producing Breg cells in endotoxic shock.