RANKL-RANK signaling in osteoclastogenesis and bone disease

Hundreds of millions of people worldwide are affected by bone-related diseases, such as osteoporosis and rheumatoid arthritis. Understanding the molecular mechanisms of bone metabolism is crucial for developing novel drugs for treating such diseases. In particular, genetic experiments showing that the receptor activator of NF-kappaB (RANK), its ligand RANKL, and the decoy receptor OPG are essential, central regulators of osteoclast development and osteoclast function were significant turning points in our understanding of bone diseases. RANKL-RANK signaling activates a variety of downstream signaling pathways required for osteoclast development. Moreover, molecular cross-talk between RANKL-RANK and other ligand-receptor systems fine-tunes bone homeostasis in normal physiology and disease. Designing novel drugs that target RANKL-RANK and their signaling pathways in osteoclasts could potentially revolutionize the treatment of many diseases associated with bone loss such as arthritis, tooth loss, cancer metastases or osteoporosis.     

Rac1 negatively regulates lipopolysaccharide-induced IL-23 p19 expression in human macrophages and dendritic cells and NF-kappaB p65 trans activation plays a novel role

IL-23 is a heterodimeric cytokine composed of a unique p19 subunit and of a p40 subunit that is also common to IL-12. We defined the distinct signaling mechanisms that regulate the LPS-mediated induction of IL-23 p19 and p40 in human macrophages and dendritic cells. We found that the overexpression of dominant-negative Rac1 (N17Rac1) enhanced LPS-induced IL-23 p19 expression but did not alter p40 expression or IL-12 p70 production in PMA-treated THP-1 macrophages and in human monocyte-derived dendritic cells. Although the inhibition of either p38 MAPK or JNK enhanced LPS-induced p19 expression, N17Rac1 did not influence either p38 MAPK or JNK activation. By contrast, N17Rac1 augmented both NF-kappaB gene expression and p65 trans activation stimulated by LPS without affecting the degradation of IkappaB-alpha or DNA binding to NF-kappaB. Furthermore, small interference RNA of NF-kappaB p65 attenuated cellular amounts of p65 and suppressed LPS-induced p19 expression but did not affect p40 expression. Our findings indicate that Rac1 negatively controls LPS-induced IL-23 p19 expression through an NF-kappaB p65 trans activation-dependent, IkappaB-independent pathway and that NF-kappaB p65 regulates LPS-induced IL-23 p19, but not p40, expression, which causes differences in the control of IL-23 p19 and p40 expression by Rac1.     

B cell antigen receptor and CD40 differentially regulate CD22 tyrosine phosphorylation

Cell surface molecules on lymphocytes positively or negatively modulate the Ag receptor signaling, and thus regulate the fate of the cell. CD22 is a B cell-specific cell surface protein that contains multiple ITIMs in the cytoplasmic tail, and critically regulates B cell activation and survival. CD22 regulation on B cell signaling is complex because CD22 can have both positive and negative roles in various contexts. We generated phosphospecific polyclonal Abs reacting four major CD22 tyrosine motifs (Y762, Y807, Y822, and Y842) and analyzed the pattern and intensity of phosphorylation of these tyrosine residues. The tyrosine motifs, Y762, Y822, and Y842, are considered as ITIM, whereas the other, Y807, is suggested to be important for Grb2 recruitment. Approximately 10% of the four tyrosine residues were constitutively phosphorylated. Upon anti-IgM ligation, CD22 Y762 underwent most rapid phosphorylation, whereas all four tyrosine residues were eventually phosphorylated equally at approximately 35% of all CD22 molecules in the cell. By contrast, anti-CD40 stimulation specifically up-regulated anti-IgM-induced phosphorylation of tyrosines within two ITIM motifs, Y762 and Y842, which was consistent with in vivo finding of the negative role of CD22 in CD40 signaling. Thus, CD22 phosphorylation is not only quantitatively but also qualitatively regulated by different stimulations, which may determine the outcome of B cell signaling.     

Unusual extra space at the active site and high activity for acetylated hydroxyproline of prolyl aminopeptidase from Serratia marcescens

The prolyl aminopeptidase complexes of Ala-TBODA [2-alanyl-5-tert-butyl-(1, 3, 4)-oxadiazole] and Sar-TBODA [2-sarcosyl-5-tert-butyl-(1, 3, 4)-oxadiazole] were analyzed by X-ray crystallography at 2.4 angstroms resolution. Frames of alanine and sarcosine residues were well superimposed on each other in the pyrrolidine ring of proline residue, suggesting that Ala and Sar are recognized as parts of this ring of proline residue by the presence of a hydrophobic proline pocket at the active site. Interestingly, there was an unusual extra space at the bottom of the hydrophobic pocket where proline residue is fixed in the prolyl aminopeptidase. Moreover, 4-acetyloxyproline-betaNA (4-acetyloxyproline beta-naphthylamide) was a better substrate than Pro-betaNA. Computer docking simulation well supports the idea that the 4-acetyloxyl group of the substrate fitted into that space. Alanine scanning mutagenesis of Phe139, Tyr149, Tyr150, Phe236, and Cys271, consisting of the hydrophobic pocket, revealed that all of these five residues are involved significantly in the formation of the hydrophobic proline pocket for the substrate. Tyr149 and Cys271 may be important for the extra space and may orient the acetyl derivative of hydroxyproline to a preferable position for hydrolysis. These findings imply that the efficient degradation of collagen fragment may be achieved through an acetylation process by the bacteria.     

Angelmarin, a novel anti-cancer agent able to eliminate the tolerance of cancer cells to nutrient starvation

The CH(2)Cl(2)-soluble extract of Angelica pubescens was found to kill PANC-1 cancer cells preferentially under nutrition starvation at a concentration of 50 microg/ml, with virtually no cytotoxicity under nutrient-rich conditions. Further bioassay-guided fractionation and isolation led to the isolation of a novel compound named angelmarin as the primary compound responsible for the preferential cytotoxicity; the compound exhibited 100% preferential cytotoxicity against PANC-1 cells at a concentration of 0.01 microg/ml.     

Smell intensity monitoring using metal oxide semiconductor odor sensors during intravenous olfaction test

The intravenous olfaction test using prosultiamine (PST) solution is simple to perform and has been used clinically in Japan. We monitored intranasal intensity of smell continuously in real time under various conditions of administration using metal oxide semiconductor odor sensors and established an optimal PST injection procedure. In this study, we found that 1) although there was fluctuation in the pattern of intensity of increase in smell in the PST original solution test, the pattern of increase in intranasal smell intensity could be stabilized by prolonging the injection time to 40 s and 2) dilution of PST with physiological saline was effective in preventing angialgia during intravenous injection. It appears that PST administration is best performed by adding 10 ml of saline to 10 mg (2 ml) of PST and injecting the resulting 12-ml solution (6x dilution) and that the best respiratory cycle for testing is once in every 2 s.     

Reproductive swarming of sympatric nereidid polychaetes in an estuary of the Omuta-gawa river in Kyushu, Japan, with special reference to simultaneous swarming of two Hediste species

Habitat differences and spatial and temporal separation in reproductive swarming among sympatric nereidid polychaetes were examined in an estuary of the Omuta-gawa River, Kyushu, Japan by annual periodical sampling from December 2003 to January 2005. Benthic adults of Tylorrhynchus osawai and Hediste diadroma occupied mainly the upper reaches of the estuary, whereas those of H. japonica usually inhabited the middle reaches, though their distributions overlapped. Reproductive swarming of mature adults occurred in the estuary just after high tide at night during spring tides in four nereidids: H. japonica (in the middle and lower reaches from late December to late February), H. diadroma (throughout the whole estuary from middle December to late April), T. osawai (in the middle reaches and another estuary from late October to late December), and Nectoneanthes oxypoda sensu Imajima, 1972 (in the lower reaches in late April and early May). This result shows that temporal separation of reproductive swarming may act as a reproductive isolation mechanism among these nereidids, except for H. japonica and H. diadroma. Simultaneous swarming and mass-spawning of the two Hediste species were commonly observed in the middle and lower reaches from late December to early February, suggesting the absence of a pre-spawning barrier to reproductive isolation between them. We found no difference in spawning behavior between H. japonica and H. diadroma. Males of both species seemed to participate in swarming earlier than females.     

Resolvin E1 dampens airway inflammation and hyperresponsiveness in a murine model of asthma

Resolvin E1 (RvE1; 5S, 12R, 18R-trihydroxyeicosapentaenoic acid) is an anti-inflammatory lipid mediator derived from the omega-3 fatty acid eicosapentaenoic acid (EPA). It has been recently shown that RvE1 is involved in the resolution of inflammation. However, it is not known whether RvE1 is involved in the resolution of asthmatic inflammation. To investigate the anti-inflammatory effect of RvE1 in asthma, a murine model of asthma was studied. After RvE1 was administered to mice intraperitoneally, there were decreases in: airway eosinophil and lymphocyte recruitment, specific Th2 cytokine, IL-13, ovalbumin-specific IgE, and airway hyperresponsiveness (AHR) to inhaled methacholine. Moreover, RvE1-treated mice had significantly lower mucus scores compared to vehicle-treated mice based on the number of goblet cells stained with periodic acid-schiff (PAS). These findings provide evidence that RvE1 is a pivotal counterregulatory signal in allergic inflammation and offer novel multi-pronged therapeutic approaches for human asthma.