Previous studies have indicated that the amount of RANKL expressed on the cell surface of osteoblasts or bone marrow stromal cells (BMSCs) is considered an important factor determining the extent of osteoclast activation. However, subcellular trafficking of RANKL and its regulatory mechanisms in osteoblastic cells is still unclear. In this study, we showed that RANKL is predominantly localized in lysosomal organelles, but little is found on the cell surface of osteoblastic cells. We also showed that RANKL is relocated to the plasma membrane in response to stimulation with RANK-Fc-coated beads, indicating that the lysosomal organelles where RANKL is localized function as secretory lysosomes. In addition, using a protein pull-down method, we identified vacuolar protein sorting (Vps)33a as interacting with the cytoplasmic tail of RANKL. Furthermore, knockdown of Vps33a expression reduced the lysosomal storage of RANKL and caused the accumulation of newly synthesized RANKL in the Golgi apparatus, indicating that Vps33a is involved in transporting RANKL from the Golgi apparatus to secretory lysosomes. We also showed that suppression of Vps33a affects the cell surface expression level of RANKL and disrupts the regulated behavior of RANKL. These results suggest that RANKL storage in secretory lysosomes is important to control osteoclast activation and to maintain bone homeostasis.

The effects of magnesium ions on a 32-mer ribozyme (R32) were examined by high resolution NMR spectroscopy. In solution, R32 (without its substrate) consisted of a GAAA loop, stem II, a non-Watson-Crick 3-base pair duplex and a 4-base pair duplex that included a wobble G:U base pair. When an uncleavable substrate RNA (RdC11) was added to R32 without Mg2+ ions, a complex did not form between R32 and RdC11 because the substrate recognition regions of R32 formed intramolecular base pairs (the recognition arms were closed). By contrast, in the presence of Mg2+ ions, the R32-RdC11 complex was formed. Moreover, titration of mixtures of R32 and RdC11 with Mg2+ ions also induced the ribozyme-substrate interaction. Elevated concentrations (1.0 M) of monovalent Na+ ions could not induce the formation of the R32-RdC11 complex. These data suggest that Mg2+ ions are not only important as the true catalysts in the function of ribozyme-type metalloenzymes, but they also induce the structural change in the R32 hammerhead ribozyme that is necessary for establishment of the active form of the ribozyme-substrate complex.

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The authors propose a magnetic actuator for use as a navigation system for capsule endoscopes. The actuator is composed of a capsule dummy, a permanent magnet inside the capsule, and an external spiral structure. The device rotates and propels wirelessly when exposed to an external rotational magnetic field. In this study we measured the effect of the spiral shape on the velocity and thrust force properties. According to our experimental results, the actuator obtained a maximum velocity and thrust force when the spiral angle was set at 45 degrees, the number of spirals was set at 4, and the spiral-height was set at 1-mmf. We also conducted a motion test in the large intestine of a pig placed on a 30 degrees slope. The actuator passed through a 700 mm length of the intestine in about 300 s. The device also managed to travel up and down the 30 degrees slope with no difficulty whatsoever. Our results demonstrate the great potential of this actuator for use as a navigation system for capsule endoscopes.

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