Knockdown of mechanosensitive adaptor Hic-5 ameliorates post-traumatic osteoarthritis in rats through repression of MMP-13.

Osteoarthritis (OA) is the most common joint disease associated with articular cartilage destruction. Matrix metalloproteinase-13 (MMP-13) has an essential role in OA pathogenesis by degradation of collagen II, a major component of articular cartilage. Hydrogen peroxide-inducible clone-5 (Hic-5; TGFB1I1), a transforming growth factor-ß-inducible mechanosensor, has previously been reported to promote OA pathogenesis by upregulating MMP-13 expression in mouse osteoarthritic lesions. In our current study, immunohistochemical analysis showed that Hic-5 protein expression was increased in human OA cartilage compared with normal cartilage. Functional experiments demonstrated that Hic-5 and MMP-13 expression was increased by mechanical stress, and mechanical stress-induced MMP-13 expression was suppressed by Hic-5 siRNA in human chondrocytes. Moreover, intracellular localization of Hic-5 shifted to the nucleus from focal adhesions in human chondrocytes subjected to mechanical stress, and nuclear Hic-5 increased MMP-13 gene expression. In vivo, intra-articular injection of Hic-5 siRNA decreased the Osteoarthritis Research Society International score and MMP-13 protein expression in articular cartilage of OA rats. Our findings suggest that Hic-5 regulates transcription of MMP-13 in human chondrocytes, and Hic-5 may be a novel therapeutic target for OA because OA progression was suppressed by intra-articular injection of Hic-5 siRNA in rats.

Characterization of a WD-repeat family protein WDR3 in the brain of WDR3 hetero knockout mice.

The nuclear protein WDR3 is a member of the WD-repeat family and is a component of the 18S pre-rRNA processing complex. However, the expression and function of WDR3 in the brain remains unknown. To characterize WDR3 in the adult mouse brain, we developed Wdr3 heterozygous knockout (WDR3-HKO) mice. Notably, no homozygous Wdr3 knockout mice were born, suggesting that complete absence of WDR3 causes lethal abnormalities during embryogenesis. Brain Wdr3 mRNA expression was significantly reduced to 60% in the WDR3-HKO mice compared to wild type (WT) mice, while the expression of 18S rRNA did not decline. Using immunohistochemistry and X-gal staining, we demonstrated that WDR3 is widely expressed in the mouse brain, especially in the hippocampus, habenular nucleus, and cerebellum. We observed no differences in body weight during adulthood or developmental weight gain between the WDR3-HKO and WT mice. Interestingly, WDR3-HKO mice exhibited a slight but significant increase in spontaneous locomotor activity compared to WT littermates. In conclusion, the WDR3-HKO mice showed no significant phenotypic changes. Further studies are required to explore the behavioral characteristics of WDR3-HKO mice.

Biochemical characterization of mitochondria from adult worms and plerocercoid larvae of Spirometra mansoni shows mixed populations of anaerobic and aerobic mitochondria.

The mitochondria of adult and plerocercoid Spirometra mansoni were characterized in isolated mitochondria and in situ by electron microscopic histochemistry with special attention to the respiratory chain. Although the specific activities of the constituent enzyme complexes of succinate oxidase are fairly similar in adult and plerocercoid mitochondria, those of succinate oxidase and NADH-FRD are approximately 4- and 25-fold higher in adult mitochondria than in plerocercoid mitochondria, respectively. Quinone analysis by high performance liquid chromatography and mass spectrometry showed that adult and plerocercoid mitochondria contained both rhodoquinone-10 and ubiquinone-10 at concentrations of 4.98 and 0.106 nmol mg-1 for adult, and 0.677 and 0.137 nmol mg-1 for plerocercoid, respectively. Inhibition studies on the succinate-oxidase system of adult mitochondria showed that they possessed both cyanide-sensitive and -insensitive succinate oxidases, the latter of which produces hydrogen peroxide. Adult mitochondria, when NADH was used as a substrate, were shown to produce hydrogen peroxide, and the production of hydrogen peroxide decreased to undetectable levels in the presence of fumarate. The specific activities of NADH-fumarate reductase and cytochrome c oxidase were significantly higher in mature proglottids than in immature and gravid proglottids. Isopycnic density-gradient centrifugation analyses and in situ electron microscopic histochemistry revealed that both adult and plerocercoid mitochondria were heterogeneous in terms of respiratory function and physicochemical properties. The physiological significance of adult and plerocercoid mitochondria is discussed in relation to the oxygen tension of their parasitic habitats.

Fluorogenic Probes for Accurate in Situ Imaging of Viral and Mammalian Sialidases.

Sialidases are widely distributed in nature and are involved in many physiological and pathological processes. Sialidases are expressed and work in various tissues and organelles. Clarification of the localization of sialidases is very helpful as a way to understand their functions. We previously developed a novel fluorogenic probe for sialidases, BTP3-Neu5Ac, that visualized the localization of sialidase activity in live cells and tissues by precipitating the hydrophobic fluorescent compound; however, for the purpose of accurate fluorescence imaging of sialidase-expressing cells or the distribution of intracellular sialidase activity, BTP3-Neu5Ac was inadequate in imaging performance. We report the design and development of a sialidase imaging probe that improves the sensitivity and accuracy of in situ fluorescence imaging performance as well as increases the hydrophobicity by attaching linear unsaturated hydrocarbon chains into the hydrophobic fluorescent compound of BTP3-Neu5Ac. The newly developed probe showed low diffusivity and high brightness for fluorescence imaging, and it enabled sensitive and highly accurate imaging of viral sialidase in virus-infected cells and sialidase-expressing cells as well as mammalian sialidase in the rat brain. The probe also enabled the fluorescence imaging of intracellular viral sialidase in live-virus-infected cells. The newly developed probe is expected to be a useful tool that will contribute to the progress of research on sialidases in various fields such as research on viruses and brains.

A visible-light-induced overall water-splitting photocatalyst: conduction-band-controlled silver tantalate.

Visible-light-induced water splitting was achieved by increasing the visible-light sensitivity of AgTaO3, by the introduction of Nb, generating AgTa1-xNbxO3. After grafting NiO onto AgTa0.7Nb0.3O3, simultaneous, catalytic liberation of H2 and O2 at a molar ratio of ∼2 : 1 was achieved from pure water under only visible-light irradiation.