Isolation and characterization of the ecdysone receptor and its heterodimeric partner ultraspiracle through development in Sciara coprophila

Regulation of DNA replication is critical, and loss of control can lead to DNA amplification. Naturally occurring, developmentally regulated DNA amplification occurs in the DNA puffs of the late larval salivary gland giant polytene chromosomes in the fungus fly, Sciara coprophila. The steroid hormone ecdysone induces DNA amplification in Sciara, and the amplification origin of DNA puff II/9A contains a putative binding site for the ecdysone receptor (EcR). We report here the isolation, cloning, and characterizing of two ecdysone receptor isoforms in Sciara (ScEcR-A and ScEcR-B) and the heterodimeric partner, ultraspiracle (ScUSP). ScEcR-A is the predominant isoform in larval tissues and ScEcR-B in adult tissues, contrary to the pattern in Drosophila. Moreover, ScEcR-A is produced at amplification but is absent just prior. We discuss these results in relation to the model of ecdysone regulation of DNA amplification.     

In vivo administration of 1,25-dihydroxyvitamin D3 suppresses the expression of RANKL mRNA in bone of thyroparathyroidectomized rats constantly infused with PTH

It is known that pharmacological or toxic doses of vitamin D induce bone resorption both in vivo and in vitro, whereas physiological doses of the vitamin have a protective effect on bone in vivo. To investigate the discrepancies of the dose-dependent effect of vitamin D on bone resorption, we examined the in vivo effect of 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] on the expression of the receptor activator of nuclear factor-kappaB (NF-kappaB) ligand (RANKL) and osteoprotegerin (OPG) mRNAs in bone of thyroparathyroidectomized (TPTX) rats infused with or without parathyroid hormone (PTH). Continuous infusion of 50 ng/h of PTH greatly increased the expression of RANKL mRNA in bone of TPTX rats. Expression of OPG mRNA was not altered by PTH infusion. When graded doses of 1,25(OH)(2)D(3) was daily administered orally for 14 days to normocalcemic TPTX rats constantly infused with PTH, 0.01 and 0.1 microg/kg of 1,25(OH)(2)D(3) inhibited the PTH-induced RANKL mRNA expression, but 0.5 microg/kg of the vitamin did not inhibit it. Regulator of G protein signaling-2 (RGS-2) gene expression was suppressed by 1,25(OH)(2)D(3) dose-dependently, but PTH/PTHrP receptor mRNA expression was not altered. Bone morphometric analyses revealed that 1,25(OH)(2)D(3) suppressed PTH-induced osteoclast number in vivo. These results suggest that pharmacological or toxic doses of 1,25(OH)(2)D(3) stimulate bone resorption by inducing RANKL, but a certain range of physiological doses of the vitamin inhibit PTH-induced bone resorption, the latter mechanism appeared to be mediated, at least in part, by the suppression of the PTH/PTHrP receptor-mediated signaling.     

Effect of ischemic preconditioning on cerebral blood flow after subsequent lethal ischemia in gerbils

Ischemic tolerance, the phenomenon where a sublethal ischemic preconditioning protects the brain against a subsequent lethal ischemia, has been widely studied. Studies have been done on cerebral blood flow levels prior to the lethal ischemia, but the hemodynamic pattern after global ischemia with ischemic preconditioning has not been reported. Sequential changes in regional cerebral blood flow (rCBF) in gerbil hippocampus after 5 min global ischemia with or without 2 min ischemic preconditioning were studied to determine if ischemic preconditioning affects rCBF. Four different treatments were given: (1) sham-operated, (2) 2 min ischemia, (3) non-preconditioned, and (4) preconditioned. Groups (1) and (2) (both groups n = 5) were given a 24-h recovery period and the rCBF was measured for baseline values. 24 h after sham-operation (3) and 2 min ischemia (4), gerbils were subjected to 5 min ischemia followed by 1 h, 6 h, 1-day or 7-day reperfusion periods (all groups n = 5). Although no regional difference was observed in the recovery pattern of rCBF, the values of rCBF were significantly higher in the preconditioned group throughout whole brain regions including hippocampus. These results indicate that ischemic preconditioning facilitated the recovery of rCBF after 5 min global ischemia. It needs further study to determine whether the protecting effects of preconditioning relate to the early recovery of rCBF or not. However, our results could be interpreted that the early recovery of rCBF may lead to benefits for cell survival in the CA1 neuron, probably facilitating other protecting mechanisms.     

Involvement of the endocannabinoid system in periodontal healing

Endocannabinoids including anandamide (AEA) and 2-arachidonoylglycerol (2-AG) are important lipid mediators for immunosuppressive effects and for appropriate homeostasis via their G-protein-coupled cannabinoid (CB) receptors in mammalian organs and tissues, and may be involved in wound healing in some organs. The physiological roles of endocannabinoids in periodontal healing remain unknown. We observed upregulation of the expression of CB1/CB2 receptors localized on fibroblasts and macrophage-like cells in granulation tissue during wound healing in a wound-healing model in rats, as well as an increase in AEA levels in gingival crevicular fluid after periodontal surgery in human patients with periodontitis. In-vitro, the proliferation of human gingival fibroblasts (HGFs) by AEA was significantly attenuated by AM251 and AM630, which are selective antagonists of CB1 and CB2, respectively. CP55940 (CB1/CB2 agonist) induced phosphorylation of the extracellular-regulated kinases (ERK) 1/2, p38 mitogen-activated protein kinase (p38MAPK), and Akt in HGFs. Wound closure by CP55940 in an in-vitro scratch assay was significantly suppressed by inhibitors of MAP kinase kinase (MEK), p38MAPK, and phosphoinositol 3-kinase (PI3-K). These findings suggest that endocannabinoid system may have an important role in periodontal healing.     

A CD36-related Transmembrane Protein Is Coordinated with an Intracellular Lipid-binding Protein in Selective Carotenoid Transport for Cocoon Coloration

The transport pathway of specific dietary carotenoids from the midgut lumen to the silk gland in the silkworm, Bombyx mori, is a model system for selective carotenoid transport because several genetic mutants with defects in parts of this pathway have been identified that manifest altered cocoon pigmentation. In the wild-type silkworm, which has both genes, Yellow blood (Y) and Yellow cocoon (C), lutein is transferred selectively from the hemolymph lipoprotein to the silk gland cells where it is accumulated into the cocoon. The Y gene encodes an intracellular carotenoid-binding protein (CBP) containing a lipid-binding domain known as the steroidogenic acute regulatory protein-related lipid transfer domain. Positional cloning and transgenic rescue experiments revealed that the C gene encodes Cameo2, a transmembrane protein gene belonging to the CD36 family genes, some of which, such as the mammalian SR-BI and the fruit fly ninaD, are reported as lipoprotein receptors or implicated in carotenoid transport for visual system. In C mutant larvae, Cameo2 expression was strongly repressed in the silk gland in a specific manner, resulting in colorless silk glands and white cocoons. The developmental profile of Cameo2 expression, CBP expression, and lutein pigmentation in the silk gland of the yellow cocoon strain were correlated. We hypothesize that selective delivery of lutein to specific tissue requires the combination of two components: 1) CBP as a carotenoid transporter in cytosol and 2) Cameo2 as a transmembrane receptor on the surface of the cells.     

Convergent signaling in the regulation of connective tissue growth factor in malignant mesothelioma: TGFβ signaling and defects in the Hippo signaling cascade

Malignant mesothelioma (MM) is a neoplasm that arises from serosal surfaces of the pleural, peritoneal and pericardial cavities with worldwide incidence, much of which is caused by asbestos exposure. Patients suffer from pain and dyspnea due to direct invasion of the chest wall, lungs and vertebral or intercostal nerves by masses of thick fibrotic tumors. Although there has been recent progress in the clinical treatment, current therapeutic approaches do not provide satisfactory results. Therefore, development of a molecularly targeted therapy for MM is urgently required. Our recent studies suggest that normal mesothelial and MM cell growth is promoted by TGFβ, and that TGFβ signaling together with intrinsic disturbances in neurofibromatosis type 2 (NF2) and Hippo signaling cascades in MM cells converges upon further expression of connective tissue growth factor (CTGF). The formation of a YAP-TEAD4–Smad3-p300 complex on the specific CTGF promoter site with an adjacent TEAD and Smad binding motif is a critical and synergistic event caused by the dysregulation of these two distinct cascades. Furthermore, we demonstrated the functional importance of CTGF through the mouse studies and human histological analyses, which may elucidate the clinical features of MM with severe fibrosis in the thoracic cavity.     

Brünnich's guillemots (Uria lomvia) maintain high temperature in the body core during dives

A major challenge for diving birds, reptiles, and mammals is regulating body temperature while conserving oxygen through a reduction in metabolic processes. To gain insight into how these needs are met, we measured dive depth and body temperatures at the core or periphery between the skin and abdominal muscles simultaneously in freely diving Brünnich's guillemots (Uria lomvia), an arctic seabird, using an implantable data logger (16-mm diameter, 50-mm length, 14-g mass, Little Leonardo Ltd., Tokyo). Guillemots exhibited increased body core temperatures, but decreased peripheral temperatures, during diving. Heat conservation within the body core appeared to result from the combined effect of peripheral vasoconstriction and a high wing beat frequency that generates heat. Conversely, the observed tissue hypothermia in the periphery should reduce metabolic processes as well as heat loss to the water. These physiological effects are likely one of the key physiological adaptations that makes guillemots to perform as an efficient predator in arctic waters.