αVβ3 Integrin ligands enhance volume-sensitive calcium influx in mechanically stretched osteocytes

We propose that specific osteocyte–matrix interactions regulate the volume-sensitive calcium influx pathway, which we have shown is mediated by stretch-activated cation channels (SA-Cat) and is essential for the stretch-activated anabolic response in bone. The current study measured the hypotonic swelling-induced increase in cytosolic calcium concentration, [Ca²⁺]_i, in rat osteocytes, and found that cells adherent to different matrices behave differently. Osteopontin and vitronectin, matrix molecules that bind the α_Vβ₃ integrin, induced larger responses to the hypotonic swelling than other matrix molecules that bind other integrins. Addition of echistatin, which is a soluble α_Vβ₃ ligand, significantly enhanced the hypotonic [Ca²⁺]_i increase in addition to inducing an immediate increase in [Ca²⁺]_i by itself. These results strongly support the contention that α_Vβ₃ integrin signaling in osteocytes interacts with that in mechanotransduction, which is downstream of SA-Cat.     

Histomorphometric Evaluation of Cortical Bone Thickness Surrounding Miniscrew for Orthodontic Anchorage

Background: Recently, the use of miniscrews as an anchorage device has become a routine approach in the orthodontic field. However, there is no report that has analyzed the healing process of the miniscrew, such as the thickness of the cortical bone, in the past. Purpose: In the present study, to histologically assess the healing process of the osseous tissue surrounding miniscrews used as an orthodontic anchorage, the change in the thickness of the cortical bone was analyzed after 3, 6, and 12 weeks after the placement. Furthermore, the change in the bone‐implant contact in different regions of the miniscrew during the initial healing period was also investigated. Materials and Methods: Ninety‐six miniscrews were placed in eight beagle dogs. After 3, 6, and 12 weeks of healing, a force of 200–300 g was applied to the force‐applied groups for 12 weeks. Non‐forced groups remained in the jaw without force application. Results: In the non‐forced groups, a significant amount of cortical bone was formed at the head of the miniscrew at the initial stage of the healing process in the maxilla. However, less cortical bone formation was observed in the mandible. After the force application, increased bone formation was observed within 1 mm of the miniscrew compared to other regions in both jaws. In the mandible, significantly less cortical bone was observed 3 and 6 weeks after the force application. Bone‐implant contact revealed that the osseous tissue surrounding the miniscrew matured from the apex toward the head of the miniscrew. Conclusion: We suggest that this sufficient amount of cortical bone at the initial stage of healing enables the immediate loading in miniscrews to resist against orthodontic force. Furthermore, less amount of cortical bone formed at the head of the miniscrew may be one reason for the higher failure rate in the mandible.     

Thrombophilic dysfibrinogen Tokyo V with the amino acid substitution of gammaAla327Thr: formation of fragile but fibrinolysis-resistant fibrin clots and its relevance to arterial thromboembolism

Thrombophilic dysfibrinogen Tokyo V was identified in a 43-year-old man with recurrent thromboembolism. Based on analyses of the patient fibrinogen genes, the amino acid sequence of the aberrant fibrinogen peptide, and deglycosylation experiments, fibrinogen Tokyo V was shown to have an amino acid substitution of gamma Ala327Thr and possibly extra glycosylation at gamma Asn325 because the mutation confers the N-linked glycosylation consensus sequence Asn-X-Thr. The mutation resulted in impaired function and hypofibrinogenemia (hypodysfibrinogen). Polymerization of fibrin monomers derived from patient fibrinogen was severely impaired with a partial correction in the presence of calcium, resulting in very low clottability. Additionally, a large amount of soluble cross-linked fibrin was formed upon thrombin treatment in the presence of factor XIII and calcium. However, Tokyo V-derived fibrin was resistant to degradation by tissue plasminogen activator (tPA)-catalyzed plasmin digestion. The structure of Tokyo V fibrin appeared severely perturbed, since there are large pores inside the tangled fibrin networks and fiber ends at the boundaries. Taken together, these data suggest that Tokyo V fibrin clots are fragile, so that fibrinolysis-resistant insoluble fibrin and soluble fibrin polymers may be released to the circulation, partly accounting for the recurrent embolic episodes in the patient.     

17β estradiol recruits GluN2B‐containing NMDARs and ERK during induction of long‐term potentiation at temporoammonic‐CA1 synapses

When circulating 17β estradiol (E2) is elevated to proestrous levels, hippocampus‐dependent learning and memory is enhanced in female rodents, nonhuman primates, and women due to heightened synaptic function at hippocampal synapses. We previously reported that proestrous‐like levels of E2 administered to young adult ovariectomized (OVX) female rats increases the magnitude of LTP at CA3 Schaffer collateral (SC)‐CA1 synapses only when dendritic spine density, the NMDAR/AMPAR ratio, and current mediated by GluN2B‐containing NMDA receptors (NMDARs) are simultaneously increased. We also reported that this increase in GluN2B‐mediated NMDAR current in area CA1 is causally related to the E2‐induced increase in novel object recognition, tying together heightened synaptic function with improved learning and memory. In addition to SC inputs, innervation from the entorhinal cortex in the temporoammonic (TA) pathway onto CA1 distal dendrites in stratum lacunosum‐moleculare is critical for spatial memory formation and retrieval. It is not known whether E2 modulates TA‐CA1 synapses similarly to SC‐CA1 synapses. Here, we report that 24 hours post‐E2 injection, dendritic spine density on CA1 pyramidal cell distal dendrites and current mediated by GluN2B‐containing NMDARs at TA‐CA1 synapses is increased, similarly to our previous findings at SC‐CA1 synapses. However, in contrast to SC‐CA1 synapses, AMPAR transmission at TA‐CA1 synapses is significantly increased, and there is no effect on the LTP magnitude. Pharmacological blockade of GluN2B‐containing NMDARs or ERK activation, which occurs downstream of synaptic but not extrasynaptic GluN2B‐containing NMDARs, attenuates the LTP magnitude only in slices from E2‐treated rats. These data show that E2 recruits a causal role for GluN2B‐containing NMDARs and ERK signaling in the induction of LTP, cellular mechanisms not required for LTP induction at TA‐CA1 synapses in vehicle‐treated OVX female rats. © 2015 Wiley Periodicals, Inc.     

Activation of protein kinase C from guinea-pig and rat ventricular muscles by musclide-A1 and its (2R, 5S)-diastereomer

The activating effects of musclide-A1, 6-methyl-2,5-heptanediol 5-(hydrogen sulphate) (la), and its (2R, 5S)- and (2R, 5R)-diastereomers on ventricular protein kinase C from guinea-pig and rat were compared with other natural compounds derived from musk and non-natural related compounds. The (2R, 5S)-diastereomer was most potent among musk-derived compounds and its related non-natural compounds. The activation by la (20 μM) was produced in the presence of low [Ca²⁺]₀ and phosphatidylserine like 1,2-dioctanoylglycerol (2 μM). The activation pattern for peaks eluted by a hydroxyapatite column demonstrated that 1a activates ventricular-specific protein kinase C peak II, suggesting one mechanism for the beta-adrenergic cardiotonic potentiation by 1a.     

Nucleic acids induce the formation of a carcinogen, 2-amino-1-methyl-6-phenylimidazo[4, 5-b]pyridine (PhIP) in a model system

Effects of nucleic acids on the formation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine(PhIP) were studied in a model system. When a mixture of a certainamount of DNA or RNA, creatinine (1 mmol) and phenylalanine(1 mmol) in 10 ml of 50 mM phosphate buffer, pH 7.4, was heatedat 60°C for 4 weeks in a screw-capped vial, PhIP was produced,and the yield of PhIP was dependent on the heating time as wellas the dose of nucleic acid added to the mixture. However, PhIPwas not detectable in the mixtures without the presence of nucleicacids. Both deoxyribonucleotides and ribonucleotides testedinduced the formation of PhIP under the presence of phenylalanineand creatinine, although bases of nucleic acids such as adenineand guanine did not induce PhIP formation. Moreover, we confirmedthat 2-deoxy-D-ribose as well as D-ribose induced the formationof PhIP in the presence of creatinine and phenylalanine. Theseresults indicate that nucleic acids can induce the formationof PhIP in the presence of creatinine and phenylalanine in themodel system. Our data also suggest that pentose in nucleicacids may participate in PhIP forming reactions.