Bone Degeneration and Its Recovery in SMP30/GNL-Knockout Mice.

Senescence marker protein-30 (SMP30) decreases androgen-independently with aging and is a lactone-hydrolyzing enzyme gluconolactonase (GNL) that is involved in vitamin C biosynthesis. In the present study, bone properties of SMP30/GNL knockout (KO) mice with deficiency in vitamin C synthesis were investigated to reveal the effects of SMP30/GNL and exogenous vitamin C supplementation on bone formation. Mineral content (BMC) and mineral density (BMD) of the mandible and femur of SMP30/GNL KO and wild-type mice at 2 and 3 months of age with or without vitamin C supplementation were measured by dual-energy X-ray absorptiometry. Body and bone weight of both age groups decreased and became significantly lower than those of wild-type mice. The bones of SMP30/GNL KO mice were rough and porous, with BMC and BMD significantly below wild-type. Oral supplementation with vitamin C eliminated differences in body weight, bone weight, BMC, and BMD between SMP30/GNL KO and wild-type mice at each age. These results indicate that bone degeneration in SMP30/GNL KO mice was caused by lack of vitamin C, and that this mouse strain is an appropriate model for bone metabolism in humans, which have no ability to synthesize vitamin C.

EAD and DAD mechanisms analyzed by developing a new human ventricular cell model.

It has long been suggested that the Ca(2+)-mechanisms are largely involved in generating the early afterdepolarization (EAD) as well as the delayed afterdepolarization (DAD). This view was examined in a quantitative manner by applying the lead potential analysis to a new human ventricular cell model. In this ventricular cell model, the tight coupled LCC-RyR model (CaRU) based on local control theory (Hinch et al. 2004) and ion channel models mostly based on human electrophysiological data were included to reproduce realistic Ca(2+) dynamics as well as the membrane excitation. Simultaneously, the Ca(2+) accumulation near the Ca(2+) releasing site was incorporated as observed in real cardiac myocytes. The maximum rate of ventricular repolarization (-1.02 mV/ms) is due to IK1 (-0.55 mV/ms) and the rest is provided nearly equally by INCX (-0.20 mV/ms), INaL (-0.16 mV/ms) and INaT (-0.13 mV/ms). These INaL and INaT components are due to closure of the voltage gate, which remains partially open during the plateau potential. DADs could be evoked by applying high-frequency stimulations supplemented by a partial Na(+)/K(+) pump inhibition, or by a microinjection of Ca(2+). EADs was evoked by retarding the inactivation of INaL. The lead potential (VL) analysis revealed that IK1 and IKr played the primary role to reverse the AP repolarization to depolarizing limb of EAD. ICaL and INCX amplified EAD, while the remaining currents partially antagonized dVL/dt. The maximum rate of rise of EAD was attributable to the rapid activation of both ICaL (45.5%) and INCX (54.5%).

Music exposure induced prolongation of cardiac allograft survival and generated regulatory CD4⁺ cells in mice.

In clinical practice, music has been used to decrease stress, heart rate, and blood pressure and to provide a distraction from disease symptoms. We investigated sound effects on alloimmune responses in murine heart transplantation. Naïve and eardrum-ruptured CBA/N (CBA, H2(K)) underwent transplantation of a C57BL/6 (B6, H2(b)) heart and were exposed to 1 of 3 types of music-opera (La Traviata), classical (Mozart), and New Age (Enya)-or 1 of 6 different single sound frequencies for 7 days. An adoptive transfer study was performed to determine whether regulatory cells were generated in allograft recipients. Cell-proliferation, cytokine, and flow cytometry assessments were also performed. CBA recipients of a B6 graft exposed to opera and classical music had significantly prolonged allograft survival (median survival times [MSTs], 26.5 and 20 days, respectively), whereas those exposed to 6 single sound frequencies and New Age did not (MSTs, 7, 8, 9, 8, 8, 8, and 11 days, respectively). Untreated and eardrum-ruptured CBA rejected B6 grafts acutely (MSTs, 7 and 8.5 days, respectively). Adoptive transfer of whole splenocytes, CD4(+) cells, and CD4(+)CD25(+) cells from opera-exposed primary recipients resulted in significantly prolonged allograft survival in naive secondary recipients (MSTs, 36, 68, and >50 days, respectively). Cell-proliferation, interleukin (IL)-2 and interferon-γ were suppressed in opera-exposed mice, whereas IL-4 and IL-10 from opera-exposed recipients were up-regulated. Flow cytometry studies showed an increased CD4(+)CD25(+)Foxp3(+) cell population in splenocytes from opera-exposed mice. In conclusion, exposure to some types of music may induce prolonged survival of fully allogeneic cardiac allografts and generate CD4(+)CD25(+)Foxp3(+) regulatory cells.

Specific antibodies to Porphyromonas gingivalis Lys-gingipain by DNA vaccination inhibit bacterial binding to hemoglobin and protect mice from infection.

Lys-gingipain (KGP), a lysine-specific cysteine proteinase, is one of the major virulence factors of Porphyromonas gingivalis. Here we examined the involvement of the catalytic domain of KGP (KGP(cd)) in hemoglobin binding by P. gingivalis, using a specific immunoglobulin G (IgG) elicited by the administration of plasmid DNA encoding KGP(cd) or the catalytic domain of Arg-gingipain (RGP(cd)). The pSeq2A/kgp(cd) and pSeq2B/rgp(cd) plasmids were constructed by the ligation of kgp(cd) and rgp(cd) DNA fragments, respectively. Female BALB/c mice were immunized with each of these plasmids. pSeq2A/kgp(cd) elicited a strong response to recombinant KGP(cd) (rKGP(cd)), as well as to comparably produced rRGP(cd)-reactive antibodies. The serum antibodies elicited by pSecTag2B/rgp(cd) also cross-reacted with rKGP(cd) as well as rRGP(cd). Anti-KGP(cd) IgG significantly inhibited hemoglobin binding by P. gingivalis. Furthermore, the inhibition of hemoglobin binding was markedly enhanced by a combination of anti-KGP(cd) and anti-fimbriae. Anti-RGP(cd) IgG showed a negligible inhibitory effect, while both anti-KGP(cd) and anti-RGP(cd) IgGs showed significant inhibitory effects on Lys- and Arg-specific proteolytic activities and on the growth of P. gingivalis under iron-restricted conditions where supplemented hemoglobin was the sole iron source. Immunized mice were challenged by intraperitoneal inoculation with P. gingivalis. All nonimmunized mice died within 72 h; however, vaccination with pSeq2A/kgp(cd) and pSeq2B/rgp(cd) prevented inflammatory responses and prolonged the survival rate of immunized mice by 43 and 27%, respectively. These results suggest that KGP(cd) acts as a hemoglobin-binding protein and can also be useful as an immunogen inducing a protective response to P. gingivalis infection.