Cluster-Based Haldane State in an Edge-Shared Tetrahedral Spin-Cluster Chain: Fedotovite K_{2}Cu_{3}O(SO_{4})_{3}.

Fedotovite K_{2}Cu_{3}O(SO_{4})_{3} is a candidate of new quantum spin systems, in which the edge-shared tetrahedral (EST) spin clusters consisting of Cu^{2+} are connected by weak intercluster couplings forming a one-dimensional array. Comprehensive experimental studies by magnetic susceptibility, magnetization, heat capacity, and inelastic neutron scattering measurements reveal the presence of an effective S=1 Haldane state below T≅4 K. Rigorous theoretical studies provide an insight into the magnetic state of K_{2}Cu_{3}O(SO_{4})_{3}: an EST cluster makes a triplet in the ground state and a one-dimensional chain of the EST induces a cluster-based Haldane state. We predict that the cluster-based Haldane state emerges whenever the number of tetrahedra in the EST is even.

[Electroablation for the treatment of spontaneous pneumothorax using SOFT COAG electrosurgical output system].

We evaluated the validity of the SOFT COAG electrosurgical output system for the treatment of spontaneous pneumothorax. From April 2008 to May 2010, we compared 64 patients who had undergone bullae resection using endoscopic linear staplers, to 20 patients subjected to electroablation of the bullae using the SOFT COAG output system. There was no significant difference between the 2 groups in terms of operation time, bleeding, and mean duration of postoperative chest tube drainage. Postoperative recurrence was apparent in 3 cases for the linear stapler, and in 2 cases for SOFT COAG. Electroablation using the SOFT COAG output system was suggested to be valid for treatment of spontaneous pneumothorax.

Possible roles of arachidonic acid and its metabolites in induction of tissue plasminogen activator (t-PA) production in human fibroblast, IMR-90 cells by proteose peptone.

Proteose peptone (p.peptone) had an ability to induce tissue plasminogen activator (t-PA) production by human embryonic lung fibroblast, IMR-90 cells. We previously demonstrated that the induction was closely related to the activation of phospholipase A2 in the cells stimulated by p.peptone. In this report, we describe the involvement of arachidonate metabolism in the induction. The induction was inhibited in a dose-dependent manner by 5,8,11,14-eicosatetraenoic acid (ETYA), an inhibitor of both cycloxygenase and lypoxygenase, and also by nordihydroguaiaretic acid (NDGA), which in low concentrations selectively inhibits lipoxygenase. However, indomethacin, a specific inhibitor of cycloxygenase, had no effect on the induction. 5-hydroxyeicosatetraenoic acid (5-HETE), which is an arachidonate metabolite derived from lipoxygenase pathway, had an inductive effect, but prostaglandin E1 (PGE1), which is a metabolite from cycloxygenase pathway, had no effect on t-PA production by the cells. These results suggest that arachidonate metabolism is involved in the induction of t-PA production in IMR-90 cells by p.peptone, and that arachidonate metabolite(s) from lipoxygenase pathway is responsible for the induction.

Role of Ca2+ in t-PA production by human embryonic lung diploid fibroblast, IMR-90 cells, stimulated by proteose peptone.

Proteose peptone (p.peptone) remarkably induced tissue plasminogen activator (t-PA) activity in the conditioned medium of confluently cultured human embryonic lung diploid fibroblast, IMR-90 cells, in a dose-dependent manner. t-PA activity correlated well with the amount of t-PA antigen found in the conditioned medium of IMR-90 cells stimulated by p.peptone. t-PA production by IMR-90 cells stimulated by p.peptone was dependent on extracellular Ca2+ concentration and maximum t-PA production required approximately 3.6 mM extracellular Ca2+. Conversely, elimination of Ca2+ from the culture medium by EGTA, Ca2+ chelate agent, strongly inhibited t-PA production induced by p.peptone. t-PA production induced by p.peptone was inhibited in a dose-dependent manner by Verapamil, which inhibits Ca2+ uptake through the slow channels and also by W-7, an inhibitor of calmodulin. These results suggested that influx of extracellular Ca2+ into IMR-90 cells was caused by p.peptone and induced t-PA production by the cells.

Participation of phospholipase A2 in induction of tissue plasminogen activator (t-PA) production by human fibroblast, IMR-90 cells, stimulated by proteose peptone.

Proteose peptone (p.peptone) had an ability to induce tissue plasminogen activator(t-PA) production by human embryonic lung fibroblast, IMR-90 cells. The induction was dependent on extracellular Ca2+ concentration. The stimulation of p.peptone caused uptake of 45Ca2+ by the cells. The presences of both p.peptone and Ca2+ in medium were necessary for the continuous induction of t-PA production. Hydrocortisone and dexamethasone inhibited t-PA production induced by p.peptone. In addition, the inhibitors of phospholipase A2, quinacrine and 4-bromophenacylbromide, respectively inhibited t-PA production as well as glucocorticoids. Conversely, melittin, an activator of phospholipase A2, induced t-PA production in a dose-dependent manner. Exogenous phospholipase A2 strongly induced t-PA production and also arachidonic acid moderately did in a dose-dependent manner. P.peptone stimulated the release of radioactive arachidonic acid from 3H-arachidonic acid-labeled IMR-90 cells under the presence of Ca2+. These results suggest that the induction of t-PA production by p.peptone is closely related to the activity of phospholipase A2, that is, the release of arachidonic acid from phospholipids in cell membrane.

Factors affecting the production of nitrile hydratase by thermophilic Bacillus smithii SC-J05-1.

Ammonium present in the medium was found to control the production of nitrile hydratases (NHase) by a moderate thermophile, Bacillus smithii SC-J05-1. The enzyme production is initiated in the absence of ammonium and suppressed in the presence of ammonium. In addition to cobalt ions as the coordinated metal ion, manganese ions are also required for NHase production.

High-density culture of FM-3A cells using a bioreactor with an external tangential-flow filtration device.

A novel bioreactor system developed for high-density cultures of suspended mammalian cells is described using a tangential-flow filtration device outside the culture vessel to separate viable cells from spent medium. The filtration device is based on thin porous microfiltration membranes with a pore size of 0.20-0.65 microns. Because cells have a diameter of about 10-20 microns, they cannot permeate these membranes with the spent medium. So, allowing a perfusion culture to be created using this system. In most membrane filtration systems, clogging of the membranes has made long-term operation difficult. In this system, however, high pressure is not applied directly to the membrane, thus minimizing clogging. Also, clogging of the membrane was prevented by washing the membrane surface once a day, and increasing the membrane surface area. With this system, FM-3A cells were cultured and maintained at a high density of 3.0 x 10(7) cells/ml for two weeks, and a continuous culture was supported for as long as 34 days.

Receptor-linked antigen delivery system. Importance of autologous alpha 2-macroglobulin in the development of peptide vaccine.

We have hijacked a process of the receptor-mediated endocytosis to transport peptide antigens into antigen presenting cells (APCs) for the purpose of increasing the level of antigen presentation (named Receptor-Linked Antigen Delivery System (R-LADS)). By coupling an endogenous plasma proteinase inhibitor alpha 2-macroglobulin (alpha 2M) to a synthetic peptide having a partial sequence of HIV-1 envelope protein, alpha 2M was made to carry the peptide into APCs as a part of the normal alpha 2M cycle, which resulted in an increased production of specific antibodies against the peptide (Mitsuda, S., Nakagawa, T., Osada, T., Shimamoto, T., Nakazato, H. and Ikai, A. (1993) Biochem. Biophys. Res. Commun. 194, 1155-1160). We demonstrate here that this procedure becomes a more efficient tool for antibody production when autologous transporter protein was used. By using murine alpha 2M (m alpha 2M) instead of heterologous human alpha 2M (h alpha 2M) when mice were experimental animals, we were able to dramatically enhance the production level of anti-HIV-1 peptide antibodies and shorten the period which is needed for antibody production. We aim to develop effective peptide vaccines by further improving this system.

A receptor-mediated antigen delivery and incorporation system. Administration of alpha 2-macroglobulin-cytochrome c conjugate induced high concentrations of antibodies against cytochrome c in mice.

Specific receptors for alpha 2-macroglobulin (alpha 2M) are found on the plasma membrane of macrophages (M phi s), one of antigen presenting cells. So far, a receptor-mediated effective uptake by M phi of foreign antigens which were linked to alpha 2M has been shown to provoke a remarkable increase in the proliferation of T lymphocytes and in the production of antibodies in vitro. Such results encouraged us to develop a new type of vaccine using a receptor-mediated antigen delivery and incorporation system based on alpha 2M and its receptor interaction. In this report, we applied the system to experimental animals. Yeast cytochrome c was used as an antigen to see if the system worked in vivo as well as in vitro. Cytochrome c was conjugated to alpha 2M through the action of trypsin and intraperitoneally administered to mice. The titer induced in mice was measured by enzyme linked immunosorbent assay (ELISA). The production of antibodies against cytochrome c was significantly increased when the protein was given in conjugated forms with alpha 2M.

Characterization of a unique scrambled peptide derived from the CD4 CDR3-related region which shows substantial activity for blocking HIV-1 infection.

We have previously identified CD4 peptides that exhibited blocking activity on human immunodeficiency virus type 1 (HIV-1) infection, i.e. CD4(68-130) and CD4(66-92) which include the region corresponding to the third complementarity-determining region of IgG. Here we describe a unique peptide derived from CD4(66-92), altered in amino acid sequence but not in composition, which was found to have increased anti-HIV-1 activity. The acidic amino acid residues in this scrambled peptide, S1, localized at the N-terminus, while in the native peptide they clustered at the C-terminus. On the other hand, a second scrambled peptide, S2, in which the acidic amino acid residues were fully dispersed, did not show any anti-HIV-1 activity. However, we could not identify any correlation between CD4(66-92) and S1 peptides by their hydrophobic or circular dichroism spectrum analyses. The results provide insight into the mechanisms of HIV-1 gp120 and CD4 interaction and may be useful as a new approach to AIDS therapy.

Purification and characterization of tissue plasminogen activator secreted by human embryonic lung diploid fibroblasts, IMR-90 cells.

The anti-urokinase-IgG-resistant plasminogen activator secreted by human embryonic lung diploid fibroblasts, IMR-90 cells (ATCC, CCL186) was purified to homogeneity from serum-free conditioned medium by a four-step procedure. The fibroblast plasminogen activator was identified as tissue plasminogen activator (t-PA) by the N-terminal sequence of the purified material and the complete amino acid sequence deduced from its complementary DNA (cDNA). The apparent molecular weight was the range of 64,000 to 68,000 by SDS-PAGE and was in the range of 69,000 to 72,000 by gel filtration. The fibroblast t-PA showed a stricter substrate specificity than urokinase in enzymatic hydrolysis of various chromogenic substrates. Compared to urokinase, the fibrobrast t-PA was more stable by heating at 95 degrees C for five min and was stable from pH 5 to 10. The fibrorast t-PA had a higher affinity for fibrin than urokinase.