Validation of a New Methodology to Create Oral Drugs beyond the Rule of 5 for Intracellular Tough Targets.

Establishing a technological platform for creating clinical compounds inhibiting intracellular protein-protein interactions (PPIs) can open the door to many valuable drugs. Although small molecules and antibodies are mainstream modalities, they are not suitable for a target protein that lacks a deep cavity for a small molecule to bind or a protein found in intracellular space out of an antibody's reach. One possible approach to access these targets is to utilize so-called middle-size cyclic peptides (defined here as those with a molecular weight of 1000-2000 g/mol). In this study, we validated a new methodology to create oral drugs beyond the rule of 5 for intracellular tough targets by elucidating structural features and physicochemical properties for drug-like cyclic peptides and developing library technologies to afford highly N-alkylated cyclic peptide hits. We discovered a KRAS inhibitory clinical compound (LUNA18) as the first example of our platform technology.

Methiothepin downregulates SNAP-23 and inhibits degranulation of rat basophilic leukemia cells and mouse bone marrow-derived mast cells.

In the present study, we found that methiothepin (a nonselective 5-hydroxytryptamine [5-HT] receptor antagonist) inhibited antigen-induced degranulation in rat basophilic leukemia cells and mouse bone marrow-derived mast cells. Although antigen stimulation induces release of histamine and serotonin (5-HT) by exocytosis and mast cells express several types of 5-HT receptor, the detailed role of these receptors remains unclear. Here, pretreatment of cells with methiothepin attenuated increased intracellular Ca2+ concentration, phosphorylated critical upstream signaling components (Src family tyrosine kinases, Syk, and PLCγ1), and suppressed TNF-α secretion via inhibition of Akt (a Ser/Thr kinase activated by PI3K)and ERK phosphorylation. Furthermore, it inhibited PMA/ionomycin-induced degranulation; this finding suggested that methiothepin affected downstream signaling. IκB kinase ß phosphorylates synaptosomal associated protein 23, which regulates the fusion events of the secretory granule/plasma membrane after mast cell activation, resulting in degranulation. We showed that methiothepin blocked PMA/ionomycin-induced phosphorylation of synaptosomal associated protein 23 by inhibiting its interaction with IκB kinase ß. Together with the results of selective 5-HT antagonists, it is suggested that methiothepin inhibits mast cell degranulation by downregulating upstream signaling pathways and exocytotic fusion machinery through mainly 5-HT1A receptor. Our findings provide that 5-HT antagonists may be used to relieve allergic reactions.

Calcineurin B1 Deficiency Reduces Proliferation, Increases Apoptosis, and Alters Secretion in Enteric Glial Cells of Mouse Small Intestine in Culture.

To investigate the roles of calcineurin (CN) in glial cells, we previously generated conditional knockout (CKO) mice lacking CNB1 in glial cells. Because these CKO mice showed dysfunction and inflammation of the small intestine in addition to growth impairment and postweaning death, we have focused on enteric glial cells (EGCs) in the small intestine. In this study, we examined the effects of CNB1 deficiency on the proliferation and survival of EGCs and the expression and secretion of EGC-derived substances in culture to reveal the mechanisms of how CNB1 deficiency leads to dysfunction and inflammation of the small intestine. In primary myenteric cultures of the small intestine, EGCs from the CKO mice showed reduced proliferation and increased apoptosis compared with EGCs from control mice. In purified EGC cultures from the CKO mice, Western blot analysis showed increased expression of S100B, iNOS, GFAP, and GDNF, and increased phosphorylation of NF-κB p65. In the supernatants of purified EGC cultures from the CKO mice, ELISA showed reduced secretion of TGF-ß1. In contrast, GDNF secretion was not altered in purified EGC cultures from the CKO mice. Furthermore, treatment with an S100B inhibitor partially rescued the CKO mice from growth impairment and postweaning death in vivo. In conclusion, CNB1 deficiency leads to reduced proliferation and increased apoptosis of EGCs and abnormal expression and secretion of EGC-derived substances, which may contribute to dysfunction and inflammation of the small intestine.

Ferrimagnetic Ordering and Spin-Glass State in Diluted GdFeO3-Type Perovskites (Lu0.5Mn0.5)(Mn1-xTix)O3 with x = 0.25, 0.50, and 0.75.

ABO3 perovskite materials with small cations at the A site, especially those with ordered cation arrangements, have attracted a great deal of interest because they show unusual physical properties and deviations from the general characteristics of perovskites. In this work, perovskite solid solutions (Lu0.5Mn0.5)(Mn1-xTix)O3 with x = 0.25, 0.50, and 0.75 were synthesized by means of a high-pressure, high-temperature method at approximately 6 GPa and approximately 1550 K. All the samples crystallize in the GdFeO3-type perovskite structure (space group Pnma) and have random distributions of the small Lu3+ and Mn2+ cations at the A site and Mn4+/3+/2+ and Ti4+ cations at the B site, as determined by Rietveld analysis of high-quality synchrotron X-ray powder diffraction data. Lattice parameters are a = 5.4431 Å, b = 7.4358 Å, c = 5.1872 Å (for x = 0.25); a = 5.4872 Å, b = 7.4863 Å, c = 5.2027 Å (for x = 0.50); and a = 5.4772 Å, b = 7.6027 Å, c = 5.2340 Å (for x = 0.75). Despite a significant dilution of the A and B sublattices by non-magnetic Ti4+ cations, the x = 0.25 and 0.50 samples show long-range ferrimagnetic order below TC = 89 K and 36 K, respectively. Mn cations at both A and B sublattices are involved in the long-range magnetic order. The x = 0.75 sample shows a spin-glass transition at TSG = 6 K and a large frustration index of approximately 22. A temperature-independent dielectric constant was observed for x = 0.50 (approximately 32 between 5 and 150 K) and for x = 0.75 (approximately 50 between 5 and 250 K).

A Simple Method for Purified Primary Culture of Enteric Glial Cells from Mouse Small Intestine.

Enteric glial cells (EGCs) have been recognized as an important cell type constituting the enteric nervous system. EGCs control intestinal function and homeostasis through interactions with enteric neurons, epithelial cells and immune cells. To clarify the roles of EGCs in intestinal function and homeostasis, especially through secretion of and response to physiologically active substances, purified EGCs in primary culture have great advantages as an experimental tool. However, contamination by other cell types, fibroblasts in particular, is problematic in conventional primary myenteric culture. Previous methods to purify primary EGCs take a long time (over one month), are expensive, and are labor intensive. In the present study, we sought to purify primary EGCs from mouse small intestine by a simpler method than previous ones. After trying various protocols, we have established a method combining serum-free treatment and scraping fibroblasts off with a pipette tip. With our method, a purity of more than 90% EGCs was achieved after 14-d primary culture. Thus, our method is useful for investigating the roles of EGCs in intestinal function and homeostasis in detail in vitro.

Temperature-dependent electronic structure of bixbyite α-Mn2O3 and the importance of a subtle structural change on oxygen electrocatalysis.

Bixbyite α -Mn2O3 is an inexpensive Earth-abundant mineral that can be used to drive both oxygen evolution (OER) and oxygen reduction reactions (ORR) in alkaline conditions. It possesses a subtle orthorhombic → cubic phase change near room temperature that suppresses Jahn-Teller distortions and presents a unique opportunity to study how atomic structure affects the electronic structure and catalytic activity at a temperature range that is easily accessible in OER/ORR experiments. Previously, we observed that heat-treated α -Mn2O3 had a better performance as a bifunctional catalyst in the oxygen evolution (OER) and oxygen reduction reactions (ORR) (Dalton Trans. 2016, 45, 18,494-18,501). We hypothesized that heat-treatment pinned the material into a more electrochemically active cubic phase. In this manuscript, we use high-resolution X-ray diffraction to collect the temperature-dependent structures of α -Mn2O3, and then input them into ab initio calculations. The electronic structure calculations indicate that the orthorhombic → cubic phase transition causes the Mn 3d and O 2p bands to overlap and mix covalently, transforming α -Mn2O3 from a semiconductor to a semimetal. This subtle change in structure also modifies Mn-O-Mn bond distances, which may improve the activity of the material in oxygen electrochemistry. OER and ORR experiments were performed using the same electrode at various temperatures. They show a jump in the exchange current density near the phase change temperature, demonstrating the higher activity of the cubic phase.

Promotion of Dendritic Differentiation of Cerebellar Purkinje Cells by Ca2+/calmodulin-dependent Protein Kinase IIα, IIß and IV and Possible Involvement of CREB Phosphorylation.

Cerebellar Purkinje cells develop the most elaborate dendritic trees among neurons in the brain. To examine the role of Ca2+/calmodulin-dependent protein kinase (CaMK) IIα, IIß and IV in the dendritic differentiation of Purkinje cells, we introduced siRNA against these CaMKs into Purkinje cells in cerebellar cell cultures using a single-cell electroporation technique. Single-cell electroporation enables us to transfer siRNA into specific cells within a heterogeneous cell population. In addition, we can easily and reliably transfer multiple types of siRNA into a cell simply by loading them together in one micropipette. Any one of the siRNA against CaMKIIα, IIß and IV (single knockdown) or any combinations of two of the siRNA against these CaMKs (double knockdown) had no significant effects on the dendritic differentiation of Purkinje cells. However, the combination of all three siRNA against these CaMKs (triple knockdown) inhibited the branching of Purkinje cell dendrites. Furthermore, the triple knockdown reduced the phosphorylation of CREB in Purkinje cells. These findings suggest the promotion of dendritic differentiation of Purkinje cells by CaMKIIα, IIß and IV and the possible involvement of phosphorylation of CREB as a common substrate of these CaMKs.

High-Pressure Synthesis, Crystal Structures, and Properties of A-Site Columnar-Ordered Quadruple Perovskites NaRMn2Ti4O12 with R = Sm, Eu, Gd, Dy, Ho, Y.

The formation of NaRMn2Ti4O12 compounds (R = rare earth) under high pressure (about 6 GPa) and high temperature (about 1750 K) conditions was studied. Such compounds with R = Sm, Eu, Gd, Dy, Ho, Y adopt an A-site columnar-ordered quadruple-perovskite structure with the generic chemical formula A2A'A″B4O12. Their crystal structures were studied by powder synchrotron X-ray and neutron diffraction between 1.5 and 300 K. They maintain a paraelectric structure with centrosymmetric space group P42/nmc (No. 137) at all temperatures, in comparison with the related CaMnTi2O6 perovskite, in which a ferroelectric transition occurs at 630 K. The centrosymmetric structure was also confirmed by second-harmonic generation. It has a cation distribution of [Na+R3+]A[Mn2+]A'[Mn2+]A″[Ti4+4]BO12 (to match with the generic chemical formula) with statistical distributions of Na+ and R3+ at the large A site and a strongly split position of Mn2+ at the square-planar A' site. We found a C-type long-range antiferromagnetic structure of Mn2+ ions at the A' and A″ sites below TN = 12 K for R = Dy and found that the presence of Dy3+ disturbs the long-range ordering of Mn2+ below a second transition at lower temperatures. The first magnetic transition occurs below 8-13 K in all compounds, but the second magnetic transition occurs only for R = Dy, Sm, Eu. All compounds show large dielectric constants of a possible extrinsic origin similar to that of CaCu3Ti4O12. NaRMn2Ti4O12 with R = Er-Lu crystallized in the GdFeO3-type Pnma perovskite structure, and NaRMn2Ti4O12 with R = La, Nd contained two perovskite phases: an AA'3B4O12-type Im3̅ phase and a GdFeO3-type Pnma phase.

Study of Polycrystalline Bulk Sr3OsO6 Double-Perovskite Insulator: Comparison with 1000 K Ferromagnetic Epitaxial Films.

Polycrystalline Sr3OsO6, which is an ordered double-perovskite insulator, is synthesized via solid-state reaction under high-temperature and high-pressure conditions of 1200 °C and 6 GPa. The synthesis enables us to conduct a comparative study of the bulk form of Sr3OsO6 toward revealing the driving mechanism of 1000 K ferromagnetism, which has recently been discovered for epitaxially grown Sr3OsO6 films. Unlike the film, the bulk is dominated by antiferromagnetism rather than ferromagnetism. Therefore, robust ferromagnetic order appears only when Sr3OsO6 is under the influence of interfaces. A specific heat capacity of 39.6(9) × 10-3 J mol-1 K-2 is found at low temperatures (<17 K). This value is remarkably high, suggesting the presence of possible Fermionic-like excitations at the magnetic ground state. Although the bulk and film forms of Sr3OsO6 share the same lattice basis and electrically insulating state, the magnetism is entirely different between them.

Emerging Atomic Energy Levels in Zero-Dimensional Silicon Quantum Dots.

Driven by the emergence of colloidal semiconductor quantum dots (QDs) of tunable emission wavelengths, characteristic of exciton absorption peaks, outstanding photostability and solution processability in device fabrication have become a key tool in the development of nanomedicine and optoelectronics. Diamond cubic crystalline silicon (Si) QDs, with a diameter larger than 2 nm, terminated with hydrogen atoms are known to exhibit bulk-inherited spin and valley properties. Herein, we demonstrate a newly discovered size region of Si QDs, in which a fast radiative recombination on the order of hundreds of picoseconds is responsible for photoluminescence (PL). Despite retaining a crystallographic structure like the bulk, controlling their diameters in the 1.1-1.7 nm range realizes the strong PL with continuous spectral tunability in the 530-580 nm window, the narrow spectral line widths without emission tails, and the fast relaxation of photogenerated carriers. In contrast, QDs with diameters greater than 1.8 nm display the decay times on the microsecond order as well as the previous Si QDs. In addition to the five-orders-of-magnitude variation in the PL decay time, a systematic study on the temperature dependence of PL properties suggests that the energy structure of the smaller QDs does not retain an indirect band gap character. It is discussed that a 1.7 nm diameter is critical to undergo changes in energy structure from bulky to molecular configurations.

Spin-Glass Magnetic Properties of A-Site Columnar-Ordered Quadruple Perovskites Y2MnGa(Mn4-xGax)O12 with 0 ≤ x ≤ 3.

Y2MnGa(Mn4-xGax)O12 solid solutions were synthesized at high pressure of ∼6 GPa and high temperature of ∼1570 K for the 0 ≤ x ≤ 3 compositional range. Synchrotron X-ray and neutron powder diffraction were used to study the crystal structures and cation distributions. These solutions adopt the parent structure of the A-site columnar-ordered quadruple perovskite family with space group P42/nmc (No. 137). They have lattice parameters of a = 7.36095 Å and c = 7.753 84 Å (x = 0), a = 7.361 68 Å and c = 7.716 16 Å (x = 1), a = 7.360 34 Å and c = 7.67142 Å (x = 2), and a = 7.363 93 Å and c = 7.616 85 Å (x = 3) at room temperature. The x = 0 sample has a cation distribution of [Y3+2]A[Mn3+]A'[Ga3+0.68Mn2+0.32]A″[Mn3.68Ga0.32]BO12 with a preferred localization of Ga3+ in the tetrahedral A″ site and with a small amount of Ga3+ in the octahedral B site. A complete triple A-site order, [Y3+2]A[Mn3+]A'[Ga3+]A″[Mn3+4-xGa3+x]BO12, is realized for x ≥ 1. All samples demonstrate spin-glass-like magnetic properties, and the absence of a long-range magnetic order at the ground state at 1.5 K was confirmed by neutron diffraction for the x = 1 sample. First-principles calculations indicated the spin-glass-like magnetic ordering is derived from the Ga substitution to the B sites and gave evidence that the ideal cation distribution could produce robust ferromagnetism in this family of perovskites.

Calcineurin B1 Deficiency in Glial Cells Reduces Gastrointestinal Motility and Results in Maldigestion and/or Malabsorption in Mice.

Calcineurin is a Ca2+/calmodulin-dependent protein phosphatase abundantly expressed in the nervous system. To investigate the roles of calcineurin in glial cells, we previously generated glial calcineurin B1-conditional knockout (CKO) mice and found that these mice displayed dysfunction of enteric glial cells, mucosal degeneration and inflammation in the small intestine, and growth retardation and postweaning death, suggesting a novel role of calcineurin in enteric glial cells. Although these findings raised a possibility that abnormalities in calcineurin B1-deficient enteric glial cells may cause dysregulation of gastrointestinal motility and result in maldigestion and/or malabsorption in the CKO mice, these issues remain to be elucidated. In the present study, we showed that gastrointestinal motility was reduced in the CKO mice. Degeneration of mucosal epithelium was observed in the small intestine. Glucose levels were decreased in serum, whereas starch, glucose, and lipid levels were increased in feces. Thus, calcineurin B1 deficiency in glial cells reduces gastrointestinal motility, induces mucosal epithelium degeneration in the small intestine, and results in maldigestion and/or malabsorption in mice, further supporting the important role of calcineurin in enteric glial cells.

Valence Variations by B-Site Doping in A-Site Columnar-Ordered Quadruple Perovskites Sm2MnMn(Mn4- xTi x)O12 with 1 ≤ x ≤ 3.

Sm2MnMn(Mn4- xTi x)O12 with 1 ≤ x ≤ 3 were prepared by a high-pressure, high-temperature method at 6 GPa and about 1570-1670 K. They belong to a family of A-site columnar-ordered quadruple perovskites A2A'A″B4O12, where A' is a site with a square-planar coordination and A″ is a site with a tetrahedral coordination. Their crystal structures were investigated using synchrotron X-ray and neutron powder diffraction. They crystallize in space group P42/ nmc (No. 137) with a = 7.41172 Å and c = 7.97131 Å for x = 1, a = 7.54945 Å and c = 7.76756 Å for x = 2, and a = 7.63949 Å and c = 7.70339 Å for x = 3 at 295 K. The determined charge and cation distributions are [Sm3+1.88Mn2+0.12]A[Mn3+]A'[Mn2+0.88Sm3+0.12]A″[Mn3+3Ti4+]BO12 for x = 1, [Sm3+1.91Mn2+0.09]A[Mn2+]A'[Mn2+0.91Sm3+0.09]A″[Mn3+2Ti4+2]BO12 for x = 2, and [Sm3+1.88Mn2+0.12]A[Mn2+0.88Sm3+0.12]A'[Mn2+]A″[Mn2+Ti4+3]BO12 for x = 3. Mn and Ti are distributed randomly in one B site in all compounds with the average oxidation state changing from +3.25 to +3.5 per one B atom, and such flexibility is realized because Mn at the A' site can change its oxidation state between +2 and +3. Sm and Mn are slightly disordered between the A and A″ sites for x = 1 and 2, and between the A and A' sites for x = 3. The x = 1 sample shows spin-canted antiferromagnetic properties with TN = 27 K, and the x = 2 sample, with TN = 62 K. On the other hand, the x = 3 sample is a ferrimagnet, confirmed by neutron diffraction, with TC = 40 K. The x = 3 sample shows relaxor-like dielectric properties below 220 K.

Intrinsic Triple Order in A-site Columnar-Ordered Quadruple Perovskites: Proof of Concept.

There is an emerging topic in the science of perovskite materials: A-site columnar-ordered A2 A'A''B4 O12 quadruple perovskites, which have an intrinsic triple order at the A sites. However, in many examples reported so far, A' and A'' cations are the same, and the intrinsic triple order is hidden. Here, we investigate structural properties of Dy2 CuMnMn4 O12 (1) and Ho2 MnGaMn4 O12 (2) by neutron and X-ray powder diffraction and prove the triple order at the A sites. The cation distributions determined are [Ho2 ]A [Mn]A' [Ga0.66 Mn0.34 ]A'' [Mn3.66 Ga0.34 ]B O12 and [Dy2 ]A [Cu0.73 Mn0.27 ]A' [Mn0.80 Dy0.20 ]A'' [Mn1.89 Cu0.11 ]B1 [Mn2 ]B2 O12 . There are clear signatures of Jahn-Teller distortions in 1 and 2, and the orbital pattern is combined with an original type of charge ordering in 1. Columnar-ordered quadruple perovskites represent a new playground to study complex interactions between different electronic degrees of freedom. No long-range magnetic order was found in 2 by neutron diffraction, and its magnetic properties in low fields are dominated by an impurity with negative magnetization or magnetization reversal. On the other hand, 1 shows three magnetic transitions at 21, 125, and 160 K.

High-Pressure Synthesis, Crystal Structure, and Semimetallic Properties of HgPbO3.

The crystal structure of HgPbO3 was studied using single-crystal X-ray diffraction and powder synchrotron X-ray diffraction. The structure was well characterized as a centrosymmetric model with a space group of R-3 m [hexagonal setting: a = 5.74413(6) Å and c = 7.25464(8) Å] rather than as a noncentrosymmetric model as was expected. It was found that Pb4+ is octahedrally coordinated by six oxygen atoms as usual, while Hg2+ is coordinated by three oxygen atoms in a planar manner, this being a very rare coordination of Hg in a solid-state material. The magnetic and electronic transport properties were investigated in terms of the magnetic susceptibility, magnetization, Hall coefficient, and specific heat capacity of polycrystalline HgPbO3. Although HgPbO3 has a carrier concentration (=7.3-8.5 × 1020 cm-3) that is equal to that of metallic oxides, the very weak temperature dependence of the electrical resistivity (residual-resistivity ratio ∼1.5), the significant diamagnetism (= -1.02 × 10-4 emu mol-1 at 300 K) that is in the same order of that of Bi powder and the remarkably small Sommerfeld coefficient [=1.6(1) × 10-3 J mol-1 K-2] implied that it is semimetallic in nature. HgPbO3 does not have a cage structure; nevertheless, at temperatures below approximately 50 K, it clearly exhibits phonon excitation of an anharmonic vibrational mode that is as significant as those of RbOs2O6. The mechanism of the anharmonic mode of the HgPbO3 has yet to be identified, however.

Sciatic Neuralgia Caused by May-Thurner Syndrome.

BACKGROUND: Sciatic neuralgia (SN) is a type of pain commonly associated with lumbosacral radiculopathy. May-Thurner syndrome (MTS) is a disease characterized by venous congestion in the left common iliac vein (LCIV) due to right common iliac artery compression. A case of MTS with SN as the first presenting symptom is described. CASE DESCRIPTION: A 53-year-old man gradually developed left SN; however, radiologic examination showed no causal findings. Conservative medical treatment did not result in satisfactory pain relief. During the follow-up period, edema and brownish skin pigmentation were noted on the left crural region. Follow-up magnetic resonance imaging of lumbar spine revealed vascular enlargement around the spine and compression of the LCIV by the right common iliac artery. Furthermore, the left S1 nerve root was compressed by enlarged veins, which was thought to be the cause of the left S1 radiculopathy and SN. After the stenosed part of the LCIV was expanded, the pain and edema in the left leg disappeared. CONCLUSIONS: This is the first report of SN associated with MTS. MTS should be considered as one of the differential diagnoses of SN. Thus, symptoms such as pain, edema, and skin hyperpigmentation on the left leg should be closely noted.

High-Pressure Synthesis, Structures, and Properties of Trivalent A-Site-Ordered Quadruple Perovskites RMn7O12 (R = Sm, Eu, Gd, and Tb).

A-site-ordered quadruple perovskites RMn7O12 with R = Sm, Eu, Gd, and Tb were synthesized at high pressure and high temperature (6 GPa and ∼1570 K), and their structural, magnetic, and dielectric properties are reported. They crystallize in space group I2/ m at room temperature. All four compounds exhibit a high-temperature phase transition to the cubic Im3̅ structure at ∼664 K (Sm), 663 K (Eu), 657 K (Gd), and 630 K (Tb). They all show one magnetic transition at TN1 ≈ 82-87 K at zero magnetic field, but additional magnetic transitions below TN2 ≈ 12 K were observed in SmMn7O12 and EuMn7O12 at high magnetic fields. Very weak kinklike dielectric anomalies were observed at TN1 in all compounds. We also observed pyroelectric current peaks near 14 K and frequency-dependent sharp steps in dielectric constant (near 18-35 K)-these anomalies are probably caused by dielectric relaxation, and they are not related to any ferroelectric transitions. TbMn7O12 shows signs of nonstoichiometry expressed as (Tb1- xMn x)Mn7O12, and these samples exhibit negative magnetization or magnetization reversal effects of an extrinsic origin on zero-field-cooled curves in intermediate temperature ranges. The crystal structures of SmMn7O12 and EuMn7O12 were refined from neutron powder diffraction data at 100 K, and the crystal structures of GdMn7O12 and (Tb0.88Mn0.12)Mn7O12 were studied by synchrotron X-ray powder diffraction at 295 K.

Calcineurin B1 Deficiency in Glial Cells Induces Mucosal Degeneration and Inflammation in Mouse Small Intestine.

Although calcineurin is abundantly expressed in the nervous system and involved in neurite extension and synaptic plasticity in neurons, little is known about its roles in glial cells. To investigate the roles of calcineurin in glial cells, we generated glial calcineurin B1-conditional knockout (CKO) mice and analyzed the abnormalities in the small intestine. The CKO mice were generated by crossing floxed calcineurin B1 mice with glial fibrillary acidic protein (GFAP)-Cre mice. The CKO mice exhibited growth retardation approximately from the third postnatal week and died mostly within the fourth postnatal week. The small intestine of the CKO mice was thin and hemorrhagic. The mucosal layer was degenerated and GFAP expression was reduced in the CKO small intestine. These pathological changes were associated with inflammation and increased intestinal permeability. In contrast, no apparent abnormalities were observed in the large intestine of the CKO mice. Nuclear factor of activated T cells failed to translocate into the nucleus after stimulation in enteric glial cells of the CKO small intestine. In conclusion, the calcineurin B1 deficiency in glial cells impairs the small intestine and leads to malnutrition and eventual death in mice, suggesting that calcineurin plays a novel and important role in enteric glial cells.

Development of 3D imaging technique of reconstructed human epidermis with immortalized human epidermal cell line.

The epidermis, the outermost layer of the skin, retains moisture and functions as a physical barrier against the external environment. Epidermal cells are continuously replaced by turnover, and thus to understand in detail the dynamic cellular events in the epidermis, techniques to observe live tissues in 3D are required. Here, we established a live 3D imaging technique for epidermis models. We first obtained immortalized human epidermal cell lines which have a normal differentiation capacity and fluorescence-labelled cytoplasm or nuclei. The reconstituted 3D epidermis was prepared with these lines. Using this culture system, we were able to observe the structure of the reconstituted epidermis live in 3D, which was similar to an in vivo epidermis, and evaluate the effect of a skin irritant. This technique may be useful for dermatological science and drug development.

Mn Self-Doping of Orthorhombic RMnO3 Perovskites: (R0.667Mn0.333)MnO3 with R = Er-Lu.

Orthorhombic rare-earth trivalent manganites RMnO3 (R = Er-Lu) were self-doped with Mn to form (R0.667Mn0.333)MnO3 compositions, which were synthesized by a high-pressure, high-temperature method at 6 GPa and about 1670 K from R2O3 and Mn2O3. The average oxidation state of Mn is 3+ in (R0.667Mn0.333)MnO3. However, Mn enters the A site in the oxidation state of 2+, creating the average oxidation state of 3.333+ at the B site. The presence of Mn2+ was confirmed by hard X-ray photoelectron spectroscopy measurements. Crystal structures were studied by synchrotron powder X-ray diffraction. (R0.667Mn0.333)MnO3 crystallizes in space group Pnma with a = 5.50348(2) Å, b = 7.37564(1) Å, and c = 5.18686(1) Å for (Lu0.667Mn0.333)MnO3 at 293 K, and they are isostructural with the parent RMnO3 manganites. Compared with RMnO3, (R0.667Mn0.333)MnO3 exhibits enhanced Néel temperatures of about TN1 = 106-110 K and ferrimagnetic or canted antiferromagnetic properties. Compounds with R = Er and Tm show additional magnetic transitions at about TN2 = 9-16 K. (Tm0.667Mn0.333)MnO3 exhibits a magnetization reversal or negative magnetization effect with a compensation temperature of about 16 K.