Histological and lectin histochemical studies in the vomeronasal organ of the Korean black goat, Capra hircus coreanae.

We examined the localization of olfactory marker protein (OMP), protein gene product9.5 (PGP9.5), and glycan diversity in the vomeronasal organ (VNO) of the Korean black goat (Capra hircus coreanae) during the prenatal and postnatal periods using immunohistochemistry and lectin histochemistry. In fetal and 1-day-old goats, OMP was occasionally identified in receptor cells of the VNO sensory epithelium, and PGP9.5 was localized in both the sensory and non-sensory epithelia. In VNO from adult goats, OMP was abundant in the sensory epithelium and scarce in single cells of the non-sensory epithelium. These results suggest that OMP production is initiated in the VNO sensory epithelium (VNE) during the fetal stage, and that its activity is increased in adult VNO receptor cells and solitary cells in the non-sensory epithelium (VNSE). Furthermore, the free borders of the sensory epithelia were positive for 7 lectins, and 6 lectins were moderately and/or highly abundant in receptor cells. Supporting and basal cells, and nerve bundles had similar expression patterns. In VNE, 7 lectins were observed in the free border, and 6 in ciliated, goblet, and basal cells, and in gland acini. The intensities of WGA, LCA, and PNA were high in VSE receptor cells, and the intensity of PNA was high in ciliated cells of the VNSE. The other 3 lectins showed similar patterns throughout development. Collectively, these results confirm that the Korean black goat VNO starts developing during the late fetal stages and differentiates further after birth.

Amelioration of experimental autoimmune encephalomyelitis by Ishige okamurae.

Experimental autoimmune encephalomyelitis (EAE) is a T-cell-mediated autoimmune central nervous system disease characterized by inflammation with oxidative stress. The aim of this study was to evaluate an anti-inflammatory effect of Ishige okamurae on EAE-induced paralysis in rats. An ethanolic extract of I. okamurae significantly delayed the first onset and reduced the duration and severity of hind-limb paralysis. The neuropathological and immunohistochemical findings in the spinal cord were in agreement with these clinical results. T-cell proliferation assay revealed that the ethyl-acetate fraction of I. okamurae suppressed the proliferation of myelin basic protein reactive T cells from EAE affected rats. Flow cytometric analysis showed TCRαß+ T cells was significantly reduced in the spleen of EAE rats with I. okamurae treatment with concurrent decrease of inflammatory mediators including tumor necrosis factor-α and cyclooxygenase-2. Collectively, it is postulated that I. okamurae ameliorates EAE paralysis with suppression of T-cell proliferation as well as decrease of pro-inflammatory mediators as far as rat EAE is concerned.

Anti-complement component 5 antibody targeting MG4 domain inhibits choroidal neovascularization.

Age-related macular degeneration (AMD) is one of the main causes of visual impairment in adults. Visual deterioration is more prominent in neovascular AMD with choroidal neovascularization (CNV). Clinical and postmortem studies suggested that complement system activation might induce CNV. In this study, we demonstrated that an anti-mouse complement component 5 (C5) antibody targeting MG4 domain of ß chain effectively inhibited CNV which was induced by laser photocoagulation in mice. The targeted epitope of this anti-C5 antibody was different from that of currently utilized anti-C5 antibody (eculizumab) in the MG7 domain in which a single nucleotide polymorphism (R885H/C) results in poor response to eculizumab. Even with targeting MG4 domain, this anti-C5 antibody reduced production of C5a, monocyte chemoattractant protein-1 and vascular endothelial growth factor to prevent infiltration of F4/80-positive cells into CNV lesions and formation of CNV. Furthermore, anti-C5 antibody targeting MG4 domain induced no definite toxicity in normal retina. These results demonstrated that anti-C5 antibody targeting MG4 domain inhibited CNV in neovascular AMD.

Evolution of crystal structures in GeTe during phase transition.

We investigated changes in the crystal structure of GeTe during its phase transition. Using density functional theory (DFT) calculations, four possible crystal structures were identified: R3m, P1, Cm, and Fm3m. Among these, P1 and Cm were examined here for the first time. By calculating the internal energy of the crystal volume change, we verified that P1, R3m, and Cm can coexist in crystalline GeTe. The X-ray diffraction spectra of annealed and laser-irradiated GeTe films revealed coexisting P1 or R3m and Cm. In addition, we confirmed that Cm transforms into P1 or R3m after laser irradiation. The presence of these new structures was revealed in the crystal Raman spectra. Many of the Raman peaks in the crystalized GeTe could be explained by the coexistence of various structures. By calculating the band gaps of these structures, we also found that a structural transformation induces a change in the crystal resistance, owing to differences in the band gaps of individual structures. The generation of new crystal structures suggests a facile phase change and instability during the structural transformation.

Next-generation sequencing enables the discovery of more diverse positive clones from a phage-displayed antibody library.

Phage display technology provides a powerful tool to screen a library for a binding molecule via an enrichment process. It has been adopted as a critical technology in the development of therapeutic antibodies. However, a major drawback of phage display technology is that because the degree of the enrichment cannot be controlled during the bio-panning process, it frequently results in a limited number of clones. In this study, we applied next-generation sequencing (NGS) to screen clones from a library and determine whether a greater number of clones can be identified using NGS than using conventional methods. Three chicken immune single-chain variable fragment (scFv) libraries were subjected to bio-panning on prostate-specific antigen (PSA). Phagemid DNA prepared from the original libraries as well as from the Escherichia coli pool after each round of bio-panning was analyzed using NGS, and the heavy chain complementarity-determining region 3 (HCDR3) sequences of the scFv clones were determined. Subsequently, through two-step linker PCR and cloning, the entire scFv gene was retrieved and analyzed for its reactivity to PSA in a phage enzyme immunoassay. After four rounds of bio-panning, the conventional colony screening method was performed for comparison. The scFv clones retrieved from NGS analysis included all clones identified by the conventional colony screening method as well as many additional clones. The enrichment of the HCDR3 sequence throughout the bio-panning process was a positive predictive factor for the selection of PSA-reactive scFv clones.

Impact of acupuncture by using life-energy (qi) oriental needle on the paralysis of rats with experimental autoimmune encephalomyelitis.

In acupuncture, adaptation to energy flows in body cycles is the key to health and therapy. From the evolution of our thinking about acupuncture, we developed the Life-Energy (Qi) oriental needle (Qi needle). It contains a rotating electromagnetic wave and has a strong affinity for the meridians. We report for the first time on the effect of acupuncture by using a Qi needle (Qi acupuncture) on rat experimental autoimmune encephalomyelitis, a model of human demyelinating multiple sclerosis. Both Qi acupuncture (QA) and general acupuncture (GA) were used on the limbs, at the shaoshang (LU11) and zhongchong (PC9) acupoints, of rats from one day post-immunization (dpi) to 12 dpi. The therapy in the QA groups significantly blocked the onset of EAE paralysis (3/13, 77%, p < 0.05) while all rats in the control EAE groups (12/15) and GA groups (11/13) showed EAE paralysis. In addition, the duration of paralysis was shortened in QA groups (1.5 ± 0.5 days) compared with those of the vehicle (5.5 ± 0.2 days) and GA groups (3.6 ± 1.1 days). The numbers of inflammatory cells and CD4(+) T cells in the QA treated EAE group were significantly reduced compared with those of the EAE control and EAE with GA (p < 0.05). Collectively, the present findings suggest that QA ameliorates the paralysis in rats in an EAE model. The precise mechanism of the amelioration and human studies, however, needs further study.

Immunohistochemical study of arginase-1 in the spinal cords of Lewis rats with experimental autoimmune encephalomyelitis.

Arginase-1, a marker for M2 phenotype alternatively activated macrophages, inhibits inflammation and is associated with phagocytosis of cell debris and apoptotic cells. We analyzed the expression of arginase-1, a competitive enzyme of inducible nitric oxide synthase (iNOS), in the spinal cords of Lewis rats with experimental autoimmune encephalomyelitis (EAE). Western blot analysis showed that both arginase-1 and iNOS significantly increased in the spinal cords of rats at the peak stage of EAE compared with the expression level in control animals (p<0.05) and declined thereafter. Immunofluorescent staining demonstrated that increased expression of arginase-1 in EAE spinal cords was confirmed in macrophages as well as in some neurons and astrocytes that were constitutively positive for arginase-1 in normal spinal cords. A semiquantitative analysis by immunofluorescence showed that in EAE lesions, an increased level of arginase-1 immunoreactivity was matched with ED1-positive macrophages, which were also positive for activin A, a marker for the M2 phenotype. Taking all of these findings into consideration, we postulate that the increased level of arginase-1, which is partly from M2 macrophages, contributes to the modulation of neuroinflammation in EAE lesions, possibly through the reduction of nitric oxide in the lesion via competition with iNOS for the use of L-arginine.

Diphlorethohydroxycarmalol, isolated from the brown algae Ishige okamurae, protects against radiation-induced cell damage in mice.

The aim of this study was to evaluate the radioprotective effects of diphlorethohydroxycarmalol (DPHC), isolated from the brown algae Ishige okamurae, in mice subjected to gamma irradiation. DPHC significantly decreased the level of radiation-induced intracellular reactive oxygen species in cultured Chinese hamster lung fibroblast (V79-4) cells (p < 0.05), enhanced cell viability that decreased after exposure to γ-rays, and reduced radiation-induced apoptosis in the V79-4 cells. Pretreatment with DPHC (100 mg/kg) in mice prior to irradiation significantly protected the intestinal crypt cells in the jejunum (p < 0.01) and maintained villi height (p < 0.01), compared with those of the vehicle-treated irradiated group. Mice pretreated with DPHC also exhibited dose-dependent increases in the bone marrow cell viability. The dose-reduction factor for gamma irradiation in the DPHC-pretreated mice was 2.05 at 3.5 days after irradiation. These results suggest that DHPC plays a role in protecting cells from irradiation-induced apoptosis, through the scavenging of reactive oxygen species in vitro, and that DPHC significantly protected intestinal progenitor cells and bone marrows cells that were decreased by gamma irradiation in vivo.