Neurotrophic activity of honokiol on the cultures of fetal rat cortical neurons

Honokiol, a main biphenyl neolignan of the traditional crude medicine, Magnoliae cortex, was found to show neurotrophic activity on the cultures of rat cortical neurons at concentration from 0.1 to 10 microM. In the cortical neurons cultured in serum-free medium supplemented with B27, honokiol could promote neurite outgrowth. In addition, the survival and growth of neurons were significantly enhanced by adding honokiol to the primary cultures in serum-free medium supplemented with N2. Its neurotrophic activity was comparable to 40 ng mL(-1) of bFGF at concentration of 10 microM.     

Probiotic Bifidobacterium breve Induces IL-10-Producing Tr1 Cells in the Colon

Specific intestinal microbiota has been shown to induce Foxp3⁺ regulatory T cell development. However, it remains unclear how development of another regulatory T cell subset, Tr1 cells, is regulated in the intestine. Here, we analyzed the role of two probiotic strains of intestinal bacteria, Lactobacillus casei and Bifidobacterium breve in T cell development in the intestine. B. breve, but not L. casei, induced development of IL-10-producing Tr1 cells that express cMaf, IL-21, and Ahr in the large intestine. Intestinal CD103⁺ dendritic cells (DCs) mediated B. breve-induced development of IL-10-producing T cells. CD103⁺ DCs from Il10 ^−/−, Tlr2 ^−/−, and Myd88 ^−/− mice showed defective B. breve-induced Tr1 cell development. B. breve-treated CD103⁺ DCs failed to induce IL-10 production from co-cultured Il27ra ^−/− T cells. B. breve treatment of Tlr2 ^−/− mice did not increase IL-10-producing T cells in the colonic lamina propria. Thus, B. breve activates intestinal CD103⁺ DCs to produce IL-10 and IL-27 via the TLR2/MyD88 pathway thereby inducing IL-10-producing Tr1 cells in the large intestine. Oral B. breve administration ameliorated colitis in immunocompromised mice given naïve CD4⁺ T cells from wild-type mice, but not Il10 ^−/− mice. These findings demonstrate that B. breve prevents intestinal inflammation through the induction of intestinal IL-10-producing Tr1 cells.     

Anabolic-androgenic steroid does not enhance compensatory muscle hypertrophy but significantly diminish muscle damages in the rat surgical ablation model

Cellular responses in the compensatory hypertrophied (plantaris) muscle induced by surgical ablation of synergistic muscles (soleus and gastrocnemius) were determined during 10-week anabolic androgenic steroid (AAS) treatment. Adult Wistar male rats were divided randomly into the Control and Steroid groups, and contralateral surgery was performed. Nandrolone decanoate was administered to the Steroid group. [³H]thymidine and [¹⁴C]leucine labeling were used to determine the serial changes in cellular mitotic activity and amino acid uptake. Myogenic cells and cellular responses in blood vessels and nerve fibers were analyzed by immunohistochemistry. Significantly lower cellular mitotic activity associated with lower volume of muscle fiber necrosis was observed in the Steroid group during the first week. However, amino acid uptake and final muscle wet weight gain did not differ between the groups. Marked activation/proliferation of muscular, vascular, and peripheral nerve-related cells was seen with the inflammatory responses in both groups. However, this activation was dependent on the volume of muscle fiber damage and was not preferentially accelerated by AAS loading. These results indicated that AAS loading significantly diminished muscle fiber damages, but they did not accelerate final muscle wet weight gain and activation of myogenic, vascular, and peripheral nerve related cells in the compensatory enlarged muscles.     

Prion removal by nanofiltration under different experimental conditions.

Manufacturing processes used in the production of biopharmaceutical or biological products should be evaluated for their ability to remove potential contaminants, including TSE agents. In the present study, we have evaluated scrapie prion protein (PrP Sc) removal in the presence of different starting materials, using virus removal filters of different pore sizes. Following 75 nm filtration, PrP Sc was detected in the filtrate by Western blot (WB) analysis when a "super-sonicated" microsomal fraction derived from hamster adapted scrapie strain 263K (263K MF) was used as the spike material. In contrast, no PrP Sc was detected when an untreated 263K MF was used. By using spike materials prepared in a manner designed to optimize the particle size distribution within the preparation, only 15 nm filtration was shown to remove PrP Sc to below the limits of detection of the WB assays used under all the experimental conditions. However, infectious PrP Sc was recovered following 15 nm filtration under one experimental condition. The results obtained suggest that the nature of the spike preparation is an important factor in evaluating the ability of filters to remove prions, and that procedures designed to minimize the particle size distribution of the prion spike, such as the "super-sonication" or detergent treatments described herein, should be used for the preparation of the spike materials.     

Sex pheromone of Ostrinia sp. newly found on the leopard plant Farfugium japonicum

Recently, larvae of Ostrinia were found feeding on the leopard plant Farfugium japonicum (Asteraceae), previously unrecorded as a host plant of this genus. The adult moths that developed from these borers were morphologically similar to, but distinct from, Ostrinia zaguliaevi, a monophagous species specialized for feeding on another Asteraceae plant, the butterbur Petasites japonicus. Although the taxonomical status of the moth feeding on F. japonicum is to be determined, distinct morphological differences in the adults strongly suggest this to be a new species (hereafter referred to as O. sp.). To gain an insight into the reproductive isolation between O. sp. and other members of the genus Ostrinia, the female sex pheromone and the males’ response to it were investigated using samples collected from F. japonicum. (Z)‐9‐tetradecenyl acetate (Z9‐14:OAc), (Z)‐11‐tetradecenyl acetate (Z11‐14:OAc), (E)‐11‐tetradecenyl acetate (E11‐14:OAc), tetradecyl acetate, and (Z)‐11‐hexadecenyl acetate were identified as candidates for sex pheromone components by analyses using gas chromatographs coupled to a mass spectrometer (GC‐MS) and electroantennographic detector (GC‐EAD). A series of bioassays of male responses in a wind‐tunnel and a field cage indicated that the former three compounds are essential for attracting males, and the latter two have no synergistic effect on the attraction. We therefore concluded that Z9‐14:OAc, Z11‐14:OAc and E11‐14:OAc are the sex pheromone components of O. sp. Although the same three compounds are used as the sex pheromone components of O. zaguliaevi and another congener, Ostrinia zealis, the blend proportions differed greatly among the three (Z9‐14:OAc/Z11‐14:OAc/E11‐14:OAc = 18/76/6 in O. sp., 45/50/5 in O. zaguliaevi and 70/6/24 in O. zealis). Differences in sex pheromones could contribute to the reproductive isolation between O. sp. and the other two Ostrinia species if males of each species exhibit a narrow window of response to their own blend ratio.     

Design, synthesis, and structure-activity relationship of new isobenzofuranone ligands of protein kinase C

Protein kinase C (PKC) is a family of enzymes, which play important roles in intracellular signal transduction. We have designed novel PKC ligands having an isobenzofuranone template, based on the proposed interaction of DAG (1,2-diacyl-sn-glycerol) with the PKCdelta C1B ligand-binding domain. Several isobenzofuranone derivatives were synthesized and their PKCalpha binding activities were evaluated. The pivaloyl derivative 1f was found to be a strong PKCalpha ligand, and the structure-activity relationship is well explained by our proposed binding model.     

Rice alpha-mannosidase digesting the high mannose glycopeptide of glutelin

α -Mannosidase (EC 3.2.1.24) from rice dry seeds was purified to homogeneity. Optimum pH and K_m for pNP- α -Man hydrolysis were pH 4.3–4.5 and 1.04 m M , respectively. The enzyme digested mannobioses such as Man α -1,2Man, Man α -1,6Man, Man α -1,3Man but Man α -1,4Man. Zn²⁺ ion was required for the activity, whereas EDTA and swainsonine inhibited the activity by 80 and 96%, respectively. The rice storage protein, glutelin was prepared and its basic subunits were shown to have high mannose-type sugar chains by two-dimensional mapping using NH₂-P and C₁₈ silica columns. They were Man₉GlcNAc₂, Man₈GlcNAc₂, Man₇GlcNAc₂, Man₆GlcNAc₂ and Man₅GlcNAc₂. All these oligosaccharides were digested by the purified α -mannosidase, and Man-GlcNAc₂ and mannose were formed. Glycopeptides, having these high mannose-type sugar chains, could also be digested by the α -mannosidase. Subunits were prepared from glutelin basic subunit and the richest subunit among them, subunit 2 (isoform 2), was digested by the α -mannosidase. Isoform 2 was digested by V8 protease only partially and slowly. However, isoform 2, pre-treated with the α -mannosidase, was rapidly and completely digested by V8 protease.     

Characteristics and distribution of endogenous RFamide-related peptide-1

We have recently identified RFamide-related peptide (RFRP) gene that would encode three peptides (i.e., RFRP-1, -2, and -3) in human and bovine, and demonstrated that synthetic RFRP-1 and -3 act as specific agonists for a G protein-coupled receptor OT7T022. However, molecular characteristics and tissue distribution of endogenous RFRPs have not been determined yet. In this study, we prepared a monoclonal antibody for the C-terminal portion of rat RFRP-1. As this antibody could recognize a consensus sequence among the C-terminal portions of rat, human, and bovine RFRP-1, we purified endogenous RFRP-1 from bovine hypothalamus on the basis of immunoreactivity to the antibody. The purified bovine endogenous RFRP-1 was found to have 35-amino-acid length that corresponds to 37-amino-acid length in human and rat. We subsequently constructed a sandwich enzyme immunoassay using the monoclonal antibody and a polyclonal antibody for the N-terminal portion of rat RFRP-1, and analyzed the tissue distribution of endogenous RFRP-1 in rats. Significant levels of RFRP-1 were detected only in the central nervous system, and the highest concentration of RFRP-1 was detected in the hypothalamus. RFRP-1-positive nerve cells were detected in the rat hypothalamus by immunohistochemical analyses using the monoclonal antibody. In culture, RFRP-1 lowered cAMP production in Chinese hamster ovary cells expressing OT7T022 and it was abolished by pre-treatment with pertussis toxin, suggesting that OT7T022 couples G(i)/G(o) in the signal transduction pathway.     

Triterpenoids from Viburnum suspensum

Three triterpenoids, 3-oxo-11,13(18)-oleanadien-28-oic acid, 24-hydroxy-3-oxo-11,13(18)-oleanadien-28-oic acid, 6β-hydroxy-3-oxo-11,13(18)-oleanadien-28-oic acid have been isolated together with the previously known virgatic acid, vibsanin B and 3-hydroxyvibsanin E from the leaves of Viburnum suspensum. Their structures were determined by spectroscopic methods and by comparison of their NMR spectral data with those of the previously known 11,13(18)-oleanadien-3β-ol.     

Functional relation among RecQ family helicases RecQL1, RecQL5, and BLM in cell growth and sister chromatid exchange formation

Human RECQL1 and RECQL5 belong to the RecQ family that includes Bloom syndrome, Werner syndrome, and Rothmund-Thomson syndrome causative genes. Cells derived from individuals suffering from these syndromes show significant levels of genomic instability. However, neither RECQL1 nor RECQL5 has been related to a disease, and nothing is known about the functions of RecQL1 and RecQL5. We generated here RECQL1(-/-), RECQL5(-/-), RECQL1(-/-)/RECQL5(-/-), RECQL1(-/-)/BLM(-/-), and RECQL5(-/-)/BLM(-/-) cells from chicken B-lymphocyte line DT40 cells. Although BLM(-/-) DT40 cells showed a slow-growth phenotype, a higher sensitivity to methyl methanesulfonate than the wild type, and an approximately 10-fold increase in the frequency of sister chromatid exchange (SCE) compared to wild-type cells, RECQL1(-/-), RECQL5(-/-), and RECQL1(-/-)/RECQL5(-/-) cells showed no significant difference from the wild-type cells in growth, sensitivity to DNA-damaging agents, and the frequency of SCE. However, both RECQL1(-/-)/BLM(-/-) and RECQL5(-/-)/BLM(-/-) cells grew more slowly than BLM(-/-) cells because of the increase in the population of dead cells, indicating that RecQL1 and RecQL5 are somehow involved in cell viability under the BLM function-impaired condition. Surprisingly, RECQL5(-/-)/BLM(-/-) cells showed a higher frequency of SCE than BLM(-/-) cells, indicating that RecQL5 suppresses SCE under the BLM function-impaired condition.