Anti-androgens with full antagonistic activity toward human prostate tumor LNCaP cells with mutated androgen receptor

Anti-androgens were designed based on the principle of inhibiting the folding of helix 12 of the nuclear androgen receptor. The prepared anti-androgens exhibited full antagonistic activity toward human prostate tumor LNCaP cells with T877A point-mutated nuclear androgen receptor, as far as examined, towards which other known anti-androgens, including hydroxyflutamide, are inactive or act as androgen agonists.     

Contribution of periodontal pathogens on tongue dorsa analyzed with real-time PCR to oral malodor

Oral malodor is considered to originate primarily from tongue microbiota populations. However, the relationship between oral malodor and tongue microbiota remains unclear. In this study, tongue periodontal pathogens were analyzed via real-time PCR, and the association between oral malodor and tongue periodontal pathogens, including Porphyromonas gingivalis, Tannerella forsythia, Prevotella intermedia, Prevotella nigrescens and Treponema denticola, was examined. The subject population consisted of 29 individuals with and 10 healthy persons without oral malodor. Oral malodor was assessed by organoleptic test and volatile sulfur compound (VSC) levels as measured by gas chromatography. Real-time PCR was conducted for anaerobes in tongue biofilm samples employing a LightCycler system; furthermore, bacterial proportion served as a quantitative parameter. Among the five anaerobes, only T. forsythia displayed higher proportions in malodor subjects than corresponding values in healthy controls. Proportions of P. intermedia and P. nigrescens correlated strongly with hydrogen sulfide concentration. Proportions of P. gingivalis and P. nigrescens also exhibited strong correlation with methyl mercaptan concentration. The correlation coefficient between the proportion of the total of the five anaerobes and total VSC level (r = 0.88) was greater than that between bacterial proportion and organoleptic score (r = 0.29). When a linear regression analysis was performed utilizing the proportion of each of the five periodontal pathogens as an independent variable, the explanatory power of these independent variables revealed 81% for total VSC level and 16% for organoleptic score. These results suggest that these five periodontal pathogens on tongue dorsa may contribute greatly to VSC production.     

Structural features of the glycogen branching enzyme encoding genes from aspergilli

A maltose binding protein, p78, was purified to homogeneity from Aspergillus nidulans by a single column chromatography step on cross-linked amylose. The partial amino acid sequence was highly homologous to the glycogen branching enzymes (GBEs) of human and yeast, and p78 did show branching enzyme activity. The genomic gene and its cDNA encoding GBE (p78) were isolated from the A. nidulans genomic and cDNA libraries. Furthermore, a cDNA encoding A. oryzae GBE was entirely sequenced. A. nidulans GBE shared overall and significant amino acid sequence identity with GBEs from A. oryzae (83.9%), Saccharomyces cerevisiae (61.1%) and human (63.0%), and with starch branching enzymes from green plants (55–56%).     

Quality Control of Photosystem II: Lipid Peroxidation Accelerates Photoinhibition under Excessive Illumination

Environmental stresses lower the efficiency of photosynthesis and sometimes cause irreversible damage to plant functions. When spinach thylakoids and Photosystem II membranes were illuminated with excessive visible light (100–1,000 µmol photons m⁻¹ s⁻¹) for 10 min at either 20°C or 30°C, the optimum quantum yield of Photosystem II decreased as the light intensity and temperature increased. Reactive oxygen species and endogenous cationic radicals produced through a photochemical reaction at and/or near the reaction center have been implicated in the damage to the D1 protein. Here we present evidence that lipid peroxidation induced by the illumination is involved in the damage to the D1 protein and the subunits of the light-harvesting complex of Photosystem II. This is reasoned from the results that considerable lipid peroxidation occurred in the thylakoids in the light, and that lipoxygenase externally added in the dark induced inhibition of Photosystem II activity in the thylakoids, production of singlet oxygen, which was monitored by electron paramagnetic resonance spin trapping, and damage to the D1 protein, in parallel with lipid peroxidation. Modification of the subunits of the light-harvesting complex of Photosystem II by malondialdehyde as well as oxidation of the subunits was also observed. We suggest that mainly singlet oxygen formed through lipid peroxidation under light stress participates in damaging the Photosystem II subunits.     

Activation of the Cpx phosphorelay signal transduction system in acidic phospholipid-deficient pgsA mutant cells of Escherichia coli

Low-intensity pulsed ultrasound (LIPUS) has been used as a safe and effective modality to enhance fracture healing. As the most abundant cells in bone, osteocytes orchestrate biological activities of effector cells via direct cell-to-cell contacts and by soluble factors. In this study, we have used the osteocytic MLO-Y4 cells to study the effects of conditioned medium from LIPUS-stimulated MLO-Y4 cells on proliferation and differentiation of osteoblastic MC3T3-E1 cells. Conditioned media from LIPUS-stimulated MLO-Y4 cells (LIPUS-Osteocyte-CM) were collected and added on MC3T3-E1 cell cultures. MC3T3-E1 cells cultured in LIPUS-Osteocyte-CM demonstrated a significant inhibition of proliferation and an increased alkaline phosphatase activity. The results of PGE(2) and NO assay showed that LIPUS could enhance PGE(2) and NO secretion from MLO-Y4 cells at all time points within 24h after LIPUS stimulation. We conclude that LIPUS regulates proliferation and differentiation of osteoblasts through osteocytes in vitro. Increased secretion of PGE(2) from osteocytes may play a role in this effect.     

Efficient decoding of the UAG triplet as a full-fledged sense codon enhances the growth of a prfA-deficient strain of Escherichia coli

We previously reassigned the amber UAG stop triplet as a sense codon in Escherichia coli by expressing a UAG-decoding tRNA and knocking out the prfA gene, encoding release factor 1. UAG triplets were left at the ends of about 300 genes in the genome. In the present study, we showed that the detrimental effect of UAG reassignment could be alleviated by increasing the efficiency of UAG translation instead of reducing the number of UAGs in the genome. We isolated an amber suppressor tRNA(Gln) variant displaying enhanced suppression activity, and we introduced it into the prfA knockout strain, RFzero-q, in place of the original suppressor tRNA(Gln). The resulting strain, RFzero-q3, translated UAG to glutamine almost as efficiently as the glutamine codons, and it proliferated faster than the parent RFzero-q strain. We identified two major factors in this growth enhancement. First, the sucB gene, which is involved in energy regeneration and has two successive UAG triplets at the end, was expressed at a higher level in RFzero-q3 than RFzero-q. Second, the ribosome stalling that occurred at UAG in RFzero-q was resolved in RFzero-q3. The results revealed the importance of "backup" stop triplets, UAA or UGA downstream of UAG, to avoid the deleterious impact of UAG reassignment on the proteome.     

Role of epithelial-stem cell interactions during dental cell differentiation

Epithelial-mesenchymal interactions regulate the growth and morphogenesis of ectodermal organs such as teeth. Dental pulp stem cells (DPSCs) are a part of dental mesenchyme, derived from the cranial neural crest, and differentiate into dentin forming odontoblasts. However, the interactions between DPSCs and epithelium have not been clearly elucidated. In this study, we established a mouse dental pulp stem cell line (SP) comprised of enriched side population cells that displayed a multipotent capacity to differentiate into odontogenic, osteogenic, adipogenic, and neurogenic cells. We also analyzed the interactions between SP cells and cells from the rat dental epithelial SF2 line. When cultured with SF2 cells, SP cells differentiated into odontoblasts that expressed dentin sialophosphoprotein. This differentiation was regulated by BMP2 and BMP4, and inhibited by the BMP antagonist Noggin. We also found that mouse iPS cells cultured with mitomycin C-treated SF2-24 cells displayed an epithelial cell-like morphology. Those cells expressed the epithelial cell markers p63 and cytokeratin-14, and the ameloblast markers ameloblastin and enamelin, whereas they did not express the endodermal cell marker Gata6 or mesodermal cell marker brachyury. This is the first report of differentiation of iPS cells into ameloblasts via interactions with dental epithelium. Co-culturing with dental epithelial cells appears to induce stem cell differentiation that favors an odontogenic cell fate, which may be a useful approach for tooth bioengineering strategies.     

Mucosal T cells expressing high levels of IL-7 receptor are potential targets for treatment of chronic colitis

The IL-7/IL-7R-dependent signaling pathway plays a crucial role in regulating the immune response in intestinal mucosa. Here we demonstrate the pivotal role of this pathway in the development and treatment of chronic colitis. T cells expressing high levels of IL-7R were substantially infiltrated in the chronic inflamed mucosa of TCR alpha-chain knockout mice and IL-7 transgenic mice. Transfer of mucosal T cells expressing high levels of IL-7R, but not T cells expressing low levels of IL-7R, from these mice into recombinase-activating gene-2(-/-) mice induced chronic colitis. Selective elimination of T cells expressing high levels of IL-7R by administrating small amounts of toxin-conjugated anti-IL-7R Ab completely ameliorated established, ongoing colitis. These findings provide evidence that therapeutic approaches targeting mucosal T cells expressing high levels of IL-7R are effective in the treatment of chronic intestinal inflammation and may be feasible for use in the therapy of human inflammatory bowel disease.