Distinct photo-oxidation-induced cell death pathways lead to selective killing of human breast cancer cells

Lack of effective treatments for aggressive breast cancer is still a major global health problem. We have previously reported that photodynamic therapy using methylene blue as photosensitizer (MB-PDT) massively kills metastatic human breast cancer, marginally affecting healthy cells. In this study, we aimed to unveil the molecular mechanisms behind MB-PDT effectiveness and specificity towards tumor cells. Through lipidomics and biochemical approaches, we demonstrated that MB-PDT efficiency and specificity rely on polyunsaturated fatty acid-enriched membranes and on the better capacity to deal with photo-oxidative damage displayed by non-tumorigenic cells. We found out that, in tumorigenic cells, lysosome membrane permeabilization is accompanied by ferroptosis and/or necroptosis. Our results also pointed at a cross-talk between lysosome-dependent cell death (LDCD) and necroptosis induction after photo-oxidation, and contributed to broaden the understanding of MB-PDT-induced mechanisms and specificity in breast cancer cells. Therefore, we demonstrated that efficient approaches could be designed on the basis of lipid composition and metabolic features for hard-to-treat cancers. The results further reinforce MB-PDT as a therapeutic strategy for highly aggressive human breast cancer cells.Subject terms: Lipidomics, Necroptosis, Stress signalling, Breast cancer     

FLOWERING LOCUS T regulates stomatal opening

Stomatal pores surrounded by a pair of guard cells in the plant epidermis control gas exchange for photosynthesis in response to light, CO(2), and phytohormone abscisic acid. Phototropins (phot1 and phot2) are plant blue-light receptor kinases and mediate stomatal opening via activation of the plasma membrane H(+)-ATPase. However, the signaling mechanism from phototropins to the H(+)-ATPase has yet to be determined. Here, we show that FLOWERING LOCUS T (FT) is expressed in guard cells and regulates stomatal opening. We isolated an scs (suppressor of closed-stomata phenotype in phot1 phot2) 1-1 mutant of Arabidopsis thaliana and showed that scs1-1 carries a novel null early flowering 3 (elf3) allele in a phot1 phot2 background. scs1-1 (elf3 phot1 phot2 triple mutant) had an open-stomata phenotype with high H(+)-ATPase activity and showed increased levels of FT mRNA in guard cells. Transgenic plants overexpressing FT in guard cells showed open stomata, whereas a loss-of-function FT allele, ft-1, exhibited closed stomata and failed to activate the H(+)-ATPase in response to blue light. Our results define a new cell-autonomous role for FT and demonstrate that the flowering time genes ELF3 and FT are involved in the regulation of H(+)-ATPase by blue light in guard cells.     

Simple Resolution of Enantiomeric NMR Signals of α‐Amino Acids by Using Samarium(III) Nitrate With L‐Tartarate

Readily available L‐tartaric acid, which is a bidentate ligand with two chiral centers forming a seven‐membered chelate ring, was applied to the chiral ligand for the chiral nuclear magnetic resonance (NMR) shift reagent of samarium(III) formed in situ. This simple method does not cause serious signal broadening in the high magnetic field. Enantiomeric ¹³C and ¹H NMR signals and enantiotopic ¹H NMR signals of α‐amino acids were successfully resolved at pH 8.0 and the 1:3 molar ratio of Sm(NO₃)₃:L‐tartaric acid. It is elucidated that the enantiomeric signal resolution is attributed to the anisotropic magnetic environment for the enantiomers induced by the chiral L‐tartarato samarium(III) complex rather than differences in stability of the diastereomeric substrate adducts. The present ¹³C NMR signal resolution was also effective for the practical simultaneous analysis of plural kinds of DL‐amino acids. Chirality 27:353–357, 2015.© 2015 Wiley Periodicals, Inc.     

Age-related decrease in cold-activated brown adipose tissue and accumulation of body fat in healthy humans

Brown adipose tissue (BAT) can be identified by (18)F-fluorodeoxyglucose (FDG)-positron emission tomography (PET) combined with X-ray computed tomography (CT) in adult humans. The objective of this study was to clarify the relationship between BAT and adiposity in healthy adult humans, particularly to test the idea that decreased BAT activity may be associated with body fat accumulation with age. One hundred and sixty-two healthy volunteers aged 20-73 years (103 males and 59 females) underwent FDG-PET/CT after 2-h cold exposure at 19 °C with light clothing. Cold-activated BAT was detected in 41% of the subjects (BAT-positive). Compared with the BAT-negative group, the BAT-positive group was younger (P < 0.01) and showed a lower BMI (P < 0.01), body fat content (P < 0.01), and abdominal fat (P < 0.01). The incidence of cold-activated BAT decreased with age (P < 0.01), being more than 50% in the twenties, but less than 10% in the fifties and sixties. The adiposity-related parameters showed some sex differences, but increased with age in the BAT-negative group (P < 0.01), while they remained unchanged from the twenties to forties in the BAT-positive group, in both sexes. These results suggest that decreased BAT activity may be associated with accumulation of body fat with age.     

A non-mammalian type opsin 5 functions dually in the photoreceptive and non-photoreceptive organs of birds

A mammalian type opsin 5 (neuropsin) is a recently identified ultraviolet (UV)-sensitive pigment of the retina and other photosensitive organs in birds. Two other opsin 5-related molecules have been found in the genomes of non-mammalian vertebrates. However, their functions have not been examined as yet. Here, we identify the molecular properties of a second avian opsin 5, cOpn5L2 (chicken opsin 5-like 2), and its localization in the post-hatch chicken. Spectrophotometric analysis and radionucleotide-binding assay have revealed that cOpn5L2 is a UV-sensitive bistable pigment that couples with the Gi subtype of guanine nucleotide-binding protein (G protein). As a bistable pigment, it also shows the direct binding ability to agonist all-trans-retinal to activate G protein. The absorption maxima of UV-light-absorbing and visible light-absorbing forms were 350 and 521 nm, respectively. Expression analysis showed relatively high expression of cOpn5L2 mRNA in the adrenal gland, which is not photoreceptive but an endocrine organ, while lower expression was found in the brain and retina. At the protein level, cOpn5L2 immunoreactive cells were present in the chromaffin cells of the adrenal gland. In the brain, cOpn5L2 immunoreactive cells were found in the paraventricular and supraoptic nuclei of the anterior hypothalamus, known for photoreceptive deep brain areas. In the retina, cOpn5L2 protein was localized to subsets of cells in the ganglion cell layer and the inner nuclear layer. These results suggest that the non-mammalian type opsin 5 (Opn5L2) functions as a second UV sensor in the photoreceptive organs, while it might function as chemosensor using its direct binding ability to agonist all-trans-retinal in non-photoreceptive organs such as the adrenal gland of birds.     

Nucleosome exclusion from the interspecies-conserved central AT-rich region of the Ars insulator

The Ars insulator is a boundary element identified in the upstream region of the arylsulfatase (HpArs) gene in the sea urchin, Hemicentrotus pulcherrimus, and possesses the ability to both block enhancer-promoter communications and protect transgenes from silent chromatin. To understand the molecular mechanism of the Ars insulator, we investigated the correlation between chromatin structure, DNA structure and insulator activity. Nuclease digestion of nuclei isolated from sea urchin embryos revealed the presence of a nuclease-hypersensitive site within the Ars insulator. Analysis of micrococcal nuclease-sensitive sites in the Ars insulator, reconstituted with nucleosomes, showed the exclusion of nucleosomes from the central AT-rich region. Furthermore, the central AT-rich region in naked DNA was sensitive to nucleotide base modification by diethylpyrocarbonate (DEPC). These observations suggest that non-B-DNA structures in the central AT-rich region may inhibit nucleosomal formation, which leads to nuclease hypersensitivity. Furthermore, comparison of nucleotide sequences between the HpArs gene and its ortholog in Strongylocentrotus purpuratus revealed that the central AT-rich region of the Ars insulator is conserved, and this conserved region showed significant enhancer blocking activity. These results suggest that the central AT-rich nucleosome-free region plays an important role in the function of the Ars insulator.     

Dendritic Cells from Oral Cavity Induce Foxp3+ Regulatory T Cells upon Antigen Stimulation

Evidence is accumulating that dendritic cells (DCs) from the intestines have the capacity to induce Foxp3⁺CD4⁺ regulatory T cells (T-regs) and regulate immunity versus tolerance in the intestines. However, the contribution of DCs to controlling immunity versus tolerance in the oral cavity has not been addressed. Here, we report that DCs from the oral cavity induce Foxp3⁺ T-regs as well as DCs from intestine. We found that oral-cavity-draining cervical lymph nodes contained higher frequencies of Foxp3⁺ T-regs and ROR-γt⁺ CD4⁺T cells than other lymph nodes. The high frequency of Foxp3⁺ T-regs in the oral-cavity-draining cervical lymph nodes was not dependent on the Toll like receptor (TLR) adaptor molecules, Myd88 and TICAM-1 (TRIF). In contrast, the high frequency of ROR-γt⁺ CD4⁺T cells relies on Myd88 and TICAM-1. In vitro data showed that CD11c⁺ DCs from oral-cavity-draining cervical lymph nodes have the capacity to induce Foxp3⁺ T-regs in the presence of antigen. These data suggest that, as well as in the intestinal environment, antigen-presenting DCs may play a vital role in maintaining tolerance by inducing Foxp3⁺ T-regs in the oral cavity.     

Development and Validation of a New Questionnaire Assessing Quality of Life in Adults with Hypopituitarism: Adult Hypopituitarism Questionnaire (AHQ)

Objective

To develop and validate the Adult Hypopituitarism Questionnaire (AHQ) as a disease-specific, self-administered questionnaire for evaluation of quality of life (QOL) in adult patients with hypopituitarism.

Methods

We developed and validated this new questionnaire, using a standardized procedure which included item development, pilot-testing and psychometric validation. Of the patients who participated in psychometric validation, those whose clinical conditions were judged to be stable were asked to answer the survey questionnaire twice, in order to assess test-retest reliability.

Results

Content validity of the initial questionnaire was evaluated via two pilot tests. After these tests, we made minor revisions and finalized the initial version of the questionnaire. The questionnaire was constructed with two domains, one psycho-social and the other physical. For psychometric assessment, analyses were performed on the responses of 192 adult patients with various types of hypopituitarism. The intraclass correlations of the respective domains were 0.91 and 0.95, and the Cronbach’s alpha coefficients were 0.96 and 0.95, indicating adequate test-retest reliability and internal consistency for each domain. For known-group validity, patients with hypopituitarism due to hypothalamic disorder showed significantly lower scores in 11 out of 13 sub-domains compared to those who had hypopituitarism due to pituitary disorder. Regarding construct validity, the domain structure was found to be almost the same as that initially hypothesized. Exploratory factor analysis (n = 228) demonstrated that each domain consisted of six and seven sub-domains.

Conclusion

The AHQ showed good reliability and validity for evaluating QOL in adult patients with hypopituitarism.     

Soy peptides enhance heterologous membrane protein productivity during the exponential growth phase of Saccharomyces cerevisiae

In this study, the production of eight G protein-coupled receptors by Saccharomyces cerevisiae was compared using two types of media, one of which contained soy peptides and the other free amino acids. Yeast cell growth improved in the medium with soy peptides, and the expression levels of six of the receptors increased during the exponential phase by an average of 2.3-fold as against the free amino acid-based medium. The enhancement of protein expression by soy peptides can be explained by alleviation of metabolite stress due to amino acid source depletion caused by heterologous protein expression.     

Validation of the Brazilian versions of two inventories for measuring oral health‐related quality of life of edentulous subjects

doi: 10.1111/j.1741‐2358.2010.00417.x
Validation of the Brazilian versions of two inventories for measuring oral health‐related quality of life of edentulous subjects Objectives: To analyse the validity of the Brazilian versions of OHIP‐EDENT and GOHAI as assessment tools of edentulous subjects’ OHRQoL. Background: Inventories for measuring oral health‐related quality of life (OHRQoL) are important in clinical studies regarding oral rehabilitation. However, there is a need for comprehensive validation after translation into different cultural settings. Materials and methods: The sample comprised of 100 complete denture wearers (29 men, 71 women, mean age of 65.2 ± 9.9 years). The associations between each OHRQoL inventory and other variables served as measurements of construct validity. Data analysis comprised the Spearman correlation test as well as multiple regression using the OHRQoL inventories as dependent variables and the other scales as determinants. Results: Both OHRQoL inventories showed good correlation with denture satisfaction, whereas lower correlation coefficients were found among the inventories and the HAD subscales. Denture satisfaction alone explained 48% and 39% of the variance found for the OHIP‐EDENT and GOHAI, respectively, as assessed by multiple regression. A smaller effect was found for OHIP‐EDENT. Conclusion: Both OHIP‐EDENT and GOHAI showed good construct validity for measurement of OHRQoL of edentulous subjects.