The spectroscopic analysis for binding of amphipathic and antimicrobial model peptides containing pyrenylalanine and tryptophan to lipid bilayer

The binding of basic amphipathic fluorescent peptides to lipid bilayers was studied in relation to their antimicrobial activity. Four fluorescent peptides containing pyrenylalanine or tryptophan in an amphipathic basic peptide (4(4] consisting of four repeated units of tetrapeptide, -L-Leu-L-Ala-L-Arg-L-Leu-, were found to have antimicrobial activities against Gram-positive bacteria and to take conformations with fairly high alpha-helical content both in aqueous solutions and liposomes. The fluorescence spectroscopic data suggested that the pyrenylalanine-peptide existed as a monomer in methanol or liposomes but as an oligomer in aqueous solutions to form an excimer between pyrenylalanyl residues. Upon binding with liposomes, the fluorescence spectra of the tryptophan-containing peptide shifted to a shorter wavelength, indicating the change in the state of tryptophan from hydrophilic environment to hydrophobic one. The analytical data for the quenching of tryptophan fluorescence by I- anion suggest that the tryptophan residue in the peptide is not deeply buried in the hydrophobic core of the bilayers. Based on these findings, it is suggested that the peptides may interact with liposomes in such a manner that they lie parallel to the surface of the lipid bilayers with their hydrophobic regions shallowly in the amphipathic moiety of the bilayers.     

Cyclic peptides. VI. Asymmetric hydrogenation of dehydroalanine or dehydroaminobutanoic acid residue in cyclodipeptides.

Several cyclo(-L-aminoacyl-deltaAla-) (aminoacyl = Ala, Val, Leu, Phe, Pro and Lys (epsilon-Ac)) were prepared by tosylation and successive detosylation of cyclo(-L-aminoacyl-L-Ser-), which were synthesized via the Nitecki and Fischer methods. Hydrogenation of the double bond of dehydroalanine residues in cyclodipeptides was carried out using Pd black in methanol at 1-atm pressure and room temperature. The degree of asymmetric hydrogenation was assessed by determining the amounts of L- and D-alanine by a modified Manning and Moore procedure. When L-valine was used as a chiral source, L-alanine residue with chiral induction of 98.4% was derived from cyclo(-L-Val-deltaAla-). L-Amino acids other than L-proline also were effective in inducing remarkable asymmetric hydrogenation. Hydrogenation of alpha,beta-dehydro-alpha-aminobutanoic acid residues in cyclodipeptides produced L-alpha-aminobutanoic acid residues with effective chiral induction to the same extent as observed with dehydroalanine residues. Optically pure l-alanine was prepared from cyclo(-L-Lys(epsilon-Ac)-deltaAla-) via asymmetric hydrogenation. A mechanism of chiral induction is discussed.     

Regulated genes in mesenchymal stem cells and gastric cancer

AIM: To investigate the genes regulated in mesenchymal stem cells(MSCs) and diffuse-type gastric cancer(GC),gene expression was analyzed. METHODS: Gene expression of MSCs and diffuse-type GC cells were analyzed by microarray. Genes related to stem cells, cancer and the epithelial-mesenchymal transition(EMT) were extracted from human gene lists using Gene Ontology and reference information. Gene panels were generated, and messenger RNA gene expression in MSCs and diffuse-type GC cells was analyzed. Cluster analysis was performed using the NCSS software.RESULTS: The gene expression of regulator of G-protein signaling 1(RGS1) was up-regulated in diffuse-type GC cells compared with MSCs. A panel of stem-cell related genes and genes involved in cancer or the EMT were examined. Stem-cell related genes, such as growth arrest-specific 6, musashi RNA-binding protein 2 and hairy and enhancer of split 1(Drosophila), NOTCH family genes and Notch ligands, such as delta-like 1(Drosophila) and Jagged 2, were regulated.CONCLUSION: Expression of RGS1 is up-regulated, and genes related to stem cells and NOTCH signaling are altered in diffuse-type GC compared with MSCs.     

Peripheral Opioid Antagonist Enhances the Effect of Anti-Tumor Drug by Blocking a Cell Growth-Suppressive Pathway In Vivo

The dormancy of tumor cells is a major problem in chemotherapy, since it limits the therapeutic efficacy of anti-tumor drugs that only target dividing cells. One potential way to overcome chemo-resistance is to “wake up” these dormant cells. Here we show that the opioid antagonist methylnaltrexone (MNTX) enhances the effect of docetaxel (Doc) by blocking a cell growth-suppressive pathway. We found that PENK, which encodes opioid growth factor (OGF) and suppresses cell growth, is predominantly expressed in diffuse-type gastric cancers (GCs). The blockade of OGF signaling by MNTX releases cells from their arrest and boosts the effect of Doc. In comparison with the use of Doc alone, the combined use of Doc and MNTX significantly prolongs survival, alleviates abdominal pain, and diminishes Doc-resistant spheroids on the peritoneal membrane in model mice. These results suggest that blockade of the pathways that suppress cell growth may enhance the effects of anti-tumor drugs.     

Identification and Characterization of CXCR4-Positive Gastric Cancer Stem Cells

Diffuse-type solid tumors are often composed of a high proportion of rarely proliferating (i.e., dormant) cancer cells, strongly indicating the involvement of cancer stem cells (CSCs) Although diffuse-type gastric cancer (GC) patients have a poor prognosis due to high-frequent development of peritoneal dissemination (PD), it is limited knowledge that the PD-associated CSCs and efficacy of CSC-targeting therapy in diffuse-type GC. In this study, we established highly metastatic GC cell lines by in vivo selection designed for the enrichment of PD-associated GC cells. By microarray analysis, we found C-X-C chemokine receptor type 4 (CXCR4) can be a novel marker for highly metastatic CSCs, since CXCR4-positive cells can grow anchorage-independently, initiate tumors in mice, be resistant to cytotoxic drug, and produce differentiated daughter cells. In clinical samples, these CXCR4-positive cells were found from not only late metastasis stage (accumulated ascites) but also earlier stage (peritoneal washings). Moreover, treatment with transforming growth factor-β enhanced the anti-cancer effect of docetaxel via induction of cell differentiation/asymmetric cell division of the CXCR4-positive gastric CSCs even in a dormant state. Therefore, differentiation inducers hold promise for obtaining the maximum therapeutic outcome from currently available anti-cancer drugs through re-cycling of CSCs.     

Development and Evaluation of a Parenting Resilience Elements Questionnaire (PREQ) Measuring Resiliency in Rearing Children with Developmental Disorders

We developed a parenting resilience elements questionnaire (PREQ) measuring the degree to which mothers possess elements that aid in adapting to challenges and difficulties related to children with developmental disorders (DD). A total of 424 parents of children with DD were recruited from five medical institutes. Psychometric properties of PREQ were evaluated using data of 363 mothers of children with DD. Furthermore, multiple regression analysis was performed, predicting depressive symptoms and parenting behavior with PREQ subscales, a general health questionnaire, and the total difficulties score of a strength and difficulties questionnaire. Factor analysis revealed three reliable factors: “knowledge of the child’s characteristics,” “perceived social supports,” and “positive perceptions of parenting.” Moreover, multiple regression analysis showed that “knowledge of the child’s characteristics” was associated with parenting behavior, whereas “perceived social supports” predicted depressive symptoms; “positive perceptions of parenting” influenced both parenting behavior and depressive symptoms. These findings indicated that the PREQ may be used as a scale measuring resiliency in mothers of children with DD and is useful for evaluating their parenting ability in clinical interventions.     

Importance of mitochondrial dysfunction in oxidative stress response: A comparative study of gene expression profiles

Mitochondria are considered to play an important role in oxidative stress response since they are a source of reactive oxygen species and are also targeted by these species. This study examined the mitochondrial conditions in cells of epithelial origin that were exposed to H(2)O(2) and found a decline in the membrane potential along with a specific loss of UQCRC1, a sub-unit of complex III, suggesting that mitochondrial dysfunction occurs upon exposure to oxidative stress. This observation led to the hypothesis that certain cellular responses to oxidative stress occurred because of mitochondrial dysfunction. When mitochondria-less (pseudo ρ0) cells were examined as a model of mitochondrial dysfunction, striking similarities were found in their cellular responses compared with those found in cells exposed to oxidative stress, including changes in gene expression and gelatinolytic enzyme activities, thus suggesting that cellular responses to oxidative stress were partly mediated by mitochondrial dysfunction. This possibility was further validated by microarray analysis, which suggested that almost one-fourth of the cellular responses to oxidative stress were mediated by mitochondrial dysfunction that accompanies oxidative stress, thereby warranting a therapeutic strategy that targets mitochondria for the treatment of oxidative stress-associated diseases.