Oxidative Stress and Longevity in Okinawa: An Investigation of Blood Lipid Peroxidation and Tocopherol in Okinawan Centenarians

Background. The Free Radical Theory of Aging mechanistically links oxidative stress to aging. Okinawa has among the world's longest-lived populations but oxidative stress in this population has not been well characterized. Methods. We compared plasma lipid peroxide (LPO) and vitamin E—plasma and intracellular tocopherol levels (total α, β, and γ), in centenarians with younger controls. Results. Both LPO and vitamin E tocopherols were lower in centenarians, with the exception of intracellular β-tocopherol, which was significantly higher in centenarians versus younger controls. There were no significant differences between age groups for tocopherol: cholesterol and tocopherol: LPO ratios. Correlations were found between α-Tocopherol and LPO in septuagenarians but not in centenarians. Conclusions. The low plasma level of LPO in Okinawan centenarians, compared to younger controls, argues for protection against oxidative stress in the centenarian population and is consistent with the predictions of the Free Radical Theory of Aging. However, the present work does not strongly support a role for vitamin E in this phenomenon. The role of intracellular β-tocopherol deserves additional study. More research is needed on the contribution of oxidative stress and antioxidants to human longevity

A Recipe for Successful Collaboration: Shared Creative Work Experiences (SCrWE) Among Co-Researchers

In this article, we discuss our home cooking school as one example of a strategy we call "Shared Creative Work Experience" (SCrWE, pronounced "screwy"): Planned, playful extra-research activity during which collaborators engage in and reflect on creative work (e.g., cooking, sewing, painting, building, writing, performing, designing, gardening) that yields a product of some sort (e.g., a meal, a quilt, a painting, a shelf, a poem, a play, a game, a communal garden). Through SCrWE, we argue, collaborators playfully but deliberately create disequilibrium, shift perspectives, and unsettle power dynamics, ultimately preparing for productive, meaningful research partnerships. By creating a space for co-researchers to experience shared creative work, we aim to disrupt taken-for-granted assumptions and invite co-researchers to embrace ambiguity together. Grounded conceptually in aesthetic experiential play and the notion of the social imagination, SCrWE helps research teams identify potential sources of substantive, procedural, and affective conflict and then explore these conflicts in productive ways. Using techniques of collaborative autoethnography, we weave together recipes, photos, and scholarly writing to illuminate our experiences. We conclude by describing the steps for developing a SCrWE and include reflective questions to help research team members uncover their ontological, epistemological, and axiological commitments, ultimately leading to more meaningful research partnerships.In diesem Artikel stellen wir unsere Kochschule als Beispiel für eine Strategie vor, die wir "Shared Creative Work Experience" (SCrWE, ausgesprochen "screwy") nennen: geplante, spielerische Aktivitäten außerhalb der Forschung, bei denen die Teilnehmer*innen kreative Arbeit (z.B. Kochen, Nähen, Malen, Bauen, Schreiben, Darstellen, Gestalten, Gärtnern) verrichten und darüber reflektieren, die zu einem Produkt führt (z.B. einer Mahlzeit, einem Quilt, einem Gemälde, einem Regal, einem Gedicht, einem Theaterstück, einem Spiel, einem Gemeinschaftsgarten). Durch SCrWE schaffen die Beteiligten spielerisch, aber absichtlich ein Ungleichgewicht, verschieben die Perspektiven und bringen die Machtdynamik ins Wanken, was letztlich eine produktive, sinnvolle Forschungspartnerschaft vorbereitet. Mittels eines Raums, in dem Forscher*innen gemeinsam kreativ arbeiten können, wollen wir gewohnte Annahmen durchbrechen und dazu einladen, sich gemeinsam auf Mehrdeutigkeit einzulassen. SCrWE basiert konzeptionell auf dem ästhetischen Erfahrungsspiel und dem Konzept der sozialen Vorstellungskraft und hilft Forschungsteams dabei, potenzielle Quellen für inhaltliche, prozessuale und affektive Konflikte zu identifizieren und diese dann auf produktive Weise zu erkunden. Mithilfe der Techniken der kollaborativen Autoethnografie verweben wir Rezepte, Fotos und wissenschaftliche Texte, um unsere Erfahrungen zu beleuchten. Abschließend beschreiben wir die Schritte zur Entwicklung eines SCrWE und stellen Reflexionsfragen, die den Mitgliedern von Forschungsteams helfen sollen, ihre ontologischen, epistemologischen und axiologischen Verpflichtungen zu erkennen, was letztlich zu sinnvollerem Zusammenarbeiten führt

Comparative mode of action of the antimicrobial peptide melimine and its derivative Mel4 against Pseudomonas aeruginosa

Melimine and Mel4 are chimeric cationic peptides with broad-spectrum antimicrobial activity. They have been shown to be highly biocompatible in animal models and human clinical trials. The current study examined the mechanism of action of these two antimicrobial peptides against P. aeruginosa. The effect of the peptides of endotoxin neutralization, and their interactions with cytoplasmic membranes using DiSC(3)-5 and Sytox green, Syto-9 and PI dyes were analysed. Release of ATP and DNA/RNA were determined using ATP luminescence and increase in OD 260 nm. The bacteriolytic ability of the peptides was determined by measuring decreases in OD 620 nm. Both the peptides neutralized LPS suggesting their interaction with lipid A. Cytoplasmic membrane was disrupted within 30 seconds, which correlated with reductions in cellular viability. At 2 minutes melimine or Mel4, released 75% and 36% cellular ATP respectively (P < 0.001). Membrane permeabilization started 5 minutes with simultaneous release of DNA/RNA. Flow cytometry demonstrated 52% and 18% bacteria were stained with PI after 30 minutes. Overall, melimine showed higher capacity for membrane disruption compared to Mel4 (P < 0.001). The findings of this study have been summarized as a timeline of bactericidal activity, suggesting that the peptides permeabilized P. aeruginosa within 5 minutes, started lysis within 2 hours of exposure

Transcription of telomeric DNA leads to high levels of homologous recombination and t-loops

The formation of DNA loops at chromosome ends (t-loops) and the transcription of telomeres producing G-rich RNA (TERRA) represent two central features of telomeres. To explore a possible link between them we employed artificial human telomeres containing long arrays of TTAGGG repeats flanked by the T7 or T3 promoters. Transcription of these DNAs generates a high frequency of t-loops within individual molecules and homologous recombination events between different DNAs at their telomeric sequences. T-loop formation does not require a single strand overhang, arguing that both terminal strands insert into the preceding duplex. The loops are very stable and some RNase H resistant TERRA remains at the t-loop, likely adding to their stability. Transcription of DNAs containing TTAGTG or TGAGTG repeats showed greatly reduced loop formation. While in the cell multiple pathways may lead to t-loop formation, the pathway revealed here does not depend on the shelterins but rather on the unique character of telomeric DNA when it is opened for transcription. Hence, telomeric sequences may have evolved to facilitate their ability to loop back on themselves

Extending healthy ageing: nutrient sensitive pathway and centenarian population

Ageing is a challenge for any living organism and human longevity is a complex phenotype. With increasing life expectancy, maintaining long-term health, functionality and well-being during ageing has become an essential goal. To increase our understanding of how ageing works, it may be advantageous to analyze the phenotype of centenarians, perhaps one of the best examples of successful ageing. Healthy ageing involves the interaction between genes, the environment, and lifestyle factors, particularly diet. Besides evaluating specific gene-environment interactions in relation to exceptional longevity, it is important to focus attention on modifiable lifestyle factors such as diet and nutrition to achieve extension of health span. Furthermore, a better understanding of human longevity may assist in the design of strategies to extend the duration of optimal human health. In this article we briefly discuss relevant topics on ageing and longevity with particular focus on dietary patterns of centenarians and nutrient-sensing pathways that have a pivotal role in the regulation of life span. Finally, we also discuss the potential role of Nrf2 system in the pro-ageing signaling emphasizing its phytohormetic activation

Mode of action of the antimicrobial peptide Mel4 is independent of Staphylococcus aureus cell membrane permeability

Mel4 is a novel cationic peptide with potent activity against Gram-positive bacteria. The current study examined the anti-staphylococcal mechanism of action of Mel4 and its precursor peptide melimine. The interaction of peptides with lipoteichoic acid (LTA) and with the cytoplasmic membrane using DiSC(3)-5, Sytox green, Syto-9 and PI dyes were studied. Release of ATP and DNA/RNA from cells exposed to the peptides were determined. Bacteriolysis and autolysin-activated cell death were determined by measuring decreases in OD620nm and killing of Micrococcus lysodeikticus cells by cell-free media. Both peptides bound to LTA and rapidly dissipated the membrane potential (within 30 seconds) without affecting bacterial viability. Disturbance of the membrane potential was followed by the release of ATP (50% of total cellular ATP) by melimine and by Mel4 (20%) after 2 minutes exposure (p<0.001). Mel4 resulted in staphylococcal cells taking up PI with 3.9% cells predominantly stained after 150 min exposure, whereas melimine showed 34% staining. Unlike melimine, Mel4 did not release DNA/RNA. Cell-free media from Mel4 treated cells hydrolysed peptidoglycan and produced greater zones of inhibition against M. lysodeikticus lawn than melimine treated samples. These findings suggest that pore formation is unlikely to be involved in Mel4-mediated membrane destabilization for staphylococci, since there was no significant Mel4-induced PI staining and DNA/RNA leakage. It is likely that the S. aureus killing mechanism of Mel4 involves the release of autolysins followed by cell death. Whereas, membrane interaction is the primary bactericidal activity of melimine, which includes membrane depolarization, pore formation, release of cellular contents leading to cell death

In Vitro and In Vivo Evaluation of Novel Ciprofloxacin-Releasing Silicone Hydrogel Contact Lenses

Hui, A., Willcox, M., & Jones, L. (2014). In Vitro and In Vivo Evaluation of Novel Ciprofloxacin-Releasing Silicone Hydrogel Contact Lenses. Investigative Opthalmology & Visual Science, 55(8), 4896. https://doi.org/10.1167/iovs.14-14855Purpose.: The purpose of this study was to evaluate ciprofloxacin-releasing silicone hydrogel contact lens materials in vitro and in vivo for the treatment of microbial keratitis. Methods.: Model silicone hydrogel contact lens materials were manufactured using a molecular imprinting technique to modify ciprofloxacin release kinetics. Various contact lens properties, including light transmission and surface wettability, were determined, and the in vitro ciprofloxacin release kinetics elucidated using fluorescence spectrophotometry. The materials then were evaluated for their ability to inhibit Pseudomonas aeruginosa growth in vitro and in an in vivo rabbit model of microbial keratitis. Results.: Synthesized lenses had similar material properties to commercial contact lens materials. There was a decrease in light transmission in the shorter wavelengths due to incorporation of the antibiotic, but over 80% light transmission between 400 and 700 nm. Modified materials released for more than 8 hours, significantly longer than unmodified controls (P 0.05), which is significantly less than corneas treated with unmodified control lenses or those that received no treatment at all (P < 0.05). Conclusions.: These novel contact lenses designed for the extended release of ciprofloxacin may be beneficial to supplement or augment future treatments of sight-threatening microbial keratitis.Supported by the Natural Science and Engineering Research Council of Canada (NSERC)20/20 Network for the Development of Advanced Ophthalmic Materialsand by an NSERC Alexander Graham Bell Doctoral Scholarshipthe Ezell Fellowship from the American Optometric Foundationand the Endeavour Research Grant from the Australian Government (AH)

Association between possession of ExoU and antibiotic resistance in pseudomonas aeruginosa

© 2018 Subedi et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Virulent strains of Pseudomonas aeruginosa are often associated with an acquired cytotoxic protein, exoenzyme U (ExoU) that rapidly destroys the cell membranes of host cells by its phospholipase activity. Strains possessing the exoU gene are predominant in eye infections and are more resistant to antibiotics. Thus, it is essential to understand treatment options for these strains. Here, we have investigated the resistance profiles and genes associated with resistance for fluoroquinolone and beta-lactams. A total of 22 strains of P. aeruginosa from anterior eye infections, microbial keratitis (MK), and the lungs of cystic fibrosis (CF) patients were used. Based on whole genome sequencing, the prevalence of the exoU gene was 61.5% in MK isolates whereas none of the CF isolates possessed this gene. Overall, higher antibiotic resistance was observed in the isolates possessing exoU. Of the exoU strains, all except one were resistant to fluoroquinolones, 100% were resistant to beta-lactams. 75% had mutations in quinolone resistance determining regions (T81I gyrA and/or S87L parC) which correlated with fluoroquinolone resistance. In addition, exoU strains had mutations at K76Q, A110T, and V126E in ampC, Q155I and V356I in ampR and E114A, G283E, and M288R in mexR genes that are associated with higher beta-lactamase and efflux pump activities. In contrast, such mutations were not observed in the strains lacking exoU. The expression of the ampC gene increased by up to nine-fold in all eight exoU strains and the ampR was upregulated in seven exoU strains compared to PAO1. The expression of mexR gene was 1.4 to 3.6 fold lower in 75% of exoU strains. This study highlights the association between virulence traits and antibiotic resistance in pathogenic P. aeruginosa

Poly-ε-Lysine or Mel4 Antimicrobial Surface Modification on a Novel Peptide Hydrogel Bandage Contact Lens

Microbial keratitis (MK) is a serious issue in many countries and is often caused by contact lens wear. Antimicrobial peptides (AMPs) are a potentially useful tool for creating antimicrobial surfaces in light of increasing antibiotic resistance. Poly-ε-lysine (pεK) is an AMP that has been used extensively as a food preservative and Mel4 has recently been synthesized and studied as an antimicrobial coating for contact lenses. A hydrogel synthesized of pεK cross-linked with biscarboxylic acids provides a potential lens material which has many surface free amines, that can be subsequently used to attach additional AMPs, creating an antimicrobial lens. The aim of this study is to investigate pεK hydrogels against a clinical strain of Pseudomonas aeruginosa (P. aeruginosa) for preventing or treating MK. Covalent attachment of AMPs is investigated and confirmed by fluorescently tagged peptides. Bound pεK effectively reduces the number of adherent P. aeruginosa in vitro (>3 log). In ex vivo studies positive antimicrobial activity is observed on bare pεK hydrogels and those with additionally bound pεK or Mel4; lenses allow the maintenance of the corneal epithelium. A pεK hydrogel contact lens with additional AMPs can be a therapeutic tool to reduce the incidence of MK