Dosimetric uncertainties impact on cell survival curve with low energy proton.

There is an increasing number of radiobiological experiments being conducted with low energy protons (less than 5 MeV) for radiobiological studies due to availability of sub-millimetre focused beam. However, low energy proton has broad microdosimetric spectra which can introduce dosimetric uncertainty. In this work, we quantify the impact of this dosimetric uncertainties on the cell survival curve and how it affects the estimation of the alpha and beta parameters in the LQ formalism. Monte Carlo simulation is used to generate the microdosimetric spectra in a micrometer-sized water sphere under proton irradiation. This is modelled using radiobiological experiment set-up at the Centre of Ion Beam Application (CIBA) in National University of Singapore. Our results show that the microdosimetric spectra can introduce both systematic and random shifts in dose and cell survival; this effect is most pronounced with low energy protons. The alpha and beta uncertainties can be up to 10% and above 30%, respectively for low energy protons passing through thin cell target (about 10 microns). These uncertainties are non-negligible and show that care must be taken in using the cell survival curve and its derived parameters for radiobiological models.

Application of optogenetic Amyloid-ß distinguishes between metabolic and physical damages in neurodegeneration.

The brains of Alzheimer's disease patients show a decrease in brain mass and a preponderance of extracellular Amyloid-ß plaques. These plaques are formed by aggregation of polypeptides that are derived from the Amyloid Precursor Protein (APP). Amyloid-ß plaques are thought to play either a direct or an indirect role in disease progression, however the exact role of aggregation and plaque formation in the aetiology of Alzheimer's disease (AD) is subject to debate as the biological effects of soluble and aggregated Amyloid-ß peptides are difficult to separate in vivo. To investigate the consequences of formation of Amyloid-ß oligomers in living tissues, we developed a fluorescently tagged, optogenetic Amyloid-ß peptide that oligomerizes rapidly in the presence of blue light. We applied this system to the crucial question of how intracellular Amyloid-ß oligomers underlie the pathologies of A. We use Drosophila, C. elegans and D. rerio to show that, although both expression and induced oligomerization of Amyloid-ß were detrimental to lifespan and healthspan, we were able to separate the metabolic and physical damage caused by light-induced Amyloid-ß oligomerization from Amyloid-ß expression alone. The physical damage caused by Amyloid-ß oligomers also recapitulated the catastrophic tissue loss that is a hallmark of late AD. We show that the lifespan deficit induced by Amyloid-ß oligomers was reduced with Li+ treatment. Our results present the first model to separate different aspects of disease progression.

Alzheimer's disease is a progressive condition that damages the brain over time. The cause is not clear, but a toxic molecule called Amyloid-ß peptide seems to play a part. It builds up in the brains of people with Alzheimer's disease, forming hard clumps called plaques. Yet, though the plaques are a hallmark of the disease, experimental treatments designed to break them down do not seem to help. This raises the question ­ do Amyloid-ß plaques actually cause Alzheimer's disease? Answering this question is not easy. One way to study the effect of amyloid plaques is to inject clumps of Amyloid-ß peptides into model organisms. This triggers Alzheimer's-like brain damage, but it is not clear why. It remains difficult to tell the difference between the damage caused by the injected Amyloid-ß peptides and the damage caused by the solid plaques that they form. For this, researchers need a way to trigger plaque formation directly inside animal brains. This would make it possible to test the effects of plaque-targeting treatments, like the drug lithium. Optogenetics is a technique that uses light to control molecules in living animals. Hsien, Kaur et al. have now used this approach to trigger plaque formation by fusing light-sensitive proteins to Amyloid-ß peptides in worms, fruit flies and zebrafish. This meant that the peptides clumped together to form plaques whenever the animals were exposed to blue light. This revealed that, while both the Amyloid-ß peptides and the plaques caused damage, the plaques were much more toxic. They damaged cell metabolism and caused tissue loss that resembled late Alzheimer's disease in humans. To find out whether it was possible to test Alzheimer's treatments in these animals, Hsien, Kaur et al. treated them with the drug, lithium. This increased their lifespan, reversing some of the damage caused by the plaques. Alzheimer's disease affects more than 46.8 million people worldwide and is the sixth leading cause of death in the USA. But, despite over 50 years of research, there is no cure. This new plaque-formation technique allows researchers to study the effects of amyloid plaques in living animals, providing a new way to test Alzheimer's treatments. This could be of particular help in studies of experimental drugs that aim to reduce plaque formation.

Modeling Alzheimer's and Other Age Related Human Diseases in Embryonic Systems.

Modeling human disease in animals is an important strategy to discover potential methods of intervention. We suggest that there is much to be gained by employing a multi-model approach that takes advantage of different animal systems used in the laboratory simultaneously. We use the example of modeling Alzheimer's disease in Drosophila melanogaster, Caenorhabditis elegans, and Danio rerio to illustrate how such an approach can be employed to investigate the pathophysiology of the disease.

Ginseng and Ginkgo Biloba Effects on Cognition as Modulated by Cardiovascular Reactivity: A Randomised Trial.

BACKGROUND: There is some evidence to suggest that ginseng and Ginkgo biloba can improve cognitive performance, however, very little is known about the mechanisms associated with such improvement. Here, we tested whether cardiovascular reactivity to a task is associated with cognitive improvement. METHODOLOGY/PRINCIPAL FINDINGS: Using a double-blind, placebo controlled, crossover design, participants (N = 24) received two doses of Panax Ginseng (500, 1000 mg) or Ginkgo Biloba (120, 240 mg) (N = 24), and underwent a series of cognitive tests while systolic, diastolic, and heart rate readings were taken. Ginkgo Biloba improved aspects of executive functioning (Stroop and Berg tasks) in females but not in males. Ginseng had no effect on cognition. Ginkgo biloba in females reversed the initial (i.e. placebo) increase in cardiovascular reactivity (systolic and diastolic readings increased compared to baseline) to cognitive tasks. This effect (reversal) was most notable after those tasks (Stroop and Iowa) that elicited the greatest cardiovascular reactivity during placebo. In males, although ginkgo also decreased cardiovascular readings, it did so from an initial (placebo) blunted response (i.e. decrease or no change from baseline) to cognitive tasks. Ginseng, on the contrary, increased cardiovascular readings compared to placebo. CONCLUSIONS/SIGNIFICANCE: These results suggest that cardiovascular reactivity may be a mechanism by which ginkgo but not ginseng, in females is associated with certain forms of cognitive improvement. TRIAL REGISTRATION: ClinicalTrials.gov NCT02386852.

Obesity in pregnancy.

The prevalence of obesity has reached alarming proportions globally, and continues to rise in both developed and developing countries. Maternal obesity has become one of the most commonly occurring risk factors in obstetric practice. The 2003-2005 report of the Confidential Enquiries into Maternal Deaths in the United Kingdom highlighted obesity as a significant risk for maternal death [1]. More than half of all women who died from direct or indirect causes were either overweight or obese. For the mother, obesity increases the risk of obstetric complications during the antenatal, intrapartum and postnatal period, as well as contributing to technical difficulties with fetal assessment. The offspring of obese mothers also have a higher rate of perinatal morbidity and an increased risk of long-term health problems.

Multicenter prospective randomized study of single-incision mini-sling vs tension-free vaginal tape-obturator in management of female stress urinary incontinence: a minimum of 1-year follow-up.

OBJECTIVE: To compare an adjustable anchored single incision mini-sling (SIMS-Ajust) vs tension-free vaginal tape-obturator (TVT-O) in the management of female stress urinary incontinence (SUI) with a minimum of 1-year follow-up. METHODS: We conducted a multicenter prospective randomized control trial (RCT) performed in 6 United Kingdom centers in the period between October 2009 and October 2011. Women were randomized to either SIMS-Ajust (C. R. Bard) performed under local anesthesia or TVT-O (Ethicon Inc.) performed under general anesthesia. Women completed validated symptom-severity and quality of life (QOL) questionnaires preoperatively and at 1 year. In addition, women completed the Patient Global Impression of Improvement (PGI-I) and underwent the Cough Stress Test (CST) at 1 year. The primary outcome at 12 months was the patient-reported success rate. Secondary outcomes included objective cure, reoperation rate, impact on women's urinary symptoms, QOL, and sexual function. Data was analyzed using SPSS-19 with significance level set at 5%. RESULTS: One hundred thirty-seven women were randomized into 2 groups: the SIMS-Ajust group (n = 69) and the TVT-O group (n = 68). At 1 year, there were no significant differences in the patient-reported success rate (odds ratio [OR] 0.895, 95% confidence interval [CI] 0.344-2.330, P = 1.000), objectives success rate (OR 0.929, 95% CI 0.382-2.258, P = 1.00), and reoperation rates (OR 0.591, 95% CI 0.136-2.576, P = .721) between the SIMS-Ajust and the TVT-O groups, respectively. A comparable number of women reported cure/improvement of urgency (P = .658), significant improvement in QOL (P = .190), and sexual function (P = .699) in both groups. CONCLUSION: Adjustable-anchored SIMS (Ajust) is associated with comparable patient-reported and objective success rates when compared to standard midurethral sling (SMUS, TVT-O) at a minimum of 1-year follow-up. The results should be interpreted with caution due to the relatively small cohort size. Long-term follow-up of this RCT is required to ascertain the durability of these results.

Hybrid-drug design targeting Pseudomonas aeruginosa dihydropteroate synthase and dihydrofolate reductase

In this study, we successfully present the dual-target design hypothesis to inhibit both dihydropteroate synthase (DHPS) and dihydrofolate reductase (DHFR) enzymes using a novel scheme that integrates our previous antibiotic-phytochemical interaction data, fragment combination and knowledge-based methods. Both the enzymes are well established antibacterial targets from folate biosynthesis pathway and their synergistic modulation by a single hybrid entity may have profound therapeutic benefits. Evaluation of the designed hybrid compounds based on their physico-chemical properties has indicated them as promising drug candidates with drug-like pharmacotherapeutic profiles. In addition, the stereo-electronic properties such as HOMO, LUMO and MEP maps calculated by quantum chemical methods gave a good correlation with the common pharmacophoric features required for dual-site interactions. Furthermore, docking and dynamics simulation studies reveal that the designed hybrid compounds have favorable binding affinity and stability in both pterin-binding site of DHPS and folate-binding site of DHFR by forming strong hydrogen bonds and hydrophobic interactions with key active-site residues. Looking forward this study could serve as a prospective lead in the process of new natural-product based hybrid-drugs development.

Bactericidal antibiotic-phytochemical combinations against methicillin resistant Staphylococcus aureus

Methicillin resistant Staphylococcus aureus (MRSA) infection is a global concern nowadays. Due to its multi-drug resistant nature, treatment with conventional antibiotics does not assure desired clinical outcomes. Therefore, there is a need to find new compounds and/or alternative methods to get arsenal against the pathogen. Combination therapies using conventional antibiotics and phytochemicals fulfill both requirements. In this study, the efficacy of different phytochemicals in combination with selected antibiotics was tested against 12 strains of S. aureus (ATCC MRSA 43300, ATCC methicillin sensitive S. aureus or MSSA 29213 and 10 MRSA clinical strains collected from National University Hospital, Singapore). Out of the six phytochemicals used, tannic acid was synergistic with fusidic acid, minocycline, cefotaxime and rifampicin against most of strains tested and additive with ofloxacin and vancomycin. Quercetin showed synergism with minocycline, fusidic acid and rifampicin against most of the strains. Gallic acid ethyl ester showed additivity against all strains in combination with all antibiotics under investigation except with vancomycin where it showed indifference effect. Eugenol, menthone and caffeic acid showed indifference results against all strains in combination with all antibiotics. Interestingly, no antagonism was observed within these interactions. Based on the fractional inhibitory concentration indices, synergistic pairs were further examined by time-kill assays to confirm the accuracy and killing rate of the combinations over time. The two methods concurred with each other with 92% accuracy and the combinatory pairs were effective throughout the 24 hours of assay. The study suggests a possible incorporation of effective phytochemicals in combination therapies for MRSA infections.

Comparison of protein expression profiles of different stages of lymph nodes metastasis in breast cancer.

Breast cancer is the most common cancer among women worldwide. Breast cancer metastasis primarily happens through lymphatic system, where the extent of lymph node metastasis is the major factor influencing staging, prognosis and therapeutic decision of the disease. We aimed to study the protein expression changes in different N (regional lymph nodes) stages of breast cancer. Protein expression profiles of breast cancerous and adjacent normal tissues were mapped by proteomics approach that comprises of two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and tandem mass spectrometry (LC-MS/MS) analysis. Calreticulin and tropomyosin alpha 3 chains were the common up-regulated proteins in N0, N1 and N2 stages of breast cancer. Potential biomarker for each N stage was HSP 70 for N0, 80 k protein H precursor and PDI for N1 stage while 78 kDa glucose-regulated protein was found useful for N2 stage. In addition, significant up-regulation of PDI A3 was detected only in the metastasized breast cancer. The up-regulation expression of these proteins in cancerous tissues can potentially use as indicators for diagnosis, treatment and prognosis of different N stages of breast cancer.

Biodegradation of Low Density Polythene (LDPE) by Pseudomonas Species.

Low density polythene (LDPE) is the most widely used packaging material primarily because of its excellent mechanical properties, barrier properties against water, light weight, low cost and high energy effectiveness. However, due to its ubiquitous nature, and resistance to biodegradability, the disposal strategies are crucial and need attention. Recently, microorganisms have become the focus of interest for environmental friendly disposal of plastic and polymer-based waste. This manuscript aims to investigate the extent of biodegradability of LDPE by four different strains of Pseudomonas bacteria-Pseudomonas aeruginosa PAO1 (ATCC 15729), Pseudomonas aeruginosa (ATCC 15692), Pseudomonas putida (KT2440 ATCC 47054) and Pseudomonas syringae (DC3000 ATCC 10862). Degradation of LDPE was determined by weight loss of the sample, morphological changes, mechanical and spectroscopic variations. The eluted compounds after degradation were analysed by gas chromatography coupled with mass spectroscopy. Our results show that Pseudomonas spp. can degrade LDPE films.

Bactericidal antibiotic-phytochemical combinations against methicillin resistant Staphylococcus aureus.

Methicillin resistant Staphylococcus aureus (MRSA) infection is a global concern nowadays. Due to its multi-drug resistant nature, treatment with conventional antibiotics does not assure desired clinical outcomes. Therefore, there is a need to find new compounds and/or alternative methods to get arsenal against the pathogen. Combination therapies using conventional antibiotics and phytochemicals fulfill both requirements. In this study, the efficacy of different phytochemicals in combination with selected antibiotics was tested against 12 strains of S. aureus (ATCC MRSA 43300, ATCC methicillin sensitive S. aureus or MSSA 29213 and 10 MRSA clinical strains collected from National University Hospital, Singapore). Out of the six phytochemicals used, tannic acid was synergistic with fusidic acid, minocycline, cefotaxime and rifampicin against most of strains tested and additive with ofloxacin and vancomycin. Quercetin showed synergism with minocycline, fusidic acid and rifampicin against most of the strains. Gallic acid ethyl ester showed additivity against all strains in combination with all antibiotics under investigation except with vancomycin where it showed indifference effect. Eugenol, menthone and caffeic acid showed indifference results against all strains in combination with all antibiotics. Interestingly, no antagonism was observed within these interactions. Based on the fractional inhibitory concentration indices, synergistic pairs were further examined by time-kill assays to confirm the accuracy and killing rate of the combinations over time. The two methods concurred with each other with 92% accuracy and the combinatory pairs were effective throughout the 24 hours of assay. The study suggests a possible incorporation of effective phytochemicals in combination therapies for MRSA infections.

Maggot metabolites and their combinatory effects with antibiotic on Staphylococcus aureus.

BACKGROUND: Maggot therapy has been in practice for effective debridement, disinfection and healing of chronic wounds. Due to their antiseptic action during wound healing, their metabolites have been investigated in the past for antibacterial activity. They have been particularly useful for treatment of wounds infected with multi-drug resistant Staphylococcus aureus. Antibiotics, on the other hand, can predispose bacteria to develop resistance. Substances that are able to modulate or delay the occurrence of resistance in bacteria are under investigation by many researchers around the world. In the present study, antibacterial activity in excretions/secretions (ES) from maggots of Lucilia cuprina blowfly was demonstrated. The extracts were also screened in combination with antibiotic, ciprofloxacin. METHODS: L. cuprina blowfly maggots were reared for extraction of its metabolites. The ES extracted was screened against S. aureus, alone and in combination with ciprofloxacin, both for short term and long term exposure analysis. A microchannel-based device and system was used for experiments instead of conventional techniques. RESULTS: The original ES had shown partial bacterial growth inhibition. However, in combination with ciprofloxacin, at sub-inhibitory concentrations, certain combinations revealed anti-staphylococcal activity, with bacterial reduction of up to 50%, after 24 hours. The six day study on S. aureus exposed to ES-ciprofloxacin combination suggested a potential delay in development of adaptive resistance as opposed to when ciprofloxacin was used as single agent. CONCLUSIONS: The combination effect of ES and ciprofloxacin at sub-MIC levels showed enhanced antibacterial activity compared to the effect of ES and ciprofloxacin as single agents. Based on the results of ES-ciprofloxacin combinations, a more effective means of treatment for S. aureus can be proposed.

Insights into antifolate activity of phytochemicals against Pseudomonas aeruginosa.

Pseudomonas aeruginosa is an opportunistic drug resistant pathogen. Drug interaction studies for phytochemicals (protocatechuic acid (PA), gallic acid (GA), quercetin (QUER), and myricetin (MYR)) in combination with antifolates (sulfamethoxazole (SMX) and trimethoprim (TMP)) are presented. Our results show that the combinations of SMX and phytochemicals are synergistic, whereas TMP in combination with phytochemicals results in additive mode of interaction. Molecular docking of phytochemicals in the active site of modeled P. aeruginosa dihydrofolate reductase (DHFR), an important enzyme in the folic acid biosynthesis pathway, shows that the phytochemicals QUER and MYR dock in the active site of P. aeruginosa DHFR with promoted binding at the NADP site, PA, and GA dock in the active site of P. aeruginosa DHFR with promoted binding at the folate binding site. Possible mode of action of these phytochemicals as anti-DHFR compounds in this bacterium is suggested. Taken together, the above findings provide novel insights to mode of interactions of these phytochemicals with antibiotics and may have significance as prospective leads in the development of antipseudomonal drug developments.

Cloning and targeted disruption of two lipopolysaccharide biosynthesis genes, kdsA and waaG, of Pseudomonas aeruginosa PAO1 by site-directed mutagenesis.

The emergence of antibiotic resistance in bacterial pathogens poses a great challenge to public health and emphasizes the need for new antimicrobial targets. The recent development of microbial genomics and the availability of genome sequences allows for the identification of essential genes which could be novel and potential targets for antibacterial drugs. However, these predicted targets need experimental validation to confirm essentiality. Here, we report on experimental validation of a two potential targets in the lipopolysaccharide (LPS) biosynthesis pathway of the pathogen Pseudomonas aeruginosa PAO1 using insertion duplication. Two genes, kdsA and waaG, from LPS encoding proteins 2-dehydro-3-deoxyphosphooctonate aldolase and UDP-glucose (heptosyl) LPS α-1,3-glucosyltransferase were selected as putative target candidates for the gene disruption experiments using plasmid insertion mutagenesis to determine essentiality. The introduction of a selectable ampicillin and kanamycin resistance marker into the chromosome resulted in lack of recovery of antibiotic-resistant colonies suggesting the essentiality of these genes for the survival of P. aeruginosa. Several molecular analyses were carried out in order to confirm the essentiality of these genes. We propose that the above two validated drug targets are essential and can be screened for functional inhibitors for the discovery of novel therapeutic compounds against antibiotic-resistant opportunistic pathogen P. aeruginosa.

Activity and interactions of antibiotic and phytochemical combinations against Pseudomonas aeruginosa in vitro.

In this study the in vitro activities of seven antibiotics (ciprofloxacin, ceftazidime, tetracycline, trimethoprim, sulfamethoxazole, polymyxin B and piperacillin) and six phytochemicals (protocatechuic acid, gallic acid, ellagic acid, rutin, berberine and myricetin) against five P. aeruginosa isolates, alone and in combination are evaluated. All the phytochemicals under investigation demonstrate potential inhibitory activity against P. aeruginosa. The combinations of sulfamethoxazole plus protocatechuic acid, sulfamethoxazole plus ellagic acid, sulfamethoxazole plus gallic acid and tetracycline plus gallic acid show synergistic mode of interaction. However, the combinations of sulfamethoxazole plus myricetin shows synergism for three strains (PA01, DB5218 and DR3062). The synergistic combinations are further evaluated for their bactericidal activity against P. aeruginosa ATCC strain using time-kill method. Sub-inhibitory dose responses of antibiotics and phytochemicals individually and in combination are presented along with their interaction network to suggest on the mechanism of action and potential targets for the phytochemicals under investigation. The identified synergistic combinations can be of potent therapeutic value against P. aeruginosa infections. These findings have potential implications in delaying the development of resistance as the antibacterial effect is achieved with lower concentrations of both drugs (antibiotics and phytochemicals).

Infrared authentication of ginseng species: the use of the 2-6PC rule.

The quality of herbal products is important for ensuring efficacy and consumer safety. Traditional methods of authenticating herbs like ginseng via their morphology are hardly reliable. Different chemical constituents in herbs like ginseng tend to exhibit characteristic IR fingerprints that enable their identification. We previously introduced an IR-based protocol known as the "2-6PC rule" to categorize and identify ginseng and its products, as well as distinguishing it from morphological fakes. Here, we describe the use of this rule as a rapid and effective means of analyzing the IR spectral fingerprints of the biologically active components of ginseng, as well as distinguishing among its species. Our results show that Panax ginseng, P. quinquefolius, and P. notoginseng can be differentiated from each other. Our results also indicate the presence of starch, carbohydrates, calcium oxalate, and ginsenosides Re and Rg1 in commercial ginseng roots sold in Singapore. This work effectively demonstrates the usefulness of the 2-6PC rule as a rapid screening tool in the authentication of ginseng species.

Activity of Chitosans in combination with antibiotics in Pseudomonas aeruginosa.

Chitosan and its derivative water soluble Chitosan oligosaccharide are used in a variety of applications in pharmaceutical preparations. In this study, 2 wild (ATCC 15729 and PAO1) and 2 mutant strains (PT121 and PT149) of P. aeruginosa are investigated for drug-drug interactions in vitro. 10 antimicrobial agents (antibiotics) are combined with different degree of deacetylated Chitosans and Chitosan oligosaccharide. All the chitosans show synergistic activity with sulfamethoxazole, a sulfonamide antimicrobial agent. It is interesting to observe that the MIC value for the MexEF-OprN overexpressing mutant strain of P. aeruginosa is 5 fold higher than the other strains under investigation suggesting a possible role of this efflux pump in Sulfamethoxazole efflux. The findings suggest on the use of chitosans as enhancing agent in combination with antibiotics in pharmaceutical preparations.

In vitro combinations of antibiotics and phytochemicals against Pseudomonas aeruginosa.

BACKGROUND AND PURPOSE: Antibiotic combinations are used to enhance antibacterial efficacy and to prevent the development of resistance. In this study, the in vitro activities of antibiotic and phytochemical combinations against Pseudomonas aeruginosa were tested by the fractional inhibitory concentration method, derived from the minimal inhibitory concentrations (MICs) of the agents in combination. METHODS: The antimicrobial activity of phytochemicals, alone and in combination with antibiotics, was evaluated using the checkerboard assay and time-kill curve methods. RESULTS: There was synergism between gentamicin and caffeic acid, and sulfadiazine and the 3 phytochemicals under investigation (protocatechuic acid, quercetin, caffeic acid). The MIC of sulfadiazine was 256 microg/mL, and of gentamicin was 2 microg/mL. When gentamicin was combined with one-quarter the MIC of caffeic acid, the MIC of gentamicin was reduced 4-fold. When sulfadiazine was tested with one-quarter the MIC of protocatechuic acid, quercetin, and caffeic acid, the MIC was reduced 4-fold in combination with each of the drugs. CONCLUSIONS: These results indicate the potential efficacy of phytochemicals in combination with antibiotics for enhancing total biological activity.