Characterization of ten microsatellite loci in midget faded rattlesnake (Crotalus oreganus concolor)

Science.gov (United States)

Oyler-McCance, Sara J.; Parker, Joshua M.

2010-01-01

Primers for 10 microsatellite loci were developed for midget faded rattlesnake (Crotalus oreganus concolor), a small bodied subspecies of the Western Rattlesnake, which is found in the Colorado Plateau of eastern Utah, western Colorado and southwestern Wyoming. In a screen of 23 individuals from the most northern portion of the subspecies range in southwestern Wyoming, the 10 loci were found to have levels of variability ranging from 4 to 11 alleles. No loci were found to be linked, although one locus revealed significant departures from Hardy–Weinberg equilibrium. These microsatellite loci will be applicable for population genetic analyses, which will ultimately aid in management efforts for this rare subspecies of rattlesnake.

Body size evolution in insular speckled rattlesnakes (Viperidae: Crotalus mitchellii.

Directory of Open Access Journals (Sweden)

Jesse M Meik

2010-03-01

Full Text Available Speckled rattlesnakes (Crotalus mitchellii inhabit multiple islands off the coast of Baja California, Mexico. Two of the 14 known insular populations have been recognized as subspecies based primarily on body size divergence from putative mainland ancestral populations; however, a survey of body size variation from other islands occupied by these snakes has not been previously reported. We examined body size variation between island and mainland speckled rattlesnakes, and the relationship between body size and various island physical variables among 12 island populations. We also examined relative head size among giant, dwarfed, and mainland speckled rattlesnakes to determine whether allometric differences conformed to predictions of gape size (and indirectly body size evolving in response to shifts in prey size.Insular speckled rattlesnakes show considerable variation in body size when compared to mainland source subspecies. In addition to previously known instances of gigantism on Angel de la Guarda and dwarfism on El Muerto, various degrees of body size decrease have occurred frequently in this taxon, with dwarfed rattlesnakes occurring mostly on small, recently isolated, land-bridge islands. Regression models using the Akaike information criterion (AIC showed that mean SVL of insular populations was most strongly correlated with island area, suggesting the influence of selection for different body size optima for islands of different size. Allometric differences in head size of giant and dwarf rattlesnakes revealed patterns consistent with shifts to larger and smaller prey, respectively.Our data provide the first example of a clear relationship between body size and island area in a squamate reptile species; among vertebrates this pattern has been previously documented in few insular mammals. This finding suggests that selection for body size is influenced by changes in community dynamics that are related to graded differences in area over

Cytoskeleton, endoplasmic reticulum and nucleus alterations in CHO-K1 cell line after Crotalus durissus terrificus (South American rattlesnake venom treatment

Directory of Open Access Journals (Sweden)

B. P. Tamieti

2007-01-01

Full Text Available Snake venoms are toxic to a variety of cell types. However, the intracellular damages and the cell death fate induced by venom are unclear. In the present work, the action of the South American rattlesnake Crotalus durissus terrificus venom on CHO-K1 cell line was analyzed. The cells CHO-K1 were incubated with C. d. terrificus venom (10, 50 and 100g/ml for 1 and 24 hours, and structural alterations of actin filaments, endoplasmic reticulum and nucleus were assessed using specific fluorescent probes and agarose gel electrophoresis for DNA fragmentation. Significant structural changes were observed in all analyzed structures. DNA fragmentation was detected suggesting that, at the concentrations used, the venom induced apoptosis.

Physiological importance of the coronary arterial blood supply to the rattlesnake heart

DEFF Research Database (Denmark)

Hagensen, Mette; Abe, Augusto S.; Falk, Erling

2008-01-01

Pa and 58.2±2.2 beats min-1, respectively, during activity and the ECG was not affected. This was not different from sham-operated snakes. Thus, while the outer compact layer of the rattlesnake heart clearly has an extensive coronary supply, rattlesnakes sustain a high blood pressure and heart rate during...

Complete Bouguer gravity and aeromagnetic maps of the Rattlesnake Roadless Area, Missoula County, Montana

Science.gov (United States)

Kulik, Dolores M.

1986-01-01

The Rattlesnake Roadless Area was identified by the U.S. Forest Service as a possible addition to the Wilderness System.  This 120 square mile area is north of Missoula, Mont. (see index map); Rattlesnake Creek forms the major drainage to the south.

Spatial ecology of timber rattlesnakes on the hardwood ecosystem experiment: pre-treatment results

Science.gov (United States)

Brian J. MacGowan; Zachary J. Walker

2013-01-01

The timber rattlesnake (Crotalus horridus) is a species of conservation concern throughout much of its geographic range and may serve as a sentinel species in investigations of the effects of timber harvesting on forest reptiles. Our objective was to determine the effect of even-aged timber management regimes on timber rattlesnake home range and...

Quantitative characterization of pyrimidine dimer excision from UV-irradiated DNA (excision capacity) by cell-free extracts of the yeast Saccharomyces cerevisiae

International Nuclear Information System (INIS)

Bekker, M.L.; Kaboev, O.K.; Akhmedov, A.T.; Luchkina, L.A.

1984-01-01

Cell-free extracts from wild-type yeast (RAD + ) and from rad mutants belonging to the RAD3 epistatic group (rad1-1, rad2-1, rad3-1, rad4-1) contain activities catalyzing the excision of pyrimidine dimers (PD) from purified ultraviolet-irradiated DNA which was not pre-treated with exogenous UV-endonuclease. The level of these activities in cell-free extracts from rad mutants did not differ from that in wild-type extract and was close to the in vivo excision capacity of the latter calculated from the LD 37 (about 10 4 PD per haploid genome). (Auth.)

Enterococcus faecium LKE12 Cell-Free Extract Accelerates Host Plant Growth via Gibberellin and Indole-3-Acetic Acid Secretion.

Science.gov (United States)

Lee, Ko-Eun; Radhakrishnan, Ramalingam; Kang, Sang-Mo; You, Young-Hyun; Joo, Gil-Jae; Lee, In-Jung; Ko, Jae-Hwan; Kim, Jin-Ho

2015-09-01

The use of microbial extracts containing plant hormones is a promising technique to improve crop growth. Little is known about the effect of bacterial cell-free extracts on plant growth promotion. This study, based on phytohormonal analyses, aimed at exploring the potential mechanisms by which Enterococcus faecium LKE12 enhances plant growth in oriental melon. A bacterial strain, LKE12, was isolated from soil, and further identified as E. faecium by 16S rDNA sequencing and phylogenetic analysis. The plant growth-promoting ability of an LKE12 bacterial culture was tested in a gibberellin (GA)-deficient rice dwarf mutant (waito-C) and a normal GA biosynthesis rice cultivar (Hwayongbyeo). E. faecium LKE12 significantly improved the length and biomass of rice shoots in both normal and dwarf cultivars through the secretion of an array of gibberellins (GA1, GA3, GA7, GA8, GA9, GA12, GA19, GA20, GA24, and GA53), as well as indole-3-acetic acid (IAA). To the best of our knowledge, this is the first study indicating that E. faecium can produce GAs. Increases in shoot and root lengths, plant fresh weight, and chlorophyll content promoted by E. faecium LKE12 and its cell-free extract inoculated in oriental melon plants revealed a favorable interaction of E. faecium LKE12 with plants. Higher plant growth rates and nutrient contents of magnesium, calcium, sodium, iron, manganese, silicon, zinc, and nitrogen were found in cell-free extract-treated plants than in control plants. The results of the current study suggest that E. faecium LKE12 promotes plant growth by producing GAs and IAA; interestingly, the exogenous application of its cell-free culture extract can be a potential strategy to accelerate plant growth.

A study of bacterial contamination of rattlesnake venom

Directory of Open Access Journals (Sweden)

E. Garcia-Lima

1987-03-01

Full Text Available The authors studied the bacterial contamination of rattlesnake venom isolated from snakes in captivity and wild snakes caught recently. The captive snakes showed a relatively high incidence of bacterial contamination of their venom.

Formation of Lignans(-)-Secoisolariciresinol and (-)-Matairesinol with Forsythia intermedia Cell-Free Extracts

Science.gov (United States)

Umezawa, Toshiaki; Davin, Laurence B.; Lewis, Norman G.

1991-01-01

In vivo labeling experiments of Forsythia intermedia plant tissue with [8-(C-14)]- and [9,9-(2)H2,OC(2)H3]coniferyl alcohols revealed that the lignans, (-)-secoisolariciresinol and (-)-matairesinol, were derived from two coniferyl alcohol molecules; no evidence for the formation of the corresponding (+)-enantiomers was found. Administration of (+/-)-[Ar-(H-3)] secoisolariciresinols to excised shoots of F.intermedia resulted in a significant conversion into (-)-matairesinol; again, the (+)-antipode was not detected. Experiments using cell-free extracts of F.intermedia confirmed and extended these findings. In the presence of NAD(P)H and H2O2, the cell-free extracts catalyzed the formation of (-)- secoisolariciresinol, with either [8-(C-14)]- or [9,9-(2)H2,OC(2)H3]coniferyl alcohols as substrates. The (+)- enantiomer was not formed. Finally, when either (-)-[Ar-(H-3)] or (+/-)-[Ar-(H-2)]secoisolariciresinols were used as substrates, in the presence of NAD(P), only (-)- and not (+)-matairesinol formation occurred. The other antipode, (+)-secoisolariciresinol, did not serve as a substrate for the formation of either (+)- or (-)-matairesinol. Thus, in F.intermedia, the formation of the lignan, (-)-secoisolariciresinol, occurs under strict stereochemical control, in a reaction or reactions requiring NAD(P)H and H2O2 as cofactors. This stereoselectivity is retained in the subsequent conversion into (-)-matairesinol, since (+)-secoisolariciresinol is not a substrate. These are the first two enzymes to be discovered in lignan formation.

Analysis of signal transduction in cell-free extracts and rafts of Xenopus eggs.

Science.gov (United States)

Tokmakov, Alexander A; Iwasaki, Tetsushi; Sato, Ken-Ichi; Fukami, Yasuo

2010-05-01

Intracellular signaling during egg activation/fertilization has been extensively studied using intact eggs, which can be manipulated by microinjection of different mRNAs, proteins, or chemical drugs. Furthermore, egg extracts, which retain high CSF activity (CSF-arrested extracts), were developed for studying fertilization/activation signal transduction, which have significant advantages as a model system. The addition of calcium to CSF-arrested extracts initiates a plethora of signaling events that take place during egg activation. Hence, the signaling downstream of calcium mobilization has been successfully studied in the egg extracts. Moreover, despite disruption of membrane-associated signaling compartments and ordered compartmentalization during extract preparation, CSF-arrested extracts can be successfully used to study early signaling events, which occur upstream of calcium release during egg activation/fertilization. In combination with the CSF-arrested extracts, activated egg rafts can reproduce some events of egg activation, including PLCgamma activation, IP3 production, transient calcium release, MAPK inactivation, and meiotic exit. This becomes possible due to complementation of the sperm-induced egg activation signaling machinery present in the rafts with the components of signal transduction system localized in the extracts. Herein, we describe protocols for studying molecular mechanisms of egg fertilization/activation using cell-free extracts and membrane rafts prepared from metaphase-arrested Xenopus eggs.

Measuring strand discontinuity-directed mismatch repair in yeast Saccharomyces cerevisiae by cell-free nuclear extracts.

Science.gov (United States)

Yuan, Fenghua; Lai, Fangfang; Gu, Liya; Zhou, Wen; El Hokayem, Jimmy; Zhang, Yanbin

2009-05-01

Mismatch repair corrects biosynthetic errors generated during DNA replication, whose deficiency causes a mutator phenotype and directly underlies hereditary non-polyposis colorectal cancer and sporadic cancers. Because of remarkably high conservation of the mismatch repair machinery between the budding yeast (Saccharomyces cerevisiae) and humans, the study of mismatch repair in yeast has provided tremendous insights into the mechanisms of this repair pathway in humans. In addition, yeast cells possess an unbeatable advantage over human cells in terms of the easy genetic manipulation, the availability of whole genome deletion strains, and the relatively low cost for setting up the system. Although many components of eukaryotic mismatch repair have been identified, it remains unclear if additional factors, such as DNA helicase(s) and redundant nuclease(s) besides EXO1, participate in eukaryotic mismatch repair. To facilitate the discovery of novel mismatch repair factors, we developed a straightforward in vitro cell-free repair system. Here, we describe the practical protocols for preparation of yeast cell-free nuclear extracts and DNA mismatch substrates, and the in vitro mismatch repair assay. The validity of the cell-free system was confirmed by the mismatch repair deficient yeast strain (Deltamsh2) and the complementation assay with purified yeast MSH2-MSH6.

Extracting histones for the specific purpose of label-free MS.

Science.gov (United States)

Govaert, Elisabeth; Van Steendam, Katleen; Scheerlinck, Ellen; Vossaert, Liesbeth; Meert, Paulien; Stella, Martina; Willems, Sander; De Clerck, Laura; Dhaenens, Maarten; Deforce, Dieter

2016-12-01

Extracting histones from cells is the first step in studies that aim to characterize histones and their post-translational modifications (hPTMs) with MS. In the last decade, label-free quantification is more frequently being used for MS-based histone characterization. However, many histone extraction protocols were not specifically designed for label-free MS. While label-free quantification has its advantages, it is also very susceptible to technical variation. Here, we adjust an established histone extraction protocol according to general label-free MS guidelines with a specific focus on minimizing sample handling. These protocols are first evaluated using SDS-PAGE. Hereafter, a selection of extraction protocols was used in a complete histone workflow for label-free MS. All protocols display nearly identical relative quantification of hPTMs. We thus show that, depending on the cell type under investigation and at the cost of some additional contaminating proteins, minimizing sample handling can be done during histone isolation. This allows analyzing bigger sample batches, leads to reduced technical variation and minimizes the chance of in vitro alterations to the hPTM snapshot. Overall, these results allow researchers to determine the best protocol depending on the resources and goal of their specific study. Data are available via ProteomeXchange with identifier PXD002885. © 2016 The Authors. Proteomics Published by Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

An evaluation of aquifer intercommunication between the unconfined and Rattlesnake Ridge aquifers on the Hanford Site

International Nuclear Information System (INIS)

Jensen, E.J.

1987-10-01

During 1986, Pacific Northwest Laboratory conducted a study of a portion of the Rattlesnake Ridge aquifer (confined aquifer) that lies beneath the B Pond - Gable Mountain Pond area of the Hanford Site. The purpose was to determine the extent of intercommunication between the unconfined aquifer and the uppermost regionally extensive confined aquifer, referred to as the Rattlesnake Ridge aquifer. Hydraulic head data and chemical data were collected from the ground water in the study area during December 1986. The hydraulic head data were used to determine the effects caused by water discharged to the ground from B Pond on both the water table of the unconfined aquifer and the potentiometric surface of the confined aquifer. The chemical data were collected to determine the extent of chemical constituents migrating from the unconfined aquifer to the confined aquifer. Analysis of chemical constituents in the Rattlesnake Ridge aquifer demonstrated that communication between the unconfined and confined aquifers had occurred. However, the levels of contaminants found in the Rattlesnake Ridge aquifer during this study were below the DOE Derived Concentration Guides

Cell-Free, De Nova Synthesis of Poliovirus

Science.gov (United States)

Molla, Akhteruzzaman; Paul, Aniko V.; Wimmer, Eckard

1991-12-01

Cell-free translation of poliovirus RNA in an extract of uninfected human (HeLa) cells yielded viral proteins through proteolysis of the polyprotein. In the extract, newly synthesized proteins catalyzed poliovirus-specific RNA synthesis, and formed infectious poliovirus de novo. Newly formed virions were neutralized by type-specific antiserum, and infection of human cells with them was prevented by poliovirus receptor-specific antibodies. Poliovirus synthesis was increased nearly 70-fold when nucleoside triphosphates were added, but it was abolished in the presence of inhibitors of translation or viral genome replication. The ability to conduct cell-free synthesis of poliovirus will aid in the study of picornavirus proliferation and in the search for the control of picornaviral disease.

Effects of the canine rattlesnake vaccine in moderate to severe cases of canine crotalid envenomation

Directory of Open Access Journals (Sweden)

Leonard MJ

2014-10-01

Full Text Available McGee J Leonard,1 Catherine Bresee,2 Andrew Cruikshank1 1Animal Specialty and Emergency Center, Los Angeles, CA, USA; 2The Biostatistics and Bioinformatics Research Center, Cedars-Sinai Medical Center, Los Angeles, CA, USA Abstract: This is a retrospective multicenter study (2006–2012 examining a population of dogs with moderate to severe crotalid envenomation for protective effects of the canine rattlesnake vaccine. Five nonacademic emergency and referral veterinary hospitals in Southern California were involved in the study and contributed records regarding a total of 82 client-owned dogs that were treated for naturally occurring rattlesnake envenomation. All dogs received antivenin (Crotalidae polyvalent, with dosages ranging from one to three vials (mean: 1.3±0.6. Fourteen dogs (17% had a history of prior vaccination against crotalid venom. In univariate logistic regression modeling, cases with lower body weight (P=0.0001 or higher snakebite severity scores (P<0.0001 were associated with greater morbidity. No statistically significant difference in morbidity or mortality between vaccinated and unvaccinated dogs was found. The findings of this study did not identify a significantly protective effect of previous vaccination in the cases of moderate to severe rattlesnake envenomation that require treatment with antivenin. Keywords: rattlesnake envenomation, vaccine, antivenin, canine

Characterization of cell-free extracts from fenpropathrin-degrading strain Bacillus cereus ZH-3 and its potential for bioremediation of pyrethroid-contaminated soils.

Science.gov (United States)

Liu, Jie; Huang, Wenwen; Han, Haitao; She, Changchun; Zhong, Guohua

2015-08-01

Synthetic pyrethroid fenpropathrin has received increasing attention because of its environmental contamination and toxic effects on non-target organisms including human beings. Here we report the degradation characteristics of cell-free extracts from fenpropathrin-degrading strain Bacillus cereus ZH-3 and its potential for pyrethroid bioremediation in soils. 50mg·L(-1) of fenpropathrin was decreased to 20.6mg·L(-1) by the enzymatic extracts (869.4mg·L(-1)) within 30min. Kinetic constants Km and Vm were determined to be 1006.7nmol·L(-1) and 56.8nmol·min(-1), respectively. Degradation products were identified as 3-phenoxybenzaldehyde, α-hydroxy-3-phenoxy-benzeneacetonitrile and phenol by gas chromatography-mass spectrometry (GC-MS). In addition to degradation of fenpropathrin, the cell-free extracts could degrade other pyrethroids including beta-cypermethrin, cyfluthrin, deltamethrin and cypermethrin. Additionally, the reaction conditions were optimized. In the sterile and non-sterile soils, 50mg·kg(-1) of fenpropathrin was reduced to 15.3 and 13.9mg·L(-1) in 1d, respectively. Sprayed 100 and 300mg·kg(-1) of fenpropathrin emulsifiable concentrate (EC), up to 84.6% and 92.1% of soil fenpropathrin were removed from soils within 7d, respectively. Taken together, our results depict the biodegradation characteristics of cell-free extracts from B. cereus ZH-3, highlight its promising potential in bioremediation of pyrethroid-contaminated soils and also provide new insights into the utilization of degrading microbes. Copyright © 2015. Published by Elsevier B.V.

Seasonal variation in hormonal responses of timber rattlesnakes (Crotalus horridus) to reproductive and environmental stressors.

Science.gov (United States)

Lutterschmidt, William I; Lutterschmidt, Deborah I; Mason, Robert T; Reinert, Howard K

2009-08-01

Data addressing adrenocortical modulation across taxonomic groups are limited, especially with regard to how female reproductive condition influences the sensitivity of the hypothalamus-pituitary-adrenal axis. We investigated seasonal and reproductive variation in basal and stress-induced hormone profiles in a population of free-ranging timber rattlesnakes (Crotalus horridus) in north-central Pennsylvania during spring (i.e., May), summer (i.e., July), and early fall (i.e., September). Baseline corticosterone concentrations varied seasonally and were significantly lower during the summer sampling period in July. We observed a significant negative relationship between baseline corticosterone and testosterone in male snakes, while baseline corticosterone and estradiol tended to be positively correlated in females. Treatment of snakes with 1 h of capture stress significantly increased corticosterone across all seasons. However, there was a significant interaction between corticosterone responses to capture stress and season, suggesting that adrenocortical function is modulated seasonally. Because elevated corticosterone may be associated with reproduction, we asked whether hormonal stress responses vary with female reproductive condition. Although sample sizes are low, reproductive snakes had significantly higher baseline and stress-induced corticosterone concentrations than non-reproductive or post-parturient females. Further, despite similar baseline corticosterone concentrations between non-reproductive and post-parturient rattlesnakes, post-parturient females responded to capture stress with a significantly higher increase in corticosterone. Collectively, these data suggest that the sensitivity of the hypothalamus-pituitary-adrenal axis varies both seasonally and with changing reproductive states.

The coupling of the neutron transport application RATTLESNAKE to the nuclear fuels performance application BISON under the MOOSE framework

Energy Technology Data Exchange (ETDEWEB)

Gleicher, Frederick N.; Williamson, Richard L.; Ortensi, Javier; Wang, Yaqi; Spencer, Benjamin W.; Novascone, Stephen R.; Hales, Jason D.; Martineau, Richard C.

2014-10-01

The MOOSE neutron transport application RATTLESNAKE was coupled to the fuels performance application BISON to provide a higher fidelity tool for fuel performance simulation. This project is motivated by the desire to couple a high fidelity core analysis program (based on the self-adjoint angular flux equations) to a high fidelity fuel performance program, both of which can simulate on unstructured meshes. RATTLESNAKE solves self-adjoint angular flux transport equation and provides a sub-pin level resolution of the multigroup neutron flux with resonance treatment during burnup or a fast transient. BISON solves the coupled thermomechanical equations for the fuel on a sub-millimeter scale. Both applications are able to solve their respective systems on aligned and unaligned unstructured finite element meshes. The power density and local burnup was transferred from RATTLESNAKE to BISON with the MOOSE Multiapp transfer system. Multiple depletion cases were run with one-way data transfer from RATTLESNAKE to BISON. The eigenvalues are shown to agree well with values obtained from the lattice physics code DRAGON. The one-way data transfer of power density is shown to agree with the power density obtained from an internal Lassman-style model in BISON.

Responses of timber rattlesnakes to fire: Lessons from two prescribed burns

Science.gov (United States)

Steven J. Beaupre; Lara E. Douglas

2012-01-01

Timber rattlesnakes (Crotalus horridus) are excellent model organisms for understanding the effects of large scale habitat manipulations because of their low-energy lifestyle, rapid response to changes in resource environment, uniform diet (small mammals), and simple behaviors. We present two case studies that illustrate interactions between timber...

Free radical scavenging and anti-oxidative activities of an ethanol-soluble pigment extract prepared from fermented Zijuan Pu-erh tea.

Science.gov (United States)

Fan, Jiang Ping; Fan, Chong; Dong, Wen Min; Gao, Bin; Yuan, Wei; Gong, Jia Shun

2013-09-01

An ethanol-soluble pigment extract was separated from fermented Zijuan Pu-erh tea. The compositions of the ethanol soluble pigment extract were analyzed by high-performance liquid chromatography-tandem mass spectroscopy (HPLC-MS/MS). The extract was prepared into a series of ethanol solutions and analyzed for free radical-scavenging activities (against two free radicals: 1,1-diphenyl-2-picrylhydrazyl (DPPH) and (2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO)) and in vitro anti-oxidative properties. Electron spin resonance spectroscopy showed that the peaks of DPPH and TEMPO decreased with increasing extract concentration, suggesting that the extract had excellent free radical-scavenging activities. In vitro cell culture suggested that, at 50-200 mg/L, the extract had no measurable effect on the viability of vascular endothelial cells (ECV340) but produced significant protective effects for cells that underwent oxidative injuries due to hydrogen peroxide (H₂O₂) treatment. Compared with the H₂O₂ treatment alone cells group, 200 mg/L of the extract increased the activity of superoxide dismutase (SOD) in cells by 397.3%, and decreased the concentration of malondialdehyde (MDA) and the activity of lactate acid dehydrogenase (LDH) by 47.8% and 69.6%, respectively. These results suggest that the extract has excellent free radical scavenging and anti-oxidative properties. Copyright © 2013 Elsevier Ltd. All rights reserved.

Biological Synthesis of Silver Nanoparticles by Cell-Free Extract of Spirulina platensis

Directory of Open Access Journals (Sweden)

Gaurav Sharma

2015-01-01

Full Text Available The present study explores biological synthesis of silver nanoparticles (AgNPs using the cell-free extract of Spirulina platensis. Biosynthesised AgNPs were characterised by UV-Vis spectroscopy, SEM, TEM, and FTIR analysis and finally evaluated for antibacterial activity. Extracellular synthesis using aqueous extract of S. platensis showed the formation of well scattered, highly stable, spherical AgNPs with an average size of 30–50 nm. The size and morphology of the nanoparticles were confirmed by SEM and TEM analysis. FTIR and UV-Vis spectra showed that biomolecules, proteins and peptides, are mainly responsible for the formation and stabilisation of AgNPs. Furthermore, the synthesised nanoparticles exhibited high antibacterial activity against pathogenic Gram-negative, that is, Escherichia coli, MTCC-9721; Proteus vulgaris, MTCC-7299; Klebsiella pneumoniae, MTCC-9751, and Gram-positive, that is, Staphylococcus aureus, MTCC-9542; S. epidermidis, MTCC-2639; Bacillus cereus, MTCC-9017, bacteria. The AgNPs had shown maximum zone of inhibition (ZOI that is 31.3±1.11 in P. vulgaris. Use of such a microalgal system provides a simple, cost-effective alternative template for the biosynthesis of nanomaterials of silver in a large scale that could be of great use in biomedical applications.

Activity cycles and foraging behaviors of free-ranging sidewinder rattlesnakes (Crotalus cerastes): the ontogeny of hunting in a precocial vertebrate.

Science.gov (United States)

Clark, Rulon W; Dorr, Scott W; Whitford, Malachi D; Freymiller, Grace A; Putman, Breanna J

2016-06-01

Predators often employ a complex series of behaviors to overcome antipredator defenses and effectively capture prey. Although hunting behaviors can improve with age and experience, many precocial species are necessarily effective predators from birth. Additionally, many predators experience innate ontogenetic shifts in predatory strategies as they grow, allowing them to adapt to prey more appropriate for their increased size and energetic needs. Understanding how the relative roles of innate age-specific adaptation and learning have evolved requires information on how predation behavior develops in situ, in free-ranging predators. However, most of the research on the ontogeny of predation behavior is based on laboratory studies of captive animals, largely due to the difficulty of following newborn individuals in nature. Here, we take advantage of the unique tracks left by juveniles of a precocial viperid, the sidewinder rattlesnake (Crotalus cerastes), which we used to follow free-ranging snakes in the field. We recorded details of their ambush hunting behavior, and compared the behaviors of these juveniles to adult snakes that we monitored in the field via radio telemetry. Although juvenile and adult behaviors were similar in most respects, we did find that adults chose more effective ambush sites, which may be due to their increased experience. We also found that juveniles (but typically not adults) perform periodic tail undulations while in ambush, and that juveniles displayed slightly different activity cycles. Both of these latter differences are likely the result of age-specific adaptations for juveniles' greater reliance on lizards versus small mammals as prey. We also compared the general predatory behavior of sidewinders to that of other species in the genus Crotalus. These findings will provide important baseline field information for more detailed empirical research on the ontogeny of predation behavior in precocial vertebrates. Copyright © 2016

Uracil excision repair in Mycobacterium tuberculosis cell-free extracts.

Science.gov (United States)

Kumar, Pradeep; Bharti, Sanjay Kumar; Varshney, Umesh

2011-05-01

Uracil excision repair is ubiquitous in all domains of life and initiated by uracil DNA glycosylases (UDGs) which excise the promutagenic base, uracil, from DNA to leave behind an abasic site (AP-site). Repair of the resulting AP-sites requires an AP-endonuclease, a DNA polymerase, and a DNA ligase whose combined activities result in either short-patch or long-patch repair. Mycobacterium tuberculosis, the causative agent of tuberculosis, has an increased risk of accumulating uracils because of its G + C-rich genome, and its niche inside host macrophages where it is exposed to reactive nitrogen and oxygen species, two major causes of cytosine deamination (to uracil) in DNA. In vitro assays to study DNA repair in this important human pathogen are limited. To study uracil excision repair in mycobacteria, we have established assay conditions using cell-free extracts of M. tuberculosis and M. smegmatis (a fast-growing mycobacterium) and oligomer or plasmid DNA substrates. We show that in mycobacteria, uracil excision repair is completed primarily via long-patch repair. In addition, we show that M. tuberculosis UdgB, a newly characterized family 5 UDG, substitutes for the highly conserved family 1 UDG, Ung, thereby suggesting that UdgB might function as backup enzyme for uracil excision repair in mycobacteria. Copyright © 2011 Elsevier Ltd. All rights reserved.

Spore coat protein synthesis in cell-free systems from sporulating cells of Bacillus subtilis.

Science.gov (United States)

Nakayama, T; Munoz, L E; Sadaie, Y; Doi, R H

1978-09-01

Cell-free systems for protein synthesis were prepared from Bacillus subtilis 168 cells at several stages of sporulation. Immunological methods were used to determine whether spore coat protein could be synthesized in the cell-free systems prepared from sporulating cells. Spore coat protein synthesis first occurred in extracts from stage t2 cells. The proportion of spore coat protein to total proteins synthesized in the cell-free systems was 2.4 and 3.9% at stages t2 and t4, respectively. The sodium dodecyl sulfate-urea-polyacrylamide gel electrophoresis patterns of immunoprecipitates from the cell-free systems showed the complete synthesis of an apparent spore coat protein precursor (molecular weight, 25,000). A polypeptide of this weight was previously identified in studies in vivo (L.E. Munoz, Y. Sadaie, and R.H. Doi, J. Biol. Chem., in press). The synthesis in vitro of polysome-associated nascent spore coat polypeptides with varying molecular weights up to 23,000 was also detected. These results indicate that the spore coat protein may be synthesized as a precursor protein. The removal of proteases in the crude extracts by treatment with hemoglobin-Sepharose affinity techniques may be preventing the conversion of the large 25,000-dalton precursor to the 12,500-dalton mature spore coat protein.

Estragole and methyl-eugenol-free extract of Artemisia dracunculus possesses immunomodulatory effects

Directory of Open Access Journals (Sweden)

Seyyed Meysam Abtahi Froushani

2016-08-01

Full Text Available Objective: Some evidence suggests that chronic uptake of estragole and methyl-eugenol, found in the essential oil of Artemisia dracunculus (tarragon, may be associated with an increased risk of hepato-carcinogenicity. The present study was conducted to investigate the immumodulatory and anti-inflammatory potentials of estragole and methyl-eugenol free extract of tarragon. Materials and Methods: Aqueous, hydroalcoholic, methanol and hexane extracts of dried and milled tarragon was prepared and analyzed by GC-MS. The estragole and methyl-eugenol free extract was characterized and used for evaluation of immunity in NMRI mice after challenging with sheep red blood cells. Results: It was shown that the aqueous extract of tarragon was free from potentially harmful estragole or methyl-eugenol. Moreover, the immunomodulatory effect of the aqueous extract of tarragon (100 mg/kg for 21 consecutive days was investigated. The extract significantly increased the level of anti-sheep red blood cells (SRBC (antibody and simultaneously decreased the level of cellular immunity in the treatment group. Moreover, tarragon caused a significant reduction in the production of pro-inflammatory IL-17 and IFN-γ in parallel with a reduction in the ratio of INF-γ to Il-10 or IL-17 to IL-10 in the splenocytes. In addition, the levels of the respiratory burst and nitric oxide production in peritoneal macrophages were significantly decreased. Additionally, the phagocytosis potential of macrophages was significantly increased in treated mice. Conclusion: These data showed that the aqueous extract of tarragon may be used as a natural source to modulate the immune system, because it can inhibit pro-inflammatory cytokines and induce anti-inflammatory macrophages.

Graded Heterojunction Engineering for Hole-Conductor-Free Perovskite Solar Cells with High Hole Extraction Efficiency and Conductivity.

Science.gov (United States)

Li, Bo; Zhang, Yanan; Zhang, Luyuan; Yin, Longwei

2017-10-01

Despite great progress in the photovoltaic conversion efficiency (PCE) of inorganic-organic hybrid perovskite solar cells (PSCs), the large-scale application of PSCs still faces serious challenges due to the poor-stability and high-cost of the spiro-OMeTAD hole transport layer (HTL). It is of great fundamental importance to rationally address the issues of hole extraction and transfer arising from HTL-free PSCs. Herein, a brand-new PSC architecture is designed by introducing multigraded-heterojunction (GHJ) inorganic perovskite CsPbBr x I 3- x layers as an efficient HTL. The grade adjustment can be achieved by precisely tuning the halide proportion and distribution in the CsPbBr x I 3- x film to reach an optimal energy alignment of the valance and conduction band between MAPbI 3 and CsPbBr x I 3- x . The CsPbBr x I 3- x GHJ as an efficient HTL can induce an electric field where a valance/conduction band edge is leveraged to bend at the heterojunction interface, boosting the interfacial electron-hole splitting and photoelectron extraction. The GHJ architecture enhances the hole extraction and conduction efficiency from the MAPbI 3 to the counter electrode, decreases the recombination loss during the hole transfer, and benefits in increasing the open-circuit voltage. The optimized HTL-free PCS based on the GHJ architecture demonstrates an outstanding thermal stability and a significantly improved PCE of 11.33%, nearly 40% increase compared with 8.16% for pure HTL-free devices. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Roads, interrupted dispersal, and genetic diversity in timber rattlesnakes.

Science.gov (United States)

Clark, Rulon W; Brown, William S; Stechert, Randy; Zamudio, Kelly R

2010-08-01

Anthropogenic habitat modification often creates barriers to animal movement, transforming formerly contiguous habitat into a patchwork of habitat islands with low connectivity. Roadways are a feature of most landscapes that can act as barriers or filters to migration among local populations. Even small and recently constructed roads can have a significant impact on population genetic structure of some species, but not others. We developed a research approach that combines fine-scale molecular genetics with behavioral and ecological data to understand the impacts of roads on population structure and connectivity. We used microsatellite markers to characterize genetic variation within and among populations of timber rattlesnakes (Crotalus horridus) occupying communal hibernacula (dens) in regions bisected by roadways. We examined the impact of roads on seasonal migration, genetic diversity, and gene flow among populations. Snakes in hibernacula isolated by roads had significantly lower genetic diversity and higher genetic differentiation than snakes in hibernacula in contiguous habitat. Genetic-assignment analyses revealed that interruption to seasonal migration was the mechanism underlying these patterns. Our results underscore the sizeable impact of roads on this species, despite their relatively recent construction at our study sites (7 to 10 generations of rattlesnakes), the utility of population genetics for studies of road ecology, and the need for mitigating effects of roads.

Gross and histologic features of gastritis due to Ophidascaris arndti in tropical rattlesnakes ( Crotalus durissus

Directory of Open Access Journals (Sweden)

É.M. Mello

Full Text Available ABSTRACT The tropical rattlesnake (Crotalus durissus is a snake of great importance for biomedical industry since its poison is used for the production of antiophidic serum and researches. Several conditions related to animal health, such as diseases and parasites, which can promote the reduction of poison production by these snakes should be investigated. Accordingly, the aim of this study was to characterize the gross and microscopic lesions related to the presence of Ophidascaris arndti in stomachs of tropical rattlesnakes. The gastrointestinal tract of thirty-five South American rattlesnakes captured in Southeastern region of Brazil were analyzed and nineteen animals showed infestation by the parasites, found in the small intestine and, especially, in the stomach of the hosts. Grossly, lesions were characterized by mucosal ulcers occasionally associated with hemorrhage. Microscopic alterations included histiocytic granulomas, fibrosis, necrosis, and hemorrhage. Based on these findings, the diagnosis of a parasitic granulomatous disease was made. The lesions may be related to the cause of death in captivity snakes, since the lesions can promote secondary infections by opportunistic bacteria. Moreover, the intense inflammatory response accompanied by fibrosis may be related to poor functioning of the gastric snakes, which it may exhibit frequent regurgitation of the food.

Controlling cell-free metabolism through physiochemical perturbations.

Science.gov (United States)

Karim, Ashty S; Heggestad, Jacob T; Crowe, Samantha A; Jewett, Michael C

2018-01-01

Building biosynthetic pathways and engineering metabolic reactions in cells can be time-consuming due to complexities in cellular metabolism. These complexities often convolute the combinatorial testing of biosynthetic pathway designs needed to define an optimal biosynthetic system. To simplify the optimization of biosynthetic systems, we recently reported a new cell-free framework for pathway construction and testing. In this framework, multiple crude-cell extracts are selectively enriched with individual pathway enzymes, which are then mixed to construct full biosynthetic pathways on the time scale of a day. This rapid approach to building pathways aids in the study of metabolic pathway performance by providing a unique freedom of design to modify and control biological systems for both fundamental and applied biotechnology. The goal of this work was to demonstrate the ability to probe biosynthetic pathway performance in our cell-free framework by perturbing physiochemical conditions, using n-butanol synthesis as a model. We carried out three unique case studies. First, we demonstrated the power of our cell-free approach to maximize biosynthesis yields by mapping physiochemical landscapes using a robotic liquid-handler. This allowed us to determine that NAD and CoA are the most important factors that govern cell-free n-butanol metabolism. Second, we compared metabolic profile differences between two different approaches for building pathways from enriched lysates, heterologous expression and cell-free protein synthesis. We discover that phosphate from PEP utilization, along with other physiochemical reagents, during cell-free protein synthesis-coupled, crude-lysate metabolic system operation inhibits optimal cell-free n-butanol metabolism. Third, we show that non-phosphorylated secondary energy substrates can be used to fuel cell-free protein synthesis and n-butanol biosynthesis. Taken together, our work highlights the ease of using cell-free systems to explore

A study of bacterial contamination of rattlesnake venom

Directory of Open Access Journals (Sweden)

E. Garcia-Lima

1987-03-01

Full Text Available The authors studied the bacterial contamination of rattlesnake venom isolated from snakes in captivity and wild snakes caught recently. The captive snakes showed a relatively high incidence of bacterial contamination of their venom.Os autores estudaram a contaminação bacteriana do veneno dë cascavéis mantidas em cativeiro e das recentemente capturadas. Verificaram que os venenos dos animais cativos apresentaram alta incidência de contaminação e os tidos como recentemente capturados estavam com baixa contaminação aparente.

Extraction method for high free radical scavenging activity of Siamese neem tree flowers

Directory of Open Access Journals (Sweden)

Worarat Chaisawangwong

2009-10-01

Full Text Available Siamese neem tree (Azadirachta indica A. Juss. var. siamensis Valeton is a medicinal plant found in Thailand. Youngleaves and young flowers of this plant are commonly consumed as a bitter tonic vegetable. The flowers are also used fortreatment of fever. The flower extract has been reported to exhibit in vitro free radical scavenging activity and can inhibitlipid peroxidation of bronchogenic cancer cell line. Active compounds in the flowers are flavonoids such as rutin andquercetin. The content of these compounds in the crude extract depends on the method of extraction. Therefore, the appropriateextraction method promoting high yield of total flavonoids and high free radical scavenging activity was investigated inthis study. Six different extraction methods, i.e. maceration, percolation, decoction, soxhlet extraction, ultrasonic extraction(UE, and microwave assisted extraction (MA were carried out for extracting dried powder of Siamese neem tree young flowers. The solvent used for maceration, percolation, and soxhlet extraction was 50% ethanol, while distilled water was used for decoction and MA, and both solvents were used for UE. The content of crude extract, free radical scavenging activity, and total flavonoids content of each extract were investigated and compared. Comparing the various extraction methods, decoction provided an extract containing a high amount of total flavonoids (17.54 mgRE/g extract and promoting the highest scavenging activity at EC50 11.36 g/ml. Decoction is also simple, cheap, and convenient and could be used in developing countries. Thus, it should be the recommended extraction method for the flowers of Siamese neem tree for furtherdevelopment of antioxidant pharmaceutical preparations.

Morulustatin, A Disintegrin that Inhibits ADP-Induced Platelet Aggregation, Isolated from the Mexican Tamaulipan Rock Rattlesnake (Crotalus lepidus morulus)

Science.gov (United States)

Borja, Miguel; Galan, Jacob Anthony; Cantu, Esteban; Zugasti-Cruz, Alejandro; Rodríguez-Acosta, Alexis; Lazcano, David; Lucena, Sara; Suntravat, Montamas; Sánchez, y Elda Eliza

2016-01-01

The Tamaulipan rock rattlesnake (Crotalus lepidus morulus) is a montane snake that occurs in the humid pine-oak forest and the upper cloud forest of the Sierra Madre Oriental in southwestern Tamaulipas, central Nuevo Leon, and southeastern Coahuila in Mexico. Venom from this rattlesnake was fractionated by High-Performance Liquid Chromatography for the purpose of discovering disintegrin molecules. Disintegrins are non-enzymatic, small molecular weight peptides that interfere with cell-cell and cell-matrix interactions by binding to various cell receptors. Eleven fractions were collected by anion exchange chromatography and pooled into six groups (I, II, III, IV, V, and VI). Proteins of the six groups were analyzed by SDS-PAGE and western blot using antibodies raised against a disintegrin. The antibodies recognized different protein bands in five (II, III, IV, V, and VI) of six groups in a molecular mass range of 7 to 105 kDa. Western blot analysis revealed fewer protein bands in the higher molecular mass range and two bands in the disintegrin weight range in group II compared with the other four groups. Proteins in group II were further separated into nine fractions using reverse phase C18 chromatography. Fraction 4 inhibited platelet aggregation and was named morulustatin, which exhibited a single band with a molecular mass of approximately 7 kDa. Mass spectrometry analysis of fraction 4 revealed the identification of disintegrin peptides LRPGAQCADGLCCDQCR (MH+ 2035.84) and AGEECDCGSPANCCDAATCK (MH+ 2328.82). Morulustatin inhibited ADP-induced platelet aggregation in human whole blood and was concentration-dependent with an IC50 of 89.5 nM ± 12. PMID:28713196

Assessing the influence of mechanical ventilation on blood gases and blood pressure in rattlesnakes

DEFF Research Database (Denmark)

Bertelsen, Mads Frost; Buchanan, Rasmus; Jensen, Heidi Meldgaard

2014-01-01

OBJECTIVE: To characterize the impact of mechanical positive pressure ventilation on heart rate (HR), arterial blood pressure, blood gases, lactate, glucose, sodium, potassium and calcium concentrations in rattlesnakes during anesthesia and the subsequent recovery period. STUDY DESIGN: Prospectiv...

Wound healing potential of adipose tissue stem cell extract.

Science.gov (United States)

Na, You Kyung; Ban, Jae-Jun; Lee, Mijung; Im, Wooseok; Kim, Manho

2017-03-25

Adipose tissue stem cells (ATSCs) are considered as a promising source in the field of cell therapy and regenerative medicine. In addition to direct cell replacement using stem cells, intercellular molecule exchange by stem cell secretory factors showed beneficial effects by reducing tissue damage and augmentation of endogenous repair. Delayed cutaneous wound healing is implicated in many conditions such as diabetes, aging, stress and alcohol consumption. However, the effects of cell-free extract of ATSCs (ATSC-Ex) containing secretome on wound healing process have not been investigated. In this study, ATSC-Ex was topically applied on the cutaneous wound and healing speed was examined. As a result, wound closure was much faster in the cell-free extract treated wound than control wound at 4, 6, 8 days after application of ATSC-Ex. Dermal fibroblast proliferation, migration and extracellular matrix (ECM) production are critical aspects of wound healing, and the effects of ATSC-Ex on human dermal fibroblast (HDF) was examined. ATSC-Ex augmented HDF proliferation in a dose-dependent manner and migration ability was enhanced by extract treatment. Representative ECM proteins, collagen type I and matrix metalloproteinase-1, are significantly up-regulated by treatment of ATSC-Ex. Our results suggest that the ATSC-Ex have improving effect of wound healing and can be the potential therapeutic candidate for cutaneous wound healing. Copyright © 2017 Elsevier Inc. All rights reserved.

The impact of roads on the timber rattlesnake (Crotalus horridus) in eastern Texas

Science.gov (United States)

D. Craig Rudolph; Shirley J. Burgdorf; Richard N. Conner; James G. Dickson

1998-01-01

Roads and associated vehicular traffic have the potential to significantly impact vertebrate populations. In eastern Texas we compared the densities of paved and unpaved roads within 2 and 4 km radii of timber rattlesnake (Crotalus horridus) ocations and of random points. Road networks were significantly more dense at random points than at snake...

Assess Current and Potential Salmonid Production in Rattlesnake Creek Associated with Restoration Efforts; US Geological Survey Reports, 2002-2003 Annual Report.

Energy Technology Data Exchange (ETDEWEB)

Connolly, Patrick J. (US Geological Survey, Columbia River Research Laboratory, Western Fisheries Research Center, Cook, WA)

2003-12-01

This project was designed to document existing habitat conditions and fish populations within the Rattlesnake Creek watershed (White Salmon River subbasin, Washington) before major habitat restoration activities are implemented and prior to the reintroduction of salmon and steelhead above Condit Dam. Returning adult salmon Oncorhynchus spp. and steelhead O. mykiss have not had access to Rattlesnake Creek since 1913. An assessment of resident trout populations should serve as a good surrogate for evaluation of factors that would limit salmon and steelhead production in the watershed. Personnel from United States Geological Survey's Columbia River Research Laboratory (USGS-CRRL) attend to three main objectives of the Rattlesnake Creek project. The first is to characterize stream and riparian habitat conditions. This effort includes measures of water quality, water quantity, stream habitat, and riparian conditions. The second objective is to determine the status of fish populations in the Rattlesnake Creek drainage. To accomplish this, we derived estimates of salmonid population abundance, determined fish species composition, assessed distribution and life history attributes, obtained tissue samples for genetic analysis, and assessed fish diseases in the watershed. The third objective is to use the collected habitat and fisheries information to help identify and prioritize areas in need of restoration. As this report covers the second year of at least a three-year study, it is largely restricted to describing our efforts and findings for the first two objectives.

Mangifera indica L. extract protects T cells from activation-induced cell death.

Science.gov (United States)

Hernández, Patricia; Delgado, Rene; Walczak, Henning

2006-09-01

The aqueous stem bark extract of Mangifera indica L. (Vimang) has been reported to have antioxidant properties. AIDS is characterized by up-regulation of CD95 ligand (CD95L) expression and enhancement of activation-induced cell death (AICD). Recent studies demonstrate oxidative signals combined with simultaneous calcium (Ca(2+)) influx into the cytosol are required for induction of CD95L expression. In this study we show that M. indica extract attenuated anti-CD3-induced accumulation of reactive oxygen species (ROS) and intracellular free Ca(2+) and consequently, downregulates CD95L mRNA expression and CD95-mediated AICD. In addition, TCR triggering caused an elevation in the antioxidant enzyme manganous superoxide dismutase (Mn-SOD) and the increase in c-Jun N-terminal kinase (JNK) phosphorylation, both effects being prevented by M. indica extract. We provide a number of evidences regarding how M. indica extract enhance T-cell survival by inhibiting AICD, a finding associated with a decrease in oxidative stress generated through the TCR signaling pathway in activated T cells.

Draft Genome Sequence of Clostridium mangenotii TR, Isolated from the Fecal Material of a Timber Rattlesnake

Science.gov (United States)

Cochran, Philip A.; Dowd, Scot E.; Andersen, Kylie; Anderson, Nichole; Brennan, Rachel; Brook, Nicole; Callaway, Tracie; Diamante, Kimberly; Duberstine, Annie; Fitch, Karla; Freiheit, Heidi; Godlewski, Chantel; Gorman, Kelly; Haubrich, Mark; Hernandez, Mercedes; Hirtreiter, Amber; Ivanoski, Beth; Jaminet, Xochitl; Kirkpatrick, Travis; Kratowicz, Jennifer; Latus, Casey; Leable, Tiegen; Lingafelt, Nicole; Lowe, DeAnna; Lowrance, Holly; Malsack, Latiffa; Mazurkiewicz, Julie; Merlos, Persida; Messley, Jamie; Montemurro, Dawn; Nakitare, Samora; Nelson, Christine; Nye, Amber; Pazera, Valerie; Pierangeli, Gina; Rellora, Ashley; Reyes, Angelica; Roberts, Jennifer; Robins, Shadara; Robinson, Jeshannah; Schultz, Alissa; Seifert, Sara; Sigler, Elona; Spangler, Julie; Swift, Ebony; TenCate, Rebecca; Thurber, Jessica; Vallee, Kristin; Wamboldt, Jennifer; Whitten, Shannon; Woods, De’andrea; Wright, Amanda; Yankunas, Darin

2014-01-01

Here, we report the draft genome sequence of Clostridium mangenotii strain TR, which was isolated from the fecal material of a timber rattlesnake. This bacterium is nonpathogenic but contains 68 genes involved in virulence, disease, and defense. PMID:24407632

Aloe vera extract activity on human corneal cells.

Science.gov (United States)

Woźniak, Anna; Paduch, Roman

2012-02-01

Ocular diseases are currently an important problem in modern societies. Patients suffer from various ophthalmologic ailments namely, conjunctivitis, dry eye, dacryocystitis or degenerative diseases. Therefore, there is a need to introduce new treatment methods, including medicinal plants usage. Aloe vera [Aloe barbadensis Miller (Liliaceae)] possesses wound-healing properties and shows immunomodulatory, anti-inflammatory or antioxidant activities. NR uptake, MTT, DPPH• reduction, Griess reaction, ELISA and rhodamine-phalloidin staining were used to test toxicity, antiproliferative activity, reactive oxygen species (ROS) reduction, nitric oxide (NO) and cytokine level, and distribution of F-actin in cells, respectively. The present study analyzes the effect of Aloe vera extracts obtained with different solvents on in vitro culture of human 10.014 pRSV-T corneal cells. We found no toxicity of ethanol, ethyl acetate and heptane extracts of Aloe vera on human corneal cells. No ROS reducing activity by heptane extract and trace action by ethanol (only at high concentration 125 µg/ml) extract of Aloe vera was observed. Only ethyl acetate extract expressed distinct free radical scavenging effect. Plant extracts decreased NO production by human corneal cells as compared to untreated controls. The cytokine (IL-1β, IL-6, TNF-α and IL-10) production decreased after the addition of Aloe vera extracts to the culture media. Aloe vera contains multiple pharmacologically active substances which are capable of modulating cellular phenotypes and functions. Aloe vera ethanol and ethyl acetate extracts may be used in eye drops to treat inflammations and other ailments of external parts of the eye such as the cornea.

Assess Current and Potential Salmonid Production in Rattlesnake Creek in Association with Restoration Efforts, US Geological Survey Report, 2004-2005 Annual Report.

Energy Technology Data Exchange (ETDEWEB)

Allen, M. Brady; Connolly, Patrick J.; Jezorek, Ian G. (US Geological Survey, Western Fisheries Research Center, Columbia River Research Laboratory, Cook, WA)

2006-06-01

This project was designed to document existing habitat conditions and fish populations within the Rattlesnake Creek watershed (White Salmon River subbasin, Washington) before major habitat restoration activities are implemented and prior to the reintroduction of salmon and steelhead above Condit Dam. Returning adult salmon Oncorhynchus spp. and steelhead O. mykiss have not had access to Rattlesnake Creek since 1913. An assessment of resident trout populations should serve as a good surrogate for evaluation of factors that would limit salmon and steelhead production in the watershed. Personnel from United States Geological Survey's Columbia River Research Laboratory (USGS-CRRL) attended to three main objectives of the Rattlesnake Creek project. The first objective was to characterize stream and riparian habitat conditions. This effort included measures of water quality, water quantity, stream habitat, and riparian conditions. The second objective was to determine the status of fish populations in the Rattlesnake Creek drainage. To accomplish this, we derived estimates of salmonid population abundance, determined fish species composition, assessed distribution and life history attributes, obtained tissue samples for genetic analysis, and assessed fish diseases in the watershed. The third objective was to use the collected habitat and fisheries information to help identify and prioritize areas in need of restoration. As this report covers the fourth year of a five-year study, it is largely restricted to describing our efforts and findings for the first two objectives.

Assess Current and Potential Salmonid Production in Rattlesnake Creek in Association with Restoration Effors; US Geological Survey Reports, 2003-2004 Annual Report.

Energy Technology Data Exchange (ETDEWEB)

Allen, M. Brady; Connolly, Patrick J.; Munz, Carrie S. (US Geological Survey, Western Fisheries Research Center, Columbia River Research Laboratory, Cook, WA)

2006-02-01

This project was designed to document existing habitat conditions and fish populations within the Rattlesnake Creek watershed (White Salmon River subbasin, Washington) before major habitat restoration activities are implemented and prior to the reintroduction of salmon and steelhead above Condit Dam. Returning adult salmon Oncorhynchus spp. and steelhead O. mykiss have not had access to Rattlesnake Creek since 1913. An assessment of resident trout populations should serve as a good surrogate for evaluation of factors that would limit salmon and steelhead production in the watershed. Personnel from United States Geological Survey's Columbia River Research Laboratory (USGS-CRRL) attend to three main objectives of the Rattlesnake Creek project. The first is to characterize stream and riparian habitat conditions. This effort includes measures of water quality, water quantity, stream habitat, and riparian conditions. The second objective is to determine the status of fish populations in the Rattlesnake Creek drainage. To accomplish this, we derived estimates of salmonid population abundance, determined fish species composition, assessed distribution and life history attributes, obtained tissue samples for genetic analysis, and assessed fish diseases in the watershed. The third objective was to use the collected habitat and fisheries information to help identify and prioritize areas in need of restoration. As this report covers the third year of at least a five-year study, it is largely restricted to describing our efforts and findings for the first two objectives.

Wound healing potential of adipose tissue stem cell extract

International Nuclear Information System (INIS)

Na, You Kyung; Ban, Jae-Jun; Lee, Mijung; Im, Wooseok; Kim, Manho

2017-01-01

Adipose tissue stem cells (ATSCs) are considered as a promising source in the field of cell therapy and regenerative medicine. In addition to direct cell replacement using stem cells, intercellular molecule exchange by stem cell secretory factors showed beneficial effects by reducing tissue damage and augmentation of endogenous repair. Delayed cutaneous wound healing is implicated in many conditions such as diabetes, aging, stress and alcohol consumption. However, the effects of cell-free extract of ATSCs (ATSC-Ex) containing secretome on wound healing process have not been investigated. In this study, ATSC-Ex was topically applied on the cutaneous wound and healing speed was examined. As a result, wound closure was much faster in the cell-free extract treated wound than control wound at 4, 6, 8 days after application of ATSC-Ex. Dermal fibroblast proliferation, migration and extracellular matrix (ECM) production are critical aspects of wound healing, and the effects of ATSC-Ex on human dermal fibroblast (HDF) was examined. ATSC-Ex augmented HDF proliferation in a dose-dependent manner and migration ability was enhanced by extract treatment. Representative ECM proteins, collagen type I and matrix metalloproteinase-1, are significantly up-regulated by treatment of ATSC-Ex. Our results suggest that the ATSC-Ex have improving effect of wound healing and can be the potential therapeutic candidate for cutaneous wound healing. - Highlights: • Topical application of ATSC-Ex results in faster wound closure than normal wound in vivo. • ATSC-Ex enhances dermal fibroblast proliferation, migration and extracellular matrix production. • This study suggests that ATSC-Ex is an effective source to augment wound healing.

Solvent-free microwave extraction of essential oil from Melaleuca leucadendra L.

Directory of Open Access Journals (Sweden)

Widya Ismanto Aviarina

2018-01-01

Full Text Available Cajuput (Melaleuca leucadendra L. oil is one of potential commodity that provides an important role for the country’s foreign exchange but the extraction of these essential oil is still using conventional method such as hydrodistillation which takes a long time to produce essential oil with good quality. Therefore it is necessary to optimize the extraction process using a more effective and efficient method. So in this study the extraction is done using solvent-free microwave extraction method that are considered more effective and efficient than conventional methods. The optimum yield in the extraction of cajuput oil using solvent-free microwave extraction method is 1.0674%. The optimum yield is obtained on the feed to distiller (F/D ratio of 0.12 g/mL with microwave power of 400 W. In the extraction of cajuput oil using solvent-free microwave extraction method is performed first-order and second-order kinetics modelling. Based on kinetics modelling that has been done, it can be said that the second-order kinetic model (R2 = 0.9901 can be better represent experimental results of extraction of cajuput oil that using solvent-free microwave extraction method when compared with the first-order kinetic model (R2 = 0.9854.

Apoptotic and free radical scavenging properties of the methanolic extract of Gentianella alborosea.

Science.gov (United States)

Acero, Nuria; Llinares, Francisco; Galán de Mera, Antonio; Oltra, Beatriz; Muñoz-Mingarro, Dolores

2006-09-01

Gentianella alborosea ("Hercampure") is a Peruvian species used in folk medicine for the treatment of a variety of health disorders. We tested the free radical scavenging (DPPH) and induction of apoptosis on a human uterus tumor cell line (HeLa) by its methanolic extract. The results showed a noticeable radical scavenging activity and a dose-dependent apoptotic effect.

Extracts from Calendula officinalis offer in vitro protection against H2 O2 induced oxidative stress cell killing of human skin cells.

Science.gov (United States)

Alnuqaydan, Abdullah M; Lenehan, Claire E; Hughes, Rachel R; Sanderson, Barbara J

2015-01-01

The in vitro safety and antioxidant potential of Calendula officinalis flower head extracts was investigated. The effect of different concentrations (0.125, 0.5, 1.0, 2.0 and 5.0% (v/v)) of Calendula extracts on human skin cells HaCaT in vitro was explored. Doses of 1.0% (v/v) (0.88 mg dry weight/mL) or less showed no toxicity. Cells were also exposed to the Calendula extracts for either 4, 24 or 48 h before being exposed to an oxidative insult (hydrogen peroxide H2 O2 ) for 1 h. Using the MTT cytotoxicity assay, it was observed that two independent extracts of C. officinalis gave time-dependent and concentration-dependent H2 O2 protection against induced oxidative stress in vitro using human skin cells. Pre-incubation with the Calendula extracts for 24 and 48 h increased survival relative to the population without extract by 20% and 40% respectively following oxidative challenge. The antioxidant potential of the Calendula extracts was confirmed using a complimentary chemical technique, the DPPH(●) assay. Calendula extracts exhibited free radical scavenging abilities. This study demonstrates that Calendula flower extracts contain bioactive and free radical scavenging compounds that significantly protect against oxidative stress in a human skin cell culture model. Copyright © 2014 John Wiley & Sons, Ltd.

Interface engineering for efficient fullerene-free organic solar cells

Energy Technology Data Exchange (ETDEWEB)

Shivanna, Ravichandran; Narayan, K. S., E-mail: rajaram@jncasr.ac.in, E-mail: narayan@jncasr.ac.in [Chemistry and Physics of Materials Unit, Jawaharlal Nehru Centre for Advanced Scientific Research, Bangalore 560064 (India); Rajaram, Sridhar, E-mail: rajaram@jncasr.ac.in, E-mail: narayan@jncasr.ac.in [International Centre for Materials Science, Jawaharlal Nehru Centre for Advanced Scientific Research, Bangalore 560064 (India)

2015-03-23

We demonstrate the role of zinc oxide (ZnO) morphology and addition of an acceptor interlayer to achieve high efficiency fullerene-free bulk heterojunction inverted organic solar cells. Nanopatterning of the ZnO buffer layer enhances the effective light absorption in the active layer, and the insertion of a twisted perylene acceptor layer planarizes and decreases the electron extraction barrier. Along with an increase in current homogeneity, the reduced work function difference and selective transport of electrons prevent the accumulation of charges and decrease the electron-hole recombination at the interface. These factors enable an overall increase of efficiency to 4.6%, which is significant for a fullerene-free solution-processed organic solar cell.

The extraction of essential oil from patchouli leaves (Pogostemon cablin Benth) using microwave hydrodistillation and solvent-free microwave extraction methods

Science.gov (United States)

Putri, D. K. Y.; Kusuma, H. S.; Syahputra, M. E.; Parasandi, D.; Mahfud, M.

2017-12-01

Patchouli plant (Pogostemon cablin Benth) is one of the important essential oil-producing plant, contributes more than 50% of total exports of Indonesia’s essential oil. However, the extraction of patchouli oil that has been done in Indonesia is generally still used conventional methods that require enormous amount of energy, high solvent usage, and long time of extraction. Therefore, in this study, patchouli oil extraction was carried out by using microwave hydrodistillation and solvent-free microwave extraction methods. Based on this research, it is known that the extraction of patchouli oil using microwave hydrodistillation method with longer extraction time (240 min) only produced patchouli oil’s yield 1.2 times greater than solvent-free microwave extraction method which require faster extraction time (120 min). Otherwise the analysis of electric consumption and the environmental impact, the solvent-free microwave extraction method showed a smaller amount when compared with microwave hydrodistillation method. It is conclude that the use of solvent-free microwave extraction method for patchouli oil extraction is suitably method as a new green technique.

Escherichia coli cell-free protein synthesis and isotope labeling of mammalian proteins.

Science.gov (United States)

Terada, Takaho; Yokoyama, Shigeyuki

2015-01-01

This chapter describes the cell-free protein synthesis method, using an Escherichia coli cell extract. This is a cost-effective method for milligram-scale protein production and is particularly useful for the production of mammalian proteins, protein complexes, and membrane proteins that are difficult to synthesize by recombinant expression methods, using E. coli and eukaryotic cells. By adjusting the conditions of the cell-free method, zinc-binding proteins, disulfide-bonded proteins, ligand-bound proteins, etc., may also be produced. Stable isotope labeling of proteins can be accomplished by the cell-free method, simply by using stable isotope-labeled amino acid(s) in the cell-free reaction. Moreover, the cell-free protein synthesis method facilitates the avoidance of stable isotope scrambling and dilution over the recombinant expression methods and is therefore advantageous for amino acid-selective stable isotope labeling. Site-specific stable isotope labeling is also possible with a tRNA molecule specific to the UAG codon. By the cell-free protein synthesis method, coupled transcription-translation is performed from a plasmid vector or a PCR-amplified DNA fragment encoding the protein. A milligram quantity of protein can be produced with a milliliter-scale reaction solution in the dialysis mode. More than a thousand solution structures have been determined by NMR spectroscopy for uniformly labeled samples of human and mouse functional domain proteins, produced by the cell-free method. Here, we describe the practical aspects of mammalian protein production by the cell-free method for NMR spectroscopy. © 2015 Elsevier Inc. All rights reserved.

Intraspecific venom variation in the medically significant Southern Pacific Rattlesnake (Crotalus oreganus helleri): biodiscovery, clinical and evolutionary implications.

Science.gov (United States)

Sunagar, Kartik; Undheim, Eivind A B; Scheib, Holger; Gren, Eric C K; Cochran, Chip; Person, Carl E; Koludarov, Ivan; Kelln, Wayne; Hayes, William K; King, Glenn F; Antunes, Agosthino; Fry, Bryan Grieg

2014-03-17

Due to the extreme variation of venom, which consequently results in drastically variable degrees of neutralization by CroFab antivenom, the management and treatment of envenoming by Crotalus oreganus helleri (the Southern Pacific Rattlesnake), one of the most medically significant snake species in all of North America, has been a clinician's nightmare. This snake has also been the subject of sensational news stories regarding supposed rapid (within the last few decades) evolution of its venom. This research demonstrates for the first time that variable evolutionary selection pressures sculpt the intraspecific molecular diversity of venom components in C. o. helleri. We show that myotoxic β-defensin peptides (aka: crotamines/small basic myotoxic peptides) are secreted in large amounts by all populations. However, the mature toxin-encoding nucleotide regions evolve under the constraints of negative selection, likely as a result of their non-specific mode of action which doesn't enforce them to follow the regime of the classic predator-prey chemical arms race. The hemorrhagic and tissue destroying snake venom metalloproteinases (SVMPs) were secreted in larger amounts by the Catalina Island and Phelan rattlesnake populations, in moderate amounts in the Loma Linda population and in only trace levels by the Idyllwild population. Only the Idyllwild population in the San Jacinto Mountains contained potent presynaptic neurotoxic phospholipase A2 complex characteristic of Mohave Rattlesnake (Crotalus scutulatus) and Neotropical Rattlesnake (Crotalus durissus terrificus). The derived heterodimeric lectin toxins characteristic of viper venoms, which exhibit a diversity of biological activities, including anticoagulation, agonism/antagonism of platelet activation, or procoagulation, appear to have evolved under extremely variable selection pressures. While most lectin α- and β-chains evolved rapidly under the influence of positive Darwinian selection, the β-chain lectin of

Airborne plutonium and americium concentrations measured from the top of Rattlesnake Mountain

International Nuclear Information System (INIS)

Sehmel, G.A.

1979-01-01

Airborne plutonium-239+240 and americium-241 blowing from offsite was measured in an initial experiment at the top of Rattlesnake Mountain. Average airborne concentration measured was similar to fallout concentrations. Airborne plutonium concentrations were independent of wind speed for seven wind speed increments between 0.5 and 31 m/sec. In contrast the airborne americium concentration was a minimum at a wind speed of approximately 7 m/sec. Similarly, the airborne solids concentration in μg/m 3 was a minimum at an intermediate wind speed increment of 7 to 11 m/sec

Electric shocks are ineffective in treatment of lethal effects of rattlesnake envenomation in mice.

Science.gov (United States)

Johnson, E K; Kardong, K V; Mackessy, S P

1987-01-01

Electrical shocks, even crudely delivered from 'stun guns' and gasoline engine spark plugs, have been reported to be effective in the treatment of snake bite. We thus applied similar electric shocks to mice artificially injected with reconstituted rattlesnake venom at various LD50 multiples. Those envenomated mice treated with electric shock survived no better than the controls. We thus found no evidence that electric shocks crudely administered had any life saving effect in mice.

Antioxidant and free radical scavenging activities of plant extracts ...

African Journals Online (AJOL)

Twenty-two species of medicinal plants collected in the Mexican state of Morelos were selected to evaluate their free radical scavenging and antioxidant activities. The extracts from the aerial parts of the plants were obtained using hexane, acetone and methanol (66 extracts). The initial qualitative screening of antioxidants ...

Analysis of Free Amino Acids in Different Extracts of Orthosiphon stamineus Leaves by High-Performance Liquid Chromatography Combined with Solid-Phase Extraction.

Science.gov (United States)

Shafaei, Armaghan; Halim, Nor Hidayah Ab; Zakaria, Norhidayah; Ismail, Zhari

2017-10-01

Orthosiphon stamineus (OS) Benth is a medicinal plant and native in Southeast Asia. Previous studies have shown that OS leaves possess antioxidant, cytotoxic, diuretic, antihypertensive, and uricosuric effects. These beneficial effects have been attributed to the presence of primary and secondary metabolites such as polyphenols, amino acids, and flavonoids. To develop and validate an high-performance liquid chromatography (HPLC)-diode array detector (DAD) method combined with solid-phase extraction that involves precolumn derivatization with O -phthaladehyde for simultaneous analysis of free amino acids in OS leaves extracts. OS leaves were extracted with water (OS-W), ethanol (OS-E), methanol (OS-M), 50% ethanol (OS-EW), and 50% methanol (OS-MW). The extracts were treated by C18 cartridge before derivatization, resulting in great improvement of separation by Zorbox Eclipse XDB-C 18 column. The HPLC-DAD method was successfully developed and validated for analyzing the contents of free amino acids in OS extracts. The results showed that l-aspartic acid with 0.93 ± 0.01 nmol/mg was the major free amino acid in OS-W extract. However, in OS-E, OS-M, OS-EW, and OS-MW, l-glutamic acid with 3.53 ± 0.16, 2.17 ± 0.10, 4.01 ± 0.12, and 2.49 ± 0.12 nmol/mg, respectively, was the major free amino acid. Subsequently, l-serine, which was detected in OS-W, OS-E, and OS-M, was the minor free amino acid with 0.33 ± 0.02, 0.12 ± 0.01, and 0.06 ± 0.01 nmol/mg, respectively. However, l-threonine with 0.26 ± 0.02 and 0.19 ± 0.08 nmol/mL in OS-EW and OS-MW, respectively, had the lowest concentration compared with other amino acid components. All validation parameters of the developed method indicate that the method is reliable and efficient to simultaneously determine the free amino acids content for routine analysis of OS extracts. The HPLC-DAD method combined with solid phase extraction was successfully developed and validated for simultaneous determination and

Bioreduction of trivalent aurum to nano-crystalline gold particles by active and inactive cells and cell-free extract of Aspergillus oryzae var. viridis

International Nuclear Information System (INIS)

Binupriya, A.R.; Sathishkumar, M.; Vijayaraghavan, K.; Yun, S.-I.

2010-01-01

Bioreduction efficacy of both active (AB) and inactive (IB) cells/biomass of Aspergillus oryzae var. viridis and their respective cell-free extracts (ACE and ICE) to convert trivalent aurum to gold nanoparticles were tested in the present study. Strong plasmon resonance of gold nanoparticles was observed between 540 and 560 nm in the samples obtained from AB, IB, ACE and ICE. Transmission electron microscopy (TEM), field emission scanning electron microscopy (FE-SEM), energy dispersive X-ray (EDX) and X-ray diffraction (XRD) were performed to examine the formation of gold nanoparticles. Comparing all four forms of A. oryzae var. viridis, ICE showed high gold nanoparticle productivity. The nanoparticles formed were quite uniform in shape and ranged in size from 10 to 60 nm. In addition some triangle, pentagon and hexagon-shaped nanoplates with size range of 30-400 nm were also synthesized especially at lower pH. Organics from the inactive cells are believed to be responsible for reduction of trivalent aurum to nano-sized gold particles. Organic content of the ICE was found to be double the amount of ACE. High productivity of gold nanoparticles by metabolic-independent process opens up an interesting area of nanoparticle synthesis using waste fungal biomass from industries.

Bioreduction of trivalent aurum to nano-crystalline gold particles by active and inactive cells and cell-free extract of Aspergillus oryzae var. viridis

Energy Technology Data Exchange (ETDEWEB)

Binupriya, A.R. [Department of Food Science and Technology, College of Agriculture and Life Science, Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Sathishkumar, M., E-mail: cvemuthu@nus.edu.sg [Singapore-Delft Water Alliance, National University of Singapore, 2 Engineering Drive 2, Singapore 117577 (Singapore); Vijayaraghavan, K. [Singapore-Delft Water Alliance, National University of Singapore, 2 Engineering Drive 2, Singapore 117577 (Singapore); Yun, S.-I., E-mail: siyun@chonbuk.ac.kr [Department of Food Science and Technology, College of Agriculture and Life Science, Chonbuk National University, Jeonju 561-756 (Korea, Republic of)

2010-05-15

Bioreduction efficacy of both active (AB) and inactive (IB) cells/biomass of Aspergillus oryzae var. viridis and their respective cell-free extracts (ACE and ICE) to convert trivalent aurum to gold nanoparticles were tested in the present study. Strong plasmon resonance of gold nanoparticles was observed between 540 and 560 nm in the samples obtained from AB, IB, ACE and ICE. Transmission electron microscopy (TEM), field emission scanning electron microscopy (FE-SEM), energy dispersive X-ray (EDX) and X-ray diffraction (XRD) were performed to examine the formation of gold nanoparticles. Comparing all four forms of A. oryzae var. viridis, ICE showed high gold nanoparticle productivity. The nanoparticles formed were quite uniform in shape and ranged in size from 10 to 60 nm. In addition some triangle, pentagon and hexagon-shaped nanoplates with size range of 30-400 nm were also synthesized especially at lower pH. Organics from the inactive cells are believed to be responsible for reduction of trivalent aurum to nano-sized gold particles. Organic content of the ICE was found to be double the amount of ACE. High productivity of gold nanoparticles by metabolic-independent process opens up an interesting area of nanoparticle synthesis using waste fungal biomass from industries.

Ultraviolet-endonuclease activity in cell extracts of Saccharomyces cerevisiae mutants defective in excision of pyrimidine dimers

International Nuclear Information System (INIS)

Bekker, M.L.; Kaboev, O.K.; Akhmedov, A.T.; Luchkina, L.A.

1980-01-01

Cell-free extracts of ultraviolet-sensitive mutants of Saccharomyces cerevisiae defective in excision of pyrimidine dimers, rad1, rad2, rad3, rad4, rad10, and rad16, as well as the extracts of the wild-type strain RAD+, display ultraviolet-endonuclease activity

Cell-free synthetic biology: thinking outside the cell.

Science.gov (United States)

Hodgman, C Eric; Jewett, Michael C

2012-05-01

Cell-free synthetic biology is emerging as a powerful approach aimed to understand, harness, and expand the capabilities of natural biological systems without using intact cells. Cell-free systems bypass cell walls and remove genetic regulation to enable direct access to the inner workings of the cell. The unprecedented level of control and freedom of design, relative to in vivo systems, has inspired the rapid development of engineering foundations for cell-free systems in recent years. These efforts have led to programmed circuits, spatially organized pathways, co-activated catalytic ensembles, rational optimization of synthetic multi-enzyme pathways, and linear scalability from the micro-liter to the 100-liter scale. It is now clear that cell-free systems offer a versatile test-bed for understanding why nature's designs work the way they do and also for enabling biosynthetic routes to novel chemicals, sustainable fuels, and new classes of tunable materials. While challenges remain, the emergence of cell-free systems is poised to open the way to novel products that until now have been impractical, if not impossible, to produce by other means. Copyright © 2011 Elsevier Inc. All rights reserved.

Cytotoxicity screening of Bangladeshi medicinal plant extracts on pancreatic cancer cells

Directory of Open Access Journals (Sweden)

Abbasi Atiya

2010-09-01

Full Text Available Abstract Background There has been a long standing interest in the identification of medicinal plants and derived natural products for developing cancer therapeutics. Our study focuses upon pancreatic cancer, due to its high mortality rate, that is attributed in part to the lack of an effective chemotherapeutic agent. Previous reports on the use of medicinal plant extracts either alone or alongside conventional anticancer agents in the treatment of this cancer have shown promising results. This work aims to investigate the therapeutic properties of a library of medicinal plants from Bangladesh. Methods 56 extracts of 44 unique medicinal plants were studied. The extracts were screened for cytotoxicity against the pancreatic adenocarcinoma cell line Panc-1, using a label-free biosensor assay. The top cytotoxic extracts identified in this screen were tested on two additional pancreatic cancer cell lines (Mia-Paca2 and Capan-1 and a fibroblast cell line (Hs68 using an MTT proliferation assay. Finally, one of the most promising extracts was studied using a caspase-3 colorimetric assay to identify induction of apoptosis. Results Crude extracts of Petunia punctata, Alternanthera sessilis, and Amoora chittagonga showed cytotoxicity to three cancer cell lines with IC50 values ranging between 20.3 - 31.4 μg/mL, 13.08 - 34.9 μg/mL, and 42.8 - 49.8 μg/mL, respectively. Furthermore, treatment of Panc-1 cells with Petunia punctata was shown to increase caspase-3 activity, indicating that the observed cytotoxicity was mediated via apoptosis. Only Amoora chittagonga showed low cytotoxicity to fibroblast cells with an IC50 value > 100 μg/mL. Conclusion Based upon the initial screening work reported here, further studies aimed at the identification of active components of these three extracts and the elucidation of their mechanisms as cancer therapeutics are warranted.

Stevia rebaudiana ethanolic extract exerts better antioxidant properties and antiproliferative effects in tumour cells than its diterpene glycoside stevioside.

Science.gov (United States)

López, Víctor; Pérez, Sergio; Vinuesa, Arturo; Zorzetto, Christian; Abian, Olga

2016-04-01

Steviol glycosides are currently being used as natural sweeteners by the food industry and Stevia rebaudiana has long been used as a sweet plant in South America for patients suffering from diabetes. In this study, a Stevia rebaudiana ethanolic extract (SREE) was prepared, analysed and tested for antioxidant activity in terms of free radical scavenging properties and antiproliferative effects in cervix (HeLa), pancreatic (MiaPaCa-2) and colonic (HCT116) cancer cells. The antiproliferative mechanism was confirmed by testing the effects on cyclin D1-CDK4. Bioassays were also performed for the diterpene glycoside stevioside. Our results demonstrate that the extract acts as an antioxidant being able to scavenge free radicals, but this activity was not due to stevioside. The extract also induced cell death in the three cell lines, being more active against cervix cancer cells (HeLa); however, the concentration of stevioside needed to produce antiproliferative effects was higher than the amount of steviol glycosides found in a lower dose of extract inducing cell death. In addition, the extract clearly inhibited CDK4 whereas stevioside did not, concluding that the antiproliferative activity of stevia may be due to inhibition of cyclin-dependent kinases performed by other compounds of the extract.

Vascular effects and electrolyte homeostasis of the natriuretic peptide isolated from Crotalus oreganus abyssus (North American Grand Canyon rattlesnake) venom

NARCIS (Netherlands)

Da Silva, S.L.; Dias-Junior, C.A.; Baldasso, P.A.; Damico, D.C.; Carvalho, B.M.; Garanto, A.; Acosta, G.; Oliveira, E.; Albericio, F.; Soares, A.M.; Marangoni, S.; Resende, R.R.

2012-01-01

Crotalus oreganus abyssus is a rattlesnake that is usually found in the Grand Canyon, United States of America. Knowledge regarding the composition of C. o. abyssus venom is scarce. New natriuretic peptides (NPs) have been isolated and characterized from the venoms of members of the Crotalinae

Partial characterization of the venom of the Peruvian rattlesnake Crotalus durissus terrificus

Directory of Open Access Journals (Sweden)

César Remuzgo

2014-06-01

Full Text Available The venom of the rattlesnake Crotalus durissus terrificus from the region of Sandia, Puno, has been investigated for its protein content and some enzymatic activities, using for it the whole venom as well as the fractions obtained by gel filtration chromatography in Sephadex G-100. The protein percentage calculated by the method of Lowry was of 68,6% for the whole venom; 3 peaks were obtained during the fractionation; the first showed proteolytic activity, the second, amidolytic, clotting and phospholipase A2 activities, while the third, another proteolytic activity. Acetylcholinesterase activity was not found while L-aminoacid oxidase activity was found only in the whole venom.

Sensationalistic journalism and tales of snakebite: are rattlesnakes rapidly evolving more toxic venom?

Science.gov (United States)

Hayes, William K; Mackessy, Stephen P

2010-03-01

Recent reports in the lay press have suggested that bites by rattlesnakes in the last several years have been more severe than those in the past. The explanation, often citing physicians, is that rattlesnakes are evolving more toxic venom, perhaps in response to anthropogenic causes. We suggest that other explanations are more parsimonious, including factors dependent on the snake and factors associated with the bite victim's response to envenomation. Although bites could become more severe from an increased proportion of bites from larger or more provoked snakes (ie, more venom injected), the venom itself evolves much too slowly to explain the severe symptoms occasionally seen. Increased snakebite severity could also result from a number of demographic changes in the victim profile, including age and body size, behavior toward the snake (provocation), anatomical site of bite, clothing, and general health including asthma prevalence and sensitivity to foreign antigens. Clinical management of bites also changes perpetually, rendering comparisons of snakebite severity over time tenuous. Clearly, careful study taking into consideration many factors will be essential to document temporal changes in snakebite severity or venom toxicity. Presently, no published evidence for these changes exists. The sensationalistic coverage of these atypical bites and accompanying speculation is highly misleading and can produce many detrimental results, such as inappropriate fear of the outdoors and snakes, and distraction from proven snakebite management needs, including a consistent supply of antivenom, adequate health care, and training. We urge healthcare providers to avoid propagating misinformation about snakes and snakebites. Copyright (c) 2010 Wilderness Medical Society. Published by Elsevier Inc. All rights reserved.

Free Radical Scavenging Activities of Methanol Extract and ...

African Journals Online (AJOL)

Michael Horsfall

fractions were subjected to in vitro antioxidant evaluation using the DPPH free radical scavenging ... The crude extract has IC-50 value for radical scavenging activity of 5μg/mL which was .... effective concentration of the sample that is required.

Molecular exclusion chromatographic analysis on 60Co irradiated rattlesnake

International Nuclear Information System (INIS)

Murata, Y.; Rogero, J.R.

1988-07-01

Ionizing radiations are sufficiently energetic to be able of breaking chemical bonds and as result of that molecules of substances present in the irradiated will be chemically changed and their biological properties affected also. In this paper the effects of gamma radiation of Co-60 on the rattlesnake venom were studied. A pool of positive crotamina Crotalus durissus terrificus venom was dissolved in 0,15 M NaCl and irradiated with Co-60. Doses of 100, 250, 500, 750, 1500 and 2000 Gy were applied at the dose rate of 1190 Gy/h. With doses over 500 Gy the solutions became turbid, suggesting the presence of aggregates and structural changes of the proteins. The concentration of proteins from the filtered solution was measured by the Lowry method. One ml samples were measured at 230 nm. The partition coefficients and the areas of the three main fraction obtained were calculated. (author) [pt

Hybrid PN-SN Calculations with SAAF for the Multiscale Transport Capability in Rattlesnake

Energy Technology Data Exchange (ETDEWEB)

Wang, Yaqi; Schunert, Sebastian; DeHart, Mark; Martineau, Richard

2016-05-01

Two interface conditions, the Lagrange multiplier method and the upwinding method, for hybrid \\pn-\\sn calculations is proposed for the self-adjoint angular flux (SAAF) formulation of the transport equation using the continuous finite element method (FEM) for spatial discretization. These interface conditions are implemented in Rattlesnake, the radiation transport application built on MOOSE, for the on-going multiscale transport simulation effort at INL. For smoothing the solution at the interface for the Lagrange multiplier method, a method based on \\sn Lagrange interpolation on the sphere is proposed. Numerical results indicate that the interface conditions give the expected convergence.

Effect of microbial cell-free meat extract on the growth of spoilage bacteria.

Science.gov (United States)

Nychas, G-J E; Dourou, D; Skandamis, P; Koutsoumanis, K; Baranyi, J; Sofos, J

2009-12-01

This study examined the effect of microbial cell-free meat extract (CFME) derived from spoiled meat, in which quorum sensing (QS) compounds were present, on the growth kinetics (lag phase, and growth rate) of two spoilage bacteria, Pseudomonas fluorescens and Serratia marcescens. Aliquots of CFME from spoiled meat were transferred to Brain Heart Infusion broth inoculated with 10(3) CFU ml(-1) of 18 h cultures of Ps. fluorescens or Ser. marcescens, both fresh meat isolates; CFME derived from unspoiled fresh meat ('clean' meat) served as a control. Changes in impedance measurements were monitored for 48 h, and the detection time (Tdet) was recorded. It was found that in the absence of CFME containing QS compounds the Tdet was shorter (P meat. The rate of growth of Ps. fluorescens, recorded as the maximum slope rate of conductance changes (MSrCC), after Tdet, was higher (P meat. Similar results in MSrCC of impedance changes were obtained for Ser. marcescens. The study indicated that the growth rate (expressed in MSrCC units) of meat spoilage bacteria in vitro was enhanced in samples supplemented with CFME containing QS compounds compared to control samples (i.e., without CFME or with CFME from 'clean' meat). This behaviour may explain the dominant role of these two bacteria in the spoilage of meat. These results illustrate the potential effect of signalling compounds released during storage of meat on the behaviour of meat spoilage bacteria. Understanding such interactions may assist in the control of fresh meat quality and the extension of its shelf life.

Efficient production of isotopically labeled proteins by cell-free synthesis: A practical protocol

Energy Technology Data Exchange (ETDEWEB)

Torizawa, Takuya; Shimizu, Masato [Crest, Jst (Japan); Taoka, Masato [Tokyo Metropolitan University, Graduate School of Science (Japan); Miyano, Hiroshi [Ajinomoto Co., Inc. Institute of Life Sciences (Japan); Kainosho, Masatsune [Crest, Jst (Japan)], E-mail: kainosho@nmr.chem.metro-u.ac.jp

2004-11-15

We provide detailed descriptions of our refined protocols for the cell-free production of labeled protein samples for NMR spectroscopy. These methods are efficient and overcome two critical problems associated with the use of conventional Escherichia coli extract systems. Endogenous amino acids normally present in E. coli S30 extracts dilute the added labeled amino acids and degrade the quality of NMR spectra of the target protein. This problem was solved by altering the protocol used in preparing the S30 extract so as to minimize the content of endogenous amino acids. The second problem encountered in conventional E. coli cell-free protein production is non-uniformity in the N-terminus of the target protein, which can complicate the NMR spectra. This problem was solved by adding a DNA sequence to the construct that codes for a cleavable N-terminal peptide tag. Addition of the tag serves to increase the yield of the protein as well as to ensure a homogeneous protein product following tag cleavage. We illustrate the method by describing its stepwise application to the production of calmodulin samples with different stable isotope labeling patterns for NMR analysis.

Efficient production of isotopically labeled proteins by cell-free synthesis: A practical protocol

International Nuclear Information System (INIS)

Torizawa, Takuya; Shimizu, Masato; Taoka, Masato; Miyano, Hiroshi; Kainosho, Masatsune

2004-01-01

We provide detailed descriptions of our refined protocols for the cell-free production of labeled protein samples for NMR spectroscopy. These methods are efficient and overcome two critical problems associated with the use of conventional Escherichia coli extract systems. Endogenous amino acids normally present in E. coli S30 extracts dilute the added labeled amino acids and degrade the quality of NMR spectra of the target protein. This problem was solved by altering the protocol used in preparing the S30 extract so as to minimize the content of endogenous amino acids. The second problem encountered in conventional E. coli cell-free protein production is non-uniformity in the N-terminus of the target protein, which can complicate the NMR spectra. This problem was solved by adding a DNA sequence to the construct that codes for a cleavable N-terminal peptide tag. Addition of the tag serves to increase the yield of the protein as well as to ensure a homogeneous protein product following tag cleavage. We illustrate the method by describing its stepwise application to the production of calmodulin samples with different stable isotope labeling patterns for NMR analysis

Free energy analysis of cell spreading.

Science.gov (United States)

McEvoy, Eóin; Deshpande, Vikram S; McGarry, Patrick

2017-10-01

In this study we present a steady-state adaptation of the thermodynamically motivated stress fiber (SF) model of Vigliotti et al. (2015). We implement this steady-state formulation in a non-local finite element setting where we also consider global conservation of the total number of cytoskeletal proteins within the cell, global conservation of the number of binding integrins on the cell membrane, and adhesion limiting ligand density on the substrate surface. We present a number of simulations of cell spreading in which we consider a limited subset of the possible deformed spread-states assumed by the cell in order to examine the hypothesis that free energy minimization drives the process of cell spreading. Simulations suggest that cell spreading can be viewed as a competition between (i) decreasing cytoskeletal free energy due to strain induced assembly of cytoskeletal proteins into contractile SFs, and (ii) increasing elastic free energy due to stretching of the mechanically passive components of the cell. The computed minimum free energy spread area is shown to be lower for a cell on a compliant substrate than on a rigid substrate. Furthermore, a low substrate ligand density is found to limit cell spreading. The predicted dependence of cell spread area on substrate stiffness and ligand density is in agreement with the experiments of Engler et al. (2003). We also simulate the experiments of Théry et al. (2006), whereby initially circular cells deform and adhere to "V-shaped" and "Y-shaped" ligand patches. Analysis of a number of different spread states reveals that deformed configurations with the lowest free energy exhibit a SF distribution that corresponds to experimental observations, i.e. a high concentration of highly aligned SFs occurs along free edges, with lower SF concentrations in the interior of the cell. In summary, the results of this study suggest that cell spreading is driven by free energy minimization based on a competition between decreasing

Full Core TREAT Kinetics Demonstration Using Rattlesnake/BISON Coupling Within MAMMOTH

Energy Technology Data Exchange (ETDEWEB)

Ortensi, Javier [Idaho National Lab. (INL), Idaho Falls, ID (United States); DeHart, Mark D. [Idaho National Lab. (INL), Idaho Falls, ID (United States); Gleicher, Frederick N. [Idaho National Lab. (INL), Idaho Falls, ID (United States); Wang, Yaqi [Idaho National Lab. (INL), Idaho Falls, ID (United States); Alberti, Anthony L. [Oregon State Univ., Corvallis, OR (United States); Palmer, Todd S. [Oregon State Univ., Corvallis, OR (United States)

2015-08-01

This report summarizes key aspects of research in evaluation of modeling needs for TREAT transient simulation. Using a measured TREAT critical measurement and a transient for a small, experimentally simplified core, Rattlesnake and MAMMOTH simulations are performed building from simple infinite media to a full core model. Cross sections processing methods are evaluated, various homogenization approaches are assessed and the neutronic behavior of the core studied to determine key modeling aspects. The simulation of the minimum critical core with the diffusion solver shows very good agreement with the reference Monte Carlo simulation and the experiment. The full core transient simulation with thermal feedback shows a significantly lower power peak compared to the documented experimental measurement, which is not unexpected in the early stages of model development.

Reactivation in UV inactivated Escherichia coli by cell-free extracts of propionic acid bacteria

International Nuclear Information System (INIS)

Vorob'eva, L.I.; Nikitenko, G.V.; Khodzhaev, E.Yu.; Ponomareva, G.M.

1993-01-01

For the first time reactivation of cell extraction of three strains of Propionibacterium shermanii in UV inactivated not filament-forming strain Escherichia colli AB 1157 is shown. Reactivation was demonstrated in prencubated and postincubated test-culture and increased as survival of E.coli decreased in a range 1,8-0,006%. The factor (factores) of defense in dialysable, thermolable and is present as in a fraction of nucleoproteins and nucleic acids so in a fraction of soluble proteins. The extracts were inactivated by incubation with proteinase K and trypsin, partly decreased activity by incubation with alpha-amylase and selected nuclease but not with lipase. Polypeltide nature of reactivative factor is supposed

Natural history of a northern population of twin-spotted rattlesnakes, Crotalus pricei

Science.gov (United States)

Prival, D.B.; Goode, M.J.; Swann, D.E.; Schwalbe, C.R.; Schroff, M.J.

2002-01-01

The twin-spotted rattlesnake (Crotalus pricei) is a small-bodied pitviper that has received little attention in the literature to date. The species reaches the northern limit of its range in southeastern Arizona, where it inhabits higher elevations than any of the state's 10 other rattlesnake species. During 1997-2000, we captured, measured, and marked 127 C. pricei in Arizona's Chiricahua Mountains between 2530 and 2900 m elevation. We also used radiotelemetry to track the movements of 16 C. pricei in the study area during 1997-1998. Mean (?? SE) snout-vent length of C. pricei was 387.8 ?? 8.3 mm (range = 168-572), and mean mass was 53.5 ?? 3.3 g (range = 3.6-188.5). Based on fecal analyses, lizards constituted the bulk of prey (74%), but the diet of C. pricei also included mammals, birds, and a conspecific. Mating was concentrated in August and early September and parturition took place during late July and August. Mean number of embryos was 3.94 ?? 0.34 (range = 1-6) and female reproduction appeared biennial or less frequent. Based on shed and growth rates, female C. pricei develop embryos at 4-5 years of age. Gravid females maintained warmer body temperatures relative to substrate temperature than nongravid females or males, presumably by spending more time basking than other snakes. Radiotelemetry revealed that movement patterns varied from year to year, as males moved over six times farther per week during the 1998 monsoon season (July to September) than during the 1997 monsoon season. Additionally, use of talus slopes by males decreased during 1998. During dry years, such as 1998, males may be forced off talus into cooler microclimates where resources are less concentrated than on talus.

Neem leaf extract as potential modifier of free radical interaction with RBC membrane

International Nuclear Information System (INIS)

Patil, Shilpa M.; Kulkarni, Satish G.; Rane, Charusheela

2004-01-01

Full text: Neem tree (Azadirachta indica) is well known as village pharmacy due to its multifaceted medicinal action in curing countless health problems. The leaf extract of Neem is known to purify blood by removing toxins and neutralizing free radicals. Using gamma irradiation as free radical generation process the efficacy of neem leaf extract in combating free radical attack on RBC membrane has been explored. The results narrating role of concentration and dose dependency in this radio modulation will be discussed

Anti-Ageing Effects of Sonchus oleraceus L. (pūhā) Leaf Extracts on H2O2-Induced Cell Senescence

OpenAIRE

Zong-Quan Ou; Thomas Rades; Arlene McDowell

2015-01-01

Antioxidants protect against damage from free radicals and are believed to slow the ageing process. Previously, we have reported the high antioxidant activity of 70% methanolic Sonchus oleraceus L. (Asteraceae) leaf extracts. We hypothesize that S. oleraceus extracts protect cells against H2O2-induced senescence by mediating oxidative stress. Premature senescence of young WI-38 cells was induced by application of H2O2. Cells were treated with S. oleraceus extracts before or after H2O2 stress...

Green Synthesis and Antimicrobial Activities of Silver Nanoparticles using Cell Free-Extracts of Enterococcus species

Directory of Open Access Journals (Sweden)

Iyabo C. OLADIPO

2017-06-01

Full Text Available Cell-free extracts of six strains of Enterococcus species obtained from fermented foods were used for the green synthesis of silver nanoparticles (AgNPs, which was characterized by UV-Vis spectroscopy, Fourier-transform infrared spectroscopy (FTIR and transmission electron microscopy (TEM. The biosynthesized AgNPs were dark brown in colour having surface plasmon resonance in the range of 420-442 nm. The spherical shaped AgNPs had sizes of 4-55 nm, whose formations were facilitated by proteins as indicated by the presence of peaks 1,635-1,637 and 3,275-3,313 cm-1 in the FTIR spectra. The energy dispersive x-ray (EDX showed prominent presence of silver in the AgNPs colloidal solution, while the selected area electron diffraction was typified by the face-centred crystalline nature of silver. The particles inhibited the growth of multi-drug resistant clinical isolates of Escherichia coli, Klebsiella pneumoniae and Proteus vulgaris, and also potentiated the activities of ampicillin, ciprofloxacin and cefuroxime in the AgNPs-antibiotic synergy studies. In addition, the prospective relevance of the particles as nanopreservative in paints was demonstrated with the inhibition of growth of Staphylococcus aureus, Pseudomonas aeruginosa, Aspergillus niger and A. flavus in AgNPs-paint admixture. This report further demonstrates the green synthesis of AgNPs by strains of Enterococcus species.

Formation of hydroxyl radical from San Joaquin Valley particles extracted in a cell-free surrogate lung fluid

Directory of Open Access Journals (Sweden)

H. Shen

2011-09-01

Full Text Available Previous studies have suggested that the adverse health effects from ambient particulate matter (PM are linked to the formation of reactive oxygen species (ROS by PM in cardiopulmonary tissues. While hydroxyl radical (^•OH is the most reactive of the ROS species, there are few quantitative studies of ^•OH generation from PM. Here we report on ^•OH formation from PM collected at an urban (Fresno and rural (Westside site in the San Joaquin Valley (SJV of California. We quantified ^•OH in PM extracts using a cell-free, phosphate-buffered saline (PBS solution with or without 50 μM ascorbate (Asc. The results show that generally the urban Fresno PM generates much more ^•OH than the rural Westside PM. The presence of Asc at a physiologically relevant concentration in the extraction solution greatly enhances ^•OH formation from all the samples. Fine PM (PM_2.5 generally makes more ^•OH than the corresponding coarse PM (PM_cf, i.e. with diameters of 2.5 to 10 μm normalized by air volume collected, while the coarse PM typically generates more ^•OH normalized by PM mass. ^•OH production by SJV PM is reduced on average by (97 ± 6 % when the transition metal chelator desferoxamine (DSF is added to the extraction solution, indicating a dominant role of transition metals. By measuring calibration curves of ^•OH generation from copper and iron, and quantifying copper and iron concentrations in our particle extracts, we find that PBS-soluble copper is primarily responsible for ^•OH production by the SJV PM, while iron often makes a significant contribution. Extrapolating our results to expected burdens of PM-derived ^•OH in human lung lining fluid suggests that typical daily PM exposures in the San Joaquin Valley are unlikely to result in a high amount of pulmonary ^•OH, although high

Effect of kinetic properties of extraction systems on separation of some elements by liquid chromatography method with free fixed phase

International Nuclear Information System (INIS)

Fedotov, P.S.; Maryutina, T.A.; Pichugin, A.A.; Spivakov, B.Ya.

1993-01-01

Effect of kinetic properties of a series of extraction systems on the separation of certain elements by the method of liquid chromatography with free fixed phase is considered. Chromatographic behaviour of europium 3 and iron 3 ions when using systems based on di-2-ethylhexylphosphovers acid and tetraphenylmethylenediphosphine dioxide is investigated. Kinetic properties of the extraction systems used are studied by diffusion cell method with mixing, europium 3 and iron 3 mass transfer coefficients are determined

Modifier free supercritical fluid extraction of uranium from sintered UO2, soil and ore samples

International Nuclear Information System (INIS)

Kanekar, A.S.; Pathak, P.N.; Acharya, R.; Mohapatra, P.K.; Manchanda, V.K.

2011-01-01

Direct extraction of uranium from different samples viz. sintered UO 2 , soil and ores was carried out by modifier free supercritical fluid using tri-n-butyl phosphate-nitric acid (TBP-HNO 3 ) adduct as extractant. These studies showed that pre-equilibration with more concentrated nitric acid helps in better dissolution and extraction of uranium from sintered UO 2 samples. Modifier free supercritical fluid extraction appears attractive with respect to minimization of secondary wastes. This method resulted 80-100% extraction of uranium from different soil/ore samples. The results were confirmed by performing neutron activation analysis of original (before extraction) and residue (after extraction) samples. (author)

Anti-ageing effects of Sonchus oleraceus L. (pūhā) leaf extracts on H₂O₂-induced cell senescence.

Science.gov (United States)

Ou, Zong-Quan; Rades, Thomas; McDowell, Arlene

2015-03-12

Antioxidants protect against damage from free radicals and are believed to slow the ageing process. Previously, we have reported the high antioxidant activity of 70% methanolic Sonchus oleraceus L. (Asteraceae) leaf extracts. We hypothesize that S. oleraceus extracts protect cells against H2O2-induced senescence by mediating oxidative stress. Premature senescence of young WI-38 cells was induced by application of H2O2. Cells were treated with S. oleraceus extracts before or after H2O2 stress. The senescence- associated β-galactosidase (SA-β-gal) activity was used to indicate cell senescence. S. oleraceus extracts showed higher cellular antioxidant activity than chlorogenic acid in WI-38 cells. S. oleraceus extracts suppressed H2O2 stress-induced premature senescence in a concentration-dependent manner. At 5 and 20 mg/mL, S. oleraceus extracts showed better or equivalent effects of reducing stress-induced premature senescence than the corresponding ascorbic acid treatments. These findings indicate the potential of S. oleraceus extracts to be formulated as an anti-ageing agent.

Antioxidant Properties of Berberis aetnensis C. Presl (Berberidaceae Roots Extract and Protective Effects on Astroglial Cell Cultures

Directory of Open Access Journals (Sweden)

Agata Campisi

2014-01-01

Full Text Available Berberis aetnensis C. Presl (Berberidaceae is a bushy-spiny shrub common on Mount Etna (Sicily. We demonstrated that the alkaloid extract of roots of B. aetnensis C. Presl contains prevalently berberine and berbamine, possesses antimicrobial properties, and was able to counteract the upregulation evoked by glutamate of tissue transglutaminase in primary rat astroglial cell cultures. Until now, there are no reports regarding antioxidant properties of B. aetnensis C. Presl collected in Sicily. Air-dried, powdered roots of B. aetnensis C. Presl were extracted, identified, and quantified by HPLC. We assessed in cellular free system its effect on superoxide anion, radicals scavenging activity of antioxidants against free radicals like the 1,1-diphenyl-2-picrylhydrazyl radical, and the inhibition of xanthine oxidase activity. In primary rat astroglial cell cultures, exposed to glutamate, we evaluated the effect of the extract on glutathione levels and on intracellular production of reactive oxygen species generated by glutamate. The alkaloid extract of B. aetnensis C. Presl inhibited superoxide anion, restored to control values, the decrease of GSH levels, and the production of reactive oxygen species. Potent antioxidant activities of the alkaloid extract of roots of B. aetnensis C. Presl may be one of the mechanisms by which the extract is effective against health disorders associated to oxidative stress.

Antioxidant properties of Berberis aetnensis C. Presl (Berberidaceae) roots extract and protective effects on astroglial cell cultures.

Science.gov (United States)

Campisi, Agata; Acquaviva, Rosaria; Bonfanti, Roberta; Raciti, Giuseppina; Amodeo, Andrea; Mastrojeni, Silvana; Ragusa, Salvatore; Iauk, Liliana

2014-01-01

Berberis aetnensis C. Presl (Berberidaceae) is a bushy-spiny shrub common on Mount Etna (Sicily). We demonstrated that the alkaloid extract of roots of B. aetnensis C. Presl contains prevalently berberine and berbamine, possesses antimicrobial properties, and was able to counteract the upregulation evoked by glutamate of tissue transglutaminase in primary rat astroglial cell cultures. Until now, there are no reports regarding antioxidant properties of B. aetnensis C. Presl collected in Sicily. Air-dried, powdered roots of B. aetnensis C. Presl were extracted, identified, and quantified by HPLC. We assessed in cellular free system its effect on superoxide anion, radicals scavenging activity of antioxidants against free radicals like the 1,1-diphenyl-2-picrylhydrazyl radical, and the inhibition of xanthine oxidase activity. In primary rat astroglial cell cultures, exposed to glutamate, we evaluated the effect of the extract on glutathione levels and on intracellular production of reactive oxygen species generated by glutamate. The alkaloid extract of B. aetnensis C. Presl inhibited superoxide anion, restored to control values, the decrease of GSH levels, and the production of reactive oxygen species. Potent antioxidant activities of the alkaloid extract of roots of B. aetnensis C. Presl may be one of the mechanisms by which the extract is effective against health disorders associated to oxidative stress.

Screening of various botanical extracts for antioxidant activity using DPPH free radical method.

Science.gov (United States)

Waqas, Muhammad Khurram; Saqib, Najam-Us; Rashid, Saeed-Ur; Shah, Pervaiz Akhtar; Akhtar, Naveed; Murtaza, Ghulam

2013-01-01

Aiming at the exploration of herbal use by society, crude extracts of the seeds of some commonly used medicinal plants (Vitis vinifera, Tamarindus indica and Glycin max) were screened for their free radical scavenging properties using ascorbic acid as standard antioxidant. Free radical scavenging activity was evaluated using 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical. The overall antioxidant activity of grape seeds (Vitis vinifera) was the strongest, followed in descending order by soybean (Glycin max) and tamarind (Tamarindus indica). The seeds extract of Vitis vinifera, Glycin max and Tamarindus indica showed 85.61%, 83.45% and 79.26%, DPPH scavenging activity respectively.

Dynamic quantitative analysis of adherent cell cultures by means of lens-free video microscopy

Science.gov (United States)

Allier, C.; Vincent, R.; Navarro, F.; Menneteau, M.; Ghenim, L.; Gidrol, X.; Bordy, T.; Hervé, L.; Cioni, O.; Bardin, S.; Bornens, M.; Usson, Y.; Morales, S.

2018-02-01

We present our implementation of lens-free video microscopy setup for the monitoring of adherent cell cultures. We use a multi-wavelength LED illumination together with a dedicated holographic reconstruction algorithm that allows for an efficient removal of twin images from the reconstructed phase image for densities up to those of confluent cell cultures (>500 cells/mm2). We thereby demonstrate that lens-free video microscopy, with a large field of view ( 30 mm2) can enable us to capture the images of thousands of cells simultaneously and directly inside the incubator. It is then possible to trace and quantify single cells along several cell cycles. We thus prove that lens-free microscopy is a quantitative phase imaging technique enabling estimation of several metrics at the single cell level as a function of time, for example the area, dry mass, maximum thickness, major axis length and aspect ratio of each cell. Combined with cell tracking, it is then possible to extract important parameters such as the initial cell dry mass (just after cell division), the final cell dry mass (just before cell division), the average cell growth rate, and the cell cycle duration. As an example, we discuss the monitoring of a HeLa cell cultures which provided us with a data-set featuring more than 10 000 cell cycle tracks and more than 2x106 cell morphological measurements in a single time-lapse.

Anti-Ageing Effects of Sonchus oleraceus L. (pūhā) Leaf Extracts on H2O2-Induced Cell Senescence

DEFF Research Database (Denmark)

Ou, Zong-Quan; Rades, Thomas; McDowell, Arlene

2015-01-01

Antioxidants protect against damage from free radicals and are believed to slow the ageing process. Previously, we have reported the high antioxidant activity of 70% methanolic Sonchus oleraceus L. (Asteraceae) leaf extracts. We hypothesize that S. oleraceus extracts protect cells against H2O2......-induced senescence by mediating oxidative stress. Premature senescence of young WI-38 cells was induced by application of H2O2. Cells were treated with S. oleraceus extracts before or after H2O2 stress. The senescence- associated β-galactosidase (SA-β-gal) activity was used to indicate cell senescence. S....... oleraceus extracts showed higher cellular antioxidant activity than chlorogenic acid in WI-38 cells. S. oleraceus extracts suppressed H2O2 stress-induced premature senescence in a concentration-dependent manner. At 5 and 20 mg/mL, S. oleraceus extracts showed better or equivalent effects of reducing stress...

Scaling Up ITO-free solar cells

DEFF Research Database (Denmark)

Galagan, Yulia; Coenen, Erica W. C.; Zimmermann, Birger

2014-01-01

Indium-tin-oxide-free (ITO-free) polymer solar cells with composite electrodes containing current-collecting grids and a semitransparent poly(3,4-ethylenedioxythiophene):polystyrenesulfonate) (PEDOT:PSS) conductor are demonstrated. The up-scaling of the length of the solar cell from 1 to 6 cm...... resistances. The performance of ITO-free organic solar cells with different dimensions and different electrode resistances are evaluated for different light intensities. The current generation and electric potential distribution are found to not be uniformly distributed in large-area devices at simulated 1...

Cell-Free Systems Based on CHO Cell Lysates: Optimization Strategies, Synthesis of "Difficult-to-Express" Proteins and Future Perspectives.

Directory of Open Access Journals (Sweden)

Lena Thoring

Full Text Available Nowadays, biotechnological processes play a pivotal role in target protein production. In this context, Chinese Hamster Ovary (CHO cells are one of the most prominent cell lines for the expression of recombinant proteins and revealed as a safe host for nearly 40 years. Nevertheless, the major bottleneck of common in vivo protein expression platforms becomes obvious when looking at the production of so called "difficult-to-express" proteins. This class of proteins comprises in particular several ion channels and multipass membrane proteins as well as cytotoxic proteins. To enhance the production of "difficult-to-express" proteins, alternative technologies were developed, mainly based on translationally active cell lysates. These so called "cell-free" protein synthesis systems enable an efficient production of different classes of proteins. Eukaryotic cell-free systems harboring endogenous microsomal structures for the synthesis of functional membrane proteins and posttranslationally modified proteins are of particular interest for future applications. Therefore, we present current developments in cell-free protein synthesis based on translationally active CHO cell extracts, underlining the high potential of this platform. We present novel results highlighting the optimization of protein yields, the synthesis of various "difficult-to-express" proteins and the cotranslational incorporation of non-standard amino acids, which was exemplarily demonstrated by residue specific labeling of the glycoprotein Erythropoietin and the multimeric membrane protein KCSA.

Stiffness-Independent Highly Efficient On-Chip Extraction of Cell-Laden Hydrogel Microcapsules from Oil Emulsion into Aqueous Solution by Dielectrophoresis.

Science.gov (United States)

Huang, Haishui; Sun, Mingrui; Heisler-Taylor, Tyler; Kiourti, Asimina; Volakis, John; Lafyatis, Gregory; He, Xiaoming

2015-10-28

A dielectrophoresis (DEP)-based method achieves highly efficient on-chip extraction of cell-laden microcapsules of any stiffness from oil into aqueous solution. The hydrogel microcapsules can be extracted into the aqueous solution by DEP and interfacial tension forces with no trapped oil, while the encapsulated cells are free from electrical damage due to the Faraday cage effect. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The Cytotoxic, Antibacterial and Free Radical Scavenging Activities of Crude Extracts of Matricaria chamomilla, Salvadora persica and Artemisia annua

KAUST Repository

Seddek, Ahmed

2011-12-01

The discovery of drugs from natural sources has been a rapidly growing science in this era. Plants used for medicinal purposes have been usually studied as rich sources of bioactive chemical compounds that can be used as medications. Several plant-derived drugs have been approved so far. Cancer and infectious diseases have been common targets for the science of drug discovery, due to the high mortality rates caused by these diseases all over the world. Several plant-derived compounds are being marketed now as anti-cancer agents. However, finding novel antimicrobial and anti-cancer compounds has become an important goal to overcome the problems of existing anti-cancer and antimicrobial agents, such as resistance and non-selectivity. In this thesis project, an attempt to find out useful biological activities of the crude extracts of some plants used traditionally for medicinal purposes in Saudi Arabia has been made. Matricaria chamomilla, Salvadora persica and Artemisia annua have been selected for study, based on the literature review performed. These plants were screened for three biological activities; anti-cancer, anti-bacterial and free radical scavenging activities. The experimental part of the study consisted of some common in-vitro techniques, such as cytotoxicity and cell viability assays, disk diffusion assay and 1,1-diphenyl-2-picryl-hydrazyl assay. In addition, the crude extract of Matricaria chamomilla has undergone chemical fractionation and four solvent fractions were obtained using column chromatography. The crude extract of Matricaria chamomilla showed a promising anti-bacterial activity against Escherichia coli and a very promising free radical scavenging activity that was comparable to ascorbic acid, an important anti-oxidant. The four solvent fractions obtained from that extract showed that these activities were produced by more than one compound belonging to different solvent fractions. In addition, the crude extract of Artemisia annua showed

Micro-electromembrane extraction across free liquid membranes. Extractions of basic drugs from undiluted biological samples

Czech Academy of Sciences Publication Activity Database

Kubáň, Pavel; Boček, Petr

2014-01-01

Roč. 1337, Apr (2014), s. 32-39 ISSN 0021-9673 R&D Projects: GA ČR(CZ) GA13-05762S Institutional support: RVO:68081715 Keywords : micro-electromembrane extraction * free liquid membranes * biological samples Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.169, year: 2014

Cytotoxicity of extracts of spices to cultured cells.

Science.gov (United States)

Unnikrishnan, M C; Kuttan, R

1988-01-01

The cytotoxicity of the extracts from eight different spices used in the Indian diet was determined using Dalton's lymphoma ascites tumor cells and human lymphocytes in vitro and Chinese Hamster Ovary cells and Vero cells in tissue culture. Alcoholic extracts of the spices were found to be more cytotoxic to these cells than their aqueous extracts. Alcoholic extracts of several spices inhibited cell growth at concentrations of 0.2-1 mg/ml in vitro and 0.12-0.3 mg/ml in tissue culture. Ginger, pippali (native to India; also called dried catkins), pepper, and garlic showed the highest activity followed by asafetida, mustard, and horse-gram (native to India). These extracts also inhibited the thymidine uptake into DNA.

Antiulcerogenic Potential Activity of Free and Nanoencapsulated Passiflora serratodigitata L. Extracts

Directory of Open Access Journals (Sweden)

Marc Strasser

2014-01-01

Full Text Available This paper provides evidence that the leaves and stem of Passiflora serratodigitata L. dry crude extract (DCE, ethylacetate fraction (EAF, and residual water fraction show potential antiulcerogenic activity. Interestingly, the polymeric nanocapsule loaded with EAF had 10-fold more activity than the free EAF. Furthermore, the polymer nanoparticles provided homogeneous colloidal drug delivery systems and allowed overcoming challenges such as poor aqueous solubility as well as the physical-chemical instability of the organic extract, which presented 90% (w/w of the flavonoid content. The entrapment efficiency of the total flavonoid was 90.6 ± 2.5% (w/v for the DCE and 79.9 ± 2.7% (w/v for the EAF. This study shows that nanoencapsulation improves both the physicochemical properties and the efficacy of the herbal formulations. Therefore, free and encapsulated extracts have the potential to be suitable drug design candidates for the therapeutic management of ulcer.

free radical scavenging and cytotoxic activity of standardized extracts

African Journals Online (AJOL)

Ledy

cancer cell line HeLa of five commercial standardized extracts rich in phenolic compounds: red wine ... and used as feedstock in nutritional supplement manu- .... pharmaceutical, food, or cosmetic products, as ..... cancer stem-like cell.

Scaling up ITO-Free solar cells

NARCIS (Netherlands)

Galagan, Y.O.; Coenen, E.W.C.; Zimmermann, B.; Slooff, L.H.; Verhees, W.J.H.; Veenstra, S.C.; Kroon, J.M.; Jørgensen, M.; Krebs, F.C.; Andriessen, H.A.J.M.

2014-01-01

Indium-tin-oxide-free (ITO-free) polymer solar cells with composite electrodes containing current-collecting grids and a semitransparent poly(3,4-ethylenedioxythiophene):polystyrenesulfonate) (PEDOT:PSS) conductor are demonstrated. The up-scaling of the length of the solar cell from 1 to 6 cm and

Efficient cell-free expression with the endogenous E. Coli RNA polymerase and sigma factor 70

Directory of Open Access Journals (Sweden)

Noireaux Vincent

2010-06-01

Full Text Available Abstract Background Escherichia coli cell-free expression systems use bacteriophage RNA polymerases, such as T7, to synthesize large amounts of recombinant proteins. These systems are used for many applications in biotechnology, such as proteomics. Recently, informational processes have been reconstituted in vitro with cell-free systems. These synthetic approaches, however, have been seriously limited by a lack of transcription modularity. The current available cell-free systems have been optimized to work with bacteriophage RNA polymerases, which put significant restrictions to engineer processes related to biological information. The development of efficient cell-free systems with broader transcription capabilities is required to study complex informational processes in vitro. Results In this work, an efficient cell-free expression system that uses the endogenous E. coli RNA polymerase only and sigma factor 70 for transcription was prepared. Approximately 0.75 mg/ml of Firefly luciferase and enhanced green fluorescent protein were produced in batch mode. A plasmid was optimized with different regulatory parts to increase the expression. In addition, a new eGFP was engineered that is more translatable in cell-free systems than the original eGFP. The protein production was characterized with three different adenosine triphosphate (ATP regeneration systems: creatine phosphate (CP, phosphoenolpyruvate (PEP, and 3-phosphoglyceric acid (3-PGA. The maximum protein production was obtained with 3-PGA. Preparation of the crude extract was streamlined to a simple routine procedure that takes 12 hours including cell culture. Conclusions Although it uses the endogenous E. coli transcription machinery, this cell-free system can produce active proteins in quantities comparable to bacteriophage systems. The E. coli transcription provides much more possibilities to engineer informational processes in vitro. Many E. coli promoters/operators specific to sigma

A Simple and Rapid Method for Preparing a Cell-Free Bacterial Lysate for Protein Synthesis.

Directory of Open Access Journals (Sweden)

Nitzan Krinsky

Full Text Available Cell-free protein synthesis (CFPS systems are important laboratory tools that are used for various synthetic biology applications. Here, we present a simple and inexpensive laboratory-scale method for preparing a CFPS system from E. coli. The procedure uses basic lab equipment, a minimal set of reagents, and requires less than one hour to process the bacterial cell mass into a functional S30-T7 extract. BL21(DE3 and MRE600 E. coli strains were used to prepare the S30-T7 extract. The CFPS system was used to produce a set of fluorescent and therapeutic proteins of different molecular weights (up to 66 kDa. This system was able to produce 40-150 μg-protein/ml, with variations depending on the plasmid type, expressed protein and E. coli strain. Interestingly, the BL21-based CFPS exhibited stability and increased activity at 40 and 45°C. To the best of our knowledge, this is the most rapid and affordable lab-scale protocol for preparing a cell-free protein synthesis system, with high thermal stability and efficacy in producing therapeutic proteins.

A Simple and Rapid Method for Preparing a Cell-Free Bacterial Lysate for Protein Synthesis

Science.gov (United States)

Kaduri, Maya; Shainsky-Roitman, Janna; Goldfeder, Mor; Ivanir, Eran; Benhar, Itai; Shoham, Yuval; Schroeder, Avi

2016-01-01

Cell-free protein synthesis (CFPS) systems are important laboratory tools that are used for various synthetic biology applications. Here, we present a simple and inexpensive laboratory-scale method for preparing a CFPS system from E. coli. The procedure uses basic lab equipment, a minimal set of reagents, and requires less than one hour to process the bacterial cell mass into a functional S30-T7 extract. BL21(DE3) and MRE600 E. coli strains were used to prepare the S30-T7 extract. The CFPS system was used to produce a set of fluorescent and therapeutic proteins of different molecular weights (up to 66 kDa). This system was able to produce 40–150 μg-protein/ml, with variations depending on the plasmid type, expressed protein and E. coli strain. Interestingly, the BL21-based CFPS exhibited stability and increased activity at 40 and 45°C. To the best of our knowledge, this is the most rapid and affordable lab-scale protocol for preparing a cell-free protein synthesis system, with high thermal stability and efficacy in producing therapeutic proteins. PMID:27768741

Differentiation of Dental Pulp Stem Cells into Neuron-Like Cells in Serum-Free Medium

Directory of Open Access Journals (Sweden)

Shahrul Hisham Zainal Ariffin

2013-01-01

Full Text Available Dental pulp tissue contains dental pulp stem cells (DPSCs. Dental pulp cells (also known as dental pulp-derived mesenchymal stem cells are capable of differentiating into multilineage cells including neuron-like cells. The aim of this study was to examine the capability of DPSCs to differentiate into neuron-like cells without using any reagents or growth factors. DPSCs were isolated from teeth extracted from 6- to 8-week-old mice and maintained in complete medium. The cells from the fourth passage were induced to differentiate by culturing in medium without serum or growth factors. RT-PCR molecular analysis showed characteristics of Cd146+, Cd166+, and Cd31− in DPSCs, indicating that these cells are mesenchymal stem cells rather than hematopoietic stem cells. After 5 days of neuronal differentiation, the cells showed neuron-like morphological changes and expressed MAP2 protein. The activation of Nestin was observed at low level prior to differentiation and increased after 5 days of culture in differentiation medium, whereas Tub3 was activated only after 5 days of neuronal differentiation. The proliferation of the differentiated cells decreased in comparison to that of the control cells. Dental pulp stem cells are induced to differentiate into neuron-like cells when cultured in serum- and growth factor-free medium.

An Extraordinary Accumulation of (-)-Pinoresinol in Cell-Free Extracts of Forsythia intermedia: Evidence for Enantiospecific Reduction of (+)-Pinoresinol

Science.gov (United States)

Katayama, Takeshi; Davin, Laurence B.; Lewis, Norman G.

1992-01-01

Stereoselective and enantiospecific transformation mechanisms in lignan biogenesis are only now yielding to scientific inquiry: it has been shown that soluble cell-free preparations from Forsythia intermedia catalysis the formation of the enantiomerically pure lignan, (-)-secoisolariciresinol, when incubated with coniferyl alcohol in the presence of NAD(P)H and H2O2. Surprisingly, (-)-pinoresinol also accumulates in this soluble cell-free assay mixture in greater than 96% enantiomeric excess, even though it is not the naturally occurring antipode present in Forsythia sp. But these soluble cell-free preparations do not engender stereoselective coupling; instead, racemic pinoresinols are first formed, catalysed by an H2O2-dependent peroxidase reaction. An enantiospecific NAD(P)H reductase then converts (+)- pinoresinol, and not the (-)-antipode, into (-)-secoisolariciresinol. Stereoselective syntheis of(+)-pinoresinol from E-coniferyl alcohol is, however, catalysed by an insoluble enzyme preparation in F. suspensa, obtained following removal of readily soluble and ionically bound enzymes; no exogenously supplied cofactors were required other than oxygen, although the reaction was stimulated by NAD-malate addition. Thus, the overall biochemical pathway to enantiomerically pure (-)-secoisolariciresinol has been delineated.

Anti-Ageing Effects of Sonchus oleraceus L. (pūhā Leaf Extracts on H2O2-Induced Cell Senescence

Directory of Open Access Journals (Sweden)

Zong-Quan Ou

2015-03-01

Full Text Available Antioxidants protect against damage from free radicals and are believed to slow the ageing process. Previously, we have reported the high antioxidant activity of 70% methanolic Sonchus oleraceus L. (Asteraceae leaf extracts. We hypothesize that S. oleraceus extracts protect cells against H2O2-induced senescence by mediating oxidative stress. Premature senescence of young WI-38 cells was induced by application of H2O2. Cells were treated with S. oleraceus extracts before or after H2O2 stress. The senescence- associated β-galactosidase (SA-β-gal activity was used to indicate cell senescence. S. oleraceus extracts showed higher cellular antioxidant activity than chlorogenic acid in WI-38 cells. S. oleraceus extracts suppressed H2O2 stress-induced premature senescence in a concentration-dependent manner. At 5 and 20 mg/mL, S. oleraceus extracts showed better or equivalent effects of reducing stress-induced premature senescence than the corresponding ascorbic acid treatments. These findings indicate the potential of S. oleraceus extracts to be formulated as an anti-ageing agent.

The Effect of Mercury Vapor and the Role of Green Tea Extract on Brain Cells

Directory of Open Access Journals (Sweden)

Dhona Afriza

2013-09-01

Full Text Available Mercury is a wellknown toxic metal that is capable to induce free radical-induced oxidative stress. It can cause human disease including brain disorders. Objective: To identify the effect of mercury vapor inhalation on brain cells and the role of green tea extract (Camellia sinensis as antioxidant on the brain cells exposed to mercury. Methods: Fourty-eight male Mus musculus were divided into 8 groups, which were given treatment for 3 and 6 weeks. Group A did not receive any treatment and served as a negative control. Group B was a positive control exposed to Mercury. Group C was exposed to Mercury and treated with 26μg/g green tea extract. Group D was exposed to mercury and treated with 52μg/g green tea extract. All animals in the Group B, C, D were exposed to mercury through inhalation for 4 hours daily. The effect of mercury on the brain cells were examined histopathologically. Results: The numbers of necrotic cells counted in the green tea-treated mice group were significantly lower than those untreated group (p<0,05. Conclusion: Mercury vapor inhalation may cause necrosis on brain cells. Administration of green tea extract as an antioxidant reduced the amount of mercury-induced necrotic brain cells in mice.DOI: 10.14693/jdi.v20i2.151

The adrenergic retulation of the cardiovascular system in the South American rattlesnake, Crotalus durissus

DEFF Research Database (Denmark)

Galli, G.L.J.; Jensen, Nini Skovgaard; Abe, A.S.

2007-01-01

The present study investigates adrenergic regulation of the systemic and pulmonary circulations of the anaesthetised South American rattlesnake, Crotalus durissus. Haemodynamic measurements were made following bolus injections of adrenaline and adrenergic antagonists administered through a systemic...... arterial catheter. Adrenaline caused a marked systemic vasoconstriction that was abolished by phentolamine, indicating this response was mediated through α-adrenergic receptors. Injection of phentolamine gave rise to a pronounced vasodilatation (systemic conductance (Gsys) more than doubled), while...... injection of propranolol caused a systemic vasoconstriction, pointing to a potent α-adrenergic, and a weaker β-adrenergic tone in the systemic vasculature of Crotalus. Overall, the pulmonary vasculature was far less responsive to adrenergic stimulation than the systemic circulation. Adrenaline caused...

Mechanisms of bacterial membrane permeabilization by crotalicidin (Ctn) and its fragment Ctn(15-34), antimicrobial peptides from rattlesnake venom.

Science.gov (United States)

Pérez-Peinado, Clara; Dias, Susana Almeida; Domingues, Marco M; Benfield, Aurélie H; Freire, João Miguel; Rádis-Baptista, Gandhi; Gaspar, Diana; Castanho, Miguel A R B; Craik, David J; Henriques, Sónia Troeira; Veiga, Ana Salomé; Andreu, David

2018-02-02

Crotalicidin (Ctn), a cathelicidin-related peptide from the venom of a South American rattlesnake, possesses potent antimicrobial, antitumor, and antifungal properties. Previously, we have shown that its C-terminal fragment, Ctn(15-34), retains the antimicrobial and antitumor activities but is less toxic to healthy cells and has improved serum stability. Here, we investigated the mechanisms of action of Ctn and Ctn(15-34) against Gram-negative bacteria. Both peptides were bactericidal, killing ∼90% of Escherichia coli and Pseudomonas aeruginosa cells within 90-120 and 5-30 min, respectively. Studies of ζ potential at the bacterial cell membrane suggested that both peptides accumulate at and neutralize negative charges on the bacterial surface. Flow cytometry experiments confirmed that both peptides permeabilize the bacterial cell membrane but suggested slightly different mechanisms of action. Ctn(15-34) permeabilized the membrane immediately upon addition to the cells, whereas Ctn had a lag phase before inducing membrane damage and exhibited more complex cell-killing activity, probably because of two different modes of membrane permeabilization. Using surface plasmon resonance and leakage assays with model vesicles, we confirmed that Ctn(15-34) binds to and disrupts lipid membranes and also observed that Ctn(15-34) has a preference for vesicles that mimic bacterial or tumor cell membranes. Atomic force microscopy visualized the effect of these peptides on bacterial cells, and confocal microscopy confirmed their localization on the bacterial surface. Our studies shed light onto the antimicrobial mechanisms of Ctn and Ctn(15-34), suggesting Ctn(15-34) as a promising lead for development as an antibacterial/antitumor agent. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Dynamic imaging of cell-free and cell-associated viral capture in mature dendritic cells.

Science.gov (United States)

Izquierdo-Useros, Nuria; Esteban, Olga; Rodriguez-Plata, Maria T; Erkizia, Itziar; Prado, Julia G; Blanco, Julià; García-Parajo, Maria F; Martinez-Picado, Javier

2011-12-01

Dendritic cells (DCs) capture human immunodeficiency virus (HIV) through a non-fusogenic mechanism that enables viral transmission to CD4(+) T cells, contributing to in vivo viral dissemination. Although previous studies have provided important clues to cell-free viral capture by mature DCs (mDCs), dynamic and kinetic insight on this process is still missing. Here, we used three-dimensional video microscopy and single-particle tracking approaches to dynamically dissect both cell-free and cell-associated viral capture by living mDCs. We show that cell-free virus capture by mDCs operates through three sequential phases: virus binding through specific determinants expressed in the viral particle, polarized or directional movements toward concrete regions of the cell membrane and virus accumulation in a sac-like structure where trapped viral particles display a hindered diffusive behavior. Moreover, real-time imaging of cell-associated viral transfer to mDCs showed a similar dynamics to that exhibited by cell-free virus endocytosis leading to viral accumulation in compartments. However, cell-associated HIV type 1 transfer to mDCs was the most effective pathway, boosted throughout enhanced cellular contacts with infected CD4(+) T cells. Our results suggest that in lymphoid tissues, mDC viral uptake could occur either by encountering cell-free or cell-associated virus produced by infected cells generating the perfect scenario to promote HIV pathogenesis and impact disease progression. © 2011 John Wiley & Sons A/S.

Comparison of microwave hydrodistillation and solvent-free microwave extraction of essential oil from Melaleuca leucadendra Linn

Science.gov (United States)

Ismanto, A. W.; Kusuma, H. S.; Mahfud, M.

2017-12-01

The comparison of solvent-free microwave extraction (SFME) and microwave hydrodistillation (MHD) in the extraction of essential oil from Melaleuca leucadendra Linn. was examined. Dry cajuput leaves were used in this study. The purpose of this study is also to determine optimal condition (microwave power). The relative electric consumption of SFME and MHD methods are both showing 0,1627 kWh/g and 0,3279 kWh/g. The results showed that solvent-free microwave extraction methods able to reduce energy consumption and can be regarded as a green technique for extraction of cajuput oil.

Effects of food supplementation on the physiological ecology of female Western diamond-backed rattlesnakes (Crotalus atrox).

Science.gov (United States)

Taylor, Emily N; Malawy, Michael A; Browning, Dawn M; Lemar, Shea V; DeNardo, Dale F

2005-06-01

Food availability is an important factor in the life histories of organisms because it is often limiting and thus can affect growth, mass change, reproduction, and behaviors such as thermoregulation, locomotion, and mating. Experimental studies in natural settings allow researchers to examine the effects of food on these parameters while animals are free to behave naturally. The wide variation among organisms in energy demands and among environmental food resources suggest that responses to changes in food availability may vary among organisms. Since most supplemental feeding field experiments have been conducted on species with high energy demands, we conducted a supplemental feeding study on free-ranging, female Western diamond-backed rattlesnakes (Crotalus atrox), a species with low energy demands and infrequent reproductive investment. Snakes were offered thawed rodents 1-4 times per week. Over two active seasons, we collected data on surface activity, home range size, growth, mass change, and reproduction of supplementally fed and control snakes. Fed and control snakes did not differ in surface activity levels (proportion of time encountered above versus below ground) or home range size. Fed snakes grew and gained mass faster, and had a dramatically higher occurrence of reproduction than control snakes. Also, fed snakes were in better body condition following reproduction than snakes that were not fed. However, litter characteristics such as offspring number and size were not increased by feeding, suggesting that these characteristics may be fixed. These data experimentally demonstrate that food availability can directly impact some life history traits (i.e., growth and reproduction for C. atrox), but not others (i.e., surface activity and home range size for C. atrox). The relationship between food availability and life history traits is affected in a complex way by ecological traits and physiological constraints, and thus interspecific variation in this

Tobacco BY-2 cell-free lysate: an alternative and highly-productive plant-based in vitro translation system.

Science.gov (United States)

Buntru, Matthias; Vogel, Simon; Spiegel, Holger; Schillberg, Stefan

2014-05-03

Cell-free protein synthesis is a rapid and efficient method for the production of recombinant proteins. Usage of prokaryotic cell-free extracts often leads to non-functional proteins. Eukaryotic counterparts such as wheat germ extract (WGE) and rabbit reticulocyte lysate (RLL) may improve solubility and promote the correct folding of eukaryotic multi-domain proteins that are difficult to express in bacteria. However, the preparation of WGEs is complex and time-consuming, whereas RLLs suffer from low yields. Here we report the development of a novel cell-free system based on tobacco Bright Yellow 2 (BY-2) cells harvested in the exponential growth phase. The highly-productive BY-2 lysate (BYL) can be prepared quickly within 4-5 h, compared to 4-5 d for WGE. The efficiency of the BYL was tested using three model proteins: enhanced yellow fluorescent protein (eYFP) and two versions of luciferase. The added mRNA was optimized by testing different 5' and 3' untranslated regions (UTRs). The protein yield in batch and dialysis reactions using BYL was much higher than that of a commercial Promega WGE preparation, achieving a maximum yield of 80 μg/mL of eYFP and 100 μg/mL of luciferase, compared to only 45 μg/mL of eYFP and 35 μg/mL of luciferase in WGEs. In dialysis reactions, the BYL yielded about 400 μg/mL eYFP, representing up to 50% more of the target protein than the Promega WGE, and equivalent to the amount using 5Prime WGE system. Due to the high yield and the short preparation time the BYL represents a remarkable improvement over current eukaryotic cell-free systems.

Danshen extract circumvents drug resistance and represses cell growth in human oral cancer cells.

Science.gov (United States)

Yang, Cheng-Yu; Hsieh, Cheng-Chih; Lin, Chih-Kung; Lin, Chun-Shu; Peng, Bo; Lin, Gu-Jiun; Sytwu, Huey-Kang; Chang, Wen-Liang; Chen, Yuan-Wu

2017-12-29

Danshen is a common traditional Chinese medicine used to treat neoplastic and chronic inflammatory diseases in China. However, the effects of Danshen on human oral cancer cells remain relatively unknown. This study investigated the antiproliferative effects of a Danshen extract on human oral cancer SAS, SCC25, OEC-M1, and KB drug-resistant cell lines and elucidated the possible underlying mechanism. We investigated the anticancer potential of the Danshen extract in human oral cancer cell lines and an in vivo oral cancer xenograft mouse model. The expression of apoptosis-related molecules was evaluated through Western blotting, and the concentration of in vivo apoptotic markers was measured using immunohistochemical staining. The antitumor effects of 5-fluorouracil and the Danshen extract were compared. Cell proliferation assays revealed that the Danshen extract strongly inhibited oral cancer cell proliferation. Cell morphology studies revealed that the Danshen extract inhibited the growth of SAS, SCC25, and OEC-M1 cells by inducing apoptosis. The Flow cytometric analysis indicated that the Danshen extract induced cell cycle G0/G1 arrest. Immunoblotting analysis for the expression of active caspase-3 and X-linked inhibitor of apoptosis protein indicated that Danshen extract-induced apoptosis in human oral cancer SAS cells was mediated through the caspase pathway. Moreover, the Danshen extract significantly inhibited growth in the SAS xenograft mouse model. Furthermore, the Danshen extract circumvented drug resistance in KB drug-resistant oral cancer cells. The study results suggest that the Danshen extract could be a potential anticancer agent in oral cancer treatment.

ACVP-05: Virus Genetic Analysis from Cell-Free Plasma, Virally Infected Cells or Tissues and Cultured Supernatant Via Single Genome Amplification and Direct Sequencing | Frederick National Laboratory for Cancer Research

Science.gov (United States)

The Viral Evolution Core within the AIDS and Cancer Virus Program will extract viral RNA/DNA from cell-free or cell-associated samples. Complementary (cDNA) will be generated as needed, and cDNA or DNA will be diluted to a single copy prior to nested

[Effects of the maca extract on the ultrastructures of mitochondria in the spinal nerve cell and exercise endurance].

Science.gov (United States)

Yu, Fa-Rong; Yang, Bo; Li, Zuo-Ping; Lian, Xiu-Zhen; Xie, Ming-Ren; Li, Deng-Lou; Zhang, Shi-Shuang

2017-06-08

To investigate the effects of maca extract on the ultrastructures of mitochondria in the spinal nerve cell and exercise endurance. The Wistar rats were randomly divided into 5 groups, including the control group (no swimming), the swimming group (free swimming), and 3 treatment groups treated with the maca extract at the doses of 4.0, 5.3 and 8.0 g/kg body weight. The animals in swimming and treatment groups were then for free swimming in the circulating water flow daily for 15 days. On the 16 th day after swimming endurance, the spinal and muscular tissues were collected from all groups. The mitochondrial ultrastructures of the neurons of the spinal cells were observed with the projection electron microscope, and the levels of the glycogen, malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and Ca 2+ in muscle tissues were determined by the RIA method. When rats were treated with maca extract (at 4.0, 5.3, 8.0 g/kg body weight), the total swimming time and the swimming duration before sinking were increased by 19.83%, 60.28%, 77.55%, and 55.34%, 73.91%, 94.47%, respectively, compared with the simple swimming group( P maca extract (4.0, 5.3, 8.0 g/kg body weight) were reduced by 7.79%, 18.18%, 31.17%, 16.95%, 27.34%, 43.31% and 13.51%, 23.19%, 43.15%, respectively. Our results demonstrated the protective effects of maca extract on the mitochondria of spinal cell and suggested that maca extract could improve the muscle antioxidant activity by increasing the levels of SOD, GSH-Px, and muscle glycogen.

The green synthesis of gold nanoparticles using the ethanol extract pf black tea and its tannin free fraction

International Nuclear Information System (INIS)

Banoee, M.; Mokhtari, N.; Akhavan Sepahi, A.; Jafari Fesharaki, P.; Monsef-Esfahani, H. R.; Ehsanfar, Z.; Khoshayand, M. R.; Shahverdi, A. R.

2010-01-01

In this research the ethanol extract of black tea and its tannin free fraction used for green synthesis of gold nanoparticles. All the extracts were used separately for the synthesis of gold nanoparticles through the reduction of aqueous AuCl 4 - . Transmission electron microscopy and visible absorption spectroscopy confirmed the reduction of gold ions to gold nanoparticles. The ethanol extract of black tea and its tannin free ethanol extract produced gold nanoparticles in the size ranges of 2.5-27.5 nm and 1.25-17.5 nm with an average size of 10 nm and 3 nm, respectively. The prepared colloid gold nanoparticles, using the ethanol extract of black tea, did not show the appropriate stability during storage time (24 hours) at 4 d eg C . In contrast, gold colloids, which were synthesized by a tannin free fraction showed no particle aggregation during short and long storage times at the same conditions. To the best of our knowledge, this is the first report on the rapid synthesis of gold nanoparticles using ethanol extract of black tea and its tannin free fraction.

[Inhibition effects of black rice pericarp extracts on cell proliferation of PC-3 cells].

Science.gov (United States)

Jiang, Weiwei; Yu, Xudong; Ren, Guofeng

2013-05-01

To observe the inhibitive effects of black rice pericarp extracts on cell proliferation of human prostate cancer cell PC-3 and to explore its effecting mechanism. The black rice pericarp extract was used to treat the PC-3 cells. The inhibitory effect of black rice pericarp extract on cells proliferation of PC-3 was tested by MTT method. Cell apoptosis rates and cell cycle were measured by flow cytometric assay (FCM). Western blot was used to study the protein expression levels of p38, p-p38, JNK, p-JNK. A dose-dependent and time-dependent proliferation inhibition of black rice pericarp extract was demonstrated in PC-3. The most prominent experiment condition was inhibitory concentration with 300microg/ml and treated for 72 h. The experiment result of flow cytometry analysis demonstrates that the apoptosis rate of PC-3 cells increased along with the increasing of black rice pericarp extract concentration, and a G1-S cell cycle arrest was induced in a dose-dependent manner. After PC-3 cell was treated with black rice pericarp extract for 72 h, the expressions of p-p38, p-JNK protein increased. Black rice pericarp extract could inhibit proliferation, change the cell cycle distributions and induce apoptosis in human prostatic cancer cell PC-3. Its inhibitory effect may be through promoting activation of the JNK, p38 signaling pathway. These results suggest that black rice pericarp extract maybe has an inhibitory effect on prostatic cancer.

Environmentally Friendly Plasma-Treated PEDOT:PSS as Electrodes for ITO-Free Perovskite Solar Cells.

Science.gov (United States)

Vaagensmith, Bjorn; Reza, Khan Mamun; Hasan, Md Nazmul; Elbohy, Hytham; Adhikari, Nirmal; Dubey, Ashish; Kantack, Nick; Gaml, Eman; Qiao, Qiquan

2017-10-18

Solution processed poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) transparent electrodes (TEs) offer great potential as a low cost alternative to expensive indium tin oxide (ITO). However, strong acids are typically used for enhancing the conductivity of PEDOT:PSS TEs, which produce processing complexity and environmental issues. This work presents an environmentally friendly acid free approach to enhance the conductivity of PEDOT:PSS using a light oxygen plasma treatment, in addition to solvent blend additives and post treatments. The plasma treatment was found to significantly reduce the sheet resistance of PEDOT:PSS TEs from 85 to as low as 15 Ω sq -1 , which translates to the highest reported conductivity of 5012 S/cm for PEDOT:PSS TEs. The plasma treated PEDOT:PSS TE resulted in an ITO-free perovskite solar cell efficiency of 10.5%, which is the highest reported efficiency for ITO-free perovskite solar cells with a PEDOT:PSS electrode that excludes the use of acid treatments. This research presents the first demonstration of this technology. Moreover, the PEDOT:PSS TEs enabled better charge extraction from the perovskite solar cells and reduced hysteresis in the current density-voltage (J-V) curves.

Oral epithelial cells are susceptible to cell-free and cell-associated HIV-1 infection in vitro

International Nuclear Information System (INIS)

Moore, Jennifer S.; Rahemtulla, Firoz; Kent, Leigh W.; Hall, Stacy D.; Ikizler, Mine R.; Wright, Peter F.; Nguyen, Huan H.; Jackson, Susan

2003-01-01

Epithelial cells lining the oral cavity are exposed to HIV-1 through breast-feeding and oral-genital contact. Genital secretions and breast milk of HIV-1-infected subjects contain both cell-free and cell-associated virus. To determine if oral epithelial cells can be infected with HIV-1 we exposed gingival keratinocytes and adenoid epithelial cells to cell-free virus and HIV-1-infected peripheral blood mononuclear cells and monocytes. Using primary isolates we determined that gingival keratinocytes are susceptible to HIV-1 infection via cell-free CD4-independent infection only. R5 but not X4 viral strains were capable of infecting the keratinocytes. Further, infected cells were able to release infectious virus. In addition, primary epithelial cells isolated from adenoids were also susceptible to infection; both cell-free and cell-associated virus infected these cells. These data have potential implications in the transmission of HIV-1 in the oral cavity

The influence of protein free calf blood extract eye gel on dry eye after pterygium surgery

Directory of Open Access Journals (Sweden)

Cai-Ni Ji

2013-07-01

Full Text Available AIM: To investigate the influence of protein free calf blood extract eye gel on dry eye after pterygium surgery. METHODS: Thirty six patients(40 eyeswith primary nasal pterygium were enrolled in this study, which were divided into study group and control group randomly, with 20 eyes in each group. All patients received pterygium excision and limbal stem cell autograft surgery and tobramicin dexamethasone eye drops after surgery. Patients of the study group received protein free calf blood extract eye gel while those of the control group received 0.1% sodium hyaluronate eye drops furthermore. Ocular surface disease index(OSDIquestionnaire, tear film break-up time(BUTand Schirmer's Ⅰ test Ⅰ(SⅠtwere carried before and 3 months after surgery to evaluate the dry eye degree of the patients. RESULTS: There was no statistical difference between the age, gender and size of the pterygium of the study and control groups preoperatively. There was no statistical difference between the OSDI(2.33±1.02 vs 2.32±0.93, BUT(8.80±2.48 vs 8.35±2.28seconds and SⅠt(4.30±2.30 vs 4.40±2.44of the two groups preoperatively. There was statistical difference between the OSDI(1.45±0.47 vs 1.81±0.60, BUT(11.20±2.07 vs 9.50±2.40seconds and SⅠt(8.35±3.13 vs 6.35±2.18of the two groups 3 months postoperatively, which was also different from that of the preoperative data correspondingly. CONCLUSION: Protein free calf blood extract eye gel could reduce the dry eye after pterygium surgery.

Label-free cell-cycle analysis by high-throughput quantitative phase time-stretch imaging flow cytometry

Science.gov (United States)

Mok, Aaron T. Y.; Lee, Kelvin C. M.; Wong, Kenneth K. Y.; Tsia, Kevin K.

2018-02-01

Biophysical properties of cells could complement and correlate biochemical markers to characterize a multitude of cellular states. Changes in cell size, dry mass and subcellular morphology, for instance, are relevant to cell-cycle progression which is prevalently evaluated by DNA-targeted fluorescence measurements. Quantitative-phase microscopy (QPM) is among the effective biophysical phenotyping tools that can quantify cell sizes and sub-cellular dry mass density distribution of single cells at high spatial resolution. However, limited camera frame rate and thus imaging throughput makes QPM incompatible with high-throughput flow cytometry - a gold standard in multiparametric cell-based assay. Here we present a high-throughput approach for label-free analysis of cell cycle based on quantitative-phase time-stretch imaging flow cytometry at a throughput of > 10,000 cells/s. Our time-stretch QPM system enables sub-cellular resolution even at high speed, allowing us to extract a multitude (at least 24) of single-cell biophysical phenotypes (from both amplitude and phase images). Those phenotypes can be combined to track cell-cycle progression based on a t-distributed stochastic neighbor embedding (t-SNE) algorithm. Using multivariate analysis of variance (MANOVA) discriminant analysis, cell-cycle phases can also be predicted label-free with high accuracy at >90% in G1 and G2 phase, and >80% in S phase. We anticipate that high throughput label-free cell cycle characterization could open new approaches for large-scale single-cell analysis, bringing new mechanistic insights into complex biological processes including diseases pathogenesis.

Snake venomics of Crotalus tigris: the minimalist toxin arsenal of the deadliest Nearctic rattlesnake venom. Evolutionary Clues for generating a pan-specific antivenom against crotalid type II venoms [corrected].

Science.gov (United States)

Calvete, Juan J; Pérez, Alicia; Lomonte, Bruno; Sánchez, Elda E; Sanz, Libia

2012-02-03

We report the proteomic and antivenomic characterization of Crotalus tigris venom. This venom exhibits the highest lethality for mice among rattlesnakes and the simplest toxin proteome reported to date. The venom proteome of C. tigris comprises 7-8 gene products from 6 toxin families; the presynaptic β-neurotoxic heterodimeric PLA(2), Mojave toxin, and two serine proteinases comprise, respectively, 66 and 27% of the C. tigris toxin arsenal, whereas a VEGF-like protein, a CRISP molecule, a medium-sized disintegrin, and 1-2 PIII-SVMPs each represent 0.1-5% of the total venom proteome. This toxin profile really explains the systemic neuro- and myotoxic effects observed in envenomated animals. In addition, we found that venom lethality of C. tigris and other North American rattlesnake type II venoms correlates with the concentration of Mojave toxin A-subunit, supporting the view that the neurotoxic venom phenotype of crotalid type II venoms may be described as a single-allele adaptation. Our data suggest that the evolutionary trend toward neurotoxicity, which has been also reported for the South American rattlesnakes, may have resulted by pedomorphism. The ability of an experimental antivenom to effectively immunodeplete proteins from the type II venoms of C. tigris, Crotalus horridus , Crotalus oreganus helleri, Crotalus scutulatus scutulatus, and Sistrurus catenatus catenatus indicated the feasibility of generating a pan-American anti-Crotalus type II antivenom, suggested by the identification of shared evolutionary trends among South and North American Crotalus species.

The emerging age of cell-free synthetic biology.

Science.gov (United States)

Smith, Mark Thomas; Wilding, Kristen M; Hunt, Jeremy M; Bennett, Anthony M; Bundy, Bradley C

2014-08-25

The engineering of and mastery over biological parts has catalyzed the emergence of synthetic biology. This field has grown exponentially in the past decade. As increasingly more applications of synthetic biology are pursued, more challenges are encountered, such as delivering genetic material into cells and optimizing genetic circuits in vivo. An in vitro or cell-free approach to synthetic biology simplifies and avoids many of the pitfalls of in vivo synthetic biology. In this review, we describe some of the innate features that make cell-free systems compelling platforms for synthetic biology and discuss emerging improvements of cell-free technologies. We also select and highlight recent and emerging applications of cell-free synthetic biology. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Efficient cell-free production of olfactory receptors: detergent optimization, structure, and ligand binding analyses.

Science.gov (United States)

Kaiser, Liselotte; Graveland-Bikker, Johanna; Steuerwald, Dirk; Vanberghem, Mélanie; Herlihy, Kara; Zhang, Shuguang

2008-10-14

High-level production of membrane proteins, particularly of G protein-coupled receptors (GPCRs) in heterologous cell systems encounters a number of difficulties from their inherent hydrophobicity in their transmembrane domains, which frequently cause protein aggregation and cytotoxicity and thus reduce the protein yield. Recent advances in cell-free protein synthesis circumvent those problems to produce membrane proteins with a yield sometimes exceeding the cell-based approach. Here, we report cell-free production of a human olfactory receptor 17-4 (hOR17-4) using the wheat germ extract. Using the simple method, we also successful produced two additional olfactory receptors. To obtain soluble olfactory receptors and to increase yield, we directly added different detergents in varying concentrations to the cell-free reaction. To identify a purification buffer system that maintained the receptor in a nonaggregated form, we developed a method that uses small-volume size-exclusion column chromatography combined with rapid and sensitive dot-blot detection. Different buffer components including salt concentration, various detergents and detergent concentration, and reducing agent and its concentrations were evaluated for their ability to maintain the cell-free produced protein stable and nonaggregated. The purified olfactory receptor displays a typical a alpha-helical CD spectrum. Surface plasmon resonance measurements were used to show binding of a known ligand undecanal to hOR17-4. Our approach to produce a high yield of purified olfactory receptor is a milestone toward obtaining a large quantity of olfactory receptors for designing bionic sensors. Furthermore, this simple approach may be broadly useful not only for other classes of GPCRs but also for other membrane proteins.

Antinociceptive and free radical scavenging activities of Cocos nucifera L. (Palmae) husk fiber aqueous extract.

Science.gov (United States)

Alviano, Daniela S; Rodrigues, Karen F; Leitão, Suzana G; Rodrigues, Marcio L; Matheus, Maria Eline; Fernandes, Patrícia D; Antoniolli, Angelo R; Alviano, Celuta S

2004-06-01

In the current study, the analgesic and free radical scavenging properties of an aqueous extract from the husk fiber of Cocos nucifera L. (Palmae) were demonstrated by the use of in vivo and in vitro models. The orally administered Cocos nucifera aqueous extract (200 or 400 mg/kg) inhibited the acetic acid-induced writhing response in mice. Tail flick and hot plate assays demonstrated that treatment of animals with this plant extract at 200 mg/kg induced attenuation in the response to a heat stimulus. A LD(50) of 2.30 g/kg was obtained in acute toxicity tests. Topic treatment of rabbits with the Cocos nucifera extract indicated that it does not induce any significant dermic or ocular irritation. In vitro experiments using the 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) photometric assay demonstrated that this plant extract also possesses free radical scavenging properties.

Radioprotective effects of hawthorn fruit extract against gamma irradiation in mouse bone marrow cells

International Nuclear Information System (INIS)

Hosseinimehr, S.J.; Azadbakht, M.; Mousavi, S.M.; Mahmoudzadeh, A.; Akhlaghpoor, S.

2007-01-01

The radioprotective effect of hawthorn (Crataegus microphylla) fruit extract against genotoxicity induced by gamma irradiation has been investigated in mouse bone marrow cells. A single intraperitoneal (ip) administration of hawthorn extract at doses of 25, 50, 100 and 200 mg/kg 1 h prior to gamma irradiation (2 Gy) reduced the frequencies of micronucleated polychromatic erythrocytes (MnPCEs). All four doses of hawthorn extract significantly reduced the frequencies of MnPCEs and increased the PCE/PCE+NCE ratio (polychromatic erythrocyte/polychromatic erythrocyte+normochromatic erythrocyte) in mice bone marrow compared with the non drug-treated irradiated control (p<0.02-0.00001). The maximum reduction in MnPCEs was observed in mice treated with extract at a dose of 200 mg/kg. Administration of amifostine at dose 100 mg/kg and hawthorn at dose 200 mg/kg reduced the frequency of MnPCE almost 4.8 and 5.7 fold; respectively, after being exposed to 2 Gy of gamma rays, compare with the irradiated control group. Crataegus extract exhibited concentration-dependent activity on 1, 1-diphenyl 2-picrylhydrazyl free radical showing that Crataegus contained high amounts of phenolic compounds and the high performance liquid chromatography (HPLC) analysis determined that it contained chlorogenic acid, epicatechin and hyperoside. It appeared that hawthorn extract with antioxidant activity reduced the genotoxicity induced by gamma irradiation in bone marrow cells. (author)

Preparative scale production of functional mouse aquaporin 4 using different cell-free expression modes.

Directory of Open Access Journals (Sweden)

Lei Kai

Full Text Available The continuous progress in the structural and functional characterization of aquaporins increasingly attracts attention to study their roles in certain mammalian diseases. Although several structures of aquaporins have already been solved by crystallization, the challenge of producing sufficient amounts of functional proteins still remains. CF (cell free expression has emerged in recent times as a promising alternative option in order to synthesize large quantities of membrane proteins, and the focus of this report was to evaluate the potential of this technique for the production of eukaryotic aquaporins. We have selected the mouse aquaporin 4 as a representative of mammalian aquaporins. The protein was synthesized in an E. coli extract based cell-free system with two different expression modes, and the efficiencies of two modes were compared. In both, the P-CF (cell-free membrane protein expression as precipitate mode generating initial aquaporin precipitates as well as in the D-CF (cell-free membrane protein expression in presence of detergent mode, generating directly detergent solubilized samples, we were able to obtain mg amounts of protein per ml of cell-free reaction. Purified aquaporin samples solubilized in different detergents were reconstituted into liposomes, and analyzed for the water channel activity. The calculated P(f value of proteoliposome samples isolated from the D-CF mode was 133 µm/s at 10°C, which was 5 times higher as that of the control. A reversible inhibitory effect of mercury chloride was observed, which is consistent with previous observations of in vitro reconstituted aquaporin 4. In this study, a fast and convenient protocol was established for functional expression of aquaporins, which could serve as basis for further applications such as water filtration.

The ability of Abelmoschus manihot L. leaf extract in scavenging of free radical DPPH and total flavonoid determination

Science.gov (United States)

Sudewi, S.; Lolo, W. A.; Warongan, M.; Rifai, Y.; Rante, H.

2017-11-01

Abelmoschus manihot L. has reported to have flavonoids content. This study aims were to determine the ability of A. manihot extract in counteracting free radical DPPH and determine the content of total flavonoids. A. manihot leaf was taken from 2 regions in North Sulawesi, namely Tomohon and Kotamobagu. The maceration was carried out to extract the active compound in a 96% ethanol solvent. Free radical scavenging analysis was carried out by DPPH and determination of its total flavonoid in the extract was measured using spectrophotometri method. The results showed that A. manihot extract from Tomohon and Kotamobagu could counteract free radical of DPPH with value of free radical activity of 88.151 and 88.801 %, respectively. A. manihot leaf from Kotamobagu has higher total flavonoids content 61.763 mg/g compare to Tomohon 46.679 mg/g which presented as quercetin. A. manihot has antioxidant activity.

Evidence for a Rad18-independent frameshift mutagenesis pathway in human cell-free extracts.

Directory of Open Access Journals (Sweden)

Régine Janel-Bintz

Full Text Available Bypass of replication blocks by specialized DNA polymerases is crucial for cell survival but may promote mutagenesis and genome instability. To gain insight into mutagenic sub-pathways that coexist in mammalian cells, we examined N-2-acetylaminofluorene (AAF-induced frameshift mutagenesis by means of SV40-based shuttle vectors containing a single adduct. We found that in mammalian cells, as previously observed in E. coli, modification of the third guanine of two target sequences, 5'-GGG-3' (3G and 5'-GGCGCC-3' (NarI site, induces -1 and -2 frameshift mutations, respectively. Using an in vitro assay for translesion synthesis, we investigated the biochemical control of these events. We showed that Pol eta, but neither Pol iota nor Pol zeta, plays a major role in the frameshift bypass of the AAF adduct located in the 3G sequence. By complementing PCNA-depleted extracts with either a wild-type or a non-ubiquitinatable form of PCNA, we found that this Pol eta-mediated pathway requires Rad18 and ubiquitination of PCNA. In contrast, when the AAF adduct is located within the NarI site, TLS is only partially dependent upon Pol eta and Rad18, unravelling the existence of alternative pathways that concurrently bypass this lesion.

studies on solvent extraction of free hydrogen cyanide from river water

African Journals Online (AJOL)

A method for free and strongly complexed cyanide measurement in river water was developed. Recovery tests from solution with and without river water, using various solvent combinations and background control were investigated to obtain an accurate and precise extraction method for the measurement of hydrogen ...

Supercritical fluid extraction of uranium and thorium using modifier free delivery of ligands

International Nuclear Information System (INIS)

Sujatha, K.; Kumar, R.; Sivaraman, N.; Srinivasan, T.G.; Vasudeva Rao, P.R.

2009-01-01

The modifier free controlled delivery of octyl (phenyl)-N,N-diisobutylcarbamoylmethy phosphineoxide (CMPO) using supercritical carbon dioxide was established for the extraction of uranyl nitrate as well as uranyl nitrate sorbed on tissue paper matrix and the results were compared with modifier method. The preferential extraction of uranium over thorium was also demonstrated using di (2-ethylhexyl)isobutyramide (D2EHIBA). (author)

Comparison of the free and bound phenolic profiles and cellular antioxidant activities of litchi pulp extracts from different solvents

Science.gov (United States)

2014-01-01

Background The phenolic contents and antioxidant activities of fruits could be underestimated if the bound phenolic compounds are not considered. In the present study, the extraction efficiencies of various solvents were investigated in terms of the total content of the free and bound phenolic compounds, as well as the phenolic profiles and antioxidant activities of the extracts. Methods Five different solvent mixtures were used to extract the free phenolic compounds from litchi pulp. Alkaline and acidic hydrolysis methods were compared for the hydrolysis of bound phenolic compounds from litchi pulp residue. The phenolic compositions of the free and bound fractions from the litchi pulp were identified using HPLC-DAD. The antioxidant activities of the litchi pulp extracts were determined by oxygen radical absorbance capacity (ORAC) and cellular antioxidant activity (CAA) assays. Results Of the solvents tested, aqueous acetone extracted the largest amount of total free phenolic compounds (210.7 mg GAE/100 g FW) from litchi pulp, followed sequentially by aqueous mixtures of methanol, ethanol and ethyl acetate, and water itself. The acid hydrolysis method released twice as many bound phenolic compounds as the alkaline hydrolysis method. Nine phenolic compounds were detected in the aqueous acetone extract. In contrast, not all of these compounds were found in the other four extracts. The classification and content of the bound phenolic compounds released by the acid hydrolysis method were higher than those achieved by the alkaline hydrolysis. The aqueous acetone extract showing the highest ORAC value (3406.9 μmol TE/100 g FW) for the free phenolic extracts. For the CAA method, however, the aqueous acetone and methanol extracts (56.7 and 55.1 μmol QE/100 g FW) showed the highest levels of activity of the five extracts tested. The ORAC and CAA values of the bound phenolic compounds obtained by acid hydrolysis were 2.6- and 1.9-fold higher than those obtained using the

Origanum vulgare leaf extract protects mice bone marrow cells against ionizing radiation

Directory of Open Access Journals (Sweden)

Reza Ghasemnezhad Targhi

2016-11-01

Full Text Available Objective: Ionizing radiation produces free radicals which induce DNA damage and cell death. Origanum vulgare leaf extract (OVLE is a natural compound and its capability of scavenging free radicals and its antioxidant activity have been demonstrated by many researchers. In this study, using micronucleus assay, radioprotective effect of OVLE against clastogenic and cytotoxic effect of gamma irradiation has been investigated in mice bone marrow cells. Materials and Methods: OVLE was injected intraperitoneally to the BALB/c mice 1hr prior to gamma irradiation (3Gy at the doses of 100 and 200 mg/kg. Twenty four hours after irradiation or treatment, animals were killed and smears were prepared from the bone marrow cells. The slides were stained with May Grunwald–Giemsa method and analyzed microscopically. The frequency of micronucleated polychromatic erythrocytes (MnPCEs, micronucleated normochromatic erythrocyte (MnNCEs and cell proliferation ratio PCE/PCE+NCE (polychromatic erythrocyte/polychromatic erythrocyte + normochromatic erythrocyte were calculated. Results: The results showed that gamma irradiation (3Gy increased the frequency of MnPCEs, MnNCEs and  reduced the PCE/PCE+NCE ratio in mice bone marrow compared to the non-irradiated control group (p< 0.0001. Injection of OVLE significantly reduced the frequency of MnPCEs (p< 0.0001 and MnNCEs (p< 0.05 and increased the PCE/PCE+NCE ratio as compared to the irradiated control group (p< 0.05. Conclusion: It seems that OVLE with its antioxidant properties and its capability of scavenging free radicals and reactive oxygen species can reduce the cytotoxic effects of gamma irradiation in mice bone marrow cells.

Hydrologic test results for the Rattlesnake Ridge interbed and Pomona basalt flow top at Borehole DB-15

International Nuclear Information System (INIS)

Strait, S.R.; Brown, W.R.

1983-07-01

This report presents results and description of hydrologic test activities for the Rattlesnake Ridge interbed and Pomona basalt flow top at Borehole DB-15. Hydrologic tests conducted include constant discharge air-lift and constant discharge submersible pumping tests. An observed hydraulic head for the test interval was 409 ± 1 feet above mean sea level. Transmissivity values determined from hydrologic tests performed, ranged between 493 and 469 ft 2 /day. The best estimate of transmissivity is 480 ft 2 /day. The best estimate of equivalent hydraulic conductivity, based on an effective test thickness of 56 feet is 8.6 ft/day. 4 refs., 7 figs., 2 tabs

Cell-free protein synthesis: applications in proteomics and biotechnology.

Science.gov (United States)

He, Mingyue

2008-01-01

Protein production is one of the key steps in biotechnology and functional proteomics. Expression of proteins in heterologous hosts (such as in E. coli) is generally lengthy and costly. Cell-free protein synthesis is thus emerging as an attractive alternative. In addition to the simplicity and speed for protein production, cell-free expression allows generation of functional proteins that are difficult to produce by in vivo systems. Recent exploitation of cell-free systems enables novel development of technologies for rapid discovery of proteins with desirable properties from very large libraries. This article reviews the recent development in cell-free systems and their application in the large scale protein analysis.

Protective effect of hexane and ethanol extract of piper longum L. On gentamicin-induced hair cell loss in neonatal cultures.

Science.gov (United States)

Yadav, Mukesh Kumar; Choi, June; Song, Jae-Jun

2014-03-01

Gentamicin (GM) is a commonly used aminoglycoside antibiotic that generates free oxygen radicals within the inner ear, which can cause vestibulo-cochlear toxicity and permanent damage to the sensory hair cells and neurons. Piper longum L. (PL) is a well-known spice and traditional medicine in Asia and Pacific islands, which has been reported to exhibit a wide spectrum of activity, including antioxidant activity. In this study, we evaluated the effect of hexane:ethanol (2:8) PL extract (subfraction of PL [SPL] extract) on GM-induced hair cell loss in basal, middle and apical regions in a neonatal cochlea cultures. The protective effects of SPL extract were measured by phalloidin staining of cultures from postnatal day 2-3 mice with GM-induced hair cell loss. The anti-apoptosis activity of SPL extract was measured using double labeling by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and myosin-7a staining. The radical-scavenging activity of SPL extract was assessed using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay. SPL extract at a concentration of 1 µg/mL significantly inhibited GM-induced hair cell loss at basal and middle region of cochlea, while 5 µg/mL was effective against apical region hair cell loss. The protective effect of SPL extract was concentration dependent and hair cells retained their stereocilia in explants treated with SPL extract prior to treatment with 0.3 mM GM. SPL extract decreased GM-induced apoptosis of hair cells as assessed by TUNEL staining. The outer hair and inner hair counts were not decreased in SPL extract treated groups in compare to GM treated explants. Additionally, SPL extract showed concentration dependent radical scavenging activity in a DPPH assay. An anti-apoptosis effect and potent radical scavenger activity of SPL extract protects from GM-induced hair cell loss at basal, middle and apical regions in neonatal cochlea cultures.

High yield cell-free production of integral membrane proteins without refolding or detergents.

Science.gov (United States)

Wuu, Jessica J; Swartz, James R

2008-05-01

Integral membrane proteins act as critical cellular components and are important drug targets. However, difficulties in producing membrane proteins have hampered investigations of structure and function. In vivo production systems are often limited by cell toxicity, and previous in vitro approaches have required unnatural folding pathways using detergents or lipid solutions. To overcome these limitations, we present an improved cell-free expression system which produces high yields of integral membrane proteins without the use of detergents or refolding steps. Our cell-free reaction activates an Escherichia coli-derived cell extract for transcription and translation. Purified E. coli inner membrane vesicles supply membrane-bound components and the lipid environment required for insertion and folding. Using this system, we demonstrated successful synthesis of two complex integral membrane transporters, the tetracycline pump (TetA) and mannitol permease (MtlA), in yields of 570+/-50 microg/mL and 130+/-30 microg/mL of vesicle-associated protein, respectively. These yields are up to 400 times typical in vivo concentrations. Insertion and folding of these proteins are verified by sucrose flotation, protease digestion, and activity assays. Whereas TetA incorporates efficiently into vesicle membranes with over two-thirds of the synthesized protein being inserted, MtlA yields appear to be limited by insufficient concentrations of a membrane-associated chaperone.

Metabolite extraction from adherently growing mammalian cells for metabolomics studies: optimization of harvesting and extraction protocols.

Science.gov (United States)

Dettmer, Katja; Nürnberger, Nadine; Kaspar, Hannelore; Gruber, Michael A; Almstetter, Martin F; Oefner, Peter J

2011-01-01

Trypsin/ethylenediaminetetraacetic acid (EDTA) treatment and cell scraping in a buffer solution were compared for harvesting adherently growing mammalian SW480 cells for metabolomics studies. In addition, direct scraping with a solvent was tested. Trypsinated and scraped cell pellets were extracted using seven different extraction protocols including pure methanol, methanol/water, pure acetone, acetone/water, methanol/chloroform/water, methanol/isopropanol/water, and acid-base methanol. The extracts were analyzed by GC-MS after methoximation/silylation and derivatization with propyl chloroformate, respectively. The metabolic fingerprints were compared and 25 selected metabolites including amino acids and intermediates of energy metabolism were quantitatively determined. Moreover, the influence of freeze/thaw cycles, ultrasonication and homogenization using ceramic beads on extraction yield was tested. Pure acetone yielded the lowest extraction efficiency while methanol, methanol/water, methanol/isopropanol/water, and acid-base methanol recovered similar metabolite amounts with good reproducibility. Based on overall performance, methanol/water was chosen as a suitable extraction solvent. Repeated freeze/thaw cycles, ultrasonication and homogenization did not improve overall metabolite yield of the methanol/water extraction. Trypsin/EDTA treatment caused substantial metabolite leakage proving it inadequate for metabolomics studies. Gentle scraping of the cells in a buffer solution and subsequent extraction with methanol/water resulted on average in a sevenfold lower recovery of quantified metabolites compared with direct scraping using methanol/water, making the latter one the method of choice to harvest and extract metabolites from adherently growing mammalian SW480 cells.

Homogenate-assisted Vacuum-powered Bubble Extraction of Moso Bamboo Flavonoids for On-line Scavenging Free Radical Capacity Analysis.

Science.gov (United States)

Sun, Yinnan; Yang, Kui; Cao, Qin; Sun, Jinde; Xia, Yu; Wang, Yinhang; Li, Wei; Ma, Chunhui; Liu, Shouxin

2017-07-11

A homogenate-assisted vacuum-powered bubble extraction (HVBE) method using ethanol was applied for extraction of flavonoids from Phyllostachys pubescens (P. pubescens) leaves. The mechanisms of homogenate-assisted extraction and vacuum-powered bubble generation were discussed in detail. Furthermore, a method for the rapid determination of flavonoids by HPLC was established. HVBE followed by HPLC was successfully applied for the extraction and quantification of four flavonoids in P. pubescens , including orientin, isoorientin, vitexin, and isovitexin. This method provides a fast and effective means for the preparation and determination of plant active components. Moreover, the on-line antioxidant capacity, including scavenging positive ion and negative ion free radical capacity of different fractions from the bamboo flavonoid extract was evaluated. Results showed that the scavenging DPPH ˙ free radical capacity of vitexin and isovitexin was larger than that of isoorientin and orientin. On the contrary, the scavenging ABTS⁺ ˙ free radical capacity of isoorientin and orientin was larger than that of vitexin and isovitexin.

Chemopreventive effects of free and bound phenolics associated to steep waters (nejayote) obtained after nixtamalization of different maize types.

Science.gov (United States)

Rojas-García, Carlos; García-Lara, Silverio; Serna-Saldivar, Sergio O; Gutiérrez-Uribe, Janet A

2012-03-01

Free and bound phenolics extracts from nejayote solids were obtained after optimally lime-cooking blue, normal white, red, normal yellow, high-carotenoid and quality protein maize types. The extraction yield ranged from 4.47 to 10.05%. Bound phenolics extracts had higher content of total phenolics, antioxidant activity and ferulic acid compared to the free phenolics extracts. In general, free phenolics extracts were less cytotoxic than the bound phenolics counterparts. Bound phenolics extracts had higher induction of quinone reductase (QR) and particularly the normal yellow nejayote exerted the highest chemopreventive index tested in Hepa1c1c7 cells. When tested for monofunctional phase 2 induction capacity in BPrc1 cells, the bound phenolics extracts of blue, normal white and quality protein nejayotes were better inducers than the normal yellow counterpart. Particularly, the free phenolics extract of the white maize nejayote induced BPrc1 cells QR and exerted a higher chemopreventive index compared to the bound phenolics extract. Therefore, the nejayote of the normal white maize was the best source of monofunctional phase 2 enzyme inducers.

Application of 2k Full Factorial Design in Optimization of Solvent-Free Microwave Extraction of Ginger Essential Oil

Directory of Open Access Journals (Sweden)

Mumtaj Shah

2014-01-01

Full Text Available The solvent-free microwave extraction of essential oil from ginger was optimized using a 23 full factorial design in terms of oil yield to determine the optimum extraction conditions. Sixteen experiments were carried out with three varying parameters, extraction time, microwave power, and type of sample for two levels of each. A first order regression equation best fits the experimental data. The predicted values calculated by the regression model were in good agreement with the experimental values. The results showed that the extraction time is the most prominent factor followed by microwave power level and sample type for extraction process. An average of 0.25% of ginger oil can be extracted using current setup. The optimum conditions for the ginger oil extraction using SFME were the extraction time 30 minutes, microwave power level 640 watts, and sample type, crushed sample. Solvent-free microwave extraction proves a green and promising technique for essential oil extraction.

Radioprotective effects of Asparagus racemosus extracts against free radical damage in rat liver mitochondria

International Nuclear Information System (INIS)

Boloor, K.K.; Kamat, J.P.; Devasagayam, T.P.A.; Venkatachalam, S.R.

2000-01-01

The possible antioxidant effect of the extracts of Asparagus racemosus against membrane damage induced by free radicals generated during γ-radiation was examined in rat liver/brain mitochondria. These extracts displayed significant antioxidant properties in mitochondria against oxidation of both lipids and proteins as assessed by lipid peroxidation, protein oxidation and depletion of thiols. The inhibitory effect of the extracts, rich in polysaccharides like galactose, was more than that of the established antioxidants glutathione and ascorbic acid. (author)

Artificial intelligence in label-free microscopy biological cell classification by time stretch

CERN Document Server

Mahjoubfar, Ata; Jalali, Bahram

2017-01-01

This book introduces time-stretch quantitative phase imaging (TS-QPI), a high-throughput label-free imaging flow cytometer developed for big data acquisition and analysis in phenotypic screening. TS-QPI is able to capture quantitative optical phase and intensity images simultaneously, enabling high-content cell analysis, cancer diagnostics, personalized genomics, and drug development. The authors also demonstrate a complete machine learning pipeline that performs optical phase measurement, image processing, feature extraction, and classification, enabling high-throughput quantitative imaging that achieves record high accuracy in label -free cellular phenotypic screening and opens up a new path to data-driven diagnosis. • Demonstrates how machine learning is used in high-speed microscopy imaging to facilitate medical diagnosis; • Provides a systematic and comprehensive illustration of time stretch technology; • Enables multidisciplinary application, including industrial, biomedical, and artificial intell...

Evaluation of in vitro anti-inflammatory effects of crude ginger and rosemary extracts obtained through supercritical CO2 extraction on macrophage and tumor cell line: the influence of vehicle type.

Science.gov (United States)

Justo, Oselys Rodriguez; Simioni, Patricia Ucelli; Gabriel, Dirce Lima; Tamashiro, Wirla Maria da Silva Cunha; Rosa, Paulo de Tarso Vieira; Moraes, Ângela Maria

2015-10-29

Numerous plants from have been investigated due to their anti-inflammatory activity and, among then, extracts or components of ginger (Zingiber officinale Roscoe) and rosemary (Rosmarinus officinalis L.), sources of polyphenolic compounds. 6-gingerol from ginger rhizome and carnosic acid and carnosol from rosemary leaves present anti-tumor, anti-inflammatory and antioxidant activities. However, the evaluation of the mechanisms of action of these and other plant extracts is limited due to their high hydrophobicity. Dimethylsulfoxide (DMSO) is commonly used as a vehicle of liposoluble materials to mammalian cells in vitro, presenting enhanced cell penetration. Liposomes are also able to efficiently deliver agents to mammalian cells, being capable to incorporate in their structure not only hydrophobic molecules, but also hydrophilic and amphiphilic compounds. Another strategy is based on the use of Pluronic F-68, a biocompatible low-foaming, non-ionic surfactant, to disperse hydrophobic components. Here, these three delivery approaches were compared to analyze their influence on the in vitro anti-inflammatory effects of ginger and rosemary extracts, at different concentrations, on primary mammalian cells and on a tumor cell line. Ginger and rosemary extracts free of organic solvents were obtained by supercritical fluid extraction and dispersed in DMSO, Pluronic F-68 or liposomes, in variable concentrations. Cell viability, production of inflammatory mediators and nitric oxide (NO) release were measured in vitro on J774 cell line and murine macrophages primary culture stimulated with bacterial lipopolysaccharide and interferon-γ after being exposed or not to these extracts. Ginger and rosemary extracts obtained by supercritical CO2 extraction inhibited the production of pro-inflammatory cytokines and the release of NO by peritoneal macrophages and J774 cells. The delivery vehicles influenced the anti-inflammatory effects. Comparatively, the ginger extract showed the

Reconstitution of the cellular response to DNA damage in vitro using damage-activated extracts from mammalian cells

International Nuclear Information System (INIS)

Roper, Katherine; Coverley, Dawn

2012-01-01

In proliferating mammalian cells, DNA damage is detected by sensors that elicit a cellular response which arrests the cell cycle and repairs the damage. As part of the DNA damage response, DNA replication is inhibited and, within seconds, histone H2AX is phosphorylated. Here we describe a cell-free system that reconstitutes the cellular response to DNA double strand breaks using damage-activated cell extracts and naïve nuclei. Using this system the effect of damage signalling on nuclei that do not contain DNA lesions can be studied, thereby uncoupling signalling and repair. Soluble extracts from G1/S phase cells that were treated with etoposide before isolation, or pre-incubated with nuclei from etoposide-treated cells during an in vitro activation reaction, restrain both initiation and elongation of DNA replication in naïve nuclei. At the same time, H2AX is phosphorylated in naïve nuclei in a manner that is dependent upon the phosphatidylinositol 3-kinase-like protein kinases. Notably, phosphorylated H2AX is not focal in naïve nuclei, but is evident throughout the nucleus suggesting that in the absence of DNA lesions the signal is not amplified such that discrete foci can be detected. This system offers a novel screening approach for inhibitors of DNA damage response kinases, which we demonstrate using the inhibitors wortmannin and LY294002. -- Highlights: ► A cell free system that reconstitutes the response to DNA damage in the absence of DNA lesions. ► Damage-activated extracts impose the cellular response to DNA damage on naïve nuclei. ► PIKK-dependent response impacts positively and negatively on two separate fluorescent outputs. ► Can be used to screen for inhibitors that impact on the response to damage but not on DNA repair. ► LY294002 and wortmannin demonstrate the system's potential as a pathway focused screening approach.

Antiproliferative Properties Against Human Breast, Cervical and Ovarian Cancer Cell Lines, and Antioxidant Capacity of Leaf Aqueous Ethanolic Extract from Cotinus coggygria Scop.

Directory of Open Access Journals (Sweden)

Gospodinova Z.

2017-10-01

Full Text Available Cotinus coggygria Scop. leaf aqueous ethanolic extract was examined for its in vitro antiproliferative and antioxidant activity. Antiproliferative effect was assessed on four human gynecological cancer cell lines: breast (MCF7, T47D, cervical (HeLa and ovarian (A2780 and compared to the cell growth inhibitory effect on non-cancerous breast epithelial cell line MCF10A using MTT cell proliferation assay. Radical scavenging assay with DPPH was applied to evaluate antioxidant potential of the extract. The obtained results showed that the herb inhibited cell growth of all of the tested cancer cell lines and the highest was the cytostatic effect on A2780 cells with a half maximal inhibitory concentration (IC50 value of 30.8 μg/ml. For the other cell lines the IC50 values were in the range of 55-122.7 μg/ml. Additionally, the extract exerted considerably weaker reduction in cell proliferation of the non-cancerous cell line MCF10A compared to cancer cells, which indicates for antiproliferative selectivity. C. coggygria extract showed high free radical scavenging activity with an IC50 value of 11.2 μg/ml. The obtained data provide evidence for pharmacological potential of the tested extract and future more detailed studies concerning the molecular mechanisms of the anticancer effect of the herb are needed.

Wound healing activity and mechanisms of action of an antibacterial protein from the venom of the eastern diamondback rattlesnake (Crotalus adamanteus.

Directory of Open Access Journals (Sweden)

Ramar Perumal Samy

Full Text Available Basic phospholipase A2 was identified from the venom of the eastern diamondback rattlesnake. The Crotalus adamanteus toxin-II (CaTx-II induced bactericidal effects (7.8 µg/ml on Staphylococcus aureus, while on Burkholderia pseudomallei (KHW, and Enterobacter aerogenes were killed at 15.6 µg/ml. CaTx-II caused pore formation and membrane damaging effects on the bacterial cell wall. CaTx-II was not cytotoxic on lung (MRC-5, skin fibroblast (HEPK cells and in mice. CaTx-II-treated mice showed significant wound closure and complete healing by 16 days as compared to untreated controls (**P<0.01. Histological examination revealed enhanced collagen synthesis and neovascularization after treatment with CaTx-II versus 2% Fusidic Acid ointment (FAO treated controls. Measurement of tissue cytokines revealed that interleukin-1 beta (IL-1β expression in CaTx-II treated mice was significantly suppressed versus untreated controls. In contrast, cytokines involved in wound healing and cell migration i.e., monocyte chemotactic protein-1 (MCP-1, fibroblast growth factor-basic (FGF-b, chemokine (KC, granulocyte-macrophage colony-stimulating factor (GM-CSF were significantly enhanced in CaTx-II treated mice, but not in the controls. CaTx-II also modulated nuclear factor-kappa B (NF-κB activation during skin wound healing. The CaTx-II protein highlights distinct snake proteins as a potential source of novel antimicrobial agents with significant therapeutic application for bacterial skin infections.

Ecology and conservation of an endangered rattlesnake, Sistrurus catenatus, in Missouri, USA

Energy Technology Data Exchange (ETDEWEB)

Seigel, R A

1986-01-01

The life history and ecology of the massasauga Sistrurus catenatus, an endangered rattlesnake, was studied from 1979 to 1983 at the Squaw Creek National Wildlife Refuge, Missouri. Except for tail length, this population exhibits little sexual dimorphism. Massasaugas are active from April to October, and are primarily diurnal, except in summer. Snakes are found mainly in a cordgrass prairie in spring and autumn, utilizing drier, upland areas in summer. Mean brood size for this population was 6.35, and there was a significant positive relationship between brood size and female body size. Although this species apparently demonstrates considerable geographic variation in reproductive potential, the significance of this variation cannot be currently assessed. Growth rates, estimated from size-frequency data, suggest an age of maturity of 3-4 years for females. Massasaugas at Squaw Creek feed mainly on rodents and other snakes. Current refuge practices, such as controlled burning of the prairie and unrestricted visitor usage, may have significant negative impacts on Sistrurus populations. Recommendations for mitigating these impacts are provided.

Cell-free synthetic biology for environmental sensing and remediation.

Science.gov (United States)

Karig, David K

2017-06-01

The fields of biosensing and bioremediation leverage the phenomenal array of sensing and metabolic capabilities offered by natural microbes. Synthetic biology provides tools for transforming these fields through complex integration of natural and novel biological components to achieve sophisticated sensing, regulation, and metabolic function. However, the majority of synthetic biology efforts are conducted in living cells, and concerns over releasing genetically modified organisms constitute a key barrier to environmental applications. Cell-free protein expression systems offer a path towards leveraging synthetic biology, while preventing the spread of engineered organisms in nature. Recent efforts in the areas of cell-free approaches for sensing, regulation, and metabolic pathway implementation, as well as for preserving and deploying cell-free expression components, embody key steps towards realizing the potential of cell-free systems for environmental sensing and remediation. Copyright © 2017 The Author. Published by Elsevier Ltd.. All rights reserved.

Number of infection events per cell during HIV-1 cell-free infection.

Science.gov (United States)

Ito, Yusuke; Remion, Azaria; Tauzin, Alexandra; Ejima, Keisuke; Nakaoka, Shinji; Iwasa, Yoh; Iwami, Shingo; Mammano, Fabrizio

2017-07-26

HIV-1 accumulates changes in its genome through both recombination and mutation during the course of infection. For recombination to occur, a single cell must be infected by two HIV strains. These coinfection events were experimentally demonstrated to occur more frequently than would be expected for independent infection events and do not follow a random distribution. Previous mathematical modeling approaches demonstrated that differences in target cell susceptibility can explain the non-randomness, both in the context of direct cell-to-cell transmission, and in the context of free virus transmission (Q. Dang et al., Proc. Natl. Acad. Sci. USA 101:632-7, 2004: K. M. Law et al., Cell reports 15:2711-83, 2016). Here, we build on these notions and provide a more detailed and extensive quantitative framework. We developed a novel mathematical model explicitly considering the heterogeneity of target cells and analysed datasets of cell-free HIV-1 single and double infection experiments in cell culture. Particularly, in contrast to the previous studies, we took into account the different susceptibility of the target cells as a continuous distribution. Interestingly, we showed that the number of infection events per cell during cell-free HIV-1 infection follows a negative-binomial distribution, and our model reproduces these datasets.

Protective effect of aqueous extract from Spirulina platensis against cell death induced by free radicals

OpenAIRE

Radhakrishnan Ammu; Lim Yen-Wei; Chu Wan-Loy; Lim Phaik-Eem

2010-01-01

Abstract Background Spirulina is a commercial alga well known to contain various antioxidants, especially phycocyanin. Apart from being sold as a nutraceutical, Spirulina is incorporated as a functional ingredient in food products and beverages. Most of the previous reports on antioxidant activity of Spirulina were based on chemical rather than cell-based assays. The primary objective of this study was to assess the antioxidant activity of aqueous extract from Spirulina based on its protectiv...

Apoptotic induction of skin cancer cell death by plant extracts.

Science.gov (United States)

Thuncharoen, Walairat; Chulasiri, Malin; Nilwarangkoon, Sirinun; Nakamura, Yukio; Watanapokasin, Ramida

2013-01-01

The aim of the present study was to investigate the effects of plant extracts on cancer apoptotic induction. Human epidermoid carcinoma A431 cell line, obtained from the American Type Culture Collection (ATCC, Manassas, VA), was maintained in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) at 37 degrees C, 5% carbon dioxide (CO2). Plant extract solutions were obtained from S & J international enterprises public company limited. These plant extracts include 50% hydroglycol extracts from Etlingera elatior (Jack) R.M.Smith (torch ginger; EE), Rosa damascene (damask rose; DR) and Rafflesia kerrii Meijer (bua phut; RM). The cell viability, time and dose dependency were determined by MTT (3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) assay. A431 cells were treated with the plant extracts and stained with Hoechst 33342 fluorescent staining dye. Cell viability was demonstrated by the inhibitory concentration 50% (IC50). The anti-proliferative effects were shown to be dependent on time and dose. Typical characteristics of apoptosis which are cell morphological changes and chromatin condensation were clearly observed. The plant extracts was shown to be effective for anti-proliferation and induction of apoptosis cell death in skin cancer cells. Therefore, mechanisms underlying the cell death and its potential use for treatment of skin cancer will be further studied.

Multiscale Capability in Rattlesnake using Contiguous Discontinuous Discretization of Self-Adjoint Angular Flux Equation

Energy Technology Data Exchange (ETDEWEB)

Zheng, Weixiong [Texas A & M Univ., College Station, TX (United States); Wang, Yaqi [Idaho National Lab. (INL), Idaho Falls, ID (United States); DeHart, Mark D. [Idaho National Lab. (INL), Idaho Falls, ID (United States)

2016-09-01

In this report, we present a new upwinding scheme for the multiscale capability in Rattlesnake, the MOOSE based radiation transport application. Comparing with the initial implementation of multiscale utilizing Lagrange multipliers to impose strong continuity of angular flux on interface of in-between subdomains, this scheme does not require the particular domain partitioning. This upwinding scheme introduces discontinuity of angular flux and resembles the classic upwinding technique developed for solving first order transport equation using discontinuous finite element method (DFEM) on the subdomain interfaces. Because this scheme restores the causality of radiation streaming on the interfaces, significant accuracy improvement can be observed with moderate increase of the degrees of freedom comparing with the continuous method over the entire solution domain. Hybrid SN-PN is implemented and tested with this upwinding scheme. Numerical results show that the angular smoothing required by Lagrange multiplier method is not necessary for the upwinding scheme.

ESR detection of free radicals in polyphenolic extracts from wine grapes, olives and green tea

International Nuclear Information System (INIS)

Troup, G.J.; Hutton, D.R.; Romani, A.; Mulinacci, N.; Vincieri, F.F.; Hunter, C.R.; Hewitt, D.G.

1998-01-01

Full text: Polyphenols are widespread in vegetables and fruits and they play an important role in human diet and health: these compounds act mainly as antioxidants and radical scavengers. In this work we have detected free radicals in the following natural polyphenols: Endotelon, an anthocyanic grapeskin extract; malvidin 3,5-O-diglucosides (malvin); oleuropein, an olive polyphenol; a commercial green tea extract, and pure epigallocatechingallate EGCG. The investigation was performed using a Varian E-12 ESR Spectrometer (∼9.1 GHz) at room temperature. All except the green tea extract gave single unstructured lines of ∼ 10 gauss linewidth. The tea extract signal showed 3 lines, one ∼ 20 gauss wide, one ∼ 10 gauss wide, and one ∼ 2-3 gauss wide. Saturation behaviour of these lines at room and liquid N 2 temperature showed them to be from different radicals About 50% of the extract is represented by epigallocatechingallate (EGCG). Using the pure sample EGCG it was possible to assign the appropriate radical, which corresponded with the broadest and strongest 'single' signal (∼20g wide). The presence of the free radicals in the solid extracts shows that the appropriate molecules can act as radical scavengers by forming stable radicals

DNA excision repair in cell extracts from human cell lines exhibiting hypersensitivity to DNA-damaging agents

International Nuclear Information System (INIS)

Hansson, J.; Keyse, S.M.; Lindahl, T.; Wood, R.D.

1991-01-01

Whole cell extracts from human lymphoid cell lines can perform in vitro DNA repair synthesis in plasmids damaged by agents including UV or cis-diamminedichloroplatinum(II) (cis-DDP). Extracts from xeroderma pigmentosum (XP) cells are defective in repair synthesis. We have now studied in vitro DNA repair synthesis using extracts from lymphoblastoid cell lines representing four human hereditary syndromes with increased sensitivity to DNA-damaging agents. Extracts of cell lines from individuals with the sunlight-sensitive disorders dysplastic nevus syndrome or Cockayne's syndrome (complementation groups A and B) showed normal DNA repair synthesis in plasmids with UV photoproducts. This is consistent with in vivo measurements of the overall DNA repair capacity in such cell lines. A number of extracts were prepared from two cell lines representing the variant form of XP (XP-V). Half of the extracts prepared showed normal levels of in vitro DNA repair synthesis in plasmids containing UV lesions, but the remainder of the extracts from the same cell lines showed deficient repair synthesis, suggesting the possibility of an unusually labile excision repair protein in XP-V. Fanconi's anemia (FA) cells show cellular hypersensitivity to cross-linking agents including cis-DDP. Extracts from cell lines belonging to two different complementation groups of FA showed normal DNA repair synthesis in plasmids containing cis-DDP or UV adducts. Thus, there does not appear to be an overall excision repair defect in FA, but the data do not exclude a defect in the repair of interstrand DNA cross-links

Generation of hematopoietic stem cells from human embryonic stem cells using a defined, stepwise, serum-free, and serum replacement-free monolayer culture method.

Science.gov (United States)

Kim, So-Jung; Jung, Ji-Won; Ha, Hye-Yeong; Koo, Soo Kyung; Kim, Eung-Gook; Kim, Jung-Hyun

2017-03-01

Embryonic stem cells (ESCs) can be expanded infinitely in vitro and have the potential to differentiate into hematopoietic stem cells (HSCs); thus, they are considered a useful source of cells for HSC production. Although several technical in vitro methods for engineering HSCs from pluripotent stem cells have been developed, clinical application of HSCs engineered from pluripotent stem cells is restricted because of the possibility of xenogeneic contamination resulting from the use of murine materials. Human ESCs (CHA-hES15) were cultured on growth factor-reduced Matrigel-coated dishes in the mTeSR1 serum-free medium. When the cells were 70% confluent, we initiated HSC differentiation by three methods involving (1) knockout serum replacement (KSR), cytokines, TGFb1, EPO, and FLT3L; (2) KSR, cytokines, and bFGF; or (3) cytokines and bFGF. Among the three differentiation methods, the minimal number of cytokines without KSR resulted in the greatest production of HSCs. The optimized method resulted in a higher proportion of CD34 + CD43 + hematopoietic progenitor cells (HPCs) and CD34 + CD45 + HPCs compared to the other methods. In addition, the HSCs showed the potential to differentiate into multiple lineages of hematopoietic cells in vitro . In this study, we optimized a two-step, serum-free, animal protein-free, KSR-free, feeder-free, chemically defined monolayer culture method for generation of HSCs and hematopoietic stem and progenitor cells (HSPCs) from human ESCs.

Arraying proteins by cell-free synthesis.

Science.gov (United States)

He, Mingyue; Wang, Ming-Wei

2007-10-01

Recent advances in life science have led to great motivation for the development of protein arrays to study functions of genome-encoded proteins. While traditional cell-based methods have been commonly used for generating protein arrays, they are usually a time-consuming process with a number of technical challenges. Cell-free protein synthesis offers an attractive system for making protein arrays, not only does it rapidly converts the genetic information into functional proteins without the need for DNA cloning, but also presents a flexible environment amenable to production of folded proteins or proteins with defined modifications. Recent advancements have made it possible to rapidly generate protein arrays from PCR DNA templates through parallel on-chip protein synthesis. This article reviews current cell-free protein array technologies and their proteomic applications.

Inhibitory Effects of Adlay Extract on Melanin Production and Cellular Oxygen Stress in B16F10 Melanoma Cells

Directory of Open Access Journals (Sweden)

Huey-Chun Huang

2014-09-01

Full Text Available The aim of this study was to determine the effects of adlay extract on melanin production and the antioxidant characteristics of the extract. The seeds were extracted by the supercritical fluid CO2 extraction (SFE method. The effect of adlay extract on melanin production was evaluated using mushroom tyrosinase activity assay, intracellular tyrosinase activity, antioxidant properties and melanin content. Those assays were performed spectrophotometrically. In addition, the expression of melanogenesis-related proteins was determined by western blotting. The results revealed that the adlay extract suppressed intracellular tyrosinase activity and decreased the amount of melanin in B16F10 cells. The adlay extract decreased the expression of microphthalmia-associated transcription factor (MITF, tyrosinase, tyrosinase related protein-1 (TRP-1 and tyrosinase related protein-2 (TRP-2. The extract also exhibited antioxidant characteristics such as free radical scavenging capacity and reducing power. It effectively decreased intracellular reactive oxygen species (ROS levels in B16F10 cells. We concluded that the adlay extract inhibits melanin production by down-regulation of MITF, tyrosinase, TRP-1 and TRP-2. The antioxidant properties of the extract may also contribute to the inhibition of melanogenesis. The adlay extract can therefore be applied as an inhibitor of melanogenesis and could also act as a natural antioxidant in skin care products.

Temperature and UV light affect the activity of marine cell-free enzymes

Directory of Open Access Journals (Sweden)

B. Thomson

2017-09-01

Full Text Available Microbial extracellular enzymatic activity (EEA is the rate-limiting step in the degradation of organic matter in the oceans. These extracellular enzymes exist in two forms: cell-bound, which are attached to the microbial cell wall, and cell-free, which are completely free of the cell. Contrary to previous understanding, cell-free extracellular enzymes make up a substantial proportion of the total marine EEA. Little is known about these abundant cell-free enzymes, including what factors control their activity once they are away from their sites (cells. Experiments were run to assess how cell-free enzymes (excluding microbes respond to ultraviolet radiation (UVR and temperature manipulations, previously suggested as potential control factors for these enzymes. The experiments were done with New Zealand coastal waters and the enzymes studied were alkaline phosphatase (APase, β-glucosidase, (BGase, and leucine aminopeptidase (LAPase. Environmentally relevant UVR (i.e. in situ UVR levels measured at our site reduced cell-free enzyme activities by up to 87 % when compared to controls, likely a consequence of photodegradation. This effect of UVR on cell-free enzymes differed depending on the UVR fraction. Ambient levels of UV radiation were shown to reduce the activity of cell-free enzymes for the first time. Elevated temperatures (15 °C increased the activity of cell-free enzymes by up to 53 % when compared to controls (10 °C, likely by enhancing the catalytic activity of the enzymes. Our results suggest the importance of both UVR and temperature as control mechanisms for cell-free enzymes. Given the projected warming ocean environment and the variable UVR light regime, it is possible that there could be major changes in the cell-free EEA and in the enzymes contribution to organic matter remineralization in the future.

Evaluation of the damage of cell wall and cell membrane for various extracellular polymeric substance extractions of activated sludge.

Science.gov (United States)

Guo, Xuesong; Liu, Junxin; Xiao, Benyi

2014-10-20

Extracellular polymeric substances (EPS) are susceptible to contamination by intracellular substances released during the extraction of EPS owing to the damage caused to microbial cell structures. The damage to cell walls and cell membranes in nine EPS extraction processes of activated sludge was evaluated in this study. The extraction of EPS (including proteins, carbohydrates and DNA) was the highest using the NaOH extraction method and the lowest using formaldehyde extraction. All nine EPS extraction methods in this study resulted in cell wall and membrane damage. The damage to cell walls, evaluated by 2-keto-3-deoxyoctonate (KDO) and N-acetylglucosamine content changes in extracted EPS, was the most significant in the NaOH extraction process. Formaldehyde extraction showed a similar extent of damage to cell walls to those detected in the control method (centrifugation), while those in the formaldehyde-NaOH and cation exchange resin extractions were slightly higher than those detected in the control. N-acetylglucosamine was more suitable than KDO for the evaluation of cell wall damage in the EPS extraction of activated sludge. The damage to cell membranes was characterized by two fluorochromes (propidium iodide and FITC Annexin V) with flow cytometry (FCM) measurement. The highest proportion of membrane-damaged cells was detected in NaOH extraction (26.54% of total cells) while membrane-damaged cells comprised 8.19% of total cells in the control. Copyright © 2014 Elsevier B.V. All rights reserved.

Effects of Ginkgo biloba extract on free radical metabolism of liver in ...

African Journals Online (AJOL)

This study investigated the effect of Ginkgo biloba extract on Free Radical Metabolism of Liver in mice during endurance exercise. Forty-eight mice were divided into the quiet group and the exercised group. And the two groups were both grouped again, including the control group and the drug-treated group.

Neuroprotective activity of Leontice leontopetalum extract against H2O2-stimulated oxidative stress in PC12 cells

Directory of Open Access Journals (Sweden)

S. Sahranavard*

2017-11-01

Full Text Available Background and objectives: Neuronal toxicity can be induced by oxidative stress via free radicals production. In recent years, great interest has been expressed to the traditional and herbal medicines. The purpose of this study was to elucidate the neuroprotective activity of Leontice leontopetalum methanol extract against H2O2-stimulated oxidative stress in PC12 cells. Methods: The plantLeontice leontopetalum was selected based on the ethnobotanical approach, which is used traditionally for the treatment of diseases related to inflammation and pain in Turkmen Sahra, Iran. Cytotoxicity of different concentrations of the methanol extract against PC12 cells was evaluated by MTT assay. Then PC12 cells were exposed to H2O2 in the presence or absence of the extract. In the next step, the total protein concentration was measured via Bradford assay and cyclooxygenase inhibition was determined by a screening assay kit. Nitrite accumulated in culture medium of supernatant was measured by Griess reaction. Results: Our results indicated that the methanol extract of Leontice leontopetalum significantly inhibited cyclooxygenase activity in the presence of H2O2; however, it was not able to inhibit nitric oxide generation in the stimulated PC12 cells. Conclusion: The results suggested that Leontice leontopetalum may be useful in reducing risk of neurodegenerative related diseases such as Alzheimer Disease.

Antioxidative and in vitro antiproliferative activity of Arctium lappa root extracts

Science.gov (United States)

2011-01-01

Background Arctium lappa, known as burdock, is widely used in popular medicine for hypertension, gout, hepatitis and other inflammatory disorders. Pharmacological studies indicated that burdock roots have hepatoprotective, anti-inflammatory, free radical scavenging and antiproliferative activities. The aim of this study was to evaluate total phenolic content, radical scavenging activity by DPPH and in vitro antiproliferative activity of different A. lappa root extracts. Methods Hot and room temperature dichloromethanic, ethanolic and aqueous extracts; hydroethanolic and total aqueous extract of A. lappa roots were investigated regarding radical scavenging activity by DPPH, total phenolic content by Folin-Ciocalteau method and antiproliferative in vitro activity was evaluated in human cancer cell lines. The hydroethanolic extract analyzed by high-resolution electrospray ionization mass spectroscopy. Results Higher radical scavenging activity was found for the hydroethanolic extract. The higher phenolic contents were found for the dichloromethane, obtained both by Soxhlet and maceration extraction and hydroethanolic extracts. The HRESI-MS demonstrated the presence of arctigenin, quercetin, chlorogenic acid and caffeic acid compounds, which were identified by comparison with previous data. The dichloromethane extracts were the only extracts that exhibited activity against cancer cell lines, especially for K562, MCF-7 and 786-0 cell lines. Conclusions The hydroethanolic extracts exhibited the strongest free radical scavenging activity, while the highest phenolic content was observed in Soxhlet extraction. Moreover, the dichloromethanic extracts showed selective antiproliferative activity against K562, MCF-7 and 786-0 human cancer cell lines. PMID:21429215

Antioxidative and in vitro antiproliferative activity of Arctium lappa root extracts.

Science.gov (United States)

Predes, Fabricia S; Ruiz, Ana L T G; Carvalho, João E; Foglio, Mary A; Dolder, Heidi

2011-03-23

Arctium lappa, known as burdock, is widely used in popular medicine for hypertension, gout, hepatitis and other inflammatory disorders. Pharmacological studies indicated that burdock roots have hepatoprotective, anti-inflammatory, free radical scavenging and antiproliferative activities. The aim of this study was to evaluate total phenolic content, radical scavenging activity by DPPH and in vitro antiproliferative activity of different A. lappa root extracts. Hot and room temperature dichloromethanic, ethanolic and aqueous extracts; hydroethanolic and total aqueous extract of A. lappa roots were investigated regarding radical scavenging activity by DPPH, total phenolic content by Folin-Ciocalteau method and antiproliferative in vitro activity was evaluated in human cancer cell lines. The hydroethanolic extract analyzed by high-resolution electrospray ionization mass spectroscopy. Higher radical scavenging activity was found for the hydroethanolic extract. The higher phenolic contents were found for the dichloromethane, obtained both by Soxhlet and maceration extraction and hydroethanolic extracts. The HRESI-MS demonstrated the presence of arctigenin, quercetin, chlorogenic acid and caffeic acid compounds, which were identified by comparison with previous data. The dichloromethane extracts were the only extracts that exhibited activity against cancer cell lines, especially for K562, MCF-7 and 786-0 cell lines. The hydroethanolic extracts exhibited the strongest free radical scavenging activity, while the highest phenolic content was observed in Soxhlet extraction. Moreover, the dichloromethanic extracts showed selective antiproliferative activity against K562, MCF-7 and 786-0 human cancer cell lines. © 2011 Predes et al; licensee BioMed Central Ltd.

Antioxidative and in vitro antiproliferative activity of Arctium lappa root extracts

Directory of Open Access Journals (Sweden)

Carvalho João E

2011-03-01

Full Text Available Abstract Background Arctium lappa, known as burdock, is widely used in popular medicine for hypertension, gout, hepatitis and other inflammatory disorders. Pharmacological studies indicated that burdock roots have hepatoprotective, anti-inflammatory, free radical scavenging and antiproliferative activities. The aim of this study was to evaluate total phenolic content, radical scavenging activity by DPPH and in vitro antiproliferative activity of different A. lappa root extracts. Methods Hot and room temperature dichloromethanic, ethanolic and aqueous extracts; hydroethanolic and total aqueous extract of A. lappa roots were investigated regarding radical scavenging activity by DPPH, total phenolic content by Folin-Ciocalteau method and antiproliferative in vitro activity was evaluated in human cancer cell lines. The hydroethanolic extract analyzed by high-resolution electrospray ionization mass spectroscopy. Results Higher radical scavenging activity was found for the hydroethanolic extract. The higher phenolic contents were found for the dichloromethane, obtained both by Soxhlet and maceration extraction and hydroethanolic extracts. The HRESI-MS demonstrated the presence of arctigenin, quercetin, chlorogenic acid and caffeic acid compounds, which were identified by comparison with previous data. The dichloromethane extracts were the only extracts that exhibited activity against cancer cell lines, especially for K562, MCF-7 and 786-0 cell lines. Conclusions The hydroethanolic extracts exhibited the strongest free radical scavenging activity, while the highest phenolic content was observed in Soxhlet extraction. Moreover, the dichloromethanic extracts showed selective antiproliferative activity against K562, MCF-7 and 786-0 human cancer cell lines.

Curcuma longa extract as a histological dye for collagen fibres and red blood cells

Science.gov (United States)

Avwioro, O G; Onwuka, S K; Moody, J O; Agbedahunsi, J M; Oduola, T; Ekpo, O E; Oladele, A A

2007-01-01

Crude ethanolic extract and column chromatographic fractions of the Allepey cultivar of Curcuma longa Roxb, commonly called turmeric (tumeric) in commerce, were used as a stain for tissue sections. Staining was carried out under basic, acidic and neutral media conditions. Inorganic and organic dissolution solvents were used. The stain was used as a counterstain after alum and iron haematoxylins. C. longa stained collagen fibres, cytoplasm, red blood cells and muscle cells yellow. It also stained in a fashion similar to eosin, except for its intense yellow colour. Preliminary phytochemical evaluation of the active column fraction revealed that it contained flavonoids, free anthraquinone and deoxy sugar. A cheap, natural dye can thus be obtained from C. longa. PMID:17451535

Male Snakes Allocate Time and Energy according to Individual Energetic Status: Body Condition, Steroid Hormones, and Reproductive Behavior in Timber Rattlesnakes, Crotalus horridus.

Science.gov (United States)

Lind, Craig M; Beaupre, Steven J

2015-01-01

Life-history theory predicts that organisms will hedge current reproductive investment against potential costs in terms of survivorship and future fecundity. However, little is known regarding the endocrine mechanisms underlying bet-hedging strategies in free-ranging male vertebrates. We examined the relationships among individual energetic status, steroid hormones, mate search, and reproductive behavior in free-ranging male timber rattlesnakes. Snakes were monitored over four active seasons in order to test two hypotheses: (1) males adjust the amount of time and energy allocated toward reproduction according to the level of individual energy stores, and (2) observed condition-dependent reproductive allocation is associated with circulating concentrations of steroid hormones (testosterone and corticosterone) thought to regulate reproductive behaviors in vertebrates. A positive relationship between body condition and testosterone was observed in both the field and the laboratory. Male mate search effort was positively correlated with both body condition and testosterone. Body condition and testosterone concentrations were negatively related to time allocated toward foraging during the breeding season. A strong effect of year was observed in the analysis of testosterone and search effort, suggesting that multiple environmental factors impact hormone production and reproductive investment. Corticosterone was not related to any measured variable. Therefore, our results did not indicate a clear role of corticosterone in mediating observed relationships between energetic status and behavior. Observed relationships are consistent with the hypothesis that males allocate time and energy toward reproduction according to individual energetic status and that testosterone plays a role in mediating the trade-off between current reproductive investment and residual reproductive value.

Simultaneous micro-electromembrane extractions of anions and cations using multiple free liquid membranes and acceptor solutions

Czech Academy of Sciences Publication Activity Database

Kubáň, Pavel; Boček, Petr

2016-01-01

Roč. 908, FEB (2016), s. 113-120 ISSN 0003-2670 R&D Projects: GA ČR(CZ) GA13-05762S Institutional support: RVO:68081715 Keywords : free liquid membranes * micro-electromembrane extraction * simultaneous extractions Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.950, year: 2016

Simultaneous micro-electromembrane extractions of anions and cations using multiple free liquid membranes and acceptor solutions

Czech Academy of Sciences Publication Activity Database

Kubáň, Pavel; Boček, Petr

2016-01-01

Roč. 908, FEB (2016), s. 113-120 ISSN 0003-2670 R&D Projects: GA ČR(CZ) GA13-05762S Institutional support: RVO:68081715 Keywords : free liquid membranes * micro-electromembrane extraction * simultaneous extractions Subject RIV: CB - Analytical Chemistry , Separation Impact factor: 4.950, year: 2016

Influence of serum extraction from the culture medium and of sublethal X-ray irradiation upon microvilli and invaginations of the membrane of Ehrlich ascites tumor cells in monolayer culture

International Nuclear Information System (INIS)

Laudenbach, G.; Pfab, R.; Hess, F.; Schachtschabel, D.O.

1984-01-01

In order to find out modifications of microvilli and invaginations, the cellular surfaces of Ehrlich ascites tumor cells in monolayer culture (basal medium of Eagle + 10% fetal calf serum) were investigated with the aid of electron-microscopic cross-sections. The tumor cells had been cultured without serum 24 hours prior to investigation or irradiated with 2 Gy. Morphometric evaluation after cell culture in a serum-free medium showed a reduced number of microvilli and a diminution of sections of microvilli. As already described before, a reduction of cell proliferation, of the microtubule-microfilament system, and of the endocytosis activity occurs under these serum-free conditions. The number of invaginations (related to a constant membrane part) was reduced by nearly 50% after serum extraction. Similarly to serum extraction, sublethal X-ray irradiation reduced the sections of microvilli, whereas the number of microvilli increased slightly. Contrary to the effect of serum extraction, the irradiated cells showed twice as many invaginations as the non-irradiated control cells. These differences in the surface structures are interpreted as a result of modified growth stimulations (+- serum) and radiogenic reparation processes. (orig.) [de

Prenatal Cell-Free DNA Screening

Science.gov (United States)

... poses no physical risks for you or your baby. While prenatal cell-free DNA screening might cause anxiety, it might help you avoid the need for more invasive tests, treatment or monitoring during your pregnancy. Keep in mind, however, that ...

Portulaca oleracea extract can inhibit nodule formation of colon cancer stem cells by regulating gene expression of the Notch signal transduction pathway.

Science.gov (United States)

Jin, Heiying; Chen, Li; Wang, Shuiming; Chao, Deng

2017-07-01

To investigate whether Portulaca oleracea extract affects tumor formation in colon cancer stem cells and its chemotherapy sensitivity. In addition, to analyze associated genetic changes within the Notch signal transduction pathway. Serum-free cultures of colon cancer cells (HT-29) and HT-29 cancer stem cells were treated with the chemotherapeutic drug 5-fluorouracil to assess sensitivity. Injections of the stem cells were also given to BALB/c mice to confirm tumor growth and note its characteristics. In addition, the effect of different concentrations of P. oleracea extract was tested on the growth of HT-29 colon cancer cells and HT-29 cancer stem cells, as determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method. The effects of P. oleracea extract on the expression of β-catenin, Notch1, and Notch2 in the HT-29 cells were studied using reverse transcription polymerase chain reaction and Western blotting. The tumor volume of the HT29 cells was two times larger than that of HT29 cancer stem cells. Treatment with P. oleracea extract inhibited the proliferation of both HT-29 cancer cells and HT-29 cancer stem cells at doses from 0.07 to 2.25 µg/mL. Apoptosis of HT-29 cancer cells and HT-29 cancer stem cells was assessed by flow cytometry; it was enhanced by the addition of P. oleracea extract. Finally, treatment with P. oleracea extract significantly downregulated the expression of the Notch1 and β-catenin genes in both cell types. The results of this study show that P. oleracea extract inhibits the growth of colon cancer stem cells in a dose-dependent manner. Furthermore, it inhibits the expression of the Notch1 and β-catenin genes. Taken together, this suggests that it may elicit its effects through regulatory and target genes that mediate the Notch signal transduction pathway.

Cytoprotective Mechanisms Mediated by Polyphenols from Chilean Native Berries against Free Radical-Induced Damage on AGS Cells.

Science.gov (United States)

Ávila, Felipe; Theoduloz, Cristina; López-Alarcón, Camilo; Dorta, Eva; Schmeda-Hirschmann, Guillermo

2017-01-01

The prevalence of cytoprotective mechanisms induced by polyphenols such as activation of intracellular antioxidant responses (ICM) and direct free radical scavenging was investigated in native Chilean species of strawberries, raspberries, and currants. Human gastric epithelial cells were co- and preincubated with polyphenolic-enriched extracts (PEEs) from Chilean raspberries ( Rubus geoides ), strawberries ( Fragaria chiloensis ssp. chiloensis f . chiloensis ), and currants ( Ribes magellanicum ) and challenged with peroxyl and hydroxyl radicals. Cellular protection was determined in terms of cell viability, glyoxalase I and glutathione s-transferases activities, and carboxymethyl lysine (CML) and malondialdehyde levels. Our results indicate that cytoprotection induced by ICM was the prevalent mechanism for Rubus geoides and F. chiloensis . This agreed with increased levels of glyoxalase I and glutathione S-transferase activities in cells preincubated with PEEs. ORAC index indicated that F. chiloensis was the most efficient peroxyl radical scavenger. Moreover, ICM mediated by F. chiloensis was effective in protecting cells from CML accumulation in contrast to the protective effects induced by free radical scavenging. Our results indicate that although both polyphenol-mediated mechanisms can exert protective effects, ICM was the most prevalent in AGS cells. These results suggest a potential use of these native berries as functional food.

The reducibility of heLa cell viability by Sargassum polycystum extracts

Science.gov (United States)

Firdaus, M.; Setijawati, D.; Islam, I.; Nursyam, H.; Kartikaningsih, H.; Yufidasari, H. S.; Prihanto, A. A.; Nurdiani, R.; Jaziri, A. A.

2018-04-01

Cervical cancer is the second largest cause of death-related cancer in women. The efficacy of cancer drugs is still low. Bioactive of brown seaweed has been studied by in vitro and in vivo as anticancer. The aim of this study was to evaluate the cytotoxicity of Sargassum polycystum extracts on HeLa cell, to recognize bioactive on extract and estimate the interaction between the bioactive and target protein. S. polycystum was found from Talango Island waters and HeLa cell was obtained from Indonesian Science Institute. Sample was extracted by ethanol, ethyl acetate and hexane, concentrated and finally, extracts were assayed on HeLa cell. The viability of this cell was quantified on ELISA-Reader. The bioactive compounds of the extract were elucidated by GC-MS. The interaction between bioactive and target protein was evaluated by using in silico method. The result showed that the lowest viability of HeLa cell on n-hexane extracts treatment. The n-hexane extract of this seaweed contained benzenepropanoic acid. This compound reduced HeLa cell viability by reducing of thrombin concentration. In conclusion, the benzene propanoic acid of S. polycystum was the cytotoxic agent and it is potential agent for anti-cervical cancer.

A Study of the Effects of Gas Well Compressor Noise on Breeding Bird Populations of the Rattlesnake Canyon Habitat Management Area, San Juan County, New Mexico

Energy Technology Data Exchange (ETDEWEB)

LaGory, K.E.; Chang, Young-Soo; Chun, K.C.; Reeves, T.; Liebich, R.; Smith, K.

2001-06-04

This report, conducted from May through July 2000, addressed the potential effect of compressor noise on breeding birds in gas-production areas administered by the FFO, specifically in the Rattlesnake Canyon Habitat Management Area northeast of Farmington, New Mexico. The study was designed to quantify and characterize noise output from these compressors and to determine if compressor noise affected bird populations in adjacent habitat during the breeding season.

Delineating the Rattlesnake Springs, New Mexico Watershed Using Precision Gravity Techniques

Science.gov (United States)

Doser, D. I.; Boykov, N. D.; Baker, M. R.; Kaip, G. M.; Langford, R. P.

2009-12-01

Rattlesnake Springs serves as the sole domestic water source for Carlsbad Caverns National Park. The recent development of oil and gas leases and agricultural lands surrounding the springs has led to concern about contamination of the fracture controlled aquifer system. We have conducted a series of precision gravity surveys (station spacing 200 to 300 m in a 4 x 4 km area), combined with other geophysical studies and geologic mapping, to delineate possible fracture systems in the gypsum and carbonate bedrock that feed the spring system. Our combined results suggest several pathways for water to enter the springs. A series of WNW-ESE striking features are apparent in our gravity data that appear to align with relict spring valleys we have mapped to the west of the springs. A self potential survey indicates that water is entering the springs at a shallow level from the northwest direction. However, gravity data also indicate a north-south trending fracture system could be providing a pathway for water to enter from the south. This is consistent with drawdown tests conducted in the 1950’s and 1960’s on irrigation wells located to the south of the springs. The north-south fracture system appears related to a basin bounding fault system observed in the regional gravity data.

The correlation between cell-free DNA and tumour burden was estimated by PET/CT in patients with advanced NSCLC

DEFF Research Database (Denmark)

Nygaard, A D; Holdgaard, Paw; Spindler, K-L G

2014-01-01

Background:Cell-free DNA (cfDNA) circulating in the blood holds a possible prognostic value in malignant diseases. Under malignant conditions, the level of cfDNA increases but the biological mechanism remains to be fully understood. We aimed to examine the correlation between cfDNA and total tumour...... burden defined by positron emission tomography (PET) parameters.Methods:Patients with advanced non-small cell lung cancer (NSCLC) were enrolled into a prospective biomarker trial. Before treatment, plasma was extracted and the level of cfDNA was determined by qPCR. An (18)F-fluorodeoxyglucose ((18)F...... analysis. MTV>the median was associated with a significantly shorter OS (P=0.02). There was no significant difference in OS according to TLG (P=0.08).Conclusion:Cell-free DNA may not be a simple measure of tumour burden, but seems to reflect more complex mechanisms of tumour biology, making it attractive...

Label-free single-cell separation and imaging of cancer cells using an integrated microfluidic system

DEFF Research Database (Denmark)

Antfolk, Maria; Kim, Soo Hyeon; Koizumi, Saori

2017-01-01

, an integrated system is presented that efficiently eliminates this risk by integrating label-free separation with single cell arraying of the target cell population, enabling direct on-chip tumor cell identification and enumeration. Prostate cancer cells (DU145) spiked into a sample with whole blood...... a fully integrated system for rapid label-free separation and on-chip phenotypic characterization of circulating tumor cells from peripheral venous blood in clinical practice....

Quantum mechanics of a free particle on a plane with an extracted point

International Nuclear Information System (INIS)

Kowalski, K.; Podlaski, K.; Rembielinski, J.

2002-01-01

A detailed study of a quantum free particle on a pointed plane is presented in this paper. In particular, some questions posed in the very recent paper by M. A. Cirone et al, Phys. Rev. A 65, 022101 (2002) are clarified. Namely, the topological effects related to extracting a point from a plane are indicated. The proposed results are introduced concerning self-adjoint extensions of operators describing the free particle on a pointed plane as well as the role played by discrete symmetries in the analysis of such extensions

Phytochemical Characteristics, Free Radical Scavenging Activities, and Neuroprotection of Five Medicinal Plant Extracts

Directory of Open Access Journals (Sweden)

Chia Lin Chang

2012-01-01

Full Text Available The objective of this study was to determine phytochemical characteristics, chemiluminescence antioxidant capacities, and neuroprotective effects on PC12 cells for methanol extracts of Spatholobus suberectus, Uncaria rhynchophylla, Alpinia officinarum, Drynaria fortunei, and Crataegus pinnatifida. The C. pinnatifida extract (CPE afforded the greatest yield and total phenolic content. The S. suberectus extract (SSE yielded the greatest total flavonoid content. The U. rhynchophylla extract (URE produced the greatest total tannin content, and the A. officinarum extract (AOE produced the greatest total triterpenoid content. The D. fortunei extract, assayed using horseradish peroxidase-luminol-hydrogen peroxide (H2O2, and AOE using pyrogallol-luminol assay each exhibited better antioxidant activity than the L-ascorbic acid and 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid did. The CPE, SSE, and URE presented neurogrowth effects and neuroprotective activities on H2O2-induced PC12 cell death at 0.5–5.0 μg/mL. The CPE represents a promising medicinal plant source for the treatment of H2O2-induced neurodegenerative disease, because of its useful phytochemical characteristics.

Phytochemical Characteristics, Free Radical Scavenging Activities, and Neuroprotection of Five Medicinal Plant Extracts

Science.gov (United States)

Chang, Chia Lin; Lin, Che San; Lai, Guia Hung

2012-01-01

The objective of this study was to determine phytochemical characteristics, chemiluminescence antioxidant capacities, and neuroprotective effects on PC12 cells for methanol extracts of Spatholobus suberectus, Uncaria rhynchophylla, Alpinia officinarum, Drynaria fortunei, and Crataegus pinnatifida. The C. pinnatifida extract (CPE) afforded the greatest yield and total phenolic content. The S. suberectus extract (SSE) yielded the greatest total flavonoid content. The U. rhynchophylla extract (URE) produced the greatest total tannin content, and the A. officinarum extract (AOE) produced the greatest total triterpenoid content. The D. fortunei extract, assayed using horseradish peroxidase-luminol-hydrogen peroxide (H2O2), and AOE using pyrogallol-luminol assay each exhibited better antioxidant activity than the L-ascorbic acid and 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid did. The CPE, SSE, and URE presented neurogrowth effects and neuroprotective activities on H2O2-induced PC12 cell death at 0.5–5.0 μg/mL. The CPE represents a promising medicinal plant source for the treatment of H2O2-induced neurodegenerative disease, because of its useful phytochemical characteristics. PMID:21845204

Pathophysiological consequences of hemolysis. Role of cell-free hemoglobin

Directory of Open Access Journals (Sweden)

Tomasz Misztal

2011-09-01

Full Text Available Abundant hemolysis is associated with a number of inherent and acquired diseases including sickle-cell disease (SCD, polycythemia, paroxysmal nocturnal hemoglobinuria (PNH and drug-induced hemolytic anemia. Despite different etiopathology of hemolytic diseases, many concomitant symptoms are comparable and include e.g. hypertension, hemoglobinuria and hypercoagulation state. Studies in the last years have shown a growing list of mechanisms lying at the basis of those symptoms, in particular irreversible reaction between cell-free hemoglobin (Hb and nitric oxide (NO – endogenous vasorelaxant and anti-thrombotic agent. Saturation of protective physiological cell-free Hb-scavenging mechanisms results in accumulation of Hb in plasma and hemoglobinemia. Extensive hemoglobinemia subsequently leads to hemoglobinuria, which may cause kidney damage and development of Fanconi syndrome. A severe problem in patients with SCD and PNH is pulmonary and systemic hypertension. It may lead to circulation failure, including stroke, and it is related to abolition of NO bioavailability for vascular smooth muscle cells. Thrombotic events are the major cause of death in SCD and PNH. It ensues from lack of platelet inhibition evoked by Hb-mediated NO scavenging. A serious complication that affects patients with excessive hemolysis is erectile dysfunction. Also direct cytotoxic, prooxidant and proinflammatory effects of cell-free hemoglobin and heme compose the clinical picture of hemolytic diseases. The pathophysiological role of plasma Hb, mechanisms of its elimination, and direct and indirect (via NO scavenging deleterious effects of cell-free Hb are presented in detail in this review. Understanding the critical role of hemolysis and cell-free Hb is important in the perspective of treating patients with hemolytic diseases and to design new effective therapies in future.

Mango (Mangifera indica L.) peel extract fractions from different cultivars differentially affect lipid accumulation in 3T3-L1 adipocyte cells.

Science.gov (United States)

Taing, Meng-Wong; Pierson, Jean-Thomas; Shaw, Paul N; Dietzgen, Ralf G; Roberts-Thomson, Sarah J; Gidley, Michael J; Monteith, Gregory R

2013-02-26

Plant phytochemicals are increasingly recognised as sources of bioactive molecules which may have potential benefit in many health conditions. In mangoes, peel extracts from different cultivars exhibit varying effects on adipogenesis in the 3T3-L1 adipocyte cell line. In this study, the effects of preparative HPLC fractions of methanol peel extracts from Irwin, Nam Doc Mai and Kensington Pride mangoes were evaluated. Fraction 1 contained the most hydrophilic components while subsequent fractions contained increasingly more hydrophobic components. High content imaging was used to assess mango peel fraction effects on lipid accumulation, nuclei count and nuclear area in differentiating 3T3-L1 cells. For all three mango cultivars, the more hydrophilic peel fractions 1-3 inhibited lipid accumulation with greater potency than the more hydrophobic peel fractions 4. For all three cultivars, the more lipophilic fraction 4 had concentrations that enhanced lipid accumulation greater than fractions 1-3 as assessed by lipid droplet integrated intensity. The potency of this fraction 4 varied significantly between cultivars. Using mass spectrometry, five long chain free fatty acids were detected in fraction 4; these were not present in any other peel extract fractions. Total levels varied between cultivars, with Irwin fraction 4 containing the highest levels of these free fatty acids. Lipophilic components appear to be responsible for the lipid accumulation promoting effects of some mango extracts and are the likely cause of the diverse effects of peel extracts from different mango cultivars on lipid accumulation.

Epigenetic reprogramming of breast cancer cells with oocyte extracts

Directory of Open Access Journals (Sweden)

Kumari Rajendra

2011-01-01

Full Text Available Abstract Background Breast cancer is a disease characterised by both genetic and epigenetic alterations. Epigenetic silencing of tumour suppressor genes is an early event in breast carcinogenesis and reversion of gene silencing by epigenetic reprogramming can provide clues to the mechanisms responsible for tumour initiation and progression. In this study we apply the reprogramming capacity of oocytes to cancer cells in order to study breast oncogenesis. Results We show that breast cancer cells can be directly reprogrammed by amphibian oocyte extracts. The reprogramming effect, after six hours of treatment, in the absence of DNA replication, includes DNA demethylation and removal of repressive histone marks at the promoters of tumour suppressor genes; also, expression of the silenced genes is re-activated in response to treatment. This activity is specific to oocytes as it is not elicited by extracts from ovulated eggs, and is present at very limited levels in extracts from mouse embryonic stem cells. Epigenetic reprogramming in oocyte extracts results in reduction of cancer cell growth under anchorage independent conditions and a reduction in tumour growth in mouse xenografts. Conclusions This study presents a new method to investigate tumour reversion by epigenetic reprogramming. After testing extracts from different sources, we found that axolotl oocyte extracts possess superior reprogramming ability, which reverses epigenetic silencing of tumour suppressor genes and tumorigenicity of breast cancer cells in a mouse xenograft model. Therefore this system can be extremely valuable for dissecting the mechanisms involved in tumour suppressor gene silencing and identifying molecular activities capable of arresting tumour growth. These applications can ultimately shed light on the contribution of epigenetic alterations in breast cancer and advance the development of epigenetic therapies.

Cell-free synthetic biology for in vitro prototype engineering.

Science.gov (United States)

Moore, Simon J; MacDonald, James T; Freemont, Paul S

2017-06-15

Cell-free transcription-translation is an expanding field in synthetic biology as a rapid prototyping platform for blueprinting the design of synthetic biological devices. Exemplar efforts include translation of prototype designs into medical test kits for on-site identification of viruses (Zika and Ebola), while gene circuit cascades can be tested, debugged and re-designed within rapid turnover times. Coupled with mathematical modelling, this discipline lends itself towards the precision engineering of new synthetic life. The next stages of cell-free look set to unlock new microbial hosts that remain slow to engineer and unsuited to rapid iterative design cycles. It is hoped that the development of such systems will provide new tools to aid the transition from cell-free prototype designs to functioning synthetic genetic circuits and engineered natural product pathways in living cells. © 2017 The Author(s).

Inhibition of the DHT-induced PSA secretion by Verbascum xanthophoeniceum and Serenoa repens extracts in human LNCaP prostate epithelial cells.

Science.gov (United States)

Marcoccia, D; Georgiev, M I; Alipieva, K I; Lorenzetti, S

2014-08-08

Verbascum xanthophoeniceum is a mullein plant, typical of Balkan region and some parts of Turkey, traditionally used as phytotherapeutic agent due to its anti-inflammatory properties. It is rich in phenylethanoid and iridoid metabolites whose anti-inflammatory properties are under characterization. The role of Verbascum xanthophoeniceum crude methanolic extract and its isolated phenylethanoid glycoside verbascoside have been evaluated, in comparison to a saw palmetto extract, on a human in vitro model of androgen-regulated prostate epithelium, the LNCaP cell line. Cytotoxicity and DHT-induced free and total PSA secretion have been thoroughly studied. We have found that similar to saw palmetto, Verbascum xanthophoeniceum extract and its isolated phenylethanoid glycoside verbascoside have no cytotoxicity in human LNCaP prostate epithelial cells, whereas an inhibitory effect on the DHT-induced free and total PSA secretion, a recognized anti-androgen like activity, has been shown in case of both Verbascum xanthophoeniceum extract and pure verbascoside. Furthermore, in the absence of the endogenous androgen DHT, an androgen-like activity in Verbascum xanthophoeniceum is detectable as it is for saw palmetto, suggesting that a mixed androgen-antiandrogen activity is present. For the first time, Serenoa repens and Verbascum xanthophoeniceum extracts have shown an absence of cytotoxicity and an inhibitory effect on DHT-induced PSA secretion in an in vitro model of human prostate epithelium, whereas the phenylethanoid glycoside verbascoside appeared to explain only part of the Verbascum xanthophoeniceum inhibitory activity on PSA secretion. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Inverted polymer solar cells with Nafion® as the hole extraction layer: efficiency and lifetime studies

Science.gov (United States)

Manceau, Matthieu; Berson, Solenn

2014-01-01

The use of Nafion® as the hole extraction layer in polymer solar cells is demonstrated in this work. Inverted devices were built on plastic foil with the following architecture: PET/ITO/ZnO/P3HT:PCBM/Nafion®/Ag. The Nafion® was processed from a surfactant-free solution in alcoholic solvents on top of the active layer. Optimization of film thickness and annealing yielded fully functional devices with power conversion efficiency similar to others referenced, along with good operational stability.

Crosslinked Remote-Doped Hole-Extracting Contacts Enhance Stability under Accelerated Lifetime Testing in Perovskite Solar Cells.

Science.gov (United States)

Xu, Jixian; Voznyy, Oleksandr; Comin, Riccardo; Gong, Xiwen; Walters, Grant; Liu, Min; Kanjanaboos, Pongsakorn; Lan, Xinzheng; Sargent, Edward H

2016-04-13

A crosslinked hole-extracting electrical contact is reported, which simultaneously improves the stability and lowers the hysteresis of perovskite solar cells. Polymerizable monomers and crosslinking processes are developed to obviate in situ degradation of the under lying perovskite. The crosslinked material is band-aligned with perovskite. The required free carrier density is induced by a high-work-function metal oxide layer atop the device, following a remote-doping strategy. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Nanoparticles of Selaginella doederleinii leaf extract inhibit human lung cancer cells A549

Science.gov (United States)

Syaefudin; Juniarti, A.; Rosiyana, L.; Setyani, A.; Khodijah, S.

2016-01-01

The aim of the present study is to evaluate cytotoxicity effect of nanoparticles of Selaginella doederleinii (S. doederleinii) leaves extract. S. doederleinii was extracted by maceration method using 70%(v/v) ethanol as solvent. Phytochemical content was analyzed qualitatively by using Harborne and Thin Layer Chromatography (TLC) methods. Nanoparticle extract was prepared by ionic gelation using chitosan as encapsulant agent. Anticancer activity was performed by using 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. The results showed that S. doederleinii contains of flavonoids. Nanoparticle of S. doederleinii leaves extract greatly inhibited A549 cells growth (cancer cells), with IC50 of 3% or 1020 μg/ml. These nanoparticles extract also inhibited the growth of Chang cells (normal cells), with IC50 of 4% or 1442 μg/ml. The effective concentration of nanoparticles extract which inhibits cancer cells without harming the normal cells is 0.5% or 167 μg/ml. Further studies are needed to obtain the concentration of nanoparticles extract which can selectively suppress cancer cells.

Cell-free DNA: A Neglected Source for Antibiotic Resistance Genes Spreading from WWTPs.

Science.gov (United States)

Zhang, Yan; Li, Aolin; Dai, Tianjiao; Li, Feifei; Xie, Hui; Chen, Lujun; Wen, Donghui

2018-01-02

Cell-associated ARGs in wastewater treatment plants (WWTPs) has been concerned, however, cell-free ARGs in WWTPs was rarely studied. In this study, the abundances of four representative ARGs, sulII, tetC, bla PSE-1 , and ermB, in a large municipal WWTP were investigated in both cell-associated and cell-free fractions. Cell-associated ARGs was the dominant ARGs fraction in the raw wastewater. After biological treatment, sludge settling, membrane filtration, and disinfection, cell-associated ARGs were substantially reduced, though the ratios of ARG/16S rRNA gene were increased with disinfection. Cell-free ARGs persisted in the WWTP with a removal of 0.36 log to 2.68 logs, which was much lower than the removal of cell-associated ARGs (3.21 logs to 4.14 logs). Therefore, the abundance ratio of cell-free ARGs to cell-associated ARGs increased from 0.04-1.59% to 2.00-1895.08% along the treatment processes. After 25-day-storage, cell-free ARGs in both biological effluent and disinfection effluent increased by 0.14 log to 1.99 logs and 0.12 log to 1.77 logs respectively, reflecting the persistence and low decay rate of cell-free ARGs in the discharge water. Therefore, cell-free ARGs might be a kind of important but previously neglected pollutant from WWTPs, which added potential risks to the effluent receiving environments.

Vector-free and transgene-free human iPS cells differentiate into functional neurons and enhance functional recovery after ischemic stroke in mice.

Directory of Open Access Journals (Sweden)

Osama Mohamad

Full Text Available Stroke is a leading cause of human death and disability in the adult population in the United States and around the world. While stroke treatment is limited, stem cell transplantation has emerged as a promising regenerative therapy to replace or repair damaged tissues and enhance functional recovery after stroke. Recently, the creation of induced pluripotent stem (iPS cells through reprogramming of somatic cells has revolutionized cell therapy by providing an unlimited source of autologous cells for transplantation. In addition, the creation of vector-free and transgene-free human iPS (hiPS cells provides a new generation of stem cells with a reduced risk of tumor formation that was associated with the random integration of viral vectors seen with previous techniques. However, the potential use of these cells in the treatment of ischemic stroke has not been explored. In the present investigation, we examined the neuronal differentiation of vector-free and transgene-free hiPS cells and the transplantation of hiPS cell-derived neural progenitor cells (hiPS-NPCs in an ischemic stroke model in mice. Vector-free hiPS cells were maintained in feeder-free and serum-free conditions and differentiated into functional neurons in vitro using a newly developed differentiation protocol. Twenty eight days after transplantation in stroke mice, hiPS-NPCs showed mature neuronal markers in vivo. No tumor formation was seen up to 12 months after transplantation. Transplantation of hiPS-NPCs restored neurovascular coupling, increased trophic support and promoted behavioral recovery after stroke. These data suggest that using vector-free and transgene-free hiPS cells in stem cell therapy are safe and efficacious in enhancing recovery after focal ischemic stroke in mice.

Culture in embryonic kidney serum and xeno-free media as renal cell carcinoma and renal cell carcinoma cancer stem cells research model.

Science.gov (United States)

Krawczyk, Krzysztof M; Matak, Damian; Szymanski, Lukasz; Szczylik, Cezary; Porta, Camillo; Czarnecka, Anna M

2018-04-01

The use of fetal bovine serum hinders obtaining reproducible experimental results and should also be removed in hormone and growth factor studies. In particular hormones found in FBS act globally on cancer cell physiology and influence transcriptome and metabolome. The aim of our study was to develop a renal carcinoma serum free culture model optimized for (embryonal) renal cells in order to select the best study model for downstream auto-, para- or endocrine research. Secondary aim was to verify renal carcinoma stem cell culture for this application. In the study, we have cultured renal cell carcinoma primary tumour cell line (786-0) as well as human kidney cancer stem cells in standard 2D monolayer cultures in Roswell Park Memorial Institute Medium or Dulbecco's Modified Eagle's Medium and Complete Human Kidney Cancer Stem Cell Medium, respectively. Serum-free, animal-component free Human Embryonic Kidney 293 media were tested. Our results revealed that xeno-free embryonal renal cells optimized culture media provide a useful tool in RCC cancer biology research and at the same time enable effective growth of RCC. We propose bio-mimic RCC cell culture model with specific serum-free and xeno-free medium that promote RCC cell viability.

Cell-Free Expression, Purification, and Characterization of the Functional β2-Adrenergic Receptor for Multianalyte Detection of β-Agonists.

Science.gov (United States)

Wang, Jian; Liu, Yuan; Zhang, Junhua; Han, Zhengzheng; Wang, Wei; Liu, Yang; Wei, Dong; Huang, Wei

2017-11-01

Large-scale expression of β 2 -adrenergic receptor (β 2 -AR) in functional form is necessary for establishment of receptor assays for detecting illegally abused β-adrenergic agonists (β-agonists). Cell-based heterologous expression systems have manycritical difficulties in synthesizing this membrane protein, such as low protein yields and aberrant folding. To overcome these challenges, the main objective of the present work was to synthesize large amounts of functional β 2 -AR in a cell-free system based on Escherichia coli extracts. A codon-optimized porcine β 2 -AR gene (codon adaptation index: 0.96) suitable for high expression in E. coli was synthesized and transcribed to the cell-free system, which contributed to increase the expression up to 1.1 mg/ml. After purification using Ni-affinity chromatography, the bioactivity of the purified receptor was measured by novel enzyme-linked receptor assays. It was determined that the relative affinities of the purified β 2 -AR for β-agonists in descending order were as follows: clenbuterol > salbutamol > ractopamine. Moreover, their IC 50 values were 45.99, 60.38, and 78.02 µg/liter, respectively. Although activity of the cell-free system was slightly lower than activity of systems based on insect and mammalian cells, this system should allow production of β 2 -AR in bulk amounts sufficient for the development of multianalyte screening methods for detecting β-agonist residues.

Cytoprotective Mechanisms Mediated by Polyphenols from Chilean Native Berries against Free Radical-Induced Damage on AGS Cells

Directory of Open Access Journals (Sweden)

Felipe Ávila

2017-01-01

Full Text Available The prevalence of cytoprotective mechanisms induced by polyphenols such as activation of intracellular antioxidant responses (ICM and direct free radical scavenging was investigated in native Chilean species of strawberries, raspberries, and currants. Human gastric epithelial cells were co- and preincubated with polyphenolic-enriched extracts (PEEs from Chilean raspberries (Rubus geoides, strawberries (Fragaria chiloensis ssp. chiloensis f. chiloensis, and currants (Ribes magellanicum and challenged with peroxyl and hydroxyl radicals. Cellular protection was determined in terms of cell viability, glyoxalase I and glutathione s-transferases activities, and carboxymethyl lysine (CML and malondialdehyde levels. Our results indicate that cytoprotection induced by ICM was the prevalent mechanism for Rubus geoides and F. chiloensis. This agreed with increased levels of glyoxalase I and glutathione S-transferase activities in cells preincubated with PEEs. ORAC index indicated that F. chiloensis was the most efficient peroxyl radical scavenger. Moreover, ICM mediated by F. chiloensis was effective in protecting cells from CML accumulation in contrast to the protective effects induced by free radical scavenging. Our results indicate that although both polyphenol-mediated mechanisms can exert protective effects, ICM was the most prevalent in AGS cells. These results suggest a potential use of these native berries as functional food.

Extraction of Active Enzymes from "Hard-to-Break-Cells"

DEFF Research Database (Denmark)

Ottaviani, Alessio; Tesauro, Cinzia; Fjelstrup, S

We present the utilization of a rolling circle amplification (RCA) based assay to investigate the extraction efficiency of active enzymes from a class of “hard-to-break” cells, yeast Saccaramyces cerevisiae. Current analyses of microorganisms, such as pathogenic bacteria, parasites or particular...... life stages of microorganisms (e.g. spores from bacteria or fungi) is hampered by the lack of efficient lysis protocols that preserve the activity and integrity of the cellular content. Presented herein is a flexible scheme to screen lysis protocols for active enzyme extraction. We also report a gentle...... yet effective approach for extraction of active enzymes by entrapping cells in microdroplets. Combined effort of optimized extraction protocols and effective analytical approaches is expected to generate impact in future disease diagnosis and environmental safety....

Antiproliferative study of B. javanica extracts against head and neck cancer cells

International Nuclear Information System (INIS)

Mohd Noor Hidayat Adenan; Zainah Adam; Shafii Khamis; Fazliana Mohd Saaya

2014-01-01

Brucea javanica or locally known as Meladapahit, are being used in Malaysia as traditional medicine mainly for the treatment of diabetes mellitus and hypertension. In order to study the potential use of this plant for cancer treatment, we have prepared crude extracts of the leaves and fruits, and assessed them for antiproliferative activities against head and neck cancer cell line which is HTB-43. The dried and ground leaves and fruits of the plant were successively extracted using hexane, chloroform, methanol and water, respectively. Inhibition of growth of the cultured cancer cells line was measured using a standard Micro culture Tetrazolium Technique (MTT) assay. The crude extracts were also subjected to toxicity test using brine shrimp lethality assay. Most of the tested crude extracts exhibited significant antiproliferative activities against the HTB-43 cell with IC 50 ranging from 8.46 μg/ml to 47.25 μg/ml. The chloroform extract from the leaves gave the highest antiproliferative activity (IC 50 , 8.46 μg/ml). Hexane extract from the fruits, aqueous and hexane extracts from B. javanica leaves showed low antiproliferative activities to the HTB-43 cell line with an IC 50 values >100 μg/ml. The chloroform extracts from fruits and leaves and methanol extract from fruits induced toxicity against brine shrimps with LC 50 values of 118.7 μg/ml, 512.44 μg/ml and 75.27 μg/ml respectively. It indicated that bioactive components presence in the crude extracts for its pharmacologic effects against head and neck cancer cells. Methanolic extract of Brucea javanica fruit was selected as the most effective extract to inhibit the growth of head and neck cancer cells (HTB-43) by the two different assays used. (author)

Radio protective effect of black mulberry extract on radiation-induced damage in bone marrow cells and liver in the rat

Science.gov (United States)

Ghasemnezhad Targhi, Reza; Homayoun, Mansour; Mansouri, Somaieh; Soukhtanloo, Mohammad; Soleymanifard, Shokouhozaman; Seghatoleslam, Masoumeh

2017-01-01

Ionizing radiation by producing free radicals induces tissue oxidative stress and has clastogenic and cytotoxic effects. The radio protective effect of black mulberry extract (BME) has been investigated on liver tissue and bone marrow cells in the rat. Intraperitoneal (ip) administration of 200 mg/kg BME three days before and three days after 3 Gy and 6 Gy gamma irradiation significantly reduced the frequencies of micro nucleated polychromatic erythrocytes (MnPCEs) and micro nucleated norm chromatic erythrocyte (MnNCEs) and increased PCE/PCE+NCE ratio in rat bone marrow compared to the non-treated irradiated groups. Moreover, this concentration of BME extract decreased the level of malondialdehyde (MDA) and superoxide dismutase (SOD), as well as enhanced the total thiol content and catalase activity in rat's liver compared to the non-treated irradiated groups. It seems that BME extract with antioxidant activity reduced the genotoxicity and cytotoxicity induced by gamma irradiation in bone marrow cells and liver in the rat.

Evaluation of the intestinal permeability of rosemary (Rosmarinus officinalis L. extract polyphenols and terpenoids in Caco-2 cell monolayers.

Directory of Open Access Journals (Sweden)

Almudena Pérez-Sánchez

Full Text Available Rosemary (Rosmarinus officinalis is grown throughout the world and is widely used as a medicinal herb and to season and preserve food. Rosemary polyphenols and terpenoids have attracted great interest due to their potential health benefits. However, complete information regarding their absorption and bioavailability in Caco-2 cell model is scarce. The permeation properties of the bioactive compounds (flavonoids, diterpenes, triterpenes and phenylpropanoids of a rosemary extract (RE, obtained by supercritical fluid extraction, was studied in Caco-2 cell monolayer model, both in a free form or liposomed. Compounds were identified and quantitated by liquid chromatography coupled to quadrupole time-of-flight with electrospray ionization mass spectrometry analysis (HPLC-ESI-QTOF-MS, and the apparent permeability values (Papp were determined, for the first time in the extract, for 24 compounds in both directions across cell monolayer. For some compounds, such as triterpenoids and some flavonoids, Papp values found were reported for the first time in Caco-2 cells.Our results indicate that most compounds are scarcely absorbed, and passive diffusion is suggested to be the primary mechanism of absorption. The use of liposomes to vehiculize the extract resulted in reduced permeability for most compounds. Finally, the biopharmaceutical classification (BCS of all the compounds was achieved according to their permeability and solubility data for bioequivalence purposes. BCS study reveal that most of the RE compounds could be classified as classes III and IV (low permeability; therefore, RE itself should also be classified into this category.

Evaluation of the intestinal permeability of rosemary (Rosmarinus officinalis L.) extract polyphenols and terpenoids in Caco-2 cell monolayers

Science.gov (United States)

Arráez-Román, David; González-Álvarez, Isabel; Ibáñez, Elena; Segura-Carretero, Antonio; Bermejo, Marival; Micol, Vicente

2017-01-01

Rosemary (Rosmarinus officinalis) is grown throughout the world and is widely used as a medicinal herb and to season and preserve food. Rosemary polyphenols and terpenoids have attracted great interest due to their potential health benefits. However, complete information regarding their absorption and bioavailability in Caco-2 cell model is scarce. The permeation properties of the bioactive compounds (flavonoids, diterpenes, triterpenes and phenylpropanoids) of a rosemary extract (RE), obtained by supercritical fluid extraction, was studied in Caco-2 cell monolayer model, both in a free form or liposomed. Compounds were identified and quantitated by liquid chromatography coupled to quadrupole time-of-flight with electrospray ionization mass spectrometry analysis (HPLC-ESI-QTOF-MS), and the apparent permeability values (Papp) were determined, for the first time in the extract, for 24 compounds in both directions across cell monolayer. For some compounds, such as triterpenoids and some flavonoids, Papp values found were reported for the first time in Caco-2 cells.Our results indicate that most compounds are scarcely absorbed, and passive diffusion is suggested to be the primary mechanism of absorption. The use of liposomes to vehiculize the extract resulted in reduced permeability for most compounds. Finally, the biopharmaceutical classification (BCS) of all the compounds was achieved according to their permeability and solubility data for bioequivalence purposes. BCS study reveal that most of the RE compounds could be classified as classes III and IV (low permeability); therefore, RE itself should also be classified into this category. PMID:28234919

Agnus castus extracts inhibit prolactin secretion of rat pituitary cells.

Science.gov (United States)

Sliutz, G; Speiser, P; Schultz, A M; Spona, J; Zeillinger, R

1993-05-01

In our studies on prolactin inhibition by plant extracts we focused on the effects of extracts of Vitex agnus castus and its preparations on rat pituitary cells under basal and stimulated conditions in primary cell culture. Both extracts from Vitex agnus castus as well as synthetic dopamine agonists (Lisuride) significantly inhibit basal as well as TRH-stimulated prolactin secretion of rat pituitary cells in vitro and as a consequence inhibition of prolactin secretion could be blocked by adding a dopamine receptor blocker. Therefore because of its dopaminergic effect Agnus castus could be considered as an efficient alternative phytotherapeutic drug in the treatment of slight hyperprolactinaemia.

Development of serum-free media for lepidopteran insect cell lines.

Science.gov (United States)

Agathos, Spiros N

2007-01-01

Lepidopteran insect cell culture technology has progressed to the point of becoming an essential part of one of the most successful eukaryotic expression systems and is increasingly used industrially on a large scale. Therefore, there is a constant need for convenient and low-cost culture media capable of supporting good insect cell growth and ensuring high yield of baculovirus as well as the strong expression of recombinant proteins. Vertebrate sera or invertebrate hemolymph were essential supplements in first-generation insect cell media. These supplements, however, are cumbersome and expensive for routine large-scale culture; thus, their use is now circumvented by substituting the essential growth factors present in these supplements with serum-free substances. Such non-serum supplements are typically of non-animal origin and include protein hydrolysates, lipid emulsions, and specialized substances (e.g., surfactants and shear damage protecting chemicals). These supplements need to complement the defined, synthetic basal medium to ensure that the fundamental nutritional needs of the cells are satisfied. Although there is a significant number of proprietary serum-free and low-protein or protein-free media on the market, the lack of information concerning their detailed composition is a drawback in their adoption for different applications, including their adaptation to the metabolic and kinetic analysis and monitoring of a given insect cell based bioprocess. Hence, there is wide appeal for formulating serum-free media based on a rational assessment of the metabolic requirements of the lepidopteran cells during both the growth and the production phases. Techniques such as statistical experimental design and genetic algorithms adapted to the cellular behavior and the bioreactor operation mode (batch, fed-batch, or perfusion) permit the formulation of versatile serum- and protein-free media. These techniques are illustrated with recent developments of serum-free

In vitro free radical scavenging activity of ethanolic extract of the whole plant of Evolvulus alsinoides (L.) L.

Science.gov (United States)

Gomathi, Duraisamy; Ravikumar, Ganesan; Kalaiselvi, Manokaran; Vidya, Balasubramaniam; Uma, Chandrasekar

2015-06-01

To identify the free radical scavenging activity of ethanolic extract of Evolvulus alsinoides. The free radical scavenging activity was evaluated by in vitro methods like reducing power assay, total antioxidant activity, 2,2-diphenyl-1-picrylhydrazyl (DPPH) reduction, superoxide radical scavenging activity, 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS(+)) scavenging activity, hydroxyl radical scavenging assay, and nitric oxide radical scavenging assay, which were studied by using ascorbic acid as standard. The extract showed significant activities in all antioxidant assays compared with the reference antioxidant ascorbic acid. The total antioxidant activity as well as the reducing power was also found to increase in a dose-dependent manner. Evolvulus alsinoides may act as a chemopreventive agent, providing antioxidant properties and offering effective protection from free radicals.

Free-zone electrophoresis of animal cells. 1: Experiments on cell-cell interactions

Science.gov (United States)

Todd, P. W.; Hjerten, S.

1985-01-01

The electrophoretically migrating zones wasa monitored. The absence of fluid flows in the direction of migration permits direct measurement of electrophoretic velocities of any material. Sedimentation is orthogonal to electrokinetic motion and the effects of particle-particle interaction on electrophoretic mobility is studied by free zone electrophoresis. Fixed erythrocytes at high concentrations, mixtures of fixed erythrocytes from different animal species, and mixtures of cultured human cells were studied in low ionic strength buffers. The electrophoretic velocity of fixed erythrocytes was not altered by increasing cell concentration or by the mixing of erythrocytes from different species. When zones containing cultured human glial cells and neuroblastoma cells are permitted to interact during electrophoresis, altered migration patterns occur. It is found that cell-cell interactions depends upon cell type.

Bergenin Content and Free Radical Scavenging Activity of Bergenia Extracts. .

Science.gov (United States)

Hendrychová, Helena; Martin, Jan; Tůmová, Lenka; Kočevar-Glavač, Nina

2015-07-01

Our research was focused on the evaluation of bergenin content and free radical scavenging activity of extracts prepared from three different species of Bergenia - B. crassifolia (L.) Fritsch., B. ciliata (Haw.) Sternb. and B. x ornata Stein. collected during different seasons. Using an HPLC method, the highest total amount of bergenin was revealed in the leaves of B. x ornata and B. crassifolia (4.9 - 5.1 mg x g(-1)). Free radical scavenging power was determined by two methods--FRAP and NADH. The best free radical scavengers were B. crassifolia (FRAP: 6.7 - 15.9 mg GAE. 100g(-1); NADH: 20.3 - 50.9%) and B. ornata (FRAP: 13.7 - 15.2 mg GAE. 100g(-1); NADH: 29.3 - 31.1%). The lowest content of bergenin and the weakest radical scavenger was B. ciliata (bergenin: 3.1 mg x g(-1); FRAP: 5.5 - 11.0 mg GAE.100g(-1); NADH: 23.2 - 25.6%). The presence of a large percentage of bergenin is responsible for the radical scavenging activity, as shown by the results from the FRAP and NADH assays. Significant, positive correlation was found between bergenin content and radical scavenging activity in both methods.

Dynamic Modeling of Cell-Free Biochemical Networks Using Effective Kinetic Models

Directory of Open Access Journals (Sweden)

Joseph A. Wayman

2015-03-01

Full Text Available Cell-free systems offer many advantages for the study, manipulation and modeling of metabolism compared to in vivo processes. Many of the challenges confronting genome-scale kinetic modeling can potentially be overcome in a cell-free system. For example, there is no complex transcriptional regulation to consider, transient metabolic measurements are easier to obtain, and we no longer have to consider cell growth. Thus, cell-free operation holds several significant advantages for model development, identification and validation. Theoretically, genome-scale cell-free kinetic models may be possible for industrially important organisms, such as E. coli, if a simple, tractable framework for integrating allosteric regulation with enzyme kinetics can be formulated. Toward this unmet need, we present an effective biochemical network modeling framework for building dynamic cell-free metabolic models. The key innovation of our approach is the integration of simple effective rules encoding complex allosteric regulation with traditional kinetic pathway modeling. We tested our approach by modeling the time evolution of several hypothetical cell-free metabolic networks. We found that simple effective rules, when integrated with traditional enzyme kinetic expressions, captured complex allosteric patterns such as ultrasensitivity or non-competitive inhibition in the absence of mechanistic information. Second, when integrated into network models, these rules captured classic regulatory patterns such as product-induced feedback inhibition. Lastly, we showed, at least for the network architectures considered here, that we could simultaneously estimate kinetic parameters and allosteric connectivity from synthetic data starting from an unbiased collection of possible allosteric structures using particle swarm optimization. However, when starting with an initial population that was heavily enriched with incorrect structures, our particle swarm approach could converge

A Vitex agnus-castus extract inhibits cell growth and induces apoptosis in prostate epithelial cell lines.

Science.gov (United States)

Weisskopf, M; Schaffner, W; Jundt, G; Sulser, T; Wyler, S; Tullberg-Reinert, H

2005-10-01

Extracts of Vitex agnus-castus fruits (VACF) are described to have beneficial effects on disorders related to hyperprolactinemia (cycle disorders, premenstrual syndrome). A VACF extract has recently been shown to exhibit antitumor activities in different human cancer cell lines. In the present study, we explored the antiproliferative effects of a VACF extract with a particular focus on apoptosis-inducing and potential cytotoxic effects. Three different human prostate epithelial cell lines (BPH-1, LNCaP, PC-3) representing different disease stages and androgen responsiveness were chosen. The action of VACF on cell viability was assessed using the WST-8-tetrazolium assay. Cell proliferation in cells receiving VACF alone or in combination with a pan-caspase inhibitor (Z-VAD-fmk) was quantified using a Crystal Violet assay. Flow cytometric cell cycle analysis and measurement of DNA fragmentation using an ELISA method were used for studying the induction of apoptosis. Lactate dehydrogenase (LDH) activity was determined as a marker of cytotoxicity. The extract inhibited proliferation of all three cell lines in a concentration-dependent manner with IC (50) values below 10 microg/mL after treatment for 48 h. Cell cycle analysis and DNA fragmentation assays suggest that part of the cells were undergoing apoptosis. The VACF-induced decrease in cell number was partially inhibited by Z-VAD-fmk, indicating a caspase-dependent apoptotic cell death. However, the concentration-dependent LDH activity of VACF treated cells indicated cytotoxic effects as well. These data suggest that VACF contains components that inhibit proliferation and induce apoptosis in human prostate epithelial cell lines. The extract may be useful for the prevention and/or treatment not only of benign prostatic hyperplasia but also of human prostate cancer.

Effect of Robola and Cabernet Sauvignon extracts on platelet activating factor enzymes activity on U937 cells.

Science.gov (United States)

Xanthopoulou, M N; Asimakopoulos, D; Antonopoulou, S; Demopoulos, C A; Fragopoulou, E

2014-12-15

A number of studies support the anti-atherogenic effect of wine compounds. The scope of this study was to examine the effect of a red (Cabernet Sauvignon-CS) and a white (Robola-R) wine, as well as resveratrol and quercetin, on the platelet activating factor (PAF) biosynthetic enzymes, acetyl-CoA:lyso-PAF acetyltransferase (lyso-PAF-AT) and DTT-insensitive CDP-choline 1-alkyl-2-acetyl-sn-glycerol cholinephosphotransferase (PAF-CPT), and its main catabolic enzyme (PAF acetylhydrolase; PAF-AH), on U937 cells, in cell free and in intact cell experiments. In cell free experiments, phenolic compounds and wine extracts inhibited PAF biosynthetic enzymes, however in higher concentrations than intact cell experiments. In the latter cases, polar lipids of both wines inhibited in the same order of magnitude the action of lyso-PAF-AT and of PAF-CPT. The water fractions possessed a dual action, in lower concentrations they activated both enzymes, while in higher concentrations only inhibited PAF-CPT. All fractions either did not affect or slightly activated PAF-AH activity. In conclusion, wine compounds may exert their anti-inflammatory activity by reducing PAF levels through modulation of the PAF metabolic enzymes. Copyright © 2014 Elsevier Ltd. All rights reserved.

Watch Out for the “Living Dead”: Cell-Free Enzymes and Their Fate

Directory of Open Access Journals (Sweden)

Federico Baltar

2018-01-01

Full Text Available Microbes are the engines driving biogeochemical cycles. Microbial extracellular enzymatic activities (EEAs are the “gatekeepers” of the carbon cycle. The total EEA is the sum of cell-bound (i.e., cell-attached, and dissolved (i.e., cell-free enzyme activities. Cell-free enzymes make up a substantial proportion (up to 100% of the total marine EEA. Although we are learning more about how microbial diversity and function (including total EEA will be affected by environmental changes, little is known about what factors control the importance of the abundant cell-free enzymes. Since cell-attached EEAs are linked to the cell, their fate will likely be linked to the factors controlling the cell’s fate. In contrast, cell-free enzymes belong to a kind of “living dead” realm because they are not attached to a living cell but still are able to perform their function away from the cell; and as such, the factors controlling their activity and fate might differ from those affecting cell-attached enzymes. This article aims to place cell-free EEA into the wider context of hydrolysis of organic matter, deal with recent studies assessing what controls the production, activity and lifetime of cell-free EEA, and what their fate might be in response to environmental stressors. This perspective article advocates the need to go “beyond the living things,” studying the response of cells/organisms to different stressors, but also to study cell-free enzymes, in order to fully constrain the future and evolution of marine biogeochemical cycles.

Watch Out for the “Living Dead”: Cell-Free Enzymes and Their Fate

Science.gov (United States)

Baltar, Federico

2018-01-01

Microbes are the engines driving biogeochemical cycles. Microbial extracellular enzymatic activities (EEAs) are the “gatekeepers” of the carbon cycle. The total EEA is the sum of cell-bound (i.e., cell-attached), and dissolved (i.e., cell-free) enzyme activities. Cell-free enzymes make up a substantial proportion (up to 100%) of the total marine EEA. Although we are learning more about how microbial diversity and function (including total EEA) will be affected by environmental changes, little is known about what factors control the importance of the abundant cell-free enzymes. Since cell-attached EEAs are linked to the cell, their fate will likely be linked to the factors controlling the cell’s fate. In contrast, cell-free enzymes belong to a kind of “living dead” realm because they are not attached to a living cell but still are able to perform their function away from the cell; and as such, the factors controlling their activity and fate might differ from those affecting cell-attached enzymes. This article aims to place cell-free EEA into the wider context of hydrolysis of organic matter, deal with recent studies assessing what controls the production, activity and lifetime of cell-free EEA, and what their fate might be in response to environmental stressors. This perspective article advocates the need to go “beyond the living things,” studying the response of cells/organisms to different stressors, but also to study cell-free enzymes, in order to fully constrain the future and evolution of marine biogeochemical cycles. PMID:29354095

Evaluation of cell cytotoxic effect on herbal extracts mixtures

International Nuclear Information System (INIS)

Kim, Yong Soo; Gwon, Hui Jeong; Choi, Bo Ram; Lim, Youn Mook; Nho, Young Chang

2009-01-01

Herbal extracts (HE) such as Houttuynia cordata Thunb., Eucommia ulimoides, Plantago asiatica var., Morus alba L., and Ulmus davidiana var., are known to suppress an atopic dermatitis like skin lesions. In this study, to evaluate the cell cytotoxicity effect on L929, HaCaT and HMC-1 cell by the HE, the herbs were extracted with distilled water (at 75 .deg. C) and then the HE mixtures were freeze-dried for 5 days and sterilized with γ-rays. The cytotoxicity was measured by Cell Counting Kit-8 (CCK-8) assay. The result showed that the HE mixtures did not significantly affect cell viability and had no toxicity on the cells. These findings indicate that the HE mixtures can be used as a potential therapeutic agent

Evaluation of cell cytotoxic effect on herbal extracts mixtures

Energy Technology Data Exchange (ETDEWEB)

Kim, Yong Soo; Gwon, Hui Jeong; Choi, Bo Ram; Lim, Youn Mook; Nho, Young Chang [Korea Atomic Energy Research Institute, Jeongeup (Korea, Republic of)

2009-12-15

Herbal extracts (HE) such as Houttuynia cordata Thunb., Eucommia ulimoides, Plantago asiatica var., Morus alba L., and Ulmus davidiana var., are known to suppress an atopic dermatitis like skin lesions. In this study, to evaluate the cell cytotoxicity effect on L929, HaCaT and HMC-1 cell by the HE, the herbs were extracted with distilled water (at 75 .deg. C) and then the HE mixtures were freeze-dried for 5 days and sterilized with {gamma}-rays. The cytotoxicity was measured by Cell Counting Kit-8 (CCK-8) assay. The result showed that the HE mixtures did not significantly affect cell viability and had no toxicity on the cells. These findings indicate that the HE mixtures can be used as a potential therapeutic agent.

Parameter-free extraction of EMCD from an energy-filtered diffraction datacube using multivariate curve resolution

International Nuclear Information System (INIS)

Muto, S.; Tatsumi, K.; Rusz, J.

2013-01-01

We present a parameter-free method of extraction of the electron magnetic circular dichroism spectra from energy-filtered diffraction patterns measured on a crystalline specimen. The method is based on a multivariate curve resolution technique. The main advantage of the proposed method is that it allows extraction of the magnetic signal regardless of the symmetry and orientation of the crystal, as long as there is a sufficiently strong magnetic component of the signal in the diffraction plane. This method essentially overcomes difficulties in extraction of the EMCD signal caused by complexity of dynamical diffraction effects. - Highlights: ► New method of extraction of EMCD signal using statistical methods (multivariate curve resolution). ► EMCD can be extracted quantitatively regardless of symmetry of crystal or its orientation. ► First principles simulation of EFDIF datacube, including dynamical diffraction effects

Catalytic reduction of 4-nitrophenol using gold nanoparticles biosynthesized by cell-free extracts of Aspergillus sp. WL-Au

Energy Technology Data Exchange (ETDEWEB)

Shen, Wenli; Qu, Yuanyuan, E-mail: qyy@dlut.edu.cn; Pei, Xiaofang; Li, Shuzhen; You, Shengnan; Wang, Jingwei; Zhang, Zhaojing; Zhou, Jiti

2017-01-05

Highlights: • A green process for AuNPs synthesis was achieved by fungus Aspergillus. • Uniform spherical AuNPs with well dispersity and stability were biosynthesized. • The biogenic AuNPs possessed remarkable catalytic activities for 4-NP reduction. - Abstract: A facile one-pot eco-friendly process for synthesis of gold nanoparticles (AuNPs) with high catalytic activity was achieved using cell-free extracts of Aspergillus sp. WL-Au as reducing, capping and stabilizing agents. The surface plasmon resonance band of UV–vis spectrum at 532 nm confirmed the presence of AuNPs. Transmission electron microscopy images showed that quite uniform spherical AuNPs were synthesized and the average size of nanoparticles increased from 4 nm to 29 nm with reaction time. X-ray diffraction analysis verified the formation of nano-crystalline gold particles. Fourier transform infrared spectra showed the presence of functional groups on the surface of biosynthesized AuNPs, such as O−H, N−H, C=O, C−H, C−OH and C−O−C groups, which increased the stability of AuNPs. The biogenic AuNPs could serve as a highly efficient catalyst for 4-nitrophenol reduction. The reaction rate constant was linearly correlated with the concentration of AuNPs, which increased from 0.59 min{sup −1} to 1.51 min{sup −1} with the amount of AuNPs increasing form 1.46 × 10{sup −6} to 17.47 × 10{sup −6} mmol. Moreover, the as-synthesized AuNPs exhibited a remarkable normalized catalytic activity (4.04 × 10{sup 5} min{sup −1} mol{sup −1}), which was much higher than that observed for AuNPs synthesized by other biological and conventional chemical methods.

Catalytic reduction of 4-nitrophenol using gold nanoparticles biosynthesized by cell-free extracts of Aspergillus sp. WL-Au

International Nuclear Information System (INIS)

Shen, Wenli; Qu, Yuanyuan; Pei, Xiaofang; Li, Shuzhen; You, Shengnan; Wang, Jingwei; Zhang, Zhaojing; Zhou, Jiti

2017-01-01

Highlights: • A green process for AuNPs synthesis was achieved by fungus Aspergillus. • Uniform spherical AuNPs with well dispersity and stability were biosynthesized. • The biogenic AuNPs possessed remarkable catalytic activities for 4-NP reduction. - Abstract: A facile one-pot eco-friendly process for synthesis of gold nanoparticles (AuNPs) with high catalytic activity was achieved using cell-free extracts of Aspergillus sp. WL-Au as reducing, capping and stabilizing agents. The surface plasmon resonance band of UV–vis spectrum at 532 nm confirmed the presence of AuNPs. Transmission electron microscopy images showed that quite uniform spherical AuNPs were synthesized and the average size of nanoparticles increased from 4 nm to 29 nm with reaction time. X-ray diffraction analysis verified the formation of nano-crystalline gold particles. Fourier transform infrared spectra showed the presence of functional groups on the surface of biosynthesized AuNPs, such as O−H, N−H, C=O, C−H, C−OH and C−O−C groups, which increased the stability of AuNPs. The biogenic AuNPs could serve as a highly efficient catalyst for 4-nitrophenol reduction. The reaction rate constant was linearly correlated with the concentration of AuNPs, which increased from 0.59 min −1 to 1.51 min −1 with the amount of AuNPs increasing form 1.46 × 10 −6 to 17.47 × 10 −6 mmol. Moreover, the as-synthesized AuNPs exhibited a remarkable normalized catalytic activity (4.04 × 10 5 min −1 mol −1 ), which was much higher than that observed for AuNPs synthesized by other biological and conventional chemical methods.

A novel validation algorithm allows for automated cell tracking and the extraction of biologically meaningful parameters.

Directory of Open Access Journals (Sweden)

Daniel H Rapoport

Full Text Available Automated microscopy is currently the only method to non-invasively and label-free observe complex multi-cellular processes, such as cell migration, cell cycle, and cell differentiation. Extracting biological information from a time-series of micrographs requires each cell to be recognized and followed through sequential microscopic snapshots. Although recent attempts to automatize this process resulted in ever improving cell detection rates, manual identification of identical cells is still the most reliable technique. However, its tedious and subjective nature prevented tracking from becoming a standardized tool for the investigation of cell cultures. Here, we present a novel method to accomplish automated cell tracking with a reliability comparable to manual tracking. Previously, automated cell tracking could not rival the reliability of manual tracking because, in contrast to the human way of solving this task, none of the algorithms had an independent quality control mechanism; they missed validation. Thus, instead of trying to improve the cell detection or tracking rates, we proceeded from the idea to automatically inspect the tracking results and accept only those of high trustworthiness, while rejecting all other results. This validation algorithm works independently of the quality of cell detection and tracking through a systematic search for tracking errors. It is based only on very general assumptions about the spatiotemporal contiguity of cell paths. While traditional tracking often aims to yield genealogic information about single cells, the natural outcome of a validated cell tracking algorithm turns out to be a set of complete, but often unconnected cell paths, i.e. records of cells from mitosis to mitosis. This is a consequence of the fact that the validation algorithm takes complete paths as the unit of rejection/acceptance. The resulting set of complete paths can be used to automatically extract important biological parameters

Insights to the effects of free cells on community structure of attached cells and chalcopyrite bioleaching during different stages.

Science.gov (United States)

Feng, Shoushuai; Yang, Hailin; Wang, Wu

2016-01-01

The effects of free cells on community structure of attached cells and chalcopyrite bioleaching by Acidithiobacillus sp. during different stages were investigated. The attached cells of Acidithiobacillus thiooxidans owned the community advantage from 14thd to the end of bioprocess in the normal system. The community structure of attached cells was greatly influenced in the free cells-deficient systems. Compared to A. thiooxidans, the attached cells community of Acidithiobacillus ferrooxidans had a higher dependence on its free cells. Meanwhile, the analysis of key biochemical parameters revealed that the effects of free cells on chalcopyrite bioleaching in different stages were diverse, ranging from 32.8% to 64.3%. The bioleaching contribution of free cells of A. ferrooxidans in the stationary stage (8-14thd) was higher than those of A. thiooxidans, while the situation was gradually reversed in the jarosite passivation inhibited stage (26-40thd). These results may be useful in guiding chalcopyrite bioleaching. Copyright © 2015 Elsevier Ltd. All rights reserved.

Anticancer Effects of Salvia miltiorrhiza Alcohol Extract on Oral Squamous Carcinoma Cells

Directory of Open Access Journals (Sweden)

Wen-Hung Wang

2017-01-01

Full Text Available Researchers have reported significant effects from Danshen (Salvia miltiorrhiza in terms of inhibiting tumor cell proliferation and promoting apoptosis in breast cancer, hepatocellular carcinomas, promyelocytic leukemia, and clear cell ovary carcinomas. Here we report our data indicating that Danshen extracts, especially alcohol extract, significantly inhibited the proliferation of the human oral squamous carcinoma (OSCC cell lines HSC-3 and OC-2. We also observed that Danshen alcohol extract activated the caspase-3 apoptosis executor by impeding members of the inhibitor of apoptosis (IAP family, but not by regulating the Bcl-2-triggered mitochondrial pathway in OSCC cells. Our data also indicate that the extract exerted promising effects in vivo, with HSC-3 tumor xenograft growth being suppressed by 40% and 69% following treatment with Danshen alcohol extract at 50 and 100 mg/kg, respectively, for 34 days. Combined, our results indicate appreciable anticancer activity and significant potential for Danshen alcohol extract as a natural antioxidant and herbal human oral cancer chemopreventive drug.

Plant Phenolics Extraction from Flos Chrysanthemi: Response Surface Methodology Based Optimization and the Correlation Between Extracts and Free Radical Scavenging Activity.

Science.gov (United States)

Wu, Yanfang; Wang, Xinsheng; Xue, Jintao; Fan, Enguo

2017-11-01

Huaiju is one of the most famous and widely used Flos Chrysanthemi (FC) for medicinal purposes in China. Although various investigations aimed at phenolics extraction from other FC have been reported, a thorough optimization of the phenolics extraction conditions from Huaiju has not been achieved. This work applied the widely used response surface methodology (RSM) to investigate the effects of 3 independent variables including ethanol concentration (%), extraction time (min), and solvent-to-material ratio (mL/g) on the ultrasound-assisted extraction (UAE) of phenolics from FC. The data suggested the optimal UAE condition was an ethanol concentration of 75.3% and extraction time of 43.5 min, whereas the ratio of solvent to material has no significant effect. When the free radical scavenging ability was used as an indicator for a successful extraction, a similar optimal extraction was achieved with an ethanol concentration of 72.8%, extraction time of 44.3 min, and the ratio of solvent to material was 29.5 mL/g. Furthermore, a moderate correlation between the antioxidant activity of TP extract and the content of extracted phenolic compounds was observed. Moreover, a well consistent of the experimental values under optimal conditions with those predicted values suggests RSM successfully optimized the UAE conditions for phenolics extraction from FC. The work of the research investigated the plant phenolics in Flos Chrysanthemi and antioxidant capacities. These results of this study can support the development of antioxidant additive and relative food. © 2017 The Authors. Journal of Food Science published by Wiley Periodicals, Inc. on behalf of Institute of Food Technologists.

An optimized rapid bisulfite conversion method with high recovery of cell-free DNA.

Science.gov (United States)

Yi, Shaohua; Long, Fei; Cheng, Juanbo; Huang, Daixin

2017-12-19

Methylation analysis of cell-free DNA is a encouraging tool for tumor diagnosis, monitoring and prognosis. Sensitivity of methylation analysis is a very important matter due to the tiny amounts of cell-free DNA available in plasma. Most current methods of DNA methylation analysis are based on the difference of bisulfite-mediated deamination of cytosine between cytosine and 5-methylcytosine. However, the recovery of bisulfite-converted DNA based on current methods is very poor for the methylation analysis of cell-free DNA. We optimized a rapid method for the crucial steps of bisulfite conversion with high recovery of cell-free DNA. A rapid deamination step and alkaline desulfonation was combined with the purification of DNA on a silica column. The conversion efficiency and recovery of bisulfite-treated DNA was investigated by the droplet digital PCR. The optimization of the reaction results in complete cytosine conversion in 30 min at 70 °C and about 65% of recovery of bisulfite-treated cell-free DNA, which is higher than current methods. The method allows high recovery from low levels of bisulfite-treated cell-free DNA, enhancing the analysis sensitivity of methylation detection from cell-free DNA.

Cytotoxic activity of water extracts of Trichilia hirta leaves on human tumor cells

International Nuclear Information System (INIS)

Hernandez Sosa, Edgar; Mora Gonzalez, Nestor; Morris Quevedo, Humberto J

2013-01-01

Trichilia hirta L. (Meliaceae) is traditionally used by patients suffering from cancer as an antitumoral resource. Therefore, the objectives of this study were to evaluate the cytotoxic activity of water extracts of Trichilia hirta leaves on tumour cells and identify through a phytochemical screening the principal families of phytocomponents contained in these extracts. The cytotoxic activity of these extracts was also evaluated on human melanoma cells (SK-mel-3) and human breast carcinoma (T-47D). The African green monkey kidney (AGMK) cells Cercopithecus aethiops (Vero) were used as a non-tumour cells control. The results showed the presence of triterpenes/steroids, saponins, coumarins, reductor sugars, phenols and tannins, flavonoids and carbohydrates/glycosides in the extracts. The water leaf extracts showed cytotoxic activity mainly on tumour cells, which contributes to explain the referred recovery by patients suffering form cancer that traditionally consume these extracts

Label-free quantitative cell division monitoring of endothelial cells by digital holographic microscopy

Science.gov (United States)

Kemper, Björn; Bauwens, Andreas; Vollmer, Angelika; Ketelhut, Steffi; Langehanenberg, Patrik; Müthing, Johannes; Karch, Helge; von Bally, Gert

2010-05-01

Digital holographic microscopy (DHM) enables quantitative multifocus phase contrast imaging for nondestructive technical inspection and live cell analysis. Time-lapse investigations on human brain microvascular endothelial cells demonstrate the use of DHM for label-free dynamic quantitative monitoring of cell division of mother cells into daughter cells. Cytokinetic DHM analysis provides future applications in toxicology and cancer research.

Fruit extract from a Sechium edule hybrid induce apoptosis in leukaemic cell lines but not in normal cells.

Science.gov (United States)

Aguiñiga-Sánchez, Itzen; Soto-Hernández, Marcos; Cadena-Iñiguez, Jorge; Ruíz-Posadas, Lucero del Mar; Cadena-Zamudio, Jorge David; González-Ugarte, Ana Karen; Steider, Benny Weiss; Santiago-Osorio, Edelmiro

2015-01-01

The antiproliferative potential of a crude extract from the chayote hybrid H-837-07-GISeM® and its potential for apoptosis induction were assessed in leukaemic cell lines and normal mouse bone marrow mononuclear cells (BM-MNCs). The extract strongly inhibited the proliferation of the P388, J774, and WEHI-3 cell lines (with an IC50 below 1.3 μg·mL(-1)), reduced cell viability, and induced apoptotic body production, phosphatidylserine translocation, and DNA fragmentation. However, the extract had no effect on BM-MNCs. We postulate that these properties make the extract a good candidate for an anti-tumour agent for clinical use.

Validation of cell-free culture using scanning electron microscopy (SEM) and gene expression studies.

Science.gov (United States)

Yang, R; Elankumaran, Y; Hijjawi, N; Ryan, U

2015-06-01

A cell-free culture system for Cryptosporidium parvum was analysed using scanning electron microscopy (SEM) to characterise life cycle stages and compare gene expression in cell-free culture and cell culture using HCT-8 cells. Cryptosporidium parvum samples were harvested at 2 h, 8 h, 14 h, 26 h, 50 h, 74 h, 98 h, 122 h and 170 h, chemically fixed and specimens were observed using a Zeiss 1555 scanning electron microscope. The presence of sporozoites, trophozoites and type I merozoites were identified by SEM. Gene expression in cell culture and cell-free culture was studied using reverse transcriptase quantitative PCR (RT-qPCR) of the sporozoite surface antigen protein (cp15), the glycoprotein 900 (gp900), the Cryptosporidium oocyst wall protein (COWP) and 18S ribosomal RNA (rRNA) genes in both cell free and conventional cell culture. In cell culture, cp15 expression peaked at 74 h, gp900 expression peaked at 74 h and 98 h and COWP expression peaked at 50 h. In cell-free culture, CP15 expression peaked at 98 h, gp900 expression peaked at 74 h and COWP expression peaked at 122 h. The present study is the first to compare gene expression of C. parvum in cell culture and cell-free culture and to characterise life cycle stages of C. parvum in cell-free culture using SEM. Findings from this study showed that gene expression patterns in cell culture and cell-free culture were similar but in cell-free culture, gene expression was delayed for CP15 and COWP in cell free culture compared with the cell culture system and was lower. Although three life cycle stageswere conclusively identified, improvements in SEM methodology should lead to the detection of more life cycle stages. Copyright © 2015 Elsevier Inc. All rights reserved.

The Effect of Sericin from Various Extraction Methods on Cell Viability and Collagen Production

Directory of Open Access Journals (Sweden)

Pornanong Aramwit

2010-05-01

Full Text Available Silk sericin (SS can accelerate cell proliferation and attachment; however, SS can be extracted by various methods, which result in SS exhibiting different physical and biological properties. We found that SS produced from various extraction methods has different molecular weights, zeta potential, particle size and amino acid content. The MTT assay indicated that SS from all extraction methods had no toxicity to mouse fibroblast cells at concentrations up to 40 μg/mL after 24 h incubation, but SS obtained from some extraction methods can be toxic at higher concentrations. Heat-degraded SS was the least toxic to cells and activated the highest collagen production, while urea-extracted SS showed the lowest cell viability and collagen production. SS from urea extraction was severely harmful to cells at concentrations higher than 100 μg/mL. SS from all extraction methods could still promote collagen production in a concentration-dependent manner, even at high concentrations that are toxic to cells.

Effect of Melanin Free Ink Extracted From Squid (Loligo sp.) on Proximate and Sensory Characteristics of Soft-Bone Milkfish (Chanos chanos) During Storage

Science.gov (United States)

Winarni Agustini, Tri; Hadiyanto; Wijayanti, Ima; Amalia, Ulfah; Benjakul, Soottawat

2018-02-01

Antioxidant could be extracted and isolated from squid inks. Squid ink in the form of melanin free ink (MFI) could be act as an electron donor which can stabilize free radicals in lipid oxidation. This study was carried out to assess the antioxidant activity of squid inks converted into MFI in different dilution and to optimize the extraction conditions for the application of MFI as an antioxidative agent on fish product. Three different types of MFI extracts i.e : pure squid ink, squid ink with 5 times dilution and squid ink with 10 times dilutions by using cooled ionized water (4°C). The ink was then centrifuged at 18.000 x g for 30 minutes at cooled centrifuge (4°C) followed by DPPH analysis. The results showed that the IC50 of MFI extracts were 2.84 ppm; 1.11 ppm and 0.34 ppm, respectively (p squid ink with 10 times dilution in extraction of MFI had the highest value in free radical inhibitory. Although the IC50 of three different dilutions are equally low, and are considered as very strong antioxidative agent, however, it showed that the MFI extracted from squid ink had the ability to prevent free radical

IMPLEMENTATION OF THE IMPROVED QUASI-STATIC METHOD IN RATTLESNAKE/MOOSE FOR TIME-DEPENDENT RADIATION TRANSPORT MODELLING

Energy Technology Data Exchange (ETDEWEB)

Zachary M. Prince; Jean C. Ragusa; Yaqi Wang

2016-02-01

Because of the recent interest in reactor transient modeling and the restart of the Transient Reactor (TREAT) Facility, there has been a need for more efficient, robust methods in computation frameworks. This is the impetus of implementing the Improved Quasi-Static method (IQS) in the RATTLESNAKE/MOOSE framework. IQS has implemented with CFEM diffusion by factorizing flux into time-dependent amplitude and spacial- and weakly time-dependent shape. The shape evaluation is very similar to a flux diffusion solve and is computed at large (macro) time steps. While the amplitude evaluation is a PRKE solve where the parameters are dependent on the shape and is computed at small (micro) time steps. IQS has been tested with a custom one-dimensional example and the TWIGL ramp benchmark. These examples prove it to be a viable and effective method for highly transient cases. More complex cases are intended to be applied to further test the method and its implementation.

Rapid and label-free separation of Burkitt's lymphoma cells from red blood cells by optically-induced electrokinetics.

Directory of Open Access Journals (Sweden)

Wenfeng Liang

Full Text Available Early stage detection of lymphoma cells is invaluable for providing reliable prognosis to patients. However, the purity of lymphoma cells in extracted samples from human patients' marrow is typically low. To address this issue, we report here our work on using optically-induced dielectrophoresis (ODEP force to rapidly purify Raji cells' (a type of Burkitt's lymphoma cell sample from red blood cells (RBCs with a label-free process. This method utilizes dynamically moving virtual electrodes to induce negative ODEP force of varying magnitudes on the Raji cells and RBCs in an optically-induced electrokinetics (OEK chip. Polarization models for the two types of cells that reflect their discriminate electrical properties were established. Then, the cells' differential velocities caused by a specific ODEP force field were obtained by a finite element simulation model, thereby established the theoretical basis that the two types of cells could be separated using an ODEP force field. To ensure that the ODEP force dominated the separation process, a comparison of the ODEP force with other significant electrokinetics forces was conducted using numerical results. Furthermore, the performance of the ODEP-based approach for separating Raji cells from RBCs was experimentally investigated. The results showed that these two types of cells, with different concentration ratios, could be separated rapidly using externally-applied electrical field at a driven frequency of 50 kHz at 20 Vpp. In addition, we have found that in order to facilitate ODEP-based cell separation, Raji cells' adhesion to the OEK chip's substrate should be minimized. This paper also presents our experimental results of finding the appropriate bovine serum albumin concentration in an isotonic solution to reduce cell adhesion, while maintaining suitable medium conductivity for electrokinetics-based cell separation. In short, we have demonstrated that OEK technology could be a promising tool for

Culture of human dental pulp cells at variable times post-tooth extraction

Directory of Open Access Journals (Sweden)

BENÍCIO Daniela Ferreira Araújo

2018-02-01

Full Text Available Abstract The aim of this study was to investigate the viability of human dental pulp cells from extracted teeth kept at standard room temperature and atmospheric pressure for different periods of time. Twenty-one healthy permanent teeth were used. They were divided into five groups according to the expected time from extraction to processing. One group was tested immediately after extraction; the other groups were each tested at one of the following time points: 30 minutes, 1 hour, 2 hours, and 5 hours post-extraction. Cell morphology was analysed by light microscopy; cell proliferation was analysed using MTT assay and by counting the viable cells in a haemocytometer. Similar results were observed in all groups (p < 0.05. A delay of up to five hours for tooth processing and tissue collection does not preclude the establishment of dental pulp cell cultures, affect the morphology of these cells, or reduce their proliferative potential.

Cell-free mitochondrial DNA copy number variation in head and neck squamous cell carcinoma: A study of non-invasive biomarker from Northeast India.

Science.gov (United States)

Kumar, Manish; Srivastava, Shilpee; Singh, Seram Anil; Das, Anup Kumar; Das, Ganesh Chandra; Dhar, Bishal; Ghosh, Sankar Kumar; Mondal, Rosy

2017-10-01

Head and neck squamous cell carcinoma is the most commonly diagnosed cancer worldwide. The lifestyle, food habits, and customary practices manifest the Northeast Indian population toward higher susceptibility to develop head and neck squamous cell carcinoma. Here, we have investigated the association of smoke and smokeless tobacco, and alcohol with copy number variation of cell-free mitochondrial DNA and cell-free nuclear DNA in cases and controls. Cell-free DNA from plasma was isolated from 50 head and neck squamous cell carcinoma cases and 50 controls with informed written consent using QIAamp Circulating Nucleic Acid Kit. Real-time polymerase chain reaction was done for copy number variation in cell-free mitochondrial DNA and cell-free nuclear DNA. Receiver operating characteristic curve analysis was performed to evaluate the diagnostic application between the two study groups using clinicopathological parameters. The levels of cell-free nuclear DNA and cell-free mitochondrial DNA of cases in association with smoke and smokeless tobacco, alcohol with smoking (p squamous cell carcinoma cases and controls, we distinguished cell-free mitochondrial DNA (cutoff: 19.84 raw Ct; sensitivity: 84%; specificity: 100%; p < 0.001) and cell-free nuclear DNA (cutoff: 463,282 genomic equivalent/mL; sensitivity: 53%; specificity: 87%; p < 0.001). The copy number variation in cases (cell-free nuclear DNA: 5451.66 genomic equivalent/mL and cell-free mitochondrial DNA: 29,103,476.15 genomic equivalent/mL) and controls (cell-free nuclear DNA: 1650.9 genomic equivalent/mL and cell-free mitochondrial DNA: 9,189,312.54 genomic equivalent/mL), respectively. Our result indicates that the cell-free mitochondrial DNA content is highly associated with smoke and smokeless tobacco, betel quid chewing, and alcohol which shows greater promises, holding the key characteristics of diagnostic biomarkers, that is, minimal invasiveness, high specificity, and sensitivity.

Mesostructured platinum-free anode and carbon-free cathode catalysts for durable proton exchange membrane fuel cells.

Science.gov (United States)

Cui, Xiangzhi; Shi, Jianlin; Wang, Yongxia; Chen, Yu; Zhang, Lingxia; Hua, Zile

2014-01-01

As one of the most important clean energy sources, proton exchange membrane fuel cells (PEMFCs) have been a topic of extensive research focus for decades. Unfortunately, several critical technique obstacles, such as the high cost of platinum electrode catalysts, performance degradation due to the CO poisoning of the platinum anode, and carbon corrosion by oxygen in the cathode, have greatly impeded its commercial development. A prototype of a single PEMFC catalyzed by a mesostructured platinum-free WO3/C anode and a mesostructured carbon-free Pt/WC cathode catalysts is reported herein. The prototype cell exhibited 93% power output of a standard PEMFC using commercial Pt/C catalysts at 50 and 70 °C, and more importantly, CO poisoning-free and carbon corrosion-resistant characters of the anode and cathode, respectively. Consequently, the prototype cell demonstrated considerably enhanced cell operation durability. The mesostructured electrode catalysts are therefore highly promising in the future development and application of PEMFCs. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Inhibition of vascular smooth muscle cell proliferation by Gentiana lutea root extracts.

Directory of Open Access Journals (Sweden)

Rushendhiran Kesavan

Full Text Available Gentiana lutea belonging to the Gentianaceae family of flowering plants are routinely used in traditional Serbian medicine for their beneficial gastro-intestinal and anti-inflammatory properties. The aim of the study was to determine whether aqueous root extracts of Gentiana lutea consisting of gentiopicroside, gentisin, bellidifolin-8-O-glucoside, demethylbellidifolin-8-O-glucoside, isovitexin, swertiamarin and amarogentin prevents proliferation of aortic smooth muscle cells in response to PDGF-BB. Cell proliferation and cell cycle analysis were performed based on alamar blue assay and propidium iodide labeling respectively. In primary cultures of rat aortic smooth muscle cells (RASMCs, PDGF-BB (20 ng/ml induced a two-fold increase in cell proliferation which was significantly blocked by the root extract (1 mg/ml. The root extract also prevented the S-phase entry of synchronized cells in response to PDGF. Furthermore, PDGF-BB induced ERK1/2 activation and consequent increase in cellular nitric oxide (NO levels were also blocked by the extract. These effects of extract were due to blockade of PDGF-BB induced expression of iNOS, cyclin D1 and proliferating cell nuclear antigen (PCNA. Docking analysis of the extract components on MEK1, the upstream ERK1/2 activating kinase using AutoDock4, indicated a likely binding of isovitexin to the inhibitor binding site of MEK1. Experiments performed with purified isovitexin demonstrated that it successfully blocks PDGF-induced ERK1/2 activation and proliferation of RASMCs in cell culture. Thus, Gentiana lutea can provide novel candidates for prevention and treatment of atherosclerosis.

Inhibition of vascular smooth muscle cell proliferation by Gentiana lutea root extracts.

Science.gov (United States)

Kesavan, Rushendhiran; Potunuru, Uma Rani; Nastasijević, Branislav; T, Avaneesh; Joksić, Gordana; Dixit, Madhulika

2013-01-01

Gentiana lutea belonging to the Gentianaceae family of flowering plants are routinely used in traditional Serbian medicine for their beneficial gastro-intestinal and anti-inflammatory properties. The aim of the study was to determine whether aqueous root extracts of Gentiana lutea consisting of gentiopicroside, gentisin, bellidifolin-8-O-glucoside, demethylbellidifolin-8-O-glucoside, isovitexin, swertiamarin and amarogentin prevents proliferation of aortic smooth muscle cells in response to PDGF-BB. Cell proliferation and cell cycle analysis were performed based on alamar blue assay and propidium iodide labeling respectively. In primary cultures of rat aortic smooth muscle cells (RASMCs), PDGF-BB (20 ng/ml) induced a two-fold increase in cell proliferation which was significantly blocked by the root extract (1 mg/ml). The root extract also prevented the S-phase entry of synchronized cells in response to PDGF. Furthermore, PDGF-BB induced ERK1/2 activation and consequent increase in cellular nitric oxide (NO) levels were also blocked by the extract. These effects of extract were due to blockade of PDGF-BB induced expression of iNOS, cyclin D1 and proliferating cell nuclear antigen (PCNA). Docking analysis of the extract components on MEK1, the upstream ERK1/2 activating kinase using AutoDock4, indicated a likely binding of isovitexin to the inhibitor binding site of MEK1. Experiments performed with purified isovitexin demonstrated that it successfully blocks PDGF-induced ERK1/2 activation and proliferation of RASMCs in cell culture. Thus, Gentiana lutea can provide novel candidates for prevention and treatment of atherosclerosis.

Anticancer Effect of Ginger Extract against Pancreatic Cancer Cells Mainly through Reactive Oxygen Species-Mediated Autotic Cell Death

Science.gov (United States)

Akimoto, Miho; Iizuka, Mari; Kanematsu, Rie; Yoshida, Masato; Takenaga, Keizo

2015-01-01

The extract of ginger (Zingiber officinale Roscoe) and its major pungent components, [6]-shogaol and [6]-gingerol, have been shown to have an anti-proliferative effect on several tumor cell lines. However, the anticancer activity of the ginger extract in pancreatic cancer is poorly understood. Here, we demonstrate that the ethanol-extracted materials of ginger suppressed cell cycle progression and consequently induced the death of human pancreatic cancer cell lines, including Panc-1 cells. The underlying mechanism entailed autosis, a recently characterized form of cell death, but not apoptosis or necroptosis. The extract markedly increased the LC3-II/LC3-I ratio, decreased SQSTM1/p62 protein, and enhanced vacuolization of the cytoplasm in Panc-1 cells. It activated AMPK, a positive regulator of autophagy, and inhibited mTOR, a negative autophagic regulator. The autophagy inhibitors 3-methyladenine and chloroquine partially prevented cell death. Morphologically, however, focal membrane rupture, nuclear shrinkage, focal swelling of the perinuclear space and electron dense mitochondria, which are unique morphological features of autosis, were observed. The extract enhanced reactive oxygen species (ROS) generation, and the antioxidant N-acetylcystein attenuated cell death. Our study revealed that daily intraperitoneal administration of the extract significantly prolonged survival (P = 0.0069) in a peritoneal dissemination model and suppressed tumor growth in an orthotopic model of pancreatic cancer (P < 0.01) without serious adverse effects. Although [6]-shogaol but not [6]-gingerol showed similar effects, chromatographic analyses suggested the presence of other constituent(s) as active substances. Together, these results show that ginger extract has potent anticancer activity against pancreatic cancer cells by inducing ROS-mediated autosis and warrants further investigation in order to develop an efficacious candidate drug. PMID:25961833

Anticancer Effect of Ginger Extract against Pancreatic Cancer Cells Mainly through Reactive Oxygen Species-Mediated Autotic Cell Death.

Directory of Open Access Journals (Sweden)

Miho Akimoto

Full Text Available The extract of ginger (Zingiber officinale Roscoe and its major pungent components, [6]-shogaol and [6]-gingerol, have been shown to have an anti-proliferative effect on several tumor cell lines. However, the anticancer activity of the ginger extract in pancreatic cancer is poorly understood. Here, we demonstrate that the ethanol-extracted materials of ginger suppressed cell cycle progression and consequently induced the death of human pancreatic cancer cell lines, including Panc-1 cells. The underlying mechanism entailed autosis, a recently characterized form of cell death, but not apoptosis or necroptosis. The extract markedly increased the LC3-II/LC3-I ratio, decreased SQSTM1/p62 protein, and enhanced vacuolization of the cytoplasm in Panc-1 cells. It activated AMPK, a positive regulator of autophagy, and inhibited mTOR, a negative autophagic regulator. The autophagy inhibitors 3-methyladenine and chloroquine partially prevented cell death. Morphologically, however, focal membrane rupture, nuclear shrinkage, focal swelling of the perinuclear space and electron dense mitochondria, which are unique morphological features of autosis, were observed. The extract enhanced reactive oxygen species (ROS generation, and the antioxidant N-acetylcystein attenuated cell death. Our study revealed that daily intraperitoneal administration of the extract significantly prolonged survival (P = 0.0069 in a peritoneal dissemination model and suppressed tumor growth in an orthotopic model of pancreatic cancer (P < 0.01 without serious adverse effects. Although [6]-shogaol but not [6]-gingerol showed similar effects, chromatographic analyses suggested the presence of other constituent(s as active substances. Together, these results show that ginger extract has potent anticancer activity against pancreatic cancer cells by inducing ROS-mediated autosis and warrants further investigation in order to develop an efficacious candidate drug.

Antioxidant Potential of Selected Korean Edible Plant Extracts

Directory of Open Access Journals (Sweden)

Yaejin Woo

2017-01-01

Full Text Available This study aimed to evaluate the antioxidant activity of various plant extracts. A total of 94 kinds of edible plant extracts obtained from the Korea Plant Extract Bank were screened for cytotoxicity, following which the total phenolic content of 24 shortlisted extracts was determined. Of these, extracts from three plants, namely, Castanea crenata (CC leaf, Camellia japonica (CJ fruit, and Viburnum dilatatum (VD leaf, were examined for antioxidant capabilities by measuring radical scavenging activity, ferric reducing/antioxidant power, and lipid peroxidation inhibitory activity. In addition, cellular antioxidant activities of the three extracts were assessed by a cell-based dichlorofluorescein assay and antioxidant response element (ARE reporter activity assay. The results demonstrated that all three extracts concentration-dependently scavenged free radicals, inhibited lipid peroxidation, reduced the cellular level of reactive oxygen species, and increased ARE-luciferase activity, indicating antioxidant enzyme-inducing potential. In particular, CJ extract showed significantly greater antioxidative activity and antimigratory effect in a breast cancer cell line compared to CC and VD extracts. Hence, CJ extract deserves further study for its in vivo functionality or biologically active constituents.

Free radical scavenging and total antioxidant capacity of root extracts of Anchomanes difformis Engl. (Araceae).

Science.gov (United States)

Aliyu, Abubakar B; Ibrahim, Mohammed A; Musa, Aliyu M; Musa, Aisha O; Kiplimo, Joyce J; Oyewale, Adebayo O

2013-01-01

Antioxidants activities from plants sources have attracted a wide range of interest across the world in recent times. This is due to growing concern for safe and alternative sources of antioxidants. The free radical scavenging activity using 1,1-diphenyl-2-picrylhydrazyl radical (DPPH), reducing power assay, total antioxidant capacity of the phosphomolybdenum method and the total phenolics content using the Folin-Ciocalteu reagent were carried out on the acetone, n-butanol and methanol root extracts of Anchomanes difformis. The results of the total phenolics content expressed in mg/100 g of gallic acid equivalent (GAE) showed that the n-butanol extract has significantly (p extracts. All the extracts displayed strong concentration dependent radical scavenging activity. It was also observed that the n-butanol extract showed higher activity of 70.87% and 78.59% at low concentrations of 31.25 microg/mL and 62.5 microg/mL, respectively, than methanol and acetone extracts. The results also showed that the n-butanol extract has strongest reducing ability which is comparable to that of gallic acid at all the concentrations tested. Phytochemical screening on the extracts revealed the presence of flavonoids, saponins, and tannins. The results suggest that n-butanol extract of the plant is very rich in antioxidant compounds worthy of further investigations.

Advancement on Lead-Free Organic-Inorganic Halide Perovskite Solar Cells: A Review.

Science.gov (United States)

Sani, Faruk; Shafie, Suhaidi; Lim, Hong Ngee; Musa, Abubakar Ohinoyi

2018-06-14

Remarkable attention has been committed to the recently discovered cost effective and solution processable lead-free organic-inorganic halide perovskite solar cells. Recent studies have reported that, within five years, the reported efficiency has reached 9.0%, which makes them an extremely promising and fast developing candidate to compete with conventional lead-based perovskite solar cells. The major challenge associated with the conventional perovskite solar cells is the toxic nature of lead (Pb) used in the active layer of perovskite material. If lead continues to be used in fabricating solar cells, negative health impacts will result in the environment due to the toxicity of lead. Alternatively, lead free perovskite solar cells could give a safe way by substituting low-cost, abundant and non toxic material. This review focuses on formability of lead-free organic-inorganic halide perovskite, alternative metal cations candidates to replace lead (Pb), and possible substitutions of organic cations, as well as halide anions in the lead-free organic-inorganic halide perovskite architecture. Furthermore, the review gives highlights on the impact of organic cations, metal cations and inorganic anions on stability and the overall performance of lead free perovskite solar cells.

Free radical scavenging activity of ethanolic extracts from herbs and spices commercialized in Brazil

Directory of Open Access Journals (Sweden)

Lilian Regina Barros Mariutti

2008-12-01

Full Text Available Ethanolic extracts from 23 different dried herbs and spices commercialized in Brazil were investigated for their free radical scavenging properties using the stable free radicals 2,2'-diphenyl-β-picrylhydrazyl (DPPH• and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid (ABTS•+, and Trolox as reference (TEAC for both radicals. The kinetic curves for both radicals showed to follow the first order kinetics model and the decay rate constant (k obs was calculated. For all the samples, the two methods showed a good linear TEAC correlation, indicating that the average reactivity of the compounds present in the ethanolic extracts was similar. Sage and rosemary extracts showed the highest free radical scavenging capacities, while onion showed negligible activity and colorifico, one of the most consumed spices in Brazil, showed low ABTS•+ scavenging activity. Three distinct situations were found for the extracts concerning the DPPH• scavenging capacities: (1 extracts, like rosemary and laurel, that presented the same efficient concentrations (EC50 but differed in the TEAC values and velocities of action (k obs, (2 extracts, such as garlic and basil, that showed similar EC50 and TEAC values, but different k obs values and (3 extracts that reacted at the same velocities but completely differed in the free radical scavenging capacities, like black pepper, savory, nutmeg, rosemary and sage. Similar considerations could be done for the ABTS•+ results. For the first time the ABTS•+ scavenging activity for allspice, basil, cardamom, chives, colorifico, cumin, dill, laurel, marjoram, parsley and tarragon was reported.Extratos etanólicos de 23 ervas e condimentos desidratados comercializados no Brasil foram analisados quanto as suas propriedades antioxidantes utilizando os radicais 2,2'-difenil-β-picrilhidrazil (DPPH• e ácido 2,2'-azino-bis(3-etilbenzotiazolina-6-sulfônico (ABTS•+, Trolox foi usado como referência para ambos radicais

Cytotoxic Effects of Alcoholic Extract of Dorema Glabrum Seed on Cancerous Cells Viability

Directory of Open Access Journals (Sweden)

Maryam Bannazadeh Amirkhiz

2013-08-01

Full Text Available Purpose: In the present study cytotoxic effects of the alcoholic extract of Dorema Glabrum seed on viability of WEHI-164 cells, mouse Fibrosarcoma cell line and L929 normal cells were compared with the cytotoxic effects of Taxol (anticancer and apoptosis inducer drug. Methods: To find out the plant extract cytotoxic effects, MTT test and DNA fragmentation assay, the biochemical hallmark of apoptosis were performed on cultured and treated cells. Results: According to the findings the alcoholic extract of Dorema Glabrum seed can alter cells morphology and because of chromatin condensation and other changes they shrink and take a spherical shape, and lose their attachment too. So the plant extract inhibits cell growth albeit in a time and dose dependent manner and results in degradation of chromosomal DNA. Conclusion: Our data well established the anti-proliferative effect of methanolic extract of Dorema Glabrum seed and clearly showed that the plant extract can induce apoptosis and not necrosis in vitro, but the mechanism of its activities remained unknown. These results demonstrated that Dorema Glabrum seed might be a novel and attractive therapeutic candidate for tumor treatment in clinical practices.

The value of cell-free DNA for molecular pathology.

Science.gov (United States)

Stewart, Caitlin M; Kothari, Prachi D; Mouliere, Florent; Mair, Richard; Somnay, Saira; Benayed, Ryma; Zehir, Ahmet; Weigelt, Britta; Dawson, Sarah-Jane; Arcila, Maria E; Berger, Michael F; Tsui, Dana Wy

2018-04-01

Over the past decade, advances in molecular biology and genomics techniques have revolutionized the diagnosis and treatment of cancer. The technological advances in tissue profiling have also been applied to the study of cell-free nucleic acids, an area of increasing interest for molecular pathology. Cell-free nucleic acids are released from tumour cells into the surrounding body fluids and can be assayed non-invasively. The repertoire of genomic alterations in circulating tumour DNA (ctDNA) is reflective of both primary tumours and distant metastatic sites, and ctDNA can be sampled multiple times, thereby overcoming the limitations of the analysis of single biopsies. Furthermore, ctDNA can be sampled regularly to monitor response to treatment, to define the evolution of the tumour genome, and to assess the acquisition of resistance and minimal residual disease. Recently, clinical ctDNA assays have been approved for guidance of therapy, which is an exciting first step in translating cell-free nucleic acid research tests into clinical use for oncology. In this review, we discuss the advantages of cell-free nucleic acids as analytes in different body fluids, including blood plasma, urine, and cerebrospinal fluid, and their clinical applications in solid tumours and haematological malignancies. We will also discuss practical considerations for clinical deployment, such as preanalytical factors and regulatory requirements. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Solvent extraction of Au(III) for preparation of a carrier-free multitracer and an Au tracer from an Au target

International Nuclear Information System (INIS)

Weginwar, R.G.; Kobayashi, Y.; Ambe, S.; Liu, B.; Enomoto, S.; Ambe, F.

1996-01-01

Separation of Au(III) and various carrier-free radionuclides by solvent extraction was investigated using an Au target irradiated by an energetic heavy-ion beam. Percentage extraction of Au(III) and coextraction of the radionuclides were determined with varying parameters such as kinds of solvent, molarity of HCl or pH, and Au concentration. Under the conditions where Au(III) was effectively extracted, namely extraction with ethyl acetate or isobutyl methyl ketone from 3 mol*dm -3 HCl, carrier-free radionuclides of many elements were found to be more or less coextracted. Coextraction of radionuclides of some elements was found to increase with an increase in the concentration of Au(III). This finding is ascribed to the formation of strong association of the complex of these elements with chloroauric acid. In order to avoid serious loss of these elements by the extraction, lowering of the Au(III) concentration or the use of a masking agent such as sodium citrate is necessary. Gold(III) was shown to be effectively back extracted with a 0.1 mol*dm -3 aqueous solution of 2-amino-2-hydroxymethyl-1,3-propanediol. Thus, a radiochemical procedure has been established for preparing a carrier-free multitracer and an Au tracer with carrier form from an Au target irradiated with a heavy-ion beam. Both tracers are now used individually for chemical and biological experiments. (author). 22 refs., 3 figs., 2 tabs

Insights into cell-free therapeutic approach: Role of stem cell "soup-ernatant".

Science.gov (United States)

Raik, Shalini; Kumar, Ajay; Bhattacharyya, Shalmoli

2018-03-01

Current advances in medicine have revolutionized the field of regenerative medicine dramatically with newly evolved therapies for repair or replacement of degenerating or injured tissues. Stem cells (SCs) can be harvested from different sources for clinical therapeutics, which include fetal tissues, umbilical cord blood, embryos, and adult tissues. SCs can be isolated and differentiated into desired lineages for tissue regeneration and cell replacement therapy. However, several loopholes need to be addressed properly before this can be extended for large-scale therapeutic application. These include a careful approach for patient safety during SC treatments and tolerance of recipients. SC treatments are associated with a number of risk factors and require successful integration and survival of transplanted cells in the desired microenvironment with concurrent tissue regeneration. Recent studies have focused on developing alternatives that can replace the cell-based therapy using paracrine factors. The development of stem "cell free" therapies can be devoted mainly to the use of soluble factors (secretome), extracellular vesicles, and mitochondrial transfer. The present review emphasizes on the paradigms related to the use of SC-based therapeutics and the potential applications of a cell-free approach as an alternative to cell-based therapy in the area of regenerative medicine. © 2017 International Union of Biochemistry and Molecular Biology, Inc.

Limitations of climatic data for inferring species boundaries: insights from speckled rattlesnakes.

Directory of Open Access Journals (Sweden)

Jesse M Meik

Full Text Available Phenotypes, DNA, and measures of ecological differences are widely used in species delimitation. Although rarely defined in such studies, ecological divergence is almost always approximated using multivariate climatic data associated with sets of specimens (i.e., the "climatic niche"; the justification for this approach is that species-specific climatic envelopes act as surrogates for physiological tolerances. Using identical statistical procedures, we evaluated the usefulness and validity of the climate-as-proxy assumption by comparing performance of genetic (nDNA SNPs and mitochondrial DNA, phenotypic, and climatic data for objective species delimitation in the speckled rattlesnake (Crotalus mitchellii complex. Ordination and clustering patterns were largely congruent among intrinsic (heritable traits (nDNA, mtDNA, phenotype, and discordance is explained by biological processes (e.g., ontogeny, hybridization. In contrast, climatic data did not produce biologically meaningful clusters that were congruent with any intrinsic dataset, but rather corresponded to regional differences in atmospheric circulation and climate, indicating an absence of inherent taxonomic signal in these data. Surrogating climate for physiological tolerances adds artificial weight to evidence of species boundaries, as these data are irrelevant for that purpose. Based on the evidence from congruent clustering of intrinsic datasets, we recommend that three subspecies of C. mitchellii be recognized as species: C. angelensis, C. mitchellii, and C. Pyrrhus.

Polyphenol-rich apple (Malus domestica L.) peel extract attenuates arsenic trioxide induced cardiotoxicity in H9c2 cells via its antioxidant activity.

Science.gov (United States)

Vineetha, Vadavanath Prabhakaran; Girija, Seetharaman; Soumya, Rema Sreenivasan; Raghu, Kozhiparambil Gopalan

2014-03-01

Evidences suggest that apple peel has a wide range of polyphenols having antioxidant activity and its consumption has been linked with improved health benefits. Arsenic trioxide (ATO) is a very effective drug for the treatment of acute promyelocytic leukemia (APL) but it leads to cardiotoxicity mediated through alterations in various cardiac ion channels and by increasing the intracellular calcium level and reactive oxygen species (ROS). The aim of the present investigation was to study the effect of methanolic extract of apple peel (APME) and aqueous extract of apple peel (APAE) on ATO (5 μM) induced toxicity in the H9c2 cardiac myoblast cell line. We estimated the cellular status of innate antioxidant enzymes, level of ROS, mitochondrial superoxide, glutathione and intracellular calcium with ATO and apple peel extracts. Prior to the cell line based study, we had evaluated the antioxidant potential of apple peel extract by 1,1-diphenyl-2-picrylhydrazyl (DPPH), total reducing power (TRP), superoxide anion and hydroxyl radical scavenging activity, in addition to quantifying total phenolic and flavonoid content. Both the extracts showed considerable antioxidant activity in cell-free chemical assays. In addition, both APME and APAE prevented the alteration in antioxidant status induced by ATO in H9c2 cells. Significant differential alterations had been observed in the activity of lactate dehydrogenase, superoxide dismutase, catalase, glutathione, glutathione peroxidase, thioredoxin reductase, xanthine oxidase, calcium overload and caspase 3 activity with ATO. The overall result revealed the protective property of polyphenol-rich apple peel extract against ATO induced cardiac toxicity via its antioxidant activity.

Generation of clinical-grade human induced pluripotent stem cells in Xeno-free conditions.

Science.gov (United States)

Wang, Juan; Hao, Jie; Bai, Donghui; Gu, Qi; Han, Weifang; Wang, Lei; Tan, Yuanqing; Li, Xia; Xue, Ke; Han, Pencheng; Liu, Zhengxin; Jia, Yundan; Wu, Jun; Liu, Lei; Wang, Liu; Li, Wei; Liu, Zhonghua; Zhou, Qi

2015-11-12

Human induced pluripotent stem cells (hiPSCs) are considered as one of the most promising seed cell sources in regenerative medicine. Now hiPSC-based clinical trials are underway. To ensure clinical safety, cells used in clinical trials or therapies should be generated under GMP conditions, and with Xeno-free culture media to avoid possible side effects like immune rejection that induced by the Xeno reagents. However, up to now there are no reports for hiPSC lines developed completely under GMP conditions using Xeno-free reagents. Clinical-grade human foreskin fibroblast (HFF) cells used as feeder cells and parental cells of the clinical-grade hiPSCs were isolated from human foreskin tissues and cultured in Xeno-free media. Clinical-grade hiPSCs were derived by integration-free Sendai virus-based reprogramming kit in Xeno-free pluriton™ reprogramming medium or X medium. Neural cells and cardiomyocytes differentiation were conducted following a series of spatial and temporal specific signals induction according to the corresponding lineage development signals. Biological safety evaluation of the clinical-grade HFF cells and hiPSCs were conducted following the guidance of the "Pharmacopoeia of the People's Republic of China, Edition 2010, Volume III". We have successfully derived several integration-free clinical-grade hiPSC lines under GMP-controlled conditions and with Xeno-free reagents culture media in line with the current guidance of international and national evaluation criteria. As for the source of hiPSCs and feeder cells, biological safety evaluation of the HFF cells have been strictly reviewed by the National Institutes for Food and Drug Control (NIFDC). The hiPSC lines are pluripotent and have passed the safety evaluation. Moreover, one of the randomly selected hiPSC lines was capable of differentiating into functional neural cells and cardiomyocytes in Xeno-free culture media. The clinical-grade hiPSC lines therefore could be valuable sources for

Solvent-Free Microwave-Assisted Extraction of Polyphenols from Olive Tree Leaves: Antioxidant and Antimicrobial Properties

Directory of Open Access Journals (Sweden)

Selin Şahin

2017-06-01

Full Text Available Response surface methodology (RSM and artificial neural networks (ANN were evaluated and compared in order to decide which method was the most appropriate to predict and optimize total phenolic content (TPC and oleuropein yields in olive tree leaf (Olea europaea extracts, obtained after solvent-free microwave-assisted extraction (SFMAE. The SFMAE processing conditions were: microwave irradiation power 250–350 W, extraction time 2–3 min, and the amount of sample 5–10 g. Furthermore, the antioxidant and antimicrobial activities of the olive leaf extracts, obtained under optimal extraction conditions, were assessed by several in vitro assays. ANN had better prediction performance for TPC and oleuropein yields compared to RSM. The optimum extraction conditions to recover both TPC and oleuropein were: irradiation power 250 W, extraction time 2 min, and amount of sample 5 g, independent of the method used for prediction. Under these conditions, the maximal yield of oleuropein (0.060 ± 0.012 ppm was obtained and the amount of TPC was 2.480 ± 0.060 ppm. Moreover, olive leaf extracts obtained under optimum SFMAE conditions showed antibacterial activity against S. aureus and S. epidermidis, with a minimum inhibitory concentration (MIC value of 1.25 mg/mL.

Solvent-Free Microwave-Assisted Extraction of Polyphenols from Olive Tree Leaves: Antioxidant and Antimicrobial Properties.

Science.gov (United States)

Şahin, Selin; Samli, Ruya; Tan, Ayşe Seher Birteksöz; Barba, Francisco J; Chemat, Farid; Cravotto, Giancarlo; Lorenzo, José M

2017-06-24

Response surface methodology (RSM) and artificial neural networks (ANN) were evaluated and compared in order to decide which method was the most appropriate to predict and optimize total phenolic content (TPC) and oleuropein yields in olive tree leaf ( Olea europaea ) extracts, obtained after solvent-free microwave-assisted extraction (SFMAE). The SFMAE processing conditions were: microwave irradiation power 250-350 W, extraction time 2-3 min, and the amount of sample 5-10 g. Furthermore, the antioxidant and antimicrobial activities of the olive leaf extracts, obtained under optimal extraction conditions, were assessed by several in vitro assays. ANN had better prediction performance for TPC and oleuropein yields compared to RSM. The optimum extraction conditions to recover both TPC and oleuropein were: irradiation power 250 W, extraction time 2 min, and amount of sample 5 g, independent of the method used for prediction. Under these conditions, the maximal yield of oleuropein (0.060 ± 0.012 ppm) was obtained and the amount of TPC was 2.480 ± 0.060 ppm. Moreover, olive leaf extracts obtained under optimum SFMAE conditions showed antibacterial activity against S. aureus and S. epidermidis , with a minimum inhibitory concentration (MIC) value of 1.25 mg/mL.

Antioxidant Activity of Ixora parviflora in a Cell/Cell-Free System and in UV-Exposed Human Fibroblasts

Directory of Open Access Journals (Sweden)

Hsiu-Mei Chiang

2011-07-01

Full Text Available Polyphenols and flavonoids possess a variety of biological activities including antioxidant and anti-tumor activities. Ixora parviflora is a member of the flavonoid-rich Rubiaceae family of flowering plants and used as folk medicine in India. The aim of this study was to investigate the antioxidant activity of Ixora parviflora extract (IPE in a cell-free system and erythrocytes, and the ability of IPE to inhibit reactive oxygen species (ROS generation in human fibroblasts (Hs68 after ultraviolet (UV exposure. Various in vitro antioxidant assays were employed in this study. The extraction yield of IPE was 17.4 ± 3.9%, the total phenolic content of IPE was 26.2 μg gallic acid equivalent (GAE/mg leaves dry weight and the total flavonoids content was 54.2 ± 4.4 μg quercetin equvalent (QE/mg extract. The content of chlorogenic acid was 9.7 ± 1.2 mg/g extract. IPE at 1000 μg/mL exhibited a reducing capacity of 90.5 ± 0.6%, a 1,1-diphenyl-2-picrylhydrazy (DPPH radical scavenging activity of 96.0 ± 0.4%, a ferrous chelating activity of 72.2 ± 3.5%, a hydroxyl radical scavenging activity of 96.8 ± 1.4%, and a hydrogen peroxide scavenging activity of 99.5 ± 3.3%. IPE at 500 μg/mL also possessed inhibitory activity against 2,2'-azobis (2-methylpropionamidine dihydrochloride (AAPH-induced hemolysis of erythrocytes (89.4 ± 1.8% and resulted in a 52.9% reduction in ROS generation in UV-exposed fibroblasts. According to our findings, IPE is a potent antioxidant and a potential anti-photoaging agent.

F NMR measurement of intracellular free calcium in human red blood cells

International Nuclear Information System (INIS)

Gupta, R.K.; Schanne, F.A.X.

1986-01-01

Optical techniques for the measurement of intracellular Ca are not readily applicable to the human red cell because of the intense absorption of hemoglobin. The authors have therefore examined the use of 19 F NMR of 5,5'-difluoro-1,2-bis(o-aminophenoxy) ethane-N,N,N',N'-tetra acetic acid (5FBAPTA) introduced non-disruptively by intracellular hydrolysis of the membrane-permeant acetoxymethyl ester derivative. 19 F NMR spectra of 5FBAPTA-containing erythrocytes at 188 MHz displayed two well resolved resonances corresponding to the free and Ca-bound forms of the chelator, the resonance of the free form being ten-fold larger than that of the Ca-bound form. Addition of the ionophore A23187 resulted in the disappearance of the resonance of the free anion and a quantitative increase in the intensity of the resonance of the Ca-complex. From these data, and a K/sub D/ of 708 nM for the Ca-5FBAPTA complex, the authors estimate red cell free Ca to be 70 nM, which is in the range of values obtained for other cells, despite the fact that the human red cell, which lacks intracellular organelles for storing Ca, possesses only 1 μmol total Ca/1. cells in comparison to mmols of total Ca found in other cells. The authors ability to use 19 F NMR to measure free Ca in the red blood cell paves the way for future NMR studies of red cell free Ca concentrations in human essential hypertension as well as in other diseases states in which alterations in cellular Ca homeostasis may be involved

Dye-sensitized solar cells with natural dyes extracted from achiote seeds

Energy Technology Data Exchange (ETDEWEB)

Gomez-Ortiz, N.M.; Vazquez-Maldonado, I.A.; Azamar-Barrios, J.A.; Oskam, G. [Departamento de Fisica Aplicada, CINVESTAV-IPN, Merida, Yuc. 97310 (Mexico); Perez-Espadas, A.R.; Mena-Rejon, G.J. [Laboratorio de Quimica Organica de Investigacion, Facultad de Quimica, Universidad Autonoma de Yucatan, Merida, Yuc. 97150 (Mexico)

2010-01-15

We have explored the application of natural dyes extracted from the seeds of the achiote shrub (Bixa orellana L.) in dye-sensitized solar cells (DSCs). The main pigments are bixin and norbixin, which were obtained by separation and purification from the dark-red extract (annatto). The dyes were characterized using {sup 1}H-NMR, FTIR spectroscopy, and UV-Vis spectrophotometry. Solar cells were prepared using TiO{sub 2} and ZnO nanostructured, mesoporous films and the annatto, bixin, and norbixin as sensitizers. The best results were obtained with bixin-sensitized TiO{sub 2} solar cells with efficiencies of up to 0.53%, illustrating the importance of purification of dyes from natural extracts. (author)

The effect of different conventional methods of extraction on Marantodes pumila var. pumila leaves in relation to free radical scavenging activity

Science.gov (United States)

Hamid, Noor Huda Abd; Latip, Jalifah; Markom, Masturah

2018-04-01

Marantodes pumila is one of the important herbs used in traditional medicine. However, there is no report on the optimum method for the preparation of M. pumila extract enriched with bioactive compounds. Therefore, this study aim to establish the suitable method in the preparation of M. pumila extracts enriched with antioxidative compounds. Different conventional extraction methods such as maceration, reflux and Soxhlet in solvents of different polarities i.e. ethanol, 50 % aqueous ethanol and water were employed to prepare extracts. Constituents of each extract were profiled using RP-HPLC followed by the free radical scavenging assay using 2,2 diphenyl-1-picrylhydrazyl (DPPH) to determine the antioxidant activity. Among all extraction methods, maceration in 50 % aqueous ethanol gave the highest total percentage yield (22.41 %) while all extraction methods were able to extract the marker compounds, gallic acid (GA) and methyl gallate (MGA). The free radical scavenging activity of extracts varied from IC50 11.93 µg/mL (50 % aqueous ethanol via reflux) to 64.15 µg/mL (water via maceration). All extracts showed better scavenging activity as compared to synthetic antioxidant, BHT. The variation in the IC50 values is expected due to the presence of different types and/or concentration of bioactive compounds in each extract. This study may provide a preliminary insight for the preparation of standardized extracts of M. pumila enriched with bioactive compounds.

Rapid Detection of Cell-Free Mycobacterium tuberculosis DNA in Tuberculous Pleural Effusion.

Science.gov (United States)

Che, Nanying; Yang, Xinting; Liu, Zichen; Li, Kun; Chen, Xiaoyou

2017-05-01

Tuberculous pleurisy is one of the most common types of extrapulmonary tuberculosis, but its diagnosis remains difficult. In this study, we report for the first time on the detection of cell-free Mycobacterium tuberculosis DNA in pleural effusion and an evaluation of a newly developed molecular assay for the detection of cell-free Mycobacterium tuberculosis DNA. A total of 78 patients with pleural effusion, 60 patients with tuberculous pleurisy, and 18 patients with alternative diseases were included in this study. Mycobacterial culture, the Xpert MTB/RIF assay, the adenosine deaminase assay, the T-SPOT.TB assay, and the cell-free Mycobacterium tuberculosis DNA assay were performed on all the pleural effusion samples. The cell-free Mycobacterium tuberculosis DNA assay and adenosine deaminase assay showed significantly higher sensitivities of 75.0% and 68.3%, respectively, than mycobacterial culture and the Xpert MTB/RIF assay, which had sensitivities of 26.7% and 20.0%, respectively ( P pleural effusion showed the highest sensitivity of 95.0% but the lowest specificity of 38.9%. The cell-free Mycobacterium tuberculosis DNA assay detected as few as 1.25 copies of IS 6110 per ml of pleural effusion and showed good accordance of the results between repeated tests ( r = 0.978, P = 2.84 × 10 -10 ). These data suggest that the cell-free Mycobacterium tuberculosis DNA assay is a rapid and accurate molecular test which provides direct evidence of Mycobacterium tuberculosis etiology. Copyright © 2017 American Society for Microbiology.

Ginseng Berry Extract Promotes Maturation of Mouse Dendritic Cells.

Directory of Open Access Journals (Sweden)

Wei Zhang

Full Text Available Ginseng extract has been shown to possess certain anti-virus, anti-tumor and immune-activating effects. However, the immunostimulatory effect of ginseng berry extract (GB has been less well characterized. In this study, we investigated the effect of GB on the activation of mouse dendritic cells (DCs in vitro and in vivo. GB treatment induced up-regulation of co-stimulatory molecules in bone marrow-derived DCs (BMDCs. Interestingly, GB induced a higher degree of co-stimulatory molecule up-regulation than ginseng root extract (GR at the same concentrations. Moreover, in vivo administration of GB promoted up-regulation of CD86, MHC class I and MHC class II and production of IL-6, IL-12 and TNF-α in spleen DCs. GB also promoted the generation of Th1 and Tc1 cells. Furthermore, Toll like receptor 4 (TLR4 and myeloid differentiation primary response 88 (MyD88 signaling pathway were essential for DC activation induced by GB. In addition, GB strongly prompted the proliferation of ovalbumin (OVA-specific CD4 and CD8 T cells. Finally, GB induced DC activation in tumor-bearing mice and the combination of OVA and GB treatment inhibited B16-OVA tumor cell growth in C57BL/6 mice. These results demonstrate that GB is a novel tumor therapeutic vaccine adjuvant by promoting DC and T cell activation.

Cytotoxic and apoptotic activities of black widow spiderling extract against HeLa cells.

Science.gov (United States)

Peng, Xiaozhen; Dai, Zhipan; Lei, Qian; Liang, Long; Yan, Shuai; Wang, Xianchun

2017-06-01

Black widow spiders contain toxic components not only in the venom glands but also in other parts of the spider body, including the legs and abdomen. Additionally, both the eggs and newborn spiderlings of the black widow spider contain venom. It is important to investigate their potential effects on cancer cells. In the present study, the effects of newborn black widow spiderling extract on human HeLa cells were evaluated in vitro . When applied at different concentrations, the total extract decreased HeLa cell viability in a dose-dependent manner, with an IC 50 value of 158 µg/ml. Flow cytometry indicated that treatment of HeLa cells with the total extract of the spiderlings induced apoptosis in HeLa cells in a dose-dependent manner and led to cell cycle arrest in the S-phase. Additionally, application of the total extract at different concentrations increased apoptosis-related caspase 3 activity in a dose-dependent manner. HeLa cells treated with the total extract appeared to be morphologically changed, exhibiting membrane blebbing, nuclear fragmentation and condensation of chromatin. Further separation and activity screening demonstrated that the cytotoxic and apoptotic activities of the total extract were attributable mainly to its high molecular mass proteins, one of which was purified and characterized to determine its anti-tumor activities on HeLa cells. The results of the present study therefore have expanded understanding regarding the effect of spider toxins on cancer cells and suggested that components of black widow spiderlings may be developed as a promising novel agent to treat cancer.

Content of intrinsic disorder influences the outcome of cell-free protein synthesis.

Science.gov (United States)

Tokmakov, Alexander A; Kurotani, Atsushi; Ikeda, Mariko; Terazawa, Yumiko; Shirouzu, Mikako; Stefanov, Vasily; Sakurai, Tetsuya; Yokoyama, Shigeyuki

2015-09-11

Cell-free protein synthesis is used to produce proteins with various structural traits. Recent bioinformatics analyses indicate that more than half of eukaryotic proteins possess long intrinsically disordered regions. However, no systematic study concerning the connection between intrinsic disorder and expression success of cell-free protein synthesis has been presented until now. To address this issue, we examined correlations of the experimentally observed cell-free protein expression yields with the contents of intrinsic disorder bioinformatically predicted in the expressed sequences. This analysis revealed strong relationships between intrinsic disorder and protein amenability to heterologous cell-free expression. On the one hand, elevated disorder content was associated with the increased ratio of soluble expression. On the other hand, overall propensity for detectable protein expression decreased with disorder content. We further demonstrated that these tendencies are rooted in some distinct features of intrinsically disordered regions, such as low hydrophobicity, elevated surface accessibility and high abundance of sequence motifs for proteolytic degradation, including sites of ubiquitination and PEST sequences. Our findings suggest that identification of intrinsically disordered regions in the expressed amino acid sequences can be of practical use for predicting expression success and optimizing cell-free protein synthesis.

Intercellular signaling through secreted proteins induces free-energy gradient-directed cell movement.

Science.gov (United States)

Kravchenko-Balasha, Nataly; Shin, Young Shik; Sutherland, Alex; Levine, R D; Heath, James R

2016-05-17

Controlling cell migration is important in tissue engineering and medicine. Cell motility depends on factors such as nutrient concentration gradients and soluble factor signaling. In particular, cell-cell signaling can depend on cell-cell separation distance and can influence cellular arrangements in bulk cultures. Here, we seek a physical-based approach, which identifies a potential governed by cell-cell signaling that induces a directed cell-cell motion. A single-cell barcode chip (SCBC) was used to experimentally interrogate secreted proteins in hundreds of isolated glioblastoma brain cancer cell pairs and to monitor their relative motions over time. We used these trajectories to identify a range of cell-cell separation distances where the signaling was most stable. We then used a thermodynamics-motivated analysis of secreted protein levels to characterize free-energy changes for different cell-cell distances. We show that glioblastoma cell-cell movement can be described as Brownian motion biased by cell-cell potential. To demonstrate that the free-energy potential as determined by the signaling is the driver of motion, we inhibited two proteins most involved in maintaining the free-energy gradient. Following inhibition, cell pairs showed an essentially random Brownian motion, similar to the case for untreated, isolated single cells.

Antiradical and cytotoxic activity of different Helichrysum plicatum flower extracts.

Science.gov (United States)

Bigović, Dubravka; Savikin, Katarina; Janković, Teodora; Menković, Nebojsa; Zdunić, Gordana; Stanojković, Tatjana; Djurić, Zorica

2011-06-01

Flowers of Helichrysum plicatum were extracted under different experimental conditions, and their antioxidant activity was determined by DPPH radical scavenging assay. Extracts obtained with higher concentration of ethyl acetate (90% or 100%) were found to contain the greatest amount of total phenolics (> 250 mg gallic acid equivalents/g of dried extract), and high correlation between total phenolic content and antiradical activity was observed (r = -0.79). Based on the total phenolic content and antiradical activity, some extracts were selected for investigation of cytotoxic activity toward PC3, HeLa and K562 human cancer cell lines in vitro. All tested extracts exhibited moderate activity against HeLa cells (41.9-42.1 microg/mL), whereas the extract obtained with 100% ethyl acetate was the most active against K562 and PC3 cell lines (25.9 and 39.2 microg/mL, respectively). Statistical analysis revealed significant correlation between total phenolic content and cytotoxic activity against PC3 and K562 cells. HPLC identification of phenolic compounds from the extracts indicated the presence of apigenin, naringenin and kaempferol as free aglycones, and glycosides of apigenin, naringenin, quercetin and kaempferol. Among aglycones, kaempferol displayed moderate cytostatic activity against all cell lines (24.8-64.7 microM).

Synthetic Biology Outside the Cell: Linking Computational Tools to Cell-Free Systems

International Nuclear Information System (INIS)

Lewis, Daniel D.; Villarreal, Fernando D.; Wu, Fan; Tan, Cheemeng

2014-01-01

As mathematical models become more commonly integrated into the study of biology, a common language for describing biological processes is manifesting. Many tools have emerged for the simulation of in vivo synthetic biological systems, with only a few examples of prominent work done on predicting the dynamics of cell-free synthetic systems. At the same time, experimental biologists have begun to study dynamics of in vitro systems encapsulated by amphiphilic molecules, opening the door for the development of a new generation of biomimetic systems. In this review, we explore both in vivo and in vitro models of biochemical networks with a special focus on tools that could be applied to the construction of cell-free expression systems. We believe that quantitative studies of complex cellular mechanisms and pathways in synthetic systems can yield important insights into what makes cells different from conventional chemical systems.

Synthetic Biology Outside the Cell: Linking Computational Tools to Cell-Free Systems

Energy Technology Data Exchange (ETDEWEB)

Lewis, Daniel D. [Integrative Genetics and Genomics, University of California Davis, Davis, CA (United States); Department of Biomedical Engineering, University of California Davis, Davis, CA (United States); Villarreal, Fernando D.; Wu, Fan; Tan, Cheemeng, E-mail: cmtan@ucdavis.edu [Department of Biomedical Engineering, University of California Davis, Davis, CA (United States)

2014-12-09

As mathematical models become more commonly integrated into the study of biology, a common language for describing biological processes is manifesting. Many tools have emerged for the simulation of in vivo synthetic biological systems, with only a few examples of prominent work done on predicting the dynamics of cell-free synthetic systems. At the same time, experimental biologists have begun to study dynamics of in vitro systems encapsulated by amphiphilic molecules, opening the door for the development of a new generation of biomimetic systems. In this review, we explore both in vivo and in vitro models of biochemical networks with a special focus on tools that could be applied to the construction of cell-free expression systems. We believe that quantitative studies of complex cellular mechanisms and pathways in synthetic systems can yield important insights into what makes cells different from conventional chemical systems.

Synthetic Biology Outside the Cell: Linking Computational Tools to Cell-Free Systems

Directory of Open Access Journals (Sweden)

Daniel eLewis

2014-12-01

Full Text Available As mathematical models become more commonly integrated into the study of biology, a common language for describing biological processes is manifesting. Many tools have emerged for the simulation of in vivo systems, with only a few examples of prominent work done on predicting the dynamics of cell-free systems. At the same time, experimental biologists have begun to study dynamics of in vitro systems encapsulated by amphiphilic molecules, opening the door for the development of a new generation of biomimetic systems. In this review, we explore both in vivo and in vitro models of biochemical networks with a special focus on tools that could be applied to the construction of cell-free expression systems. We believe that quantitative studies of complex cellular mechanisms and pathways in synthetic systems can yield important insights into what makes cells different from conventional chemical systems.

Huaier Extract Induces Autophagic Cell Death by Inhibiting the mTOR/S6K Pathway in Breast Cancer Cells.

Directory of Open Access Journals (Sweden)

Xiaolong Wang

Full Text Available Huaier extract is attracting increased attention due to its biological activities, including antitumor, anti-parasite and immunomodulatory effects. Here, we investigated the role of autophagy in Huaier-induced cytotoxicity in MDA-MB-231, MDA-MB-468 and MCF7 breast cancer cells. Huaier treatment inhibited cell viability in all three cell lines and induced various large membranous vacuoles in the cytoplasm. In addition, electron microscopy, MDC staining, accumulated expression of autophagy markers and flow cytometry revealed that Huaier extract triggered autophagy. Inhibition of autophagy attenuated Huaier-induced cell death. Furthermore, Huaier extract inhibited the mammalian target of the rapamycin (mTOR/S6K pathway in breast cancer cells. After implanting MDA-MB-231 cells subcutaneously into the right flank of BALB/c nu/nu mice, Huaier extract induced autophagy and effectively inhibited xenograft tumor growth. This study is the first to show that Huaier-induced cytotoxicity is partially mediated through autophagic cell death in breast cancer cells through suppression of the mTOR/S6K pathway.

Huaier Aqueous Extract Induces Hepatocellular Carcinoma Cells Arrest in S Phase via JNK Signaling Pathway

Directory of Open Access Journals (Sweden)

Chengshuo Zhang

2015-01-01

Full Text Available Huaier aqueous extract, the main active constituent of Huaier proteoglycan, has antihepatocarcinoma activity in experimental and clinical settings. However, the potential and associated antihepatoma mechanisms of Huaier extract are not yet fully understood. Therefore, in this study, we aimed to elucidate the inhibitory proliferation effect of Huaier extract on apoptosis and cycle of HepG2 and Bel-7402 cells. Our data demonstrated that incubation with Huaier extract resulted in a marked decrease in cell viability dose-dependently. Flow cytometric analysis showed that a 48 h treatment of Huaier extract caused cell apoptosis. Typical apoptotic nucleus alterations were observed with fluorescence microscope after Hoechst staining. Immunoblot analysis further demonstrated that Huaier extract activated caspase 3 and PARP. Additionally, Huaier extract inhibited the activity of p-ERK, p-p38, and p-JNK in terms of MAPK. Furthermore, Huaier extract induced HCC cells arrest in S phase and decreased the cycle related protein expression of β-catenin and cyclin D1. Studies with JNK specific inhibitor, SP600125, showed that Huaier extract induced S phase arrest and decreased β-catenin and cyclin D1 expression via JNK signaling pathway. In conclusion, we verify that Huaier extract causes cell apoptosis and induces hepatocellular carcinoma cells arrest in S phase via JNK pathway, which advances our understanding on the molecular mechanisms of Huaier extract in hepatocarcinoma management.

Effect of Azadirachta indica leaves extract on acetic acid-induced colitis in rats:Role of antioxidants, free radicals and myeloperoxidase

Directory of Open Access Journals (Sweden)

Ghatule RR

2012-10-01

Full Text Available Objective: To evaluate the healing effects of extract of dried leaves of Azadirachta indica (Neem on acetic acid-induced colitis in rats. Neem tree is known as ‘arishtha ’ in Sanskrit, meaning ‘reliever of sicknesses ’. Methods: 50% ethanolic extract of Azadirachta indica leaves was administered orally, once daily for 14 days in rats after the induction of colitis with acetic acid and 500 mg/kg dose of extract was found to have an optimal effect against acetic acid-induced colonic damage score, weight and adhesions (Macroscopic. Effect of Azadirachta indica extract was then further studied on various physical (mucous/blood in stool, food and water intake and body weight changes, colonic mucosal damage and inflammation (microscopic, antibacterial and biochemical parameters viz. i antioxidants (superoxide dismutase, catalase and reduced glutathione and ii free radicals (nitric oxide and lipid peroxidation and myeloperoxidase (acute inflammatory marker activities in acetic acid-induced colitis. Results: Azadirachta indica extract decreased colonic mucosal damage and inflammation (macroscopic and microscopic, mucous/bloody diarrhea, fecal frequency and increased body weight. Azadirachta indica extract showed intestinal antibacterial activity and enhanced the antioxidants but decreased free radicals and myeloperoxidase activities. Acute toxicity study indicated no mortality or other ANS or CNS related adverse effects even with 5.0 g/kg dose (10 times of effective dose indicating its safety. Conclusions: Azadirachta indica seemed to be safe and effective in colitis by its predominant effect on promoting antioxidant status and decreasing intestinal bacterial load, free radicals and myeloperoxidase responsible for tissue damage and delayed healing.

Heat-induced chemical and color changes of extractive-free Black Locust (Rosinia Pseudoacacia) wood

Science.gov (United States)

Yao Chen; Jianmin Gao; Yongming Fan; Mandla A. Tshabalala; Nicole M. Stark

2012-01-01

To investigate chemical and color changes of the polymeric constituents of black locust (Robinia pseudoacacia) wood during heat treatment, extractive-free wood flour was conditioned to 30% initial moisture content (MC) and heated for 24 h at 120 °C in either an oxygen or nitrogen atmosphere. The color change was measured using the CIELAB color system. Chemical changes...

Inverted polymer solar cells with Nafion® as the hole extraction layer: efficiency and lifetime studies

International Nuclear Information System (INIS)

Manceau, Matthieu; Berson, Solenn

2014-01-01

The use of Nafion ®  as the hole extraction layer in polymer solar cells is demonstrated in this work. Inverted devices were built on plastic foil with the following architecture: PET/ITO/ZnO/P3HT:PCBM/Nafion ® /Ag. The Nafion ®  was processed from a surfactant-free solution in alcoholic solvents on top of the active layer. Optimization of film thickness and annealing yielded fully functional devices with power conversion efficiency similar to others referenced, along with good operational stability. (paper)

Positive cell-free fetal DNA testing for trisomy 13 reveals confined placental mosaicism.

Science.gov (United States)

Hall, April L; Drendel, Holli M; Verbrugge, Jennifer L; Reese, Angela M; Schumacher, Katherine L; Griffith, Christopher B; Weaver, David D; Abernathy, Mary P; Litton, Christian G; Vance, Gail H

2013-09-01

We report on a case in which cell-free fetal DNA was positive for trisomy 13 most likely due to confined placental mosaicism. Cell-free fetal DNA testing analyzes DNA derived from placental trophoblast cells and can lead to incorrect results that are not representative of the fetus. We sought to confirm commercial cell-free fetal DNA testing results by chorionic villus sampling and amniocentesis. These results were followed up by postnatal chromosome analysis of cord blood and placental tissue. First-trimester cell-free fetal DNA test results were positive for trisomy 13. Cytogenetic analysis of chorionic villus sampling yielded a mosaic karyotype of 47,XY,+13[10]/46,XY[12]. G-banded analysis of amniotic fluid was normal, 46,XY. Postnatal cytogenetic analysis of cord blood was normal. Karyotyping of tissues from four quadrants of the placenta demonstrated mosaicism for trisomy 13 in two of the quadrants and a normal karyotype in the other two. Our case illustrates several important aspects of this new testing methodology: that cell-free fetal DNA may not be representative of the fetal karyotype; that follow-up with diagnostic testing of chorionic villus sampling and/or amniotic fluid for abnormal test results should be performed; and that pretest counseling regarding the full benefits, limitations, and possible testing outcomes of cell-free fetal DNA screening is important.

Selectivity of Pinus sylvestris extract and essential oil to estrogen-insensitive breast cancer cells Pinus sylvestris against cancer cells.

Science.gov (United States)

Hoai, Nguyen Thi; Duc, Ho Viet; Thao, Do Thi; Orav, Anne; Raal, Ain

2015-10-01

So far, the anticancer action of pine tree extracts has mainly been shown for the species distributed widely around the Asian countries. Therefore, this study was performed to examine the potential cytotoxicity of Scots pine (Pinus sylvestris L.) native also to the European region and growing widely in Estonia. The cytotoxic activity of methanol extract and essential oil of Scots pine needles was determined by sulforhodamine B assay in different human cancer cell lines. This needle extract was found to suppress the viability of several human cancer cell lines showing some selectivity to estrogen receptor negative breast cancer cells, MDA-MB-231(half maximal inhibitory concentration [IC50] 35 μg/ml) in comparison with estrogen receptor-positive breast cancer cells, MCF-7 (IC50 86 μg/ml). It is the strongest cytotoxic effect at all measured, thus far for the needles and leaves extracts derived from various pine species, and is also the first study comparing the anticancer effects of pine tree extracts on molecularly different human breast cancer cells. The essential oil showed the stronger cytotoxic effect to both negative and positive breast cancer cell lines (both IC50 29 μg/ml) than pine extract (IC50 42 and 80 μg/ml, respectively). The data from this report indicate that Scots pine needles extract and essential oil exhibits some potential as chemopreventive or chemotherapeutic agent for mammary tumors unresponsive to endocrine treatment.

Analysis of the Interactions of Botanical Extract Combinations Against the Viability of Prostate Cancer Cell Lines

Directory of Open Access Journals (Sweden)

Lynn S. Adams

2006-01-01

Full Text Available Herbal medicines are often combinations of botanical extracts that are assumed to have additive or synergistic effects. The purpose of this investigation was to compare the effect of individual botanical extracts with combinations of extracts on prostate cell viability. We then modeled the interactions between botanical extracts in combination isobolographically. Scutellaria baicalensis, Rabdosia rubescens, Panax-pseudo ginseng, Dendranthema morifolium, Glycyrrhiza uralensis and Serenoa repens were collected, taxonomically identified and extracts prepared. Effects of the extracts on cell viability were quantitated in prostate cell lines using a luminescent ATP cell viability assay. Combinations of two botanical extracts of the four most active extracts were tested in the 22Rv1 cell line and their interactions assessed using isobolographic analysis. Each extract significantly inhibited the proliferation of prostate cell lines in a time- and dose-dependent manner except repens. The most active extracts, baicalensis, D. morifolium, G. uralensis and R. rubescens were tested as two-extract combinations. baicalensis and D. morifolium when combined were additive with a trend toward synergy, whereas D. morifolium and R. rubescens together were additive. The remaining two-extract combinations showed antagonism. The four extracts together were significantly more effective than the two-by-two combinations and the individual extracts alone. Combining the four herbal extracts significantly enhanced their activity in the cell lines tested compared with extracts alone. The less predictable nature of the two-way combinations suggests a need for careful characterization of the effects of each individual herb based on their intended use.

Differentiation of oligodendrocyte progenitor cells from dissociated monolayer and feeder-free cultured pluripotent stem cells.

Science.gov (United States)

Yamashita, Tomoko; Miyamoto, Yuki; Bando, Yoshio; Ono, Takashi; Kobayashi, Sakurako; Doi, Ayano; Araki, Toshihiro; Kato, Yosuke; Shirakawa, Takayuki; Suzuki, Yutaka; Yamauchi, Junji; Yoshida, Shigetaka; Sato, Naoya

2017-01-01

Oligodendrocytes myelinate axons and form myelin sheaths in the central nervous system. The development of therapies for demyelinating diseases, including multiple sclerosis and leukodystrophies, is a challenge because the pathogenic mechanisms of disease remain poorly understood. Primate pluripotent stem cell-derived oligodendrocytes are expected to help elucidate the molecular pathogenesis of these diseases. Oligodendrocytes have been successfully differentiated from human pluripotent stem cells. However, it is challenging to prepare large amounts of oligodendrocytes over a short amount of time because of manipulation difficulties under conventional primate pluripotent stem cell culture methods. We developed a proprietary dissociated monolayer and feeder-free culture system to handle pluripotent stem cell cultures. Because the dissociated monolayer and feeder-free culture system improves the quality and growth of primate pluripotent stem cells, these cells could potentially be differentiated into any desired functional cells and consistently cultured in large-scale conditions. In the current study, oligodendrocyte progenitor cells and mature oligodendrocytes were generated within three months from monkey embryonic stem cells. The embryonic stem cell-derived oligodendrocytes exhibited in vitro myelinogenic potency with rat dorsal root ganglion neurons. Additionally, the transplanted oligodendrocyte progenitor cells differentiated into myelin basic protein-positive mature oligodendrocytes in the mouse corpus callosum. This preparative method was used for human induced pluripotent stem cells, which were also successfully differentiated into oligodendrocyte progenitor cells and mature oligodendrocytes that were capable of myelinating rat dorsal root ganglion neurons. Moreover, it was possible to freeze, thaw, and successfully re-culture the differentiating cells. These results showed that embryonic stem cells and human induced pluripotent stem cells maintained in a

Effects of Curcuma longa Extract on Telomerase Activity in Lung and Breast Cancer Cells

Directory of Open Access Journals (Sweden)

Nosratollah Zarghami

2014-10-01

Full Text Available Background: The purpose of this study is to evaluate the effect of Curcuma longa extract on the telomerase gene expression in QU-DB lung cancer and T47D breast cancer cell lines. Materials and Methods: The present study is an experimental research. Using 3 different phases n-hexane, dichloromethane and methanol, total extract of Curcuma longa in a serial dilution was prepared and three phases was analyzed for determining which phase has more curcuminoids. Then the extract cytotoxicity effect was tested on breast cancer cell line (T47D, and lung cancer cell line (QU-DB by 24, 48 and 72 h MTT (Dimethyl thiazolyl diphenyl tetrazolium assay. Then, the cells were treated with serial concentrations of the extract. Finally, total protein was extracted from the control and test groups, its quantity was determined and telomeric repeat amplification protocol (TRAP assay was performed for measurement of possible inhibition of the telomerase activity. Results: Cell viability and MTT-based cytotoxicity assay show that the total extract of Curcuma longa has cytotoxic effect with different IC50s in breast and lung cancer cell lines. Analysis of TRAP assay also shows a significant reduction in telomerase activity on both cancer cells with different levels. Conclusion: Curcuma longa extract has anti-proliferation and telomerase inhibitory effects on QU-DB lung cancer and T47D breast cancer cells with differences in levels of telomerase inhibition.

CYTOTOXIC AND ANTIOXIDANT ACTIVITY OF BUCKWHEAT HULL EXTRACTS

Directory of Open Access Journals (Sweden)

Martina Danihelová

2013-02-01

Full Text Available Buckwheat contains many prophylactic compounds that are concentrated mainly in outer layers of buckwheat grain. The aim of this study was to prepare buckwheat hull extracts. Ten buckwheat cultivars were screened for their antioxidant and anticancer properties. Total polyphenol content was determined using Folin-Ciocalteau's reagent. Antioxidant activity was established by the method of binding free radical DPPH. Cytotoxic properties were measured on human cervical cancer cells HeLa using mitochondrial cytotoxic test (MTT. Total polyphenol content ranged from 166.67 to 635.31 mg GAE/100 g DW. The highest content displayed tartary buckwheat cultivar Madawaska (0.64% of hulls weight. Among common buckwheat the richest in polyphenols were cultivars Bamby and KASHO-2. The best free radical binding antioxidant activity was found for cultivars with highest polyphenol content. This relationship was not observed for cytotoxic action on human cervical cancer cells. The best growth inhibitory activity on HeLa cancer cells displayed common buckwheat cultivars Bamby and KASHO-2 (up to 50%, extract concentration 100 µg/ml. This was not found for tartary buckwheat cultivar Madawaska.

Effects of Ligusticum porteri (Osha) Root Extract on Human Promyelocytic Leukemia Cells

OpenAIRE

Nguyen, Khanh; Sparks, Jean; Omoruyi, Felix

2017-01-01

Background: Ligusticum porteri roots have been traditionally used in folk medicine, but the scientific basis is unclear. Objective: To investigate the cytotoxicity, antioxidant, and immunomodulatory effects of L. porteri root extract on human promyelocytic leukemia (HL-60) cells and H2O2-induced oxidative damaged HL-60 cells. Materials and Methods: HL-60 cells were incubated with different concentrations of root extract, and cells were harvested for viability assays on day 3 and 7. Cytokine l...

The Effect of Salvia Rhytidea Extract on the Number of Cells of Different Layers of Cerebellar Cortex Following Ischemia Reperfusion in Rats

Directory of Open Access Journals (Sweden)

M Farahmand

2016-09-01

Full Text Available Background & aim: Salvia has anti-oxidant oxygen free radicals which are generated during the interruption and reestablishment of ischemia reperfusion.  The aim of study was to investigate the effect of Salvia Rhytidea extract on the number of cells of different layers of cerebellar cortex following ischemia reperfusion in rats. Methods: In the present experimental study, 35 adult male rats were randomly divided into 7 groups of 5: Group 1 (control-: Sampling without ischemia. Group 2 (control +: Cerebellar ischemia with administration of normal saline. Group 3(sham: Manipulation without ischemia with normal saline administration. Group 4   received (3.2 mg/kg aqueous and alcoholic Salvia extract 2 hours after ischemia. Group 5 received 50 mg/kg silymarin drug, 2 hours after ischemia. Group 6 received 3.2 mg/kg aqueous and alcoholic Salvia extract 72, 48, 24 and 0 h before ischemia and group 7 received silymarin drug (50 mg/kg, 0, 24, 48, and 72, hrs. before ischemia. 24 hrs. following reperfusion, the rats were euthanized and samples of the cerebellum were obtained. By using routine histological technique, the sections were stained by H&E. The measurement of cell count in cerebellar cortex were accomplished. Data were evaluated with One-Way ANOVA and Tukey diagnostic tests. Results: A significant decrease was observed in the number of neural cells in granular layer in the non-treated ischemia group and in the groups which received Salvia extract and silymarin, two hours after the ischemia (p< 0.05. No significant decrease was observed in the number of cells of this layer in the groups which received salvia extract before ischemia. But regarding the cell number of molecular and purkinje layers in above groups, no significant difference was observed compared to the control group (P˃0.05. However, no significant differences was seen in the number of cells layers compared to the control group (P˃0.05. Conclusion: Finally, administration of

Artemesia annua extract prevents glyoxal-induced cell injury in ...

African Journals Online (AJOL)

Purpose: To investigate the effect of Artemesia annua extract on glyoxal-induced injury in retinal microvascular endothelial cells (HRECs). Methods: HRECs were cultured in a medium containing 500 μM glyoxal or glyoxal plus 50μM Artemesia annua extract, or in the medium alone for 24 h. Apoptosis was analysed by flow ...

Direct quantification of cell-free, circulating DNA from unpurified plasma.

Science.gov (United States)

Breitbach, Sarah; Tug, Suzan; Helmig, Susanne; Zahn, Daniela; Kubiak, Thomas; Michal, Matthias; Gori, Tommaso; Ehlert, Tobias; Beiter, Thomas; Simon, Perikles

2014-01-01

Cell-free DNA (cfDNA) in body tissues or fluids is extensively investigated in clinical medicine and other research fields. In this article we provide a direct quantitative real-time PCR (qPCR) as a sensitive tool for the measurement of cfDNA from plasma without previous DNA extraction, which is known to be accompanied by a reduction of DNA yield. The primer sets were designed to amplify a 90 and 222 bp multi-locus L1PA2 sequence. In the first module, cfDNA concentrations in unpurified plasma were compared to cfDNA concentrations in the eluate and the flow-through of the QIAamp DNA Blood Mini Kit and in the eluate of a phenol-chloroform isoamyl (PCI) based DNA extraction, to elucidate the DNA losses during extraction. The analyses revealed 2.79-fold higher cfDNA concentrations in unpurified plasma compared to the eluate of the QIAamp DNA Blood Mini Kit, while 36.7% of the total cfDNA were found in the flow-through. The PCI procedure only performed well on samples with high cfDNA concentrations, showing 87.4% of the concentrations measured in plasma. The DNA integrity strongly depended on the sample treatment. Further qualitative analyses indicated differing fractions of cfDNA fragment lengths in the eluate of both extraction methods. In the second module, cfDNA concentrations in the plasma of 74 coronary heart disease patients were compared to cfDNA concentrations of 74 healthy controls, using the direct L1PA2 qPCR for cfDNA quantification. The patient collective showed significantly higher cfDNA levels (mean (SD) 20.1 (23.8) ng/ml; range 5.1-183.0 ng/ml) compared to the healthy controls (9.7 (4.2) ng/ml; range 1.6-23.7 ng/ml). With our direct qPCR, we recommend a simple, economic and sensitive procedure for the quantification of cfDNA concentrations from plasma that might find broad applicability, if cfDNA became an established marker in the assessment of pathophysiological conditions.

Comparing handheld and hands-free cell phone usage behaviors while driving.

Science.gov (United States)

Soccolich, Susan A; Fitch, Gregory M; Perez, Miguel A; Hanowski, Richard J

2014-01-01

The goal of this study was to compare cell phone usage behaviors while driving across 3 types of cell phones: handheld (HH) cell phones, portable hands-free (PHF) cell phones, and integrated hands-free (IHF) cell phones. Naturalistic driving data were used to observe HH, PHF, and IHF usage behaviors in participants' own vehicles without any instructions or manipulations by researchers. In addition to naturalistic driving data, drivers provided their personal cell phone call records. Calls during driving were sampled and observed in naturalistically collected video. Calls were reviewed to identify cell phone type used for, and duration of, cell phone subtasks, non-cell phone secondary tasks, and other use behaviors. Drivers in the study self-identified as HH, PHF, or IHF users if they reported using that cell phone type at least 50% of the time. However, each sampled call was classified as HH, PHF, or IHF if the talking/listening subtask was conducted using that cell phone type, without considering the driver's self-reported group. Drivers with PHF or IHF systems also used HH cell phones (IHF group used HH cell phone in 53.2% of the interactions, PHF group used HH cell phone for 55.5% of interactions). Talking/listening on a PHF phone or an IHF phone was significantly longer than talking/listening on an HH phone (P phone call task for HH phones was significantly longer in duration than the end phone call task for PHF and IHF phones. Of all the non-cell phone-related secondary tasks, eating or drinking was found to occur significantly more often during IHF subtasks (0.58%) than in HH subtasks (0.15%). Drivers observed to reach for their cell phone mostly kept their cell phone in the cup holder (36.3%) or in their seat or lap (29.0% of interactions); however, some observed locations may have required drivers to move out of position. Hands-free cell phone technologies reduce the duration of cell phone visual-manual tasks compared to handheld cell phones. However

Protection of ionizing radiation-induced cytogenetic damage by hydroalcoholic extract of Cynodon dactylon in Chinese hamster lung fibroblast cells and human peripheral blood lymphocytes.

Science.gov (United States)

Rao, Bola Sadashiva Satish; Upadhya, Dinesh; Adiga, Satish Kumar

2008-01-01

The radiomodulatory potential of hydroalcoholic extract of a medicinal plant Cynodon dactylon (family: Poaceae) against radiation-induced cytogenetic damage was analyzed using Chinese hamster lung fibroblast (V79) cells and human peripheral blood lymphocytes (HPBLs) growing in vitro. Induction of micronuclei was used as an index of cytogenetic damage, evaluated in cytokinesis blocked binucleate cells. The hydroalcoholic Cynodon dactylon extract (CDE) rendered protection against the radiation-induced DNA damage, as evidenced by the significant (p<0.001) reduction in micronucleated binucleate cells (MNBNC%) after various doses of CDE treatment in V79 cells and HPBLs. The optimum dose of CDE (40 and 50 microg/ml in HPBLs and V79 cells, respectively) with the greatest reduction in micronuclei was further used in combination with various doses of gamma radiation (0.5, 1, 2, 3, and 4 Gy) exposed 1 h after CDE treatment. A linear dose-dependent MNBNC% increase in radiation alone group was observed, while 40/50 microg/ml CDE significantly resulted in the reduction of MNBNC%, compared to the respective radiation alone groups. CDE resulted in a dose-dependent increase in free radical scavenging ability against various free radicals, viz., 2, 2-diphenyl-2-picryl-hydrazyl (DPPH); 2, 2-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS); superoxide anion (O2*-); hydroxyl radical (OH*) and nitric oxide radical (NO*) generated in vitro. Also, an excellent (70%) inhibition of lipid peroxidation in vitro was observed at a dose of 300 microg/ml CDE, attaining the saturation point at higher doses. The present findings demonstrated the radioprotective effect of CDE, also rendering protection against radiation-induced genomic instability and DNA damage. The observed radioprotective effect may be partly attributed to the free radical scavenging and antilipid peroxidative potential of CDE.

Extracts of human atherosclerotic lesions modify LDL inducing enhanced macrophage uptake

International Nuclear Information System (INIS)

Hoff, H.F.; O'Neill, J.

1986-01-01

Both an LDL-like fraction isolated from human aortic plaques and LDL incubated with cultured aortic endothelial or smooth muscle cells have been shown to be internalized by macrophages in vitro in an unregulated fashion leading to foam cell formation. Lipid peroxidation induced by free radicals released from cells was shown to be responsible for cell-modified LDL. The authors incubated LDL with a supernatant fraction of leached, i.e. non-homogenized, extracts of aortic plaques for one hour at 37 0 C, to determine whether extracellular components present in arteries were also capable of modifying LDL. Extract-treated LDL showed the following changes relative to untreated LDL: 1) increased electrophretic mobility, 2) altered pattern of B-100 on SDS-PAGE, i.e. presence of a doublet with higher M/sub r/ than B-100, and 3) enhanced uptake by cultured mouse peritoneal macrophages as measured by increased degradation of 125 I-LDL, and increased stimulation of cholesterol esterification using 14 C-oleate. Extracts from homogenized plaques and grossly normal intima induced similar changes. The modification was tissue specific in that extracts of arteries but not of liver, muscle or skin modified LDL. Protease degradation of LDL during incubation was probably not responsible since inhibitors did not prevent modification. It is possible that products of lipid peroxidation present in extracellular lipid of arteries may propagate free radicals or be incorporated into LDL, leading to modifications similar to those found in cell-modified LDL

21 CFR 182.40 - Natural extractives (solvent-free) used in conjunction with spices, seasonings, and flavorings.

Science.gov (United States)

2010-04-01

... conjunction with spices, seasonings, and flavorings. 182.40 Section 182.40 Food and Drugs FOOD AND DRUG... in conjunction with spices, seasonings, and flavorings. Natural extractives (solvent-free) used in conjunction with spices, seasonings, and flavorings that are generally recognized as safe for their intended...

21 CFR 582.40 - Natural extractives (solvent-free) used in conjunction with spices, seasonings, and flavorings.

Science.gov (United States)

2010-04-01

... conjunction with spices, seasonings, and flavorings. 582.40 Section 582.40 Food and Drugs FOOD AND DRUG... in conjunction with spices, seasonings, and flavorings. Natural extractives (solvent-free) used in conjunction with spices, seasonings, and flavorings that are generally recognized as safe for their intended...

Investigation of cell-free DNA in canine plasma and its relation to disease.

Science.gov (United States)

Burnett, Deborah L; Cave, Nicholas J; Gedye, Kristene R; Bridges, Janis P

2016-09-01

DNA is released from dying cells during apoptosis and necrosis. This cell-free DNA (cfDNA) diffuses into the plasma where it can be measured. In humans, an increase in cfDNA correlates with disease severity and prognosis. It was hypothesized that when DNA in canine plasma was measured by emission fluorometry without prior DNA extraction, the concentration of cfDNA would increase with disease severity. The diseased population consisted of 97 client-owned dogs. The clinically normal population consisted of nine client-owned dogs presenting for 'wellness screens', and 15 colony-owned Harrier Hounds. Plasma cfDNA was measured by fluorometry without prior DNA extraction. The effects of ex vivo storage conditions were evaluated in plasma from two clinically normal dogs. In all other dogs, plasma was separated within two hours of collection. The association between the cfDNA concentration in hospitalized dogs and a variety of clinical, clinicopathological and outcome variables was tested. The concentration of cfDNA was reliably measured when plasma was separated within two hours of blood collection. The diseased dogs had significantly higher cfDNA than clinically normal dogs (P Dogs that did not survive to discharge had significantly higher cfDNA concentrations than survivors (P = 0.02). Conclusions/Clinical Importance: The concentration of cfDNA in the plasma of diseased dogs is associated with disease severity and prognosis. Measurement of canine cfDNA could be a useful non-specific disease indicator and prognostic tool.

Advanced feeder-free generation of induced pluripotent stem cells directly from blood cells.

Science.gov (United States)

Trokovic, Ras; Weltner, Jere; Nishimura, Ken; Ohtaka, Manami; Nakanishi, Mahito; Salomaa, Veikko; Jalanko, Anu; Otonkoski, Timo; Kyttälä, Aija

2014-12-01

Generation of validated human induced pluripotent stem cells (iPSCs) for biobanking is essential for exploring the full potential of iPSCs in disease modeling and drug discovery. Peripheral blood mononuclear cells (PBMCs) are attractive targets for reprogramming, because blood is collected by a routine clinical procedure and is a commonly stored material in biobanks. Generation of iPSCs from blood cells has previously been reported using integrative retroviruses, episomal Sendai viruses, and DNA plasmids. However, most of the published protocols require expansion and/or activation of a specific cell population from PBMCs. We have recently collected a PBMC cohort from the Finnish population containing more than 2,000 subjects. Here we report efficient generation of iPSCs directly from PBMCs in feeder-free conditions in approximately 2 weeks. The produced iPSC clones are pluripotent and transgene-free. Together, these properties make this novel method a powerful tool for large-scale reprogramming of PBMCs and for iPSC biobanking. ©AlphaMed Press.

Cell-free transfer of sterols by plant fractions

International Nuclear Information System (INIS)

Morre, D.J.; Wilkinson, F.E.; Morre, D.M.; Moreau, P.; Sandelius, A.S.; Penel, C.; Greppin, H.

1990-01-01

Microsomes from etiolated hypocotyls of soybean or leaves of light-grown spinach radiolabeled in vivo with [ 3 H]acetate or in vitro with [ 3 H]squalene or [ 3 H]cholesterol as donor transferred radioactivity to unlabeled acceptor membranes immobilized on nitrocellulose. Most efficient transfer was with plasma membrane or tonoplast as the acceptor. The latter were highly purified by aqueous two-phase partition (plasma membrane) and preparative free-flow electrophoresis (tonoplast and plasma membrane). Plasma membrane- and tonoplast-free microsomes and purified mitochondria were less efficient acceptors. Sterol transfer was verified by thin-layer chromatography of extracted lipids. Transfer was time- and temperature-dependent, required ATP but was not promoted by cytosol. The nature of the donor (endoplasmic reticulum, Golgi apparatus or both) and of the transfer mechanism is under investigation

Influence of Porcine Intervertebral Disc Matrix on Stem Cell Differentiation

Directory of Open Access Journals (Sweden)

Hans-Lothar Fuchsbauer

2011-08-01

Full Text Available For back disorders, cell therapy is one approach for a real regeneration of a degenerated nucleus pulposus. Human mesenchymal stem cells (hMSC could be differentiated into nucleus pulposus (NP-like cells and used for cell therapy. Therefore it is necessary to find a suitable biocompatible matrix, which supports differentiation. It could be shown that a differentiation of hMSC in a microbial transglutaminase cross-linked gelatin matrix is possible, but resulted in a more chondrocyte-like cell type. The addition of porcine NP extract to the gelatin matrix caused a differentiation closer to the desired NP cell phenotype. This concludes that a hydrogel containing NP extract without any other supplements could be suitable for differentiation of hMSCs into NP cells. The NP extract itself can be cross-linked by transglutaminase to build a hydrogel free of NP atypical substrates. As shown by side-specific biotinylation, the NP extract contains molecules with free glutamine and lysine residues available for the transglutaminase.

Effects of Puerariae Radix extract on the activity of antioxidant

Directory of Open Access Journals (Sweden)

Young-Joon Eun

2007-12-01

Full Text Available Objective : The objective of this study was to investigate the antioxidative effects of Puerariae Radix extract. Method : Total antioxidant capacity (TAC, Total antioxidant response (TAR, Total phenolic content, Reactive oxygen species (ROS, 1,1-Diphenyl-2-picrylhydrazyl (DPPH free radical scavenging activities, lipid peroxidation were examined. Result : Total antioxidant status was examined by total antioxidant capacity(TAC and total antioxidant response(TAR against potent free radical reactions. TAC and TAR of Puerariae Radix extract at the concentration of 5 mg/㎖ were 2.02 and 1.50 mM Trolox equivalents, respectively. Total phenolic content of Puerariae Radix extract at the concentration of 5mg/㎖ was 2.29 mM gallic acid equivalent. Concentration of Puerariae Radix extract at which DPPH radical scavenging activity was inhibited by 50% was 5.91 mg/㎖ as compared to 100% by pyrogallol solution as a reference. The inhibitory effect of the extract on lipid peroxidation was examined using rat liver mitochondria induced by FeSO4/ascorbic acid. Puerariae Radix extract at the concentration of 1 mg/㎖ slightly but significantly decreased TBARS concentration. The extract further prevented lipid peroxidation in a dose-dependent manner. The effect of Puerariae Radix extract on reactive oxygen species (ROS generation was examined using cell-free system induced by hydrogen peroxide/FeSO4. Addition of 1 mg/㎖ of Puerariae Radix extract significantly reduced dichloroflurescein (DCF fluorescence. The extract caused concentration-dependent attenuation of the increase in DCF fluorescence, indicating that the extract significantly prevented ROS generation in vitro. Thus antioxidant effects of Puerariae Radix extract seem to be due to, at least in part, the prevention from free radicals-induced oxidation, followed by inhibition of lipid peroxidation. Conclusion : As a result, Puerariae Radix seems to have antioxitative effect and antioxidant compount.

Progress on lipid extraction from wet algal biomass for biodiesel production.

Science.gov (United States)

Ghasemi Naghdi, Forough; González González, Lina M; Chan, William; Schenk, Peer M

2016-11-01

Lipid recovery and purification from microalgal cells continues to be a significant bottleneck in biodiesel production due to high costs involved and a high energy demand. Therefore, there is a considerable necessity to develop an extraction method which meets the essential requirements of being safe, cost-effective, robust, efficient, selective, environmentally friendly, feasible for large-scale production and free of product contamination. The use of wet concentrated algal biomass as a feedstock for oil extraction is especially desirable as it would avoid the requirement for further concentration and/or drying. This would save considerable costs and circumvent at least two lengthy processes during algae-based oil production. This article provides an overview on recent progress that has been made on the extraction of lipids from wet algal biomass. The biggest contributing factors appear to be the composition of algal cell walls, pre-treatments of biomass and the use of solvents (e.g. a solvent mixture or solvent-free lipid extraction). We compare recently developed wet extraction processes for oleaginous microalgae and make recommendations towards future research to improve lipid extraction from wet algal biomass. © 2016 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

Cancer-suppressive potential of extracts of endemic plant Helichrysum zivojinii: effects on cell migration, invasion and angiogenesis.

Science.gov (United States)

Matić, Ivana Z; Aljancić, Ivana; Vajs, Vlatka; Jadranin, Milka; Gligorijević, Nevenka; Milosavljević, Slobodan; Juranić, Zorica D

2013-09-01

Helichrysum zivojinii Cernjavski & Soska is an endemic plant species that grows in the National Park Galicica in Macedonia. Five extracts were isolated as fractions from the aerial parts of the plant: a n-hexane extract (1), a dichloromethane extract (2), an ethyl-acetate extract (3), a n-butanol extract (4) and a methanol extract (5). A dose-dependent cytotoxic activity of the extracts on MDA-MB-231 and EA.hy926 cells was observed. Extracts exhibited more pronounced cytotoxic actions on MDA-MB-231 cells than on EA.hy926 cells. The n-hexane extract (1), at a non-toxic concentration, exhibited an inhibitory effect on the migration as well the invasiveness of MDA-MB-231 cells. The dichloromethane extract (2), at a non-toxic concentration, demonstrated inhibition of MDA-MB-231 cells invasion. Each of the five extracts applied at non-toxic concentrations inhibited migration of EA.hy926 cells. The prominent inhibitory effect of the n-hexane extract on EA.hy926 cells migration was associated with a notable anti-angiogenic action of this extract. The other four tested extracts demonstrated mild anti-angiogenic activity. Our data highlight the prominent anticancer potential of n-hexane (1) and dichloromethane (2) extracts, which could be attributed to their very pronounced and selective cytotoxic activities as well as their anti-invasive and anti-angiogenic properties.

Remodeling of ribosomal genes in somatic cells by Xenopus egg extract

Energy Technology Data Exchange (ETDEWEB)

Ostrup, Olga, E-mail: osvarcova@gmail.com [Institute of Basic Animal and Veterinary Sciences, Faculty of Life Sciences, University of Copenhagen, Frederiksberg C (Denmark); Stem Cell Epigenetics Laboratory, Institute of Basic Medical Sciences, Faculty of Medicine, University of Oslo, Oslo (Norway); Norwegian Center for Stem Cell Research, Oslo (Norway); Hyttel, Poul; Klaerke, Dan A. [Institute of Basic Animal and Veterinary Sciences, Faculty of Life Sciences, University of Copenhagen, Frederiksberg C (Denmark); Collas, Philippe, E-mail: philc@medisin.uio.no [Stem Cell Epigenetics Laboratory, Institute of Basic Medical Sciences, Faculty of Medicine, University of Oslo, Oslo (Norway); Norwegian Center for Stem Cell Research, Oslo (Norway)

2011-09-02

Highlights: {yields} Xenopus egg extract remodels nuclei and alter cell growth characteristics. {yields} Ribosomal genes are reprogrammed within 6 h after extract exposure. {yields} rDNA reprogramming involves promoter targeting of SNF2H remodeling complex. {yields} Xenopus egg extract does not initiate stress-related response in somatic cells. {yields} Aza-cytidine elicits a stress-induced response in reprogrammed cells. -- Abstract: Extracts from Xenopus eggs can reprogram gene expression in somatic nuclei, however little is known about the earliest processes associated with the switch in the transcriptional program. We show here that an early reprogramming event is the remodeling of ribosomal chromatin and gene expression. This occurs within hours of extract treatment and is distinct from a stress response. Egg extract elicits remodeling of the nuclear envelope, chromatin and nucleolus. Nucleolar remodeling involves a rapid and stable decrease in ribosomal gene transcription, and promoter targeting of the nucleolar remodeling complex component SNF2H without affecting occupancy of the transcription factor UBF and the stress silencers SUV39H1 and SIRT1. During this process, nucleolar localization of UBF and SIRT1 is not altered. On contrary, azacytidine pre-treatment has an adverse effect on rDNA remodeling induced by extract and elicits a stress-type nuclear response. Thus, an early event of Xenopus egg extract-mediated nuclear reprogramming is the remodeling of ribosomal genes involving nucleolar remodeling complex. Condition-specific and rapid silencing of ribosomal genes may serve as a sensitive marker for evaluation of various reprogramming methods.

Cellular toxicity of calf blood extract on human corneal epithelial cells in vitro.

Science.gov (United States)

Park, Young Min; Kim, Su Jin; Han, Young Sang; Lee, Jong Soo

2015-01-01

To investigate the biologic effects of the calf blood extract on corneal epithelial cells in vitro. The effects on corneal epithelial cells were evaluated after 1, 4, 12, and 24 h of exposure to various concentrations of calf blood extract (3, 5, 8 and 16%). The MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) assay was performed to measure levels of cellular metabolic activity. The lactate dehydrogenase (LDH) assay was performed to determine the extent of cellular damage. Cellular morphology was examined using phase-contrast microscopy. The scratch wound assay was performed to quantify the migration of corneal epithelial cells. At the 3 and 5% concentrations of calf blood extract, MTT values were similar to those observed in the control group. However, at a concentration of 8 and 16%, cellular metabolic activity was significantly decreased after 4 h of exposure to calf blood extract. After 12 h of exposure to 8 and 16% concentrations of calf blood extract, LDH activity and cellular morphological damage to the corneal epithelial cells were significantly increased. There was no evidence of cellular migration after 12 h exposure to 5% or higher concentration of calf blood extract because of cellular toxicity. Compared with normal corneal epithelial cells, the cellular activity was decreased, and toxicity was increased after over 12 h of exposure to more than 5% concentration of calf blood extract. Further clinical studies will be necessary to determine the optimal concentration and exposure time for the topical application of eye drops containing calf blood extract.

SurfCut: Free-Boundary Surface Extraction

KAUST Repository

Algarni, Marei Saeed Mohammed; Sundaramoorthi, Ganesh

2016-01-01

We present SurfCut, an algorithm for extracting a smooth simple surface with unknown boundary from a noisy 3D image and a seed point. In contrast to existing approaches that extract smooth simple surfaces with boundary, our method requires less user

Cranberry and Grape Seed Extracts Inhibit the Proliferative Phenotype of Oral Squamous Cell Carcinomas

Directory of Open Access Journals (Sweden)

Kourt Chatelain

2011-01-01

Full Text Available Proanthocyanidins, compounds highly concentrated in dietary fruits, such as cranberries and grapes, demonstrate significant cancer prevention potential against many types of cancer. The objective of this study was to evaluate cranberry and grape seed extracts to quantitate and compare their anti-proliferative effects on the most common type of oral cancer, oral squamous cell carcinoma. Using two well-characterized oral squamous cell carcinoma cell lines, CAL27 and SCC25, assays were performed to evaluate the effects of cranberry and grape seed extract on phenotypic behaviors of these oral cancers. The proliferation of both oral cancer cell lines was significantly inhibited by the administration of cranberry and grape seed extracts, in a dose-dependent manner. In addition, key regulators of apoptosis, caspase-2 and caspase-8, were concomitantly up-regulated by these treatments. However, cranberry and grape seed extracts elicited differential effects on cell adhesion, cell morphology, and cell cycle regulatory pathways. This study represents one of the first comparative investigations of cranberry and grape seed extracts and their anti-proliferative effects on oral cancers. Previous findings using purified proanthocyanidin from grape seed extract demonstrated more prominent growth inhibition, as well as apoptosis-inducing, properties on CAL27 cells. These observations provide evidence that cranberry and grape seed extracts not only inhibit oral cancer proliferation but also that the mechanism of this inhibition may function by triggering key apoptotic regulators in these cell lines. This information will be of benefit to researchers interested in elucidating which dietary components are central to mechanisms involved in the mediation of oral carcinogenesis and progression.

Rapid acquisition and model-based analysis of cell-free transcription–translation reactions from nonmodel bacteria

Science.gov (United States)

Wienecke, Sarah; Ishwarbhai, Alka; Tsipa, Argyro; Aw, Rochelle; Kylilis, Nicolas; Bell, David J.; McClymont, David W.; Jensen, Kirsten; Biedendieck, Rebekka

2018-01-01

Native cell-free transcription–translation systems offer a rapid route to characterize the regulatory elements (promoters, transcription factors) for gene expression from nonmodel microbial hosts, which can be difficult to assess through traditional in vivo approaches. One such host, Bacillus megaterium, is a giant Gram-positive bacterium with potential biotechnology applications, although many of its regulatory elements remain uncharacterized. Here, we have developed a rapid automated platform for measuring and modeling in vitro cell-free reactions and have applied this to B. megaterium to quantify a range of ribosome binding site variants and previously uncharacterized endogenous constitutive and inducible promoters. To provide quantitative models for cell-free systems, we have also applied a Bayesian approach to infer ordinary differential equation model parameters by simultaneously using time-course data from multiple experimental conditions. Using this modeling framework, we were able to infer previously unknown transcription factor binding affinities and quantify the sharing of cell-free transcription–translation resources (energy, ribosomes, RNA polymerases, nucleotides, and amino acids) using a promoter competition experiment. This allows insights into resource limiting-factors in batch cell-free synthesis mode. Our combined automated and modeling platform allows for the rapid acquisition and model-based analysis of cell-free transcription–translation data from uncharacterized microbial cell hosts, as well as resource competition within cell-free systems, which potentially can be applied to a range of cell-free synthetic biology and biotechnology applications. PMID:29666238

Alpinia pricei Rhizome Extracts Induce Cell Cycle Arrest in Human Squamous Carcinoma KB Cells and Suppress Tumor Growth in Nude Mice

Directory of Open Access Journals (Sweden)

You-Cheng Hseu

2011-01-01

Full Text Available Alpinia pricei has been shown to induce apoptosis in human squamous carcinoma (KB cells. In this study, we report the effectiveness of the ethanol (70% extracts of A. pricei rhizome (AP extracts in terms of tumor regression as determined using both in vitro cell culture and in vivo athymic nude mice models of KB cells. We found that the AP extract (25–200 μg/mL treatment decreased the proliferation of KB cells by arresting progression through the G2/M phase of the cell cycle. This cell cycle blockade was associated with reductions in cyclin A and B1, Cdc2, and Cdc25C, and increased p21/WAF1, Wee1, p53 and phospho-p53 (p-p53 in a dose- and time-dependent manner. Moreover, we found that AP extract treatment decreased metalloproteinase-9 (MMP-9 and urokinase plasminogen activator (u-PA expression, while expression of their endogenous inhibitors, tissue inhibitor of MMP-1 (TIMP-1 and plasminogen activator inhibitor-1 (PAI-1, were increased in KB cells. Furthermore, AP extract treatment effectively delayed tumor incidence in nude mice inoculated with KB cells and reduced the tumor burden. AP extract treatment also induced apoptotic DNA fragmentation, as detected by in situ TUNEL staining. Thus, A. pricei may possess antitumor activity in human squamous carcinoma (KB cells.

Label-free and live cell imaging by interferometric scattering microscopy.

Science.gov (United States)

Park, Jin-Sung; Lee, Il-Buem; Moon, Hyeon-Min; Joo, Jong-Hyeon; Kim, Kyoung-Hoon; Hong, Seok-Cheol; Cho, Minhaeng

2018-03-14

Despite recent remarkable advances in microscopic techniques, it still remains very challenging to directly observe the complex structure of cytoplasmic organelles in live cells without a fluorescent label. Here we report label-free and live-cell imaging of mammalian cell, Escherischia coli , and yeast, using interferometric scattering microscopy, which reveals the underlying structures of a variety of cytoplasmic organelles as well as the underside structure of the cells. The contact areas of the cells attached onto a glass substrate, e.g. , focal adhesions and filopodia, are clearly discernible. We also found a variety of fringe-like features in the cytoplasmic area, which may reflect the folded structures of cytoplasmic organelles. We thus anticipate that the label-free interferometric scattering microscopy can be used as a powerful tool to shed interferometric light on in vivo structures and dynamics of various intracellular phenomena.

Pumpkin seed extract: Cell growth inhibition of hyperplastic and cancer cells, independent of steroid hormone receptors.

Science.gov (United States)

Medjakovic, Svjetlana; Hobiger, Stefanie; Ardjomand-Woelkart, Karin; Bucar, Franz; Jungbauer, Alois

2016-04-01

Pumpkin seeds have been known in folk medicine as remedy for kidney, bladder and prostate disorders since centuries. Nevertheless, pumpkin research provides insufficient data to back up traditional beliefs of ethnomedical practice. The bioactivity of a hydro-ethanolic extract of pumpkin seeds from the Styrian pumpkin, Cucurbita pepo L. subsp. pepo var. styriaca, was investigated. As pumpkin seed extracts are standardized to cucurbitin, this compound was also tested. Transactivational activity was evaluated for human androgen receptor, estrogen receptor and progesterone receptor with in vitro yeast assays. Cell viability tests with prostate cancer cells, breast cancer cells, colorectal adenocarcinoma cells and a hyperplastic cell line from benign prostate hyperplasia tissue were performed. As model for non-hyperplastic cells, effects on cell viability were tested with a human dermal fibroblast cell line (HDF-5). No transactivational activity was found for human androgen receptor, estrogen receptor and progesterone receptor, for both, extract and cucurbitin. A cell growth inhibition of ~40-50% was observed for all cell lines, with the exception of HDF-5, which showed with ~20% much lower cell growth inhibition. Given the receptor status of some cell lines, a steroid-hormone receptor independent growth inhibiting effect can be assumed. The cell growth inhibition for fast growing cells together with the cell growth inhibition of prostate-, breast- and colon cancer cells corroborates the ethnomedical use of pumpkin seeds for a treatment of benign prostate hyperplasia. Moreover, due to the lack of androgenic activity, pumpkin seed applications can be regarded as safe for the prostate. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Free radical scavenging and anti-edematogenic activities of Paullinia elegans Cambess., Sapindaceae, leaves extracts

Directory of Open Access Journals (Sweden)

Rodrigo N. Guimarães

Full Text Available Ethanol extract of the leaves of Paullinia elegans Cambess., Sapindaceae, and its hexane, chloroform, ethyl acetate, and hydroethanol fractions were evaluated for their antiedematogenic and free radical scavenging activities. The ethanol extract and the hexane fraction produced statistically significant inhibition (74.4 and 76.0%, respectively of the ear edema induced by croton oil in mice, observed at doses of 5 mg/ear. The ethyl acetate and hydroethanol fractions showed significant radical scavenging effect in the DPPH assay, with IC50 of 36.7 and 30.1 µg/mL, respectively. Fractionation of the extracts through chromatographic methods afforded epifriedelanol, oleanolic acid 3-O-acetyl, a mixture of stigmasterol 3-β-O-glucopyranoside and sitosterol 3-β-O-glucopyranoside, kaempferol 3,7-O-α-dirhamnopyranoside, kaempeferol-3-O-α-rhamnopyranoside and 2-O-methyl-chiro-inositol. The compounds were identified on the basis of their NMR spectral data and comparison with those of literature.

Serum-Free Media and the Immunoregulatory Properties of Mesenchymal Stem Cells In Vivo and In Vitro

Directory of Open Access Journals (Sweden)

Mei Wu

2014-02-01

Full Text Available Background: Mesenchymal stem cells are capable of self-renewal and multi-lineage differentiation. They are used extensively to treat several diseases. Traditionally, mesenchymal stem cells are cultured in serum-containing media, typically supplemented with fetal bovine serum (FBS. However, the variability of FBS is likely to skew experimental results. Although serum-free media used to expand mesenchymal stem cells has facilitated remarkable achievements, immunomodulation of these cells in under serum-free conditions is poorly understood. We hypothesized that mesenchymal stem cells expanded in serum-free media will retain powerful immunoregulatory functions in vitro and in vivo. Design and Methods: Immunosuppressive activity and the immunomodulatory cytokines produced by mesenchymal stem cells in serum-free media were characterized in vitro. Immunomodulation by serum-free mesenchymal stem cell expansion in monocrotaline-induced pulmonary hypertension was explored in vivo. Results: Similar to cells in serum-containing media, mesenchymal stem cells expanded in serum-free media inhibited proliferation and apoptosis of CD4+T cells. They also exhibited strong immunosuppressive activities and secreted high levels of immunomodulatory cytokines such as PGE2, IDO1, COX2, IL-6, and IL-1β, but not HGF. On the other hand, growth of mesenchymal stem cells in serum-free media attenuated pulmonary vascular remodeling and inhibited mRNA expression of proinflammatory cytokines TNF-α, IFN-γ, IL-6, IL-1β, and IL-18. Conclusions: Mesenchymal stem cells in serum-free media maintained powerful immunomodulatory function in vitro and in vivo; serum-free media may replace serum-containing media for basic research and clinical applications.

Population Characteristics and Seasonal Movement Patterns of the Rattlesnake Hills Elk Herd - Status Report 2000

Energy Technology Data Exchange (ETDEWEB)

Tiller, B.L.; Zufelt, R.K.; Turner, S.; Cadwell, L.L.; Bender, L.; Turner, G.K.

2000-10-10

Population characteristics of the Rattlesnake Hills elk herd indicate reduced herd growth rates from the 1980s compared to the 1990s (McCorquodale 1988; Eberhardt 1996). However, the population continued to grow approximately 25% annually through the 1990s, reaching a high of 838 animals in summer 1999. Calf recruitment rates appear to be cyclic and are likely related to reduced calf survival during the first weeks of life; however, late-term abortions may also have occurred. The cause(s) could be predator-related and/or a function of shifts in nutritional condition (age-class distributions, assuming older-age cows are less likely to recruit calves, major climate shifts) or changes in the human-related disturbances during gestation, and/or calf rearing periods. In fall 1999 and spring 2000, the population was reduced from 838 individuals to 660 individuals. The primary controlling factors were modified hunting seasons on private and state lands and the large-scale roundup conducted in spring 2000. Continued removal of animals (particularly females) within the population will be pivotal to maintain the population at a level that minimizes land damage complaints, animal-vehicle collisions, use of central Hanford areas, and deterioration of natural resources.

Ceratonia siliqua L. hydroethanolic extract obtained by ultrasonication: antioxidant activity, phenolic compounds profile and effects in yogurts functionalized with their free and microencapsulated forms.

Science.gov (United States)

Rached, Irada; Barros, Lillian; Fernandes, Isabel P; Santos-Buelga, Celestino; Rodrigues, Alírio E; Ferchichi, Ali; Barreiro, Maria Filomena; Ferreira, Isabel C F R

2016-03-01

Bioactive extracts were obtained from powdered carob pulp through an ultrasound extraction process and then evaluated in terms of antioxidant activity. Ten minutes of ultrasonication at 375 Hz were the optimal conditions leading to an extract with the highest antioxidant effects. After its chemical characterization, which revealed the preponderance of gallotannins, the extract (free and microencapsulated) was incorporated in yogurts. The microspheres were prepared using an extract/sodium alginate ratio of 100/400 (mg mg(-1)) selected after testing different ratios. The yogurts with the free extract exhibited higher antioxidant activity than the samples added with the encapsulated extracts, showing the preserving role of alginate as a coating material. None of the forms significantly altered the yogurt's nutritional value. This study confirmed the efficiency of microencapsulation to stabilize functional ingredients in food matrices maintaining almost the structural integrity of polyphenols extracted from carob pulp and furthermore improving the antioxidant potency of the final product.

Interface Engineering Based on Liquid Metal for Compact-Layer-free, Fully Printable Mesoscopic Perovskite Solar Cells.

Science.gov (United States)

Zhang, Yumin; Zhao, Jianhong; Zhang, Jin; Jiang, Xixi; Zhu, Zhongqi; Liu, Qingju

2018-05-09

A printing process for the fabrication of perovskite solar cells (PSCs) exhibits promising future application in the photovoltaic industry due to its low-cost and eco-friendly preparation. In mesoscopic carbon-based PSCs, however, compared to conventional ones, the hole-transport-layer-free PSCs often lead to inefficient hole extraction. Here, we used liquid metal (LM, Galinstan) as an interface modifier material in combination with a carbon electrode. Considering the high conductivity and room-temperature fluidity, it is found that LMs are superior in improving hole extraction and, more importantly, LMs tend to be reserved at the interface between ZrO 2 and carbon for enhancing the contact property. Correspondingly, the carrier transfer resistance was decreased at the carbon/perovskite interface. As optimized content, the triple mesoscopic PSCs based on mixed-cation perovskite with a power conversion efficiency of 13.51% was achieved, involving a 26% increase compared to those without LMs. This work opens new techniques for LMs in optoelectronics and printing.

SurfCut: Free-Boundary Surface Extraction

KAUST Repository

Algarni, Marei Saeed Mohammed

2016-09-15

We present SurfCut, an algorithm for extracting a smooth simple surface with unknown boundary from a noisy 3D image and a seed point. In contrast to existing approaches that extract smooth simple surfaces with boundary, our method requires less user input, i.e., a seed point, rather than a 3D boundary curve. Our method is built on the novel observation that certain ridge curves of a front propagated using the Fast Marching algorithm are likely to lie on the surface. Using the framework of cubical complexes, we design a novel algorithm to robustly extract such ridge curves and form the surface of interest. Our algorithm automatically cuts these ridge curves to form the surface boundary, and then extracts the surface. Experiments show the robustness of our method to errors in the data, and that we achieve higher accuracy with lower computational cost than comparable methods. © Springer International Publishing AG 2016.

Extraction of nucleic acids from yeast cells and plant tissues using ethanol as medium for sample preservation and cell disruption.

Science.gov (United States)

Linke, Bettina; Schröder, Kersten; Arter, Juliane; Gasperazzo, Tatiana; Woehlecke, Holger; Ehwald, Rudolf

2010-09-01

Here we report that dehydrated ethanol is an excellent medium for both in situ preservation of nucleic acids and cell disruption of plant and yeast cells. Cell disruption was strongly facilitated by prior dehydration of the ethanol using dehydrated zeolite. Following removal of ethanol, nucleic acids were extracted from the homogenate pellet using denaturing buffers. The method provided DNA and RNA of high yield and integrity. Whereas cell wall disruption was essential for extraction of DNA and large RNA molecules, smaller molecules such as tRNAs could be selectively extracted from undisrupted, ethanol-treated yeast cells. Our results demonstrate the utility of absolute ethanol for sample fixation, cell membrane and cell wall disruption, as well as preservation of nucleic acids during sample storage.

Progress in Electrolyte-Free Fuel Cells

International Nuclear Information System (INIS)

Lu, Yuzheng; Zhu, Bin; Cai, Yixiao; Kim, Jung-Sik; Wang, Baoyuan; Wang, Jun; Zhang, Yaoming; Li, Junjiao

2016-01-01

Solid oxide fuel cell (SOFC) represents a clean electrochemical energy conversion technology with characteristics of high conversion efficiency and low emissions. It is one of the most important new energy technologies in the future. However, the manufacture of SOFCs based on the structure of anode/electrolyte/cathode is complicated and time-consuming. Thus, the cost for the entire fabrication and technology is too high to be affordable, and challenges still hinder commercialization. Recently, a novel type of electrolyte-free fuel cell (EFFC) with single component was invented, which could be the potential candidate for the next generation of advanced fuel cells. This paper briefly introduces the EFFC, working principle, performance, and advantages with updated research progress. A number of key R&D issues about EFFCs have been addressed, and future opportunities and challenges are discussed.

Progress in Electrolyte-Free Fuel Cells

Energy Technology Data Exchange (ETDEWEB)

Lu, Yuzheng [Jiangsu Provincial Key Laboratory of Solar Energy Science and Technology, School of Energy and Environment, Southeast University, Nanjing (China); Zhu, Bin, E-mail: binzhu@kth.se [Faculty of Physics and Electronic Technology, Hubei Collaborative Innovation Center for Advanced Organic Materials, Hubei University, Wuhan (China); Department of Energy Technology, Royal Institute of Technology KTH, Stockholm (Sweden); Cai, Yixiao [Ångström Laboratory, Department of Engineering Sciences, Uppsala University, Uppsala (Sweden); Kim, Jung-Sik [Department of Aeronautical and Automotive Engineering, Loughborough University, Loughborough (United Kingdom); Wang, Baoyuan [Faculty of Physics and Electronic Technology, Hubei Collaborative Innovation Center for Advanced Organic Materials, Hubei University, Wuhan (China); Department of Energy Technology, Royal Institute of Technology KTH, Stockholm (Sweden); Wang, Jun, E-mail: binzhu@kth.se; Zhang, Yaoming [Jiangsu Provincial Key Laboratory of Solar Energy Science and Technology, School of Energy and Environment, Southeast University, Nanjing (China); Li, Junjiao [Nanjing Yunna Nano Technology Co., Ltd., Nanjing (China)

2016-05-02

Solid oxide fuel cell (SOFC) represents a clean electrochemical energy conversion technology with characteristics of high conversion efficiency and low emissions. It is one of the most important new energy technologies in the future. However, the manufacture of SOFCs based on the structure of anode/electrolyte/cathode is complicated and time-consuming. Thus, the cost for the entire fabrication and technology is too high to be affordable, and challenges still hinder commercialization. Recently, a novel type of electrolyte-free fuel cell (EFFC) with single component was invented, which could be the potential candidate for the next generation of advanced fuel cells. This paper briefly introduces the EFFC, working principle, performance, and advantages with updated research progress. A number of key R&D issues about EFFCs have been addressed, and future opportunities and challenges are discussed.

Cell-free DNA, inflammation, and the initiation of spontaneous term labor.

Science.gov (United States)

Herrera, Christina A; Stoerker, Jay; Carlquist, John; Stoddard, Gregory J; Jackson, Marc; Esplin, Sean; Rose, Nancy C

2017-11-01

Hypomethylated cell-free DNA from senescent placental trophoblasts may be involved in the activation of the inflammatory cascade to initiate labor. To determine the changes in cell-free DNA concentrations, the methylation ratio, and inflammatory markers between women in labor at term vs women without labor. In this prospective cohort study, eligible participants carried a nonanomalous singleton fetus. Women with major medical comorbidity, preterm labor, progesterone use, aneuploidy, infectious disease, vaginal bleeding, abdominal trauma, or invasive procedures during the pregnancy were excluded. Maternal blood samples were collected at 28 weeks, 36 weeks, and at admission for delivery. Total cell-free DNA concentration, methylation ratio, and interleukin-6 were analyzed. The primary outcome was the difference in methylation ratio in women with labor vs without labor. Secondary outcomes included the longitudinal changes in these biomarkers corresponding to labor status. A total of 55 women were included; 20 presented in labor on admission and 35 presented without labor. Women in labor had significantly greater methylation ratio (P = .001) and interleukin-6 (P < .001) on admission for delivery than women without labor. After we controlled for body mass index and maternal age, methylation ratio (adjusted relative risk, 1.38; 95% confidence interval, 1.13 to 1.68) and interleukin-6 (adjusted relative risk, 1.12, 95% confidence interval, 1.07 to 1.17) remained greater in women presenting in labor. Total cell-free DNA was not significantly different in women with labor compared with women without. Longitudinally, total cell-free DNA (P < .001 in labor, P = .002 without labor) and interleukin-6 (P < .001 in labor, P = .01 without labor) increased significantly across gestation in both groups. The methylation ratio increased significantly in women with labor from 36 weeks to delivery (P = .02). Spontaneous labor at term is associated with a greater cell-free DNA

Murraya koenigii leaf extract inhibits proteasome activity and induces cell death in breast cancer cells.

Science.gov (United States)

Noolu, Bindu; Ajumeera, Rajanna; Chauhan, Anitha; Nagalla, Balakrishna; Manchala, Raghunath; Ismail, Ayesha

2013-01-09

Inhibition of the proteolytic activity of 26S proteasome, the protein-degrading machine, is now considered a novel and promising approach for cancer therapy. Interestingly, proteasome inhibitors have been demonstrated to selectively kill cancer cells and also enhance the sensitivity of tumor cells to chemotherapeutic agents. Recently, polyphenols/flavonoids have been reported to inhibit proteasome activity. Murraya koenigii Spreng, a medicinally important herb of Indian origin, has been used for centuries in the Ayurvedic system of medicine. Here we show that Murraya koenigii leaves (curry leaves), a rich source of polyphenols, inhibit the proteolytic activity of the cancer cell proteasome, and cause cell death. Hydro-methanolic extract of curry leaves (CLE) was prepared and its total phenolic content [TPC] determined by, the Folin-Ciocalteau's method. Two human breast carcinoma cell lines: MCF-7 and MDA-MB-231 and a normal human lung fibroblast cell line, WI-38 were used for the studies. Cytotoxicity of the CLE was assessed by the MTT assay. We studied the effect of CLE on growth kinetics using colony formation assay. Growth arrest was assessed by cell cycle analysis and apoptosis by Annexin-V binding using flow cytometry. Inhibition of the endogenous 26S proteasome was studied in intact cells and cell extracts using substrates specific to 20S proteasomal enzymes. CLE decreased cell viability and altered the growth kinetics in both the breast cancer cell lines in a dose-dependent manner. It showed a significant arrest of cells in the S phase albeit in cancer cells only. Annexin V binding data suggests that cell death was via the apoptotic pathway in both the cancer cell lines. CLE treatment significantly decreased the activity of the 26S proteasome in the cancer but not normal cells. Our study suggests M. koenigii leaves to be a potent source of proteasome inhibitors that lead to cancer cell death. Therefore, identification of active component(s) from the leaf

Investigation of the cytotoxic and antileishmanal effects of fagonia indica l. extract and extract mediated silver nanoparticles (AgNPs)

International Nuclear Information System (INIS)

Ullah, I.; Shinwari, Z. K.; Khalil, A. T.

2017-01-01

Leishmaniasis is one of the globally neglected tropical disease cause by protozoan parasite of the genus Leishmania. In Pakistan, cutaneous leishmaniasis is more sporadic in the Afghan refugee camps, which is concern for the local villager and Pakistani population. In the current study an approache was made to synthesise biogenic silver nanoparticles using Fagonia indica leaf extract. Furthermore, the antileishmanial activity of the nanoparticles was evaluated compared to the crude extracts against Leishmania tropica which is the causative agent of cutenious leishmaniasis. MTT cell viability assay was used to determine the non toxicological concentration of the extract and nanoparticle in macrophage cell lines (J774), and the antileishmanial activity. We found that silver nanoparticles are not toxic to macropage cell above 30 μg/ml. where as the IC/sub 50/ against leishmania parsites was calculated as 8.16+-0.63 μg/ml and 4.8+-0.819 μg/ml for extract and AgNPs respectively. We also determine the infection index of the parasite in the macrophage cell. The infectivity of parasites also decreases as compared to control group after activation of macrophages. We further, evaluate the mechanism of growth inhibition using Griess reagent for the estimation of nitrogen oxide. We found that both the extract and AgNPs produce an elevated level of nitrogen oxide free radical. These radical produce oxidative stress in the cell that lead to the reduced metabolic activities of the parasites and ultimate death. Overall, the results indicate that Fagonia indica leves extract and AgNPs are potent antileishmanial agents. (author)

Applications of cell-free protein synthesis in synthetic biology: Interfacing bio-machinery with synthetic environments.

Science.gov (United States)

Lee, Kyung-Ho; Kim, Dong-Myung

2013-11-01

Synthetic biology is built on the synthesis, engineering, and assembly of biological parts. Proteins are the first components considered for the construction of systems with designed biological functions because proteins carry out most of the biological functions and chemical reactions inside cells. Protein synthesis is considered to comprise the most basic levels of the hierarchical structure of synthetic biology. Cell-free protein synthesis has emerged as a powerful technology that can potentially transform the concept of bioprocesses. With the ability to harness the synthetic power of biology without many of the constraints of cell-based systems, cell-free protein synthesis enables the rapid creation of protein molecules from diverse sources of genetic information. Cell-free protein synthesis is virtually free from the intrinsic constraints of cell-based methods and offers greater flexibility in system design and manipulability of biological synthetic machinery. Among its potential applications, cell-free protein synthesis can be combined with various man-made devices for rapid functional analysis of genomic sequences. This review covers recent efforts to integrate cell-free protein synthesis with various reaction devices and analytical platforms. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Reference Proteome Extracts for Mass Spec Instrument Performance Validation and Method Development

Science.gov (United States)

Rosenblatt, Mike; Urh, Marjeta; Saveliev, Sergei

2014-01-01

Biological samples of high complexity are required to test protein mass spec sample preparation procedures and validate mass spec instrument performance. Total cell protein extracts provide the needed sample complexity. However, to be compatible with mass spec applications, such extracts should meet a number of design requirements: compatibility with LC/MS (free of detergents, etc.)high protein integrity (minimal level of protein degradation and non-biological PTMs)compatibility with common sample preparation methods such as proteolysis, PTM enrichment and mass-tag labelingLot-to-lot reproducibility Here we describe total protein extracts from yeast and human cells that meet the above criteria. Two extract formats have been developed: Intact protein extracts with primary use for sample preparation method development and optimizationPre-digested extracts (peptides) with primary use for instrument validation and performance monitoring

Investigating free radical generation in HepG2 cells using immuno-spin trapping.

Science.gov (United States)

Horinouchi, Yuya; Summers, Fiona A; Ehrenshaft, Marilyn; Kawazoe, Kazuyoshi; Tsuchiya, Koichiro; Tamaki, Toshiaki; Mason, Ronald P

2014-10-01

Oxidative stress can induce the generation of free radicals, which are believed to play an important role in both physiological and pathological processes and a number of diseases such as cancer. Therefore, it is important to identify chemicals which are capable of inducing oxidative stress. In this study, we evaluated the ability of four environmental chemicals, aniline, nitrosobenzene (NB), N,N-dimethylaniline (DMA) and N,N-dimethyl-4-nitrosoaniline (DMNA), to induce free radicals and cellular damage in the hepatoma cell line HepG2. Cytotoxicity was assessed using lactate dehydrogenase (LDH) assays and morphological changes were observed using phase contrast microscopy. Free radicals were detected by immuno-spin trapping (IST) in in-cell western experiments or in confocal microscopy experiments to determine the subcellular localization of free radical generation. DMNA induced free radical generation, LDH release and morphological changes in HepG2 cells whereas aniline, NB and DMA did not. Confocal microscopy showed that DMNA induced free radical generation mainly in the cytosol. Preincubation of HepG2 cells with N-acetylcysteine and 2,2'-dipyridyl significantly prevented free radical generation upon subsequent incubation with DMNA, whereas preincubation with apocynin and dimethyl sulfoxide did not. These results suggest that DMNA induces oxidative stress and that reactive oxygen species, metals and free radical generation play a critical role in DMNA-induced cytotoxicity. Copyright © 2014. Published by Elsevier Inc.

Catalytical Properties of Free and Immobilized Aspergillus niger Tannase

OpenAIRE

Abril Flores-Maltos; Luis V. Rodríguez-Durán; Jacqueline Renovato; Juan C. Contreras; Raúl Rodríguez; Cristóbal N. Aguilar

2011-01-01

A fungal tannase was produced, recovered, and immobilized by entrapment in calcium alginate beads. Catalytical properties of the immobilized enzyme were compared with those of the free one. Tannase was produced intracellularly by the xerophilic fungus Aspergillus niger GH1 in a submerged fermentation system. Enzyme was recovered by cell disruption and the crude extract was partially purified. The catalytical properties of free and immobilized tannase were evaluated using tannic acid and methy...

Achillea millefolium L. hydroethanolic extract inhibits growth of human tumor cell lines by interfering with cell cycle and inducing apoptosis.

Science.gov (United States)

Pereira, Joana M; Peixoto, Vanessa; Teixeira, Alexandra; Sousa, Diana; Barros, Lillian; Ferreira, Isabel C F R; Vasconcelos, M Helena

2018-06-05

The cell growth inhibitory activity of the hydroethanolic extract of Achillea millefolium was studied in human tumor cell lines (NCI-H460 and HCT-15) and its mechanism of action was investigated. The GI 50 concentration was determined with the sulforhodamine B assay and cell cycle and apoptosis were analyzed by flow cytometry following incubation with PI or Annexin V FITC/PI, respectively. The expression levels of proteins involved in cell cycle and apoptosis were analyzed by Western blot. The extracts were characterized regarding their phenolic composition by LC-DAD-ESI/MS. 3,5-O-Dicaffeoylquinic acid, followed by 5-O-caffeoylquinic acid, were the main phenolic acids, while, luteolin-O-acetylhexoside and apigenin-O-acetylhexoside were the main flavonoids. This extract decreased the growth of the tested cell lines, being more potent in HCT-15 and then in NCI-H460 cells. Two different concentrations of the extract (75 and 100 μg/mL) caused alterations in cell cycle profile and increased apoptosis levels in HCT-15 and NCI-H460 cells. Moreover, the extract caused an increase in p53 and p21 expression in NCI-H460 cells (which have wt p53), and reduced XIAP levels in HCT-15 cells (with mutant p53). This work enhances the importance of A. millefolium as source of bioactive phenolic compounds, particularly of XIAP inhibitors. Copyright © 2018 Elsevier Ltd. All rights reserved.

Acacia catechu ethanolic bark extract induces apoptosis in human oral squamous carcinoma cells

Directory of Open Access Journals (Sweden)

Thangavelu Lakshmi

2017-01-01

Full Text Available Oral cancer is in approximately 30% of all cancers in India. This study was conducted to evaluate the cytotoxic activity of ethanolic extract of Acacia catechu bark (ACB against human squamous cell carcinoma cell line-25 (SCC-25. Cytotoxic effect of ACB extract was determined by 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium Bromide assay. A. catechu extract was treated SCC-25 cells with 25 and 50 μg/mL for 24 h. Apoptosis markers such as caspases-8 and 9, bcl-2, bax, and cytochrome c (Cyt-c were done by RT-PCR. Morphological changes of ACB treated cells were evaluated using acridine orange/ethidium bromide (AO/EB dual staining. Nuclear morphology and DNA fragmentation were evaluated using propidium iodide (PI staining. Further, cell cycle analysis was performed using flow cytometry. A. catechu treatment caused cytotoxicity in SCC-25 cells with an IC50 of 52.09 μg/mL. Apoptotic marker gene expressions were significantly increased on ACB treatment. Staining with AO/EB and PI shows membrane blebbing and nuclear membrane distortion, respectively, and it confirms the apoptosis induction in SCC-25 cells. These results suggest that ACB extract can be used as a modulating agent in oral squamous cell carcinoma.

Preparation of ubiquitin-conjugated proteins using an insect cell-free protein synthesis system.

Science.gov (United States)

Suzuki, Takashi; Ezure, Toru; Ando, Eiji; Nishimura, Osamu; Utsumi, Toshihiko; Tsunasawa, Susumu

2010-01-01

Ubiquitination is one of the most significant posttranslational modifications (PTMs). To evaluate the ability of an insect cell-free protein synthesis system to carry out ubiquitin (Ub) conjugation to in vitro translated proteins, poly-Ub chain formation was studied in an insect cell-free protein synthesis system. Poly-Ub was generated in the presence of Ub aldehyde (UA), a de-ubiquitinating enzyme inhibitor. In vitro ubiquitination of the p53 tumor suppressor protein was also analyzed, and p53 was poly-ubiquitinated when Ub, UA, and Mdm2, an E3 Ub ligase (E3) for p53, were added to the in vitro reaction mixture. These results suggest that the insect cell-free protein synthesis system contains enzymatic activities capable of carrying out ubiquitination. CBB-detectable ubiquitinated p53 was easily purified from the insect cell-free protein synthesis system, allowing analysis of the Ub-conjugated proteins by mass spectrometry (MS). Lys 305 of p53 was identified as one of the Ub acceptor sites using this strategy. Thus, we conclude that the insect cell-free protein synthesis system is a powerful tool for studying various PTMs of eukaryotic proteins including ubiqutination presented here.

Cytotoxic Effect of the Genus Sinularia Extracts on Human SCC25 and HaCaT Cells

International Nuclear Information System (INIS)

Wang, G.H.; Chou, T.H.; Liang, C.H.; Lin, R.J.; Sheu, J.H.; Wang, S.H.

2009-01-01

Soft corals of the genus Sinularia are being increasingly adopted to treat a wide variety of disease processes. However, the mechanism underlying its activity against human oral cancer cells is poorly understood. This study evaluates the cyototoxicity effects of the genus Sinularia extracts (S. grandilobata, S. parva, S. triangula, S. scabra, S. nanolobata and S. gibberosa) by SCC25 and HaCaT cells. The cell adhesion assay indicates that extracts reduce the cell attachment. Extracts exhibit a dose-dependent cytotoxic effect using MTS assay.Treatment of extracts to observe the morphological alterations in cells, membrane blebbing, nuclear condensation, and apoptotic bodies is demonstrated. Flow cytometry shows that extracts sensitized the cells in the G0/G1 and G2/M phases with a concomitant significantly increased sub-G1 fraction, suggesting cell death by apoptosis. Extracts of the genus Sinularia thus apparently cause apoptosis of SCC25 and HaCaT cells, and warrant further research investigating the possible antioral cancer compounds in these soft corals.

A single-cell and feeder-free culture system for monkey embryonic stem cells.

Science.gov (United States)

Ono, Takashi; Suzuki, Yutaka; Kato, Yosuke; Fujita, Risako; Araki, Toshihiro; Yamashita, Tomoko; Kato, Hidemasa; Torii, Ryuzo; Sato, Naoya

2014-01-01

Primate pluripotent stem cells (PSCs), including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), hold great potential for research and application in regenerative medicine and drug discovery. To maximize primate PSC potential, a practical system is required for generating desired functional cells and reproducible differentiation techniques. Much progress regarding their culture systems has been reported to date; however, better methods would still be required for their practical use, particularly in industrial and clinical fields. Here we report a new single-cell and feeder-free culture system for primate PSCs, the key feature of which is an originally formulated serum-free medium containing FGF and activin. In this culture system, cynomolgus monkey ESCs can be passaged many times by single-cell dissociation with traditional trypsin treatment and can be propagated with a high proliferation rate as a monolayer without any feeder cells; further, typical PSC properties and genomic stability can be retained. In addition, it has been demonstrated that monkey ESCs maintained in the culture system can be used for various experiments such as in vitro differentiation and gene manipulation. Thus, compared with the conventional culture system, monkey ESCs grown in the aforementioned culture system can serve as a cell source with the following practical advantages: simple, stable, and easy cell maintenance; gene manipulation; cryopreservation; and desired differentiation. We propose that this culture system can serve as a reliable platform to prepare primate PSCs useful for future research and application.

Toxicity of extracts from disposable chopsticks, toothpicks, and paper cups on L-929 cells.

Science.gov (United States)

Li, Juntao; Chen, Sifan; Li, Wenxue; Yang, Guangyu; Zhu, Wei

2015-04-01

To evaluate the toxicity of extracts from disposable chopsticks, toothpicks, and paper cups on L-929 cells. We followed national standards to prepare the extracts from disposable chopsticks, toothpicks, and paper cups used for the cell culture media, and the morphology of L-929 cells was observed with an optical microscope. The loss rate for adherent cells was evaluated with the trypan blue exclusion method, and cell proliferation was determined using the WST-1 assay. Compared with the control group, the cells cultured in media containing the extracts showed signs of apoptosis and necrosis after culturing for 4 or 7 days, and the loss rate for adherent cells was significantly increased (P disposable chopsticks, toothpicks, and paper cups can affect the growth and proliferation of L-929 cells and are potentially toxic to humans.

Fermentation characteristics of polysaccharide fractions extracted from the cell walls of soya bean cotyledons

NARCIS (Netherlands)

Laar, van H.; Tamminga, S.; Williams, B.A.; Verstegen, M.W.A.; Schols, H.A.

2000-01-01

Full-fat soya beans were separated into hulls and cotyledons. After separation the cell wall fraction was extracted from the cotyledons. These purified cell walls were sequentially extracted with 0.05 M cyclohexane-trans-1,2-diamine-N,N,N ,N -tetraacetate (CDTA) 0.05 M NH4 oxalate (extract 1), 0.05

Mangosteen peel extract reduces formalin-induced liver cell death in rats

Directory of Open Access Journals (Sweden)

Afiana Rohmani

2014-08-01

Full Text Available Background Formalin is a xenobiotic that is now commonly used as a preservative in the food industry. The liver is an organ that has the highest metabolic capacity as compared to other organs. Mangosteen or Garcinia mangostana Linn (GML peel contains xanthones, which are a source of natural antioxidants. The purpose of this study was to evaluate the effect of mangosteen peel extract on formalin-induced liver cell mortality rate and p53 protein expression in Wistar rats. Methods Eighteen rats received formalin orally for 2 weeks, and were subsequently divided into 3 groups, consisting of the formalin-control group receiving a placebo and treatment groups 1 and 2, which were treated with mangosteen peel extract at doses of 200 and 400 mg/kgBW/day, respectively. The treatment was carried out for 1 week, and finally the rats were terminated. The differences in liver cell mortality rate and p53 protein expression were analyzed. Results One-way ANOVA analysis showed significant differences in liver cell mortality rate among the three groups (p=0.004. The liver cell mortality rate in the treatment group receiving 400 mg/kgBW/day extract was lower than that in the formalin-control group. There was no p53 expression in all groups. Conclusions Garcinia mangostana Linn peel extract reduced the mortality rate of liver cells in rats receiving oral formalin. Involvement of p53 expression in liver cell mortality in rats exposed to oral formalin is presumably negligible.

Cytotoxic effects of Mangifera indica L. kernel extract on human breast cancer (MCF-7 and MDA-MB-231 cell lines) and bioactive constituents in the crude extract.

Science.gov (United States)

Abdullah, Al-Shwyeh Hussah; Mohammed, Abdulkarim Sabo; Abdullah, Rasedee; Mirghani, Mohamed Elwathig Saeed; Al-Qubaisi, Mothanna

2014-06-25

Waterlily Mango (Mangifera indica L.) is thought to be antioxidant-rich, conferred by its functional phytochemicals. The potential anticancer effects of the ethanolic kernel extract on breast cancer cells (MDA-MB-231 and MCF-7) using MTT, anti-proliferation, neutral red (NR) uptake and lactate dehydrogenase (LDH) release assays were evaluated. Cytological studies on the breast cancer cells were also conducted, and phytochemical analyses of the extract were carried out to determine the likely bioactive compounds responsible for such effects. Results showed the extract induced cytotoxicity in MDA-MB-231 cells and MCF-7 cells with IC50 values of 30 and 15 μg/mL, respectively. The extract showed significant toxicity towards both cell lines, with low toxicity to normal breast cells (MCF-10A). The cytotoxic effects on the cells were further confirmed by the NR uptake, antiproliferative and LDH release assays. Bioactive analyses revealed that many bioactives were present in the extract although butylated hydroxytoluene, a potent antioxidant, was the most abundant with 44.65%. M. indica extract appears to be more cytoxic to both estrogen positive and negative breast cancer cell lines than to normal breast cells. Synergistic effects of its antioxidant bioactives could have contributed to the cytotoxic effects of the extract. The extract of M. indica, therefore, has potential anticancer activity against breast cancer cells. This potential is worth studying further, and could have implications on future studies and eventually management of human breast cancers.

Anaerobic Aryl Reductive Dehalogenation of Halobenzoates by Cell Extracts of “Desulfomonile tiedjei”

OpenAIRE

DeWeerd, Kim A.; Suflita, Joseph M.

1990-01-01

We studied the transformation of halogenated benzoates by cell extracts of a dehalogenating anaerobe, “Desulfomonile tiedjei.” We found that cell extracts possessed aryl reductive dehalogenation activity. The activity was heat labile and dependent on the addition of reduced methyl viologen, but not on that of reduced NAD, NADP, flavin mononucleotide, flavin adenine dinucleotide, desulfoviridin, cytochrome c3, or benzyl viologen. Dehalogenation activity in extracts was stimulated by formate, C...

Effects of maple (Acer) plant part extracts on proliferation, apoptosis and cell cycle arrest of human tumorigenic and non-tumorigenic colon cells.

Science.gov (United States)

González-Sarrías, Antonio; Li, Liya; Seeram, Navindra P

2012-07-01

Phenolic-enriched extracts of maple sap and syrup, obtained from the sugar and red maple species (Acer saccharum Marsh, A. rubrum L., respectively), are reported to show anticancer effects. Despite traditional medicinal uses of various other parts of these plants by Native Americans, they have not been investigated for anticancer activity. Here leaves, stems/twigs, barks and sapwoods of both maple species were evaluated for antiproliferative effects against human colon tumorigenic (HCT-116, HT-29, Caco-2) and non-tumorigenic (CCD-18Co) cells. Extracts were standardized to total phenolic and ginnalin-A (isolated in our laboratory) levels. Overall, the extracts inhibited the growth of the colon cancer more than normal cells (over two-fold), their activities increased with their ginnalin-A levels, with red > sugar maple extracts. The red maple leaf extract, which contained the highest ginnalin-A content, was the most active extract (IC₅₀  = 35 and 16 µg/mL for extract and ginnalin-A, respectively). The extracts were not cytotoxic nor did they induce apoptosis of the colon cancer cells. However, cell cycle analyses revealed that the antiproliferative effects of the extracts were mediated through cell cycle arrest in the S-phase. The results from the current study suggest that these maple plant part extracts may have potential anticolon cancer effects. Copyright © 2011 John Wiley & Sons, Ltd.

Effects of biodegradable Mg–6Zn alloy extracts on apoptosis of intestinal epithelial cells

International Nuclear Information System (INIS)

Wang Zhanhui; Yan Jun; Li Jianan; Zheng Qi; Wang Zhigang; Zhang Xiaonong; Zhang Shaoxiang

2012-01-01

Highlights: ► We evaluated the effects of Mg–6Zn alloys on apoptosis of IEC-6 cells. ► The apoptosis was evaluated by investigating the expression of caspase-1 and Bcl-2. ► The IEC-6 cells displayed better cell functions in 60% or 20% extract. ► The conspicuous alkaline environment is disadvantageous to apoptosis of IEC cells. ► The excessive Mg concentration is disadvantageous to apoptosis of IEC-6 cells. - Abstract: In this study, intestinal epithelial cells (IEC)-6 were cultured in different concentration extracts of Mg–6Zn alloys for different time periods. To achieve a total of three concentrations (100%, 60% and 20% concentration), the extracts were serially diluted with Dulbecco's modified Eagle medium High Glucose to observe a dose–response relationship. We studied the indirect effects of Mg–6Zn alloys on IEC-6 cells apoptosis. The apoptosis of IEC-6 cells was measured using flow cytometry. And the apoptosis of IEC-6 cells was evaluated by investigating the expression of caspase-1and Bcl-2 using real-time polymerase chain reaction (PCR) and Western blotting tests. It was found that the levels of apoptosis in IEC-6 cells cultured in 100% Mg–6Zn alloy extracts were significantly higher than those in 60% and 20% extracts; the 100% extract can down-regulate expression of Bcl-2 after culture. The in vitro results indicated that the conspicuous alkaline environment and excessive Mg concentration, even Zn concentration caused by rapid corrosion of Mg–6Zn alloys promote IEC-6 cells apoptosis, although further experiments will be necessary to formally prove our conclusions. Therefore, the adjustment of the degradation rate is needed for using Mg–Zn alloy as a surgical suture material.

Cannabidiol rather than Cannabis sativa extracts inhibit cell growth and induce apoptosis in cervical cancer cells.

Science.gov (United States)

Lukhele, Sindiswa T; Motadi, Lesetja R

2016-09-01

Cervical cancer remains a global health related issue among females of Sub-Saharan Africa, with over half a million new cases reported each year. Different therapeutic regimens have been suggested in various regions of Africa, however, over a quarter of a million women die of cervical cancer, annually. This makes it the most lethal cancer amongst black women and calls for urgent therapeutic strategies. In this study we compare the anti-proliferative effects of crude extract of Cannabis sativa and its main compound cannabidiol on different cervical cancer cell lines. To achieve our aim, phytochemical screening, MTT assay, cell growth analysis, flow cytometry, morphology analysis, Western blot, caspase 3/7 assay, and ATP measurement assay were conducted. Results obtained indicate that both cannabidiol and Cannabis sativa extracts were able to halt cell proliferation in all cell lines at varying concentrations. They further revealed that apoptosis was induced by cannabidiol as shown by increased subG0/G1 and apoptosis through annexin V. Apoptosis was confirmed by overexpression of p53, caspase 3 and bax. Apoptosis induction was further confirmed by morphological changes, an increase in Caspase 3/7 and a decrease in the ATP levels. In conclusion, these data suggest that cannabidiol rather than Cannabis sativa crude extracts prevent cell growth and induce cell death in cervical cancer cell lines.

Extract of mouse embryonic stem cells induces the expression of pluripotency genes in human adipose tissue-derived stem cells.

Science.gov (United States)

Salehi, Paria Motamen; Foroutan, Tahereh; Javeri, Arash; Taha, Masoumeh Fakhr

2017-11-01

In some previous studies, the extract of embryonic carcinoma cells (ECCs) and embryonic stem cells (ESCs) have been used to reprogram somatic cells to more dedifferentiated state. The aim of this study was to investigate the effect of mouse ESCs extract on the expression of some pluripotency markers in human adipose tissue-derived stem cells (ADSCs). Human ADSCs were isolated from subcutaneous abdominal adipose tissue and characterized by flow cytometric analysis for the expression of some mesenchymal stem cell markers and adipogenic and osteogenic differentiation. Frequent freeze-thaw technique was used to prepare cytoplasmic extract of ESCs. Plasma membranes of the ADSCs were reversibly permeabilized by streptolysin-O (SLO). Then the permeabilized ADSCs were incubated with the ESC extract and cultured in resealing medium. After reprogramming, the expression of some pluripotency genes was evaluated by RT-PCR and quantitative real-time PCR (qPCR) analyses. Third-passaged ADSCs showed a fibroblast-like morphology and expressed mesenchymal stem cell markers. They also showed adipogenic and osteogenic differentiation potential. QPCR analysis revealed a significant upregulation in the expression of some pluripotency genes including OCT4 , SOX2 , NANOG , REX1 and ESG1 in the reprogrammed ADSCs compared to the control group. These findings showed that mouse ESC extract can be used to induce reprogramming of human ADSCs. In fact, this method is applicable for reprogramming of human adult stem cells to a more pluripotent sate and may have a potential in regenerative medicine.

Cell disruption and lipid extraction for microalgal biorefineries: A review.

Science.gov (United States)

Lee, Soo Youn; Cho, Jun Muk; Chang, Yong Keun; Oh, You-Kwan

2017-11-01

The microalgae-based biorefinement process has attracted much attention from academic and industrial researchers attracted to its biofuel, food and nutraceutical applications. In this paper, recent developments in cell-disruption and lipid-extraction methods, focusing on four biotechnologically important microalgal species (namely, Chlamydomonas, Haematococcus, Chlorella, and Nannochloropsis spp.), are reviewed. The structural diversity and rigidity of microalgal cell walls complicate the development of efficient downstream processing methods for cell-disruption and subsequent recovery of intracellular lipid and pigment components. Various mechanical, chemical and biological cell-disruption methods are discussed in detail and compared based on microalgal species and status (wet/dried), scale, energy consumption, efficiency, solvent extraction, and synergistic combinations. The challenges and prospects of the downstream processes for the future development of eco-friendly and economical microalgal biorefineries also are outlined herein. Copyright © 2017 Elsevier Ltd. All rights reserved.

Stem cell recovering effect of copper-free GHK in skin.

Science.gov (United States)

Choi, Hye-Ryung; Kang, Youn-A; Ryoo, Sun-Jong; Shin, Jung-Won; Na, Jung-Im; Huh, Chang-Hun; Park, Kyoung-Chan

2012-11-01

The peptide Gly-His-Lys (GHK) is a naturally occurring copper(II)-chelating motifs in human serum and cerebrospinal fluid. In industry, GHK (with or without copper) is used to make hair and skin care products. Copper-GHK plays a physiological role in the process of wound healing and tissue repair by stimulating collagen synthesis in fibroblasts. We also reported that copper-GHK promotes the survival of basal stem cells in the skin. However, the effects of copper-free GHK (GHK) have not been investigated well. In this study, the effects of GHK were studied using cultured normal human keratinocytes and skin equivalent (SE) models. In monolayer cultured keratinocytes, GHK increased the proliferation of keratinocytes. When GHK was added during the culture of SE models, the basal cells became more cuboidal than control model. In addition, there was linear and intense staining of α6 and β1 integrin along the basement membrane. The number of p63 and proliferating cell nuclear antigen positive cells was also significantly increased in GHK-treated SEs than in control SEs. Western blot and slide culture experiment showed that GHK increased the expression of integrin by keratinocytes. All these results showed that GHK increased the stemness and proliferative potential of epidermal basal cells, which is associated with increased expression of integrin. In conclusion, copper-free GHK showed similar effects with copper-GHK. Thus, it can be said that copper-free GHK can be used in industry to obtain the effects of copper-GHK in vivo. Further study is necessary to explore the relationship between copper-free GHK and copper-GHK. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd.

Chemical Characterization and in Vitro Cytotoxicity on Squamous Cell Carcinoma Cells of Carica Papaya Leaf Extracts

Directory of Open Access Journals (Sweden)

Thao T. Nguyen

2015-12-01

Full Text Available In traditional medicine, Carica papaya leaf has been used for a wide range of therapeutic applications including skin diseases and cancer. In this study, we investigated the in vitro cytotoxicity of aqueous and ethanolic extracts of Carica papaya leaves on the human oral squamous cell carcinoma SCC25 cell line in parallel with non-cancerous human keratinocyte HaCaT cells. Two out of four extracts showed a significantly selective effect towards the cancer cells and were found to contain high levels of phenolic and flavonoid compounds. The chromatographic and mass spectrometric profiles of the extracts obtained with Ultra High Performance Liquid Chromatography-Quadrupole Time of Flight-Mass Spectrometry were used to tentatively identify the bioactive compounds using comparative analysis. The principal compounds identified were flavonoids or flavonoid glycosides, particularly compounds from the kaempferol and quercetin families, of which several have previously been reported to possess anticancer activities. These results confirm that papaya leaf is a potential source of anticancer compounds and warrant further scientific investigation to validate the traditional use of papaya leaf to treat cancer.

Effects of Extraction Solvents on the Quantification of Free Amino Acids in Lyophilised Brewer’s Yeast

Directory of Open Access Journals (Sweden)

Andreea STĂNILĂ

2018-05-01

Full Text Available The aim of this work was to test some solvents in order to improve the free amino acids extraction from lyophilised brewer’s yeast. The brewer’ yeast was treated with four types of extraction solvents: Solvent I – acetonitrile 25%/HCl 0.01M (ACN; Solvent II – ethanol 80%; solvent III – HCl 0.05M/deionized water (1/1 volume; Solvent IV – HCl 0.05M/ethanol 80% (1/1 volume. The supernatants were analysed by HPLC-DAD-ESI-MS method. Acetonitrile provided the less quantities and number of amino acids extracted due to its weaker polarity. Solvent II and IV (ethanol, respectively acidified ethanol, which have an increased polarity, extracted 15 amino acids due to the addition of HCl in solvent IV. Solvent III (acidified water proved to be the best extraction solvent for the amino acids from brewer’s yeast providing the separation of 17 compounds: GLN, ASN, SER, GLY, ALA, ORN, PRO, HIS, LYS, GLU, TRP, LEU, PHE, ILE, AAA, HPHE, TYR.

Progress in Electrolyte-Free Fuel Cells

Directory of Open Access Journals (Sweden)

Yuzheng eLu

2016-05-01

Full Text Available Solid Oxide Fuel Cell (SOFC represents a clean electrochemical energy conversion technology with characteristics of high conversion efficiency and low emissions. It is one of the most important new energy technologies in the future. However, the manufacture of SOFCs based on the structure of anode/electrolyte/cathode is complicated and time-consuming. Thus, the cost for the entire fabrication and technology is too high to be affordable and challenges still hinder commercialization. Recently, a novel type of Electrolyte -free fuel cell (EFFC with single component was invented which could be the potential candidate for the next generation of advanced fuel cells. This paper briefly introduces the EFFC, working principle, performance and advantages with updated research progress. A number of key R&D issues about EFFCs have been addressed and future opportunities and challenges are discussed.

Cell-free protein synthesis for structure determination by X-ray crystallography.

Science.gov (United States)

Watanabe, Miki; Miyazono, Ken-ichi; Tanokura, Masaru; Sawasaki, Tatsuya; Endo, Yaeta; Kobayashi, Ichizo

2010-01-01

Structure determination has been difficult for those proteins that are toxic to the cells and cannot be prepared in a large amount in vivo. These proteins, even when biologically very interesting, tend to be left uncharacterized in the structural genomics projects. Their cell-free synthesis can bypass the toxicity problem. Among the various cell-free systems, the wheat-germ-based system is of special interest due to the following points: (1) Because the gene is placed under a plant translational signal, its toxic expression in a bacterial host is reduced. (2) It has only little codon preference and, especially, little discrimination between methionine and selenomethionine (SeMet), which allows easy preparation of selenomethionylated proteins for crystal structure determination by SAD and MAD methods. (3) Translation is uncoupled from transcription, so that the toxicity of the translation product on DNA and its transcription, if any, can be bypassed. We have shown that the wheat-germ-based cell-free protein synthesis is useful for X-ray crystallography of one of the 4-bp cutter restriction enzymes, which are expected to be very toxic to all forms of cells retaining the genome. Our report on its structure represents the first report of structure determination by X-ray crystallography using protein overexpressed with the wheat-germ-based cell-free protein expression system. This will be a method of choice for cytotoxic proteins when its cost is not a problem. Its use will become popular when the crystal structure determination technology has evolved to require only a tiny amount of protein.

Andrographis paniculata extracts and major constituent diterpenoids inhibit growth of intrahepatic cholangiocarcinoma cells by inducing cell cycle arrest and apoptosis.

Science.gov (United States)

Suriyo, Tawit; Pholphana, Nanthanit; Rangkadilok, Nuchanart; Thiantanawat, Apinya; Watcharasit, Piyajit; Satayavivad, Jutamaad

2014-05-01

Andrographis paniculata is an important herbal medicine widely used in several Asian countries for the treatment of various diseases due to its broad range of pharmacological activities. The present study reports that A. paniculata extracts potently inhibit the growth of liver (HepG2 and SK-Hep1) and bile duct (HuCCA-1 and RMCCA-1) cancer cells. A. paniculata extracts with different contents of major diterpenoids, including andrographolide, 14-deoxy-11,12-didehydroandrographolide, neoandrographolide, and 14-deoxyandrographolide, exhibited a different potency of growth inhibition. The ethanolic extract of A. paniculata at the first true leaf stage, which contained a high amount of 14-deoxyandrographolide but a low amount of andrographolide, showed a cytotoxic effect to cancer cells about 4 times higher than the water extract of A. paniculata at the mature leaf stage, which contained a high amount of andrographolide but a low amount of 14-deoxyandrographolide. Andrographolide, not 14-deoxy-11,12-didehydroandrographolide, neoandrographolide, or 14-deoxyandrographolide, possessed potent cytotoxic activity against the growth of liver and bile duct cancer cells. The cytotoxic effect of the water extract of A. paniculata at the mature leaf stage could be explained by the present amount of andrographolide, while the cytotoxic effect of the ethanolic extract of A. paniculata at the first true leaf stage could not. HuCCA-1 cells showed more sensitivity to A. paniculata extracts and andrographolide than RMCCA-1 cells. Furthermore, the ethanolic extract of A. paniculata at the first true leaf stage increased cell cycle arrest at the G0/G1 and G2/M phases, and induced apoptosis in both HuCCA-1 and RMCCA-1 cells. The expressions of cyclin-D1, Bcl-2, and the inactive proenzyme form of caspase-3 were reduced by the ethanolic extract of A. paniculata in the first true leaf stage treatment, while a proapoptotic protein Bax was increased. The cleavage of poly (ADP

Determination of the potency of a novel saw palmetto supercritical CO2 extract (SPSE) for 5α-reductase isoform II inhibition using a cell-free in vitro test system.

Science.gov (United States)

Pais, Pilar; Villar, Agustí; Rull, Santiago

2016-01-01

The nicotinamide adenine dinucleotide phosphate-dependent membrane protein 5α-reductase catalyses the conversion of testosterone to the most potent androgen - 5α-dihydrotestosterone. Two 5α-reductase isoenzymes are expressed in humans: type I and type II. The latter is found primarily in prostate tissue. Saw palmetto extract (SPE) has been used extensively in the treatment of lower urinary tract symptoms secondary to benign prostatic hyperplasia (BPH). The pharmacological effects of SPE include the inhibition of 5α-reductase, as well as anti-inflammatory and antiproliferative effects. Clinical studies of SPE have been inconclusive - some have shown significant results, and others have not - possibly the result of varying bioactivities of the SPEs used in the studies. To determine the in vitro potency in a cell-free test system of a novel SP supercritical CO2 extract (SPSE), an inhibitor of the 5α-reductase isoenzyme type II. The inhibitory potency of SPSE was compared to that of finasteride, an approved 5α-reductase inhibitor, on the basis of the enzymatic conversion of the substrate androstenedione to the 5α-reduced product 5α-androstanedione. By concentration-dependent inhibition of 5α-reductase type II in vitro (half-maximal inhibitory concentration 3.58±0.05 μg/mL), SPSE demonstrated competitive binding toward the active site of the enzyme. Finasteride, the approved 5α-reductase inhibitor tested as positive control, led to 63%-75% inhibition of 5α-reductase type II. SPSE effectively inhibits the enzyme that has been linked to BPH, and the amount of extract required for activity is comparatively low. It can be confirmed from the results of this study that SPSE has bioactivity that promotes prostate health at a level that is superior to that of many other phytotherapeutic extracts. The bioactivity of SPSE corresponds favorably to that reported for the hexane extract used in a large number of positive BPH clinical trials, as well as to finasteride

Effects of Ligusticum porteri (Osha) Root Extract on Human Promyelocytic Leukemia Cells.

Science.gov (United States)

Nguyen, Khanh; Sparks, Jean; Omoruyi, Felix

2017-01-01

Ligusticum porteri roots have been traditionally used in folk medicine, but the scientific basis is unclear. To investigate the cytotoxicity, antioxidant, and immunomodulatory effects of L. porteri root extract on human promyelocytic leukemia (HL-60) cells and H 2 O 2 -induced oxidative damaged HL-60 cells. HL-60 cells were incubated with different concentrations of root extract, and cells were harvested for viability assays on day 3 and 7. Cytokine levels (interferon-gamma [IFN-γ], interleukin-2 [IL-2], and interleukin-10 [IL-10]) and antioxidant indexes (malondialdehyde [MDA], reduced glutathione [GSH], superoxide dismutase [SOD], and catalase [CAT]) in H 2 O 2 -induced-stressed HL-60 were measured after 2 days. The viability of HL-60 challenged with H 2 O 2 declined by 42% compared to unstressed cells. After 7 days of incubation with 200 or 400 μg/mL L. porteri , the viability of HL-60 cells was two-fold higher than the control. Stressed HL-60 cells treated with 100, 200, and 400 μg/mL L. porteri reduced the lipid peroxidation by 12%-13%. We noted an increase in GSH levels, SOD and CAT activities in stressed HL-60 supplemented with 400 μg/mL root extract. Treatment with 400 μg/mL L. porteri significantly ( P effect against the oxidation of reduced glutathione (GSH)Treatment with L. porteri root extract may be effective in preventing oxidative damage through increasing the activities of antioxidant enzymes (superoxide dismutase [SOD] and catalase [CAT]) in acute promyelocytic leukemia cells.

Vochysia rufa Stem Bark Extract Protects Endothelial Cells against High Glucose Damage

Directory of Open Access Journals (Sweden)

Neire Moura de Gouveia

2017-02-01

Full Text Available Background: Increased oxidative stress by persistent hyperglycemia is a widely accepted factor in vascular damage responsible for type 2 diabetes complications. The plant Vochysia rufa (Vr has been used in folk medicine in Brazil for the treatment of diabetes. Thus; the protective effect of a Vr stem bark extract against a challenge by a high glucose concentration on EA.hy926 (EA endothelial cells is evaluated. Methods: Vegetal material is extracted with distilled water by maceration and evaporated until dryness under vacuum. Then; it is isolated by capillary electrophoresis–tandem mass spectrometry. Cell viability is evaluated on EA cells treated with 0.5–100 µg/mL of the Vr extract for 24 h. The extract is diluted at concentrations of 5, 10 and 25 µg/mL and maintained for 24 h along with 30 mM of glucose to evaluate its protective effect on reduced glutathione (GSH; glutathione peroxidase (GPx and reductase (GR and protein carbonyl groups. Results: V. rufa stem bark is composed mainly of sugars; such as inositol; galactose; glucose; mannose; sacarose; arabinose and ribose. Treatment with Vr up to 100 µg/mL for 24 h did not affect cell viability. Treatment of EA cells with 30 mM of glucose for 24 h significantly increased the cell damage. EA cells treated with 30 mM of glucose showed a decrease of GSH concentration and increased Radical Oxygen Species (ROS and activity of antioxidant enzymes and protein carbonyl levels; compared to control. Co-treatment of EA with 30 mM glucose plus 1–10 μg/mL Vr significantly reduced cell damage while 5–25 μg/mL Vr evoked a significant protection against the glucose insult; recovering ROS; GSH; antioxidant enzymes and carbonyls to baseline levels. Conclusion: V. rufa extract protects endothelial cells against oxidative damage by modulating ROS; GSH concentration; antioxidant enzyme activity and protein carbonyl levels.

A powder-free surgical glove bag for retraction of the gallbladder during laparoscopic cholecystectomy.

Science.gov (United States)

Holme, Jørgen Bendix; Mortensen, Frank Viborg

2005-08-01

To test the use of a simple and cheap powder-free glove bag to extract the gallbladder during laparoscopic cholecystectomy (LC). The medical records of 142 consecutive patients who had their gallbladder removed using a powder-free glove bag were reviewed. No complications in the form of bile or stone spillage during extraction were observed. The absence of complications and the low cost make routine use of the glove bag a wise option for extracting the gallbladder during LC. The use of the glove bag seems to reduce the risk of contamination with bacteria, bile, and gallstones and may reduce contamination by malignant cells in case of unexpected gallbladder carcinoma.

Impacts of tomato extract on the mice fibrosarcoma cells

Directory of Open Access Journals (Sweden)

Shirzad Hedayatollah

2013-01-01

Full Text Available Introduction: The anticancer effect of tomato lycopene has been approved in some cancers. This study was aimed to determine the prohibitive and therapeutic effects of tomato extract on the growth of fibrosarcoma in mice. Materials and Methods: In this experimental study 3 groups of 10 male Balb/c mice were injected subcutaneously with 5×105 WEHI-164 tumor cells in the chest area. Prevention group was fed tomato extract (5 mg for a 4 week period (from 2 weeks before tumor cell injection up to 2 weeks after injection and the treatment group was fed simultaneously with tumor cell injection up to two weeks after injection daily by an oral gastric tube. The tumors areas were measured and recorded on days 10, 12, 14, 16, 18, 20 and 22. The data were analyzed using Kruskal-Wallis and Mann-Whitney tests. Results: The results showed that the tumor areas in control group were significantly more after the intervention than two groups of treatment and prevention (p<0.05. The difference was not statistically significant between the two groups of prevention and treatment. Conclusion: With emphasize on antioxidant of tomato, it seems that tomato extract has an important role in prevention and control fibrosarcoma growth.

Efficient indium-tin-oxide free inverted organic solar cells based on aluminum-doped zinc oxide cathode and low-temperature aqueous solution processed zinc oxide electron extraction layer

International Nuclear Information System (INIS)

Chen, Dazheng; Zhang, Chunfu; Wang, Zhizhe; Zhang, Jincheng; Tang, Shi; Wei, Wei; Sun, Li; Hao, Yue

2014-01-01

Indium-tin-oxide (ITO) free inverted organic solar cells (IOSCs) based on aluminum-doped zinc oxide (AZO) cathode, low-temperature aqueous solution processed zinc oxide (ZnO) electron extraction layer, and poly(3-hexylthiophene-2, 5-diyl):[6, 6]-phenyl C 61 butyric acid methyl ester blend were realized in this work. The resulted IOSC with ZnO annealed at 150 °C shows the superior power conversion efficiency (PCE) of 3.01%, if decreasing the ZnO annealing temperature to 100 °C, the obtained IOSC also shows a PCE of 2.76%, and no light soaking issue is observed. It is found that this ZnO film not only acts as an effective buffer layer but also slightly improves the optical transmittance of AZO substrates. Further, despite the relatively inferior air-stability, these un-encapsulated AZO/ZnO IOSCs show comparable PCEs to the referenced ITO/ZnO IOSCs, which demonstrates that the AZO cathode is a potential alternative to ITO in IOSCs. Meanwhile, this simple ZnO process is compatible with large area deposition and plastic substrates, and is promising to be widely used in IOSCs and other relative fields.

Extraction of Natural Antioxidants from the Thelephora ganbajun Mushroom by an Ultrasound-Assisted Extraction Technique and Evaluation of Antiproliferative Activity of the Extract against Human Cancer Cells

Directory of Open Access Journals (Sweden)

Dong-Ping Xu

2016-10-01

Full Text Available The Thelephora ganbajun mushroom has been found to be a potential rich source of natural antioxidants. In this study, an ultrasound-assisted extraction (UAE technique together with GRAS (generally recognized as safe solvents (ethanol and water was used to maximize the extraction of antioxidants from Thelephora ganbajun. Five extraction parameters (ethanol concentration, solvent to solid ratio, extraction time, temperature and ultrasound power were investigated by single-factor experiments, and then a central composite rotatable design was employed to study interaction of three key extraction parameters. The optimum conditions were as follows: 57.38% ethanol, 70.15 mL/g solvent to solid ratio, 10.58 min extraction time, 40 °C extraction temperature and 500 W ultrasound power. Under the optimum conditions, the antioxidant activity obtained was 346.98 ± 12.19 µmol Trolox/g DW, in accordance with the predicted value of 344.67 µmol Trolox/g DW. Comparison of UAE with conventional maceration and Soxhlet extraction, the UAE method showed stronger extract efficiency in a shorter extraction time. These results showed that UAE was an effective technique to extract antioxidants from Thelephora ganbajun. Furthermore, the extracts obtained under the optimized conditions exhibited antiproliferative activities toward human lung (A549, breast (MCF-7, liver (HepG2 and colon (HT-29 cancer cells, especially for liver and lung cancer cells. In addition, rutin, 2-hydrocinnamic acid and epicatechin were identified in the extract, which might contribute to antioxidant and antiproliferative activities.

Extraction of Natural Antioxidants from the Thelephora ganbajun Mushroom by an Ultrasound-Assisted Extraction Technique and Evaluation of Antiproliferative Activity of the Extract against Human Cancer Cells.

Science.gov (United States)

Xu, Dong-Ping; Zheng, Jie; Zhou, Yue; Li, Ya; Li, Sha; Li, Hua-Bin

2016-10-01

The Thelephora ganbajun mushroom has been found to be a potential rich source of natural antioxidants. In this study, an ultrasound-assisted extraction (UAE) technique together with GRAS (generally recognized as safe) solvents (ethanol and water) was used to maximize the extraction of antioxidants from Thelephora ganbajun . Five extraction parameters (ethanol concentration, solvent to solid ratio, extraction time, temperature and ultrasound power) were investigated by single-factor experiments, and then a central composite rotatable design was employed to study interaction of three key extraction parameters. The optimum conditions were as follows: 57.38% ethanol, 70.15 mL/g solvent to solid ratio, 10.58 min extraction time, 40 °C extraction temperature and 500 W ultrasound power. Under the optimum conditions, the antioxidant activity obtained was 346.98 ± 12.19 µmol Trolox/g DW, in accordance with the predicted value of 344.67 µmol Trolox/g DW. Comparison of UAE with conventional maceration and Soxhlet extraction, the UAE method showed stronger extract efficiency in a shorter extraction time. These results showed that UAE was an effective technique to extract antioxidants from Thelephora ganbajun . Furthermore, the extracts obtained under the optimized conditions exhibited antiproliferative activities toward human lung (A549), breast (MCF-7), liver (HepG2) and colon (HT-29) cancer cells, especially for liver and lung cancer cells. In addition, rutin, 2-hydrocinnamic acid and epicatechin were identified in the extract, which might contribute to antioxidant and antiproliferative activities.

Live cell imaging reveals different modes of cytotoxic action of extracts derived from commonly used luting cements.

Science.gov (United States)

Trumpaitė-Vanagienė, Rita; Čebatariūnienė, Alina; Tunaitis, Virginijus; Pūrienė, Alina; Pivoriūnas, Augustas

2018-02-01

To compare cytotoxicity of extracts derived from commonly used luting cements: Hoffmann's Zinc Phosphate (ZPC), GC Fuji Plus Resin Modified Glass Ionomer (RMGIC) and 3M ESPE RelyX Unicem Resin Cement (RC) on primary human gingival fibroblasts (HGFs). HGFs were exposed to different concentrations of the ZPC, RMGIC and RC extracts. The cytotoxicity was assessed with the PrestoBlue Cell Viability Reagent and viable cells were counted by a haemocytometer using the trypan blue exclusion test. In order to determine the primary mechanism of the cell death induced by extracts from different luting cements, the real-time monitoring of caspase-3/-7 activity and membrane integrity of cells was employed. The extracts from the RMGIC and ZPC decreased the metabolic activity and numbers of viable cells. Unexpectedly, the extracts from the RC evoked only small effects on the metabolic activity of HGFs with a decreasing number of viable cells in a dose-and time-dependent manner. The live cell imaging revealed that the apoptosis was the primary mechanism of a cell death induced by the extracts derived from the RMGIC, whereas the extracts from the RC and ZPC induced a cell death through a necrotic and caspase-independent pathway. The apoptosis was the primary mechanism of the cell death induced by the extracts derived from the RMGIC, whereas the extracts from the RC and ZPC induced a cell death via a necrotic pathway. We suggest that metabolic assays commonly used to assess the cytotoxicity of luting cements should be validated by alternative methods. Copyright © 2017 Elsevier Ltd. All rights reserved.

Glioprotective Effects of Ashwagandha Leaf Extract against Lead Induced Toxicity

Directory of Open Access Journals (Sweden)

Praveen Kumar

2014-01-01

Full Text Available Withania somnifera (Ashwagandha, also known as Indian Ginseng, is a well-known Indian medicinal plant due to its antioxidative, antistress, antigenotoxic, and immunomodulatory properties. The present study was designed to assess and establish the cytoprotective potential of Ashwagandha leaf aqueous extract against lead induced toxicity. Pretreatment of C6 cells with 0.1% Ashwagandha extract showed cytoprotection against 25 μM to 400 μM concentration of lead nitrate. Further pretreatment with Ashwagandha extract to lead nitrate exposed cells (200 μM resulted in normalization of glial fibrillary acidic protein (GFAP expression as well as heat shock protein (HSP70, mortalin, and neural cell adhesion molecule (NCAM expression. Further, the cytoprotective efficacy of Ashwagandha extract was studied in vivo. Administration of Ashwagandha extract provided significant protection to lead induced altered antioxidant defense that may significantly compromise normal cellular function. Ashwagandha also provided a significant protection to lipid peroxidation (LPx levels, catalase, and superoxide dismutase (SOD but not reduced glutathione (GSH contents in brain tissue as well as peripheral organs, liver and kidney, suggesting its ability to act as a free radical scavenger protecting cells against toxic insult. These results, thus, suggest that Ashwagandha water extract may have the potential therapeutic implication against lead poisoning.

Ultrasound-assisted extraction of phenolic compounds from Cratoxylum formosum ssp. formosum leaves using central composite design and evaluation of its protective ability against H2O2-induced cell death.

Science.gov (United States)

Yingngam, Bancha; Monschein, Marlene; Brantner, Adelheid

2014-09-01

To optimize the processing parameters for phenolic compounds extracted from Cratoxylum formosum ssp. formosum leaves using an ultrasound-assisted extraction and to evaluate its protective ability against H2O2-induced cell death. The influence of three independent variables including ethanol concentration (%), extraction temperature (°C) and extraction time (min) on the extraction yield of phenolic compounds were optimized using a central composite design-based response surface methodology. The obtained extract was assessed for its antioxidant activity by DPPH(•) and ABTS(•)(+) methods. Cellular protective ability against H2O2-induced cell death was evaluated on HEK293 cells using the MTT assay. The optimal conditions to achieve maximal yields of phenolic compounds were ethanol concentration of 50.33% (v/v), temperature of 65 °C, and extractiontion time of 15 min. The yield of phenolic compounds was (40.00±1.00) mg gallic acid equivalent/g dry powder which matched well with the values predicted from the proposed model. These conditions resulted in a higher efficiency concerning the extraction of phenolics compared to a conventional heat reflux extraction by providing shorter extraction time and reduced energy consumption. 5-O-caffeoylquinic acid identified by high performance liquid chromatography-diode array detector-electron spin ionization-mass spectrometry was the major compound in the obtained extract [(41.66±0.07) mg/g plant extract]. The obtained extract showed a strong ability to scavenge both DPPH(•) and ABTS(•)(+) free radicals and exhibited additionally good ability to protect HEK293 cells death against oxidative stress. These results indicate the suitability of ultrasound-assisted extraction for the extraction of phenolic compounds from Cratoxylum formosum ssp. formosum leaves. This phenolic-enriched extract can be used as valuable antioxidant source for health benefits. Copyright © 2014 Hainan Medical College. Published by Elsevier B.V. All

Determination of the potency of a novel saw palmetto supercritical CO2 extract (SPSE for 5α-reductase isoform II inhibition using a cell-free in vitro test system

Directory of Open Access Journals (Sweden)

Pais P

2016-04-01

Full Text Available Pilar Pais, Agustí Villar, Santiago Rull Euromed, Barcelona, Spain Background: The nicotinamide adenine dinucleotide phosphate-dependent membrane protein 5α-reductase catalyses the conversion of testosterone to the most potent androgen – 5α-dihydrotestosterone. Two 5α-reductase isoenzymes are expressed in humans: type I and type II. The latter is found primarily in prostate tissue. Saw palmetto extract (SPE has been used extensively in the treatment of lower urinary tract symptoms secondary to benign prostatic hyperplasia (BPH. The pharmacological effects of SPE include the inhibition of 5α-reductase, as well as anti-inflammatory and antiproliferative effects. Clinical studies of SPE have been inconclusive – some have shown significant results, and others have not – possibly the result of varying bioactivities of the SPEs used in the studies. Purpose: To determine the in vitro potency in a cell-free test system of a novel SP supercritical CO2 extract (SPSE, an inhibitor of the 5α-reductase isoenzyme type II. Materials and methods: The inhibitory potency of SPSE was compared to that of finasteride, an approved 5α-reductase inhibitor, on the basis of the enzymatic conversion of the substrate androstenedione to the 5α-reduced product 5α-androstanedione. Results: By concentration-dependent inhibition of 5α-reductase type II in vitro (half-maximal inhibitory concentration 3.58±0.05 µg/mL, SPSE demonstrated competitive binding toward the active site of the enzyme. Finasteride, the approved 5α-reductase inhibitor tested as positive control, led to 63%–75% inhibition of 5α-reductase type II. Conclusion: SPSE effectively inhibits the enzyme that has been linked to BPH, and the amount of extract required for activity is comparatively low. It can be confirmed from the results of this study that SPSE has bioactivity that promotes prostate health at a level that is superior to that of many other phytotherapeutic extracts. The

Serum-free media formulations are cell line-specific and require optimization for microcarrier culture.

Science.gov (United States)

Tan, Kah Yong; Teo, Kim Leng; Lim, Jessica F Y; Chen, Allen K L; Choolani, Mahesh; Reuveny, Shaul; Chan, Jerry; Oh, Steve Kw

2015-08-01

Mesenchymal stromal cells (MSCs) are being investigated as potential cell therapies for many different indications. Current methods of production rely on traditional monolayer culture on tissue-culture plastic, usually with the use of serum-supplemented growth media. However, the monolayer culturing system has scale-up limitations and may not meet the projected hundreds of billions to trillions batches of cells needed for therapy. Furthermore, serum-free medium offers several advantages over serum-supplemented medium, which may have supply and contaminant issues, leading to many serum-free medium formulations being developed. We cultured seven MSC lines in six different serum-free media and compared their growth between monolayer and microcarrier culture. We show that (i) expansion levels of MSCs in serum-free monolayer cultures may not correlate with expansion in serum-containing media; (ii) optimal culture conditions (serum-free media for monolayer or microcarrier culture) differ for each cell line; (iii) growth in static microcarrier culture does not correlate with growth in stirred spinner culture; (iv) and that early cell attachment and spreading onto microcarriers does not necessarily predict efficiency of cell expansion in agitated microcarrier culture. Current serum-free media developed for monolayer cultures of MSCs may not support MSC proliferation in microcarrier cultures. Further optimization in medium composition will be required for microcarrier suspension culture for each cell line. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Microscale to manufacturing scale-up of cell-free cytokine production--a new approach for shortening protein production development timelines.

Science.gov (United States)

Zawada, James F; Yin, Gang; Steiner, Alexander R; Yang, Junhao; Naresh, Alpana; Roy, Sushmita M; Gold, Daniel S; Heinsohn, Henry G; Murray, Christopher J

2011-07-01

Engineering robust protein production and purification of correctly folded biotherapeutic proteins in cell-based systems is often challenging due to the requirements for maintaining complex cellular networks for cell viability and the need to develop associated downstream processes that reproducibly yield biopharmaceutical products with high product quality. Here, we present an alternative Escherichia coli-based open cell-free synthesis (OCFS) system that is optimized for predictable high-yield protein synthesis and folding at any scale with straightforward downstream purification processes. We describe how the linear scalability of OCFS allows rapid process optimization of parameters affecting extract activation, gene sequence optimization, and redox folding conditions for disulfide bond formation at microliter scales. Efficient and predictable high-level protein production can then be achieved using batch processes in standard bioreactors. We show how a fully bioactive protein produced by OCFS from optimized frozen extract can be purified directly using a streamlined purification process that yields a biologically active cytokine, human granulocyte-macrophage colony-stimulating factor, produced at titers of 700 mg/L in 10 h. These results represent a milestone for in vitro protein synthesis, with potential for the cGMP production of disulfide-bonded biotherapeutic proteins. Copyright © 2011 Wiley Periodicals, Inc.

Murraya koenigii leaf extract inhibits proteasome activity and induces cell death in breast cancer cells

Directory of Open Access Journals (Sweden)

Noolu Bindu

2013-01-01

Full Text Available Abstract Background Inhibition of the proteolytic activity of 26S proteasome, the protein-degrading machine, is now considered a novel and promising approach for cancer therapy. Interestingly, proteasome inhibitors have been demonstrated to selectively kill cancer cells and also enhance the sensitivity of tumor cells to chemotherapeutic agents. Recently, polyphenols/flavonoids have been reported to inhibit proteasome activity. Murraya koenigii Spreng, a medicinally important herb of Indian origin, has been used for centuries in the Ayurvedic system of medicine. Here we show that Murraya koenigii leaves (curry leaves, a rich source of polyphenols, inhibit the proteolytic activity of the cancer cell proteasome, and cause cell death. Methods Hydro-methanolic extract of curry leaves (CLE was prepared and its total phenolic content [TPC] determined by, the Folin-Ciocalteau’s method. Two human breast carcinoma cell lines: MCF-7 and MDA-MB-231 and a normal human lung fibroblast cell line, WI-38 were used for the studies. Cytotoxicity of the CLE was assessed by the MTT assay. We studied the effect of CLE on growth kinetics using colony formation assay. Growth arrest was assessed by cell cycle analysis and apoptosis by Annexin-V binding using flow cytometry. Inhibition of the endogenous 26S proteasome was studied in intact cells and cell extracts using substrates specific to 20S proteasomal enzymes. Results CLE decreased cell viability and altered the growth kinetics in both the breast cancer cell lines in a dose-dependent manner. It showed a significant arrest of cells in the S phase albeit in cancer cells only. Annexin V binding data suggests that cell death was via the apoptotic pathway in both the cancer cell lines. CLE treatment significantly decreased the activity of the 26S proteasome in the cancer but not normal cells. Conclusions Our study suggests M. koenigii leaves to be a potent source of proteasome inhibitors that lead to cancer cell death

Parameter extraction of different fuel cell models with transferred adaptive differential evolution

International Nuclear Information System (INIS)

Gong, Wenyin; Yan, Xuesong; Liu, Xiaobo; Cai, Zhihua

2015-01-01

To improve the design and control of FC (fuel cell) models, it is important to extract their unknown parameters. Generally, the parameter extraction problems of FC models can be transformed as nonlinear and multi-variable optimization problems. To extract the parameters of different FC models exactly and fast, in this paper, we propose a transferred adaptive DE (differential evolution) framework, in which the successful parameters of the adaptive DE solving previous problems are properly transferred to solve new optimization problems in the similar problem-domains. Based on this framework, an improved adaptive DE method (TRADE, in short) is presented as an illustration. To verify the performance of our proposal, TRADE is used to extract the unknown parameters of two types of fuel cell models, i.e., PEMFC (proton exchange membrane fuel cell) and SOFC (solid oxide fuel cell). The results of TRADE are also compared with those of other state-of-the-art EAs (evolutionary algorithms). Even though the modification is very simple, the results indicate that TRADE can extract the parameters of both PEMFC and SOFC models exactly and fast. Moreover, the V–I characteristics obtained by TRADE agree well with the simulated and experimental data in all cases for both types of fuel cell models. Also, it improves the performance of the original adaptive DE significantly in terms of both the quality of final solutions and the convergence speed in all cases. Additionally, TRADE is able to provide better results compared with other EAs. - Highlights: • A framework of transferred adaptive differential evolution is proposed. • Based on the framework, an improved differential evolution (TRADE) is presented. • TRADE obtains very promising results to extract the parameters of PEMFC and SOFC models

The best time of cytotoxicity for extracted cell wall from Lactobacillus casei and paracasei in K562 cell line

Directory of Open Access Journals (Sweden)

Riki M

2013-02-01

Full Text Available Background: The aim of this study was to evaluate the effect of extracted cell walls from Lactobacillus casei and Lactobacillus paracasei as probiotic bacteria (isolated from common carp intestine on K562 and the role of cell concentration on the results of MTT [3-(4,5-Dimethylthiazol-2-yl2,5- Diphenyl tetrazolium Bromide] test.Methods: For this purpose, bacteria were cultured in specific medium (MRS broth at anaerobic condition for 24-48 hour. After incubation period culture medium was centri-fuged, then the cells were washed twice with PBS buffer to remove additional medium. Finally, collected bacterial cell disrupted by Sonication and cell walls were separated from other components by centrifugation. After that, different concentrations of cell walls (500, 1000, 2000 and 4000 µg/ml were prepared in RPMI medium for each bacteria, separately. Then anticancer properties of the cell walls were determined in vitro at 12, 24, 48 and 72 h, also the effect of K562 concentration was assayed with MTT technique.Results: The results showed extracted cell wall from both probiotic statistically (P=0.098 have anti turmeric properties in K562 and their properties will arise in relation with concentration. As well as, we found that the number of cell had not any affect on the result of MTT assay.Conclusion: We conclude that the cytotoxicity property of extracted cell wall is related in the type of bacteria, but this anticancer property would warrant further study on the clinical application of extracted cell wall.

Characterization of a Sea Buckthorn Extract and Its Effect on Free and Encapsulated Lactobacillus casei.

Science.gov (United States)

Pop, Oana Lelia; Dulf, Francisc Vasile; Cuibus, Lucian; Castro-Giráldez, Marta; Fito, Pedro J; Vodnar, Dan Cristian; Coman, Cristina; Socaciu, Carmen; Suharoschi, Ramona

2017-11-24

Probiotics are bacteria that can provide health benefits to consumers and are suitable to be added to a variety of foods. In this research, viability of immobilized Lactobacillus casei in alginate with or without sea buckthorn lipid extract were studied during heat treatment and with an in vitro gastrointestinal model. The characterization of the lipid extract was also done using the UV-Vis spectrometry (UV-Vis), high-performance liquid chromatography photodiode array detection method (HPLC-PDA), gas chromatography coupled with mass spectrometry (GS-MS) and Cryo scanning electron microscopy (Cryo-SEM). During heat treatment, the entrapped probiotic cells proved high viability (>6 CFU log/g), even at temperatures above 50 °C. The rich in monounsaturated fatty acids sea buckthorn fraction improved the in vitro digestion passage regarding the probiotic viability. The survival of the probiotic cells was 15% higher after 2 h in the acidic medium of the simulated gastric fluid in the sample where L. casei was encapsulated with the sea buckthorn extract compared with the samples where no extract was added. Thus, this approach may be effective for the future development of probiotic-supplemented foods as foods with health welfare for the consumers.

Characterization of a Sea Buckthorn Extract and Its Effect on Free and Encapsulated Lactobacillus casei

Directory of Open Access Journals (Sweden)

Oana Lelia Pop

2017-11-01

Full Text Available Probiotics are bacteria that can provide health benefits to consumers and are suitable to be added to a variety of foods. In this research, viability of immobilized Lactobacillus casei in alginate with or without sea buckthorn lipid extract were studied during heat treatment and with an in vitro gastrointestinal model. The characterization of the lipid extract was also done using the UV-Vis spectrometry (UV-Vis, high-performance liquid chromatography photodiode array detection method (HPLC-PDA, gas chromatography coupled with mass spectrometry (GS-MS and Cryo scanning electron microscopy (Cryo-SEM. During heat treatment, the entrapped probiotic cells proved high viability (>6 CFU log/g, even at temperatures above 50 °C. The rich in monounsaturated fatty acids sea buckthorn fraction improved the in vitro digestion passage regarding the probiotic viability. The survival of the probiotic cells was 15% higher after 2 h in the acidic medium of the simulated gastric fluid in the sample where L. casei was encapsulated with the sea buckthorn extract compared with the samples where no extract was added. Thus, this approach may be effective for the future development of probiotic-supplemented foods as foods with health welfare for the consumers.

Continuous nucleus extraction by optically-induced cell lysis on a batch-type microfluidic platform.

Science.gov (United States)

Huang, Shih-Hsuan; Hung, Lien-Yu; Lee, Gwo-Bin

2016-04-21

The extraction of a cell's nucleus is an essential technique required for a number of procedures, such as disease diagnosis, genetic replication, and animal cloning. However, existing nucleus extraction techniques are relatively inefficient and labor-intensive. Therefore, this study presents an innovative, microfluidics-based approach featuring optically-induced cell lysis (OICL) for nucleus extraction and collection in an automatic format. In comparison to previous micro-devices designed for nucleus extraction, the new OICL device designed herein is superior in terms of flexibility, selectivity, and efficiency. To facilitate this OICL module for continuous nucleus extraction, we further integrated an optically-induced dielectrophoresis (ODEP) module with the OICL device within the microfluidic chip. This on-chip integration circumvents the need for highly trained personnel and expensive, cumbersome equipment. Specifically, this microfluidic system automates four steps by 1) automatically focusing and transporting cells, 2) releasing the nuclei on the OICL module, 3) isolating the nuclei on the ODEP module, and 4) collecting the nuclei in the outlet chamber. The efficiency of cell membrane lysis and the ODEP nucleus separation was measured to be 78.04 ± 5.70% and 80.90 ± 5.98%, respectively, leading to an overall nucleus extraction efficiency of 58.21 ± 2.21%. These results demonstrate that this microfluidics-based system can successfully perform nucleus extraction, and the integrated platform is therefore promising in cell fusion technology with the goal of achieving genetic replication, or even animal cloning, in the near future.

Effect on cell surface hydrophobicity and susceptibility of Helicobacter pylori to medicinal plant extracts.

Science.gov (United States)

Annuk, H; Hirmo, S; Türi, E; Mikelsaar, M; Arak, E; Wadström, T

1999-03-01

Effects on aqueous extracts of medicinal plants on ten Helicobacter pylori strains were studied by the salt aggregation test to determine the possibility to modulate their cell surface hydrophobicity and by an agar diffusion assay for detection of antimicrobial activity. It was established that aqueous extracts of bearberry and cowberry leaves enhance cell aggregation of all H. pylori strains tested by the salt aggregation test, and the extract of bearberry possessed a remarkable bacteriostatic activity. Pure tannic acid showed a result similar to that of bearberry and cowberry extracts which contained a large amount of tannins. In contrast, extracts of wild camomile and pineapple-weed, which blocked aggregation of H. pylori, contained small amounts of tannins and did not reveal any antimicrobial activity. Tannic acid seems to be the component of bearberry and cowberry aqueous extracts with the highest activity to decrease cell surface hydrophobicity as well as in antibacterial activity against H. pylori.

IRON AND FREE RADICAL OXIDATIONS IN CELL MEMBRANES

Science.gov (United States)

Schafer, Freya Q.; Yue Qian, Steven; Buettner, Garry R.

2013-01-01

Brain tissue being rich in polyunsaturated fatty acids, is very susceptible to lipid peroxidation. Iron is well known to be an important initiator of free radical oxidations. We propose that the principal route to iron-mediated lipid peroxidations is via iron-oxygen complexes rather than the reaction of iron with hydrogen peroxide, the Fenton reaction. To test this hypothesis, we enriched leukemia cells (K-562 and L1210 cells) with docosahexaenoic acid (DHA) as a model for brain tissue, increasing the amount of DHA from approximately 3 mole % to 32 mole %. These cells were then subjected to ferrous iron and dioxygen to initiate lipid peroxidation in the presence or absence of hydrogen peroxide. Lipid-derived radicals were detected using EPR spin trapping with α-(4-pyridyl-1-oxide)-N-t-butylnitrone (POBN). As expected, lipid-derived radical formation increases with increasing cellular lipid unsaturation. Experiments with Desferal demonstrate that iron is required for the formation of lipid radicals from these cells. Addition of iron to DHA-enriched L1210 cells resulted in significant amounts of radical formation; radical formation increased with increasing amount of iron. However, the exposure of cells to hydrogen peroxide before the addition of ferrous iron did not increase cellular radical formation, but actually decreased spin adduct formation. These data suggest that iron-oxygen complexes are the primary route to the initiation of biological free radical oxidations. This model proposes a mechanism to explain how catalytic iron in brain tissue can be so destructive. PMID:10872752

Evaluation of the biological activity of sunflower hull extracts

Energy Technology Data Exchange (ETDEWEB)

Taha, F. S.; Wagdy, S. M.; Hassanein, M. M. M.; Hamed, S. F.

2012-11-01

This work was planned with the aim of adding value to sunflower seed hulls, a waste product of the oil industry by preparing a sunflower hull phenolic extract rich in chlorogenic acid (CGA). In order to fulfill this goal, the optimization for the extraction of a phenolic extract from the hulls was investigated. The parameters studied were: type of solvent, solvent to water ratio and hull to solvent ratio. In addition, the solvent mixtures were also studied. The resulting phenolic extracts were evaluated for their biological activities. This included phenolic content determination, evaluation of the antioxidant and antimicrobial activities. Chlorogenic acid was determined in two chosen hull extracts using the UV spectrophotometric method and HPLC analysis. The anti carcinogenic activity of the two chosen extracts was tested on seven different cell line carcinomas. The results revealed that all the phenolic extracts of sunflower hull studied contain between 190-312.5 mg phenolics/ 100 g hulls. The highest phenolic extraction was achieved with 80% methanol (1:30, hull to solvent, w/v ratio) and methanol to ethanol to water (7:7:6 v/v/v) mixture with values of 312.5 and 306.5 mg phenolics/100 g hulls, respectively. The free radical scavenging activity and antioxidant activity of all the samples ranged from 33.6-72.6%. The highest antioxidant activity and free radical scavenging activity were achieved by the same extracts that possessed the highest phenolic content, namely methanol to ethanol to water extract and 80% methanol with values 71.8 and 72.6%, 68.2 and 70.9% respectively, compared to 77.9 and 76.9% respectively for TBHQ. All the phenolic extracts possessed antimicrobial activity but to different levels against different pathogenic bacteria. The two chosen extracts also possessed anti carcinogenic activity, which differed among varying cell line carcinomas. The HPLC analysis indicated that chlorogenic acid was the main phenolic acid in the extract. Thus it can

Anticancer potential of Hericium erinaceus extracts against particular human cancer cell lines

Directory of Open Access Journals (Sweden)

Younis AM

2017-06-01

Full Text Available Cancer is a leading cause of death worldwide. Cancer resulted in 8.2 million human deaths in 2012. It is expected that annual cancer cases will rise from 14 million in 2013 to 22 million within the next two decades. Mushrooms are extensively used as nutritional supplements in many countries. Moreover, mushrooms have many medicinal properties, including anticancer activity. In this study, the anticancer activity of different polar and non-polar extracts of Hericium erinaceus were evaluated against different human cancer cell lines including human liver carcinoma (Hep G2, the human colonic epithelial carcinoma (HCT 116, the human cervical cancer cells (HeLa and the human breast adenocarcinoma (MCF-7 using 3-(4,5-Dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT assay. Furthermore, as a control, the cytotoxicity effect of the different extracts were tested against isolated mouse hepatocytes. It was observed that the extracts by water and methanol from fresh and lyophilized fruiting bodies of H. erinaceus had the strongest anticancer effect. In contrast, the extracts by ether and ethyl acetate from mycelia and broth of H. erinaceus showed lower anticancer activity against the tested carcinoma cell lines. The highest anticancer activity was recorded for aqueous extract of lyophilized fruiting bodies with half maximal inhibitory concentration (IC50 values of 6.1±0.2, 5.1±0.1, 5.7±0.2 and 5.8±0.3 µg/ml against Hep G2, HCT 116, HeLa and MCF-7 cells, respectively with non-significant effect on the normal mouse hepatocytes. To summarise, polar extracts of H. erinaceus can be good sources for isolating natural anticancer compounds. I recommend further chemical studies to isolate the active principles of the extract of H. erinaceus evaluated in the present.