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Vitro Conditions (vitro + condition)
Selected AbstractsEvaluation of plant oils for suppression of crown rot disease and improvement of shelf life of banana (Musa spp.INTERNATIONAL JOURNAL OF FOOD SCIENCE & TECHNOLOGY, Issue 5 2010AAA subgroup, cv. Robusta) Summary Fourteen plant oils were evaluated to control the crown rot disease caused by Lasiodiplodia theobromae and Colletotrichum musae. Five of these, viz. Ocimum sanctum, Cymbopogan citratus, C. martinii, C. nardus and Pelargonium graveolens oils completely arrested the mycelial growth of both test pathogens at their lowest concentration compared to other oils. Besides, these plant oils have also inhibited the activity of cellulolytic and pectinolytic enzymes produced by these pathogens effectively under in vitro condition. The treatment of banana fruit var. Robusta (Cavendish-AAA) with oils of O. sanctum, C. citratus, C. nardus and C. martinii not only reduced the crown rot severity significantly, but also increased the shelf life of banana fruits. However, under low-temperature storage (14 °C) condition, O. sanctum oil increased the shelf life of banana fruits up to 48 days without affecting their organoleptic properties. Hence, O. sanctum oil could be used as an alternative to chemical fungicides for the management of crown rot disease. [source] Effect of Mechanical Stretching on Expressions of Muscle Specific Transcription Factors MyoD, Myf-5, Myogenin and MRF4 in Proliferated MyoblastsANATOMIA, HISTOLOGIA, EMBRYOLOGIA, Issue 4 2009S. Abe Summary We examined expression of four important members of myogenic regulatory factors (MRFs) in the myoblasts both at mRNA and protein levels, which were subjected to mechanical stretching in in vitro condition. Our results showed that MyoD expression existed both in the stretch and in the control group at all time periods of the mechanical stimulus. Myf-5 expressed only at early stage of the stretch group. Although mRNA and protein expressions of myogenin and MRF4 were detected both in the stretch and in the control group at 12 h after the stretching, their expressions were only shown in the stretch group at 24 h after the mechanical stimulus. However, at 36 and 48 h, none of the MRFs examined except MyoD appeared in both groups. Our results suggest that the MRFs are up-regulated upon mechanical stimulus and each member plays a different major role for either proliferation or differentiation of the myoblasts. [source] Osteogenic Evaluation of Glutaraldehyde Crosslinked Gelatin Composite with Fetal Rat Calvarial Culture ModelARTIFICIAL ORGANS, Issue 8 2001Hwa-Chang Liu Abstract: The cytotoxicity of the synthetic bone substitute composed of tricalcium phosphate and glutaraldehyde crosslinked gelatin (GTG) were evaluated by osteoblast cell culture. In a previous study, the GTG composites were soaked in distilled water for 1, 2, 4, 7, 14, 28, and 42 days, and then the solutions (or extracts) were cocultured with osteoblasts to evaluate the cytotoxicity of GTG composites by alive cell counting. In this study, the extracts were cocultured with the osteoblasts; thereafter, the concentration of transforming growth factor-, (TGF-,1) and prostaglandin E2 (PGE2) in the medium was analyzed to strictly reflect the biological effects of GTG composites on the growth of osteoblasts. In order to investigate the osteoconductive potential of the GTG composites on new bone formation in a relative short term, a model of neonatal rat calvarial organ culture was designed prior to animal experiments. Three experimental materials of 4, 8, and 12% GTG composites were evaluated by fetal rat calvarial organ culture for their ability for bone regeneration. Deproteinized bovine and porcine cancellous bone matrixes were used as the controlled materials. All the organ culture units were maintained in cultured medium for 5 weeks. Following the culture period, the morphology of tissue was observed under an optical microscope, and the quantitative evaluation of the new generation bone was determined by using a semiautomatic histomorphometeric method. Except in the initial 4 days, the concentration of TGF-,1 of 4% and 8% GTG composites was higher than that of the blank group for all the other experimental time periods. The PGE2 concentration for 4% and 8% GTG composites was lower than that of the blank group. It revealed that the 4% and 8% GTG composites would not lead to inflammation and would promote osteoblast growth. The morphology and activity of the osteoblasts were not transformed or changed by the 2 GTG composites. For the 12% GTG composite, the performance of the in vitro condition was inferior to the blank group and the other 2 GTG composites. Although the concentration of TGF-,1 and PGE2 was gradually back to normal after 14 days, the morphology of the osteoblasts was abnormal with features such as contracted cytoplast structures. The osteoblast was damaged perhaps in the initial stage. We suggested that the 4% and 8% GTG composites should be soaked in distilled water at least for 4 days before medical applications. The 12% GTG composite and the composites with a concentration of glutaraldehyde solution higher than 12% were not recommended as a medical prostheses in any condition. The fetal rat calvaria culture also showed the same results with the analysis of TGF-,1 and PGE2. From the study, we could predict the results of animal experiments in the future. [source] Villin-type headpiece domains show a wide range of F-actin-binding affinitiesCYTOSKELETON, Issue 1 2002D. Vardar Abstract The villin-type "headpiece" domain is a modular motif found at the extreme C-terminus of larger "core" domains in over 25 cytoskeletal proteins in plants and animals. Although headpiece is classified as an F-actin-binding domain, it has been suggested that some expressed fusion-proteins containing headpiece may lack F-actin-binding in vivo. To determine the intrinsic F-actin affinity of headpiece domains, we quantified the F-actin affinity of seven headpiece domains and three N-terminal truncations, under identical in vitro conditions. The constructs are folded and adopt the native headpiece structure. However, they show a wide range of affinities that can be grouped into high, low, and nonspecific-binding categories. Computer models of the structure and charged surface potential of these headpiece domains suggest features important for high F-actin affinity. We conclude that not all headpiece domains are intrinsically F-actin-binding motifs, and suggest that the surface charge distribution may be an important element for F-actin recognition. Cell Motil. Cytoskeleton 52:9,21, 2002. © 2002 Wiley-Liss, Inc. [source] d -Alanyl ester depletion of teichoic acids in Lactobacillus reuteri 100-23 results in impaired colonization of the mouse gastrointestinal tractENVIRONMENTAL MICROBIOLOGY, Issue 7 2007Jens Walter Summary The dlt operon of Gram-positive bacteria encodes proteins required for the incorporation of d -alanine esters into cell wall-associated teichoic acids (TA). d -Alanylation of TA has been shown to be important for acid tolerance, resistance to antimicrobial peptides, adhesion, biofilm formation, and virulence of a variety of pathogenic organisms. The aim of this study was to determine the importance of d -alanylation for colonization of the gastrointestinal tract by Lactobacillus reuteri 100-23. Insertional inactivation of the dltA gene resulted in complete depletion of d -alanine substitution of lipoteichoic acids. The dlt mutant had similar growth characteristics as the wild type under standard in vitro conditions, but formed lower population sizes in the gastrointestinal tract of ex- Lactobacillus -free mice, and was almost eliminated from the habitat in competition experiments with the parental strain. In contrast to the wild type, the dlt mutant was unable to form a biofilm on the forestomach epithelium during gut colonization. Transmission electron microscope observations showed evidence of cell wall damage of mutant bacteria present in the forestomach. The dlt mutant had impaired growth under acidic culture conditions and increased susceptibility to the cationic peptide nisin relative to the wild type. Ex vivo adherence of the dlt mutant to the forestomach epithelium was not impaired. This study showed that d -alanylation is an important cell function of L. reuteri that seems to protect this commensal organism against the hostile conditions prevailing in the murine forestomach. [source] Mechanism of DNA damage by cadmium and interplay of antioxidant enzymes and agentsENVIRONMENTAL TOXICOLOGY, Issue 2 2007Veera L. D. Badisa Abstract Cadmium is an environmental toxicant, which causes cancer in different organs. It was found that it damages DNA in the various tissues and cultured cell lines. To investigate the mechanism of DNA damage, we have studied the effect of cadmium-induced DNA damage in plasmid pBR322 DNA, and the possible ameliorative effects of antioxidative agents under in vitro conditions. It was observed that cadmium alone did not cause DNA damage. However, it caused DNA damage in the presence of hydrogen peroxide, in a dose dependent manner, because of production of hydroxyl radicals. Findings from this study show the conversion of covalently closed circular double-stranded pBR 322 DNA to the open circular and linear forms of DNA when treated with 10 ,M cadmium and various concentrations of H2O2. The conversion was due to nicking in DNA strands. The observed rate of DNA strand breakage was dependent on H2O2 concentration, temperature, and time. Metallothionein I failed to prevent cadmium-induced DNA nicking in the presence of H2O2. Of the two antioxidant enzymes (catalase and superoxide dismutase) studied, only catalase conferred significant (50,60%) protection. EDTA and DMSO exhibited protection similar to catalase, while mannitol showed only about 20% protection against DNA damage. Ethyl alcohol failed to ameliorate cadmium-induced DNA strands break. From this study, it is plausible to infer that cadmium in the presence of hydrogen peroxide causes DNA damage probably by the formation of hydroxyl ions. These results may indicate that cadmium in vivo could play a major role in the DNA damage induced by oxidative stress. © 2007 Wiley Periodicals, Inc. Environ Toxicol 22: 144,151, 2007. [source] Somatodendritic autoreceptor regulation of serotonergic neurons: dependence on l -tryptophan and tryptophan hydroxylase-activating kinasesEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 4 2005Rong-Jian Liu Abstract The somatodendritic 5-HT1A autoreceptor has been considered a major determinant of the output of the serotonin (5-HT) neuronal system. However, recent studies in brain slices from the dorsal raphe nucleus have questioned the relevance of 5-HT autoinhibition under physiological conditions. In the present study, we found that the difficulty in demonstrating 5-HT tonic autoinhibition in slice results from in vitro conditions that are unfavorable for sustaining 5-HT synthesis. Robust, tonic 5-HT1A autoinhibition can be restored by reinstating in vivo 5-HT synthesizing conditions with the initial 5-HT precursor l -tryptophan and the tryptophan hydroxylase co-factor tetrahydrobiopterin (BH4). The presence of tonic autoinhibition under these conditions was revealed by the disinhibitory effect of a low concentration of the 5-HT1A antagonist WAY 100635. Neurons showing an autoinhibitory response to l -tryptophan were confirmed immunohistochemically to be serotonergic. Once conditions for tonic autoinhibition had been established in raphe slice, we were able to show that 5-HT autoinhibition is critically regulated by the tryptophan hydroxylase-activating kinases calcium/calmodulin protein kinase II (CaMKII) and protein kinase A (PKA). In addition, at physiological concentrations of l -tryptophan, there was an augmentation of 5-HT1A receptor-mediated autoinhibition when the firing of 5-HT cells activated with increasing concentrations of the ,1 adrenoceptor agonist phenylephrine. Increased calcium influx at higher firing rates, by activating tryptophan hydroxylase via CaMKII and PKA, can work together with tryptophan to enhance negative feedback control of the output of the serotonergic system. [source] Human enamel dissolution in citric acid as a function of pH in the range 2.30,pH,6.30 , a nanoindentation studyEUROPEAN JOURNAL OF ORAL SCIENCES, Issue 3 2003Michele E. Barbour The objective of this study was to investigate the dissolution of human enamel in citric acid solutions over a wide range of pH. The in vitro conditions are considered to be relevant to soft drink-induced enamel erosion. Nanoindentation was used to investigate changes in the nanomechanical properties of polished enamel surfaces after exposure to citric acid solutions. Solutions used had 38.1 mmol l,1 citric acid and pH greater than 2.3 but less than 6.3 (2.30 pH 6.30). Samples were exposed to rapidly stirred, constant composition solutions for 120 s. Statistically significant changes in enamel hardness and reduced elastic modulus were observed after exposure to all solutions. There was an approximately linear dependence of enamel hardness on solution pH for 2.90 pH 6.30. Below pH 2.90, enamel is thought to have reached the lowest possible hardness value. The reduction in enamel dissolution caused by an increase in pH of a soft drink is likely to be small. Product modification to reduce the erosive potential of drinks may require additional methods such as addition of calcium salts. [source] Respiration of nitrous oxide in suboxic soilEUROPEAN JOURNAL OF SOIL SCIENCE, Issue 3 2009B. Vieten Summary Reduction of nitrous oxide (N2O) is an autonomous respiratory pathway. Nitrous oxide is an alternative electron acceptor to O2 when intensive biological activity and reduced diffusivity result in an O2 deficit. Hypoxic or anoxic micro sites may form even in well-aerated soils, and provide a sink for N2O diffusing through the gas-filled pore space. We reproduced similar in vitro conditions in suboxic (0.15% O2) flow-through incubation experiments with samples from a Stagnosol and from a Histosol. Apparent half-saturation constants (km) for N2O reduction were similar for both soils and were, on average, 3.8 ,mol mol,1 at 5°C, 5.1 ,mol mol,1 at 10°C, and 6.9 ,mol mol,1 at 20°C. Respiration of N2O was estimated to contribute a maximum proportion of 1.7% to total respiration in the Stagnosol (pH 7.0) and 0.9% in the Histosol (pH 2.9). [source] The polypeptide chain release factor eRF1 specifically contacts the s4UGA stop codon located in the A site of eukaryotic ribosomesFEBS JOURNAL, Issue 10 2001Laurent Chavatte It has been shown previously [Brown, C.M. & Tate, W.P. (1994) J. Biol. Chem.269, 33164,33170.] that the polypeptide chain release factor RF2 involved in translation termination in prokaryotes was able to photocrossreact with mini-messenger RNAs containing stop signals in which U was replaced by 4-thiouridine (s4U). Here, using the same strategy we have monitored photocrosslinking to eukaryotic ribosomal components of 14-mer mRNA in the presence of , and 42-mer mRNA in the presence of tRNAAsp (tRNAAsp gene transcript). We show that: (a) both 14-mer and 42-mer mRNAs crossreact with ribosomal RNA and ribosomal proteins. The patterns of the crosslinked ribosomal proteins are similar with both mRNAs and sensitive to ionic conditions; (b) the crosslinking patterns obtained with 42-mer mRNAs show characteristic modification upon addition of tRNAAsp providing evidence for appropriate mRNA phasing onto the ribosome. Similar changes are not detected with the 14-mer pairs; (c) when eukaryotic polypeptide chain release factor 1 (eRF1) is added to the ribosome·tRNAAsp complex it crossreacts with the 42-mer mRNA containing the s4UGA stop codon located in the A site, but not with the s4UCA sense codon; this crosslink involves the N-terminal and middle domains of eRF1 but not the C domain which interacts with eukaryotic polypeptide chain release factor 3 (eRF3); (d) addition of eRF3 has no effect on the yield of eRF1,42-mer mRNA crosslinking and eRF3 does not crossreact with 42-mer mRNA. These experiments delineate the in vitro conditions allowing optimal phasing of mRNA on the eukaryotic ribosome and demonstrate a direct and specific contact of ,core' eRF1 and s4UGA stop codon within the ribosomal A site. [source] Bioaccessibility studies of ferro-chromium alloy particles for a simulated inhalation scenario: A comparative study with the pure metals and stainless steelINTEGRATED ENVIRONMENTAL ASSESSMENT AND MANAGEMENT, Issue 3 2010Klara Midander Abstract The European product safety legislation, REACH, requires that companies that manufacture, import, or use chemicals demonstrate safe use and high level of protection of their products placed on the market from a human health and environmental perspective. This process involves detailed assessment of potential hazards for various toxicity endpoints induced by the use of chemicals with a minimum use of animal testing. Such an assessment requires thorough understanding of relevant exposure scenarios including material characteristics and intrinsic properties and how, for instance, physical and chemical properties change from the manufacturing phase, throughout use, to final disposal. Temporary or permanent adverse health effects induced by particles depend either on their shape or physical characteristics, and/or on chemical interactions with the particle surface upon human exposure. Potential adverse effects caused by the exposure of metal particles through the gastrointestinal system, the pulmonary system, or the skin, and their subsequent potential for particle dissolution and metal release in contact with biological media, show significant gaps of knowledge. In vitro bioaccessibility testing at conditions of relevance for different exposure scenarios, combined with the generation of a detailed understanding of intrinsic material properties and surface characteristics, are in this context a useful approach to address aspects of relevance for accurate risk and hazard assessment of chemicals, including metals and alloys and to avoid the use of in vivo testing. Alloys are essential engineering materials in all kinds of applications in society, but their potential adverse effects on human health and the environment are very seldom assessed. Alloys are treated in REACH as mixtures of their constituent elements, an approach highly inappropriate because intrinsic properties of alloys generally are totally different compared with their pure metal components. A large research effort was therefore conducted to generate quantitative bioaccessibility data for particles of ferro-chromium alloys compared with particles of the pure metals and stainless steel exposed at in vitro conditions in synthetic biological media of relevance for particle inhalation and ingestion. All results are presented combining bioaccessibility data with aspects of particle characteristics, surface composition, and barrier properties of surface oxides. Iron and chromium were the main elements released from ferro-chromium alloys upon exposure in synthetic biological media. Both elements revealed time-dependent release processes. One week exposures resulted in very small released particle fractions being less than 0.3% of the particle mass at acidic conditions and less than 0.001% in near pH-neutral media. The extent of Fe released from ferro-chromium alloy particles was significantly lower compared with particles of pure Fe, whereas Cr was released to a very low and similar extent as from particles of pure Cr and stainless steel. Low release rates are a result of a surface oxide with passive properties predominantly composed of chromium(III)-rich oxides and silica and, to a lesser extent, of iron(II,III)oxides. Neither the relative bulk alloy composition nor the surface composition can be used to predict or assess the extent of metals released in different synthetic biological media. Ferro-chromium alloys cannot be assessed from the behavior of their pure metal constituents. Integr Environ Assess Manag 2010;6:441,455. © 2009 SETAC [source] Differential response to phytoestrogens in endocrine sensitive and resistant breast cancer cells in vitroINTERNATIONAL JOURNAL OF CANCER, Issue 3 2006Jane L. Limer Abstract Women approaching menopause increasingly investigate alternatives to hormone replacement therapy. Plant phytoestrogens are being promoted as "natural" alternatives but there is a lack of substantive data to advocate their safe use in breast cancer patients receiving tamoxifen (TAM), or in those who have relapsed. The aim of our study was to investigate the proliferative effects and mode of action of the phytoestrogens genistein, daidzein and coumestrol on TAM-sensitive (-s) and resistant (-r) breast cancer cells under in vitro conditions designed to mimic the hormonal environment of the pre- and post-menopausal breast. At physiological concentrations (<10 ,M) and under reduced estrogen (E2) conditions, genistein was mitogenic to TAM-s cells with TAM-r cells generally refractory. Daidzein and coumestrol were growth stimulatory irrespective of TAM sensitivity. Transcriptional activity was ERE-mediated. Combining phytoestrogens with E2 (simulating the pre-menopausal breast environment) had no effect on growth of TAM-s or TAM-r cells. Addition of 4-HT mimicked the hormonal environment in post-menopausal breast cancer patients receiving TAM. The growth inhibitory effects of 4-HT were abrogated in TAM-s cells when combined with genistein and coumestrol, and to a lesser extent, daidzein, where significant growth stimulatory effects were observed. In TAM-r cells, proliferation did not exceed control values. At phytoestrogen concentrations above 10 ,M, growth inhibitory effects were seen, irrespective of estrogenic environment or cell sensitivity to TAM. Our in vitro data suggests that phytoestrogens could have potentially adverse mitogenic effects on tumour cells and should probably be avoided by patients who remain sensitive to TAM or in those with pre-existing and possibly undiagnosed breast tumours. © 2006 Wiley-Liss, Inc. [source] Influence of the dietary potassium content on transepithelial potassium transport in rat jejunumJOURNAL OF ANIMAL PHYSIOLOGY AND NUTRITION, Issue 3 2000R. Cermak Summary In a recent study, we found that the distal rat jejunum is able to secrete K+ under in vitro conditions. The question therefore arises as to whether the small intestine might participate in K+ homeostasis. Consequently, this study examined the influence of the dietary K+ content on transepithelial K+ transport in rat jejunum. Rats were fed two diets differing in K+ content (control diet 4.0 g K+/kg, low K+ diet (LK) 0.27 g K+/kg). After a minimal feeding period of 7 days, distal jejunal sheets were mounted in Ussing chambers and unidirectional 86Rb+ fluxes (as a marker for K+ transport) were measured under short-circuit conditions. Jejunum obtained from rats fed the control diet showed a net K+ secretion of 200 nmol Rb+/h/cm2. Unidirectional Rb+ fluxes were smaller in distal jejunum from rats fed the LK diet. In these tissues, glucose-induced short-circuit current and tissue conductance were also smaller than in controls. However, net Rb+ fluxes were not significantly different in small intestine from K+ -restricted rats compared with jejunum from control animals. Based on the observation that the dietary K+ content does not affect transepithelial net K+ transport, we conclude that transcellular K+ secretion by the small intestine is not involved in K+ homeostasis. [source] Response surface methodology study of the combined effects of temperature, pH, and aw on the growth rate of Trichoderma asperellumJOURNAL OF APPLIED MICROBIOLOGY, Issue 4 2007B.A.D. Begoude Abstract Aims:, To evaluate the influence of environmental parameters (water activity aw, temperature, and pH) on the radial growth rate of Trichoderma asperellum (strains PR10, PR11, PR12, and 659-7), an antagonist of Phytophthora megakarya, the causal agent of cocoa black pod disease. Methods and Results:, The radial growth of four strains of T. asperellum was monitored for 30 days on modified PDA medium. Six levels of aw (0·995, 0·980, 0·960, 0·930, 0·910, and 0·880) were combined with three values of pH (4·5, 6·5, and 8·5) and three incubation temperatures (20, 25, and 30°C). Whatever the strain, mycelial growth rate was optimal at aw between 0·995 and 0·980, independently of the temperature and pH. Each strain appeared to be very sensitive to aw reduction. In addition, all four strains were able to grow at all temperatures and pH values (4·5,8·5) tested, highest growth rate being observed at 30°C and at pH 4·5,6·5. The use of response surface methodology to model the combined effects of aw, temperature, and pH on the radial growth rate of the T. asperellum strains confirmed the observed results. In our model, growth of the T. asperellum strains showed a greater dependence on aw than on temperature or pH under in vitro conditions. Conclusion:,aw is a crucial environmental factor. Low aw can prevent growth of T. asperellum strains under some conditions. The observed and predicted radial growth rate of strain PR11 showed its greater capacity to support low aw (0·93) as compared with other tested strains at 20°C. This is in agreement with its better protective level when applied in medium-scale trials on cocoa plantations. Significance and Impact of the Study:, This study should contribute towards improving the biocontrol efficacy of T. asperellum strains used against P. megakarya. Integrated into a broader study of the impact of environmental factors on the biocontrol agent,pathogen system, this work should help to build a more rational control strategy, possibly involving the use of a compatible adjuvant protecting T. asperellum against desiccation. [source] Characterization of ACC deaminase gene in Pseudomonas entomophila strain PS-PJH isolated from the rhizosphere soilJOURNAL OF BASIC MICROBIOLOGY, Issue 2 2010Seralathan Kamala-Kannan Abstract The enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase cleaves the ethylene precursor ACC into ,-ketobutyrate and ammonia. The decreased level of ethylene allows the plant to be more resistant to a wide environmental stress including plant pathogens. In the present study, we characterized the ACC deaminase activity of a Pseudomonas entomophila strain PS-PJH isolated from the red pepper rhizosphere region of red pepper grown at Jinan, Korea. The isolate produced 23.8 ± 0.4 ,mol of ,-ketobutyrate/mg of protein/h during ACC deamination under in vitro conditions. Polymerase chain reaction for acdS gene showed that the isolated P. entomophila strain PS-PJH carry sequences similar to the known acdS genes. Results of the multiple sequence alignment revealed >99% identity (nucleotide and amino acid) with acdS gene of Pseudomonas putida strains AM15 and UW4. The isolated bacteria promoted 43.3 and 34.1% of growth in Raphanus sativus and Lactuca sativa plants, respectively. Based on the 16S,23S internal transcribed spacer region sequences, the isolate was identified as P. entomophila. To the best of our knowledge this is the first study to report the acdS gene in P. entomophila. (© 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [source] The role of thyroid hormone on phenylhydrazine hydrochloride mediated inhibitory effects on blood acetylcholinesterase: An in vivo and in vitro studyJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 4 2002Mitali Banerjee Abstract A novel phenomenon of protective counteraction by thyroid hormone has been demonstrated in phenylhydrazine hydrochloride (PHH) induced insult on blood acetylcholinesterase (AChE, EC 3.1.1.7) activity, in both, in vivo and in vitro conditions. Injection of PHH (20 ,g/g) to juvenile male rats for three consecutive days caused a 48% decrease (p < 0.001) in the total blood AChE activity on the third day (i.e. 24 h after injections for three consecutive days) in comparison to the control animals. Simultaneous injections of thyroxine (T4) 1 or 2 ,g/g with PHH (20 ,g/g) showed a recovery in AChE activity by 27% (p < 0.02) and 55% (p < 0.001), respectively, in comparison to the only PHH-injected animals. T4 at 1, 2 and 4 ,g/g doses showed unchanged levels in comparison to the untreated controls. In our in vitro system, incubations of the RBCs in PHH (2 mM) containing medium also showed an inhibition of 44% (p < 0.001) of the RBC membrane AChE activity in comparison to the control conditions. A recovery of 23,81% of the enzyme activity was observed after simultaneous use of T4 (1 nM,100 nM) or T3 (0.1 nM,100 nM), or triiodothyroacetic acid (TRIAC) (100 nM) with PHH (2 mM) in a dose-dependent manner with a potency profile of T3 > T4 > TRIAC. Incubation of RBCs only with T4, T3, or TRIAC at 0.1,100 nM concentration did not cause any alteration in the membrane AChE activity in comparison to control conditions. Thus, thyroid hormone distinctly demonstrated a counteraction or protective nature of action on the PHH-induced inhibition of total blood and RBC membrane AChE activity. © 2002 Wiley Periodicals, Inc. J Biochem Mol Toxicol 16:162,168, 2002; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.10039 [source] Comparison of potential protective effects of melatonin, indole-3-propionic acid, and propylthiouracil against lipid peroxidation caused by potassium bromate in the thyroid glandJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 1 2005Malgorzata Karbownik Abstract Potassium bromate (KBrO3) is a prooxidant and carcinogen, inducing thyroid tumors. Melatonin and indole-3-propionic acid (IPA) are effective antioxidants. Some antioxidative effects of propylothiouracil (PTU),a thyrostatic drug,have been found. The aim of the study was to compare protective effects of melatonin, IPA, and PTU against lipid peroxidation in the thyroids, collected from rats treated with KBrO3, and in homogenates of porcine thyroids, incubated in the presence of KBrO3. Wistar rats were administered KBrO3 (110 mg/kg b.w., i.p., on the 10th day of the experiment) and/or melatonin, or IPA (0.0645 mmol/kg b.w., i.p., twice daily, for 10 days), or PTU (0.025% solution in drinking water, for 10 days). Homogenates of porcine thyroids were incubated for 30 min in the presence of KBrO3 (5 mM) plus one of the antioxidants: melatonin (0.01, 0.1, 0.5, 1.0, 5.0, 7.5 mM), or IPA (0.01, 0.1, 0.5, 1.0, 5.0, 7.5, 10.0 mM), or PTU (0.01, 0.1, 0.5, 1.0, 5.0, 7.5, 10.0 mM). The level of lipid peroxidation products (MDA,+,4-HDA) was measured spectrophotometrically in thyroid homogenates. In vivo pretreatment with either melatonin or with IPA or with PTU decreased lipid peroxidation caused by KBrO3,injections in rat thyroid gland. Under in vitro conditions, PTU (5.0, 7.5, and 10.0 mM), but neither melatonin nor IPA, reduced KBrO3 -related lipid peroxidation in the homogenates of porcine thyroids. In conclusion, melatonin and IPA may be of great value as protective agents under conditions of exposure to KBrO3. © 2005 Wiley-Liss, Inc. [source] Conjugation of isometamidium chloride to antibodies and the use of the conjugate against the haemoflagellate, Cryptobia salmositica Katz, 1951: an immuno-chemotherapeutic strategyJOURNAL OF FISH DISEASES, Issue 8 2001B F Ardelli The trypanocidal drug isometamidium chloride (Samorin) was conjugated to polyclonal and monoclonal antibodies produced against the pathogenic haemoflagellate Cryptobia salmositica. Under in vitro conditions the unconjugated drug normally accumulates rapidly in the kinetoplast in the parasite; however, once it was conjugated to antibodies (either polyclonal or monoclonal) it was found throughout the parasite. Isometamidium conjugated to polyclonal antibodies lysed C. salmositica under in vitro conditions, but parasites were not agglutinated. In contrast, isometamidium conjugated to monoclonal antibodies (against a 200 kDa surface membrane glycoprotein) did not lyse C. salmositica, but parasites were agglutinated. Because of the low efficacy of the monoclonal conjugate against the parasite in vitro, its cryptobiocidal effect was not evaluated further. The infectivity of C. salmositica (incubated either in culture medium or whole blood) was reduced in fish after in vitro exposure to isometamidium conjugated to polyclonal antibodies. Parasitaemias were reduced in infected chinook salmon, Oncorhynchus tshawytscha, after treatment with isometamidium conjugated to polyclonal antibodies. [source] Use of Bean Sprout Enterobacteriaceae Isolates as Biological Control Agents of Pseudomonas fluorescensJOURNAL OF FOOD SCIENCE, Issue 1 2004K. ENOMOTOArticle first published online: 28 JUN 200 ABSTRACT: Bean sprouts were cultivated under in vitro conditions as a model system to study the mechanism of bacteria-mediated spoilage in bean sprouts. Pseudomonas fluorescens, or Erwinia spp., were inoculated onto sprouts at several stages during cultivation. Five strains of Enterobacteriaceae isolated from the native microflora of sprouts prevented Pseudomonas -mediated spoilage by co-inoculating these cultures on seeds that were soaking in water. The population of P. fluorescens in co-inoculated liquid medium culture with a strain (B1) decreased slightly. The results indicated that the Enterobacteriaceae isolates tested played an important role in preventing Pseudomonas -mediated spoilage by growing competitively with P. fluorescens. [source] The effect of oxygen tension on the in vitro assay of human osteoblastic connective tissue progenitor cellsJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 10 2008Sandra M. Villarruel Abstract Connective tissue progenitors (CTPs) are defined as the heterogeneous set of stem and progenitor cells that reside in native tissues and are capable of proliferation and differentiation into one or more connective tissue phenotypes. CTPs play important roles in tissue formation, repair, and remodeling. Therefore, in vitro assays of CTP prevalence and biological potential have important scientific and clinical relevance. This study evaluated oxygen tension as an important variable in optimizing in vitro conditions for quantitative assays of human CTPs. Bone marrow aspirates were collected from 20 human subjects and cultured using established medium conditions at ambient oxygen tensions of 1, 5, 10, and 20%. Colony-forming efficiency (CFE), proliferation, and colony density were assessed. CFE and proliferation were greatest at 5% O2. Traditional conditions using atmospheric oxygen tension (20% O2) reduced CFE by as much as 32%. CFE and proliferation at 1% O2 were less than 5% O2 but comparable to that seen at 20% O2, suggesting that CTPs are relatively resilient under hypoxic conditions, a fact that may be relevant to their function in wound repair and their potential use in tissue engineering applications involving transplantation into settings of moderate to severe hypoxia. These data demonstrate that optimization of quantitative assays for CTPs will require control of oxygen tension. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:1390,1397, 2008 [source] Osmotic Potential Effects on In Vitro Growth, Morphology and Pathogenicity of Macrophomina phaseolinaJOURNAL OF PHYTOPATHOLOGY, Issue 7-8 2003D. Cervantes-García Abstract The influence of osmotic potential (,s) on in vitro growth and morphology of four isolates of Macrophomina phaseolina was studied using potato-glucose-agar adjusted to different osmotic potentials with KCl, NaCl and sucrose. The effect of NaCl on M. phaseolina pathogenicity in common bean was determined and then related to the pathogenicity of each isolate on seeds of 12 differential common bean cultivars. The three sources of ,s reduced M. phaseolina growth under in vitro conditions, whereby NaCl caused the highest negative effects. The high solute concentrations reduced the synthesis of mycelial pigments and the size and shape of microsclerotia. Pathogenicity of M. phaseolina in common bean seeds was reduced by high concentrations of NaCl but no relationship was found between the tolerance to high ,s concentrations under in vitro conditions and M. phaseolina pathogenicity in common bean seeds. [source] Inhibitory effect of phenolics extracted from sorghum genotypes on Aspergillus parasiticus (NRRL 2999) growth and aflatoxin productionJOURNAL OF THE SCIENCE OF FOOD AND AGRICULTURE, Issue 6 2007CV Ratnavathi Abstract The inhibitory activity of bioactive polyphenols present in six sorghum genotypes,two red (AON 486 and IS 620), two yellow (LPJ and IS 17779) and two white (SPV 86 and SPV 462) varieties,on Aspergillus parasiticus (NRRL 2999) growth and aflatoxin production was evaluated. In the first experiment the production of aflatoxins in the six sorghum genotypes after removal of surface phenolics by acidic methanol treatment was studied and compared with that in untreated grains. Aflatoxin production was found to be fourfold higher in treated grains. The total phenols and bioactive polyphenols extracted by acidic methanol were quantified using the Folin,Denis method and the bovine serum albumin,benzidine conjugate procedure respectively. In the second experiment the effect of extracted sorghum phenolics under in vitro conditions on fungal growth and aflatoxin production was studied at two concentrations (0.01% and 0.1%) of phenolics. Extracted phenolics added to yeast extract sucrose (YES) medium at 0.1% concentration showed an inhibitory effect on aflatoxin production. At 0.01% phenolic concentration, aflatoxin production was minimal on day 3 after infection. At other time points the aflatoxin content was similar to that in the control. At 9 days after infection the fungal biomass in IS 620 was significantly lower than that in the control. At 0.1% phenolic concentration, aflatoxin production was minimal and the red genotype IS 620 showed maximum resistance. Fungal biomass was lowest at all growth stages in IS 620 as compared with the control. Polyphenol oxidase (PPO) activity was not detected in A. parasiticus grown on YES medium (control). PPO activity was not induced in A. parasiticus by the addition of phenolics to the liquid culture medium (no PPO activity was detected in the culture medium). The inhibitory activity of bioactive polyphenols could be attributed to the lack of PPO enzyme in this fungus. Copyright © 2007 Society of Chemical Industry [source] Effect of tannic acid on in vitro enzymatic hydrolysis of some protein sourcesJOURNAL OF THE SCIENCE OF FOOD AND AGRICULTURE, Issue 5 2003Tomás F Martínez Abstract The pH-stat system has been used to assess the effect of tannic acid (TA) on solubility and in vitro enzyme hydrolysis of different proteins. Added TA (from 10 to 50 g kg,1) decreased the extent of hydrolysis of bovine serum albumin. Enzymic hydrolysis of casein, pea meal, soybean meal, and haemoglobin (HB) was increased, as measured by total amino acids released and by the degree of hydrolysis. SDS-PAGE confirmed the results of the in vitro enzymatic hydrolysis. These findings suggest that, under in vitro conditions, when simulating the gastrointestinal environment of domestic mammals, the negative effects of TA described from in vivo experiments are not necessarily due to reduced hydrolysis of proteins. Copyright © 2003 Society of Chemical Industry [source] How does polyspermy happen in mammalian oocytes?MICROSCOPY RESEARCH AND TECHNIQUE, Issue 4 2003Wei-Hua Wang Abstract Polyspermy is one of the most commonly observed abnormal types of fertilization in mammalian oocytes. In vitro fertilization (IVF) provides approaches to study the mechanisms by which oocytes block polyspermic fertilization. Accumulated data indicate that oocyte, sperm and insemination conditions are all related to the occurrence of polyspermic fertilization. A high proportion of immature and aged oocytes showed polyspermy as compared with mature oocytes. Preincubation of oocytes and/or sperm with oviductal epithelial cells or collected oviductal fluid before IVF reduces polyspermic penetration. Recently, it was found that an abnormal zona pellucida is one of main causes of polyspermy in human eggs. A high proportion of polyspermy has resulted from the use of a high concentration of capacitated spermatozoa at the site of fertilization, irrespective of in the in vivo or in vitro environment. Oviductal secretions or oviductal epithelial cells themselves can regulate the number of spermatozoa reaching or binding to the zona pellucida thus reducing multiple sperm penetration. Suboptimal in vitro conditions, such as supplementations in IVF media, pH, and temperature during IVF, also induce polyspermic fertilization in some mammals. Species-specific differences are present regarding the relationship between insemination conditions and polyspermy. Microsc. Res. Tech. 61:335,341, 2003. © 2003 Wiley-Liss, Inc. [source] Cross-species hybridization of a Borrelia burgdorferi DNA array reveals infection- and culture-associated genes of the unsequenced genome of the relapsing fever agent Borrelia hermsiiMOLECULAR MICROBIOLOGY, Issue 3 2004Jianmin Zhong Summary The known genome sequence of Borrelia burgdorferi, an agent of Lyme borreliosis, was used to study the genetic content and gene expression in B. hermsii, another spirochete pathogen and a cause of relapsing fever. Cross-species hybridization of a DNA array representing 1628 open reading frames (ORF) of B. burgdorferi with genomic DNA of B. hermsii indicated that the latter organism has at least 81% of the chromosomal genes and 43% of the plasmid genes of B. burgdorferi. We then carried out quantitative hybridization of the arrays with multiple replicates of cDNA produced from B. hermsii cells growing in the blood of infected mice or in culture medium that was adjusted to the same pH, temperature and a spirochete density as infected blood. Of 642 B. burgdorferi ORFs hybridized by all replicates under both conditions, 12 (1.9%) demonstrated differential expression by a regularized t -test and stringent criteria. BBP07 and BBG30, two plasmid-borne ORFs with the greatest measurable difference in expression between in vivo and in vitro conditions, putatively encode proteins of unknown function. Orthologues of BBP07 in B. hermsii were identified, and increased expression in infected mice was demonstrated by quantitative reverse-transcriptase polymerase chain reaction. [source] Higher expression of hyaluronan binding protein 1 (HABP1/p32/gC1qR/SF2) during follicular development and cumulus oocyte complex maturation in ratMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 3 2008Sonu Chand Thakur Abstract Ovulation is a complex process of releasing a fertilizable oocyte and depends on the proper formation of an extracellular hyaluronan rich matrix by the cumulus oocyte complex (COC). The formation of a HA rich matrix is dependent on the synthesis and organization of HA in the presence of several biomolecules that mediate its crosslinking. To gain an insight into the follicular maturation and COC expansion, we have studied the expression of hyaluronan binding protein 1 (HABP1), which is known to interact specifically with hyaluronan. The level of HABP1 increased markedly during ovulation after gonadotropin stimulation, and the overexpression was seen in mural granulosa cells, expanding cumulus cells and follicular fluid. However, HABP1 could not be detected in the luteal cells of corpus luteum after ovulation. Such increased expression of HABP1 was observed both during in vivo and in vitro conditions of COC expansion. The level of HABP1 transcript was upregulated up to fivefold after COC expansion as compared to compact COC. Immunofluorescence analysis showed HABP1 to be localized in the cytoplasm and extracellular matrix, suggesting its role in ECM organization. The cultured expanded COC treated with hyaluronidase for different time periods showed the gradual dispersion of COC, which coincide with the loss of HABP1 from the matrix suggesting that HABP1 is bound to hyaluronan. These results indicate that HABP1 expressed in rat COCs during maturation may facilitate the formation of the HA matrix in the extracellular space around the oocyte with cumulus expansion during maturation. Mol. Reprod. Dev. 75: 429,438, 2008. © 2007 Wiley-Liss, Inc. [source] Photodynamic effect of hypericin in primary cultures of human umbilical endothelial cells and glioma cell linesPHYTOTHERAPY RESEARCH, Issue 6 2009Viktória Stupáková Abstract Hypericin is the most powerful naturally occurring photosensitizer and as such there is renaissant interest in the potentials of this compound for anticancer photodynamic therapy (PDT). The purpose of this study was to investigate the hypericin-mediated photodynamic therapy effects on normal human umbilical endothelial cells (HUVECs) in comparison with cancer human glioma cell lines U-87 MG and U-373 MG, in in vitro conditions. The data suggest that endothelial cells as well as glioma cell lines are sensitive only to photoactivated hypericin. The inhibitory effects of photoactivated hypericin did not differ in endothelial compared with tumor cells in cytotoxicity MTT and DNA fragmentation assays. However, an important difference in sensitivity was found between the above mentioned cell types in migration and metalloproteinases inhibition assays performed as cell function tests. The findings in both function tests were supported by the high sensitivity of endothelial cells in an additional angiogenesis test of tubular formation in vitro. Copyright © 2009 John Wiley & Sons, Ltd. [source] In vivo and in vitro activities of the seed extract of Piper guineense Schum. and Thonn. against skin and gill monogenean parasites of gold,sh (Carassius auratus auratus)PHYTOTHERAPY RESEARCH, Issue 10 2004A. P. Ekanem Abstract Methanol extracts of the seeds of Piper guineense (Piperaceae) were active against gold,sh (Carassius auratus auratus L. Pisces Cyprinidae) monogenean parasites. The seed extract of P. guineense was administered at different concentrations (0.5,2.0 mg/L) under in vivo and in vitro conditions. There was a higher ef,cacy of the effects of the extracts against ,sh parasites under in vitro situations than under in vivo. Three major compounds (piperanine, N -isobutyl (E,E)-2,4 decadienamide and ,,,, -dihydrowasanine) were identi,ed from the seed extract of Piper guineense by LC-MS analysis. Copyright © 2004 John Wiley & Sons, Ltd. [source] Protective effect of green tea polyphenol (-)-epigallocatechin gallate and other antioxidants on lipid peroxidation in gerbil brain homogenatesPHYTOTHERAPY RESEARCH, Issue 3 2003Seong-Ryong Lee Abstract The aim of this study was to compare the protective effects of green tea polyphenol (-)-epigallocatechin gallate (EGCG) and other well-known antioxidants on the lipid peroxidation in gerbil brain homogenates. Oxidative stress was induced by H2O2 (10 mM) or ferrous ammonium sulfate (5 µM) and lipid peroxidation was studied. Hydrogen peroxide and ferrous ions are capable of oxidizing a wide range of substrates and causing biological damage. The reaction, referred to as the Fenton process, is complex and can generate both hydroxyl radicals and higher oxidation states of the iron. Thiobarbituric acid-reactive substances (TBA-RS) were used as a marker of lipid peroxidation. EGCG, trolox, lipoic acid, and melatonin reduced H2O2 - or ferrous ion-induced lipid peroxidation in a concentration-dependant manner. In reducing the H2O2 -induced lipid peroxidation, IC50 values of antioxidants were as follows: EGCG (0.66 µM), trolox (37.08 µM), lipoic acid (7.88 mM), and melatonin (19.11 mM). In reducing the ferrous ion-induced lipid peroxidation, IC50 values of antioxidants were as follows: EGCG (3.32 µM), trolox (75.65 µM), lipoic acid (7.63 mM), and melatonin (15.48 mM). Under the in vitro conditions of this experiment, EGCG was the most potent antioxidant in inhibiting H2O2 or ferrous ion-induced lipid peroxidation in the gerbil brain homogenates. Copyright © 2003 John Wiley & Sons, Ltd. [source] Co-action of temperature and phosphate in inducing turion formation in Spirodela polyrhiza (Great duckweed),PLANT CELL & ENVIRONMENT, Issue 9 2002K.-J. Appenroth Abstract Increased phosphate concentration, higher temperature and addition of glucose all increased the number of fronds and turions of the duckweed Spirodela polyrhiza formed under in vitro conditions. Increasing the number of turions by increasing the plant biomass does not mean that the developmental process (switch of the programme of the primordia from vegetative fronds toward resting turions) has been specifically influenced. The specific turion yield (STY; number of turions formed by one frond) and the time of onset of turion formation have been used as more specific measures of turion induction. At more than 30 µm initial phosphate the STY was increased by lower temperature (15 °C) and became independent of the phosphate concentration. Between 10 and 30 µm and at higher temperatures (25 °C) the STY was increased by lower phosphate levels. The stimulatory effect of lower temperature was more pronounced than that of lower phosphate concentrations. Decreased phosphate concentration highly accelerated the formation of the first turions. The influence of low temperature was small at lower phosphate concentration but became dominant at higher concentrations (especially in autotrophic cultures). Low phosphate levels (e.g. 10 µm) and low temperatures (e.g. 15 °C) both represent specific turion-inducing factors having significant interactive effects. In S. polyrhiza, these signals may replace the interactive effects of photoperiods and low temperature known from other hydrophytes in turion induction under natural conditions. [source] | ||||||||||||||||||||||||||||||||||