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Sample records for inhibits in-vitro candida

  1. A novel antifungal is active against Candida albicans biofilms and inhibits mutagenic acetaldehyde production in vitro.

    PubMed

    Nieminen, Mikko T; Novak-Frazer, Lily; Rautemaa, Vilma; Rajendran, Ranjith; Sorsa, Timo; Ramage, Gordon; Bowyer, Paul; Rautemaa, Riina

    2014-01-01

    The ability of C. albicans to form biofilms is a major virulence factor and a challenge for management. This is evident in biofilm-associated chronic oral-oesophageal candidosis, which has been shown to be potentially carcinogenic in vivo. We have previously shown that most Candida spp. can produce significant levels of mutagenic acetaldehyde (ACH). ACH is also an important mediator of candidal biofilm formation. We have also reported that D,L-2-hydroxyisocaproic acid (HICA) significantly inhibits planktonic growth of C. albicans. The aim of the present study was to investigate the effect of HICA on C. albicans biofilm formation and ACH production in vitro. Inhibition of biofilm formation by HICA, analogous control compounds or caspofungin was measured using XTT to measure biofilm metabolic activity and PicoGreen as a marker of biomass. Biofilms were visualised by scanning electron microscopy (SEM). ACH levels were measured by gas chromatography. Transcriptional changes in the genes involved in ACH metabolism were measured using RT-qPCR. The mean metabolic activity and biomass of all pre-grown (4, 24, 48 h) biofilms were significantly reduced after exposure to HICA (p<0.05) with the largest reductions seen at acidic pH. Caspofungin was mainly active against biofilms pre-grown for 4 h at neutral pH. Mutagenic levels (>40 µM) of ACH were detected in 24 and 48 h biofilms at both pHs. Interestingly, no ACH production was detected from D-glucose in the presence of HICA at acidic pH (p<0.05). Expression of genes responsible for ACH catabolism was up-regulated by HICA but down-regulated by caspofungin. SEM showed aberrant hyphae and collapsed hyphal structures during incubation with HICA at acidic pH. We conclude that HICA has potential as an antifungal agent with ability to inhibit C. albicans cell growth and biofilm formation. HICA also significantly reduces the mutagenic potential of C. albicans biofilms, which may be important when treating bacterial-fungal biofilm

  2. In vitro modification of Candida albicans invasiveness.

    PubMed

    Fontenla de Petrino, S E; de Jorrat, M E; Sirena, A; Valdez, J C; Mesón, O

    1986-05-01

    Candida albicans produces germ-tubes (GT) when it is incubated in animal or human serum. This dimorphism is responsible for its invasive ability. The purpose of the present paper is (1) to evaluate the ability of rat peritoneal macrophages to inhibit GT production of ingested Candida albicans, obtained from immunized rats and then activated in vitro with Candida-induced lymphokines; (2) to determinate any possible alteration of phagocytic and candidacidal activities. The phagocytes were obtained from rats immunized with viable C. albicans. Some of them were exposed to Candida-induced lymphokines in order to activate the macrophages in vitro. The monolayers of activated, immune and normal macrophages were infected with a C. albicans suspension during 4 hr. Activated macrophages presented not only the highest phagocytic and candidacidal activities but a noticeable inhibition of GT formation and incremented candidacidal activity.

  3. In vitro modification of Candida albicans invasiveness.

    PubMed

    Fontenla de Petrino, S E; de Jorrat, M E; Sirena, A; Valdez, J C; Mesón, O

    1986-05-01

    Candida albicans produces germ-tubes (GT) when it is incubated in animal or human serum. This dimorphism is responsible for its invasive ability. The purpose of the present paper is (1) to evaluate the ability of rat peritoneal macrophages to inhibit GT production of ingested Candida albicans, obtained from immunized rats and then activated in vitro with Candida-induced lymphokines; (2) to determinate any possible alteration of phagocytic and candidacidal activities. The phagocytes were obtained from rats immunized with viable C. albicans. Some of them were exposed to Candida-induced lymphokines in order to activate the macrophages in vitro. The monolayers of activated, immune and normal macrophages were infected with a C. albicans suspension during 4 hr. Activated macrophages presented not only the highest phagocytic and candidacidal activities but a noticeable inhibition of GT formation and incremented candidacidal activity. PMID:3523254

  4. Chitin biosynthesis in Candida albicans grown in vitro and in vivo and its inhibition by nikkomycin Z.

    PubMed Central

    Chapman, T; Kinsman, O; Houston, J

    1992-01-01

    An N-acetyl-D-[14C]glucosamine radiolabel incorporation assay has been used to monitor chitin biosynthesis in whole cells of Candida albicans both in vitro and in vivo in two different mouse infection models, one using the peritoneal cavity as a chamber in which to add and retrieve cells and the other using infected kidneys. Specific labeling of chitin in alkali-insoluble material was confirmed by chitinase digestion, analysis of acid hydrolysates, and the use of nikkomycin Z as a probe. Nikkomycin Z was shown to strongly inhibit chitin biosynthesis in C. albicans grown in vitro and in vivo in both models. This demonstrates that nikkomycin Z-susceptible chitin synthase activity is present in C. albicans when the fungus is in its pathogenic state in vivo. The limited use of nikkomycin as a therapeutic agent is discussed. PMID:1416881

  5. Specific inhibition of in vitro Candida-induced lymphocyte proliferation by polysaccharidic antigens present in the serum of patients with chronic mucocutaneous candidiasis.

    PubMed Central

    Fischer, A; Ballet, J J; Griscelli, C

    1978-01-01

    A specific inhibitory activity of in vitro proliferative responses of normal human lymphocytes to Candida metabolic antigen was found in the serum of 6 out of 23 children with chronic mucocutaneous candidiasis. In each of the six patients, the presence of an inhibitory activity was associated with Candida-specific cellular defects, characterized by a negative-skin test and a lack of in vitro lymphocyte proliferation. The presence of a circulating inhibitor was detected during relapses of the disease and disappeared under antifungal therapy. This inhibitory effect was not associated with any toxicity on tested lymphocytes. The factor was shown to be nondialysable, thermostable, nonprecipitable with ammonium sulfate and absorbable on anti-Candida antibodies or concanavalin A-coupled agarose columns. Altogether, these results suggest that the inhibitory factor is not an immunoglobulin, but rather a polysaccharidic antigen of Candida albicans. An inhibition of Candida-induced proliferative response of normal human lymphocytes was also obtained by addition of polysacharide antigens or purified mannans from C. albicans to cultures. Candida polysaccharidic antigens appeared, therefore, to be involved in specific depression of cellular functions observed in chronic candidiasis. PMID:361754

  6. Bifidobacterium longum-fermented broccoli supernatant inhibited the growth of Candida albicans and some pathogenic bacteria in vitro.

    PubMed

    Suido, Hirohisa; Miyao, Manabu

    2008-06-01

    The aim of this study is to develop a growth inhibitory material against some pathogenic microorganisms, using beneficial bacteria such as Bifidobacterium species and certain types of vegetables which can be good substrates for the growth of the beneficial bacteria. At first, various vegetable juices were screened for the growth promotion of Bifidobacterium longum etc. Among the vegetables tested, broccoli (Brassica oleracea L. var. botrytis L.) and cabbage (Brassica oleracea L. var. capitata L.) showed excellent growth promoting activities for B. longum. Secondly, the B. longum-fermented broccoli (BFB) and Lactobacillus pentosus-fermented broccoli (LFB) supernatants were prepared and the growth inhibitory activities against Candida albicans were determined. Both of them showed dose-dependent, growth inhibitory effects, and the effect of BFB was superior to LFB. It was thought that the superior effect of BFB could be mainly attributed to the acids, especially acetic acid, produced by B. longum. BFB also inhibited some pathogenic bacteria such as Streptococcus mutans and Porphylomonas gingivalis. In conclusion, broccoli was found to be a good growth-promoting substance for B. longum. The fermented product, BFB, appears to be a usable material that inhibits the growth of C. albicans and some pathogenic bacteria. PMID:18661679

  7. Symbiotic Plant Peptides Eliminate Candida albicans Both In Vitro and in an Epithelial Infection Model and Inhibit the Proliferation of Immortalized Human Cells

    PubMed Central

    Ördögh, Lilla; Vörös, Andrea; Nagy, István; Kondorosi, Éva

    2014-01-01

    The increasing number of multidrug-resistant microbes now emerging necessitates the identification of novel antimicrobial agents. Plants produce a great variety of antimicrobial peptides including hundreds of small, nodule-specific cysteine-rich NCR peptides that, in the legume Medicago truncatula, govern the differentiation of endosymbiotic nitrogen fixing bacteria and, in vitro, can display potent antibacterial activities. In this study, the potential candidacidal activity of 19 NCR peptides was investigated. Cationic NCR peptides having an isoelectric point above 9 were efficient in killing Candida albicans, one of the most common fungal pathogens of humans. None of the tested NCR peptides were toxic for immortalized human epithelial cells at concentrations that effectively killed the fungus; however, at higher concentrations, some of them inhibited the division of the cells. Furthermore, the cationic peptides successfully inhibited C. albicans induced human epithelial cell death in an in vitro coculture model. These results highlight the therapeutic potential of cationic NCR peptides in the treatment of candidiasis. PMID:25243129

  8. Synthetic antimicrobial β-peptide in dual-treatment with fluconazole or ketoconazole enhances the in vitro inhibition of planktonic and biofilm Candida albicans.

    PubMed

    Mora-Navarro, Camilo; Caraballo-León, Jean; Torres-Lugo, Madeline; Ortiz-Bermúdez, Patricia

    2015-12-01

    Fungal infections are a pressing concern for human health worldwide, particularly for immunocompromised individuals. Current challenges such as the elevated toxicity of common antifungal drugs and the emerging resistance towards these could be overcome by multidrug therapy. Natural antimicrobial peptides, AMPs, in combination with other antifungal agents are a promising avenue to address the prevailing challenges. However, they possess limited biostability and susceptibility to proteases, which has significantly hampered their development as antifungal therapies. β-peptides are synthetic materials designed to mimic AMPs while allowing high tunability and increased biostability. In this work, we report for the first time the inhibition achieved in Candida albicans when treated with a mixture of a β-peptide model and fluconazole or ketoconazole. This combination treatment enhanced the biological activity of these azoles in planktonic and biofilm Candida, and also in a fluconazole-resistant strain. Furthermore, the in vitro cytotoxicity of the dual treatment was evaluated towards the human hepatoma cell line, HepG2, a widely used model derived from liver tissue, which is primarily affected by azoles. Analyses based on the LA-based method and the mass-action law principle, using a microtiter checkerboard approach, revealed synergism of the combination treatment in the inhibition of planktonic C. albicans. The dual treatment proved to be fungicidal at 48 and 72 h. Interestingly, it was also found that the viability of HepG2 was not significantly affected by the dual treatments. Finally, a remarkable enhancement in the inhibition of the highly azole-resistant biofilms and fluconazole resistant C. albicans strain was obtained.

  9. In Vitro Models for Candida Biofilm Development.

    PubMed

    Krom, Bastiaan P; Willems, Hubertine M E

    2016-01-01

    Development of Candida spp. biofilms on medical devices such as catheters and voice prosthesis has been recognized as an increasing clinical problem. Different in vitro models are presented with increasing complexity. Each model system can be utilized for analysis of new active compounds to prevent or treat Candida biofilms as well as to study molecular processes involved in biofilm formation. Susceptibility studies of clinical isolates are generally performed in a simple 96-well model system similar to the CLSI standard. In the present chapter, optimized conditions that promote biofilm formation within individual wells of microtiter plates are described. In addition, the method has proven useful in preparing C. albicans biofilms for investigation by a variety of microscopic and molecular techniques. A more realistic and more complex biofilm system is presented by the Amsterdam Active Attachment (AAA) model. In this 24-well model all crucial steps of biofilm formation: adhesion, proliferation, and maturation, can be simulated on various surfaces, while still allowing a medium throughput approach. This model has been applied to study susceptibility, complex molecular mechanisms as well as interspecies (Candida-bacterium) interactions. Finally, a realistic microfluidics channel system is presented to follow dynamic processes in biofilm formation. In this Bioflux-based system, molecular mechanisms as well as dynamic processes can be studied at a high time-resolution.

  10. In Vitro Anti-Candida Activity of Zataria multiflora Boiss

    PubMed Central

    Dabbagh, Muhammad Ali; Fouladi, Zahra

    2007-01-01

    Zataria multiflora Boiss known as Avishan Shirazi (in Iran) is one of the valuable Iranian medicinal plants. The aim of study was to evaluate anti-Candida activity of Z. multiflora against different species of Candida in vitro. Anti-Candida activity of the aqueous, ethanolic and methanolic maceration extract of the aerial parts of Z. multiflora Boiss was studied in vitro. Anti-Candida activity against Candida species was done using serial dilutions of extracts in Sabouraud's dextrose agar. Minimal inhibitory concentration (MIC) of the methanolic and ethanolic extracts was 70.7 and 127 mg l−1, respectively. Aqueous extract showed no remarkable activity against Candida species. We conclude that methanolic extract of the aerial parts of Z. multiflora Boiss has more anti-Candida effect at 70.7 mg l−1 compared to ethanolic extract 127 mg l−1. In addition, the isolates of Candida parapsilosis were more susceptible to methanolic extract than other tested species. PMID:17965766

  11. In Vitro Activity of Caspofungin against Candida albicans Biofilms

    PubMed Central

    Bachmann, Stefano P.; VandeWalle, Kacy; Ramage, Gordon; Patterson, Thomas F.; Wickes, Brian L.; Graybill, John R.; López-Ribot, José L.

    2002-01-01

    Most manifestations of candidiasis are associated with biofilm formation on biological or inanimate surfaces. Candida albicans biofilms are recalcitrant to treatment with conventional antifungal therapies. Here we report on the activity of caspofungin, a new semisynthetic echinocandin, against C. albicans biofilms. Caspofungin displayed potent in vitro activity against sessile C. albicans cells within biofilms, with MICs at which 50% of the sessile cells were inhibited well within the drug's therapeutic range. Scanning electron microscopy and confocal scanning laser microscopy were used to visualize the effects of caspofungin on preformed C. albicans biofilms, and the results indicated that caspofungin affected the cellular morphology and the metabolic status of cells within the biofilms. The coating of biomaterials with caspofungin had an inhibitory effect on subsequent biofilm development by C. albicans. Together these findings indicate that caspofungin displays potent activity against C. albicans biofilms in vitro and merits further investigation for the treatment of biofilm-associated infections. PMID:12384370

  12. Growth inhibition of Candida by human oral epithelial cells.

    PubMed

    Steele, C; Leigh, J; Swoboda, R; Fidel, P L

    2000-11-01

    Oropharyngeal candidiasis (OPC) caused by Candida albicans is a significant problem in human immunodeficiency virus (HIV)-infected persons. Recognizing the paucity of information on innate and/or adaptive mucosal host defenses against C. albicans, we recently reported that human and nonhuman primate and mouse vaginal epithelial cells inhibit the growth of C. albicans in vitro. In the present study, oral epithelial cells collected from saliva of healthy volunteers and a purified oral epithelial cell line were found to inhibit blastoconidia and/or hyphal growth of several Candida species. Cell contact was a strict requirement for the epithelial cell anti-Candida activity; neither saliva nor culture supernatants alone inhibited Candida growth, and addition of saliva to the coculture did not modulate the epithelial cell activity. Finally, epithelial cell anti-Candida activity was significantly lower in HIV-infected persons with OPC. Together, these results suggest that oral epithelial cells may play a role in innate resistance against OPC.

  13. Sodium houttuyfonate and EDTA-Na₂ in combination effectively inhibits Pseudomonas aeruginosa, Staphylococcus aureus and Candida albicans in vitro and in vivo.

    PubMed

    Huang, Weifeng; Duan, Qiangjun; Li, Fang; Shao, Jing; Cheng, Huijuan; Wu, Daqiang

    2015-01-01

    Sodium houttuyfonate (SH) has been found to be mildly anti-pathogenetic against the planktonic and biofilm phenotypes of Pseudomonas aeruginosa. Here, we have attempted to investigate further the anti-pathogenicity of SH and EDTA-Na₂ in combination and in vitro and in vivo against planktonic and biofilm phenotypes of Pseudomonas aeruginosa (16 strains), Staphylococcus aureus (13 strains) and Candida albicans (13 strains). The antimicrobial activity of SH against all three pathogens increased dramatically when it was combined with EDTA-Na₂, in vitro. Toxic reactions to the drugs when administered orally were insignificant in mice; no abnormalities were observed in the internal organs, such as the lungs and kidneys. Finally, the results of in vivo studies indicate that SH could extend the lifespan of infected animals when administered in combination with EDTA-Na2. Therefore, the results of the present study lead us to suggest that SH could be a promising antimicrobial agent and that SH combined with EDTA-Na₂ has the potential to be an excellent choice of drug for combating bacterial infections clinically. PMID:25467165

  14. Sodium houttuyfonate and EDTA-Na₂ in combination effectively inhibits Pseudomonas aeruginosa, Staphylococcus aureus and Candida albicans in vitro and in vivo.

    PubMed

    Huang, Weifeng; Duan, Qiangjun; Li, Fang; Shao, Jing; Cheng, Huijuan; Wu, Daqiang

    2015-01-01

    Sodium houttuyfonate (SH) has been found to be mildly anti-pathogenetic against the planktonic and biofilm phenotypes of Pseudomonas aeruginosa. Here, we have attempted to investigate further the anti-pathogenicity of SH and EDTA-Na₂ in combination and in vitro and in vivo against planktonic and biofilm phenotypes of Pseudomonas aeruginosa (16 strains), Staphylococcus aureus (13 strains) and Candida albicans (13 strains). The antimicrobial activity of SH against all three pathogens increased dramatically when it was combined with EDTA-Na₂, in vitro. Toxic reactions to the drugs when administered orally were insignificant in mice; no abnormalities were observed in the internal organs, such as the lungs and kidneys. Finally, the results of in vivo studies indicate that SH could extend the lifespan of infected animals when administered in combination with EDTA-Na2. Therefore, the results of the present study lead us to suggest that SH could be a promising antimicrobial agent and that SH combined with EDTA-Na₂ has the potential to be an excellent choice of drug for combating bacterial infections clinically.

  15. In vitro pharmacodynamic modelling of anidulafungin against Candida spp.

    PubMed

    Gil-Alonso, Sandra; Jauregizar, Nerea; Ortega, Ignacio; Eraso, Elena; Suárez, Elena; Quindós, Guillermo

    2016-03-01

    The aim of this study was to fit anidulafungin in vitro static time-kill data from nine strains of Candida with a pharmacodynamic (PD) model in order to describe the antifungal activity of this drug against Candida spp. Time-kill data from strains of Candida albicans, Candida glabrata and Candida parapsilosis clades were best fit using an adapted sigmoidal Emax model and resulted in a set of PD parameters (Emax, EC50 and Hill factor) for each fungal strain. The data were analysed with NONMEM 7. Anidulafungin was effective in a species- and concentration-dependent manner against the strains of C. glabrata and C. parapsilosis clades as observed with the EC50 estimates. Maximum killing rate constant (Emax) values were higher against C. glabrata and C. parapsilosis complex strains. In conclusion, we demonstrated that the activity of anidulafungin against Candida can be accurately described using an adapted sigmoidal Emax model.

  16. In vitro activity of eugenol against Candida albicans biofilms.

    PubMed

    He, Miao; Du, Minquan; Fan, Mingwen; Bian, Zhuan

    2007-03-01

    Most manifestations of candidiasis are associated with biofilm formation occurring on the surfaces of host tissues and medical devices. Candida albicans is the most frequently isolated causative pathogen of candidiasis, and the biofilms display significantly increased levels of resistance to the conventional antifungal agents. Eugenol, the major phenolic component of clove essential oil, possesses potent antifungal activity. The aim of this study was to investigate the effects of eugenol on preformed biofilms, adherent cells, subsequent biofilm formation and cell morphogenesis of C. albicans. Eugenol displayed in vitro activity against C. albicans cells within biofilms, when MIC(50) for sessile cells was 500 mg/L. C. albicans adherent cell populations (after 0, 1, 2 and 4 h of adherence) were treated with various concentrations of eugenol (0, 20, 200 and 2,000 mg/L). The extent of subsequent biofilm formation were then assessed with the tetrazolium salt reduction assay. Effect of eugenol on morphogenesis of C. albicans cells was observed by scanning electron microscopy (SEM). The results indicated that the effect of eugenol on adherent cells and subsequent biofilm formation was dependent on the initial adherence time and the concentration of this compound, and that eugenol can inhibit filamentous growth of C. albicans cells. In addition, using human erythrocytes, eugenol showed low hemolytic activity. These results indicated that eugenol displayed potent activity against C. albicans biofilms in vitro with low cytotoxicity and therefore has potential therapeutic implication for biofilm-associated candidal infections. PMID:17356790

  17. In vitro shoot growth of Brugmansia x candida Pers.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to improve the growth of in vitro shoot cultures of Brugmansia × candida 'Creamsickle'. Several mineral nutrient experiments were conducted to determine the effect of NH4+, NO3-, K+, FeSO4/EDTA, ZnSO4, MnSO4, and CuSO4 on quality, leaf width and length, size and weigh...

  18. The In Vitro Antimicrobial Activities of Metabolites from Lactobacillus Strains on Candida Species Implicated in Candida Vaginitis

    PubMed Central

    Ogunshe, Adenike A O; Omotoso, Mopelola A; Bello, Victoria B

    2011-01-01

    Background: Research from developing countries, such as Nigeria, on Lactobacillus species in the female urogenital tract and their role as a barrier to vaginal infection is limited. Therefore, the aim of this study was to assess the clinical biotherapeutic potential of indigenous Lactobacillus species. Methods: Antimicrobial metabolites production were characterised using simple and easily reproducible qualitative and quantitative methods. The in vitro inhibitory effect of Lactobacillus antimicrobials on vulvovaginal candidiasis–associated Candida species was investigated using modified agar spot and agar well-diffusion methods. Results: The maximum levels of lactic acid, hydrogen peroxide, and diacetyl from 20 vaginal Lactobacillus strains from diseased subjects were 1.46 mg/L, 1.36 mmol/L, and 1.72 mg/L respectively. From the 4 healthy subjects, the maximum level of lactic acid was 1.08 mg/L; hydrogen peroxide, 1.36 mmol/L; and diacetyl, 0.86 mg/L. The maximum productions of these substances occurred between 72 and 120 hours of incubation. The in vitro antagonistic activities of vaginal L. acidophilus, L. fermentum, L. brevis, L. plantarum, L. casei, L. delbrueckii, and L. jensenii from diseased subjects inhibited a maximum of 5.71% of the 35 Candida species tested, while vaginal L. acidophilus and L. plantarum from healthy subjects inhibited between 57.1% and 68.6% of Candida species in vitro. Conclusion: Antimicrobial-producing lactobacilli can be considered as adjunct biotherapeutic candidates for the treatment of vulvovaginal candidiasis. PMID:22589669

  19. Chrysin Inhibits Lymphangiogenesis in Vitro.

    PubMed

    Prangsaengtong, Orawin; Athikomkulchai, Sirivan; Xu, Jiuxiang; Koizumi, Keiichi; Inujima, Akiko; Shibahara, Naotoshi; Shimada, Yutaka; Tadtong, Sarin; Awale, Suresh

    2016-01-01

    The induction of lymphangiogenesis is an important process to promote cancer growth and cancer metastasis via the lymphatic system. Identifying the compounds that can prevent lymphangiogenesis for cancer therapy is urgently required. Chrysin, 5,7-dihydroxyflavone, a natural flavone extracted from Thai propolis, was used to investigate the effect on the lymphangiogenesis process of TR-LE, rat lymphatic endothelial cells. In this study, maximal nontoxic doses of chrysin on TR-LE cells were selected by performing a proliferation assay. The process of lymphangiogenesis in vitro was determined by cord formation assay, adhesion assay and migration assay. Chrysin at a nontoxic dose (25 μM) significantly inhibited cord formation, cell adhesion and migration of TR-LE cells when compared with the control group. We also found that chrysin significantly induced vascular endothelial growth factor C (VEGF-C) mRNA expression and nitric oxide (NO) production in TR-LE cells which was involved in decreasing the cord formation of TR-LE cells. In conclusion, we report for the first time that chrysin inhibited the process of lymphangiogenesis in an in vitro model. This finding may prove to be a natural compound for anti-lymphangiogenesis that could be developed for use in cancer therapy. PMID:27040620

  20. In vitro avarol does affect the growth of Candida sp.

    PubMed

    Pejin, Boris; Ciric, Ana; Markovic, Dejan; Tommonaro, Giuseppina; Sokovic, Marina

    2016-09-01

    This work extends in vitro screening of antimicrobial activity of avarol, the marine natural product firstly isolated from the Mediterranean sponge Dysidea avara. Its anticandidial activity was evaluated by microdilution method against eight Candida strains, two ATCC and six clinical ones. At a different extent this compound was proven to be active against all the strains tested (MIC 0.8-6.0 μg/mL and MFC 1.6-12.0 μg/mL, respectively). According to the best of our knowledge, this is the first report on avarol activity towards any yeast strain which may be of relevance for Alzheimer's disease. Indeed, avarol derivatives showing moderate AChE activity should be screened for anticandidial activity both in vitro and in vivo. PMID:26405848

  1. Oral administration of the broad-spectrum antibiofilm compound toremifene inhibits Candida albicans and Staphylococcus aureus biofilm formation in vivo.

    PubMed

    De Cremer, Kaat; Delattin, Nicolas; De Brucker, Katrijn; Peeters, Annelies; Kucharíková, Soña; Gerits, Evelien; Verstraeten, Natalie; Michiels, Jan; Van Dijck, Patrick; Cammue, Bruno P A; Thevissen, Karin

    2014-12-01

    We here report on the in vitro activity of toremifene to inhibit biofilm formation of different fungal and bacterial pathogens, including Candida albicans, Candida glabrata, Candida dubliniensis, Candida krusei, Pseudomonas aeruginosa, Staphylococcus aureus, and Staphylococcus epidermidis. We validated the in vivo efficacy of orally administered toremifene against C. albicans and S. aureus biofilm formation in a rat subcutaneous catheter model. Combined, our results demonstrate the potential of toremifene as a broad-spectrum oral antibiofilm compound. PMID:25288093

  2. The in vitro activity of vaginal Lactobacillus with probiotic properties against Candida.

    PubMed

    Strus, Magdalena; Kucharska, Agnieszka; Kukla, Grazyna; Brzychczy-Włoch, Monika; Maresz, Katarzyna; Heczko, Piotr B

    2005-06-01

    Lactobacilli, the predominant vaginal microorganisms in healthy premenopausal women, control other members of the vaginal microflora and thus protect against bacterial vaginosis and urinary tract infections. It has been claimed that some lactobacilli are also protective against Candida vaginitis. Little is known, however, about the mechanisms by which these lactobacilli can control vaginal populations of Candida and prevent vaginitis. To address this question, vaginal Lactobacillus strains with known antagonistic properties against bacteria were tested for their cell surface properties, adhesion to vaginal cell lines in vitro and antagonistic activities against Candida. A small proportion of the lactobacilli tested adhered strongly to cultured vaginal epithelial cells and inhibited growth of Candida albicans but not of C. pseudotropicalis. This anticandidal activity was in some Lactobacillus strains related to hydrogen peroxide (H2O2) production, but catalase treatment did not suppress this activity in other Lactobacillus strains, suggesting alternative mechanism(s). Moreover, tested vaginal Candida strains were resistant to relatively high concentrations of H2O2 that markedly exceeded those produced by even the most active Lactobacillus strains.

  3. Polyketide Glycosides from Bionectria ochroleuca Inhibit Candida albicans Biofilm Formation

    PubMed Central

    2015-01-01

    One of the challenges presented by Candida infections is that many of the isolates encountered in the clinic produce biofilms, which can decrease these pathogens’ susceptibilities to standard-of-care antibiotic therapies. Inhibitors of fungal biofilm formation offer a potential solution to counteracting some of the problems associated with Candida infections. A screening campaign utilizing samples from our fungal extract library revealed that a Bionectria ochroleuca isolate cultured on Cheerios breakfast cereal produced metabolites that blocked the in vitro formation of Candida albicans biofilms. A scale-up culture of the fungus was undertaken using mycobags (also known as mushroom bags or spawn bags), which afforded four known [TMC-151s C–F (1–4)] and three new [bionectriols B–D (5–7)] polyketide glycosides. All seven metabolites exhibited potent biofilm inhibition against C. albicans SC5314, as well as exerted synergistic antifungal activities in combination with amphotericin B. In this report, we describe the structure determination of the new metabolites, as well as compare the secondary metabolome profiles of fungi grown in flasks and mycobags. These studies demonstrate that mycobags offer a useful alternative to flask-based cultures for the preparative production of fungal secondary metabolites. PMID:25302529

  4. In vitro interactions between anidulafungin and nonsteroidal anti-inflammatory drugs on biofilms of Candida spp.

    PubMed

    Rosato, Antonio; Catalano, Alessia; Carocci, Alessia; Carrieri, Antonio; Carone, Addolorata; Caggiano, Giuseppina; Franchini, Carlo; Corbo, Filomena; Montagna, Maria Teresa

    2016-03-01

    Candida spp. are responsible for many biomaterial-related infections; they give rise to infective pathologies typically associated with biofilm formation. We recently reported that the echinocandin anidulafungin (ANF) showed a strong in vitro activity against both planktonic and biofilms cells. Herein, we report the antifungal activities of ANF alone and in association with some non-steroidal anti-inflammatory drugs (NSAIDs) against nine Candida strain biofilms: four Candida albicans, two Candida glabrata and three Candida guilliermondii. The activity of ANF was assessed using an in vitro microbiological model relevant for clinical practice. ANF proved oneself to be active against biofilms cells, and a clear-cut synergism was found against Candida species biofilms when ANF was used in combination with three NSAIDs: aspirin, diclofenac, ibuprofen. The positive synergism against Candida spp. of ANF in association with aspirin or the other NSAIDs proved to be a very effective antifungal treatment (FICI<0.5). These results may provide the starting point for new combination therapies of ANF with NSAIDs against Candida biofilm pathologies.

  5. In vitro synergism of simvastatin and fluconazole against Candida species.

    PubMed

    Menezes, Everardo Albuquerque; Vasconcelos Júnior, Antônio Alexandre de; Silva, Carlla Lorena Façanha; Plutarco, Fábio Ximenes; Cunha, Maria da Conceição dos Santos Oliveira; Cunha, Francisco Afrânio

    2012-08-01

    Systemic fungal infections are responsible for high mortality rates. Several species of fungi may be involved, but Candida spp. is the most prevalent. Simvastatin is used to lower cholesterol and also exhibits antifungal action. The aim of this study was to evaluate the synergistic action of simvastatin with fluconazole against strains of Candida spp. Susceptibility testing was performed according to protocol M27-A3, by broth microdilution method and the synergistic effect of simvastatin and fluconazole was calculated based on FICI (Fractional Inhibitory Concentration Index). Eleven strains were evaluated, and simvastatin showed a synergistic effect with fluconazole against 10 (91%) of the Candida spp. strains tested. Simvastatin may be a valuable drug in the treatment of systemic infections caused by Candida spp.

  6. Praziquantel inhibits Schistosoma mansoni attachment in vitro.

    PubMed

    da-Silva, S P; Noel, F

    1990-01-01

    Male adult Schistosoma mansoni worms were placed in a glass dish containing Tyrode solution and observed for 15 min after addition of praziquantel (0.01 to 1 microM). Praziquantel promoted a concentration- and time-dependent inhibition of sucker-mediated attachment of the worm. Attachment inhibition was correlated with shortening of the parasite. We propose that the rapid and total inhibition of worm attachment observed in vitro with 1 microM praziquantel indicates that therapeutic concentrations of this drug should promote a rapid hepatic shift, in vivo, which may facilitate host tissue reaction. PMID:2101049

  7. Histone Deacetylases and Their Inhibition in Candida Species

    PubMed Central

    Garnaud, Cécile; Champleboux, Morgane; Maubon, Danièle; Cornet, Muriel; Govin, Jérôme

    2016-01-01

    Fungi are generally benign members of the human mucosal flora or live as saprophytes in the environment. However, they can become pathogenic, leading to invasive and life threatening infections in vulnerable patients. These invasive fungal infections are regarded as a major public health problem on a similar scale to tuberculosis or malaria. Current treatment for these infections is based on only four available drug classes. This limited therapeutic arsenal and the emergence of drug-resistant strains are a matter of concern due to the growing number of patients to be treated, and new therapeutic strategies are urgently needed. Adaptation of fungi to drug pressure involves transcriptional regulation, in which chromatin dynamics and histone modifications play a major role. Histone deacetylases (HDACs) remove acetyl groups from histones and actively participate in controlling stress responses. HDAC inhibition has been shown to limit fungal development, virulence, biofilm formation, and dissemination in the infected host, while also improving the efficacy of existing antifungal drugs toward Candida spp. In this article, we review the functional roles of HDACs and the biological effects of HDAC inhibitors on Candida spp., highlighting the correlations between their pathogenic effects in vitro and in vivo. We focus on how HDAC inhibitors could be used to treat invasive candidiasis while also reviewing recent developments in their clinical evaluation. PMID:27547205

  8. Histone Deacetylases and Their Inhibition in Candida Species.

    PubMed

    Garnaud, Cécile; Champleboux, Morgane; Maubon, Danièle; Cornet, Muriel; Govin, Jérôme

    2016-01-01

    Fungi are generally benign members of the human mucosal flora or live as saprophytes in the environment. However, they can become pathogenic, leading to invasive and life threatening infections in vulnerable patients. These invasive fungal infections are regarded as a major public health problem on a similar scale to tuberculosis or malaria. Current treatment for these infections is based on only four available drug classes. This limited therapeutic arsenal and the emergence of drug-resistant strains are a matter of concern due to the growing number of patients to be treated, and new therapeutic strategies are urgently needed. Adaptation of fungi to drug pressure involves transcriptional regulation, in which chromatin dynamics and histone modifications play a major role. Histone deacetylases (HDACs) remove acetyl groups from histones and actively participate in controlling stress responses. HDAC inhibition has been shown to limit fungal development, virulence, biofilm formation, and dissemination in the infected host, while also improving the efficacy of existing antifungal drugs toward Candida spp. In this article, we review the functional roles of HDACs and the biological effects of HDAC inhibitors on Candida spp., highlighting the correlations between their pathogenic effects in vitro and in vivo. We focus on how HDAC inhibitors could be used to treat invasive candidiasis while also reviewing recent developments in their clinical evaluation. PMID:27547205

  9. Miltefosine is effective against Candida albicans and Fusarium oxysporum nail biofilms in vitro.

    PubMed

    Machado Vila, Taissa Vieira; Sousa Quintanilha, Natália; Rozental, Sonia

    2015-11-01

    Onychomycosis is a fungal nail infection that represents ∼50 % of all nail disease cases worldwide. Clinical treatment with standard antifungals frequently requires long-term systemic therapy to avoid chronic disease. Onychomycosis caused by non-dermatophyte moulds, such as Fusarium spp., and yeasts, such as Candida spp., is particularly difficult to treat, possibly due to the formation of drug-resistant fungal biofilms on affected areas. Here, we show that the alkylphospholipid miltefosine, used clinically against leishmaniasis and cutaneous breast metastases, has potent activity against biofilms of Fusarium oxysporum and Candida albicans formed on human nail fragments in vitro. Miltefosine activity was compared with that of commercially available antifungals in the treatment of biofilms at two distinct developmental phases: formation and maturation (pre-formed biofilms). Drug activity towards biofilms formed on nail fragments and on microplate surfaces (microdilution assays) was evaluated using XTT [2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide] assays, and drug effects on fingernail biofilms were analysed by scanning electron microscopy (SEM). For F. oxysporum, miltefosine at 8 μg ml- 1 inhibited biofilm formation by 93%, whilst 256 μg ml- 1 reduced the metabolic activity of pre-formed nail biofilms by 93%. Treatment with miltefosine at 1000 μg ml- 1 inhibited biofilm formation by 89% and reduced the metabolic activity of pre-formed C. albicans biofilms by 99%. SEM analyses of biofilms formed on fingernail fragments showed a clear reduction in biofilm biomass after miltefosine treatment, in agreement with XTT results. Our results show that miltefosine has potential as a therapeutic agent against onychomycosis and should be considered for in vivo efficacy studies, especially in topical formulations for refractory disease treatment. PMID:26404553

  10. Miltefosine is effective against Candida albicans and Fusarium oxysporum nail biofilms in vitro.

    PubMed

    Machado Vila, Taissa Vieira; Sousa Quintanilha, Natália; Rozental, Sonia

    2015-11-01

    Onychomycosis is a fungal nail infection that represents ∼50 % of all nail disease cases worldwide. Clinical treatment with standard antifungals frequently requires long-term systemic therapy to avoid chronic disease. Onychomycosis caused by non-dermatophyte moulds, such as Fusarium spp., and yeasts, such as Candida spp., is particularly difficult to treat, possibly due to the formation of drug-resistant fungal biofilms on affected areas. Here, we show that the alkylphospholipid miltefosine, used clinically against leishmaniasis and cutaneous breast metastases, has potent activity against biofilms of Fusarium oxysporum and Candida albicans formed on human nail fragments in vitro. Miltefosine activity was compared with that of commercially available antifungals in the treatment of biofilms at two distinct developmental phases: formation and maturation (pre-formed biofilms). Drug activity towards biofilms formed on nail fragments and on microplate surfaces (microdilution assays) was evaluated using XTT [2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide] assays, and drug effects on fingernail biofilms were analysed by scanning electron microscopy (SEM). For F. oxysporum, miltefosine at 8 μg ml- 1 inhibited biofilm formation by 93%, whilst 256 μg ml- 1 reduced the metabolic activity of pre-formed nail biofilms by 93%. Treatment with miltefosine at 1000 μg ml- 1 inhibited biofilm formation by 89% and reduced the metabolic activity of pre-formed C. albicans biofilms by 99%. SEM analyses of biofilms formed on fingernail fragments showed a clear reduction in biofilm biomass after miltefosine treatment, in agreement with XTT results. Our results show that miltefosine has potential as a therapeutic agent against onychomycosis and should be considered for in vivo efficacy studies, especially in topical formulations for refractory disease treatment.

  11. Simvastatin inhibits planktonic cells and biofilms of Candida and Cryptococcus species.

    PubMed

    Brilhante, Raimunda Sâmia Nogueira; Caetano, Erica Pacheco de; Oliveira, Jonathas Sales; Castelo-Branco, Débora de Souza Collares Maia; Souza, Elizabeth Ribeiro Yokobatake; Alencar, Lucas Pereira de; Cordeiro, Rossana de Aguiar; Bandeira, Tereza de Jesus Pinheiro Gomes; Sidrim, José Júlio Costa; Rocha, Marcos Fábio Gadelha

    2015-01-01

    The antifungal activity of some statins against different fungal species has been reported. Thus, at the first moment, the in vitro antifungal activity of simvastatin, atorvastatin and pravastatin was tested against Candida spp. and Cryptococcus spp. Then, in a second approach, considering that the best results were obtained for simvastatin, this drug was evaluated in combination with antifungal drugs against planktonic growth and tested against biofilms of Candida spp. and Cryptococcus spp. Drug susceptibility testing was performed using the microdilution broth method, as described by the Clinical and Laboratory Standards Institute. The interaction between simvastatin and antifungals against planktonic cells was analyzed by calculating the fractional inhibitory concentration index. Regarding biofilm susceptibility, simvastatin was tested against growing biofilm and mature biofilm of one strain of each tested yeast species. Simvastatin showed inhibitory effect against Candida spp. and Cryptococcus spp. with minimum inhibitory concentration values ranging from 15.6 to 1000 mg L(-1) and from 62.5 to 1000 mg L(-1), respectively. The combination of simvastatin with itraconazole and fluconazole showed synergism against Candida spp. and Cryptococcus spp., while the combination of simvastatin with amphotericin B was synergistic only against Cryptococcus spp. Concerning the biofilm assays, simvastatin was able to inhibit both growing biofilm and mature biofilm of Candida spp. and Cryptococcus spp. The present study showed that simvastatin inhibits planktonic cells and biofilms of Candida and Cryptococcus species.

  12. [In vitro nystatin sensitivity of vaginal isolates of Candida spp].

    PubMed

    Andreu, C M; Medina, Y E; Gonzáles, T C; Llanes, D M

    2001-01-01

    The minimum inhibitory concentration (MIC) of nistatine, one of the most used antifungal agents for this micosis, was determined in 68 Candida strains isolated from vaginal smears. Candida albicans represented 75% of the total strains whereas C. parapsilosis, C. krusei and C. glabrata were much less frequently found. The predisposing factors were pregnancy and antibacterial treatment whereas leukorrhea and itching were the prevailing symptoms in most of the cases. MIC values from the use of a broth dilution method ranged from 0,5-8mg/mL and the geometric mean was 1.36mg/ mL. For C. albicans, MIC was 4mg/mL due to two strains that showed the highest MIC values (8 mg/mL). Similarly, the strains showed low MIC values, this means that therapeutic failures are not inherent to the emergence of resistant strains.

  13. In vitro activities of natural products against oral Candida isolates from denture wearers

    PubMed Central

    2011-01-01

    Background Candida-associated denture stomatitis is a frequent infectious disease. Treatment of this oral condition is difficult because failures and recurrences are common. The aim of this study was to test the in vitro antifungal activity of pure constituents of essentials oils. Methods Eight terpenic derivatives (carvacrol, farnesol, geraniol, linalool, menthol, menthone, terpinen-4-ol, and α-terpineol), a phenylpropanoid (eugenol), a phenethyl alcohol (tyrosol) and fluconazole were evaluated against 38 Candida isolated from denture-wearers and 10 collection Candida strains by the CLSI M27-A3 broth microdilution method. Results Almost all the tested compounds showed antifungal activity with MIC ranges of 0.03-0.25% for eugenol and linalool, 0.03-0.12% for geraniol, 0.06-0.5% for menthol, α-terpineol and terpinen-4-ol, 0.03-0.5% for carvacrol, and 0.06-4% for menthone. These compounds, with the exception of farnesol, menthone and tyrosol, showed important in vitro activities against the fluconazole-resistant and susceptible-dose dependent Candida isolates. Conclusions Carvacrol, eugenol, geraniol, linalool and terpinen-4-ol were very active in vitro against oral Candida isolates. Their fungistatic and fungicidal activities might convert them into promising alternatives for the topic treatment of oral candidiasis and denture stomatitis. PMID:22118215

  14. Ocimum sanctum essential oil inhibits virulence attributes in Candida albicans.

    PubMed

    Khan, Amber; Ahmad, Aijaz; Xess, Immaculata; Khan, Luqman A; Manzoor, Nikhat

    2014-03-15

    Candida albicans is an opportunistic human fungal pathogen which causes disease mainly in immunocompromised patients. Activity of hydrolytic enzymes is essential for virulence of C. albicans and so is the capacity of these cells to undergo transition from yeast to mycelial form of growth. Ocimum sanctum is cultivated worldwide for its essential oil which exhibits medicinal properties. This work evaluates the anti-virulence activity of O. sanctum essential oil (OSEO) on 22 strains of C. albicans (including a standard strain ATCC 90028) isolated from both HIV positive and HIV negative patients. Candida isolates were exposed to sub-MICs of OSEO. In vitro secretion of proteinases and phospholipases was evaluated by plate assay containing BSA and egg yolk respectively. Morphological transition from yeast to filamentous form was monitored microscopically in LSM. For genetic analysis, respective genes associated with morphological transition (HWP1), proteinase (SAP1) and phospholipase (PLB2) were also investigated by Real Time PCR (qRT-PCR). Results were analyzed using Student's t-test. OSEO inhibits morphological transition in C. albicans and had a significant inhibitory effect on extracellular secretion of proteinases and phospholipases. Expression profile of respective selected genes associated with C. albicans virulence by qRT-PCR showed a reduced expression of HWP1, SAP1 and PLB2 genes in cells treated with sub-inhibitory concentrations of OSEO. This work suggests that OSEO inhibits morphological transition in C. albicans and decreases the secretion of hydrolytic enzymes involved in the early stage of infection as well as down regulates the associated genes. Further studies will assess the clinical application of OSEO and its constituents in the treatment of fungal infections. PMID:24252340

  15. In vitro phagocytosis of several Candida berkhout species by murine leukocytes.

    PubMed

    Fontenla de Petrino, S E; Bibas Bonet de Jorrat, M E; Sirena, A

    1985-03-01

    In vitro phagocytosis of thirteen Candida berkhout species by rat leukocytes was studied to assess a possible correlation between pathogenicity and phagocytosis Yeast-leukocyte suspensions were mixed up for 3 h and phagocytic index, germ-tube formation and leukocyte candidacidal activity were evaluated. Highest values for phagocytosis were reached in all cases at the end of the first hour. Leukocyte candidacidal activity was absent. Only C. albicans produced germ-tubes. The various phagocytosis indices were determined depending on the Candida species assayed. Under these conditions, the more pathogenic species presented the lower indices of phagocytosis. It is determined that the in vitro phagocytic index may bear a close relationship with the pathogenicity of the Candida berkhout.

  16. In vitro phagocytosis of several Candida berkhout species by murine leukocytes.

    PubMed

    Fontenla de Petrino, S E; Bibas Bonet de Jorrat, M E; Sirena, A

    1985-03-01

    In vitro phagocytosis of thirteen Candida berkhout species by rat leukocytes was studied to assess a possible correlation between pathogenicity and phagocytosis Yeast-leukocyte suspensions were mixed up for 3 h and phagocytic index, germ-tube formation and leukocyte candidacidal activity were evaluated. Highest values for phagocytosis were reached in all cases at the end of the first hour. Leukocyte candidacidal activity was absent. Only C. albicans produced germ-tubes. The various phagocytosis indices were determined depending on the Candida species assayed. Under these conditions, the more pathogenic species presented the lower indices of phagocytosis. It is determined that the in vitro phagocytic index may bear a close relationship with the pathogenicity of the Candida berkhout. PMID:3990766

  17. In Vitro and In Vivo Activity of a Novel Antifungal Small Molecule against Candida Infections

    PubMed Central

    Yuen, Kwok Yong; Wang, Yu; Yang, Dan; Samaranayake, Lakshman Perera

    2014-01-01

    Candida is the most common fungal pathogen of humans worldwide and has become a major clinical problem because of the growing number of immunocompromised patients, who are susceptible to infection. Moreover, the number of available antifungals is limited, and antifungal-resistant Candida strains are emerging. New and effective antifungals are therefore urgently needed. Here, we discovered a small molecule with activity against Candida spp. both in vitro and in vivo. We screened a library of 50,240 small molecules for inhibitors of yeast-to-hypha transition, a major virulence attribute of Candida albicans. This screening identified 20 active compounds. Further examination of the in vitro antifungal and anti-biofilm properties of these compounds, using a range of Candida spp., led to the discovery of SM21, a highly potent antifungal molecule (minimum inhibitory concentration (MIC) 0.2 – 1.6 µg/ml). In vitro, SM21 was toxic to fungi but not to various human cell lines or bacterial species and was active against Candida isolates that are resistant to existing antifungal agents. Moreover, SM21 was relatively more effective against biofilms of Candida spp. than the current antifungal agents. In vivo, SM21 prevented the death of mice in a systemic candidiasis model and was also more effective than the common antifungal nystatin at reducing the extent of tongue lesions in a mouse model of oral candidiasis. Propidium iodide uptake assay showed that SM21 affected the integrity of the cell membrane. Taken together, our results indicate that SM21 has the potential to be developed as a novel antifungal agent for clinical use. PMID:24465737

  18. In vitro and in vivo activity of a novel antifungal small molecule against Candida infections.

    PubMed

    Wong, Sarah Sze Wah; Kao, Richard Yi Tsun; Yuen, Kwok Yong; Wang, Yu; Yang, Dan; Samaranayake, Lakshman Perera; Seneviratne, Chaminda Jayampath

    2014-01-01

    Candida is the most common fungal pathogen of humans worldwide and has become a major clinical problem because of the growing number of immunocompromised patients, who are susceptible to infection. Moreover, the number of available antifungals is limited, and antifungal-resistant Candida strains are emerging. New and effective antifungals are therefore urgently needed. Here, we discovered a small molecule with activity against Candida spp. both in vitro and in vivo. We screened a library of 50,240 small molecules for inhibitors of yeast-to-hypha transition, a major virulence attribute of Candida albicans. This screening identified 20 active compounds. Further examination of the in vitro antifungal and anti-biofilm properties of these compounds, using a range of Candida spp., led to the discovery of SM21, a highly potent antifungal molecule (minimum inhibitory concentration (MIC) 0.2-1.6 µg/ml). In vitro, SM21 was toxic to fungi but not to various human cell lines or bacterial species and was active against Candida isolates that are resistant to existing antifungal agents. Moreover, SM21 was relatively more effective against biofilms of Candida spp. than the current antifungal agents. In vivo, SM21 prevented the death of mice in a systemic candidiasis model and was also more effective than the common antifungal nystatin at reducing the extent of tongue lesions in a mouse model of oral candidiasis. Propidium iodide uptake assay showed that SM21 affected the integrity of the cell membrane. Taken together, our results indicate that SM21 has the potential to be developed as a novel antifungal agent for clinical use. PMID:24465737

  19. Comparison of the in vitro Effect of Chemical and Herbal Mouthwashes on Candida albicans

    PubMed Central

    Talebi, Somayeh; Sabokbar, Azar; Riazipour, Majid; Saffari, Mohsen

    2014-01-01

    Background: During the recent decades research has focused to find scientific evidence for the effects of herbal medicines. Researchers are interested in herbal remedies for medication and aim to substitute herbal material instead of chemical formula with limited side effects for human being. Objectives: The aim of the current study was to compare the in vitro effect of herbal and chemical mouthwashes against Candida albicans. Materials and Methods: In this research, we used a standard strain of C. albicans, PTCC 5027. The suspension was made by a fresh culture of C. albicans (24 hours) and the optical density (turbidity equating to a McFarland standard of 0.5) was read at 530 nm. The C. albicans suspension was cultured on Sabouraud dextrose agar plate. Next, two wells were filled with mouthwashes and after incubation at 30ºC for 24 hours, the inhibition zone was measured. Minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) of mouthwashes were determined. Data were analyzed using the SPSS software, independent T-tests and one-sided variance analysis (ANOVA-one way). Results: Based on these findings on agar diffusion with (P = 0.764), MIC and MFC tests (P = 0.879), there were no significant differences between the antifungal effect of herbal and chemical mouthwashes. Conclusions: This study showed that, chemical mouthwashes acted better than herbal mouthwashes and among different chemical mouthwashes, Oral B was most effective. PMID:25741429

  20. In Vitro and In Vivo Activities of Pterostilbene against Candida albicans Biofilms

    PubMed Central

    Li, De-Dong; Zhao, Lan-Xue; Mylonakis, Eleftherios; Hu, Gan-Hai; Zou, Yong; Huang, Tong-Kun; Yan, Lan

    2014-01-01

    Pterostilbene (PTE) is a stilbene-derived phytoalexin that originates from several natural plant sources. In this study, we evaluated the activity of PTE against Candida albicans biofilms and explored the underlying mechanisms. In 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) reduction assays, biofilm biomass measurement, confocal laser scanning microscopy, and scanning electron microscopy, we found that ≤16 μg/ml PTE had a significant effect against C. albicans biofilms in vitro, while it had no fungicidal effect on planktonic C. albicans cells, which suggested a unique antibiofilm effect of PTE. Then we found that PTE could inhibit biofilm formation and destroy the maintenance of mature biofilms. At 4 μg/ml, PTE decreased cellular surface hydrophobicity (CSH) and suppressed hyphal formation. Gene expression microarrays and real-time reverse transcription-PCR showed that exposure of C. albicans to 16 μg/ml PTE altered the expression of genes that function in morphological transition, ergosterol biosynthesis, oxidoreductase activity, and cell surface and protein unfolding processes (heat shock proteins). Filamentation-related genes, especially those regulated by the Ras/cyclic AMP (cAMP) pathway, including ECE1, ALS3, HWP1, HGC1, and RAS1 itself, were downregulated upon PTE treatment, indicating that the antibiofilm effect of PTE was related to the Ras/cAMP pathway. Then, we found that the addition of exogenous cAMP reverted the PTE-induced filamentous growth defect. Finally, with a rat central venous catheter infection model, we confirmed the in vivo activity of PTE against C. albicans biofilms. Collectively, PTE had strong activities against C. albicans biofilms both in vitro and in vivo, and these activities were associated with the Ras/cAMP pathway. PMID:24514088

  1. Oxidative stress of photodynamic antimicrobial chemotherapy inhibits Candida albicans virulence

    NASA Astrophysics Data System (ADS)

    Kato, Ilka Tiemy; Prates, Renato Araujo; Tegos, George P.; Hamblin, Michael R.; Simões Ribeiro, Martha

    2011-03-01

    Photodynamic antimicrobial chemotherapy (PACT) is based on the principal that microorganisms will be inactivated using a light source combined to a photosensitizing agent in the presence of oxygen. Oxidative damage of cell components occurs by the action of reactive oxygen species leading to cell death for microbial species. It has been demonstrated that PACT is highly efficient in vitro against a wide range of pathogens, however, there is limited information for its in vivo potential. In addition, it has been demonstrated that sublethal photodynamic inactivation may alter the virulence determinants of microorganisms. In this study, we explored the effect of sublethal photodynamic inactivation to the virulence factors of Candida albicans. Methylene Blue (MB) was used as photosensitizer for sublethal photodynamic challenge on C. albicans associated with a diode laser irradiation (λ=660nm). The parameters of irradiation were selected in causing no reduction of viable cells. The potential effects of PACT on virulence determinants of C. albicans cells were investigated by analysis of germ tube formation and in vivo pathogenicity assays. Systemic infection was induced in mice by the injection of fungal suspension in the lateral caudal vein. C. albicans exposed to sublethal photodynamic inactivation formed significantly less germ tube than untreated cells. In addition, mice infected with C. albicans submitted to sublethal PACT survived for a longer period of time than mice infected with untreated cells. The oxidative damage promoted by sublethal photodynamic inactivation inhibited virulence determinants and reduced in vivo pathogenicity of C. albicans.

  2. In Vitro Activity of Tea Tree Oil Vaginal Suppositories against Candida spp. and Probiotic Vaginal Microbiota.

    PubMed

    Di Vito, Maura; Mattarelli, Paola; Modesto, Monica; Girolamo, Antonietta; Ballardini, Milva; Tamburro, Annunziata; Meledandri, Marcello; Mondello, Francesca

    2015-10-01

    The aim of this work is to evaluate the in vitro microbicidal activity of vaginal suppositories (VS) containing tea tree oil (TTO-VS) towards Candida spp. and vaginal probiotics. A total of 20 Candida spp. strains, taken from patients with vaginitis and from an established type collection, including reference strains, were analysed by using the CLSI microdilution method. To study the action of VS towards the beneficial vaginal microbiota, the sensitivity of Bifidobacterium animalis subsp. lactis (DSM 10140) and Lactobacillus spp. (Lactobacillus casei R-215 and Lactobacillus acidophilus R-52) was tested. Both TTO-VS and TTO showed fungicidal activity against all strains of Candida spp. whereas placebo-VS or the Aloe gel used as controls were ineffective. The study of fractional fungicidal concentrations (FFC) showed synergistic interaction with the association between Amphotericin B and TTO (0.25 to 0.08 µg/ml, respectively) against Candida albicans. Instead, the probiotics were only affected by TTO concentration ≥ 4% v/v, while, at concentrations < 2% v/v, they remained viable. TTO-VS exhibits, in vitro, a selective fungicidal action, slightly affecting only the Bifidobacteriun animalis strain growth belonging to the vaginal microbiota. In vivo studies are needed to confirm the efficacy to prevent acute or recurrent vaginal candidiasis. PMID:26235937

  3. In Vitro Activity of Tea Tree Oil Vaginal Suppositories against Candida spp. and Probiotic Vaginal Microbiota.

    PubMed

    Di Vito, Maura; Mattarelli, Paola; Modesto, Monica; Girolamo, Antonietta; Ballardini, Milva; Tamburro, Annunziata; Meledandri, Marcello; Mondello, Francesca

    2015-10-01

    The aim of this work is to evaluate the in vitro microbicidal activity of vaginal suppositories (VS) containing tea tree oil (TTO-VS) towards Candida spp. and vaginal probiotics. A total of 20 Candida spp. strains, taken from patients with vaginitis and from an established type collection, including reference strains, were analysed by using the CLSI microdilution method. To study the action of VS towards the beneficial vaginal microbiota, the sensitivity of Bifidobacterium animalis subsp. lactis (DSM 10140) and Lactobacillus spp. (Lactobacillus casei R-215 and Lactobacillus acidophilus R-52) was tested. Both TTO-VS and TTO showed fungicidal activity against all strains of Candida spp. whereas placebo-VS or the Aloe gel used as controls were ineffective. The study of fractional fungicidal concentrations (FFC) showed synergistic interaction with the association between Amphotericin B and TTO (0.25 to 0.08 µg/ml, respectively) against Candida albicans. Instead, the probiotics were only affected by TTO concentration ≥ 4% v/v, while, at concentrations < 2% v/v, they remained viable. TTO-VS exhibits, in vitro, a selective fungicidal action, slightly affecting only the Bifidobacteriun animalis strain growth belonging to the vaginal microbiota. In vivo studies are needed to confirm the efficacy to prevent acute or recurrent vaginal candidiasis.

  4. In vitro investigations on the mode of action of the hydroxypyridone antimycotics rilopirox and piroctone on Candida albicans.

    PubMed

    Sigle, H C; Schäfer-Korting, M; Korting, H C; Hube, B; Niewerth, M

    2006-05-01

    Rilopirox and piroctone belong to the class of hydroxypyridone antimycotics. This class is not related to other antimycotics. In contrast to azole antimycotics and polyene antimycotics the mode of action of hydroxypyridone antimycotics is not fully understood. Inhibition of cellular uptake of essential compounds as well as loss of other compounds seems to be only a secondary effect of a primary not known action of these drugs. The antifungal effect in vitro depends on the medium used. The hyphal induction of Candida albicans is inhibited by hydroxypyridone antimycotics, but this effect is compensated by iron ions. A damage of the cell membrane and a direct influence on adenosine triphosphate synthesis, respectively, do not seem to be part of the mode of action. But there are clear hints that reactive oxygen species (ROS) and available metabolic activity are important parts of the mode of action of the hydroxypyridone antimycotics rilopirox and piroctone. PMID:16681805

  5. Inhibition of dioscin on Saprolegnia in vitro.

    PubMed

    Liu, Lei; Shen, Yu-Feng; Liu, Guang-Lu; Ling, Fei; Liu, Xin-Yang; Hu, Kun; Yang, Xian-Le; Wang, Gao-Xue

    2015-12-01

    As one of the most serious pathogens in the freshwater aquatic environment, Saprolegnia can induce a high mortality rate during the fish egg incubation period. This study investigated the anti-Saprolegnia activity of a total of 108 plants on Saprolegnia parasitica in vitro and Dioscorea collettii was selected for further studies. By loading on an open silica gel column and eluting with petroleum ether-ethyl acetate-methanol, dioscin (C45H72O16) was isolated from D. collettii. Saprolegnia parasitica growth was inhibited significantly when dioscin concentration was more than 2.0 mg L(-1). When compared with formalin and hydrogen peroxide, dioscin showed a higher inhibitory effect. As potential inhibition mechanisms, dioscin could cause the S. parasitica mycelium morphologic damage, dense folds, or disheveled protuberances observed by field emission scanning electron microscopy and the influx of Propidium iodide. The structural changes in the treated mycelium were indicative of an efficient anti-Saprolegnia activity of dioscin. The oxidative stress results showed that dioscin also accumulated reactive oxygen species excessively and increased total antioxidant and superoxide dismutase activity. These situations could render S. parasitica more vulnerable to oxidative damage. Additionally, when dioscin concentration was less than 2.0 mg L(-1), the survival rate of embryos was more than 70%. Therefore, the use of dioscin could be a viable way of preventing and controlling saprolegniasis.

  6. Inhibition of dioscin on Saprolegnia in vitro.

    PubMed

    Liu, Lei; Shen, Yu-Feng; Liu, Guang-Lu; Ling, Fei; Liu, Xin-Yang; Hu, Kun; Yang, Xian-Le; Wang, Gao-Xue

    2015-12-01

    As one of the most serious pathogens in the freshwater aquatic environment, Saprolegnia can induce a high mortality rate during the fish egg incubation period. This study investigated the anti-Saprolegnia activity of a total of 108 plants on Saprolegnia parasitica in vitro and Dioscorea collettii was selected for further studies. By loading on an open silica gel column and eluting with petroleum ether-ethyl acetate-methanol, dioscin (C45H72O16) was isolated from D. collettii. Saprolegnia parasitica growth was inhibited significantly when dioscin concentration was more than 2.0 mg L(-1). When compared with formalin and hydrogen peroxide, dioscin showed a higher inhibitory effect. As potential inhibition mechanisms, dioscin could cause the S. parasitica mycelium morphologic damage, dense folds, or disheveled protuberances observed by field emission scanning electron microscopy and the influx of Propidium iodide. The structural changes in the treated mycelium were indicative of an efficient anti-Saprolegnia activity of dioscin. The oxidative stress results showed that dioscin also accumulated reactive oxygen species excessively and increased total antioxidant and superoxide dismutase activity. These situations could render S. parasitica more vulnerable to oxidative damage. Additionally, when dioscin concentration was less than 2.0 mg L(-1), the survival rate of embryos was more than 70%. Therefore, the use of dioscin could be a viable way of preventing and controlling saprolegniasis. PMID:26472687

  7. Microdilution in vitro Antifungal Susceptibility Patterns of Candida Species, From Mild Cutaneous to Bloodstream Infections

    PubMed Central

    Rezazadeh, Elham; Sabokbar, Azar; Moazeni, Maryam; Rezai, Mohammad Sadegh; Badali, Hamid

    2016-01-01

    Background Candida species, as opportunistic organisms, can cause various clinical manifestations, ranging from mild cutaneous infections to systemic candidiasis in otherwise healthy individuals. Remarkably, the incidence and mortality rates of candidemia have significantly increased worldwide, even after advances in medical interventions and the development of novel antifungal drugs. Objectives Given the possible resistance to antifungal agents, susceptibility testing can be useful in defining the activity spectrum of antifungals and determining the appropriate treatment regime. Materials and Methods The in vitro susceptibilities of molecularly identified Candida strains (n = 150) belonging to seven species recovered from clinical specimens, including vaginal, cutaneous, sputum, bronchoalveolar lavage (BAL), and blood samples, were determined for six antifungal drugs (amphotericin B, fluconazole, itraconazole, voriconazole, posaconazole, and caspofungin), based on the clinical and laboratory standards institute’s M27-A3 and M27-S4 documents. Results Candida albicans was the most frequently isolated species (44.66%), followed by non-albicans Candida, including C. glabrata (20%), C. parapsilosis (13.33%), C. krusei (8%), C. tropicalis (7.3%), C. dubliniensis (4%), and C. africana (3.33%). Posaconazole had the lowest geometric mean minimum inhibitory concentration (MIC) (0.0122 µg/ml), followed by amphotericin B (0.0217 µg/mL), voriconazole (0.1022 µg/mL), itraconazole (0.1612 µg/mL), caspofungin (0.2525 µg/mL), and fluconazole (0.4874 µg/mL) against all isolated Candida species. Candida africana and C. parapsilosis were significantly more susceptible to fluconazole, compared to C. albicans and other Candida species (P < 0.001). However, their clinical effectiveness in the treatment of Candida infections remains to be determined. Conclusions These findings highlight the importance of precise and correct species identification of clinical yeast isolates via

  8. Species Distribution and In Vitro Antifungal Susceptibility of Vulvovaginal Candida Isolates in China

    PubMed Central

    Wang, Feng-Juan; Zhang, Dai; Liu, Zhao-Hui; Wu, Wen-Xiang; Bai, Hui-Hui; Dong, Han-Yu

    2016-01-01

    Background: Vulvovaginal candidiasis (VVC) was a common infection associated with lifelong harassment of woman's social and sexual life. The purpose of this study was to describe the species distribution and in vitro antifungal susceptibility of Candida species (Candida spp.) isolated from patients with VVC over 8 years. Methods: Species which isolated from patients with VVC in Peking University First Hospital were identified using chromogenic culture media. Susceptibility to common antifungal agents was determined using agar diffusion method based on CLSI M44-A2 document. SPSS software (version 14.0, Inc., Chicago, IL, USA) was used for statistical analysis, involving statistical description and Chi-square test. Results: The most common strains were Candida (C.) albicans, 80.5% (n = 1775) followed by C. glabrata, 18.1% (n = 400). Nystatin exhibited excellent activity against all species (<4% resistant [R]). Resistance to azole drugs varied among different species. C. albicans: clotrimazole (3.1% R) < fluconazole (16.6% R) < itraconazole (51.5% R) < miconazole (54.0% R); C. glabrata: miconazole (25.6% R) < clotrimazole (50.5% R) < itraconazole (61.9% R) < fluconazole (73.3% R); Candida krusei: clotrimazole (0 R) < fluconazole (57.7% R) < miconazole (73.1% R) < itraconazole (83.3% R). The susceptibility of fluconazole was noticeably decreasing among all species in the study period. Conclusions: Nystatin was the optimal choice for the treatment of VVC at present. The species distribution and in vitro antifungal susceptibility of Candida spp. isolated from patients with VVC had changed over time. PMID:27174323

  9. Characterization of a secretory proteinase of Candida parapsilosis and evidence for the absence of the enzyme during infection in vitro.

    PubMed Central

    Rüchel, R; Böning, B; Borg, M

    1986-01-01

    The opportunistic yeastlike fungi of the genus Candida comprise three species which are proteolytic in vitro. Among them, C. albicans and C. tropicalis are of foremost medical importance. However, a strict correlation between extracellular proteolytic activity and virulence is opposed by the low virulence of the third proteolytic species, C. parapsilosis. We purified the secretory acid proteinase of C. parapsilosis (clinical isolate 265). The enzyme is a carboxyl proteinase (EC 3.4.23) like all other secretory Candida proteinases handled so far. Proteinase 265 is distinguished by a lower molecular weight (approximately 33,000); it has increased hydrophobicity, which accounts for inhibition of the enzyme by hemin, and required the presence of nonionic detergent in the initial steps of purification. The enzyme already undergoes alkaline denaturation at neutrality. Its activity is thus confined to the acid microenvironment of the fungal cell wall. Within this range, the enzyme may degrade immunoglobulins like immunoglobulin A1 (IgA1), IgA2, and secretory IgA. No indication was found for glycosylation of proteinase 265 and the related enzyme of C. albicans CBS 2730. However, the comparable proteinase of C. tropicalis 293 was identified as a manno protein. Antiserum against proteinase 265 cross-reacted strongly with corresponding enzymes from other Candida species. Antisera against proteinases of C. albicans and C. tropicalis reacted only weakly with proteinase 265. Thus, secretory Candida proteinases are likely to possess common and species-specific antigenic sites. In contrast to C. albicans, infection of phagocytes by C. parapsilosis 265 was not accompanied by secretion of fungal proteinase. This lack of induction of the enzyme under conditions of infection may account for the low virulence of most isolates of C. parapsilosis. Images PMID:3525413

  10. Effectiveness of Photodynamic Therapy for the Inactivation of Candida spp. on Dentures: In Vitro Study

    PubMed Central

    Mima, Ewerton Garcia de Oliveira; Ribeiro, Daniela Garcia; Dovigo, Livia Nordi; Vergani, Carlos Eduardo; Bagnato, Vanderlei Salvador

    2011-01-01

    Abstract Objective: This in vitro study evaluated the effectiveness of photodynamic therapy (PDT) for the inactivation of different species of Candida on maxillary complete dentures. Background data: The treatment of denture stomatitis requires the inactivation of Candida spp. on dentures. PDT has been reported as an effective method for Candida inactivation. Methods: Reference strains of C. albicans, C. glabrata, C. tropicalis, C. dubliniensis and C. krusei were tested. Thirty-four dentures were fabricated in a standardized procedure and subjected to ethylene oxide sterilization. The dentures were individually inoculated with one of the strains and incubated at 37°C for 24 h. Dentures submitted to PDT (P+L+) were individually sprayed with 50 mg/L of Photogem® (PS) and, after 30 min, illuminated by LED light for 26 min (37.5 J/cm2). Additional dentures were treated only with PS (P+L-) or light (P-L+) or neither (P-L-). Samples of serial dilutions were spread on Sabouraud dextrose agar and incubated at 37°C for 48 h. The colonies were counted and the values of log (cfu/mL) were analyzed by Kruskall-Wallis and Dunn tests (p<0.05). Results: For all species of Candida, PDT resulted in significant reduction (p<0.05) of cfu/mL values from dentures when compared with P-L- (reductions from 1.73 to 3.99 log10). Significant differences (p<0.05), but lower reductions, were also observed for P+L- and P-L+when compared with P-L- for some species of Candida. Conclusions: PDT was an effective method for reducing Candida spp. on dentures. PMID:21916614

  11. Inhibitory activity in vitro of probiotic lactobacilli against oral Candida under different fermentation conditions.

    PubMed

    Jiang, Q; Stamatova, I; Kari, K; Meurman, J H

    2015-01-01

    Clinical studies have shown that probiotics positively affect oral health by decreasing gum bleeding and/or reducing salivary counts of certain oral pathogens. Our aim was to investigate the inhibitory effect of six probiotic lactobacilli against opportunistic oral Candida species. Sugar utilisation by both lactobacilli and Candida was also assessed. Agar overlay assay was utilised to study growth inhibition of Candida albicans, Candida glabrata and Candida krusei by Lactobacillus rhamnosus GG, Lactobacillus casei Shirota, Lactobacillus reuteri SD2112, Lactobacillus brevis CD2, Lactobacillus bulgaricus LB86 and L. bulgaricus LB Lact. The inhibitory effect was measured at pH 5.5, 6.4, and 7.2, respectively, and in the presence of five different carbohydrates in growth medium (glucose, fructose, lactose, sucrose, and sorbitol). Growth and final pH values were measured at two-hour time points to 24 h. L. rhamnosus GG showed the strongest inhibitory activity in fructose and glucose medium against C. albicans, followed by L. casei Shirota, L. reuteri SD2112 and L. brevis CD2. None of the lactobacilli tested affected the growth of C. krusei. Only L. rhamnosus GG produced slight inhibitory effect on C. glabrata. The lower pH values led to larger inhibition zones. Sugar fermentation profiles varied between the strains. L. casei Shirota grew in the presence of all sugars tested, whereas L. brevis CD2 could utilise only glucose and fructose. All Candida species metabolised the available sugars but the most rapid growth was observed with C. glabrata. The results suggest that commercially available probiotics differ in their inhibitory activity and carbohydrate utilisation; the above properties are modified by different pH values and sugars with more pronounced inhibition at lower pH.

  12. Inhibition of Candida albicans by Lactobacillus acidophilus.

    PubMed

    Collins, E B; Hardt, P

    1980-05-01

    Candida albicans grew at pH 4.6 or above in nutrient broth containing 5% glucose but was retarded at pH 7.7 by filtrates of Lactobacillus acidophilus grown in casitone broth. Vaginal implantation of nonfermented acidophilus milk, yogurt, or low-fat milk for preventing recurrence of monilia vaginitis subsequent to treatment with Nystatin was studied with 30 women. Reinfections within 3 mo according to product received were: no milk product, 3; yogurt, 1; nonfermented acidophilus milk, 1; and low-fat milk, 0. PMID:6771309

  13. The influence of photodynamic therapy parameters on the inactivation of Candida spp: in vitro and in vivo studies

    NASA Astrophysics Data System (ADS)

    Alves, F.; Mima, E. G.; Dovigo, L. N.; Bagnato, V. S.; Jorge, J. H.; de Souza Costa, C. A.; Pavarina, A. C.

    2014-04-01

    The influence of parameters of photodynamic therapy (PDT), such as pre-irradiation time (PIT), on the inactivation of Candida spp. was assessed in vitro and in vivo. Suspensions of Candida albicans, Candida tropicalis, Candida krusei and Candida glabrata were treated with Photogem®, incubated for 5, 10 or 15 min and illuminated with a blue LED light. Colonies were cultivated and log values of CFU ml-1 were analyzed by ANOVA and Kruskall-Wallis test. For in vivo evaluation, immunosuppressed mice were inoculated with C. albicans. PDT was performed on the dorsum of the tongue by topical administration of Photogem® and illumination after 5, 10 or 15 min. C. albicans was recovered from the tongue and the number of CFU ml-1 was analyzed by ANOVA and Tukey test. Animals were killed and the tongues were surgically removed for histological analysis. Susceptibility of Candida spp. suspensions to PDT was in decreasing order: C. albicans = C. tropicalis < C. krusei < C. glabrata. No significant difference was observed among the different PIT (p > 0.05), both in vivo and in vitro. A significant reduction (p < 0.05) of log(CFU ml-1) of C. albicans from tongues of mice was observed with no adverse effects in the tissue. PDT was effective to inactivate in vitroCandida spp. and for reduction of C. albicans in vivo, independently of the PIT used.

  14. In vitro effects of glycyrrhetinic acid on the growth of clinical isolates of Candida albicans.

    PubMed

    Pellati, Donatella; Fiore, Cristina; Armanini, Decio; Rassu, Mario; Bertoloni, Giulio

    2009-04-01

    Compounds derived from Glycyrrhiza glabra L. root have been used widely for centuries for their numerous therapeutic properties. The present study aimed to test the in vitro activity against Candida albicans strains of the compound 18-beta glycyrrhetinic acid (18-beta GA), derived from the root of Glycyrrhiza species. This antimicrobial activity was assessed using the National Committee for Clinical Laboratory Standards (NCCLS) method on C. albicans strains that were isolated from patients with recurrent vulvovaginal candidiasis (RVVC). The in vitro growth of the C. albicans strains was markedly reduced, in a pH-dependent manner, by relatively low doses (6.2 microg/mL) of 18-beta GA. The results demonstrate that 18-beta GA is a promising biological alternative for the topical treatment of recurrent vulvovaginal candidiasis (RVVC). PMID:19067381

  15. [Effects of combined addition of atovaquone and lithium on the in vitro cell growth of the pathogenic yeast Candida albicans].

    PubMed

    Minagawa, Nobuko; Uehara, Mariko; Seki, Shiori; Nitta, Ayumi; Kogawara, Kento

    2010-02-01

    Atovaquone, an analog of ubiquinone, binds tightly to the ubiquinol oxidation site (Qo site) of parasite cytochrome bc(1) complex to inhibit electron transport at concentrations far lower than those at which the mammalian system is affected. The mode of action is thought similar to that of myxothiazol. To treat Pneumocystis jirovecii and Plasmodium falciparum infections, atovaquone has been used worldwide whereas it is unapproved in Japan. Since the pathogenic Candida species fungi seem resistant to atovaquone, this drug is not clinically available for candidosis, particularly deep mycosis. We examined the effects of atovaquone on cellular respiration and in vitro growth of C. albicans to explore a new therapeutic possibility for fungal infections. Atovaquone strongly inhibited glucose-dependent cellular respiration similarly to antimycin A, stigmatellin, and myxothiazol, specific bc(1) complex inhibitors. However, atovaquone suppressed glucose-dependent cell growth to a much lesser extent versus the comparator agents. When added alone, lithium exerted slight growth inhibition. The combined addition of lithium with atovaquone showed a significant increase in inhibition of growth. Although the way lithium acts synergistically with atovaquone remains to be elucidated, our results suggest a new therapeutic possibility of this combination for the treatment of candidosis.

  16. Effect of Streptococcus salivarius K12 on the in vitro growth of Candida albicans and its protective effect in an oral candidiasis model.

    PubMed

    Ishijima, Sanae A; Hayama, Kazumi; Burton, Jeremy P; Reid, Gregor; Okada, Masashi; Matsushita, Yuji; Abe, Shigeru

    2012-04-01

    Oral candidiasis is often accompanied by severe inflammation, resulting in a decline in the quality of life of immunosuppressed individuals and elderly people. To develop a new oral therapeutic option for candidiasis, a nonpathogenic commensal oral probiotic microorganism, Streptococcus salivarius K12, was evaluated for its ability to modulate Candida albicans growth in vitro, and its therapeutic activity in an experimental oral candidiasis model was tested. In vitro inhibition of mycelial growth of C. albicans was determined by plate assay and fluorescence microscopy. Addition of S. salivarius K12 to modified RPMI 1640 culture medium inhibited the adherence of C. albicans to the plastic petri dish in a dose-dependent manner. Preculture of S. salivarius K12 potentiated its inhibitory activity for adherence of C. albicans. Interestingly, S. salivarius K12 was not directly fungicidal but appeared to inhibit Candida adhesion to the substratum by preferentially binding to hyphae rather than yeast. To determine the potentially anti-infective attributes of S. salivarius K12 in oral candidiasis, the probiotic was administered to mice with orally induced candidiasis. Oral treatment with S. salivarius K12 significantly protected the mice from severe candidiasis. These findings suggest that S. salivarius K12 may inhibit the process of invasion of C. albicans into mucous surfaces or its adhesion to denture acrylic resins by mechanisms not associated with the antimicrobial activity of the bacteriocin. S. salivarius K12 may be useful as a probiotic as a protective tool for oral care, especially with regard to candidiasis. PMID:22267663

  17. Effect of Streptococcus salivarius K12 on the In Vitro Growth of Candida albicans and Its Protective Effect in an Oral Candidiasis Model

    PubMed Central

    Hayama, Kazumi; Burton, Jeremy P.; Reid, Gregor; Okada, Masashi; Matsushita, Yuji; Abe, Shigeru

    2012-01-01

    Oral candidiasis is often accompanied by severe inflammation, resulting in a decline in the quality of life of immunosuppressed individuals and elderly people. To develop a new oral therapeutic option for candidiasis, a nonpathogenic commensal oral probiotic microorganism, Streptococcus salivarius K12, was evaluated for its ability to modulate Candida albicans growth in vitro, and its therapeutic activity in an experimental oral candidiasis model was tested. In vitro inhibition of mycelial growth of C. albicans was determined by plate assay and fluorescence microscopy. Addition of S. salivarius K12 to modified RPMI 1640 culture medium inhibited the adherence of C. albicans to the plastic petri dish in a dose-dependent manner. Preculture of S. salivarius K12 potentiated its inhibitory activity for adherence of C. albicans. Interestingly, S. salivarius K12 was not directly fungicidal but appeared to inhibit Candida adhesion to the substratum by preferentially binding to hyphae rather than yeast. To determine the potentially anti-infective attributes of S. salivarius K12 in oral candidiasis, the probiotic was administered to mice with orally induced candidiasis. Oral treatment with S. salivarius K12 significantly protected the mice from severe candidiasis. These findings suggest that S. salivarius K12 may inhibit the process of invasion of C. albicans into mucous surfaces or its adhesion to denture acrylic resins by mechanisms not associated with the antimicrobial activity of the bacteriocin. S. salivarius K12 may be useful as a probiotic as a protective tool for oral care, especially with regard to candidiasis. PMID:22267663

  18. Effect of Streptococcus salivarius K12 on the in vitro growth of Candida albicans and its protective effect in an oral candidiasis model.

    PubMed

    Ishijima, Sanae A; Hayama, Kazumi; Burton, Jeremy P; Reid, Gregor; Okada, Masashi; Matsushita, Yuji; Abe, Shigeru

    2012-04-01

    Oral candidiasis is often accompanied by severe inflammation, resulting in a decline in the quality of life of immunosuppressed individuals and elderly people. To develop a new oral therapeutic option for candidiasis, a nonpathogenic commensal oral probiotic microorganism, Streptococcus salivarius K12, was evaluated for its ability to modulate Candida albicans growth in vitro, and its therapeutic activity in an experimental oral candidiasis model was tested. In vitro inhibition of mycelial growth of C. albicans was determined by plate assay and fluorescence microscopy. Addition of S. salivarius K12 to modified RPMI 1640 culture medium inhibited the adherence of C. albicans to the plastic petri dish in a dose-dependent manner. Preculture of S. salivarius K12 potentiated its inhibitory activity for adherence of C. albicans. Interestingly, S. salivarius K12 was not directly fungicidal but appeared to inhibit Candida adhesion to the substratum by preferentially binding to hyphae rather than yeast. To determine the potentially anti-infective attributes of S. salivarius K12 in oral candidiasis, the probiotic was administered to mice with orally induced candidiasis. Oral treatment with S. salivarius K12 significantly protected the mice from severe candidiasis. These findings suggest that S. salivarius K12 may inhibit the process of invasion of C. albicans into mucous surfaces or its adhesion to denture acrylic resins by mechanisms not associated with the antimicrobial activity of the bacteriocin. S. salivarius K12 may be useful as a probiotic as a protective tool for oral care, especially with regard to candidiasis.

  19. Glycerol inhibition of ruminal lipolysis in vitro

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Supplemental glycerol inhibits rumen lipolysis, a prerequisite for rumen biohydrogenation, which is responsible for the saturation of dietary fatty acids consumed by ruminant animals. Feeding excess glycerol, however, adversely affects dry matter digestibility. To more clearly define the effect of...

  20. Effects of Treated versus Untreated Polystyrene on Caspofungin In Vitro Activity against Candida Species

    PubMed Central

    Fothergill, Annette W.; McCarthy, Dora I.; Albataineh, Mohammad T.; Sanders, Carmita; McElmeel, Maria

    2016-01-01

    Significant interlaboratory variability is observed in testing the caspofungin susceptibility of Candida species by both the CLSI and EUCAST broth microdilution methodologies. We evaluated the influence of treated versus untreated polystyrene microtiter trays on caspofungin MICs using 209 isolates of four Candida species, including 16 C. albicans and 11 C. glabrata isolates with defined FKS mutations. Caspofungin MICs were also determined using the commercially available YeastOne and Etest assays and 102 isolates. All C. glabrata isolates had caspofungin MICs of ≥0.5 μg/ml, the clinical breakpoint for caspofungin resistance in this species, measured using trays made of treated polystyrene, regardless of the FKS status. In contrast, susceptible isolates could readily be distinguished from resistant/non-wild-type isolates when caspofungin MICs were measured using untreated polystyrene trays and both the YeastOne and Etest assays. Similar results were also observed for C. krusei isolates, as all isolates had caspofungin MICs above the threshold for resistance measured using treated polystyrene trays. In contrast, C. albicans isolates could be correctly identified as susceptible or resistant when caspofungin MICs were measured with treated or untreated trays and with the YeastOne and Etest assays. MICs falsely elevated above the resistance breakpoint were also not observed for C. tropicalis isolates. These results demonstrated that the use of treated polystyrene may be one factor that leads to falsely elevated caspofungin in vitro susceptibility results and that this may also be a greater issue for some Candida species than for others. PMID:26763959

  1. In Vitro Susceptibility and Trailing Growth Effect of Clinical Isolates of Candida Species to Azole Drugs

    PubMed Central

    Zomorodian, Kamiar; Bandegani, Azadeh; Mirhendi, Hossein; Pakshir, Keyvan; Alinejhad, Navvab; Poostforoush Fard, Ali

    2016-01-01

    Background: Emergence of resistance to respective antifungal drugs is a primary concern for the treatment of candidiasis. Hence, determining antifungal susceptibility of the isolated yeasts is of special importance for effective therapy. For this purpose, the clinical laboratory standard institute (CLSI) has introduced a broth microdilution method to determine minimum inhibitory concentration (MIC). However, the so-called “Trailing effect” phenomenon might sometimes pose ambiguity in the interpretation of the results. Objectives: The present study aimed to determine the in vitro susceptibility of clinical isolates of Candida against azoles and the frequency of the Trailing effect. Materials and Methods: A total of 193 Candida isolates were prospectively collected and identified through the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Using a broth microdilution test, according to the guidelines of CLSI M27-A3, antifungal susceptibilities of the isolated yeasts against Fluconazole (FLU), Itraconazole (ITR), Ketoconazole (KET) and Voriconazole (VOR) were assessed. Moreover, trailing growth was determined when a susceptible MIC was incubated for 24 hours, and turned into a resistant one after 48 hours of incubation. Results: Among the tested antifungal drugs in this study, the highest rate of resistance was observed against ITR (28.5%) followed by VOR (26.4%), FLU (20.8%) and KET (1.5%). The trailing effect was induced in 27 isolates (14.0%) by VOR, in 26 isolates (13.5%) by ITR, in 24 isolates (12.4%) by FLU, and in 19 isolates (9.8%) by KET. Conclusions: The monitoring of antifungal susceptibilities of Candida species isolated from clinical sources is highly recommended for the efficient management of patients. Moreover, the trailing effect should be taken into consideration once the interpretation of the results is intended. PMID:27127587

  2. [In vitro antifungal resistance in Candida albicans from HIV-infected patients with and without oral candidosis.].

    PubMed

    Ceballos Salobreña, A; Gaitán Cepeda, L A; Orihuela Cañada, F; Olea Barrionuevo, D; Ceballos García, L; Quindós, G

    1999-12-01

    The main purpose of this study has been to determine the in vitro antifungal susceptibility of clinical isolates from HIV-infected or AIDS patients, depending on the presence of oral candidosis. The oral cavity of 307 HIV-infected or AIDS patients was examined and an oral swab was cultured on Sabouraud glucose agar and studied by conventional mycological methods. In vitro antifungal susceptibility to amphotericin B, nystatin, fluconazole, itraconazole and ketoconazole was tested by disk diffusion with Neo-Sensitabs tablets (Rosco Diagnostica, Dinamarca). One hundred and thirty five Candida albicans isolates (91 serotype A, 38 serotype B, three C. albicans variety stellatoidea and three untyped isolates), three Candida krusei and two Candida glabrata were obtained. All the isolates were susceptible to nystatin and amphotericin B. However, 7.9% isolates were resistant to fluconazole and 2.9% isolates were resistant to ketoconazole or itraconazole. Nearly all C. krusei and C. glabrata isolates, 31% patients with candidosis and 20% Candida-colonized patients showed decreased susceptibility to azoles. This study shows that polyenes had a great in vitro efficacy against clinical isolates from HIV-infected patients and that in vitro resistance to azoles is not as high as observed in other countries.

  3. Chemical characterization by GC-MS and in vitro activity against Candida albicans of volatile fractions prepared from Artemisia dracunculus, Artemisia abrotanum, Artemisia absinthium and Artemisia vulgaris

    PubMed Central

    2014-01-01

    Background A large number of essential oils is reported to have significant activity against Candida albicans. But the different chemical composition influences the degree of their activity. The intention of this study was to investigate the chemical composition and the activity against Candida albicans of volatile oils obtained from Artemisia dracunculus, A. abrotanum, A. absinthium and A. vulgaris (Asteraceae). The aim of the study was to identify new chemical compounds that have effect against C. albicans. The essential oils were obtained by hydrodistillation or extraction with dichloromethane (a new procedure we developed trying to obtain better, more separated compounds) from air dried above ground plant material and analyzed by GC-MS. Additionally commercial essential oils from the same species were tested. The Candida albicans inhibition studies were carried out by the paper disc diffusion method. Results The essential oils shared common components but presented differences in composition and showed variable antifungal activity. Davanone and derivatives thereof, compounds with silphiperfolane skeleton, estragole, davanone oil, β-thujone, sabinyl acetate, herniarin, cis-chrysanthenyl acetate, 1,8-cineol, and terpineol were the main components of Artemisia volatiles. Conclusions Among the volatile fractions tested those from A. abrotanum containing davanone or silphiperfolane derivatives showed the highest antifungal activity. The in vitro tests revealed that the Artemisia oils are promising candidates for further research to develop novel anti-candida drugs. PMID:24475951

  4. In vitro antifungal synergy between amphiphilic aminoglycoside K20 and azoles against Candida species and Cryptococcus neoformans.

    PubMed

    Shrestha, Sanjib K; Grilley, Michelle; Anderson, Thomas; Dhiman, Christine; Oblad, John; Chang, Cheng-Wei T; Sorensen, Kevin N; Takemoto, Jon Y

    2015-11-01

    Several azoles are widely used to treat human fungal infections. Increasing resistance to these azoles has prompted exploration of their synergistic antifungal activities when combined with other agents. The amphiphilic aminoglycoside, K20, was recently shown to inhibit filamentous fungi, yeasts and heterokonts, but not bacteria. In this study, in vitro synergistic growth inhibition by combinations of K20 and azoles (fluconazole, itraconazole, voriconazole, clotrimazole, or posaconazole) were examined against Candida species and Cryptococcus neoformans. Checkerboard microbroth dilution, time-kill curve, and disk diffusion assays revealed that K20 has synergistic inhibitory activities with all five azoles against C. albicans including azole-resistant C. albicans strains ATCC 64124 and ATCC 10231. Four (fluconazole, itraconazole, clotrimazole, posaconazole) and three (fluconazole, itraconazole, voriconazole) azoles were synergistically inhibitory with K20 against C. lusitaniae and C. tropicalis, respectively. Only posaconazole showed synergy with K20 against two Cryptococcus neoformans strains (90-26 and VR-54). Time-kill curves with azole-resistant C. albicans 64124 and azole-sensitive C. albicans MYA-2876 confirmed the K20-azole synergistic interactions with a ≥ 2 log10 decrease in colony-forming units (CFU)/ml compared with the corresponding azoles alone. These results suggest that combinations of K20 and azoles offer a possible strategy for developing therapies against candidiasis.

  5. Ecology of Candida albicans gut colonization: inhibition of Candida adhesion, colonization, and dissemination from the gastrointestinal tract by bacterial antagonism.

    PubMed Central

    Kennedy, M J; Volz, P A

    1985-01-01

    Antibiotic-treated and untreated Syrian hamsters were inoculated intragastrically with Candida albicans to determine whether C. albicans could opportunistically colonize the gastrointestinal tract and disseminate to visceral organs. Antibiotic treatment decreased the total population levels of the indigenous bacterial flora and predisposed hamsters to gastrointestinal overgrowth and subsequent systemic dissemination by C. albicans in 86% of the animals. Both control hamsters not given antibiotics and antibiotic-treated animals reconventionalized with an indigenous microflora showed significantly lower gut populations of C. albicans, and C. albicans organisms were cultured from the visceral organs of 0 and 10% of the animals, respectively. Conversely, non-antibiotic-treated hamsters inoculated repeatedly with C. albicans had high numbers of C. albicans in the gut, and viable C. albicans was recovered from the visceral organs of 53% of the animals. Examination of the mucosal surfaces from test and control animals indicated further that animals which contained a complex indigenous microflora had significantly lower numbers of C. albicans associated with their gut walls than did antibiotic-treated animals. The ability of C. albicans to associate with intestinal mucosal surfaces also was tested by an in vitro adhesion assay. The results indicate that the indigenous microflora reduced the mucosal association of C. albicans by forming a dense layer of bacteria in the mucus gel, out-competing yeast cells for adhesion sites, and producing inhibitor substances (possibly volatile fatty acids, secondary bile acids, or both) that reduced C. albicans adhesion. It is suggested, therefore, that the indigenous intestinal microflora suppresses C. albicans colonization and dissemination from the gut by inhibiting Candida-mucosal association and reducing C. albicans population levels in the gut. Images PMID:3897061

  6. Glycerol inhibition of ruminal lipolysis in vitro.

    PubMed

    Edwards, H D; Anderson, R C; Miller, R K; Taylor, T M; Hardin, M D; Smith, S B; Krueger, N A; Nisbet, D J

    2012-09-01

    Supplemental glycerol inhibits rumen lipolysis, a prerequisite for rumen biohydrogenation, which is responsible for the saturation of dietary fatty acids consumed by ruminant animals. Feeding excess glycerol, however, adversely affects dry matter digestibility. To more clearly define the effect of supplemental glycerol on rumen lipolysis, mixed populations of ruminal bacteria were incubated with 6 or 20% glycerol (vol/vol). After 48-h anaerobic incubation of mixed culture rumen fluid, rates of free fatty acid production (nmol/mL per h) for the 6 and 20% glycerol-supplemented samples were decreased by 80 and 86%, respectively, compared with rates from nonsupplemented control cultures (12.4±1.0; mean ± SE). Conversely, assay of the prominent ruminal lipase-producing bacteria Anaerovibrio lipolyticus 5S, Butyrivibrio fibrisolvens 49, and Propionibacterium species avidum and acnes revealed no effect of 2 or 10% (vol/vol) added glycerol on lipolytic activity by these organisms. Supplementing glycerol at 6% on a vol/vol basis, equivalent to supplementing glycerol at approximately 8 to 15% of diet dry matter, effectively reduced lipolysis. However, the mechanism of glycerol inhibition of ruminal lipolysis remains to be demonstrated. PMID:22916923

  7. A novel renal epithelial cell in vitro assay to assess Candida albicans virulence

    PubMed Central

    Szabo, Edina K; MacCallum, Donna M

    2014-01-01

    Candida albicans, an opportunistic fungal pathogen, can cause severe systemic infections in susceptible patient groups. Systemic candidiasis is mainly studied in the mouse intravenous challenge model, where progressive infection correlates with increased early renal chemokine levels. To develop a new in vitro assay to assess C. albicans virulence, which reflects the events occurring in the murine infection model, renal M-1 cortical collecting duct epithelial cells were evaluated as the early producers of cytokines in response to C. albicans. We show that renal epithelial cells respond only to live C. albicans cells capable of forming hyphae, producing chemokines KC and MIP-2, with levels correlating with epithelial cell damage. By assaying epithelial cell responses to strains of known virulence in the murine intravenous challenge model we demonstrate that renal epithelial cells can discriminate between virulent and attenuated strains. This simple, novel assay is a useful initial screen for altered virulence of C. albicans mutants or clinical isolates in vitro and provides an alternative to the mouse systemic infection model. PMID:24225657

  8. Standardized Method for In Vitro Antifungal Susceptibility Testing of Candida albicans Biofilms

    PubMed Central

    Ramage, Gordon; Vande Walle, Kacy; Wickes, Brian L.; López-Ribot, José L.

    2001-01-01

    Candida albicans is implicated in many biomaterial-related infections. Typically, these infections are associated with biofilm formation. Cells in biofilms display phenotypic traits that are dramatically different from those of their free-floating planktonic counterparts and are notoriously resistant to antimicrobial agents. Consequently, biofilm-related infections are inherently difficult to treat and to fully eradicate with normal treatment regimens. Here, we report a rapid and highly reproducible microtiter-based colorimetric assay for the susceptibility testing of fungal biofilms, based on the measurement of metabolic activities of the sessile cells by using a formazan salt reduction assay. The assay was used for in vitro antifungal susceptibility testing of several C. albicans strains grown as biofilms against amphotericin B and fluconazole and the increased resistance of C. albicans biofilms against these antifungal agents was demonstrated. Because of its simplicity, compatibility with a widely available 96-well microplate platform, high throughput, and automation potential, we believe this assay represents a promising tool for the standardization of in vitro antifungal susceptibility testing of fungal biofilms. PMID:11502517

  9. Effects of Candida norvegensis Live Cells on In vitro Oat Straw Rumen Fermentation

    PubMed Central

    Ruiz, Oscar; Castillo, Yamicela; Arzola, Claudio; Burrola, Eduviges; Salinas, Jaime; Corral, Agustín; Hume, Michael E.; Murillo, Manuel; Itza, Mateo

    2016-01-01

    This study evaluated the effect of Candida norvegensis (C. norvegensis) viable yeast culture on in vitro ruminal fermentation of oat straw. Ruminal fluid was mixed with buffer solution (1:2) and anaerobically incubated with or without yeast at 39°C for 0, 4, 8, 16, and 24 h. A fully randomized design was used. There was a decrease in lactic acid (quadratic, p = 0.01), pH, (quadratic, p = 0.02), and yeasts counts (linear, p<0.01) across fermentation times. However, in vitro dry matter disappearance (IVDMD) and ammonia-N increased across fermentation times (quadratic; p<0.01 and p<0.02, respectively). Addition of yeast cells caused a decrease in pH values compared over all fermentation times (p<0.01), and lactic acid decreased at 12 h (p = 0.05). Meanwhile, yeast counts increased (p = 0.01) at 12 h. C. norvegensis increased ammonia-N at 4, 8, 12, and 24 h (p<0.01), and IVDMD of oat straw increased at 8, 12, and 24 h (p<0.01) of fermentation. Yeast cells increased acetate (p<0.01), propionate (p<0.03), and butyrate (p<0.03) at 8 h, while valeriate and isovaleriate increased at 8, 12, and 24 h (p<0.01). The yeast did not affect cellulolytic bacteria (p = 0.05), but cellulolytic fungi increased at 4 and 8 h (p<0.01), whereas production of methane decreased (p<0.01) at 8 h. It is concluded that addition of C. norvegensis to in vitro oat straw fermentation increased ruminal fermentation parameters as well as microbial growth with reduction of methane production. Additionally, yeast inoculum also improved IVDMD. PMID:26732446

  10. Characterization of In Vitro Resistance Development to the Novel Echinocandin CD101 in Candida Species.

    PubMed

    Locke, Jeffrey B; Almaguer, Amanda L; Zuill, Douglas E; Bartizal, Ken

    2016-10-01

    CD101 is a novel echinocandin with a long half-life undergoing clinical development for treatment of candidemia/invasive candidiasis and vulvovaginal candidiasis. The potential for and mechanisms underlying the development of resistance to CD101 in Candida species were investigated by using spontaneous resistance and serial passage selection methodologies. Four Candida spp. (C. albicans, C. glabrata, C. parapsilosis, and C. krusei) were chosen for resistance characterization with CD101, anidulafungin, and caspofungin. The frequency of spontaneous, single-step mutations conferring reduced susceptibility to CD101 at 1× the agar growth inhibition concentration was low across all species, with median frequencies ranging from 1.35 × 10(-8) to 3.86 × 10(-9), similar to ranges generated for anidulafungin and caspofungin. Serial passage of Candida spp. on agar plates containing drug gradients demonstrated a low potential for resistance development, with passage 20 CD101-selected strains possessing increases in MICs equivalent to or lower than those for the majority of strains generated under selection with anidulafungin and caspofungin. A total of 12 fks "hot spot" mutations were identified, typically in strains with the highest MIC shifts. Cross-resistance was broadly observed among the 3 echinocandins evaluated, with no CD101-selected mutants (with or without fks hot spot mutations) exhibiting reduced susceptibility to CD101 but not also to anidulafungin and/or caspofungin. Consistent with currently approved echinocandins, CD101 demonstrates a low potential for resistance development, which could be further enhanced in vivo by the high maximum concentration of drug in serum (Cmax)/area under the concentration-time curve (AUC) plasma drug exposure achieved with once-weekly dosing of CD101.

  11. In vitro sensitivity of Candida spp. to hematoporphyrin monomethyl ether-mediated photodynamic inactivation

    NASA Astrophysics Data System (ADS)

    Wang, Yucheng; Wang, Ying; Wu, Sumin; Gu, Ying

    2014-11-01

    Background: An increasing prevalence of Candida infections has emerged with the wide use of immune-suppressants and antibiotics. Current antifungal drugs exhibit low efficiency and high toxicity to the normal organs. Photodynamic inactivation (PDI) provides an alternative therapeutic strategy involving the use of photosensitizer (PS) and light irradiation. This study evaluated PDI effects against strains of C. albicans, C. parapsilosis, C. krusei and C. glabrata, using the PS of hematoporphyrin monomethyl ether (HMME), which is a second-generation PS clinically approved in China. Methods: Suspensions (~106 CFU/ml) were incubated with seven HMME concentrations (0.25~50 μM) for 30 min followed by 532-nm laser irradiation for 10 min at 40 mW/cm2. Viability of cells was assayed by CFU counting. Furthermore, fetal calf serum (10%) and singlet oxygen quencher sodium azide (100mM) were respectively added to the suspension of C. krusei to evaluate their roles in PDI process. Results: Among the four species, C. albicans was the most sensitive to PDI; 4 log10 killing was achieved at the concentration of 7.5 μM. C. glabrata was the most resistant; 3 log10 killing was not obtained even at PS concentration of 50 μM. PDI effects against C. krusei were inhibited by both serum and sodium azide. Conclusions: HMME-mediated PDI was able to effectively kill Candida in our experimental conditions, mainly through a Type Ⅱ photoprocess. However, the effects could be intensively reversed by the presence of serum. Thus, there might be a long way before HMME can be used in fighting against Candida in real infectious foci.

  12. Characterization of In Vitro Resistance Development to the Novel Echinocandin CD101 in Candida Species

    PubMed Central

    Almaguer, Amanda L.; Zuill, Douglas E.; Bartizal, Ken

    2016-01-01

    CD101 is a novel echinocandin with a long half-life undergoing clinical development for treatment of candidemia/invasive candidiasis and vulvovaginal candidiasis. The potential for and mechanisms underlying the development of resistance to CD101 in Candida species were investigated by using spontaneous resistance and serial passage selection methodologies. Four Candida spp. (C. albicans, C. glabrata, C. parapsilosis, and C. krusei) were chosen for resistance characterization with CD101, anidulafungin, and caspofungin. The frequency of spontaneous, single-step mutations conferring reduced susceptibility to CD101 at 1× the agar growth inhibition concentration was low across all species, with median frequencies ranging from 1.35 × 10−8 to 3.86 × 10−9, similar to ranges generated for anidulafungin and caspofungin. Serial passage of Candida spp. on agar plates containing drug gradients demonstrated a low potential for resistance development, with passage 20 CD101-selected strains possessing increases in MICs equivalent to or lower than those for the majority of strains generated under selection with anidulafungin and caspofungin. A total of 12 fks “hot spot” mutations were identified, typically in strains with the highest MIC shifts. Cross-resistance was broadly observed among the 3 echinocandins evaluated, with no CD101-selected mutants (with or without fks hot spot mutations) exhibiting reduced susceptibility to CD101 but not also to anidulafungin and/or caspofungin. Consistent with currently approved echinocandins, CD101 demonstrates a low potential for resistance development, which could be further enhanced in vivo by the high maximum concentration of drug in serum (Cmax)/area under the concentration-time curve (AUC) plasma drug exposure achieved with once-weekly dosing of CD101. PMID:27480852

  13. Brassinosteroids inhibit in vitro angiogenesis in human endothelial cells.

    PubMed

    Rárová, Lucie; Zahler, Stefan; Liebl, Johanna; Kryštof, Vladimír; Sedlák, David; Bartůněk, Petr; Kohout, Ladislav; Strnad, Miroslav

    2012-11-01

    Antiangiogenic activity of the brassinosteroid plant hormones (BRs) and their derivative cholestanon was investigated in human umbilical vein endothelial cells (HUVEC) and in human microvascular endothelial cells (HMEC-1). 24-Epibrassinolide and 28-homocastasterone from group of 21 tested natural BRs inhibited migration of HUVEC cells. Seven tested BRs decreased the number of tubes significantly. Synthetic analogue cholestanon inhibited angiogenesis in vitro more effectively than natural BRs. Because of the similarity of BRs to human steroids, we have also studied interactions of BRs with human steroid receptors. Synthetic BRs cholestanon showed agonistic effects on estrogen-receptor-α, estrogen-receptor-β and androgen receptor. Of the natural BRs, 24-epibrassinolide was found to be a weak antagonist of estrogen-receptor-α (ERα). Our results provide the first evidence that large group of BRs can inhibit in vitro angiogenesis of primary endothelial cells. BRs constitute a novel group of human steroid receptor activators or inhibitors with capacity to inhibit angiogenesis.

  14. In Vitro Fungicidal Activities of Anidulafungin, Caspofungin, and Micafungin against Candida glabrata, Candida bracarensis, and Candida nivariensis Evaluated by Time-Kill Studies

    PubMed Central

    Gil-Alonso, Sandra; Jauregizar, Nerea; Cantón, Emilia; Eraso, Elena

    2015-01-01

    Anidulafungin, caspofungin, and micafungin killing activities against Candida glabrata, Candida bracarensis, and Candida nivariensis were evaluated by the time-kill methodology. The concentrations assayed were 0.06, 0.125, and 0.5 μg/ml, which are achieved in serum. Anidulafungin and micafungin required between 13 and 26 h to reach the fungicidal endpoint (99.9% killing) against C. glabrata and C. bracarensis. All echinocandins were less active against C. nivariensis. PMID:25801575

  15. Geranylgeranylacetone inhibits ovarian cancer progression in vitro and in vivo

    SciTech Connect

    Hashimoto, Kae; Morishige, Ken-ichirou . E-mail: mken@gyne.med.osaka-u.ac.jp; Sawada, Kenjiro; Ogata, Seiji; Tahara, Masahiro; Shimizu, Shoko; Sakata, Masahiro; Tasaka, Keiichi; Kimura, Tadashi

    2007-04-27

    Geranylgeranylacetone (GGA), an isoprenoid compound, is an anti-ulcer drug developed in Japan. In our previous study, GGA was shown to inhibit ovarian cancer invasion by attenuating Rho activation [K. Hashimoto, K. Morishige, K. Sawada, M. Tahara, S. Shimizu, M. Sakata, K. Tasaka, Y. Murata, Geranylgeranylacetone inhibits lysophosphatidic acid-induced invasion of human ovarian carcinoma cells in vitro. Cancer 103 (2005) 1529-1536.]. In the present study, GGA treatment inhibited ovarian cancer progression in vitro and suppressed the tumor growth and ascites in the in vivo ovarian cancer model. In vitro analysis, treatment of cancer cells by GGA resulted in the inhibition of cancer cell proliferation, the inactivation of Ras, and the suppression of tyrosine phosphorylation of mitogen-activated protein kinase (MAPK). In conclusion, this is the first report that GGA inhibited ovarian cancer progression and the anti-tumor effect by GGA is, at least in part, derived not only from the suppression of Rho activation but also Ras-MAPK activation.

  16. In vivo and in vitro acquisition of resistance to voriconazole by Candida krusei.

    PubMed

    Ricardo, Elisabete; Miranda, Isabel M; Faria-Ramos, Isabel; Silva, Raquel M; Rodrigues, Acácio Gonçalves; Pina-Vaz, Cidália

    2014-08-01

    Candida krusei is an important agent of opportunistic infections that often displays resistance to several antifungals. We describe here the in vivo acquisition of resistance to voriconazole (VRC) by C. krusei isolates recovered from a leukemia patient during a long period of VRC therapy. In order to mimic the in vivo development of VRC resistance, a susceptible C. krusei isolate was exposed daily to 1 μg/ml of VRC in vitro. Interestingly, after 5 days of exposure to VRC, a MIC of 4 μg/ml was achieved; this value remained constant after 25 additional days of treatment with VRC and also after 30 consecutive days of incubation in VRC-free medium. Our objective was to determine the associated molecular resistance mechanisms, such as expression of efflux pump genes and ERG11 gene mutations, among the resistant strains. Synergistic effects between the efflux blocker tacrolimus (FK506) and VRC were found in all of the resistant strains. Moreover, ABC1 gene expression increased over time in both the in vivo- and in vitro-induced resistant strains, in contrast to the ABC2 and ERG11 genes, whose expression was invariably lower and constant. ERG11 gene sequencing showed two different types of mutations, i.e., heterozygosity at T1389T/C, corresponding to synonymous mutations, in C. krusei strains and a missense mutation at position T418C, resulting in a change from Tyr to His, among resistant C. krusei clinical isolates. This study highlights the relevance of ATP-dependent efflux pump (namely, Abc1p) activity in VRC resistance and describes new mutations in the ERG11 gene among resistant C. krusei clinical isolates. PMID:24867987

  17. Evaluation of the in vitro antimicrobial properties of ultraviolet A/riboflavin mediated crosslinking on Candida albicans and Fusarium solani

    PubMed Central

    Sun, Bing; Li, Zhi-Wei; Yu, Hai-Qun; Tao, Xiang-Chen; Zhang, Yong; Mu, Guo-Ying

    2014-01-01

    AIM To evaluate the antimicrobial properties of ultraviolet A (UVA) (365 nm)/riboflavin against Candida albicans and Fusarium solani. METHODS Two fungus isolates were cultured in vitro and prepared with 10-fold serial PBS dilutions of cell concentration. For each dilution of fungus suspension, the concentration (colony-forming units/mL, CFU/mL) and the inactivation ratio of fungal cells were evaluated under 4 conditions: no treatment (control), UVA (365 nm)/riboflavin, riboflavin, and UVA (365 nm). RESULTS The cell concentration decreased in UVA (365 nm)/riboflavin group for Candida albicans at each dilution and Fusarium solani at dilutions of 104, 103, 102 CFU/mL, when compared with that in control, riboflavin, and UVA (365 nm) groups (P<0.01). No difference of cell concentration was detected amongst the culture of control, riboflavin, and UVA (365 nm) groups for the two fungus. There is a negative correlation between suspension concentration (log-transformed) and the inactivation ratio in UVA (365 nm)/riboflavin group for Candida albicans and Fusarium solani (P<0.01). CONCLUSION According to the standard protocol of corneal collagen cross-linking, UVA (365 nm)/riboflavin combination treatment is found to moderately inactivate the viability of Candida albicans and Fusarium solani in vitro. The inactivation ratio was found to increase with the decrease of cell concentration under UVA (365 nm)/riboflavin condition. PMID:24790859

  18. Suppression of polymorphonuclear (PMN) and monocyte-mediated inhibition of Candida albicans growth by delta-9-tetrahydrocannabinol

    SciTech Connect

    Djeu, J.Y.; Parapanios, A.; Halkias, D.; Friedman, H.

    1986-03-05

    This study was an in vitro attempt to identify the effector cells responsible for growth inhibition of the opportunistic fungus, candida albicans, and to determine if THC or another marijuana derivatives, 11-hydroxyTHC, would adversely affect their function. Using a 24h radiolabel assay, the authors found that growth inhibition of C. albicans was primarily mediated by PMN and monocytes that could be isolated normal human peripheral blood. Both effector cell types caused almost complete inhibition of Candida growth at effector/target ratio of 300/1 and inhibition was often still seen at 30/1-. Incubation of PMN, PBL, or monocytes for 1 hr at 37C with THC or 11-hydroxyTHC caused a marked suppression of function in all 3 cell populations. Maximal suppression was obtained with 7.5-10..mu..g/ml of the drugs in medium containing 10% fetal bovine serum (FBS) or with 2-4..mu..g/ml in 1% FBS. These drug concentrations did not affect lymphoid cell viability or candida growth in the absence of lymphoid effector cells. Marijuana derivatives, therefore, are doubly dangerous in that opportunistic fungi such as C. albicans can grow in their presence while the effector cells that control fungal growth are readily inactivated.

  19. The inhibition of Candida species by selected essential oils and their synergism with amphotericin B.

    PubMed

    Rosato, Antonio; Vitali, Cesare; Gallo, Daniela; Balenzano, Luca; Mallamaci, Rosanna

    2008-08-01

    In this work we highlight a possible synergistic anti-Candida effect between Melaleuca alternifolia, Origanum vulgare and Pelargonium graveolens essential oils and the antifungal compound Amphotericin B. The antifungal activity was assessed using the agar dilution method in eleven Candida strains. The results obtained indicate the occurrence of a synergistic interaction between the essential oils under study and Amphotericin B. P. graveolens essential oil appeared to be the most effective, inhibiting all the Candida species evaluated by this study. PMID:18579358

  20. In vitro antimicrobial properties of coconut oil on Candida species in Ibadan, Nigeria.

    PubMed

    Ogbolu, D O; Oni, A A; Daini, O A; Oloko, A P

    2007-06-01

    The emergence of antimicrobial resistance, coupled with the availability of fewer antifungal agents with fungicidal actions, prompted this present study to characterize Candida species in our environment and determine the effectiveness of virgin coconut oil as an antifungal agent on these species. In 2004, 52 recent isolates of Candida species were obtained from clinical specimens sent to the Medical Microbiology Laboratory, University College Hospital, Ibadan, Nigeria. Their susceptibilities to virgin coconut oil and fluconazole were studied by using the agar-well diffusion technique. Candida albicans was the most common isolate from clinical specimens (17); others were Candida glabrata (nine), Candida tropicalis (seven), Candida parapsilosis (seven), Candida stellatoidea (six), and Candida krusei (six). C. albicans had the highest susceptibility to coconut oil (100%), with a minimum inhibitory concentration (MIC) of 25% (1:4 dilution), while fluconazole had 100% susceptibility at an MIC of 64 microg/mL (1:2 dilution). C. krusei showed the highest resistance to coconut oil with an MIC of 100% (undiluted), while fluconazole had an MIC of > 128 microg/mL. It is noteworthy that coconut oil was active against species of Candida at 100% concentration compared to fluconazole. Coconut oil should be used in the treatment of fungal infections in view of emerging drug-resistant Candida species.

  1. In vitro antimicrobial properties of coconut oil on Candida species in Ibadan, Nigeria.

    PubMed

    Ogbolu, D O; Oni, A A; Daini, O A; Oloko, A P

    2007-06-01

    The emergence of antimicrobial resistance, coupled with the availability of fewer antifungal agents with fungicidal actions, prompted this present study to characterize Candida species in our environment and determine the effectiveness of virgin coconut oil as an antifungal agent on these species. In 2004, 52 recent isolates of Candida species were obtained from clinical specimens sent to the Medical Microbiology Laboratory, University College Hospital, Ibadan, Nigeria. Their susceptibilities to virgin coconut oil and fluconazole were studied by using the agar-well diffusion technique. Candida albicans was the most common isolate from clinical specimens (17); others were Candida glabrata (nine), Candida tropicalis (seven), Candida parapsilosis (seven), Candida stellatoidea (six), and Candida krusei (six). C. albicans had the highest susceptibility to coconut oil (100%), with a minimum inhibitory concentration (MIC) of 25% (1:4 dilution), while fluconazole had 100% susceptibility at an MIC of 64 microg/mL (1:2 dilution). C. krusei showed the highest resistance to coconut oil with an MIC of 100% (undiluted), while fluconazole had an MIC of > 128 microg/mL. It is noteworthy that coconut oil was active against species of Candida at 100% concentration compared to fluconazole. Coconut oil should be used in the treatment of fungal infections in view of emerging drug-resistant Candida species. PMID:17651080

  2. Quantitative Investigation of Efficiency of Ultraviolet and Visible Light in Eradication of Candida albicans In Vitro

    PubMed Central

    Maver-Bišćanin, Mirela; Mravak-Stipetić, Marinka; Bukovski, Suzana; Bišćanin, Alen

    2014-01-01

    Abstract Objective: The aim of this study was to quantitatively investigate the efficiency of the ultraviolet (UV) and visible light in eradication of Candida albicans in vitro; in particular, to determine, for selected wavelengths, the specific eradication coefficients and thresholds in terms of energy density levels required to effect 3.0log10 and 4.0log10 reduction. Background data: Oral candidosis is the most common infection of the oral cavity and is caused by Candida species. The widespread use of topical and systemic antifungal agents as conventional treatment for oral candidosis has resulted in the development of resistance in C. albicans. Therefore, it has become necessary to develop alternative therapies for the treatment of oral candidosis. Methods: C. albicans ATCC® 90028™ was irradiated with 254 nm, 365 nm, 406 nm, 420 nm, and broadband Xe spectrum. For each wavelength, a fit of experimental data (survival fraction vs. applied energy density) with an exponential decay function enabled estimation of the specific eradication coefficients and thresholds. Results: Based on estimated specific efficiencies (Δ) and eradication thresholds (ET) of the investigated wavelengths, the ranking in eradication efficiency of C. albicans (most to least effective) is: 254 nm (Δ=6.1 mJ/cm−2, ET99.99=56 mJ/cm−2), broadband Xe spectrum (Δ=27.7 mJ/cm−2, ET99.99=255 mJ/cm−2), 365 nm (Δ=4.3 J/cm−2, ET99.99=39 J/cm−2), 420 nm (Δ=0.65 J/cm−2, ET99.99=6 J/cm−2), and 406 nm (Δ=11.4 J/cm−2, ET99.99=104 J/cm−2). Conclusions: The results provide insight into the wavelength-dependent dynamics of eradication of C. albicans. For each investigated wavelength, the eradication coefficient and corresponding eradication threshold were estimated. The observed different eradication efficiencies are consequence of different spectrally dependent inactivation mechanisms. The established methodology enables unambiguous quantitative

  3. In vitro antifungal and antibiofilm activities of halogenated quinoline analogues against Candida albicans and Cryptococcus neoformans.

    PubMed

    Zuo, Ran; Garrison, Aaron T; Basak, Akash; Zhang, Peilan; Huigens, Robert W; Ding, Yousong

    2016-08-01

    With the increasing prevalence of fungal infections coupled with emerging drug resistance, there is an urgent need for new and effective antifungal agents. Here we report the antifungal activities of 19 diverse halogenated quinoline (HQ) small molecules against Candida albicans and Cryptococcus neoformans. Four HQ analogues inhibited C. albicans growth with a minimum inhibitory concentration (MIC) of 100 nM, whilst 16 analogues effectively inhibited C. neoformans at MICs of 50-780 nM. Remarkably, two HQ analogues eradicated mature C. albicans and C. neoformans biofilms [minimum biofilm eradication concentration (MBEC) = 6.25-62.5 µM]. Several active HQs were found to penetrate into fungal cells, whilst one inactive analogue was unable to, suggesting that HQs elicit their antifungal activities through an intracellular mode of action. HQs are a promising class of small molecules that may be useful in future antifungal treatments. PMID:27256584

  4. Quinacrine inhibits Candida albicans growth and filamentation at neutral pH.

    PubMed

    Kulkarny, Vibhati V; Chavez-Dozal, Alba; Rane, Hallie S; Jahng, Maximillian; Bernardo, Stella M; Parra, Karlett J; Lee, Samuel A

    2014-12-01

    Candida albicans is a common cause of catheter-related bloodstream infections (CR-BSI), in part due to its strong propensity to form biofilms. Drug repurposing is an approach that might identify agents that are able to overcome antifungal drug resistance within biofilms. Quinacrine (QNC) is clinically active against the eukaryotic protozoan parasites Plasmodium and Giardia. We sought to investigate the antifungal activity of QNC against C. albicans biofilms. C. albicans biofilms were incubated with QNC at serially increasing concentrations (4 to 2,048 μg/ml) and assessed using a 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) assay in a static microplate model. Combinations of QNC and standard antifungals were assayed using biofilm checkerboard analyses. To define a mechanism of action, QNC was assessed for the inhibition of filamentation, effects on endocytosis, and pH-dependent activity. High-dose QNC was effective for the prevention and treatment of C. albicans biofilms in vitro. QNC with fluconazole had no interaction, while the combination of QNC and either caspofungin or amphotericin B demonstrated synergy. QNC was most active against planktonic growth at alkaline pH. QNC dramatically inhibited filamentation. QNC accumulated within vacuoles as expected and caused defects in endocytosis. A tetracycline-regulated VMA3 mutant lacking vacuolar ATPase (V-ATPase) function demonstrated increased susceptibility to QNC. These experiments indicate that QNC is active against C. albicans growth in a pH-dependent manner. Although QNC activity is not biofilm specific, QNC is effective in the prevention and treatment of biofilms. QNC antibiofilm activity likely occurs via several independent mechanisms: vacuolar alkalinization, inhibition of endocytosis, and impaired filamentation. Further investigation of QNC for the treatment and prevention of biofilm-related Candida CR-BSI is warranted. PMID:25288082

  5. Antifungal activity of naphthoquinoidal compounds in vitro against fluconazole-resistant strains of different Candida species: a special emphasis on mechanisms of action on Candida tropicalis.

    PubMed

    Neto, João B A; da Silva, Cecília R; Neta, Maria A S; Campos, Rosana S; Siebra, Janaína T; Silva, Rose A C; Gaspar, Danielle M; Magalhães, Hemerson I F; de Moraes, Manoel O; Lobo, Marina D P; Grangeiro, Thalles B; Carvalho, Tatiane S C; Diogo, Emilay B T; da Silva Júnior, Eufrânio N; Rodrigues, Felipe A R; Cavalcanti, Bruno C; Júnior, Hélio V N

    2014-01-01

    In recent decades, the incidence of candidemia in tertiary hospitals worldwide has substantially increased. These infections are a major cause of morbidity and mortality; in addition, they prolong hospital stays and raise the costs associated with treatment. Studies have reported a significant increase in infections by non-albicans Candida species, especially C. tropicalis. The number of antifungal drugs on the market is small in comparison to the number of antibacterial agents available. The limited number of treatment options, coupled with the increasing frequency of cross-resistance, makes it necessary to develop new therapeutic strategies. The objective of this study was to evaluate and compare the antifungal activities of three semisynthetic naphthofuranquinone molecules against fluconazole-resistant Candida spp. strains. These results allowed to us to evaluate the antifungal effects of three naphthofuranquinones on fluconazole-resistant C. tropicalis. The toxicity of these compounds was manifested as increased intracellular ROS, which resulted in membrane damage and changes in cell size/granularity, mitochondrial membrane depolarization, and DNA damage (including oxidation and strand breakage). In conclusion, the tested naphthofuranquinones (compounds 1-3) exhibited in vitro cytotoxicity against fluconazole-resistant Candida spp. strains.

  6. In Vitro and In Vivo Antifungal Activity of Lichochalcone-A against Candida albicans Biofilms

    PubMed Central

    Seleem, Dalia; Benso, Bruna; Noguti, Juliana; Pardi, Vanessa; Murata, Ramiro Mendonça

    2016-01-01

    Oral candidiasis (OC) is an opportunistic fungal infection with high prevalence among immunocompromised patients. Candida albicans is the most common fungal pathogen responsible for OC, often manifested in denture stomatitis and oral thrush. Virulence factors, such as biofilms formation and secretion of proteolytic enzymes, are key components in the pathogenicity of C. albicans. Given the limited number of available antifungal therapies and the increase in antifungal resistance, demand the search for new safe and effective antifungal treatments. Lichochalcone-A is a polyphenol natural compound, known for its broad protective activities, as an antimicrobial agent. In this study, we investigated the antifungal activity of lichochalcone-A against C. albicans biofilms both in vitro and in vivo. Lichochalcone-A (625 μM; equivalent to 10x MIC) significantly reduced C. albicans (MYA 2876) biofilm growth compared to the vehicle control group (1% ethanol), as indicated by the reduction in the colony formation unit (CFU)/ml/g of biofilm dry weight. Furthermore, proteolytic enzymatic activities of proteinases and phospholipases, secreted by C. albicans were significantly decreased in the lichochalcone-A treated biofilms. In vivo model utilized longitudinal imaging of OC fungal load using a bioluminescent-engineered C. albicans (SKCa23-ActgLUC) and coelenterazine substrate. Mice treated with lichochalcone-A topical treatments exhibited a significant reduction in total photon flux over 4 and 5 days post-infection. Similarly, ex vivo analysis of tongue samples, showed a significant decrease in CFU/ml/mg in tongue tissue sample of lichochalcone-A treated group, which suggest the potential of lichochalcone-A as a novel antifungal agent for future clinical use. PMID:27284694

  7. In vitro evaluation of antifungal activity of monolaurin against Candida albicans biofilms.

    PubMed

    Seleem, Dalia; Chen, Emily; Benso, Bruna; Pardi, Vanessa; Murata, Ramiro M

    2016-01-01

    Monolaurin (also known as glycerol monolaurate) is a natural compound found in coconut oil and is known for its protective biological activities as an antimicrobial agent. The nature of oral candidiasis and the increased antifungal resistance demand the search for novel antifungal therapeutic agents. In this study, we examine the antifungal activity of monolaurin against Candida albicans biofilms (strain ATCC:SC5314/MYA2876) in vitro and investigate whether monolaurin can alter gene expression of host inflammatory cytokines, IL-1α and IL-1β. In a co-culture model, oral fibroblast cells were cultured simultaneously with C. albicans for 24 hrs followed by the exposure to treatments of monolaurin (3.9-2,500 µM), positive control fluconazole (32.2 µM), and vehicle control group (1% ethanol), which was a model used to evaluate the cytotoxicity of monolaurin on fibroblasts as well as to analyze morphological characteristics of biofilms through fluorescence microscopy. In addition, the co-culture model was used for RNA extraction of oral fibroblasts to assess gene expression of host inflammatory cytokines, using quantitative real-time PCR. Our results showed the MIC and MFC of monolaurin were in the range 62.5-125 µM and 125-250 µM, respectively. Biofilm antifungal assay showed significant reduction in Log (CFU/ml) of biofilms treated with 1,250 and 2,500 µM of 1-monolaurin when compared to the control groups . There was also a significant down-regulation of IL-1α and IL-1β in the co-culture treated with monolaurin. It can be concluded that monolaurin has a potential antifungal activity against C. albicans and can modulate the pro-inflammatory response of the host. PMID:27366648

  8. In Vitro and In Vivo Antifungal Activity of Lichochalcone-A against Candida albicans Biofilms.

    PubMed

    Seleem, Dalia; Benso, Bruna; Noguti, Juliana; Pardi, Vanessa; Murata, Ramiro Mendonça

    2016-01-01

    Oral candidiasis (OC) is an opportunistic fungal infection with high prevalence among immunocompromised patients. Candida albicans is the most common fungal pathogen responsible for OC, often manifested in denture stomatitis and oral thrush. Virulence factors, such as biofilms formation and secretion of proteolytic enzymes, are key components in the pathogenicity of C. albicans. Given the limited number of available antifungal therapies and the increase in antifungal resistance, demand the search for new safe and effective antifungal treatments. Lichochalcone-A is a polyphenol natural compound, known for its broad protective activities, as an antimicrobial agent. In this study, we investigated the antifungal activity of lichochalcone-A against C. albicans biofilms both in vitro and in vivo. Lichochalcone-A (625 μM; equivalent to 10x MIC) significantly reduced C. albicans (MYA 2876) biofilm growth compared to the vehicle control group (1% ethanol), as indicated by the reduction in the colony formation unit (CFU)/ml/g of biofilm dry weight. Furthermore, proteolytic enzymatic activities of proteinases and phospholipases, secreted by C. albicans were significantly decreased in the lichochalcone-A treated biofilms. In vivo model utilized longitudinal imaging of OC fungal load using a bioluminescent-engineered C. albicans (SKCa23-ActgLUC) and coelenterazine substrate. Mice treated with lichochalcone-A topical treatments exhibited a significant reduction in total photon flux over 4 and 5 days post-infection. Similarly, ex vivo analysis of tongue samples, showed a significant decrease in CFU/ml/mg in tongue tissue sample of lichochalcone-A treated group, which suggest the potential of lichochalcone-A as a novel antifungal agent for future clinical use. PMID:27284694

  9. In vitro expression of Candida albicans alcohol dehydrogenase genes involved in acetaldehyde metabolism.

    PubMed

    Bakri, M M; Rich, A M; Cannon, R D; Holmes, A R

    2015-02-01

    Alcohol consumption is a risk factor for oral cancer, possibly via its conversion to acetaldehyde, a known carcinogen. The oral commensal yeast Candida albicans may be one of the agents responsible for this conversion intra-orally. The alcohol dehydrogenase (Adh) family of enzymes are involved in acetaldehyde metabolism in yeast but, for C. albicans it is not known which family member is responsible for the conversion of ethanol to acetaldehyde. In this study we determined the expression of mRNAs from three C. albicans Adh genes (CaADH1, CaADH2 and CaCDH3) for cells grown in different culture media at different growth phases by Northern blot analysis and quantitative reverse transcription polymerase chain reaction. CaADH1 was constitutively expressed under all growth conditions but there was differential expression of CaADH2. CaADH3 expression was not detected. To investigate whether CaAdh1p or CaAdh2p can contribute to alcohol catabolism in C. albicans, each gene from the reference strain C. albicans SC5314 was expressed in Saccharomyces cerevisiae. Cell extracts from an CaAdh1p-expressing S. cerevisiae recombinant, but not an CaAdh2p-expressing recombinant, or an empty vector control strain, possessed ethanol-utilizing Adh activity above endogenous S. cerevisiae activity. Furthermore, expression of C. albicans Adh1p in a recombinant S. cerevisiae strain in which the endogenous ScADH2 gene (known to convert ethanol to acetaldehyde in this yeast) had been deleted, conferred an NAD-dependent ethanol-utilizing, and so acetaldehyde-producing, Adh activity. We conclude that CaAdh1p is the enzyme responsible for ethanol use under in vitro growth conditions, and may contribute to the intra-oral production of acetaldehyde.

  10. In vitro evaluation of antifungal activity of monolaurin against Candida albicans biofilms

    PubMed Central

    Benso, Bruna; Pardi, Vanessa

    2016-01-01

    Monolaurin (also known as glycerol monolaurate) is a natural compound found in coconut oil and is known for its protective biological activities as an antimicrobial agent. The nature of oral candidiasis and the increased antifungal resistance demand the search for novel antifungal therapeutic agents. In this study, we examine the antifungal activity of monolaurin against Candida albicans biofilms (strain ATCC:SC5314/MYA2876) in vitro and investigate whether monolaurin can alter gene expression of host inflammatory cytokines, IL-1α and IL-1β. In a co-culture model, oral fibroblast cells were cultured simultaneously with C. albicans for 24 hrs followed by the exposure to treatments of monolaurin (3.9–2,500 µM), positive control fluconazole (32.2 µM), and vehicle control group (1% ethanol), which was a model used to evaluate the cytotoxicity of monolaurin on fibroblasts as well as to analyze morphological characteristics of biofilms through fluorescence microscopy. In addition, the co-culture model was used for RNA extraction of oral fibroblasts to assess gene expression of host inflammatory cytokines, using quantitative real-time PCR. Our results showed the MIC and MFC of monolaurin were in the range 62.5–125 µM and 125–250 µM, respectively. Biofilm antifungal assay showed significant reduction in Log (CFU/ml) of biofilms treated with 1,250 and 2,500 µM of 1-monolaurin when compared to the control groups . There was also a significant down-regulation of IL-1α and IL-1β in the co-culture treated with monolaurin. It can be concluded that monolaurin has a potential antifungal activity against C. albicans and can modulate the pro-inflammatory response of the host. PMID:27366648

  11. Self-assembled amphotericin B-loaded polyglutamic acid nanoparticles: preparation, characterization and in vitro potential against Candida albicans

    PubMed Central

    Zia, Qamar; Khan, Aijaz Ahmed; Swaleha, Zubair; Owais, Mohammad

    2015-01-01

    In the present study, we developed a self-assembled biodegradable polyglutamic acid (PGA)-based formulation of amphotericin B (AmB) and evaluated its in vitro antifungal potential against Candida albicans. The AmB-loaded PGA nanoparticles were prepared in-house and had a mean size dimension of around 98±2 nm with a zeta potential of −35.2±7.3 mV. Spectroscopic studies revealed that the drug predominantly acquires an aggregated form inside the formulation with an aggregation ratio above 2. The PGA-based AmB formulation was shown to be highly stable in phosphate-buffered saline as well as in serum (only 10%–20% of the drug was released after 10 days). The AmB–PGA nanoparticles were less toxic to red blood cells (<15% lysis at an AmB concentration of 100 μg/mL after 24 hours) when compared with Fungizone®, a commercial antifungal product. An MTT assay showed that the viability of mammalian cells (KB and RAW 264.7) was negligibly affected at AmB concentrations as high as 200 μg/mL. Histopathological examination of mouse kidney revealed no signs of tissue necrosis. The AmB–PGA formulation showed potent antimicrobial activity similar to that of Fungizone against C. albicans. Interestingly, AmB-bearing PGA nanoparticles were found to inhibit biofilm formation to a considerable extent. In summary, AmB–PGA nanoparticles showed highly attenuated toxicity when compared with Fungizone, while retaining equivalent active antifungal properties. This study indicates that the AmB–PGA preparation could be a promising treatment for various fungal infections. PMID:25784804

  12. Comparison of the in vitro activity of echinocandins against Candida albicans, Candida dubliniensis, and Candida africana by time-kill curves.

    PubMed

    Gil-Alonso, Sandra; Jauregizar, Nerea; Cantón, Emilia; Eraso, Elena; Quindós, Guillermo

    2015-05-01

    Candida albicans remains the most common fungal pathogen. This species is closely related to 2 phenotypically similar cryptic species, Candida dubliniensis and Candida africana. This study aims to compare the antifungal activities of echinocandins against 7 C. albicans, 5 C. dubliniensis, and 2 C. africana strains by time-kill methodology. MIC values were similar for the 3 species; however, differences in killing activity were observed among species, isolates, and echinocandins. Echinocandins produced weak killing activity against the 3 species. In all drugs, the fungicidal endpoint (99.9% mortality) was reached at ≤31 h with ≥0.5 μg/mL for anidulafungin in 4 C. albicans and 1 C. dubliniensis, for caspofungin in 1 C. albicans and 2 C. dubliniensis, and for micafungin in 4 C. albicans and 1 C. dubliniensis. None of echinocandins showed lethality against C. africana. Identification of these new cryptic species and time-kill studies would be recommendable when echinocandin treatment fails.

  13. An in vitro evaluation of Candida tropicalis infectivity using human cell monolayers.

    PubMed

    Negri, Melyssa; Botelho, Cláudia; Silva, Sónia; Lopes, Luís Miguel Reis Henriques; Henriques, Mariana; Azeredo, Joana; Oliveira, Rosário

    2011-09-01

    The aim of this study was to investigate the interaction of Candida tropicalis with three different human cell lines: TCC-SUP (epithelial cells from urinary bladder), HeLa (epithelial cells from cervical carcinoma) and Caco-2 (epithelial cells from colorectal adenocarcinoma). In particular we sought to assess the degree of cell damage and activity reduction induced by C. tropicalis adhesion and the role of secreted aspartyl proteinase (SAP) gene expression in this process. Two C. tropicalis strains were used: the reference strain ATCC 750 and a clinical isolate from urine (U69). The ability of C. tropicalis to adhere to a confluent layer of human cells was determined using an adaptation of the crystal violet staining method; cell damage and cell activity inhibition induced by the adhesion of C. tropicalis were assessed by measuring lactate dehydrogenase and tetrazolium salt (MTS) reduction, respectively. C. tropicalis SAP gene expression was determined by real-time PCR. Both C. tropicalis strains were able to adhere to the different human cells, although in a strain- and cell-line-dependent manner. Concerning the cellular response to C. tropicalis, the highest inhibition of cell activity was obtained for Caco-2, followed by TCC-SUP and HeLa cells. The highest percentage of cell damage (around 14 %) was observed for TCC-SUP cells in contact with the U69 isolate and for Caco-2 in contact with the reference strain. Real-time PCR analysis revealed a wide range of expression profiles of SAP genes for both C. tropicalis strains in contact with the different types of epithelial cells. SAPT3 was the gene expressed at the highest level for both C. tropicalis strains in contact with the three human epithelial cell lines. The results highlight that the response of human cells to C. tropicalis adhesion, as well as production of SAPs, is dependent on both the strain and the epithelial cell line.

  14. Natural cell-mediated cytotoxicity against Candida albicans induced by cyclophosphamide: nature of the in vitro cytotoxic effector.

    PubMed Central

    Baccarini, M; Bistoni, F; Puccetti, P; Garaci, E

    1983-01-01

    We have recently reported the in vivo modulation of resistance to experimental Candida albicans infection by cyclophosphamide (150 mg/kg intraperitoneally) in mice and have shown that increased resistance to the microbial challenge occurs 12 to 21 days after treatment with the drug (Bistoni et al., Infect. Immun. 40: 46-55, 1983). The event is accompanied by the appearance of a highly candidacidal cell population in the spleen and the activation of a subpopulation of natural cytotoxic effectors reactive in vitro against YAC-1 tumor cells. We now provide evidence that these anti-YAC-1 cytotoxic effectors are clearly distinct from the cyclophosphamide-induced candidacidal effectors, which seem to belong to a macrophage-monocyte lineage. The enhanced cytotoxic activity induced by cyclophosphamide was not restricted to C. albicans but was also exerted against a panel of Candida strains. PMID:6352489

  15. Identification of the aminocatechol A-3253 as an in vitro poison of DNA topoisomerase I from Candida albicans.

    PubMed Central

    Fostel, J; Montgomery, D

    1995-01-01

    The aminocatechol A-3253 is active against several pathogenic fungi, including Candida albicans, Cryptococcus albidus, and Aspergillus niger. A-3253 interferes with both the in vitro biosynthesis of (1,3)-beta-glucan and the activity of topoisomerases I isolated from Candida spp. It is likely that one or more of the enzymes involved in glucan biosynthesis rather than topoisomerase I is the primary intracellular target of A-3253, since a strain of Saccharomyces cerevisiae lacking topoisomerase I is as susceptible to A-3253 as cells containing wild-type levels of topoisomerase I. However, the interaction of A-3253 with topoisomerase I in vitro is of interest since the Candida topoisomerase is more susceptible to A-3253 than is the topoisomerase I isolated from human HeLa cells. A-3253 is both a reversible inhibitor of topoisomerase I catalysis and a reversible poison of topoisomerase I, and in both reactions the fungal topoisomerase I is more susceptible than the human topoisomerase I to A-3253. In contrast, an earlier study found that the human topoisomerase I is more susceptible than the fungal topoisomerase to camptothecin (J. M. Fostel, D. A. Montgomery, and L. L. Shen, Antimicrob. Agents Chemother. 36:2131-2138, 1992). Taken together with the response to camptothecin, the greater susceptibility of the Candida topoisomerase I to A-3253 suggests that there are structural differences between the human and fungal type I topoisomerases which can likely be exploited to allow for the development of antifungal agents which act against the fungal topoisomerase and which have minimal activity against the human enzyme. PMID:7793856

  16. Favipiravir (T-705) inhibits in vitro norovirus replication.

    PubMed

    Rocha-Pereira, J; Jochmans, D; Dallmeier, K; Leyssen, P; Nascimento, M S J; Neyts, J

    2012-08-10

    Human noroviruses are the primary cause of foodborne gastroenteritis. Potent and safe inhibitors are needed for the treatment/prophylaxis of norovirus infections. We demonstrate that Favipiravir [T-705, a drug in advanced clinical development for the treatment of infections with the influenza virus] inhibits in vitro murine norovirus replication. Time-of-drug addition studies reveal that T-705 exerts its activity at a time-point that coincides with onset of viral RNA synthesis, which is in line with the viral polymerase as the presumed target.

  17. Nucleosomes Inhibit Cas9 Endonuclease Activity in Vitro.

    PubMed

    Hinz, John M; Laughery, Marian F; Wyrick, John J

    2015-12-01

    During Cas9 genome editing in eukaryotic cells, the bacterial Cas9 enzyme cleaves DNA targets within chromatin. To understand how chromatin affects Cas9 targeting, we characterized Cas9 activity on nucleosome substrates in vitro. We find that Cas9 endonuclease activity is strongly inhibited when its target site is located within the nucleosome core. In contrast, the nucleosome structure does not affect Cas9 activity at a target site within the adjacent linker DNA. Analysis of target sites that partially overlap with the nucleosome edge indicates that the accessibility of the protospacer-adjacent motif (PAM) is the critical determinant of Cas9 activity on a nucleosome.

  18. The effect of parenteral alimentation fluid, undiluted with saline or fresh sera, on the growth of Candida albicans in vitro at 37 degrees C.

    PubMed

    Lederer, T; Rippon, J; Baldwin, S; Pachman, L M

    1975-02-28

    Parenteral alimentation is often complicated by Candida albicans infection which may be fatal. This study investigated the effect of alimentation fluid (Aminosol) on C. albicans' growth in vitro. It was found that concentrated Aminosol (1400 millisomoles) maintained C. albicans in a viable state but inhibited replication. Dilution of alimentation fluid to physiological concentrations (300 milliosmoles) with either saline or aged pooled normal sera promoted in vitro growth of C. albicans which was equivalent to that obtained in BHI broth and was slightly less than that obtained in Sabouraud's broth. The effects of fresh sera with full complement activity were also investigated. In fresh sera appropriately diluted with physiological saline, some clumping of the yeasts was observed and all formed germ tubes. Growth as defined by budding or the formation of hyphae was inhibited. When Aminosol was diluted to 300 milliosomoles with fresh sera, all yeasts were noted to be in clumps with germ tubes as well as continually growing hyphae. Growth was approximately equal to that seen in Aminosol similarly diluted with saline.

  19. High-Frequency, In Vitro Reversible Switching of Candida lusitaniae Clinical Isolates from Amphotericin B Susceptibility to Resistance

    PubMed Central

    Yoon, Stephanie A.; Vazquez, Jose A.; Steffan, Paul E.; Sobel, Jack D.; Akins, Robert A.

    1999-01-01

    Recent studies have revealed an increase in the incidence of serious infections caused by non-albicans Candida species. Candida lusitaniae is of special interest because of its sporadic resistance to amphotericin B (AmB). The present in vitro study demonstrated that, unlike other Candida species, C. lusitaniae isolates frequently generated AmB-resistant lineages form previously susceptible colonies. Cells switching from a resistant colony to a susceptible phenotype were also detected after treatment with either UV light, heat shock, or exposure to whole blood, all of which increased the frequency of switching. In some C. lusitaniae lineages, after a cell switched to a resistant phenotype, the resistant phenotype was stable; in other lineages, colonies were composed primarily of AmB-susceptible cells. Although resistant and susceptible lineages were identical in many aspects, their cellular morphologies were dramatically different. Switching mechanisms that involve exposure to antifungals may have an impact on antifungal therapeutic strategies as well as on standardized susceptibility testing of clinical yeast specimens. PMID:10103188

  20. Novel protein inhibits in vitro precipitation of calcium carbonate.

    PubMed

    Burgess, S K; Carey, D M; Oxendine, S L

    1992-09-01

    Organic molecules both coexist and interact with inorganic crystal lattices in biomineralizing tissues. Mineral precipitation and crystal morphology are tightly regulated by the actions of these molecules. Polyacrylamide gel electrophoresis studies on water soluble extracts from the cuticle of Callinectes sapidus (Atlantic blue crab) reveal the presence, in unmineralized nascent premolt cuticle, of proteins which are absent in the mineralized postmolt cuticle. In the present studies, homogenates from both premolt and postmolt C. sapidus cuticles have been tested for their effect on the in vitro precipitation of calcium carbonate. The role of protein in this process was determined by heat pretreatment and trypsin pretreatment of the cuticle homogenates prior to the precipitation assay. The results from these experiments indicate that proteins, with molecular weights of approximately 75,000 and between 10,000 and 20,000, concentrated in the C. sapidus premolt cuticle, inhibit calcium carbonate precipitation in vitro. The inhibitory activity of these proteins appears to be a result of specific interactions since trypsin, myoglobin, and ovalbumin are not inhibitory. The presence of lower amounts of these inhibitory proteins in C. sapidus postmolt cuticle may be responsible for the subsequent mineralization of this tissue.

  1. Curcumin associated magnetite nanoparticles inhibit in vitro melanoma cell growth.

    PubMed

    de Souza, Fernanda França; dos Santos, Michelly Christine; dos Passos, Debora Cristina Silva; Lima, Emilia Celma de Oliveira; Guillo, Lidia Andreu

    2011-09-01

    Curcumin is a natural product possessing therapeutic properties but the low water solubility of this compound limits its use. We have successfully incorporated curcumin into a bilayer of dodecanoic acid attached to magnetite nanoparticles in an effort to maximize solubility and delivery efficiency. Curcumin/magnetite nanoparticles were characterized using diffused reflectance infra-red fourier transform spectroscopy (DRIFTS) and X-ray powder diffraction (XRD). Moreover curcumin associated magnetite nanoparticles inhibited in vitro melanoma cell growth. An inhibitory concentration (IC50) of 66.0 +/- 3.0 microM (48 +/- 2.2 microg-iron/mL) was observed for the curcumin/magnetite nanoparticles. Fluorescent microscopy revealed that curcumin associated magnetite nanoparticles were internalized by the melanoma cells and remained in the cytoplasm. The curcumin/magnetic nanoparticles synthesized in this study possess magnetic and water solubility properties making this a novel curcumin formulation with therapeutic potential.

  2. In vitro antifungal activity of silver nanoparticles against fluconazole-resistant Candida species.

    PubMed

    Artunduaga Bonilla, Jhon J; Paredes Guerrero, Daissy J; Sánchez Suárez, Clara I; Ortiz López, Claudia C; Torres Sáez, Rodrigo G

    2015-11-01

    Nowadays, current advances in nanotechnology constitute a promising alternative in the development of new antimicrobial agents. Silver nanoparticles (AgNPs) are some very interesting products currently provided by available nanotechnology for control of microbial infection. In the present study, AgNPs were synthesized by eco-friendly method, using cysteine as a reducing agent. Also, antifungal activity against Candida species with resistance to fluconazole was evaluated through determination of Minimum Inhibitory Concentration (MIC50) according to protocol M27-A3 of Clinical and Laboratory Standards Institute (CLSI) and Minimum Fungicide Concentration (MFC). This study was carried out with strains Candida krusei and Candida glabrata. As a result, the formation of spherical nanoparticles was obtained with mean sizes of 19 nm and positive surface charge. Values of MIC50 were 0.1 µg ml(-1) AgNPs for the studied species, and MFC were 0.25 and 0.5 µg ml(-1) for C. glabrata and C. krusei, respectively. The AgNPs synthesized showed cytotoxic effect in 50% of Murine Fibroblast Cells (CC50) at a mean concentrations of 10 µg ml(-1) (100 times higher than MIC50). Consequently, AgNPs could be considered as an alternative potential in the development of new antifungal agents with minimum cytotoxicity in fibroblasts and lethal action on Candida species with resistance to conventional antifungal compounds.

  3. The inhibition of Candida albicans by selected essential oils and their major components.

    PubMed

    Tampieri, Maria Paola; Galuppi, Roberta; Macchioni, Fabio; Carelle, Maria Stella; Falcioni, Laura; Cioni, Pier Luigi; Morelli, Ivano

    2005-04-01

    Many volatile oils are known to possess antifungal properties and are potentially applicable as antimycotic agents. By studying the efficacy of essential oils against different pathogenic mycetes, we have evaluated the in-vitro inhibiting activity of some essential oils and their main constituents against a strain of Candida albicans. Sixteen commercial essential oils and forty-two pure constituents (alcohols, aldehydes, ketons, phenols and hydrocarbons), were tested by using a semisolid agar antifungal susceptibility (SAAS) method. Gas chromatography/mass spectroscopy analyses of the oils tested were performed. The essential oils of Origanum vulgare, Satureja montana, Mentha piperita, Cinnamomum verum, Cymbopogon flexuosus showed maximum inhibitory activity (MIC = 500 ppm) after 7 days. According to the results of the examination of pure constituents, beta-phellandrene proved to be the most interesting component among cyclic monoterpenic hydrocarbons as it showed a strong activity (MIC = 50 ppm). The most active of phenols was carvacrol (MIC 100 ppm). The open-chain alcohol 1-decanol was the most active of alcohols at 50 ppm. Finally, among aldehydes, a strong activity was shown by trans-cynnamaldehyde (MIC 50 ppm).

  4. In vitro antifungal activity of 2-(2'-hydroxy-5'-aminophenyl)benzoxazole in Candida spp. strains.

    PubMed

    Daboit, Tatiane Caroline; Stopiglia, Cheila Denise Ottonelli; Carissimi, Mariana; Corbellini, Valeriano Antonio; Stefani, Valter; Scroferneker, Maria Lúcia

    2009-11-01

    The development of azole antifungals has allowed for the treatment of several fungal infections. However, the use of these compounds is restricted because of their hepatotoxicity or because they need to be administered together with other drugs in order to prevent resistance to monotherapy. Benzoxazole derivatives are among the most thriving molecular prototypes for the development of antifungal agents. 2-(2'-hydroxyphenyl) benzoxazoles are versatile molecules that emit fluorescence and have antibacterial, antiviral and antifungal properties. 2-(2'-hydroxy-5'-aminophenyl) benzoxazole (HAMBO) was tested against Candida yeast. The inhibition provided by HAMBO was lower than that of fluconazole, showing low antifungal activity against Candida spp., but equivalent to that of benzoxazoles tested in similar studies. HAMBO showed fungistatic activity against all analysed strains. This class of novel benzoxazole compounds may be used as template to produce better antifungal drugs.

  5. In vitro inhibition of mitochondrial respiratory rate by antidepressants.

    PubMed

    Hroudová, Jana; Fišar, Zdeněk

    2012-09-18

    Mitochondria represent a possible drug target with unexplored therapeutic and toxicological potential. The possibility was suggested that antidepressants, mood stabilizers and other drugs may show some therapeutic and/or toxic effects through their action on mitochondrial functions. There are no sufficient data about the effect of these drugs on mitochondrial respiration in the brain. We investigated the in vitro effects of amitriptyline, fluoxetine, tianeptine, ketamine, lithium, valproate, olanzapine, chlorpromazine and propranolol on mitochondrial respiration in crude mitochondrial fractions of pig brains. Respiration was energized using substrates of complex I or complex II and dose dependent drug-induced changes in mitochondrial respiratory rate were measured by high-resolution respirometry. Antidepressants, but not mood stabilizers, ketamine and propranolol were found to inhibit mitochondrial respiratory rate. The effective dose of antidepressants reaching half the maximal respiratory rate was in the range of 0.07-0.46 mmol/L. Partial inhibition was found for all inhibitors. Differences between individual drugs with similar physicochemical properties indicate selectivity of drug-induced changes in mitochondrial respiratory rate. Our findings suggest that mood stabilizers do not interfere with brain mitochondrial respiration, whereas direct mitochondrial targeting is involved in mechanisms of action of pharmacologically different antidepressants. PMID:22842584

  6. In vitro inhibition of mitochondrial respiratory rate by antidepressants.

    PubMed

    Hroudová, Jana; Fišar, Zdeněk

    2012-09-18

    Mitochondria represent a possible drug target with unexplored therapeutic and toxicological potential. The possibility was suggested that antidepressants, mood stabilizers and other drugs may show some therapeutic and/or toxic effects through their action on mitochondrial functions. There are no sufficient data about the effect of these drugs on mitochondrial respiration in the brain. We investigated the in vitro effects of amitriptyline, fluoxetine, tianeptine, ketamine, lithium, valproate, olanzapine, chlorpromazine and propranolol on mitochondrial respiration in crude mitochondrial fractions of pig brains. Respiration was energized using substrates of complex I or complex II and dose dependent drug-induced changes in mitochondrial respiratory rate were measured by high-resolution respirometry. Antidepressants, but not mood stabilizers, ketamine and propranolol were found to inhibit mitochondrial respiratory rate. The effective dose of antidepressants reaching half the maximal respiratory rate was in the range of 0.07-0.46 mmol/L. Partial inhibition was found for all inhibitors. Differences between individual drugs with similar physicochemical properties indicate selectivity of drug-induced changes in mitochondrial respiratory rate. Our findings suggest that mood stabilizers do not interfere with brain mitochondrial respiration, whereas direct mitochondrial targeting is involved in mechanisms of action of pharmacologically different antidepressants.

  7. First Human Model of In Vitro Candida albicans Persistence within Granuloma for the Reliable Study of Host-Fungi Interactions

    PubMed Central

    Alvarez-Rueda, Nidia; Albassier, Marjorie; Allain, Sophie; Deknuydt, Florence; Altare, Frédéric; Le Pape, Patrice

    2012-01-01

    Backgound The balance between human innate immune system and Candida albicans virulence signaling mechanisms ultimately dictates the outcome of fungal invasiveness and its pathology. To better understand the pathophysiology and to identify fungal virulence-associated factors in the context of persistence in humans, complex models are indispensable. Although fungal virulence factors have been extensively studied in vitro and in vivo using different immune cell subsets and cell lines, it is unclear how C. albicans survives inside complex tissue granulomas. Methodology/Principal Finding We developed an original model of in vitro human granuloma, reproducing the natural granulomatous response to C. albicans. Persistent granulomas were obtained when the ratio of phagocytes to fungi was high. This in vitro fungal granuloma mimics natural granulomas, with infected macrophages surrounded by helper and cytotoxic T lymphocytes. A small proportion of granulomas exhibited C. albicans hyphae. Histological and time-lapse analysis showed that C. albicans blastoconidia were located within the granulomas before hyphae formation. Using staining techniques, fungal load calculations, as well as confocal and scanning electron microscopy, we describe the kinetics of fungal granuloma formation. We provide the first direct evidence that C. albicans are not eliminated by immunocompetent cells inside in vitro human granulomas. In fact, after an initial candicidal period, the remaining yeast proliferate and persist under very complex immune responses. Conclusions/Significance Using an original in vitro model of human fungal granuloma, we herein present the evidence that C. albicans persist and grow into immunocompetent granulomatous structures. These results will guide us towards a better understanding of fungal invasiveness and, henceforth, will also help in the development of better strategies for its control in human physiological conditions. PMID:22768252

  8. Leptin inhibits testosterone secretion from adult rat testis in vitro.

    PubMed

    Tena-Sempere, M; Pinilla, L; González, L C; Diéguez, C; Casanueva, F F; Aguilar, E

    1999-05-01

    Leptin, the product of the ob gene, has emerged recently as a pivotal signal in the regulation of fertility. Although the actions of leptin in the control of reproductive function are thought to be exerted mainly at the hypothalamic level, the potential direct effects of leptin at the pituitary and gonadal level have been poorly characterised. In the present study, we first assessed the ability of leptin to regulate testicular testosterone secretion in vitro. Secondly, we aimed to evaluate whether leptin can modulate basal gonadotrophin and prolactin (PRL) release by incubated hemi-pituitaries from fasted male rats. To attain the first goal, testicular slices from prepubertal and adult rats were incubated with increasing concentrations (10(-9)-10(-7) M) of recombinant leptin. Assuming that in vitro testicular responsiveness to leptin may be dependent on the background leptin levels, testicular tissue from both food-deprived and normally-fed animals was used. Furthermore, leptin modulation of stimulated testosterone secretion was evaluated by incubation of testicular samples with different doses of leptin in the presence of 10 IU human chorionic gonadotrophin (hCG). In addition, analysis of leptin actions on pituitary function was carried out using hemi-pituitaries from fasted adult male rats incubated in the presence of increasing concentrations (10(-9)-10(-7) M) of recombinant leptin. Serum testosterone levels, and basal and hCG-stimulated testosterone secretion by incubated testicular tissue were significantly decreased by fasting in prepubertal and adult male rats. However, a significant reduction in circulating LH levels was only evident in adult fasted rats. Doses of 10(-9)-10(-7) M leptin had no effect on basal or hCG-stimulated testosterone secretion by testes from prepubertal rats, regardless of the nutritional state of the donor animal. In contrast, leptin significantly decreased basal and hCG-induced testosterone secretion by testes from fasted and fed

  9. Leptin inhibits testosterone secretion from adult rat testis in vitro.

    PubMed

    Tena-Sempere, M; Pinilla, L; González, L C; Diéguez, C; Casanueva, F F; Aguilar, E

    1999-05-01

    Leptin, the product of the ob gene, has emerged recently as a pivotal signal in the regulation of fertility. Although the actions of leptin in the control of reproductive function are thought to be exerted mainly at the hypothalamic level, the potential direct effects of leptin at the pituitary and gonadal level have been poorly characterised. In the present study, we first assessed the ability of leptin to regulate testicular testosterone secretion in vitro. Secondly, we aimed to evaluate whether leptin can modulate basal gonadotrophin and prolactin (PRL) release by incubated hemi-pituitaries from fasted male rats. To attain the first goal, testicular slices from prepubertal and adult rats were incubated with increasing concentrations (10(-9)-10(-7) M) of recombinant leptin. Assuming that in vitro testicular responsiveness to leptin may be dependent on the background leptin levels, testicular tissue from both food-deprived and normally-fed animals was used. Furthermore, leptin modulation of stimulated testosterone secretion was evaluated by incubation of testicular samples with different doses of leptin in the presence of 10 IU human chorionic gonadotrophin (hCG). In addition, analysis of leptin actions on pituitary function was carried out using hemi-pituitaries from fasted adult male rats incubated in the presence of increasing concentrations (10(-9)-10(-7) M) of recombinant leptin. Serum testosterone levels, and basal and hCG-stimulated testosterone secretion by incubated testicular tissue were significantly decreased by fasting in prepubertal and adult male rats. However, a significant reduction in circulating LH levels was only evident in adult fasted rats. Doses of 10(-9)-10(-7) M leptin had no effect on basal or hCG-stimulated testosterone secretion by testes from prepubertal rats, regardless of the nutritional state of the donor animal. In contrast, leptin significantly decreased basal and hCG-induced testosterone secretion by testes from fasted and fed

  10. An easy and economical in vitro method for the formation of Candida albicans biofilms under continuous conditions of flow.

    PubMed

    Uppuluri, Priya; Lopez-Ribot, Jose L

    2010-01-01

    Candida albicans can develop biofilms on medical devices and these biofilms are most often nourished by a continuous flow of body fluids and subjected to shear stress forces. While many C. albicans biofilm studies have been carried out using in vitro static models, more limited information is available for biofilms developed under conditions of flow. We have previously described a simple flow biofilm model (SFB) for the development of C. albicans biofilms under conditions of continuous media flow. Here, we recount in detail from a methodological perspective, this model that can be assembled easily using materials commonly available in most microbiological laboratories. The entire procedure takes approximately two days to complete. Biofilms developed using this system are robust, and particularly suitable for studies requiring large amounts of biofilm cells for downstream analyses. This methodology simplifies biofilm formation under continuous replenishment of nutrients. Moreover, this technique mimics in vivo flow conditions, thereby making it physiologically more relevant than the currently dominant static models.

  11. In vitro synergistic activity of diketopiperazines alone and in combination with amphotericin B or clotrimazole against Candida albicans.

    PubMed

    Kumar, S Nishanth; Nambisan, Bala; Mohandas, C; Sundaresan, A

    2013-11-01

    The synergistic anticandidal activity of three diketopiperazines [cyclo-(L-Pro-L-Leu) (1), cyclo-(D-Pro-L-Leu) (2), and cyclo-(D-Pro-L-Tyr) (3)] purified from a Bacillus sp. N strain associated with entomopathogenic nematode Rhabditis (Oscheius) in combination with amphotericin B and clotrimazole was investigated using the macrodilution method. The minimum inhibitory concentration and minimum fungicidal concentration of the diketopiperazines was compared with that of the standard antibiotics. The synergistic anticandidal activities of diketopiperazines with amphotericin B or clotrimazole were assessed using the checkerboard and time-kill methods. The results of the present study showed that the combined effects of diketopiperazines with amphotericin B or clotrimazole predominantly recorded synergistic (<0.5). Time-kill study showed that the growth of the Candida was completely attenuated after 12-24 h of treatment with 50:50 ratios of diketopiperazines and antibiotics. These results suggest that diketopiperazines combined with antibiotics may be microbiologically beneficial and not antagonistic. These findings have potential implications in delaying the development of resistance as the anticandidal effect is achieved with lower concentrations of both drugs (diketopiperazines and antibiotics). The cytotoxicity of diketopiperazines was also tested against two normal human cell lines (L231 lung epithelial and FS normal fibroblast) and no cytotoxicity was recorded for diketopiperazines up to 200 μg/mL. The in vitro synergistic activity of diketopiperazines with antibiotics against Candida albicans is reported here for the first time.

  12. Reduced inhibition of Candida albicans adhesion by saliva from patients receiving oral cancer therapy.

    PubMed Central

    Umazume, M; Ueta, E; Osaki, T

    1995-01-01

    The effect of saliva on the adhesion of Candida albicans to epithelial cells was examined in vitro by using saliva from healthy controls and patients with oral squamous cell carcinoma. The adhesion of C. albicans to established epithelial tumor cells was reduced by 40% by salivary treatment of the C. albicans or epithelial cells. The inhibitory activity of saliva was almost completely abolished by anti-secretory immunoglobulin A antibody, concanavalin A, and mannose. Compared with saliva from healthy individuals, that from patients who had received chemoradiotherapy for oral carcinoma showed reduced suppression of C. albicans adhesion, which accompanied decreased salivary secretory immunoglobulin A and lactoferrin concentrations. A greater number of C. albicans cells adhered to buccal cells obtained from patients who had received chemoradiotherapy than to those from healthy individuals. Treatment of either epithelial cells or C. albicans with anticancer drugs induced an increase in adherence of epithelial cells and yeast cells. In contrast, concanavalin A- and mannose-pretreated C. albicans exhibited reduced adhesion to epithelial cells. No further decrease of C. albicans adhesion was observed when both epithelial cells and yeast phase C. albicans were treated with mannose. In conclusion, the inhibition of C. albicans adhesion by saliva depends largely on mannose residues on salivary glycoproteins and mannose is one of the binding ligands on both C. albicans and epithelial cells. In addition, anticancer therapy may induce oral C. albicans overgrowth by decreasing salivation and the concentrations of glycoproteins in saliva inhibiting C. albicans adhesion and by increasing the adhesive properties of both C. albicans and oral epithelial cells. PMID:7714204

  13. Quercetin inhibits rhinovirus replication in vitro and in vivo.

    PubMed

    Ganesan, Shyamala; Faris, Andrea N; Comstock, Adam T; Wang, Qiong; Nanua, Suparna; Hershenson, Marc B; Sajjan, Uma S

    2012-06-01

    Rhinovirus (RV), which is responsible for the majority of common colds, also causes exacerbations in patients with asthma and chronic obstructive pulmonary disease. So far, there are no drugs available for treatment of rhinovirus infection. We examined the effect of quercetin, a plant flavanol on RV infection in vitro and in vivo. Pretreatment of airway epithelial cells with quercetin decreased Akt phosphosphorylation, viral endocytosis and IL-8 responses. Addition of quercetin 6h after RV infection (after viral endocytosis) reduced viral load, IL-8 and IFN responses in airway epithelial cells. This was associated with decreased levels of negative and positive strand viral RNA, and RV capsid protein, abrogation of RV-induced eIF4GI cleavage and increased phosphorylation of eIF2α. In mice infected with RV, quercetin treatment decreased viral replication as well as expression of chemokines and cytokines. Quercetin treatment also attenuated RV-induced airway cholinergic hyperresponsiveness. Together, our results suggest that quercetin inhibits RV endocytosis and replication in airway epithelial cells at multiple stages of the RV life cycle. Quercetin also decreases expression of pro-inflammatory cytokines and improves lung function in RV-infected mice. Based on these observations, further studies examining the potential benefits of quercetin in the prevention and treatment of RV infection are warranted.

  14. Further evidence that naphthoquinone inhibits Toxoplasma gondii growth in vitro.

    PubMed

    da Silva, Luciana Lemos Rangel; Portes, Juliana de Araujo; de Araújo, Marlon Heggdorne; Silva, Jéssica Lays Sant'ana; Rennó, Magdalena Nascimento; Netto, Chaquip Daher; da Silva, Alcides José Monteiro; Costa, Paulo Roberto Ribeiro; De Souza, Wanderley; Seabra, Sergio Henrique; DaMatta, Renato Augusto

    2015-12-01

    Toxoplasmosis is a widely disseminated disease caused by Toxoplasma gondii, an intracellular protozoan parasite. Standard treatment causes many side effects, such as depletion of bone marrow cells, skin rashes and gastrointestinal implications. Therefore, it is necessary to find chemotherapeutic alternatives for the treatment of this disease. It was shown that a naphthoquinone derivative compound is active against T. gondii, RH strain, with an IC50 around 2.5 μM. Here, three different naphthoquinone derivative compounds with activity against leukemia cells and breast carcinoma cell were tested against T. gondii (RH strain) infected LLC-MK2 cell line. All the compounds were able to inhibit parasite growth in vitro, but one of them showed an IC50 activity below 1 μM after 48 h of treatment. The compounds showed low toxicity to the host cell. In addition, these compounds were able to induce tachyzoite-bradyzoite conversion confirmed by morphological changes, Dolichus biflorus lectin cyst wall labeling and characterization of amylopectin granules in the parasites by electron microscopy analysis using the Thierry technique. Furthermore, the compounds induced alterations on the ultrastructure of the parasite. Taken together, our results point to the naphthoquinone derivative (LQB 151) as a potential compound for the development of new drugs for the treatment of toxoplasmosis. PMID:26335616

  15. Thiol-dependent inhibition of RNA synthesis in vitro by acridines: structure-inhibition relationships.

    PubMed

    Gniazdowski, M; Szmigiero, L; Wilmańska, D

    1982-01-01

    In the presence of sulfhydryl compounds an anticancer drug, 1-nitro-9-aminoalkylacridine derivative, forms with DNA irreversible, probably covalent, complexes of decreased template properties. Five 9-substituted 1-nitro-9-aminoacridine derivatives of cytostatic activity show irreversible thiol-dependent inhibitory effects on the RNA synthesis in vitro system while equal inhibition is observed both in the presence and in the absence of dithiothreitol with biologically inactive analogues of nitrocrine. In the absence of sulfhydryl compounds the inhibition depends on the planarity of the acridine ring. Hence, both 1-nitro-9-aminoalkylacridine and tetrahydroacridine derivatives show low inhibitory effect. PMID:6174208

  16. Triazine Inhibits Toxoplasma gondii tachyzoites in vitro and in vivo.

    PubMed

    Mui, Ernest J; Jacobus, David; Milhous, Wilbur K; Schiehser, Guy; Hsu, Honghue; Roberts, Craig W; Kirisits, Michael J; McLeod, Rima

    2005-08-01

    The triazine WR99210 [4,6-diamino-1,2-dihydro-2,2-dimethyl-1-(2,4,5-trichlorophenoxypropyloxy)-1,3,5 triazine] inhibits Toxoplasma gondii in vitro at nanomolar levels (P < 0.05). The 50% inhibitory concentration (IC(50)) was approximately 50 nM. It is a potent inhibitor in vitro and is also effective in vivo. Administration of WR99210 parenterally (i.e., intraperitoneally) reduced the mean number of RH strain tachyzoites present in peritoneal fluid substantially 4 days after intraperitoneal infection of mice. There was a mean of approximately 35 million parasites in control mice as contrasted with approximately 2 million parasites in mice treated with 1.25 mg WR99210/kg of body weight in a representative experiment (P < 0.05). In addition the prodrug PS-15 N'-[3-(2,4, 5-trichlorophenoxy)propyloxy]-N9-(1-methylethyl) imidocarbonimidicdiamide is converted to 4,6-diamino-1,2-dihydro-2,2-dimethyl-1-(2,4,5-trichlorophenoxypropyloxy)-1,3,5 triazine in vivo when the prodrug is administered orally. PS-15 administered by gavage also reduced intraperitoneal RH strain T. gondii tachyzoite numbers. WR99210 has high efficacy and relatively low toxicity because of its substantial effect on T. gondii dihydrofolate reductase (DHFR) but not the mammalian host DHFR. Amino acid sequences of T. gondii, Plasmodium falciparum, and Homo sapiens DHFRs were compared. It is of interest that of the DHFR amino acids considered to be interacting with WR99210 in P. falciparum within interatomic distances within 3 to 5 A, four of eight were shared with T. gondii DHFR. H. sapiens also shared four amino acids thought to be interacting with WR99210. Efficacy of intraperitoneal administration of WR99210 and peroral administration of PS-15 demonstrate the potential usefulness of this class of compounds in treatment of toxoplasmosis administered either parenterally or perorally. The recent development program for this class of antimicrobials as antimalarials makes our proof of principle of improved

  17. Triazine Inhibits Toxoplasma gondii Tachyzoites In Vitro and In Vivo

    PubMed Central

    Mui, Ernest J.; Jacobus, David; Milhous, Wilbur K.; Schiehser, Guy; Hsu, Honghue; Roberts, Craig W.; Kirisits, Michael J.; McLeod, Rima

    2005-01-01

    The triazine WR99210 [4,6-diamino-1,2-dihydro-2,2-dimethyl-1-(2,4,5-trichlorophenoxypropyloxy)-1,3,5 triazine] inhibits Toxoplasma gondii in vitro at nanomolar levels (P < 0.05). The 50% inhibitory concentration (IC50) was approximately 50 nM. It is a potent inhibitor in vitro and is also effective in vivo. Administration of WR99210 parenterally (i.e., intraperitoneally) reduced the mean number of RH strain tachyzoites present in peritoneal fluid substantially 4 days after intraperitoneal infection of mice. There was a mean of approximately 35 million parasites in control mice as contrasted with approximately 2 million parasites in mice treated with 1.25 mg WR99210/kg of body weight in a representative experiment (P < 0.05). In addition the prodrug PS-15 N′-[3-(2,4, 5-trichlorophenoxy)propyloxy]-N9-(1-methylethyl) imidocarbonimidicdiamide is converted to 4,6-diamino-1,2-dihydro-2,2-dimethyl-1-(2,4,5-trichlorophenoxypropyloxy)-1,3,5 triazine in vivo when the prodrug is administered orally. PS-15 administered by gavage also reduced intraperitoneal RH strain T. gondii tachyzoite numbers. WR99210 has high efficacy and relatively low toxicity because of its substantial effect on T. gondii dihydrofolate reductase (DHFR) but not the mammalian host DHFR. Amino acid sequences of T. gondii, Plasmodium falciparum, and Homo sapiens DHFRs were compared. It is of interest that of the DHFR amino acids considered to be interacting with WR99210 in P. falciparum within interatomic distances within 3 to 5 Å, four of eight were shared with T. gondii DHFR. H. sapiens also shared four amino acids thought to be interacting with WR99210. Efficacy of intraperitoneal administration of WR99210 and peroral administration of PS-15 demonstrate the potential usefulness of this class of compounds in treatment of toxoplasmosis administered either parenterally or perorally. The recent development program for this class of antimicrobials as antimalarials makes our proof of principle of improved

  18. In vitro susceptibility of 137 Candida sp. isolates from HIV positive patients to several antifungal drugs.

    PubMed

    Magaldi, S; Mata, S; Hartung, C; Verde, G; Deibis, L; Roldán, Y; Marcano, C

    2001-01-01

    Oropharyngeal candidiasis caused by various species of Candida is one of the most common infections in HIV seropositive or AIDS patients. Drug resistance among these yeasts is an increasing problem. We studied the frequency of resistance profile to fluconazole, itraconazole, ketoconazole, amphotericin B and terbinafine of 137 isolates of Candida sp. From HIV positive or AIDS patients with oropharyngeal candidiasis at Instituto de Inmunología, U.C.V. and the Hospital "Jose Ignacio Baldó", Caracas Venezuela, using the well diffusion susceptibility test (Magaldi et al.). We found that nearly 10% of C. albicans isolates were primarily fluconazole resistant, 45% of C. albicans isolates from patients with previous treatment were resistant to fluconazole, of which 93% showed cross-resistance to itraconazole, and even about 30% of C. tropicalis (n = 13) were resistant to fluconazole and/or itraconazole. To this respect, several recent reports have been described antifungal cross-resistance among azoles. Therefore, we consider that C. tropicalis should be added to the growing list of yeast in which antifungal drug resistance is common. This report could be useful for therapeutic aspect in AIDS patients with oral candidiasis.

  19. Effect of in vivo administration of recombinant murine gamma interferon on in vitro lymphoproliferative responses following immunization with Candida albicans.

    PubMed

    Garner, R E; Kuruganti, U; al-Hussaini, L A; Czarniecki, C W; Domer, J E

    1992-05-01

    The effect of in vivo administration of recombinant murine gamma interferon (rMuIFN-gamma) on in vitro proliferation of lymphocytes to Candida antigens and lectins was examined in naive CBA/J mice and in similar mice colonized with Candida albicans by intragastric (i.g.) intubation and/or inoculated intradermally (i.d.) with the fungus. Lymph node lymphocyte and splenic lymphocyte (splenocyte) responses to soluble cytoplasmic substances derived from C. albicans varied with the route of inoculation of the fungus, the sex of the animal, and the presence or absence of rMuIFN-gamma treatment. In the absence of rMuIFN-gamma treatment, lymphoid cells from lymph nodes draining the site of the i.d. lesion responded well to soluble cytoplasmic substances. Colonization of the gut of female mice with C. albicans either had no effect or promoted better lymph node responses when such animals were also challenged i.d., whereas gut colonization of males followed by i.d. challenge appeared to have a suppressive influence on the level of proliferation in response to antigens in vitro. Antigen-specific splenocyte responses could be detected as well, and they were best in animals inoculated i.g.-i.d. or i.d. only. With the exception of lymph node lymphocytes from male mice, treatment of infected animals, regardless of the route of infection, with rMuIFN-gamma frequently resulted in lowered responses to antigens when comparable treatment groups were examined. With respect to mitogen stimulation, infection with C. albicans, especially i.g. or i.g.-i.d., resulted in a population of lymph node lymphocytes with lower-than-normal responses to concanavalin A but higher-than-normal responses to lipopolysaccharide (LPS). Splenocyte responses to mitogens were not altered as dramatically as the responses of lymph node lymphocytes, but splenocytes from female mice had a suppressed response regardless of the route of exposure to C. albicans, and those from mice which were maximally stimulated

  20. Effect of in vivo administration of recombinant murine gamma interferon on in vitro lymphoproliferative responses following immunization with Candida albicans.

    PubMed Central

    Garner, R E; Kuruganti, U; al-Hussaini, L A; Czarniecki, C W; Domer, J E

    1992-01-01

    The effect of in vivo administration of recombinant murine gamma interferon (rMuIFN-gamma) on in vitro proliferation of lymphocytes to Candida antigens and lectins was examined in naive CBA/J mice and in similar mice colonized with Candida albicans by intragastric (i.g.) intubation and/or inoculated intradermally (i.d.) with the fungus. Lymph node lymphocyte and splenic lymphocyte (splenocyte) responses to soluble cytoplasmic substances derived from C. albicans varied with the route of inoculation of the fungus, the sex of the animal, and the presence or absence of rMuIFN-gamma treatment. In the absence of rMuIFN-gamma treatment, lymphoid cells from lymph nodes draining the site of the i.d. lesion responded well to soluble cytoplasmic substances. Colonization of the gut of female mice with C. albicans either had no effect or promoted better lymph node responses when such animals were also challenged i.d., whereas gut colonization of males followed by i.d. challenge appeared to have a suppressive influence on the level of proliferation in response to antigens in vitro. Antigen-specific splenocyte responses could be detected as well, and they were best in animals inoculated i.g.-i.d. or i.d. only. With the exception of lymph node lymphocytes from male mice, treatment of infected animals, regardless of the route of infection, with rMuIFN-gamma frequently resulted in lowered responses to antigens when comparable treatment groups were examined. With respect to mitogen stimulation, infection with C. albicans, especially i.g. or i.g.-i.d., resulted in a population of lymph node lymphocytes with lower-than-normal responses to concanavalin A but higher-than-normal responses to lipopolysaccharide (LPS). Splenocyte responses to mitogens were not altered as dramatically as the responses of lymph node lymphocytes, but splenocytes from female mice had a suppressed response regardless of the route of exposure to C. albicans, and those from mice which were maximally stimulated

  1. Inhibition of alcoholic fermentation by substrate and ethanol. [Candida pseudotropicalis

    SciTech Connect

    Maulin, H.B.; Galzy, P.

    1980-11-01

    The effect of ethanol and sugars on rates of fermentation was studied. A strain of Candida pseudotropicalis was used. The specific rate of fermentation was determined by using the Warburg manometer. The effect of ethanol was formulated as an exponential function of ethanol concentration, but the empirical constant was different when glucose or lactose was used as a substrate. The effects of both ethanol and substrate were formulated. It was demonstrated that when lactose and glucose were present in the medium with a small amount of alcohol, a synergistic effect on the rate of fermentation appeared. This phenomenon considerably limits the rate of fermentation.

  2. In Vitro Activity of Miltefosine against Candida albicans under Planktonic and Biofilm Growth Conditions and In Vivo Efficacy in a Murine Model of Oral Candidiasis.

    PubMed

    Vila, Taissa Vieira Machado; Chaturvedi, Ashok K; Rozental, Sonia; Lopez-Ribot, Jose L

    2015-12-01

    The generation of a new antifungal against Candida albicans biofilms has become a major priority, since biofilm formation by this opportunistic pathogenic fungus is usually associated with an increased resistance to azole antifungal drugs and treatment failures. Miltefosine is an alkyl phospholipid with promising antifungal activity. Here, we report that, when tested under planktonic conditions, miltefosine displays potent in vitro activity against multiple fluconazole-susceptible and -resistant C. albicans clinical isolates, including isolates overexpressing efflux pumps and/or with well-characterized Erg11 mutations. Moreover, miltefosine inhibits C. albicans biofilm formation and displays activity against preformed biofilms. Serial passage experiments confirmed that miltefosine has a reduced potential to elicit resistance, and screening of a library of C. albicans transcription factor mutants provided additional insight into the activity of miltefosine against C. albicans growing under planktonic and biofilm conditions. Finally, we demonstrate the in vivo efficacy of topical treatment with miltefosine in the murine model of oropharyngeal candidiasis. Overall, our results confirm the potential of miltefosine as a promising antifungal drug candidate, in particular for the treatment of azole-resistant and biofilm-associated superficial candidiasis. PMID:26416861

  3. In Vitro Activity of Miltefosine against Candida albicans under Planktonic and Biofilm Growth Conditions and In Vivo Efficacy in a Murine Model of Oral Candidiasis

    PubMed Central

    Chaturvedi, Ashok K.; Rozental, Sonia

    2015-01-01

    The generation of a new antifungal against Candida albicans biofilms has become a major priority, since biofilm formation by this opportunistic pathogenic fungus is usually associated with an increased resistance to azole antifungal drugs and treatment failures. Miltefosine is an alkyl phospholipid with promising antifungal activity. Here, we report that, when tested under planktonic conditions, miltefosine displays potent in vitro activity against multiple fluconazole-susceptible and -resistant C. albicans clinical isolates, including isolates overexpressing efflux pumps and/or with well-characterized Erg11 mutations. Moreover, miltefosine inhibits C. albicans biofilm formation and displays activity against preformed biofilms. Serial passage experiments confirmed that miltefosine has a reduced potential to elicit resistance, and screening of a library of C. albicans transcription factor mutants provided additional insight into the activity of miltefosine against C. albicans growing under planktonic and biofilm conditions. Finally, we demonstrate the in vivo efficacy of topical treatment with miltefosine in the murine model of oropharyngeal candidiasis. Overall, our results confirm the potential of miltefosine as a promising antifungal drug candidate, in particular for the treatment of azole-resistant and biofilm-associated superficial candidiasis. PMID:26416861

  4. In Vitro Activity of Miltefosine against Candida albicans under Planktonic and Biofilm Growth Conditions and In Vivo Efficacy in a Murine Model of Oral Candidiasis.

    PubMed

    Vila, Taissa Vieira Machado; Chaturvedi, Ashok K; Rozental, Sonia; Lopez-Ribot, Jose L

    2015-12-01

    The generation of a new antifungal against Candida albicans biofilms has become a major priority, since biofilm formation by this opportunistic pathogenic fungus is usually associated with an increased resistance to azole antifungal drugs and treatment failures. Miltefosine is an alkyl phospholipid with promising antifungal activity. Here, we report that, when tested under planktonic conditions, miltefosine displays potent in vitro activity against multiple fluconazole-susceptible and -resistant C. albicans clinical isolates, including isolates overexpressing efflux pumps and/or with well-characterized Erg11 mutations. Moreover, miltefosine inhibits C. albicans biofilm formation and displays activity against preformed biofilms. Serial passage experiments confirmed that miltefosine has a reduced potential to elicit resistance, and screening of a library of C. albicans transcription factor mutants provided additional insight into the activity of miltefosine against C. albicans growing under planktonic and biofilm conditions. Finally, we demonstrate the in vivo efficacy of topical treatment with miltefosine in the murine model of oropharyngeal candidiasis. Overall, our results confirm the potential of miltefosine as a promising antifungal drug candidate, in particular for the treatment of azole-resistant and biofilm-associated superficial candidiasis.

  5. Candida tropicalis from veterinary and human sources shows similar in vitro hemolytic activity, antifungal biofilm susceptibility and pathogenesis against Caenorhabditis elegans.

    PubMed

    Brilhante, Raimunda Sâmia Nogueira; Oliveira, Jonathas Sales de; Evangelista, Antônio José de Jesus; Serpa, Rosana; Silva, Aline Lobão da; Aguiar, Felipe Rodrigues Magalhães de; Pereira, Vandbergue Santos; Castelo-Branco, Débora de Souza Collares Maia; Pereira-Neto, Waldemiro Aquino; Cordeiro, Rossana de Aguiar; Sidrim, José Júlio Costa; Rocha, Marcos Fábio Gadelha

    2016-08-30

    The aim of this study was to evaluate the in vitro hemolytic activity and biofilm antifungal susceptibility of veterinary and human Candida tropicalis strains, as well as their pathogenesis against Caenorhabditis elegans. Twenty veterinary isolates and 20 human clinical isolates of C. tropicalis were used. The strains were evaluated for their hemolytic activity and biofilm production. Biofilm susceptibility to itraconazole, fluconazole, voriconazole, amphotericin B and caspofungin was assessed using broth microdilution assay. The in vivo evaluation of strain pathogenicity was investigated using the nematode C. elegans. Hemolytic factor was observed in 95% of the strains and 97.5% of the isolates showed ability to form biofilm. Caspofungin and amphotericin B showed better results than azole antifungals against mature biofilms. Paradoxical effect on mature biofilm metabolic activity was observed at elevated concentrations of caspofungin (8-64μg/mL). Azole antifungals were not able to inhibit mature C. tropicalis biofilms, even at the higher tested concentrations. High mortality rates of C. elegans were observed when the worms were exposed to with C. tropicalis strains, reaching up to 96%, 96h after exposure of the worms to C. tropicalis strains. These results reinforce the high pathogenicity of C. tropicalis from veterinary and human sources and show the effectiveness of caspofungin and amphotericin B against mature biofilms of this species. PMID:27527785

  6. Acetylcholine Protects against Candida albicans Infection by Inhibiting Biofilm Formation and Promoting Hemocyte Function in a Galleria mellonella Infection Model.

    PubMed

    Rajendran, Ranjith; Borghi, Elisa; Falleni, Monica; Perdoni, Federica; Tosi, Delfina; Lappin, David F; O'Donnell, Lindsay; Greetham, Darren; Ramage, Gordon; Nile, Christopher

    2015-08-01

    Both neuronal acetylcholine and nonneuronal acetylcholine have been demonstrated to modulate inflammatory responses. Studies investigating the role of acetylcholine in the pathogenesis of bacterial infections have revealed contradictory findings with regard to disease outcome. At present, the role of acetylcholine in the pathogenesis of fungal infections is unknown. Therefore, the aim of this study was to determine whether acetylcholine plays a role in fungal biofilm formation and the pathogenesis of Candida albicans infection. The effect of acetylcholine on C. albicans biofilm formation and metabolism in vitro was assessed using a crystal violet assay and phenotypic microarray analysis. Its effect on the outcome of a C. albicans infection, fungal burden, and biofilm formation were investigated in vivo using a Galleria mellonella infection model. In addition, its effect on modulation of host immunity to C. albicans infection was also determined in vivo using hemocyte counts, cytospin analysis, larval histology, lysozyme assays, hemolytic assays, and real-time PCR. Acetylcholine was shown to have the ability to inhibit C. albicans biofilm formation in vitro and in vivo. In addition, acetylcholine protected G. mellonella larvae from C. albicans infection mortality. The in vivo protection occurred through acetylcholine enhancing the function of hemocytes while at the same time inhibiting C. albicans biofilm formation. Furthermore, acetylcholine also inhibited inflammation-induced damage to internal organs. This is the first demonstration of a role for acetylcholine in protection against fungal infections, in addition to being the first report that this molecule can inhibit C. albicans biofilm formation. Therefore, acetylcholine has the capacity to modulate complex host-fungal interactions and plays a role in dictating the pathogenesis of fungal infections.

  7. Acetylcholine Protects against Candida albicans Infection by Inhibiting Biofilm Formation and Promoting Hemocyte Function in a Galleria mellonella Infection Model

    PubMed Central

    Rajendran, Ranjith; Borghi, Elisa; Falleni, Monica; Perdoni, Federica; Tosi, Delfina; Lappin, David F.; O'Donnell, Lindsay; Greetham, Darren; Ramage, Gordon

    2015-01-01

    Both neuronal acetylcholine and nonneuronal acetylcholine have been demonstrated to modulate inflammatory responses. Studies investigating the role of acetylcholine in the pathogenesis of bacterial infections have revealed contradictory findings with regard to disease outcome. At present, the role of acetylcholine in the pathogenesis of fungal infections is unknown. Therefore, the aim of this study was to determine whether acetylcholine plays a role in fungal biofilm formation and the pathogenesis of Candida albicans infection. The effect of acetylcholine on C. albicans biofilm formation and metabolism in vitro was assessed using a crystal violet assay and phenotypic microarray analysis. Its effect on the outcome of a C. albicans infection, fungal burden, and biofilm formation were investigated in vivo using a Galleria mellonella infection model. In addition, its effect on modulation of host immunity to C. albicans infection was also determined in vivo using hemocyte counts, cytospin analysis, larval histology, lysozyme assays, hemolytic assays, and real-time PCR. Acetylcholine was shown to have the ability to inhibit C. albicans biofilm formation in vitro and in vivo. In addition, acetylcholine protected G. mellonella larvae from C. albicans infection mortality. The in vivo protection occurred through acetylcholine enhancing the function of hemocytes while at the same time inhibiting C. albicans biofilm formation. Furthermore, acetylcholine also inhibited inflammation-induced damage to internal organs. This is the first demonstration of a role for acetylcholine in protection against fungal infections, in addition to being the first report that this molecule can inhibit C. albicans biofilm formation. Therefore, acetylcholine has the capacity to modulate complex host-fungal interactions and plays a role in dictating the pathogenesis of fungal infections. PMID:26092919

  8. Demonstration of synergy with fluconazole and either ibuprofen, sodium salicylate, or propylparaben against Candida albicans in vitro.

    PubMed Central

    Scott, E M; Tariq, V N; McCrory, R M

    1995-01-01

    The combination of fluconazole with either ibuprofen, sodium salicylate, or propylparaben resulted in synergistic activity (fractional inhibitory index, < 0.5) against Candida albicans NCYC 620 in a microdilution checkerboard assay. Synergism between miconazole and ibuprofen was also demonstrated. In three or four clinical isolates of C. albicans from AIDS patients, the combination of fluconazole and ibuprofen was synergistic. Preparation of the inoculum and the growth conditions used were those recommended by the National Committee for Clinical Laboratory Standards for susceptibility testing. A visual estimation of total inhibition of growth and determination of an 80% reduction in the optical density at 492 nm compared with those for the control were taken as endpoints for the calculation of synergy, and a good correlation between both estimates was demonstrated. PMID:8592988

  9. Antifungal Effect of Lavender Essential Oil (Lavandula angustifolia) and Clotrimazole on Candida albicans: An In Vitro Study.

    PubMed

    Behmanesh, Fereshteh; Pasha, Hajar; Sefidgar, Ali Asghar; Taghizadeh, Mohsen; Moghadamnia, Ali Akbar; Adib Rad, Hajar; Shirkhani, Leyla

    2015-01-01

    Background. The treatment of candidiasis infections is an important problem in the health care system. This study aimed to investigate the in vitro effect of lavender essential oil and clotrimazole on isolated C. albicans from vaginal candidiasis. Materials and Methods. In this clinical trial, C. albicans isolated from the vaginal discharge samples was obtained. Results. The pairwise comparison showed that lavender and clotrimazole had a significant difference; this difference in the lavender group was lower than clotrimazole. But, after 48 hours, there was no difference seen between groups. There was a significant difference between clotrimazole and DMSO groups. Comparing the changes between groups based on the same dilution, at 24 h and 48 h in clotrimazole group, showed a significant difference two times in the fungal cell count that its average during 48 h was less than 24 h. A significant difference was observed between the two periods in lavender group, only at the dilutions of 1/20 and 1/80. The average fungal cell count after 48 h was also lower in lavender group. Conclusions. Given that the lavender has antifungal activity, this can be used as an antifungal agent. However, more clinical studies are necessary to validate its use in candida infection. PMID:26550521

  10. In vitro activity of essential oils extracted from condiments against fluconazole-resistant and -sensitive Candida glabrata.

    PubMed

    Soares, I H; Loreto, É S; Rossato, L; Mario, D N; Venturini, T P; Baldissera, F; Santurio, J M; Alves, S H

    2015-09-01

    In the present study, the antifungal activity of essential oils obtained from Origanum vulgare (oregano), Cinnamomum zeylanicum (cinnamon), Lippia graveolens (Mexican oregano), Thymus vulgaris (thyme), Salvia officinalis (sage), Rosmarinus officinalis (rosemary), Ocimum basilicum (basil) and Zingiber officinale (ginger) were assessed against Candida glabrata isolates. One group contained 30 fluconazole-susceptible C. glabrata isolates, and the second group contained fluconazole-resistant isolates derived from the first group after the in vitro induction of fluconazole-resistance, for a total of 60 tested isolates. The broth microdilution methodology was used. Concentrations of 50μg/mL, 100μg/mL, 200μg/mL, 400μg/mL, 800μg/mL, 1600μg/mL and 3200μg/mL of the essential oils were used, and the minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) were determined. Thyme, sage, rosemary, basil and ginger essential oils showed no antifungal activity at the tested concentrations. Antimicrobial activity less than or equal to 3200μg/mL was observed for oregano, Mexican oregano and cinnamon essential oils. Both the oregano and Mexican oregano essential oils showed high levels of antifungal activity against the fluconazole-susceptible C. glabrata group, whereas the cinnamon essential oil showed the best antifungal activity against the fluconazole-resistant C. glabrata isolates.

  11. In vitro activity of essential oils extracted from condiments against fluconazole-resistant and -sensitive Candida glabrata.

    PubMed

    Soares, I H; Loreto, É S; Rossato, L; Mario, D N; Venturini, T P; Baldissera, F; Santurio, J M; Alves, S H

    2015-09-01

    In the present study, the antifungal activity of essential oils obtained from Origanum vulgare (oregano), Cinnamomum zeylanicum (cinnamon), Lippia graveolens (Mexican oregano), Thymus vulgaris (thyme), Salvia officinalis (sage), Rosmarinus officinalis (rosemary), Ocimum basilicum (basil) and Zingiber officinale (ginger) were assessed against Candida glabrata isolates. One group contained 30 fluconazole-susceptible C. glabrata isolates, and the second group contained fluconazole-resistant isolates derived from the first group after the in vitro induction of fluconazole-resistance, for a total of 60 tested isolates. The broth microdilution methodology was used. Concentrations of 50μg/mL, 100μg/mL, 200μg/mL, 400μg/mL, 800μg/mL, 1600μg/mL and 3200μg/mL of the essential oils were used, and the minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) were determined. Thyme, sage, rosemary, basil and ginger essential oils showed no antifungal activity at the tested concentrations. Antimicrobial activity less than or equal to 3200μg/mL was observed for oregano, Mexican oregano and cinnamon essential oils. Both the oregano and Mexican oregano essential oils showed high levels of antifungal activity against the fluconazole-susceptible C. glabrata group, whereas the cinnamon essential oil showed the best antifungal activity against the fluconazole-resistant C. glabrata isolates. PMID:26281965

  12. Candida albicans is able to use M cells as a portal of entry across the intestinal barrier in vitro.

    PubMed

    Albac, Sandrine; Schmitz, Antonin; Lopez-Alayon, Carolina; d'Enfert, Christophe; Sautour, Marc; Ducreux, Amandine; Labruère-Chazal, Catherine; Laue, Michael; Holland, Gudrun; Bonnin, Alain; Dalle, Frederic

    2016-02-01

    Candida albicans is the most frequent yeast responsible for systemic infections in humans. These infections mainly originate from the gastrointestinal tract where C. albicans can invade the gut epithelial barrier to gain access to the bloodstream. Along the gut, pathogens can use Microfold (M) cells as a portal of entry to cross the epithelial barrier. M cells are specialized cells mainly located in the follicule-associated epithelium of Peyer patches. In this study, we used scanning electron and fluorescence microscopy, adhesion and invasion assays and fungal mutants to investigate the interactions of C. albicans with M cells obtained in an established in vitro model whereby enterocyte-like Caco-2 cells co-cultured with the Raji B cell line undergo a phenotypic switch to morphologically and functionally resembling M cells. Our data demonstrate that C. albicans co-localizes with and invades preferentially M cells, providing evidence that the fungus can use M cells as a portal of entry into the intestinal barrier. In addition to active penetration, F-actin dependent endocytosis contributes to internalization of the fungus into M cells through a mechanism involving hypha-associated invasins including Ssa1 and Als3. PMID:26242223

  13. In vitro activity of xanthorrhizol isolated from the rhizome of Javanese turmeric (Curcuma xanthorrhiza Roxb.) against Candida albicans biofilms.

    PubMed

    Rukayadi, Yaya; Hwang, Jae-Kwan

    2013-07-01

    The purpose of this study was to investigate the activity of xanthorrhizol isolated from Curcuma xanthorrhiza Roxb. on Candida albicans biofilms at adherent, intermediate, and mature phase of growth. C. albicans biofilms were formed in flat-bottom 96-well microtiter plates. The biofilms of C. albicans at different phases of development were exposed to xanthorrhizol at different concentrations (0.5 µg/mL-256 µg/mL) for 24 h. The metabolic activity of cells within the biofilms was quantified using the XTT reduction assay. Sessile minimum inhibitory concentrations (SMICs) were determined at 50% and 80% reduction in the biofilm OD₄₉₀ compared to the control wells. The SMIC₅₀ and SMIC₈₀ of xanthorrhizol against 18 C. albicans biofilms were 4--16 µg/mL and 8--32 µg/mL, respectively. The results demonstrated that the activity of xanthorrhizol in reducing C. albicans biofilms OD₄₉₀ was dependent on the concentration and the phase of growth of biofilm. Xanthorrhizol at concentration of 8 µg/mL completely reduced in biofilm referring to XTT-colorimetric readings at adherent phase, whereas 32 µg/mL of xanthorrhizol reduced 87.95% and 67.48 % of biofilm referring to XTT-colorimetric readings at intermediate and mature phases, respectively. Xanthorrhizol displayed potent activity against C. albicans biofilms in vitro and therefore might have potential therapeutic implication for biofilm-associated candidal infections.

  14. An In Vitro Model for Oral Mixed Biofilms of Candida albicans and Streptococcus gordonii in Synthetic Saliva

    PubMed Central

    Montelongo-Jauregui, Daniel; Srinivasan, Anand; Ramasubramanian, Anand K.; Lopez-Ribot, Jose L.

    2016-01-01

    As a member of the normal human oral microbiota, the fungus Candida albicans is often found in association with Streptococcus gordonii, a member of dental plaque forming bacteria. Evidence suggests that S. gordonii serves as a facilitator of C. albicans adherence to dental tissues, which represents a clinically relevant problem, particularly for immunocompromised individuals that could subsequently develop fungal infections. In this study we describe the development of a relatively simple and economical in vitro model that allows for the growth of mixed bacterial/fungal biofilms in 96-well microtiter plates. We have applied this method to test and compare the growth characteristics of single and dual species biofilms in traditional microbiological media versus a synthetic saliva medium (basal medium mucin, BMM) that more closely resembles physiological conditions within the oral cavity. Results indicated a synergistic effect for the formation of biofilms when both microorganisms were seeded together under all conditions tested. The structural and architectural features of the resulting biofilms were further characterized using scanning electron microscopy and confocal scanning laser microscopy. We also performed drug susceptibility assays against single and mixed species biofilms using commonly used antifungals and antibacterial antibiotics, both in monotherapy and in combination therapy, for a direct comparison of resistance against antimicrobial treatment. As expected, mixed species biofilms displayed higher levels of resistance to antimicrobial treatment at every dose tested in both traditional media and BMM synthetic saliva, as compared to single-species biofilms. PMID:27242712

  15. Antifungal Effect of Lavender Essential Oil (Lavandula angustifolia) and Clotrimazole on Candida albicans: An In Vitro Study

    PubMed Central

    Behmanesh, Fereshteh; Pasha, Hajar; Sefidgar, Ali Asghar; Taghizadeh, Mohsen; Moghadamnia, Ali Akbar; Adib Rad, Hajar; Shirkhani, Leyla

    2015-01-01

    Background. The treatment of candidiasis infections is an important problem in the health care system. This study aimed to investigate the in vitro effect of lavender essential oil and clotrimazole on isolated C. albicans from vaginal candidiasis. Materials and Methods. In this clinical trial, C. albicans isolated from the vaginal discharge samples was obtained. Results. The pairwise comparison showed that lavender and clotrimazole had a significant difference; this difference in the lavender group was lower than clotrimazole. But, after 48 hours, there was no difference seen between groups. There was a significant difference between clotrimazole and DMSO groups. Comparing the changes between groups based on the same dilution, at 24 h and 48 h in clotrimazole group, showed a significant difference two times in the fungal cell count that its average during 48 h was less than 24 h. A significant difference was observed between the two periods in lavender group, only at the dilutions of 1/20 and 1/80. The average fungal cell count after 48 h was also lower in lavender group. Conclusions. Given that the lavender has antifungal activity, this can be used as an antifungal agent. However, more clinical studies are necessary to validate its use in candida infection. PMID:26550521

  16. Chloride anion transporters inhibit growth of methicillin-resistant Staphylococcus aureus (MRSA) in vitro.

    PubMed

    Share, Andrew I; Patel, Khushali; Nativi, Cristina; Cho, Eun J; Francesconi, Oscar; Busschaert, Nathalie; Gale, Philip A; Roelens, Stefano; Sessler, Jonathan L

    2016-06-18

    A series of aminopyrrolic receptors were tested as anion transporters using POPC liposome model membranes. Many were found to be effective Cl(-) transporters and to inhibit clinical strains of Staphylococcus aureus growth in vitro. The best transporters proved effective against the methicillin-resistant Staphylococcus aureus (MRSA) strains, Mu50 and HP1173. Tris-thiourea tren-based chloride transporters were also shown to inhibit the growth of S. aureus in vitro.

  17. Copper, aluminum, iron and calcium inhibit human acetylcholinesterase in vitro.

    PubMed

    Pohanka, Miroslav

    2014-01-01

    Acetylcholinesterase (AChE) is an important part of cholinergic nerves where it participates in termination of neurotransmission. AChE can be inhibited by e.g. some Alzheimer disease drugs, nerve agents, and secondary metabolites. In this work, metal salts aluminum chloride, calcium chloride, cupric chloride, ferric chloride, potassium chloride, magnesium chloride and sodium chloride were tested for their ability to inhibit AChE. Standard Ellman assay based on human recombinant AChE was done and inhibition was measured using Dixon plot. No inhibition was proved for sodium, potassium and magnesium ions. However, aluminum, cupric, ferric and calcium ions were able to inhibit AChE via noncompetitive mechanism of inhibition. Though the inhibition is much weaker when compared to e.g. drugs with noncompetitive mechanism of action, biological relevance of the findings can be anticipated. PMID:24473150

  18. Evaluation of Antimicrobial and Antifungal Efficacy of Inter Appointment Intracanal Medicaments against Enterococcus and Candida albicans: An In Vitro Study

    PubMed Central

    Gupta, Shubh Prabhat; Bhati, Manisha; Jhajharia, Kapil; Patel, Hardik; Paliwal, Ashutosh; Franklin, Sundeep

    2015-01-01

    Background: Microorganisms and their by-products in pulpal and periapical diseases are to be considered as the primary etiological agents of the pulpal necrosis and apical periodontitis. Enterococcus faecalis, which is the most common organism isolated from failed root canals, is a Gram-positive facultative anaerobe. Yeasts can be detected in 7-18% of infected root canals. Materials and Methods: Designed to evaluate the antimicrobial and antifungal efficacy of chlorhexidine gutta-percha (CHX-GP), and calcium hydroxide GP points against E. faecalis and Candida albicans. The test materials used are divided into 3 groups; Group A - Control, Regular GP, Group B – CHX-GP, Group C - Calcium hydroxide GP. Detail Method is explained in the article. Results: There was a significant difference in the inhibition of E. faecalis, in different materials at each time interval period (P < 0.05). Similarly, the inhibition of C. albicans in the different materials at each time period was found to be statistically significant (P < 0.05). Calcium hydroxide GP did not exhibit any antimicrobial effect on any of tested microorganisms for all the time periods. Conclusion: CHX-GP had the maximum effect on the test microorganisms, with the maximum efficacy on E. faecalis followed by C. albicans. Regular GP also had a significant efficacy on test microorganisms, with the maximum efficacy on E. faecalis followed by C. albicans. Calcium hydroxide GP did not have any effect on any test microorganisms during the entire test periods. PMID:26124609

  19. In vitro Candida albicans biofilm induced proteinase activity and SAP8 expression correlates with in vivo denture stomatitis severity.

    PubMed

    Ramage, Gordon; Coco, Brent; Sherry, Leighann; Bagg, Jeremy; Lappin, David F

    2012-07-01

    Denture stomatitis is a common inflammatory disorder of the palatal mucosa amongst denture wearers. The pathological changes are induced by Candida albicans biofilm on the fitting surface of the upper denture, and different individuals experience different levels of disease. C. albicans is known to produce secreted aspartyl proteinases (SAPs) to aid adhesion, invasion and tissue destruction. We hypothesised that differential expression and activity of SAPs from denture stomatitis isolates results in different levels of disease amongst denture wearers. We selected C. albicans isolates from asymptomatic controls and three different severities of disease [Newton’s type (NT) 0, I, II and III]. We assessed biofilm formation and proteinase activity for each biofilm and investigated the transcriptional profile of SAPs 1, 2, 5, 6 and 8 from early (12 h) and mature (24 h) biofilms. There were no significant differences between isolates with respect to biofilm formation, whereas proteinase activity normalised to biofilm growth was significantly increased in the diseased groups (p < 0.0001). Proteinase activity correlated strongly with SAP expression (p < 0.0001). SAP8 expression was the greatest, followed by SAP5, 6, 2 and 1. The diseased groups showed the greatest levels of SAP expression, with significant differences also observed between the groups (p < 0.005). All SAPs except SAP5 were expressed in greater amounts in the mature biofilms compared to early biofilms. Overall, this study suggests that SAP activity in biofilms determined in vitro may help to explain differences in disease severity. SAP8 has been shown for the first time to play a prominent role in biofilms.

  20. Statin Drugs Markedly Inhibit Testosterone Production by Rat Leydig Cells In Vitro: Implications for Men

    EPA Science Inventory

    Statin drugs lower blood cholesterol by inhibiting hepatic 3-hydroxy-3-methylglutaryl-Coenzyme-A reductase. During drug development it was shown that statins inhibit production of cholesterol in the testis. We evaluated testosterone production in vitro, using highly purified rat ...

  1. Small molecules inhibit growth, viability and ergosterol biosynthesis in Candida albicans.

    PubMed

    Rajput, Sandeep B; Karuppayil, S Mohan

    2013-12-01

    The aim of this work was to evaluate the anti-Candida efficacy of twenty five molecules of plant origin. Based on their MICs, effective molecules were categorized into four categories. Susceptibility testing of test compounds was carried out by standard methodology (M27-A2) as per CLSI guidelines. Minimum Fungicidal Concentration (MFC) was determined as the lowest concentration of drug killing 99.9% cells. Effect on sterol profile was evaluated by sterol quantitation method. Among the screened molecules, cinnamaldehyde, piperidine, citral, furfuraldehyde and indole were potent inhibitors of growth and viability. Exposure of Candida cells to cinnamaldehyde, piperidine, citral, furfuraldehyde, indole, α- and β- pinene at MIC's, altered ergosterol profile. Our results indicate that the molecules altering sterol profile may exert their antifungal effect through inhibition of ergosterol biosynthesis and could be good candidates for fungal specific drug development. PMID:23449869

  2. In vitro Effects of Lemongrass Extract on Candida albicans Biofilms, Human Cells Viability, and Denture Surface.

    PubMed

    Madeira, Petrus L B; Carvalho, Letícia T; Paschoal, Marco A B; de Sousa, Eduardo M; Moffa, Eduardo B; da Silva, Marcos A Dos Santos; Tavarez, Rudys de Jesus Rodolfo; Gonçalves, Letícia M

    2016-01-01

    The purpose of this study was to investigate whether immersion of a denture surface in lemongrass extract (LGE) has effects on C. albicans biofilms, human cell viability and denture surface. Minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC) were performed for LGE against C. albicans. For biofilm analysis, discs were fabricated using a denture acrylic resin with surface roughness standardization. C. albicans biofilms were developed on saliva-coated discs, and the effects of LGE at MIC, 5XMIC, and 10XMIC were investigated during biofilm formation and after biofilm maturation. Biofilms were investigated for cell counting, metabolic activity, and microscopic analysis. The cytotoxicity of different concentrations of LGE to peripheral blood mononuclear cells (PBMC) was analyzed using MTT. The effects of LGE on acrylic resin were verified by measuring changes in roughness, color and flexural strength after 28 days of immersion. Data were analyzed by ANOVA, followed by a Tukey test at a 5% significance level. The minimal concentration of LGE required to inhibit C. albicans growth was 0.625 mg/mL, while MFC was 2.5 mg/mL. The presence of LGE during biofilm development resulted in a reduction of cell counting (p < 0.05), which made the MIC sufficient to reduce approximately 90% of cells (p < 0.0001). The exposure of LGE after biofilm maturation also had a significant antifungal effect at all concentrations (p < 0.05). When compared to the control group, the exposure of PBMC to LGE at MIC resulted in similar viability (p > 0.05). There were no verified differences in color perception, roughness, or flexural strength after immersion in LGE at MIC compared to the control (p > 0.05). It could be concluded that immersion of the denture surface in LGE was effective in reducing C. albicans biofilms with no deleterious effects on acrylic properties at MIC. MIC was also an effective and safe concentration for use. PMID:27446818

  3. In vitro Effects of Lemongrass Extract on Candida albicans Biofilms, Human Cells Viability, and Denture Surface

    PubMed Central

    Madeira, Petrus L. B.; Carvalho, Letícia T.; Paschoal, Marco A. B.; de Sousa, Eduardo M.; Moffa, Eduardo B.; da Silva, Marcos A. dos Santos; Tavarez, Rudys de Jesus Rodolfo; Gonçalves, Letícia M.

    2016-01-01

    The purpose of this study was to investigate whether immersion of a denture surface in lemongrass extract (LGE) has effects on C. albicans biofilms, human cell viability and denture surface. Minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC) were performed for LGE against C. albicans. For biofilm analysis, discs were fabricated using a denture acrylic resin with surface roughness standardization. C. albicans biofilms were developed on saliva-coated discs, and the effects of LGE at MIC, 5XMIC, and 10XMIC were investigated during biofilm formation and after biofilm maturation. Biofilms were investigated for cell counting, metabolic activity, and microscopic analysis. The cytotoxicity of different concentrations of LGE to peripheral blood mononuclear cells (PBMC) was analyzed using MTT. The effects of LGE on acrylic resin were verified by measuring changes in roughness, color and flexural strength after 28 days of immersion. Data were analyzed by ANOVA, followed by a Tukey test at a 5% significance level. The minimal concentration of LGE required to inhibit C. albicans growth was 0.625 mg/mL, while MFC was 2.5 mg/mL. The presence of LGE during biofilm development resulted in a reduction of cell counting (p < 0.05), which made the MIC sufficient to reduce approximately 90% of cells (p < 0.0001). The exposure of LGE after biofilm maturation also had a significant antifungal effect at all concentrations (p < 0.05). When compared to the control group, the exposure of PBMC to LGE at MIC resulted in similar viability (p > 0.05). There were no verified differences in color perception, roughness, or flexural strength after immersion in LGE at MIC compared to the control (p > 0.05). It could be concluded that immersion of the denture surface in LGE was effective in reducing C. albicans biofilms with no deleterious effects on acrylic properties at MIC. MIC was also an effective and safe concentration for use. PMID:27446818

  4. Nesfatin-1 inhibits ovarian epithelial carcinoma cell proliferation in vitro

    SciTech Connect

    Xu, Yang; Pang, Xiaoyan; Dong, Mei; Wen, Fang Zhang, Yi

    2013-11-01

    Highlights: •Nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest. •Nesfatin-1 enhances HO-8910 cell apoptosis. •Nesfatin-1 inhibits HO-8910 cell proliferation via mTOR and RhoA/ROCK signaling pathway. •The first report of nesfatin-1-mediated proliferation in ovarian epithelial carcinoma. -- Abstract: Nesfatin-1, an 82-amino-acid peptide derived from a 396-amino-acid precursor protein nucleobindin 2 (NUCB2), was originally identified in hypothalamic nuclei involved in the regulation of food intake. It was recently reported that nesfatin-1 is a novel depot specific adipokine preferentially produced by subcutaneous tissue, with obesity- and food deprivation-regulated expression. Although a relation between ovarian cancer mortality and obesity has been previously established, a role of nesfatin-1 in ovarian epithelial carcinoma remains unknown. The aim of the present study is to examine the effect of nesfatin-1 on ovary carcinoma cells proliferation. We found that nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest, this inhibition could be abolished by nesfatin-1 neutralizing antibody. Nesfatin-1 enhances HO-8910 cell apoptosis, activation of mammalian target of rapamycin (mTOR) and RhoA/ROCK signaling pathway block the effects of nesfatin-1-induced apoptosis, therefore reverses the inhibition of HO-8910 cell proliferation by nesfatin-1. In conclusion, the present study demonstrated that nesfatin-1 can inhibit the proliferation in human ovarian epithelial carcinoma cell line HO-8910 cells through inducing apoptosis via mTOR and RhoA/ROCK signaling pathway. This study provides a novel regulatory signaling pathway of nesfatin-1-regulated ovarian epithelial carcinoma growth and may contribute to ovarian cancer prevention and therapy, especially in obese patients.

  5. Inhibition of Plasmodium falciparum dihydropteroate synthetase and growth in vitro by sulfa drugs.

    PubMed Central

    Zhang, Y; Meshnick, S R

    1991-01-01

    The Michaelis-Menten inhibitory constants (Kis) and the concentrations required for 50% inhibition of the Plasmodium falciparum dihydropteroate synthetase were determined for six sulfa drugs. These drugs inhibited the in vitro growth of P. falciparum (50% lethal concentration) at concentrations of 30 to 500 nM; these concentrations were 100 to 1,000 times lower than the concentrations required for 50% inhibition and Kis (6 to 500 microM). The uptake of p-aminobenzoic acid was not inhibited by the sulfa drugs. However, infected erythrocytes took up more labeled sulfamethoxazole than did uninfected erythrocytes. Thus, the concentration of sulfa drugs by malaria parasites may explain how sulfa drugs inhibit in vitro growth of parasites through the inhibition of dihydropteroate synthetase. PMID:2024960

  6. The FGF-2-Derived Peptide FREG Inhibits Melanoma Growth In Vitro and In Vivo

    PubMed Central

    Aguzzi, Maria S; Faraone, Debora; D'Arcangelo, Daniela; De Marchis, Francesco; Toietta, Gabriele; Ribatti, Domenico; Parazzoli, Alberto; Colombo, Paolo; Capogrossi, Maurizio C; Facchiano, Antonio

    2011-01-01

    Previous data report that fibroblast growth factor-2 (FGF-2)-derived peptide FREG potently inhibits FGF-2-dependent angiogenesis in vitro and in vivo. Here, we show that FREG inhibits up to 70% in vitro growth and invasion/migration of smooth muscle and melanoma cells. Such inhibition is mediated by platelet-derived growth factor-receptor-α (PDGF-Rα); in fact, proliferation and migration were restored upon PDGF-Rα neutralization. Further experiments demonstrated that FREG interacts with PDGF-Rα both in vitro and in vivo and stimulates its phosphorylation. We have previously shown that overexpressing PDGF-Rα strongly inhibits melanoma growth in vivo; we, therefore, hypothesized that PDGF-Rα agonists may represent a novel tool to inhibit melanoma growth in vivo. To support this hypothesis, FREG was inoculated intravenously (i.v.) in a mouse melanoma model and markedly inhibited pulmonary metastases formation. Immunohistochemical analyses showed less proliferation, less angiogenesis, and more apoptosis in metastasized lungs upon FREG treatment, as compared to untreated controls. Finally, in preliminary acute toxicity studies, FREG showed no toxicity signs in healthy animals, and neither microscopic nor macroscopic toxicity at the liver, kidney, and lungs level. Altogether, these data indicate that FREG systemic treatment strongly inhibits melanoma metastases development and indicate for the first time that agonists of PDGF-Rα may control melanoma both in vitro and in vivo. PMID:20924364

  7. The FGF-2-derived peptide FREG inhibits melanoma growth in vitro and in vivo.

    PubMed

    Aguzzi, Maria S; Faraone, Debora; D'Arcangelo, Daniela; De Marchis, Francesco; Toietta, Gabriele; Ribatti, Domenico; Parazzoli, Alberto; Colombo, Paolo; Capogrossi, Maurizio C; Facchiano, Antonio

    2011-02-01

    Previous data report that fibroblast growth factor-2 (FGF-2)-derived peptide FREG potently inhibits FGF-2-dependent angiogenesis in vitro and in vivo. Here, we show that FREG inhibits up to 70% in vitro growth and invasion/migration of smooth muscle and melanoma cells. Such inhibition is mediated by platelet-derived growth factor-receptor-α (PDGF-Rα); in fact, proliferation and migration were restored upon PDGF-Rα neutralization. Further experiments demonstrated that FREG interacts with PDGF-Rα both in vitro and in vivo and stimulates its phosphorylation. We have previously shown that overexpressing PDGF-Rα strongly inhibits melanoma growth in vivo; we, therefore, hypothesized that PDGF-Rα agonists may represent a novel tool to inhibit melanoma growth in vivo. To support this hypothesis, FREG was inoculated intravenously (i.v.) in a mouse melanoma model and markedly inhibited pulmonary metastases formation. Immunohistochemical analyses showed less proliferation, less angiogenesis, and more apoptosis in metastasized lungs upon FREG treatment, as compared to untreated controls. Finally, in preliminary acute toxicity studies, FREG showed no toxicity signs in healthy animals, and neither microscopic nor macroscopic toxicity at the liver, kidney, and lungs level. Altogether, these data indicate that FREG systemic treatment strongly inhibits melanoma metastases development and indicate for the first time that agonists of PDGF-Rα may control melanoma both in vitro and in vivo. PMID:20924364

  8. Mechanisms involved in the inhibition of glycolysis by cyanide and antimycin A in Candida albicans and its reversal by hydrogen peroxide. A common feature in Candida species.

    PubMed

    Peña, Antonio; Sánchez, Norma Silvia; González-López, Omar; Calahorra, Martha

    2015-12-01

    In Candida albicans, cyanide and antimycin A inhibited K(+) transport, not only with ethanol-O2 as the substrate, but also with glucose. The reason for this was that they inhibited not only respiration, but also fermentation, decreasing ATP production. Measurements of oxygen levels in cell suspensions allowed identification of the electron pathways involved. NADH fluorescence levels increased in the presence of the inhibitors, indirectly indicating lower levels of NAD(+) and so pointing to glyceraldehyde-3-phosphate dehydrogenase as the limiting step responsible for the inhibition of glycolysis, which was confirmed by the levels of glycolytic intermediaries. The cyanide effect could be reversed by hydrogen peroxide, mainly due to an activity by which H2O2 can be reduced by electrons flowing from NADH through a pathway that can be inhibited by antimycin A, and appears to be a cytochrome c peroxidase. Therefore, the inhibition of glycolysis by the respiratory inhibitors seems to be due to the decreased availability of NAD(+), resulting in a decreased activity of glyceraldehyde-3-phosphate dehydrogenase. Compartmentalization of pyridine nucleotides in favor of the mitochondria can contribute to explaining the low fermentation capacity of C. albicans. Similar results were obtained with three C. albicans strains, Candida dubliniensis and, to a lower degree, Candida parapsilosis.

  9. Inhibition of catalase activity in vitro by diesel exhaust particles

    SciTech Connect

    Mori, Yoki; Murakami, Sumika; Sagae, Toshiyuki

    1996-02-09

    The effect of diesel exhaust particles (DEP) on the activity of catalase, an intracellular anti-oxidant, was investigated because H{sub 2}O{sub 2} is a cytotoxic oxidant, and catalase released from alveolar cells is an important antioxidant in the epithelial lining fluid in the lung. DEP inhibited the activity of bovine liver catalase dose-dependently, to 25-30% of its original value. The inhibition of catalase by DEP was observed only in the presence of anions such as Cl{sup {minus}}, Br{sup {minus}}, or thiocyanate. Other anions, such as CH{sub 3}COO{sup {minus}} or SO{sub 4}{sup {minus}}, and cations such as K{sup +}, Na{sup +}, Mg{sup 2+}, or Fe{sup 2+}, did not affect the activity of catalase, even in the presence of DEP extract. Catalase from guinea pig alveolar cells and catalase from red blood cells were also inhibited by DEP extracts, as was catalase from bovine liver. These results suggest that DEP taken up in the lung and located on alveolar spaces might cause cell injury by inhibiting the activity of catalase in epithelial lining fluid, enhancing the toxicity of H{sub 2}O{sub 2} generated from cells in addition to that of O{sub 2}{sup {minus}} generated by the chemical reaction of DEP with oxygen. 10 refs., 6 figs.

  10. Inhibition of human fibroblast growth in vitro by a snake oil.

    PubMed

    Datubo-Brown, D D; Blight, A

    1990-03-01

    The inhibitory effects of boa constrictor fat (BCF) oil on the growth kinetics of keloid and normal dermal fibroblasts were tested in fibroblasts cultures. BCF significantly (p less than 0.0001) inhibited the in vitro growth of both keloid and normal dermal fibroblasts. Although the active ingredient(s) in this snake oil is not yet determined, it is postulated that fatty acids which are the main constituents of the oil may in part account for this observed in vitro effect.

  11. In Vitro Assessment of Dual Drug Combinations To Inhibit Growth of Neisseria gonorrhoeae

    PubMed Central

    Pettus, Kevin; Sharpe, Samera

    2015-01-01

    The development of resistance to first-line antimicrobial therapies has led to recommendations for combination therapies for the treatment of gonorrhea infection. Recent studies have shown the success of combination therapies in treating patients, but few have reported on the in vitro activities of these drug combinations. An in vitro assessment of azithromycin in combination with gentamicin demonstrated inhibition of growth and suggests that clinical trials may be warranted to assess the utility of this combination in treating gonorrhea infections. PMID:25624328

  12. Cadmium inhibits brain calmodulin: In vitro and in vivo studies

    SciTech Connect

    Vig, P.J.S. ); Bhatia, M.; Gill, K.D.; Nath, R. )

    1989-10-01

    The toxic effects of divalent cations especially Cd{sup 2+}, are related to their chemical and physical characteristics viz. ion polarizability, electronic structure and the hard and soft characteristics. Cadmium ion, a hazardous environmental pollutant in areas in which cadmium nickel batteries are manufactured, accumulates in tissues such as kidney, liver, brain or bone with a very slow turnover and can cause pathophysiological disorders. Calmodulin (CaM), a Ca{sup 2+} binding protein is found in many if not most eukaryotic cells. The protein mediates the control of a large number of enzymes by Ca{sup 2+} fluxes and alterations in these fluxes have been postulated to be involved in steps leading to irreversible cell damage. The present study demonstrates an interaction of Cd{sup 2+} with CaM resulting in an impairment of CaM-dependent phosphodiesterase (PDE) stimulation in vitro. Biological activity of CaM is also affected in brain (cerebral cortex) of Cd-exposed rats in vivo.

  13. Downregulation of amyloid precursor protein inhibits neurite outgrowth in vitro

    PubMed Central

    1995-01-01

    The amyloid precursor protein (APP) is a transmembrane protein expressed in several cell types. In the nervous system, APP is expressed by glial and neuronal cells, and several lines of evidence suggest that it plays a role in normal and pathological phenomena. To address the question of the actual function of APP in normal developing neurons, we undertook a study aimed at blocking APP expression using antisense oligonucleotides. Oligonucleotide internalization was achieved by linking them to a vector peptide that translocates through biological membranes. This original technique, which is very efficient and gives direct access to the cell cytosol and nucleus, allowed us to work with extracellular oligonucleotide concentrations between 40 and 200 nM. Internalization of antisense oligonucleotides overlapping the origin of translation resulted in a marked but transient decrease in APP neosynthesis that was not observed with the vector peptide alone, or with sense oligonucleotides. Although transient, the decrease in APP neosynthesis was sufficient to provoke a distinct decrease in axon and dendrite outgrowth by embryonic cortical neurons developing in vitro. The latter decrease was not accompanied by changes in the spreading of the cell bodies. A single exposure to coupled antisense oligonucleotides at the onset of the culture was sufficient to produce significant morphological effects 6, 18, and 24 h later, but by 42 h, there were no remaining significant morphologic changes. This report thus demonstrates that amyloid precursor protein plays an important function in the morphological differentiation of cortical neurons in primary culture. PMID:7876315

  14. Inhibition of leucocytic lysosomal enzymes by glycosaminoglycans in vitro.

    PubMed Central

    Avila, J L; Convit, J

    1975-01-01

    1. A lysosomal fraction was separated by density-gradient centrifugation from a highly purified human polymorphonuclear leucocyte suspension. 2. Some 23 different lysosomal enzymes were assayed for activity in the presence of various concentrations of glycosaminoglycans. 3. The 21 acid hydrolases assayed were strongly inhibited to different degrees by low (0-12 mmol/l) concentrations of glycosaminoglycans in a pH-dependent manner. Thus inhibitions were stronger below pH4.5, with activity returning to control values at about pH5.0. 4. On a molar basis, the inhibitory activity for the several glycosaminoglycans studied was: heparin greater than chondroitin sulphate greater than hyaluronic acid. 5. Once the glycosaminoglycan-acid hydrolase complex was formed, it was partially dissociated by slight elevations in the pH of the incubation medium, by increasing the ionic strength of the incubation medium, or by adding several cationic proteins (e.g. histone, protamine). 6. As leucocytic lysosomes contain large amounts of chondroitin sulphate, and have a strongly acid intragranular pH, we suggest that glycosaminoglycans may modify lysosomal function through the formation of complexes with lysosomal enzymes, by inhibiting the digestive activity of the acid hydrolases when the intralysosomal pH is below their pI. PMID:2162

  15. Gellan sulfate inhibits Plasmodium falciparum growth and invasion of red blood cells in vitro

    PubMed Central

    Recuenco, Frances Cagayat; Kobayashi, Kyousuke; Ishiwa, Akiko; Enomoto-Rogers, Yukiko; Fundador, Noreen Grace V.; Sugi, Tatsuki; Takemae, Hitoshi; Iwanaga, Tatsuya; Murakoshi, Fumi; Gong, Haiyan; Inomata, Atsuko; Horimoto, Taisuke; Iwata, Tadahisa; Kato, Kentaro

    2014-01-01

    Here, we assessed the sulfated derivative of the microbial polysaccharide gellan gum and derivatives of λ and κ-carrageenans for their ability to inhibit Plasmodium falciparum 3D7 and Dd2 growth and invasion of red blood cells in vitro. Growth inhibition was assessed by means of flow cytometry after a 96-h exposure to the inhibitors and invasion inhibition was assessed by counting ring parasites after a 20-h exposure to them. Gellan sulfate strongly inhibited invasion and modestly inhibited growth for both P. falciparum 3D7 and Dd2; both inhibitory effects exceeded those achieved with native gellan gum. The hydrolyzed λ-carrageenan and oversulfated κ-carrageenan were less inhibitory than their native forms. In vitro cytotoxicity and anticoagulation assays performed to determine the suitability of the modified polysaccharides for in vivo studies showed that our synthesized gellan sulfate had low cytotoxicity and anticoagulant activity. PMID:24740150

  16. Caffeine inhibits hepatitis C virus replication in vitro.

    PubMed

    Batista, Mariana N; Carneiro, Bruno M; Braga, Ana Cláudia S; Rahal, Paula

    2015-02-01

    Hepatitis C is considered the major cause of cirrhosis and hepatocellular carcinoma. Conventional treatment is not effective against some hepatitis C virus (HCV) genotypes; therefore, new treatments are needed. Coffee and, more recently, caffeine, have been found to have a beneficial effect in several disorders of the liver, including those manifesting abnormal liver biochemistry, cirrhosis and hepatocellular carcinoma. Caffeine acts directly by delaying fibrosis, thereby improving the function of liver cellular pathways and interfering with pathways used by the HCV replication cycle. In the current study, the direct relationship between caffeine and viral replication was evaluated. The Huh-7.5 cell line was used for transient infections with FL-J6/JFH-5'C19Rluc2AUbi and to establish a cell line stably expressing SGR-Feo JFH-1. Caffeine efficiently inhibited HCV replication in a dose-dependent manner at non-cytotoxic concentrations and demonstrated an IC50 value of 0.7263 mM after 48 h of incubation. These data demonstrate that caffeine may be an important new agent for anti-HCV therapies due to its efficient inhibition of HCV replication at non-toxic concentrations.

  17. In vitro activities of terbinafine in combination with fluconazole and itraconazole against isolates of Candida albicans with reduced susceptibility to azoles.

    PubMed Central

    Barchiesi, F; Falconi Di Francesco, L; Scalise, G

    1997-01-01

    A checkerboard microdilution method was applied to study the in vitro interaction of terbinafine with either fluconazole and itraconazole against 30 strains of Candida albicans. Synergy was observed in 40% of the terbinafine-fluconazole interactions and in 43% of the terbinafine-itraconazole interactions, while antagonism was not observed. Even when only additivity was achieved, the combinations still showed beneficial effects since at least twofold reductions in the MICs of both drugs were found in 100% of the terbinafine-fluconazole interactions and in 76% of the terbinafine-itraconazole interactions. PMID:9257768

  18. An Optimized Lock Solution Containing Micafungin, Ethanol and Doxycycline Inhibits Candida albicans and Mixed C. albicans – Staphyloccoccus aureus Biofilms

    PubMed Central

    Lown, Livia; Peters, Brian M.; Walraven, Carla J.; Noverr, Mairi C.; Lee, Samuel A.

    2016-01-01

    Candida albicans is a major cause of catheter-related bloodstream infections and is associated with high morbidity and mortality. Due to the propensity of C. albicans to form drug-resistant biofilms, the current standard of care includes catheter removal; however, reinsertion may be technically challenging or risky. Prolonged exposure of an antifungal lock solution within the catheter in conjunction with systemic therapy has been experimentally attempted for catheter salvage. Previously, we demonstrated excellent in vitro activity of micafungin, ethanol, and high-dose doxycycline as single agents for prevention and treatment of C. albicans biofilms. Thus, we sought to investigate optimal combinations of micafungin, ethanol, and/or doxycycline as a lock solution. We performed two- and three-drug checkerboard assays to determine the in vitro activity of pairwise or three agents in combination for prevention or treatment of C. albicans biofilms. Optimal lock solutions were tested for activity against C. albicans clinical isolates, reference strains and polymicrobial C. albicans-S. aureus biofilms. A solution containing 20% (v/v) ethanol, 0.01565 μg/mL micafungin, and 800 μg/mL doxycycline demonstrated a reduction of 98% metabolic activity and no fungal regrowth when used to prevent fungal biofilm formation; however there was no advantage over 20% ethanol alone. This solution was also successful in inhibiting the regrowth of C. albicans from mature polymicrobial biofilms, although it was not fully bactericidal. Solutions containing 5% ethanol with low concentrations of micafungin and doxycycline demonstrated synergistic activity when used to prevent monomicrobial C. albicans biofilm formation. A combined solution of micafungin, ethanol and doxycycline is highly effective for the prevention of C. albicans biofilm formation but did not demonstrate an advantage over 20% ethanol alone in these studies. PMID:27428310

  19. An Optimized Lock Solution Containing Micafungin, Ethanol and Doxycycline Inhibits Candida albicans and Mixed C. albicans - Staphyloccoccus aureus Biofilms.

    PubMed

    Lown, Livia; Peters, Brian M; Walraven, Carla J; Noverr, Mairi C; Lee, Samuel A

    2016-01-01

    Candida albicans is a major cause of catheter-related bloodstream infections and is associated with high morbidity and mortality. Due to the propensity of C. albicans to form drug-resistant biofilms, the current standard of care includes catheter removal; however, reinsertion may be technically challenging or risky. Prolonged exposure of an antifungal lock solution within the catheter in conjunction with systemic therapy has been experimentally attempted for catheter salvage. Previously, we demonstrated excellent in vitro activity of micafungin, ethanol, and high-dose doxycycline as single agents for prevention and treatment of C. albicans biofilms. Thus, we sought to investigate optimal combinations of micafungin, ethanol, and/or doxycycline as a lock solution. We performed two- and three-drug checkerboard assays to determine the in vitro activity of pairwise or three agents in combination for prevention or treatment of C. albicans biofilms. Optimal lock solutions were tested for activity against C. albicans clinical isolates, reference strains and polymicrobial C. albicans-S. aureus biofilms. A solution containing 20% (v/v) ethanol, 0.01565 μg/mL micafungin, and 800 μg/mL doxycycline demonstrated a reduction of 98% metabolic activity and no fungal regrowth when used to prevent fungal biofilm formation; however there was no advantage over 20% ethanol alone. This solution was also successful in inhibiting the regrowth of C. albicans from mature polymicrobial biofilms, although it was not fully bactericidal. Solutions containing 5% ethanol with low concentrations of micafungin and doxycycline demonstrated synergistic activity when used to prevent monomicrobial C. albicans biofilm formation. A combined solution of micafungin, ethanol and doxycycline is highly effective for the prevention of C. albicans biofilm formation but did not demonstrate an advantage over 20% ethanol alone in these studies. PMID:27428310

  20. Strontium Promotes Cementoblasts Differentiation through Inhibiting Sclerostin Expression In Vitro

    PubMed Central

    Bao, Xingfu; Liu, Xianjun; Zhang, Yi; Cui, Yue; Yao, Jindan

    2014-01-01

    Cementogenesis, performed by cementoblasts, is important for the repair of root resorption caused by orthodontic treatment. Based on recent studies, strontium has been applied for osteoporosis treatment due to its positive effect on osteoblasts. Although promising, the effect of strontium on cementoblasts is still unclear. So the aim of this research was to clarify and investigate the effect of strontium on cementogenesis via employing cementoblasts as model. A series of experiments including MTT, alkaline phosphatase activity, gene analysis, alizarin red staining, and western blot were carried out to evaluate the proliferation and differentiation of cementoblasts. In addition, expression of sclerostin was checked to analyze the possible mechanism. Our results show that strontium inhibits the proliferation of cementoblasts with a dose dependent manner; however, it can promote the differentiation of cementoblasts via downregulating sclerostin expression. Taking together, strontium may facilitate cementogenesis and benefit the treatment of root resorption at a low dose. PMID:25003114

  1. Strontium promotes cementoblasts differentiation through inhibiting sclerostin expression in vitro.

    PubMed

    Bao, Xingfu; Liu, Xianjun; Zhang, Yi; Cui, Yue; Yao, Jindan; Hu, Min

    2014-01-01

    Cementogenesis, performed by cementoblasts, is important for the repair of root resorption caused by orthodontic treatment. Based on recent studies, strontium has been applied for osteoporosis treatment due to its positive effect on osteoblasts. Although promising, the effect of strontium on cementoblasts is still unclear. So the aim of this research was to clarify and investigate the effect of strontium on cementogenesis via employing cementoblasts as model. A series of experiments including MTT, alkaline phosphatase activity, gene analysis, alizarin red staining, and western blot were carried out to evaluate the proliferation and differentiation of cementoblasts. In addition, expression of sclerostin was checked to analyze the possible mechanism. Our results show that strontium inhibits the proliferation of cementoblasts with a dose dependent manner; however, it can promote the differentiation of cementoblasts via downregulating sclerostin expression. Taking together, strontium may facilitate cementogenesis and benefit the treatment of root resorption at a low dose.

  2. Strontium promotes cementoblasts differentiation through inhibiting sclerostin expression in vitro.

    PubMed

    Bao, Xingfu; Liu, Xianjun; Zhang, Yi; Cui, Yue; Yao, Jindan; Hu, Min

    2014-01-01

    Cementogenesis, performed by cementoblasts, is important for the repair of root resorption caused by orthodontic treatment. Based on recent studies, strontium has been applied for osteoporosis treatment due to its positive effect on osteoblasts. Although promising, the effect of strontium on cementoblasts is still unclear. So the aim of this research was to clarify and investigate the effect of strontium on cementogenesis via employing cementoblasts as model. A series of experiments including MTT, alkaline phosphatase activity, gene analysis, alizarin red staining, and western blot were carried out to evaluate the proliferation and differentiation of cementoblasts. In addition, expression of sclerostin was checked to analyze the possible mechanism. Our results show that strontium inhibits the proliferation of cementoblasts with a dose dependent manner; however, it can promote the differentiation of cementoblasts via downregulating sclerostin expression. Taking together, strontium may facilitate cementogenesis and benefit the treatment of root resorption at a low dose. PMID:25003114

  3. In vitro inhibition of lysine decarboxylase activity by organophosphate esters.

    PubMed

    Wang, Sufang; Wan, Bin; Zhang, Lianying; Yang, Yu; Guo, Liang-Hong

    2014-12-01

    Organophosphate esters (OPEs), a major group of organophosphorus flame retardants, are regarded as emerging environmental contaminants of health concern. Amino acid decarboxylases catalyze the conversion of amino acids into polyamines that are essential for cell proliferation, hypertrophy and tissue growth. In this paper, inhibitory effect of twelve OPEs with aromatic, alkyl or chlorinated alkyl substituents on the activity of lysine decarboxylase (LDC) was assessed quantitatively with an economic and label-free fluorescence sensor and cell assay. The sensor comprises a macrocyclic host (cucurbit[7]uril) and a fluorescent dye (acridine orange) reporter. The twelve OPEs were found to vary in their capacity to inhibit LDC activity. Alkyl group substituted OPEs had no inhibitory effect. By contrast, six OPEs substituted with aromatic or chlorinated alkyl groups inhibited LDC activity significantly with IC50 ranging from 1.32 μM to 9.07 μM. Among them, the inhibitory effect of tri-m-cresyl phosphate (TCrP) was even more effective as an inhibitor than guanosine 5'-diphosphate-3'-diphosphate (ppGpp) (1.60 μM), an LDC natural inhibitor in vivo. Moreover, at non-cytotoxic concentrations, these six OPEs showed perceptible inhibitory effects on LDC activity in PC12 living cells, and led to a marked loss in the cadaverine content. Molecular docking analysis of the LDC/OPE complexes revealed that different binding modes contribute to the difference in their inhibitory effect. Our finding suggested that LDC, as a new potential biological target of OPEs, might be implicated in toxicological and pathogenic mechanism of OPEs. PMID:25264276

  4. Evaluation of antifungal activity of white-colored mineral trioxide aggregate on different strains of Candida albicans in vitro

    PubMed Central

    Bhardwaj, Archana; Bhardwaj, Abhishek; Rao, Nageshwar

    2014-01-01

    Aim: The purpose of this study was to evaluate the antifungal action of various concentrations of white mineral trioxide aggregate (MTA) against seven different strains of Candida albicans using the tube dilution test. Materials and Methods: Fresh mix of MTA was prepared at concentrations of 100, 50, 25, and 12.5 mg/ml and added to a broth tube containing Sabouraud's liquid medium. A total of 1287 broth tubes were prepared and divided into experimental and control groups. Stock cultures of seven strains of C. albicans were obtained. Fresh inoculate of the microorganism was prepared by growing overnight cultures. Aliquots of the test C. albicans were taken and added to the test tubes. All tubes were incubated at 37°C for 1-, 24-, 72-, and 168-h time periods. At each time period, the presence of C. albicans colonies was assessed. Statistical analysis used: Differences among the groups were statistically analyzed using Kruskal–Wallis and Mann–Whitney U tests. Results: Results showed that one strain showed resistance even after 3 days at the lower MTA concentrations of 12.5 and 25 mg/ml. Growth reoccurred with three strains at MTA concentration of 12.5 mg/ml after 7 days. A significant difference was found between strain 3 and other strains at MTA concentrations of 12.5 and 25 mg/ml at the 3-days time period and between tubes containing 12.5 mg/ml and tubes containing higher concentrations of MTA at the 7-days time period. Conclusion: White MTA in concentrations of 100 and 50 mg/ml is effective in inhibiting the seven tested strains of C. albicans for periods up to 1 week. PMID:24944454

  5. Terpenoids inhibit Candida albicans growth by affecting membrane integrity and arrest of cell cycle.

    PubMed

    Zore, Gajanan B; Thakre, Archana D; Jadhav, Sitaram; Karuppayil, S Mohan

    2011-10-15

    Anti-Candida potential of six terpenoids were evaluated in this study against various isolates of Candida albicans (n=39) and non-C. albicans (n=9) that are differentially susceptible to fluconazole. All the six terpenoids tested, showed excellent activity and were equally effective against isolates of Candida sps., tested in this study. Linalool and citral were the most effective ones, inhibiting all the isolates at ≤0.064% (v/v). Five among the six terpenoids tested were fungicidal. Time dependent kill curve assay showed that MFCs of linalool and eugenol were highly toxic to C. albicans, killing 99.9% inoculum within seven min of exposure, while that of citronellal, linalyl acetate and citral required 15min, 1h and 2h, respectively. FIC index values (Linalool - 0.140, benzyl benzoate - 0.156, eugenol - 0.265, citral - 0.281 and 0.312 for linalyl acetate and citronellal) and isobologram obtained by checker board assay showed that all the six terpenoids tested exhibit excellent synergistic activity with fluconazole against a fluconazole resistant strain of C. albicans. Terpenoids tested arrested C. albicans cells at different phases of the cell cycle i.e. linalool and LA at G1, citral and citronellal at S phase and benzyl benzoate at G2-M phase and induced apoptosis. Linalool, citral, citronellal and benzyl benzoate caused more than 50% inhibition of germ tube induction at 0.008%, while eugenol and LA required 0.032 and 0.016% (v/v) concentrations, respectively. MICs of all the terpenoids for the C. albicans growth were non toxic to HeLa cells. Terpenoids tested exhibited excellent activity against C. albicans yeast and hyphal form growth at the concentrations that are non toxic to HeLa cells. Terpenoids tested in this study may find use in antifungal chemotherapy, not only as antifungal agents but also as synergistic agents along with conventional drugs like fluconazole.

  6. [In vitro biofilm formation and relationship with antifungal resistance of Candida spp. isolated from vaginal and intrauterine device string samples of women with vaginal complaints].

    PubMed

    Calışkan, Seyda; Keçeli Özcan, Sema; Cınar, Selvi; Corakçı, Aydın; Calışkan, Eray

    2011-10-01

    Intrauterin device (IUD) application is a widely used effective, safe and economic method for family planning. However IUD use may cause certain changes in vaginal ecosystem and may disturb microflora leading to increased colonization of various opportunistic pathogen microorganisms. The aims of this study were (i) to detect the biofilm production characteristics of Candida spp. isolated from vaginal and IUD string samples of women with IUDs, and (ii) to investigate the relationship between biofilm production and antifungal resistance. A total of 250 women (mean age: 34.4 ± 7.6 years) admitted to gynecology outpatient clinics with vaginal symptoms (discharge and itching) were included in the study. The patients have been implanted CuT380a type IUDs for a mean duration of 59.8 ± 42.4 months. Without removing IUD, string samples were obtained by cutting and simultaneous vaginal swab samples were also collected. Isolated Candida spp. were identified by conventional methods and API 20C AUX (BioMerieux, Fransa) system. Minimal inhibitory concentrations (MIC) of fluconazole, itraconazole and amphotericin B were determined by broth microdilution method according to the CLSI guidelines. Biofilm formation was evaluated by crystal violet staining and XTT-reduction assays, and the isolates which yielded positive results in both of the methods were accepted as biofilm-producers. In the study, Candida spp. were isolated from 33.2% (83/250) of the vaginal and 34% (85/250) of the IUD string samples, C.albicans being the most frequently detected species (54 and 66 strains for the samples, respectively). The total in vitro biofilm formation rate was 25% (21/83) for vaginal isolates and 44.7% (38/85) for IUD string isolates. Biofilm formation rate of vaginal C.albicans isolates was significantly lower than vaginal non-albicans Candida spp. (14.8% and 44.8%, respectively; p= 0.003). Biofilm formation rate of C.albicans strains isolated from vaginal and IUD string samples were found

  7. Tests in vitro and in pots with certain chemicals for inhibition of Pseudomonas solanacearum.

    PubMed

    el-Goorani, M A; Abo-el-Dahab, M K; Wagih, E E

    1978-01-01

    Twenty one isolates of Pseudomonas solanacearum E. F. Smith (Race 3) from various parts of Egypt were inhibited in vitro by Ampicillin, Chloramphenicol, Kanamycin, Oxytetracycline. Tetracycline, Penicillin G, Streptomycin, Nabam (Dithane A-40), Maneb (Dithane M-22), Zinc-ion maneb complex (Dithane M-45), and the insecticide Chlorthion. On the basis of in vitro-sensitivity to the selected 11 chemicals the 21 isolates could not be separated into different groups. Carbendazin (Bavistin), Benomyl, Drazoxolon (Mil-Col), and Temik proved ineffective in inhibiting the in vitro growth of all isolates at all tested concentrations. Preliminary investigations indicate that drenching the soil with solutions of Dithane M-22 (0.25% w/v, Dithane M-45 (0.25%) w/v, or insecticide Chlorthion (50 microgram/ml active ingredient) decreased the incidence of potato wilt disease that developed in soil, artificially infested with P. solanacearum. PMID:696045

  8. In vitro and in vivo stimulation of neutrophil migration and lymphocyte transformation by thiamine related to inhibition of the peroxidase/H2O2/halide system.

    PubMed Central

    Theron, A; Anderson, R; Grabow, G; Meiring, J L

    1981-01-01

    The effects of thiamine on neutrophil functions and mitogen-induced lymphocyte transformation were investigated in vitro and in vivo in adult volunteers following the injection of 50 mg thiamine intramuscularly. Thiamine caused stimulation of neutrophil motility in vitro and in vivo and increased lymphocyte transformation in vivo. Enhancement of these functions was related to inhibition of neutrophil post-phagocytic iodination of Candida albicans by the MPO/H2O2/halide system. The horseradish peroxidase/-H2O2/125 I-mediated iodination of bovine serum albumin was also inhibited by thiamine concentrations which caused increased neutrophil motility. It was found that preincubation of neutrophils and lymphocytes with the horseradish peroxidase/H2O2/halide system caused considerable inhibition of the migratory and proliferative responses respectively. Inclusion of thiamine at concentrations which were found to inhibit the peroxidase/-H2O2/halide system protected the neutrophil migratory and lymphocyte proliferative responses from inactivation by this system. It is suggested that thiamine may cause increased neutrophil migration and lymphocyte transformation by protecting these cells from toxic oxidative products generated by the peroxidase/H2O2/halide system. PMID:6273033

  9. Inhibiting the immunoproteasome exacerbates the pathogenesis of systemic Candida albicans infection in mice

    PubMed Central

    Mundt, Sarah; Basler, Michael; Buerger, Stefanie; Engler, Harald; Groettrup, Marcus

    2016-01-01

    Apart from its role in MHC class I antigen processing, the immunoproteasome has recently been implicated in the modulation of T helper cell differentiation under polarizing conditions in vitro and in the pathogenesis of autoimmune diseases in vivo. In this study, we investigated the influence of LMP7 on T helper cell differentiation in response to the fungus Candida albicans. We observed a strong effect of ONX 0914, an LMP7-selective inhibitor of the immunoproteasome, on IFN-γ and IL-17A production by murine splenocytes and human peripheral blood mononuclear cells (PBMCs) stimulated with C. albicans in vitro. Using a murine model of systemic candidiasis, we could confirm reduced generation of IFN-γ- and IL-17A-producing cells in ONX 0914 treated mice in vivo. Interestingly, ONX 0914 treatment resulted in increased susceptibility to systemic candidiasis, which manifested at very early stages of infection. Mice treated with ONX 0914 showed markedly increased kidney and brain fungal burden which resulted in enhanced neutrophil recruitment and immunopathology. Together, these results strongly suggest a role of the immunoproteasome in promoting proinflammatory T helper cells in response to C. albicans but also in affecting the innate antifungal immunity in a T helper cell-independent manner. PMID:26776888

  10. Curcumin inhibits the replication of enterovirus 71 in vitro

    PubMed Central

    Qin, Ying; Lin, Lexun; Chen, Yang; Wu, Shuo; Si, Xiaoning; Wu, Heng; Zhai, Xia; Wang, Yan; Tong, Lei; Pan, Bo; Zhong, Xiaoyan; Wang, Tianying; Zhao, Wenran; Zhong, Zhaohua

    2014-01-01

    Human enterovirus 71 (EV71) is the main causative pathogen of hand, foot, and mouth disease (HFMD) in children. The epidemic of HFMD has been a public health problem in Asia-Pacific region for decades, and no vaccine and effective antiviral medicine are available. Curcumin has been used as a traditional medicine for centuries to treat a diversity of disorders including viral infections. In this study, we demonstrated that curcumin showed potent antiviral effect again EV71. In Vero cells infected with EV71, the addition of curcumin significantly suppressed the synthesis of viral RNA, the expression of viral protein, and the overall production of viral progeny. Similar with the previous reports, curcumin reduced the production of ROS induced by viral infection. However, the antioxidant property of curcumin did not contribute to its antiviral activity, since N-acetyl-l-cysteine, the potent antioxidant failed to suppress viral replication. This study also showed that extracellular signal-regulated kinase (ERK) was activated by either viral infection or curcumin treatment, but the activated ERK did not interfere with the antiviral effect of curcumin, indicating ERK is not involved in the antiviral mechanism of curcumin. Unlike the previous reports that curcumin inhibited protein degradation through ubiquitin–proteasome system (UPS), we found that curcumin had no impact on UPS in control cells. However, curcumin did reduce the activity of proteasomes which was increased by viral infection. In addition, the accumulation of the short-lived proteins, p53 and p21, was increased by the treatment of curcumin in EV71-infected cells. We further probed the antiviral mechanism of curcumin by examining the expression of GBF1 and PI4KB, both of which are required for the formation of viral replication complex. We found that curcumin significantly reduced the level of both proteins. Moreover, the decreased expression of either GBF1 or PI4KB by the application of siRNAs was

  11. Cinnamon extract inhibits tau aggregation associated with Alzheimer’s Disease in vitro

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An aqueous extract of Ceylon cinnamon (C. zeylanicum) was found to inhibit tau aggregation and filament formation, hallmarks of Alzheimer’s disease (AD) in vitro using brain cells taken from patients who died with AD. The extract also promoted complete disassembly of recombinant tau filaments, and ...

  12. INHIBITION OF THE SECONDARY ANTIBODY RESPONSE IN VITRO BY SALICYLATE AND GENTISATE

    PubMed Central

    Ambrose, Charles Tesch

    1966-01-01

    Salicylate inhibition of the secondary antibody response initiated in vitro on day 0 has been studied in cultures of rabbit lymph node fragments. Levels of 1.25 to 1.5 mM (0.20 to 0.24 mg/ml) sodium salicylate present in serum-free medium throughout an 18- or 21-day culture period completely inhibit the secondary response. This inhibition is largely accomplished by the drug's action during the first 9 days, which corresponds to the inductive phase for this culture system. Relatively little inhibition is produced by adding the drug only after day 9, although over 90% of the antibody produced during a 21-day experiment is synthesized after day 9. Studies with media of different pH's show that this inhibition is more correctly a function of the nonionized salicylic acid concentration in the medium than of the total salicylate concentration. Arguments are presented against the possibility that salicylate at the levels used here inhibits antibody synthesis by uncoupling oxidative phosphorylation. Acetylsalicylic acid (aspirin) produces the same degree of inhibition in vitro as do equimolar concentrations of sodium salicylate. Gentisate (5-hydroxysalicylate) is 15-fold more effective in producing 50% inhibition than salicylate; its temporal pattern of inhibition is similar to that of salicylate. PMID:5922744

  13. Commercial and Plant Extract Denture Cleansers in Prevention of Candida albicans Growth on Soft Denture Reliner: In Vitro Study

    PubMed Central

    Dhaded, Sunil; Joshi, Shalini

    2016-01-01

    Objective To evaluate and compare the efficacy of two plant extracts and two commercially available denture cleansers against candida albicans adherent to soft denture reline material. Materials and Methods In this study 60 specimens of soft denture reliner material specimens were fabricated with dimensions 10x10x2 mm. The sterile specimens were inoculated by immersion in Sabourand broth containing Candida albicans for 16 hours at 37°C in an incubator. Then the specimens were washed and immersed in denture cleansers which were divided into group five groups from Group I-V for CD Clean®, Nigella sativa, thyme essential oil, Fittydent® and distilled water respectively, for 8 hours at room temperature. Then they were washed, fixed with methanol and stained with crystal violet. Candida cells adherent to the specimens were counted under microscope. The number of cells adherent to test samples were compared with that adherent to control. Results The effectiveness of Fittydent® was more than CD Clean® in reducing the adherent candida albicans and the difference was statically significant (p = <0.001). Both thyme essential oil and nigella sativa were almost same in effectiveness against candida albicans but the difference was not statically significant (p= 0.79). Post-hoc Tukey’s test was performed which indicated that Fittydent® was the most effective amongst the denture cleansers tested in this study, followed by thyme essential oil, nigella sativa and CD Clean®. Conclusion The results of the study showed that all denture cleansers used in the study were significantly effective. The study indicated that Fittydent is more effective amongst the denture cleansers because of its mechanism of action; however the plant extracts used in this study were also significantly effective against candida albicans. PMID:27042584

  14. Arsenic trioxide and reduced glutathione act synergistically to augment inhibition of thyroid peroxidase activity in vitro.

    PubMed

    Palazzolo, Dominic L; Ely, Emily A

    2015-05-01

    Thyroid peroxidase (TPO) is the enzyme involved in thyroid hormone synthesis. Arsenic trioxide (As2O3) is known to inhibit TPO activity in vitro. This inhibition is believed to occur when As2O3 binds to TPO's free sulfhydryl groups. Reduced glutathione (GSH) is also known to inhibit TPO activity in vitro. This inhibition may occur because GSH acts as a competitive substrate for hydrogen peroxide, or possibly reduce the oxidized form of iodide, requirements for TPO action. On the other hand, one could speculate that GSH reduces arsenic-induced TPO inhibition by interacting directly with arsenic or TPO, consequently limiting arsenic's ability to inhibit TPO activity. Since GSH is known to inhibit thyroid hormone synthesis while at the same time it is also known to be an important antioxidant preventing cellular damage induced by oxidative stress and protecting the thyroid gland from oxidative damage induced by arsenic, we wanted to determine if a combination of As2O3 and reduced GSH would either attenuate or augment the As2O3-induced inhibition on TPO activity. Using an in vitro system, TPO was assayed spectrophotometrically in the presence of As2O3 (0.01, 0.1, 1, and 10 ppm), GSH (0.1, 1, 5, and 10 ppm), and As2O3 (0.1 ppm) and GSH (0.01, 0.1, 1, or 10 ppm) combinations. Our results show that 0.1, 1.0, and 10 ppm As2O3 inhibit TPO activity. Similarly, 5 and 10 ppm GSH also inhibit TPO activity. When 0.1 ppm As2O3 (i.e., the lowest dose of arsenic able to partially inhibit TPO activity) is combined with 0.01, 0.1, 1.0, or 10 ppm GSH inhibition of in vitro TPO activity is augmented as indicated by complete inhibition of TPO. The mechanism of this augmentation and whether it translates to living systems remains unclear.

  15. In vitro evaluation of adherence of Candida albicans, Candida glabrata, and Streptococcus mutans to an acrylic resin modified by experimental coatings.

    PubMed

    Izumida, Fernanda Emiko; Moffa, Eduardo Buozi; Vergani, Carlos Eduardo; Machado, Ana Lúcia; Jorge, Janaína Habib; Giampaolo, Eunice Teresinha

    2014-01-01

    This study evaluated the effect of experimental coatings, containing zwitterion or hydrophilic monomers, on the adherence of Candida albicans, Candida glabrata, and Streptococcus mutans to an acrylic resin. Acrylic samples (smooth or rough surfaces) were left untreated (control) or coated with one of the following experimental coatings: 3-hydroxypropylmethacrylate (HP) or sulfobetaine methacrylate (S), at concentrations of 25, 30, or 35%. Half of the specimens were coated with saliva. The adhesion test was performed by incubating specimens in C. albicans, C. glabrata, and S. mutans suspensions at 37°C for 90 min. The number of adhered microorganisms was determined by metabolic activity (XTT) and by cell viability (CFU). All coated specimens exhibited lower absorbance and CFU values compared to control specimens. Saliva and roughness did not promote microorganism adherence. An XPS analysis confirmed the modification in the chemical composition of the coatings in the experimental samples. These experimental coatings significantly reduced the adherence of C. albicans, C. glabrata and S. mutans to acrylic resin.

  16. Inhibition of human low-density lipoprotein oxidation in vitro by ginger extracts.

    PubMed

    Gunathilake, K D Prasanna P; Rupasinghe, H P Vasantha

    2014-04-01

    Oxidative modification of low-density lipoprotein (LDL) is thought to play a key role in atherosclerotic plaque formation. Currently, there is a renewed interest in ginger because of its antioxidants and cardioprotective properties. The effects of ethanol, methanol, ethyl acetate, and hexane solvent extracts of ginger and pure major ginger constituents on Cu(2+)-induced oxidation of human LDL in vitro were examined. The LDL oxidation inhibition by ethanol, methanol, ethyl acetate, and hexane extracts of ginger was 71%, 76%, 67%, and 67%, respectively, at their optimum extraction conditions. Inhibition of LDL oxidation by water extracts of ginger, which was prepared by ultrasonic-assisted extraction conditions of 52°C for 15 min, was about 43%. Phenolic bioactives of ginger-6-gingerols, 8-gingerols, 10-gingerols, and 6-shogaol-seem to be strong inhibitors of Cu(+2)-induced LDL oxidation. Overall, ginger extracts, including the water extract possess the antioxidant activities to inhibit human LDL oxidation in vitro.

  17. Diverse role of microbially bioconverted product of cabbage (Brassica oleracea) by Pseudomonas syringe pv. T1 on inhibiting Candida species.

    PubMed

    Bajpai, Vivek K; Kang, Sun Chul; Park, Euiho; Jeon, Weon-Tai; Baek, Kwang-Hyun

    2011-02-01

    This study was carried out to evaluate the anticandidal effects of bioconverted product, obtained from the microbial conversion of cabbage (Brassica oleracea) by a bacterial strain Pseudomonas syringe pv. T1 (Ps-T1) against various isolates of Candida species. The diameters of zones of inhibition of bioconverted product of cabbage (10 μl, corresponding to 500 μg/disc) against Candida albicans KACC 30003 and 30062, Candida geochares KACC 30061, Candida saitoana KACC 41238 and Candida glabrata P00368 were found between 10±1 and 16±0.8 mm. The bioconverted product was tested for the minimum inhibitory and minimum fungicidal concentration values against the tested pathogens which were found in the range of 125-500 and 125-500 μg/ml, respectively. On the viable counts of the tested fungal pathogens, the bioconverted product showed a remarkable anticandidal effect. Also the study of using scanning electron microscopy on the morphology of C.albicans KACC 30062 revealed potential detrimental effect of bioconverted product at MIC concentration. The results of this study suggest that bioconverted product of cabbage by Ps-T1 holds potential therapeutic value and medicinal significance to control Candida species.

  18. ROCK Inhibition Facilitates In Vitro Expansion of Glioblastoma Stem-Like Cells

    PubMed Central

    Tilson, Samantha G.; Haley, Elizabeth M.; Triantafillu, Ursula L.; Dozier, David A.; Langford, Catherine P.; Gillespie, G. Yancey; Kim, Yonghyun

    2015-01-01

    Due to their stem-like characteristics and their resistance to existing chemo- and radiation therapies, there is a growing appreciation that cancer stem cells (CSCs) are the root cause behind cancer metastasis and recurrence. However, these cells represent a small subpopulation of cancer cells and are difficult to propagate in vitro. Glioblastoma is an extremely deadly form of brain cancer that is hypothesized to have a subpopulation of CSCs called glioblastoma stem cells (GSCs; also called brain tumor initiating cells, BTICs). We propose the use of selective Rho-kinase (ROCK) inhibitors, Y-27632 and fasudil, to promote GSC/BTIC-like cell survival and propagation in vitro. ROCK inhibitors have been implicated in suppressing apoptosis, and it was hypothesized that they would increase the number of GSC/BTIC-like cells grown in vitro and improve cloning efficiencies. Indeed, our data demonstrate that transient and continuous supplementation of non-toxic concentrations of Y-27632 and fasudil inhibited apoptosis, enhanced the cells’ ability to form spheres, and increased stem cell marker expressing GSC/BTIC-like cell subpopulation. Our data indicated that pharmacological and genetic (siRNA) inhibitions of the ROCK pathway facilitates in vitro expansion of GSC/BTIC-like cells. Thus, ROCK pathway inhibition shows promise for future optimization of CSC culture media. PMID:26167936

  19. Partial inhibition of in vitro pollen germination by simulated solar ultraviolet-B radiation

    SciTech Connect

    Flint, S.D.; Caldwell, M.M.

    1984-01-01

    Pollen from four temperate-latitude taxa were treated with UV radiation in a portion of the UV-B (280-320 nm) waveband during in vitro germination. Inhibition of germination was noted in this pollen compared to samples treated identically except for the exclusion of the UV-B portion of the spectrum. Levels similar to maximum solar UV-B found in temperate-latitude areas failed to inhibit pollen germination significantly, while levels similar to maximum solar UV-B found in equatorial alpine locations caused partial inhibition of germination in three of the four taxa examined.

  20. Inhibition on Candida albicans biofilm formation using divalent cation chelators (EDTA).

    PubMed

    Ramage, Gordon; Wickes, Brian L; López-Ribot, José L

    2007-12-01

    Candida albicans can readily form biofilms on both inanimate and biological surfaces. In this study we investigated a means of inhibiting biofilm formation using EDTA (Ethylenediaminetetra-acetic acid), a divalent cation chelating agent, which has been shown to affect C. albicans filamentation. Candida albicans biofilms were formed in 96-well microtitre plates. Cells were allowed to adhere for 1, 2, and 4 h at 37 degrees C, washed in PBS, and then treated with different concentrations of EDTA (0, 2.5, 25, and 250 mM). EDTA was also added to the standardized suspension prior to adding to the microtiter plate and to a preformed 24 h biofilm. All plates were then incubated at 37 degrees C for an additional 24 h to allow for biofilm formation. The extent and characteristics of biofilm formation were then microscopically assessed and with a semi-quantitative colorimetric technique based on the use of an XTT-reduction assay. Northern blot analysis of the hyphal wall protein (HWP1) expression was also monitored in planktonic and biofilm cells treated with EDTA. Microscopic analysis and colorimetric readings revealed that filamentation and biofilm formation were inhibited by EDTA in a concentration dependent manner. However, preformed biofilms were minimally affected by EDTA (maximum of 31% reduction at 250 mM). The HWP1 gene expression was reduced in EDTA-treated planktonic and biofilm samples. These results indicate that EDTA inhibits C. albicans biofilm formation are most likely through its inhibitory effect on filamentation and indicates the potential therapeutic effects of EDTA. This compound may serve a non-toxic means of preventing biofilm formation on infections with a C. albicans biofilm etiology. PMID:17909983

  1. Sustained Nitric Oxide-Releasing Nanoparticles Induce Cell Death in Candida albicans Yeast and Hyphal Cells, Preventing Biofilm Formation In Vitro and in a Rodent Central Venous Catheter Model

    PubMed Central

    Ahmadi, Mohammed S.; Lee, Hiu Ham; Sanchez, David A.; Friedman, Adam J.; Tar, Moses T.; Davies, Kelvin P.; Nosanchuk, Joshua D.

    2016-01-01

    Candida albicans is a leading nosocomial pathogen. Today, candidal biofilms are a significant cause of catheter infections, and such infections are becoming increasingly responsible for the failure of medical-implanted devices. C. albicans forms biofilms in which fungal cells are encased in an autoproduced extracellular polysaccharide matrix. Consequently, the enclosed fungi are protected from antimicrobial agents and host cells, providing a unique niche conducive to robust microbial growth and a harbor for recurring infections. Here we demonstrate that a recently developed platform comprised of nanoparticles that release therapeutic levels of nitric oxide (NO-np) inhibits candidal biofilm formation, destroys the extracellular polysaccharide matrices of mature fungal biofilms, and hinders biofilm development on surface biomaterials such as the lumen of catheters. We found NO-np to decrease both the metabolic activity of biofilms and the cell viability of C. albicans in vitro and in vivo. Furthermore, flow cytometric analysis found NO-np to induce apoptosis in biofilm yeast cells in vitro. Moreover, NO-np behave synergistically when used in combination with established antifungal drug therapies. Here we propose NO-np as a novel treatment modality, especially in combination with standard antifungals, for the prevention and/or remediation of fungal biofilms on central venous catheters and other medical devices. PMID:26810653

  2. Waikialoid A Suppresses Hyphal Morphogenesis and Inhibits Biofilm Development in Pathogenic Candida albicans

    PubMed Central

    Wang, Xiaoru; You, Jianlan; King, Jarrod B.; Powell, Douglas R.; Cichewicz, Robert H.

    2012-01-01

    A chemically prolific strain of Aspergillus was isolated from a soil sample collected near Waikiki Beach, Honolulu, Hawaii. The fungus produced several secondary metabolites that were purified and placed in our natural products library, which was later screened for substances capable of inhibiting biofilm formation by Candida albicans. It was determined that one of the secondary metabolites from the Hawaiian fungal isolate, a new complex prenylated indole alkaloid named waikialoid A (1), inhibited biofilm formation with an IC50 value of 1.4 μM. Another structurally unrelated, presumably polyketide metabolite, waikialide A (15), also inhibited C. albicans biofilm formation, but was much less potent (IC50 value of 32.4 μM). Microscopy studies revealed that compound 1 also inhibited C. albicans hyphal morphogenesis. While metabolite 1 appears ineffective at disrupting preformed biofilms, the accumulated data indicate that the new compound may exert its activity against C. ablicans during the early stages of surface colonization involving cell adherence, hyphal development, and/or biofilm assembly. Unlike some other stephacidin/notoamide compounds, metabolite 1 was not cytotoxic to fungi or human cells (up to 200 μM), which makes this an intriguing model compound for studying the adjunctive use of biofilm inhibitors in combination with standard antifungal antibiotics. PMID:22400916

  3. Waikialoid A suppresses hyphal morphogenesis and inhibits biofilm development in pathogenic Candida albicans.

    PubMed

    Wang, Xiaoru; You, Jianlan; King, Jarrod B; Powell, Douglas R; Cichewicz, Robert H

    2012-04-27

    A chemically prolific strain of Aspergillus was isolated from a soil sample collected near Waikiki Beach, Honolulu, Hawaii. The fungus produced several secondary metabolites, which were purified and placed in our natural products library and were later screened for substances capable of inhibiting biofilm formation by Candida albicans. It was determined that one of the secondary metabolites from the Hawaiian fungal isolate, a new complex prenylated indole alkaloid named waikialoid A (1), inhibited biofilm formation with an IC(50) value of 1.4 μM. Another structurally unrelated, presumably polyketide metabolite, waikialide A (15), also inhibited C. albicans biofilm formation, but was much less potent (IC(50) value of 32.4 μM). Microscopy studies revealed that compound 1 also inhibited C. albicans hyphal morphogenesis. While metabolite 1 appears ineffective at disrupting preformed biofilms, the accumulated data indicate that the new compound may exert its activity against C. albicans during the early stages of surface colonization involving cell adherence, hyphal development, and/or biofilm assembly. Unlike some other stephacidin/notoamide compounds, metabolite 1 was not cytotoxic to fungi or human cells (up to 200 μM), which makes this an intriguing model compound for studying the adjunctive use of biofilm inhibitors in combination with standard antifungal antibiotics. PMID:22400916

  4. Recombinant tumour necrosis factor inhibits malaria parasites in vivo but not in vitro.

    PubMed Central

    Taverne, J; Tavernier, J; Fiers, W; Playfair, J H

    1987-01-01

    As tumour necrosis serum kills malarial parasites in vitro and inhibits the multiplication of some species of Plasmodium in mice, we examined the effect of recombinant mouse tumour necrosis factor (rTNF) on P. yoelii both in vitro and in vivo. Parasites incubated overnight with rTNF showed no loss of viability, but repeated injection of rTNF into infected mice reduced parasitaemia and significantly prolonged survival of mice infected with a lethal variant of the parasite. We conclude that TNF acts on blood-stage malaria in vivo via a host cell and that another molecule in tumour necrosis serum is involved in killing the parasite in vitro. PMID:3621669

  5. Lefty inhibits in vitro decidualization by regulating P57 and cyclin D1 expressions.

    PubMed

    Li, Hong; Li, Hui; Bai, Liang; Yu, Hua

    2014-12-01

    Endometrial decidualization is highly important for successful construction and maintenance of embryo implantation and pregnancy. Lefty gene at different menstrual cycle phases has different expressions, indicating its regulatory significance. To study the mechanism of Lefty in decidualization, human endometrial stromal cells (hESCs) were cultured and induced with medroxyprogesterone acetate (MPA) and 8-bromoadenosine-cAMP (8-Br-cAMP) in vitro as a research model. Our results showed that Lefty1 overexpression inhibited MPA- and 8-Br-cAMP-induced hESC decidualization and significantly reduced the secretion of prolactin (PRL) and insulin-like growth factor-binding protein 1 (IGFBP-1). With the inhibition of Lefty1 expression, hESC decidualization induced by MPA and 8-Br-cAMP became more remarkable, and the secretions of PRL and IGFBP-1 were higher too. Further tests indicated that during the process of decidualization, P57 expression increased, whereas cyclin D1 expression decreased. Although Lefty1 overexpression did not significantly change the expressions of P57 and cyclin D1, inhibition of Lefty1 expression resulted in more evident changes in P57 and cyclin D1 expressions. Meanwhile, cell cycle examination showed that Lefty1 overexpression reduced the cell cycle arrest at G1/S phase in the in vitro hESC decidualization model. Therefore, Lefty1 could regulate the cell cycle via modulating the expressions of P57 and cyclin D1 and then inhibit the decidualization in vitro. PMID:25339094

  6. Inhibition of Mycobacterial Growth In Vitro following Primary but Not Secondary Vaccination with Mycobacterium bovis BCG

    PubMed Central

    Fletcher, Helen A.; Tanner, Rachel; Wallis, Robert S.; Meyer, Joel; Manjaly, Zita-Rose; Harris, Stephanie; Satti, Iman; Silver, Richard F.; Hoft, Dan; Kampmann, Beate; Walker, K. Barry; Dockrell, Hazel M.; Fruth, Uli; Barker, Lew; McShane, Helen

    2013-01-01

    Despite the widespread use of the Mycobacterium bovis BCG vaccine, there are more than 9 million new cases of tuberculosis (TB) every year, and there is an urgent need for better TB vaccines. TB vaccine candidates are selected for evaluation based in part on the detection of an antigen-specific gamma interferon (IFN-γ) response. The measurement of mycobacterial growth in blood specimens obtained from subjects immunized with investigational TB vaccines may be a better in vitro correlate of in vivo vaccine efficacy. We performed a clinical study with 30 United Kingdom adults who were followed for 6 months to evaluate the abilities of both a whole-blood- and a novel peripheral blood mononuclear cell (PBMC)-based mycobacterial growth inhibition assay to measure a response to primary vaccination and revaccination with BCG. Using cryopreserved PBMCs, we observed a significant improvement in mycobacterial growth inhibition following primary vaccination but no improvement in growth inhibition following revaccination with BCG (P < 0.05). Mycobacterial growth inhibition following primary BCG vaccination was not correlated with purified protein derivative (PPD) antigen-specific IFN-γ enzyme-linked immunospot (ELISPOT) responses. We demonstrate that a mycobacterial growth inhibition assay can detect improved capacity to control growth following primary immunization, but not revaccination, with BCG. This is the first study to demonstrate that an in vitro growth inhibition assay can identify a difference in vaccine responses by comparing both primary and secondary BCG vaccinations, suggesting that in vitro growth inhibition assays may serve as better surrogates of clinical efficacy than the assays currently used for the assessment of candidate TB vaccines. PMID:23986316

  7. Synergistic Inhibition of Angiogenesis by Artesunate and Captopril In Vitro and In Vivo

    PubMed Central

    Krusche, Benjamin; Arend, Joachim; Efferth, Thomas

    2013-01-01

    Inhibition of angiogenesis represents one major strategy of cancer chemotherapy. In the present investigation, we investigated the synergism of artesunate and captopril to inhibit angiogenesis. Artesunate is an antimalarial derivative of artemisinin from the Chinese medicinal plant, Artemisia annua L., which also reveals profound anticancer activity in vitro and in vivo. Captopril is an angiotensin I-converting (ACE) inhibitor, which is well established in Western academic medicine. Both compounds inhibited migration of human umbilical vein endothelial cells (HUVECs) in vitro. The combination of both drugs resulted in synergistically inhibited migration. Whereas artesunate inhibited HUVEC growth in the XTT assay, captopril did not, indicating independent modes of action. We established a chorioallantoic membrane (CAM) assay of quail embryos (Coturnix coturnix L.) and a computer-based evaluation routine for quantitative studies on vascularization processes in vivo. Artesunate and captopril inhibited blood vessel formation and growth. For the first time, we demonstrated that both drugs revealed synergistic effects when combined. These results may also have clinical impact, since cardiovascular diseases and cancer frequently occur together in older cancer patients. Therefore, comorbid patients may take advantage, if they take captopril to treat cardiovascular symptoms and artesunate to treat cancer. PMID:24223058

  8. Kinetic characterization and in vitro toxicity evaluation of a luciferase less susceptible to HPV chemical inhibition.

    PubMed

    Kim-Choi, Eileen; Danilo, Christiane; Kelly, Jeffrey; Carroll, Ronnie; Shonnard, David; Rybina, Irina

    2006-12-01

    Enzyme-based in vitro toxicity assays are often susceptible to inhibition by test compounds. A mutant luciferase selected to be less susceptible to inhibition by chloroform (CNBLuc03-06) and other high production volume (HPV) chemicals, consisting of three point mutations was created and characterized. The mutant luciferase was less inhibited by chloroform, other HPV chemicals and common surfactant release reagents (Triton-X and SDS) compared to the wild-type. Inhibition was shown to be competitive. CNBLuc03-06 was a factor of 1.5-3.2 more active than wild type and exhibited a much higher affinity for ATP. CNBLuc03-06 was more thermostable than wild-type and also more active at pH values higher than 10. Both luciferases exhibited a significant tradeoff between activation and susceptibility to chemical inhibition in the presences of the reducing agent DTT. Inhibition to HPV chemicals was eliminated using an "optimum" formulation of DTT and co-solvent ethanol. The performance of CNBLuc03-06 in cell-based in vitro toxicity assays was shown to be superior to the current commercial formulation.

  9. Synergistic inhibition of angiogenesis by artesunate and captopril in vitro and in vivo.

    PubMed

    Krusche, Benjamin; Arend, Joachim; Efferth, Thomas

    2013-01-01

    Inhibition of angiogenesis represents one major strategy of cancer chemotherapy. In the present investigation, we investigated the synergism of artesunate and captopril to inhibit angiogenesis. Artesunate is an antimalarial derivative of artemisinin from the Chinese medicinal plant, Artemisia annua L., which also reveals profound anticancer activity in vitro and in vivo. Captopril is an angiotensin I-converting (ACE) inhibitor, which is well established in Western academic medicine. Both compounds inhibited migration of human umbilical vein endothelial cells (HUVECs) in vitro. The combination of both drugs resulted in synergistically inhibited migration. Whereas artesunate inhibited HUVEC growth in the XTT assay, captopril did not, indicating independent modes of action. We established a chorioallantoic membrane (CAM) assay of quail embryos (Coturnix coturnix L.) and a computer-based evaluation routine for quantitative studies on vascularization processes in vivo. Artesunate and captopril inhibited blood vessel formation and growth. For the first time, we demonstrated that both drugs revealed synergistic effects when combined. These results may also have clinical impact, since cardiovascular diseases and cancer frequently occur together in older cancer patients. Therefore, comorbid patients may take advantage, if they take captopril to treat cardiovascular symptoms and artesunate to treat cancer. PMID:24223058

  10. In vitro antifungal activity of fluconazole and voriconazole against non-Candida yeasts and yeast-like fungi clinical isolates.

    PubMed

    Mandras, Narcisa; Roana, Janira; Scalas, Daniela; Fucale, Giacomo; Allizond, Valeria; Banche, Giuliana; Barbui, Anna; Li Vigni, Nicolò; Newell, Vance A; Cuffini, Anna Maria; Tullio, Vivian

    2015-10-01

    The risk of opportunistic infections caused by non-Candida yeasts and yeast-like fungi is increasingly common, mainly in immunocompromised patients. Appropriate first-line therapy has not been defined and standardized, mainly due to the low number of cases reported. To improve empirical treatment guidelines, we describe the susceptibility profile to fluconazole and voriconazole of 176 non-Candida yeasts and yeast-like fungi collected from hospitals in Piedmont, North West Italy from January 2009 to December 2013. The results showed that most isolates are susceptible to voriconazole (94%), but less susceptible to fluconazole (78%), suggesting that voriconazole could be used as first-line therapy in infections caused by these fungi.

  11. Candida albicans Inhibits Pseudomonas aeruginosa Virulence through Suppression of Pyochelin and Pyoverdine Biosynthesis

    PubMed Central

    Lopez-Medina, Eduardo; Fan, Di; Coughlin, Laura A.; Ho, Evi X.; Lamont, Iain L.; Reimmann, Cornelia; Hooper, Lora V.; Koh, Andrew Y.

    2015-01-01

    Bacterial-fungal interactions have important physiologic and medical ramifications, but the mechanisms of these interactions are poorly understood. The gut is host to trillions of microorganisms, and bacterial-fungal interactions are likely to be important. Using a neutropenic mouse model of microbial gastrointestinal colonization and dissemination, we show that the fungus Candida albicans inhibits the virulence of the bacterium Pseudomonas aeruginosa by inhibiting P. aeruginosa pyochelin and pyoverdine gene expression, which plays a critical role in iron acquisition and virulence. Accordingly, deletion of both P. aeruginosa pyochelin and pyoverdine genes attenuates P. aeruginosa virulence. Heat-killed C. albicans has no effect on P. aeruginosa, whereas C. albicans secreted proteins directly suppress P. aeruginosa pyoverdine and pyochelin expression and inhibit P. aeruginosa virulence in mice. Interestingly, suppression or deletion of pyochelin and pyoverdine genes has no effect on P. aeruginosa’s ability to colonize the GI tract but does decrease P. aeruginosa’s cytotoxic effect on cultured colonocytes. Finally, oral iron supplementation restores P. aeruginosa virulence in P. aeruginosa and C. albicans colonized mice. Together, our findings provide insight into how a bacterial-fungal interaction can modulate bacterial virulence in the intestine. Previously described bacterial-fungal antagonistic interactions have focused on growth inhibition or colonization inhibition/modulation, yet here we describe a novel observation of fungal-inhibition of bacterial effectors critical for virulence but not important for colonization. These findings validate the use of a mammalian model system to explore the complexities of polymicrobial, polykingdom infections in order to identify new therapeutic targets for preventing microbial disease. PMID:26313907

  12. Candida albicans Inhibits Pseudomonas aeruginosa Virulence through Suppression of Pyochelin and Pyoverdine Biosynthesis.

    PubMed

    Lopez-Medina, Eduardo; Fan, Di; Coughlin, Laura A; Ho, Evi X; Lamont, Iain L; Reimmann, Cornelia; Hooper, Lora V; Koh, Andrew Y

    2015-08-01

    Bacterial-fungal interactions have important physiologic and medical ramifications, but the mechanisms of these interactions are poorly understood. The gut is host to trillions of microorganisms, and bacterial-fungal interactions are likely to be important. Using a neutropenic mouse model of microbial gastrointestinal colonization and dissemination, we show that the fungus Candida albicans inhibits the virulence of the bacterium Pseudomonas aeruginosa by inhibiting P. aeruginosa pyochelin and pyoverdine gene expression, which plays a critical role in iron acquisition and virulence. Accordingly, deletion of both P. aeruginosa pyochelin and pyoverdine genes attenuates P. aeruginosa virulence. Heat-killed C. albicans has no effect on P. aeruginosa, whereas C. albicans secreted proteins directly suppress P. aeruginosa pyoverdine and pyochelin expression and inhibit P. aeruginosa virulence in mice. Interestingly, suppression or deletion of pyochelin and pyoverdine genes has no effect on P. aeruginosa's ability to colonize the GI tract but does decrease P. aeruginosa's cytotoxic effect on cultured colonocytes. Finally, oral iron supplementation restores P. aeruginosa virulence in P. aeruginosa and C. albicans colonized mice. Together, our findings provide insight into how a bacterial-fungal interaction can modulate bacterial virulence in the intestine. Previously described bacterial-fungal antagonistic interactions have focused on growth inhibition or colonization inhibition/modulation, yet here we describe a novel observation of fungal-inhibition of bacterial effectors critical for virulence but not important for colonization. These findings validate the use of a mammalian model system to explore the complexities of polymicrobial, polykingdom infections in order to identify new therapeutic targets for preventing microbial disease. PMID:26313907

  13. In vitro inhibition of cytochrome P450 3A4 by Aronia melanocarpa constituents.

    PubMed

    Bräunlich, Marie; Christensen, Hege; Johannesen, Siri; Slimestad, Rune; Wangensteen, Helle; Malterud, Karl E; Barsett, Hilde

    2013-01-01

    Extracts, subfractions, isolated anthocyanins and procyanidins, and two phenolic acids from aronia [Aronia melanocarpa] were investigated for their CYP3A4 inhibitory effects, using midazolam as the probe substrate and recombinant insect cell microsomes expressing CYP3A4 as the enzyme source. Procyanidin B5 was a considerably stronger CYP3A4 inhibitor in vitro than the isomeric procyanidin B2 and comparable to bergamottin, a known CYP3A4 inhibitor from grapefruit juice. The inhibitory activity of proanthocyanidin-containing fractions was correlated to the degree of polymerization. Among the anthocyanins, cyanidin 3-arabinoside showed stronger CYP3A4 inhibition than cyanidin 3-galactoside and cyanidin 3-glucoside. Thus, the ability to inhibit CYP3A4 in vitro seems to be influenced by the sugar unit linked to the anthocyanidin.

  14. Decontamination efficacy of erbium:yttrium-aluminium-garnet and diode laser light on oral Candida albicans isolates of a 5-day in vitro biofilm model.

    PubMed

    Sennhenn-Kirchner, Sabine; Schwarz, Peter; Schliephake, Henning; Konietschke, Frank; Brunner, Edgar; Borg-von Zepelin, Margarete

    2009-05-01

    The different forms of superficial and systemic candidiasis are often associated with biofilm formation on surfaces of host tissues or medical devices. The biofilm formation of Candida spp., in general, necessitates significantly increased amounts of antifungal agents for therapy. Often the therapeutic effect is doubtful. A 5-day biofilm model with oral Candida isolates was established according to Chandra et al. (J Dent Res 80:903-908, 2001) on glass and titanium surfaces and was modified by Sennhenn-Kirchner et al. (Z Zahnärztl Implantol 3:45-51, 2007) to investigate different aspects unanswered in the field of dentistry. In this model, the efficacy of erbium:yttrium-aluminium-garnet (Er:YAG) light (2940 nm, 100 mJ, 10 Hz, 300 micros pulsed mode applied for 80 s) and diode laser light (810 nm, 1 W, continuous wave mode applied for 20 s with four repetitions after 30 s pauses each) was evaluated and compared to untreated controls. The photometric evaluation of the samples was completed by observations on morphological changes of yeast cells grown in the biofilm. Compared to the untreated controls Candida cells grown in mature in vitro biofilms were significantly reduced by both wavelengths investigated. Comparison between the different methods of laser treatment additionally revealed a significantly greater effect of the Er:YAG over the diode laser. Scanning electron microscopy findings proved that the diode laser light was effective in direct contact mode. In contrast, in the areas without direct contact, the fungal cells were left almost unchanged. The Er:YAG laser damaged the fungal cells to a great extent wherever it was applied.

  15. [In vitro antimicrobial, antiadherent and antifungal activity of Brazilian medicinal plants on oral biofilm microorganisms and strains of the genus Candida].

    PubMed

    Alves, Pollianna Muniz; Queiroz, Lélia Maria Guedes; Pereira, Jozinete Vieira; Pereira, Maria do Socorro Vieira

    2009-01-01

    The antimicrobial, antifungal and antiadherent activity of aroeira-do-sertão, mallow and guava tree on oral biofilm microorganisms and oral candidiasis was evaluated in vitro. The extracts were shown to be effective in inhibiting the growth of bacteria of the oral biofilm and fungi of oral candidiasis, thus suggesting that these extracts can be used as alternative means of dental therapy.

  16. Parsley extract inhibits in vitro and ex vivo platelet aggregation and prolongs bleeding time in rats.

    PubMed

    Gadi, Dounia; Bnouham, Mohamed; Aziz, Mohammed; Ziyyat, Abderrahim; Legssyer, Abdelkhaleq; Legrand, Chantal; Lafeve, Françoise Fauvel; Mekhfi, Hassane

    2009-08-17

    Many cardiovascular diseases are associated with an increase in blood platelet activity. In Morocco, parsley (Petroselinum crispum, Apiaceae) is one of the medicinal herbs used to treat cardiovascular diseases such as arterial hypertension. In this study, crude aqueous extract (CAE) of parsley was evaluated for its anti-platelet activity in experimental animals on platelet aggregation in vitro and ex vivo; and on bleeding time in vivo. The in vitro aggregation was monitored after pre-incubation of platelets with CAE. The bleeding time and ex vivo aggregation were performed after oral treatment. CAE inhibited dose dependently platelet aggregation in vitro induced by thrombin, ADP, collagen and epinephrine. The oral administration of CAE (3g/kg) inhibited significantly (p<0.001) platelet aggregation ex vivo and prolonged bleeding time (p<0.001) without changes in the platelet amount. The prolongation of bleeding time by CAE may be attributed to the observed inhibition of platelet aggregation. These effects could be related in part to the polyphenolic compounds present in the extract. These results support the hypothesis that the dietary intake of parsley may be benefit in the normalization of platelet hyperactivation, in the nutritional prevention of cardiovascular diseases and are potentially interesting in the development of new prevention strategies.

  17. The thermal potentiation of acetaminophen-inhibited PMN oxidative metabolism in vitro.

    PubMed

    Shalabi, E A; al-Tuwaijri, A S

    1996-08-01

    The effect of high temperatures (39, 41, and 43 degrees C) on acetaminophen (AM-) induced inhibition of the oxidative respiratory burst of polymorphonuclear leukocytes (PMNs) in vitro has been examined. Whole blood or isolated human PMNs were exposed to various temperatures in vitro in the presence or absence of AM for 0-90 min. Phagocyte membrane-bound NADPH oxidase was studied using the luminol chemiluminescence (CL) response and the superoxide dismutase inhibitable reduction of ferricytochrome C. The NADPH oxidase was stimulated by phorbol myristate acetate (PMA). The results showed that high temperatures (39-43 degrees C) potentiate the AM inhibitory effect on CL peak response of phagocytes in a temperature-dependent manner. Furthermore, the inhibition of superoxide (O2-) production induced by AM was potentiated by incubating the cells at 39 or 43 degrees C at different time intervals. These studies suggest that high temperatures significantly potentiate the AM inhibitory effect on oxidative metabolism of PMNs in vitro. These actions of AM may influence the outcome in patients with infectious febrile conditions. PMID:8866041

  18. The Effect of Cyclooxygenase Inhibition on Tendon-Bone Healing in an In Vitro Coculture Model

    PubMed Central

    Schwarting, Tim; Pretzsch, Sebastian; Debus, Florian; Ruchholtz, Steffen; Lechler, Philipp

    2015-01-01

    The effects of cyclooxygenase (COX) inhibition following the reconstruction of the anterior cruciate ligament remain unclear. We examined the effects of selective COX-2 and nonselective COX inhibition on bone-tendon integration in an in vitro model. We measured the dose-dependent effects of ibuprofen and parecoxib on the viability of lipopolysaccharide- (LPS-) stimulated and unstimulated mouse MC3T3-E1 and 3T3 cells, the influence on gene expression at the osteoblast, interface, and fibroblast regions measured by quantitative PCR, and cellular outgrowth assessed on histological sections. Ibuprofen led to a dose-dependent suppression of MC3T3 cell viability, while parecoxib reduced the viability of 3T3 cultures. Exposure to ibuprofen significantly suppressed expression of Alpl (P < 0.01), Bglap (P < 0.001), and Runx2 (P < 0.01), and although parecoxib reduced expression of Alpl (P < 0.001), Fmod (P < 0.001), and Runx2 (P < 0.01), the expression of Bglap was increased (P < 0.01). Microscopic analysis showed a reduction in cellular outgrowth in LPS-stimulated cultures following exposure to ibuprofen and parecoxib. Nonselective COX inhibition and the specific inhibition of COX-2 led to region-specific reductions in markers of calcification and cell viability. We suggest further in vitro and in vivo studies examining the biologic and biomechanical effects of selective and nonselective COX inhibition. PMID:26063979

  19. An aqueous pomegranate peel extract inhibits neutrophil myeloperoxidase in vitro and attenuates lung inflammation in mice.

    PubMed

    Bachoual, Rafik; Talmoudi, Wifak; Boussetta, Tarek; Braut, Françoise; El-Benna, Jamel

    2011-06-01

    Punica granatum peel aqueous extract (PGE) is widely used to treat disorders such as inflammation, ulcers and infections, but its pharmacological target is not known. In this study we investigated the effect of PGE on human neutrophil reactive oxygen species (ROS) production in vitro and on LPS-induced lung inflammation in vivo in mice. Neutrophils were isolated and ROS generation was measured by luminol-amplified chemiluminescence. Superoxide anion generation was detected by the cytochrome c reduction assay. H(2)O(2) was detected by DCFH fluorescence assay. Myeloperoxidase (MPO) activity was measured by the tetramethyl benzidine oxidation method. Lung inflammation was induced in mice by LPS instillation. PGE inhibited luminol-amplified chemiluminescence of resting neutrophils and N-formyl-methionyl-leucyl-phenylalanine (fMLF)- or phorbol myristate acetate (PMA)-stimulated neutrophils, in a concentration-dependent manner. PGE had no effect on superoxide anion generation, suggesting that it does not directly inhibit NADPH oxidase activity or activation pathways, or scavenge superoxide anions. PGE did not scavenge H(2)O(2) but directly inhibited myeloperoxidase activity in vitro. In vivo studies showed that PGE also attenuated LPS-induced lung inflammation in mice. So this study reveals that PGE inhibits neutrophil MPO activity and attenuates LPS-induced lung inflammation in mice. Inhibition of MPO activity by PGE could explain its anti-inflammatory action. PMID:21376769

  20. An aqueous pomegranate peel extract inhibits neutrophil myeloperoxidase in vitro and attenuates lung inflammation in mice.

    PubMed

    Bachoual, Rafik; Talmoudi, Wifak; Boussetta, Tarek; Braut, Françoise; El-Benna, Jamel

    2011-06-01

    Punica granatum peel aqueous extract (PGE) is widely used to treat disorders such as inflammation, ulcers and infections, but its pharmacological target is not known. In this study we investigated the effect of PGE on human neutrophil reactive oxygen species (ROS) production in vitro and on LPS-induced lung inflammation in vivo in mice. Neutrophils were isolated and ROS generation was measured by luminol-amplified chemiluminescence. Superoxide anion generation was detected by the cytochrome c reduction assay. H(2)O(2) was detected by DCFH fluorescence assay. Myeloperoxidase (MPO) activity was measured by the tetramethyl benzidine oxidation method. Lung inflammation was induced in mice by LPS instillation. PGE inhibited luminol-amplified chemiluminescence of resting neutrophils and N-formyl-methionyl-leucyl-phenylalanine (fMLF)- or phorbol myristate acetate (PMA)-stimulated neutrophils, in a concentration-dependent manner. PGE had no effect on superoxide anion generation, suggesting that it does not directly inhibit NADPH oxidase activity or activation pathways, or scavenge superoxide anions. PGE did not scavenge H(2)O(2) but directly inhibited myeloperoxidase activity in vitro. In vivo studies showed that PGE also attenuated LPS-induced lung inflammation in mice. So this study reveals that PGE inhibits neutrophil MPO activity and attenuates LPS-induced lung inflammation in mice. Inhibition of MPO activity by PGE could explain its anti-inflammatory action.

  1. The Effect of Cyclooxygenase Inhibition on Tendon-Bone Healing in an In Vitro Coculture Model.

    PubMed

    Schwarting, Tim; Pretzsch, Sebastian; Debus, Florian; Ruchholtz, Steffen; Lechler, Philipp

    2015-01-01

    The effects of cyclooxygenase (COX) inhibition following the reconstruction of the anterior cruciate ligament remain unclear. We examined the effects of selective COX-2 and nonselective COX inhibition on bone-tendon integration in an in vitro model. We measured the dose-dependent effects of ibuprofen and parecoxib on the viability of lipopolysaccharide- (LPS-) stimulated and unstimulated mouse MC3T3-E1 and 3T3 cells, the influence on gene expression at the osteoblast, interface, and fibroblast regions measured by quantitative PCR, and cellular outgrowth assessed on histological sections. Ibuprofen led to a dose-dependent suppression of MC3T3 cell viability, while parecoxib reduced the viability of 3T3 cultures. Exposure to ibuprofen significantly suppressed expression of Alpl (P < 0.01), Bglap (P < 0.001), and Runx2 (P < 0.01), and although parecoxib reduced expression of Alpl (P < 0.001), Fmod (P < 0.001), and Runx2 (P < 0.01), the expression of Bglap was increased (P < 0.01). Microscopic analysis showed a reduction in cellular outgrowth in LPS-stimulated cultures following exposure to ibuprofen and parecoxib. Nonselective COX inhibition and the specific inhibition of COX-2 led to region-specific reductions in markers of calcification and cell viability. We suggest further in vitro and in vivo studies examining the biologic and biomechanical effects of selective and nonselective COX inhibition. PMID:26063979

  2. ROCK inhibition enhances neurite outgrowth in neural stem cells by upregulating YAP expression in vitro.

    PubMed

    Jia, Xu-Feng; Ye, Fei; Wang, Yan-Bo; Feng, Da-Xiong

    2016-06-01

    Spontaneous axonal regeneration of neurons does not occur after spinal cord injury because of inhibition by myelin and other inhibitory factors. Studies have demonstrated that blocking the Rho/Rho-kinase (ROCK) pathway can promote neurite outgrowth in spinal cord injury models. In the present study, we investigated neurite outgrowth and neuronal differentiation in neural stem cells from the mouse subventricular zone after inhibition of ROCK in vitro. Inhibition of ROCK with Y-27632 increased neurite length, enhanced neuronal differentiation, and upregulated the expression of two major signaling pathway effectors, phospho-Akt and phospho-mitogen-activated protein kinase, and the Hippo pathway effector YAP. These results suggest that inhibition of ROCK mediates neurite outgrowth in neural stem cells by activating the Hippo signaling pathway. PMID:27482229

  3. ROCK inhibition enhances neurite outgrowth in neural stem cells by upregulating YAP expression in vitro

    PubMed Central

    Jia, Xu-feng; Ye, Fei; Wang, Yan-bo; Feng, Da-xiong

    2016-01-01

    Spontaneous axonal regeneration of neurons does not occur after spinal cord injury because of inhibition by myelin and other inhibitory factors. Studies have demonstrated that blocking the Rho/Rho-kinase (ROCK) pathway can promote neurite outgrowth in spinal cord injury models. In the present study, we investigated neurite outgrowth and neuronal differentiation in neural stem cells from the mouse subventricular zone after inhibition of ROCK in vitro. Inhibition of ROCK with Y-27632 increased neurite length, enhanced neuronal differentiation, and upregulated the expression of two major signaling pathway effectors, phospho-Akt and phospho-mitogen-activated protein kinase, and the Hippo pathway effector YAP. These results suggest that inhibition of ROCK mediates neurite outgrowth in neural stem cells by activating the Hippo signaling pathway. PMID:27482229

  4. In vitro activities of new triazole antifungal agents, posaconazole and voriconazole, against oral Candida isolates from patients suffering from denture stomatitis.

    PubMed

    Marcos-Arias, Cristina; Eraso, Elena; Madariaga, Lucila; Carrillo-Muñoz, Alfonso Javier; Quindós, Guillermo

    2012-01-01

    Denture stomatitis is often treated with antifungal agents but recurrences or new episodes are common, and certain episodes can be resistant. New triazoles, such as posaconazole and voriconazole, may represent useful alternatives for management. In vitro activities of amphotericin B, nystatin, miconazole, fluconazole, itraconazole, posaconazole and voriconazole against 150 oral Candida (101 C. albicans, 18 C. tropicalis, 12 C. glabrata, 11 C. guilliermondii, 4 C. parapsilosis, 2 Saccharomyces cerevisiae, 1 C. dubliniensis and 1 C. krusei) from 100 denture wearers were tested by the CLSI M27-A3 method. Resistant isolates were retested by Sensititre YeastOne and Etest. Most antifungal agents were very active. However, 4 C. glabrata (33.3%), 2 C. tropicalis (11.1%), 6 C. albicans (5.6%) and 1 C. krusei were resistant to itraconazole. Posaconazole was active against 143 yeast isolates (95.3%): 6 C. albicans (5.9%) and 1 C. tropicalis (5.6%) were resistant. Geometric mean MICs were 0.036 μg/ml for C. parapsilosis, 0.062 μg/ml for C. albicans, 0.085 μg/ml for C. tropicalis, 0.387 μg/ml for C. guilliermondii and 0.498 μg/ml for C. glabrata. Voriconazole was active against 148 isolates (98.7%) with geometric mean MICs ranging from 0.030 μg/ml for C. parapsilosis, 0.042 μg/ml for C. albicans, 0.048 μg/ml for C. tropicalis, 0.082 μg/ml for C. guilliermondii, to 0.137 μg/ml for C. glabrata. Only 2 C. albicans (2%) were resistant to voriconazole showing cross-resistance to other azoles. Posaconazole and voriconazole have excellent in vitro activities against all Candida isolates and could represent useful alternatives for recalcitrant or recurrent candidiasis.

  5. Seminal plasma proteins inhibit in vitro- and cooling-induced capacitation in boar spermatozoa.

    PubMed

    Vadnais, Melissa L; Roberts, Kenneth P

    2010-01-01

    Dilute boar seminal plasma (SP) has been shown to inhibit in vitro capacitation and cooling-induced capacitation-like changes in boar spermatozoa, as assessed by the ability of the spermatozoa to undergo an ionophore-induced acrosome reaction. We hypothesised that the protein component of SP is responsible for this effect. To test this hypothesis, varying concentrations of total SP protein or SP proteins fractionated by heparin binding were assayed for their ability to inhibit in vitro capacitation, as well as cooling- and cryopreservation-induced capacitation-like changes. In vitro capacitation and cooling-induced capacitation-like changes were prevented by 10% whole SP, as well as by total proteins extracted from SP at concentrations greater than 500 microg mL(-1). No amount of SP protein was able to prevent cryopreservation-induced capacitation-like changes. Total SP proteins were fractionated based on their heparin-binding properties and the heparin-binding fraction was shown to possess capacitation inhibitory activity at concentrations as low as 250 microg mL(-1). The proteins in the heparin-binding fraction were subjected to mass spectrometry and identified. The predominant proteins were three members of the spermadhesin families, namely AQN-3, AQN-1 and AWN, and SP protein pB1. We conclude that one or more of these heparin-binding SP proteins is able to inhibit in vitro capacitation and cooling-induced capacitation-like changes, but not cryopreservation-induced capacitation-like changes, in boar spermatozoa. PMID:20591323

  6. A Trypsin Inhibitor from Tecoma stans Leaves Inhibits Growth and Promotes ATP Depletion and Lipid Peroxidation in Candida albicans and Candida krusei

    PubMed Central

    Patriota, Leydianne L. S.; Procópio, Thamara F.; de Souza, Maria F. D.; de Oliveira, Ana Patrícia S.; Carvalho, Lidiane V. N.; Pitta, Maira G. R.; Rego, Moacyr J. B. M.; Paiva, Patrícia M. G.; Pontual, Emmanuel V.; Napoleão, Thiago H.

    2016-01-01

    Tecoma stans (yellow elder) has shown medicinal properties and antimicrobial activity. Previous reports on antifungal activity of T. stans preparations and presence of trypsin inhibitor activity from T. stans leaves stimulated the investigation reported here. In this work, we proceeded to the purification and characterization of a trypsin inhibitor (TesTI), which was investigated for anti-Candida activity. Finally, in order to determine the potential of TesTI as a new natural chemotherapeutic product, its cytotoxicity to human peripheral blood mononuclear cells (PBMCs) was evaluated. TesTI was isolated from saline extract by ammonium sulfate fractionation followed by ion exchange and gel filtration chromatographies. Antifungal activity was evaluated by determining the minimal inhibitory (MIC) and fungicide (MFC) concentrations using fungal cultures containing only yeast form or both yeast and hyphal forms. Candida cells treated with TesTI were evaluated for intracellular ATP levels and lipid peroxidation. Cytotoxicity of TesTI to PBMCs was evaluated by MTT assay. TesTI (39.8 kDa, pI 3.41, Ki 43 nM) inhibited similarly the growth of both C. albicans and C. krusei culture types at MIC of 100 μg/mL. The MFCs were 200 μg/mL for C. albicans and C. krusei. Time-response curves revealed that TesTI (at MIC) was more effective at inhibiting the replication of C. albicans cells. At MIC, TesTI promoted reduction of ATP levels and lipid peroxidation in the Candida cells, being not cytotoxic to PBMCs. In conclusion, TesTI is an antifungal agent against C. albicans and C. krusei, without toxicity to human cells. PMID:27199940

  7. A Trypsin Inhibitor from Tecoma stans Leaves Inhibits Growth and Promotes ATP Depletion and Lipid Peroxidation in Candida albicans and Candida krusei.

    PubMed

    Patriota, Leydianne L S; Procópio, Thamara F; de Souza, Maria F D; de Oliveira, Ana Patrícia S; Carvalho, Lidiane V N; Pitta, Maira G R; Rego, Moacyr J B M; Paiva, Patrícia M G; Pontual, Emmanuel V; Napoleão, Thiago H

    2016-01-01

    Tecoma stans (yellow elder) has shown medicinal properties and antimicrobial activity. Previous reports on antifungal activity of T. stans preparations and presence of trypsin inhibitor activity from T. stans leaves stimulated the investigation reported here. In this work, we proceeded to the purification and characterization of a trypsin inhibitor (TesTI), which was investigated for anti-Candida activity. Finally, in order to determine the potential of TesTI as a new natural chemotherapeutic product, its cytotoxicity to human peripheral blood mononuclear cells (PBMCs) was evaluated. TesTI was isolated from saline extract by ammonium sulfate fractionation followed by ion exchange and gel filtration chromatographies. Antifungal activity was evaluated by determining the minimal inhibitory (MIC) and fungicide (MFC) concentrations using fungal cultures containing only yeast form or both yeast and hyphal forms. Candida cells treated with TesTI were evaluated for intracellular ATP levels and lipid peroxidation. Cytotoxicity of TesTI to PBMCs was evaluated by MTT assay. TesTI (39.8 kDa, pI 3.41, K i 43 nM) inhibited similarly the growth of both C. albicans and C. krusei culture types at MIC of 100 μg/mL. The MFCs were 200 μg/mL for C. albicans and C. krusei. Time-response curves revealed that TesTI (at MIC) was more effective at inhibiting the replication of C. albicans cells. At MIC, TesTI promoted reduction of ATP levels and lipid peroxidation in the Candida cells, being not cytotoxic to PBMCs. In conclusion, TesTI is an antifungal agent against C. albicans and C. krusei, without toxicity to human cells. PMID:27199940

  8. In Vitro Inhibition of Hepatitis C Virus by Antisense Oligonucleotides in PBMC Compared to Hepatoma Cells

    PubMed Central

    Youssef, Samar Samir; Fahmy, Ahmed Mohamed; Omran, Moataza Hassan; Mohamed, Amr Saad; El Desouki, Mohamed Ali; El-Awady, Mostafa K.

    2014-01-01

    Aim. To assess the efficiency of phosphorothioate antisense oligodeoxynucleotide 1 (S-ODN1) on HCV translation inhibition in PBMC compared to hepatoma cells in vitro for the first time. Materials and Methods. The study included 34 treatment naive HCV patients. IRES domain III and IV sequence variations were tested in 45 clones from 9 HCV patients. PBMC of HCV positive patients were subjected to S-ODN in vitro. Concomitantly HepG2 cells infected by the same patient's serum were also treated with S-ODN1 for 24 and 48 hours. Cellular RNA was tested for HCV plus and minus strands by reverse transcription polymerase chain reaction (RT-PCR). Results. Sequence variations were seen in HCV IRES domain III only while domain IV was conserved among all the tested patient's clones. S-ODN1 successfully inhibited HCV translation in HepG2 cells, while in PBMC inhibition was partial. Conclusion. HCV IRES domain IV is more conserved than domain IIId in genotype 4 HCV patients. S-ODN against HCV IRES domain IV was not efficient to inhibit HCV translation in PBMC under the study conditions. Further studies testing other S-ODN targeting other HCV IRES domains in PBMC should be done. PMID:24991538

  9. Potassium humate inhibits complement activation and the production of inflammatory cytokines in vitro

    SciTech Connect

    van Rensburg, C.E.J.; Naude, P.J.

    2009-08-15

    The effects of brown coal derived potassium humate on lymphocyte proliferation, cytokine production and complement activation were investigated in vitro. Potassium humate increased lymphocyte proliferation of phytohaemaglutinin A (PHA) and pokeweed mitogen (PWM) stimulated mononuclear lymphocytes (MNL) in vitro from concentrations of 20 to 80 {mu} g/ml, in a dose dependant manner. On the other hand potassium humate, at 40 {mu} g/ml, significantly inhibited the release of TNF-alpha, IL-1 beta, IL-6 and IL-10 by PHA stimulated MNL. Regarding complement activation it was found that potassium humate inhibits the activation of both the alternative and classical pathways without affecting the stability of the red blood cell membranes. These results indicate that the anti-inflammatory potential of potassium humate could be partially due to the inhibition of pro-inflammatory cytokines responsible for the initiation of these reactions as well as inhibition of complement activation. The increased lymphocyte proliferation observed, might be due to increased IL-2 production as previously been documented.

  10. A new small molecule inhibits streptococcus mutans biofilms in vitro and in vivo

    PubMed Central

    Pan, Wenting; Fan, Mingwen; Wu, Hui; Melander, Christian; Liu, Chang

    2015-01-01

    Aims The aim of this study is to identify new small molecules that can inhibit Streptococcus mutans biofilms by in-vitro and in-vivo model. Methods and Results We evaluated the effect of a small molecule 2-amino-imidazole/triazole conjugate (2-AI/T) on the formation of S. mutans biofilms by culturing in 96-well plates. Toxicity was assessed through cell culture and intragastrically administering to mice. The anti-biofilm and anti-caries effects were investigated in vivo. The inhibitive mechanism was detected by isobaric tag for relative and absolute quantitation (itraq) and RT-QPCR. In vitro and in vivo study revealed that 2-AI/T significantly inhibited biofilm formation of S. mutans and is more so than inhibiting planktonic cells without toxicity. The ribosome and histidine metabolism pathways of S. mutans were significantly regulated by this compound. Conclusions These results suggest that the 2-AI/T conjugate is a potent inhibitor that can be potentially developed into a new drug to treat and prevent dental caries. Significance and Impact of the Study This is the first study to use small molecule from marine natural products, to protect from dental cariesin vivo. It has potential broad range application in clinical caries prevention, or as a bioactive ingredient for food applications. PMID:26294263

  11. Inhibition of influenza A virus infection in vitro by saliphenylhalamide-loaded porous silicon nanoparticles.

    PubMed

    Bimbo, Luis M; Denisova, Oxana V; Mäkilä, Ermei; Kaasalainen, Martti; De Brabander, Jef K; Hirvonen, Jouni; Salonen, Jarno; Kakkola, Laura; Kainov, Denis; Santos, Hélder A

    2013-08-27

    Influenza A viruses (IAVs) cause recurrent epidemics in humans, with serious threat of lethal worldwide pandemics. The occurrence of antiviral-resistant virus strains and the emergence of highly pathogenic influenza viruses have triggered an urgent need to develop new anti-IAV treatments. One compound found to inhibit IAV, and other virus infections, is saliphenylhalamide (SaliPhe). SaliPhe targets host vacuolar-ATPase and inhibits acidification of endosomes, a process needed for productive virus infection. The major obstacle for the further development of SaliPhe as antiviral drug has been its poor solubility. Here, we investigated the possibility to increase SaliPhe solubility by loading the compound in thermally hydrocarbonized porous silicon (THCPSi) nanoparticles. SaliPhe-loaded nanoparticles were further investigated for the ability to inhibit influenza A infection in human retinal pigment epithelium and Madin-Darby canine kidney cells, and we show that upon release from THCPSi, SaliPhe inhibited IAV infection in vitro and reduced the amount of progeny virus in IAV-infected cells. Overall, the PSi-based nanosystem exhibited increased dissolution of the investigated anti-IAV drug SaliPhe and displayed excellent in vitro stability, low cytotoxicity, and remarkable reduction of viral load in the absence of organic solvents. This proof-of-principle study indicates that PSi nanoparticles could be used for efficient delivery of antivirals to infected cells.

  12. Inhibition of influenza A virus infection in vitro by saliphenylhalamide-loaded porous silicon nanoparticles.

    PubMed

    Bimbo, Luis M; Denisova, Oxana V; Mäkilä, Ermei; Kaasalainen, Martti; De Brabander, Jef K; Hirvonen, Jouni; Salonen, Jarno; Kakkola, Laura; Kainov, Denis; Santos, Hélder A

    2013-08-27

    Influenza A viruses (IAVs) cause recurrent epidemics in humans, with serious threat of lethal worldwide pandemics. The occurrence of antiviral-resistant virus strains and the emergence of highly pathogenic influenza viruses have triggered an urgent need to develop new anti-IAV treatments. One compound found to inhibit IAV, and other virus infections, is saliphenylhalamide (SaliPhe). SaliPhe targets host vacuolar-ATPase and inhibits acidification of endosomes, a process needed for productive virus infection. The major obstacle for the further development of SaliPhe as antiviral drug has been its poor solubility. Here, we investigated the possibility to increase SaliPhe solubility by loading the compound in thermally hydrocarbonized porous silicon (THCPSi) nanoparticles. SaliPhe-loaded nanoparticles were further investigated for the ability to inhibit influenza A infection in human retinal pigment epithelium and Madin-Darby canine kidney cells, and we show that upon release from THCPSi, SaliPhe inhibited IAV infection in vitro and reduced the amount of progeny virus in IAV-infected cells. Overall, the PSi-based nanosystem exhibited increased dissolution of the investigated anti-IAV drug SaliPhe and displayed excellent in vitro stability, low cytotoxicity, and remarkable reduction of viral load in the absence of organic solvents. This proof-of-principle study indicates that PSi nanoparticles could be used for efficient delivery of antivirals to infected cells. PMID:23889734

  13. Ginger's (Zingiber officinale Roscoe) inhibition of rat colonic adenocarcinoma cells proliferation and angiogenesis in vitro.

    PubMed

    Brown, Amy C; Shah, Chirag; Liu, Jessie; Pham, Jimmy T H; Zhang, Jian Gang; Jadus, Martin R

    2009-05-01

    Ginger's (Zingiber officinale Roscoe) natural bioactives, specifically ginger extract and 6-gingerol, were measured for their in vitro inhibition of two key aspects of colon cancer biology--cancer cell proliferation and angiogenic potential of endothelial cell tubule formation. Ginger extract was obtained via column distillation, while the 6-gingerol was purchased from Calbiochem. Antiproliferation activity was assessed through tritiated thymidine ([(3)H]Tdr) incorporation studies of YYT colon cancer cells; the anti-angiogenic ability of gingerol was assessed by a Matrigel assays using MS1 endothelial cells. These selected ginger bioactives had: 1) a direct effect on YYT rat cancer cell proliferation (6-1.5% ginger extract; 100-4 microM 6-gingerol); 2) an indirect effect on MS1 endothelial cell function either at the level of endothelial cell proliferation or through inhibition of MS1 endothelial cell tube formation (100-0.8 microM). Compound 6-gingerol was most effective at lower doses in inhibiting endothelial cell tube formation. These in vitro studies show that 6-gingerol has two types of antitumor effects: 1) direct colon cancer cell growth suppression, and 2) inhibition of the blood supply of the tumor via angiogenesis. Further research is warranted to test 6-gingerol in animal studies as a potential anticancer plant bioactive in the complementary treatment of cancer. PMID:19117330

  14. Inhibitory effect of ebselen on cerebral acetylcholinesterase activity in vitro: kinetics and reversibility of inhibition.

    PubMed

    Martini, Franciele; Bruning, César Augusto; Soares, Suelen Mendonca; Nogueira, Cristina Wayne; Zeni, Gilson

    2015-01-01

    Ebselen is a synthetic organoselenium compound that has been considered a potential pharmacological agent with low toxicity, showing antioxidant, anti-inflammatory and neuroprotective effects. It is bioavailable, blood-brain barrier permeant and safe based on cellular toxicity and Phase I-III clinical trials. There is evidence that ebselen inhibits acetylcholinesterase (AChE) activity, an enzyme that plays a key role in the cholinergic system by hydrolyzing acetylcholine (ACh), in vitro and ex vivo. This system has a well-known relationship with cognitive process, and AChE inhibitors, such as donepezil and galantamine, have been used to treat cognitive deficits, mainly in the Alzheimer's Disease (AD). However, these drugs have poor bioavailability and a number of side effects, including gastrointestinal upsets and hepatotoxicity. In this way, this study aimed to evaluate the effect of ebselen on cerebral AChE activity in vitro and to determine the kinetic profile and the reversibility of inhibition by dialysis. Ebselen inhibited the cerebral AChE activity with an IC50 of 29 µM, similar to IC50 found with pure AChE from electric eel, demonstrating a mixed and reversible inhibition of AChE, since it increased Km and decreased Vmax. The AChE activity was recovered within 60 min of dialysis. Therefore, the use of ebselen as a therapeutic agent for treatment of AD should be considered, although memory behavior tasks are needed to support such hypothesis. PMID:25312723

  15. Inhibition of Candida albicans Biofilm Formation by Farnesol, a Quorum-Sensing Molecule

    PubMed Central

    Ramage, Gordon; Saville, Stephen P.; Wickes, Brian L.; López-Ribot, José L.

    2002-01-01

    Farnesol is a quorum-sensing molecule that inhibits filamentation in Candida albicans. Both filamentation and quorum sensing are deemed to be important factors in C. albicans biofilm development. Here we examined the effect of farnesol on C. albicans biofilm formation. C. albicans adherent cell populations (after 0, 1, 2, and 4 h of adherence) and preformed biofilms (24 h) were treated with various concentrations of farnesol (0, 3, 30, and 300 μM) and incubated at 37°C for 24 h. The extent and characteristics of biofilm formation were then assessed microscopically and with a semiquantitative colorimetric technique based on the use of 2,3-bis(2-methoxy-4-nitro-5-sulfo-phenyl)-2H-tetrazolium-5-carboxanilide. The results indicated that the effect of farnesol was dependent on the concentration of this compound and the initial adherence time, and preincubation with 300 μM farnesol completely inhibited biofilm formation. Supernatant media recovered from mature biofilms inhibited the ability of planktonic C. albicans to form filaments, indicating that a morphogenetic autoregulatory compound is produced in situ in biofilms. Northern blot analysis of RNA extracted from cells in biofilms indicated that the levels of expression of HWP1, encoding a hypha-specific wall protein, were decreased in farnesol-treated biofilms compared to the levels in controls. Our results indicate that farnesol acts as a naturally occurring quorum-sensing molecule which inhibits biofilm formation, and we discuss its potential for further development and use as a novel therapeutic agent. PMID:12406738

  16. Bismuth oxide aqueous colloidal nanoparticles inhibit Candida albicans growth and biofilm formation

    PubMed Central

    Hernandez-Delgadillo, Rene; Velasco-Arias, Donaji; Martinez-Sanmiguel, Juan Jose; Diaz, David; Zumeta-Dube, Inti; Arevalo-Niño, Katiushka; Cabral-Romero, Claudio

    2013-01-01

    Multiresistance among microorganisms to common antimicrobials has become one of the most significant concerns in modern medicine. Nanomaterials are a new alternative to successfully treat the multiresistant microorganisms. Nanostructured materials are used in many fields, including biological sciences and medicine. Recently, it was demonstrated that the bactericidal activity of zero-valent bismuth colloidal nanoparticles inhibited the growth of Streptococcus mutans; however the antimycotic potential of bismuth nanostructured derivatives has not yet been studied. The main objective of this investigation was to analyze the fungicidal activity of bismuth oxide nanoparticles against Candida albicans, and their antibiofilm capabilities. Our results showed that aqueous colloidal bismuth oxide nanoparticles displayed antimicrobial activity against C. albicans growth (reducing colony size by 85%) and a complete inhibition of biofilm formation. These results are better than those obtained with chlorhexidine, nystatin, and terbinafine, the most effective oral antiseptic and commercial antifungal agents. In this work, we also compared the antimycotic activities of bulk bismuth oxide and bismuth nitrate, the precursor metallic salt. These results suggest that bismuth oxide colloidal nanoparticles could be a very interesting candidate as a fungicidal agent to be incorporated into an oral antiseptic. Additionally, we determined the minimum inhibitory concentration for the synthesized aqueous colloidal Bi2O3 nanoparticles. PMID:23637533

  17. Terpenoids of plant origin inhibit morphogenesis, adhesion, and biofilm formation by Candida albicans.

    PubMed

    Raut, Jayant S; Shinde, Ravikumar B; Chauhan, Nitin M; Karuppayil, S Mohan

    2013-01-01

    Biofilm-related infections caused by Candida albicans and associated drug resistant micro-organisms are serious problems for immunocompromised populations. Molecules which can prevent or remove biofilms are needed. Twenty-eight terpenoids of plant origin were analysed for their activity against growth, virulence attributes, and biofilms of C. albicans. Eighteen molecules exhibited minimum inhibitory concentrations of <2 mg ml(-1) for planktonic growth. Selected molecules inhibited yeast to hyphal dimorphism at low concentrations (0.031-0.5 mg ml(-1)), while adhesion to a solid surface was prevented at 0.5-2 mg ml(-1). Treatment with 14 terpenoids resulted in significant (p < 0.05) inhibition of biofilm formation, and of these, linalool, nerol, isopulegol, menthol, carvone, α-thujone, and farnesol exhibited biofilm-specific activity. Eight terpenoids were identified as inhibitors of mature biofilms. This study demonstrated the antibiofilm potential of terpenoids, which need to be further explored as therapeutic strategy against biofilm associated infections of C. albicans. PMID:23216018

  18. Coptis chinensis and Myrobalan (Terminalia chebula) Can Synergistically Inhibit Inflammatory Response In Vitro and In Vivo

    PubMed Central

    Cui, Enhui; Zhi, Xiaoyan; Chen, Ying; Gao, Yuanyuan; Fan, Yunpeng; Zhang, Weimin; Ma, Wuren; Hou, Weifeng; Guo, Chao; Song, Xiaoping

    2014-01-01

    Objectives. To investigate the anti-inflammatory effect of Coptis chinensis plus myrobalan (CM) in vitro and in vivo. Methods. The inflammation in mouse peritoneal macrophages was induced by lipopolysaccharide (LPS). Animal models were established by using ear swelling and paw edema of mouse induced by xylene and formaldehyde, respectively. In vitro, cytotoxicity, the phagocytosis of macrophages, the levels of nitric oxide (NO), induced nitric oxide synthase (iNOS), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) in cell supernatant were detected. In vivo, swelling rate and edema inhibitory rate of ear and paw were observed using CM-treated mice. Results. At 150–18.75 μg·mL−1, CM had no cytotoxicity and could significantly promote the growth and the phagocytosis of macrophages and inhibit the overproduction of NO, iNOS, TNF-α, and IL-6 in macrophages induced by LPS. In vivo, pretreatment with CM, the ear swelling, and paw edema of mice could be significantly inhibited in a dose-dependent manner, and the antiedema effect of CM at high dose was better than dexamethasone. Conclusion. Our results demonstrated that Coptis chinensis and myrobalan possessed synergistically anti-inflammatory activities in vitro and in vivo, which indicated that CM had therapeutic potential for the prevention and treatment of inflammation-mediated diseases. PMID:25587343

  19. Noscapine inhibits human hepatocellular carcinoma growth through inducing apoptosis in vitro and in vivo.

    PubMed

    Xu, G; Niu, Z; Dong, J; Zhao, Y; Zhang, Y; Li, X

    2016-01-01

    Noscapine, a phthalideisoquinoline alkaloid derived from opium, has been demonstrated as a promising anti-tumor compound against various cancers. However, the anti-cancer activity of noscapine in hepatocellular carcinoma has not been defined. In this study, we investigate the inhibitive effects of noscapine on human hepatocellular carcinoma (HCC) using both in vitro and in vivo models. In vitro proliferation assay showed that noscapine suppressed HepG2 and Huh7 cells in dose- and time-dependent manners. With a mouse xenograft model, noscapine showed notable inhibition on HCC tumor growth in vivo without suppression of body weight. Moreover, apoptotic induction and regulation of related signalings by noscapine were examined by nuclear DNA staining, TUNEL, and western blotting assays. Results showed that noscapine induced apoptosis in HCC cells both in vitro and in vivo. Further studies indicated that noscapine induced antive-capsase-3, cleavage PARP, and decreased the ratio of Bcl-2/Bax. Hence, these data indicates that noscapine selectively suppresses HCC cell growth through apoptosis induction, providing evidence for application of noscapine as a novel agent against human hepatocellular carcinoma. PMID:27468876

  20. Berry polyphenols inhibit α-amylase in vitro: identifying active components in rowanberry and raspberry.

    PubMed

    Grussu, Dominic; Stewart, Derek; McDougall, Gordon J

    2011-03-23

    Polyphenol-rich extracts from a range of berries inhibited α-amylase in vitro, but the most effective were from raspberry and rowanberry (IC50 values of 21.0 and 4.5 μg/mL, respectively). The inhibitory components were examined by different approaches. Extracts from yellow and red raspberries were equally able to inhibit α-amylase. Because the yellow raspberry extracts effectively lacked anthocyanins, this suggested that they were not crucial for amylase inhibition. Notably, however, higher levels of other phenolic components in yellow raspberries (particularly, ellagitannins) did not increase amylase inhibition. Amylase inhibition in rowanberry was recovered in a fraction enriched in proanthocyanidins (PACs). Inhibition was ameliorated by bovine serum albumin, suggesting that PACs acted by binding to amylase. Co-incubation of rowanberry PACs with acarbose reduced the concentration of acarbose required for effective amylase inhibition. Such synergistic interactions could have implications for the current clinical use of acarbose for postprandial glycaemic control in type-2 diabetics.

  1. Correlation between in vivo and in vitro studies of modulation of resistance to experimental Candida albicans infection by cyclophosphamide in mice.

    PubMed Central

    Bistoni, F; Baccarini, M; Blasi, E; Marconi, P; Puccetti, P; Garaci, E

    1983-01-01

    Mice receiving a single injection of cyclophosphamide (150 mg/kg) 1 to 6 days before inoculation with viable Candida albicans showed an increased susceptibility to the challenge accompanied by a reduction in peripheral blood polymorphonuclear leukocytes and lymphocytes as well as in spleen cellularity. Several immunological in vitro functions also appeared to be dramatically depressed. Most of these hematological and functional parameters returned to control values by day 9 after cyclophosphamide administration, at a time when resistance to C. albicans infection appeared to be unchanged. However, when exposure to cyclophosphamide occurred 12 to 21 days before inoculation with the live yeast, enhanced resistance was observed with the majority of the animals surviving challenge. To gain some insight into the mechanisms underlying this late increase in resistance to C. albicans infection after cyclophosphamide administration, we analyzed a series of immunological functions, including the in vitro candidacidal activity of polymorphonuclear neutrophils and plastic-adherent and nonadherent spleen cells as well as the activity of natural killer cells and alloreactive T lymphocytes. The results show that a numerical rebound of blood polymorphonuclear neutrophils and the appearance of a highly candidacidal cell population in the spleen may be among the factors underlying the late increase in resistance to C. albicans after administration of cyclophosphamide. PMID:6339410

  2. A human serum mannose-binding protein inhibits in vitro infection by the human immunodeficiency virus

    PubMed Central

    1989-01-01

    In vitro infection by the human immunodeficiency virus (HIV) of CD4+ H9 lymphoblasts is inhibited by a mannose-binding protein (MBP) purified from human serum. In addition, MBP is able to selectively bind to HIV- infected H9 cells and HIV-infected cells from the monocyte cell line U937. These results indicate MBP most likely recognizes high mannose glycans known to be present on gp120 in the domain that is recognized by CD4 and thereby inhibits viral entry to susceptible cells. In support of this contention, recombinant gp120 binds directly to MBP; the binding is saturable, mannan inhibitable, removed by N-glycanase treatment, and dependent on divalent cations. PMID:2909656

  3. Herbicide clomazone does not inhibit in vitro geranylgeranyl synthesis from mevalonate.

    PubMed

    Weimer, M R; Balke, N E; Buhler, D D

    1992-02-01

    Clomazone reduced the chlorophyll and carotenoid contents of spinach (Spinacia oleracea L.), barley (Hordeum vulgare L.), velvetleaf (Abutilon theophrasti Medik.), and soybean (Glycine max L. Merr.) seedlings. The order of species sensitivity was velvetleaf > spinach > barley > soybean. Clomazone (100 micromolar) did not affect the in vitro activities of spinach isopentenyl pyrophosphate isomerase or prenyl transferase. Clomazone also did not affect the synthesis of isopentenyl pyrophosphate from mevalonic acid. Thus, clomazone had no direct in vitro effect on the synthesis of geranylgeranyl pyrophosphate from mevalonic acid. Greening seedlings of both soybean and velvetleaf metabolized clomazone. No qualitative differences in the metabolites were detected between soybean and velvetleaf. Thus, differential metabolism of clomazone to a toxic chemical that inhibits terpenoid synthesis is unlikely. Clomazone has either a mode of action not yet identified or a metabolite that is selective in that it is much more active in sensitive than tolerant species.

  4. Herbicide Clomazone Does Not Inhibit In Vitro Geranylgeranyl Synthesis from Mevalonate 1

    PubMed Central

    Weimer, Monte R.; Balke, Nelson E.; Buhler, Douglas D.

    1992-01-01

    Clomazone reduced the chlorophyll and carotenoid contents of spinach (Spinacia oleracea L.), barley (Hordeum vulgare L.), velvetleaf (Abutilon theophrasti Medik.), and soybean (Glycine max L. Merr.) seedlings. The order of species sensitivity was velvetleaf > spinach > barley > soybean. Clomazone (100 micromolar) did not affect the in vitro activities of spinach isopentenyl pyrophosphate isomerase or prenyl transferase. Clomazone also did not affect the synthesis of isopentenyl pyrophosphate from mevalonic acid. Thus, clomazone had no direct in vitro effect on the synthesis of geranylgeranyl pyrophosphate from mevalonic acid. Greening seedlings of both soybean and velvetleaf metabolized clomazone. No qualitative differences in the metabolites were detected between soybean and velvetleaf. Thus, differential metabolism of clomazone to a toxic chemical that inhibits terpenoid synthesis is unlikely. Clomazone has either a mode of action not yet identified or a metabolite that is selective in that it is much more active in sensitive than tolerant species. PMID:16668657

  5. In vitro inhibition of murine macrophage migration by Bordetella pertussis lymphocytosis-promoting factor.

    PubMed Central

    Meade, B D; Kind, P D; Ewell, J B; McGrath, P P; Manclark, C R

    1984-01-01

    Lymphocytosis promoting factor (LPF) of Bordetella pertussis is a protein toxin which may have a role in the pathogenesis of pertussis. Since macrophages have an important role in the control of respiratory infections, the in vitro effects of LPF on macrophages from C3H/HeN and C3H/HeJ mice and on a murine macrophage-like cell line, RAW264, were examined. LPF inhibited random migration of resident peritoneal macrophages as well as the chemotaxis of peritoneal macrophages and the cell line. Fifty percent inhibition of chemotaxis occurred at 0.2 to 0.3 ng of LPF per ml for the macrophages and at 1 to 2 ng of LPF per ml for the cell line. When LPF was either heated at 80 degrees C for 5 min or premixed with specific antibodies, it failed to inhibit migration. At 20 ng/ml, LPF inhibited chemotaxis by more than 80% and also decreased Fc-mediated phagocytosis by 25 to 35%. At this dose, LPF was not a chemoattractant for murine macrophages and did not reduce macrophage viability, adherence, or opsonized zymosan-stimulated superoxide release. When LPF-treated macrophages were added to tissue culture dishes and then examined microscopically after 4 h, the LPF-treated cells adhered but failed to spread and elongate as well as control macrophages. These data indicate that LPF specifically inhibits macrophage migration in vitro and suggest that a possible role for LPF in pathogenesis is to inhibit migration of macrophages to the site of B. pertussis infection. Images PMID:6088394

  6. Milk thistle extract and silymarin inhibit lipopolysaccharide induced lamellar separation of hoof explants in vitro.

    PubMed

    Reisinger, Nicole; Schaumberger, Simone; Nagl, Veronika; Hessenberger, Sabine; Schatzmayr, Gerd

    2014-10-01

    The pathogenesis of laminitis is not completely identified and the role of endotoxins (lipopolysaccharides, LPS) in this process remains unclear. Phytogenic substances, like milk thistle (MT) and silymarin, are known for their anti-inflammatory and antioxidant properties and might therefore have the potential to counteract endotoxin induced effects on the hoof lamellar tissue. The aim of our study was to investigate the influence of endotoxins on lamellar tissue integrity and to test if MT and silymarin are capable of inhibiting LPS-induced effects in an in vitro/ex vivo model. In preliminary tests, LPS neutralization efficiency of these phytogenics was determined in an in vitro neutralization assay. Furthermore, tissue explants gained from hooves of slaughter horses were tested for lamellar separation after incubation with different concentrations of LPS. By combined incubation of explants with LPS and either Polymyxin B (PMB; positive control), MT or silymarin, the influence of these substances on LPS-induced effects was assessed. In the in vitro neutralization assay, MT and silymarin reduced LPS concentrations by 64% and 75%, respectively, in comparison PMB reduced 98% of the LPS concentration. In hoof explants, LPS led to a concentration dependent separation. Accordantly, separation force was significantly decreased by 10 µg/mL LPS. PMB, MT and silymarin could significantly improve tissue integrity of explants incubated with 10 µg/mL LPS. This study showed that LPS had a negative influence on the structure of hoof explants in vitro. MT and silymarin reduced endotoxin activity and inhibited LPS-induced effects on the lamellar tissue. Hence, MT and silymarin might be used to support the prevention of laminitis and should be further evaluated for this application.

  7. Milk thistle extract and silymarin inhibit lipopolysaccharide induced lamellar separation of hoof explants in vitro.

    PubMed

    Reisinger, Nicole; Schaumberger, Simone; Nagl, Veronika; Hessenberger, Sabine; Schatzmayr, Gerd

    2014-10-01

    The pathogenesis of laminitis is not completely identified and the role of endotoxins (lipopolysaccharides, LPS) in this process remains unclear. Phytogenic substances, like milk thistle (MT) and silymarin, are known for their anti-inflammatory and antioxidant properties and might therefore have the potential to counteract endotoxin induced effects on the hoof lamellar tissue. The aim of our study was to investigate the influence of endotoxins on lamellar tissue integrity and to test if MT and silymarin are capable of inhibiting LPS-induced effects in an in vitro/ex vivo model. In preliminary tests, LPS neutralization efficiency of these phytogenics was determined in an in vitro neutralization assay. Furthermore, tissue explants gained from hooves of slaughter horses were tested for lamellar separation after incubation with different concentrations of LPS. By combined incubation of explants with LPS and either Polymyxin B (PMB; positive control), MT or silymarin, the influence of these substances on LPS-induced effects was assessed. In the in vitro neutralization assay, MT and silymarin reduced LPS concentrations by 64% and 75%, respectively, in comparison PMB reduced 98% of the LPS concentration. In hoof explants, LPS led to a concentration dependent separation. Accordantly, separation force was significantly decreased by 10 µg/mL LPS. PMB, MT and silymarin could significantly improve tissue integrity of explants incubated with 10 µg/mL LPS. This study showed that LPS had a negative influence on the structure of hoof explants in vitro. MT and silymarin reduced endotoxin activity and inhibited LPS-induced effects on the lamellar tissue. Hence, MT and silymarin might be used to support the prevention of laminitis and should be further evaluated for this application. PMID:25290524

  8. Acetate-mediated growth inhibition in sterol 14alpha-demethylation-deficient cells of Candida albicans.

    PubMed

    Shimokawa, O; Nakayama, H

    1999-01-01

    Candida albicans is a fungus thought to be viable in the presence of a deficiency in sterol 14alpha-demethylation. We showed in a strain of this species that the deficiency, caused either by a mutation or by an azole antifungal agent, made the cells susceptible to growth inhibition by acetate included in the culture medium. Studies with a mutant demonstrated that the inhibition was complete at a sodium acetate concentration of 0.24 M (20 g/liter) and was evident even at a pH of 8, the latter result indicating the involvement of acetate ions rather than the undissociated form of acetic acid. In fluconazole-treated cells, sterol profiles determined by thin-layer chromatography revealed that the minimum sterol 14alpha-demethylation-inhibitory concentrations (MDICs) of the drug, thought to be the most important parameter for clinical purposes, were practically identical in the media with and without 0.24 M acetate and were equivalent to the MIC in the acetate-supplemented medium. The acetate-mediated growth inhibition of azole-treated cells was confirmed with two additional strains of C. albicans and four different agents, suggesting the possibility of generalization. From these results, it was surmised that the acetate-containing medium may find use in azole susceptibility testing, for which there is currently no method capable of measuring MDICs directly for those fungi whose viability is not lost as a result of sterol 14alpha-demethylation deficiency. Additionally, the acetate-supplemented agar medium was found to be useful in detecting reversions from sterol 14alpha-demethylation deficiency to proficiency. PMID:9869573

  9. β-Asarone, an active principle of Acorus calamus rhizome, inhibits morphogenesis, biofilm formation and ergosterol biosynthesis in Candida albicans.

    PubMed

    Rajput, Sandeep B; Karuppayil, S Mohan

    2013-01-15

    Anti-Candida potential of Acorus calamus rhizome and its active principle, β-asarone, was evaluated against the human fungal pathogen, Candida albicans. β-Asarone exhibited promising growth inhibitory activity at 0.5mg/ml and it was fungicidal at 8 mg/ml. Time dependant kill curve assay showed that MFC of β-asarone was highly toxic to C. albicans, killing 99.9% inoculum within 120 min of exposure. β-Asarone caused significant inhibition of C. albicans morphogenesis and biofilm development at sub-inhibitory concentrations. Our data indicate that the growth inhibitory activity of β-asarone might be through inhibition of ergosterol biosynthesis. Hemolytic assay showed that β-asarone is non-toxic, even at concentrations approaching MIC value. Our results suggest that β-asarone may be safe as a topical antifungal agent. PMID:23123225

  10. Lithocarpus polystachyus REHD leaf aqueous extract inhibits human breast cancer growth in vitro and in vivo.

    PubMed

    Lin, Caiyu; Wang, Li; Wang, Hong; Fang, Shengtao; Zhang, Quanbo; Yang, Liuqi; Guo, Huijie; Lin, Ping; Zhang, Jie; Wang, Xiujie

    2014-01-01

    Lithocarpus polystachyus leaves have been used as tea beverage and folk medicine for healthy care in the Southwest of China. The purpose of this study is to investigate the anticancer activity of Lithocarpus polystachyus Rehd leaf aqueous extract (LPAE) and to explore the possible mechanism of its activity. Growth inhibition effects of LPAE breast cancer were tested in vitro and in vivo. The possible mechanism of its activity was analyzed with cell biological and molecular biological assays. After LPAE treatment, the proliferation and colony formation of cancer cells decreased; apoptotic cells increased; DNA fragmentations were evident; mRNA and protein expressions of PPARγ, Bax, and caspase-3 genes increased and expressions of cyclin D1 and Bcl-2 genes decreased; in vivo experiment, LPAE inhibited human beast cancer growth. The findings in this experimental study suggested that LPAE has potential cytotoxic and apoptotic effects on human breast cancer cells in vitro and inhibits the cancer growth in vivo, and its mechanism of activity might be associated with apoptosis induction of cancer cells through upregulation of the mRNA and protein expressions of PPARγ, Bax, and capase-3 genes and downregulation of the expressions of cyclin D1 and Bcl-2 genes.

  11. Inhibition of RNA synthesis in vitro by acridines--relation between structure and activity.

    PubMed

    Piestrzeniewicz, M K; Wilmańska, D; Studzian, K; Szemraj, J; Czyz, M; Denny, W A; Gniazdowski, M

    1998-01-01

    The effects of acridine derivatives (proflavine and 2,7-dialkyl derivatives, diacridines and triacridines, 9-aminoacridine carboxamides, and 9-anilinoacridine, amsacrine and its congeners) on overall RNA synthesis in vitro, on synthesis of initiating oligonucleotides and the binding of the enzyme to DNA were studied. The primary mechanism of action is related to inhibition of the enzyme binding to DNA. The acridines (intercalating or non-intercalating and bis-intercalating ligands) assayed here differ in the properties of their complexes with DNA. Correlation is generally observed between inhibition of RNA synthesis in vitro and cytotoxicity in cell cultures for di- and triacridines and 9-aminoacridine carboxamide derivatives. No relationship was found between the effect on RNA polymerase system and biological effects for amsacrine and its derivatives in contrast to the other series of acridines studied here. The aniline ring seems to decrease the inhibitory potency of a ligand. The discrepancy between the biological effect and RNA synthesis inhibition may be due to a different mechanism of cytotoxicity action of amsacrine which is a potent topoisomerase II poison. PMID:9679327

  12. In vitro actinomycete biofilm development and inhibition by the polyene antibiotic, nystatin, on IUD copper surfaces.

    PubMed

    Shanmughapriya, Santhanam; Francis, Arumugam Lency; Kavitha, Senthil; Natarajaseenivasan, Kalimuthusamy

    2012-01-01

    The presence of intrauterine contraceptive devices (IUDs) gives a solid surface for attachment and an ideal niche for biofilm to form and flourish. Pelvic actinomycosis is often associated with the use of IUDs. Treatment of IUD-associated pelvic actinomycosis requires the immediate removal of the IUD. Therefore, this article presents in vitro evidence to support the use of novel antibiotics in the treatment of actinomycete biofilms. Twenty one clinical actinomycetes isolates from endocervical swabs of IUD wearers were assessed for their biofilm forming ability. An in vitro biofilm model with three isolates, Streptomyces strain A4, Nocardia strain C15 and Nocardia strain C17 was subjected to treatment with nystatin. Inhibition of biofilm formation by nystatin was found to be concentration dependent, with MBIC50 values in the range 0.08-0.16 mg ml(-1). Furthermore, at a concentration of 0.16 mg ml(-1), nystatin inhibited the twitching motility of the isolates, providing evidence for a possible mechanism of biofilm inhibition.

  13. A natural small molecule voacangine inhibits angiogenesis both in vitro and in vivo

    SciTech Connect

    Kim, Yonghyo; Jung, Hye Jin; Kwon, Ho Jeong

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer Voacangine exhibits potent anti-angiogenic activity both in vitro and in vivo. Black-Right-Pointing-Pointer Voacangine inhibits tumor-induced angiogenesis by suppressing HIF-1{alpha}. Black-Right-Pointing-Pointer Voacangine could be the basis for the development of novel anti-angiogenic agents. -- Abstract: Angiogenesis, the formation of new blood vessels from pre-existing ones, plays a critical role in normal and pathological phenotypes, including solid tumor growth and metastasis. Accordingly, the development of new anti-angiogenic agents is considered an efficient strategy for the treatment of cancer and other human diseases linked with angiogenesis. We have identified voacangine, isolated from Voacanga africana, as a novel anti-angiogenic agent. Voacangine inhibits the proliferation of HUVECs at an IC{sub 50} of 18 {mu}M with no cytotoxic effects. Voacangine significantly suppressed in vitro angiogenesis, such as VEGF-induced tube formation and chemoinvasion. Moreover, the compound inhibits in vivo angiogenesis in the chorioallantoic membrane at non-toxic doses. In addition, voacangine decreased the expression levels of hypoxia inducible factor-1{alpha} and its target gene, VEGF, in a dose-dependent manner. Taken together, these results suggest that the naturally occurring compound, voacangine, is a novel anti-angiogenic compound.

  14. [6]-Gingerol, a pungent ingredient of ginger, inhibits angiogenesis in vitro and in vivo.

    PubMed

    Kim, Eok-Cheon; Min, Jeong-Ki; Kim, Tae-Yoon; Lee, Shin-Jeong; Yang, Hyun-Ok; Han, Sanghwa; Kim, Young-Myeong; Kwon, Young-Guen

    2005-09-23

    [6]-Gingerol, a pungent ingredient of ginger (Zingiber officinale Roscoe, Zingiberaceae), has anti-bacterial, anti-inflammatory, and anti-tumor-promoting activities. Here, we describe its novel anti-angiogenic activity in vitro and in vivo. In vitro, [6]-gingerol inhibited both the VEGF- and bFGF-induced proliferation of human endothelial cells and caused cell cycle arrest in the G1 phase. It also blocked capillary-like tube formation by endothelial cells in response to VEGF, and strongly inhibited sprouting of endothelial cells in the rat aorta and formation of new blood vessel in the mouse cornea in response to VEGF. Moreover, i.p. administration, without reaching tumor cytotoxic blood levels, to mice receiving i.v. injection of B16F10 melanoma cells, reduced the number of lung metastasis, with preservation of apparently healthy behavior. Taken together, these results demonstrate that [6]-gingerol inhibits angiogenesis and may be useful in the treatment of tumors and other angiogenesis-dependent diseases. PMID:16081047

  15. TQ inhibits hepatocellular carcinoma growth in vitro and in vivo via repression of Notch signaling

    PubMed Central

    Ke, Xiquan; Zhao, Yan; Lu, Xinlan; Wang, Zhe; Liu, Yuanyuan; Ren, Mudan; Lu, Guifang; Zhang, Dan; Sun, Zhenguo; Xu, Zhipeng; Song, Jee Hoon; Cheng, Yulan; Meltzer, Stephen J.; He, Shuixiang

    2015-01-01

    Thymoquinone (TQ) has been reported to possess anti-tumor activity in various types of cancer. However, its effects and molecular mechanism of action in hepatocellular carcinoma (HCC) are still not completely understood. We observed that TQ inhibited tumor cell growth in vitro, where treatment with TQ arrested the cell cycle in G1 by upregulating p21 and downregulating cyclinD1 and CDK2 expression; moreover, TQ induced apoptosis by decreasing expression of Bcl-2 and increasing expression of Bax. Simultaneously, TQ demonstrated a suppressive impact on the Notch pathway, where overexpression of NICD1 reversed the inhibitory effect of TQ on cell proliferation, thereby attenuating the repressive effects of TQ on the Notch pathway, cyclinD1, CDK2 and Bcl-2, and also diminishing upregulation of p21 and Bax. In a xenograft model, TQ inhibited HCC growth in nude mice; this inhibitory effect in vivo, as well as of HCC cell growth in vitro, was associated with a discernible decline in NICD1 and Bcl-2 levels and a dramatic rise in p21 expression. In conclusion, TQ inhibits HCC cell growth by inducing cell cycle arrest and apoptosis, achieving these effects by repression of the Notch signaling pathway, suggesting that TQ represents a potential preventive or therapeutic agent in HCC patients. PMID:26416455

  16. Trifluoperazine inhibits the MgATPase activity and in vitro motility of conventional and unconventional myosins.

    PubMed

    Sellers, James R; Wang, Fei; Chantler, Peter D

    2003-01-01

    Trifluoperazine, a calmodulin antagonist, has recently been shown to inhibit the MgATPase activity of scallop myosin in the absence of light chain dissociation (Patel et al. (2000) J Biol Chem 275: 4880-4888). To investigate the generality of this observation and the mechanism by which it occurs, we have examined the ability of trifluoperazine to inhibit the enzymatic properties of other conventional and unconventional myosins. We show that trifluoperazine can inhibit the actin-activated MgATPase activity of rabbit skeletal muscle myosin II heavy meromyosin (HMM), phosphorylated turkey gizzard smooth muscle myosin II HMM, phosphorylated human nonmuscle myosin IIA HMM and myosin V subfragment-1 (S1). In all cases half maximal inhibition occurred at 50-75 microM trifluoperazine while light chains (myosin II) or calmodulin (myosin V) remained associated with the heavy chains. In vitro motility of all myosins tested was completely inhibited by trifluoperazine. Chymotryptic digestion of baculovirus-expressed myosin V HMM possessing only two calmodulin binding sites yielded a minimal motor fragment with no bound calmodulin. The MgATPase of this fragment was inhibited by trifluoperazine over the same range of concentrations as the S1 fragment of myosin.

  17. Inhibition of Breast Cancer Cell Proliferation and In Vitro Tumorigenesis by a New Red Apple Cultivar

    PubMed Central

    Schiavano, Giuditta Fiorella; De Santi, Mauro; Brandi, Giorgio; Fanelli, Mirco; Bucchini, Anahi; Giamperi, Laura; Giomaro, Giovanna

    2015-01-01

    Purpose The aim of this study was to evaluate the antiproliferative activity in breast cancer cells and the inhibition of tumorigenesis in pre-neoplastic cells of a new apple cultivar with reddish pulp, called the Pelingo apple. Methods The antiproliferative activity was evaluated in MCF-7 and MDA-MB-231 human breast cancer cells. The inhibition of tumorigenesis was performed in JB6 promotion-sensitive (P+) cells. Results Results showed that Pelingo apple juice is characterized by a very high polyphenol content and strongly inhibited breast cancer cell proliferation. Its antiproliferative activity was found to be higher than the other five apple juices tested. Pelingo juice induced cell accumulation in the G2/M phase of the cell cycle and autophagy through overexpression of p21, inhibition of extracellular signal-regulated kinases 1/2 (ERK1/2) activity and an increase in lipidated microtubule-associated protein-1 light chain-3 beta (LC3B). Remarkably, Pelingo juice inhibited the 12-o-tetra-decanoyl-phorbol-13-acetate (TPA)-induced tumorigenesis of JB6 P+ cells, suppressing colony formation in semi-solid medium and TPA-induced ERK1/2 phosphorylation. Conclusions Our data indicate that the Pelingo apple is rich in food components that can markedly inhibit in vitro tumorigenesis and growth of human breast cancer cells and could provide natural bioactive non-nutrient compounds, with potential chemopreventive activity. PMID:26284516

  18. Interleukin-1 and tumor necrosis factor alpha inhibit repair of the porcine meniscus in vitro

    PubMed Central

    Hennerbichler, Alfred; Moutos, Franklin T.; Hennerbichler, Diana; Weinberg, J. Brice; Guilak, Farshid

    2011-01-01

    OBJECTIVE Injury or removal of the knee meniscus leads to progressive joint degeneration, and current surgical therapies for meniscal tears seek to maximally preserve meniscal structure and function. However, the factors that influence intrinsic repair of the meniscus are not well understood. The goal of this study was to investigate the capacity of meniscus tissue to repair a simulated defect in vitro and to examine the effect of pro-inflammatory cytokines on this process. METHODS Cylindrical explants were harvested from the outer one-third of medial porcine menisci. To simulate a full-thickness defect, a central core was removed and reinserted immediately into the defect. Explants were cultured for 2, 4, or 6 weeks in serum-containing media in the presence or absence of interleukin-1 (IL-1) or tumor necrosis factor alpha (TNF-alpha), and meniscal repair was investigated using mechanical testing and fluorescence confocal microscopy. RESULTS Meniscal lesions in untreated samples showed a significant capacity for intrinsic repair in vitro, with increasing cell accumulation and repair strength over time in culture. In the presence of IL-1 or TNF-alpha, no repair was observed despite the presence of abundant viable cells. CONCLUSIONS This study demonstrates that the meniscus exhibits an intrinsic repair response in vitro. However, the presence of pro-inflammatory cytokines completely inhibited repair. These findings suggest that increased levels of pro-inflammatory cytokines post-injury or under arthritic conditions may inhibit meniscal repair. Therefore, inhibition of these cytokines may provide a means of accelerating repair of damaged or injured menisci in vivo. PMID:17448702

  19. Combined Application of Essential Oil Compounds and Bacteriophage to Inhibit Growth of Staphylococcus aureus In Vitro.

    PubMed

    Ghosh, Anisha; Ricke, Steven C; Almeida, Giselle; Gibson, Kristen E

    2016-04-01

    Staphylococcus aureus is considered an important human pathogen. This study aimed to investigate the combination of essential oil compounds (EOCs) and bacteriophage as alternative antimicrobials to control S. aureus in vitro. Here, four EOCs (alpha-pinene, 3-carene, (+)-limonene, (1S)-(-)-β-pinene) were evaluated by disc diffusion assay (DDA) and growth inhibition assay (GIA) to determine inhibitory effects against five strains of S. aureus. Phage adsorption assays were performed with phage K up to 120 h at 6, 13, and 37 °C to determine lytic activity. Combinations of phage K and EOCs against S. aureus were also evaluated at 37 °C. Alpha-pinene exhibited significantly greater inhibition towards S. aureus strains when compared to other EOCs tested by the DDA. GIAs indicate that all S. aureus strains exhibited significantly reduced growth (P < 0.006) over a 48-h period when exposed to EOCs. Phage adsorption assays indicate that phage K has high lytic activity at 37 °C with at least a 1.5-log increase in the number of plaque-forming units (PFU) over 6 h when compared to 6 and 13 °C. S. aureus strains showed significantly reduced growth (P < 0.05) when treated with combined phage K and EOCs. Results from the combined effect of EOC and phage indicate that phage alone inhibits S. aureus in vitro at 37 °C as effectively as EOCs alone or in combination with phage with variability between strains. The results from DDA, GIA, and phage adsorption assays indicate that select EOCs and phage K can be used as antimicrobials against S. aureus in vitro with potential application in situ. PMID:26719188

  20. Fibroblast contraction of collagen lattices in vitro: inhibition by chronic inflammatory cell mediators.

    PubMed

    Ehrlich, H P; Wyler, D J

    1983-09-01

    Fibroblast-populated collagen lattices (FPCL), prepared in petri dishes with serum-containing culture medium and incubated at 37 degrees C, undergo progressive and symmetric contraction (reduction in size) over a period of days. The in vitro contraction process requires viable cells with intact cytoskeletal elements, is associated with cell elongation, and is believed to represent a fibroblast function which also occurs in vivo during wound healing and tissue fibrosis. We report that soluble mediators elaborated by chronic inflammatory cells cultured in vitro, when added to FPCL, inhibit lattice contraction. Granulomas, isolated from the liver of Schistosoma mansoni-infected mice, secrete a factor(s) with an estimated molecular weight between 13,700 and 43,000 daltons (gel filtration: Sephadex G-200) and pI = 6 (preparative isoelectrofocusing in granular gel) which inhibits lattice contraction but is not toxic to fibroblasts. Supernatants (cell-free conditioned culture medium) of cultured macrophages isolated from these granulomas also contain this activity. The contraction inhibitory activity present in granuloma culture supernatants is abrogated by the addition of indomethacin to the lattices, while the addition of prostaglandin E2 (PGE2) alone to lattices inhibits contraction. Furthermore, culture supernatants interfere with fibroblast elongation in lattices. We propose that the ability of fibroblasts to contract collagen lattices in vitro and a fibrotic mass in vivo may be regulated by soluble products of chronic inflammatory cells, including macrophages. This process may be mediated by fibroblast-derived prostaglandins which alter cytoskeletal functions and has implications for understanding regulation of tissue fibrogenesis in a variety of diseases. PMID:6885932

  1. An Engineered Arginase FC Protein Inhibits Tumor Growth In Vitro and In Vivo

    PubMed Central

    Li, Lihua; Wang, Yan; Chen, Jun; Cheng, Bi; Hu, Jiehua; Zhou, Yuehua; Gao, Xin; Gao, Liucun; Mei, Xifan; Sun, Meiyan; Zhang, Zhuomei; Song, Haifeng

    2013-01-01

    Arginine is a semiessential amino acid required for the growth of melanoma and hepatocellular carcinoma, and the enzymatic removal of arginine by pegylated arginine deiminase (ADI) or arginase is being tested clinically. Here, we report a genetically engineered arginase FC fusion protein exhibiting a prolonged half-life and enhanced efficacy. The use of this enzyme to treat different tumor lines both inhibited cell proliferation and impaired cellular migration in vitro and in vivo. Our data reinforce the hypothesis that nutritional depletion is a key strategy for cancer treatment. PMID:23737831

  2. [In vitro antimicrobial, antiadherent and antifungal activity of Brazilian medicinal plants on oral biofilm microorganisms and strains of the genus Candida].

    PubMed

    Alves, Pollianna Muniz; Queiroz, Lélia Maria Guedes; Pereira, Jozinete Vieira; Pereira, Maria do Socorro Vieira

    2009-01-01

    The antimicrobial, antifungal and antiadherent activity of aroeira-do-sertão, mallow and guava tree on oral biofilm microorganisms and oral candidiasis was evaluated in vitro. The extracts were shown to be effective in inhibiting the growth of bacteria of the oral biofilm and fungi of oral candidiasis, thus suggesting that these extracts can be used as alternative means of dental therapy. PMID:19448949

  3. Antioxidant and Preventive Effects of Extract from Nymphaea candida Flower on In Vitro Immunological Liver Injury of Rat Primary Hepatocyte Cultures

    PubMed Central

    Zhao, Jun; Liu, Tao; Ma, Long; Yan, Ming; Gu, Zhengyi; Huang, Yi; Xu, Fang; Zhao, Yu

    2011-01-01

    Nymphaea candida is traditional Uighur medicine that is commonly used to treat head pains, cough, hepatitis and hypertension in Xinjiang of China. In this article, the extract of N. candida was measured for antioxidant activity, using 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals scavenging assay and reducing power determination, and compared with those of the positive controls of butylated hydroxytoluene (BHT) and gallic acid (GA). The active extract was further purified by liquid-liquid partition to afford four fractions, of which the ethyl acetate-soluble (EA) fraction (NCE) exhibited the strongest antioxidant capacity with IC50 value of 12.6 μg/mL for DPPH. Thirteen phenolic compounds were isolated from this fraction, and they all showed significant antioxidant activities in DPPH model system. Furthermore, NCE showed potent antioxidant capacity with IC50 value of 59.32 μg/mL, 24.48 μg/mL and 86.85 μg/mL, for O2−, ·OH and H2O2 radicals, respectively. Moreover, NCE on BCG plus LPS-induced immunological liver injury was evaluated using primary cultured rat hepatocytes. NCE produced significant hepatoprotective effects as evidenced by decreased supernatant enzyme activities (AST—aspartate transaminase, P <  .01; ALT—alanine transferase, P <  .01) and nitric oxide (NO, P <  .01) production. These results revealed the in vitro antioxidant and hepatoprotective activities of NCE against immunological liver injury. Further investigations are necessary to verify these activities in vivo. PMID:19196740

  4. Mycoplasma Contamination Revisited: Mesenchymal Stromal Cells Harboring Mycoplasma hyorhinis Potently Inhibit Lymphocyte Proliferation In Vitro

    PubMed Central

    Zinöcker, Severin; Wang, Meng-Yu; Gaustad, Peter; Kvalheim, Gunnar; Rolstad, Bent; Vaage, John T.

    2011-01-01

    Background Mesenchymal stromal cells (MSC) have important immunomodulatory effects that can be exploited in the clinical setting, e.g. in patients suffering from graft-versus-host disease after allogeneic stem cell transplantation. In an experimental animal model, cultures of rat T lymphocytes were stimulated in vitro either with the mitogen Concanavalin A or with irradiated allogeneic cells in mixed lymphocyte reactions, the latter to simulate allo-immunogenic activation of transplanted T cells in vivo. This study investigated the inhibitory effects of rat bone marrow-derived MSC subsequently found to be infected with a common mycoplasma species (Mycoplasma hyorhinis) on T cell activation in vitro and experimental graft-versus-host disease in vivo. Principal Findings We found that M. hyorhinis infection increased the anti-proliferative effect of MSC dramatically, as measured by both radiometric and fluorimetric methods. Inhibition could not be explained solely by the well-known ability of mycoplasmas to degrade tritiated thymidine, but likely was the result of rapid dissemination of M. hyorhinis in the lymphocyte culture. Conclusions This study demonstrates the potent inhibitory effect exerted by M. hyorhinis in standard lymphocyte proliferation assays in vitro. MSC are efficient vectors of mycoplasma infection, emphasizing the importance of monitoring cell cultures for contamination. PMID:21264307

  5. Inhibition of wheat bran and it's active compoments on α-glucosidase in vitro

    PubMed Central

    Tu, Jie; Chen, Jun; Zhu, Shuyun; Zhang, Chunxiao; Chen, Hua; Liu, Youbing

    2013-01-01

    Background: Wheat bran is a traditional Chinese medicine; however, it is mostly used as feedstuff in China. Wheat bran is widely accepted as an important ingredient in many low-glycemic index foods in modern western societies; however, its glycemic control mechanism is unknown. Objective: To determine potent α-glucosidase inhibitory compounds from wheat bran and to identify the inhibition on α-glucosidase. Materials and Methods: Ethanolic extract of wheat bran was prepared to evaluate the inhibitory activity on α-glucosidase, then fractionation of the extract was guided by in vitro enzyme-inhibition assay, and the potent α-glucosidase inhibitory compounds were identified by high performance liquid chromatography and atmospheric pressure chemical ionization-mass spectrometry; finally the enzyme inhibition process was studied using the Michaelis-Menton and the Lineweaver-Burk equations. Results: Both baker's yeast and rat intestinal enzymes were mostly inhibited (87.9% and 66.8% inhibition, respectively) at concentration 0.6 mg/mL of the ethanolic extract of wheat bran. The petroleum ether fraction in the ethanolic extract of wheat bran showed significant activity against rat intestinal α-glucosidase, and revealed a dose-dependent effect. The inhibition was 76.57% at 0.3 mg/mL and 100% at 0.6 mg/mL. The active fraction 13 of petroleum ether fraction was identified as alkylresorcinols (ARs). ARs showed strong inhibition towards α-glucosidase and its IC50 value was found to be 37.58 μg/mL. The enzyme kinetic studies showed that, in the presence of ARs, the Michaelis-Menton constant (Km) remains constant whereas the maximal velocity (Vmax) decreases, revealing a non-competitive type of inhibition. Conclusion: The therapeutic potentiality of ARs in the management of the postprandial hyperglycemia will proliferate the utilization of wheat bran in controlling type 2 diabetes. PMID:24124282

  6. Comparison of Antimicrobial Activity of Chlorhexidine, Coconut Oil, Probiotics, and Ketoconazole on Candida albicans Isolated in Children with Early Childhood Caries: An In Vitro Study.

    PubMed

    Shino, Beena; Peedikayil, Faizal C; Jaiprakash, Shyamala R; Ahmed Bijapur, Gufran; Kottayi, Soni; Jose, Deepak

    2016-01-01

    Background. Early childhood caries (ECC) is associated with early colonisation and high levels of cariogenic microorganisms. With C. albicans being one of those, there is a need to determine the effectiveness of various chemotherapeutic agents against it. The study is aimed at isolating Candida species in children with ECC and at studying the antifungal effect of coconut oil, probiotics, Lactobacillus, and 0.2% chlorhexidine on C. albicans in comparison with ketoconazole. Materials and Methods. Samples were collected using sterile cotton swabs, swabbed on the tooth surfaces from children with ECC of 3 to 6 yrs and streaked on Sabouraud dextrose agar (HI Media) plates and incubated in a 5% CO2 enriched atmosphere at 37°C for 24 hours. Candida was isolated and its susceptibility to probiotics, chlorhexidine, ketoconazole, and coconut oil was determined using Disc Diffusion method. Results. The mean zone of inhibition for chlorhexidine was 21.8 mm, whereas for coconut oil it was 16.8 mm, for probiotics it was 13.5 mm, and for ketoconazole it was 22.3 mm. The difference between the groups was not statistically significant (Chi-square value 7.42, P value 0.06). Conclusion. Chlorhexidine and coconut oil have shown significant antifungal activity which is comparable with ketoconazole.

  7. Comparison of Antimicrobial Activity of Chlorhexidine, Coconut Oil, Probiotics, and Ketoconazole on Candida albicans Isolated in Children with Early Childhood Caries: An In Vitro Study.

    PubMed

    Shino, Beena; Peedikayil, Faizal C; Jaiprakash, Shyamala R; Ahmed Bijapur, Gufran; Kottayi, Soni; Jose, Deepak

    2016-01-01

    Background. Early childhood caries (ECC) is associated with early colonisation and high levels of cariogenic microorganisms. With C. albicans being one of those, there is a need to determine the effectiveness of various chemotherapeutic agents against it. The study is aimed at isolating Candida species in children with ECC and at studying the antifungal effect of coconut oil, probiotics, Lactobacillus, and 0.2% chlorhexidine on C. albicans in comparison with ketoconazole. Materials and Methods. Samples were collected using sterile cotton swabs, swabbed on the tooth surfaces from children with ECC of 3 to 6 yrs and streaked on Sabouraud dextrose agar (HI Media) plates and incubated in a 5% CO2 enriched atmosphere at 37°C for 24 hours. Candida was isolated and its susceptibility to probiotics, chlorhexidine, ketoconazole, and coconut oil was determined using Disc Diffusion method. Results. The mean zone of inhibition for chlorhexidine was 21.8 mm, whereas for coconut oil it was 16.8 mm, for probiotics it was 13.5 mm, and for ketoconazole it was 22.3 mm. The difference between the groups was not statistically significant (Chi-square value 7.42, P value 0.06). Conclusion. Chlorhexidine and coconut oil have shown significant antifungal activity which is comparable with ketoconazole. PMID:27051559

  8. Comparison of Antimicrobial Activity of Chlorhexidine, Coconut Oil, Probiotics, and Ketoconazole on Candida albicans Isolated in Children with Early Childhood Caries: An In Vitro Study

    PubMed Central

    Ahmed Bijapur, Gufran; Kottayi, Soni; Jose, Deepak

    2016-01-01

    Background. Early childhood caries (ECC) is associated with early colonisation and high levels of cariogenic microorganisms. With C. albicans being one of those, there is a need to determine the effectiveness of various chemotherapeutic agents against it. The study is aimed at isolating Candida species in children with ECC and at studying the antifungal effect of coconut oil, probiotics, Lactobacillus, and 0.2% chlorhexidine on C. albicans in comparison with ketoconazole. Materials and Methods. Samples were collected using sterile cotton swabs, swabbed on the tooth surfaces from children with ECC of 3 to 6 yrs and streaked on Sabouraud dextrose agar (HI Media) plates and incubated in a 5% CO2 enriched atmosphere at 37°C for 24 hours. Candida was isolated and its susceptibility to probiotics, chlorhexidine, ketoconazole, and coconut oil was determined using Disc Diffusion method. Results. The mean zone of inhibition for chlorhexidine was 21.8 mm, whereas for coconut oil it was 16.8 mm, for probiotics it was 13.5 mm, and for ketoconazole it was 22.3 mm. The difference between the groups was not statistically significant (Chi-square value 7.42, P value 0.06). Conclusion. Chlorhexidine and coconut oil have shown significant antifungal activity which is comparable with ketoconazole. PMID:27051559

  9. Experiment research on inhibition of glioma with sTRAIL in vitro.

    PubMed

    Dou, Yihe; Wang, Yangang; Xu, Jian; Li, Zhaojian; Sun, Peng; Meng, Qinghai

    2014-06-01

    We report that adenovirus mediated TNF-related apoptosis-inducing ligand (TRAIL) influenced the cell growth and cell cycle in the glioma cells in vitro. After being infected with the Ad-sTRAIL, U251 cell growth was inhibited. The expression of sTRAIL was detected using immunofluorescence. The higher rate of apoptosis was demonstrated using short-term microculture tetrazoliun (MTT) assay and flow cytometry. The rate of Ad-sTRAIL-inducing U251 cell apoptosis was increased depending on the dosage and the time. The apoptosis of G0/G1 and S phase cells was more significant than that of the control groups. The growth and proliferation of U251 cell line was inhibited after the infection of Ad-sTRAIL. It is dose- and time dependent. PMID:24156316

  10. Artemisinin inhibits in vitro and in vivo invasion and metastasis of human hepatocellular carcinoma cells.

    PubMed

    Weifeng, Tan; Feng, Shen; Xiangji, Luo; Changqing, Su; Zhiquan, Qiu; Huazhong, Zeng; Peining, Yan; Yong, Yu; Mengchao, Wu; Xiaoqing, Jiang; Wan-Yee, Lau

    2011-01-15

    Artemisinin (ART) is isolated from the medicinal plant Artemisia annua L. To determine its effects on the invasion and metastasis of tumors, the human hepatocellular carcinoma (HCC) cell lines HepG2 and SMMC-7721 were treated with different concentrations of ART. Starting at 12.5μM, ART had inhibitory effects in migration and invasion assays that increased at higher concentrations. The inhibitory effect also became stronger with time, from 24 to 72h. ART significantly inhibited the in vivo metastatic abilities of the HepG2 HCC cell line. ART inhibited the invasion and metastasis of HCC cells both in vitro and in vivo by reducing the level of the MMP2 metalloproteinase, and by inducing the TIMP2 protein. ART activated Cdc42, which enhanced E-cadherin activity, resulting in greater cell-cell adhesion, and significantly reduced metastasis. PMID:20739158

  11. Proanthocyanidin-rich extracts from cranberry fruit (Vaccinium macrocarpon Ait.) selectively inhibit the growth of human pathogenic fungi Candida spp. and Cryptococcus neoformans.

    PubMed

    Patel, Kunal D; Scarano, Frank J; Kondo, Miwako; Hurta, Robert A R; Neto, Catherine C

    2011-12-28

    Cranberry ( Vaccinium macrocarpon ) has been shown in clinical studies to reduce infections caused by Escherichia coli and other bacteria, and proanthocyanidins are believed to play a role. The ability of cranberry to inhibit the growth of opportunistic human fungal pathogens that cause oral, skin, respiratory, and systemic infections has not been well-studied. Fractions from whole cranberry fruit were screened for inhibition of five Candida species and Cryptococcus neoformans , a causative agent of fungal meningitis. Candida glabrata , Candida lusitaniae , Candida krusei , and Cryptococcus neoformans showed significant susceptibility to treatment with cranberry proanthocyanidin fractions in a broth microdilution assay, with minimum inhibitory concentrations as low as 1 μg/mL. MALDI-TOF MS analysis of subfractions detected epicatechin oligomers of up to 12 degrees of polymerization. Those containing larger oligomers caused the strongest inhibition. This study suggests that cranberry has potential as an antifungal agent.

  12. Baicalein Inhibits Staphylococcus aureus Biofilm Formation and the Quorum Sensing System In Vitro

    PubMed Central

    Wang, Ke; Hou, Changchun; Cai, Shuangqi; Huang, Yingying; Du, Zhongye; Huang, Hong; Kong, Jinliang; Chen, Yiqiang

    2016-01-01

    Biofilm formed by Staphylococcus aureus significantly enhances antibiotic resistance by inhibiting the penetration of antibiotics, resulting in an increasingly serious situation. This study aimed to assess whether baicalein can prevent Staphylococcus aureus biofilm formation and whether it may have synergistic bactericidal effects with antibiotics in vitro. To do this, we used a clinically isolated strain of Staphylococcus aureus 17546 (t037) for biofilm formation. Virulence factors were detected following treatment with baicalein, and the molecular mechanism of its antibiofilm activity was studied. Plate counting, crystal violet staining, and fluorescence microscopy revealed that 32 μg/mL and 64 μg/mL baicalein clearly inhibited 3- and 7-day biofilm formation in vitro. Moreover, colony forming unit count, confocal laser scanning microscopy, and scanning electron microscopy showed that vancomycin (VCM) and baicalein generally enhanced destruction of biofilms, while VCM alone did not. Western blotting and real-time quantitative polymerase chain reaction analyses (RTQ-PCR) confirmed that baicalein treatment reduced staphylococcal enterotoxin A (SEA) and α-hemolysin (hla) levels. Most strikingly, real-time qualitative polymerase chain reaction data demonstrated that 32 μg/mL and 64 μg/mL baicalein downregulated the quorum-sensing system regulators agrA, RNAIII, and sarA, and gene expression of ica, but 16 μg/mL baicalein had no effect. In summary, baicalein inhibited Staphylococcus aureus biofilm formation, destroyed biofilms, increased the permeability of vancomycin, reduced the production of staphylococcal enterotoxin A and α-hemolysin, and inhibited the quorum sensing system. These results support baicalein as a novel drug candidate and an effective treatment strategy for Staphylococcus aureus biofilm-associated infections. PMID:27128436

  13. Baicalein Inhibits Staphylococcus aureus Biofilm Formation and the Quorum Sensing System In Vitro.

    PubMed

    Chen, Yan; Liu, Tangjuan; Wang, Ke; Hou, Changchun; Cai, Shuangqi; Huang, Yingying; Du, Zhongye; Huang, Hong; Kong, Jinliang; Chen, Yiqiang

    2016-01-01

    Biofilm formed by Staphylococcus aureus significantly enhances antibiotic resistance by inhibiting the penetration of antibiotics, resulting in an increasingly serious situation. This study aimed to assess whether baicalein can prevent Staphylococcus aureus biofilm formation and whether it may have synergistic bactericidal effects with antibiotics in vitro. To do this, we used a clinically isolated strain of Staphylococcus aureus 17546 (t037) for biofilm formation. Virulence factors were detected following treatment with baicalein, and the molecular mechanism of its antibiofilm activity was studied. Plate counting, crystal violet staining, and fluorescence microscopy revealed that 32 μg/mL and 64 μg/mL baicalein clearly inhibited 3- and 7-day biofilm formation in vitro. Moreover, colony forming unit count, confocal laser scanning microscopy, and scanning electron microscopy showed that vancomycin (VCM) and baicalein generally enhanced destruction of biofilms, while VCM alone did not. Western blotting and real-time quantitative polymerase chain reaction analyses (RTQ-PCR) confirmed that baicalein treatment reduced staphylococcal enterotoxin A (SEA) and α-hemolysin (hla) levels. Most strikingly, real-time qualitative polymerase chain reaction data demonstrated that 32 μg/mL and 64 μg/mL baicalein downregulated the quorum-sensing system regulators agrA, RNAIII, and sarA, and gene expression of ica, but 16 μg/mL baicalein had no effect. In summary, baicalein inhibited Staphylococcus aureus biofilm formation, destroyed biofilms, increased the permeability of vancomycin, reduced the production of staphylococcal enterotoxin A and α-hemolysin, and inhibited the quorum sensing system. These results support baicalein as a novel drug candidate and an effective treatment strategy for Staphylococcus aureus biofilm-associated infections.

  14. β-catenin knockdown inhibits the proliferation of human glioma cells in vitro and in vivo

    PubMed Central

    WANG, ZHONG; CHEN, QIANXUE

    2016-01-01

    β-catenin is a crucial oncogene that is capable of regulating cancer progression. The aim of the present study was to clarify whether β-catenin was associated with the proliferation and progress of glioma. In order to knockdown the expression of β-catenin in human U251 glioma cells, three pairs of small interfering (si)RNA were designed and synthesized and the most effective siRNA was selected and used for silencing the endogenous β-catenin, which was detected by western blot analysis and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Proliferation was subsequently detected using a methylthiazolyl-tetrazolium bromide assay and the results demonstrated that knockdown of β-catenin significantly inhibited the proliferation of U251 cells in a time- and dose-dependent manner (P<0.01). Cell apoptosis rate was analyzed using flow cytometry and Annexin V-fluorescein isothiocyanate/propidium iodide staining demonstrated that β-catenin siRNA significantly increased the apoptosis of U251 cells (P<0.01). Furthermore, the results of an in vitro scratch assay demonstrated that β-catenin silencing suppressed the proliferation of U251 cells, as compared with the control group (P<0.01). In vivo, β-catenin expression levels in U251 cells were significantly inhibited (P<0.01) following β-catenin short hairpin (sh)RNA lentiviral-vector transfection, as detected by western blot analysis and RT-qPCR. Tumorigenicity experiments demonstrated that β-catenin inhibition significantly increased the survival rate of nude mice. The results of the present study demonstrated that knockdown of β-catenin expression significantly inhibited the progression of human glioma cancer cells, in vitro and in vivo; thus suggesting that β-catenin silencing may be a novel therapy for the treatment of human glioma. PMID:26998037

  15. Baicalein Inhibits Staphylococcus aureus Biofilm Formation and the Quorum Sensing System In Vitro.

    PubMed

    Chen, Yan; Liu, Tangjuan; Wang, Ke; Hou, Changchun; Cai, Shuangqi; Huang, Yingying; Du, Zhongye; Huang, Hong; Kong, Jinliang; Chen, Yiqiang

    2016-01-01

    Biofilm formed by Staphylococcus aureus significantly enhances antibiotic resistance by inhibiting the penetration of antibiotics, resulting in an increasingly serious situation. This study aimed to assess whether baicalein can prevent Staphylococcus aureus biofilm formation and whether it may have synergistic bactericidal effects with antibiotics in vitro. To do this, we used a clinically isolated strain of Staphylococcus aureus 17546 (t037) for biofilm formation. Virulence factors were detected following treatment with baicalein, and the molecular mechanism of its antibiofilm activity was studied. Plate counting, crystal violet staining, and fluorescence microscopy revealed that 32 μg/mL and 64 μg/mL baicalein clearly inhibited 3- and 7-day biofilm formation in vitro. Moreover, colony forming unit count, confocal laser scanning microscopy, and scanning electron microscopy showed that vancomycin (VCM) and baicalein generally enhanced destruction of biofilms, while VCM alone did not. Western blotting and real-time quantitative polymerase chain reaction analyses (RTQ-PCR) confirmed that baicalein treatment reduced staphylococcal enterotoxin A (SEA) and α-hemolysin (hla) levels. Most strikingly, real-time qualitative polymerase chain reaction data demonstrated that 32 μg/mL and 64 μg/mL baicalein downregulated the quorum-sensing system regulators agrA, RNAIII, and sarA, and gene expression of ica, but 16 μg/mL baicalein had no effect. In summary, baicalein inhibited Staphylococcus aureus biofilm formation, destroyed biofilms, increased the permeability of vancomycin, reduced the production of staphylococcal enterotoxin A and α-hemolysin, and inhibited the quorum sensing system. These results support baicalein as a novel drug candidate and an effective treatment strategy for Staphylococcus aureus biofilm-associated infections. PMID:27128436

  16. SIRT1 inhibition in pancreatic cancer models: contrasting effects in vitro and in vivo.

    PubMed

    Oon, Chern Ein; Strell, Carina; Yeong, Keng Yoon; Östman, Arne; Prakash, Jai

    2015-06-15

    Gemcitabine remains the standard treatment for pancreatic cancer, although most patients acquire resistance to the therapy. Up-regulated in pancreatic cancer, SIRT1 is involved in tumorigenesis and drug resistance. However the mechanism through which SIRT1 regulates drug sensitivity in cancer cells is mainly unknown. We hypothesise that inhibiting SIRT1 activity may increase sensitivity of pancreatic cancer cells to gemcitabine treatment through the regulation of apototic cell death, cell cycle, epithelial-mesenschymal-transition (EMT) and senescence. We demonstrate that gemcitabine or 6-Chloro-2,3,4,9-tetrahydro-1 H-Carbazole-1-carboxamide (EX527) SIRT1 inhibitor reduces PANC-1 cell proliferation in vitro. EX527 enhanced sensitivity of PANC-1 cells to gemcitabine treatment through increased apoptosis. However, EX527 displayed no beneficial effect either as a monotreatment or in combination with gemcitabine in the modulation of cell cycle progression. Combination treatment did not reverse the two phenomena known to affect drug sensitivity, namely EMT and senescence, which are both induced by gemcitabine. Unexpectedly, EX527 promoted PANC-1 xenograft tumour growth in SCID mice compared to control group. Dual tX527 and gemcitabine displayed no synergistic effect compared to gemcitabine alone. The study reveals that SIRT1 is involved in chemoresistance and that inhibiting SIRT1 activity with EX527 sensitised PANC-1 cells to gemcitabine treatment in vitro. Sensitisation of cells is shown to be mainly through induction of micronuclei formation as a result of DNA damage and apoptosis in vitro. However, the absence of positive combinatorial effects in vivo indicates possible effects on cells of the tumor microenvironment and suggests caution regarding the clinical relevance of tissue culture findings with EX527.

  17. Terminalia bellirica Extract Inhibits Low-Density Lipoprotein Oxidation and Macrophage Inflammatory Response in Vitro.

    PubMed

    Tanaka, Miori; Kishimoto, Yoshimi; Saita, Emi; Suzuki-Sugihara, Norie; Kamiya, Tomoyasu; Taguchi, Chie; Iida, Kaoruko; Kondo, Kazuo

    2016-01-01

    The deciduous tree Terminalia bellirica found in Southeast Asia is extensively used in traditional Indian Ayurvedic medicine for the treatment of hypertension, rheumatism, and diabetes. The anti-atherogenic effect of Terminalia bellirica fruit has not been fully elucidated. Here, we investigated the effect of Terminalia bellirica extract (TBE) on low-density lipoprotein (LDL) oxidation and inflammation in macrophages. TBE showed 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity (EC50: 7.2 ± 1.2 μg/mL) and 15-lipoxygenase inhibitory activity. TBE also significantly inhibited free radical-induced LDL oxidation compared to the solvent control in vitro. In THP-1 macrophages, TBE treatment resulted in significant decreases of the mRNA expression of tumor necrosis factor-alpha (TNF-α), interleukin-1beta (IL-1β), and lectin-like oxidized LDL receptor-1 (LOX-1). TBE also reduced matrix metalloproteinase (MMP)-9 secretion and intracellular reactive oxygen species (ROS) production in THP-1 macrophages. These results show that TBE has the inhibitory effects on LDL oxidation and macrophage inflammatory response in vitro, suggesting that its in vivo use might inhibit atherosclerosis plaque progression. PMID:27314393

  18. Anthocyanin Induces Apoptosis of DU-145 Cells In Vitro and Inhibits Xenograft Growth of Prostate Cancer

    PubMed Central

    Ha, U-Syn; Bae, Woong Jin; Kim, Su Jin; Yoon, Byung Il; Hong, Sung Hoo; Lee, Ji Youl; Hwang, Tae-Kon; Hwang, Sung Yeoun; Wang, Zhiping

    2015-01-01

    Purpose To investigate the effects of anthocyanins extracted from black soybean, which have antioxidant activity, on apoptosis in vitro (in hormone refractory prostate cancer cells) and on tumor growth in vivo (in athymic nude mouse xenograft model). Materials and Methods The growth and viability of DU-145 cells treated with anthocyanins were assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and apoptosis was assessed by DNA laddering. Immunoblotting was conducted to evaluate differences in the expressions of p53, Bax, Bcl, androgen receptor (AR), and prostate specific antigen (PSA). To study the inhibitory effects of anthocyanins on tumor growth in vivo, DU-145 tumor xenografts were established in athymic nude mice. The anthocyanin group was treated with daily oral anthocyanin (8 mg/kg) for 14 weeks. After 2 weeks of treatment, DU-145 cells (2×106) were inoculated subcutaneously into the right flank to establish tumor xenografts. Tumor dimensions were measured twice a week using calipers and volumes were calculated. Results Anthocyanin treatment of DU-145 cells resulted in 1) significant increase in apoptosis in a dose-dependent manner, 2) significant decrease in p53 and Bcl-2 expressions (with increased Bax expression), and 3) significant decrease in PSA and AR expressions. In the xenograft model, anthocyanin treatment significantly inhibit tumor growth. Conclusion This study suggests that anthocyanins from black soybean inhibit the progression of prostate cancer in vitro and in a xenograft model. PMID:25510742

  19. Sinusoidal electromagnetic fields promote bone formation and inhibit bone resorption in rat femoral tissues in vitro.

    PubMed

    Zhou, Jian; Ma, Xiao-Ni; Gao, Yu-Hai; Yan, Juan-Li; Shi, Wen-Gui; Xian, Cory J; Chen, Ke-Ming

    2016-01-01

    Effects of sinusoidal electromagnetic fields (SEMFs) on bone metabolism have not yet been well defined. The present study investigated SEMF effects on bone formation and resorption in rat femur bone tissues in vitro. Cultured femur diaphyseal (cortical bone) and metaphyseal (trabecular bone) tissues were treated with 50 Hz 1.8 mT SEMFs 1.5 h per day for up to 12 days and treatment effects on bone formation and resorption markers and associated gene expression were examined. Treatment with SEMFs caused a significant increase in alkaline phosphatase (ALP) activity and inhibited the tartrate-resistant acid phosphatase (TRACP) activity in the femoral diaphyseal or metaphyseal tissues. SEMFs also significantly increased levels of mRNA expression of osterix (OSX), insulin-like growth factor (IGF-1) and ALP in the bone tissues. SEMF treatment decreased glucose content and increased lactic acid contents in the culture conditioned medium. In addition, treatment with SEMFs decreased mRNA expression levels of bone resorption-related genes TRACP, macrophage colony stimulating factor (M-CSF) and cathepsin K (CTSK) in the cultured bone tissues. In conclusion, the current study demonstrated that treatment with 1.8 mT SEMFs at 1.5 h per day promoted bone formation, increased metabolism and inhibited resorption in both metaphyseal and diaphyseal bone tissues in vitro.

  20. Inhibition of adenovirus multiplication by inosine pranobex and interferon α in vitro

    PubMed Central

    Lasek, Witold; Janyst, Michał; Młynarczyk, Grażyna

    2016-01-01

    There are no specific antivirals designed for adenoviral infections. Due to many cases of adenovirus infections worldwide, epidemic nature of some types of adenoviruses, and growing number of patients with severe adenoviral infections resulting from dysfunction the immune system, the need for searching an effective and safe therapy is increasing. Inosine pranobex exerts antiviral effects which are both direct and secondary to immunomodulatory activity. In the present study we evaluated in vitro effect of inosine pranobex and interferon α (IFN-α) on replication of HAdV-2 and HAdV-5. The effectiveness of inosine pranobex under these conditions has not been previously reported. In conducted study we reported that inosine pranobex reduced the titer of infectious HAdV-2 and HAdV-5 in vitro. Higher concentrations of IP strongly inhibited multiplication of viruses. Combination of inosine pranobex and IFN-α display higher efficacy than either treatment alone and suggest that both agents may increase therapeutic effectiveness without augmenting toxic effects. Combination index calculations showed that inosine pranobex and INF-α synergistically inhibit HAdV-2 and HAdV-5 titers in A549 cells. PMID:26862302

  1. Cilengitide inhibits proliferation and differentiation of human endothelial progenitor cells in vitro

    SciTech Connect

    Loges, Sonja; Butzal, Martin; Otten, Jasmin; Schweizer, Michaela; Fischer, Uta; Bokemeyer, Carsten; Hossfeld, Dieter K.; Schuch, Gunter; Fiedler, Walter . E-mail: fiedler@uke.uni-hamburg.de

    2007-06-15

    Bone marrow derived hematopoietic stem cells can function as endothelial progenitor cells. They are recruited to malignant tumors and differentiate into endothelial cells. This mechanism of neovascularization termed vasculogenesis is distinct from proliferation of pre-existing vessels. To better understand vasculogenesis we developed a cell culture model with expansion and subsequent endothelial differentiation of human CD133{sup +} progenitor cells in vitro. {alpha}{sub v}{beta}{sub 3}-integrins are expressed by endothelial cells and play a role in the attachment of endothelial cells to the extracellular matrix. We investigated the effect of Cilengitide, a peptide-like, high affinity inhibitor of {alpha}{sub v}{beta}{sub 3}- and {alpha}{sub v}{beta}{sub 5}-integrins in our in vitro system. We could show expression of {alpha}{sub v}{beta}{sub 3}-integrin on 60 {+-} 9% of non-adherent endothelial progenitors and on 91 {+-} 7% of differentiated endothelial cells. {alpha}{sub v}{beta}{sub 3}-integrin was absent on CD133{sup +} hematopoietic stem cells. Cilengitide inhibited proliferation of CD133{sup +} cells in a dose-dependent manner. The development of adherent endothelial cells from expanded CD133{sup +} cells was reduced even stronger by Cilengitide underlining its effect on integrin mediated cell adhesion. Expression of endothelial antigens CD144 and von Willebrand factor on differentiating endothelial precursors was decreased by Cilengitide. In summary, Cilengitide inhibits proliferation and differentiation of human endothelial precursor cells underlining its anti-angiogenic effects.

  2. Linarin Inhibits the Acetylcholinesterase Activity In-vitro and Ex-vivo.

    PubMed

    Feng, Xinchi; Wang, Xin; Liu, Youping; Di, Xin

    2015-01-01

    Linarin is a flavone glycoside in the plants Flos chrysanthemi indici, Buddleja officinalis, Cirsium setosum, Mentha arvensis and Buddleja davidii, and has been reported to possess analgesic, antipyretic, anti-inflammatory and neuroprotective activities. In this paper, linarin was investigated for its AChE inhibitory potential both in-vitro and ex-vivo. Ellman's colorimetric method was used for the determination of AChE inhibitory activity in mouse brain. In-vitro assays revealed that linarin inhibited AChE activity with an IC50 of 3.801 ± 1.149 μM. Ex-vivo study showed that the AChE activity was significantly reduced in both the cortex and hippocampus of mice treated intraperitoneally with various doses of linarin (35, 70 and 140 mg/Kg). The inhibition effects produced by high dose of linarin were the same as that obtained after huperzine A treatment (0.5 mg/Kg). Molecular docking study revealed that both 4'-methoxyl group and 7-O-sugar moiety of linarin played important roles in ligand-receptor binding and thus they are mainly responsible for AChE inhibitory activity. In view of its potent AChE inhibitory activity, linarin may be a promising therapeutic agent for the treatment of some diseases associated with AChE, such as glaucoma, myasthenia gravis, gastric motility and Alzheimer's disease.

  3. Linarin Inhibits the Acetylcholinesterase Activity In-vitro and Ex-vivo

    PubMed Central

    Feng, Xinchi; Wang, Xin; Liu, Youping; Di, Xin

    2015-01-01

    Linarin is a flavone glycoside in the plants Flos chrysanthemi indici, Buddleja officinalis, Cirsium setosum, Mentha arvensis and Buddleja davidii, and has been reported to possess analgesic, antipyretic, anti-inflammatory and neuroprotective activities. In this paper, linarin was investigated for its AChE inhibitory potential both in-vitro and ex-vivo. Ellman’s colorimetric method was used for the determination of AChE inhibitory activity in mouse brain. In-vitro assays revealed that linarin inhibited AChE activity with an IC50 of 3.801 ± 1.149 μM. Ex-vivo study showed that the AChE activity was significantly reduced in both the cortex and hippocampus of mice treated intraperitoneally with various doses of linarin (35, 70 and 140 mg/Kg). The inhibition effects produced by high dose of linarin were the same as that obtained after huperzine A treatment (0.5 mg/Kg). Molecular docking study revealed that both 4’-methoxyl group and 7-O-sugar moiety of linarin played important roles in ligand-receptor binding and thus they are mainly responsible for AChE inhibitory activity. In view of its potent AChE inhibitory activity, linarin may be a promising therapeutic agent for the treatment of some diseases associated with AChE, such as glaucoma, myasthenia gravis, gastric motility and Alzheimer’s disease. PMID:26330885

  4. Unfractionated Heparin Promotes Osteoclast Formation in Vitro by Inhibiting Osteoprotegerin Activity

    PubMed Central

    Li, Binghan; Lu, Dan; Chen, Yuqing; Zhao, Minghui; Zuo, Li

    2016-01-01

    Heparin has been proven to enhance bone resorption and induce bone loss. Since osteoclasts play a pivotal role in bone resorption, the effect of heparin on osteoclastogenesis needs to be clarified. Since osteocytes are the key modulator during osteoclastogenesis, we evaluated heparin’s effect on osteoclastogenesis in vitro by co-culturing an osteocyte cell line (MLO-Y4) and pre-osteoclasts (RAW264.7). In this co-culture system, heparin enhanced osteoclastogenesis and osteoclastic bone resorption while having no influence on the production of RANKL (receptor activator of NFκB ligand), M-CSF (macrophage colony-stimulating factor), and OPG (osteoprotegerin), which are three main regulatory factors derived from osteocytes. According to previous studies, heparin could bind specifically to OPG and inhibit its activity, so we hypothesized that this might be a possible mechanism of heparin activity. To test this hypothesis, osteoclastogenesis was induced using recombinant RANKL or MLO-Y4 supernatant. We found that heparin has no effect on RANKL-induced osteoclastogenesis (contains no OPG). However, after incubation with OPG, the capacity of MLO-Y4 supernatant for supporting osteoclast formation was increased. This effect disappeared after OPG was neutralized and reappeared after OPG was replenished. These results strongly suggest that heparin promotes osteocyte-modulated osteoclastogenesis in vitro, at least partially, through inhibiting OPG activity. PMID:27110777

  5. ppGpp inhibits peptide elongation cycle of chloroplast translation system in vitro.

    PubMed

    Nomura, Yuhta; Takabayashi, Taito; Kuroda, Hiroshi; Yukawa, Yasushi; Sattasuk, Kwanchanok; Akita, Mitsuru; Nozawa, Akira; Tozawa, Yuzuru

    2012-01-01

    Chloroplasts possess common biosynthetic pathways for generating guanosine 3',5'-(bis)pyrophosphate (ppGpp) from GDP and ATP by RelA-SpoT homolog enzymes. To date, several hypothetical targets of ppGpp in chloroplasts have been suggested, but they remain largely unverified. In this study, we have investigated effects of ppGpp on translation apparatus in chloroplasts by developing in vitro protein synthesis system based on an extract of chloroplasts isolated from pea (Pisum sativum). The chloroplast extracts showed stable protein synthesis activity in vitro, and the activity was sensitive to various types of antibiotics. We have demonstrated that ppGpp inhibits the activity of chloroplast translation in dose-effective manner, as does the toxic nonhydrolyzable GTP analog guanosine 5'-(β,γ-imido)triphosphate (GDPNP). We further examined polyuridylic acid-directed polyphenylalanine synthesis as a measure of peptide elongation activity in the pea chloroplast extract. Both ppGpp and GDPNP as well as antibiotics, fusidic acid and thiostrepton, inhibited the peptide elongation cycle of the translation system, but GDP in the similar range of the tested ppGpp concentration did not affect the activity. Our results thus show that ppGpp directly affect the translation system of chloroplasts, as they do that of bacteria. We suggest that the role of the ppGpp signaling system in translation in bacteria is conserved in the translation system of chloroplasts.

  6. Aged black garlic extract induces inhibition of gastric cancer cell growth in vitro and in vivo.

    PubMed

    Wang, Xin; Jiao, Fei; Wang, Qin-Wen; Wang, Juan; Yang, Ke; Hu, Rong-Rong; Liu, Han-Chen; Wang, Hong-Yang; Wang, Yi-Shan

    2012-01-01

    There is mounting evidence that garlic extracts possess significant anticancer actions. However, no studies have been reported on the effects of aged black garlic extracts (ABGE) on gastric cancer in vitro or in vivo. To examine the potential action of ABGE against gastric cancer, the present study evaluated its effect on the inhibition of cell proliferation and induction of apoptosis in SGC-7901 human gastric cancer cells. Additionally, we performed an in vivo study by inoculating the murine foregastric carcinoma cell line in Kunming mice and treating them with various doses of ABGE (0, 200, 400 and 800 mg/kg, intraperitoneally) for 2 weeks. Dose-dependent apoptosis was detected in ABGE-treated cells in in vitro studies. In tumor-bearing mice, significant antitumor effects of ABGE were observed, such as growth inhibition of inoculated tumors. Further investigation of serum superoxide dismutases, glutathione peroxidase, interleukin-2 and the increased indices of spleen and thymus indicated that the anticancer action of ABGE may be partly due to its antioxidant and immunomodulative effects. PMID:21922142

  7. Terminalia bellirica Extract Inhibits Low-Density Lipoprotein Oxidation and Macrophage Inflammatory Response in Vitro

    PubMed Central

    Tanaka, Miori; Kishimoto, Yoshimi; Saita, Emi; Suzuki-Sugihara, Norie; Kamiya, Tomoyasu; Taguchi, Chie; Iida, Kaoruko; Kondo, Kazuo

    2016-01-01

    The deciduous tree Terminalia bellirica found in Southeast Asia is extensively used in traditional Indian Ayurvedic medicine for the treatment of hypertension, rheumatism, and diabetes. The anti-atherogenic effect of Terminalia bellirica fruit has not been fully elucidated. Here, we investigated the effect of Terminalia bellirica extract (TBE) on low-density lipoprotein (LDL) oxidation and inflammation in macrophages. TBE showed 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity (EC50: 7.2 ± 1.2 μg/mL) and 15-lipoxygenase inhibitory activity. TBE also significantly inhibited free radical-induced LDL oxidation compared to the solvent control in vitro. In THP-1 macrophages, TBE treatment resulted in significant decreases of the mRNA expression of tumor necrosis factor-alpha (TNF-α), interleukin-1beta (IL-1β), and lectin-like oxidized LDL receptor-1 (LOX-1). TBE also reduced matrix metalloproteinase (MMP)-9 secretion and intracellular reactive oxygen species (ROS) production in THP-1 macrophages. These results show that TBE has the inhibitory effects on LDL oxidation and macrophage inflammatory response in vitro, suggesting that its in vivo use might inhibit atherosclerosis plaque progression. PMID:27314393

  8. Unfractionated Heparin Promotes Osteoclast Formation in Vitro by Inhibiting Osteoprotegerin Activity.

    PubMed

    Li, Binghan; Lu, Dan; Chen, Yuqing; Zhao, Minghui; Zuo, Li

    2016-01-01

    Heparin has been proven to enhance bone resorption and induce bone loss. Since osteoclasts play a pivotal role in bone resorption, the effect of heparin on osteoclastogenesis needs to be clarified. Since osteocytes are the key modulator during osteoclastogenesis, we evaluated heparin's effect on osteoclastogenesis in vitro by co-culturing an osteocyte cell line (MLO-Y4) and pre-osteoclasts (RAW264.7). In this co-culture system, heparin enhanced osteoclastogenesis and osteoclastic bone resorption while having no influence on the production of RANKL (receptor activator of NFκB ligand), M-CSF (macrophage colony-stimulating factor), and OPG (osteoprotegerin), which are three main regulatory factors derived from osteocytes. According to previous studies, heparin could bind specifically to OPG and inhibit its activity, so we hypothesized that this might be a possible mechanism of heparin activity. To test this hypothesis, osteoclastogenesis was induced using recombinant RANKL or MLO-Y4 supernatant. We found that heparin has no effect on RANKL-induced osteoclastogenesis (contains no OPG). However, after incubation with OPG, the capacity of MLO-Y4 supernatant for supporting osteoclast formation was increased. This effect disappeared after OPG was neutralized and reappeared after OPG was replenished. These results strongly suggest that heparin promotes osteocyte-modulated osteoclastogenesis in vitro, at least partially, through inhibiting OPG activity. PMID:27110777

  9. In vitro adhesion of Candida glabrata to denture base acrylic resin modified by glow-discharge plasma treatment.

    PubMed

    Zamperini, Camila Andrade; Carneiro, Haline de Lima; Rangel, Elidiane Cipriano; Cruz, Nilson Cristino; Vergani, Carlos Eduardo; Machado, Ana Lucia

    2013-03-01

    This study evaluated the potential of plasma treatments to modify the surface chemistry and hydrophobicity of a denture base acrylic resin to reduce the Candida glabrata adhesion. Specimens (n = 54) with smooth surfaces were made and divided into three groups (n = 18): control - non-treated; experimental groups - submitted to plasma treatment (Ar/50 W; AAt/130 W). The effects of these treatments on chemical composition and surface topography of the acrylic resin were evaluated. Surface free energy measurements (SFE) were performed after the treatments and after 48 h of immersion in water. For each group, half (n = 9) of the specimens were preconditionated with saliva before the adhesion assay. The number of adhered C. glabrata was evaluated by cell counting after crystal violet staining. The Ar/50 W and AAt/130 W treatments altered the chemistry composition, hydrophobicity and topography of acrylic surface. The Ar/50 W group showed significantly lower C. glabrata adherence than the control group, in the absence of saliva. After preconditioning with saliva, C. glabrata adherence in experimental and control groups did not differ significantly. There were significant changes in the SFE after immersion in water. The results demonstrated that Ar/50 W treated surfaces have potential for reducing C. glabrata adhesion to denture base resins and deserve further investigation, especially to tailor the parameters to prolong the increased wettability. PMID:22809146

  10. Trends in frequency and in vitro antifungal susceptibility patterns of Candida isolates from women attending the STD outpatients clinic of a tertiary care hospital in Northern Italy during the years 2002-2007.

    PubMed

    Asticcioli, Sara; Sacco, Laura; Daturi, Rossana; Matti, Cecilia; Nucleo, Elisabetta; Zara, Francesca; Pagani, Laura

    2009-04-01

    Vulvovaginal candidiasis is a common mucosal infection caused by saprophytic and opportunistic yeasts belonging to the Candida genus. 518 vaginal swabs, with positive fungal culture were collected from unselected women attending the Sexually Transmitted Disease clinic of an Italian tertiary care hospital over a six year period to determine the pathogen prevalence in vulvovaginal candidiasis and to evaluate in vitro the antifungal susceptibilities of yeast recovered by Sensititre YeastOne antifungal panel plates according to CLSI document M27-A2. The isolates belonging to the genus Candida were 495 (95.5%) with Candida albicans percentage equal to 61.2%. Voriconazole was highly active (MIC50 0.008; MIC90 0.5 microg/ml), regardless of the species tested. On the contrary, fluconazole susceptibility was based upon the species. The intrinsic resistance to fluconazole of C. krusei was confirmed.

  11. Production of mouse monoclonal antibodies which inhibit in vitro adherence of Entamoeba histolytica trophozoites.

    PubMed Central

    Ravdin, J I; Petri, W A; Murphy, C F; Smith, R D

    1986-01-01

    Adherence by axenic Entamoeba histolytica trophozoites to mammalian cells is mediated by an N-acetylgalactosamine (GalNAc)-inhibitable adhesin on the surface of the parasite. We isolated 35 hybridoma cell lines producing antibodies to E. histolytica as indicated by ELISA with sonicated amebic protein or by immunofluorescence assay with fixed whole trophozoites. Tissue culture supernatants were further screened for subcloning by the ability to bind to Chinese hamster ovary (CHO) cells which were first exposed to a partially purified soluble preparation of the amebic GalNAc-inhibitable lectin. Eight tissue culture supernatants were positive in this assay. Antibodies from four subcloned cell lines (D3-14, H8-5, I12-2, and I1-21) inhibited amebic adherence to CHO cells (P less than 0.01). Of the original 35 tissue culture supernatants, 3 also inhibited amebic adherence (P less than 0.01; F1, F14, and J10); monoclonal antibodies in these supernatants did not bind to lectin-exposed CHO cells. Three purified monoclonal antibodies (H8-5, I12-2, and I1-21) inhibited amebic adherence at greater than or equal to 2 micrograms/10(4) amebae (P less than 0.05). None of these inhibitory monoclonal antibodies immunoprecipitated with a soluble amebic protein preparation following sodium dodecyl sulfate-polyacrylamide gel electrophoresis under denaturing conditions. Monoclonal antibodies which inhibit in vitro adherence by E. histolytica will be useful in purification of the GalNAc-inhibitable lectin. PMID:2873102

  12. Inhibition of the adenine nucleotide translocator by N-acetyl perfluorooctane sulfonamides in vitro

    SciTech Connect

    O'Brien, Timothy M. Oliveira, Paulo J.; Wallace, Kendall B.

    2008-03-01

    N-alkyl perfluorooctane sulfonamides have been widely used as surfactants on fabrics and papers, fire retardants, and anti-corrosion agents, among many other commercial applications. The global distribution and environmental persistence of these compounds has generated considerable interest regarding potential toxic effects. We have previously reported that perfluorooctanesulfonamidoacetate (FOSAA) and N-ethylperfluorooctanesulfonamidoacetate (N-EtFOSAA) induce the mitochondrial permeability transition (MPT) in vitro. In this study we tested the hypothesis that FOSAA and N-EtFOSAA interact with the adenine nucleotide translocator (ANT) resulting in a functional inhibition of the translocator and induction of the MPT. Respiration and membrane potential of freshly isolated liver mitochondria from Sprague-Dawley rats were measured using an oxygen electrode and a tetraphenylphosphonium-selective (TPP{sup +}) electrode, respectively. Mitochondrial swelling was measured spectrophotometrically. The ANT ligands bongkregkic acid (BKA) and carboxyatractyloside (cATR) inhibited uncoupling of mitochondrial respiration caused by 10 {mu}M N-EtFOSAA, 40 {mu}M FOSAA, and the positive control 8 {mu}M oleic acid. ADP-stimulated respiration and depolarization of mitochondrial membrane potential were inhibited by cATR, FOSAA, N-EtFOSAA, and oleic acid, but not by FCCP. BKA inhibited calcium-dependent mitochondrial swelling induced by FOSAA, N-EtFOSAA, and oleic acid. Seventy-five micromolar ADP also inhibited swelling induced by the test compounds, but cATR induced swelling was not inhibited by ADP. Results of this investigation indicate that N-acetyl perfluorooctane sulfonamides interact directly with the ANT to inhibit ADP translocation and induce the MPT, one or both of which may account for the metabolic dysfunction observed in vivo.

  13. In vitro synergic efficacy of the combination of Nystatin with the essential oils of Origanum vulgare and Pelargonium graveolens against some Candida species.

    PubMed

    Rosato, Antonio; Vitali, Cesare; Piarulli, Monica; Mazzotta, Manuela; Argentieri, Maria Pia; Mallamaci, Rosanna

    2009-10-01

    In this study we investigated a synergistic effect between the essential oils Origanum vulgare, Pelargonium graveolens and Melaleuca alternifolia and the antifungal compound Nystatin. Nystatin is considered a drug of choice in the treatment of fungal infections, but it can cause some considerable problems through its side effects, such as renal damage. Finding a new product that can reduce the Nystatin dose via combination is very important. Our findings showed an experimental occurrence of a synergistic interaction between two of these essential oils and Nystatin. The essential oil O. vulgare appeared to be the most effective, inhibiting all the Candida species evaluated in this study. Some combinations of Nystatin and P. graveolens essential oil did not have any synergistic interactions for some of the strains considered. Associations of Nystatin with M. alternifolia essential oil had only an additive effect. PMID:19616925

  14. In vitro synergic efficacy of the combination of Nystatin with the essential oils of Origanum vulgare and Pelargonium graveolens against some Candida species.

    PubMed

    Rosato, Antonio; Vitali, Cesare; Piarulli, Monica; Mazzotta, Manuela; Argentieri, Maria Pia; Mallamaci, Rosanna

    2009-10-01

    In this study we investigated a synergistic effect between the essential oils Origanum vulgare, Pelargonium graveolens and Melaleuca alternifolia and the antifungal compound Nystatin. Nystatin is considered a drug of choice in the treatment of fungal infections, but it can cause some considerable problems through its side effects, such as renal damage. Finding a new product that can reduce the Nystatin dose via combination is very important. Our findings showed an experimental occurrence of a synergistic interaction between two of these essential oils and Nystatin. The essential oil O. vulgare appeared to be the most effective, inhibiting all the Candida species evaluated in this study. Some combinations of Nystatin and P. graveolens essential oil did not have any synergistic interactions for some of the strains considered. Associations of Nystatin with M. alternifolia essential oil had only an additive effect.

  15. Induction of sister chromatid exchanges and inhibition of cellular proliferation in vitro. I. Caffeine

    SciTech Connect

    Guglielmi, G.E.; Vogt, T.F.; Tice, R.R.

    1982-01-01

    While many agents have been examined for their ability to induce SCE's, complete dose-response information has often been lacking. We have reexamined the ability of one such compound - caffeine - to induce SCEs and also to inhibit cellular proliferation in human peripheral lymphocytes in vitro. An acute exposure to caffeine prior to the DNA synthetic period did not affect either SCE frequency or the rate of cellular proliferation. Chronic exposure to caffeine throughout the culture period lead to both a dose-dependent increase in SCEs (SCE/sub d/ or doubling dose = 2.4 mM; SCE/sub 10/ or the dose capable of inducing 10 SCE = 1.4 mM) and a dose-dependent inhibition of cellular proliferation (IC/sub 50/ or the 50% inhibition concentration = 2.6 mM). The relative proportion of first generation metaphase cells, an assessment of proliferative inhibiton, increased linearly with increasing caffeine concentrations. However, SCE frequency increased nonlinearly over the same range of caffeine concentrations. Examination of the ratio of nonsymmetrical to symmetrical SCEs in third generation metaphase cells indicated that caffeine induced SCEs in equal frequency in each of three successive generations. The dependency of SCE induction and cellular proliferative inhibition on caffeine's presence during the DNA synthetic period suggests that caffeine may act as an antimetabolite in normal human cells.

  16. Crude Aloe vera Gel Shows Antioxidant Propensities and Inhibits Pancreatic Lipase and Glucose Movement In Vitro

    PubMed Central

    Taukoorah, Urmeela; Mahomoodally, M. Fawzi

    2016-01-01

    Aloe vera gel (AVG) is traditionally used in the management of diabetes, obesity, and infectious diseases. The present study aimed to investigate the inhibitory potential of AVG against α-amylase, α-glucosidase, and pancreatic lipase activity in vitro. Enzyme kinetic studies using Michaelis-Menten (Km) and Lineweaver-Burk equations were used to establish the type of inhibition. The antioxidant capacity of AVG was evaluated for its ferric reducing power, 2-diphenyl-2-picrylhydrazyl hydrate scavenging ability, nitric oxide scavenging power, and xanthine oxidase inhibitory activity. The glucose entrapment ability, antimicrobial activity, and total phenolic, flavonoid, tannin, and anthocyanin content were also determined. AVG showed a significantly higher percentage inhibition (85.56 ± 0.91) of pancreatic lipase compared to Orlistat. AVG was found to increase the Michaelis-Menten constant and decreased the maximal velocity (Vmax) of lipase, indicating mixed inhibition. AVG considerably inhibits glucose movement across dialysis tubes and was comparable to Arabic gum. AVG was ineffective against the tested microorganisms. Total phenolic and flavonoid contents were 66.06 ± 1.14 (GAE)/mg and 60.95 ± 0.97 (RE)/mg, respectively. AVG also showed interesting antioxidant properties. The biological activity observed in this study tends to validate some of the traditional claims of AVG as a functional food. PMID:26880905

  17. In vitro developmental toxicity test detects inhibition of stem cell differentiation by silica nanoparticles

    SciTech Connect

    Park, Margriet V.D.Z. Annema, Wijtske; Salvati, Anna; Lesniak, Anna; Elsaesser, Andreas; Barnes, Clifford; McKerr, George; Howard, C. Vyvyan; Lynch, Iseult; Dawson, Kenneth A.; Piersma, Aldert H.; Jong, Wim H. de

    2009-10-01

    While research into the potential toxic properties of nanomaterials is now increasing, the area of developmental toxicity has remained relatively uninvestigated. The embryonic stem cell test is an in vitro screening assay used to investigate the embryotoxic potential of chemicals by determining their ability to inhibit differentiation of embryonic stem cells into spontaneously contracting cardiomyocytes. Four well characterized silica nanoparticles of various sizes were used to investigate whether nanomaterials are capable of inhibition of differentiation in the embryonic stem cell test. Nanoparticle size distributions and dispersion characteristics were determined before and during incubation in the stem cell culture medium by means of transmission electron microscopy (TEM) and dynamic light scattering. Mouse embryonic stem cells were exposed to silica nanoparticles at concentrations ranging from 1 to 100 {mu}g/ml. The embryonic stem cell test detected a concentration dependent inhibition of differentiation of stem cells into contracting cardiomyocytes by two silica nanoparticles of primary size 10 (TEM 11) and 30 (TEM 34) nm while two other particles of primary size 80 (TEM 34) and 400 (TEM 248) nm had no effect up to the highest concentration tested. Inhibition of differentiation of stem cells occurred below cytotoxic concentrations, indicating a specific effect of the particles on the differentiation of the embryonic stem cells. The impaired differentiation of stem cells by such widely used particles warrants further investigation into the potential of these nanoparticles to migrate into the uterus, placenta and embryo and their possible effects on embryogenesis.

  18. The rhizobacterium Arthrobacter agilis produces dimethylhexadecylamine, a compound that inhibits growth of phytopathogenic fungi in vitro.

    PubMed

    Velázquez-Becerra, Crisanto; Macías-Rodríguez, Lourdes I; López-Bucio, José; Flores-Cortez, Idolina; Santoyo, Gustavo; Hernández-Soberano, Christian; Valencia-Cantero, Eduardo

    2013-12-01

    Plant diseases caused by fungal pathogens such as Botrytis cinerea and the oomycete Phytophthora cinnamomi affect agricultural production worldwide. Control of these pests can be done by the use of fungicides such as captan, which may have deleterious effects on human health. This study demonstrates that the rhizobacterium Arthrobacter agilis UMCV2 produces volatile organic compounds that inhibit the growth of B. cinerea in vitro. A single compound from the volatile blends, namely dimethylhexadecylamine (DMHDA), could inhibit the growth of both B. cinerea and P. cinnamomi when supplied to the growth medium in low concentrations. DMHDA also inhibited the growth of beneficial fungi Trichoderma virens and Trichoderma atroviride but at much higher concentrations. DMHDA-related aminolipids containing 4, 8, 10, 12, and 14 carbons in the alkyl chain were tested for their inhibitory effect on the growth of the pathogens. The results show that the most active compound from those tested was dimethyldodecylamine. This effect correlates with a decrease in the number of membrane lipids present in the mycelium of the pathogen including eicosanoic acid, (Z)-9-hexadecenoic acid, methyl ester, and (Z)-9-octadecenoic acid, methyl ester. Strawberry leaflets treated with DMHDA were not injured by the compound. These data indicate that DMHDA and related compounds, which can be produced by microorganisms may effectively inhibit the proliferation of certain plant pathogens. PMID:23674267

  19. Crude Aloe vera Gel Shows Antioxidant Propensities and Inhibits Pancreatic Lipase and Glucose Movement In Vitro.

    PubMed

    Taukoorah, Urmeela; Mahomoodally, M Fawzi

    2016-01-01

    Aloe vera gel (AVG) is traditionally used in the management of diabetes, obesity, and infectious diseases. The present study aimed to investigate the inhibitory potential of AVG against α-amylase, α-glucosidase, and pancreatic lipase activity in vitro. Enzyme kinetic studies using Michaelis-Menten (K m ) and Lineweaver-Burk equations were used to establish the type of inhibition. The antioxidant capacity of AVG was evaluated for its ferric reducing power, 2-diphenyl-2-picrylhydrazyl hydrate scavenging ability, nitric oxide scavenging power, and xanthine oxidase inhibitory activity. The glucose entrapment ability, antimicrobial activity, and total phenolic, flavonoid, tannin, and anthocyanin content were also determined. AVG showed a significantly higher percentage inhibition (85.56 ± 0.91) of pancreatic lipase compared to Orlistat. AVG was found to increase the Michaelis-Menten constant and decreased the maximal velocity (V max) of lipase, indicating mixed inhibition. AVG considerably inhibits glucose movement across dialysis tubes and was comparable to Arabic gum. AVG was ineffective against the tested microorganisms. Total phenolic and flavonoid contents were 66.06 ± 1.14 (GAE)/mg and 60.95 ± 0.97 (RE)/mg, respectively. AVG also showed interesting antioxidant properties. The biological activity observed in this study tends to validate some of the traditional claims of AVG as a functional food. PMID:26880905

  20. An in vitro approach to estimate putative inhibition of buprenorphine and norbuprenorphine glucuronidation.

    PubMed

    Oechsler, Stephanie; Skopp, Gisela

    2010-05-01

    An in vitro inhibition study was performed to investigate potential drug-drug interactions on glucuronidation of buprenorphine (BUP) and norbuprenorphine (NBUP), which represents the major elimination pathway of the drug using cDNA-expressed uridine 5'-diphosphate glucuronosyltransferases (UGTs) and human liver microsomes (HLMs). Following identification of major UGT enzymes for BUP and NBUP glucuronidation, substrates were incubated with drugs (amitriptyline, nortriptyline, lamotrigine, oxazepam, and temazepam), which are extensively cleared by glucuronidation as well as are often used during maintenance treatment. To evaluate the inhibitory potential, the half maximal inhibitor concentration (IC(50)), the inhibition constant (K (i)), and the inhibitor concentration (K (I)) that yield half the maximum rate of inactivation and the enzyme inactivation rate constant (k (inact)) were determined, if appropriate. Amitriptyline and temazepam are inhibitors of NBUP glucuronidation (UGT1A3, HLMs), whereas BUP glucuronidation was affected by amitriptyline (HLMs), oxazepam, and temazepam (UGT2B7). Additionally, BUP inhibits NBUP glucuronidation (UGT1A1, 1A3, HLMs) and vice versa (UGT1A3). A decrease in the metabolic clearance of NBUP may increase the risk of adverse effects such as respiratory depression. Further investigations are needed to evaluate whether inhibition of BUP and NBUP glucuronidation contributes to adverse events. PMID:20111869

  1. Properties of silver and copper nanoparticle-containing aqueous solutions and evaluation of their in vitro activity against Candida albicans and Staphylococcus aureus biofilms

    NASA Astrophysics Data System (ADS)

    Montes Aguirre, Melissa Mariluz

    Most microorganisms grow on surfaces as biofilms rather than as individual planktonic cells, and cells within biofilms show high levels of resistance against antimicrobial drugs. Thereby biofilm formation complicates treatment and contributes to high morbidity and mortality rates associated with infections. This study explores the physical, optical, and nano-structural properties of selected nanoparticles dispersed in aqueous solutions (nanoparticulate colloidal water or nanofluids) and examines their in vitro activity against microbial biofilms. Silver and copper nanofluids of various concentrations were prepared and studied. Their surface energies, surface charge and surface plasmonic resonance properties were obtained using contact angle measurement, zeta potential and optical spectrometer, respectively. The temperature dependence of the surface plasmon resonance behavior was also determined for the selected nanoparticulate aqueous solutions. A model of biofilm formation on the wells of microtiter plates was used to determine the in vitro activity of the nanoparticle preparations against both fungal (Candida albicans) and bacterial (Staphylococcus aureus) biofilms. Scanning electron microscopy (SEM) was used to observe the nanoparticle interactions with microbial cells. Results show that silver nanofluid has higher surface energy than that of the copper, the surface energy increases as the concentration of silver nanoparticles increases; and both nanoparticles in liquid are positively charged. The interaction between silver nanoparticles and water molecules produces notable changes on the usual temperature properties of water. Altogether, effectiveness of silver nanoparticle-containing liquids in controlling biofilm formation is observed and reported. For a given size of silver nanoparticles studied, it is found that the effective concentrations of silver nanoparticles against microbial biofilms are far lower than their cytotoxic concentrations, indicating an

  2. Persea americana Glycolic Extract: In Vitro Study of Antimicrobial Activity against Candida albicans Biofilm and Cytotoxicity Evaluation

    PubMed Central

    Jesus, D.; Oliveira, J. R.; Oliveira, F. E.; Higa, K. C.; Junqueira, J. C.; Jorge, A. O. C.; Back-Brito, G. N.; Oliveira, L. D.

    2015-01-01

    This study evaluated the antifungal activity of Persea americana extract on Candida albicans biofilm and its cytotoxicity in macrophage culture (RAW 264.7). To determine the minimum inhibitory concentration (MIC), microdilution in broth (CLSI M27-S4 protocol) was performed. Thereafter, the concentrations of 12.5, 25, 50, 100, and 200 mg/mL (n = 10) with 5 min exposure were analyzed on mature biofilm in microplate wells for 48 h. Saline was used as control (n = 10). After treatment, biofilm cells were scraped off and dilutions were plated on Sabouraud dextrose agar. After incubation (37°C/48 h), the values of colony forming units per milliliter (CFU/mL) were converted to log10 and analyzed (ANOVA and Tukey test, 5%). The cytotoxicity of the P. americana extract was evaluated on macrophages by MTT assay. The MIC of the extract was 6.25 mg/mL and with 12.5 mg/mL there was elimination of 100% of planktonic cultures. Regarding the biofilms, a significant reduction (P < 0.001) of the biofilm at concentrations of 50 (0.580 ± 0.209 log10), 100 (0.998 ± 0.508 log10), and 200 mg/mL (1.093 ± 0.462 log10) was observed. The concentrations of 200 and 100 mg/mL were cytotoxic for macrophages, while the concentrations of 50, 25, and 12.5 mg/mL showed viability higher than 55%. PMID:26605376

  3. Persea americana Glycolic Extract: In Vitro Study of Antimicrobial Activity against Candida albicans Biofilm and Cytotoxicity Evaluation.

    PubMed

    Jesus, D; Oliveira, J R; Oliveira, F E; Higa, K C; Junqueira, J C; Jorge, A O C; Back-Brito, G N; Oliveira, L D

    2015-01-01

    This study evaluated the antifungal activity of Persea americana extract on Candida albicans biofilm and its cytotoxicity in macrophage culture (RAW 264.7). To determine the minimum inhibitory concentration (MIC), microdilution in broth (CLSI M27-S4 protocol) was performed. Thereafter, the concentrations of 12.5, 25, 50, 100, and 200 mg/mL (n = 10) with 5 min exposure were analyzed on mature biofilm in microplate wells for 48 h. Saline was used as control (n = 10). After treatment, biofilm cells were scraped off and dilutions were plated on Sabouraud dextrose agar. After incubation (37°C/48 h), the values of colony forming units per milliliter (CFU/mL) were converted to log10 and analyzed (ANOVA and Tukey test, 5%). The cytotoxicity of the P. americana extract was evaluated on macrophages by MTT assay. The MIC of the extract was 6.25 mg/mL and with 12.5 mg/mL there was elimination of 100% of planktonic cultures. Regarding the biofilms, a significant reduction (P < 0.001) of the biofilm at concentrations of 50 (0.580 ± 0.209 log10), 100 (0.998 ± 0.508 log10), and 200 mg/mL (1.093 ± 0.462 log10) was observed. The concentrations of 200 and 100 mg/mL were cytotoxic for macrophages, while the concentrations of 50, 25, and 12.5 mg/mL showed viability higher than 55%. PMID:26605376

  4. Persea americana Glycolic Extract: In Vitro Study of Antimicrobial Activity against Candida albicans Biofilm and Cytotoxicity Evaluation.

    PubMed

    Jesus, D; Oliveira, J R; Oliveira, F E; Higa, K C; Junqueira, J C; Jorge, A O C; Back-Brito, G N; Oliveira, L D

    2015-01-01

    This study evaluated the antifungal activity of Persea americana extract on Candida albicans biofilm and its cytotoxicity in macrophage culture (RAW 264.7). To determine the minimum inhibitory concentration (MIC), microdilution in broth (CLSI M27-S4 protocol) was performed. Thereafter, the concentrations of 12.5, 25, 50, 100, and 200 mg/mL (n = 10) with 5 min exposure were analyzed on mature biofilm in microplate wells for 48 h. Saline was used as control (n = 10). After treatment, biofilm cells were scraped off and dilutions were plated on Sabouraud dextrose agar. After incubation (37°C/48 h), the values of colony forming units per milliliter (CFU/mL) were converted to log10 and analyzed (ANOVA and Tukey test, 5%). The cytotoxicity of the P. americana extract was evaluated on macrophages by MTT assay. The MIC of the extract was 6.25 mg/mL and with 12.5 mg/mL there was elimination of 100% of planktonic cultures. Regarding the biofilms, a significant reduction (P < 0.001) of the biofilm at concentrations of 50 (0.580 ± 0.209 log10), 100 (0.998 ± 0.508 log10), and 200 mg/mL (1.093 ± 0.462 log10) was observed. The concentrations of 200 and 100 mg/mL were cytotoxic for macrophages, while the concentrations of 50, 25, and 12.5 mg/mL showed viability higher than 55%.

  5. Lipopeptides from Bacillus subtilis AC7 inhibit adhesion and biofilm formation of Candida albicans on silicone.

    PubMed

    Ceresa, Chiara; Rinaldi, Maurizio; Chiono, Valeria; Carmagnola, Irene; Allegrone, Gianna; Fracchia, Letizia

    2016-10-01

    Candida albicans is the major fungus that colonises medical implants, causing device-associated infections with high mortality. Antagonistic bacterial products with interesting biological properties, such as biosurfactants, have recently been considered for biofilm prevention. This study investigated the activity of lipopeptide biosurfactant produced by Bacillus subtilis AC7 (AC7 BS) against adhesion and biofilm formation of C. albicans on medical-grade silicone elastomeric disks (SEDs). Chemical analysis, stability, surface activities of AC7 BS crude extract and physicochemical characterisation of the coated silicone disk surfaces were also carried out. AC7 BS showed a good reduction of water surface tension, low critical micelle concentration, good emulsification activity, thermal resistance and pH stability. Co-incubation with 2 mg ml(-1) AC7 BS significantly reduced adhesion and biofilm formation of three C. albicans strains on SEDs in a range of 67-69 % and of 56-57 %, respectively. On pre-coated SEDs, fungal adhesion and biofilm formation were reduced by 57-62 % and 46-47 %, respectively. Additionally, AC7 BS did not inhibit viability of C. albicans strains in both planktonic and sessile form. Chemical analysis of the crude extract revealed the presence of two families of lipopeptides, principally surfactin and a lower percentage of fengycin. The evaluation of surface wettability indicated that AC7 BS coating of SEDs surface was successful although uneven. AC7 BS significantly prohibits the initial deposition of C. albicans and slows biofilm growth, suggesting a potential role of biosurfactant coatings for preventing fungal infection associated with silicone medical devices. PMID:27444239

  6. Collaborative investigation of broth microdilution and semisolid agar dilution for in vitro susceptibility testing of Candida albicans.

    PubMed

    Shawar, R; Paetznick, V; Witte, Z; Ensign, L G; Anaissie, E; LaRocco, M

    1992-08-01

    A study was performed in two laboratories to evaluate the effect of growth medium and test methodology on inter- and intralaboratory variations in the MICs of amphotericin B (AMB), flucytosine (5FC), fluconazole (FLU), itraconazole (ITRA), and the triazole Sch 39304 (SCH) against 14 isolates of Candida albicans. Testing was performed by broth microdilution and semisolid agar dilution with the following media, buffered to pH 7.0 with morpholinepropanesulfonic acid (MOPS): buffered yeast nitrogen base (BYNB), Eagle's minimal essential medium (EMEM), RPMI 1640 medium (RPMI), and synthetic amino acid medium for fungi (SAAMF). Inocula were standardized spectrophotometrically, and endpoints were defined by the complete absence of growth for AMB and by no more than 25% of the growth in the drug-free control for all other agents. Comparative analyses of median MICs, as determined by each test method, were made for all drug-medium combinations. Both methods yielded similar (+/- 1 twofold dilution) median MICs for AMB in EMEM and RPMI, 5FC in all media, and FLU in EMEM, RPMI, and SAAMF. In contrast, substantial between-method variations in median MICs were seen for AMB in BYNB and SAAMF, FLU In BYNB, and ITRA and SCH in all media. Interlaboratory concordance of median MICs was good for AMB, 5FC, and FLU but poor for ITRA and SCH in all media. Endpoint determinations were analyzed by use of kappa statistical analyses for evaluating the strength of observer agreement. Moderate to almost perfect interlaboratory agreement occurred with AMB and 5FC in all media and with FLU in EMEM, RPMI, and SAAMF, irrespective of the test method. Slight to almost perfect interlaboratory agreement occurred with ITRA and SCH in EMEM, RPMI, and SAAMF when tested by semisolid agar dilution but not broth microdilution. Kappa values assessing intralaboratory agreement between methods were high for 5FC in all media, for AMB in BYNB, ENEM, and RPMI, and for FLU in EMEM, RPMI, and SAAMF. One laboratory

  7. Collaborative investigation of broth microdilution and semisolid agar dilution for in vitro susceptibility testing of Candida albicans.

    PubMed Central

    Shawar, R; Paetznick, V; Witte, Z; Ensign, L G; Anaissie, E; LaRocco, M

    1992-01-01

    A study was performed in two laboratories to evaluate the effect of growth medium and test methodology on inter- and intralaboratory variations in the MICs of amphotericin B (AMB), flucytosine (5FC), fluconazole (FLU), itraconazole (ITRA), and the triazole Sch 39304 (SCH) against 14 isolates of Candida albicans. Testing was performed by broth microdilution and semisolid agar dilution with the following media, buffered to pH 7.0 with morpholinepropanesulfonic acid (MOPS): buffered yeast nitrogen base (BYNB), Eagle's minimal essential medium (EMEM), RPMI 1640 medium (RPMI), and synthetic amino acid medium for fungi (SAAMF). Inocula were standardized spectrophotometrically, and endpoints were defined by the complete absence of growth for AMB and by no more than 25% of the growth in the drug-free control for all other agents. Comparative analyses of median MICs, as determined by each test method, were made for all drug-medium combinations. Both methods yielded similar (+/- 1 twofold dilution) median MICs for AMB in EMEM and RPMI, 5FC in all media, and FLU in EMEM, RPMI, and SAAMF. In contrast, substantial between-method variations in median MICs were seen for AMB in BYNB and SAAMF, FLU In BYNB, and ITRA and SCH in all media. Interlaboratory concordance of median MICs was good for AMB, 5FC, and FLU but poor for ITRA and SCH in all media. Endpoint determinations were analyzed by use of kappa statistical analyses for evaluating the strength of observer agreement. Moderate to almost perfect interlaboratory agreement occurred with AMB and 5FC in all media and with FLU in EMEM, RPMI, and SAAMF, irrespective of the test method. Slight to almost perfect interlaboratory agreement occurred with ITRA and SCH in EMEM, RPMI, and SAAMF when tested by semisolid agar dilution but not broth microdilution. Kappa values assessing intralaboratory agreement between methods were high for 5FC in all media, for AMB in BYNB, ENEM, and RPMI, and for FLU in EMEM, RPMI, and SAAMF. One laboratory

  8. Collaborative investigation of broth microdilution and semisolid agar dilution for in vitro susceptibility testing of Candida albicans.

    PubMed

    Shawar, R; Paetznick, V; Witte, Z; Ensign, L G; Anaissie, E; LaRocco, M

    1992-08-01

    A study was performed in two laboratories to evaluate the effect of growth medium and test methodology on inter- and intralaboratory variations in the MICs of amphotericin B (AMB), flucytosine (5FC), fluconazole (FLU), itraconazole (ITRA), and the triazole Sch 39304 (SCH) against 14 isolates of Candida albicans. Testing was performed by broth microdilution and semisolid agar dilution with the following media, buffered to pH 7.0 with morpholinepropanesulfonic acid (MOPS): buffered yeast nitrogen base (BYNB), Eagle's minimal essential medium (EMEM), RPMI 1640 medium (RPMI), and synthetic amino acid medium for fungi (SAAMF). Inocula were standardized spectrophotometrically, and endpoints were defined by the complete absence of growth for AMB and by no more than 25% of the growth in the drug-free control for all other agents. Comparative analyses of median MICs, as determined by each test method, were made for all drug-medium combinations. Both methods yielded similar (+/- 1 twofold dilution) median MICs for AMB in EMEM and RPMI, 5FC in all media, and FLU in EMEM, RPMI, and SAAMF. In contrast, substantial between-method variations in median MICs were seen for AMB in BYNB and SAAMF, FLU In BYNB, and ITRA and SCH in all media. Interlaboratory concordance of median MICs was good for AMB, 5FC, and FLU but poor for ITRA and SCH in all media. Endpoint determinations were analyzed by use of kappa statistical analyses for evaluating the strength of observer agreement. Moderate to almost perfect interlaboratory agreement occurred with AMB and 5FC in all media and with FLU in EMEM, RPMI, and SAAMF, irrespective of the test method. Slight to almost perfect interlaboratory agreement occurred with ITRA and SCH in EMEM, RPMI, and SAAMF when tested by semisolid agar dilution but not broth microdilution. Kappa values assessing intralaboratory agreement between methods were high for 5FC in all media, for AMB in BYNB, ENEM, and RPMI, and for FLU in EMEM, RPMI, and SAAMF. One laboratory

  9. IN VITRO CONAZOLE EXPOSURE INHIBITS TESTOSTERONE PRODUCTION IN THE ADULT AND NEONATAL RAT TESTIS THROUGH THE INHIBITION OF CYP17 ACTIVITY

    EPA Science Inventory

    IN VITRO CONAZOLE EXPOSURE INHIBITS TESTOSTERONE PRODUCTION IN THE ADULT AND NEONATAL RAT TESTIS THROUGH THE INHIBITION OF CYP17 ACTIVITY

    Chad R. Blystone1, David J. Dix2, and John C. Rockett2
    1Department of Environmental and Molecular Toxicology, NC State University, R...

  10. Thymoquinone inhibits proliferation in gastric cancer via the STAT3 pathway in vivo and in vitro

    PubMed Central

    Zhu, Wen-Qian; Wang, Jun; Guo, Xu-Feng; Liu, Zhou; Dong, Wei-Guo

    2016-01-01

    AIM: To elucidate the mechanism of thymoquinone (TQ)-induced apoptosis in human gastric cancer cells in vitro and in vivo. METHODS: HGC27, BGC823, and SGC7901 cells were cultured in vitro and treated with TQ (0, 10, 25, 50, 75, 100, 125 μmol/L) for 12 h, 24 h, and 36 h, and then the proliferation inhibitory rates were detected by methylthiazole tetrazolium assay. Apoptosis was observed after Hoechst staining. The protein expressions of signal transducer and activator of transcription (STAT)3, p-STAT3, STAT5, p-STAT5, phospho-janus-activated kinase 2 (JAK2), JAK2, p-Src, Src, glyceraldehyde-3-phosphate dehydrogenase, lamin-A, survivin, Cyclin D, Bcl-2, Bax, peroxisome proliferator activated receptor, and caspase-3,7,9 were detected by western blot. Cell cycle and apoptosis were determined with flow cytometry. TQ induced dose-dependent apoptotic cell death in HGC27 cells was measured by Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) analysis and Hoechst 33258. RESULTS: TQ inhibited the phosphorylation of STAT3 but not STAT5. TQ-induced downregulation of STAT3 activation was associated with a reduction in JAK2 and c-Src activity. TQ also downregulated the expression of STAT3-regulated genes, such as Bcl-2, cyclin D, survivin, and vascular endothelial growth factor, and activated caspase-3,7,9. Consistent with the in vitro results, TQ was significantly effective as an antitumor agent in a xenograft tumor mouse model. CONCLUSION: This study provides strong evidence that downregulation of the STAT3 signaling pathway mediates TQ-induced apoptosis in gastric cancer. PMID:27122665

  11. Secreted frizzled related proteins inhibit fibrosis in vitro but appear redundant in vivo

    PubMed Central

    2014-01-01

    Background The pathogenesis of pulmonary fibrosis remains poorly understood. The Wnt signaling pathway regulates fibrogenesis in different organs. Here, we studied the role of two extracellular Wnt antagonists, secreted frizzled-related protein-1 (SFRP1) and frizzled-related protein (FRZB) on lung fibrosis in vitro and in vivo. For this purpose, we used an alveolar epithelial cell line and a lung fibroblast cell line, and the bleomycin-induced lung fibrosis model, respectively. Results During the course of bleomycin-induced lung fibrosis, Sfrp1 and Frzb expression are upregulated. Expression of Sfrp1 appears much higher than that of Frzb. In vitro, recombinant SFRP1, but not FRZB, counteracts the transforming growth factor β1 (TGFβ1)-induced upregulation of type I collagen expression both in pulmonary epithelial cells and fibroblasts. Both SFRP1 and FRZB inhibit the TGFβ1-induced increase of active β-catenin, but do not influence the TGFβ1-induced phosphorylation levels of SMAD3, positioning Wnt signaling activity downstream of the active TGFβ signal in lung fibroblasts, but not in alveolar epithelial cells. In vivo, Sfrp1 −/− and Frzb −/− mice showed identical responses to bleomycin in the lung compared to wild-type controls. Conclusions Although SFRP1 counteracts the effect of TGFβ1 in pulmonary cells in vitro; loss of neither SFRP1 nor FRZB alters fibrotic outcomes in the lungs in vivo. The lack of in vivo effect in the absence of specific SFRPs suggests functional redundancy within this family of Wnt antagonists. PMID:25317206

  12. Raddeanin A induces human gastric cancer cells apoptosis and inhibits their invasion in vitro

    SciTech Connect

    Xue, Gang; Zou, Xi; Zhou, Jin-Yong; Sun, Wei; Wu, Jian; Xu, Jia-Li; Wang, Rui-Ping

    2013-09-20

    Highlights: •Raddeanin A is a triterpenoid saponin in herb medicine Anemone raddeana Regel. •Raddeanin A can inhibit 3 kinds of gastric cancer cells’ proliferation and invasion. •Caspase-cascades’ activation indicates apoptosis induced by Raddeanin A. •MMPs, RECK, Rhoc and E-cad are involved in Raddeanin A-induced invasion inhibition. -- Abstract: Raddeanin A is one of the triterpenoid saponins in herbal medicine Anemone raddeana Regel which was reported to suppress the growth of liver and lung cancer cells. However, little was known about its effect on gastric cancer (GC) cells. This study aimed to investigate its inhibitory effect on three kinds of different differentiation stage GC cells (BGC-823, SGC-7901 and MKN-28) in vitro and the possible mechanisms. Proliferation assay and flow cytometry demonstrated Raddeanin A’s dose-dependent inhibitory effect and determined its induction of cells apoptosis, respectively. Transwell assay, wounding heal assay and cell matrix adhesion assay showed that Raddeanin A significantly inhibited the abilities of the invasion, migration and adhesion of the BGC-823 cells. Moreover, quantitative real time PCR and Western blot analysis found that Raddeanin A increased Bax expression while reduced Bcl-2, Bcl-xL and Survivin expressions and significantly activated caspase-3, caspase-8, caspase-9 and poly-ADP ribose polymerase (PARP). Besides, Raddeanin A could also up-regulate the expression of reversion inducing cysteine rich protein with Kazal motifs (RECK), E-cadherin (E-cad) and down-regulate the expression of matrix metalloproteinases-2 (MMP-2), MMP-9, MMP-14 and Rhoc. In conclusion, Raddeanin A inhibits proliferation of human GC cells, induces their apoptosis and inhibits the abilities of invasion, migration and adhesion, exhibiting potential to become antitumor drug.

  13. Inhibition of Helicobacter pylori growth in vitro by Bulgarian propolis: preliminary report.

    PubMed

    Boyanova, Lyudmila; Derejian, Sirigan; Koumanova, Radka; Katsarov, Nikolai; Gergova, Galina; Mitov, Ivan; Nikolov, Rossen; Krastev, Zacharii

    2003-05-01

    Bee glue (propolis) possesses antimicrobial, anti-inflammatory, anaesthetic and immunostimulating activities. The aim of the study was to evaluate the inhibitory effect of Bulgarian propolis on Helicobacter pylori growth in vitro. Activity of 30% ethanolic extract of propolis (EEP) against 38 clinical isolates of H. pylori was evaluated by using the agar-well diffusion method. Ethanol was used as a control. In addition, the effect of propolis on the growth of 26 H. pylori and 18 Campylobacter strains was tested by the disc diffusion method. Mean diameters of H. pylori growth inhibition by the agar-well diffusion method, using 30, 60 or 90 microl EEP or 30 microl ethanol per well, were 17.8, 21.2, 28.2 and 8.5 mm, respectively. EEP was significantly more active than ethanol against H. pylori (P < 0.001). The results obtained by the disc diffusion method were similar. The use of moist propolis discs resulted in mean diameters of growth inhibition of 21.4 mm for H. pylori and 13.6 mm for Campylobacter spp. Dried propolis discs exhibited antibacterial effect against 73.1% of H. pylori isolates, with a considerable zone of growth inhibition (> or = 15 mm) in 36.4% of isolates. Using dried propolis discs resulted in mean diameters of growth inhibition of 12.4 mm for H. pylori and 11.6 mm for Campylobacter spp. In conclusion, Bulgarian propolis possesses considerable antibacterial activity against H. pylori, and can also inhibit the growth of Campylobacter jejuni and Campylobacter coli. The potential of propolis in the prevention or treatment of H. pylori infection is worth further extensive evaluation.

  14. alpha-Chaconine inhibits angiogenesis in vitro by reducing matrix metalloproteinase-2.

    PubMed

    Lu, Ming-Kun; Chen, Pei-Hsieng; Shih, Yuan-Wei; Chang, Ya-Ting; Huang, En-Tze; Liu, Cheng-Ruei; Chen, Pin-Shern

    2010-01-01

    alpha-Chaconine, a naturally occurring steroidal glycoalkaloid in potato sprouts, was found to possess anti-carcinogenic properties, such as inhibiting proliferation, migration, invasion, and inducing apoptosis of tumor cells. However, the effect of alpha-chaconine on tumor angiogenesis remains unclear. In the present study, we examined the effect of alpha-chaconine on angiogenesis in vitro. Data demonstrated that alpha-chaconine inhibited proliferation of bovine aortic endothelial cells (BAECs) in a dose-dependent manner. When treated with non-toxic doses of alpha-chaconine, cell migration, invasion and tube formation were markedly suppressed. Furthermore, alpha-chaconine reduced the expression and activity of matrix metalloproteinase-2 (MMP-2), which is involved in angiogenesis. Our biochemical assays indicated that alpha-chaconine potently suppressed the phosphorylation of c-Jun N-terminal kinase (JNK), phosphatidylinositide-3 kinase (PI3K) and Akt, while it did not affect phosphorylation of extracellular signal regulating kinase (ERK) and p38. In addition, alpha-chaconine significantly increased the cytoplasmic level of inhibitors of kappaBalpha (IkappaBalpha) and decreased the nuclear level of nuclear factor kappa B (NF-kappaB), suggesting that alpha-chaconine could inhibit NF-kappaB activity. Furthermore, the treatment of inhibitors specific for JNK (SP600125), PI3K (LY294002) or NF-kappaB (pyrrolidine dithiocarbamate) to BAECs reduced tube formation. Taken together, the results suggested that alpha-chaconine inhibited migration, invasion and tube formation of BAECs by reducing MMP-2 activities, as well as JNK and PI3K/Akt signaling pathways and inhibition of NF-kappaB activity. These findings reveal a new therapeutic potential for alpha-chaconine on anti-angiogenic therapy.

  15. An in vitro screening with emerging contaminants reveals inhibition of carboxylesterase activity in aquatic organisms.

    PubMed

    Solé, Montserrat; Sanchez-Hernandez, Juan C

    2015-12-01

    Pharmaceuticals and personal care products (PPCPs) form part of the new generation of pollutants present in many freshwater and marine ecosystems. Although environmental concentrations of these bioactive substances are low, they cause sublethal effects (e.g., enzyme inhibition) in non-target organisms. However, little is known on metabolism of PPCPs by non-mammal species. Herein, an in vitro enzyme trial was performed to explore sensitivity of carboxylesterase (CE) activity of aquatic organisms to fourteen PPCPs. The esterase activity was determined in the liver of Mediterranean freshwater fish (Barbus meridionalis and Squalius laietanus), coastal marine fish (Dicentrarchus labrax and Solea solea), middle-slope fish (Trachyrhynchus scabrus), deep-sea fish (Alepocephalus rostratus and Cataetix laticeps), and in the digestive gland of a decapod crustacean (Aristeus antennatus). Results showed that 100μM of the lipid regulators simvastatin and fenofibrate significantly inhibited (30-80% of controls) the CE activity of all target species. Among the personal care products, nonylphenol and triclosan were strong esterase inhibitors in most species (36-68% of controls). Comparison with literature data suggests that fish CE activity is as sensitive to inhibition by some PPCPs as that of mammals, although their basal activity levels are lower than in mammals. Pending further studies on the interaction between PPCPs and CE activity, we postulate that this enzyme may act as a molecular sink for certain PPCPs in a comparable way than that described for the organophosphorus pesticides. PMID:26562051

  16. Inhibition of adenovirus DNA synthesis in vitro by sera from patients with systemic lupus erythematosus

    SciTech Connect

    Horwitz, M.S.; Friefeld, B.R.; Keiser, H.D.

    1982-12-01

    Sera containing antinuclear antibodies from patients with systemic lupus erythematosus (SLE) and related disorders were tested for their effect on the synthesis of adenovirus (Ad) DNA in an in vitro replication system. After being heated at 60/sup 0/C for 1 h, some sera from patients with SLE inhibited Ad DNA synthesis by 60 to 100%. Antibodies to double-stranded DNA were present in 15 of the 16 inhibitory sera, and inhibitory activity copurified with anti-double-stranded DNA in the immunoglobulin G fraction. These SLE sera did not inhibit the DNA polymerases ..cap alpha.., BETA, ..gamma.. and had no antibody to the 72,000-dalton DNA-binding protein necessary for Ad DNA synthesis. The presence of antibodies to single-stranded DNA and a variety of saline-extractable antigens (Sm, Ha, nRNP, and rRNP) did not correlate with SLE serum inhibitory activity. Methods previously developed for studying the individual steps in Ad DNA replication were used to determine the site of inhibition by the SLE sera that contained antibody to double-stranded DNA. Concentrations of the SLE inhibitor that decreased the elongation of Ad DNA by greater than 85% had no effect on either the initiation of Ad DNA synthesis or the polymerization of the first 26 deoxyribonucleotides.

  17. An in vitro screening with emerging contaminants reveals inhibition of carboxylesterase activity in aquatic organisms.

    PubMed

    Solé, Montserrat; Sanchez-Hernandez, Juan C

    2015-12-01

    Pharmaceuticals and personal care products (PPCPs) form part of the new generation of pollutants present in many freshwater and marine ecosystems. Although environmental concentrations of these bioactive substances are low, they cause sublethal effects (e.g., enzyme inhibition) in non-target organisms. However, little is known on metabolism of PPCPs by non-mammal species. Herein, an in vitro enzyme trial was performed to explore sensitivity of carboxylesterase (CE) activity of aquatic organisms to fourteen PPCPs. The esterase activity was determined in the liver of Mediterranean freshwater fish (Barbus meridionalis and Squalius laietanus), coastal marine fish (Dicentrarchus labrax and Solea solea), middle-slope fish (Trachyrhynchus scabrus), deep-sea fish (Alepocephalus rostratus and Cataetix laticeps), and in the digestive gland of a decapod crustacean (Aristeus antennatus). Results showed that 100μM of the lipid regulators simvastatin and fenofibrate significantly inhibited (30-80% of controls) the CE activity of all target species. Among the personal care products, nonylphenol and triclosan were strong esterase inhibitors in most species (36-68% of controls). Comparison with literature data suggests that fish CE activity is as sensitive to inhibition by some PPCPs as that of mammals, although their basal activity levels are lower than in mammals. Pending further studies on the interaction between PPCPs and CE activity, we postulate that this enzyme may act as a molecular sink for certain PPCPs in a comparable way than that described for the organophosphorus pesticides.

  18. Probiotic strains and their combination inhibit in vitro adhesion of pathogens to pig intestinal mucosa.

    PubMed

    Collado, M C; Grześkowiak, Łukasz; Salminen, Seppo

    2007-09-01

    The aim of this study was to investigate in vitro the protective effect of commercial probiotic strains (Bifidobacterium lactis Bb12 and Lactobacillus rhamnosus LGG) alone and in combination on the adhesion of pathogenic strains as Salmonella, Clostridium, and Escherichia coli to pig intestinal mucus obtained from different intestinal regions. In combination, probiotic strains enhanced each other's adhesion, mainly in large intestinal mucus. Treatment of intestinal mucus with Bb12 and LGG, alone or in combination, significantly reduced (P < 0.05) the adhesion of the tested pathogens. The ability to inhibit pathogen adhesion appears to depend on the specific probiotics and pathogens and on the mucosal site. B. lactis Bb12 and L. rhamnosus LGG in combination revealed a better ability to inhibit adhesion of all pathogens tested to pig intestinal mucus than probiotic strains. Probiotic combinations could be useful for counteracting disease-associated aberrations in intestinal microbiota. Specific protective probiotics could be selected for particular pig pathogens. Probiotic strains from human origin and intended for human use also adhere to pig intestinal mucus and are able to displace and inhibit pathogens. PMID:17657533

  19. 3',5'Di-O-trityluridine inhibits in vitro flavivirus replication.

    PubMed

    De Burghgraeve, Tine; Selisko, Barbara; Kaptein, Suzanne; Chatelain, Grégory; Leyssen, Pieter; Debing, Yannick; Jacobs, Michael; Van Aerschot, Arthur; Canard, Bruno; Neyts, Johan

    2013-05-01

    The dengue fever virus (DENV) and the yellow fever virus (YFV) are members of the genus flavivirus in the family Flaviviridae. An estimated 50-100 million cases of DENV infections occur each year and approximately half a million patients require hospitalization. There is no vaccine or effective antiviral treatment available. There is an urgent need for potent and safe inhibitors of DENV replication; ideally such compounds should have broad-spectrum activity against flaviviruses. We here report on the in vitro activity of 3',5'di-O-trityluridine on flavivirus replication. The compound results in a dose-dependent inhibition of (i) DENV- and YFV-induced cytopathic effect (CPE) (EC₅₀ values in the low micromolar range for the 4 DENV serotypes), (ii) RNA replication (DENV-2 EC₅₀=1.5 μM; YFV-17D EC₅₀=0.83 μM) and (iii) viral antigen production. Antiviral activity was also demonstrated in DENV subgenomic replicons (which do not encode the structural viral proteins) (EC₅₀=2.3 μM), indicating that the compound inhibits intracellular events of the viral replication cycle. Preliminary data indicate that the molecule may inhibit the viral RNA-dependent RNA polymerase. PMID:23470860

  20. Inhibition of calcium oxalate crystallisation in vitro by an extract of Bergenia ciliata

    PubMed Central

    Saha, Sarmistha; Verma, Ramtej J.

    2013-01-01

    Objective To evaluate the effectiveness of an extract obtained from the rhizomes of Bergenia ciliata (Saxifragaceae) on the inhibition of calcium oxalate (CaOx) crystallisation in vitro. Materials and methods A hydro-alcoholic extract (30:70, v/v) of rhizomes of B. ciliata was prepared at different concentrations (1–10 mg/mL). The crystallisation of CaOx monohydrate (COM) was induced in a synthetic urine system. The nucleation and aggregation of COM crystals were measured using spectrophotometric methods. The rates of nucleation and aggregation were evaluated by comparing the slope of the turbidity of a control system with that of one exposed to the extract. The results were compared with a parallel study conducted with a marketed poly-herbal combination, Cystone, under identical concentrations. Crystals generated in the urine were also analysed by light microscopy. Statistical differences and percentage inhibitions were calculated and assessed. Results The extract of B. ciliata was significantly more effective in inhibiting the nucleation and aggregation of COM crystals in a dose-dependent manner than was Cystone. Moreover, the extract induced more CaOx dihydrate crystals, with a significant reduction in the number and size of COM crystals. Conclusion An extract of the traditional herb B. ciliata has an excellent inhibitory activity on crystalluria and therefore might be beneficial in dissolving urinary stones. However, further study in animal models of urolithiasis is needed to evaluate its potential anti-urolithiatic activity. PMID:26558080

  1. Marine compounds inhibit growth of multiple myeloma in vitro and in vivo

    PubMed Central

    Steiner, Normann; Ribatti, Domenico; Willenbacher, Wolfgang; Jöhrer, Karin; Kern, Johann; Marinaccio, Christian; Aracil, Miguel; García-Fernández, Luis F.; Gastl, Guenther

    2015-01-01

    Purpose The prognosis of patients with multiple myeloma (MM) is still dismal despite recent improvements achieved by introducing new therapeutic agents. However, there remains an urgent need for progress in myeloma drug development. We here show that novel marine-derived compounds can exert potent anti-myeloma activity. Experimental Design Nine marine-derived compounds were applied at low nM concentrations (0.1-100 nM) to MM cell lines (OPM-2, NCI-H929, U266, RPMI-8226), to primary human myeloma cells and to peripheral blood mononuclear cells. Apoptosis was determined by flow cytometry. In addition, eGFP-transgenic MM cell lines growing with mesenchymal cells from bone marrow were used to visualize tumors by fluorescence stereomicroscopy. Anti-myelomaactivities were studied in vitro in 3D spheroids and in vivo in myeloma xenografts on chicken embryos. Tumor size was analyzed by measuring GFP content with a GFP ELISA. Anti-angiogenic activities of compounds were tested in an in vivo gelatin sponge assay with conditioned media from primary bone marrow-derived endothelial cells. Results We identified a subset of marine compounds with strong anti-myeloma activity in vitro and in vivo. Moreover, some of the compounds inhibited myeloma-related angiogenesis in the in vivo gelatin sponge assay. They merit further drug development to improve treatment options for MM. PMID:25860931

  2. Microtubule protein ADP-ribosylation in vitro leads to assembly inhibition and rapid depolymerization

    SciTech Connect

    Scaife, R.M. ); Wilson, L. ); Purich, D.L. )

    1992-01-14

    Bovine brain microtubule protein, containing both tubulin and microtubule-associated proteins, undergoes ADP-ribosylation in the presence of ({sup 14}C)NAD{sup +} and a turkey erythrocyte mono-ADP-ribosyltransferase in vitro. The modification reaction could be demonstrated in crude brain tissue extracts where selective ADP-ribosylation of both the {alpha} and {beta} chains of tubulin and of the high molecular weight microtubule-associated protein MAP-2 occurred. In experiments with purified microtubule protein, tubulin dimer, the high molecular weight microtubule-associated protein MAP-2, and another high molecular weight microtubule-associated protein which may be a MAP-1 species were heavily labeled. Tubulin and MAP-2 incorporated ({sup 14}C)ADP-ribose to an average extent of approximately 2.4 and 30 mol of ADP-ribose/mol of protein, respectively. Assembly of microtubule protein into microtubules in vitro was inhibited by ADP-ribosylation, and incubation of assembled steady-state microtubules with ADP-ribosyltransferase and NAD{sup +} resulted in rapid depolymerization of the microtubules. Thus, the eukaryotic enzyme can ADP-ribosylate tubulin and microtubule-associated proteins to much greater extents than previously observed with cholera and pertussis toxins, and the modification can significantly modulate microtubule assembly and disassembly.

  3. Skeletal unloading inhibits the in vitro proliferation and differentiation of rat osteoprogenitor cells

    NASA Technical Reports Server (NTRS)

    Kostenuik, P. J.; Halloran, B. P.; Morey-Holton, E. R.; Bikle, D. D.

    1997-01-01

    Loss of weight bearing in the growing rat decreases bone formation, osteoblast numbers, and bone maturation in unloaded bones. These responses suggest an impairment of osteoblast proliferation and differentiation. To test this assumption, we assessed the effects of skeletal unloading using an in vitro model of osteoprogenitor cell differentiation. Rats were hindlimb elevated for 0 (control), 2, or 5 days, after which their tibial bone marrow stromal cells (BMSCs) were harvested and cultured. Five days of hindlimb elevation led to significant decreases in proliferation, alkaline phosphatase (AP) enzyme activity, and mineralization of BMSC cultures. Differentiation of BMSCs was analyzed by quantitative competitive polymerase chain reaction of cDNA after 10, 15, 20, and 28 days of culture. cDNA pools were analyzed for the expression of c-fos (an index of proliferation), AP (an index of early osteoblast differentiation), and osteocalcin (a marker of late differentiation). BMSCs from 5-day unloaded rats expressed 50% less c-fos, 61% more AP, and 35% less osteocalcin mRNA compared with controls. These data demonstrate that cultured osteoprogenitor cells retain a memory of their in vivo loading history and indicate that skeletal unloading inhibits proliferation and differentiation of osteoprogenitor cells in vitro.

  4. In vitro toxicities of experimental jet fuel system ice-inhibiting agents.

    PubMed

    Geiss, K T; Frazier, J M

    2001-07-01

    One research emphasis within the Department of Defense has been to seek the replacement of operational compounds with alternatives that pose less potential risk to human and ecological systems. Alternatives to glycol ethers, such as diethylene glycol monomethyl ether (M-DE), were investigated for use as jet fuel system ice-inhibiting agents (FSIIs). This group of chemicals includes three derivatives of 1,3-dioxolane-4-methanol (M-1, M-2, and M-3) and a 1,3-dioxane (M-27). In addition, M-DE was evaluated as a reference compound. Our approach was to implement an in vitro test battery based on primary rat hepatocyte cultures to perform initial toxicity evaluations. Hepatocytes were exposed to experimental chemicals (0, 0.001, 0.01, 0.1, 1, 10 mM dosages) for periods up to 24 h. Samples were assayed for lactate dehydrogenase (LDH) release, MTT dye reduction activity, glutathione level, and rate of protein synthesis as indicators of toxicity. Of the compounds tested, M-1, especially at the 10-mM dose, appeared to be more potent than the other chemicals, as measured by these toxicity assays. M-DE, the current FSII, elicited little response in the toxicity assays. Although some variations in toxicity were observed at the 10-mM dose, the in vitro toxicities of the chemicals tested (except for M-1) were not considerably greater than that of M-DE.

  5. Downregulation of the CXCR4 receptor inhibits cervical carcinoma metastatic behavior in vitro and in vivo

    PubMed Central

    SEKUŁA, MAŁGORZATA; MIEKUS, KATARZYNA; MAJKA, MARCIN

    2014-01-01

    Cervical carcinoma is frequently diagnosed among women, particularly in low and middle income countries. In this study, we investigated the role of the SDF-1/CXCR4 axis during cervical carcinoma growth and progression in vitro and in vivo. Downregulation of CXCR4 receptor using an RNA interference system led to almost complete inhibition of the receptor expression, activation and function. CXCR4 receptor silencing led to decreased ability to signal, to induce migration and to form holoclone-like colonies, with no influence on viability/proliferation of the cells. CXCR4-deficient cells had also significantly lower levels of MMP-9. Interestingly, downregulation of CXCR4 expression resulted in reduced tumor growth in vivo. Tumors generated by CXCR4-deficient cells had also lower expression of the proliferation marker Ki-67 and decreased ability to engraft into lungs and spleen. Taken together, our results indicate that CXCR4 receptor may play an important role during cervical carcinoma invasion. In our study CXCR4 influenced invasive properties of cervical carcinoma cells both in vitro and in vivo. PMID:24728301

  6. The novel JNK inhibitor AS602801 inhibits cancer stem cells in vitro and in vivo

    PubMed Central

    Okada, Masashi; Kuramoto, Kenta; Takeda, Hiroyuki; Watarai, Hikaru; Sakaki, Hirotsugu; Seino, Shizuka; Seino, Manabu; Suzuki, Shuhei; Kitanaka, Chifumi

    2016-01-01

    A phase 2 clinical trial investigating the efficacy and safety of AS602801, a newly developed JNK inhibitor, in the treatment of inflammatory endometriosis is complete. We are now examining whether AS602801 acts against human cancer cells in vitro and in vivo. In vitro, AS602801 exhibited cytotoxicity against both serum-cultured non-stem cancer cells and cancer stem cells derived from human pancreatic cancer, non-small cell lung cancer, ovarian cancer and glioblastoma at concentrations that did not decrease the viability of normal human fibroblasts. AS602801 also inhibited the self-renewal and tumor-initiating capacity of cancer stem cells surviving AS602801 treatment. Cancer stem cells in established xenograft tumors were reduced by systemic administration of AS602801 at a dose and schedule that did not adversely affect the health of the tumor-bearing mice. These findings suggest AS602801 is a promising anti-cancer stem cell agent, and further investigation of the utility of AS602801 in the treatment of cancer seems warranted. PMID:27027242

  7. Mechanisms of transthyretin inhibition of β-amyloid aggregation in vitro.

    PubMed

    Li, Xinyi; Zhang, Xin; Ladiwala, Ali Reza A; Du, Deguo; Yadav, Jay K; Tessier, Peter M; Wright, Peter E; Kelly, Jeffery W; Buxbaum, Joel N

    2013-12-11

    Tissue-specific overexpression of the human systemic amyloid precursor transthyretin (TTR) ameliorates Alzheimer's disease (AD) phenotypes in APP23 mice. TTR-β-amyloid (Aβ) complexes have been isolated from APP23 and some human AD brains. We now show that substoichiometric concentrations of TTR tetramers suppress Aβ aggregation in vitro via an interaction between the thyroxine binding pocket of the TTR tetramer and Aβ residues 18-21 (nuclear magnetic resonance and epitope mapping). The K(D) is micromolar, and the stoichiometry is <1 for the interaction (isothermal titration calorimetry). Similar experiments show that engineered monomeric TTR, the best inhibitor of Aβ fibril formation in vitro, did not bind Aβ monomers in liquid phase, suggesting that inhibition of fibrillogenesis is mediated by TTR tetramer binding to Aβ monomer and both tetramer and monomer binding of Aβ oligomers. The thousand-fold greater concentration of tetramer relative to monomer in vivo makes it the likely suppressor of Aβ aggregation and disease in the APP23 mice. PMID:24336709

  8. GBT440 Inhibits Sickling of Sickle Cell Trait Blood Under In Vitro Conditions Mimicking Strenuous Exercise

    PubMed Central

    Dufu, Kobina; Lehrer-Graiwer, Josh; Ramos, Eleanor; Oksenberg, Donna

    2016-01-01

    In sickle cell trait (SCT), hemoglobin A (HbA) and S (HbS) are co-expressed in each red blood cell (RBC). While homozygous expression of HbS (HbSS) leads to polymerization and sickling of RBCs resulting in sickle cell disease (SCD) characterized by hemolytic anemia, painful vaso-occlusive episodes and shortened life-span, SCT is considered a benign condition usually with minor or no complications related to sickling. However, physical activities that cause increased tissue oxygen demand, dehydration and/or metabolic acidosis leads to increased HbS polymerization and life-threatening complications including death. We report that GBT440, an agent being developed for the treatment of SCD, increases the affinity of oxygen for Hb and inhibits in vitro polymerization of a mixture of HbS and HbA that simulates SCT blood. Moreover, GBT440 prevents sickling of SCT blood under in vitro conditions mimicking strenuous exercise with hypoxia, dehydration and acidosis. Together, our results indicate that GBT440 may have the potential to protect SCT individuals from sickling-related complications during conditions that favor HbS polymerization. PMID:27757216

  9. Soluble form of canine transferrin receptor inhibits canine parvovirus infection in vitro and in vivo.

    PubMed

    Wen, Jiexia; Pan, Sumin; Liang, Shuang; Zhong, Zhenyu; He, Ying; Lin, Hongyu; Li, Wenyan; Wang, Liyue; Li, Xiujin; Zhong, Fei

    2013-01-01

    Canine parvovirus (CPV) disease is an acute, highly infectious disease threatening the dog-raising industry. So far there are no effective therapeutic strategies to control this disease. Although the canine transferrin receptor (TfR) was identified as a receptor for CPV infection, whether extracellular domain of TfR (called soluble TfR (sTfR)) possesses anti-CPV activities remains elusive. Here, we used the recombinant sTfR prepared from HEK293T cells with codon-optimized gene structure to investigate its anti-CPV activity both in vitro and in vivo. Our results indicated that codon optimization could significantly improve sTfR expression in HEK293T cells. The prepared recombinant sTfR possessed a binding activity to both CPV and CPV VP2 capsid proteins and significantly inhibited CPV infection of cultured feline F81 cells and decreased the mortality of CPV-infected dogs, which indicates that the sTfR has the anti-CPV activity both in vitro and in vivo.

  10. In vitro inhibition of caprine herpesvirus 1 by acyclovir and mizoribine.

    PubMed

    Elia, G; Camero, M; Decaro, N; Lovero, A; Martella, V; Tempesta, M; Buonavoglia, C; Crescenzo, G

    2015-04-01

    Caprine herpesvirus 1 (CpHV-1) infection in goats induces genital vesicular-ulcerative lesions that strictly resemble the lesions induced by herpesvirus 2 in the human host. The immunosuppressive drug Mizoribine (MIZ) was found to increase the antiviral activity of Acyclovir (ACV) against herpesvirus infections, raising interesting perspectives on new combined therapeutic strategies. In this study the anti-CpHV-1 activity in vitro of ACV alone or in combination with MIZ was characterized. When applied alone at non-toxic concentrations, ACV had a slight effect on CpHV-1 replication while in combination with MIZ a dose-dependent inhibition of the virus yield was observed with an IC50 of ACV of 28.5 µM. These findings suggest that combined therapy of ACV and MIZ is potentially exploitable in the treatment of genital infection by herpesviruses.

  11. The Acyclic Retinoid Peretinoin Inhibits Hepatitis C Virus Replication and Infectious Virus Release in Vitro

    NASA Astrophysics Data System (ADS)

    Shimakami, Tetsuro; Honda, Masao; Shirasaki, Takayoshi; Takabatake, Riuta; Liu, Fanwei; Murai, Kazuhisa; Shiomoto, Takayuki; Funaki, Masaya; Yamane, Daisuke; Murakami, Seishi; Lemon, Stanley M.; Kaneko, Shuichi

    2014-04-01

    Clinical studies suggest that the oral acyclic retinoid Peretinoin may reduce the recurrence of hepatocellular carcinoma (HCC) following surgical ablation of primary tumours. Since hepatitis C virus (HCV) infection is a major cause of HCC, we assessed whether Peretinoin and other retinoids have any effect on HCV infection. For this purpose, we measured the effects of several retinoids on the replication of genotype 1a, 1b, and 2a HCV in vitro. Peretinoin inhibited RNA replication for all genotypes and showed the strongest antiviral effect among the retinoids tested. Furthermore, it reduced infectious virus release by 80-90% without affecting virus assembly. These effects could be due to reduced signalling from lipid droplets, triglyceride abundance, and the expression of mature sterol regulatory element-binding protein 1c and fatty acid synthase. These negative effects of Peretinoin on HCV infection may be beneficial in addition to its potential for HCC chemoprevention in HCV-infected patients.

  12. Screening of selected pesticides for inhibition of CYP19 aromatase activity in vitro.

    PubMed

    Vinggaard, A M; Hnida, C; Breinholt, V; Larsen, J C

    2000-06-01

    Many pesticides are able to block or activate the steroid hormone receptors and/or to affect the levels of sex hormones, thereby potentially affecting the development or expression of the male and female reproductive system or both. This emphasizes the relevance of screening pesticides for a wide range of hormone-mimicking effects. Twenty-two pesticides were tested for their ability to affect CYP19 aromatase activity in human placental microsomes using the classical [(3)H](2)O method. Prochloraz, imazalil, propioconazole, fenarimol, triadimenol, triadimefon (all fungicides), and dicofol (an acaricide) gave rise to a statistically significant inhibition of aromatase activity. The IC(50)s of prochloraz, imazalil, propioconazole fenarimol, triadimenol, and triadimefon were calculated from dose-response curves to be 0.04, 0.34, 6.5, 10, 21 and 32 microM, respectively. The IC(50) of dicofol was greater than 50 microM. The positive control 4-hydroxyandrostendione (1 microM) caused an inhibition of aromatase activity by 74%. The compounds, which did not affect the aromatase activity, were bromopropylate, chlorfenvinphos, chlorobenzilate, chlorpyrifos, diuron, heptachlor, iprodion, linuron, pentachlorphenol, procymidon, propyzamide, quintozen, tetrachlorvinphos and tetradifon. With the purpose of comparing the results for fenarimol obtained with the microsomal system with data from an intact cell system, an aromatase assay based on JEG-3 cells was established. 4-Hydroxyandrostendione (1 microM) inhibited the aromatase activity in JEG-3 cells by 94%. The IC(50) for fenarimol in this system was 2 microM, slightly lower than that observed in the microsomal system. For the first time, fenarimol has been demonstrated to inhibit aromatase activity in human tissues and, furthermore, propioconazole, triadimefon, and triadimenol were identified as weak aromatase inhibitors. In conclusion, seven out of 22 tested pesticides turned out to be weak to moderate aromatase inhibitors in

  13. In vitro inhibition of human cytomegalovirus replication by calcium trinatrium diethylenetriaminepentaacetic acid.

    PubMed

    Cinatl, J; Hoffmann, F; Cinatl, J; Weber, B; Scholz, M; Rabenau, H; Stieneker, F; Kabickova, H; Blasko, M; Doerr, H W

    1996-06-01

    Desferrioxamine (DFO) has been shown to inhibit human cytomegalovirus (CMV) replication in vitro. In the present study, we compared antiviral effects of DFO in human foreskin fibroblast (HFF) cells against several CMV strains with those of other chelators that interact with iron and other ions from different pools. DFO, a hydrophilic chelator, that may chelate both intracellular and extracellular ions inhibited production of CMV late antigen at 50% effective concentrations (EC50S) ranging from 6.2 to 8.9 microM. EC50S for calcium trinatrium diethylenetriaminepentaacetic acid (CaDTPA) ranged from 6.1 to 9.9 microM. EC50S for 2,2'-bipyridine (BPD), a hydrophobic chelator, which diffuses into cell membranes ranged from 65 to 72 microM. Concentrations which inhibited BrdU incorporation into cellular DNA by 50% (IC50S) ranged from 8.2 to 12.0 microM (DFO), from 65 to 89 microM (BPD), and from 139 to 249 microM (CaDTPA). CaDTPA was the only chelator which completely inhibited production of infectious virus in HFF and vascular endothelial cells at concentrations which had no significant effects on cellular DNA synthesis and growth. Addition of stoichiometric amounts of Fe3+ in the culture medium of HFF cells completely eliminated antiviral effects of DFO while antiviral effects of CaDTPA and BPD were only moderately affected. Fe2+ and Cu2+ were stronger inhibitors of CaDTPA than Fe3+; however, Mn2+ and Zn2+ completely suppressed antiviral effects of CaDTPA. The results show that CaDTPA is a novel nontoxic inhibitor of CMV replication. The antiviral activity of CaDTPA is suppressed by metal ions with a decreasing potency order of Mn2+/Zn2+ > Fe2+ > Cu2+ > Fe3+.

  14. Shikonin Exerts Antitumor Activity via Proteasome Inhibition and Cell Death Induction in vitro and in vivo

    PubMed Central

    Yang, Huanjie; Zhou, Ping; Huang, Hongbiao; Chen, Di; Ma, Ningfang; Cui, Cindy Qiuzhi; Shen, Shouxing; Dong, Weihua; Zhang, Xiaoyan; Lian, Wen; Wang, Xuejun; Dou, Q. Ping; Liu, Jinbao

    2009-01-01

    Dysregulation of the ubiquitin-proteasome pathway plays an essential role in tumor growth and development. Shikonin, a natural naphthoquinone isolated from the traditional Chinese medicine Zi Cao (gromwell), has been reported to possess tumor cell-killing activity, and results from a clinical study using a shikonin-containing mixture demonstrated its safety and efficacy for the treatment of late-stage lung cancer. In the present study, we reported that shikonin is an inhibitor of tumor proteasome activity in vitro and in vivo. Our computational modeling predicts that the carbonyl carbons C1 and C4 of shikonin potentially interact with the catalytic site of β5 chymotryptic subunit of the proteasome. Indeed, shikonin potently inhibits the chymotrypsin-like activity of purified 20S proteasome (IC50 12.5 μmol/L) and tumor cellular 26S proteasome (IC50 between 2-16 μmol/L). Inhibition of the proteasome by shikonin in murine hepatoma H22, leukemia P388 and human prostate cancer PC-3 cultures resulted in accumulation of ubiquitinated proteins and several proteasome target proapoptotic proteins (IκB-α, Bax and p27), followed by induction of cell death. Shikonin treatment resulted in tumor growth inhibition in both H22 allografts and PC-3 xenografts, associated with suppression of the proteasomal activity and induction of cell death in vivo. Finally, shikonin treatment significantly prolonged the survival period of mice bearing P388 leukemia. Our results indicate that the tumor proteasome is one of the cellular targets of shikonin, and inhibition of the proteasome activity by shikonin contributes to its anti-tumor property. PMID:19165859

  15. Epithelial Cell Secretions from the Human Female Reproductive Tract Inhibit Sexually Transmitted Pathogens and Candida albicans but not Lactobacillus

    PubMed Central

    Wira, CR; Ghosh, M; Smith, JM; Shen, L; Connor, RI; Sundstrom, P; Frechette, Gregory M.; Hill, EM; Fahey, JV

    2011-01-01

    Female reproductive tract (FRT) epithelial cells protect against potential pathogens and sexually transmitted infections. The purpose of this study was to determine if epithelial cells from the upper FRT secrete antimicrobials that inhibit reproductive tract pathogens which threaten women's health. Apical secretions from primary cultures of Fallopian tube, uterine, cervical and ectocervical epithelial cells were incubated with Neisseria gonorrhoeae, Candida albicans (yeast and hyphal forms), HIV-1, and Lactobacillus crispatus, prior to being tested for their ability to grow and/or infect target cells. Epithelial cell secretions from the upper FRT inhibit N. gonorrhoeae and both forms of Candida, as well as reduce HIV-1 (R5) infection of target cells. In contrast, none had an inhibitory effect on L. crispatus. Cytokines and chemokines analysis in uterine secretions revealed several molecules that could account for pathogen inhibition. These findings provide definitive evidence for the critical role of epithelial cells in protecting the FRT from infections, without comprising the beneficial presence of L. crispatus, which is part of the normal vaginal microflora of humans. PMID:21048705

  16. Nitric Oxide Synthase (NOS) Inhibition during Porcine In Vitro Maturation Modifies Oocyte Protein S-Nitrosylation and In Vitro Fertilization

    PubMed Central

    Romero-Aguirregomezcorta, Jon; Santa, Ángela Patricia; García-Vázquez, Francisco Alberto; Coy, Pilar; Matás, Carmen

    2014-01-01

    Nitric oxide (NO) is a molecule involved in many reproductive processes. Its importance during oocyte in vitro maturation (IVM) has been demonstrated in various species although sometimes with contradictory results. The objective of this study was to determine the effect of NO during IVM of cumulus oocyte complexes and its subsequent impact on gamete interaction in porcine species. For this purpose, IVM media were supplemented with three NOS inhibitors: NG-nitro-L-arginine methyl ester (L-NAME), NG-monomethyl-L-arginine (L-NMMA) and aminoguanidine (AG). A NO donor, S-nitrosoglutathione (GSNO), was also used. The effects on the cumulus cell expansion, meiotic resumption, zona pellucida digestion time (ZPdt) and, finally, on in vitro fertilization (IVF) parameters were evaluated. The oocyte S-nitrosoproteins were also studied by in situ nitrosylation. The results showed that after 42 h of IVM, AG, L-NAME and L-NMMA had an inhibitory effect on cumulus cell expansion. Meiotic resumption was suppressed only when AG was added, with 78.7% of the oocytes arrested at the germinal vesicle state (P<0.05). Supplementation of the IVM medium with NOS inhibitors or NO donor did not enhance the efficiency of IVF, but revealed the importance of NO in maturation and subsequent fertilization. Furthermore, protein S-nitrosylation is reported for the first time as a pathway through which NO exerts its effect on porcine IVM; therefore, it would be important to determine which proteins are nitrosylated in the oocyte and their functions, in order to throw light on the mechanism of action of NO in oocyte maturation and subsequent fertilization. PMID:25542028

  17. Thiazolidinediones inhibit the growth of PC12 cells both in vitro and in vivo

    SciTech Connect

    Kim, Sang Wan; Choi, Ok Kyung; Chang, Mee Soo; Shin, Chan Soo; Park, Kyong Soo; Kim, Seong Yeon

    2008-06-27

    Thiazolidinediones (TZDs) have recently been proposed as a therapy for PPAR{gamma}-expressing tumors. Pheochromocytoma (PHEO) is associated with high morbidity and mortality due to excess catecholamine production, and few effective drug therapies currently exist. We investigated the effects of TZDs on PHEO both in vitro and in vivo. PPAR{gamma} protein was expressed in human adrenal PHEO tissues as well as in rat PHEO cells, PC12. TZDs, including rosiglitazone (RGZ) and pioglitazone (PGZ), inhibited proliferation of PC12 cells in a dose-dependent manner and increased casapse-3 expression of PC12 cells. TZDs also reduced expression of cyclin E and cyclin-dependent kinase2. RGZ inhibited nerve growth factor-induced neurite outgrowth and reduced expression of catecholamine-synthesizing enzymes. Finally, rat PHEO growth generated by subcutaneous injection of PC12 cells was slowed in an RGZ-treated mouse. These data suggest that TZDs may be a promising therapeutic approach for medical treatment for PHEO.

  18. Sildenafil inhibits the growth of human colorectal cancer in vitro and in vivo

    PubMed Central

    Mei, Xiao-Long; Yang, Yang; Zhang, Yao-Jun; Li, Yong; Zhao, Jin-Ming; Qiu, Jian-Ge; Zhang, Wen-Ji; Jiang, Qi-Wei; Xue, You-Qiu; Zheng, Di-Wei; Chen, Yao; Qin, Wu-Ming; Wei, Meng-Ning; Shi, Zhi

    2015-01-01

    Colorectal cancer is the third most common human cancer with frequent overexpression of the cGMP-specific phosphodiesterase 5 (PDE5). In the present study, we investigated that the anticancer effect of sildenafil on human colorectal cancer in vitro and in vivo, which is a potent and selective inhibitor of PDE5 for the treatment of erectile dysfunction and pulmonary arterial hypertension in the clinic. Sildenafil significantly induced cell growth inhibition, cell cycle arrest and apoptosis of human colorectal cancer with increased intracellular reactive oxidative specie (ROS) levels, which were accompanied by obvious alterations of related proteins such as CDKs, Cyclins and PARP etc. Pretreatment with ROS scavenger N-acetyl-L-cysteine could reverse sildenafil-induced ROS accumulation and cell apoptosis. Inhibition of the activity of protein kinase G with KT-5823 could enhance sildenafil-induced apoptosis. Furthermore, sildenafil caused the reduction of xenograft models of human colorectal cancer in nude mice. Overall, these findings suggest that sildenafil has the potential to be used for treatment of human colorectal cancer. PMID:26807313

  19. Curcumin and Boswellia serrata gum resin extract inhibit chikungunya and vesicular stomatitis virus infections in vitro.

    PubMed

    von Rhein, Christine; Weidner, Tatjana; Henß, Lisa; Martin, Judith; Weber, Christopher; Sliva, Katja; Schnierle, Barbara S

    2016-01-01

    Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes chikungunya fever and has infected millions of people mainly in developing countries. The associated disease is characterized by rash, high fever, and severe arthritis that can persist for years. CHIKV has adapted to Aedes albopictus, which also inhabits temperate regions including Europe and the United States of America. CHIKV has recently caused large outbreaks in Latin America. No treatment or licensed CHIKV vaccine exists. Traditional medicines are known to have anti-viral effects; therefore, we examined whether curcumin or Boswellia serrata gum resin extract have antiviral activity against CHIKV. Both compounds blocked entry of CHIKV Env-pseudotyped lentiviral vectors and inhibited CHIKV infection in vitro. In addition, vesicular stomatitis virus vector particles and viral infections were also inhibited to the same extent, indicating a broad antiviral activity. Although the bioavailability of these compounds is rather poor, they might be used as a lead structure to develop more effective antiviral drugs or might be used topically to prevent CHIKV spread in the skin after mosquito bites.

  20. Inhibition of serotonin release by bombesin-like peptides in rat hypothalamus in vitro

    SciTech Connect

    Saporito, M.S.; Warwick, R.O. Jr.

    1989-01-01

    We investigated the activity of bombesin (BN), neuromedin-C (NM-C) and neuromedin-B (NM-B) on serotonin (5-HT) release and reuptake in rat hypothalamus (HYP) in vitro. BN and NM-C but not NM-B decreased K/sup +/ evoked /sup 3/H-5-HT release from superfused HYP slices by 25%. Bacitracin, a nonspecific peptidase inhibitor, reversed the inhibitory effect of BN on K/sup +/ evoked /sup 3/H-5-HT release. Phosphoramidon (PAN, 10 /mu/M) an endopeptidase 24.11 inhibitor, abolished the inhibitory effect of BN, but not NM-C, on K/sup +/ evoked /sup 3/H-5-HT release. The peptidyl dipeptidase A inhibitor enalaprilat (ENP, 10 /mu/M), enhanced both BN and NM-C inhibition of /sup 3/H-5-HT release. Bestatin (BST, 10 /mu/M) had no effect on BN or NM-C inhibitory activity on /sup 3/H-5-HT release. Neither BN, NM-C nor NM-B affected reuptake of /sup 3/H-5-HT into HYP synaptosomes alone or in combination with any of the peptidase inhibitors, nor did these peptides alter the ability of fluoxetine to inhibit /sup 3/H-5-HT uptake.

  1. Chronic exposure of low dose salinomycin inhibits MSC migration capability in vitro

    PubMed Central

    SCHERZAD, AGMAL; HACKENBERG, STEPHAN; FROELICH, KATRIN; RAK, KRISTEN; HAGEN, RUDOLF; TAEGER, JOHANNES; BREGENZER, MAXIMILLIAN; KLEINSASSER, NORBERT

    2016-01-01

    Salinomycin is a polyether antiprotozoal antibiotic that is used as a food additive, particularly in poultry farming. By consuming animal products, there may be a chronic human exposure to salinomycin. Salinomycin inhibits the differentiation of preadipocytes into adipocytes. As human mesenchymal stem cells (MSC) may differentiate into different mesenchymal cells, it thus appeared worthwhile to investigate whether chronic salinomycin exposure impairs the functional properties of MSC and induces genotoxic effects. Bone marrow MSC were treated with low-dose salinomycin (100 nM) (MSC-Sal) for 4 weeks, while the medium containing salinomycin was changed every other day. Functional changes were evaluated and compared to MSC without salinomycin treatment (MSC-control). MSC-Sal and MSC-control were positive for cluster of differentiation 90 (CD90), CD73 and CD44, and negative for CD34. There were no differences observed in cell morphology or cytoskeletal structures following salinomycin exposure. The differentiation into adipocytes and osteocytes was not counteracted by salinomycin, and proliferation capability was not inhibited following salinomycin exposure. The migration of MSC-Sal was attenuated significantly as compared to the MSC-control. There were no genotoxic effects after 4 weeks of salinomycin exposure. The present study shows an altered migration capacity as a sign of functional impairment of MSC induced by chronic salinomycin exposure. Further in vitro toxicological investigations, particularly with primary human cells, are required to understand the impact of chronic salinomycin consumption on human cell systems. PMID:26998269

  2. Glycogen Synthase Kinase 3 Inhibition Promotes Adult Hippocampal Neurogenesis in Vitro and in Vivo

    PubMed Central

    2012-01-01

    Glycogen synthase kinase-3 (GSK-3) is a serine/threonine kinase originally identified as a regulator of glycogen metabolism but it also plays a pivotal role in numerous cellular functions, including differentiation, cell cycle regulation, and proliferation. The dentate gyrus of the hippocampus, together with the subventricular zone of the lateral ventricles, is one of the regions in which neurogenesis takes place in the adult brain. Here, using a chemical genetic approach that involves the use of several diverse inhibitors of GSK-3 as pharmacological tools, we show that inhibition of GSK-3 induces proliferation, migration, and differentiation of neural stem cells toward a neuronal phenotype in in vitro studies. Also, we demonstrate that inhibition of GSK-3 with the small molecule NP03112, called tideglusib, induces neurogenesis in the dentate gyrus of the hippocampus of adult rats. Taken together, our results suggest that GSK-3 should be considered as a new target molecule for modulating the production and integration of new neurons in the hippocampus as a treatment for neurodegenerative diseases or brain injury and, consequently, its inhibitors may represent new potential therapeutic drugs in neuroregenerative medicine. PMID:23173075

  3. Eritadenine from Edible Mushrooms Inhibits Activity of Angiotensin Converting Enzyme in Vitro.

    PubMed

    Afrin, Sadia; Rakib, Md Abdur; Kim, Boh Hyun; Kim, Jeong Ok; Ha, Yeong Lae

    2016-03-23

    The inhibition of angiotensin converting enzyme (ACE) activity was determined in vitro by mushroom-derived eritadenine (EA), which was analyzed in 11 principal Korean edible mushrooms. EA inhibited ACE activity with 0.091 μM IC50, whereas the IC50 of captopril (CP), which is a reference compound, was 0.025 μM. Kinetic measurements of ACE reaction in the substrate of hippuryl-l-histidyl-l-leucine (HHL) with or without EA revealed that the Vmax (0.0465 O.D/30 min) was unchanged, but the the Km increased from 2.063 to 3.887 mM, indicating that EA competes with HHL for the active site. When EA was analyzed by HPLC, Lentinus edodes with a soft cap contained the highest amount EA (642.8 mg%); however, Phellinus linteus with a hard cap contained the least amount of EA (9.4 mg%). These results indicate that EA was a strong competitive inhibitor for ACE, and edible mushrooms with soft caps contained a significant amount of EA.

  4. Binding of Galanthus nivalis lectin to Chlamydia trachomatis and inhibition of in vitro infection.

    PubMed

    Amin, K; Beillevaire, D; Mahmoud, E; Hammar, L; Mårdh, P A; Fröman, G

    1995-10-01

    A glycoprotein present in Chlamydia trachomatis, serotype L1, elementary bodies (EBs) was earlier found to bind the lectin from Galanthus nivalis (GNA). In the present paper we investigate the interaction of GNA with chlamydial EBs and its effect on in vitro infectivity. The binding affinity was studied with 125I-GNA lectin. Within 15 min about 80% maximal binding was obtained. The chlamydia-GNA interaction was inhibited by alpha-methylmannoside, causing a decrease of about 50% at 1 mM. Curve fit analyses indicated two types of binding sites for GNA on the EBs. The affinity to these differed by a factor of 15. The influence of the lectin on the ability of C. trachomatis to infect McCoy cells was also investigated. There was a GNA-dependent inhibition with a 50% reduction in the number of intracellular inclusions at 0.2 microM of the lectin. The findings indicate the presence of terminal mannose structures on the chlamydial surface at or in the proximity of the cell-binding domains. Mannose-binding proteins of eukaryotic cells could be important for the initial uptake of EBs.

  5. Sialic acid glycoproteins inhibit in vitro and in vivo replication of rotaviruses.

    PubMed Central

    Yolken, R H; Willoughby, R; Wee, S B; Miskuff, R; Vonderfecht, S

    1987-01-01

    We investigated the interactions of rotaviruses with glycoproteins and cells that support rotaviral replication. We found that a wide range of naturally occurring glycoproteins, including ovalbumins and ovomucoids from chicken and turkey eggs, and mucin derived from bovine submaxillary glands, inhibit the replication of rotaviruses in MA-104 cells. Our studies further indicated that the glycoproteins bind directly to rotaviruses and that virus-glycoprotein binding is dependent largely upon interactions with sialic acid oligosaccharides. We found that accessible sialic acid oligosaccharides are required for efficient rotavirus infection of MA-104 cells, thus demonstrating that sialic acid oligosaccharides play an important role in the interactions of rotaviruses with both glycoproteins and cells that support rotaviral replication. Bovine submaxillary mucin and chicken ovoinhibitor can also prevent the shedding of rotavirus antigen and the development of rotavirus gastroenteritis in a mouse model of rotavirus infection. Our findings document that a range of glycoproteins inhibit the in vivo and in vitro replication of rotaviruses and suggest that the alteration in the quantity or chemical composition of intestinal glycoproteins is a potential means for the modulation of enteric infections. Images PMID:3025257

  6. In vitro inhibition of protein synthesis by purified cores from vaccinia virus.

    PubMed Central

    Ben-Hamida, F; Beaud, G

    1978-01-01

    The mechanism of the shutoff of cellular protein synthesis in vaccinia virus-infected cells has been investigated by using in vitro systems. Purified vaccinia cores cause inhibition of endogenous mRNA translation in nonpreincubated reticulocyte lysates and Ehrlich ascites tumor cell-free systems. Translation of viral mRNA from turnip yellow mosaic virus is also impaired in wheat germ cell-free extracts. The block induced by vaccinia cores in protein synthesis is not due to a decrease in the availability of mRNA but rather to an alteration of the cellular translational machinery. No nucleolytic activity able of digesting mRNA could be detected in purified vaccinia cores with three sensitive tests. There is a lack of inhibition in the poly(Phe)-poly(U) system, which bypasses the normal initiation process. An almost complete disaggregation of polyribosomes in the reticulocyte lysate appears when vaccinia cores are present. These results indicate that mRNA translation in a cell-free system is affected predominantly at the level of polypeptide chain initiation. PMID:272632

  7. Glycogen synthase kinase 3 inhibition promotes adult hippocampal neurogenesis in vitro and in vivo.

    PubMed

    Morales-Garcia, Jose A; Luna-Medina, Rosario; Alonso-Gil, Sandra; Sanz-Sancristobal, Marina; Palomo, Valle; Gil, Carmen; Santos, Angel; Martinez, Ana; Perez-Castillo, Ana

    2012-11-21

    Glycogen synthase kinase-3 (GSK-3) is a serine/threonine kinase originally identified as a regulator of glycogen metabolism but it also plays a pivotal role in numerous cellular functions, including differentiation, cell cycle regulation, and proliferation. The dentate gyrus of the hippocampus, together with the subventricular zone of the lateral ventricles, is one of the regions in which neurogenesis takes place in the adult brain. Here, using a chemical genetic approach that involves the use of several diverse inhibitors of GSK-3 as pharmacological tools, we show that inhibition of GSK-3 induces proliferation, migration, and differentiation of neural stem cells toward a neuronal phenotype in in vitro studies. Also, we demonstrate that inhibition of GSK-3 with the small molecule NP03112, called tideglusib, induces neurogenesis in the dentate gyrus of the hippocampus of adult rats. Taken together, our results suggest that GSK-3 should be considered as a new target molecule for modulating the production and integration of new neurons in the hippocampus as a treatment for neurodegenerative diseases or brain injury and, consequently, its inhibitors may represent new potential therapeutic drugs in neuroregenerative medicine.

  8. Autophagy inhibition radiosensitizes in vitro, yet reduces radioresponses in vivo due to deficient immunogenic signalling

    PubMed Central

    Ko, A; Kanehisa, A; Martins, I; Senovilla, L; Chargari, C; Dugue, D; Mariño, G; Kepp, O; Michaud, M; Perfettini, J-L; Kroemer, G; Deutsch, E

    2014-01-01

    Clinical oncology heavily relies on the use of radiotherapy, which often leads to merely transient responses that are followed by local or distant relapse. The molecular mechanisms explaining radioresistance are largely elusive. Here, we identified a dual role of autophagy in the response of cancer cells to ionizing radiation. On one hand, we observed that the depletion of essential autophagy-relevant gene products, such as ATG5 and Beclin 1, increased the sensitivity of human or mouse cancer cell lines to irradiation, both in vitro (where autophagy inhibition increased radiation-induced cell death and decreased clonogenic survival) and in vivo, after transplantation of the cell lines into immunodeficient mice (where autophagy inhibition potentiated the tumour growth-inhibitory effect of radiotherapy). On the other hand, when tumour proficient or deficient for autophagy were implanted in immunocompetent mice, it turned out that defective autophagy reduced the efficacy of radiotherapy. Indeed, radiotherapy elicited an anti-cancer immune response that was dependent on autophagy-induced ATP release from stressed or dying tumour cells and was characterized by dense lymphocyte infiltration of the tumour bed. Intratumoural injection of an ecto-ATPase inhibitor restored the immune infiltration of autophagy-deficient tumours post radiotherapy and improved the growth-inhibitory effect of ionizing irradiation. Altogether, our results reveal that beyond its cytoprotective function, autophagy confers immunogenic properties to tumours, hence amplifying the efficacy of radiotherapy in an immunocompetent context. This has far-reaching implications for the development of pharmacological radiosensitizers. PMID:24037090

  9. Curcumin and Boswellia serrata gum resin extract inhibit chikungunya and vesicular stomatitis virus infections in vitro.

    PubMed

    von Rhein, Christine; Weidner, Tatjana; Henß, Lisa; Martin, Judith; Weber, Christopher; Sliva, Katja; Schnierle, Barbara S

    2016-01-01

    Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes chikungunya fever and has infected millions of people mainly in developing countries. The associated disease is characterized by rash, high fever, and severe arthritis that can persist for years. CHIKV has adapted to Aedes albopictus, which also inhabits temperate regions including Europe and the United States of America. CHIKV has recently caused large outbreaks in Latin America. No treatment or licensed CHIKV vaccine exists. Traditional medicines are known to have anti-viral effects; therefore, we examined whether curcumin or Boswellia serrata gum resin extract have antiviral activity against CHIKV. Both compounds blocked entry of CHIKV Env-pseudotyped lentiviral vectors and inhibited CHIKV infection in vitro. In addition, vesicular stomatitis virus vector particles and viral infections were also inhibited to the same extent, indicating a broad antiviral activity. Although the bioavailability of these compounds is rather poor, they might be used as a lead structure to develop more effective antiviral drugs or might be used topically to prevent CHIKV spread in the skin after mosquito bites. PMID:26611396

  10. Lactobacillus crispatus inhibits the infectivity of Chlamydia trachomatis elementary bodies, in vitro study

    PubMed Central

    Nardini, Paola; Ñahui Palomino, Rogers Alberto; Parolin, Carola; Laghi, Luca; Foschi, Claudio; Cevenini, Roberto; Vitali, Beatrice; Marangoni, Antonella

    2016-01-01

    Lactobacillus species dominate the vaginal microbiota of healthy reproductive-age women and protect the genitourinary tract from the attack of several infectious agents. Chlamydia trachomatis, a leading cause of sexually transmitted disease worldwide, can induce severe sequelae, i.e. pelvic inflammatory disease, infertility and ectopic pregnancy. In the present study we investigated the interference of Lactobacillus crispatus, L. gasseri and L. vaginalis, known to be dominant species in the vaginal microbiome, with the infection process of C. trachomatis. Lactobacilli exerted a strong inhibitory effect on Chlamydia infectivity mainly through the action of secreted metabolites in a concentration/pH dependent mode. Short contact times were the most effective in the inhibition, suggesting a protective role of lactobacilli in the early steps of Chlamydia infection. The best anti-Chlamydia profile was shown by L. crispatus species. In order to delineate metabolic profiles related to anti-Chlamydia activity, Lactobacillus supernatants were analysed by 1H-NMR. Production of lactate and acidification of the vaginal environment seemed to be crucial for the activity, in addition to the consumption of the carbonate source represented by glucose. The main conclusion of this study is that high concentrations of L. crispatus inhibit infectivity of C. trachomatis in vitro. PMID:27354249

  11. CHLORHEXIDINE INHIBITS L1 CELL ADHESION MOLECULE MEDIATED NEURITE OUTGROWTH IN VITRO

    PubMed Central

    Milstone, Aaron M.; Bamford, Penny; Aucott, Susan W.; Tang, Ningfeng; White, Kimberly R.; Bearer, Cynthia F.

    2013-01-01

    Background Chlorhexidine is a skin disinfectant that reduces skin and mucous membrane bacterial colonization and inhibits organism growth. Despite numerous studies assessing chlorhexidine safety in term infants, residual concerns have limited its use in hospitalized neonates, especially low birth weight preterm infants. The aim of this study was to assess the potential neurotoxicity of chlorhexidine on the developing central nervous system using a well-established in vitro model of neurite outgrowth that includes laminin and L1 cell adhesion molecule (L1) as neurite outgrowth promoting substrates. Methods Cerebellar granule neurons are plated on either poly L-lysine, L1 or laminin. Chlorhexidine, hexachlorophene or their excipients are added to the media. Neurons are grown for 24 h, then fixed and neurite length measured. Results Chlorhexidine significantly reduced the length of neurites grown on L1 but not laminin. Chlorhexidine concentrations as low as 125 ng/ml statistically significantly reduced neurite length on L1. Hexachlorophene did not affect neurite length. Conclusion Chlorhexidine at concentrations detected in the blood following topical applications in preterm infants specifically inhibited L1 mediated neurite outgrowth of cerebellar granule neurons. It is now vital to determine whether the blood brain barrier is permeable to chlorhexidine in preterm infants. PMID:24126818

  12. Emergence of Resistance of Candida albicans to Clotrimazole in Human Immunodeficiency Virus-Infected Children: In Vitro and Clinical Correlations

    PubMed Central

    Pelletier, René; Peter, Joanne; Antin, Cynthia; Gonzalez, Corina; Wood, Lauren; Walsh, Thomas J.

    2000-01-01

    Oropharyngeal candidiasis (OPC) is a common opportunistic infection in human immunodeficiency virus (HIV)-infected patients and other immunocompromised hosts. Clotrimazole troches are widely used in the treatment of mucosal candidiasis. However, little is known about the potential contribution of clotrimazole resistance to the development of refractory mucosal candidiasis. We therefore investigated the potential emergence of resistance to clotrimazole in a prospectively monitored HIV-infected pediatric population receiving this azole. Adapting the National Committee for Clinical Laboratory Standards M27-A reference method for broth antifungal susceptibility testing of yeasts to clotrimazole, we compared MICs in macrodilution and microdilution assays. We further analyzed the correlation between these in vitro findings and the clinical response to antifungal therapy. One isolate from each of 87 HIV-infected children was studied by the macrodilution and microdilution methods. Two inoculum sizes were tested by the macrodilution method (103 and 104 CFU/ml) in order to assess the effect of inoculum size on clotrimazole MICs. The same isolates also were tested using a noncolorimetric microdilution method. Clotrimazole concentrations ranged from 0.03 to 16 μg/ml. Readings were performed after incubation for 24 and 48 h at 35°C. For 62 (71.2%) of 87 clinical isolates, the MICs were low (≤0.06 μg/ml). The MIC for 90% of the strains tested was 0.5 μg/ml, and the highest MIC was 8 μg/ml. There was no significant difference between MICs at the two inoculum sizes. There was 89% agreement (±1 tube) between the microdilution method at 24 h and the macrodilution method at 48 h. If the MIC of clotrimazole for an isolate of C. albicans was ≥0.5 μg/ml, there was a significant risk (P < 0.001) of cross-resistance to other azoles: fluconazole, ≥64 μg/ml (relative risk [RR] = 8.9); itraconazole, ≥1 μg/ml (RR = 10). Resistance to clotrimazole was highly associated with

  13. In vitro activity of essential oils extracted from plants used as spices against fluconazole-resistant and fluconazole-susceptible Candida spp.

    PubMed

    Pozzatti, Patrícia; Scheid, Liliane Alves; Spader, Tatiana Borba; Atayde, Margareth Linde; Santurio, Janio Morais; Alves, Sydney Hartz

    2008-11-01

    In the present study, the antifungal activity of selected essential oils obtained from plants used as spices was evaluated against both fluconazole-resistant and fluconazole-susceptible Candida spp. The Candida species studied were Candida albicans, Candida dubliniensis, Candida tropicalis, Candida glabrata, and Candida krusei. For comparison purposes, they were arranged in groups as C. albicans, C. dubliniensis, and Candida non-albicans. The essential oils were obtained from Cinnamomum zeylanicum Breyn, Lippia graveolens HBK, Ocimum basilicum L., Origanum vulgare L., Rosmarinus officinalis L., Salvia officinalis L., Thymus vulgaris L., and Zingiber officinale. The susceptibility tests were based on the M27-A2 methodology. The chemical composition of the essential oils was obtained by gas chromatography-mass spectroscopy and by retention indices. The results showed that cinnamon, Mexican oregano, oregano, thyme, and ginger essential oils have different levels of antifungal activity. Oregano and ginger essential oils were found to be the most and the least efficient, respectively. The main finding was that the susceptibilities of fluconazole-resistant C. albicans, C. dubliniensis, and Candida non-albicans to Mexican oregano, oregano, thyme, and ginger essential oils were higher than those of the fluconazole-susceptible yeasts (P<0.05). In contrast, fluconazole-resistant C. albicans and Candida non-albicans were less susceptible to cinnamon essential oil than their fluconazole-susceptible counterparts (P<0.05). A relationship between the yeasts' susceptibilities and the chemical composition of the essential oils studied was apparent when these 2 parameters were compared. Finally, basil, rosemary, and sage essential oils did not show antifungal activity against Candida isolates at the tested concentrations. PMID:18997851

  14. Cancer drugs inhibit morphogenesis in the human fungal pathogen, Candida albicans.

    PubMed

    Routh, Madhushree M; Chauhan, Nitin M; Karuppayil, S Mohan

    2013-01-01

    Candida infections are very common in cancer patients and it is a common practice to prescribe antifungal antibiotics along with anticancer drugs. Yeast to hyphal form switching is considered to be important in invasive candidiasis. Targeting morphogenetic switching may be useful against invasive candidiasis. In this study, we report the antimorphogenetic properties of thirty cancer drugs. PMID:24516452

  15. Ischemic postconditioning inhibits apoptosis in an in vitro proximal tubular cell model

    PubMed Central

    WENG, XIAODONG; WANG, LEI; CHEN, HUI; LIU, XIUHENG; QIU, TAO; CHEN, ZHIYUAN

    2015-01-01

    Ischemia-reperfusion is a common injury of clinical ischemic disease and surgical lesions. Ischemic postconditioning (IPO) improves the ability of organs subjected to ischemia to tolerate injury. However, renal IPO studies have been based on animal models. In order to gain insights into IPO-induced alterations at the cellular level, an in vitro model for IPO was designed using the rat proximal tubular cell line NRK-52E. This model was established by placing NRK-52E cells in ischemic conditions for 3 h, then exposing cells to three cycles of reperfusion for 10 min and finally to ischemic conditions for 10 min (postconditioning). The cells were cultured further in reperfusion conditions for 3, 6, 12 and 24 h. Flow cytometry and Hoechst were used to assess apoptosis. The protein expression of B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), caspase-3, cleaved caspase-3 and caspase-8 were analyzed by western blotting. The results demonstrated that apoptosis occurred in cells subjected to ischemia/reperfusion (I/R) alone or with postconditioning following reperfusion for 24 h. Cells subjected to I/R demonstrated increased expression of Bax, cleaved caspase-3 and caspase-8 at the end of reperfusion. However, the levels of Bax, cleaved caspase-3 and caspase-8 were significantly attenuated in cells, which had undergone IPO. In conclusion, apoptosis was observed in cells subjected to 3 h of ischemia-reperfusion injury and IPO was able to inhibit this apoptosis. IPO inhibited apoptosis by inhibiting the caspase pathway thereby exerting protective effects. PMID:25672392

  16. Ischemic postconditioning inhibits apoptosis in an in vitro proximal tubular cell model.

    PubMed

    Weng, Xiaodong; Wang, Lei; Chen, Hui; Liu, Xiuheng; Qiu, Tao; Chen, Zhiyuan

    2015-07-01

    Ischemia-reperfusion is a common injury of clinical ischemic disease and surgical lesions. Ischemic postconditioning (IPO) improves the ability of organs subjected to ischemia to tolerate injury. However, renal IPO studies have been based on animal models. In order to gain insights into IPO-induced alterations at the cellular level, an in vitro model for IPO was designed using the rat proximal tubular cell line NRK-52 E. This model was established by placing NRK-52 E cells in ischemic conditions for 3 h, then exposing cells to three cycles of reperfusion for 10 min and finally to ischemic conditions for 10 min (postconditioning). The cells were cultured further in reperfusion conditions for 3, 6, 12 and 24 h. Flow cytometry and Hoechst were used to assess apoptosis. The protein expression of B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), caspase-3, cleaved caspase-3 and caspase-8 were analyzed by western blotting. The results demonstrated that apoptosis occurred in cells subjected to ischemia/reperfusion (I/R) alone or with postconditioning following reperfusion for 24 h. Cells subjected to I/R demonstrated increased expression of Bax, cleaved caspase-3 and caspase-8 at the end of reperfusion. However, the levels of Bax, cleaved caspase-3 and caspase-8 were significantly attenuated in cells, which had undergone IPO. In conclusion, apoptosis was observed in cells subjected to 3 h of ischemia-reperfusion injury and IPO was able to inhibit this apoptosis. IPO inhibited apoptosis by inhibiting the caspase pathway thereby exerting protective effects. PMID:25672392

  17. Effect of CO2 laser on root caries inhibition around composite restorations: an in vitro study.

    PubMed

    de Melo, Jociana Bandeira; Hanashiro, Fernando Seishim; Steagall, Washington; Turbino, Miriam Lacalle; Nobre-dos-Santos, Marinês; Youssef, Michel Nicolau; de Souza-Zaroni, Wanessa Christine

    2014-03-01

    The aim of the present study was to investigate the in vitro effect of CO2 laser on the inhibition of root surface demineralization around composite resin restorations. For this purpose, 30 blocks obtained from human molar roots were divided into three groups: group 1 (negative control), cavity prepared with cylindrical diamond bur + acid etching + adhesive + composite resin restoration; group 2, cavity prepared with cylindrical diamond bur + CO2 laser (5.0 J/cm(2)) + acid etching + adhesive + composite resin; and group 3, cavity prepared with cylindrical diamond bur + CO2 laser (6.0 J/cm(2)) + acid etching + adhesive + composite resin. After this procedure, the blocks were submitted to thermal and pH cycling. Root surface demineralization around the restorations was measured by microhardness analysis. The hardness results of the longitudinally sectioned root surface were converted into percentage of mineral volume, which was used to calculate the mineral loss delta Z (ΔZ). The percentage of mineral volume, ΔZ, and the percentage of demineralization inhibition of the groups were statistically analyzed by using analysis of variance and Tukey-Kramer test. The percentage of mineral volume was higher in the irradiated groups up to 80 μm deep. The ΔZ was significantly lower in the irradiated groups than in the control group. The percentage of reduction in demineralization ranged from 19.73 to 29.21 in position 1 (50 μm), and from 24.76 to 26.73 in position 2 (100 μm), when using 6 and 5 J/cm(2), respectively. The CO2 laser was effective in inhibiting root demineralization around composite resin restorations. PMID:23291879

  18. Compound C inhibits in vitro angiogenesis and ameliorates thrombin-induced endothelial barrier failure.

    PubMed

    Gündüz, Dursun; Klewer, Matthias; Bauer, Pascal; Tanislav, Christian; Sedding, Daniel; Rohrbach, Susanne; Schulz, Rainer; Aslam, Muhammad

    2015-12-01

    Compound C (comp. C) is a cell-permeable pyrrazolopyrimidine derivative and widely used as adenosine monophosphate-activated protein kinase (AMPK) inhibitor to characterise the role of AMPK in various physiological processes. However, its AMPK-independent effects have also been reported. In the present study we investigated the effects of moderate dose (1-10μM) comp. C on endothelial cell (EC) proliferation, in vitro angiogenesis, and endothelial barrier function. Comp. C was unable to inhibit AMPK phosphorylation (activation) induced by metformin and A-769662 in ECs even at concentration of 10μM. At lower concentration (1μM), comp. C inhibited and potentiated the inhibitory effects of metformin and A-769662 on EC proliferation, migration, tube formation, and sprouting without inducing apoptosis. However, at higher concentration (10μM), it strongly induced apoptosis as measured by enhanced caspase 3/7 activity. Moreover, comp. C antagonised thrombin-induced EC hyperpermeability accompanied by activation of Rac1 and strengthening of adherens junctions (AJs). This EC barrier protective effect was not affected by the presence of AMPK activators. The data of the present study demonstrate that long-term treatment of ECs with low concentration comp. C inhibits EC proliferation and angiogenesis without induction of apoptosis. While short-term incubation antagonises thrombin-induced EC hyperpermeability presumably via Rac1-dependent strengthening of AJs. Furthermore, higher concentration of comp. C (10μM or above) is toxic for ECs and warns that this agent should be used with caution to demonstrate the AMPK-mediated effects. PMID:26522925

  19. Novel Triazine JPC-2067-B Inhibits Toxoplasma gondii In Vitro and In Vivo

    PubMed Central

    Mui, Ernest J.; Schiehser, Guy A.; Milhous, Wilbur K.; Hsu, Honghue; Roberts, Craig W.; Kirisits, Michael; Muench, Stephen; Rice, David; Dubey, J. P.; Fowble, Joseph W.; Rathod, Pradipsinh K.; Queener, Sherry F.; Liu, Susan R.; Jacobus, David P.; McLeod, Rima

    2008-01-01

    Background and Methodology Toxoplasma gondii causes substantial morbidity, mortality, and costs for healthcare in the developed and developing world. Current medicines are not well tolerated and cause hypersensitivity reactions. The dihydrotriazine JPC-2067-B (4, 6-diamino-1, 2-dihydro-2, 2-dimethyl-1-(3′(2-chloro-, 4-trifluoromethoxyphenoxy)propyloxy)-1, 3, 5-triazine), which inhibits dihydrofolate reductase (DHFR), is highly effective against Plasmodium falciparum, Plasmodium vivax, and apicomplexans related to T. gondii. JPC-2067-B is the primary metabolite of the orally active biguanide JPC-2056 1-(3′-(2-chloro-4-trifluoromethoxyphenyloxy)propyl oxy)- 5-isopropylbiguanide, which is being advanced to clinical trials for malaria. Efficacy of the prodrug JPC-2056 and the active metabolite JPC-2067-B against T. gondii and T. gondii DHFR as well as toxicity toward mammalian cells were tested. Principal Findings and Conclusions Herein, we found that JPC-2067-B is highly effective against T. gondii. We demonstrate that JPC-2067-B inhibits T. gondii growth in culture (IC50 20 nM), inhibits the purified enzyme (IC50 6.5 nM), is more efficacious than pyrimethamine, and is cidal in vitro. JPC-2067-B administered parenterally and the orally administered pro-drug (JPC-2056) are also effective against T. gondii tachyzoites in vivo. A molecular model of T. gondii DHFR-TS complexed with JPC-2067-B was developed. We found that the three main parasite clonal types and isolates from South and Central America, the United States, Canada, China, and Sri Lanka have the same amino acid sequences preserving key binding sites for the triazine. Significance JPC-2056/JPC-2067-B have potential to be more effective and possibly less toxic treatments for toxoplasmosis than currently available medicines. PMID:18320016

  20. In vitro atrazine-exposure inhibits human natural killer cell lytic granule release

    SciTech Connect

    Rowe, Alexander M.; Brundage, Kathleen M.; Barnett, John B. . E-mail: jbarnett@hsc.wvu.edu

    2007-06-01

    The herbicide atrazine is a known immunotoxicant and an inhibitor of human natural killer (NK) cell lytic function. The precise changes in NK cell lytic function following atrazine exposure have not been fully elucidated. The current study identifies the point at which atrazine exerts its affect on the stepwise process of human NK cell-mediated lyses of the K562 target cell line. Using intracellular staining of human peripheral blood lymphocytes, it was determined that a 24-h in vitro exposure to atrazine did not decrease the level of NK cell lytic proteins granzyme A, granzyme B or perforin. Thus, it was hypothesized that atrazine exposure was inhibiting the ability of the NK cells to bind to the target cell and subsequently inhibit the release of lytic protein from the NK cell. To test this hypothesis, flow cytometry and fluorescent microscopy were employed to analyze NK cell-target cell co-cultures following atrazine exposure. These assays demonstrated no significant decrease in the level of target cell binding. However, the levels of NK intracellular lytic protein retained and the amount of lytic protein released were assessed following a 4-h incubation with K562 target cells. The relative level of intracellular lytic protein was 25-50% higher, and the amount of lytic protein released was 55-65% less in atrazine-treated cells than vehicle-treated cells following incubation with the target cells. These results indicate that ATR exposure inhibits the ability of NK cells to lyse target cells by blocking lytic granule release without affecting the ability of the NK cell to form stable conjugates with target cells.

  1. Selenium and vitamin E inhibit radiogenic and chemically induced transformation in vitro via different mechanisms

    SciTech Connect

    Borek, C.; Ong, A.; Mason, H.; Donahue, L.; Biaglow, J.E.

    1986-03-01

    Results from in vivo and in vitro studies showing that antioxidants may act as anticarcinogens support the role of active oxygen in carcinogenesis and provide impetus for exploring the functions of dietary antioxidants in cancer prevention by using in vitro models. The authors examined the single and combined effects of selenium, a component of glutathione peroxidase, and vitamin E, a known antioxidant, on cell transformation induced in C3H/10T-1/2 cells by x-rays, benzo(a)pyrene, or tryptophan pyrolysate and on the levels of cellular scavenging systems peroxide destruction. Incubation of C3H/10T-1/2 cells with 2.5 ..mu..M Na/sup 2/SeO/sup 3/ (selenium) or with 7 ..mu..M ..cap alpha..-tocopherol succinate (vitamin E) 24 hr prior to exposure to x-rays or the chemical carcinogens resulted in an inhibition of transformation by each of the antioxidants with an additive-inhibitory action when the two nutrients were combined. Cellular pretreatment with selenium resulted in increased levels of cellular glutathione peroxidase, catalase, and nonprotein thiols (glutathione) and in an enhanced destruction of peroxide. The results support our earlier studies showing that free radical-mediated events play a role in radiation and chemically induced transformation. They indicate that selenium and vitamin E act alone and in additive fashion as radioprotecting and chemopreventing agents. The results further suggest that selenium confers protection in part by inducing or activating cellular free-radical scavenging systems and by enhancing peroxide breakdown while vitamin E appears to confer its protection by and alternate complementary mechanism.

  2. Comparison of a spectrophotometric microdilution method with RPMI-2% glucose with the National Committee for Clinical Laboratory Standards reference macrodilution method M27-P for in vitro susceptibility testing of amphotericin B, flucytosine, and fluconazole against Candida albicans.

    PubMed Central

    Rodríguez-Tudela, J L; Berenguer, J; Martínez-Suárez, J V; Sanchez, R

    1996-01-01

    The National Committee for Clinical Laboratory Standards has proposed a reference broth macrodilution method for in vitro antifungal susceptibility testing of yeasts (the M27-P method). This method is cumbersome and time-consuming and includes MIC endpoint determination by visual and subjective inspection of growth inhibition after 48 h of incubation. An alternative microdilution procedure was compared with the M27-P method for determination of the amphotericin B, flucytosine, and fluconazole susceptibilities of 8 American Type Culture Collection strains (6 of them were quality control or reference strains) and 50 clinical isolates of candida albicans. This microdilution method uses as culture medium RPMI 1640 supplemented with 18 g of glucose per liter (RPMI-2% glucose). Preparation of drugs, basal medium, and inocula was done by following the recommendations of the National Committee for Clinical Laboratory Standards. The MIC endpoint was calculated objectively from the turbidimetric data read at 24 h. Increased growth of C. albicans in RPMI-2% glucose and its spectrophotometric reading allowed for the rapid (24 h) and objective calculation of MIC endpoints compared with previous microdilution methods with standard RPMI 1640. Nevertheless, good agreement was shown between the M27-P method and this microdilution test. The MICs obtained for the quality control or reference strains by the microdilution method were in the ranges published for those strains. For clinical isolates, the percentages of agreement were 100% for amphotericin B and fluconazole and 98.1% for flucytosine. These data suggest that this microdilution method may serve as a less subjective and more rapid alternative to the M27-P method for antifungal susceptibility testing of yeasts. PMID:8878570

  3. INHIBITION OF IN VITRO FERTILIZATION IN THE HAMSTER BY ANTIBODIES RAISED AGAINST THE RAT SPERM PROTEIN SP22

    EPA Science Inventory

    INHIBITION OF IN VITRO FERTILIZATION IN THE HAMSTER BY ANTIBODIES RAISED AGAINST THE RAT SPERM PROTEIN SP22. SC Jeffay*, SD Perreault, KL Bobseine*, JE Welch*, GR Klinefelter, US EPA, Research Triangle Park, NC.
    SP22, a rat sperm membrane protein that is highly-correlated w...

  4. Adherence of Candida to cultured vascular endothelial cells: mechanisms of attachment and endothelial cell penetration.

    PubMed

    Rotrosen, D; Edwards, J E; Gibson, T R; Moore, J C; Cohen, A H; Green, I

    1985-12-01

    To elucidate the pathogenesis of hematogenous Candida infections, we developed an in vitro model of Candida adherence to and penetration of human endothelial cells. We enhanced or inhibited adherence in order to probe mechanisms of attachment. Adherence of Candida albicans showed a linear relation to Candida inoculum (range, 10(2)-10(5) cfu, r = .99, P less than .01) and exceeded that of less virulent Candida species and that of Saccharomyces cerevisiae (P less than .01). Candida immune serum blocked attachment (greater than 95% inhibition; P less than .001), however, this activity was abolished by immunoprecipitation of immune serum with C. albicans mannan (P less than .001) and was unaffected by immunoprecipitation with S. cerevisiae mannan or by adsorption with particulate chitin. Adherence was diminished by exposing C. albicans to heat (greater than 99% inhibition; P less than .01), UV light (98% inhibition; P less than .01), or sodium periodate (greater than 72% inhibition; P less than .01). An extract from heat-exposed C. albicans blocked adherence (greater than 51% inhibition; P less than .001). Transmission electron microscopy demonstrated that viable or killed Candida organisms were attached to endothelial cells, were enveloped by membrane processes from the endothelial cell surface, and were incorporated into the endothelial cells within phagosomes. Cytochalasin B blocked incorporation without blocking surface attachment. PMID:3905987

  5. Comparison of Results Obtained by Testing with Three Different Agar Media and by the NCCLS M27-A Method for In Vitro Testing of Fluconazole against Candida spp.

    PubMed Central

    Rubio, M. Carmen; Gil, Joaquina; de Ocáriz, Inmaculada Ramírez; Benito, Rafael; Rezusta, Antonio

    2003-01-01

    Fluconazole susceptibilities of 150 Candida isolates were determined by a 25-μg fluconazole disk diffusion agar test and compared with the microdilution NCCLS M27-A method. The agar test used three different media and was read at 24 and 48 h. When only the susceptible and nonsusceptible categories were used, disk diffusion with Müeller-Hinton agar supplemented with 2% glucose and 0.5 μg of methylene blue (MHGM) per ml had a 95.37% correlation with the MIC method at 24 h, followed by RPMI 1640-2% of glucose agar (correlation, 94%) and Shadomy medium (SHDM) (correlation, 92.6%). The growth of microcolonies inside the inhibition zones was common (>63%) in the RPMI and SHDM media and minimal with MHGM (8.7%). At 48 h, MHGM and SHDM still had a >91% correlation with the MIC, while RPMI results had dropped to 75%. The best overall agreement was obtained with C. dubliniensis (100%). PMID:12791899

  6. Flavonoids casticin and chrysosplenol D from Artemisia annua L. inhibit inflammation in vitro and in vivo

    SciTech Connect

    Li, Yu-Jie; Guo, Yan; Yang, Qing; Weng, Xiao-Gang; Yang, Lan; Wang, Ya-Jie; Chen, Ying; Zhang, Dong; Li, Qi; Liu, Xu-Cen; Kan, Xiao-Xi; Chen, Xi; Zhu, Xiao-Xin; Kmoníèková, Eva; Zídek, Zdenìk

    2015-08-01

    Background: The aim of our experiments was to investigate the anti-inflammatory properties of casticin and chrysosplenol D, two flavonoids present in Artemisia annua L. Methods: Topical inflammation was induced in ICR mice using croton oil. Mice were then treated with casticin or chrysosplenol D. Cutaneous histological changes and edema were assessed. ICR mice were intragastrically administrated with casticin or chrysosplenol D followed by intraperitoneal injection of lipopolysaccharide (LPS). Mouse Raw264.7 macrophage cells were incubated with casticin or chrysosplenol D. Intracellular phosphorylation was detected, and migration was assessed by trans-well assay. HT-29/NFκB-luc cells were incubated with casticin or chrysosplenol D in the presence or absence of LPS, and NF-κB activation was quantified. Results: In mice, administration of casticin (0.5, 1 and 1.5 μmol/cm{sup 2}) and chrysosplenol D (1 and 1.5 μmol/cm{sup 2}) inhibited croton oil-induced ear edema (casticin: 29.39–64.95%; chrysosplenol D: 37.76–65.89%, all P < 0.05) in a manner similar to indomethacin (0.5, 1 and 1.5 μmol/cm{sup 2}; 55.63–84.58%). Casticin (0.07, 0.13 and 0.27 mmol/kg) and chrysosplenol D (0.07, 0.14 and 0.28 mmol/kg) protected against LPS-induced systemic inflammatory response syndrome (SIRS) in mice (all P < 0.05), in a manner similar to dexamethasone (0.03 mmol/kg). Casticin and chrysosplenol D suppressed LPS-induced release of IL-1 beta, IL-6 and MCP-1, inhibited cell migration, and reduced LPS-induced IκB and c-JUN phosphorylation in Raw264.7 cells. JNK inhibitor SP600125 blocked the inhibitory effect of chrysosplenol D on cytokine release. Conclusions: The flavonoids casticin and chrysosplenol D from A. annua L. inhibited inflammation in vitro and in vivo. - Highlights: • We report a new activity of the flavonoids present in Artemisia annua L. • These flavonoids inhibit croton oil-induced ear edema in mice. • These flavonoids protect against LPS-induced SIRS in

  7. Inhibition of Influenza A Virus Infection In Vitro by Peptides Designed In Silico

    PubMed Central

    López-Martínez, Rogelio; Ramírez-Salinas, G. Lizbeth; Correa-Basurto, José; Barrón, Blanca L.

    2013-01-01

    Influenza A viruses are enveloped, segmented negative single-stranded RNA viruses, capable of causing severe human respiratory infections. Currently, only two types of drugs are used to treat influenza A infections, the M2 H+ ion channel blockers (amantadine and rimantadine) and the neuraminidase inhibitors (NAI) (oseltamivir and zanamivir). Moreover, the emergence of drug-resistant influenza A virus strains has emphasized the need to develop new antiviral agents to complement or replace the existing drugs. Influenza A virus has on the surface a glycoprotein named hemagglutinin (HA) which due to its important role in the initial stage of infection: receptor binding and fusion activities of viral and endosomal membranes, is a potential target for new antiviral drugs. In this work we designed nine peptides using several bioinformatics tools. These peptides were derived from the HA1 and HA2 subunits of influenza A HA with the aim to inhibit influenza A virus infection. The peptides were synthetized and their antiviral activity was tested in vitro against several influenza A viral strains: Puerto Rico/916/34 (H1N1), (H1N1)pdm09, swine (H1N1) and avian (H5N2). We found these peptides were able to inhibit the influenza A viral strains tested, without showing any cytotoxic effect. By docking studies we found evidence that all the peptides were capable to bind to the viral HA, principally to important regions on the viral HA stalk, thus could prevent the HA conformational changes required to carry out its membranes fusion activity. PMID:24146939

  8. Inhibition of RNA synthesis in vitro and cell growth by anthracycline antibiotics.

    PubMed

    Studzian, K; Wasowska, M; Piestrzeniewicz, M K; Wilmańska, D; Szmigiero, L; Oszczapowicz, I; Gniazdowski, M

    2001-01-01

    New derivatives of doxorubicin and daunorubicin with amidine group bonded to daunosamine at C-3' atom and bearing the morpholine ring attached to the amidine group have been recently synthesized. Their cytotoxic activities and effects on RNA synthesis in vitro were assayed. The drug concentrations inhibiting mouse leukaemia L1210 cell growth to 50% were about two- and three fold higher for the derivatives compared to doxorubicin and daunorubicin respectively. Inhibition of phage T7 RNA polymerase by the non-covalently interacting derivatives was also slightly lower than that by the parent compounds. As doxorubicin and daunorubicin, their amidine derivatives in the presence of dithiothreitol and Fe(III) ions are activated and covalently bind to DNA. The adducts formed affect RNA polymerase activity. Several bands corresponding to prematurely terminated RNA chains are observed by means of polyacrylamide gel electrophoresis. The patterns of bands are virtually identical for all the anthracyclines studied here and are similar to the terminations induced by actinomycin D. This observation is consistent with a notion that the adducts are formed at guanine in GpC sequences which are also binding sites of actinomycin D. A substantial difference between daunorubicin and its amidine derivative is shown by means of high performance liquid chromatography. The derivative undergoes rapid rearrangements in the presence of dithiothreitol and Fe(III) ions, while daunorubicin is stable for several hours under these conditions. The results presented here indicate that the amidine derivatives despite bulky morpholine substitution exhibit biological activity in the systems used here. PMID:11845988

  9. Trichostatin A Inhibits Retinal Pigmented Epithelium Activation in an In Vitro Model of Proliferative Vitreoretinopathy

    PubMed Central

    Greene, Whitney A.; Burke, Teresa A.; Wang, Heuy-Ching

    2016-01-01

    Abstract Purpose: Proliferative vitreoretinopathy (PVR) is a blinding disorder that develops after a retinal tear or detachment. Activation of the retinal pigmented epithelium (RPE) is implicated in PVR; however, the mechanisms leading to enhanced RPE proliferation, migration, and contraction remain largely unknown. This study utilized an in vitro model of PVR to investigate the role of acetylation in RPE activation and its contribution to the progression of this disease. Methods: ARPE-19 cells, primary cultures of porcine RPE, and induced pluripotent stem cell-derived RPE (iPS-RPE) were utilized for cellular and molecular analyses. Cells treated with transforming growth factor beta 2 (TGFβ2; 10 ng/mL) alone or in the presence of the broad-spectrum histone deacetylase (HDAC) inhibitor, trichostatin A (TSA; 0.1 μM), were assessed for contraction and migration through collagen contraction and scratch assays, respectively. Western blotting and immunofluorescence analysis were performed to assess α-smooth muscle actin (α-SMA) and β-catenin expression after TGFβ2 treatment alone or in combination with TSA. Results: TGFβ2 significantly increased RPE cell contraction in collagen matrix and this effect was inhibited in the presence of TSA (0.1 μM). In agreement with these data, immunofluorescence analysis of TSA-treated iPS-RPE wounded monolayers revealed decreased α-SMA as compared with control. Scratch assays to assess wound healing revealed TSA inhibited TGFβ2-mediated iPS-RPE cell migration. Conclusions: Our findings indicate a role of acetylation in RPE activation. Specifically, the HDAC inhibitor TSA decreased RPE cell proliferation and TGFβ2-mediated cell contraction and migration. Further investigation of pharmacological compounds that modulate acetylation may hold promise as therapeutic agents for PVR. PMID:27494828

  10. The inhibition of free radical generation by preparations of Harpagophytum procumbens in vitro.

    PubMed

    Grant, Louise; McBean, Douglas E; Fyfe, Lorna; Warnock, A Mary

    2009-01-01

    Harpagophytum procumbens (Hp), commonly known as Devil's Claw has been used as a traditional treatment for a variety of illnesses for centuries. Since the early twentieth century, it has become a popular antiinflammatory and analgesic preparation amongst European herbalists for supportive or adjuvant treatment of degenerative joint diseases. Extracts of Hp tubers have demonstrated antiinflammatory and analgesic effects in animal models of inflammation and in human trials. The mechanism(s) of action responsible for these attributes, however, remain to be elucidated. Reactive oxygen species generated in acute and chronic inflammatory diseases are known to be cytotoxic and can cause tissue damage. In this study, a root tuber extract (Hp extract) and commercially available tincture (Hp tincture) were investigated for antioxidant characteristics using in vitro test systems. Both preparations were found to effectively scavenge DPPH radical, inhibit nitrite levels in supernatants harvested from LPS-stimulated RAW 264.7 macrophages, and cause dose-dependent suppressions in the detection of fMLP- and AA-induced neutrophil MPO. The antioxidant effects demonstrated for both preparations of Hp may contribute to the antiinflammatory and analgesic properties observed for the plant. PMID:18803229

  11. In vitro antioxidant and H+, K+-ATPase inhibition activities of Acalypha wilkesiana foliage extract

    PubMed Central

    Prakash Gupta, Rajesh Kashi; Pradeepa; Hanumanthappa, Manjunatha

    2013-01-01

    Aims: The aim of this study was to evaluate the antioxidant activty and anti-acid property of Acalypha wilkesiana foliage extract. Materials and Methods: Hot and cold aqueous extracts were prepared from healthy leaves of A. wilkesiana. Free radical scavenging activity and H+, K+-ATPase inhibition activities of aqueous foliage extracts was screened by in vitro models. Statistical Analysis Used: All experiments were performed in triplicate and results are expressed as mean ± SEM. Results: A. wilkesiana hot aqueous extract (AWHE) showed significant antioxidants and free radical scavenging activity. Further, AWHE has shown a potent H+, K+-ATPase inhibitory activity (IC50: 51.5 ± 0.28 μg/ml) when compare to standard proton pump inhibitor omeprazole (56.2 ± 0.64 μg/ml); however, latter activity is equal to A. wilkesiana cold aqueous extract (AWCE). Quantitative analysis of AWHE has revealed more content of phenols and flavonoids; this is found to be the reason for good antioxidant activity over AWCE. Molecular docking was carried out against H+, K+-ATPase enzyme crystal structure to validate the anti-acid activity of A. wilkesiana major phytochemicals. Conclusions: The present study indicates that the constituents of AWHE and AWCE have good antacid and free radical scavenging activity. PMID:24082698

  12. Inhibition by cyclosporin A of rodent malaria in vivo and human malaria in vitro.

    PubMed Central

    Nickell, S P; Scheibel, L W; Cole, G A

    1982-01-01

    The development and course of normally lethal parasitemias in mice inoculated intraperitoneally with erythrocytic stages of Plasmodium yoelii or Plasmodium berghei were markedly affected by treatment with the antilymphoid drug cyclosporin A (CS-A). When the first of four daily subcutaneous 25-mg/kg doses of CS-A was given at the time of parasite inoculation, patent infections failed to develop. If begun up to 5 days earlier, this same treatment regimen prolonged the prepatent period, attenuated parasitemia, and reduced mortality. In mice with patient infections, two consecutive daily 25-mg/kg doses of CS-A were sufficient to terminate parasitemias which, after several days, reappeared but were self-limiting. This pattern of apparent cure followed by transient recrudescence remained unaltered even when daily treatment with the same drug dose was continued for 3 weeks. Recrudescence was associated with the emergence of parasite populations that were relatively resistant to CS-A and, in the case of P. yoelii, of reduced virulence. In more limited experiments, CS-A was found to be active in vitro against erythrocytic stages of the human malarial parasite palsmodium falciparum. Depending on the concentration of drug in the culture medium, parasite growth was either prevented or inhibited. PMID:6752020

  13. Compstatin analog Cp40 inhibits complement dysregulation in vitro in C3 glomerulopathy

    PubMed Central

    Zhang, Yuzhou; Shao, Dingwu; Ricklin, Daniel; Hilkin, Brieanna M.; Nester, Carla M.; Lambris, John D.; Smith, Richard J. H.

    2015-01-01

    C3 glomerulopathy (C3G) defines a group of untreatable ultra-rare renal diseases caused by uncontrolled activation of the alternative complement pathway. Nearly half of patients progress to end stage renal failure within 10 years. Cp40, a second-generation compstatin analog in clinical development, is a 14 amino-acid cyclic peptide that selectively inhibits complement activation in humans and non-human primates by binding to C3 and C3b. We hypothesized that by targeting C3 Cp40 would provide an effective treatment for C3G. By investigating its effects in vitro using multiple assays of complement activity, we show that Cp40 prevents complement-mediated lysis of sheep erythrocytes in sera from C3G patients, prevents complement dysregulation in the presence of patient-derived autoantibodies to the C3 and C5 convertases, and prevents complement dysregulation associated with disease-causing genetic mutations. In aggregate, these data suggest that Cp40 may offer a novel and promising therapeutic option to C3G patients as a disease-specific, targeted therapy. As such, Cp40 could represent a major advance in the treatment of this disease. PMID:25982307

  14. Downregulation of stathmin 1 in human gallbladder carcinoma inhibits tumor growth in vitro and in vivo

    PubMed Central

    Wang, Jiwen; Yao, Yanli; Ming, Yue; Shen, Sheng; Wu, Nan; Liu, Jiaqi; Liu, Han; Suo, Tao; Pan, Hongtao; Zhang, Dexiang; Ding, Kan; Liu, Houbao

    2016-01-01

    Gallbladder carcinoma (GBC) is a highly lethal malignancy of the gastrointestinal tract. Despite extensive research, the underlying molecular mechanism of GBC remains largely unclear. Stathmin 1 (STMN1) is an important cytosolic protein associated with microtubule stability that was reported to be involved in tumorigenesis. Up to our knowledge, its role in gallbladder carcinoma has not been analyzed. In this study, we found that STMN1 was significantly highly expressed in GBC by immunohistochemistry (IHC). Further research demonstrated that silencing of STMN1 inhibited cell growth in vitro. Moreover, knockdown of STMN1 induced apoptosis and delayed G2/M phase transformation in GBC cells. Our data support a rationale for further studies that the silencing of STMN1 may regulate the activity of p38 MAPK kinase and p53/p21 signal pathway. Besides, xenografted gallbladder carcinoma cells growth were significantly impaired after STMN1 was silenced in vivo. These results suggested that STMN1 played an important role in cell proliferation and migration. This provided a potential clue for investigating the therapeutic target in GBC. PMID:27349455

  15. Rice-produced MSP142 of Plasmodium falciparum elicits antibodies that inhibit parasite growth in vitro.

    PubMed

    Chen, Q; Liang, W; Qian, F; Qian, B; Cao, J; Zhang, D; Xu, Y; Tang, L

    2016-10-01

    Many malaria antigens contain multiple disulphide bonds involved in the formation of inhibitory B-cell epitopes. Producing properly folded malaria antigens in sufficient quantities for vaccination is often a challenge. The 42-kDa fragment of Plasmodium falciparum merozoite surface protein 1 (MSP142 ) is such a kind of malaria antigen. In this study, we investigated the expression of MSP142 in a rice system (9522, a cultivar of Oryza sativa ssp. japonica), which was used as a bioreactor for protein production. The MSP142 gene was synthesized according to rice-preferred codons and transformed into rice plants via an Agrobacterium-mediated method. The recombinant antigen was efficiently expressed in rice seeds with a level up to 1.56% of total soluble protein and was recognized by both the conformational monoclonal antibody 5.2 (mAb5.2) and the pooled sera of P. falciparum malaria patients. Rabbits were immunized intramuscularly with the purified MSP142 formulated with Freund's adjuvant. High antibody titres against MSP142 were elicited. The rabbit immune sera reacted well with the native protein of P. falciparum parasite and strongly inhibited the in vitro growth of blood-stage P. falciparum parasites, demonstrating that transgenic rice can become an efficient bioreactor for the production of malaria vaccine antigens. PMID:27493141

  16. Monoclonal Antibodies Inhibit in vitro Fibrillar Aggregation of the Alzheimer β -Amyloid Peptide

    NASA Astrophysics Data System (ADS)

    Solomon, Beka; Koppel, Rela; Hanan, Eilat; Katzav, Tamar

    1996-01-01

    The β -amyloid peptide, the hallmark of Alzheimer disease, forms fibrillar toxic aggregates in brain tissue that can be dissolved only by strong denaturing agents. To study β -amyloid formation and its inhibition, we prepared immune complexes with two monoclonal antibodies (mAbs), AMY-33 and 6F/3D, raised against β -amyloid fragments spanning amino acid residues 1-28 and 8-17 of the β -amyloid peptide chain, respectively. In vitro aggregation of β -amyloid peptide was induced by incubation for 3 h at 37 degrees C and monitored by ELISA, negative staining electron microscopy, and fluorimetric studies. We found that the mAbs prevent the aggregation of β -amyloid peptide and that the inhibitory effect appears to be related to the localization of the antibody-binding sites and the nature of the aggregating agents. Preparation of mAbs against ``aggregating epitopes,'' defined as sequences related to the sites where protein aggregation is initiated, may lead to the understanding and prevention of protein aggregation. The results of this study may provide a foundation for using mAbs in vivo to prevent the β -amyloid peptide aggregation that is associated with Alzheimer disease.

  17. Salubrinal inhibits the expression of proteoglycans and favors neurite outgrowth from cortical neurons in vitro.

    PubMed

    Barreda-Manso, M Asunción; Yanguas-Casás, Natalia; Nieto-Sampedro, Manuel; Romero-Ramírez, Lorenzo

    2015-07-01

    After CNS injury, astrocytes and mesenchymal cells attempt to restore the disrupted glia limitans by secreting proteoglycans and extracellular matrix proteins (ECMs), forming the so-called glial scar. Although the glial scar is important in sealing the lesion, it is also a physical and functional barrier that prevents axonal regeneration. The synthesis of secretory proteins in the RER is under the control of the initiation factor of translation eIF2α. Inhibiting the synthesis of secretory proteins by increasing the phosphorylation of eIF2α, might be a pharmacologically efficient way of reducing proteoglycans and other profibrotic proteins present in the glial scar. Salubrinal, a neuroprotective drug, decreased the expression and secretion of proteoglycans and other profibrotic proteins induced by EGF or TGFβ, maintaining eIF2α phosphorylated. Besides, Salubrinal also reduced the transcription of proteoglycans and other profibrotic proteins, suggesting that it induced the degradation of non-translated mRNA. In a model in vitro of the glial scar, cortical neurons grown on cocultures of astrocytes and fibroblasts with TGFβ treated with Salubrinal, showed increased neurite outgrowth compared to untreated cells. Our results suggest that Salubrinal may be considered of therapeutic value facilitating axonal regeneration, by reducing overproduction and secretion of proteoglycans and profibrotic protein inhibitors of axonal growth. PMID:25882497

  18. In vitro inhibition of feline coronavirus replication by small interfering RNAs.

    PubMed

    McDonagh, Phillip; Sheehy, Paul A; Norris, Jacqueline M

    2011-06-01

    Infection with virulent biotypes of feline coronavirus (FCoV) can result in the development of feline infectious peritonitis (FIP), a typically fatal immune mediated disease for which there is currently no effective antiviral treatment. In this study we demonstrate the ability of small interfering RNA (siRNA) mediated RNA interference (RNAi) to inhibit the replication of virulent FCoV strain FIPV WSU 79-1146 in an immortalised feline cell line. A panel of eight synthetic siRNAs targeting four different regions of the FCoV genome were tested for antiviral effects. Efficacy was determined by qRT-PCR of intracellular viral genomic and messenger RNA, TCID50 infectivity assay of extracellular virus, and direct IFA for viral protein expression. All siRNAs demonstrated an inhibitory effect on viral replication in vitro. The two most effective siRNAs, targeting the untranslated 5' leader sequence (L2) and the nucleocapsid gene (N1), resulted in a >95% reduction in extracellular viral titre. Further characterisation of these two siRNAs demonstrated their efficacy when used at low concentrations and in cells challenged with high viral loads. Taken together these findings provide important information for the potential therapeutic application of RNAi in treating FIP.

  19. Fluoxetine Induces Proliferation and Inhibits Differentiation of Hypothalamic Neuroprogenitor Cells In Vitro

    PubMed Central

    Sousa-Ferreira, Lígia; Aveleira, Célia; Botelho, Mariana; Álvaro, Ana Rita; Pereira de Almeida, Luís; Cavadas, Cláudia

    2014-01-01

    A significant number of children undergo maternal exposure to antidepressants and they often present low birth weight. Therefore, it is important to understand how selective serotonin reuptake inhibitors (SSRIs) affect the development of the hypothalamus, the key center for metabolism regulation. In this study we investigated the proliferative actions of fluoxetine in fetal hypothalamic neuroprogenitor cells and demonstrate that fluoxetine induces the proliferation of these cells, as shown by increased neurospheres size and number of proliferative cells (Ki-67+ cells). Moreover, fluoxetine inhibits the differentiation of hypothalamic neuroprogenitor cells, as demonstrated by decreased number of mature neurons (Neu-N+ cells) and increased number of undifferentiated cells (SOX-2+ cells). Additionally, fluoxetine-induced proliferation and maintenance of hypothalamic neuroprogenitor cells leads to changes in the mRNA levels of appetite regulator neuropeptides, including Neuropeptide Y (NPY) and Cocaine-and-Amphetamine-Regulated-Transcript (CART). This study provides the first evidence that SSRIs affect the development of hypothalamic neuroprogenitor cells in vitro with consequent alterations on appetite neuropeptides. PMID:24598761

  20. Boric acid inhibits germination and colonization of Saprolegnia spores in vitro and in vivo.

    PubMed

    Ali, Shimaa E; Thoen, Even; Evensen, Øystein; Skaar, Ida

    2014-01-01

    Saprolegnia infections cause severe economic losses among freshwater fish and their eggs. The banning of malachite green increased the demand for finding effective alternative treatments to control the disease. In the present study, we investigated the ability of boric acid to control saprolegniosis in salmon eggs and yolk sac fry. Under in vitro conditions, boric acid was able to decrease Saprolegnia spore activity and mycelial growth in all tested concentrations above 0.2 g/L, while complete inhibition of germination and growth was observed at a concentration of 0.8 g/L. In in vivo experiments using Atlantic salmon eyed eggs, saprolegniosis was controlled by boric acid at concentrations ranging from 0.2-1.4 g/L during continuous exposure, and at 1.0-4.0 g/L during intermittent exposure. The same effect was observed on salmon yolk sac fry exposed continuously to 0.5 g/L boric acid during the natural outbreak of saprolegniosis. During the experiments no negative impact with regard to hatchability and viability was observed in either eggs or fry, which indicate safety of use at all tested concentrations. The high hatchability and survival rates recorded following the in vivo testing suggest that boric acid is a candidate for prophylaxis and control of saprolegniosis.

  1. The Organophosphate Paraoxon and Its Antidote Obidoxime Inhibit Thrombin Activity and Affect Coagulation In Vitro

    PubMed Central

    Golderman, Valery; Shavit-Stein, Efrat; Tamarin, Ilia; Rosman, Yossi; Shrot, Shai; Rosenberg, Nurit

    2016-01-01

    Organophosphates (OPs) are potentially able to affect serine proteases by reacting with their active site. The potential effects of OPs on coagulation factors such as thrombin and on coagulation tests have been only partially characterized and potential interactions with OPs antidotes such as oximes and muscarinic blockers have not been addressed. In the current study, we investigated the in vitro interactions between coagulation, thrombin, the OP paraoxon, and its antidotes obidoxime and atropine. The effects of these substances on thrombin activity were measured in a fluorescent substrate and on coagulation by standard tests. Both paraoxon and obidoxime but not atropine significantly inhibited thrombin activity, and prolonged prothrombin time, thrombin time, and partial thromboplastin time. When paraoxon and obidoxime were combined, a significant synergistic effect was found on both thrombin activity and coagulation tests. In conclusion, paraoxon and obidoxime affect thrombin activity and consequently alter the function of the coagulation system. Similar interactions may be clinically relevant for coagulation pathways in the blood and possibly in the brain. PMID:27689805

  2. Immune thrombocytopenia: antiplatelet autoantibodies inhibit proplatelet formation by megakaryocytes and impair platelet production in vitro

    PubMed Central

    Iraqi, Muna; Perdomo, Jose; Yan, Feng; Choi, Philip Y-I; Chong, Beng H.

    2015-01-01

    Primary immune thrombocytopenia is an autoimmune disease mediated by antiplatelet autoantibodies that cause platelet destruction and suppression of platelet production. In vitro effects of autoantibodies on megakaryocyte production and maturation have been reported recently. However, the impact of these autoantibodies on crucial megakaryocyte functions, proplatelet formation and subsequent platelet release, has not been evaluated. We examined the effects of serum and IgG from 19 patients with immune thrombocytopenia using day 8 or 9 megakaryocytes (66.3 ± 10.6% CD41+), derived from cord blood hematopoietic stem cells (CD34+). The number of proplatelet-bearing megakaryocytes, the number of platelets released in the culture, total megakaryocyte numbers, ploidy pattern and caspase activation were measured at various times after treatment. After 5 days of treatment the number of proplatelet-bearing megakaryocytes was significantly decreased by 13 immune thrombocytopenia autoantibodies relative to the control group (P<0.0001) and this decrease was accompanied by a corresponding reduction of platelet release. Other features, including total megakaryocyte numbers, maturation and apoptosis, were not affected by immune thrombocytopenia antibodies. Treating the megakaryocytes with the thrombopoietin receptor agonists romiplostim and eltrombopag reversed the effect of the autoantibodies on megakaryocytes by restoring their capacity to form proplatelets. We conclude that antiplatelet antibodies in immune thrombocytopenia inhibit proplatelet formation by megakaryocytes and hence the ability of the megakaryocytes to release platelets. Treatment with either romiplostim or eltrombopag regenerates proplatelet formation from the megakaryocytes. PMID:25682608

  3. The use of sesame oil and other vegetable oils in the inhibition of human colon cancer growth in vitro.

    PubMed

    Salerno, J W; Smith, D E

    1991-01-01

    Sesame contains large quantities of the essential polyunsaturated fatty acid (PUFA), linoleic acid, in the form triglycerides. The antineoplastic properties of many PUFAs such as linoleic acid and their metabolites are known. We tested the hypothesis that natural vegetable oils, such as sesame oil and its component linoleic acid, when added to human colon adenocarcinoma cells growing in tissue culture would inhibit their growth and that normal colon cells would not be similarly affected. Three human colon cancer cell lines and one normal human colon cell line were exposed to the following: (1) pure linoleic acid; (2) lipase-digested sesame oil; (3) undigested sesame oil; (4) five additional common vegetable oils; (5) mineral oil. Linoleic acid inhibited the in vitro growth of all three malignant human colon adenocarcinoma cell lines. The normal colon cell line showed dramatically less inhibition of growth. Lipase-digested sesame oil (LDSO) and undigested sesame oil (UDSO) produced greater inhibition of growth of all three malignant colon cell lines than of the normal colon cells. Five other common vegetable oils containing various amounts of PUFAs such as corn, soybean, safflower, olive and coconut oils, all in their lipase-digested form, were found to dramatically inhibit the growth of the HT-29 malignant human colon cell line. Undigested olive and safflower oils also inhibited the HT-29 cells although not as markedly as the lipase-digested oils. Mineral oil did not inhibit the growth of HT-29 cells. Both lauric and palmitic acid, which are saturated fatty acids found in abundance in coconut oil inhibits the HT-29 cells more strongly than linoleic acid, while oleic acid did not inhibit. We conclude that many vegetable oils including sesame contain in vitro antineoplastic properties and that this finding warrants further investigation both in vitro and in vivo to assess their possible chemotherapeutic potential.

  4. Heterometallic titanium-gold complexes inhibit renal cancer cells in vitro and in vivo‡

    PubMed Central

    Sadhukha, Tanmoy; Prabha, Swayam; Sanaú, Mercedes; Rotenberg, Susan A.

    2016-01-01

    Following recent work on heterometallic titanocene-gold complexes as potential chemotherapeutics for renal cancer, we report here on the synthesis, characterization and stability studies of new titanocene complexes containing a methyl group and a carboxylate ligand (mba = S-C6H4-COO−) bound to gold(I)-phosphane fragments through a thiolate group ([(η-C5H5)2TiMe(μ-mba)Au(PR3)]. The compounds are more stable in physiological media than those previously reported and are highly cytotoxic against human cancer renal cell lines. We describe here preliminary mechanistic data involving studies on the interaction of selected compounds with plasmid (pBR322) DNA used as a model nucleic acid, and with selected protein kinases from a panel of 35 protein kinases having oncological interest. Preliminary mechanistic studies in Caki-1 renal cells indicate that the cytotoxic and anti-migration effects of the most active compound 5 ([(η-C5H5)2TiMe(μ-mba)Au(PPh3)] involve inhibition of thioredoxin reductase and loss of expression of protein kinases that drive cell migration (AKT, p90-RSK, and MAPKAPK3). The co-localization of both titanium and gold metals (1:1 ratio) in Caki-1 renal cells was demonstrated for 5 indicating the robustness of the heterometallic compound in vitro. Two compounds were selected for further in vivo studies on mice based on their selectivity in vitro against renal cancer cell lines when compared to non-tumorigenic human kidney cell lines (HEK-293T and RPTC) and the favourable preliminary toxicity profile in C57BL/6 mice. Evaluation of Caki-1 xenografts in NOD.CB17-Prkdc SCID/J mice showed an impressive tumor reduction (67%) after treatment for 28 days (3 mg/kg/every other day) with heterometallic compound 5 as compared with the previously described [(η-C5H5)2Ti{OC(O)-4-C6H4-P(Ph2)AuCI}2] 3 which was non-inhibitory. These findings indicate that structural modifications on the ligand scaffold affect the in vivo efficacy of this class of compounds. PMID

  5. Inhibition of Both Hsp70 Activity and Tau Aggregation in Vitro Best Predicts Tau Lowering Activity of Small Molecules.

    PubMed

    Martin, Mackenzie D; Baker, Jeremy D; Suntharalingam, Amirthaa; Nordhues, Bryce A; Shelton, Lindsey B; Zheng, Dali; Sabbagh, Jonathan J; Haystead, Timothy A J; Gestwicki, Jason E; Dickey, Chad A

    2016-07-15

    Three scaffolds with inhibitory activity against the heat shock protein 70 (Hsp70) family of chaperones have been found to enhance the degradation of the microtubule associated protein tau in cells, neurons, and brain tissue. This is important because tau accumulation is linked to neurodegenerative diseases including Alzheimer's disease (AD) and chronic traumatic encephalopathy (CTE). Here, we expanded upon this study to investigate the anti-tau efficacy of additional scaffolds with Hsp70 inhibitory activity. Five of the nine scaffolds tested lowered tau levels, with the rhodacyanine and phenothiazine scaffolds exhibiting the highest potency as previously described. Because phenothiazines also inhibit tau aggregation in vitro, we suspected that this activity might be a more accurate predictor of tau lowering. Interestingly, the rhodacyanines did inhibit in vitro tau aggregation to a similar degree as phenothiazines, correlating well with tau-lowering efficacy in cells and ex vivo slices. Moreover, other Hsp70 inhibitor scaffolds with weaker tau-lowering activity in cells inhibited tau aggregation in vitro, albeit at lower potencies. When we tested six well-characterized tau aggregation inhibitors, we determined that this mechanism of action was not a better predictor of tau-lowering than Hsp70 inhibition. Instead, we found that compounds possessing both activities were the most effective at promoting tau clearance. Moreover, cytotoxicity and PAINS activity are critical factors that can lead to false-positive lead identification. Strategies designed around these principles will likely yield more efficacious tau-lowering compounds. PMID:27177119

  6. Staphylococcus aureus enterotoxins A and B inhibit human and mice eosinophil chemotaxis and adhesion in vitro.

    PubMed

    Squebola-Cola, Dalize M; De Mello, Glaucia C; Anhê, Gabriel F; Condino-Neto, Antonio; DeSouza, Ivani A; Antunes, Edson

    2014-12-01

    Staphylococcus aureus aggravates the allergic eosinophilic inflammation. We hypothesized that Staphylococcus aureus-derived enterotoxins directly affect eosinophil functions. Therefore, this study investigated the effects of Staphylococcal enterotoxins A and B (SEA and SEB) on human and mice eosinophil chemotaxis and adhesion in vitro, focusing on p38 MAPK phosphorylation and intracellular Ca(2+) mobilization. Eosinophil chemotaxis was evaluated using a microchemotaxis chamber, whereas adhesion was performed in VCAM-1 and ICAM-1-coated plates. Measurement of p38 MAPK phosphorylation and intracellular Ca(2+) levels were monitored by flow cytometry and fluorogenic calcium-binding dye, respectively. Prior incubation (30 to 240 min) of human blood eosinophils with SEA (0.5 to 3 ng/ml) significantly reduced eotaxin-, PAF- and RANTES-induced chemotaxis (P<0.05). Likewise, SEB (1 ng/ml, 30 min) significantly reduced eotaxin-induced human eosinophil chemotaxis (P<0.05). The reduction of eotaxin-induced human eosinophil chemotaxis by SEA and SEB was prevented by anti-MHC monoclonal antibody (1 μg/ml). In addition, SEA and SEB nearly suppressed the eotaxin-induced human eosinophil adhesion in ICAM-1- and VCAM-1-coated plates. SEA and SEB prevented the increases of p38 MAPK phosphorylation and Ca(2+) levels in eotaxin-activated human eosinophils. In separate protocols, we evaluated the effects of SEA on chemotaxis and adhesion of eosinophils obtained from mice bone marrow. SEA (10 ng/ml) significantly reduced the eotaxin-induced chemotaxis along with cell adhesion to both ICAM-1 and VCAM-1-coated plates (P<0.05). In conclusion, the inhibition by SEA and SEB of eosinophil functions (chemotaxis and adhesion) are associated with reductions of p38 MAPK phosphorylation and intracellular Ca(2+) mobilization.

  7. Inhibition of fast axonal transport in vitro by the local anesthetics prilocaine, mepivacaine, and bupivacaine.

    PubMed

    Lavoie, P A

    1983-12-01

    The aim of the present study was to establish the concentrations of prilocaine, mepivacaine, and bupivacaine which are effective at blocking fast axonal transport, to determine whether prilocaine and mepivacaine offer a better prospect of dissociating conduction block and transport block in vivo than does lidocaine and whether bupivacaine offers a better prospect than etidocaine in the same context. Fast axonal transport of [3H]leucine-labeled proteins was studied in vitro in bullfrog spinal nerves and quantitated by liquid scintillation counting. Exposure of spinal nerves to 14 mM prilocaine reduced the quantity of 3H-labeled proteins which accumulated at a ligature by 86%, and exposure to 14 mM mepivacaine reduced it by 70%; 10 mM prilocaine reduced this same parameter by 54%, a degree of inhibition close to the 44% reduction caused by 14 mM lidocaine. The D(-) and L(+) stereoisomers of mepivacaine each reduced transport to the ligature by approximately 50% at a concentration of 14 mM. Bupivacaine reduced the accumulation of 3H-labeled proteins at the ligature by 49% at a 10 mM concentration (pH 6.2); its potency is close to that found for etidocaine in a previous study. Since prilocaine and mepivacaine are at least as potent as lidocaine as transport inhibitors and at blocking impulse conduction, these two anesthetics offer no advantage over lidocaine to achieve dissociation of conduction block from transport block in vivo. Bupivacaine appears to offer no advantage over etidocaine in the same context, as the two agents have a similar potency as local anesthetics and a similar potency as inhibitors of fast axonal transport.

  8. Carbon disulfide inhibits neurite outgrowth and neuronal migration of dorsal root ganglion in vitro.

    PubMed

    Ding, Ning; Xiang, Yujuan; Jiang, Hao; Zhang, Weiwei; Liu, Huaxiang; Li, Zhenzhong

    2011-12-01

    Carbon disulfide (CS₂) is a neurotoxic industrial solvent and widely used in the vulcanization of rubber, rayon, cellophane, and adhesives. Although the neurotoxicity of CS₂ has been recognized for over a century, the precise mechanism of neurotoxic action of CS₂ remains unknown. In the present study, a embryonic rat dorsal root ganglia (DRG) explants culture model was established. Using the organotypic DRG cultures, the direct neurotoxic effects of CS₂ on outgrowth of neurites and migration of neurons from DRG explants were investigated. The organotypic DRG cultures were exposed to different concentrations of CS₂ (0.01 mmol/L, 0.1 mmol/L, 1 mmol/L). The number of nerve fiber bundles extended from DRG explants decreased significantly in the presence of CS₂ (0.01 mmol/L, 15.00 ± 2.61, p < .05; 0.1 mmol/L, 11.17 ± 1.47, p < .001; 1 mmol/L, 8.00 ± 1.41, p < .001) as compared with that in the absence of CS₂ (17.83 ± 2.48). The number of neurons migrated from DRG explants decreased significantly in the presence of CS₂ (0.01 mmol/L, 79.50 ± 9.40, p < .01; 0.1 mmol/L, 62.50 ± 14.15, p < .001; 1 mmol/L, 34.67 ± 7.58, p < .001) as compared with that in the absence of CS₂ (99.33 ± 15.16). And also, the decreases in the number of nerve fiber bundles and migrated DRG neurons were in a dose-dependent manner of CS₂. These data implicated that CS₂ could inhibit neurite outgrowth and neuronal migration from DRG explants in vitro. PMID:21777162

  9. Thymoquinone Inhibits Murine Leukemia WEHI-3 Cells In Vivo and In Vitro

    PubMed Central

    Ali Salim, Landa Zeenelabdin; Othman, Rozana; Abdulla, Mahmood Ameen; Al-Jashamy, Karim; Mohd Ali, Hapipah; Hassandarvish, Pouya; Dehghan, Firouzeh; Ibrahim, Mohamed Yousif; Omer, Fatima Abd Elmutaal Ahmed; Mohan, Syam

    2014-01-01

    Background Thymoquinone is an active ingredient isolated from Nigella sativa (Black Seed). This study aimed to evaluate the in vitro and in vivo anti-leukemic effects of thymoquinone on WEHI-3 cells. Methodology/Principal Findings The cytotoxic effect of thymoquinone was assessed using an MTT assay, while the inhibitory effect of thymoquinone on murine WEHI-3 cell growth was due to the induction of apoptosis, as evidenced by chromatin condensation dye, Hoechst 33342 and acridine orange/propidium iodide fluorescent staining. In addition, Annexin V staining for early apoptosis was performed using flowcytometric analysis. Apoptosis was found to be associated with the cell cycle arrest at the S phase. Expression of Bax, Bcl2 and HSP 70 proteins were observed by western blotting. The effects of thymoquinone on BALB/c mice injected with WEHI-3 cells were indicated by the decrease in the body, spleen and liver weights of the animal, as compared to the control. Conclusion Thymoquinone promoted natural killer cell activities. This compound showed high toxicity against WEHI-3 cell line which was confirmed by an increase of the early apoptosis, followed by up-regulation of the anti-apoptotic protein, Bcl2, and down-regulation of the apoptotic protein, Bax. On the other hand, high reduction of the spleen and liver weight, and significant histopathology study of spleen and liver confirmed that thymoquinone inhibited WEHI-3 growth in the BALB/c mice. Results from this study highlight the potential of thymoquinone to be developed as an anti-leukemic agent. PMID:25531768

  10. Ovine tracheal muscle contraction in vitro: inhibition by calcium channel blockers gallopamil and verapamil.

    PubMed

    Jackowski, J; Chapman, G A; Abraham, W M; Ahmed, T

    1993-01-01

    We compared the inhibitory effects of calcium channel blockers, gallopamil and verapamil on acetylcholine (Ach)-induced contractions of ovine tracheal muscle in vitro. Adult sheep were sacrificed and tracheal strips were obtained by cutting the single tracheal rings from the mid-trachea. Tracheal strips were suspended in Krebs-Henseleit solution and isometric tension measured upon stimulation with cumulative doses of Ach (10(-7) to 10(-4) M) without and after pretreatment with gallopamil (10(-7) to 10(-6) M) or verapamil (10(-6) to 10(-5) M). In untreated tissues, the mean concentration of Ach required to produce 50% of maximal response (EC50) was 4.3 x 10(-6) M Ach. Both gallopamil and verapamil inhibited the Ach-induced contractions of ovine tracheal smooth muscle, by shifting the dose-response curves to Ach to the right. EC50 Ach for gallopamil (10(-6) M) and verapamil (10(-6) M) was 2.6 x 10(-5) and 5.2 x 10(-6) M, respectively. Dose ratio defined as postantagonist EC50 Ach/control EC50 Ach, was 7.7 for gallopamil and 2.0 for verapamil. Thus, the inhibitory effect of gallopamil was approximately 4-fold more potent than that of verapamil. Gallopamil was 17-fold more potent than verapamil in relaxing precontracted tracheal strips. The dose of calcium antagonists required to produce 25% relaxation (EC25) of tracheal strips precontracted with 10(-4) Ach was 3.7 x 10(-5) M for verapamil and 2.2 x 10(-6) M for gallopamil. These results indicate that gallopamil is effective against Ach-induced contractions of ovine trachealis muscles, and is more potent than verapamil. PMID:8469817

  11. Silencing of ghrelin receptor expression inhibits endometrial cancer cell growth in vitro and in vivo.

    PubMed

    Fung, Jenny N T; Jeffery, Penny L; Lee, John D; Seim, Inge; Roche, Deborah; Obermair, Andreas; Chopin, Lisa K; Chen, Chen

    2013-07-15

    Ghrelin is a 28-amino acid peptide hormone produced predominantly in the stomach but also in a range of normal cell types and tumors, where it has endocrine, paracrine, and autocrine roles. Previously, we have demonstrated that ghrelin has proliferative and antiapoptotic effects in endometrial cancer cell lines, suggesting a potential role in promoting tumor growth. In the present study, we investigated the effect of ghrelin receptor, GHSR, and gene silencing in vitro and in vivo and characterized ghrelin and GHSR1a protein expression in human endometrial tumors. GHSR gene silencing was achieved in the Ishikawa and KLE endometrial cancer cell lines, using a lentiviral short-hairpin RNA targeting GHSR. The effects of GHSR1a knockdown were further analyzed in vivo using the Ishikawa cell line in a NOD/SCID xenograft model. Cell proliferation was reduced in cultured GHSR1a knockdown Ishikawa and KLE cells compared with scrambled controls in the absence of exogenously applied ghrelin and in response to exogenous ghrelin (1,000 nM). The tumor volumes were reduced significantly in GHSR1a knockdown Ishikawa mouse xenograft tumors compared with scrambled control tumours. Using immunohistochemistry, we demonstrated that ghrelin and GHSR1a are expressed in benign and cancerous glands in human endometrial tissue specimens, although there was no correlation between the intensity of staining and cancer grade. These data indicate that downregulation of GHSR expression significantly inhibits endometrial cancer cell line and mouse xenograft tumour growth. This is the first preclinical evidence that downregulation of GHSR may be therapeutic in endometrial cancer.

  12. Tissue distribution, characterization and in vitro inhibition of B-esterases in the earwig Forficula auricularia.

    PubMed

    Malagnoux, Laure; Capowiez, Yvan; Rault, Magali

    2014-10-01

    Earwigs are important natural enemies of numerous pests in pome fruit orchards worldwide. Studying the effects of agricultural practices on these biological control agents is important for understanding its vulnerability in the field. The aim of this study was to characterize the B-esterase activities in the European earwig Forficula auricularia and to evaluate in vitro its sensitivity to organophosphate and carbamate pesticides. Acetylcholinesterase (AChE) activity was mainly measured with 1.5 mM acetylthiocholine as the substrate in the microsomal fraction of earwig heads (70% of total AChE activity). Carboxylesterase (CbE) activities were measured with three substrates [5 mM 4-nitrophenyl acetate (4-NPA), 1mM 4-nitrophenyl valerate (4-NPV), and 2 mM α-naphtyl acetate (α-NA)] to examine different isoenzymes, which were present mainly in the cytosolic fraction (about 70-88% of total activities) of all earwig tissues. CbE activity was higher than AChE activity, especially with α-NA, then 4-NPA and lastly 4-NPV. Chlorpyrifos-oxon an organophosphate, and carbaryl a carbamate pesticide, inhibited AChE and CbE activities in a concentration-dependent manner. Earwig CbE activities showed a stronger sensitivity to organophosphate than AChE, with the strongest effect for chlorpyrifos-oxon on male carboxylesterase activities. CbE and AChE showed about the same sensitivity to carbamate pesticides regardless of sex. These results suggest that B-type esterases in the European earwig F.auricularia are suitable biomarkers of pesticide exposure.

  13. Guava leaves polyphenolics-rich extract inhibits vital enzymes implicated in gout and hypertension in vitro

    PubMed Central

    Irondi, Emmanuel Anyachukwu; Agboola, Samson Olalekan; Oboh, Ganiyu; Boligon, Aline Augusti; Athayde, Margareth Linde; Shode, Francis O.

    2016-01-01

    Background/Aim: Elevated uric acid level, an index of gout resulting from the over-activity of xanthine oxidase (XO), increases the risk of developing hypertension. However, research has shown that plant-derived inhibitors of XO and angiotensin 1-converting enzyme (ACE), two enzymes implicated in gout and hypertension, respectively, can prevent or ameliorate both diseases, without noticeable side effects. Hence, this study characterized the polyphenolics composition of guava leaves extract and evaluated its inhibitory effect on XO and ACE in vitro. Materials and Methods: The polyphenolics (flavonoids and phenolic acids) were characterized using high-performance liquid chromatography (HPLC) coupled with diode array detection (DAD). The XO, ACE, and Fe2+-induced lipid peroxidation inhibitory activities, and free radicals (2,2-diphenylpicrylhydrazyl [DPPH]* and 2,2´-azino-bis-3-ethylbenzthiazoline-6-sulphonic [ABTS]*+) scavenging activities of the extract were determined using spectrophotometric methods. Results: Flavonoids were present in the extract in the order of quercetin > kaempferol > catechin > quercitrin > rutin > luteolin > epicatechin; while phenolic acids were in the order of caffeic acid > chlorogenic acid > gallic acids. The extract effectively inhibited XO, ACE and Fe2+-induced lipid peroxidation in a dose-dependent manner; having half-maximal inhibitory concentrations (IC50) of 38.24 ± 2.32 μg/mL, 21.06 ± 2.04 μg/mL and 27.52 ± 1.72 μg/mL against XO, ACE and Fe2+-induced lipid peroxidation, respectively. The extract also strongly scavenged DPPH* and ABTS*+. Conclusion: Guava leaves extract could serve as functional food for managing gout and hypertension and attenuating the oxidative stress associated with both diseases. PMID:27104032

  14. Chloroquine inhibits colon cancer cell growth in vitro and tumor growth in vivo via induction of apoptosis.

    PubMed

    Zheng, Yuzhu; Zhao, Ying-Lan; Deng, Xiaoqiang; Yang, Shengyong; Mao, Yongqiu; Li, Zhengguang; Jiang, Peidu; Zhao, Xia; Wei, Yuquan

    2009-03-01

    The present study was to investigate the anticancer effect of chloroquine on proliferation of mouse colon cancer cell line CT26 in vivo and in vitro and the possible mechanism. We found that chloroquine inhibited CT26 proliferation by concentration- and time-dependent manner. This effect was associated with apoptosis induction and decreased level of phosphorylated p42/44 mitogen-activated protein kinase and phosphorylated Akt. The in vivo study showed chloroquine-reduced tumor volume and prolonged survival time in CT26-bearing mice. These observations indicated chloroquine could inhibit CT26 proliferation by inducing apoptosis both in vitro and in vivo, providing its chemotherapeutic potential of human cancers. PMID:19194831

  15. Bayesian models trained with HTS data for predicting β-haematin inhibition and in vitro antimalarial activity

    PubMed Central

    Wicht, Kathryn J.; Combrinck, Jill M.; Smith, Peter J.; Egan, Timothy J.

    2015-01-01

    A large quantity of high throughput screening (HTS) data for antimalarial activity has become available in recent years. This includes both phenotypic and target-based activity. Realising the maximum value of these data remains a challenge. In this respect, methods that allow such data to be used for virtual screening maximise efficiency and reduce costs. In this study both in vitro antimalarial activity and inhibitory data for β-haematin formation, largely obtained from publically available sources, has been used to develop Bayesian models for inhibitors of β-haematin formation and in vitro antimalarial activity. These models were used to screen two in silico compound libraries. In the first, the 1510 U.S. Food and Drug Administration approved drugs available on PubChem were ranked from highest to lowest Bayesian score based on a training set of β-haematin inhibiting compounds active against P. falciparum that did not include any of the clinical antimalarials or close analogues. The six known clinical antimalarials that inhibit β-haematin formation were ranked in the top 2.1% of compounds. Furthermore, the in vitro antimalarial hit-rate for this prioritised set of compounds was found to be 81% in the case of the subset where activity data are available in PubChem. In the second, a library of about 5,000 commercially available compounds (AldrichCPR) was virtually screened for ability to inhibit β-haematin formation and then for in vitro antimalarial activity. A selection of 34 compounds was purchased and tested, of which 24 were predicted to be β-haematin inhibitors. The hit rate for inhibition of β-haematin formation was found to be 25% and a third of these were active against P. falciparum, corresponding to enrichments estimated at about 25- and 140-fold relative to random screening, respectively. PMID:25573118

  16. Anti-Candida-biofilm activity of micafungin is attenuated by voriconazole but restored by pharmacological inhibition of Hsp90-related stress responses.

    PubMed

    Kaneko, Yukihiro; Ohno, Hideaki; Fukazawa, Hidesuke; Murakami, Yuko; Imamura, Yoshifumi; Kohno, Shigeru; Miyazaki, Yoshitsugu

    2010-06-01

    We have conducted an in vitro evaluation of the efficacy of a voriconazole-micafungin combination against Candida albicans. When used alone, both micafungin and voriconazole decreased the metabolic activity of planktonic cells, but only micafungin displayed potent anti-biofilm activity. Their combination appeared to have an additive effect against planktonic cells. However, voriconazole significantly antagonized the fungicidal effect of micafungin against Candida biofilms. Time-lag experiments showed that pre-treatment with voriconazole induced resistance to micafungin in Candida biofilms. The micafungin-antagonizing effect of voriconazole persisted even when the biofilm was no longer exposed to voriconazole. In contrast, voriconazole addition after 24 h of micafungin treatment did not alter micafungin sensitivity. To investigate the mechanism of antagonism, we used inhibitors of Hsp90 and its effectors because Hsp90 seems to be implicated in the resistance to micafungin. These molecules reversed the voriconazole-induced resistance to micafungin which suggests that Hsp90-related stress responses are involved in the antagonism. Our results may provide clues as to the mechanism of increased drug resistance in Candida biofilms and raises concerns about the use of the voriconazole-micafungin combination in clinical settings. PMID:19958255

  17. SILIBININ INHIBITS ETHANOL METABOLISM AND ETHANOL-DEPENDENT CELL PROLIFERATION IN AN IN VITRO MODEL OF HEPATOCELLULAR CARCINOMA

    PubMed Central

    Brandon-Warner, Elizabeth; Sugg, James A.; Schrum, Laura W.; McKillop, Iain H.

    2009-01-01

    Chronic ethanol consumption is a known risk factor for developing hepatocellular carcinoma (HCC). The use of plant-derived antioxidants is gaining increasing clinical prominence as a potential therapy to ameliorate the effects of ethanol on hepatic disease development and progression. This study demonstrates silibinin, a biologically active flavanoid derived from milk thistle, inhibits cytochrome p4502E1 induction, ethanol metabolism and reactive oxygen species generation in HCC cells in vitro. These silibinin-mediated effects also inhibit ethanol-dependent increases in HCC cell proliferation in culture. PMID:19900758

  18. Candida albicans Dbf4-dependent Cdc7 kinase plays a novel role in the inhibition of hyphal development

    PubMed Central

    Lai, Wei-Chung; Chang, Tschen-wei; Wu, Chang Hao; Yang, Shu-Ya; Lee, Tai-Lin; Li, Wan Chen; Chien, Ting; Cheng, Yu-Che; Shieh, Jia-Ching

    2016-01-01

    Candida albicans is an opportunistic human fungal pathogen. The ability to switch among multiple cellular forms is key to its pathogenesis. The Dbf4-dependent protein kinase gene CDC7 is conserved due to its role in initiating DNA replication. Because a C. albicans Cdc7 (Cacdc7) homozygous null was not viable, we generated a C. albicans strain with a deleted C. albicans CDC7 (CaCDC7) allele and an expression-repressible allele. Surprisingly, cells of the strain grew as hyphae under the repressed conditions. The in vitro kinase assays confirmed that CaCdc7 (K232) and CaCdc7 (T437) are critical for catalytic and phosphoacceptor of activation activity, respectively. C. albicans cells formed hyphae when expressing either the catalytically inactive CaCdc7 (K232R) or the phosphoacceptor-deficient CaCdc7 (T437A). While CaCdc7 interacted with CaDbf4, cells of the strain in which CaCDC7 was repressed were not rescued by constitutively expressing C. albicans DBF4 or vice versa. We conclude that CaDBF4-dependent CaCDC7 is an essential gene suppressing the hyphal development. PMID:27644158

  19. Azole antifungal inhibition of buprenorphine, methadone and oxycodone in vitro metabolism.

    PubMed

    Moody, David E; Liu, Fenyun; Fang, Wenfang B

    2015-06-01

    Opioid-related mortality rates have escalated. Drug interactions may increase blood concentrations of the opioid. We therefore used human liver microsomes (HLMs) and cDNA-expressed human cytochrome P450s (rCYPs) to study in vitro inhibition of buprenorphine metabolism to norbuprenorphine (CYP3A4 and 2C8), oxycodone metabolism to noroxycodone (CYP3A4 and 2C18) and oxymorphone (CYP2D6), and methadone metabolism to R- and S-2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP; CYP3A4 and 2B6). In this study, we have examined the inhibitory effect of 12 (mostly antifungal) azoles. These compounds have a wide range of solubility; to keep organic solvent ≤1%, there was an equally wide range of highest concentration tested (e.g., itraconazole 5 µM to fluconazole 1000 µM). Inhibitors were first incubated with HLMs at three concentrations with or without preincubation of inhibitor with reducing equivalents to also screen for time-dependent inhibition (TDI). Posaconazole displayed evidence of TDI; metronidazole and albendazole had no significant effect. Azoles were next screened at the highest achievable concentration for non-CYP3A4 pathways. IC50 values (µM) were determined for most CYP3A4 pathways (ranges) and other pathways as dictated by screen results: clotrimazole (0.30 - 0.35; others >30 µM); econazole (2.2 - 4.9; 2B6 R-EDDP - 9.5, S-EDDP - 6.8; 2C8 - 6.0; 2C18 - 1.0; 2D6 - 1.2); fluconazole (7.7 - 66; 2B6 - 313, 361; 2C8 - 1240; 2C18 - 17; 2D6 - 1000); itraconazole (2.5 to >5; others >5); ketoconazole (0.032 - 0.094; 2B6 - 12, 31; 2C8 - 78; 2C18 - 0.98; 2D6 - 182); miconazole (2.3 - 7.6; 2B6 - 2.8, 2.8; 2C8 - 5.3; 2C18 - 3.1; 2D6 - 5.9); posaconazole (3.4 - 20; 2C18 - 3.8; others >30); terconazole (0.48 to >10; 2C18 - 8.1; others >10) and voriconazole (0.40 - 15; 2B6 - 2.4, 2.5; 2C8 - 170; 2C18 - 13; 2D6 >300). Modeling based on estimated Ki values and plasma concentrations from the literature suggest that the orally administered azoles, particularly

  20. In vitro and in vivo antiangiogenic activity of desacetylvinblastine monohydrazide through inhibition of VEGFR2 and Axl pathways

    PubMed Central

    Lei, Xueping; Chen, Minfeng; Nie, Qiulin; Hu, Jianyang; Zhuo, Zhenjian; Yiu, Anita; Chen, Heru; Xu, Nanhui; Huang, Maohua; Ye, Kaihe; Bai, Liangliang; Ye, Wencai; Zhang, Dongmei

    2016-01-01

    Tumor angiogenic process is regulated by multiple proangiogenic pathways, such as vascular endothelial growth factor receptor 2 (VEGFR2) and Axl receptor tyrosine kinase (Axl). Axl is one of many important factors involved in anti-VEGF resistance. Inhibition of VEGF/VEGFR2 signaling pathway alone fails to block tumor neovascularization. Therefore, discovery of novel agents targeting multiple angiogenesis pathways is in demand. Desacetylvinblastine monohydrazide (DAVLBH), a derivative of vinblastine (VLB), has been reported exhibit an anticancer activity via its cytotoxic effect. However, little attention has been paid to the antiangiogenic properties of DAVLBH. Here, we firstly reported that DAVLBH exerted a more potent antiangiogenic effect than VLB in vitro and in vivo, which was associated with inactivation of VEGF/VEGFR2 and Gas6/Axl signaling pathways. We found that DAVLBH inhibited VEGF- and Gas6-induced HUVECs proliferation, migration, tube formation and vessel sprouts formation in vitro and ex vivo. It significantly inhibited in vivo tumor angiogenesis and tumor growth in HeLa xenografts. It also inhibited Gas6-induced pericytes recruitment to endothelial tubes accompanied with a decrease in expression and activation of Axl. Besides, it could block the compensatory up-regulating expression and activation of Axl in response to bevacizumab treatment in HUVECs. Taken together, our results suggest that DAVLBH potently inhibits angiogenesis-mediated tumor growth through blockage of the activation of VEGF/VEGFR2 and Gas6/Axl pathways and it might serve as a promising antiangiogenic agent for the cancer therapy. PMID:27186435

  1. Raltegravir Has a Low Propensity To Cause Clinical Drug Interactions through Inhibition of Major Drug Transporters: an In Vitro Evaluation

    PubMed Central

    Houle, Robert; Chan, Grace Hoyee; Hafey, Mike; Rhee, Elizabeth G.; Chu, Xiaoyan

    2014-01-01

    Raltegravir (RAL) is a human immunodeficiency virus type 1 (HIV-1) integrase inhibitor approved to treat HIV infection in adults in combination with other antiretrovirals. The potential of RAL to cause transporter-related drug-drug interactions (DDIs) as an inhibitor has not been well described to date. In this study, a series of in vitro experiments were conducted to assess the inhibitory effects of RAL on major human drug transporters known to be involved in clinically relevant drug interactions, including hepatic and renal uptake transporters and efflux transporters. For hepatic uptake transporters, RAL showed no inhibition of organic anion-transporting polypeptide 1B1 (OATP1B1), weak inhibition of OATP1B3 (40% inhibition at 100 μM), and no inhibition of organic cation transporter 1 (OCT1). Studies of renal uptake transporters showed that RAL inhibited organic anion transporters 1 and 3 (OAT1 and OAT3) with 50% inhibitory concentrations (IC50s) (108 μM and 18.8 μM, respectively) well above the maximum concentration of drug in plasma (Cmax) at the clinical 400-mg dose and did not inhibit organic cation transporter 2 (OCT2). As for efflux transporters, RAL did not inhibit breast cancer resistance protein (BCRP) and showed weak inhibition of multidrug and toxin extrusion protein 1 (MATE1) (52% inhibition at 100 μM) and MATE2-K (29% inhibition at 100 μM). These studies indicate that at clinically relevant exposures, RAL does not inhibit or only weakly inhibits hepatic uptake transporters OATP1B1, OATP1B3, and OCT1, renal uptake transporters OCT2, OAT1, and OAT3, as well as efflux transporters BCRP, MATE1, and MATE2-K. The propensity for RAL to cause DDIs via inhibition of these transporters is therefore considered low. PMID:24295974

  2. Silibinin and indocyanine green-loaded nanoparticles inhibit the growth and metastasis of mammalian breast cancer cells in vitro

    PubMed Central

    Sun, Hui-ping; Su, Jing-han; Meng, Qing-shuo; Yin, Qi; Zhang, Zhi-wen; Yu, Hai-jun; Zhang, Peng-cheng; Wang, Si-ling; Li, Ya-ping

    2016-01-01

    Aim: To improve the therapeutic efficacy of cancer treatments, combinational therapies based on nanosized drug delivery system (NDDS) has been developed recently. In this study we designed a new NDDS loaded with an anti-metastatic drug silibinin and a photothermal agent indocyanine green (ICG), and investigated its effects on the growth and metastasis of breast cancer cells in vitro. Methods: Silibinin and ICG were self-assembled into PCL lipid nanoparticles (SIPNs). Their physical characteristics including the particle size, zeta potential, morphology and in vitro drug release were examined. 4T1 mammalian breast cancer cells were used to evaluate their cellular internalization, cytotoxicity, and their influences on wound healing, in vitro cell migration and invasion. Results: SIPNs showed a well-defined spherical shape with averaged size of 126.3±0.4 nm and zeta potential of −10.3±0.2 mV. NIR laser irradiation substantially increased the in vitro release of silibinin from the SIPNs (58.3% at the first 8 h, and 97.8% for the total release). Furthermore, NIR laser irradiation markedly increased the uptake of SIPNs into 4T1 cells. Under the NIR laser irradiation, both SIPNs and IPNs (PCL lipid nanoparticles loaded with ICG alone) caused dose-dependent ablation of 4T1 cells. The wound healing, migration and invasion experiments showed that SIPNs exposed to NIR laser irradiation exhibited dramatic in vitro anti-metastasis effects. Conclusion: SIPNs show temperature-sensitive drug release following NIR laser irradiation, which can inhibit the growth and metastasis of breast cancer cells in vitro. PMID:27133295

  3. Inhibition of elastase by a synthetic cotton-bound serine protease inhibitor: in vitro kinetics and inhibitor release.

    PubMed

    Edwards, J V; Bopp, A F; Batiste, S; Ullah, A J; Cohen, I K; Diegelmann, R F; Montante, S J

    1999-01-01

    A cotton-bound serine protease inhibitor of elastase (fiber-inhibitor) has been formulated for in vitro evaluation in chronic wound fluid. As a model to understand the properties of the inhibitor in wound dressings, the kinetic profile and in vitro release of the fiber-inhibitor formulation have been examined. The elastase inhibitor N-Methoxysuccinyl-Ala-Ala-Pro-Val-chloromethylketone was modified onto cotton cellulose fibers and assayed as a colloidal system. Amino acid analysis and reversed phase high performance liquid chromatography were compared as semiquantitative methods to assess elastase inhibitor release from the cotton fibers. The kinetics of inhibition was assessed on treated fibers of synthetic dressings such that a colloidal suspension of the fiber-inhibitor and elastase was employed as an assay. A dose-response relationship was observed in the kinetics of substrate hydrolysis catalyzed by three elastases: porcine pancreatic elastase, which was employed to model this approach; human leukocyte elastase; and elastase in human chronic wound fluid. Both freely dissolved and fiber-bound inhibitors were studied. The initial rates of substrate hydrolysis were inversely linear with freely dissolved inhibitor dose. The apparent first order rate constants, kobs, for the elastase-inhibitor complex were calculated from the kinetic profiles. The kobs for inhibitor bound enzyme varied as a function of inhibitor vs. enzyme concentration and based on the order of mixing of substrate, inhibitor and enzyme in the assay. Enzyme inhibition by the fiber-inhibitor was measured as inhibitor concentration at 50% inhibition (I50). I50 values measured from the colloidal assay with fiber-released inhibitor were within the same range to those for freely dissolved inhibitor. Inhibition of elastase activity in chronic wound fluid was observed with 1-5 mg of fiber-inhibitor formulation. This approach constitutes an in vitro assessment of synthetic serine protease inhibitors on

  4. Inhibition of APE1/Ref-1 redox activity with APX3330 blocks retinal angiogenesis in vitro and in vivo.

    PubMed

    Jiang, Aihua; Gao, Hua; Kelley, Mark R; Qiao, Xiaoxi

    2011-01-01

    This study examines the role of APE1/Ref-1 in the retina and its potential as a therapeutic target for inhibiting retinal angiogenesis. APE1/Ref-1 expression was quantified by Western blot. The role of APE1/Ref-1 redox function in endothelial cell in vitro angiogenesis was examined by treating retinal vascular endothelial cells (RVECs) with APX3330, a small molecule inhibitor of APE1/Ref-1 redox activity. In vitro methods included a proliferation assay, a transwell migration assay, a Matrigel tube formation assay, and a Real-Time Cell Analysis (RTCA) using the xCELLigence System. In vivo functional studies of APE1/Ref-1 were carried out by treating very low density lipoprotein (VLDL) receptor knockout mice (Vldlr(-/-)) with intravitreal injection of APX3330, and subsequent measurement of retinal angiomatous proliferation (RAP)-like neovascularization for one week. APE1/Ref-1 was highly expressed in the retina and in RVECs and pericytes in mice. APX3330 (1-10 μM) inhibited proliferation, migration and tube formation of RVECs in vitro in a dose-dependent manner. Vldlr(-/-) RVECs were more sensitive to APX3330 than wild-type RVECs. In Vldlr(-/-) mice, a single intravitreal injection of APX3330 at the onset of RAP-like neovascularization significantly reduced RAP-like neovascularization development. APE1/Ref-1 is expressed in retinal vascular cells. APX3330 inhibits RVEC angiogenesis in vitro and significantly reduces RAP-like neovascularization in Vldlr(-/-) mice. These data support the conclusion that APE1/Ref-1 redox function is required for retinal angiogenesis. Thus, APE1/Ref-1 may have potential as a therapeutic target for treating neovascular age-related macular degeneration and other neovascular diseases.

  5. Optimization of in vitro inhibition of HT-29 colon cancer cell cultures by Solanum tuberosum L. extracts.

    PubMed

    Zuber, T; Holm, D; Byrne, P; Ducreux, L; Taylor, M; Kaiser, M; Stushnoff, C

    2015-01-01

    Secondary metabolites in potato have been reported to possess bioactive properties, including growth inhibition of cancer cells. Because potatoes are widely consumed globally, potential health benefits may have broad application. Thus we investigated growth inhibition of HT-29 colon cancer cell cultures by extracts from 13 diverse genetic breeding clones. Extracts from three pigmented selections (CO97226-2R/R, CO97216-1P/P, CO04058-3RW/RW) inhibited growth of in vitro HT-29 cell cultures more effectively than other clones tested. While inhibition was highest from pigmented selections and pigmented tuber tissue sectors, not all pigmented breeding lines tested had appreciable inhibitory properties. Thus, inhibition was not uniquely linked to pigmentation. Immature tubers had the highest inhibitory properties, and in most cases mature tubers retained very low inhibition properties. Flowers and skins inhibited strongly at lower extract concentrations. An extract consisting of 7.2 mg mL⁻¹ cell culture medium was the lowest effective concentration. While raw tuber extracts inhibited most effectively, a few clones at higher concentrations retained inhibition after cooking. Heated whole tubers retained higher inhibition than heated aqueous extracts. While all aqueous extracts from the two tuber selections (CO97216-1P/P and CO97226-2R/R) inhibited HT-29 cell cultures, inhibition was significantly enhanced in purple pigmented tubers of CO97216-1P/P prepared cryogenically as liquid nitrogen powders compared to extracts from freeze dried samples. Upregulation of caspase-3 protease activity, indicative of apoptosis, was highest among the most inhibitory clone samples. The unique sectorial red pigment expressing selection (CO04058-3RW/RW) provided a model system that isolated expression in pigmented sectors, and thus eliminated developmental, environmental and genetic confounding. PMID:25338312

  6. Optimization of in vitro inhibition of HT-29 colon cancer cell cultures by Solanum tuberosum L. extracts.

    PubMed

    Zuber, T; Holm, D; Byrne, P; Ducreux, L; Taylor, M; Kaiser, M; Stushnoff, C

    2015-01-01

    Secondary metabolites in potato have been reported to possess bioactive properties, including growth inhibition of cancer cells. Because potatoes are widely consumed globally, potential health benefits may have broad application. Thus we investigated growth inhibition of HT-29 colon cancer cell cultures by extracts from 13 diverse genetic breeding clones. Extracts from three pigmented selections (CO97226-2R/R, CO97216-1P/P, CO04058-3RW/RW) inhibited growth of in vitro HT-29 cell cultures more effectively than other clones tested. While inhibition was highest from pigmented selections and pigmented tuber tissue sectors, not all pigmented breeding lines tested had appreciable inhibitory properties. Thus, inhibition was not uniquely linked to pigmentation. Immature tubers had the highest inhibitory properties, and in most cases mature tubers retained very low inhibition properties. Flowers and skins inhibited strongly at lower extract concentrations. An extract consisting of 7.2 mg mL⁻¹ cell culture medium was the lowest effective concentration. While raw tuber extracts inhibited most effectively, a few clones at higher concentrations retained inhibition after cooking. Heated whole tubers retained higher inhibition than heated aqueous extracts. While all aqueous extracts from the two tuber selections (CO97216-1P/P and CO97226-2R/R) inhibited HT-29 cell cultures, inhibition was significantly enhanced in purple pigmented tubers of CO97216-1P/P prepared cryogenically as liquid nitrogen powders compared to extracts from freeze dried samples. Upregulation of caspase-3 protease activity, indicative of apoptosis, was highest among the most inhibitory clone samples. The unique sectorial red pigment expressing selection (CO04058-3RW/RW) provided a model system that isolated expression in pigmented sectors, and thus eliminated developmental, environmental and genetic confounding.

  7. Alginate Oligosaccharides Inhibit Fungal Cell Growth and Potentiate the Activity of Antifungals against Candida and Aspergillus spp

    PubMed Central

    Tøndervik, Anne; Sletta, Håvard; Klinkenberg, Geir; Emanuel, Charlotte; Powell, Lydia C.; Pritchard, Manon F.; Khan, Saira; Craine, Kieron M.; Onsøyen, Edvar; Rye, Phil D.; Wright, Chris; Thomas, David W.; Hill, Katja E.

    2014-01-01

    The oligosaccharide OligoG, an alginate derived from seaweed, has been shown to have anti-bacterial and anti-biofilm properties and potentiates the activity of selected antibiotics against multi-drug resistant bacteria. The ability of OligoG to perturb fungal growth and potentiate conventional antifungal agents was evaluated using a range of pathogenic fungal strains. Candida (n = 11) and Aspergillus (n = 3) spp. were tested using germ tube assays, LIVE/DEAD staining, scanning electron microscopy (SEM), atomic force microscopy (AFM) and high-throughput minimum inhibition concentration assays (MICs). In general, the strains tested showed a significant dose-dependent reduction in cell growth at ≥6% OligoG as measured by optical density (OD600; P<0.05). OligoG (>0.5%) also showed a significant inhibitory effect on hyphal growth in germ tube assays, although strain-dependent variations in efficacy were observed (P<0.05). SEM and AFM both showed that OligoG (≥2%) markedly disrupted fungal biofilm formation, both alone, and in combination with fluconazole. Cell surface roughness was also significantly increased by the combination treatment (P<0.001). High-throughput robotic MIC screening demonstrated the potentiating effects of OligoG (2, 6, 10%) with nystatin, amphotericin B, fluconazole, miconazole, voriconazole or terbinafine with the test strains. Potentiating effects were observed for the Aspergillus strains with all six antifungal agents, with an up to 16-fold (nystatin) reduction in MIC. Similarly, all the Candida spp. showed potentiation with nystatin (up to 16-fold) and fluconazole (up to 8-fold). These findings demonstrate the antifungal properties of OligoG and suggest a potential role in the management of fungal infections and possible reduction of antifungal toxicity. PMID:25409186

  8. GSK-3 inhibition in vitro and in vivo enhances antitumor effect of sorafenib in renal cell carcinoma (RCC)

    SciTech Connect

    Kawazoe, Hisashi; Bilim, Vladimir N.; Ugolkov, Andrey V.; Yuuki, Kaori; Naito, Sei; Nagaoka, Akira; Kato, Tomoyuki; Tomita, Yoshihiko

    2012-07-06

    Highlights: Black-Right-Pointing-Pointer Sorafenib treatment upregulated GSK-3{beta} levels in RCC cells. Black-Right-Pointing-Pointer Pharmacologic inhibition of GSK-3 suppressed xenograft RCC tumor growth. Black-Right-Pointing-Pointer Inhibition of GSK-3 enhanced antitumor effect of sorafenib in vitro and in vivo. -- Abstract: Sorafenib is a multikinase inhibitor approved for the systemic treatment of renal cell carcinoma (RCC). However, sorafenib treatment has a limited effect due to acquired chemoresistance of RCC. Previously, we identified glycogen synthase kinase-3 (GSK-3) as a new therapeutic target in RCC. Here, we observed that sorafenib inhibits proliferation and survival of RCC cells. Significantly, we revealed that sorafenib enhances GSK-3 activity in RCC cells, which could be a potential mechanism of acquired chemoresistance. We found that pharmacological inhibition of GSK-3 potentiates sorafenib antitumor effect in vitro and in vivo. Our results suggest that combining GSK-3 inhibitor and sorafenib might be a potential new therapeutic approach for RCC treatment.

  9. Alginate as a protease inhibitor in vitro and in a model gut system; selective inhibition of pepsin but not trypsin

    PubMed Central

    Chater, Peter Ian; Wilcox, Mathew D.; Brownlee, Iain A.; Pearson, Jeffrey P.

    2015-01-01

    Alginates are widely used in the food and medical industries, including as a Gastro-Oesophagul Reflux treatment. This work investigates the inhibitory effects of alginate on the reflux aggressors trypsin and pepsin and the role of alginate-substrate binding, pH and alginate structure on inhibition. Alginates were shown to reduce pepsin activity by up to 53.9% (±9.5SD) in vitro. Strong positive correlation between alginate mannuronate residue frequency and levels of pepsin inhibition was observed. Limited inhibition of trypsin was shown. Viscometric observations of pH dependent interactions between alginate and protein suggest a mechanism whereby pH dependent ionic interactions reduce substrate availability to enzyme at acidic pH. To understand how dietary protein digestion is affected by alginate, proteolytic digestion was investigated in an in vitro model of the upper digestive tract. Significant inhibition of proteolysis was shown in the gastric phase of digestion, but not the small intestinal phase. PMID:26256170

  10. Inhibition of cytochrome P450 3A by acetoxylated analogues of resveratrol in in vitro and in silico models

    NASA Astrophysics Data System (ADS)

    Basheer, Loai; Schultz, Keren; Kerem, Zohar

    2016-08-01

    Many dietary compounds, including resveratrol, are potent inhibitors of CYP3A4. Here we examined the potential to predict inhibition capacity of dietary polyphenolics using an in silico and in vitro approaches and synthetic model compounds. Mono, di, and tri-acetoxy resveratrol were synthesized, a cell line of human intestine origin and microsomes from rat liver served to determine their in vitro inhibition of CYP3A4, and compared to that of resveratrol. Docking simulation served to predict the affinity of the synthetic model compounds to the enzyme. Modelling of the enzyme’s binding site revealed three types of interaction: hydrophobic, electrostatic and H-bonding. The simulation revealed that each of the examined acetylations of resveratrol led to the loss of important interactions of all types. Tri-acetoxy resveratrol was the weakest inhibitor in vitro despite being the more lipophilic and having the highest affinity for the binding site. The simulation demonstrated exclusion of all interactions between tri-acetoxy resveratrol and the heme due to distal binding, highlighting the complexity of the CYP3A4 binding site, which may allow simultaneous accommodation of two molecules. Finally, the use of computational modelling may serve as a quick predictive tool to identify potential harmful interactions between dietary compounds and prescribed drugs.

  11. Inhibition of cytochrome P450 3A by acetoxylated analogues of resveratrol in in vitro and in silico models.

    PubMed

    Basheer, Loai; Schultz, Keren; Kerem, Zohar

    2016-01-01

    Many dietary compounds, including resveratrol, are potent inhibitors of CYP3A4. Here we examined the potential to predict inhibition capacity of dietary polyphenolics using an in silico and in vitro approaches and synthetic model compounds. Mono, di, and tri-acetoxy resveratrol were synthesized, a cell line of human intestine origin and microsomes from rat liver served to determine their in vitro inhibition of CYP3A4, and compared to that of resveratrol. Docking simulation served to predict the affinity of the synthetic model compounds to the enzyme. Modelling of the enzyme's binding site revealed three types of interaction: hydrophobic, electrostatic and H-bonding. The simulation revealed that each of the examined acetylations of resveratrol led to the loss of important interactions of all types. Tri-acetoxy resveratrol was the weakest inhibitor in vitro despite being the more lipophilic and having the highest affinity for the binding site. The simulation demonstrated exclusion of all interactions between tri-acetoxy resveratrol and the heme due to distal binding, highlighting the complexity of the CYP3A4 binding site, which may allow simultaneous accommodation of two molecules. Finally, the use of computational modelling may serve as a quick predictive tool to identify potential harmful interactions between dietary compounds and prescribed drugs. PMID:27530542

  12. Inhibition of multixenobiotic resistance transporters (MXR) by silver nanoparticles and ions in vitro and in Daphnia magna.

    PubMed

    Georgantzopoulou, Anastasia; Cambier, Sébastien; Serchi, Tommaso; Kruszewski, Marcin; Balachandran, Yekkuni L; Grysan, Patrick; Audinot, Jean-Nicolas; Ziebel, Johanna; Guignard, Cédric; Gutleb, Arno C; Murk, AlberTinka J

    2016-11-01

    The P-glycoprotein (P-gp, ABCB1) and multidrug resistance associated protein 1 (MRP1), important members of the ABC (ATP-binding cassette) transporters, protect cells and organisms via efflux of xenobiotics and are responsible for the phenomenon of multidrug or multixenobiotic resistance (MXR). In this study we first evaluated, in vitro, the interaction of silver nanoparticles (Ag NPs, 20, 23 and 27nm), Ag 200nm particles and Ag ions (AgNO3) with MXR efflux transporters using MDCKII and the P-gp over-expressing MDCKII-MDR1 cells and calcein-AM as a substrate of the transporters. Next the in vivo modulation of MXR activity was studied in Daphnia magna juveniles with the model P-gp and MRP1 inhibitors verapamil-HCl and MK571, respectively. The common environmental contaminants perfluorooctane sulfonate and bisphenol A, previously observed to interfere with the P-gp in vitro, also inhibited the efflux of calcein in vivo. Small-sized Ag NPs (with biomolecules present on the surface) and AgNO3 inhibited the MXR activity in daphnids and MDCKII-MDR1 cells, but abcb1 gene expression remained unchanged. Both Ag NPs and dissolved ions contributed to the effects. This study provides evidence of the interference of Ag NPs and AgNO3 with the MXR activity both in vitro and in D. magna, and should be taken into account when Ag NP toxicity is assessed. PMID:27376922

  13. Inhibition of cytochrome P450 3A by acetoxylated analogues of resveratrol in in vitro and in silico models

    PubMed Central

    Basheer, Loai; Schultz, Keren; Kerem, Zohar

    2016-01-01

    Many dietary compounds, including resveratrol, are potent inhibitors of CYP3A4. Here we examined the potential to predict inhibition capacity of dietary polyphenolics using an in silico and in vitro approaches and synthetic model compounds. Mono, di, and tri-acetoxy resveratrol were synthesized, a cell line of human intestine origin and microsomes from rat liver served to determine their in vitro inhibition of CYP3A4, and compared to that of resveratrol. Docking simulation served to predict the affinity of the synthetic model compounds to the enzyme. Modelling of the enzyme’s binding site revealed three types of interaction: hydrophobic, electrostatic and H-bonding. The simulation revealed that each of the examined acetylations of resveratrol led to the loss of important interactions of all types. Tri-acetoxy resveratrol was the weakest inhibitor in vitro despite being the more lipophilic and having the highest affinity for the binding site. The simulation demonstrated exclusion of all interactions between tri-acetoxy resveratrol and the heme due to distal binding, highlighting the complexity of the CYP3A4 binding site, which may allow simultaneous accommodation of two molecules. Finally, the use of computational modelling may serve as a quick predictive tool to identify potential harmful interactions between dietary compounds and prescribed drugs. PMID:27530542

  14. Multi-drug resistant oral Candida species isolated from HIV-positive patients in South Africa and Cameroon.

    PubMed

    Dos Santos Abrantes, Pedro Miguel; McArthur, Carole P; Africa, Charlene Wilma Joyce

    2014-06-01

    Candida species are a common cause of infection in immune-compromised HIV-positive individuals, who are usually treated with the antifungal drug, fluconazole, in public hospitals in Africa. However, information about the prevalence of drug resistance to fluconazole and other antifungal agents on Candida species is very limited. This study examined 128 Candida isolates from South Africa and 126 Cameroonian Candida isolates for determination of species prevalence and antifungal drug susceptibility. The isolates were characterized by growth on chromogenic and selective media and by their susceptibility to 9 antifungal drugs tested using the TREK™ YeastOne9 drug panel (Thermo Scientific, USA). Eighty-three percent (82.8%) of South African isolates were Candida albicans (106 isolates), 9.4% were Candida glabrata (12 isolates), and 7.8% were Candida dubliniensis (10 isolates). Of the Cameroonian isolates, 73.02% were C. albicans (92 isolates); 19.05% C. glabrata (24 isolates); 3.2% Candida tropicalis (4 isolates); 2.4% Candida krusei (3 isolates); 1.59% either Candida kefyr, Candida parapsilopsis, or Candida lusitaneae (2 isolates); and 0.79% C. dubliniensis (1 isolate). Widespread C. albicans resistance to azoles was detected phenotypically in both populations. Differences in drug resistance were seen within C. glabrata found in both populations. Echinocandin drugs were more effective on isolates obtained from the Cameroon than in South Africa. A multiple-drug resistant C. dubliniensis strain isolated from the South African samples was inhibited only by 5-flucytosine in vitro on the YO9 panel. Drug resistance among oral Candida species is common among African HIV patients in these 2 countries. Regional surveillance of Candida species drug susceptibility should be undertaken to ensure effective treatment for HIV-positive patients.

  15. Comparison of in vitro and ex vivo thyroid hormone synthesis inhibition results and in vivo outcomes for a series of benzothiazoles

    EPA Science Inventory

    Assessing how in vitro data may be used to predict adverse effects in vivo is critical as efforts are advanced to incorporate in vitro assays into a risk assessment framework. Within the context of a thyroid hormone (TH) synthesis inhibition adverse outcome pathway (AOP), in vitr...

  16. Streptococcus thermophilus and its biosurfactants inhibit adhesion by Candida spp. on silicone rubber.

    PubMed

    Busscher, H J; van Hoogmoed, C G; Geertsema-Doornbusch, G I; van der Kuijl-Booij, M; van der Mei, H C

    1997-10-01

    The adhesion of yeasts, two Candida albicans and two Candida tropicalis strains isolated from naturally colonized voice prostheses, to silicone rubber with and without a salivary conditioning film in the absence and presence of adhering Streptococcus thermophilus B, a biosurfactant-releasing dairy isolate, was studied. Coverage of 1 to 4% of the surface of silicone rubber substrata with adhering S. thermophilus B gave significant reductions in the initial yeast adhesion regardless of the presence of a conditioning film. Mechanistically, this interference in yeast adhesion by S. thermophilus B was not due to direct physical effects but to biosurfactant release by the adhering bacteria, because experiments with S. thermophilus B cells that had released their biosurfactants prior to adhesion to silicone rubber and competition with yeasts did not show interference with initial yeast adhesion. The amounts of biosurfactants released were highest for mid-exponential- and early-stationary-phase bacteria (37 mg.g of cells-1 [dry weight]), but biosurfactants released by stationary-phase bacteria (14 mg.g of cells-1 [dry weight]) were the most surface active. The crude biosurfactants released were mixtures of various components, with a glycolipid-like component being the most surface active. A lipid-enriched biosurfactant fraction reduced the surface tension of an aqueous solution to about 35 mJ.m-2 at a concentration of only 0.5 mg.ml-1. The amount of biosurfactant released per S. thermophilus B cell was estimated to be sufficient to cover approximately 12 times the area of the cross section of the bacterium, making biosurfactant release a powerful defense weapon in the postadhesion competition of the bacterium with microorganisms such as yeasts. Preadsorption of biosurfactants to the silicone rubber prior to allowing yeasts to adhere was as effective against C. albicans GB 1/2 adhesion as covering 1 to 2% of the silicone rubber surface with adhering S. thermophilus B, but a

  17. [In vitro inhibition of granulopoiesis by beta-lactam antibiotics. Comparison of piperacillin, mezlocillin, ceftriaxone and ceftazidime].

    PubMed

    Marie, J P; Thevenin, D; Zittoun, R

    1986-12-20

    The mechanism of neutropenia induced by beta-lactam antibiotics was explored by studying the action of these drugs on granulopoiesis in vitro. Normal bone marrows were cultivated in the presence of increasing concentrations of piperacillin (10 marrows), mezlocillin, ceftriaxone and ceftazidime (5 marrows each) in order to find out whether these antibiotics exhibited toxicity to granulocyte-monocyte precursors. A dose-dependent inhibition of granulopoiesis was found in all cases. When the doses used were equivalent to maximum plasma concentrations in vivo, inhibition was minimal with piperacillin and mezlocillin and much more pronounced with the cephalosporins. This dose-dependent inhibition suggests that toxicity is involved in the mechanism of neutropenia induced by beta-lactam antibiotics.

  18. Boric acid destabilizes the hyphal cytoskeleton and inhibits invasive growth of Candida albicans.

    PubMed

    Pointer, Benjamin R; Boyer, Michael P; Schmidt, Martin

    2015-04-01

    Exposure of Candida albicans to sub-lethal concentrations of boric acid (BA) restricts the dimorphic fungus to its yeast morphology and prevents the formation of invasive hyphae on solid substrates. Exposure to BA causes a rapid and reversible disappearance of polarisome and Spitzenkörper in growing hyphae. In BA-treated hyphae of C. albicans, actin quickly reorganizes from cytoplasmic cables to cortical patches and cell wall growth switches from an apical to an isotropic pattern. As a result of the cytoskeletal changes, the hyphal tips broaden and directional growth of hyphae ceases in the presence of BA. An analysis of homozygous deletion strains showed that mutants with constitutive or enhanced hyphal growth (tup1, nrg1, ssn6, rbf1) are BA-sensitive, demonstrating that cellular morphology is a major determinant of BA tolerance. The screening of deletion mutants also showed that deficiencies of the main activator of hyphal gene expression, Efg1, and the Rim101-signalling cascade, leading to Efg1 activation, cause BA resistance. Taken together, the data presented show that the selective inhibitory effect on BA on C. albicans hyphae is rooted in a disruption of apical cytoskeletal elements of growing hyphae.

  19. In vitro growth of multidrug-resistant Neisseria gonorrhoeae isolates is inhibited by ETX0914, a novel spiropyrimidinetrione.

    PubMed

    Papp, John R; Lawrence, Kenneth; Sharpe, Samera; Mueller, John; Kirkcaldy, Robert D

    2016-09-01

    Antimicrobial resistance in Neisseria gonorrhoeae has severely limited the number of treatment options, and the emergence of extended-spectrum cephalosporin resistance threatens the effectiveness of the last remaining recommended treatment regimen. This study assessed the in vitro susceptibility of N. gonorrhoeae to ETX0914, a novel spiropyrimidinetrione that inhibits DNA biosynthesis. In vitro activity was determined by agar dilution against 100 N. gonorrhoeae isolates collected from men presenting with urethritis in the USA during 2012-2013 through the Gonococcal Isolate Surveillance Project. The minimum inhibitory concentration (MIC) that inhibited growth in 50% (MIC50) and 90% (MIC90) of isolates was calculated for each antimicrobial agent. ETX0914 demonstrated a high level of antimicrobial activity against N. gonorrhoeae, including isolates with decreased susceptibility or resistance to currently available agents. The ability of ETX0914 to inhibit the growth of N. gonorrhoeae was similar to ceftriaxone, which is currently recommended in combination with azithromycin to treat gonorrhoea. The data presented in this study strongly suggest that ETX0914 should be evaluated in a clinical trial for the treatment of N. gonorrhoeae.

  20. In vitro growth of multidrug-resistant Neisseria gonorrhoeae isolates is inhibited by ETX0914, a novel spiropyrimidinetrione.

    PubMed

    Papp, John R; Lawrence, Kenneth; Sharpe, Samera; Mueller, John; Kirkcaldy, Robert D

    2016-09-01

    Antimicrobial resistance in Neisseria gonorrhoeae has severely limited the number of treatment options, and the emergence of extended-spectrum cephalosporin resistance threatens the effectiveness of the last remaining recommended treatment regimen. This study assessed the in vitro susceptibility of N. gonorrhoeae to ETX0914, a novel spiropyrimidinetrione that inhibits DNA biosynthesis. In vitro activity was determined by agar dilution against 100 N. gonorrhoeae isolates collected from men presenting with urethritis in the USA during 2012-2013 through the Gonococcal Isolate Surveillance Project. The minimum inhibitory concentration (MIC) that inhibited growth in 50% (MIC50) and 90% (MIC90) of isolates was calculated for each antimicrobial agent. ETX0914 demonstrated a high level of antimicrobial activity against N. gonorrhoeae, including isolates with decreased susceptibility or resistance to currently available agents. The ability of ETX0914 to inhibit the growth of N. gonorrhoeae was similar to ceftriaxone, which is currently recommended in combination with azithromycin to treat gonorrhoea. The data presented in this study strongly suggest that ETX0914 should be evaluated in a clinical trial for the treatment of N. gonorrhoeae. PMID:27499432

  1. The fungicide Pristine® inhibits mitochondrial function in vitro but not flight metabolic rates in honey bees.

    PubMed

    Campbell, Jacob B; Nath, Rachna; Gadau, Juergen; Fox, Trevor; DeGrandi-Hoffman, Gloria; Harrison, Jon F

    2016-03-01

    Honey bees and other pollinators are exposed to fungicides that act by inhibiting fungal mitochondria. Here we test whether a common fungicide (Pristine®) inhibits the function of mitochondria of honeybees, and whether consumption of ecologically-realistic concentrations can cause negative effects on the mitochondria of flight muscles, or the capability for flight, as judged by CO2 emission rates and thorax temperatures during flight. Direct exposure of mitochondria to Pristine® levels above 5 ppm strongly inhibited mitochondrial oxidation rates in vitro. However, bees that consumed pollen containing Pristine® at ecologically-realistic concentrations (≈ 1 ppm) had normal flight CO2 emission rates and thorax temperatures. Mitochondria isolated from the flight muscles of the Pristine®-consuming bees had higher state 3 oxygen consumption rates than control bees, suggesting that possibly Pristine®-consumption caused compensatory changes in mitochondria. It is likely that the lack of a strong functional effect of Pristine®-consumption on flight performance and the in vitro function of flight muscle mitochondria results from maintenance of Pristine® levels in the flight muscles at much lower levels than occur in the food, probably due to metabolism and detoxification. As Pristine® has been shown to negatively affect feeding rates and protein digestion of honey bees, it is plausible that Pristine® consumption negatively affects gut wall function (where mitochondria may be exposed to higher concentrations of Pristine®).

  2. Taxifolin prevents diabetic cardiomyopathy in vivo and in vitro by inhibition of oxidative stress and cell apoptosis.

    PubMed

    Sun, Xiao; Chen, Rong-chang; Yang, Zhi-hong; Sun, Gui-bo; Wang, Min; Ma, Xiao-jun; Yang, Li-juan; Sun, Xiao-bo

    2014-01-01

    Diabetic cardiomyopathy has been increasingly recognized as an important cause of heart failure in diabetic patients. Excessive oxidative stress has been suggested to play a critical role in the development of diabetic cardiomyopathy. The objective of this study was to investigate the potential protective effects and mechanisms of taxifolin on cardiac function of streptozotocin-induced diabetic mice and on hyperglycemia-induced apoptosis of H9c2 cardiac myoblasts. In vivo study revealed that taxifolin improved diastolic dysfunction, ameliorated myocardium structure abnormality, inhibited myocyte apoptosis and enhanced endogenous antioxidant enzymes activities. Interestingly, taxifolin reduced angiotensin II level in myocardium, inhibited NADPH oxidase activity, and increased JAK/STAT3 activation. In vitro investigation demonstrated that taxifolin inhibited 33 mM glucoseinduced H9c2 cells apoptosis by decreasing intracellular ROS level. It also inhibited caspase-3 and caspase-9 activation, restored mitochondrial membrane potential, and regulated the expression of proteins related to the intrinsic pathway of apoptosis, thus inhibiting the release of cytochrome c from mitochondria into the cytoplasm. In conclusion, taxifolin exerted cardioprotective effects against diabetic cardiomyopathy by inhibiting oxidative stress and cardiac myocyte apoptosis and might be a potential agent in the treatment of diabetic cardiomyopathy. PMID:24269735

  3. Acacetin inhibits in vitro and in vivo angiogenesis and down-regulates Stat signaling and VEGF expression

    PubMed Central

    Bhat, Tariq A.; Nambiar, Dhanya; Tailor, Dhanir; Pal, Arttatrana; Agarwal, Rajesh; Singh, Rana P.

    2013-01-01

    Angiogenesis is an effective target in cancer control. The anti-angiogenic efficacy and associated mechanisms of acacetin, a plant flavone, is poorly known. In the present study, acacetin inhibited growth and survival (upto 92%, p<0.001), and capillary-like tube formation on matrigel (upto 98%, p<0.001) by human umbilical vein endothelial cells (HUVEC) in regular condition, as well as VEGF-induced and tumor cells conditioned medium-stimulated growth conditions. It caused retraction and disintegration of preformed capillary networks (upto 91%, p<0.001). HUVEC migration and invasion were suppressed by 68-100% (p<0.001). Acacetin inhibited Stat-1 (Tyr701) and Stat-3 (Tyr705) phosphorylation, and down-regulated pro-angiogenic factors including VEGF, eNOS, iNOS, MMP-2 and bFGF in HUVEC. It also suppressed nuclear localization of pStat-3 (Tyr705). Acacetin strongly inhibited capillary sprouting and networking from rat aortic rings and fertilized chicken egg chorioallantoic membrane (CAM) (~71%, p<0.001). Furthermore, it suppressed angiogenesis in matrigel plugs implanted in Swiss albino mice. Acacetin also inhibited tyrosine phosphorylation of Stat-1 and Stat-3, and expression of VEGF in cancer cells. Overall, acacetin inhibits Stat signaling and suppresses angiogenesis in vitro, ex vivo and in vivo, and therefore, it could be a potential agent to inhibit tumor angiogenesis and growth. PMID:23943785

  4. An extensive cocktail approach for rapid risk assessment of in vitro CYP450 direct reversible inhibition by xenobiotic exposure.

    PubMed

    Spaggiari, Dany; Daali, Youssef; Rudaz, Serge

    2016-07-01

    Acute exposure to environmental factors strongly affects the metabolic activity of cytochrome P450 (P450). As a consequence, the risk of interaction could be increased, modifying the clinical outcomes of a medication. Because toxic agents cannot be administered to humans for ethical reasons, in vitro approaches are therefore essential to evaluate their impact on P450 activities. In this work, an extensive cocktail mixture was developed and validated for in vitro P450 inhibition studies using human liver microsomes (HLM). The cocktail comprised eleven P450-specific probe substrates to simultaneously assess the activities of the following isoforms: 1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1, 2J2 and subfamily 3A. The high selectivity and sensitivity of the developed UHPLC-MS/MS method were critical for the success of this methodology, whose main advantages are: (i) the use of eleven probe substrates with minimized interactions, (ii) a low HLM concentration, (iii) fast incubation (5min) and (iv) the use of metabolic ratios as microsomal P450 activities markers. This cocktail approach was successfully validated by comparing the obtained IC50 values for model inhibitors with those generated with the conventional single probe methods. Accordingly, reliable inhibition values could be generated 10-fold faster using a 10-fold smaller amount of HLM compared to individual assays. This approach was applied to assess the P450 inhibition potential of widespread insecticides, namely, chlorpyrifos, fenitrothion, methylparathion and profenofos. In all cases, P450 2B6 was the most affected with IC50 values in the nanomolar range. For the first time, mixtures of these four insecticides incubated at low concentrations showed a cumulative inhibitory in vitro effect on P450 2B6. PMID:27105555

  5. A prostate-specific antigen-dependent fusion polypeptide inhibits growth of prostate cancer cells in vitro and in vivo

    PubMed Central

    Zhang, Xiang; Ma, Yueyun; Wei, Hua; Li, Bin; Xiao, Fengjing; Yang, Jing; Yue, Qiaohong; Yang, Angang; Hao, Xiaoke

    2016-01-01

    Polypeptide APP8 is a prostate-specific antigen (PSA)-activated prodrug that was designed to synergize the effects of the Bcl-2 homology domain 3 (BH3) peptide, K237 and the DG2 peptide. The aim of this study is to evaluate its biodistribution and anticancer effect in vitro and in vivo. In this study, APP8 and each component peptide were synthesized. The biodistribution was identified using con-focal microscopyin both PSA+ cell line and PSA- cell line in vitro. Then cell cycle, MTT and in-cell western blot were accessed to analyze the effect mechanisms. Finally, xenografts were used to confirm the anticancer effect in vivo. Here, it was shown that APP8 was hydrolyzed and BH3 was released into the nucleus, while K237 and DG2 were located predominantly in the cytoplasm, only in LNCaP cells (PSA+), but not PC3 cells (PSA-). K237 and DG2 could induce cell apoptosis through decreasing the phosphorylation of ERK-2 and Flk-1. APP8 also caused the death of LNCaP cells, and was predominantly dependent on BH3 in vitro. In addition, It was noted that as the tumor grew in vivo, APP8 could inhibit the tumor volume to 77.3%, mainly depending on K237 and DG2 via inhibition of the growth of vascular endothelial cells. Our results suggested that APP8 could promote prostate cancer cell death and stop prostate cancer growth via synergizing apoptosis induction of tumor cell and inhibition of the growth of vascular endothelial cells. It provides a novel candidate prodrug for specific therapy of prostate cancer. PMID:27293998

  6. AGE-RELATED BRAIN CHOLINESTERASE INHIBITION KINETICS FOLLOWING IN VITRO INCUBATION WITH CHLORPYRIFOS-OXON AND DIAZINON-OXON

    SciTech Connect

    Kousba, Ahmed A.; Poet, Torka S.; Timchalk, Chuck

    2007-01-01

    Chlorpyrifos and diazinon are two commonly used organophosphorus (OP) insecticides, and their primary mechanism of action involves the inhibition of acetylcholinesterase (AChE) by their metabolites chlorpyrifos-oxon (CPO) and diazinon-oxon (DZO), respectively. The study objectives were to assess the in vitro age-related inhibition kinetics of neonatal rat brain cholinesterase (ChE) by estimating the bimolecular inhibitory rate constant (ki) values for CPO and DZO. Brain ChE inhibition and ki values following CPO and DZO incubation with neonatal Sprague-Dawley rats rat brain homogenates were determined at post natal day (PND) -5, -12 and -17 and compared with the corresponding inhibition and ki values obtained in the adult rat. A modified Ellman method was utilized for measuring the ChE activity. Chlorpyrifos-oxon resulted in greater ChE inhibition than DZO consistent with the estimated ki values of both compounds. Neonatal brain ChE inhibition kinetics exhibited a marked age-related sensitivity to CPO, where the order of ChE inhibition was PND-5 > PND-7 > PND-17 with ki values of 0.95, 0.50 and 0.22 nM-1hr-1, respectively. In contrast, DZO did not exhibit an age-related inhibition of neonatal brain ChE, and the estimated ki value at all PND ages was 0.02 nM-1hr-1. These results demonstrated an age- and chemical-related OP-selective inhibition of rat brain ChE which may be critically important in understanding the potential sensitivity of juvenile humans to specific OP exposures.

  7. Monothiocarbamates Strongly Inhibit Carbonic Anhydrases in Vitro and Possess Intraocular Pressure Lowering Activity in an Animal Model of Glaucoma.

    PubMed

    Vullo, Daniela; Durante, Mariaconcetta; Di Leva, Francesco Saverio; Cosconati, Sandro; Masini, Emanuela; Scozzafava, Andrea; Novellino, Ettore; Supuran, Claudiu T; Carta, Fabrizio

    2016-06-23

    A series of monothiocarbamates (MTCs) were prepared from primary/secondary amines and COS as potential carbonic anhydrase (CA, EC 4.2.1.1) inhibitors, using the dithiocarbamates, the xanthates, and the trithiocarbonates as lead compounds. The MTCs effectively inhibited the pharmacologically relevant human (h) hCAs isoforms I, II, IX, and XII in vitro and showed KIs spanning between the low and medium nanomolar range. By means of a computational study, the MTC moiety binding mode on the CAs was explained. Furthermore, a selection of MTCs were evaluated in a normotensive glaucoma rabbit model for their intraocular pressure (IOP) lowering effects and showed interesting activity. PMID:27253845

  8. Celastrol inhibits prostaglandin E2-induced proliferation and osteogenic differentiation of fibroblasts isolated from ankylosing spondylitis hip tissues in vitro

    PubMed Central

    Zou, Yu-Cong; Yang, Xian-Wen; Yuan, Shi-Guo; Zhang, Pei; Li, Yi-Kai

    2016-01-01

    Background Heterotopic ossification on the enthesis, which develops after subsequent inflammation, is one of the most distinctive features in ankylosing spondylitis (AS). Prostaglandin E2 (PGE-2) serves as a key mediator of inflammation and bone remodeling in AS. Celastrol, a well-known Chinese medicinal herb isolated from Tripterygium wilfordii, is widely used in treating inflammatory diseases, including AS. It has been proven that it can inhibit lipopolysac-charide-induced expression of various inflammation mediators, such as PGE-2. However, the mechanism by which celastrol inhibits inflammation-induced bone forming in AS is unclear. Objective To investigate whether celastrol could inhibit isolated AS fibroblast osteogenesis induced by PGE-2. Methods Hip synovial tissues were obtained from six AS patients undergoing total hip replacement in our hospital. Fibroblasts were isolated, primarily cultured, and then treated with PGE-2 for osteogenic induction. Different doses of celastrol and indometacin were added to observe their effects on osteogenic differentiation. Cell proliferation, osteogenic markers, alizarin red staining as well as the activity of alkaline phosphatase were examined in our study. Results Celastrol significantly inhibits cell proliferation of isolated AS fibroblasts and in vitro osteogenic differentiation compared with control groups in a time- and dose-dependent manner. Conclusion Our results demonstrated that celastrol could inhibit isolated AS fibroblast proliferation and in vitro osteogenic differentiation. The interaction of PI3K/AKT signaling and Wnt protein may be involved in the process. Further studies should be performed in vivo and animal models to identify the potential effect of celastrol on the bone metabolism of AS patients. PMID:27022241

  9. Dopamine inhibits ATP-induced responses in the cat petrosal ganglion in vitro.

    PubMed

    Alcayaga, Julio; Retamal, Mauricio; Cerpa, Verónica; Arroyo, Jorge; Zapata, Patricio

    2003-03-21

    The petrosal ganglion (PG) provides sensory innervation to the carotid sinus and carotid body through the carotid (sinus) nerve (CN). Application of either acetylcholine (ACh) or adenosine 5'-triphosphate (ATP) to the PG superfused in vitro activates CN fibers. Dopamine (DA) modulates the effects of ACh. We have previously shown that DA when applied to the PG modulates the effects of ACh on carotid sinus nerve fibers. We currently report the effects of DA on the ATP-induced responses in the isolated PG in vitro. While DA had no effect on the basal activity recorded from the CN, it reduced ATP-induced responses in a dose-dependent manner, when preceding ATP applications by 30 s. Our results suggest that DA-a transmitter present in a group of PG neurons and in carotid body cells-may act as an inhibitory modulator of ATP-evoked responses in PG neurons.

  10. Comparative Evaluation of Antimicrobial Activity of QMiX, 2.5% Sodium Hypochlorite, 2% Chlorhexidine, Guava Leaf Extract and Aloevera Extract Against Enterococcus faecalis and Candida albicans – An in-vitro Study

    PubMed Central

    Krishnamma, Shoba; Peedikayil, Faizal; Aman, Shibu; Tomy, Nithya; Mariodan, Jithin Pulickal

    2016-01-01

    Introduction Debridement and disinfection of the root canal system is a critical step in endodontic treatment. Most of the irrigants presently used in the endodontic treatment can have an impact on the microbes surviving in the biofilm but none of them are able to do all of the required tasks. Researches are going on its full swing in order to produce an endodontic irrigant having ideal properties. Aim To compare the antimicrobial efficacy of different irrigants like QMiX, guava leaf extract, aloevera extract, 2.5% sodium hypochlorite and 2% chlorhexidine gluconate against Enterococcus faecalis and Candida albicans. Materials and Methods The antimicrobial activity was determined using agar diffusion test. The solutions were divided into five groups: Group I- QMiX, Group II- Guava leaf extract and Group III-Aloevera extract, Group IV–2.5% Sodium hypochlorite and Group V-2% Chlorhexidine. The zones of inhibition of growth were recorded. Results Statistical analysis was performed using one way ANOVA with post-hoc Tukey’s HSD. Values obtained were statistically analyzed (p<0.05). QMiX showed maximum inhibitory effect against Enterococcus faecalis and Candida albicans followed by, 2% chlorhexidine, 2.5% sodium hypochlorite, guava leaf extract and aloevera extract. Results obtained were statistically significant. Conclusion Guava leaf extract showed significant inhibitory effects against Enterococcus faecalis and Candida albicans. QMiX demonstrated the best results among the tested solutions and can be considered as a potential alternative to existing root canal irrigants. PMID:27437354

  11. Dexamethasone diffusion across contact lenses is inhibited by Staphylococcus epidermidis biofilms in vitro

    PubMed Central

    Brothers, Kimberly M.; Nau, Amy C.; Romanowski, Eric G.; Shanks, Robert M. Q.

    2014-01-01

    Purpose This study was designed to measure the impact of bacterial biofilms on diffusion of an ocular therapeutic through silicone hydrogel bandage lenses in vitro. Methods An assay was designed to study the passage of a commonly used steroid dexamethasone through the silicone hydrogel soft contact lenses. Diffused dexamethasone was measured using a spectrophotometer over a period of 18 hours and quantified using a standard curve. This assay was performed with control and Staphylococcus epidermidis biofilm-coated contact lenses composed of lotrafilcon A and methafilcon. Biofilms were formed in brain heart infusion broth supplemented with D-glucose. Results The presented data validate a simple in vitro model that can be used to measure penetration of a topical therapeutic through silicone hydrogel soft contact lenses. Using this model we measured a reduction of dexamethasone diffusion by up to 88% through S. epidermidis biofilm-coated silicon hydrogel lenses compared to control lenses. Conclusions The results of this in vitro study demonstrate that bacterial biofilms impede dexamethasone diffusion through silicon hydrogel contact lenses, and warrant future studies regarding the clinical benefit of using ocular therapeutics in the setting of bandage contact lens use for corneal epithelial defects. PMID:25090165

  12. Multitask Imidazolium Salt Additives for Innovative Poly(l-lactide) Biomaterials: Morphology Control, Candida spp. Biofilm Inhibition, Human Mesenchymal Stem Cell Biocompatibility, and Skin Tolerance.

    PubMed

    Schrekker, Clarissa M L; Sokolovicz, Yuri C A; Raucci, Maria G; Selukar, Balaji S; Klitzke, Joice S; Lopes, William; Leal, Claudio A M; de Souza, Igor O P; Galland, Griselda B; Dos Santos, João Henrique Z; Mauler, Raquel S; Kol, Moshe; Dagorne, Samuel; Ambrosio, Luigi; Teixeira, Mário L; Morais, Jonder; Landers, Richard; Fuentefria, Alexandre M; Schrekker, Henri S

    2016-08-24

    Candida species have great ability to colonize and form biofilms on medical devices, causing infections in human hosts. In this study, poly(l-lactide) films with different imidazolium salt (1-n-hexadecyl-3-methylimidazolium chloride (C16MImCl) and 1-n-hexadecyl-3-methylimidazolium methanesulfonate (C16MImMeS)) contents were prepared, using the solvent casting process. Poly(l-lactide)-imidazolium salt films were obtained with different surface morphologies (spherical and directional), and the presence of the imidazolium salt in the surface was confirmed. These films with different concentrations of the imidazolium salts C16MImCl and C16MImMeS presented antibiofilm activity against isolates of Candida tropicalis, Candida parapsilosis, and Candida albicans. The minor antibiofilm concentration assay enabled one to determine that an increasing imidazolium salt content promoted, in general, an increase in the inhibition percentage of biofilm formation. Scanning electron microscopy micrographs confirmed the effective prevention of biofilm formation on the imidazolium salt containing biomaterials. Lower concentrations of the imidazolium salts showed no cytotoxicity, and the poly(l-lactide)-imidazolium salt films presented good cell adhesion and proliferation percentages with human mesenchymal stem cells. Furthermore, no acute microscopic lesions were identified in the histopathological evaluation after contact between the films and pig ear skin. In combination with the good morphological, physicochemical, and mechanical properties, these poly(l-lactide)-based materials with imidazolium salt additives can be considered as promising biomaterials for use in the manufacturing of medical devices.

  13. Ascorbic acid inhibition of Candida albicans Hsp90-mediated morphogenesis occurs via the transcriptional regulator Upc2.

    PubMed

    Van Hauwenhuyse, Frédérique; Fiori, Alessandro; Van Dijck, Patrick

    2014-10-01

    Morphogenetic transitions of the opportunistic fungal pathogen Candida albicans are influenced by temperature changes, with induction of filamentation upon a shift from 30 to 37°C. Hsp90 was identified as a major repressor of an elongated cell morphology at low temperatures, as treatment with specific inhibitors of Hsp90 results in elongated growth forms at 30°C. Elongated growth resulting from a compromised Hsp90 is considered neither hyphal nor pseudohyphal growth. It has been reported that ascorbic acid (vitamin C) interferes with the yeast-to-hypha transition in C. albicans. In the present study, we show that ascorbic acid also antagonizes the morphogenetic change caused by hampered Hsp90 function. Further analysis revealed that Upc2, a transcriptional regulator of genes involved in ergosterol biosynthesis, and Erg11, the target of azole antifungals, whose expression is in turn regulated by Upc2, are required for this antagonism. Ergosterol levels correlate with elongated growth and are reduced in cells treated with the Hsp90 inhibitor geldanamycin (GdA) and restored by cotreatment with ascorbic acid. In addition, we show that Upc2 appears to be required for ascorbic acid-mediated inhibition of the antifungal activity of fluconazole. These results identify Upc2 as a major regulator of ascorbic acid-induced effects in C. albicans and suggest an association between ergosterol content and elongated growth upon Hsp90 compromise. PMID:25084864

  14. Inhibition of Osteoclastogenesis and Bone Resorption in vitro and in vivo by a prenylflavonoid xanthohumol from hops.

    PubMed

    Li, Jing; Zeng, Li; Xie, Juan; Yue, Zhiying; Deng, Huayun; Ma, Xueyun; Zheng, Chunbing; Wu, Xiushan; Luo, Jian; Liu, Mingyao

    2015-01-01

    Excessive RANKL signaling leads to superfluous osteoclast formation and bone resorption, is widespread in the pathologic bone loss and destruction. Therefore, targeting RANKL or its signaling pathway has been a promising and successful strategy for this osteoclast-related diseases. In this study, we examined the effects of xanthohumol (XN), an abundant prenylflavonoid from hops plant, on osteoclastogenesis, osteoclast resorption, and RANKL-induced signaling pathway using both in vitro and in vivo assay systems. In mouse and human, XN inhibited osteoclast differentiation and osteoclast formation at the early stage. Furthermore, XN inhibited osteoclast actin-ring formation and bone resorption in a dose-dependent manner. In ovariectomized-induced bone loss mouse model and RANKL-injection-induced bone resorption model, we found that administration of XN markedly inhibited bone loss and resorption by suppressing osteoclast activity. At the molecular level, XN disrupted the association of RANK and TRAF6, resulted in the inhibition of NF-κB and Ca(2+)/NFATc1 signaling pathway during osteoclastogenesis. As a results, XN suppressed the expression of osteoclastogenesis-related marker genes, including CtsK, Nfatc1, Trap, Ctr. Therefore, our data demonstrated that XN inhibits osteoclastogenesis and bone resorption through RANK/TRAF6 signaling pathways. XN could be a promising drug candidate in the treatment of osteoclast-related diseases such as postmenopausal osteoporosis. PMID:26620037

  15. Salinomycin inhibits proliferation and induces apoptosis of human nasopharyngeal carcinoma cell in vitro and suppresses tumor growth in vivo

    SciTech Connect

    Wu, Danxin; Zhang, Yu; Huang, Jie; Fan, Zirong; Shi, Fengrong; Wang, Senming

    2014-01-10

    Highlight: •We first evaluated the effect of salinomycin on nasopharyngeal carcinoma (NPC). •Salinomycin could inhibit Wnt/β-catenin signaling and induce apoptosis in NPC. •So salinomycin may be a good potential candidate for the chemotherapy of NPC. -- Abstract: Salinomycin (Sal) is a polyether ionophore antibiotic that has recently been shown to induce cell death in various human cancer cells. However, whether salinomycin plays a functional role in nasopharyngeal carcinoma (NPC) has not been determined to date. The present study investigated the chemotherapeutic efficacy of salinomycin and its molecular mechanisms of action in NPC cells. Salinomycin efficiently inhibited proliferation and invasion of 3 NPC cell lines (CNE-1, CNE-2, and CNE-2/DDP) and activated a extensive apoptotic process that is accompanied by activation of caspase-3 and caspase-9, and decreased mitochondrial membrane potential. Meanwhile, the protein expression level of the Wnt coreceptor lipoprotein receptor related protein 6 (LRP6) and β-catenin was down-regulated, which showed that the Wnt/β-catenin signaling was involved in salinomycin-induced apoptosis of NPC cells. In a nude mouse NPC xenograft model, the anti-tumor effect of salinomycin was associated with the downregulation of β-catenin expression. The present study demonstrated that salinomycin can effectively inhibit proliferation and invasion, and induce apoptosis of NPC cells in vitro and inhibit tumor growth in vivo, probably via the inhibition of Wnt/β-catenin signaling, suggesting salinomycin as a potential candidate for the chemotherapy of NPC.

  16. Inhibition of Osteoclastogenesis and Bone Resorption in vitro and in vivo by a prenylflavonoid xanthohumol from hops

    PubMed Central

    Li, Jing; Zeng, Li; Xie, Juan; Yue, Zhiying; Deng, Huayun; Ma, Xueyun; Zheng, Chunbing; Wu, Xiushan; Luo, Jian; Liu, Mingyao

    2015-01-01

    Excessive RANKL signaling leads to superfluous osteoclast formation and bone resorption, is widespread in the pathologic bone loss and destruction. Therefore, targeting RANKL or its signaling pathway has been a promising and successful strategy for this osteoclast-related diseases. In this study, we examined the effects of xanthohumol (XN), an abundant prenylflavonoid from hops plant, on osteoclastogenesis, osteoclast resorption, and RANKL-induced signaling pathway using both in vitro and in vivo assay systems. In mouse and human, XN inhibited osteoclast differentiation and osteoclast formation at the early stage. Furthermore, XN inhibited osteoclast actin-ring formation and bone resorption in a dose-dependent manner. In ovariectomized-induced bone loss mouse model and RANKL-injection-induced bone resorption model, we found that administration of XN markedly inhibited bone loss and resorption by suppressing osteoclast activity. At the molecular level, XN disrupted the association of RANK and TRAF6, resulted in the inhibition of NF-κB and Ca2+/NFATc1 signaling pathway during osteoclastogenesis. As a results, XN suppressed the expression of osteoclastogenesis-related marker genes, including CtsK, Nfatc1, Trap, Ctr. Therefore, our data demonstrated that XN inhibits osteoclastogenesis and bone resorption through RANK/TRAF6 signaling pathways. XN could be a promising drug candidate in the treatment of osteoclast-related diseases such as postmenopausal osteoporosis. PMID:26620037

  17. PCCG-IV inhibits the induction of long-term potentiation in the dentate gyrus in vitro.

    PubMed

    Huang, L; Breakwell, N A; Rowan, M J; Anwyl, R

    1997-08-01

    The effects of two ligands with previously established high and selective potency for metabotropic glutamate receptors (mGlu receptors) group II have been investigated on the high frequency stimulation (HFS) induced long-term potentiation of the field excitatory postsynaptic potential (EPSP) in the dentate gyrus of the rat hippocampus in vitro. The ligands investigated were (2S,1'S,2'S,3'R)-2-(2"-carboxy-3'-phenylcyclopropyl)glycine (PCCG-IV) and (R,S)-alpha-methyl-4-tetrazolylphenylglycine (MTPG). PCCG-IV (10 microM) strongly inhibited the induction of long-term potentiation of the field EPSP by high frequency stimulation. MTPG (50 microM) did not inhibit the induction of long-term potentiation, but prevented the inhibition of long-term potentiation induction by PCCG-IV. The inhibition of long-term potentiation induction by PCCG-IV is suggested to be due to an agonistic action on mGlu receptor group II, probably mGlu3 receptor, as the inhibition of long-term potentiation can be reversed by the application of MTPG, a well-known selective and potent antagonist of mGlu receptor group II. PMID:9286617

  18. Isoliquiritigenin Attenuates Adipose Tissue Inflammation in vitro and Adipose Tissue Fibrosis through Inhibition of Innate Immune Responses in Mice

    PubMed Central

    Watanabe, Yasuharu; Nagai, Yoshinori; Honda, Hiroe; Okamoto, Naoki; Yamamoto, Seiji; Hamashima, Takeru; Ishii, Yoko; Tanaka, Miyako; Suganami, Takayoshi; Sasahara, Masakiyo; Miyake, Kensuke; Takatsu, Kiyoshi

    2016-01-01

    Isoliquiritigenin (ILG) is a flavonoid derived from Glycyrrhiza uralensis and potently suppresses NLRP3 inflammasome activation resulting in the improvement of diet-induced adipose tissue inflammation. However, whether ILG affects other pathways besides the inflammasome in adipose tissue inflammation is unknown. We here show that ILG suppresses adipose tissue inflammation by affecting the paracrine loop containing saturated fatty acids and TNF-α by using a co-culture composed of adipocytes and macrophages. ILG suppressed inflammatory changes induced by the co-culture through inhibition of NF-κB activation. This effect was independent of either inhibition of inflammasome activation or activation of peroxisome proliferator-activated receptor-γ. Moreover, ILG suppressed TNF-α-induced activation of adipocytes, coincident with inhibition of IκBα phosphorylation. Additionally, TNF-α-mediated inhibition of Akt phosphorylation under insulin signaling was alleviated by ILG in adipocytes. ILG suppressed palmitic acid-induced activation of macrophages, with decreasing the level of phosphorylated Jnk expression. Intriguingly, ILG improved high fat diet-induced fibrosis in adipose tissue in vivo. Finally, ILG inhibited TLR4- or Mincle-stimulated expression of fibrosis-related genes in stromal vascular fraction from obese adipose tissue and macrophages in vitro. Thus, ILG can suppress adipose tissue inflammation by both inflammasome-dependent and -independent manners and attenuate adipose tissue fibrosis by targeting innate immune sensors. PMID:26975571

  19. Anti-neuroblastoma cell line antibodies in inflammatory demyelinating polyneuropathy: inhibition in vitro and in vivo by IV immunoglobulin.

    PubMed

    van Doorn, P A; Brand, A; Vermeulen, M

    1988-10-01

    We tested serum from 48 patients with Guillain-Barré syndrome and 42 with chronic inflammatory demyelinating polyneuropathy (CIDP) against a selected neuroblastoma cell line (NBL 108cc15). Forty-two percent of the patients showed a positive immunofluorescence test against the NBL 108cc15. These antibodies were mainly of the IgM-class; they disappeared in all seven CIDP patients retested after improvement following intravenous IgG treatment (IV-IgG) and were present in only 5% of serum from patients with other disorders. Absorption studies showed a partial homology between the NBL 108cc15 and human sciatic nerve. In vitro studies showed that IgG from pooled normal donors (IV-IgG) inhibits the reaction between serum from a CIDP patient and the NBL cell line. This inhibition may be due to neutralization of autoantibodies against nervous tissue by anti-idiotypic antibodies in IV-IgG.

  20. In vitro inhibition effect and structure-activity relationships of some saccharin derivatives on erythrocyte carbonic anhydrase I and II.

    PubMed

    Sonmez, Fatih; Bilen, Cigdem; Sumersan, Sinem; Gencer, Nahit; Isik, Semra; Arslan, Oktay; Kucukislamoglu, Mustafa

    2014-02-01

    In this study, in vitro inhibitory effects of some saccharin derivatives on purified carbonic anhydrase I and II were investigated using CO2 as a substrate. The results showed that all compounds inhibited the hCA I and hCA II enzyme activities. Among the compounds, 6-(p-tolylthiourenyl) saccharin (6m) was found to be the most active one for hCA I activity (IC50=13.67 μM) and 6-(m-methoxyphenylurenyl) saccharin (6b) was found to be the most active one for hCA II activity (IC50=6.54 μM). Structure-activity relationships (SARs) study showed that, generally, thiourea derivatives (6l--v) inhibited more hCA I and hCA II than urea derivatives (6a-k). All compounds (excluding 6c and 6r) have higher inhibitory activity on hCA II than on hCA I.

  1. Monohaloacetic acid drinking water disinfection by-products inhibit follicle growth and steroidogenesis in mouse ovarian antral follicles in vitro.

    PubMed

    Jeong, Clara H; Gao, Liying; Dettro, Tyler; Wagner, Elizabeth D; Ricke, William A; Plewa, Michael J; Flaws, Jodi A

    2016-07-01

    Water disinfection greatly reduced the incidence of waterborne diseases, but the reaction between disinfectants and natural organic matter in water leads to the formation of drinking water disinfection by-products (DBPs). DBPs have been shown to be toxic, but their effects on the ovary are not well defined. This study tested the hypothesis that monohalogenated DBPs (chloroacetic acid, CAA; bromoacetic acid, BAA; iodoacetic acid, IAA) inhibit antral follicle growth and steroidogenesis in mouse ovarian follicles. Antral follicles were isolated and cultured with either vehicle or DBPs (0.25-1.00mM of CAA; 2-15μM of BAA or IAA) for 48 and 96h. Follicle growth was measured every 24h and the media were analyzed for estradiol levels at 96h. Exposure to DBPs significantly inhibited antral follicle growth and reduced estradiol levels compared to controls. These data demonstrate that DBP exposure caused ovarian toxicity in vitro. PMID:27151372

  2. Effect of Usnic Acid on Candida orthopsilosis and C. parapsilosis

    PubMed Central

    Lucarini, Rodrigo; Mendes-Giannini, Maria Jose Soares

    2012-01-01

    The activity of usnic acid against Candida orthopsilosis and Candida parapsilosis on planktonic and biofilm conditions was investigated by using a broth microdilution and microplate methods. Potent in vitro activities against different Candida species were obtained. The metabolic activity of sessile cells of C. parapsilosis complex was reduced by 80% at four times the 80% inhibitory concentration. The in vitro studies support further efforts to determine whether usnic acid can be used clinically to cure patients with Candida infections. PMID:22006006

  3. Inhibition of gizzard shade (Dorosoma cepedianum) P4501A activity by in vivo and in vitro administration of clotrimazole

    SciTech Connect

    Levine, S.; Czosnyka, H.; Oris, J.

    1995-12-31

    The ability of the 1-substituted imidazole agent clotrimazole to inhibit in vivo and in vitro hepatic microsomal ethoxyresorufin O-deethylase (EROD) activity was investigated using gizzard shad (Dorosoma cepedianum) pre-treated with clotrimazole and then given waterborne exposures of benzo[a]pyrene (BaP). Gizzard shad pre-treated with 50 mg clotrimazole/kg and then exposed for 1 or 3 d to BaP (0.86 {micro}g/L) had significantly lower hepatic EROD activity compared to fish that were exposed to BaP alone. Following 1 and 3 d of BaP exposure, groups pre-treated with clotrimazole had 7% and 28% of the EROD activity respectively relative to groups exposed to BaP alone. In addition, there was 8x and 11x more parent BaP in whole-fish extracts from gizzard shad pre-treated with clotrimazole after 1 and 3 d of BaP exposure, respectively, compared to fish exposed to BaP alone. Clotrimazole produced a ``type 11`` binding spectral complex with BaP-treated microsomes and was an effective in vitro inhibitor of EROD activity in micromolar concentrations. The IC{sub 50} for EROD activity was 0.57 {micro}M clotrimazole. The mechanism of inhibition by clotrimazole was determined to be noncompetitive (mixed type) using kinetics experiments. Noncompetitive inhibition of monooxygenase activity is believed to arise from the interaction created via the non-bonded electrons of the nitrogen in the imidazole ring with the name moiety of cytochrome P450, resulting in inhibition of enzymatic activity. The high inhibitory potency of clotrimazole on P4501A catalyzed reactions may prove valuable for its use as a probe in determining whether chemical toxicity is manifested through a cytochrome P450-mediated pathway.

  4. Clofazimine Inhibits the Growth of Babesia and Theileria Parasites In Vitro and In Vivo.

    PubMed

    Tuvshintulga, Bumduuren; AbouLaila, Mahmoud; Davaasuren, Batdorj; Ishiyama, Aki; Sivakumar, Thillaiampalam; Yokoyama, Naoaki; Iwatsuki, Masato; Otoguro, Kazuhiko; Ōmura, Satoshi; Igarashi, Ikuo

    2016-05-01

    The present study evaluated the growth-inhibitory effects of clofazimine, currently used for treating leprosy, against Babesia bovis, B. bigemina, B. caballi, and Theileria equi in in vitro culture and against Babesia microti in mice. The 50% inhibitory concentrations (IC50s) of clofazimine against the in vitro growth of B. bovis, B. bigemina, B. caballi, and T. equi were 4.5, 3, 4.3, and 0.29 μM, respectively. In mice infected with B. microti, treatment with 20 mg/kg of body weight of clofazimine administered orally resulted in a significantly lower peak parasitemia (5.3%) than that in the control group (45.9%), which was comparable to the subcutaneous administration of 25 mg/kg diminazene aceturate, the most widely used treatment for animal piroplasmosis. Although slight anemia was observed in both clofazimine- and diminazene aceturate-treated infected mice, the level and duration of anemia were lower and shorter, respectively, than those in untreated infected mice. Using blood transfusions and PCR, we also examined whether clofazimine completely killed B. microti On day 40 postinfection, when blood analysis was performed, parasites were not found in blood smears; however, the DNA of B. microti was detected in the blood of clofazimine-treated animals and in several tissues of clofazimine- and diminazene aceturate-treated mice by PCR. The growth of parasites was observed in mice after blood transfusions from clofazimine-treated mice. In conclusion, clofazimine showed excellent inhibitory effects against Babesia and Theileria in vitro and in vivo, and further study on clofazimine is required for the future development of a novel chemotherapy with high efficacy and safety against animal piroplasmosis and, possibly, human babesiosis. PMID:26883713

  5. Clofazimine Inhibits the Growth of Babesia and Theileria Parasites In Vitro and In Vivo

    PubMed Central

    Tuvshintulga, Bumduuren; AbouLaila, Mahmoud; Davaasuren, Batdorj; Ishiyama, Aki; Sivakumar, Thillaiampalam; Yokoyama, Naoaki; Iwatsuki, Masato; Otoguro, Kazuhiko; Ōmura, Satoshi

    2016-01-01

    The present study evaluated the growth-inhibitory effects of clofazimine, currently used for treating leprosy, against Babesia bovis, B. bigemina, B. caballi, and Theileria equi in in vitro culture and against Babesia microti in mice. The 50% inhibitory concentrations (IC50s) of clofazimine against the in vitro growth of B. bovis, B. bigemina, B. caballi, and T. equi were 4.5, 3, 4.3, and 0.29 μM, respectively. In mice infected with B. microti, treatment with 20 mg/kg of body weight of clofazimine administered orally resulted in a significantly lower peak parasitemia (5.3%) than that in the control group (45.9%), which was comparable to the subcutaneous administration of 25 mg/kg diminazene aceturate, the most widely used treatment for animal piroplasmosis. Although slight anemia was observed in both clofazimine- and diminazene aceturate-treated infected mice, the level and duration of anemia were lower and shorter, respectively, than those in untreated infected mice. Using blood transfusions and PCR, we also examined whether clofazimine completely killed B. microti. On day 40 postinfection, when blood analysis was performed, parasites were not found in blood smears; however, the DNA of B. microti was detected in the blood of clofazimine-treated animals and in several tissues of clofazimine- and diminazene aceturate-treated mice by PCR. The growth of parasites was observed in mice after blood transfusions from clofazimine-treated mice. In conclusion, clofazimine showed excellent inhibitory effects against Babesia and Theileria in vitro and in vivo, and further study on clofazimine is required for the future development of a novel chemotherapy with high efficacy and safety against animal piroplasmosis and, possibly, human babesiosis. PMID:26883713

  6. Change of nitric oxide in experimental colitis and its inhibition by melatonin in vivo and in vitro

    PubMed Central

    Mei, Q; Xu, J; Xiang, L; Hu, Y; Hu, X; Xu, Z

    2005-01-01

    Aim: To investigate the change of nitric oxide (NO) in rat colitis and its inhibition by melatonin in vivo and in vitro. Methods: In vivo, rat colitis was established intracolonically with trinitrobenzenesulphonic acid (TNBS) and ethanol. The animals were randomised into five groups: control group, model group, melatonin group (2.5, 5.0, 10.0 mg/kg), and treated intracolonically with saline, saline and melatonin respectively (once a day, from day 7 after colitis was established to day 28). After the end of the experiment, the mucosal damage index (CMDI) and histology score (HS) were evaluated and the level of myeloperoxidase (MPO) and malondiadehyde (MDA) and NO in the colon tissue were measured. In vitro, the co-culture model of the inflamed colon mucosa (from the colitis) with lipopolysaccharide (LPS), and the colonocytes oxidative injury model by hydroxyl radical, were designed respectively to elucidate the inhibition of NO by melatonin. Results: After treated with TNBS/ethanol, the extent of CMDI and HS, the levels of MPO, MDA, and NO in the model group, were higher than that in the control group; melatonin ameliorated these parameters effectively. The stimulation of LPS increased the level of NO and MPO and MDA in the co-culture model of inflamed colon mucosa, and melatonin significantly reduced the level of MPO, MDA, and NO. In the coloncyte oxidative injury model by hydroxyl radical, the contents of LDH, MDA, and NO were increased; melatonin reversed this oxidative injury considerably. Conclusion: This study showed that TNBS/ethanol induced colitis was pharmacologically controlled by melatonin in vivo and in vitro. PMID:16210467

  7. High-power helium-neon laser irradiation inhibits the growth of traumatic scars in vitro and in vivo.

    PubMed

    Shu, Bin; Ni, Guo-Xin; Zhang, Lian-Yang; Li, Xiang-Ping; Jiang, Wan-Ling; Zhang, Li-Qun

    2013-05-01

    This study explored the inhibitory effect of the high-power helium-neon (He-Ne) laser on the growth of scars post trauma. For the in vitro study, human wound fibroblasts were exposed to the high-power He-Ne laser for 30 min, once per day with different power densities (10, 50, 100, and 150 mW/cm(2)). After 3 days of repeated irradiation with the He-Ne laser, fibroblast proliferation and collagen synthesis were evaluated. For in vivo evaluation, a wounded animal model of hypertrophic scar formation was established. At postoperative day 21, the high-power He-Ne laser irradiation (output power 120 mW, 6 mm in diameter, 30 min each session, every other day) was performed on 20 scars. At postoperative day 35, the hydroxyproline content, apoptosis rate, PCNA protein expression and FADD mRNA level were assessed. The in vitro study showed that the irradiation group that received the power densities of 100 and 150 mW/cm(2) showed decreases in the cell proliferation index, increases in the percentage of cells in the G0/G1 phase, and decreases in collagen synthesis and type I procollagen gene expression. In the in vivo animal studies, regions exposed to He-Ne irradiation showed a significant decrease in scar thickness as well as decreases in hydroxyproline levels and PCNA protein expression. Results from the in vitro and in vivo studies suggest that repeated irradiation with a He-Ne laser at certain power densities inhibits fibroblast proliferation and collagen synthesis, thereby inhibits the growth of hypertrophic scars.

  8. High-power helium-neon laser irradiation inhibits the growth of traumatic scars in vitro and in vivo.

    PubMed

    Shu, Bin; Ni, Guo-Xin; Zhang, Lian-Yang; Li, Xiang-Ping; Jiang, Wan-Ling; Zhang, Li-Qun

    2013-05-01

    This study explored the inhibitory effect of the high-power helium-neon (He-Ne) laser on the growth of scars post trauma. For the in vitro study, human wound fibroblasts were exposed to the high-power He-Ne laser for 30 min, once per day with different power densities (10, 50, 100, and 150 mW/cm(2)). After 3 days of repeated irradiation with the He-Ne laser, fibroblast proliferation and collagen synthesis were evaluated. For in vivo evaluation, a wounded animal model of hypertrophic scar formation was established. At postoperative day 21, the high-power He-Ne laser irradiation (output power 120 mW, 6 mm in diameter, 30 min each session, every other day) was performed on 20 scars. At postoperative day 35, the hydroxyproline content, apoptosis rate, PCNA protein expression and FADD mRNA level were assessed. The in vitro study showed that the irradiation group that received the power densities of 100 and 150 mW/cm(2) showed decreases in the cell proliferation index, increases in the percentage of cells in the G0/G1 phase, and decreases in collagen synthesis and type I procollagen gene expression. In the in vivo animal studies, regions exposed to He-Ne irradiation showed a significant decrease in scar thickness as well as decreases in hydroxyproline levels and PCNA protein expression. Results from the in vitro and in vivo studies suggest that repeated irradiation with a He-Ne laser at certain power densities inhibits fibroblast proliferation and collagen synthesis, thereby inhibits the growth of hypertrophic scars. PMID:22678421

  9. R-thanatin inhibits growth and biofilm formation of methicillin-resistant Staphylococcus epidermidis in vivo and in vitro.

    PubMed

    Hou, Zheng; Da, Fei; Liu, Baohui; Xue, Xiaoyan; Xu, Xiuli; Zhou, Ying; Li, Mingkai; Li, Zhi; Ma, Xue; Meng, Jingru; Jia, Min; Wang, Yukun; Luo, Xiaoxing

    2013-10-01

    Staphylococcus epidermidis is one of the most frequent causes of device-associated infections, because it is known to cause biofilms that grow on catheters or other surgical implants. The persistent increasing resistance of S. epidermidis and other coagulase-negative staphylococci (CoNS) has driven the need for newer antibacterial agents with innovative therapeutic strategies. Thanatin is reported to display potent antibiotic activities, especially against extended-spectrum-beta-lactamase-producing Escherichia coli. The present study aimed to investigate whether a shorter derivative peptide (R-thanatin) could be used as a novel antibacterial agent. We found that R-thanatin was highly potent in vitro against coagulase-negative staphylococci, such as S. epidermidis, S. haemolyticus, and S. hominis, and inhibited biofilm formation at subinhibitory concentrations. Properties of little toxicity to human red blood cells (hRBCs) and human umbilical vein endothelial cells, a low incidence of resistance, and relatively high stability in plasma were confirmed. Excellent in vivo protective effects were also observed using a methicillin-resistant S. epidermidis (MRSE)-induced urinary tract infection rat model. Electron microscopy and confocal laser-scanning microscopy analyses suggested that R-thanatin disturbed cell division of MRSE severely, which might be the reason for inhibition of MRSE growth. These findings indicate that R-thanatin is active against the growth and biofilm formation of MRSE in vitro and in vivo. R-thanatin might be considered as a specific drug candidate for treating CoNS infections.

  10. In vitro inhibition of proliferation of vascular smooth muscle cells by serum of rats treated with Dahuang Zhechong pill.

    PubMed

    Zhang, Yuan-Hui; Liu, Jun-Tian; Wen, Bin-Yu; Xiao, Xiang-Hua

    2007-06-13

    Dahuang Zhechong pill (DHZCP) is a famous and classical Chinese herbal prescription, which is clinically used to treat hepatic, gynecological and cardiovascular diseases in China. The aim of this study was to observe the effects of the serum of rats treated with DHZCP on the proliferation of cultured rat vascular smooth muscle cells (VSMCs) stimulated by platelet-derived growth factor (PDGF), oxidized low density lipoprotein (ox-LDL) and hyperlipidemic serum (HLS), and on DNA, protein and collagen syntheses of VSMCs induced by PDGF in vitro. VSMCs proliferation was assayed by measuring the cell viability with MTT method, and syntheses of DNA, protein and collagen were evaluated by detecting [(3)H]-thymidine, [(3)H]-leucine and [(3)H]-proline incorporations, respectively. The results showed that PDGF, ox-LDL and HLS stimulated the proliferation of rat VSMCs in vitro. The serum of rats treated with DHZCP significantly inhibited the proliferation of rat VSMCs induced by the above stimulants and the incorporations of [(3)H]-thymidine, [(3)H]-leucine and [(3)H]-proline into rat VSMCs induced by PDGF in comparison with the model control group (P<0.01). The data suggest that DHZCP is able to obviously inhibit VSMCs proliferation via interfering with syntheses of DNA and protein, and to decrease production of extracellular matrix by VSMCs through antagonizing collagen synthesis.

  11. Broad HIV-1 inhibition in vitro by vaccine-elicited CD8+ T cells in African adults

    PubMed Central

    Mutua, Gaudensia; Farah, Bashir; Langat, Robert; Indangasi, Jackton; Ogola, Simon; Onsembe, Brian; Kopycinski, Jakub T; Hayes, Peter; Borthwick, Nicola J; Ashraf, Ambreen; Dally, Len; Barin, Burc; Tillander, Annika; Gilmour, Jill; De Bont, Jan; Crook, Alison; Hannaman, Drew; Cox, Josephine H; Anzala, Omu; Fast, Patricia E; Reilly, Marie; Chinyenze, Kundai; Jaoko, Walter; Hanke, Tomáš; HIV-CORE 004 study group, the

    2016-01-01

    We are developing a pan-clade HIV-1 T-cell vaccine HIVconsv, which could complement Env vaccines for prophylaxis and be a key to HIV cure. Our strategy focuses vaccine-elicited effector T-cells on functionally and structurally conserved regions (not full-length proteins and not only epitopes) of the HIV-1 proteome, which are common to most global variants and which, if mutated, cause a replicative fitness loss. Our first clinical trial in low risk HIV-1-negative adults in Oxford demonstrated the principle that naturally mostly subdominant epitopes, when taken out of the context of full-length proteins/virus and delivered by potent regimens involving combinations of simian adenovirus and poxvirus modified vaccinia virus Ankara, can induce robust CD8+ T cells of broad specificities and functions capable of inhibiting in vitro HIV-1 replication. Here and for the first time, we tested this strategy in low risk HIV-1-negative adults in Africa. We showed that the vaccines were well tolerated and induced high frequencies of broadly HIVconsv-specific plurifunctional T cells, which inhibited in vitro viruses from four major clades A, B, C, and D. Because sub-Saharan Africa is globally the region most affected by HIV-1/AIDS, trial HIV-CORE 004 represents an important stage in the path toward efficacy evaluation of this highly rational and promising vaccine strategy.

  12. Tox-Database.net: a curated resource for data describing chemical triggered in vitro cardiac ion channels inhibition

    PubMed Central

    2012-01-01

    Background Drugs safety issues are now recognized as being factors generating the most reasons for drug withdrawals at various levels of development and at the post-approval stage. Among them cardiotoxicity remains the main reason, despite the substantial effort put into in vitro and in vivo testing, with the main focus put on hERG channel inhibition as the hypothesized surrogate of drug proarrhythmic potency. The large interest in the IKr current has resulted in the development of predictive tools and informative databases describing a drug's susceptibility to interactions with the hERG channel, although there are no similar, publicly available sets of data describing other ionic currents driven by the human cardiomyocyte ionic channels, which are recognized as an overlooked drug safety target. Discussion The aim of this database development and publication was to provide a scientifically useful, easily usable and clearly verifiable set of information describing not only IKr (hERG), but also other human cardiomyocyte specific ionic channels inhibition data (IKs, INa, ICa). Summary The broad range of data (chemical space and in vitro settings) and the easy to use user interface makes tox-database.net a useful tool for interested scientists. Database URL http://tox-database.net. PMID:22947121

  13. Broad HIV-1 inhibition in vitro by vaccine-elicited CD8+ T cells in African adults

    PubMed Central

    Mutua, Gaudensia; Farah, Bashir; Langat, Robert; Indangasi, Jackton; Ogola, Simon; Onsembe, Brian; Kopycinski, Jakub T; Hayes, Peter; Borthwick, Nicola J; Ashraf, Ambreen; Dally, Len; Barin, Burc; Tillander, Annika; Gilmour, Jill; De Bont, Jan; Crook, Alison; Hannaman, Drew; Cox, Josephine H; Anzala, Omu; Fast, Patricia E; Reilly, Marie; Chinyenze, Kundai; Jaoko, Walter; Hanke, Tomáš; HIV-CORE 004 study group, the

    2016-01-01

    We are developing a pan-clade HIV-1 T-cell vaccine HIVconsv, which could complement Env vaccines for prophylaxis and be a key to HIV cure. Our strategy focuses vaccine-elicited effector T-cells on functionally and structurally conserved regions (not full-length proteins and not only epitopes) of the HIV-1 proteome, which are common to most global variants and which, if mutated, cause a replicative fitness loss. Our first clinical trial in low risk HIV-1-negative adults in Oxford demonstrated the principle that naturally mostly subdominant epitopes, when taken out of the context of full-length proteins/virus and delivered by potent regimens involving combinations of simian adenovirus and poxvirus modified vaccinia virus Ankara, can induce robust CD8+ T cells of broad specificities and functions capable of inhibiting in vitro HIV-1 replication. Here and for the first time, we tested this strategy in low risk HIV-1-negative adults in Africa. We showed that the vaccines were well tolerated and induced high frequencies of broadly HIVconsv-specific plurifunctional T cells, which inhibited in vitro viruses from four major clades A, B, C, and D. Because sub-Saharan Africa is globally the region most affected by HIV-1/AIDS, trial HIV-CORE 004 represents an important stage in the path toward efficacy evaluation of this highly rational and promising vaccine strategy. PMID:27617268

  14. LTP-1, a novel antimitotic agent and Stat3 inhibitor, inhibits human pancreatic carcinomas in vitro and in vivo

    PubMed Central

    Huang, Han-Li; Chao, Min-Wu; Chen, Chung-Chun; Cheng, Chun-Chun; Chen, Mei-Chuan; Lin, Chao-Feng; Liou, Jing-Ping; Teng, Che-Ming; Pan, Shiow-Lin

    2016-01-01

    Pancreatic cancer is the leading cause of cancer death worldwide with a poor survival rate. The objective of this study was to determine the mechanism of action of a novel antimitotic and Stat3 inhibitor, LTP-1, on human pancreatic cancer in vitro and in vivo. We found that LTP-1 inhibited pancreatic cancer cell growth and viability with significant G2/M arrest and disruption of microtubule dynamics. LTP-1 also caused G2/M arrest-independent Stat3 dephosphorylation along with ERK activation, which indicated the possible dual function of LTP-1. Long-term treatment of LTP-1 also induced polyploidy, activated caspases, induced subG1 cell population, and therefore, triggered pancreatic cancer cell apoptosis. Finally, we used an in vivo xenograft model to demonstrate that LTP-1 suppressed the growth of pancreatic adenocarcinoma. In summary, our data suggest that LTP-1 may alter microtubule dynamics, which ultimately causes polyploidy and apoptosis, thereby inhibiting pancreatic cancer growth in vitro and in vivo. This study provides evidence that LTP-1 could be a potential therapeutic agent for further development of pancreatic cancer treatment. PMID:27278358

  15. Broad HIV-1 inhibition in vitro by vaccine-elicited CD8(+) T cells in African adults.

    PubMed

    Mutua, Gaudensia; Farah, Bashir; Langat, Robert; Indangasi, Jackton; Ogola, Simon; Onsembe, Brian; Kopycinski, Jakub T; Hayes, Peter; Borthwick, Nicola J; Ashraf, Ambreen; Dally, Len; Barin, Burc; Tillander, Annika; Gilmour, Jill; De Bont, Jan; Crook, Alison; Hannaman, Drew; Cox, Josephine H; Anzala, Omu; Fast, Patricia E; Reilly, Marie; Chinyenze, Kundai; Jaoko, Walter; Hanke, Tomáš; Hiv-Core 004 Study Group, The

    2016-01-01

    We are developing a pan-clade HIV-1 T-cell vaccine HIVconsv, which could complement Env vaccines for prophylaxis and be a key to HIV cure. Our strategy focuses vaccine-elicited effector T-cells on functionally and structurally conserved regions (not full-length proteins and not only epitopes) of the HIV-1 proteome, which are common to most global variants and which, if mutated, cause a replicative fitness loss. Our first clinical trial in low risk HIV-1-negative adults in Oxford demonstrated the principle that naturally mostly subdominant epitopes, when taken out of the context of full-length proteins/virus and delivered by potent regimens involving combinations of simian adenovirus and poxvirus modified vaccinia virus Ankara, can induce robust CD8(+) T cells of broad specificities and functions capable of inhibiting in vitro HIV-1 replication. Here and for the first time, we tested this strategy in low risk HIV-1-negative adults in Africa. We showed that the vaccines were well tolerated and induced high frequencies of broadly HIVconsv-specific plurifunctional T cells, which inhibited in vitro viruses from four major clades A, B, C, and D. Because sub-Saharan Africa is globally the region most affected by HIV-1/AIDS, trial HIV-CORE 004 represents an important stage in the path toward efficacy evaluation of this highly rational and promising vaccine strategy. PMID:27617268

  16. Red and infrared laser therapy inhibits in vitro growth of major bacterial species that commonly colonize skin ulcers.

    PubMed

    de Sousa, Natanael Teixeira Alves; Gomes, Rosana Caetano; Santos, Marcos Ferracioli; Brandino, Hugo Evangelista; Martinez, Roberto; de Jesus Guirro, Rinaldo Roberto

    2016-04-01

    Low-level laser therapy (LLLT) is used in chronic wounds due to its healing effects. However, bacterial species may colonize these wounds and the optimal parameters for effective bacterial inhibition are not clear. The aim of this study was to analyze the effect of LLLT on bacterial growth in vitro. Bacterial strains including Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa were suspended in saline solution at a concentration of 10(3) cells/ml and exposed to laser irradiation at wavelengths of 660, 830, and 904 nm at fluences of 0 (control), 3, 6, 12, 18, and 24 J/cm(2). An aliquot of the irradiated suspension was spread on the surface of petri plates and incubated at 37 °C for quantification of colony-forming unit after 24, 48, and 72 h. Laser irradiation inhibited the growth of S. aureus at all wavelengths and fluences higher than 12 J/cm(2), showing a strong correlation between increase in fluence and bacterial inhibition. However, for P. aeruginosa, LLLT inhibited growth at all wavelengths only at a fluence of 24 J/cm(2). E. coli had similar growth inhibition at a wavelength of 830 nm at fluences of 3, 6, 12, and 24 J/cm(2). At wavelengths of 660 and 904 nm, growth inhibition was only observed at fluences of 12 and 18 J/cm(2), respectively. LLLT inhibited bacterial growth at all wavelengths, for a maximum of 72 h after irradiation, indicating a correlation between bacterial species, fluence, and wavelength.

  17. Evaluation of in vitro urease and lipoxygenase inhibition activity of weight reducing tablets.

    PubMed

    Jaffary, Syed Rashid Ali; Ahmed, Syed Waseemuddin; Shakeel, Sadia; Asif, Hafiz Muhammad; Usmanghani, Khan

    2016-07-01

    Enzyme inhibition is a significant part of research in pharmaceutical field in view of the fact that these studies have directed to the innovations of drugs having remarkable performance in diverse physiological conditions. The present study was aimed to assess urease and lipoxygenase inhibitory activity of weight reducing tablets. For evaluating the urease activity indophenol method was employed using Thiourea as the model urease inhibitor. The lipoxygenase inhibition was evaluated by measuring the hydroperoxides produced in lipoxygenation reaction using a purified lipoxygenase with lionoleic acid as substrate. When formulation of the weight reducing tablets was compared at various concentrations (50, 100 and 500µg/ml). The antiurease activity and lipoxygenase inhibition activity increased in a dose dependent manner. The formulations under test have an excellent antiurease and lipoxygenase inhibition potential and prospective to be used in the cure of a variety of complications associated with the production of urease and lipoxygenase enzymes. PMID:27592490

  18. Competitive binding inhibition enzyme-linked immunosorbent assay that uses the secreted aspartyl proteinase of Candida albicans as an antigenic marker for diagnosis of disseminated candidiasis.

    PubMed

    Morrison, Christine J; Hurst, Steven F; Reiss, Errol

    2003-09-01

    The secreted aspartyl proteinases (Saps) of Candida albicans have been implicated as virulence factors associated with adherence and tissue invasion. The potential use of proteinases as markers of invasive candidiasis led us to develop a competitive binding inhibition enzyme-linked immunosorbent assay (ELISA) to detect Sap in clinical specimens. Daily serum and urine specimens were collected from rabbits that had been immunosuppressed with cyclophosphamide and cortisone acetate and infected intravenously with 10(7) C. albicans blastoconidia. Disseminated infection was confirmed by organ culture and histopathology. Although ELISA inhibition was observed when serum specimens from these rabbits were used, more significant inhibition, which correlated with disease progression, occurred when urine specimens were used. Urine collected as early as 1 day after infection resulted in significant ELISA inhibition (mean inhibition +/- standard error [SE] compared with preinfection control urine, 15.7% +/- 2.7% [P < 0.01]), and inhibition increased on days 2 through 5 (29.4% +/- 4.8% to 44.5% +/- 3.5% [P < 0.001]). Urine specimens from immunosuppressed rabbits infected intravenously with Candida tropicalis, Candida parapsilosis, Candida krusei, Cryptococcus neoformans, Aspergillus fumigatus, or Staphylococcus aureus were negative in the assay despite culture-proven dissemination. Nonimmunosuppressed rabbits receiving oral tetracycline and gentamicin treatment were given 2 x 10(8) C. albicans blastoconidia orally or intraurethrally to establish colonization of the gastrointestinal tract or bladder, respectively, without systemic dissemination; urine specimens from these rabbits also gave negative ELISA results. Dissemination to the kidney and spleen occurred in one rabbit challenged by intragastric inoculation, and urine from this rabbit demonstrated significant inhibition in the ELISA (mean inhibition +/- SE by day 3 after infection, 32.9% +/- 2.7% [P < 0.001]). The overall

  19. Beta-lactam antibiotic-induced platelet dysfunction: Evidence for irreversible inhibition of platelet activation in vitro and in vivo after prolonged exposure to penicillin

    SciTech Connect

    Burroughs, S.F.; Johnson, G.J. )

    1990-04-01

    beta-Lactam antibiotics cause platelet dysfunction with bleeding complications. Previous in vitro studies documented reversible inhibition of agonist-receptor interaction. This mechanism is inadequate to explain the effect of beta-lactam antibiotics in vivo. Platelet function does not return to normal immediately after drug treatment, implying irreversible inhibition of platelet function. We report here evidence of irreversible platelet functional and biochemical abnormalities after in vitro and in vivo exposure to beta-lactam antibiotics. Irreversible binding of (14C)-penicillin (Pen) occurred in vitro. After 24 hours' in vitro incubation with 10 to 20 mmol/L Pen, or ex vivo after antibiotic treatment, irreversible functional impairment occurred; but no irreversible inhibition of alpha 2 adrenergic receptors, measured with (3H)-yohimbine, or high-affinity thromboxane A2/prostaglandin H2 (TXA2/PGH2) receptors, measured with agonist (3H)-U46619 and antagonist (3H)-SQ29548, occurred. However, low-affinity platelet TXA2/PGH2 receptors were decreased 40% after Pen exposure in vitro or in vivo, indicating irreversible membrane alteration. Two postreceptor biochemical events were irreversibly inhibited in platelets incubated with Pen for 24 hours in vitro or ex vivo after antibiotic treatment. Thromboxane synthesis was inhibited 28.3% to 81.7%. Agonist-induced rises in cytosolic calcium ((Ca2+)i) were inhibited 40.1% to 67.5% in vitro and 26.6% to 52.2% ex vivo. Therefore, Pen binds to platelets after prolonged exposure, resulting in irreversible dysfunction attributable to inhibition of TXA2 synthesis and impairment of the rise in (Ca2+)i. The loss of low-affinity TXA2/PGH2 receptors suggests that the primary site of action of these drugs is on the platelet membrane.

  20. Inhibition of the entomopathogenic fungus Metarhizium anisopliae in vitro by the bed bug defensive secretions (E)-2-hexenal and (E)-2-octenal

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The two major aldehydes (E)-2-hexenal and (E)-2-octenal emitted as defensive secretions by bed bugs Cimex lectularius L. (Hemiptera: Cimicidae), inhibit the in vitro growth of Metarhizium anisopliae (Metsch.) Sokorin (Hypocreales: Clavicipitaceae). These chemicals inhibit fungal growth by direct con...

  1. IL-1β Irreversibly Inhibits Tenogenic Differentiation and Alters Metabolism In Injured Tendon-Derived Progenitor Cells In Vitro

    PubMed Central

    Zhang, Kairui; Asai, Shuji; Yu, Bin; Enomoto-Iwamoto, Motomi

    2015-01-01

    Tendon injuries are common, and the damaged tendon often turns into scar tissue and never completely regains the original biomechanical properties. Previous studies have reported that the mRNA levels of inflammatory cytokines such as IL-1β are remarkably up-regulated in injured tendons. To examine how IL-1β impacts tendon repair process, we isolated the injured tendon-derived progenitor cells (inTPCs) from mouse injured Achilles tendons and studied the effects of IL-1β on the inTPCs in vitro. IL-1β treatment strongly reduced expression of tendon cell markers such as scleraxis and tenomodulin, and also down-regulated gene expression of collagen 1, collagen 3, biglycan and fibromodulin in inTPCs. Interestingly, IL-1β stimulated lactate production with increases in hexokinase II and lactate dehydrogenase expression and a decrease in pyruvate dehydrogenase. Inhibition of lactate production restored IL-1β-induced down-regulation of collagen1 and scleraxis expression. Furthermore, IL-1β significantly inhibited adipogenic, chondrogenic and osteogenic differentiation of inTPCs. Interestingly, inhibition of tenogenic and adipogenic differentiation was not recovered after removal of IL-1β while chondrogenic and osteogenic differentiation abilities were not affected. These findings indicate that IL-1β strongly and irreversibly impairs tenogenic potential and alters glucose metabolism in tendon progenitors appearing in injured tendons. Inhibition of IL-1β may be beneficial for maintaining function of tendon progenitor cells during the tendon repair process. PMID:26051275

  2. Amperozide, a putative anti-psychotic drug: Uptake inhibition and release of dopamine in vitro in the rat brain

    SciTech Connect

    Eriksson, E. )

    1990-01-01

    The effects of amperozide (a diphenylbutylpiperazinecarboxamide derivative) on the uptake and release of {sup 3}H-dopamine in vitro were investigated. Amperozide inhibited the amphetamine-stimulated release of dopamine from perfused rat striatal tissue in a dose-dependent manner. With 1 and 10 {mu}m amperozide there was significant inhibition of the amphetamine-stimulated release of dopamine, to 44 and 36 % of control. In contrast, 10 {mu}M amperozide significantly strengthened the electrically stimulated release of dopamine from perfused striatal slices. Amperozide 1-10 {mu}M had no significant effect on the potassium-stimulated release of dopamine, 10 {mu}M amperozide also slightly increased the basal release of {sup 3}H-dopamine from perfused striatal tissue. These effects on various types of release are similar to those reported for uptake inhibitors. The uptake of dopamine in striatal tissue was inhibited by amperozide with IC{sub 50} values of 18 {mu}M for uptake in chopped tissue and 1.0 {mu}M for uptake in synaptosomes. Amperozide also inhibited the uptake of serotonin in synaptosomes from frontal cortex, IC{sub 50} = 0.32 {mu}M and the uptake of noradrenaline in cortical synaptosomes, IC{sub 50} = 0.78 {mu}M.

  3. Pre- and post-treatment effect of physostigmine on soman-inhibited human erythrocyte and muscle acetylcholinesterase in vitro

    SciTech Connect

    Herkert, N.M.; Schulz, S.; Wille, T.; Thiermann, H.; Hatz, R.A.; Worek, F.

    2011-05-15

    Standard treatment of organophosphorus (OP) poisoning includes administration of an antimuscarinic (e.g., atropine) and of an oxime-based reactivator. However, successful oxime treatment in soman poisoning is limited due to rapid aging of phosphylated acetylcholinesterase (AChE). Hence, the inability of standard treatment procedures to counteract the effects of soman poisoning resulted in the search for alternative strategies. Recently, results of an in vivo guinea pig study indicated a therapeutic effect of physostigmine given after soman. The present study was performed to investigate a possible pre- and post-treatment effect of physostigmine on soman-inhibited human AChE given at different time intervals before or after perfusion with soman by using a well-established dynamically working in vitro model for real-time analysis of erythrocyte and muscle AChE. The major findings were that prophylactic physostigmine prevented complete inhibition of AChE by soman and resulted in partial spontaneous recovery of the enzyme by decarbamylation. Physostigmine given as post-treatment resulted in a time-dependent reduction of the protection from soman inhibition and recovery of AChE. Hence, these date indicate that physostigmine given after soman does not protect AChE from irreversible inhibition by the OP and that the observed therapeutic effect of physostigmine in nerve agent poisoning in vivo is probably due to other factors.

  4. A novel in vitro approach for simultaneous evaluation of CYP3A4 inhibition and kinetic aqueous solubility.

    PubMed

    Pérez, José; Díaz, Caridad; Asensio, Francisco; Palafox, Alexandra; Genilloud, Olga; Vicente, Francisca

    2015-02-01

    In the early stages of the drug discovery process, evaluation of the drug metabolism and physicochemical properties of new chemical entities is crucial to prioritize those candidates displaying a better profile for further development. In terms of metabolism, drug-drug interactions mediated through CYP450 inhibition are a significant safety concern, and therefore the effect of new candidate drugs on CYP450 activity should be screened early. In the initial stages of drug discovery, when physicochemical properties such as aqueous solubility have not been optimized yet, there might be a large number of candidate compounds showing artificially low CYP450 inhibition, and consequently potential drug-drug interaction toxicity might be overlooked. In this work, we present a novel in vitro approach for simultaneous evaluation of CYP3A4 inhibition potential and kinetic aqueous solubility (NIVA-CYPI-KS). This new methodology is based on fluorogenic CYP450 activities and turbidimetric measurements for compound solubility, and it provides a significant improvement in the use of resources and a better understanding of CYP450 inhibition data.

  5. Herbal extracts of Tribulus terrestris and Bergenia ligulata inhibit growth of calcium oxalate monohydrate crystals in vitro

    NASA Astrophysics Data System (ADS)

    Joshi, V. S.; Parekh, B. B.; Joshi, M. J.; Vaidya, A. B.

    2005-02-01

    A large number of people in this world are suffering from urinary stone problem. Calcium oxalate monohydrate (COM) and calcium oxalate dihydrate (COD) containing stones (calculi) are commonly found. In the present study, COM crystals were grown by a double diffusion gel growth technique using U-tubes. The gel was prepared from hydrated sodium metasilicate solution. The gel framework acts like a three-dimensional crucible in which the crystal nuclei are delicately held in the position of their formation, and nutrients are supplied for the growth. This technique can be utilized as a simplified screening static model to study the growth, inhibition and dissolution of urinary stones in vitro. The action of putative litholytic medicinal plants, Tribulus terrestris Linn. ( T.t) and Bergenia ligulata Linn. ( B.l.), has been studied in the growth of COM crystals. Tribulus terrestris and Bergenia ligulata are commonly used as herbal medicines for urinary calculi in India. To verify the inhibitive effect, aqueous extracts of Tribulus terrestris and Bergenia ligulata were added along with the supernatant solutions. The growth was measured and compared, with and without the aqueous extracts. Inhibition of COM crystal growth was observed in the herbal extracts. Maximum inhibition was observed in Bergenia ligulata followed by Tribulus terrestris. The results are discussed.

  6. Metformin and temozolomide act synergistically to inhibit growth of glioma cells and glioma stem cells in vitro and in vivo

    PubMed Central

    Yu, Zhiyun; Zhao, Gang; Xie, Guifang; Zhao, Liyan; Chen, Yong; Yu, Hongquan; Zhang, Zhonghua; Li, Cai; Li, Yunqian

    2015-01-01

    Glioblastoma (GBM) is the most frequent and aggressive brain tumor in adults. In spite of advances in diagnosis and therapy, the prognosis of patients with GBM has remained dismal. The fast recurrence and multi-drug resistance are some of the key challenges in combating brain tumors. Glioma stem cells (GSCs) which are considered the source of relapse and chemoresistance, the need for more effective therapeutic options is overwhelming. In our present work, we found that combined treatment with temozolomide (TMZ) and metformin (MET) synergistically inhibited proliferation and induced apoptosis in both glioma cells and GSCs. Combination of TMZ and MET significantly reduced the secondary gliosphere formation and expansion of GSCs. We first demonstrated that MET effectively inhibited the AKT activation induced by TMZ, and a combination of both drugs led to enhanced reduction of mTOR, 4EBP1 and S6K phosphorylation. In addition, the combination of the two drugs was accompanied with a powerful AMP-activated protein kinase (AMPK) activation, while this pathway is not determinant. Xenografts performed in nude mice demonstrate in vivo demonstrated that combined treatment significantly reduced tumor growth rates and prolonged median survival of tumor-bearing mice. In conclusion, TMZ in combination with MET synergistically inhibits the GSCs proliferation through downregulation of AKT-mTOR signaling pathway. The combined treatment of two drugs inhibits GSCs self-renewal capability and partly eliminates GSCs in vitro and in vivo. This combined treatment could be a promising option for patients with advanced GBM. PMID:26431379

  7. Curcumin/turmeric solubilized in sodium hydroxide inhibits HNE protein modification--an in vitro study.

    PubMed

    Kurien, Biji T; Scofield, R Hal

    2007-03-21

    Free radical mediated lipid peroxidation has been implicated in multiple diseases. A major oxidation by-product of this deleterious process is 4-hydroxy-2-nonenal (HNE). HNE is cytotoxic, mutagenic and genotoxic and is involved in disease pathogenesis. Curcumin, a non-steroidal anti-inflammatory agent (occurring as the yellow pigment found in the rhizomes of the perennial herb Curcuma longa known as turmeric), has emerged as the newest "nutraceutical" agent that has been shown to be efficacious against colon cancer and other disorders, including correcting cystic fibrosis defects. Since curcumin has been reported to have anti-oxidant properties we hypothesized that it will inhibit HNE-modification of a protein substrate. Using an ELISA that employed HNE-modification of solid phase antigen following immobilization, we found that the curcumin solubilized in dilute alkali (5mM sodium hydroxide, pH 11) inhibited HNE-protein modification by 65%. Turmeric also inhibited HNE-protein modification similarly (65%) but at a much lower alkali level (130muM sodium hydroxide, pH 7.6). Alkali by itself (5mM sodium hydroxide, pH 11) was found to enhance HNE modification by as much as 267%. Curcumin/turmeric has to inhibit this alkali enhanced HNE-modification prior to inhibiting the normal HNE protein modification induced by HNE. Thus, inhibition of HNE-modification could be a mechanism by which curcumin exerts its antioxidant effects. The pH at which the inhibition of HNE modification of substrate was observed was close to the physiological pH, making this formulation of curcumin potentially useful practically.

  8. In vitro extinction learning in Hermissenda: involvement of conditioned inhibition molecules.

    PubMed

    Cavallo, Joel S; Hamilton, Brittany N; Farley, Joseph

    2014-01-01

    Extinction of a conditioned association is typically viewed as the establishment of new learning rather than the erasure of the original memory. However, recent research in the nudibranch, Hermissenda crassicornis (H.c.) demonstrated that extinction training (using repeated light-alone presentations) given 15 min, but not 23 h, after memory acquisition reversed both the cellular correlates of learning (enhanced Type B cell excitability) and the behavioral changes (reduced phototaxis) produced by associative conditioning (pairings of light, CS, and rotation, US). Here, we investigated the putative molecular signaling pathways that underlie this extinction in H.c. by using a novel in vitro protocol combined with pharmacological manipulations. After intact H.c. received either light-rotation pairings (Paired), random presentations of light and ro