Molecular docking and in vitro studies of soap nut trypsin inhibitor (SNTI) against phospholipase A2 isoforms in therapeutic intervention of inflammatory diseases.

PubMed

Sirisha, Gandreddi V D; Vijaya Rachel, K; Zaveri, Kunal; Yarla, Nagendra Sastry; Kiranmayi, P; Ganash, Magdah; Alkreathy, Huda Mohammad; Rajeh, Nisreen; Ashraf, Ghulam Md

2018-07-15

Therapeutic value of allelochemicals in inflammatory disorders and the potential drug targets need to be elucidated to alleviate tissue and vascular injury. Natural anti-inflammatory agents are known to cause minimal adverse effects. Presence of different secondary metabolites (allelochemicals), protease inhibitors like soap nut trypsin inhibitor (SNTI) from Sapindus trifoliatus and allied compounds from natural sources cannot be blithely ignored as natural therapeutics. In the present study, SNTI, a prospective protease inhibitor isolated from the seeds of Sapindus trifoliatus were subjected to docking against three isoforms of Phospholipase A 2 (PLA 2 ) molecules of the inflammatory pathways which are localized in the membrane, cytosol and pancreas. Eleven ligand molecules were selected from Sapindus trifoliatus and docked against membrane, cytosolic and pancreatic PLA 2 . Cytosolic PLA 2 showed a strong inhibition by Kampferol, a secondary metabolite from seed endosperm of Sapindus trifoliatus. SNTI showed best interaction with membrane PLA 2 in both in silico as well as in in vitro studies. SNTI showed IC 50 value of 29.02 μM in in vitro assay. Docking interaction profiles and in vitro studies validate selected molecules from Sapindus trifoliatus as immunomodulators and can mollify inflammatory responses. Copyright © 2018 Elsevier B.V. All rights reserved.

Encapsulation of Naproxen in Lipid-Based Matrix Microspheres: Characterization and Release Kinetics

PubMed Central

Bhoyar, PK; Morani, DO; Biyani, DM; Umekar, MJ; Mahure, JG; Amgaonkar, YM

2011-01-01

The objective of this study was to microencapsulate the anti-inflammatory drug (naproxen) to provide controlled release and minimizing or eliminating local side effect by avoiding the drug release in the upper gastrointestinal track. Naproxen was microencapsulated with lipid-like carnauba wax, hydrogenated castor oil using modified melt dispersion (modified congealable disperse phase encapsulation) technique. Effect of various formulation and process variables such as drug-lipid ratio, concentration of modifier, concentration of dispersant, stirring speed, stirring time, temperature of external phase, on evaluatory parameters such as size, entrapment efficiency, and in vitro release of naproxen were studied. The microspheres were characterized for particle size, scanning electron microscopy (SEM), FT-IR spectroscopy, drug entrapment efficiency, in vitro release studies, for in vitro release kinetics. The shape of microspheres was found to be spherical by SEM. The drug entrapment efficiency of various batches of microspheres was found to be ranging from 60 to 90 %w/w. In vitro drug release studies were carried out up to 24 h in pH 7.4 phosphate buffer showing 50-65% drug release. In vitro drug release from all the batches showed better fitting with the Korsmeyer-Peppas model, indicating the possible mechanism of drug release to be by diffusion and erosion of the lipid matrix. PMID:21731354

Encapsulation of naproxen in lipid-based matrix microspheres: characterization and release kinetics.

PubMed

Bhoyar, P K; Morani, D O; Biyani, D M; Umekar, M J; Mahure, J G; Amgaonkar, Y M

2011-04-01

The objective of this study was to microencapsulate the anti-inflammatory drug (naproxen) to provide controlled release and minimizing or eliminating local side effect by avoiding the drug release in the upper gastrointestinal track. Naproxen was microencapsulated with lipid-like carnauba wax, hydrogenated castor oil using modified melt dispersion (modified congealable disperse phase encapsulation) technique. Effect of various formulation and process variables such as drug-lipid ratio, concentration of modifier, concentration of dispersant, stirring speed, stirring time, temperature of external phase, on evaluatory parameters such as size, entrapment efficiency, and in vitro release of naproxen were studied. The microspheres were characterized for particle size, scanning electron microscopy (SEM), FT-IR spectroscopy, drug entrapment efficiency, in vitro release studies, for in vitro release kinetics. The shape of microspheres was found to be spherical by SEM. The drug entrapment efficiency of various batches of microspheres was found to be ranging from 60 to 90 %w/w. In vitro drug release studies were carried out up to 24 h in pH 7.4 phosphate buffer showing 50-65% drug release. In vitro drug release from all the batches showed better fitting with the Korsmeyer-Peppas model, indicating the possible mechanism of drug release to be by diffusion and erosion of the lipid matrix.

Enhanced in Vitro Anti-Tumor Activity of 5-Azacytidine by Entrapment into Solid Lipid Nanoparticles

PubMed Central

Jahanfar, Farhad; Hasani, Akbar; Shanebandi, Dariush; Rahmati, Mohammad; Hamishehkar, Hamed

2016-01-01

Purpose: In this study the effectiveness of encapsulating of 5-azacytidine into the lipid nanoparticles was investigated and in vitro effect of encapsulated 5-azacytidine studied on MCF-7 cell lines Methods: 5-azacytidine -loaded solid lipid nanoparticles were produced by double emulsification (w/o/w) method by using stearic acid as lipid matrix, soy lecithin and poloxamer 407 as surfactant and co-surfactant respectively. Particle size, zeta potential, surface morphology, entrapment efficiency and kinetic of drug release were studied. In vitro effect of 5-azacytidine on MCF-7 cell line studied by MTT assay, DAPI staining, Rhodamine B relative uptake, and also Real time RT-PCR was performed for studying difference effect of free and encapsulated drug on expression of RARß2 gene. Results: The formulation F5 with 55.84±0.46 % of entrapment efficiency shows zero order kinetic of drug release and selected for in vitro studies; the cytotoxicity of free drug and encapsulated drug in 48 h of incubation have significant difference. DAPI staining shows morphology of apoptotic nucleus in both free and encapsulated drug, Rhodamine B labeled SLNs show time dependency and accumulation of SLNs in cytoplasm. Real time qRT-PCR doesn’t show any significant difference (p>0.05) in expression of RARß2 gene in both cells treated with free or encapsulated drug. Conclusion: The results of the present study indicated that the entrapment of 5-azacytidine into SLNs enhanced its cytotoxicity performance and may pave a way for the future design of a desired dosage form for 5-azacytidine. PMID:27766220

An Alternative Method for Long-Term Culture of Chicken Embryonic Stem Cell In Vitro.

PubMed

Zhang, Li; Wu, Yenan; Li, Xiang; Wei, Shao; Xing, Yiming; Lian, Zhengxing; Han, Hongbing

2018-01-01

Chicken embryonic stem cells (cESCs) obtained from stage X embryos provide a novel model for the study of avian embryonic development. A new way to maintain cESCs for a long period in vitro still remains unexplored. We found that the cESCs showed stem cell-like properties in vitro for a long term with the support of DF-1 feeder and basic culture medium supplemented with human basic fibroblast growth factor (hbFGF), mouse stem cell factor (mSCF), and human leukemia inhibitory factor (hLIF). During the long culture period, the cESCs showed typical ES cell morphology and expressed primitive stem cell markers with a relatively stable proliferation rate and high telomerase activity. These cells also exhibited the capability to differentiate into cardiac myocytes, smooth muscle cells, neural cells, osteoblast, and adipocyte in vitro . Chimera chickens were produced by cESCs cultured for 25 passages with this new culture system. The experiments showed that DF-1 was the optimal feeder and hbFGF was an important factor for maintaining the pluripotency of cESCs in vitro .

Bacteroides fragilis induce necrosis on mice peritoneal macrophages: In vitro and in vivo assays

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vieira, J.M.B.D., E-mail: jmanya@terra.com.br; Laboratorio de Biologia de Anaerobios, IMPPG, UFRJ, Rio de Janeiro; Seabra, S.H.

Bacteroides fragilis is an anaerobic bacteria component of human intestinal microbiota and agent of infections. In the host B. fragilis interacts with macrophages, which produces toxic radicals like NO. The interaction of activated mice peritoneal macrophages with four strains of B. fragilis was evaluated on this study. Previously was shown that such strains could cause metabolic and morphologic alterations related to macrophage death. In this work propidium iodide staining showed the strains inducing macrophage necrosis in that the labeling was evident. Besides nitroblue tetrazolium test showed that B. fragilis stimulates macrophage to produce oxygen radicals. In vivo assays performed inmore » BalbC mice have results similar to those for in vitro tests as well as scanning electron microscopy, which showed the same surface pore-like structures observed in vitro before. The results revealed that B. fragilis strains studied lead to macrophage death by a process similar to necrosis.« less

The novel oral glucan synthase inhibitor SCY-078 shows in vitro activity against sessile and planktonic Candida spp.

PubMed

Marcos-Zambrano, Laura Judith; Gómez-Perosanz, Marta; Escribano, Pilar; Bouza, Emilio; Guinea, Jesús

2017-07-01

We studied the antifungal activity of SCY-078 (an orally bioavailable 1,3-β -d- glucan synthesis inhibitor), micafungin and fluconazole against the planktonic and sessile forms of 178 Candida and non- Candida isolates causing fungaemia in patients recently admitted to a large European hospital. The in vitro activity of SCY-078, micafungin and fluconazole against the planktonic form of the isolates was assessed using EUCAST EDef 7.3 and CLSI M27-A3. Antibiofilm activity was assessed using the XTT reduction assay. SCY-078 and micafungin showed potent in vitro activity against Candida and non- Candida isolates. The in vitro activity of both drugs was similar, but SYC-078 displayed significantly lower MIC values than micafungin against Candida parapsilosis and non- Candida isolates, whereas micafungin displayed significantly lower MIC values for the remaining species ( P  <0.001). In contrast, SCY-078 and micafungin showed essentially the same activity against the biofilms with the exception of Candida glabrata , in which the micafungin sessile MIC values were significantly lower ( P  <0.001). These observations were confirmed by assessing biofilm structure by scanning electron microscopy after antifungal treatment. Our study showed that the high in vitro activity of SCY-078 against invasive Candida isolates in both sessile and planktonic forms is comparable to that of micafungin. © The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Formulation and Characterization of Aceclofenac -Aloe vera Transemulgel.

PubMed

Raju, Y Prasanna; Haritha, K; Satyanarayana, Rao P; Vandana, K R; Bindu, D Thushara; Vinesha, V; Chowdary, V Harini

2015-01-01

The present research was aimed to formulate aceclofenac transemulgel using Aloe vera as gel base. The prepared formulations were subjected to physical characterization, in-vitro and in-vivo assessment. Aceclofenac, a hydrophobic potential non steroidal anti inflammatory drug, causes ulceration upon chronic oral administration, could be formulated into transemulgel to enhance therapeutic efficacy and to lower the unwanted side effects. The transemulgel was prepared from aqueous Aloe vera gel and aceclofenac emulsion. The prepared transemulgel was evaluated for its pH, viscosity, drug content, skin irritation, in-vitro diffusion and accelerated stability studies. The prepared aceclofenac-Aloe vera tranemulgel and commercial aceclofenac gel were subjected to pharmacodynamic studies in albino rats of Wistar strain employing carrageenan induced left hind paw edema method to assess the anti-inflammatory effect. The transemulgel showed a pH of 6.78 and viscosity of 18 cps. In-vitro diffusion data revealed better permeation characteristics. Topical application of formulation found no skin irritation. Stability study has proved the integrity of the formulation. The prepared aceclofenac Aloe vera transemulgel showed better in-vitro drug release when compared with the commercial aceclofenac gel formulation. Anti-inflammatory activity in treated rats showed the significant paw volume reduction at p<0.05 compared with that of control. Thus, it is concluded that aceclofenac, a potential non steroidal anti inflammatory drug, showed high therapeutic efficiency when formulated into transemulgel using aqueous Aloe vera as gel base.

In vitro and in vivo photosensitized inactivation of dermatophyte fungi by heterotricyclic dyes.

PubMed Central

Propst, C; Lubin, L

1978-01-01

The ability of three heterotricyclic dyes to photosensitize dermatophyte fungi was studied with Trichophyton mentagrophytes and Microsporum gypseum. In vitro studies showed that methylene blue, neutral red, and proflavine were capable of killing these fungi when used in conjunction with broad-spectrum light. Proflavine, however, killed both fungi most rapidly and was used for further studies. Fungal killing by proflavine plus light was dependent on dye concentration, pH, light wavelength, and light intensity. Based on the in vitro studies, a treatment regimen was developed for in vivo use on experimentally infected animals. When treatment of guinea pigs inoculated with T. mentagrophytes was begun during fungal invasion, lesion formation at inoculated sites was either prevented or substantially reduced. When treatment was begun after lesion formation, however, light-plus-dyed treated sites showed only slightly faster curing than untreated sites. PMID:669788

In vitro evaluation, in vivo quantification, and microbial diversity studies of nutritional strategies for reducing enteric methane production.

PubMed

Abdalla, Adibe Luiz; Louvandini, Helder; Sallam, Sobhy Mohamed Abdallah Hassan; Bueno, Ives Cláudio da Silva; Tsai, Siu Mui; Figueira, Antonio Vargas de Oliveira

2012-06-01

The main objective of the present work was to study nutritive strategies for lessening the CH(4) formation associated to ruminant tropical diets. In vitro gas production technique was used for evaluating the effect of tannin-rich plants, essential oils, and biodiesel co-products on CH(4) formation in three individual studies and a small chamber system to measure CH(4) released by sheep for in vivo studies was developed. Microbial rumen population diversity from in vitro assays was studied using qPCR. In vitro studies with tanniniferous plants, herbal plant essential oils derived from thyme, fennel, ginger, black seed, and Eucalyptus oil (EuO) added to the basal diet and cakes of oleaginous plants (cotton, palm, castor plant, turnip, and lupine), which were included in the basal diet to replace soybean meal, presented significant differences regarding fermentation gas production and CH(4) formation. In vivo assays were performed according to the results of the in vitro assays. Mimosa caesalpineaefolia, when supplemented to a basal diet (Tifton-85 hay Cynodon sp, corn grain, soybean meal, cotton seed meal, and mineral mixture) fed to adult Santa Ines sheep reduced enteric CH(4) emission but the supplementation of the basal diet with EuO did not affect (P > 0.05) methane released. Regarding the microbial studies of rumen population diversity using qPCR with DNA samples collected from the in vitro trials, the results showed shifts in microbial communities of the tannin-rich plants in relation to control plant. This research demonstrated that tannin-rich M. caesepineapholia, essential oil from eucalyptus, and biodiesel co-products either in vitro or in vivo assays showed potential to mitigate CH(4) emission in ruminants. The microbial community study suggested that the reduction in CH(4) production may be attributed to a decrease in fermentable substrate rather than to a direct effect on methanogenesis.

Spine micromorphology of normal and hyperhydric Mammillaria gracilis Pfeiff. (Cactaceae) shoots.

PubMed

Peharec, P; Posilović, H; Balen, B; Krsnik-Rasol, M

2010-07-01

Artificial conditions of tissue culture affect growth and physiology of crassulacean acid metabolism plants which often results in formation of hyperhydric shoots. In in vitro conditions Mammillaria gracilis Pfeiff. (Cactaceae) growth switches from organized to unorganized way, producing a habituated organogenic callus which simultaneously regenerates morphologically normal as well as altered hyperhydric shoots. In this study, influence of tissue culture conditions on morphology of cactus spines of normal and hyperhydric shoots was investigated. Spines of pot-grown Mammillaria plants and of in vitro regenerated shoots were examined with stereo microscope and scanning electron microscope. The pot-grown plants had 16-17 spines per areole. In vitro grown normal shoots, even though they kept typical shoot morphology, had lower number of spines (11-12) and altered spine morphology. This difference was even more pronounced in spine number (six to seven) and morphology of the hyperhydric shoots. Scanning electron microscopy analysis revealed remarkable differences in micromorphology of spine surface between pot-grown and in vitro grown shoots. Spines of in vitro grown normal shoots showed numerous long trichomes, which were more elongated on spines of the hyperhydric shoots; the corresponding structures on spine surface of pot-grown plants were noticed only as small protrusions. Scanning electron microscopy morphometric studies showed that the spines of pot-grown plants were significantly longer compared to the spines of shoots grown in tissue culture. Moreover, transverse section shape varies from elliptical in pot-grown plants to circular in normal and hyperhydric shoots grown in vitro. Cluster and correspondence analyses performed on the scanning electron microscope obtained results suggest great variability among spines of pot-grown plants. Spines of in vitro grown normal and hyperhydric shoots showed low level of morphological variation among themselves despite the significant difference in shoot morphology.

Radiation and chemical mutagen induced somaclonal variations through in vitro organogenesis of cotton (Gossypium hirsutum L.).

PubMed

Muthusamy, Annamalai; Jayabalan, Narayanasamy

2014-12-01

The purpose of the investigation was to induce somaclonal variations by gamma rays (GR), ethylmethane sulphonate (EMS) and sodium azide (SA) during in vitro organogenesis of cotton. The shoot tip explants were irradiated with 5-50 Gray (Gy) GR (Cobalt 60), 0.5-5.0 mM EMS and SA separately, and inoculated on Murashige and Skoog (MS) medium fortified with plant growth regulator (PGR) for organogenesis. The plantlets with well-developed root systems were acclimatized and transferred into the experimental field to screen the somaclonal variations during growth and development. The number of somaclonal variations was observed in growth of irradiated/treated shoot tips, multiplication, plantlet regeneration and growth in vitro and ex vitro. The lower doses/concentrations of mutagenic treatments showed significant enhancement in selected agronomical characters and they showed decreased trends with increasing doses/concentrations of mutagenic agents. The results of the present study revealed the influence of lower doses/concentrations of mutagenic treatments on in vitro and ex vitro growth of cotton plantlets and their significant improvement in agronomical characters which needs further imperative stability analysis. The present observations showed the platform to use lower doses/concentrations of mutagenic agents to induce variability for enhanced agronomical characters, resistant and tolerant cotton varieties.

Cherenkov radiation imaging of beta emitters: in vitro and in vivo results

NASA Astrophysics Data System (ADS)

Spinelli, Antonello E.; Boschi, Federico; D'Ambrosio, Daniela; Calderan, Laura; Marengo, Mario; Fenzi, Alberto; Menegazzi, Marta; Sbarbati, Andrea; Del Vecchio, Antonella; Calandrino, Riccardo

2011-08-01

The main purpose of this work was to investigate both in vitro and in vivo Cherenkov radiation (CR) emission coming from 18F and 32P. The main difference between 18F and 32P is mainly the number of the emitted light photons, more precisely the same activity of 32P emits more CR photons with respect to 18F. In vitro results obtained by comparing beta counter measurements with photons average radiance showed that Cherenkov luminescence imaging (CLI) allows quantitative tracer activity measurements. In order to investigate in vivo the CLI approach, we studied an experimental xenograft tumor model of mammary carcinoma (BB1 tumor cells). Cherenkov in vivo dynamic whole body images of tumor bearing mice were acquired and the tumor tissue time activity curves reflected the well-known physiological accumulation of 18F-FDG in malignant tissues with respect to normal tissues. The results presented here show that it is possible to use conventional optical imaging devices for in vitro or in vivo study of beta emitters.

[Preparation and in vitro release characteristics of vincristine sulphate loaded poly (butylcyanoacrylate) nanoparticles].

PubMed

Tan, Rong; Liu, Ying; Feng, Nianping; Zhao, Jihui

2011-06-01

To prepare vincristine sulphate loaded poly (butylcyanoacrylate) nanoparticles (VCR-PBCA-NPs) and to investigate the in vitro release charactersitics. VCR-PBCA-NPs were prepared by emulsion polymerization method, and characterized for morphology, particle size, drug encapsulation efficiency and loading efficiency. The formulation was optimized using central composite design and response surface methodology. In vitro release study of VCR-PBCA-NPs was performed by dialysis technique. Model fitting was used to determine the kinetics and to discuss the mechanism. The nanoparticles were spherical and uniform with a mean diameter of (98.9 +/- 3.05) nm. The drug encapsulation efficiency and loading efficiency were (55.23 +/- 0.96)% and (7.87 +/- 0.11)%, respectively. In vitro release results showed that 63.66% of VCR was released from VCR-PBCA-NPs in 4 h, and the Weibull model fitted VCR release pattern best. The VCR-PBCA-NPs prepared in this study showed sustained release compared with VCR solution.

Evaluation of Cytarabine Against Ewing Sarcoma Xenografts by the Pediatric Preclinical Testing Program

PubMed Central

Houghton, Peter J.; Morton, Christopher L.; Kang, Min; Reynolds, C. Patrick; Billups, Catherine A.; Favours, Edward; Payne-Turner, Debbie; Tucker, Chandra; Smith, Malcolm A.

2015-01-01

Treatment with the nucleoside analog cytarabine has been shown to mimic changes in gene expression associated with down-regulation of the EWS-FLI1 oncogene in Ewing sarcoma cell lines, selectively inhibit their growth in vitro, and cause tumor regression in athymic nude mice. For this report cytarabine was studied in vitro against a panel of 23 pediatric cancer cell lines and in vivo against 6 Ewing sarcoma xenografts. Acute lymphoblastic leukemia cell lines were the most sensitive to cytarabine in vitro (median IC50 9 nM), while Ewing sarcoma cell lines showed intermediate sensitivity (median IC50 232 nM). Cytarabine at a dose of 150 mg/kg administered daily 5× failed to significantly inhibit growth of five xenograft models, but reduced growth rate of the A673 xenograft by 50%. Cytarabine shows no differential in vitro activity against Ewing sarcoma cell lines and is ineffective in vivo against Ewing sarcoma xenografts at the dose and schedule studied. PMID:20979180

[Study on solid dispersion of precipitated calcium carbonate-based oleanolic acid].

PubMed

Yan, Hong-mei; Zhang, Zhen-hai; Jia, Xiao-bin; Jiang, Yan-rong; Sun, E

2015-05-01

Oleanolic acid-precipitated calcium carbonate solid dispersion was prepared by using solvent evaporation method. The microscopic structure and physicochemical properties of solid dispersion were analyzed using differential scanning calorimetry and scanning electron microscopy (SEM). And its in vitro release also was investigated. The properties of the precipitated calcium carbonate was studied which was as a carrier of oleanolic acid solid dispersion. Differential scanning calorimetry analysis suggested that oleanolic acid may be present in solid dispersion as amorphous substance. The in vitro release determination results of oleanolic acid-precipitated calcium carbonate (1: 5) solid dispersion showed accumulated dissolution rate of.oleanolic acid was up to 90% at 45 min. Accelerating experiment showed that content and in vitro dissolution of oleanolic acid solid dispersion did not change after storing over 6 months. The results indicated that in vitro dissolution of oleanolic acid was improved greatly by the solid dispersion with precipitated calcium carbonate as a carrier. The solid dispersion is a stabilizing system which has actual applied value.

Chromosomal Radiosensitivity in Lymphocytes of Cervix Cancer Patients—Correlation with Side Effect after Radiotherapy

NASA Astrophysics Data System (ADS)

Wegierek-Ciuk, Aneta; Lankoff, Anna; Lisowska, Halina; Banasik-Nowak, Anna; Arabski, Michał; Kedzierawski, Piotr; Florek, Agnieszka; Wojcik, Andrzej

2010-01-01

It is well known that cancer patients receiving similar radiotherapy treatments differ widely in normal tissue reactions ranging from undetectable to unacceptably severe levels. Therefore, an important goal of radiobiological research is to establish a test which would allow identifying individual radiosensitivity of patients prior to radiotherapy. The aim of the presented study is to assess the relationship between lymphocyte intrinsic radiosensitivity in vitro and early reaction of normal tissue in cervix cancer patients treated by radiotherapy. The following endpoints are analyzed in vitro: frequency of micronuclei, the kinetics of DNA repair and apoptosis. Acute normal tissue reaction to radiotherapy in the skin, bladder and rectum are scored according to the EORTC/RTOG scale. Our results show a wide inter-individual variability in chromosomal radiosensitivity in vitro. The majority of patients show a Grade 0, 1 or 2 reaction for all organs studied. No statistically significant correlation has been observed between the in vitro results in lymphocytes and the degree of early normal tissue and organ reaction.

Collaborative study for the validation of alternative in vitro potency assays for human tetanus immunoglobulin.

PubMed

Gross, S; Janssen, S W J; de Vries, B; Terao, E; Daas, A; Buchheit, K-H

2009-10-01

The European Pharmacopoeia (Ph. Eur.) monograph Human tetanus immunoglobulin (0398) gives a clear outline of the in vivo assay to be performed to determine the potency of human tetanus immunoglobulins during their development. Furthermore, it states that an in vitro method shall be validated for the batch potency estimation. Since no further guidance is given on the in vitro assay, every control laboratory concerned is free to design and validate an in-house method. At the moment there is no agreed in vitro method available. The aim of this study was to validate and compare 2 alternative in vitro assays, i.e. an enzyme-linked immunoassay (EIA) and a toxoid inhibition assay (TIA), through an international collaborative study, in view of their eventual inclusion into the Ph. Eur.. The study was run in the framework of the Biological Standardisation Programme (BSP), under the aegis of the European Commission and the Council of Europe. The collaborative study reported here involved 21 laboratories (public and industry) from 15 countries. Initially, 3 samples with low, medium and high potencies were tested by EIA and TIA. Results showed good reproducibility and repeatability of the 2 in vitro methods. The correlation of the data with the in vivo potency assigned by the manufacturers however appeared initially poor for high potency samples. Thorough re-examination of the data showed that the in vivo potencies assigned by the manufacturers had to be corrected: one for potency loss at the time of in vitro testing and one because of a reporting error. After these corrections the values obtained by in vivo and in vitro methods were in close agreement. A supplementary collaborative work was carried out to validate the 2 methods for immunoglobulin products with high potencies. Eight laboratories (public and industry) took part in this additional study to test 3 samples with medium and high potencies by EIA and TIA. Results confirmed that the 2 alternative methods are comparable in terms of assay repeatability, precision and reproducibility. In all laboratories, both methods discriminated between the low, medium and high potency samples. Analysis of the data collected in this study showed a good correlation between EIA and TIA potency estimates as well as a close agreement between values obtained by in vitro and in vivo methods. The study demonstrated that EIA and TIA are suitable quality control methods for polyclonal human tetanus immunoglobulin, which can be standardised in a quality control laboratory using a quality assurance system. Consequently, the Ph. Eur. Group of Experts 6B on Human Blood and Blood products decided in April 2009 to include both methods as examples in the Ph. Eur. monograph 0398 on Human Tetanus immunoglobulin.

Tumoricidal activity of low-energy 160-KV versus 6-MV X-rays against platinum-sensitized F98 glioma cells

PubMed Central

Lim, Sara N.; Pradhan, Anil K.; Barth, Rolf F.; Nahar, Sultana N.; Nakkula, Robin J.; Yang, Weilian; Palmer, Alycia M.; Turro, Claudia; Weldon, Michael; Bell, Erica Hlavin; Mo, Xiaokui

2015-01-01

The purposes of this study were (i) to investigate the differences in effects between 160-kV low-energy and 6-MV high-energy X-rays, both by computational analysis and in vitro studies; (ii) to determine the effects of each on platinum-sensitized F98 rat glioma and murine B16 melanoma cells; and (iii) to describe the in vitro cytotoxicity and in vivo toxicity of a Pt(II) terpyridine platinum (Typ-Pt) complex. Simulations were performed using the Monte Carlo code Geant4 to determine enhancement in absorption of low- versus high-energy X-rays by Pt and to determine dose enhancement factors (DEFs) for a Pt-sensitized tumor phantom. In vitro studies were carried out using Typ-Pt and again with carboplatin due to the unexpected in vivo toxicity of Typ-Pt. Cell survival was determined using clonogenic assays. In agreement with computations and simulations, in vitro data showed up to one log unit reduction in surviving fractions (SFs) of cells treated with 1–4 µg/ml of Typ-Pt and irradiated with 160-kV versus 6-MV X-rays. DEFs showed radiosensitization in the 50–200 keV range, which fell to approximate unity at higher energies, suggesting marginal interactions at MeV energies. Cells sensitized with 1–5 or 7 µg/ml of carboplatin and then irradiated also showed a significant decrease (P < 0.05) in SFs. However, it was unlikely this was due to increased interactions. Theoretical and in vitro studies presented here demonstrated that the tumoricidal activity of low-energy X-rays was greater than that of high-energy X-rays against Pt-sensitized tumor cells. Determining whether radiosensitization is a function of increased interactions will require additional studies. PMID:25266332

Antitubercular activity of ZnO nanoparticles prepared by solution combustion synthesis using lemon juice as bio-fuel.

PubMed

Gopala Krishna, Prashanth; Paduvarahalli Ananthaswamy, Prashanth; Trivedi, Priyanka; Chaturvedi, Vinita; Bhangi Mutta, Nagabhushana; Sannaiah, Ananda; Erra, Amani; Yadavalli, Tejabhiram

2017-06-01

In this study, we report the synthesis, structural and morphological characteristics of zinc oxide (ZnO) nanoparticles using solution combustion synthesis method where lemon juice was used as the fuel. In vitro anti-tubercular activity of the synthesized ZnO nanoparticles and their biocompatibility studies, both in vitro and in vivo were carried out. The synthesized nanoparticles showed inhibition of Mycobacterium tuberculosis H37Ra strain at concentrations as low as 12.5μg/mL. In vitro cytotoxicity study performed with normal mammalian cells (L929, 3T3-L1) showed that ZnO nanoparticles are non-toxic with a Selectivity Index (SI) >10. Cytotoxicity performed on two human cancer cell lines DU-145 and Calu-6 indicated the anti-cancer activity of ZnO nanoparticles at varied concentrations. Results of blood hemolysis indicated the biocompatibility of ZnO nanoparticles. Furthermore, in vivo toxicity studies of ZnO nanoparticles conducted on Swiss albino mice (for 14days as per the OECD 423 guidelines) showed no evident toxicity. Copyright © 2017 Elsevier B.V. All rights reserved.

Saccharomyces cerevisiae show low levels of traversal across human endothelial barrier in vitro.

PubMed

Pérez-Torrado, Roberto; Querol, Amparo

2017-01-01

Background :   Saccharomyces cerevisiae is generally considered safe, and is involved in the production of many types of foods and dietary supplements. However, some isolates, which are genetically related to strains used in brewing and baking, have shown virulent traits, being able to produce infections in humans, mainly in immunodeficient patients. This can lead to systemic infections in humans. Methods : In this work, we studied S. cerevisiae isolates in an in vitro human endothelial barrier model, comparing their behaviour with that of several strains of the related pathogens Candida glabrata and Candida albicans . Results : The results showed that this food related yeast is able to cross the endothelial barrier in vitro . However, in contrast to C. glabrata and C. albicans , S. cerevisiae showed very low levels of traversal. Conclusions : We conclude that using an in vitro human endothelial barrier model with S. cerevisiae can be useful to evaluate the safety of S. cerevisiae strains isolated from foods.

Development of low erosive carbonated fruit drinks. 1. Evaluation of two experimental orange drinks in vitro and in situ.

PubMed

Hunter, M L; Hughes, J A; Parker, D M; West, N X; Newcombe, R G; Addy, M

2003-05-01

To determine the in vitro erosive potential and in situ erosive effect of two new formulation low calorie carbonated orange drinks with that of two conventional diet products and water. In the in vitro study, six specimens of deciduous and permanent enamel were randomly allocated to each of the five products and a '4h' protocol employed. In the in situ study, 15 healthy volunteers participated in a single centre, single blind, 5-phase crossover study, conducted according to Good Clinical Practice, and employing a validated model. The in vitro erosive potential of the experimental formulations was less than that of the comparators at all time points. Conversely, the observed erosive potential of both experimental formulations was greater than that of the control. Consistent statistically significant differences were found in relation to permanent enamel only. Unfortunately, the in situ study did not produce results entirely consistent with those of the in vitro study. Notably, a generally progressive loss of enamel was observed in specimens exposed to the control. The data from the in vitro study show the experimental formulations to have low comparative erosivity. However, the methodologies in vitro and in situ somewhat unusually do not correlate in ranking the erosivity of drinks. The results of this study should therefore be viewed with caution, further research being clearly warranted.

Development of mucoadhesive patches for buccal administration of carvedilol.

PubMed

Vishnu, Y Vamshi; Chandrasekhar, K; Ramesh, G; Rao, Y Madhusudan

2007-01-01

A buccal patch for systemic administration of carvedilol in the oral cavity has been developed using two different mucoadhesive polymers. The formulations were tested for in vitro drug permeation studies, buccal absorption test, in vitro release studies, moisture absorption studies and in vitro bioadhesion studies. The physicochemical interactions between carvedilol and polymers were investigated by Fourier transform infrared (FTIR) Spectroscopy. According to FTIR the drug did not show any evidence of an interaction with the polymers used and was present in an unchanged state. XRD studies reveal that the drug is in crystalline state in the polymer matrix. The results indicate that suitable bioadhesive buccal patches with desired permeability could be prepared. Bioavailability studies in healthy pigs reveal that carvedilol has got good buccal absorption. The bioavailability of carvedilol from buccal patches has increased 2.29 folds when compared to that of oral solution. The formulation AC5 (HPMC E 15) shows 84.85 + 0.089% release and 38.69 + 6.61% permeated through porcine buccal membrane in 4 hr. The basic pharmacokinetic parameters like the C(max), T(max) and AUC(total) were calculated and showed statistically significant difference (P<0.05) when given by buccal route compared to that of oral solution.

Preparation and characterization of metoprolol tartrate containing matrix type transdermal drug delivery system.

PubMed

Malipeddi, Venkata Ramana; Awasthi, Rajendra; Ghisleni, Daniela Dal Molim; de Souza Braga, Marina; Kikuchi, Irene Satiko; de Jesus Andreoli Pinto, Terezinha; Dua, Kamal

2017-02-01

The present study aimed to develop matrix-type transdermal drug delivery system (TDDS) of metoprolol tartrate using polyvinyl pyrrolidone (PVP) and polyvinyl alcohol (PVA). The transdermal films were evaluated for physical parameters, Fourier transform infrared spectroscopy analysis (FTIR), differential scanning calorimetry (DSC), in vitro drug release, in vitro skin permeability, skin irritation test and stability studies. The films were found to be tough, non-sticky, easily moldable and possess good tensile strength. As the concentration of PVA was increased, the tensile strength of the films was also increased. Results of FTIR spectroscopy and DSC revealed the absence of any drug-polymer interactions. In vitro release of metoprolol followed zero-order kinetics and the mechanism of release was found to be diffusion rate controlled. In vitro release studies of metoprolol using Keshary-Chein (vertical diffusion cell) indicated 65.5 % drug was released in 24 h. In vitro skin permeation of metoprolol transdermal films showed 58.13 % of the drug was released after 24 h. In vitro skin permeation of metoprolol followed zero-order kinetics in selected formulations. The mechanism of release was found to be diffusion rate controlled. In a 22-day skin irritation test, tested formulation of transdermal films did not exhibit any allergic reactions, inflammation, or contact dermatitis. The transdermal films showed good stability in the 180-day stability study. It can be concluded that the TDDS of MPT can help in bypassing the first-pass effect and will provide patient improved compliance, without sacrificing the therapeutic advantages of the drugs.

Efficacy evaluations of Mimosa pudica tannin isolate (MPT) for its anti-ophidian properties.

PubMed

Ambikabothy, Jamunaa; Ibrahim, Halijah; Ambu, Stephen; Chakravarthi, Srikumar; Awang, Khalijah; Vejayan, Jaya

2011-09-01

Evaluations of the anti-snake venom efficacy of Mimosa pudica tannin isolate (MPT) obtained from root of the plant. MPT was investigated in vitro and in vivo for its efficacy against the venom of Naja kaouthia snake. In vitro: (1) mice injected i.p. with MPT pre-incubated with Naja kaouthia venom at concentrations as low as 0.625 mg/ml showed 100% survival after a 24-h observation period. (2) In the proteomics study, mice injected with MPT pre-incubated with the Naja kaouthia venom showed down-regulation of five serum proteins. (3) In the protein-dye-binding study, the percentage of Bradford dye-protein binding showed a reduction relative to the decrease in MPT concentration used to incubate with the venom. In vivo: the results from the animal studies showed that MPT had no in vivo protection against the Naja kaouthia venom (0.875 mg/kg) in four different rescue modes and in an oral pre-treatment experiment. The study indicated the promising ability of MPT to neutralize the Naja kaouthia venom in in vitro experiments but fell short in its in vivo potential. As such, the use of Mimosa pudica (Mimosaceae) as therapeutics for snake bites is questionable as all the possible in vivo rescue studies and pre-treatment of the active constituents showed no protection against the affected mice. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Plant Regeneration and Cellular Behaviour Studies in Celosia cristata Grown In Vivo and In Vitro

PubMed Central

Taha, Rosna Mat; Wafa, Sharifah Nurashikin

2012-01-01

Tissue culture studies of Celosia cristata were established from various explants and the effects of various hormones on morphogenesis of this species were examined. It was found that complete plant regeneration occurred at highest percentage on MS medium supplemented with 2.0 mg/L NAA and 1.5 mg/L BAP, with the best response showed by shoot explants. In vitro flowering was observed on MS basal medium after six weeks. The occurrence of somaclonal variation and changes in cellular behavior from in vivo and in vitro grown plants were investigated through cytological studies and image analysis. It was observed that Mitotic Index (MI), mean chromosome numbers, and mean nuclear to cell area ratio of in vitro root meristem cells were slightly higher compared to in vivo values. However, in vitro plants produced lower mean cell areas but higher nuclear areas when compared to in vivo plants. Thus, no occurrence of somaclonal variation was detected, and this was supported by morphological features of the in vitro plants. PMID:22593677

In vitro porcine blood-brain barrier model for permeability studies: pCEL-X software pKa(FLUX) method for aqueous boundary layer correction and detailed data analysis.

PubMed

Yusof, Siti R; Avdeef, Alex; Abbott, N Joan

2014-12-18

In vitro blood-brain barrier (BBB) models from primary brain endothelial cells can closely resemble the in vivo BBB, offering valuable models to assay BBB functions and to screen potential central nervous system drugs. We have recently developed an in vitro BBB model using primary porcine brain endothelial cells. The model shows expression of tight junction proteins and high transendothelial electrical resistance, evidence for a restrictive paracellular pathway. Validation studies using small drug-like compounds demonstrated functional uptake and efflux transporters, showing the suitability of the model to assay drug permeability. However, one limitation of in vitro model permeability measurement is the presence of the aqueous boundary layer (ABL) resulting from inefficient stirring during the permeability assay. The ABL can be a rate-limiting step in permeation, particularly for lipophilic compounds, causing underestimation of the permeability. If the ABL effect is ignored, the permeability measured in vitro will not reflect the permeability in vivo. To address the issue, we explored the combination of in vitro permeability measurement using our porcine model with the pKa(FLUX) method in pCEL-X software to correct for the ABL effect and allow a detailed analysis of in vitro (transendothelial) permeability data, Papp. Published Papp using porcine models generated by our group and other groups are also analyzed. From the Papp, intrinsic transcellular permeability (P0) is derived by simultaneous refinement using a weighted nonlinear regression, taking into account permeability through the ABL, paracellular permeability and filter restrictions on permeation. The in vitro P0 derived for 22 compounds (35 measurements) showed good correlation with P0 derived from in situ brain perfusion data (r(2)=0.61). The analysis also gave evidence for carrier-mediated uptake of naloxone, propranolol and vinblastine. The combination of the in vitro porcine model and the software analysis provides a useful tool to better predict BBB permeability in vivo and gain better mechanistic information about BBB permeation. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

Biodegradable injectable in situ implants and microparticles for sustained release of montelukast: in vitro release, pharmacokinetics, and stability.

PubMed

Ahmed, Tarek A; Ibrahim, Hany M; Samy, Ahmed M; Kaseem, Alaa; Nutan, Mohammad T H; Hussain, Muhammad Delwar

2014-06-01

The objective of this study was to investigate the sustained release of a hydrophilic drug, montelukast (MK), from two biodegradable polymeric drug delivery systems, in situ implant (ISI) and in situ microparticles (ISM). N-Methyl pyrrolidone (NMP), dimethyl sulfoxide (DMSO), triacetin, and ethyl acetate were selected as solvents. The release of 10% (w/v) MK from both systems containing poly-lactic-co-glycolic acid (PLGA) as the biodegradable polymer was compared. Upon contact with the aqueous medium, the PLGA in ISI and ISM systems solidified resulting in implants and microparticles, respectively. The in vitro drug release from the ISI system showed marked difference from miscible solvents (NMP and DMSO) than the partially miscible ones (triacetin and ethyl acetate), and the drug release decreased with increased PLGA concentration. In the ISM system, the initial in vitro drug release decreased with decreased ratio of polymer phase to external oil phase. In vivo studies in rats showed that ISM had slower drug release than the drug release from ISI. Also, the ISM system when compared to ISI system had significantly reduced initial burst effect. In vitro as well as the in vivo studies for both ISI and ISM systems showed sustained release of MK. The ISM system is suitable for sustained release of MK over 4-week period with a lower initial burst compared to the ISI system. Stability studies of the ISI and ISM formulations showed that MK is stable in the formulations stored at 4°C for more than 2 years.

The Escherichia coli cAMP receptor protein bound at a single target can activate transcription initiation at divergent promoters: a systematic study that exploits new promoter probe plasmids.

PubMed Central

El-Robh, Mohamed Samir; Busby, Stephen J W

2002-01-01

We report the first detailed quantitative study of divergent promoters dependent on the Escherichia coli cAMP receptor protein (CRP), a factor known to activate transcription initiation at target promoters by making direct interactions with the RNA polymerase holoenzyme. In this work, we show that CRP bound at a single target site is able to activate transcription at two divergently organized promoters. Experiments using promoter probe plasmids, designed to study divergent promoters in vivo and in vitro, show that the divergent promoters function independently. Further in vitro experiments show that two holo RNA polymerase molecules cannot be accommodated simultaneously at the divergent promoters. PMID:12350222

The use of in vitro technologies coupled with high resolution accurate mass LC-MS for studying drug metabolism in equine drug surveillance.

PubMed

Scarth, James P; Spencer, Holly A; Timbers, Sarah E; Hudson, Simon C; Hillyer, Lynn L

2010-01-01

The detection of drug abuse in horseracing often requires knowledge of drug metabolism, especially if urine is the matrix of choice. In this study, equine liver/lung microsomes/S9 tissue fractions were used to study the phase I metabolism of eight drugs of relevance to equine drug surveillance (acepromazine, azaperone, celecoxib, fentanyl, fluphenazine, mepivacaine, methylphenidate and tripelennamine). In vitro samples were analyzed qualitatively alongside samples originating from in vivo administrations using LC-MS on a high resolution accurate mass Thermo Orbitrap Discovery instrument and by LC-MS/MS on an Applied Biosystems Sciex 5500 Q Trap.Using high resolution accurate mass full-scan analysis on the Orbitrap, the in vitro systems were found to generate at least the two most abundant phase I metabolites observed in vitro for all eight drugs studied. In the majority of cases, in vitro experiments were also able to generate the minor in vivo metabolites and sometimes metabolites that were only observed in vitro. More detailed analyses of fentanyl incubates using LC-MS/MS showed that it was possible to generate good quality spectra from the metabolites generated in vitro. These data support the suggestion of using in vitro incubates as metabolite reference material in place of in vivo post-administration samples in accordance with new qualitative identification guidelines in the 2009 International Laboratory Accreditation Cooperation-G7 (ILAC-G7) document.In summary, the in vitro and in vivo phase I metabolism results reported herein compare well and demonstrate the potential of in vitro studies to compliment, refine and reduce the existing equine in vivo paradigm. © 2010 John Wiley & Sons, Ltd.

In Vitro and Clinical Evaluations of the Drug-Drug Interaction Potential of a Metabotropic Glutamate 2/3 Receptor Agonist Prodrug with Intestinal Peptide Transporter 1

PubMed Central

Long, Amanda J.; Annes, William F.; Witcher, Jennifer W.; Knadler, Mary Pat; Ayan-Oshodi, Mosun A.; Mitchell, Malcolm I.; Leese, Phillip; Hillgren, Kathleen M.

2017-01-01

Despite peptide transporter 1 (PEPT1) being responsible for the bioavailability for a variety of drugs, there has been little study of its potential involvement in drug-drug interactions. Pomaglumetad methionil, a metabotropic glutamate 2/3 receptor agonist prodrug, utilizes PEPT1 to enhance absorption and bioavailability. In vitro studies were conducted to guide the decision to conduct a clinical drug interaction study and to inform the clinical study design. In vitro investigations determined the prodrug (LY2140023 monohydrate) is a substrate of PEPT1 with Km value of approximately 30 µM, whereas the active moiety (LY404039) is not a PEPT1 substrate. In addition, among the eight known PEPT1 substrates evaluated in vitro, valacyclovir was the most potent inhibitor (IC50 = 0.46 mM) of PEPT1-mediated uptake of the prodrug. Therefore, a clinical drug interaction study was conducted to evaluate the potential interaction between the prodrug and valacyclovir in healthy subjects. No effect of coadministration was observed on the pharmacokinetics of the prodrug, valacyclovir, or either of their active moieties. Although in vitro studies showed potential for the prodrug and valacyclovir interaction via PEPT1, an in vivo study showed no interaction between these two drugs. PEPT1 does not appear to easily saturate because of its high capacity and expression in the intestine. Thus, a clinical interaction at PEPT1 is unlikely even with a compound with high affinity for the transporter. PMID:27895114

In vitro and in vivo studies of Allium sativum extract against deltamethrin-induced oxidative stress in rats brain and kidney.

PubMed

Ncir, Marwa; Saoudi, Mongi; Sellami, Hanen; Rahmouni, Fatma; Lahyani, Amina; Makni Ayadi, Fatma; El Feki, Abdelfattah; Allagui, Mohamed Salah

2017-09-18

The present study investigated the in vitro and the in vivo antioxidant capacities of Allium sativum (garlic) extract against deltamethrin-induced oxidative damage in rat's brain and kidney. The in vitro result showed that highest extraction yield was achieved with methanol (20.08%). Among the tested extracts, the methanol extract exhibited the highest total phenolic, flavonoids contents and antioxidant activity. The in vivo results showed that deltamethrin treatment caused an increase of the acetylcholinesterase level (AChE) in brain and plasma, the brain and kidney conjugated dienes and lipid peroxidation (LPO) levels as compared to control group. The antioxidant enzymes results showed that deltamethrin treatment induced a significantly decrease (p < 0.01) in brain and kidney antioxidant enzymes as catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GPx) to control group. The co-administration of garlic extract reduced the toxic effects in brain and kidney tissues induced by deltamethrin.

In vitro culture thawed human ovarian tissue: NIV versus slow freezing method.

PubMed

Xiao, Zhun; Wang, Yan; Li, Ling-Ling; Li, Shang-wei

2013-01-01

The aim of this study was to determine if the needle immersed vitrification method (NIV) can improve the growth potential of thawed ovarian tissue in vitro culture. Human ovarian cortical tissues were cryopreserved using NIV and slow freezing method. After 14 days of culture, the preservation outcomes of NIV and slow freezing groups were analyzed histologically using light microscope and apoptosis was assessed by TUNEL assay. The result showed that the percentage of morphologically abnormal primordial follicles was lower in NIV group than in slow freezing group (P < 0.05). The incidence of TUNEL-positive primordial follicles was lower in NIV group than in slow freezing group (P < 0.05). The study showed that cryopreservation of human ovarian tissue with NIV was effective in improving the growth potential of frozen-thawed ovarian tissue in vitro culture.

Combining microfluidics, optogenetics and calcium imaging to study neuronal communication in vitro.

PubMed

Renault, Renaud; Sukenik, Nirit; Descroix, Stéphanie; Malaquin, Laurent; Viovy, Jean-Louis; Peyrin, Jean-Michel; Bottani, Samuel; Monceau, Pascal; Moses, Elisha; Vignes, Maéva

2015-01-01

In this paper we report the combination of microfluidics, optogenetics and calcium imaging as a cheap and convenient platform to study synaptic communication between neuronal populations in vitro. We first show that Calcium Orange indicator is compatible in vitro with a commonly used Channelrhodopsine-2 (ChR2) variant, as standard calcium imaging conditions did not alter significantly the activity of transduced cultures of rodent primary neurons. A fast, robust and scalable process for micro-chip fabrication was developed in parallel to build micro-compartmented cultures. Coupling optical fibers to each micro-compartment allowed for the independent control of ChR2 activation in the different populations without crosstalk. By analyzing the post-stimuli activity across the different populations, we finally show how this platform can be used to evaluate quantitatively the effective connectivity between connected neuronal populations.

Preparation of osthole-loaded nano-vesicles for skin delivery: Characterization, in vitro skin permeation and preliminary in vivo pharmacokinetic studies.

PubMed

Meng, Shu; Zhang, Chong; Shi, Wei; Zhang, Xiao-Wei; Liu, Dan-Hua; Wang, Ping; Li, Jian-Xin; Jin, Ying

2016-09-20

The main objective of the present research was to evaluate the flexible nano-vesicles for transdermal delivery of osthole. The nano-vesicles were formulated, characterized and evaluated for their physicochemical properties, in vitro skin permeation and in vivo plasma concentration. The encapsulation efficiency of osthole in ethosome and transfersome was measured to be 83.3±4.8% and 80.9±3.6%, respectively. In vitro studies showed that osthole ethosome provided an enhanced transdermal flux of 6.98±1.6μg/cm(2)/h and a decreased lag time of 2.45h across porcine ear skin. Moreover, ethosome also showed increased skin deposition of the drug over transfersome (1.5-fold) and saturated solution of osthole in 35% ethanol (2.1-fold). Data from in vivo pharmacokinetic studies showed that AUC and Cmax of the osthole loaded-ethosome were remarkably increasing compared with the other formulations. Thus, osthole ethosome was considered as an effective delivery system for the drug. Copyright © 2016 Elsevier B.V. All rights reserved.

In vitro and in vivo evaluation of sanguinarine liposomes prepared by a remote loading method with three different ammonium salts.

PubMed

Ke, X; Bei, J H; Zhang, Y; Li, J

2011-04-01

Sanguinarine liposomes were prepared by a remote loading method using three different ammonium salts. A series of studies, including in vitro release, in vitro and in vivo anti-tumor effects and pharmacokinetics in rats, were conducted. The three liposomes showed pH-sensitive release characteristics in vitro, but there were obvious variations in their release profiles. Among the three liposomes, the liposomes made using ammonium citrate and phosphate possessed better anti-tumor activity in vitro and in vivo, compared with the liposome using ammonium sulfate. Pharmacokinetics test results in rats indicated that sanguinarine liposomes have notably elevated AUC (P<0.05) and markedly lower CL (P<0.05) compared with the solution, but there were no obvious differences between the three liposomes. The present study may be useful for better understanding and better choice of a suitable ammonium salt for the remote loading method.

Polymorphism and methylation patterns in Agave tequilana Weber var. 'Azul' plants propagated asexually by three different methods.

PubMed

Díaz-Martínez, Miriam; Nava-Cedillo, Alejandro; Guzmán-López, José Alfredo; Escobar-Guzmán, Rocío; Simpson, June

2012-04-01

Genetic variation in three forms of asexually propagated Agave tequilana Weber var. 'Azul' plants namely offsets, bulbils and in vitro cultured individuals was studied by AFLP analysis. Low levels of variation were observed between mother plants and offsets and a higher level between mother plant and bulbils. Families obtained from commercial plantations showed lower levels of variation in comparison to families grown as ornamentals. No variation was observed between the original explant and four generations of in vitro cultured plants. Epigenetic variation was also studied by analyzing changes in methylation patterns between mother plants and offspring in each form of asexual reproduction. Offsets and bulbils showed an overall decrease in methylation whereas in vitro cultured plants showed patterns specific to each generation: Generations 1 and 4 showed overall demethylation whereas Generations 2 and 3 showed increased methylation. Analysis of ESTs associated with transposable elements revealed higher proportions of ESTs from Ty1-copia-like, Gypsy and CACTA transposable elements in cDNA libraries obtained from pluripotent tissue suggesting a possible correlation between methylation patterns, expression of transposable element associated genes and somaclonal variation. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Polycaprolactone nanofibres loaded with 20(S)-protopanaxadiol for in vitro and in vivo anti-tumour activity study

PubMed Central

Liu, Dan-qing; Cheng, Zhi-qiang; Feng, Qing-jie; Li, He-jie; Ye, Shu-feng

2018-01-01

In this work, 20(S)-protopanaxadiol (PPD)-loaded polycaprolactone (PCL) nanofibres were successfully fabricated by the electrospinning technique using Tween 80 as a solubilizer. Firstly, smooth and continuous nanofibres were collected using suitable solvents and appropriate spinning conditions. Secondly, nanofibre mats were characterized by scanning electron microscopy, thermogravimetric (TG) analysis, Fourier transform infrared spectroscopy and mechanical testing. Finally, nanofibrous membranes were evaluated using water contact angle, in vitro drug release, biodegradation test, in vitro and in vivo anti-tumour activity and cell apoptosis assay. Scanning electron microscopic observations indicated that the diameter of the drug-loaded nanofibres increased with the increase of drug concentration. TG analysis and mechanical test showed that nanofibres were equipped with great thermal and mechanical properties. Biodegradation test exhibited that the structure of fabricated nanofibres had a certain degree of change after 15 days. An in vitro release study showed that PPD from drug-loaded nanofibres could be released in a sustained and prolonged mode. The cytotoxic effect of drug-loaded nanofibre mats examined on human laryngeal carcinoma cells (Hep-2 cells) demonstrated that the prepared nanofibres had a remarkable anti-tumour effect. Meanwhile, the drug-loaded fibre mats showed a super anti-tumour effect in an in vivo anti-tumour study. All in all, PCL nanofibres could be a potential carrier of PPD for cancer treatment. PMID:29892448

Vorinostat-eluting poly(DL-lactide-co-glycolide) nanofiber-coated stent for inhibition of cholangiocarcinoma cells

PubMed Central

Song, Yeon Hui; Kim, Chan; Kim, Jungsoo; Seo, Sol-Ji; Jeong, Young-Il; Kang, Dae Hwan

2017-01-01

Purpose The aim of this study was to fabricate a vorinostat (Zolinza™)-eluting nanofiber membrane-coated gastrointestinal (GI) stent and to study its antitumor activity against cholangiocarcinoma (CCA) cells in vitro and in vivo. Methods Vorinostat and poly(DL-lactide-co-glycolide) dissolved in an organic solvent was sprayed onto a GI stent to make a nanofiber-coated stent using an electro-spinning machine. Intact vorinostat and vorinostat released from nanofibers was used to assess anticancer activity in vitro against various CCA cells. The antitumor activity of the vorinostat-eluting nanofiber membrane-coated stent was evaluated using HuCC-T1 bearing mice. Results A vorinostat-incorporated polymer nanofiber membrane was formed on the surface of the GI stent. Vorinostat was continuously released from the nanofiber membrane over 10 days, and its release rate was higher in cell culture media than in phosphate-buffered saline. Released vorinostat showed similar anticancer activity against various CCA cells in vitro compared to that of vorinostat. Like vorinostat, vorinostat released from nanofibers induced acetylation of histone H4 and inhibited histone deacetylases 1⋅3⋅4/5/7 expression in vitro and in vivo. Furthermore, vorinostat nanofibers showed a higher tumor growth inhibition rate in HuCC-T1 bearing mice than vorinostat injections. Conclusion Vorinostat-eluting nanofiber membranes showed significant antitumor activity against CCA cells in vitro and in vivo. We suggest the vorinostat nanofiber-coated stent may be a promising candidate for CCA treatment. PMID:29089762

New Alcohol and Onyx Mixture for Embolization: Feasibility and Proof of Concept in Both In Vitro and In Vivo Models

DOE Office of Scientific and Technical Information (OSTI.GOV)

Saeed Kilani, Mohammad, E-mail: msaeedkilani@gmail.com, E-mail: mohammadalikilani@yahoo.com; Zehtabi, Fatemeh, E-mail: fatemeh.zehtabi@gmail.com; Lerouge, Sophie, E-mail: Sophie.Lerouge@etsmtl.ca

IntroductionOnyx and ethanol are well-known embolic and sclerotic agents that are frequently used in embolization. These agents present advantages and disadvantages regarding visibility, injection control and penetration depth. Mixing both products might yield a new product with different characteristics. The aim of this study is to evaluate the injectability, radiopacity, and mechanical and occlusive properties of different mixtures of Onyx 18 and ethanol in vitro and in vivo (in a swine model).Materials and MethodsVarious Onyx 18 and ethanol formulations were prepared and tested in vitro for their injectability, solidification rate and shrinkage, cohesion and occlusive properties. In vivo tests weremore » performed using 3 swine. Ease of injection, radiopacity, cohesiveness and penetration were analyzed using fluoroscopy and high-resolution CT.ResultsAll mixtures were easy to inject through a microcatheter with no resistance or blockage in vitro and in vivo. The 50%-ethanol mixture showed delayed copolymerization with fragmentation and proximal occlusion. The 75%-ethanol mixture showed poor radiopacity in vivo and was not tested in vitro. The 25%-ethanol mixture showed good occlusive properties and accepted penetration and radiopacity.ConclusionMixing Onyx and ethanol is feasible. The mixture of 25% of ethanol and 75% of Onyx 18 could be a new sclero-embolic agent. Further research is needed to study the chemical changes of the mixture, to confirm the significance of the added sclerotic effect and to find out the ideal mixture percentages.« less

New Alcohol and Onyx Mixture for Embolization: Feasibility and Proof of Concept in Both In Vitro and In Vivo Models.

PubMed

Saeed Kilani, Mohammad; Zehtabi, Fatemeh; Lerouge, Sophie; Soulez, Gilles; Bartoli, Jean Michel; Vidal, Vincent; Badran, Mohammad F

2017-05-01

Onyx and ethanol are well-known embolic and sclerotic agents that are frequently used in embolization. These agents present advantages and disadvantages regarding visibility, injection control and penetration depth. Mixing both products might yield a new product with different characteristics. The aim of this study is to evaluate the injectability, radiopacity, and mechanical and occlusive properties of different mixtures of Onyx 18 and ethanol in vitro and in vivo (in a swine model). Various Onyx 18 and ethanol formulations were prepared and tested in vitro for their injectability, solidification rate and shrinkage, cohesion and occlusive properties. In vivo tests were performed using 3 swine. Ease of injection, radiopacity, cohesiveness and penetration were analyzed using fluoroscopy and high-resolution CT. All mixtures were easy to inject through a microcatheter with no resistance or blockage in vitro and in vivo. The 50%-ethanol mixture showed delayed copolymerization with fragmentation and proximal occlusion. The 75%-ethanol mixture showed poor radiopacity in vivo and was not tested in vitro. The 25%-ethanol mixture showed good occlusive properties and accepted penetration and radiopacity. Mixing Onyx and ethanol is feasible. The mixture of 25% of ethanol and 75% of Onyx 18 could be a new sclero-embolic agent. Further research is needed to study the chemical changes of the mixture, to confirm the significance of the added sclerotic effect and to find out the ideal mixture percentages.

Ectopic bone formation during tissue-engineered cartilage repair using autologous chondrocytes and novel plasma-derived albumin scaffolds.

PubMed

Robla Costales, David; Junquera, Luis; García Pérez, Eva; Gómez Llames, Sara; Álvarez-Viejo, María; Meana-Infiesta, Álvaro

2016-10-01

The aims of this study were twofold: first, to evaluate the production of cartilaginous tissue in vitro and in vivo using a novel plasma-derived scaffold, and second, to test the repair of experimental defects made on ears of New Zealand rabbits (NZr) using this approach. Scaffolds were seeded with chondrocytes and cultured in vitro for 3 months to check in vitro cartilage production. To evaluate in vivo cartilage production, a chondrocyte-seeded scaffold was transplanted subcutaneously to a nude mouse. To check in vivo repair, experimental defects made in the ears of five New Zealand rabbits (NZr) were filled with chondrocyte-seeded scaffolds. In vitro culture produced mature chondrocytes with no extracellular matrix (ECM). Histological examination of redifferentiated in vitro cultures showed differentiated chondrocytes adhered to scaffold pores. Subcutaneous transplantation of these constructs to a nude mouse produced cartilage, confirmed by histological study. Experimental cartilage repair in five NZr showed cartilaginous tissue repairing the defects, mixed with calcified areas of bone formation. It is possible to produce cartilaginous tissue in vivo and to repair experimental auricular defects by means of chondrocyte cultures and the novel plasma-derived scaffold. Further studies are needed to determine the significance of bone formation in the samples. Copyright © 2016 European Association for Cranio-Maxillo-Facial Surgery. Published by Elsevier Ltd. All rights reserved.

In vitro anti-tubulin effects of mebendazole and fenbendazole on canine glioma cells.

PubMed

Lai, S R; Castello, S A; Robinson, A C; Koehler, J W

2017-12-01

Benzimidazole anthelmintics have reported anti-neoplastic effects both in vitro and in vivo. The purpose of this study was to evaluate the in vitro chemosensitivity of three canine glioma cell lines to mebendazole and fenbendazole. The mean inhibitory concentration (IC 50 ) (±SD) obtained from performing the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay after treating J3T, G06-A, and SDT-3G cells for 72 h with mebendazole were 0.030 ± 0.003, 0.080 ± 0.015 and 0.030 ± 0.006 μM respectively, while those for fenbendazole were 0.550  ± 0.015, 1.530 ± 0.159 and 0.690 ± 0.095 μM; treatment of primary canine fibroblasts for 72 h at IC 50 showed no significant effect. Immunofluorescence studies showed disruption of tubulin after treatment. Mebendazole and fenbendazole are cytotoxic in canine glioma cell lines in vitro and may be good candidates for treatment of canine gliomas. Further in vivo studies are required. © 2017 John Wiley & Sons Ltd.

Three-dimensional printing of Hela cells for cervical tumor model in vitro.

PubMed

Zhao, Yu; Yao, Rui; Ouyang, Liliang; Ding, Hongxu; Zhang, Ting; Zhang, Kaitai; Cheng, Shujun; Sun, Wei

2014-09-01

Advances in three-dimensional (3D) printing have enabled the direct assembly of cells and extracellular matrix materials to form in vitro cellular models for 3D biology, the study of disease pathogenesis and new drug discovery. In this study, we report a method of 3D printing for Hela cells and gelatin/alginate/fibrinogen hydrogels to construct in vitro cervical tumor models. Cell proliferation, matrix metalloproteinase (MMP) protein expression and chemoresistance were measured in the printed 3D cervical tumor models and compared with conventional 2D planar culture models. Over 90% cell viability was observed using the defined printing process. Comparisons of 3D and 2D results revealed that Hela cells showed a higher proliferation rate in the printed 3D environment and tended to form cellular spheroids, but formed monolayer cell sheets in 2D culture. Hela cells in 3D printed models also showed higher MMP protein expression and higher chemoresistance than those in 2D culture. These new biological characteristics from the printed 3D tumor models in vitro as well as the novel 3D cell printing technology may help the evolution of 3D cancer study.

In vitro tuberization of Chlorophytum Borivilianum Sant & Fern (Safed musli) as influenced by sucrose, CCC and culture systems.

PubMed

Farshad Ashraf, Mehdi; Abd Aziz, Maheran; Abdul Kadir, Mihdzar; Stanslas, Johnson; Farokhian, Elmira

2013-08-01

This study focuses on the establishment of in vitro tuberization of Chlorophytum borivilianum using solid and liquid culture systems. A high in vitro tuberization rate on solid and stationary liquid Murashige and Skoog media was observed in the presence of 60 g l⁻¹ sucrose with 950, 1,265 and 1,580 µM 2-chloroethyl-trimethylammonium chloride (CCC). Application of a higher sucrose concentration of 90 g l⁻¹ showed a negative interaction with CCC on in vitro tuber number and days to in vitro tuber induction. For economic feasibility, 950 µM CCC with 60 g l⁻¹ sucrose was chosen as the best combination for in vitro tuberization in both solid and stationary liquid media. For optimization of in vitro tuber production,a comparison between solid, stationary liquid and shake liquid culture was carried out. Liquid culture with shaking at 80 r.p.m. resulted in a >2.5-fold increase in in vitro tuber production compared with solid culture.

EFFECT OF X RADIATION ON THE AMOUNT OF PROPERDINE SERUM IN THE RAT (in French)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Verain, A.; Despaux, E.; Verain, A.

1958-01-01

The effect of x radiation on the amount of properdine in rat serum was studied in vivo and in vitro. White rats were submitted to 1000 r, and the amount of properdine before and after irradiation was determined. Serum in vitro was irradiatcd with 1 to 2 Mr. The results showed a rapid, almost constant, deerease of the properdine in the irradiated rat. This effect was found in vitro only when large radiation doses were used. (J.S.R.)

Composition and structural features of condensed tannins from Texas legumes exhibiting methane abatement activity during in vitro rumen digestion

USDA-ARS?s Scientific Manuscript database

Previous studies showed that a series of purified condensed tannins (CTs) from warm-season perennial legumes exhibited high variability in their modulation of methane production during in vitro rumen digestion. The molecular weight difference of these CTs did not provide correlation with either the ...

Effect of Citrus bergamia juice on human neuroblastoma cells in vitro and in metastatic xenograft models.

PubMed

Navarra, M; Ursino, M R; Ferlazzo, N; Russo, M; Schumacher, U; Valentiner, U

2014-06-01

Neuroblastoma is the most common extracranial pediatric solid tumor with poor prognosis in children with disseminated stage of disease. A number of studies show that molecules largely distributed in commonly consumed fruits and vegetables may have anti-tumor activity. In this study we evaluate the effect of Citrus bergamia (bergamot) juice (BJ) in vitro and in a spontaneous metastatic neuroblastoma SCID mouse model. Qualitative and quantitative characterizations of BJ flavonoid fractions were performed by RP-HPLC/PDA/MS. We show that BJ significantly affects SK-N-SH and LAN-1 cell proliferation in vitro, but fails to reduce primary tumor weight in vivo. Moreover, BJ reduced cell adhesiveness and invasion of LAN-1 and SK-N-SH cells in vitro and the number of pulmonary metastases under consideration of the number of tumor cells in the blood in mice inoculated with LAN-1 cells in vivo. These effects without any apparent sign of systemic toxicity confirm the potential clinical interest of BJ and lay the basis for further investigation in cancer. Copyright © 2014. Published by Elsevier B.V.

In vitro digestibility of goat milk and kefir with a new standardised static digestion method (INFOGEST cost action) and bioactivities of the resultant peptides.

PubMed

Nehir El, Sedef; Karakaya, Sibel; Simsek, Sebnem; Dupont, Didier; Menfaatli, Esra; Eker, Alper Tolga

2015-07-01

The hydrolysis degrees of goat milk and kefir during simulated gastrointestinal digestion and some bioactivities of the resulting peptides after fermentation and digestion were studied. A static in vitro digestion method by the COST FA1005 Action INFOGEST was used and goat milk and kefir were partially hydrolyzed during the gastric phase and had above 80% hydrolysis after duodenal digestion. There were no differences between the digestibility of goat milk and kefir (p > 0.05). Goat milk and kefir displayed about 7-fold antioxidant activity after digestion (p < 0.05). Fermentation showed no effect on the calcium-binding capacity of the samples (p > 0.05), however, after in vitro digestion calcium-binding capacity of the goat milk and kefir increased 2 and 5 fold, respectively (p < 0.05). Digested goat milk and kefir showed a higher dose-dependent inhibitory effect on α-amylase compared to undigested samples (p < 0.05). α-Glucosidase inhibitory activities and in vitro bile acid-binding capacities of the samples were not determined at the studied concentrations.

Patient-specific in vitro models for hemodynamic analysis of congenital heart disease - Additive manufacturing approach.

PubMed

Medero, Rafael; García-Rodríguez, Sylvana; François, Christopher J; Roldán-Alzate, Alejandro

2017-03-21

Non-invasive hemodynamic assessment of total cavopulmonary connection (TCPC) is challenging due to the complex anatomy. Additive manufacturing (AM) is a suitable alternative for creating patient-specific in vitro models for flow measurements using four-dimensional (4D) Flow MRI. These in vitro systems have the potential to serve as validation for computational fluid dynamics (CFD), simulating different physiological conditions. This study investigated three different AM technologies, stereolithography (SLA), selective laser sintering (SLS) and fused deposition modeling (FDM), to determine differences in hemodynamics when measuring flow using 4D Flow MRI. The models were created using patient-specific MRI data from an extracardiac TCPC. These models were connected to a perfusion pump circulating water at three different flow rates. Data was processed for visualization and quantification of velocity, flow distribution, vorticity and kinetic energy. These results were compared between each model. In addition, the flow distribution obtained in vitro was compared to in vivo. The results showed significant difference in velocities measured at the outlets of the models that required internal support material when printing. Furthermore, an ultrasound flow sensor was used to validate flow measurements at the inlets and outlets of the in vitro models. These results were highly correlated to those measured with 4D Flow MRI. This study showed that commercially available AM technologies can be used to create patient-specific vascular models for in vitro hemodynamic studies at reasonable costs. However, technologies that do not require internal supports during manufacturing allow smoother internal surfaces, which makes them better suited for flow analyses. Copyright © 2017 Elsevier Ltd. All rights reserved.

Development and In Vitro Characterization of a Gemcitabine-loaded MUC4-targeted Immunoliposome Against Pancreatic Ductal Adenocarcinoma.

PubMed

Urey, Carlos; Hilmersson, Katarzyma Said; Andersson, Bodil; Ansari, Daniel; Andersson, Roland

2017-11-01

Pancreatic Ductal adeno-carcinoma (PDAC) is a devastating disease. Gemcitabine is the standard chemotherapeutic agent against PDAC but has only limited effectiveness. The aim of the study was to develop and study the targeting affinity and in vitro antiproliferative effect of a MUC4-targeted gemcitabine-loaded immuno-liposome for treatment of PDAC. Gemcitabine-loaded immunoliposomes were developed by grafting anti-MUC4 antibodies to the liposomal surface. Targeting affinity was compared in vitro between immunoliposomes and non-targeted liposomes and anti-proliferative effect was compared in vitro between free drug, non-targeted liposomal gemcitabine and MUC4-targeted immunoliposomal gemcitabine on a MUC4-positive pancreatic cancer cell line, Capan-1. Development of a MUC4-targeted immunoliposome was confirmed and characterized by immunoblots and size characterization. The MUC4-targeted immunoliposome showed a significantly higher targeting affinity compared to the non-targeted liposomes and also showed an improved antiproliferative effect compared to free and non-targeted liposomal drug. Successful development and characterization of a MUC4-targeted immunoliposome shows promising results for a targeted treatment and improved retention of gemcitabine for treatment of PDAC. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Silibinin and indocyanine green-loaded nanoparticles inhibit the growth and metastasis of mammalian breast cancer cells in vitro.

PubMed

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

2016-07-01

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. 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. 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. SIPNs show temperature-sensitive drug release following NIR laser irradiation, which can inhibit the growth and metastasis of breast cancer cells in vitro.

Use of an in vitro human skin permeation assay to assess bioequivalence of two topical cream formulations containing butenafine hydrochloride (1%, w/w).

PubMed

Mitra, Amitava; Kim, Nanhye; Spark, Darren; Toner, Frank; Craig, Susan; Roper, Clive; Meyer, Thomas A

2016-12-01

The primary objective of this work was to investigate, using an in vitro human skin permeation study, whether changes in the excipients of butenafine hydrochloride cream would have any effect on bioperformance of the formulation. Such in vitro data would be a surrogate for any requirement of a bioequivalence (BE) study to demonstrate formulation similarity. A LC-MS/MS method for quantitation of butenafine in various matrices was developed and validated. A pilot study was performed to validate the in vitro skin permeation methodology using three cream formulations containing butenafine hydrochloride at concentrations of 0.5, 1.0 and 1.5% (w/w). Finally, a definitive in vitro human skin permeation study was conducted, comparing the extent of butenafine hydrochloride permeation from the new formulation to that from the current formulation. The results of the study comparing the two formulations showed that there was no statistically significant difference in the extent of butenafine permeation into human skin. In conclusion, these in vitro data demonstrated that the formulation change is likely to have no significant impact on the bioperformance of 1% (w/w) butenafine hydrochloride cream. Copyright © 2016 Elsevier Inc. All rights reserved.

Physiologically based pharmacokinetics model predicts the lack of inhibition by repaglinide on the metabolism of pioglitazone.

PubMed

Xiao, Qingqing; Tang, Liling; Xu, Ruijuan; Qian, Wei; Yang, Jin

2015-12-01

Repaglinide and pioglitazone are both CYP2C8 and CYP3A4 substrates. This study was to determine whether repaglinide has an inhibitory effect on the metabolism of pioglitazone in vitro, in silico and in vivo. In vitro, the effect of repaglinide on the metabolism of pioglitazone was assessed in pooled human liver microsomes. In silico, an IVIVE-PBPK linked model was built with Simcyp®. Then, a randomized, 2-phase cross-over clinical study was conducted in 12 healthy volunteers. Repaglinide showed a strong inhibitory effect on the metabolism of pioglitazone in vitro (Ki  = 0.0757 µm), [I]/Ki  > 0.1. The Simcyp® prediction ratios of AUC and Cmax between the two treatment groups were both about 1.01. The pharmacokinetics of pioglitazone in clinical trials showed no significant difference between these two treatment groups (p > 0.05). Repaglinide has no significant inhibitory effect on the metabolism of pioglitazone in vivo, which is inconsistent with the in vitro results. The lack of an inhibitory effect was partly due to extensive plasma protein binding and to the high in vivo clearance of repaglinide, for the concentration of repaglinide in vivo was far smaller than in vitro. Copyright © 2015 John Wiley & Sons, Ltd.

Catecholamines and in vitro growth of pathogenic bacteria: enhancement of growth varies greatly among bacterial species

NASA Technical Reports Server (NTRS)

Belay, Tesfaye; Aviles, Hernan; Vance, Monique; Fountain, Kimberly; Sonnenfeld, Gerald

2003-01-01

The purpose of this study was to examine the effects of catecholamines on in vitro growth of a range of bacterial species, including anaerobes. Bacteria tested included: Porphyromonas gingivalis, Bacteriodes fragilis, Shigella boydii, Shigella sonnie, Enterobacter Sp, and Salmonella choleraesuis. The results of the current study indicated that supplementation of bacterial cultures in minimal medium with norepinephrine or epinephrine did not result in increased growth of bacteria. Positive controls involving treatment of Escherichia coli with catecholamines did result in increased growth of that bacterial species. The results of the present study extend previous observations that showed differential capability of catecholamines to enhance bacterial growth in vitro.

Anticancer effects of deproteinized asparagus polysaccharide on hepatocellular carcinoma in vitro and in vivo.

PubMed

Xiang, Jianfeng; Xiang, Yanjie; Lin, Shengming; Xin, Dongwei; Liu, Xiaoyu; Weng, Lingling; Chen, Tao; Zhang, Minguang

2014-04-01

Hepatocellular carcinoma (HCC) is one of the most aggressive malignancies in the world whose chemoprevention became increasingly important in HCC treatment. Although the anticancer effects of asparagus constituents have been investigated in several cancers, its effects on hepatocellular carcinoma have not been fully studied. In this study, we investigated the anticancer effects of the deproteinized asparagus polysaccharide on the hepatocellular carcinoma cells using the in vitro and in vivo experimental model. Our data showed that deproteinized asparagus polysaccharide might act as an effective inhibitor on cell growth in vitro and in vivo and exert potent selective cytotoxicity against human hepatocellular carcinoma Hep3B and HepG2 cells. Further study showed that it could potently induce cell apoptosis and G2/M cell cycle arrest in the more sensitive Hep3B and HepG2 cell lines. Moreover, deproteinized asparagus polysaccharide potentiated the effects of mitomycin both in vitro and in vivo. Mechanistic studies revealed that deproteinized asparagus polysaccharide might exert its activity through an apoptosis-associated pathway by modulating the expression of Bax, Bcl-2, and caspase-3. In conclusion, deproteinized asparagus polysaccharide exhibited significant anticancer activity against hepatocellular carcinoma cells and could sensitize the tumoricidal effects of mitomycin, indicating that it is a potential therapeutic agent (or chemosensitizer) for liver cancer therapy.

Mucoadhesive buccal patches based on interpolymer complexes of chitosan–pectin for delivery of carvedilol

PubMed Central

Kaur, Amanpreet; Kaur, Gurpreet

2011-01-01

The study was designed to develop bioadhesive patches of carvedilol hydrochloride using chitosan (CH) and pectin (PE) interpolymer complexes and to systematically evaluate their in vitro and in vivo performances. Mucoadhesive buccal patches of carvedilol were prepared using solvent casting method. The physicochemical interaction between CH and PE was investigated by FTIR and DSC studies. The patches were evaluated for their physical characteristics like mass variation, content uniformity, folding endurance, ex vivo mucoadhesion strength, ex vivo mucoadhesion time, surface pH, in vitro drug release, in situ release study, and in vivo bioavailability study. The swelling index of the patches was found to be proportional to the PE concentration. The surface pH of all the formulated bioadhesive patches was found to lie between 6.2 and 7.2. The optimized bioadhesive patch (C1, CH:PE 20:80) showed bioadhesive strength of 22.10 ± 0.20 g, in vitro release of 98.73% and ex vivo mucoadhesion time of 451 min with in a period of 8 h. The optimized patch demonstrated good in vitro and in vivo results. The buccal delivery of carvedilol in rabbits showed a significant improvement in bioavailability of carvedilol from patches when compared to oral route. PMID:23960773

Mucoadhesive buccal patches based on interpolymer complexes of chitosan-pectin for delivery of carvedilol.

PubMed

Kaur, Amanpreet; Kaur, Gurpreet

2012-01-01

The study was designed to develop bioadhesive patches of carvedilol hydrochloride using chitosan (CH) and pectin (PE) interpolymer complexes and to systematically evaluate their in vitro and in vivo performances. Mucoadhesive buccal patches of carvedilol were prepared using solvent casting method. The physicochemical interaction between CH and PE was investigated by FTIR and DSC studies. The patches were evaluated for their physical characteristics like mass variation, content uniformity, folding endurance, ex vivo mucoadhesion strength, ex vivo mucoadhesion time, surface pH, in vitro drug release, in situ release study, and in vivo bioavailability study. The swelling index of the patches was found to be proportional to the PE concentration. The surface pH of all the formulated bioadhesive patches was found to lie between 6.2 and 7.2. The optimized bioadhesive patch (C1, CH:PE 20:80) showed bioadhesive strength of 22.10 ± 0.20 g, in vitro release of 98.73% and ex vivo mucoadhesion time of 451 min with in a period of 8 h. The optimized patch demonstrated good in vitro and in vivo results. The buccal delivery of carvedilol in rabbits showed a significant improvement in bioavailability of carvedilol from patches when compared to oral route.

Evaluation of trimethoprim-sulfamethoxazole based combination therapy against Stenotrophomonas maltophilia: in vitro effects and clinical efficacy in cancer patients.

PubMed

Araoka, Hideki; Baba, Masaru; Okada, Chikako; Abe, Masahiro; Kimura, Muneyoshi; Yoneyama, Akiko

2017-05-01

The aim of this study was to evaluate the in vitro effects and clinical efficacies of trimethoprim-sulfamethoxazole (SXT) combined with other antimicrobial agents against Stenotrophomonas maltophilia. In vitro analysis was conducted on 89 S. maltophilia strains isolated from blood and the respiratory tract between June 2012 and October 2014. Levofloxacin (LVX), ticarcillin-clavulanic acid (TIM), and minocycline (MIN) were selected for an examination of their effects when individually combined with SXT by the checkerboard method. In addition, 29 S. maltophilia bacteremia cases were reviewed and the clinical efficacies of SXT-based combination therapies were analyzed. SXT+LVX showed synergy in 21, no interactions in 61, and antagonism in 7. SXT+TIM showed synergy in 71, and no interactions in 18. SXT+MIN showed synergy in 10, and no interactions in 79. The review of clinical data indicated that a combination of SXT+fluoroquinolone was not associated with improved prognosis compared with monotherapy. The in vitro data indicated that SXT+TIM had beneficial microbiological effects and was not antagonistic. Our in vitro and clinical data analyses do not support the routine use of SXT+fluoroquinolone combination therapy for S. maltophilia infection. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Differential Inhibitory Activities of Four Plant Essential Oils on In Vitro Growth of Fusarium oxysporum f. sp. fragariae Causing Fusarium Wilt in Strawberry Plants.

PubMed

Park, Jin Young; Kim, Su Hyeon; Kim, Na Hee; Lee, Sang Woo; Jeun, Yong-Chull; Hong, Jeum Kyu

2017-12-01

The objective of this study was to determine inhibitory activities of four volatile plant essential oils (cinnamon oil, fennel oil, origanum oil and thyme oil) on in vitro growth of Fusarium oxysporum f. sp. fragariae causing Fusarium wilt of strawberry plants. Results showed that these essential oils inhibited in vitro conidial germination and mycelial growth of F. oxysporum f. sp. fragariae in a dose-dependent manner. Cinnamon oil was found to be most effective one in suppressing conidial germination while fennel oil, origanum oil and thyme oil showed moderate inhibition of conidial germination at similar levels. Cinnamon oil, origanum oil and thyme oil showed moderate antifungal activities against mycelial growth at similar levels while fennel oil had relatively lower antifungal activity against mycelial growth. Antifungal effects of these four plant essential oils in different combinations on in vitro fungal growth were also evaluated. These essential oils demonstrated synergistic antifungal activities against conidial germination and mycelial growth of F. oxysporum f. sp. fragariae in vitro. Simultaneous application of origanum oil and thyme oil enhanced their antimicrobial activities against conidial germination and fungal mycelial growth. These results underpin that volatile plant essential oils could be used in eco-friendly integrated disease management of Fusarium wilt in strawberry fields.

Preparation and evaluation of sustained release loxoprofen loaded microspheres.

PubMed

Venkatesan, P; Manavalan, R; Valliappan, K

2011-06-01

The aim of present study was to formulate and evaluate the loxoprofen loaded Sustained release microspheres by emulsion solvent evaporation technique. Ethylcellulose, a biocompatible polymer is used as the retardant material. The effects of process conditions such as drug loading, polymer type and solvent type on the characteristics of microspheres were investigated. The prepared microspheres were characterized for their particle size and drug loading and drug release. The in-vitro release studies were carried out in phosphate buffer at pH 7.4. The prepared microspheres were white, free flowing and spherical in shape. The drug-loaded microspheres showed 71.2% of entrapment and the in-vitro release studies showed that Loxoprofen microspheres of 1:3 ratios showed better sustained effect over a period of 8 hours.

Preparation and evaluation of sustained release loxoprofen loaded microspheres

PubMed Central

Venkatesan, P.; Manavalan, R.; Valliappan, K.

2011-01-01

The aim of present study was to formulate and evaluate the loxoprofen loaded Sustained release microspheres by emulsion solvent evaporation technique. Ethylcellulose, a biocompatible polymer is used as the retardant material. The effects of process conditions such as drug loading, polymer type and solvent type on the characteristics of microspheres were investigated. The prepared microspheres were characterized for their particle size and drug loading and drug release. The in-vitro release studies were carried out in phosphate buffer at pH 7.4. The prepared microspheres were white, free flowing and spherical in shape. The drug-loaded microspheres showed 71.2% of entrapment and the in-vitro release studies showed that Loxoprofen microspheres of 1:3 ratios showed better sustained effect over a period of 8 hours PMID:24826017

Antioxidative and antihypertensive activities of pig meat before and after cooking and in vitro gastrointestinal digestion: Comparison between Italian autochthonous pig Suino Nero Lucano and a modern crossbred pig.

PubMed

Simonetti, Amalia; Gambacorta, Emilio; Perna, Annamaria

2016-12-01

The aim of this study was to evaluate and compare antioxidative and antihypertensive activities of Longissimus dorsi muscle from Suino Nero Lucano (SNL) and a modern crossbred (CG) pigs, before and after cooking and in vitro gastrointestinal digestion. Pig meat showed antioxidative and antihypertensive activities, heat treatment decreased the thiols content but at the same time increased angiotensin I-converting enzyme (ACE) inhibitory activity, and in vitro gastrointestinal digestion enhanced the biological activity of meat. Autochthonous SNL meat showed a higher nutraceutical quality compared to CG meat, highlighting a greater potential beneficial physiological effect on human health. The results of this study indicate that the pig meat, in particular autochthonous pig meat, may be considered a functional food since it is a good source of antioxidative and antihypertensive peptides. Copyright © 2016 Elsevier Ltd. All rights reserved.

In vitro antioxidant/prooxidant effects of combined use of flavonoids.

PubMed

Eren-Guzelgun, B; Ince, E; Gurer-Orhan, H

2018-06-01

The present study was undertaken to investigate the individual and combined antioxidant or prooxidant effects of genistein, daidzein and quercetin in human erythrocytes and rat microsomes in vitro. Their reducing potential against oxidation of a redox sensitive fluorescent probe, their protective effect against H 2 O 2 -induced membrane lipid peroxidation and their inhibitory effect on AAPH-induced hemolysis were evaluated. Genistein and daidzein were prooxidant in erythrocytes but antioxidant in microsomes where their metabolites might have been formed which suggests the importance of metabolic capacity in in vitro models to predict the physiological situation. Quercetin showed antioxidant effects in all models and conditions. Prooxidant effect of 'genistein-daidzein mixture', at their concentrations reflecting the real life, was suppressed by addition of quercetin to the mixture. Our study shows that flavonoids can exert prooxidant effects depending on the conditions, but the mixture effect should be considered while assessing their effects and safety in humans.

Haemocytes from Crassostrea gigas and OsHV-1: A promising in vitro system to study host/virus interactions.

PubMed

Morga, Benjamin; Faury, Nicole; Guesdon, Stéphane; Chollet, Bruno; Renault, Tristan

2017-11-01

Since 2008, mass mortality outbreaks associated with the detection of particular variants of OsHV-1 have been reported in Crassostrea gigas spat and juveniles in several countries. Recent studies have reported information on viral replication during experimental infection. Viral DNA and RNA were also detected in the haemolymph and haemocytes suggesting that the virus could circulate through the circulatory system. However, it is unknown if the virus is free in the haemolymph, passively associated at the surface of haemocytes, or able to infect and replicate inside these cells inducing (or not) virion production. In the present study, we collected haemocytes from the haemolymphatic sinus of the adductor muscle of healthy C. gigas spat and exposed them in vitro to a viral suspension. Results showed that viral RNAs were detectable one hour after contact and the number of virus transcripts increased over time in association with an increase of viral DNA detection. These results suggested that the virus is able to initiate replication rapidly inside haemocytes maintained in vitro. These in vitro trials were also used to carry out a dual transcriptomic study. We analyzed concomitantly the expression of some host immune genes and 15 viral genes. Results showed an up regulation of oyster genes currently studied during OsHV-1 infection. Additionally, transmission electron microscopy examination was carried out and did not allow the detection of viral particles. Moreover, All the results suggested that the in vitro model using haemocytes can be valuable for providing new perspective on virus-oyster interactions. Copyright © 2017 Elsevier Inc. All rights reserved.

In vitro experimental environments lacking or containing soil disparately affect competition experiments of Aspergillus flavus and co-occurring fungi in maize grains.

PubMed

Falade, Titilayo D O; Syed Mohdhamdan, Sharifah H; Sultanbawa, Yasmina; Fletcher, Mary T; Harvey, Jagger J W; Chaliha, Mridusmita; Fox, Glen P

2016-07-01

In vitro experimental environments are used to study interactions between microorganisms, and to predict dynamics in natural ecosystems. This study highlights that experimental in vitro environments should be selected to match closely the natural environment of interest during in vitro studies to strengthen extrapolations about aflatoxin production by Aspergillus and competing organisms. Fungal competition and aflatoxin accumulation were studied in soil, cotton wool or tube (water-only) environments, for Aspergillus flavus competition with Penicillium purpurogenum, Fusarium oxysporum or Sarocladium zeae within maize grains. Inoculated grains were incubated in each environment at two temperature regimes (25 and 30°C). Competition experiments showed interaction between the main effects of aflatoxin accumulation and the environment at 25°C, but not so at 30°C. However, competition experiments showed fungal populations were always interacting with their environments. Fungal survival differed after the 72-h incubation in different experimental environments. Whereas all fungi incubated within the soil environment survived, in the cotton wool environment none of the competitors of A. flavus survived at 30°C. With aflatoxin accumulation, F. oxysporum was the only fungus able to interdict aflatoxin production at both temperatures. This occurred only in the soil environment and fumonisins accumulated instead. Smallholder farmers in developing countries face serious mycotoxin contamination of their grains, and soil is a natural reservoir for the associated fungal propagules, and a drying and storage surface for grains on these farms. Studying fungal dynamics in the soil environment and other environments in vitro can provide insights into aflatoxin accumulation post-harvest.

In vivo and in vitro toxicological effects of titanium dioxide nanoparticles on small intestine

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tassinari, Roberta; La Rocca, Cinzia; Tait, Sabrina

2015-06-23

In European Union, titanium dioxide (TiO{sub 2}) as bulk material is a food additive (E171) and - as nanoparticle (NP) - is used as a white pigment in several products (e.g. food, cosmetics, drugs). E171 contains approximately 36% of particles less than 100 nm in at least one dimension and TiO{sub 2} NP exposure is estimated fairly below 2.5 mg/person/day. The gastrointestinal tract is a route of entry for NPs, thus representing a potential target of effects. In in vivo study, the effects of TiO{sub 2} NP in adult rat small intestine have been evaluated by oral administration of 0 (CTRL), 1more » and 2 mg/kg body weight per day - relevant to human dietary intake. Detailed quali/quantitative histopathological analyses were performed on CTRL and treated rat samples. Scanning electron microscopy (SEM) analysis was performed on small intestine. An in vitro study on Caco-2 cells was also used in order to evaluate the potential cytotoxic effects directly on enterocytes through the lactate dehydrogenase (LDH) assay. Suspensions of TiO{sub 2} NPs for in vitro and in vivo study were characterized by EM. Histomorphometrical data showed treatment-related changes of villus height and widths in male rats. Significantly different from CTRL decreased LDH levels in the medium were detected in vitro at 24h with 2.5, 5, 10 and 20 µg/cm{sup 2} levels of TiO{sub 2} NPs. SEM analysis showed no damaged areas. Overall the results showed that enterocytes may represent a target of TiO{sub 2} NP toxicity by direct exposure both in vivo and in vitro models.« less

In vivo and in vitro toxicological effects of titanium dioxide nanoparticles on small intestine

NASA Astrophysics Data System (ADS)

Tassinari, Roberta; La Rocca, Cinzia; Stecca, Laura; Tait, Sabrina; De Berardis, Barbara; Ammendolia, Maria Grazia; Iosi, Francesca; Di Virgilio, Antonio; Martinelli, Andrea; Maranghi, Francesca

2015-06-01

In European Union, titanium dioxide (TiO2) as bulk material is a food additive (E171) and - as nanoparticle (NP) - is used as a white pigment in several products (e.g. food, cosmetics, drugs). E171 contains approximately 36% of particles less than 100 nm in at least one dimension and TiO2 NP exposure is estimated fairly below 2.5 mg/person/day. The gastrointestinal tract is a route of entry for NPs, thus representing a potential target of effects. In in vivo study, the effects of TiO2 NP in adult rat small intestine have been evaluated by oral administration of 0 (CTRL), 1 and 2 mg/kg body weight per day - relevant to human dietary intake. Detailed quali/quantitative histopathological analyses were performed on CTRL and treated rat samples. Scanning electron microscopy (SEM) analysis was performed on small intestine. An in vitro study on Caco-2 cells was also used in order to evaluate the potential cytotoxic effects directly on enterocytes through the lactate dehydrogenase (LDH) assay. Suspensions of TiO2 NPs for in vitro and in vivo study were characterized by EM. Histomorphometrical data showed treatment-related changes of villus height and widths in male rats. Significantly different from CTRL decreased LDH levels in the medium were detected in vitro at 24h with 2.5, 5, 10 and 20 µg/cm2 levels of TiO2 NPs. SEM analysis showed no damaged areas. Overall the results showed that enterocytes may represent a target of TiO2 NP toxicity by direct exposure both in vivo and in vitro models.

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.

Approved oncology drugs lack in vivo activity against Trichuris muris despite in vitro activity.

PubMed

Cowan, Noemi; Raimondo, Alessia; Keiser, Jennifer

2016-11-01

Infections with soil-transmitted helminths (STHs) are considered among the most persistent global health problems. The few available drugs have limitations including low efficacy against Trichuris trichiura infections. As a starting point toward drug repositioning, we studied a set of FDA-approved oncology drugs for activity against Trichuris muris since targets relevant to cancer therapy might have a function in helminth biology. Drugs were tested in vitro on the larval and adult stage of T. muris. Compounds active in vitro were tested in the T. muris mouse model at single oral dosages of 200-400 mg/kg. Of the 114 drugs tested in vitro, 12 showed activity against T. muris larvae (>80 % drug effect at 50 μM). Ten of these drugs were also active on the adult worm stage (>80 % drug effect at 50 μM), of which six revealed IC 50 values between 1.8 and 5.0 μM. Except for tamoxifen citrate, all in vitro active drugs were protein kinase inhibitors. None of the drugs tested in vivo showed efficacy, revealing worm burden reductions of 0-24 % and worm expulsion rates of 0-7.9 %. The promising in vitro activities of protein kinases could not be confirmed in vivo. Drug discovery against STH should be strengthened including the definition of compound progression criteria. Follow-up structure-activity relationship studies with modified compounds might be considered.

Validation of an approach to predict total-tract fiber digestibility using a standardized in vitro technique for different diets fed to high-producing dairy cows.

PubMed

Lopes, F; Ruh, K; Combs, D K

2015-04-01

The experimental objective was to validate an in vitro model to predict total-tract neutral detergent fiber (NDF) digestibility in dairy cattle. Twenty-one diets from 7 studies conducted at University of Wisconsin-Madison were analyzed for in vitro fiber digestibility. Forages varied among diets (corn, alfalfa, tall and meadow fescue, and wheat straw silages) and nutrient composition (ranges: NDF = 22.5 to 33.8%; crude protein = 15.8 to 18.9%; nonfiber carbohydrates = 38.0 to 51.0%). Total-tract NDF digestibility (TTNDFD) observed in in vivo trials was determined using different markers as described in the individual studies. The in vitro TTNDFD model predicted total-tract fiber digestibility from the proportion of total NDF potentially digestible (pdNDF), rate of pdNDF degradation, and rate of passage of pdNDF. The model predicted TTNDFD similar to in vivo measurements. The relationship between TTNDFD measured in vivo and TTNDFD predicted by the in vitro assay was significant (R(2) = 0.68). The relationship between in vitro 30-h NDF digestibility values and in vivo total-tract NDF digestibility values was not significant, whereas in vitro 48-h NDF digestibility values were correlated (R(2) = 0.30) with in vivo TTNDFD measurements. Indigestible NDF (iNDF) showed a negative relationship (R(2) = 0.40) with TTNDFD in vivo. Each 1-percentage-unit increase of iNDF resulted in a decrease of 0.96 percentage units of total-tract NDF digestibility; however, iNDF by itself was not a good predictor of TTNDFD because of the difference among the means. This study showed that an in vitro TTNDFD model that uses iNDF, pdNDF, and rates of pdNDF digestion and passage can predict (R(2) = 0.68) total-tract NDF digestibility. Most importantly, we demonstrated the ability to predict total-tract fiber digestibility from a model based on in vitro NDF degradation, which could improve our ability to optimize forage utilization and milk production. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Formulation and in vitro evaluation of xanthan gum-based bilayered mucoadhesive buccal patches of zolmitriptan.

PubMed

Shiledar, Rewathi R; Tagalpallewar, Amol A; Kokare, Chandrakant R

2014-01-30

A novel bilayered mucoadhesive buccal patch of zolmitriptan was prepared using xanthan gum (XG) as mucoadhesive polymer. Hydroxypropyl methylcellulose E-15 was used as film-former and polyvinyl alcohol (PVA) was incorporated, to increase the tensile strength of the patches. To study the effect of independent variables viz. concentrations of XG and PVA, on various dependent variables like in vitro drug release, ex vivo mucoadhesive strength and swelling index, 3(2) factorial design was employed. In vitro drug release studies of optimized formulation showed initially, rapid drug release; 43.15% within 15 min, followed by sustained release profile over 5h. Incorporation of 4% dimethyl sulfoxide enhanced drug permeability by 3.29 folds, transported 29.10% of drug after 5h and showed no buccal mucosal damage after histopathological studies. In conclusion, XG can be used as a potential drug release modifier and mucoadhesive polymer for successful formulation of zolmitriptan buccal patches. Copyright © 2013 Elsevier Ltd. All rights reserved.

Gamma radiation effects on phenolics, antioxidants activity and in vitro digestion of pistachio ( Pistachia vera) hull

NASA Astrophysics Data System (ADS)

Behgar, M.; Ghasemi, S.; Naserian, A.; Borzoie, A.; Fatollahi, H.

2011-09-01

The effect of gamma radiation (10, 20, 30, 40, 50 and 60 kGy) on tannin, total phenolics, antioxidants activity and in vitro digestion of pistachio hulls has been investigated in this study. The possibility of using the radial diffusion method based on software measurement of the rings area has also been investigated in this study. The software based method in radial diffusion method showed a higher r2 (0.995) value when compared to the traditional method. Irradiation reduced the tannin content ( P<0.01) and activity of antioxidants ( P<0.05) of pistachio hull extracts but increased the total phenolic content ( P<0.05). There was no effect of gamma irradiation on the in vitro digestion of the pistachio hull. Irradiation decreased the digestion rate of the pistachio hull at the dose of 40 kGy when compared to the control. This study showed that gamma irradiation decreased tannin and antioxidants activity of pistachio hull.

Preparation and characterization of cefditoren pivoxil-loaded liposomes for controlled in vitro and in vivo drug release

PubMed Central

Venugopalarao, Gojjala; Lakshmipathy, Rajasekhar; Sarada, Nallani Chakravarthula

2015-01-01

Background The application of antibiotics has been limited due to weak biodistribution and pharmacokinetics. Encapsulation of these drugs in lipid vesicles might be a good solution for obtaining the required properties. Liposomes are one of the most suitable drug-delivery systems to deliver the drug to the target organ and minimize the distribution of the drug to non-target tissues. Objective The study reported here aimed to develop cefditoren pivoxil liposomes by thin-film hydration, characterize them in terms of physical interactions, and undertake in vitro and in vivo release studies. Methodology The pre-formulation studies were carried out using Fourier-transform infrared spectroscopy and differential scanning calorimetry. Cefditoren pivoxil liposomal formulations were formulated by thin-film hydration using biomaterials ie, soya lecithin and cholesterol in different molar ratios. The best molar ratio was determined by in vitro studies such as entrapment efficacy, particle size distribution, and diffusion. Results From the in vitro release studies, it was found that the formulation that contained soya lecithin and cholesterol in a 1.0:0.6 molar ratio gave good entrapment of 72.33% and drug release of 92.5% at 36 hours. Further, the formulation’s zeta potential and surface morphology were examined and stability and in vivo studies were undertaken evaluating the pharmacokinetic parameters, which showed promising results. Conclusion Formulation CPL VI showed the maximum drug-loading capacity of 72.3% with good controlled release and acceptable stability when compared with the other formulations. In vivo studies in rabbits showed that the drug release from the liposomes was successfully retarded with good controlled release behavior which can be used to treat many bacterial infections with a minimal dose. PMID:26491316

In vivo and in vitro invasion in relation to phenotypic characteristics of human colorectal carcinoma cells.

PubMed Central

de Vries, J. E.; Dinjens, W. N.; De Bruyne, G. K.; Verspaget, H. W.; van der Linden, E. P.; de Bruïne, A. P.; Mareel, M. M.; Bosman, F. T.; ten Kate, J.

1995-01-01

In this study we investigated the tumorigenicity, growth pattern and spontaneous metastatic ability of a series of nine human colorectal carcinoma cell lines after subcutaneous and intracaecal xenografting in nude mice. CaCo2 cells were found to be poorly tumorigenic to non-tumorigenic in either site; the other cell lines were tumorigenic in both sites. SW1116, SW480 and SW620 did not show local invasive in the NCI-H716 and LS174T cells were both invasive in the caecum, but only NCI-H716 was invasive in the subcutis. HT29 and 5583 (S and E) cells were invasive in the caecum and from that site metastatic to the lungs and/or the liver. HT29 and 5583S cells were both invasive in the subcutis, but 5583E cells were not. Of each category of in vivo behaviour in the caecum, one cell line was further investigated with regard to invasion in vitro (into embryonic chick heart fragments), E-cadherin expression in vivo and in vitro and in vitro production of u-PA and t-PA. These parameters were chosen in view of their purported role in extracellular matrix degradation and intercellular adhesion, which are all involved in the invasive and metastatic cascade. Invasion in vitro was not predictive for invasion or metastasis in vivo. In the cell line which showed invasion in embryonic chick heart tissue, heterogeneous E-cadherin expression was observed in vitro together with a relatively high production of u-PA. The non-invasive cell lines showed in vitro homogeneous expression of E-cadherin with a relatively low production of u-PA. In vivo expression of E-cadherin was either absent or heterogeneous. We conclude that: (1) colorectal carcinoma xenografts show site-specific modification of in vivo invasive and metastatic behaviour; (2) invasion in vitro does not correlate with invasion and metastasis in vivo; (3) in vitro non-invasion might be associated with homogeneous E-cadherin expression and low production of u-PA; (4) E-cadherin expression in vitro differs from E-cadherin expression in vivo. The results support the notion that the microenvironment in which cancer cells grow is one of the factors involved in the regulation of invasive and metastatic behaviour. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:7841040

Zinc attenuates forskolin-stimulated electrolyte secretion without involvement of the enteric nervous system in small intestinal epithelium from weaned piglets.

PubMed

Feng, Zike; Carlson, Dorthe; Poulsen, Hanne Damgaard

2006-11-01

In a previous study, we found that secretagogue-stimulated electrolyte secretion was attenuated by dietary and serosal zinc in piglet small intestinal epithelium in Ussing chambers. Several studies show that the enteric nervous system (ENS) is involved in regulation of electrolyte and/or fluid transport in intestinal epithelium from many species. The aim of the present study is to examine the mechanisms behind the attenuating effect of zinc on electrolyte secretion and to study whether the ENS is involved in this effect of zinc in vitro. Twenty-four piglets (six litters of four piglets) were allocated randomly to one of two dietary treatments consisting of a basic diet supplemented with 100 mg zinc/kg (Zn(100)) or 2500 mg zinc/kg (Zn(2500)), as ZnO. All the piglets were killed at 5-6 days after weaning and in vitro experiments with small intestinal epithelium in Ussing chambers were carried out. Furthermore, zinc, copper, alkaline phosphatase (AP) and metallothionein (MT) in mucosa, liver, and plasma were measured. These measurements showed that zinc status was increased in the Zn(2500) compared to the Zn(100) fed piglets. The in vitro studies did not confirm previous findings of attenuating effects of dietary zinc and zinc in vitro on the 5-HT induced secretion. But it showed that the addition of zinc at the serosal side attenuated the forskolin (FSK) (cAMP-dependent) induced ion secretion in epithelium from piglets fed with Zn(100) diet. Blocking the ENS with lidocaine or hexamethonium apparently slightly reduced this effect of zinc in vitro, but did not remove the effect of zinc. Consequently, it is suggested that zinc attenuates the cAMP dependent ion secretion mainly due to an effect on epithelial cells rather than affecting the mucosal neuronal pathway.

Clinical-grade production of human mesenchymal stromal cells: occurrence of aneuploidy without transformation.

PubMed

Tarte, Karin; Gaillard, Julien; Lataillade, Jean-Jacques; Fouillard, Loic; Becker, Martine; Mossafa, Hossein; Tchirkov, Andrei; Rouard, Hélène; Henry, Catherine; Splingard, Marie; Dulong, Joelle; Monnier, Delphine; Gourmelon, Patrick; Gorin, Norbert-Claude; Sensebé, Luc

2010-02-25

Clinical-grade human mesenchymal stromal cells (MSCs) have been expanded in vitro for tissue engineering or immunoregulatory purposes without standardized culture conditions or release criteria. Although human MSCs show poor susceptibility for oncogenic transformation, 2 recent studies described their capacity to accumulate chromosomal instability and to give rise to carcinoma in immunocompromised mice after long-term culture. We thus investigated the immunologic and genetic features of MSCs expanded with fetal calf serum and fibroblast growth factor or with platelet lysate in 4 cell-therapy facilities during 2 multicenter clinical trials. Cultured MSCs showed a moderate expression of human leukocyte antigen-DR without alteration of their low immunogenicity or their immunomodulatory capacity. Moreover, some transient and donor-dependent recurring aneuploidy was detected in vitro, independently of the culture process. However, MSCs with or without chromosomal alterations showed progressive growth arrest and entered senescence without evidence of transformation either in vitro or in vivo.

Antioxidant Activity of Pistacia vera Fruits, Leaves and Gum Extracts

PubMed Central

Hosseinzadeh, Hossein; Sajadi Tabassi, Sayyed Abolghasem; Milani Moghadam, Negar; Rashedinia, Marzieh; Mehri, Soghra

2012-01-01

The side effects of synthetic antioxidants have been considered in different studies. Accordingly, there is an increasing interest toward the use of natural substances instead of the synthetic ones. In this study, the aqueous and ethanolic extracts of Pistacia vera leaves and fruits as well as hydroalcoholic extract of gum were tested for a possible antioxidant activity using in vitro methods. Deoxyribose assay, erythrocyte membrane lipid peroxidation and liver misrosomal non- enzymatic lipid peroxidation tests were used as an in-vitro model for determination antioxidant activity. The extract were evaluated at different concentratios: 25,100, 250, 500 and 1000 μg/mL. In all procedures, all extracts showed free radical scavenging activity. The effect of ethanolic extract of P. vera fruit at 1000 μg/mL was quite similar to positive control (DMSO 20 mM) in deoxyribose method. In two other tests, the ethanolic extracts of fruits and leaves were more effective than the aqueous extracts to inhibit malondialdehyde generation. Phytochemical tests showed the presence of flavonoids and tannins in Pistocia vera extracts. The present study showed that extracts of different part of P. vera have antioxidant activity in different in vitro methods. The ethanolic extracts of leaves and fruits showed more roles for antioxidant properties and gum hydroalcoholic extract demonstrated less antioxidant effect. PMID:24250515

Study on bioactive compounds of in vitro cultured Calculus Suis and natural Calculus Bovis.

PubMed

Wan, Tien-Chun; Cheng, Fu-Yuan; Liu, Yu-Tse; Lin, Liang-Chuan; Sakata, Ryoichi

2009-12-01

The purpose of the study was to investigate bioactive compounds of in vitro cultured Calculus Suis and natural Calculus Bovis obtained as valuable by-products from animals used for meat production. The results showed that the components of natural Calculus Bovis were rich in bilirubin and biliverdin and had higher content of essential amino acids. The major amino acids of in vitro cultured Calculus Suis were identified as glycine, alanine, glutamic acid and aspartic acid, and those for natural Calculus Bovis were found to be glutamic acid, aspartic acid, proline, and arginine. The methionine and cysteine contents of precursors for glutathione in natural Calculus Bovis were significantly higher than those of in vitro cultured Calculus Suis. The mineral contents of zinc, iron and manganese of natural Calculus Bovis were significantly higher than those of in vitro cultured Calculus Suis. The major bile acids in both products were cholic acid and dehydrocholic acid, respectively. The chenodeoxycholic and ursodeoxycholic acid content of in vitro cultured Calculus Suis was significantly higher than that of natural Calculus Bovis.

Way forward in case of a false positive in vitro genotoxicity result for a cosmetic substance?

PubMed

Doktorova, Tatyana Y; Ates, Gamze; Vinken, Mathieu; Vanhaecke, Tamara; Rogiers, Vera

2014-02-01

The currently used regulatory in vitro mutagenicity/genotoxicity test battery has a high sensitivity for detecting genotoxicants, but it suffers from a large number of irrelevant positive results (i.e. low specificity) thereby imposing the need for additional follow-up by in vitro and/or in vivo genotoxicity tests. This could have a major impact on the cosmetic industry in Europe, seen the imposed animal testing and marketing bans on cosmetics and their ingredients. Afflicted, but safe substances could therefore be lost. Using the example of triclosan, a cosmetic preservative, we describe here the potential applicability of a human toxicogenomics-based in vitro assay as a potential mechanistically based follow-up test for positive in vitro genotoxicity results. Triclosan shows a positive in vitro chromosomal aberration test, but is negative during in vivo follow-up tests. Toxicogenomics analysis unequivocally shows that triclosan is identified as a compound acting through non-DNA reactive mechanisms. This proof-of-principle study illustrates the potential of genome-wide transcriptomics data in combination with in vitro experimentation as a possible weight-of-evidence follow-up approach for de-risking a positive outcome in a standard mutagenicity/genotoxicity battery. As such a substantial number of cosmetic compounds wrongly identified as genotoxicants could be saved for the future. Copyright © 2013 Elsevier Ltd. All rights reserved.

Spermicidal activity of the hexane extract of Piper longum: an in vitro study.

PubMed

Sarwar, Abu Hasnath Md Golam; Nirala, Ranjeet Kumar; Arif, Mohammed; Khillare, Beena; Thakur, Sonu Chand

2015-01-01

This study was carried out to assess the spermicidal action of hexane extract from the fruits of Piper longum Linn. The sperm immobilisation studies showed that 20 mg/mL of hexane extract was able to immobilise sperms completely within 20 s. The sperm revival test revealed that the effects were spermicidal as sperm immobilisation effect was irreversible. There was also a significant reduction in sperm viability in the treated group in comparison to the control. The hypo-osmotic swelling of these sperms was significantly reduced, indicating that the hexane extract may probably cause injury to the sperm plasma membrane. Hence, this study showed that the hexane extract of P. longum possesses potential contraceptive spermicidal activity in vitro.

Laminin α5 substrates promote survival, network formation and functional development of human pluripotent stem cell-derived neurons in vitro.

PubMed

Hyysalo, Anu; Ristola, Mervi; Mäkinen, Meeri E-L; Häyrynen, Sergei; Nykter, Matti; Narkilahti, Susanna

2017-10-01

Laminins are one of the major protein groups in the extracellular matrix (ECM) and specific laminin isoforms are crucial for neuronal functions in the central nervous system in vivo. In the present study, we compared recombinant human laminin isoforms (LN211, LN332, LN411, LN511, and LN521) and laminin isoform fragment (LN511-E8) in in vitro cultures of human pluripotent stem cell (hPSC)-derived neurons. We showed that laminin substrates containing the α5-chain are important for neuronal attachment, viability and network formation, as detected by phase contrast imaging, viability staining, and immunocytochemistry. Gene expression analysis showed that the molecular mechanisms involved in the preference of hPSC-derived neurons for specific laminin isoforms could be related to ECM remodeling and cell adhesion. Importantly, the microelectrode array analysis revealed the widest distribution of electrophysiologically active neurons on laminin α5 substrates, indicating most efficient development of neuronal network functionality. This study shows that specific laminin α5 substrates provide a controlled in vitro culture environment for hPSC-derived neurons. These substrates can be utilized not only to enhance the production of functional hPSC-derived neurons for in vitro applications like disease modeling, toxicological studies, and drug discovery, but also for the production of clinical grade hPSC-derived cells for regenerative medicine applications. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Enhanced Bioavailability and Anticancer Effect of Curcumin-Loaded Electrospun Nanofiber: In Vitro and In Vivo Study

NASA Astrophysics Data System (ADS)

Wang, Chuan; Ma, Chao; Wu, Zhenkai; Liang, He; Yan, Peng; Song, Jia; Ma, Nan; Zhao, Qinghua

2015-11-01

Nanofibers have attracted increasing attention in drug delivery and other biomedical applications due to their some special properties. The present study aims to prepare a fiber-based nanosolid dispersion system to enhance the bioavailability of curcumin (CUR). CUR-loaded polyvinyl pyrrolidone (CUR@PVP) nanofibers were successfully prepared via electrospinning. Scanning electron microscopy (SEM) was employed to observe the morphology of the nanofibers, and the SEM image showed that the drug-loaded nanofibers were smooth, and no CUR clusters were found on the surface of the nanofibers. The results of X-ray diffraction (XRD) demonstrated that the CUR was evenly distributed in the nanofibers in an amorphous state. Fourier transform infrared (FTIR) spectroscopy analysis indicated that intermolecular hydrogen bonding occurred between the CUR and the polymer matrix. In vitro dissolution profiles showed that CUR@PVP nanofiber could be quickly dissolved in phosphate-buffered saline (PBS) solution, while negligible dissolution was observed in pure CUR sample. Importantly, in vitro cell viability assays and in vivo animal tests revealed that the nanosolid dispersion system dramatically enhanced the bioavailability and showed effective anticancer effect of the CUR.

In vitro dissolution and in vivo gamma scintigraphic evaluation of press-coated salbutamol sulfate tablets.

PubMed

Li, Wei; Shi, Cai-Hong; Sheng, Yi-Ling; Cui, Ping; Zhao, Yu-Qing; Zhang, Xiang-Rong

2013-12-01

The aim of this study was to investigate the in vitro and in vivo performance of salbutamol sulfate press-coated tablets for delayed release. The in vitro release behavior of press-coated tablets with the outer layer of PEG 6000/ Eudragit S100 blends (2:1) in pH 1.2 (0.1 mol L-1 HCl) and then pH 6.8 buffer solution was examined. Morphological change of the press-coated tablet during in vitro release was recorded with a digital camera. Release of salbutamol sulfate from press-coated tablets was less than 5 % before 3 h and was completed after 8 h in pH 6.8 phosphate buffer solution. In vivo gamma scintigraphy study carried out on healthy men indicated that the designed system released the drug in lower parts of the GI tract after a lag time of 5 hours. The results showed the capability of the system of achieving delayed release of the drug in both in vitro and in vivo gamma scintigraphy studies.

Correlation of In Vivo Versus In Vitro Benchmark Doses (BMDs) Derived From Micronucleus Test Data: A Proof of Concept Study.

PubMed

Soeteman-Hernández, Lya G; Fellows, Mick D; Johnson, George E; Slob, Wout

2015-12-01

In this study, we explored the applicability of using in vitro micronucleus (MN) data from human lymphoblastoid TK6 cells to derive in vivo genotoxicity potency information. Nineteen chemicals covering a broad spectrum of genotoxic modes of action were tested in an in vitro MN test using TK6 cells using the same study protocol. Several of these chemicals were considered to need metabolic activation, and these were administered in the presence of S9. The Benchmark dose (BMD) approach was applied using the dose-response modeling program PROAST to estimate the genotoxic potency from the in vitro data. The resulting in vitro BMDs were compared with previously derived BMDs from in vivo MN and carcinogenicity studies. A proportional correlation was observed between the BMDs from the in vitro MN and the BMDs from the in vivo MN assays. Further, a clear correlation was found between the BMDs from in vitro MN and the associated BMDs for malignant tumors. Although these results are based on only 19 compounds, they show that genotoxicity potencies estimated from in vitro tests may result in useful information regarding in vivo genotoxic potency, as well as expected cancer potency. Extension of the number of compounds and further investigation of metabolic activation (S9) and of other toxicokinetic factors would be needed to validate our initial conclusions. However, this initial work suggests that this approach could be used for in vitro to in vivo extrapolations which would support the reduction of animals used in research (3Rs: replacement, reduction, and refinement). © The Author 2015. Published by Oxford University Press on behalf of the Society of Toxicology.

In Vitro Sensitivity of Cutaneous Leishmania Promastigote Isolates Circulating in French Guiana to a Set of Drugs

PubMed Central

Ginouvès, Marine; Simon, Stéphane; Nacher, Mathieu; Demar, Magalie; Carme, Bernard; Couppié, Pierre; Prévot, Ghislaine

2017-01-01

Anti-leishmaniasis drug resistance is a common problem worldwide. The aim of this study was to inventory the general in vitro level of sensitivity of Leishmania isolates circulating in French Guiana and to highlight potential in vitro pentamidine-resistant isolates. This sensitivity study was conducted on 36 patient-promastigote isolates for seven drugs (amphotericin B, azithromycin, fluconazole, meglumine antimoniate, miltefosine, paromomycin, and pentamidine) using the Cell Counting Kit-8 viability test. The IC50 values obtained were heterogeneous. One isolate exhibited high IC50 values for almost all drugs tested. Pentamidine, which is the first-line treatment in French Guiana, showed efficacy at very low doses (mean of 0.0038 μg/mL). The concordance of the in vitro pentamidine results with the patients' clinical outcomes was 94% (K = 0.82). PMID:28167598

Quantitative determination of secoiridoid and gamma-pyrone compounds in Gentiana lutea cultured in vitro.

PubMed

Menković, N; Savikin-Fodulović, K; Momcilović, I; Grubisić, D

2000-02-01

The production of secondary metabolites was studied in shoots, roots, and hairy roots of Gentiana lutea obtained in vitro. In shoots, both secoiridoid and gamma-pyrone compounds were detected in amounts similar to those found in aerial parts of plants collected from nature. The most abundant secoiridoid was gentiopicrin while mangiferin was the main compound among the gamma-pyrones. The adventitious roots obtained in vitro showed a poor biosynthetic capacity. Upon infection with Agrobacterium rhizogenes, nine hairy root clones were established which differed in the amount of secondary metabolites.

In vitro activity of doripenem and other carbapenems against contemporary Gram-negative pathogens isolated from hospitalised patients in the Asia-Pacific region: results of the COMPACT Asia-Pacific Study.

PubMed

Christiansen, K J; Ip, M; Ker, H B; Mendoza, M; Hsu, L; Kiratisin, P; Chongthaleong, A; Redjeki, I S; Quintana, A; Flamm, R; Garcia, J; Cassettari, M; Cooper, D; Okolo, P; Morrissey, I

2010-12-01

The Comparative Activity of Carbapenems Testing (COMPACT) Study was designed to determine the in vitro potency of doripenem compared with imipenem and meropenem against a large number of contemporary Gram-negative pathogens from more than 100 centres across Europe and the Asia-Pacific region and to assess the reliability of Etest methodology for doripenem minimum inhibitory concentration (MIC) determination against these pathogens. Data from eight countries within the Asia-Pacific region, which collected 1612 bacterial isolates, are presented here. Etest methodology was found to be a reliable method for MIC determination. Doripenem showed in vitro activity similar to or better than meropenem and at least four-fold better than imipenem against Enterobacteriaceae. Against Pseudomonas aeruginosa, doripenem was also the most active of the three carbapenems in vitro. However, in vitro results do not necessarily correlate with clinical outcome. Copyright © 2010 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

In vitro transcription in E. coli crude lysates prepared on cellophane discs.

PubMed Central

Valla, S; Lindqvist, B H

1978-01-01

An in vitro RNA-synthesizing system consisting of gently lysed E. coli cells on cellophane discs is described. The system has been optimalized with respect to total RNA synthesis. Under certain standard conditions DNA dependent RNA polymerase (EC 2.7.7.6) is responsible for the majority of the RNA synthesis. The extensive rifampicin sensitivity of the synthesis indicates that most of the transcripts are initiated in vitro. The RNA synthesizing system described here has been developed with the aim of studying phage transcription in vitro. We show here that lysates of a P4 infected P2 lysogen support initiation and propagation of transcription from the P2 prophage. PMID:27767

Antibacterial, anti-inflammatory, and bone-regenerative dual-drug-loaded calcium phosphate nanocarriers-in vitro and in vivo studies.

PubMed

Madhumathi, K; Rubaiya, Y; Doble, Mukesh; Venkateswari, R; Sampath Kumar, T S

2018-05-01

A dual local drug delivery system (DDS) composed of calcium phosphate bioceramic nanocarriers aimed at treating the antibacterial, anti-inflammatory, and bone-regenerative aspects of periodontitis has been developed. Calcium-deficient hydroxyapatite (CDHA, Ca/P = 1.61) and tricalcium phosphate (β-TCP) were prepared by microwave-accelerated wet chemical synthesis method. The phase purity of the nanocarriers was confirmed by x-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FT-IR), while the transmission electron microscopy (TEM) confirmed their nanosized morphology. CDHA was selected as carrier for the antibiotic (tetracycline) while TCP was chosen as the anti-inflammatory drug (ibuprofen) carrier. Combined drug release profile was studied in vitro from CDHA/TCP (CTP) system and compared with a HA/TCP (BCP) biphasic system. The tetracycline and ibuprofen release rate was 71 and 23% from CTP system as compared to 63 and 20% from BCP system. CTP system also showed a more controlled drug release profile compared to BCP system. Modeling of drug release kinetics from CTP system indicated that the release follows Higuchi model with a non-typical Fickian diffusion profile. In vitro biological studies showed the CTP system to be biocompatible with significant antibacterial and anti-inflammatory activity. In vivo implantation studies on rat cranial defects showed greater bone healing and new bone formation in the drug-loaded CTP system compared to control (no carrier) at the end of 12 weeks. The in vitro and in vivo results suggest that the combined drug delivery platform can provide a comprehensive management for all bone infections requiring multi-drug therapy.

Formulation and in vitro evaluation of xanthan gum or carbopol 934-based mucoadhesive patches, loaded with nicotine.

PubMed

Abu-Huwaij, Rana; Obaidat, Rana M; Sweidan, Kamal; Al-Hiari, Yusuf

2011-03-01

Bilayer nicotine mucoadhesive patches were prepared and evaluated to determine the feasibility of the formulation as a nicotine replacement product to aid in smoking cessation. Nicotine patches were prepared using xanthan gum or carbopol 934 as a mucoadhesive polymers and ethyl cellulose as a backing layer. The patches were evaluated for their thickness, weight and content uniformity, swelling behavior, drug-polymers interaction, adhesive properties, and drug release. The physicochemical interactions between nicotine and the polymers were investigated by Fourier transform infrared (FTIR) spectroscopy. Mucoadhesion was assessed using two-arm balance method, and the in vitro release was studied using the Franz cell. FTIR revealed that there was an acid base interaction between nicotine and carbopol as well as nicotine and xanthan. Interestingly, the mucoadhesion and in vitro release studies indicated that this interaction was strong between the drug and carbopol whereas it was weak between the drug and xanthan. Loading nicotine concentration to non-medicated patches showed a significant decrease in the mucoadhesion strength of carbopol patches and no significant effect on the mucoadhesion strength of xanthan patches. In vitro release studies of the xanthan patches showed a reasonable fast initial release profile followed by controlled drug release over a 10-h period. © 2010 American Association of Pharmaceutical Scientists

Application of Box-Behnken design for preparation of levofloxacin-loaded stearic acid solid lipid nanoparticles for ocular delivery: Optimization, in vitro release, ocular tolerance, and antibacterial activity.

PubMed

Baig, Mirza Salman; Ahad, Abdul; Aslam, Mohammed; Imam, Syed Sarim; Aqil, Mohd; Ali, Asgar

2016-04-01

The aim of the present study was to develop and optimize levofloxacin loaded solid lipid nanoparticles for the treatment of conjunctivitis. Box-Behnken experimental design was applied for optimization of solid lipid nanoparticles. The independent variables were stearic acid as lipid (X1), Tween 80 as surfactant (X2) and sodium deoxycholate as co-surfactant (X3) while particle size (Y1) and entrapment efficiency (Y2) were the dependent variables. Further in vitro release and antibacterial activity in vitro were also performed. The optimized formulation of levofloxacin provides particle size of 237.82 nm and showed 78.71% entrapment efficiency and achieved flux 0.2,493 μg/cm(2)/h across excised goat cornea. In vitro release study showed prolonged drug release from the optimized formulation following Korsmeyer-Peppas model. Antimicrobial study revealed that the developed formulation possesses antibacterial activity against Staphylococcus aureus, and Escherichia coli equivalent to marketed eye drops. HET-CAM test demonstrated that optimized formulation was found to be non-irritant and safe for topical ophthalmic use. Our results concluded that solid lipid nanoparticles are an efficient carrier for ocular delivery of levofloxacin and other drugs. Copyright © 2015 Elsevier B.V. All rights reserved.

Comparison of skin permeability for three diclofenac topical formulations: an in vitro study.

PubMed

Folzer, E; Gonzalez, D; Singh, R; Derendorf, H

2014-01-01

Diclofenac is a hydrophilic non-steroidal anti-inflammatory drug (NSAID) widely used in humans and animals. There are limited published studies evaluating diclofenac's skin permeation following topical administration. The aim of our study was to evaluate and compare the in vitro permeation of three different diclofenac-containing formulations (patch, gel, solution) over 24 hours. These formulations were applied (n = 6 per formulation) to pig skin sandwiched between the two chambers in a static Franz diffusion cell and aliquots from the receptor medium were sampled at pre-defined time points. An HPLC method with UV detection was developed and validated with the aim of characterizing the transepidermal penetration in the in vitro system. Using this assay to determine the permeation parameters, results at 24 hours showed that the Flector patch released the highest drug amount (54.6%), whereas a lower drug amount was delivered with the Voltaren Emulgel (38.2%) and the solution (34.4%). The commercial gel showed the highest flux (39.9 +/- 0.9 microg/cm2/h) and the shortest lag-time (1.97 +/- 0.02 h). Based on these in vitro results using pig skin, the transdermal patch resulted in a long-lasting controlled release of diclofenac, while the gel had the shortest lag-time.

Reconstituted human gingival epithelium: nonsubmerged in vitro model.

PubMed

Delcourt-Huard, A; Corlu, A; Joffre, A; Magloire, H; Bonnaure-Mallet, M

1997-01-01

Many studies have shown that human gingival keratinocytes grown in submerged culture fail to attain optimal differentiation. This study reports an in vitro culture system for oral gingival epithelial cells, in which they are grown at the air-liquid interface, on polycarbonate inserts, in the presence of an NIH-3T3 feeder layer. This model was compared with two submerged culture methods for gingival keratinocytes, on type 1 collagen gel and on an NIH-3T3 feeder layer. Transmission electron microscopy showed an advanced level of stratification (over six layers of cells) for cultures grown at the air-liquid interface. Immunofluorescence and electrophoretic patterns showed the presence of cytokeratins 10 and 11 in cytoskeletal protein extracts of these cultured keratinocytes. In this air-liquid interface culture model, in the presence of NIH-3T3 feeder cells, keratinocytes can achieve an advanced level of stratification and differentiation and a resemblance to in vivo gingiva. The obtention of a highly differentiated epithelium will permit in vitro pharmacological studies and studies on the biocompatability of certain alloys with the superficial periodontium; it will also provide grafts for patients undergoing periodontal surgery.

In vitro evaluation of α-lipoic acid-loaded lipid nanocapsules for topical delivery.

PubMed

Xia, Nan; Liu, Tian; Wang, Qiang; Xia, Qiang; Bian, Xiaoli

2017-09-01

This study aimed at in vitro evaluation of α-lipoic acid-loaded lipid nanocapsules for topical delivery, which was prepared by hot high-pressure homogenisation. Stable particles could be formed and particle size was 148.54 ± 2.31 nm with polydispersity index below 0.15. Encapsulation efficiency and drug loading of α-lipoic acid were 95.23 ± 0.45% and 2.81 ± 0.37%. Antioxidant study showed α-lipoic acid could be protected by lipid nanocapsules without loss of antioxidant activity. Sustained release of α-lipoic acid from lipid nanocapsules was obtained and cumulative release was 62.18 ± 1.51%. In vitro percutaneous study showed the amount of α-lipoic acid distributed in skin was 1.7-fold than permeated. Cytotoxicity assay and antioxidant activity on L929 cells indicated this formulation had low cytotoxicity and ability of protecting cells from oxidative damage within specific concentration. These studies suggested α-lipoic acid-loaded lipid nanocapsules could be potential formulation for topical delivery.

Bioactivities examination of Cinchona leaves ethanol extracts

NASA Astrophysics Data System (ADS)

Artanti, Nina; Udin, Linar Z.; Hanafi, M.; Jamilah, Kurniasih, Ida Rahmi; Primahana, Gian; Anita, Yulia; Sundowo, Andini; Kandace, Yoice Sri

2017-01-01

Cinchona species especially the barks are commonly known for commercial production of quinine as antimalarial. Although it is also reported for treatment of depurative, whooping cough, influenza and dysentery. In this paper we reported in vitro examination of other bioactivities (antidiabetes, antioxidant and in vitro cytotoxicity) of 70% ethanol extract of Cinchona ledgeriana and C. succirubra leaves as well as qunine, quinidine, and cinchonine the major alkaloids found in Cinchona species. Antidiabetes was conducted using α-glucosidase inhibitory activity assay. Antioxidant was conducted using DPPH free radical scavenging activity assay. In vitro cytotoxic activity was concucted by microscopic observation on growth of breast cancer cell line MCF-7. The results showed that at concentration of 100 µg/ml, C. ledgeriana leaves ethanol extracts showed the best activity as antidiabetes (98% inhibitory of α-glucosidase activity) and antioxidant (92% DPPH free radical scavenging activity), whereas at the same concentration C. succirubra, quinine, quinidine and cinchonine showed very low activities of antidiabetes and antioxidant. Microscopic observation of in vitro cytotoxicity showed that C. ledgeriana also has excellent cytotoxicity to breast cancer cell line MCF-7 which better than quinine, quinidine and cinchonine, whereas C. succirubra showed low cytotoxicity. These results suggest that cinchona species have many potential as the source of drugs discovery and development other than just for malaria treatment. Therefore it is important to conduct further studies and to maintain the available Cinchona plantation in Indonesia.

In vitro and in vivo biological activities of anthocyanins from Nitraria tangutorun Bobr. fruits.

PubMed

Ma, Tao; Hu, Na; Ding, Chenxi; Zhang, Qiulong; Li, Wencong; Suo, Yourui; Wang, Honglun; Bai, Bo; Ding, Chenxu

2016-03-01

Anthocyanins are the main compounds in Nitraria tangutorun Bobr. The enrichment and purification of anthocyanins on macroporous resins were investigated. Regarding anthocyanin purification, static adsorption and desorption were studied. The optimal experimental conditions were the following: resin type: X-5; static adsorption time: 6h; desorption solution: ethanol-water-HCl (80:19:1, V/V/V; pH 1); desorption time: 40min. Furthermore, the in vitro and in vivo biological activities of the anthocyanins were evaluated. The anthocyanins showed ideal scavenging effects on free radicals in vitro, especially on 1,1-diphenyl-2-picrylhydrazyl (DPPH) and hydroxyl free radical (OH). In the animal experiment, blood lipid metabolism of hyperlipidemia rats was regulated by anthocyanin contents. The superoxide dismutase (SOD) activity and the total antioxidant capacity (TAC) of hyperlipidemia rats were also improved by anthocyanins. These results showed that anthocyanins from N. tangutorun Bobr. fruits had potential biological activities in vivo as well as in vitro. Copyright © 2015. Published by Elsevier Ltd.

In vitro leakage susceptibility of tracheoesophageal shunt prostheses in the absence and presence of a biofilm.

PubMed

Oosterhof, Janine J H; van der Mei, Henny C; Busscher, Henk J; Free, Rolien H; Kaper, Hans J; van Weissenbruch, Ranny; Albers, Frans W J

2005-04-01

Although leakage through a tracheoesophageal shunt prosthesis is the main cause of prosthesis failure in a laryngectomy patient, this has never been the subject of in vitro evaluation. The aim of this study was to compare three commercially available voice prostheses by comparison of their in vitro leakage patterns, in absence or presence of a biofilm. To compare in vitro leakage patterns, a model comprised of an artificial throat equipped with a single prosthesis coupled to a water reservoir was developed. By varying the height of the water reservoir, different pressures on the voice prosthesis can be obtained. Both in absence and presence of a biofilm, the Blom Singer voice prosthesis demonstrated the lowest leakage, followed by Groningen Low Resistance. The Provox2 showed significantly the most leakage, however, in presence of a biofilm the leakage of the Provox2 significantly decreased. Regular airflow during biofilm formation significantly increased leakage through the Provox2. Out of 746 clinical replacements, Provox2 showed 76% and Groningen Low Resistance 57% replacements due to leakage. The model used in this study showed significant differences in leakage of the three types of voice prostheses used. Leakage occurred more readily through Provox2 than through Groningen Low Resistance and Blom Singer prostheses, which is in line with clinical observations and enforces the model. (c) 2005 Wiley Periodicals, Inc.

In vitro and in vivo effects of two coconut oils in comparison to monolaurin on Staphylococcus aureus: rodent studies.

PubMed

Manohar, Vijaya; Echard, Bobby; Perricone, Nicholas; Ingram, Cass; Enig, Mary; Bagchi, Debasis; Preuss, Harry G

2013-06-01

Since monolaurin, a monoglyceride formed in the human body in small quantities, has proven effective both in vitro and in vivo against certain strains of Staphylococcus aureus, an important question arises whether consuming a substance high in lauric acid content, such as coconut oil could increase intrinsic monolaurin production to levels that would be successful in overcoming staphylococcal and other microbial invaders. Both a cup plate method and a microdilution broth culture system were employed to test bacteriostatic and bactericidal effects of the test agents in vitro. To test effectiveness in vivo, female C3H/he mice (10-12 per group) were orally administered sterile saline (regular control), vancomycin (positive control), aqueous monolaurin, or two varieties of coconut oil (refined, bleached, deodorized coconut oil and virgin coconut oil) for 1 week before bacterial challenge and 30 days after. A final group received both monolaurin and vancomycin. In contrast to monolaurin, the coconut oils did not show bactericidal activity in vitro. In vivo, the groups receiving vancomycin, monolaurin, or the combination showed some protection--50-70% survival, whereas the protection from the coconut oils were virtually the same as control--0-16% survival. Although we did not find that the two coconut oils are helpful to overcome S. aureus infections, we corroborated earlier studies showing the ability of monolaurin to do such.

Stability of total phenolic concentration and antioxidant capacity of extracts from pomegranate co-products subjected to in vitro digestion.

PubMed

Fawole, Olaniyi Amos; Opara, Umezuruike Linus

2016-09-13

Co-products obtained from pomegranate juice processing contain high levels of polyphenols with potential high added values. From value-addition viewpoint, the aim of this study was to evaluate the stability of polyphenolic concentrations in pomegranate fruit co-products in different solvent extracts and assess the effect on the total antioxidant capacity using the FRAP, DPPH˙ and ABTS(+) assays during simulated in vitro digestion. Pomegranate juice, marc and peel were extracted in water, 50 % ethanol (50%EtOH) and absolute ethanol (100%EtOH) and analysed for total phenolic concentration (TPC), total flavonoids concentration (TFC) and total antioxidant capacity in DPPH˙, ABTS(+) and FRAP assays before and after in vitro digestion. Total phenolic concentration (TPC) and total flavonoid concentration (TFC) were in the order of peel > marc > juice throughout the in vitro digestion irrespective of the extraction solvents used. However, 50 % ethanol extracted 1.1 to 12-fold more polyphenols than water and ethanol solvents depending on co-products. TPC and TFC increased significantly in gastric digests. In contrast, after the duodenal phase of in vitro digestion, polyphenolic concentrations decreased significantly (p < 0.05) compared to those obtained in gastric digests. Undigested samples and gastric digests showed strong and positive relationships between polyphenols and the antioxidant activities measured in DPPH, ABTS(+) and FRAP assays, with correlation coefficients (r (2)) ranging between 0.930-0.990. In addition, the relationships between polyphenols (TPC and TFC) and radical cation scavenging activity in ABTS(+) were moderately positive in duodenal digests. Findings from this study showed that concentration of pomegranate polyphenols and the antioxidant capacity during in vitro gastro-intestinal digestion may not reflect the pre-digested phenolic concentration. Thus, this study highlights the need to provide biologically relevant information on antioxidants by providing data reflecting their stability and activity after in vitro digestion.

[Effects of salt stress on germination and in vitro growth of pistachio (Pistacia vera L.)].

PubMed

Benmahioul, Benamar; Daguin, Florence; Kaid-Harche, Meriem

2009-08-01

In order to study the salinity tolerance of pistachio (Pistacia vera L.), embryos developed from mature seeds were isolated and cultured in vitro and subjected to different NaCl concentrations (0, 42.8, 85.5, 171.1 and 256.6 mM) for 30 days. The results showed that in vitro germination of embryonic axes was not affected by the salt concentration. However, the germinated embryo survival rates decreased from 100% for the control to 62.9% for the highest salt concentration (256.6 mM). In addition, the plantlet growth (length of aerial and root parts, number of leaf produced per embryo, as well as the production of total fresh and dry matter for both aerial parts and roots) showed significant differences according the various salt concentrations.

Heparin for clearance of peripherally inserted central venous catheter in newborns: an in vitro study

PubMed Central

Balaminut, Talita; Venturini, Danielle; da Silva, Valéria Costa Evangelista; Rossetto, Edilaine Giovanini; Zani, Adriana Valongo

2015-01-01

Objective: To compare the efficacy of two concentrations of heparin to clear the lumen of in vitro clotted neonatal peripherally inserted central catheters (PICCs). Methods: This is an in vitro, experimental quantitative study of 76 neonatal 2.0-Fr PICCs coagulated in vitro. The catheters were divided into two groups of 38 PICCs each. In both groups an infusion of low molecular weight heparin was administered with a dose of 25IU/mL for Group 1 and 50IU/mL for Group 2. The negative pressure technique was applied to the catheters of both groups at 5, 15 and 30min and at 4h to test their permeability. Kaplan-Meier survival analysis was used to verify the outcome of the groups according to time intervals. Results: The comparison between both groups in the first 5min showed that more catheters from Group 2 were cleared compared to Group 1 (57.9 vs. 21.1%, respectively). Kaplan-Meier survival analysis showed that less time was needed to clear catheters treated with 50IU/mL of heparin (p<0.001). Conclusions: The use of low molecular weight heparin at a concentration of 50IU/mL was more effective in restoring the permeability of neonatal PICCs occluded in vitro by a clot, and the use of this concentration is within the safety margin indicated by scientific literature. PMID:26116325

The in vitro isolated whole guinea pig brain as a model to study epileptiform activity patterns.

PubMed

de Curtis, Marco; Librizzi, Laura; Uva, Laura

2016-02-15

Research on ictogenesis is based on the study of activity between seizures and during seizures in animal models of epilepsy (chronic condition) or in in vitro slices obtained from naïve non-epileptic brains after treatment with pro-convulsive drugs, manipulations of the extracellular medium and specific stimulation protocols. The in vitro isolated guinea pig brain retains the functional connectivity between brain structures and maintains interactions between neuronal, glial and vascular compartments. It is a close-to-in vivo preparation that offers experimental advantages not achieved with the use of other experimental models. Neurophysiological and imaging techniques can be utilized in this preparation to study brain activity during and between seizures induced by pharmacological or functional manipulations. Cellular and network determinants of interictal and ictal discharges that reproduce abnormal patterns observed in human focal epilepsies and the associated changes in extracellular ion and blood-brain permeability can be identified and analyzed in the isolated guinea pig brain. Ictal and interictal patterns recorded in in vitro slices may show substantial differences from seizure activity recorded in vivo due to slicing procedure itself. The isolated guinea pig brain maintained in vitro by arterial perfusion combines the typical facilitated access of in vitro preparations, that are difficult to approach during in vivo experiments, with the preservation of larger neuronal networks. The in vitro whole isolated guinea pig brain preparation offers an unique experimental model to study systemic and neurovascular changes during ictogenesis. Published by Elsevier B.V.

Lecithin-coated gold nanoflowers (GNFs) for CT scan imaging applications and biochemical parameters; in vitro and in vivo studies.

PubMed

Aziz, Farooq; Bano, Khizra; Siddique, Ahmad Hassan; Bajwa, Sadia Zafar; Nazir, Aalia; Munawar, Anam; Shaheen, Ayesha; Saeed, Madiha; Afzal, Muhammad; Iqbal, M Zubair; Wu, Aiguo; Khan, Waheed S

2018-01-09

We report a novel strategy for the fabrication of lecithin-coated gold nanoflowers (GNFs) via single-step design for CT imaging application. Field-emission electron microscope confirmed flowers like morphology of the as-synthesized nanostructures. Furthermore, these show absorption peak in near-infrared (NIR) region at λ max 690 nm Different concentrations of GNFs are tested as a contrast agent in CT scans at tube voltage 135 kV and tube current 350 mA. These results are compared with same amount of iodine at same CT scan parameters. The results of in vitro CT scan study show that GNFs have good contrast enhancement properties, whereas in vivo study of rabbits CT scan shows that GNFs enhance the CT image clearly at 135 kV as compared to that of iodine. Cytotoxicity was studied and blood profile show minor increase of white blood cells and haemoglobin, whereas decrease of red blood cells and platelets.

The combi-targeting concept: in vitro and in vivo fragmentation of a stable combi-nitrosourea engineered to interact with the epidermal growth factor receptor while remaining DNA reactive.

PubMed

Qiu, Qiyu; Domarkas, Juozas; Banerjee, Ranjita; Merayo, Nuria; Brahimi, Fouad; McNamee, James P; Gibbs, Bernard F; Jean-Claude, Bertrand J

2007-01-01

JDA58 (NSC 741282), a "combi-molecule" optimized in the context of the "combi-targeting concept," is a nitrosourea moiety tethered to an anilinoquinazoline. Here, we sought to show its binary epidermal growth factor receptor (EGFR)/DNA targeting property and to study its fragmentation in vitro and in vivo. The fragmentation of JDA58 was detected in cells in vitro and in vivo by fluorescence microscopy and tandem mass spectrometry. EGFR phosphorylation and DNA damage were determined by Western blotting and comet assay, respectively. Tumor data were examined for statistical significance using the Student's t test. JDA58 inhibited EGFR tyrosine kinase (IC(50), 0.2 micromol/L) and blocked EGFR phosphorylation in human DU145 prostate cancer cells. It induced significant levels of DNA damage in DU145 cells in vitro or in vivo and showed potent antiproliferative activity both in vitro and in a DU145 xenograft model. In cell-free medium, JDA58 was hydrolyzed to JDA35, a fluorescent amine that could be observed in tumor cells both in vitro and in vivo. In tumor cells in vitro or in vivo, or in plasma collected from mice, the denitrosated species JDA41 was the predominant metabolite. However, mass spectrometric analysis revealed detectable levels of the hydrolytic product JDA35 in tumor cells both in vitro and in vivo. The results in toto suggest that growth inhibition in vitro and in vivo may be sustained by the intact combi-molecule plus JDA35 plus JDA41, three inhibitors of EGFR, and the concomitantly released DNA-damaging species. This leads to a model wherein a single molecule carries a complex multitargeted-multidrug combination.

MicroRNA-133 controls vascular smooth muscle cell phenotypic switch in vitro and vascular remodeling in vivo.

PubMed

Torella, Daniele; Iaconetti, Claudio; Catalucci, Daniele; Ellison, Georgina M; Leone, Angelo; Waring, Cheryl D; Bochicchio, Angela; Vicinanza, Carla; Aquila, Iolanda; Curcio, Antonio; Condorelli, Gianluigi; Indolfi, Ciro

2011-09-30

MicroRNA (miR)-1 and -133 play a crucial role in skeletal and cardiac muscle biology and pathophysiology. However, their expression and regulation in vascular cell physiology and disease is currently unknown. The aim of the present study was to evaluate the role, if any, of miR-1 and miR-133 in vascular smooth muscle cell (VSMC) phenotypic switch in vitro and in vivo. We demonstrate here that miR-133 is robustly expressed in vascular smooth muscle cells (VSMCs) in vitro and in vivo, whereas miR-1 vascular levels are negligible. miR-133 has a potent inhibitory role on VSMC phenotypic switch in vitro and in vivo, whereas miR-1 does not have any relevant effect per se. miR-133 expression is regulated by extracellular signal-regulated kinase 1/2 activation and is inversely correlated with VSMC growth. Indeed, miR-133 decreases when VSMCs are primed to proliferate in vitro and following vascular injury in vivo, whereas it increases when VSMCs are coaxed back to quiescence in vitro and in vivo. miR-133 loss- and gain-of-function experiments show that miR-133 plays a mechanistic role in VSMC growth. Accordingly, adeno-miR-133 reduces but anti-miR-133 exacerbates VSMC proliferation and migration in vitro and in vivo. miR-133 specifically suppresses the transcription factor Sp-1 expression in vitro and in vivo and through Sp-1 repression regulates smooth muscle gene expression. Our data show that miR-133 is a key regulator of vascular smooth muscle cell phenotypic switch in vitro and in vivo, suggesting its potential therapeutic application for vascular diseases.

The synergy of honokiol and fluconazole against clinical isolates of azole-resistant Candida albicans.

PubMed

Jin, J; Guo, N; Zhang, J; Ding, Y; Tang, X; Liang, J; Li, L; Deng, X; Yu, L

2010-09-01

To evaluate the interaction of fluconazole (FLC) and honokiol (HNK) in vitro and vivo against azole-resistant (azole-R) clinical isolates of Candida albicans. A checkerboard microdilution method was used to study the in vitro interaction of FLC and HNK in 24 azole-R clinical isolates of C. albicans. In vivo antifungal activity was performed to further analyse the interaction between FLC and HNK. In the in vitro study, synergism was observed in all 24 FLC-resistant strains tested as determined by fractional inhibitory concentration index (FICI), and in 22 strains by Delta E models. No antagonistic activity was observed in any of the strains tested. These positive interactions were also confirmed by using the time-killing test for the selected strain C. albicans YL371, which shows strong susceptible to the combination of HNK and FLC. In the in vivo study, the mice with candidiasis were treated successfully by a combination therapy of HNK with FLC, the results showed a decrease of the colony forming unit in infected and treated animals compared to the controls, at the conditions of the treatment used in this study. Synergistic activity of HNK and FLC against clinical isolates of FLC-resistant C. albicans was observed in vitro and in vivo. This report might provide a potential therapeutic method to overcome the problem of drug-resistance in C. albicans.

Comparative evaluation of rivastigmine permeation from a transdermal system in the Franz cell using synthetic membranes and pig ear skin with in vivo-in vitro correlation.

PubMed

Simon, Alice; Amaro, Maria Inês; Healy, Anne Marie; Cabral, Lucio Mendes; de Sousa, Valeria Pereira

2016-10-15

In the present study, in vitro permeation experiments in a Franz diffusion cell were performed using different synthetic polymeric membranes and pig ear skin to evaluate a rivastigmine (RV) transdermal drug delivery system. In vitro-in vivo correlations (IVIVC) were examined to determine the best model membrane. In vitro permeation studies across different synthetic membranes and skin were performed for the Exelon(®) Patch (which contains RV), and the results were compared. Deconvolution of bioavailability data using the Wagner-Nelson method enabled the fraction of RV absorbed to be determined and a point-to-point IVIVC to be established. The synthetic membrane, Strat-M™, showed a RV permeation profile similar to that obtained with pig ear skin (R(2)=0.920). Studies with Strat-M™ resulted in a good and linear IVIVC (R(2)=0.991) when compared with other synthetic membranes that showed R(2) values less than 0.90. The R(2) for pig ear skin was 0.982. Strat-M™ membrane was the only synthetic membrane that adequately simulated skin barrier performance and therefore it can be considered to be a suitable alternative to human or animal skin in evaluating transdermal drug transport, potentially reducing the number of studies requiring human or animal samples. Copyright © 2016 Elsevier B.V. All rights reserved.

Development and evaluation of nanoparticles based on mPEG-PLA for controlled delivery of vinpocetine: in vitro and in vivo studies.

PubMed

Wang, Run; Xu, Yong

2017-02-01

The aim of present study was to develop VIN-loaded mPEG-PLA nanoparticle systems. The VIN mPEG-PLA nanoparticles were prepared using an emulsion solvent evaporation method, and studied their particle size, morphology, encapsulation efficiency and drug-loading coefficient. Moreover, the nanoparticles were evaluated on the drug release behaviors in vitro and bioavailability in vivo. The results show that the spherical nanoparticles obtained were negatively charged with a zeta potential of about -23.4 mV and characterized ∼110 nm with a narrow size distribution. The encapsulation efficiency and drug loading of prepared NPs were 76.4 ± 6.3 and 9.2 ± 2.2% (n=5), respectively. The in vitro release showed that the percent of accumulated dissolution of VIN NPs in phosphate-buffered saline 6.8 over 24 h was <80%, which was almost 100% of VIN in commercial injections. The in vivo study indicated that systemic absorption of VIN was significantly enhanced by incorporating into mPEG-PLA NPs compared with VIN injection (2.87-fold in AUC 0- t ). The results suggested that the form of VIN in mPEG-PLA NPs could enter the body circulation to perform sustained release in vitro and in vivo.

Eugenia dysenterica DC. (Myrtaceae) exerts chemopreventive effects against hexavalent chromium-induced damage in vitro and in vivo.

PubMed

Ávila, Renato Ivan de; Mattos Alvarenga, Cátia Belo; Ávila, Paulo Henrique Marcelino de; Moreira, Roger Cardoso; Arruda, Andréa Fernandes; Fernandes, Thaís de Oliveira; Rodrigues, Bruna Dos Santos; Andrade, Wanessa Machado; Batista, Aline Carvalho; Paula, José Realino de; Valadares, Marize Campos

2016-11-01

Eugenia dysenterica DC. (Myrtaceae) has been widely used in the folk medicine and it presents phytochemicals constituents associated to antioxidant properties. The objective of this study was to investigate the protective effects of E. dysenterica leaf hydroalcoholic extract (EDE) in vitro and in vivo using AMJ2-C11 cells and Swiss mice exposed to hexavalent chromium [Cr(VI)], respectively. AMJ2-C11 cells were pretreated with EDE and exposed to Cr(VI) to evaluate cytotoxicity and the pathways involved in the chemopreventive effects of the extract. Mice were daily pretreated with EDE and then exposed to Cr(VI). Survival analysis, histopathological examination and determination of Cr levels in biological tissues were carried out. In vitro studies showed that pretreatment of the AMJ2-C11 cells with EDE protected against the cytotoxicity and oxidative stress induced by Cr(VI). Consequently, the pretreatment with EDE reduced reactive oxygen species and apoptosis triggered by Cr(VI), probably by a marked antioxidant and chelating activities demonstrated by EDE. Regarding in vivo studies, pretreatment for 10 days with EDE increased survival of the mice exposed to Cr(VI). In addition, EDE prevented liver and kidney pathological damages, in parallel with reduction in chromium levels found in these organs and plasma. EDE also showed a marked antioxidant potential associated with the presence of polyphenols, especially flavonoids and tannins, as confirmed by HPLC-PDA. The study showed that EDE protects against Cr(VI)-induced damage in vitro and in vivo supporting further studies for the development of therapeutic products applied to prevent the damage induced by toxic metals, especially Cr(VI).

Expanded adipose-derived stem cells suppress mixed lymphocyte reaction by secretion of prostaglandin E2.

PubMed

Cui, Lei; Yin, Shuo; Liu, Wei; Li, Ningli; Zhang, Wenjie; Cao, Yilin

2007-06-01

Multipotent mesenchymal stem cells (MSCs) in adult tissue are known to be less immunogenic and immunosuppressive. Previous study showed that primary cultures of human adipose-derived stem cells (ADSCs) shared their immunomodulatory properties with other MSCs. However, whether passaged human ADSCs can retain their immunomodulatory effect after in vitro expansion remains unknown. In addition, the mechanism of ADSC-mediated immunomodulatory effect remains to be elucidated. This study aimed to investigate these issues by using passaged human ADSCs as an in vitro study model. Flow cytometry showed that passaged ADSCs expressed human leukocyte antigen (HLA) class I but not class II molecules, which could be induced to express to a high level with interferon-gamma (IFN-gamma) treatment. The study found that passaged ADSCs could not elicit lymphocyte proliferation after co-culturing with them, even after IFN-gamma treatment. In addition, either IFN-gamma-treated or non-treated ADSCs could inhibit phytohemagglutinin (PHA)-stimulated lymphocyte proliferation. Moreover, passaged ADSCs could serve as the third-party cells to inhibited two-way mixed lymphocyte reaction (MLR). Further study using a transwell system also showed that this type of immunosuppressive effect was not cell-cell contact dependent. In defining possible soluble factors, we found that passaged ADSCs significantly increased their secretion of prostaglandin E2 (PGE2), but not transforming growth factor-beta (TGF-beta) and hepatocyte growth factor (HGF), when they were co-cultured with MLR. Furthermore, the result demonstrated that only PGE2 production inhibitor indomethacine, but not TGF-beta- and HGF-neutralizing antibodies, could significantly counteract ADSC-mediated suppression on allogeneic lymphocyte proliferation. These results indicated that in vitro expanded ADSCs retain low immunogenicity and immunosuppressive effect, and PGE2 might be the major soluble factor involved in the in vitro inhibition of allogeneic lymphocyte reaction.

Dissofilm: A Novel Approach for Delivery of Phenobarbital; Design and Characterization

PubMed Central

Yellanki, SK; Jagtap, S; Masareddy, R

2011-01-01

An attempt to develop and evaluate mouth-dissolving film of phenobarbital for quick effect in treatment of epilepsy occurring in pediatric population has been made in the present study. Suitable film formers and plasticizers are selected based on optimization studies. Effect of superdisintegrants in formulation of mouth dissolving films at different concentrations has been investigated. Films were prepared by solvent casting method. The prepared films were evaluated for physicochemical parameters, in vitro disintegration and dissolution time, in vitro release rate study, stability study, and in vivo animal safety study. The best formulation was found to be F3 showing 96.57% drug release in 14 min, following first-order kinetics. X-Ray diffraction studies show change in crystalline nature of drug in formulation. In vivo studies in hamster reports effective and safe use of formulation in animals. PMID:21897656

Transfer of in vitro expanded T lymphocytes after activation with dendritomas prolonged survival of mice challenged with EL4 tumor cells.

PubMed

Li, Jinhua; Theofanous, Leigh; Stickel, Sara; Bouton-Verville, Hilary; Burgin, Kelly E; Jakubchak, Susan; Wagner, Thomas E; Wei, Yanzhang

2007-07-01

Adoptive T cell transfer after in vitro expansion represents an attractive cancer immunotherapy. The majority of studies so far have been focusing on the expansion of tumor infiltrated lymphocytes (TIL) and some have shown very encouraging results. Recently, we have developed a unique tumor immune response activator, dendritomas, by fusion of dendritic cells and tumor cells. Animal studies and early clinical trials have shown that dendritomas are able to activate tumor specific immune responses. In this study, we hypothesized that naïve T cells can be primed with dendritomas and expanded in vitro to develop an adoptive transfer therapy for patients who do not have solid tumors, such as leukemia. T cells were isolated and purified from lymph nodes of mice. The cells were then incubated with dendritomas made from syngeneic DCs and tumor cells and expanded in vitro using Dynabeads mouse CD3/CD28 T cell expander for approximately three weeks. The in vitro primed and expanded T cells showed tumor cell specific CTL activity and increased secretion of IFN-gamma. Tumor bearing mice receiving the in vitro expanded T cells survived significantly longer than control mice. Furthermore, the depletion of regulator T cells enhanced the survival of the mice that received the adoptive transfer therapy.

A new extract of the plant calendula officinalis produces a dual in vitro effect: cytotoxic anti-tumor activity and lymphocyte activation

PubMed Central

Jiménez-Medina, Eva; Garcia-Lora, Angel; Paco, Laura; Algarra, Ignacio; Collado, Antonia; Garrido, Federico

2006-01-01

Background Phytopharmacological studies of different Calendula extracts have shown anti-inflamatory, anti-viral and anti-genotoxic properties of therapeutic interest. In this study, we evaluated the in vitro cytotoxic anti-tumor and immunomodulatory activities and in vivo anti-tumor effect of Laser Activated Calendula Extract (LACE), a novel extract of the plant Calendula Officinalis (Asteraceae). Methods An aqueous extract of Calendula Officinalis was obtained by a novel extraction method in order to measure its anti-tumor and immunomodulatory activities in vitro. Tumor cell lines derived from leukemias, melanomas, fibrosarcomas and cancers of breast, prostate, cervix, lung, pancreas and colorectal were used and tumor cell proliferation in vitro was measured by BrdU incorporation and viable cell count. Effect of LACE on human peripheral blood lymphocyte (PBL) proliferation in vitro was also analyzed. Studies of cell cycle and apoptosis were performed in LACE-treated cells. In vivo anti-tumor activity was evaluated in nude mice bearing subcutaneously human Ando-2 melanoma cells. Results The LACE extract showed a potent in vitro inhibition of tumor cell proliferation when tested on a wide variety of human and murine tumor cell lines. The inhibition ranged from 70 to 100%. Mechanisms of inhibition were identified as cell cycle arrest in G0/G1 phase and Caspase-3-induced apoptosis. Interestingly, the same extract showed an opposite effect when tested on PBLs and NKL cell line, in which in vitro induction of proliferation and activation of these cells was observed. The intraperitoneal injection or oral administration of LACE extract in nude mice inhibits in vivo tumor growth of Ando-2 melanoma cells and prolongs the survival day of the mice. Conclusion These results indicate that LACE aqueous extract has two complementary activities in vitro with potential anti-tumor therapeutic effect: cytotoxic tumor cell activity and lymphocyte activation. The LACE extract presented in vivo anti-tumoral activity in nude mice against tumor growth of Ando-2 melanoma cells. PMID:16677386

Impact of Humidity on In Vitro Human Skin Permeation Experiments for Predicting In Vivo Permeability.

PubMed

Ishida, Masahiro; Takeuchi, Hiroyuki; Endo, Hiromi; Yamaguchi, Jun-Ichi

2015-12-01

In vitro skin permeation studies have been commonly conducted to predict in vivo permeability for the development of transdermal therapeutic systems (TTSs). We clarified the impact of humidity on in vitro human skin permeation of two TTSs having different breathability and then elucidated the predictability of in vivo permeability based on in vitro experimental data. Nicotinell(®) TTS(®) 20 and Frandol(®) tape 40mg were used as model TTSs in this study. The in vitro human skin permeation experiments were conducted under humidity levels similar to those used in clinical trials (approximately 50%) as well as under higher humidity levels (approximately 95%). The skin permeability values of drugs at 95% humidity were higher than those at 50% humidity. The time profiles of the human plasma concentrations after TTS application fitted well with the clinical data when predicted based on the in vitro permeation parameters at 50% humidity. On the other hand, those profiles predicted based on the parameters at 95% humidity were overestimated. The impact of humidity was higher for the more breathable TTS; Frandol(®) tape 40mg. These results show that in vitro human skin permeation experiments should be investigated under realistic clinical humidity levels especially for breathable TTSs. © 2015 Wiley Periodicals, Inc. and the American Pharmacists Association.

Carboxylated nanodiamonds can be used as negative reference in in vitro nanogenotoxicity studies.

PubMed

Moche, H; Paget, V; Chevalier, D; Lorge, E; Claude, N; Girard, H A; Arnault, J C; Chevillard, S; Nesslany, F

2017-08-01

Nanodiamonds (NDs) are promising nanomaterials for biomedical applications. However, a few studies highlighted an in vitro genotoxic activity for detonation NDs, which was not evidenced in one of our previous work quantifying γ-H2Ax after 20 and 100 nm high-pressure high-temperature ND exposures of several cell lines. To confirm these results, in the present work, we investigated the genotoxicity of the same 20 and 100 nm NDs and added intermediate-sized NDs of 50 nm. Conventional in vitro genotoxicity tests were used, i.e., the in vitro micronucleus and comet assays that are recommended by the French National Agency for Medicines and Health Products Safety for the toxicological evaluation of nanomedicines. In vitro micronucleus and in vitro comet assays (standard and hOGG1-modified) were therefore performed in two human cell lines, the bronchial epithelial 16HBE14o- cells and the colon carcinoma T84 cells. Our results did not show any genotoxic activity, whatever the test, the cell line or the size of carboxylated NDs. Even though these in vitro results should be confirmed in vivo, they reinforce the potential interest of carboxylated NDs for biomedical applications or even as a negative reference nanoparticle in nanotoxicology. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

Comparison of 99mTc-UBI 29-41, 99mTc-Ciprofloxacin, 99mTc-Ciprofloxacin dithiocarbamate and 111In-biotin for targeting experimental Staphylococcus aureus and Escherichia coli foreign-body infections: an ex-vivo study.

PubMed

Auletta, Sveva; Baldoni, Daniela; Varani, Michela; Galli, Filippo; Hajar, Iman A; Duatti, Adriano; Ferro-Flores, Guillermina; Trampuz, Andrej; Signore, Alberto

2017-08-28

Diagnosis of implant-associated infection is challenging. Several radiopharmaceuticals have been described but direct comparisons are limited. Here we compared in vitro and in an animal model 99mTc-UBI, 99mTc-Ciprofloxacin, 99mTcN-CiproCS2 and 111In-DTPA-biotin for targeting E. coli (ATCC 25922) and S. aureus (ATCC 43335). Stability controls were performed with the labelled radiopharmaceuticals during 6 h in saline and serum. The in vitro binding to viable or killed bacteria was evaluated at 37 °C and 4 °C. For in vivo studies, Teflon cages were subcutaneously implanted in mice, followed by percutaneous infection. Biodistribution of i.v. injected radiolabelled radiopharmaceuticals were evaluated during 24 h in cages and dissected tissues. Labelling efficiency of all radiopharmaceuticals ranged between 94% and 98%, with high stability both in saline and in human serum. In vitro binding assays displayed a rapid but poor bacterial binding for all tested agents. Similar binding kinetic occurred also with heat-killed and ethanol-killed bacteria. In the tissue cage model, infection was detected at different time points: 99mTc-UBI and 99mTcN-CiproCS2 showed higher infected cage/sterile cage ratio at 24 h for both E. coli and S. aureus; 99mTc-Ciprofloxacin at 24 h for both E. coli and at 4 h for S. aureus; 111In-DTPA-biotin accumulates faster in both E. coli and S. aureus infected cages. 99mTc-UBI, 99mTcN-CiproCS2 showed poor in vitro binding but good in vivo binding to E. coli only. 111In-DTPA-biotin showed poor in vitro binding but good in vivo binding to S. aureus and poor to E. coli. 99mTc-Ciprofloxacin showed poor in vitro binding but good in vivo binding to all tested bacteria. The mechanism of accumulation in infected sites remains to be elucidated.

Chromosomal aberrations and deoxyribonucleic acid single-strand breaks in adipose-derived stem cells during long-term expansion in vitro.

PubMed

Froelich, Katrin; Mickler, Johannes; Steusloff, Gudrun; Technau, Antje; Ramos Tirado, Mario; Scherzed, Agmal; Hackenberg, Stephan; Radeloff, Andreas; Hagen, Rudolf; Kleinsasser, Norbert

2013-07-01

Adipose-derived stem cells (ASCs) are a promising mesenchymal cell source for tissue engineering approaches. To obtain an adequate cell amount, in vitro expansion of the cells may be required in some cases. To monitor potential contraindications for therapeutic applications in humans, DNA strand breaks and chromosomal aberrations in ASCs during in vitro expansion were examined. After isolation of ASC from human lipoaspirates of seven patients, in vitro expansion over 10 passages was performed. Cells from passages 1, 2, 3, 5 and 10 were used for the alkaline single-cell microgel electrophoresis (comet) assay to detect DNA single-strand breaks and alkali labile as well as incomplete excision repair sites. Chromosomal changes were examined by means of the chromosomal aberration test. During in vitro expansion, ASC showed no DNA single-strand breaks in the comet assay. With the chromosomal aberration test, however, a significant increase in chromosomal aberrations were detected. The study showed that although no DNA fragmentation could be determined, the safety of ASC cannot be ensured with respect to chromosome stability during in vitro expansion. Thus, reliable analyses for detecting ASC populations, which accumulate chromosomal aberrations or even undergo malignant transformation during extensive in vitro expansion, must be implemented as part of the safety evaluation of these cells for stem cell-based therapy. Copyright © 2013 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

In Situ and In Vitro Effects of Two Bleaching Treatments on Human Enamel Hardness.

PubMed

Henn-Donassollo, Sandrina; Fabris, Cristiane; Gagiolla, Morgana; Kerber, Ícaro; Caetano, Vinícius; Carboni, Vitor; Salas, Mabel Miluska Suca; Donassollo, Tiago Aurélio; Demarco, Flávio Fernando

2016-01-01

The aim of this study was to evaluate in vitro and in situ the effects of two bleaching treatments on human enamel surface microhardness. Sixty enamel slabs from recently extracted thirty molars were used. The specimens were polished with sandpapers under water-cooling. The enamel samples were randomly divided in four groups, treated with 10% hydrogen peroxide (HP) or Whitening Strips (WS) containing 10% hydrogen peroxide and using two conditions: in vitro or in situ model. For in situ condition, six volunteers wore an intra-oral appliance containing enamel slabs, while for in vitro condition the specimens were kept in deionized water after the bleaching protocols. The bleaching treatments were applied one-hour daily for 14 days. Similar amounts of bleaching agents were used in both conditions. Before and after bleaching treatments, microhardness was measured. Statistical analysis (ANOVA and Tukey test) showed that in the in situ condition there was no statistically significant microhardness reduction in the bleached enamel (p>0.05). Significant decrease in hardness was observed for enamel slabs bleached with both treatments in the in vitro condition (p<0.05). Regarding the bleaching agents, in situ results showed no difference between HP and WS, while in vitro WS produced the lowest hardness value. It could be concluded that there was no deleterious effect on enamel produced by any of the bleaching protocols used in the in situ model. The reduction of hardness was only observed in vitro.

Techniques to Evaluate Dental Erosion: A Systematic Review of Literature.

PubMed

Joshi, Mahasweta; Joshi, Nikhil; Kathariya, Rahul; Angadi, Prabhakar; Raikar, Sonal

2016-10-01

This article reviews different techniques for evaluating dental erosion, weighs the advantages and disadvantages of these techniques, and presents the latest trends in the study of dental erosion. In May 2014, an initial search was carried out in the PubMed/MEDLINE database of indexed journals from 1975 to 2013 using the following keywords: dental erosion; dental erosion In-vitro; and dental erosion in-vivo. Bibliographic citations from the papers found were then used to find other useful sources. The authors categorize the techniques into three classes: in-vitro, in-vivo and in-vitro/in-vivo. The article discusses the instrumentation required to use each of these techniques, as well as their rationale, merits and applications. The emergence of in-vitro/in-vivo techniques offers the potential to accurately quantify tooth wear in clinical situations. Cross-sectional as well as longitudinal studies show that these techniques will improve diagnosis, treatment planning and management of dental erosion.

In vitro growth inhibition and cytotoxicity of Euphorbia caducifolia against four human cancer cell lines and its phytochemical characterisation.

PubMed

Bano, Shaista; Siddiqui, Bina Shaheen; Farooq, Ahsana Dar; Begum, Sabira; Siddiqui, Faheema; Kashif, Muhammad; Azhar, Mudassar

2017-12-01

Several Euphorbia species have been used in folklore as cancer remedies, however, scientific studies on the cytotoxicity (in vitro studies) of Euphorbia caducifolia are lacking. In present study, anticancer potential of E. caducifolia aerial parts ethanol extract and its fractions were evaluated against human lung (NCI-H460), breast (MCF-7), prostate (PC-3) and cervical (HeLa) cancer cell lines, using sulphorhodamine-B in vitro cytotoxicity (in vitro studies) assay. The ethanol extract demonstrated growth inhibitory effect against all aforementioned cancer cell lines with IC 50 , 19-135 μg/mL and LC 50 , ~220 μg/mL, and its petroleum ether fraction obtained on bioactivity guided fraction showed highest activity with IC 50 , 28-70 μg/mL and LC 50 , 71 μg/mL against NCI-H460 and MCF-7 cell lines. Its phytochemicals were analysed by gas chromatography-mass spectrometry (GC-MS). The present study provides scientific justification for its traditional use against cancer.

Loss of Optineurin In Vivo Results in Elevated Cell Death and Alters Axonal Trafficking Dynamics

PubMed Central

Paulus, Jeremiah D.; Link, Brian A.

2014-01-01

Mutations in Optineurin have been associated with ALS, glaucoma, and Paget’s disease of bone in humans, but little is known about how these mutations contribute to disease. Most of the cellular consequences of Optineurin loss have come from in vitro studies, and it remains unclear whether these same defects would be seen in vivo. To answer this question, we assessed the cellular consequences of Optineurin loss in zebrafish embryos to determine if they showed the same defects as have been described in the in vitro studies. We found that loss of Optineurin resulted in increased cell death, as well as subtle cell morphology, cell migration and vesicle trafficking defects. However, unlike experiments on cells in culture, we found no indication that the Golgi apparatus was disrupted or that NF-κB target genes were upregulated. Therefore, we conclude that in vivo loss of Optineurin shows some, but not all, of the defects seen in in vitro work. PMID:25329564

Antibacterial Activity of the Alkaloid-Enriched Extract from Prosopis juliflora Pods and Its Influence on in Vitro Ruminal Digestion

PubMed Central

dos Santos, Edilene T.; Pereira, Mara Lúcia A.; da Silva, Camilla Flávia P.G.; Souza-Neta, Lourdes C.; Geris, Regina; Martins, Dirceu; Santana, Antônio Euzébio G.; Barbosa, Luiz Cláudio A.; Silva, Herymá Giovane O.; Freitas, Giovana C.; Figueiredo, Mauro P.; de Oliveira, Fernando F.; Batista, Ronan

2013-01-01

The purpose of this study was to assess the in vitro antimicrobial activity of alkaloid-enriched extracts from Prosopis juliflora (Fabaceae) pods in order to evaluate them as feed additives for ruminants. As only the basic chloroformic extract (BCE), whose main constituents were juliprosopine (juliflorine), prosoflorine and juliprosine, showed Gram-positive antibacterial activity against Micrococcus luteus (MIC = 25 μg/mL), Staphylococcus aureus (MIC = 50 μg/mL) and Streptococcus mutans (MIC = 50 μg/mL), its influence on ruminal digestion was evaluated using a semi-automated in vitro gas production technique, with monensin as the positive control. Results showed that BCE has decreased gas production as efficiently as monensin after 36 h of fermentation, revealing its positive influence on gas production during ruminal digestion. Since P. juliflora is a very affordable plant, this study points out this alkaloid enriched extract from the pods of Prosopis juliflora as a potential feed additive to decrease gas production during ruminal digestion. PMID:23595000

Synthesis, crystal structures, molecular docking, and in vitro biological activities evaluation of transition metal complexes with 4-(3,4-dichlorophenyl) piperazine-1-carboxylic acid

NASA Astrophysics Data System (ADS)

Chen, Zhi-Jian; Chen, Ya-Na; Xu, Chun-Na; Zhao, Shan-Shan; Cao, Qi-Yue; Qian, Shao-Song; Qin, Jie; Zhu, Hai-Liang

2016-08-01

Three novel mononuclear complexes, [MⅡ(L)2·2H2O], (M = Cu, Ni or Cd; HL = 4-(3,4-dichlorophenyl)piperazine-1-carboxylic acid)were synthesized and structurally determined by single-crystal X-ray diffraction. Molecular docking study preliminarily revealed that complex 1 had potential urease inhibitory activity. In accordance with the result of calculation, in vitro tests of the inhibitory activities of complexes 1-3 against jack bean urease showed complex 1 (IC50 = 8.17 ± 0.91 μM) had better inhibitory activities than the positive reference acetohydroxamic acid (AHA) (IC50 = 26.99 ± 1.43 μM), while complexes 2 and 3 showed no inhibitory activities., kinetics study was carried out to explore the mechanism of the inhibiting of the enzyme, and the result indicated that complex 1 was a competitive inhibitor of urease. Albumin binding experiment and in vitro toxicity evaluation of complex 1 were implemented to explore its Pharmacological properties.

Influence of the Fruit Juice Carriers on the Ability of Lactobacillus plantarum DSM20205 to Improve in Vitro Intestinal Barrier Integrity and Its Probiotic Properties.

PubMed

Valero-Cases, Estefanía; Roy, Nicole C; Frutos, María José; Anderson, Rachel C

2017-07-19

This study investigates the influence of tomato and feijoa juices as fermentable carriers of Lactobacillus plantarum (LP DSM20205) on the ability of the bacterium to improve intestinal barrier function using the trans-epithelial electrical resistance (TEER) assay in an apical anaerobic model. The survival of LP DSM20205 in different fruit juices during in vitro digestion, its adhesion capacity, and potential cytotoxic effect on Caco-2 cells were also studied. The results showed that carrier fruit juices have a significant influence on LP DSM20205 growth, survival during in vitro digestion, adhesion capacity, and TEER. All fermented samples were not cytotoxic to Caco-2 cells. The fermented tomato juice showed the largest improvement to intestinal barrier integrity. The digested fermented juices did not increase TEER, although the LP DSM20205 in these samples adhered well. Therefore, LP DSM20205 has the potential to be used as a probiotic in the production of fermented tomato and feijoa juices.

Multifunctional Cinnamic Acid Derivatives.

PubMed

Peperidou, Aikaterini; Pontiki, Eleni; Hadjipavlou-Litina, Dimitra; Voulgari, Efstathia; Avgoustakis, Konstantinos

2017-07-25

Our research to discover potential new multitarget agents led to the synthesis of 10 novel derivatives of cinnamic acids and propranolol, atenolol, 1-adamantanol, naphth-1-ol, and (benzylamino) ethan-1-ol. The synthesized molecules were evaluated as trypsin, lipoxygenase and lipid peroxidation inhibitors and for their cytotoxicity. Compound 2b derived from phenoxyphenyl cinnamic acid and propranolol showed the highest lipoxygenase (LOX) inhibition (IC 50 = 6 μΜ) and antiproteolytic activity (IC 50 = 0.425 μΜ). The conjugate 1a of simple cinnamic acid with propranolol showed the higher antiproteolytic activity (IC 50 = 0.315 μΜ) and good LOX inhibitory activity (IC 50 = 66 μΜ). Compounds 3a and 3b , derived from methoxylated caffeic acid present a promising combination of in vitro inhibitory and antioxidative activities. The S isomer of 2b also presented an interesting multitarget biological profile in vitro . Molecular docking studies point to the fact that the theoretical results for LOX-inhibitor binding are identical to those from preliminary in vitro study.

A Study on the Quality and Identity of Brazilian Pampa Biome Honey: Evidences for Its Beneficial Effects against Oxidative Stress and Hyperglycemia

PubMed Central

Cruz, L. C.; Batista, J. E. S.; Zemolin, A. P. P.; Nunes, M. E. M.; Lippert, D. B.; Royes, L. F. F.; Soares, F. A.; Pereira, A. B.; Posser, T.; Franco, J. L.

2014-01-01

We characterized, for the first time, the quality and identity of Brazilian Pampa biome honey and its antioxidant properties in vitro (FRAP, DDPH and ABTS). The potential protective effect of honey against oxidative stress induced by iron (Fe) and paraquat, (PQ) in a Drosophila melanogaster model (in vivo) was also tested. The results indicated that all honey samples tested showed antioxidant activity in vitro. Flies treated with honey showed increased lifespan and were protected against oxidative stress induced by Fe and PQ. Despite the high concentration of sugars in honey (approximately 70–80%), our results demonstrate a hypoglycemic-like effect of honey in Drosophila. Thus, this study demonstrates the high quality of Brazilian Pampa biome honey as well as its significant antioxidant activity in vitro and in vivo, pointing to the potential use of this natural product as an alternative in the therapy of oxidative stress-associated diseases. PMID:26904632

A Study on the Quality and Identity of Brazilian Pampa Biome Honey: Evidences for Its Beneficial Effects against Oxidative Stress and Hyperglycemia.

PubMed

Cruz, L C; Batista, J E S; Zemolin, A P P; Nunes, M E M; Lippert, D B; Royes, L F F; Soares, F A; Pereira, A B; Posser, T; Franco, J L

2014-01-01

We characterized, for the first time, the quality and identity of Brazilian Pampa biome honey and its antioxidant properties in vitro (FRAP, DDPH and ABTS). The potential protective effect of honey against oxidative stress induced by iron (Fe) and paraquat, (PQ) in a Drosophila melanogaster model (in vivo) was also tested. The results indicated that all honey samples tested showed antioxidant activity in vitro. Flies treated with honey showed increased lifespan and were protected against oxidative stress induced by Fe and PQ. Despite the high concentration of sugars in honey (approximately 70-80%), our results demonstrate a hypoglycemic-like effect of honey in Drosophila. Thus, this study demonstrates the high quality of Brazilian Pampa biome honey as well as its significant antioxidant activity in vitro and in vivo, pointing to the potential use of this natural product as an alternative in the therapy of oxidative stress-associated diseases.

The novel iron chelator, 2-pyridylcarboxaldehyde 2-thiophenecarboxyl hydrazone, reduces catecholamine-mediated myocardial toxicity.

PubMed

Mladĕnka, Premysl; Kalinowski, Danuta S; Haskova, Pavlína; Bobrovová, Zuzana; Hrdina, Radomír; Simůnek, Tomás; Nachtigal, Petr; Semecký, Vladimĺr; Vávrová, Jaroslava; Holeckova, Magdaléna; Palicka, Vladimir; Mazurová, Yvona; Jansson, Patric J; Richardson, Des R

2009-01-01

Iron (Fe) chelators are used clinically for the treatment of Fe overload disease. Iron also plays a role in the pathology of many other conditions, and these potentially include the cardiotoxicity induced by catecholamines such as isoprenaline (ISO). The current study examined the potential of Fe chelators to prevent ISO cardiotoxicity. This was done as like other catecholamines, ISO contains the classical catechol moiety that binds Fe and may form redox-active and cytotoxic Fe complexes. Studies in vitro used the cardiomyocyte cell line, H9c2, which was treated with ISO in the presence or absence of the chelator, desferrioxamine (DFO), or the lipophilic ligand, 2-pyridylcarboxaldehyde 2-thiophenecarboxyl hydrazone (PCTH). Both of these chelators were not cardiotoxic and significantly reduced ISO cardiotoxicity in vitro. However, PCTH was far more effective than DFO, with the latter showing activity only at a high, clinically unachievable concentration. Further studies in vitro showed that interaction of ISO with Fe(II)/(III) did not increase cytotoxic radical generation, suggesting that this mechanism was not involved. Studies in vivo were initiated using rats pretreated intravenously with DFO or PCTH before subcutaneous administration of ISO (100 mg/kg). DFO at a clinically used dose (50 mg/kg) failed to reduce catecholamine cardiotoxicity, while PCTH at an equimolar dose totally prevented catecholamine-induced mortality and reduced cardiotoxicity. This study demonstrates that PCTH reduced ISO-induced cardiotoxicity in vitro and in vivo, demonstrating that Fe plays a role, in part, in the pathology observed.

In vitro repellent effect of tea tree (Melaleuca alternifolia) and andiroba (Carapa guianensis) oils on Haemotobia irritans and Chrysomya megacephala flies.

PubMed

Klauck, V; Pazinato, R; Radavelli, W M; Volpato, A; Stefani, L M; Santos, R C V; Vaucher, R A; Boligon, A A; Athayde, M L; Da Silva, A S

2015-03-01

This study aimed to evaluate the repellent effect of tea tree (Melaleuca alternifolia) and andiroba (Carapa guianensis) essential oils on two species of flies (Haemotobia irritans and Chrysomya megacephala). For the in vitro studies, free-living adult flies were captured and reared in the laboratory. To verify the repellency effect, an apparatus was constructed where H. irritans and C. megacephala were exposed to andiroba and tea tree oils (5.0%), as well as to a known repellent (citronella, 5.0%) to validate the test. The study demonstrated that all three oils used showed in vitro repellent effect against both species of flies. It is possible to conclude that the essential oils (tea tree and andiroba) have repellent effect on these species of flies used in this study.

Development of an in vitro cell culture model to study milk to plasma ratios of therapeutic drugs.

PubMed

Athavale, Maithili A; Maitra, Anurupa; Patel, Shahnaz; Bhate, Vijay R; Toddywalla, Villi S

2013-01-01

To create an in vitro cell culture model to predict the M/P (concentration of drug in milk/concentration in maternal plasma) ratios of therapeutic drugs viz. rifampicin, theophylline, paracetamol, and aspirin. An in vitro cell culture model using CIT3 cells (mouse mammary epithelial cells) was created by culturing the cells on transwells. The cells formed an integral monolayer, allowing only transcellular transport as it happens in vivo. Functionality of the cells was confirmed through scanning electron microscopy. Time wise transfer of the study drugs from plasma to milk was studied and compared with actual (in vivo) M/P ratios obtained at reported tmax for the respective drugs. The developed model mimicked two important intrinsic factors of mammary epithelial cells viz. secretory and tight-junction properties and also the passive route of drug transport. The in vitro M/P ratios at reported tmax were 0.23, 0.61, 0.87, and 0.03 respectively, for rifampicin, theophylline, paracetamol, and salicylic acid as compared to 0.29, 0.65, 0.65, and 0.22, respectively, in vitro. Our preliminary effort to develop an in vitro physiological model showed promising results. Transfer rate of the drugs using the developed model compared well with the transfer potential seen in vivo except for salicylic acid, which was transferred in far lower concentration in vitro. The model has a potential to be developed as a non-invasive alternative to the in vitro technique for determining the transfer of therapeutic drugs into breast milk.

[Preclinical study of AV0012 early stage inhibitor of hepatitis C virus infection: I. In vitro ADME and pharmacokinetics].

PubMed

Ivashchenko, A V; Iamanushkin, P M; Mit'kin, O D; Ezhova, E V; Korzinov, O M; Shevkun, N A; Koriakova, A G; Karapetian, R N; Bychko, V V; Ivashchenko, A A; Agrba, V Z; Lapin, B A; Orlov, S V

2014-01-01

In vitro immunohistochemical investigations on the human hepatoma cell line (Huh7) infected with hepatitis C virus (HCV) strain JFH-1 showed that AV0012 compound blocks the early stages of viral infection. AV0012 also blocked viral infection spread in tissue culture through the secreted virus and through tight cell-to-cell contact. AV0012 is a specific inhibitor of HCV but not of related pestivirus, flaviviruses and other RNA-containing viruses such as bovine diarrhea (BVDV), Venezuelan equine encephalitis (strain TC-83), dengue type 2 (New Guinea), yellow fever (strain 17D), west Nile fever, parainfluenza (type 3) virus, RSV (strain A2), and Rhinovirus (type 2 strain HGP). It is established that human serum does not significantly affect the antiviral activity of AV0012 in vitro. The drug combination studies with AV0012 and interferon alpha 2a in vitro showed that the two inhibitors act additively, which makes possible the use of this combination in clinical tests. AV0012 is highly soluble and stable in aqueous solutions and murine blood plasma, has limited metabolic stability, low binding to human plasma proteins, high permeability through biological membranes, and only interacts with isoenzymes 2D6 and 3A4 of human cytochrome P450. In animal pharmacokinetic studies, AV0012 was rapidly absorbed into the blood stream upon oral administration, showed sufficiently long half-elimination times, and had high oral bioavailability that reached 92% in monkeys. Further preclinical development of AV0012 is in progress.

Mind "De GaPP": in vitro efficacy of deferiprone and gallium-protoporphyrin against Staphylococcus aureus biofilms.

PubMed

Richter, Katharina; Ramezanpour, Mahnaz; Thomas, Nicky; Prestidge, Clive A; Wormald, Peter-John; Vreugde, Sarah

2016-07-01

Biofilms are clusters of bacteria embedded in a protective matrix that frequently cause failure of medical treatments and increase the risk of recurrent infections. In particular, Staphylococcus aureus biofilms are associated with a series of chronic and nosocomial infections that are increasingly resistant to antibiotics. This study proposes a novel intervention strategy targeting the essential iron metabolism for bacterial growth, survival and pathogenesis using the compounds deferiprone (Def) and gallium-protoporphyrin (GaPP). S. aureus biofilms were challenged with Def/GaPP as single and dual treatments. In vitro anti-biofilm efficacy was assessed by the AlamarBlue viability assay and confocal microscopy. In vitro cytotoxicity of the treatments was examined by the lactate dehydrogenase assay on mouse fibroblast (L929) and human bronchial epithelial cells (Nuli-1). Def (20 mM) and GaPP (200 μg/mL) monotherapy for 2 hours showed 35% and 74% biofilm removal, respectively, whereas simultaneous Def/GaPP administration showed 55% biofilm removal. In contrast, the consecutive treatment (2 hours Def followed by 2 hours GaPP) achieved 95% biofilm removal. Cytotoxicity studies indicated no cell hazard in all treatments. This study demonstrated the in vitro efficacy of a novel treatment combination against S. aureus biofilms targeting the bacterial iron metabolism. The consecutive Def/GaPP treatment showed significantly enhanced biofilm efficacy than the individual compounds, while being not toxic to 2 cell lines. This novel treatment combination is a promising approach to combat S. aureus-associated biofilm infections having high potential for future clinical application. © 2016 ARS-AAOA, LLC.

Gravitational physiology of human immune cells: a review of in vivo, ex vivo and in vitro studies

NASA Technical Reports Server (NTRS)

Cogoli, A.

1996-01-01

The study of the function of immune cells in microgravity has been studied for more than 20 years in several laboratories. It is clear today that the immune system is depressed in more than 50% of the astronauts during and after space flight and that the activation of T lymphocytes by mitogens in vitro changes dramatically. This article gives an overview of the gravitational studies conducted by our laboratory in Spacelab, in MIR station, in sounding rockets and on the ground in the clinostat and the centrifuge. Three experimental approaches are followed in our work: (i) Ex vivo studies are performed with blood samples drawn from astronauts; (ii) in vivo studies are based on the application of seven antigens to the skin of the astronauts; (iii) in vitro studies are carried out with immune cells purified from the blood of healthy donors (not astronauts). The data from our in vivo and ex vivo studies are in agreement with those of other laboratories and show that the immunological function is depressed in the majority of astronauts as a consequence of the stress of space flight rather than by a direct influence of gravity on the cell. Immune depression may become a critical hazard on long duration flights on space stations or to other planets. In vitro experiments show that cultures of free-floating lymphocytes and monocytes undergo a dramatic depression of activation by the mitogen concanavalin A, while activation is more than doubled when the cells are attached to microcarrier beads. Such effects may be attributed to both direct and indirect effects of gravitational unloading on basic biological mechanisms of the cell. While the in vitro data are very important to clarify certain aspects of the biological mechanism of T cells activation, they are not descriptive of the changes of the immunological function of the astronauts.

Gravitational physiology of human immune cells: a review of in vivo, ex vivo and in vitro studies.

PubMed

Cogoli, A

1996-04-01

The study of the function of immune cells in microgravity has been studied for more than 20 years in several laboratories. It is clear today that the immune system is depressed in more than 50% of the astronauts during and after space flight and that the activation of T lymphocytes by mitogens in vitro changes dramatically. This article gives an overview of the gravitational studies conducted by our laboratory in Spacelab, in MIR station, in sounding rockets and on the ground in the clinostat and the centrifuge. Three experimental approaches are followed in our work: (i) Ex vivo studies are performed with blood samples drawn from astronauts; (ii) in vivo studies are based on the application of seven antigens to the skin of the astronauts; (iii) in vitro studies are carried out with immune cells purified from the blood of healthy donors (not astronauts). The data from our in vivo and ex vivo studies are in agreement with those of other laboratories and show that the immunological function is depressed in the majority of astronauts as a consequence of the stress of space flight rather than by a direct influence of gravity on the cell. Immune depression may become a critical hazard on long duration flights on space stations or to other planets. In vitro experiments show that cultures of free-floating lymphocytes and monocytes undergo a dramatic depression of activation by the mitogen concanavalin A, while activation is more than doubled when the cells are attached to microcarrier beads. Such effects may be attributed to both direct and indirect effects of gravitational unloading on basic biological mechanisms of the cell. While the in vitro data are very important to clarify certain aspects of the biological mechanism of T cells activation, they are not descriptive of the changes of the immunological function of the astronauts.

In vitro cellular adhesion and antimicrobial property of SiO2-MgO-Al2O3-K2O-B2O3-F glass ceramic.

PubMed

Kalmodia, Sushma; Molla, Atiar Rahaman; Basu, Bikramjit

2010-04-01

The aim of the present study was to examine the cellular functionality and antimicrobial properties of SiO(2)-MgO-Al(2)O(3)-K(2)O-B(2)O(3)-F glass ceramics (GC) containing fluorophlogopite as major crystalline phase. The cellular morphology and cell adhesion study using human osteoblast-like Saos-2 cells and mouse fibroblast L929 cells reveals good in vitro cytocompatibility of GC. The potential use of the GC for biomedical application was also assessed by in vitro synthesis of the alkaline phosphatase (ALP) activity of Saos-2 cells. It is proposed that B(2)O(3) actively enhances the cell adhesion and supports osteoconduction process, whereas, fluorine component significantly influences cell viability. The Saos-2 and L929 cells on GC shows extensive multidirectional network of actin cytoskeleton. The in vitro results of this study illustrate how small variation in fluorine and boron in base glass composition influences significantly the biocompatibility and antimicrobial bactericidal property, as evaluated using a range of biochemical assays. Importantly, it shows that the cell viability and osteoconduction can be promoted in glass ceramics with lower fluorine content. The underlying reasons for difference in biological properties are analyzed and reported. It is suggested that oriented crystalline morphology in the lowest fluorine containing glass ceramic enhanced cellular spreading. Overall, the in vitro cell adhesion, cell flattening, cytocompatibility and antimicrobial study of the three different compositions of glass ceramic clearly reveals that microstructure and base glass composition play an important role in enhancing the cellular functionality and antimicrobial property.

Comparative evaluation of the efficacy of a herbal mouthwash and chlorhexidine mouthwash on select periodontal pathogens: An in vitro and ex vivo study

PubMed Central

Pathan, Multazim Muradkhan; Bhat, Kishore Gajanan; Joshi, Vinayak Mahableshwar

2017-01-01

Background: Several herbal mouthwash and herbal extracts have been tested in vitro and in vivo in search of a suitable adjunct to mechanical therapy for long-term use. In this study, we aimed to look at the antimicrobial effect of the herbal mouthwash and chlorhexidine (CHX) mouthwash on select organisms in in vitro test and an ex vivo model. Materials and Methods: The antimicrobial effects were determined against standard strains of bacteria that are involved in different stages of periodontal diseases. The in vitro tests included determination of minimum inhibitory concentration (MIC) using broth dilution and agar diffusion. In the ex vivo part of the study supragingival dental plaque were obtained from 20 periodontally healthy adult volunteers. Descriptive analysis was done for the entire quantitative and qualitative variable recorded. Results: The MIC by broth dilution method found no statistically significant difference between the mouthwashes. The agar dilution method showed CHX was more effective as compared to the herbal mouthwash against standard strains of Streptococcus mutans, Streptococcus sanguinis, and Aggregatibacter actinomycetemcomitans. However, no difference was observed between the mouthwashes for Porphyromonas, Pseudomonas aeruginosa, and Fusobacterium nucleatum. The ex vivo results conclude that none of the selected mouthwashes were statistically significantly different from each other. Conclusion: In the present study, CHX showed higher levels of antimicrobial action than the herbal mouthwash against bacterial species. The results reinforce the earlier findings that the in vitro testing is sensitive to methods and due diligence is needed when extrapolating the data for further use. However, long-term use and in vivo effectiveness against the periopathogens need to be tested in well-planned clinical trials. PMID:29456300

Synergistic in vitro and in vivo antimicrobial effect of a mixture of ZnO nanoparticles and Lactobacillus fermentation liquor.

PubMed

Kuang, Huijuan; Yang, Lin; Shah, Nagendra P; Aguilar, Zoraida P; Wang, Lijun; Xu, Hengyi; Wei, Hua

2016-04-01

In this study, we investigated the antibacterial activity of ZnO nanoparticles (NPs) and Lactobacillus-fermentation liquor (LFL) against two pathogenic bacteria in vitro and in vivo. Bactericidal tests were performed on solid agar plates and quantitative real-time PCR (qPCR), and denaturing gradient gel electrophoresis (DGGE) techniques were used to examine the antibacterial activity of the mixture of ZnO NPs and LFL in vivo. The results showed that the mixture exhibited higher antibacterial activity against Salmonella typhimurium in vitro in comparison with ZnO NPs alone. The results showed that ZnO NPs and LFL significantly enhanced microbial diversity in mouse intestine which suggested a synergistic antibacterial activity against the tested pathogenic bacteria that could be used for the control of the spread and persistence of bacterial infections.

In Vitro Effects of Cooking Methods on Digestibility of Lipids and Formation of Cholesterol Oxidation Products in Pork

PubMed Central

Moon, Sung Sil

2014-01-01

This study investigated the effects of cooking methods on the digestibility of lipids and formation of cholesterol oxidation products (COPs) in pork, during in vitro human digestion. Pork patties were cooked using four different methods (oven cooking, pan frying, boiling, and microwaving), to an internal temperature of approximately 85℃. The digestibility of pork patties were then evaluated, using the in vitro human digestion model that simulated the composition (pH, minerals, surfaceactive components, and enzymes) of digestive juices in the human mouth, stomach, and small intestine. The total lipid digestibility was higher after microwave cooking, whereas pan-frying resulted in lower in vitro digestibility, compared to the other cooking methods. The microwaving method followed by in vitro digestion also showed significantly higher content of free fatty acids and thiobarbituric acid reactive substances (TBARS), compared to the other cooking methods; whereas, the pan frying and boiling methods showed the lowest. Cholesterol content was not significantly different among the cooked samples before, and after in vitro human digestion. The formation of COPs was significantly higher in the microwave-treated pork samples, compared to those cooked by the other methods, which was consistent with the trend for lipid peroxidation (TBARS). We propose that from the point of view of COPs formation and lipid oxidation, the pan-frying or boiling methods would be useful. PMID:26761168

Phosphate solubilization and chromium (VI) remediation potential of Klebsiella sp. strain CPSB4 isolated from the chromium contaminated agricultural soil.

PubMed

Gupta, Pratishtha; Kumar, Vipin; Usmani, Zeba; Rani, Rupa; Chandra, Avantika

2018-02-01

In this study, an effort was made to identify an efficient phosphate solubilizing bacterial strain from chromium contaminated agricultural soils. Based on the formation of a solubilized halo around the colonies on Pikovskaya's agar amended with chromium (VI), 10 strains were initially screened out. Out of 10, strain CPSB4, which showed significantly high solubilization zone at different chromium concentrations, was selected for further study. The strain CPSB4 showed significant plant growth promotion traits with chromium (VI) stress under in-vitro conditions in broth. The plant growth promotion activities of the strain decreased regularly, but were not completely lost with the increase in concentration of chromium up to 200 mg L -1 . On subjected to FT-IR analysis, the presence of the functional group, indicating the organic acid aiding in phosphate solubilization was identified. At an optimal temperature of 30  ° C and pH 7.0, the strain showed around 93% chromium (VI) reduction under in-vitro conditions in broth study. In soil condition, the maximum chromium (VI) reduction obtained was 95% under in-vitro conditions. The strain CPSB4 was identified as Klebsiella sp. on the basis of morphological, biochemical and 16S rRNA gene sequencing. This study shows that the diverse role of the bacterial strain CPSB4 would be useful in the chromium contaminated soil as a good bioremediation and plant growth promoting agent as well. Copyright © 2017 Elsevier Ltd. All rights reserved.

A Heterogeneous In Vitro Three Dimensional Model of Tumour-Stroma Interactions Regulating Sprouting Angiogenesis

PubMed Central

Correa de Sampaio, Pedro; Auslaender, David; Krubasik, Davia; Failla, Antonio Virgilio; Skepper, Jeremy N.; Murphy, Gillian; English, William R.

2012-01-01

Angiogenesis, the formation of new blood vessels, is an essential process for tumour progression and is an area of significant therapeutic interest. Different in vitro systems and more complex in vivo systems have been described for the study of tumour angiogenesis. However, there are few human 3D in vitro systems described to date which mimic the cellular heterogeneity and complexity of angiogenesis within the tumour microenvironment. In this study we describe the Minitumour model – a 3 dimensional human spheroid-based system consisting of endothelial cells and fibroblasts in co-culture with the breast cancer cell line MDA-MB-231, for the study of tumour angiogenesis in vitro. After implantation in collagen-I gels, Minitumour spheroids form quantifiable endothelial capillary-like structures. The endothelial cell pre-capillary sprouts are supported by the fibroblasts, which act as mural cells, and their growth is increased by the presence of cancer cells. Characterisation of the Minitumour model using small molecule inhibitors and inhibitory antibodies show that endothelial sprout formation is dependent on growth factors and cytokines known to be important for tumour angiogenesis. The model also shows a response to anti-angiogenic agents similar to previously described in vivo data. We demonstrate that independent manipulation of the different cell types is possible, using common molecular techniques, before incorporation into the model. This aspect of Minitumour spheroid analysis makes this model ideal for high content studies of gene function in individual cell types, allowing for the dissection of their roles in cell-cell interactions. Finally, using this technique, we were able to show the requirement of the metalloproteinase MT1-MMP in endothelial cells and fibroblasts, but not cancer cells, for sprouting angiogenesis. PMID:22363483

3D printed alendronate-releasing poly(caprolactone) porous scaffolds enhance osteogenic differentiation and bone formation in rat tibial defects.

PubMed

Kim, Sung Eun; Yun, Young-Pil; Shim, Kyu-Sik; Kim, Hak-Jun; Park, Kyeongsoon; Song, Hae-Ryong

2016-09-29

The aim of this study was to evaluate the in vitro osteogenic effects and in vivo new bone formation of three-dimensional (3D) printed alendronate (Aln)-releasing poly(caprolactone) (PCL) (Aln/PCL) scaffolds in rat tibial defect models. 3D printed Aln/PCL scaffolds were fabricated via layer-by-layer deposition. The fabricated Aln/PCL scaffolds had high porosity and an interconnected pore structure and showed sustained Aln release. In vitro studies showed that MG-63 cells seeded on the Aln/PCL scaffolds displayed increased alkaline phosphatase (ALP) activity and calcium content in a dose-dependent manner when compared with cell cultures in PCL scaffolds. In addition, in vivo animal studies and histologic evaluation showed that Aln/PCL scaffolds implanted in a rat tibial defect model markedly increased new bone formation and mineralized bone tissues in a dose-dependent manner compared to PCL-only scaffolds. Our results show that 3D printed Aln/PCL scaffolds are promising templates for bone tissue engineering applications.

Oocyte cryopreservation and in vitro culture affect calcium signalling during human fertilization.

PubMed

Nikiforaki, D; Vanden Meerschaut, F; Qian, C; De Croo, I; Lu, Y; Deroo, T; Van den Abbeel, E; Heindryckx, B; De Sutter, P

2014-01-01

What are the precise patterns of calcium oscillations during the fertilization of human oocytes matured either in vivo or in vitro or aged in vitro and what is the effect of cryopreservation? Human oocytes matured in vivo exhibit a specific pattern of calcium oscillations, which is affected by in vitro maturation, in vitro ageing and cryopreservation. Oscillations in cytoplasmic calcium concentration are crucial for oocyte activation and further embryonic development. While several studies have described in detail the calcium oscillation pattern during fertilization in animal models, studies with human oocytes are scarce. This was a laboratory-based study using human MII oocytes matured in vivo or in vitro either fresh or after cryopreservation with slow freezing or vitrification. Altogether, 205 human oocytes were included in the analysis. In vivo and in vitro matured human oocytes were used for this research either fresh or following vitrification/warming (V/W) and slow freezing/thawing (F/T). Human oocytes were obtained following written informed consent from patients undergoing ovarian hyperstimulation. For the calcium pattern analysis, oocytes were loaded with the ratiometric calcium indicator fluorescent dye Fura-2. Following ICSI using sperm from a single donor, intracellular calcium was measured for 16 h at 37°C under 6% CO(2). The calcium oscillation parameters were calculated for all intact oocytes that showed calcium oscillations and were analyzed using the Mann-Whitney U-test. Human in vivo MII oocytes display a specific pattern of calcium oscillations following ICSI. This pattern is significantly affected by in vitro ageing, with the calcium oscillations occurring over a longer period of time and with a lower frequency, shorter duration and higher amplitude (P < 0.05). In vitro matured oocytes from the GV and MI stage exhibit a different pattern of calcium oscillations with calcium transients being of lower frequency and shorter duration compared with in vivo matured MII. In MI oocytes that reached the MII stage within 3 h the calcium oscillations additionally appear over a longer period of time (P < 0.05). In vivo MII oocytes show a different calcium oscillation pattern following V/W with calcium oscillations occurring over a longer period of time, with a higher amplitude and a lower frequency (P < 0.05). In vitro matured oocytes, either from the GV or the MI stage, also display an altered pattern of calcium oscillations after V/W and the parameters that were similarly affected in all these oocyte groups are the frequency and the amplitude of the calcium transients. Slow freezing/thawing differentially affects the calcium oscillation pattern of in vitro matured and in vitro aged oocytes. The relationship between a specific pattern of calcium oscillations and subsequent human embryonic development could not be evaluated since the calcium indicator used and the high-intensity excitation light impair development. Furthermore, all oocytes were derived from stimulated cycles and immature oocytes were denuded prior to in vitro maturation. Our data show for the first time how calcium signalling during human fertilization is affected by oocyte in vitro maturation, in vitro ageing as well as V/W and slow freezing/thawing. The analysis of calcium oscillations could be used as an oocyte quality indicator to evaluate in vitro culture and cryopreservation techniques of human oocytes. This work was supported by a clinical research mandate from the Flemish Foundation of Scientific Research (FWO-Vlaanderen, FWO09/ASP/063) to F.V.M, a fundamental clinical research mandate from the FWO-Vlaanderen (FWO05/FKM/001) to P.D.S and a Ghent University grant (KAN-BOF E/01321/01) to B.H. The authors have no conflict of interest to declare.

Development, characterization, and in vivo assessment of mucoadhesive nanoparticles containing fluconazole for the local treatment of oral candidiasis.

PubMed

Rençber, Seda; Karavana, Sinem Yaprak; Yılmaz, Fethiye Ferda; Eraç, Bayri; Nenni, Merve; Özbal, Seda; Pekçetin, Çetin; Gurer-Orhan, Hande; Hoşgör-Limoncu, Mine; Güneri, Pelin; Ertan, Gökhan

2016-01-01

This study aimed to develop a suitable buccal mucoadhesive nanoparticle (NP) formulation containing fluconazole for the local treatment of oral candidiasis. The suitability of the prepared formulations was assessed by means of particle size (PS), polydispersity index, and zeta potential measurements, morphology analysis, mucoadhesion studies, drug entrapment efficiency (EE), in vitro drug release, and stability studies. Based on the optimum NP formulation, ex vivo drug diffusion and in vitro cytotoxicity studies were performed. Besides, evaluation of the antifungal effect of the optimum formulation was evaluated using agar diffusion method, fungicidal activity-related in vitro release study, and time-dependent fungicidal activity. The effect of the optimum NP formulation on the healing of oral candidiasis was investigated in an animal model, which was employed for the first time in this study. The zeta potential, mucoadhesion, and in vitro drug release studies of various NP formulations revealed that chitosan-coated NP formulation containing EUDRAGIT(®) RS 2.5% had superior properties than other formulations. Concerning the stability study of the selected formulation, the formulation was found to be stable for 6 months. During the ex vivo drug diffusion study, no drug was found in receptor phase, and this is an indication of local effect. The in vitro antifungal activity studies showed the in vitro efficacy of the NP against Candida albicans for an extended period. Also, the formulation had no cytotoxic effect at the tested concentration. For the in vivo experiments, infected rabbits were successfully treated with local administration of the optimum NP formulation once a day. This study has shown that the mucoadhesive NP formulation containing fluconazole is a promising candidate with once-a-day application for the local treatment of oral candidiasis.

Polyisoprene matrix for progesterone release: in vitro and in vivo studies.

PubMed

Heredia, V; Bianco, I D; Tríbulo, H; Tríbulo, R; Seoane, M Ferro; Faudone, S; Cuffini, S L; Demichelis, N A; Schalliol, H; Beltramo, D M

2009-12-01

Latex, a polyisoprene (PI) hydrophobic elastomer, was evaluated in vitro and in vivo as a matrix for intravaginal steroid hormone delivery. Matrices containing hormone were prepared by swelling latex in chloroform that contained soluble progesterone (P4). In vitro studies demonstrate that P4 release from PI follows a zero order model during at least 100 h and depends on initial load up to 10 mg cm(-2). The release of P4 from a PI matrix was found to be two times faster than from a polydimethylsiloxane (PDMS) matrix. FT-IR and X-ray powder diffraction analysis of P4 polymorphs show that when nucleated in PDMS, the hormone crystallizes only in alpha-form while in latex, crystallizes as a mixture of alpha- and beta-form. In vivo studies show that devices with a PI matrix containing 0.5 g of P4 are effective to reach plasma levels above 1 ng ml(-1) that are needed to synchronize estrous in cattle. Altogether, the results show that PI, a vulcanized polymer with a carbon-carbon backbone, can be used as a new matrix for the intravaginal administration of progesterone with improved release profile than silicone and that the matrix can influence the crystalline state of the hormone.

Physicochemical, in vitro and in vivo evaluation of flurbiprofen microemulsion.

PubMed

Naeem, Muhammad; Ur Rahman, Nisar; Tavares, Guilherme D; Barbosa, Sávio F; Chacra, Nádia B; Löbenberg, Raimar; Sarfraz, Muhammad K

2015-09-01

Flurbiprofen, a potent nonsteroidal anti-inflammatory drug, is widely used for relief of pain in patients suffering from rheumatic diseases, migraine, sore throat and primary dysmenorrheal. However, this drug has many gastrointestinal side effects produced by its oral administration, such as gastric bleeding and peptic ulcer. These effects were responsible for non-compliance among patients, which ultimately results in treatment failure. The physicochemical properties of flurbiprofen, make it a suitable candidate for transdermal drug delivery, which can overcome the drawbacks of oral administration. In this sense, microemulsions have been proved to increase the cutaneous absorption of lipophilic drugs when compared to conventional drug delivery systems. The purpose of this study was to formulate and characterize gel based microemulsions, for topical delivery of flurbiprofen. Different gel bases, containing microemulsion and hydro-alcoholic solution of flurbiprofen, were developed and compared. In vitro study showed that gels containing microemulsion had a higher permeation rate than those containing hydro-alcoholic solutions. Additionally, formulation of Carbopol-I (microemulsion) showed higher percent of inhibition of inflammation than others bases. Further, skin irritation study demonstrated that Carbopol-I was none irritating. Flurbiprofen microemulsion incorporated on Carbopol-I showed physicochemical, in vitro and in vivo characteristics suitable for the development of alternative transdermal delivery formulation.

In vitro starch digestibility, pasting and textural properties of mung bean: effect of different processing methods.

PubMed

Kaur, Maninder; Sandhu, Kawaljit Singh; Ahlawat, RavinderPal; Sharma, Somesh

2015-03-01

Mung bean was subjected to different processing conditions (soaking, germination, cooking and autoclaving) and their textural, pasting and in vitro starch digestibility characteristics were studied. A significant reduction in textural properties (hardness, cohesiveness, gumminess and chewiness) after cooking and autoclaving treatment of mung bean was observed. Flours made from differently processed mung bean showed significant differences (P < 0.05) in their pastin g characteristics. Peak and final viscosity were the highest for flour from germinated mung bean whereas those made from autoclaved mung bean showed the lowest value. in vitro starch digestibility of mung bean flours was assessed enzymatically using modified Englyst method and the parameters studied were readily digestible starch (RDS), slowly digestible starch (SDS), resistant starch (RS) and total starch (TS) content. Various processing treatments increased the RDS contents of mung bean, while the SDS content was found to be the highest for soaked and the lowest for the autoclaved sample. Germinated sample showed higher amount of digestible starch (RDS + SDS) as compared to raw and soaked samples. Flours from raw and soaked samples showed significantly low starch hydrolysis rate at all the temperatures with total hydrolysis of 29.9 and 31.2 %, respectively at 180 min whereas cooked and autoclaved samples showed high hydrolysis rates with 50.2 and 53.8 % of these hydrolyzing within 30 min of hydrolysis.

Aspartame induces angiogenesis in vitro and in vivo models.

PubMed

Yesildal, F; Aydin, F N; Deveci, S; Tekin, S; Aydin, I; Mammadov, R; Fermanli, O; Avcu, F; Acikel, C H; Ozgurtas, T

2015-03-01

Angiogenesis is the process of generating new blood vessels from preexisting vessels and is considered essential in many pathological conditions. The purpose of the present study is to evaluate the effect of aspartame on angiogenesis in vivo chick chorioallantoic membrane (CAM) and wound-healing models as well as in vitro 2,3-bis-2H-tetrazolium-5-carboxanilide (XTT) and tube formation assays. In CAM assay, aspartame increased angiogenesis in a concentration-dependent manner. Compared with the control group, aspartame has significantly increased vessel proliferation (p < 0.001). In addition, in vivo rat model of skin wound-healing study showed that aspartame group had better healing than control group, and this was statistically significant at p < 0.05. There was a slight proliferative effect of aspartame on human umbilical vein endothelial cells on XTT assay in vitro, but it was not statistically significant; and there was no antiangiogenic effect of aspartame on tube formation assay in vitro. These results provide evidence that aspartame induces angiogenesis in vitro and in vivo; so regular use may have undesirable effect on susceptible cases. © The Author(s) 2015.

Photoacoustic study of percutaneous absorption of Carbopol and transdermic gels for topic use in skin

NASA Astrophysics Data System (ADS)

Rossi, R. C. P.; de Paiva, R. F.; da Silva, M. D.; Barja, P. R.

2008-01-01

Topical medicine application has been used to treat a good number of pathological processes. Its efficacy is associated to an efficient penetration of the drug in the internal skin layers, promoting systemic effects and excluding the possibility of drug degradation by the digestive tract and hepatic elimination. This work analyzes the penetration kinetics of two soluble bases employed as vehicles for topic application: superficial gel (Carbopol 940) and transdermic (transdermal) gel. Analysis was performed with the photoacoustic technique, based upon the absorption of modulated light by a sample with subsequent conversion of the absorbed energy in heat, generating acoustic waves in the air layer adjacent to the sample. Each of the two vehicles was evaluated through in vivo (human skin) and in vitro application. Measurements in vitro employed samples of VitroSkin (synthetic material with properties similar to those of real skin, employed in the pharmaceutical industry research). Results show that the permeation was faster for the transdermal gel, both for in vivo and in vitro measurements, indicating that in vitro measurements may be utilized in qualitative, comparative permeation studies.

In vitro evaluation of whole faba bean and its seed coat as a potential source of functional food components.

PubMed

Çalışkantürk Karataş, Selen; Günay, Demet; Sayar, Sedat

2017-09-01

In vitro studies were conducted to evaluate the particular nutritional benefits of whole faba bean seed (WFB) and fava bean seed coat (FBSC). Total dietary fiber contents of WFB and FBSC were 27.5% and 82.3%, respectively. FBSC were contained much higher total phenolic substances, condensed tannins, and total antioxidant activity than WFB. Bile acid (BA)-binding capacities of in vitro digested samples and nutritionally important products produced by in vitro fermentation of digestion residues were also studied. The BA-binding capacities of WFB and FBSC were 1.94 and 37.50μmol/100mg, respectively. Total BA bound by FBSC was even higher than the positive standard cholestyramine. Lignin and other constituents of the Klason residue were found to influence BA-binding properties. Moreover, the extent of the in vitro fermentation process showed that, fermentability of FBSC residue was significantly lower than that of WFB residue. Overall, faba bean, especially its seed coat, has great potential as a functional food. Copyright © 2017 Elsevier Ltd. All rights reserved.

Ellipsometry analysis of the in vitro adsorption of tea polyphenols onto salivary pellicles.

PubMed

Joiner, Andrew; Muller, Dries; Elofsson, Ulla M; Arnebrant, Thomas

2004-12-01

The adsorption of components from black tea and of purified tea polyphenols onto a whole unstimulated salivary pellicle-like protein layer, formed in vitro on hydroxyapatite discs, was studied by in situ ellipsometry. It was found that components from black tea and the purified polyphenols epicatechin-3-gallate (ECG), epigallocatechin-3-gallate (EGCG) and theaflavin readily adsorbed onto the pellicle. Further investigations showed that under the experimental conditions of this study, no black tea- or purified polyphenol-modified pellicles were eluted by either phosphate buffer or sodium dodecyl sulphate rinses. Therefore, black tea and its polyphenol components are indicated to have a profound effect on in vitro pellicle modification. Similar effects were observed for tannic acid. Copyright Eur J Oral Sci, 2004.

Comparative evaluation of the antimicrobial activity of natural extracts of Morinda citrifolia, papain and aloe vera (all in gel formulation), 2% chlorhexidine gel and calcium hydroxide, against Enterococcus faecalis: An in vitro study.

PubMed

Bhardwaj, Anuj; Ballal, Suma; Velmurugan, Natanasabapathy

2012-07-01

A comparative evaluation of the antimicrobial activity of natural extracts of Morinda citrifolia, papain, and aloe vera (all in gel formulations), 2% chlorhexidine gel and calcium hydroxide, against Enterococcus faecalis-an in vitro study. The antimicrobial efficacy was assessed in vitro using dentin shavings collected at 2 depths of 200 and 400 μm. The total colony forming units at the end of 1, 3, and 5 days were assessed. The overall percentage inhibition of bacterial growth (200 and 400 μm depth) was 100% with chlorhexidine gel. This was followed by M. citrifolia gel (86.02%), which showed better antimicrobial efficacy as compared with aloe vera gel (78.9%), papain gel (67.3%), and calcium hydroxide (64.3%). There was no statistical difference between data at 200 and 400 μm depth. Chlorhexidine gel showed the maximum antimicrobial activity against E. faecalis, whereas calcium hydroxide showed the least. Among the natural intracanal medicaments, M. citrifolia gel consistently exhibited good inhibition up to the 5(th) day followed by aloe vera gel and papain gel.

Delivery of risperidone from gels across porcine skin in vitro and in vivo in rabbits.

PubMed

Ning, Yuming; Chen, Xiaojin; Yu, Zhenwei; Liang, Wenquan; Li, Fanzhu

2018-05-01

The purpose of this study was to develop and evaluate a transdermal delivery system for RIS using hydrogels. First, the effects of different concentrations of hydroxypropyl methylcellulose and Carbomer 934 (CBR) on RIS permeation were investigated in porcine skin. The optimized formulation was chosen as the base gel to screen for penetration enhancers. The pharmacokinetics of the optimized RIS formulation was then studied in vitro in rabbits. A formulation with 0.5% CBR showed the highest RIS permeation and was selected as the base gel. RIS permeation was further increased by incorporation of Azone, lauryl alcohol, or menthol, and the enhancing effects of the three were dose-dependent. When each enhancer combined with propylene glycol (PG) a synergistic effect was found. A combination of 6% menthol and 6% PG exhibited highest RIS in vitro penetration rate and showed a high efficiency in vivo, with a relative bioavailability of 131.53% compared with intragastric administration. These findings showed that 1% RIS in 0.5% CBR, containing a combination of 6% menthol and 6% PG, can deliver doses of RIS that are therapeutically relevant for treating patients with schizophrenia.

In Vivo and In Vitro Toxicity Evaluation of Hydroethanolic Extract of Kalanchoe brasiliensis (Crassulaceae) Leaves.

PubMed

Fonseca, Aldilane Gonçalves; Ribeiro Dantas, Luzia Leiros Sena Fernandes; Fernandes, Júlia Morais; Zucolotto, Silvana Maria; Lima, Adley Antoninni Neves; Soares, Luiz Alberto Lira; Rocha, Hugo Alexandre Oliveira; Lemos, Telma Maria Araújo Moura

2018-01-01

The species Kalanchoe brasiliensis , known as "Saião , " has anti-inflammatory, antimicrobial, and antihistamine activities. It also has the quercetin and kaempferol flavonoids, which exert their therapeutic activities. With extensive popular use besides the defined therapeutical properties, the study of possible side effects is indispensable. The objective of this study is to evaluate the toxicity in vitro and in vivo from the hydroethanolic extract of the leaves of K. brasiliensis . The action of the extract (concentrations from 0.1 to 1000 uL/100 uL) in normal and tumor cells was evaluated using the MTT method. Acute toxicity and subchronic toxicity were evaluated in mice with doses of 250 to 1000 mg/kg orally, following recognized protocols. The in vitro results indicated cytotoxic activity for 3T3 cell line (normal) and 786-0 (kidney carcinoma), showing the activity to be concentration-dependent, reaching 92.23% cell inhibition. In vivo , the extract showed no significant toxicity; only liver changes related to acute toxicity and some signs of liver damage, combining biochemical and histological data. In general, the extract showed low or no toxicity, introducing itself as safe for use with promising therapeutic potential.

In vitro cholesteatoma growth and secretion of cytokines.

PubMed

Helgaland, Tore; Engelen, Bart; Olsnes, Carla; Aarstad, Hans Jørgen; Vassbotn, Flemming S

2010-07-01

Our results show a significant difference between skin and cholesteatoma biology in vitro. Cholesteatoma disease is a process of destruction characterized by uncontrolled growth of squamous epithelial cells in the middle ear or temporal bone. The pathophysiology behind the cholesteatoma development is controversial, and the mechanisms driving the cholesteatoma growth, migration and destructive properties is still unclear. We aimed to provide a method to study the effect of various compounds on cholesteatoma and skin tissue growth, as well as to further investigate the biological differences between normal skin and cholesteatoma tissue. We have established a method to study cholesteatoma biopsy tissue in vitro. Cholesteatoma tissues from patients undergoing surgery for chronic otitis were grown in culture medium and compared to growth patterns and behaviour of normal retroauricular skin. Conditioned medium was analysed for various secreted cytokines. We found a radial outgrowth of keratinocyte epithelium from the circular biopsies. After 5 days of culture we found a significant growth of both cholesteatoma and skin-derived cells. Cholesteatoma samples showed higher growth rate as compared with skin control cultures from the same patient. Moreover, the cholesteatoma cells showed higher production of monocyte chemoattractant protein-1 (MCP-1) and interleukin (IL)-6 as compared with normal skin.

Heterogeneity of proangiogenic features in mesenchymal stem cells derived from bone marrow, adipose tissue, umbilical cord, and placenta.

PubMed

Du, Wen Jing; Chi, Ying; Yang, Zhou Xin; Li, Zong Jin; Cui, Jun Jie; Song, Bao Quan; Li, Xue; Yang, Shao Guang; Han, Zhi Bo; Han, Zhong Chao

2016-11-10

Mesenchymal stem cells (MSCs) have been widely proven effective for therapeutic angiogenesis in ischemia animal models as well as clinical vascular diseases. Because of the invasive method, limited resources, and aging problems of adult tissue-derived MSCs, more perinatal tissue-derived MSCs have been isolated and studied as promising substitutable MSCs for cell transplantation. However, fewer studies have comparatively studied the angiogenic efficacy of MSCs derived from different tissues sources. Here, we evaluated whether the in-situ environment would affect the angiogenic potential of MSCs. We harvested MSCs from adult bone marrow (BMSCs), adipose tissue (AMSCs), perinatal umbilical cord (UMSCs), and placental chorionic villi (PMSCs), and studied their "MSC identity" by flow cytometry and in-vitro trilineage differentiation assay. Then we comparatively studied their endothelial differentiation capabilities and paracrine actions side by side in vitro. Our data showed that UMSCs and PMSCs fitted well with the minimum standard of MSCs as well as BMSCs and AMSCs. Interestingly, we found that MSCs regardless of their tissue origins could develop similar endothelial-relevant functions in vitro, including producing eNOS and uptaking ac-LDL during endothelial differentiation in spite of their feeble expression of endothelial-related genes and proteins. Additionally, we surprisingly found that BMSCs and PMSCs could directly form tubular structures in vitro on Matrigel and their conditioned medium showed significant proangiogenic bioactivities on endothelial cells in vitro compared with those of AMSCs and UMSCs. Besides, several angiogenic genes were upregulated in BMSCs and PMSCs in comparison with AMSCs and UMSCs. Moreover, enzyme-linked immunosorbent assay further confirmed that BMSCs secreted much more VEGF, and PMSCs secreted much more HGF and PGE2. Our study demonstrated the heterogeneous proangiogenic properties of MSCs derived from different tissue origins, and the in vivo isolated environment might contribute to these differences. Our study suggested that MSCs derived from bone marrow and placental chorionic villi might be preferred in clinical application for therapeutic angiogenesis.

Antifungal activities of diphenyl diselenide and ebselen against echinocandin-susceptible and -resistant strains of Candida parapsilosis.

PubMed

Chassot, Francieli; Pozzebon Venturini, Tarcieli; Baldissera Piasentin, Fernanda; Morais Santurio, Janio; Estivalet Svidzinski, Terezinha Inez; Hartz Alves, Sydney

2016-10-01

We evaluated the in vitro antifungal activity of diphenyl diselenide and ebselen against echinocandin-susceptible and -resistant strains of Candida parapsilosis using the broth microdilution method. Diphenyl diselenide (MIC range =1-8 µg/mL) and ebselen (MIC range =0.25-4 µg/mL) showed in vitro activity against echinocandin-susceptible isolates. However, ebselen also showed the highest antifungal activity against echinocandin-resistant strains (MIC range =0.06-4 µg/mL). This study demonstrated that the antifungal potential of diphenyl diselenide and ebselen deserves further investigation using in vivo experimental protocols.

A new radiopharmaceutical compound (131I-PR81) for radioimmunotherapy of breast cancer: labeling of antibody and its quality control.

PubMed

Mohammadnejad, J; Rasaee, M J; Babaei, M H; Paknejad, M; Zahir, M H; Salouti, M; Rajabi, A Bitarafan; Mazidi, M

2010-01-01

PR81 is a monoclonal antibody that binds with high affinity to MUC1, which is over expressed on breast and other tumors. The objective of this study was to evaluate the application of this antibody against MUC1 as a radioimmunotherapeutical agent. Monoclonal antibody (PR81) against MUC1 was prepared, characterized, purified, and labeled with 131I. The immunoreactivity of radiolabeled mAb PR81with MUC1 (the native protein), BSA-P20 (a 20 amino acid corresponding the tandem repeat of MUC1) and MCF7 cell line were performed by RIA. In vitro stability of radiolabeled mAb in human serum was determined by thin layer chromatography (TLC). Cell toxicity and in vitro internalization studies were performed with the MCF7 cell line, and the tissue biodistribution of the radioiodinated PR81 was evaluated in normal BALB/c mice at 4, 24 and 48 hrs. The tumor imaging was performed in BALB/c mice with breast xenograft tumors at 24 and 72 hr after the complex injection. The labeling efficiency was found to be 59.9% ± 7.9%. MAb-131I conjugates showed high immunoreactivity towards MUC1 protein, BSA-P20 and MCF7 cell line. In vitro stability of the labeled product in human serum was found to be more than %50 over 24 hr. Cell toxicity and in vitro internalization studies showed that the mAb-131I conjugate inhibited 80% growth of the MCF7 cultured cell lines in vitro in a high concentration and up to %60 of the conjugate internalized after 24 h. Biodistribution studies were performed in normal BALB/c mice at 4, 24 and 48 hrs post-injection and no important accumulation was observed in vital organs. The tumors were visualized with high sensitivity after 24 and 72 hr in radioimmunoscintographical studies. These results show that the new radiopharmaceutical may be considered as a promising candidate for therapy of breast cancer.

Inhibitory effects of Agaricus blazei extracts on human myeloid leukemia cells.

PubMed

Kim, Chi-Fai; Jiang, Jing-Jing; Leung, Kwok-Nam; Fung, Kwok-Pui; Lau, Clara Bik-San

2009-03-18

Agaricus blazei has been used as an adjuvant in cancer chemotherapy and is found to inhibit the growth of various types of tumor cells. Our study has adopted a systematic and bioassay-guided approach to optimize the extraction of Agaricus blazei for anti-leukemic bioactive components. The tumor-selective growth inhibitory activity of the extracts on leukemic cell lines was evaluated in vitro and in vivo using tumor-bearing nude mice. Agaricus blazei extracts were prepared using different methods. MTT and tritiated thymidine incorporation assays were used to evaluate the in vitro anti-leukemic effects. The most potent extract was further investigated using NB-4 cells-bearing nude mice and mechanistic studies using DNA fragmentation assay and cell death detection ELISA. The JAB80E70 extract showed the most potent tumor-selective growth inhibitory activity against human leukemia NB-4 and K-562 cells. This is the first report of anti-leukemic activity of JAB80E70 in athymic nude mice bearing NB-4 cells. Using DNA fragmentation assays and cell death detection ELISA, JAB80E70 was found to induce apoptosis in NB-4 cells. However, the polysaccharide enriched fractions failed to show significant cytotoxicity on NB-4 cells in vitro. The JAB80E70 extract exhibited potent anti-leukemic effect in vitro and in vivo. The effect can be attributed, at least in part, to the induction of apoptosis. Besides, polysaccharides in Agaricus blazei may not possess direct anti-leukemic activity in vitro.

Preparation, characterization, and in vitro release study of albendazole-encapsulated nanosize liposomes

PubMed Central

Panwar, Preety; Pandey, Bhumika; Lakhera, P C; Singh, K P

2010-01-01

The purpose of the present study was to formulate effective and controlled release albendazole liposomal formulations. Albendazole, a hydrophobic drug used for the treatment of hydatid cysts, was encapsulated in nanosize liposomes. Rapid evaporation method was used for the preparation of albendazole-encapsulated conventional and PEGylated liposomes consisting of egg phosphatidylcholine (PC) and cholesterol (CH) in the molar ratios of (6:4) and PC:CH: polyethylene glycol (PEG) (5:4:1), respectively. In this study, PEGylated and conventional liposomes containing albendazole were prepared and their characteristics, such as particle size, encapsulation efficiency, and in vitro drug release were investigated. The drug encapsulation efficiency of PEGylated and conventional liposomes was 81% and 72%, respectively. The biophysical characterization of both conventional and PEG-coated liposomes were done by transmission electron microscopy and UV-vis spectrophotometry. Efforts were made to study in vitro release of albendazole. The drug release rate showed decrease in albendazole release in descending order: free albendazole, albendazole-loaded conventional liposomes, and least with albendazole-loaded PEG-liposomes. Biologically relevant vesicles were prepared and in vitro release of liposome-entrapped albendazole was determined. PMID:20309396

Cellular and Tumor Radiosensitivity is Correlated to Epidermal Growth Factor Receptor Protein Expression Level in Tumors Without EGFR Amplification;Epidermal growth factor receptor; Radiotherapy; Squamous cell carcinoma; Biomarker; Local tumor control

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kasten-Pisula, Ulla; Saker, Jarob; Eicheler, Wolfgang

2011-07-15

Purpose: There is conflicting evidence for whether the expression of epidermal growth factor receptor in human tumors can be used as a marker of radioresponse. Therefore, this association was studied in a systematic manner using squamous cell carcinoma (SCC) cell lines grown as cell cultures and xenografts. Methods and Materials: The study was performed with 24 tumor cell lines of different tumor types, including 10 SCC lines, which were also investigated as xenografts on nude mice. Egfr gene dose and the length of CA-repeats in intron 1 were determined by polymerase chain reaction, protein expression in vitro by Western blotmore » and in vivo by enzyme-linked immunosorbent assay, and radiosensitivity in vitro by colony formation. Data were correlated with previously published tumor control dose 50% data after fractionated irradiation of xenografts of the 10 SCC. Results: EGFR protein expression varies considerably, with most tumor cell lines showing moderate and only few showing pronounced upregulation. EGFR upregulation could only be attributed to massive gene amplification in the latter. In the case of little or no amplification, in vitro EGFR expression correlated with both cellular and tumor radioresponse. In vivo EGFR expression did not show this correlation. Conclusions: Local tumor control after the fractionated irradiation of tumors with little or no gene amplification seems to be dependent on in vitro EGFR via its effect on cellular radiosensitivity.« less

Evaluation of a Bioabsorbable Self-Expandable Vein Stent-Base Made of Poly(L-lactide) In Vitro and In Vivo.

PubMed

Løvdal, Alexandra Liv Vest; Calve, Sarah; Yang, Shuo; Van Alstine, William; Binkert, Christoph A; Klausen, Kasper

2017-01-01

This study was designed to evaluate performance and tissue response to a self-expandable bioabsorbable vein stent-base cut from a tube with enhanced stiffness and strength in vitro and in vivo. A diamond-shaped stent-base was cut from a sequential biaxially strained poly(L-lactide) (PLLA) tube for optimized performance. The performance of the stent-base was evaluated in a finite element analysis model, and validation was attempted in vitro through a cyclic flat-plate compression and radial force measurement. The performance of the stent-base was tested in vivo using 3 sheep with 2 implants each for 2 and 3½ weeks, respectively. In vitro the stent-base showed an elliptical deformation but no fractures. In vivo the stent-base showed adequate radial force and no migration. All implanted stent-bases showed multiple fractures not only at the predicted stress zones but at all connecting points. Fragments of the caudal stent-base stayed in the vein wall indicating sufficient tissue coverage to avoid embolization of the fractured stent pieces, whereas fragments from the cranial device remaining were few. Neointima formation was confirmed histologically at 2 and 3½ weeks. A bioabsorbable self-expandable stent-base made from PLLA for large veins seems feasible, but over time, the PLLA used in this study appears too stiff and lacks the sufficient flexibility to move with the vena cava, causing multiple fractures.

Evaluation of a Bioabsorbable Self-Expandable Vein Stent-Base Made of Poly(l-lactide) In Vitro and In Vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Løvdal, Alexandra Liv Vest, E-mail: alvlo@nanotech.dtu.dk; Calve, Sarah; Yang, Shuo

PurposeThis study was designed to evaluate performance and tissue response to a self-expandable bioabsorbable vein stent-base cut from a tube with enhanced stiffness and strength in vitro and in vivo.MethodsA diamond-shaped stent-base was cut from a sequential biaxially strained poly(l-lactide) (PLLA) tube for optimized performance. The performance of the stent-base was evaluated in a finite element analysis model, and validation was attempted in vitro through a cyclic flat-plate compression and radial force measurement. The performance of the stent-base was tested in vivo using 3 sheep with 2 implants each for 2 and 3½ weeks, respectively.ResultsIn vitro the stent-base showed anmore » elliptical deformation but no fractures. In vivo the stent-base showed adequate radial force and no migration. All implanted stent-bases showed multiple fractures not only at the predicted stress zones but at all connecting points. Fragments of the caudal stent-base stayed in the vein wall indicating sufficient tissue coverage to avoid embolization of the fractured stent pieces, whereas fragments from the cranial device remaining were few. Neointima formation was confirmed histologically at 2 and 3½ weeks.ConclusionA bioabsorbable self-expandable stent-base made from PLLA for large veins seems feasible, but over time, the PLLA used in this study appears too stiff and lacks the sufficient flexibility to move with the vena cava, causing multiple fractures.« less

Thymomodulin: biological properties and clinical applications.

PubMed

Kouttab, N M; Prada, M; Cazzola, P

1989-01-01

Thymomodulin (Ellem Industria Farmaceutica s.p.a., Milan, Italy) is a calf thymus acid lysate derivative, composed of several peptides with a molecular weight range of 1-10 kD. Thymomodulin did not exhibit any mutagenic effect. Furthermore, thymomodulin used in animal studies showed no toxicity even when used at high concentrations. Of major significance are the observations in murine and human systems that thymomodulin remains active when administered orally. In vitro and in vivo administered thymomodulin was able to induce the maturation of T-lymphocytes. Additionally, studies in vitro showed that this thymic derivative can enhance the functions of mature T-lymphocytes with cascading effects on B-cell and macrophage functions. Extensive human clinical trials with thymomodulin showed that this agent can improve the clinical symptoms observed with various disease processes, including infections, allergies and malignancies, and can improve immunological functions during ageing.

Neural differentiation of adipose-derived stem cells isolated from GFP transgenic mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fujimura, Juri; Department of Pediatrics, Nippon Medical School, Tokyo; E-mail: juri-f@nms.ac.jp

2005-07-22

Taking advantage of homogeneously marked cells from green fluorescent protein (GFP) transgenic mice, we have recently reported that adipose-derived stromal cells (ASCs) could differentiate into mesenchymal lineages in vitro. In this study, we performed neural induction using ASCs from GFP transgenic mice and were able to induce these ASCs into neuronal and glial cell lineages. Most of the neurally induced cells showed bipolar or multipolar appearance morphologically and expressed neuronal markers. Electron microscopy revealed their neuronal morphology. Some cells also showed glial phenotypes, as shown immunocytochemically. The present study clearly shows that ASCs derived from GFP transgenic mice differentiate intomore » neural lineages in vitro, suggesting that these cells might provide an ideal source for further neural stem cell research with possible therapeutic application for neurological disorders.« less

Aseptic Culture Systems of Radopholus similis for In Vitro Assays on Musa spp. and Arabidopsis thaliana

PubMed Central

Elsen, A.; Lens, K.; Nguyet, D. T. M.; Broos, S.; Stoffelen, R.; De Waele, D.

2001-01-01

Radopholus similis is one of the most damaging nematodes in bananas. Chemical control is currently the most-used method, but nematode control through genetic improvement is widely encouraged. The objective of this study was to establish an aseptic culture system for R. similis and determine whether R. similis can infect and reproduce on in vitro banana plantlets and in vitro Arabidopsis thaliana. In the study's first part, a suitable aseptic culture system was developed using alfalfa callus. Radopholus similis could penetrate and reproduce in the callus. Six weeks after inoculation with 25 females, the reproduction ratio was 26.3 and all vermiform stages were present. The reproduction ratio increased to 223.2 after 12 weeks. Results of a greenhouse test showed that R. similis did not lose its pathogenicity after culturing on alfalfa callus. In the study's second part, the infection and reproduction of the nematodes cultured on the callus were studied on both in vitro banana plantlets and A. thaliana. Radopholus similis infected and reproduced on both banana and A. thaliana. Furthermore, nematode damage was observed in the root systems of both hosts. These successful infections open new perspectives for rapid in vitro screening for resistance in banana cultivars and anti-nematode proteins expressed in A. thaliana. PMID:19266012

Progress in Assessing Air Pollutant Risks from In Vitro Exposures: Matching Ozone Dose and Effect in Human Airway Cells

PubMed Central

Hatch, Gary E.; Duncan, Kelly E.; Diaz-Sanchez, David; Schmitt, Michael T.; Ghio, Andrew J.; Carraway, Martha Sue; McKee, John; Dailey, Lisa A.; Berntsen, Jon; Devlin, Robert B.

2014-01-01

In vitro exposures to air pollutants could, in theory, facilitate a rapid and detailed assessment of molecular mechanisms of toxicity. However, it is difficult to ensure that the dose of a gaseous pollutant to cells in tissue culture is similar to that of the same cells during in vivo exposure of a living person. The goal of the present study was to compare the dose and effect of O3 in airway cells of humans exposed in vivo to that of human cells exposed in vitro. Ten subjects breathed labeled O3 (18O3, 0.3 ppm, 2 h) while exercising intermittently. Bronchial brush biopsies and lung lavage fluids were collected 1 h post exposure for in vivo data whereas in vitro data were obtained from primary cultures of human bronchial epithelial cells exposed to 0.25–1.0 ppm 18O3 for 2 h. The O3 dose to the cells was defined as the level of 18O incorporation and the O3 effect as the fold increase in expression of inflammatory marker genes (IL-8 and COX-2). Dose and effect in cells removed from in vivo exposed subjects were lower than in cells exposed to the same 18O3 concentration in vitro suggesting upper airway O3 scrubbing in vivo. Cells collected by lavage as well as previous studies in monkeys show that cells deeper in the lung receive a higher O3 dose than cells in the bronchus. We conclude that the methods used herein show promise for replicating and comparing the in vivo dose and effect of O3 in an in vitro system. PMID:24928893

Paracrine signalling of inflammatory cytokines from an in vitro blood brain barrier model upon exposure to polymeric nanoparticles.

PubMed

Raghnaill, Michelle Nic; Bramini, Mattia; Ye, Dong; Couraud, Pierre-Olivier; Romero, Ignacio A; Weksler, Babette; Åberg, Christoffer; Salvati, Anna; Lynch, Iseult; Dawson, Kenneth A

2014-03-07

Nanoparticle properties, such as small size relative to large highly modifiable surface area, offer great promise for neuro-therapeutics and nanodiagnostics. A fundamental understanding and control of how nanoparticles interact with the blood-brain barrier (BBB) could enable major developments in nanomedical treatment of previously intractable neurological disorders, and help ensure that nanoparticles not intended to reach the brain do not cause adverse effects. Nanosafety is of utmost importance to this field. However, a distinct lack of knowledge exists regarding nanoparticle accumulation within the BBB and the biological effects this may induce on neighbouring cells of the Central Nervous System (CNS), particularly in the long-term. This study focussed on the exposure of an in vitro BBB model to model carboxylated polystyrene nanoparticles (PS COOH NPs), as these nanoparticles are well characterised for in vitro experimentation and have been reported as non-toxic in many biological settings. TEM imaging showed accumulation but not degradation of 100 nm PS COOH NPs within the lysosomes of the in vitro BBB over time. Cytokine secretion analysis from the in vitro BBB post 24 h 100 nm PS COOH NP exposure showed a low level of pro-inflammatory RANTES protein secretion compared to control. In contrast, 24 h exposure of the in vitro BBB endothelium to 100 nm PS COOH NPs in the presence of underlying astrocytes caused a significant increase in pro-survival signalling. In conclusion, the tantalising possibilities of nanomedicine must be balanced by cautious studies into the possible long-term toxicity caused by accumulation of known 'toxic' and 'non-toxic' nanoparticles, as general toxicity assays may be disguising significant signalling regulation during long-term accumulation.

Multifunctional reduction-responsive SPIO&DOX-loaded PEGylated polymeric lipid vesicles for magnetic resonance imaging-guided drug delivery

NASA Astrophysics Data System (ADS)

Wang, Sheng; Yang, Weitao; Du, Hongli; Guo, Fangfang; Wang, Hanjie; Chang, Jin; Gong, Xiaoqun; Zhang, Bingbo

2016-04-01

Multifunctional superparamagnetic iron-oxide (SPIO)-based nanoparticles have been emerging as candidate nanosystems for cancer diagnosis and therapy. Here, we report the use of reduction- responsive SPIO/doxorubicin (DOX)-loaded poly(ethylene glycol) monomethyl ether (PEG)ylated polymeric lipid vesicles (SPIO&DOX-PPLVs) as a novel theranostic system for tumor magnetic resonance imaging (MRI) diagnosis and controlled drug delivery. These SPIO&DOX-PPLVs are composed of SPIOs that function as MR contrast agents for tumor enhancement and PPLVs as polymer matrices for encapsulating SPIO and antitumor drugs. The in vitro characterizations show that the SPIO&DOX-PPLVs have nanosized structures (˜80 nm), excellent colloidal stability, good biocompatibility, as well as T 2-weighted MRI capability with a relatively high T 2 relaxivity (r 2 = 213.82 mM-1 s-1). In vitro drug release studies reveal that the release rate of DOX from the SPIO&DOX-PPLVs is accelerated in the reduction environment. An in vitro cellular uptake study and an antitumor study show that the SPIO&DOX-PPLVs have magnetic targeting properties and effective antitumor activity. In vivo studies show the SPIO&DOX-PPLVs have excellent T 2-weighted tumor targeted MRI capability, image-guided drug delivery capability, and high antitumor effects. These results suggest that the SPIO&DOX-PPLVs are promising nanocarriers for MRI diagnosis and cancer therapy applications.

Comparative evaluation of antiplatelet effect of lycopene with aspirin and the effect of their combination on platelet aggregation: An in vitro study.

PubMed

Sawardekar, Swapna B; Patel, Tejal C; Uchil, Dinesh

2016-01-01

The objective was to compare antiplatelet effect of lycopene with aspirin and to study effect of combination of the two on platelet aggregation in vitro, using platelets from healthy volunteers. Platelets were harvested; platelet count of platelet-rich plasma adjusted to 2.5 Χ 10(5)/μL. Aspirin (140 μmol/L) and lycopene (4, 6, 8, 10, and 12 μmol/L) were studied in vitro against adenosine-5'- diphosphate (ADP) (2.5 μM/L) and collagen. All the concentrations of lycopene (4-12 μmol/L) exhibited reduction in maximum platelet aggregation induced by aggregating agents ADP and collagen (P < 0.01 vs. vehicle) and were comparable with aspirin. Lycopene at concentration 10 μmol/L showed maximum platelet inhibition (47.05% ± 19.56%) against ADP, whereas lycopene at concentration 8 μmol/L showed maximum platelet inhibition (54.26% ± 30.71%) against collagen. Four μmol/L of lycopene combined with 140 μmol/L and 70 μmol/L aspirin showed greater inhibition of platelets as compared to aspirin 140 μmol/L alone, against both ADP and collagen. The study favorably compares lycopene and aspirin with respect to their antiplatelet activities against ADP and collagen. Lycopene can be considered as a potential target for modifying the thrombotic and pro-inflammatory events associated with platelet activation.

[Morphogenesis in formative process in vitro from Rehmannia glutinosa].

PubMed

Xue, Jian-ping; Zhang, Ai-min; Liu, Jun; Xu, Xue-feng

2004-01-01

To study the morphogenesis in formative process of tuberous root in vitro from Rehmannia glutinosa and compare the anatomical shape of tuberous root with nature term R. glutinosa. Tuberous roots of different vegetal phase were cut and dyed, then made into paraffin cuts and observed microscope. In anatomical shape, nature R. glutinosa and tuberous root were the same, which showed that no structural variation occurred in tuberous root induced process.

Influence of acidic pH on keratinocyte function and re-epithelialisation of human in vitro wounds.

PubMed

Lönnqvist, Susanna; Emanuelsson, Peter; Kratz, Gunnar

2015-01-01

Chronic wounds are one of the greatest challenges for the healthcare system. Today, a plethora of dressings are used in the treatment of these wounds, each with specific influence on the wound environment. Due to differences in the permeability of the dressings the use will result in differences in the pH balance in the wound bed. However, little is known about how changes in the pH in the wound environment affect the different phases of the healing process. The aim of the present study was to investigate the effects of acidic pH on the regeneration phase by studying keratinocyte function in vitro and re-epithelialisation in an in vitro model of human skin. In vitro assays showed reduced viability and migration rates in human keratinocytes when pH was lowered. Real time PCR revealed differential expression of genes related to wound healing and environmental impairment. Tissue culture showed no re-epithelialisation of wounds subjected to pH 5.0 and moderate re-epithelialisation at pH 6.0, compared to controls at pH 7.4. The results indicate that lowering pH down to pH 5.0 in wounds is counterproductive in aspect of keratinocyte function which is crucial for successful wound healing.

Different concentrations of grape seed extract affect in vitro starch fermentation by porcine small and large intestinal inocula.

PubMed

Wang, Dongjie; Williams, Barbara A; Ferruzzi, Mario G; D'Arcy, Bruce R

2013-01-01

Grape seed extract (GSE) phenolics have potential health-promoting properties, either from compounds present within the extract, or metabolites resulting from gastrointestinal tract (GIT) fermentation of these compounds. This study describes how GSE affected the kinetics and end-products of starch fermentation in vitro using pig intestinal and fecal inocula. Six GSE concentrations (0, 60, 125, 250, 500, and 750 µg ml⁻¹ were fermented in vitro by porcine ileal and fecal microbiota using starch as the energy source. Cumulative gas production, and end-point short chain fatty acids and ammonia were measured. GSE phenolics altered the pattern (gas kinetics, and end-products such as SCFA and NH₄⁺) of starch fermentation by both inocula, at concentrations above 250 µg ml⁻¹ . Below this level, neither inoculum showed any significant (P > 0.05) effect of the GSE. The results show that GSE phenolics at a concentration over 250 µg ml⁻¹ can have measurable effects on microbial activity in an in vitro fermentation system, as evidenced by the changes in kinetics and end-products from starch fermentation. This suggests that fermentation patterns could be conceivably shifted in the actual GIT, though further evidence will be required from in vivo studies. Copyright © 2012 Society of Chemical Industry.

Pirfenidone-loaded liposomes for lung targeting: preparation and in vitro/in vivo evaluation

PubMed Central

Meng, Hui; Xu, Yong

2015-01-01

Background The purpose of this study was to develop novel pirfenidone (PFD)-loaded liposomes for targeting to the lung. Methods The liposomes were prepared by the film hydration method, and their in vitro/vivo characteristics were evaluated. Results The PFD liposomes appeared visually as green to yellowish suspensions and were spherical in shape. The particle size was 582.3±21.6 nm and the entrapment efficiency was relatively high (87.2%±5.7%). The liposomes showed typical sustained and prolonged drug-release behavior in vitro and fitted well with the Weibull distribution equation. The relatively slower time taken to reach a minimal plasma PFD concentration in vivo suggests that PFD liposomes have a sustained-release profile, which is consistent with the results of the in vitro release study. The PFD liposomes showed the largest area under the curve for the lung. The high distribution of PFD achieved in the lungs using this liposomal formulation may be explained by physical entrapment of the liposomes in the vascular network of the lung. Histopathological results indicated that liposomal PFD could alleviate pathological injury in lung tissue. Conclusion This liposomal formulation can enable sustained release of PFD and increase targeting to the lung. PMID:26185416

In vitro and in situ characterization of arthroscopic loop security and knot security of braided polyblend sutures: a biomechanical study.

PubMed

Armstrong, Lucas C; Chong, Alexander; Livermore, Ryan W; Prohaska, Daniel J; Doyon, Amanda N; Wooley, Paul H

2015-04-01

We conducted a study to evaluate biomechanical performance during destructive testing of several different suture materials in various arthroscopic knot configurations under both in vitro and in situ conditions. Surgeons of different levels of experience tied the knots. Three different arthroscopic knots (static surgeon's, Weston, Tennessee slider) with 3 reverse half-hitches on alternating posts were tested using Fiberwire, ForceFiber, Orthocord, and Ultrabraid suture materials under both in vitro and in situ (blood plasma at 37°C) conditions. Three surgeons of different experience levels tied the knots on a post 30 mm in circumference. A single load-to-failure test was performed. There were no significant in vitro-in situ differences for Ultrabraid in the different knot configurations or with the different experience levels. Surgeon B (intermediate experience) showed no significant differences between test conditions for any knot configuration or suture material. With Tennessee slider knots, surgeon C (least experience) showed significantly lower clinical failure load under both test conditions and had a higher percentage of complete knot slippage. Surgeon B had no knot slippage with use of Fiberwire. Both the aqueous environment and the surgeon's familiarity with certain knots have an effect on knot security.

Structural characterization and in vitro antioxidant activity of kojic dipalmitate loaded w/o/w multiple emulsions intended for skin disorders.

PubMed

Gonçalez, Maíra Lima; Marcussi, Diana Gleide; Calixto, Giovana Maria Fioramonti; Corrêa, Marcos Antonio; Chorilli, Marlus

2015-01-01

Multiple emulsions (MEs) are intensively being studied for drug delivery due to their ability to load and increase the bioavailability of active lipophilic antioxidant, such as kojic dipalmitate (KDP). The aim of this study was to structurally characterize developed MEs by determining the average droplet size (Dnm) and zeta potential (ZP), performing macroscopic and microscopic analysis and analyzing their rheological behavior and in vitro bioadhesion. Furthermore, the in vitro safety profile and antioxidant activity of KDP-loaded MEs were evaluated. The developed MEs showed a Dnm of approximately 1 micrometer and a ZP of -13 mV, and no change was observed in Dnm or ZP of the system with the addition of KDP. KDP-unloaded MEs exhibited ''shear thinning" flow behavior whereas KDP-loaded MEs exhibited Newtonian behavior, which are both characteristic of antithixotropic materials. MEs have bioadhesion properties that were not influenced by the incorporation of KDP. The results showed that the incorporation of KDP into MEs improved the safety profile of the drug. The in vitro antioxidant activity assay suggested that MEs presented a higher capacity for maintaining the antioxidant activity of KDP. ME-based systems may be a promising platform for the topical application of KDP in the treatment of skin disorders.

Preparation and In Vitro/Ex Vivo Evaluation of Moxifloxacin-Loaded PLGA Nanosuspensions for Ophthalmic Application.

PubMed

Mudgil, Meetali; Pawar, Pravin K

2013-01-01

The aim of the present investigation was to prepare a colloidal ophthalmic formulation to improve the residence time of moxifloxacin. Moxifloxacin-loaded poly(dl-lactide-co-glycolide) (PLGA) nanosuspensions were prepared by using the solvent evaporation technique. The nanosuspensions were characterised physically by using different techniques like particle size, zeta potential, FTIR, DSC, and XRD analysis. In vitro and ex vivo studies of nanosuspensions were carried out using a modified USP dissolution apparatus and all-glass Franz diffusion cells, respectively. The antibacterial activities of the nanosuspension and marketed formulations were performed against S. aureus and P. aeroginosa. The moxifloxacin-loaded PLGA nanosuspensions showed uniform particle size, ranging between 164-490 nm with negative zeta potential for all batches. The percentage entrapment efficiency of the drug-loaded nano-suspension was found to be between 84.09 to 92.05%. In vitro drug release studies suggest that all of the formulations showed extended drug release profiles and follow Korsemeyer-Peppas release kinetics. In vitro corneal permeability was found to be comparable with that of the marketed formulation across isolated goat cornea, indicating the suitability of the nanosuspension formulation in the ophthalmic delivery of moxifloxacin. The optimised nano-suspension was found to be more active against S. aureus and P. aeruginosa compared to the marketed eye drops.

Characterization and evaluation of 5-fluorouracil-loaded solid lipid nanoparticles prepared via a temperature-modulated solidification technique.

PubMed

Patel, Meghavi N; Lakkadwala, Sushant; Majrad, Mohamed S; Injeti, Elisha R; Gollmer, Steven M; Shah, Zahoor A; Boddu, Sai Hanuman Sagar; Nesamony, Jerry

2014-12-01

The aim of this research was to advance solid lipid nanoparticle (SLN) preparation methodology by preparing glyceryl monostearate (GMS) nanoparticles using a temperature-modulated solidification process. The technique was reproducible and prepared nanoparticles without the need of organic solvents. An anticancer agent, 5-fluorouracil (5-FU), was incorporated in the SLNs. The SLNs were characterized by particle size analysis, zeta potential analysis, differential scanning calorimetry (DSC), infrared spectroscopy, atomic force microscopy (AFM), transmission electron microscopy (TEM), drug encapsulation efficiency, in vitro drug release, and in vitro cell viability studies. Particle size of the SLN dispersion was below 100 nm, and that of redispersed lyophilizates was ~500 nm. DSC and infrared spectroscopy suggested that the degree of crystallinity did not decrease appreciably when compared to GMS. TEM and AFM images showed well-defined spherical to oval particles. The drug encapsulation efficiency was found to be approximately 46%. In vitro drug release studies showed that 80% of the encapsulated drug was released within 1 h. In vitro cell cultures were biocompatible with blank SLNs but demonstrated concentration-dependent changes in cell viability to 5-FU-loaded SLNs. The 5-FU-loaded SLNs can potentially be utilized in an anticancer drug delivery system.

Evaluating in Vitro Culture Medium of Gut Microbiome with Orthogonal Experimental Design and a Metaproteomics Approach.

PubMed

Li, Leyuan; Zhang, Xu; Ning, Zhibin; Mayne, Janice; Moore, Jasmine I; Butcher, James; Chiang, Cheng-Kang; Mack, David; Stintzi, Alain; Figeys, Daniel

2018-01-05

In vitro culture based approaches are time- and cost-effective solutions for rapidly evaluating the effects of drugs or natural compounds against microbiomes. The nutritional composition of the culture medium is an important determinant for effectively maintaining the gut microbiome in vitro. This study combines orthogonal experimental design and a metaproteomics approach to obtaining functional insights into the effects of different medium components on the microbiome. Our results show that the metaproteomic profile respond differently to medium components, including inorganic salts, bile salts, mucin, and short-chain fatty acids. Multifactor analysis of variance further revealed significant main and interaction effects of inorganic salts, bile salts, and mucin on the different functional groups of gut microbial proteins. While a broad regulating effect was observed on basic metabolic pathways, different medium components also showed significant modulations on cell wall, membrane, and envelope biogenesis and cell motility related functions. In particular, flagellar assembly related proteins were significantly responsive to the presence of mucin. This study provides information on the functional influences of medium components on the in vitro growth of microbiome communities and gives insight on the key components that must be considered when selecting and optimizing media for culturing ex vivo microbiotas.

In vitro and in vivo degradation of potential anti-adhesion materials: Electrospun membranes of poly(ester-amide) based on l-phenylalanine and p-(dioxanone).

PubMed

Wang, Bing; Dong, Jun; Niu, Lijing; Chen, Wenyan; Chen, Dongliang; Shen, Chengyi; Zhu, Jiang; Zhang, Xiaoming

2017-08-01

Electrospun membranes of poly(p-dioxanone-co-l-phenylalanine) (PDPA) hold potential as an anti-adhesion material. Since adjustable degradation properties are important for anti-adhesion materials, in this study, the in vitro and in vivo degradation processes of PDPA electrospun membranes were investigated in detail. The morphological analysis of these membranes revealed the main degradation conditions of PDPA membranes. The weight remaining and molecular weight variation showed that the overall degradation rate of the membranes could be adjusted by modulating the molecular structure of the PDPAs. Especially, α-chymotrypsin could catalyze the degradation process of PDPAs. Based on these results, the in vitro degradation mechanism was demonstrated, and confirmed by 1 H NMR of the hydrolysis products. Finally, the in vivo degradation and biocompatibility of different PDPAs were investigated. The kinetic study showed that the in vitro and in vivo molecular weight loss of PDPAs have the first-order characteristics. The in vivo degradation rate of the most Phe-containing PDPA-3 is the slowest, and this result relates to the biocompatibilities of PDPAs. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 1369-1378, 2017. © 2016 Wiley Periodicals, Inc.

The effect of antimicrobial agents on bond strength of orthodontic adhesives: a meta-analysis of in vitro studies.

PubMed

Altmann, A S P; Collares, F M; Leitune, V C B; Samuel, S M W

2016-02-01

Antimicrobial orthodontic adhesives aim to reduce white spot lesions' incidence in orthodontic patients, but they should not jeopardizing its properties. Systematic review and meta-analysis were performed to answer the question whether the association of antimicrobial agents with orthodontic adhesives compromises its mechanical properties and whether there is a superior antimicrobial agent. PubMed and Scopus databases. In vitro studies comparing shear bond strength of conventional photo-activated orthodontic adhesives to antimicrobial photo-activated orthodontic adhesives were considered eligible. Search terms included the following: orthodontics, orthodontic, antimicrobial, antibacterial, bactericidal, adhesive, resin, resin composite, bonding agent, bonding system, and bond strength. The searches yielded 494 citations, which turned into 467 after duplicates were discarded. Titles and abstracts were read and 13 publications were selected for full-text reading. Twelve studies were included in the meta-analysis. The global analysis showed no statistically significant difference between control and experimental groups. In the subgroup analysis, only the chlorhexidine subgroup showed a statistically significant difference, where the control groups had higher bond strength than the experimental groups. Many studies on in vitro orthodontic bond strength fail to report test conditions that could affect their outcomes. The pooled in vitro data suggest that adding an antimicrobial agent to an orthodontic adhesive system does not influence bond strength to enamel. It is not possible to state which antimicrobial agent is better to be associated. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Formulation and Optimization of Polymeric Nanoparticles for Intranasal Delivery of Lorazepam Using Box-Behnken Design: In Vitro and In Vivo Evaluation

PubMed Central

Sharma, Deepak; Maheshwari, Dipika; Rana, Ravish; Bhatia, Shanu; Singh, Manisha; Gabrani, Reema; Sharma, Sanjeev K.; Ali, Javed; Sharma, Rakesh Kumar; Dang, Shweta

2014-01-01

The aim of the present study was to optimize lorazepam loaded PLGA nanoparticles (Lzp-PLGA-NPs) by investigating the effect of process variables on the response using Box-Behnken design. Effect of four independent factors, that is, polymer, surfactant, drug, and aqueous/organic ratio, was studied on two dependent responses, that is, z-average and % drug entrapment. Lzp-PLGA-NPs were successfully developed by nanoprecipitation method using PLGA as polymer, poloxamer as surfactant and acetone as organic phase. NPs were characterized for particle size, zeta potential, % drug entrapment, drug release behavior, TEM, and cell viability. Lzp-PLGA-NPs were characterized for drug polymer interaction using FTIR. The developed NPs showed nearly spherical shape with z-average 167–318 d·nm, PDI below 0.441, and −18.4 mV zeta potential with maximum % drug entrapment of 90.1%. In vitro drug release behavior followed Korsmeyer-Peppas model and showed initial burst release of 21.7 ± 1.3% with prolonged drug release of 69.5 ± 0.8% from optimized NPs up to 24 h. In vitro drug release data was found in agreement with ex vivo permeation data through sheep nasal mucosa. In vitro cell viability study on Vero cell line confirmed the safety of optimized NPs. Optimized Lzp-PLGA-NPs were radiolabelled with Technitium-99m for scintigraphy imaging and biodistribution studies in Sprague-Dawley rats to establish nose-to-brain pathway. PMID:25126544

Evaluation of polymerization shrinkage of resin cements through in vitro and in situ experiments

NASA Astrophysics Data System (ADS)

Franco, A. P. G. O.; Karam, L. Z.; Pulido, C. A.; Gomes, O. M. M.; Kalinowski, H. J.

2014-08-01

The aim of this study was to evaluate the behavior of two types of resin cements , conventional dual and dual self adhesive, through in vitro and in situ experiments. For the in vitro assay were selected two resin cements that were handled and dispensed over a mylar strip supported by a glass plate. The Bragg grating sensors were positioned and another portion of cement. was placed, covered by another mylar strip. For the in situ experiment 16 single-rooted teeth were selected who were divided into 2 groups: group 1 - conventional dual resin cement Relyx ARC and group 2 - dual self adhesive resin cement Relyx U200 ( 3M/ESPE ). The teeth were treated and prepared to receive the intracanal posts. Two Bragg grating sensors were recorded and introduced into the root canal at different apical and coronal positions. The results showed that the in vitro experiment presented similar values of polymerization shrinkage that the in situ experiment made in cervical position; whereas Relyx ARC resulted lower values compared to Relyx U200; and cervical position showed higher shrinkage than the apical.

In vitro and in vivo antithrombotic and cytotoxicity effects of ferulic acid.

PubMed

Choi, Jun-Hui; Park, Jong-Kook; Kim, Ki-Man; Lee, Hyo-Jeong; Kim, Seung

2018-01-01

We discovered recently in vitro and in vivo antithrombotic and cytotoxicity effects of ferulic acid. The cytotoxicity assays showed that ferulic acid (∼300 μg/mL) did not cause any significant toxicity on three cell lines, platelets, leukocytes, and erythrocytes. In vitro assays showed inhibitory effects of ferulic acid on thrombin (THR)- or collagen/epinephrine-stimulated platelet activation by inhibiting platelet aggregation, and decreasing clot retraction activity. The in vitro effect of ferulic acid on THR-stimulated platelet activation was proved by the decrease in the secretion of serotonin from the platelets. The anticoagulant effects of ferulic acid were confirmed by the prolongation of the intrinsic or/and extrinsic pathways and the delay of recalcification time in plasma coagulation. Ferulic acid had antithrombotic effect in acute thromboembolism model in vivo, and decreased the expression of α IIb β 3 /FIB and phosphorylation of AKT in THR-stimulated platelet activation in vivo, and their antithrombotic efficacies hold promise for therapeutic targeting in our ongoing studies. © 2017 Wiley Periodicals, Inc.

A three-dimensional skin equivalent reflecting some aspects of in vivo aged skin.

PubMed

Diekmann, Johanna; Alili, Lirija; Scholz, Okka; Giesen, Melanie; Holtkötter, Olaf; Brenneisen, Peter

2016-01-01

Human skin undergoes morphological, biochemical and functional modifications during the ageing process. This study was designed to produce a 3-dimensional (3D) skin equivalent in vitro reflecting some aspects of in vivo aged skin. Reconstructed skin was generated by co-culturing skin fibroblasts and keratinocytes on a collagen-glycosaminoglycan-chitosan scaffold, and ageing was induced by the exposition of fibroblasts to Mitomycin-C (MMC). Recently published data showed that MMC treatment resulted in a drug-induced accelerated senescence (DIAS) in human dermal fibroblast cultures. Next to established ageing markers, histological changes were analysed in comparison with in vivo aged skin. In aged epidermis, the filaggrin expression is reduced in vivo and in vitro. Furthermore, in dermal tissue, the amount of elastin and collagen is lowered in aged skin in vivo as well as after the treatment of 3D skin equivalents with MMC in vitro. Our results show histological signs and some aspects of ageing in a 3D skin equivalent in vitro, which mimics aged skin in vivo. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Molecular docking and ex vivo and in vitro anticholinesterase activity studies of Salvia sp. and highlighted rosmarinic acid.

PubMed

Demirezer, Lütfiye Ömür; Gürbüz, Perihan; Kelicen Uğur, Emine Pelin; Bodur, Mine; Özenver, Nadire; Uz, Ayse; Güvenalp, Zühal

2015-01-01

To evaluate acetylcholinesterase (AChE) inhibitory activity and antioxidant capacity of the major molecule from Salvia sp., rosmarinic acid, as a drug candidate molecule for treatment of Alzheimer disease (AD). The AChE inhibitory activity of different extracts from Salvia trichoclada, Salvia verticillata, and Salvia fruticosa was determined by the Ellman and isolated guinea pig ileum methods, and the antioxidant capacity was determined with DPPH. The AChE inhibitory activity of the major molecule rosmarinic acid was determined by in silico docking and isolated guinea pig ileum methods. The methanol extract of Salvia trichoclada showed the highest inhibition on AChE. The same extract and rosmarinic acid showed significant contraction responses on isolated guinea pig ileum. All the extracts and rosmarinic acid showed high radical scavenging capacities. Docking results of rosmarinic acid showed high affinity to the selected target, AChE. In this study in vitro and ex vivo studies and in silico docking research of rosmarinic acid were used simultaneously for the first time. Rosmarinic acid showed promising results in all the methods tested.

Tolerogenic Dendritic Cells Generated by In Vitro Treatment With SAHA Are Not Stable In Vivo.

PubMed

Thewissen, Kristof; Broux, Bieke; Hendriks, Jerome J A; Vanhees, Mandy; Stinissen, Piet; Slaets, Helena; Hellings, Niels

2016-01-01

The aim of this study is to examine whether the histone deacetylase inhibitor, suberoylanilide hydroxamic acid (SAHA), can generate dendritic cells (DCs) with a stable tolerogenic phenotype to counteract autoimmune responses in an animal model of multiple sclerosis. We investigated if the tolerogenic potency of DCs could be increased by continuous treatment during in vitro differentiation toward DCs compared to standard 24-h in vitro treatment of already terminally differentiated DCs. We show that in vitro treatment with SAHA reduces the generation of new CD11c(+) DCs out of mouse bone marrow. SAHA-generated DCs show reduced antigen-presenting function as evidenced by a reduction in myelin endocytosis, a decreased MHC II expression, and a failure to upregulate costimulatory molecules upon LPS challenge. In addition, SAHA-generated DCs display a reduction in proinflammatory cytokines and molecules involved in apoptosis induction, inflammatory migration, and TLR signaling, and they are less immunostimulatory compared to untreated DCs. We demonstrated that the underlying mechanism involves a diminished STAT1 phosphorylation and was independent of STAT6 activation. Although in vitro results were promising, SAHA-generated DCs were not able to alleviate the development of experimental autoimmune encephalomyelitis in mice. In vitro washout experiments demonstrated that the tolerogenic phenotype of SAHA-treated DCs is reversible. Taken together, while SAHA potently boosts tolerogenic properties in DCs during the differentiation process in vitro, SAHA-generated DCs were unable to reduce autoimmunity in vivo. Our results imply that caution needs to be taken when developing DC-based therapies to induce tolerance in the context of autoimmune disease.

In vitro production of Sudanese camel (Camelus dromedarius) embryos from epididymal spermatozoa and follicular oocytes of slaughtered animals.

PubMed

Abdelkhalek, A E; Gabr, Sh A; Khalil, W A; Shamiah, Sh M; Pan, L; Qin, G; Farouk, M H

2017-03-28

Application of assisted reproductive technology in camelidea, such as artificial insemination (AI) and embryo transfer, has been slow in comparison to that for other livestock species. In Egypt, there are few attempts to establish in vitro maturation (IVM) and fertilization (IVF) techniques in dromedary camel. The present study was carried out to produce Sudanese camel embryos using in vitro matured oocytes and epididymal spermatozoa. Dromedary camel ovaries were collected from abattoirs and then, the oocytes were aspirated from all the visible follicles on the ovarian surface (~2-8 mm in a diameter). Meanwhile, Fetal Dromedary Camel Serum (FDCS) was obtained from camel fetuses after slaughtering. Thereafter, only Cumulus Oocyte Complexes (COCs) were matured in vitro in the Tissue Culture Medium (TCM-199) complemented with 10% FDCS. Spermatozoa required for in vitro fertilization were collected from testes (epididymal cauda) of the slaughtered camel bulls. The results clearly showed that the maturation rate of oocytes at metaphase II was about 59.5% while the fertilization rate was around 70.4%. Intriguingly, the embryo rates determined were 13.1%, in 2-cell; 0.0%, in 4-cell; 34.7%, in 8-16% cell; 39.1%, in morula and 13.1% in a blastocyst stage. This study represented a successful in vitro production of Sudanese dromedary camel embryos from epididymal sperm cells and in vitro matured oocytes recovered from slaughtered camels.

Targeted antitumoral dehydrocrotonin nanoparticles with L-ascorbic acid 6-stearate.

PubMed

Frungillo, Lucas; Martins, Dorival; Teixeira, Sérgio; Anazetti, Maristela Conti; Melo, Patrícia da Silva; Durán, Nelson

2009-12-01

Tumoral cells are known to have a higher ascorbic acid uptake than normal cells. Therefore, the aim of this study was to obtain polymeric nanoparticles containing the antitumoral compound trans-dehydrocrotonin (DHC) functionalized with L-ascorbic acid 6-stearate (AAS) to specifically target this system tumoral cells. Nanoparticle suspensions (NP-AAS-DHC) were prepared by the nanoprecipitation method. The systems were characterized for AAS presence by thin-layer chromatography and for drug loading (81-88%) by UV-Vis spectroscopy. To further characterize these systems, in vitro release kinetics, size distribution (100-140 nm) and Zeta potential by photon-correlation spectroscopic method were used. In vitro toxicity against HL60 cells was evaluated by tetrazolium reduction and Trypan blue exclusion assays. Cell death by apoptosis was quantified and characterized by flow cytometry and caspase activity. Zeta potential analyses showed that the system has a negatively charged outer surface and also indicate that AAS is incorporated on the external surface of the nanoparticles. In vitro release kinetics assay showed that DHC loaded in nanoparticles had sustained release behavior. In vitro toxicity assays showed that NP-AAS-DHC suspension was more effective as an antitumoral than free DHC or NP-DHC and increased apoptosis induction by receptor-mediated pathway. 2009 Wiley-Liss, Inc. and the American Pharmacists Association

Reduction of adipogenesis and lipid accumulation by Taraxacum officinale (Dandelion) extracts in 3T3L1 adipocytes: an in vitro study.

PubMed

González-Castejón, Marta; García-Carrasco, Belén; Fernández-Dacosta, Raquel; Dávalos, Alberto; Rodriguez-Casado, Arantxa

2014-05-01

In this in vitro study, we have investigated the ability of Taraxacum officinale (dandelion) to inhibit adipocyte differentiation and lipogenesis in 3T3-L1 preadipocytes. HPLC analysis of the three plant extracts used in this study-leaf and root extracts and a commercial root powder-identified caffeic and chlorogenic acids as the main phenolic constituents. Oil Red O staining and triglyceride levels analysis showed decreased lipid and triglyceride accumulation, respectively. Cytotoxicity was assessed with the MTT assay showing non-toxic effect among the concentrations tested. DNA microarray analysis showed that the extracts regulated the expression of a number of genes and long non-coding RNAs that play a major role in the control of adipogenesis. Taken together, our results indicate that the dandelion extracts used in this study may play a significant role during adipogenesis and lipid metabolism, and thus, supporting their therapeutic interest as potential candidates for the treatment of obesity. Copyright © 2013 John Wiley & Sons, Ltd.

Bioanalytical method for in vitro metabolism study of repaglinide using 96-blade thin-film solid-phase microextraction and LC-MS/MS.

PubMed

Simões, Rodrigo Almeida; Bonato, Pierina Sueli; Mirnaghi, Fatemeh S; Bojko, Barbara; Pawliszyn, Janusz

2015-01-01

A high-throughput bioanalytical method using 96-blade thin film microextraction (TFME) and LC-MS/MS for the analysis of repaglinide (RPG) and two of its main metabolites was developed and used for an in vitro metabolism study. The target analytes were extracted from human microsomal medium by a 96-blade-TFME system employing the low-cost prototype 'SPME multi-sampler' using C18 coating. Method validation showed recoveries around 90% for all analytes and was linear over the concentration range of 2-1000 ng ml(-1) for RPG and of 2-500 ng ml(-1) for each RPG metabolite. The method was applied to an in vitro metabolism study of RPG employing human liver microsomes and proved to be very useful for this purpose.

Peroxidases, lignin and anatomy during in vitro and ex vitro rooting of gardenia (Gardenia jasminoides Ellis) microshoots.

PubMed

Hatzilazarou, Stefanos P; Syros, Thomas D; Yupsanis, Traianos A; Bosabalidis, Artemios M; Economou, Athanasios S

2006-07-01

In vitro and ex vitro rooting of gardenia (Gardenia jasminoides Ellis) microshoots with or without indolic-3-butyric acid (IBA) was studied in order to improve acclimatization of microplants after root formation and transplantation. Peroxidase (POD) activity and isoforms, lignin content and anatomical observations were evaluated in the course of the three interdependent phases (induction, initiation and expression) of microshoot rooting. Microshoots treated or not treated with IBA achieved high rooting percentages both in vitro and ex vitro. At the end of the 2-week acclimatization period, the percentage of surviving microplants ranged from 80% to 100%, for in vitro and ex vitro rooted microshoots, respectively. Microshoots rooted in vitro and ex vitro showed a relationship between rooting and POD activity but in a different time course. It appeared that root formation occurred after the microshoots had reached and passed a peak of maximum enzyme activity. In all treatments, electrophoretic analysis (native PAGE) of PODs revealed the appearance of one anionic and three cationic POD isoforms (C(1), C(3) and C(4)). An additional cationic POD isoform (C(2)) appeared only in the ex vitro rooting. The lignin content was similar in microshoots rooted both in vitro and ex vitro. The sequential anatomical changes during the rooting process were similar in both in vitro and ex vitro rooting treatments. In the case of in vitro rooting, pith cells had vacuoles entirely filled with a dark substance, while in the case of ex vitro rooting, pith cells contained many amyloplasts. The origin of the adventitious roots, in both rooting conditions, was located in the cambial ring. Roots with organized tissue systems emerged from the microshoot stem 10-14 days after the root induction treatments; on day 10 for rooting in vitro, while a 4-day delay was noted in microshoots rooted ex vitro.

Efficient sequence-specific isolation of DNA fragments and chromatin by in vitro enChIP technology using recombinant CRISPR ribonucleoproteins.

PubMed

Fujita, Toshitsugu; Yuno, Miyuki; Fujii, Hodaka

2016-04-01

The clustered regularly interspaced short palindromic repeats (CRISPR) system is widely used for various biological applications, including genome editing. We developed engineered DNA-binding molecule-mediated chromatin immunoprecipitation (enChIP) using CRISPR to isolate target genomic regions from cells for their biochemical characterization. In this study, we developed 'in vitro enChIP' using recombinant CRISPR ribonucleoproteins (RNPs) to isolate target genomic regions. in vitro enChIP has the great advantage over conventional enChIP of not requiring expression of CRISPR complexes in cells. We first showed that in vitro enChIP using recombinant CRISPR RNPs can be used to isolate target DNA from mixtures of purified DNA in a sequence-specific manner. In addition, we showed that this technology can be used to efficiently isolate target genomic regions, while retaining their intracellular molecular interactions, with negligible contamination from irrelevant genomic regions. Thus, in vitro enChIP technology is of potential use for sequence-specific isolation of DNA, as well as for identification of molecules interacting with genomic regions of interest in vivo in combination with downstream analysis. © 2016 The Authors. Genes to Cells published by Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.

A reproducible accelerated in vitro release testing method for PLGA microspheres.

PubMed

Shen, Jie; Lee, Kyulim; Choi, Stephanie; Qu, Wen; Wang, Yan; Burgess, Diane J

2016-02-10

The objective of the present study was to develop a discriminatory and reproducible accelerated in vitro release method for long-acting PLGA microspheres with inner structure/porosity differences. Risperidone was chosen as a model drug. Qualitatively and quantitatively equivalent PLGA microspheres with different inner structure/porosity were obtained using different manufacturing processes. Physicochemical properties as well as degradation profiles of the prepared microspheres were investigated. Furthermore, in vitro release testing of the prepared risperidone microspheres was performed using the most common in vitro release methods (i.e., sample-and-separate and flow through) for this type of product. The obtained compositionally equivalent risperidone microspheres had similar drug loading but different inner structure/porosity. When microsphere particle size appeared similar, porous risperidone microspheres showed faster microsphere degradation and drug release compared with less porous microspheres. Both in vitro release methods investigated were able to differentiate risperidone microsphere formulations with differences in porosity under real-time (37 °C) and accelerated (45 °C) testing conditions. Notably, only the accelerated USP apparatus 4 method showed good reproducibility for highly porous risperidone microspheres. These results indicated that the accelerated USP apparatus 4 method is an appropriate fast quality control tool for long-acting PLGA microspheres (even with porous structures). Copyright © 2015 Elsevier B.V. All rights reserved.

In vitro growth of Plasmodium falciparum in neonatal blood.

PubMed

Sauerzopf, Ulrich; Honkpehedji, Yabo J; Adgenika, Ayôla A; Feugap, Elianne N; Ngoma, Ghyslain Mombo; Mackanga, Jean-Rodolphe; Lötsch, Felix; Loembe, Marguerite M; Kremsner, Peter G; Mordmüller, Benjamin; Ramharter, Michael

2014-11-18

Children below the age of six months suffer less often from malaria than older children in sub-Saharan Africa. This observation is commonly attributed to the persistence of foetal haemoglobin (HbF), which is considered not to permit growth of Plasmodium falciparum and therefore providing protection against malaria. Since this concept has recently been challenged, this study evaluated the effect of HbF erythrocytes and maternal plasma on in vitro parasite growth of P. falciparum in Central African Gabon. Umbilical cord blood and peripheral maternal blood were collected at delivery at the Albert Schweitzer Hospital in Gabon. Respective erythrocyte suspension and plasma were used in parallel for in vitro culture. In vitro growth rates were compared between cultures supplemented with either maternal or cord erythrocytes. Plasma of maternal blood and cord blood was evaluated. Parasite growth rates were assessed by the standard HRP2-assay evaluating the increase of HRP2 concentration in Plasmodium culture. Culture of P. falciparum using foetal erythrocytes led to comparable growth rates (mean growth rate = 4.2, 95% CI: 3.5 - 5.0) as cultures with maternal red blood cells (mean growth rate =4.2, 95% CI: 3.4 - 5.0) and those from non-malaria exposed individuals (mean growth rate = 4.6, 95% CI: 3.8 - 5.5). Standard in vitro culture of P. falciparum supplemented with either maternal or foetal plasma showed both significantly lower growth rates than a positive control using non-malaria exposed donor plasma. These data challenge the concept of HbF serving as intrinsic inhibitor of P. falciparum growth in the first months of life. Erythrocytes containing HbF are equally permissive to P. falciparum growth in vitro. However, addition of maternal and cord plasma led to reduced in vitro growth which may translate to protection against clinical disease or show synergistic effects with HbF in vivo. Further studies are needed to elucidate the pathophysiology of innate and acquired protection against neonatal malaria.

Sol-gel processed porous silica carriers for the controlled release of diclofenac diethylamine.

PubMed

Czarnobaj, Katarzyna; Czarnobaj, Joanna

2008-10-01

Silica xerogels doped with diclofenac diethylamine were prepared by the sol-gel method from a hydrolysed tetraethoxysilane (TEOS) solution containing diclofenac diethylamine. Two different catalysts, drying conditions and levels of water content were used to alter the microstructure of the silica xerogels. The aim of this study was to determine the rate of Diclofenac release from the silica xerogels. This in vitro study showed that the sol-gel method is useful for entrapping Diclofenac in the pores of xerogels. It also showed that, in vitro, Diclofenac is released from the silica xerogel, through the pores, by diffusion. Base-catalysed gels proved to be much more effective than acid-catalyzed gels. (c) 2008 Wiley Periodicals, Inc.

[Characterization of a human cell line from an anaplastic carcinoma of the thyroid gland].

PubMed

Gioanni, J; Zanghellini, E; Mazeau, C; Zhang, D; Courdi, A; Farges, M; Lambert, J C; Duplay, H; Schneider, M

1991-11-01

A new cell line derived from a thyroid anaplastic carcinoma, CAL 62, has been established in culture. This line is constituted by highly tumorigenic cells. Their epithelial phenotype is stable in culture. Immunochemical staining for human thyroglobulin is negative. Cytogenetic analysis showed a gain of chromosome 20, the translocation i (14q), and breakpoints of centrometric chromatine. These results are similar to those previously reported by other investigators. CAL 62 radiosensibility has been studied. The survival curve of the in vitro irradiated cells has been adjusted with a linear-quadratic model. This cell line is thus showed to be radioresistant. Cell line CAL 62 constitutes an appropriate model for in vitro studies of thyroid anaplastic carcinoma.

The MTA family proteins as novel histone H3 binding proteins.

PubMed

Wu, Meng; Wang, Lina; Li, Qian; Li, Jiwen; Qin, Jun; Wong, Jiemin

2013-01-03

The nucleosome remodeling and histone deacetylase complex (Mi2/NRD/NuRD/NURD) has a broad role in regulation of transcription, DNA repair and cell cycle. Previous studies have revealed a specific interaction between NURD and histone H3N-terminal tail in vitro that is not observed for another HDAC1/2-containing complex, Sin3A. However, the subunit(s) responsible for specific binding of H3 by NURD has not been defined. In this study, we show among several class I HDAC-containing corepressor complexes only NURD exhibits a substantial H3 tail-binding activity in vitro. We present the evidence that the MTA family proteins within the NURD complex interact directly with H3 tail. Extensive in vitro binding assays mapped the H3 tail-binding domain to the C-terminal region of MTA1 and MTA2. Significantly, although the MTA1 and MTA2 mutant proteins with deletion of the C-terminal H3 tail binding domain were assembled into the endogenous NURD complex when expressed in mammalian cells, the resulting NURD complexes were deficient in binding H3 tail in vitro, indicating that the MTA family proteins are required for the observed specific binding of H3 tail peptide by NURD in vitro. However, chromatin fractionation experiments show that the NURD complexes with impaired MTA1/2-H3 tail binding activity remained to be associated with chromatin in cells. Together our study reveals a novel histone H3-binding activity for the MTA family proteins and provides evidence that the MTA family proteins mediate the in vitro specific binding of H3 tail peptide by NURD complex. However, multiple mechanisms are likely to contribute to the chromatin association of NURD complex in cells. Our finding also raises the possibility that the MTA family proteins may exert their diverse biological functions at least in part through their direct interaction with H3 tail.

The MTA family proteins as novel histone H3 binding proteins

PubMed Central

2013-01-01

Background The nucleosome remodeling and histone deacetylase complex (Mi2/NRD/NuRD/NURD) has a broad role in regulation of transcription, DNA repair and cell cycle. Previous studies have revealed a specific interaction between NURD and histone H3N-terminal tail in vitro that is not observed for another HDAC1/2-containing complex, Sin3A. However, the subunit(s) responsible for specific binding of H3 by NURD has not been defined. Results In this study, we show among several class I HDAC-containing corepressor complexes only NURD exhibits a substantial H3 tail-binding activity in vitro. We present the evidence that the MTA family proteins within the NURD complex interact directly with H3 tail. Extensive in vitro binding assays mapped the H3 tail-binding domain to the C-terminal region of MTA1 and MTA2. Significantly, although the MTA1 and MTA2 mutant proteins with deletion of the C-terminal H3 tail binding domain were assembled into the endogenous NURD complex when expressed in mammalian cells, the resulting NURD complexes were deficient in binding H3 tail in vitro, indicating that the MTA family proteins are required for the observed specific binding of H3 tail peptide by NURD in vitro. However, chromatin fractionation experiments show that the NURD complexes with impaired MTA1/2-H3 tail binding activity remained to be associated with chromatin in cells. Conclusions Together our study reveals a novel histone H3-binding activity for the MTA family proteins and provides evidence that the MTA family proteins mediate the in vitro specific binding of H3 tail peptide by NURD complex. However, multiple mechanisms are likely to contribute to the chromatin association of NURD complex in cells. Our finding also raises the possibility that the MTA family proteins may exert their diverse biological functions at least in part through their direct interaction with H3 tail. PMID:23286669

Activity of fosfomycin against nosocomial multiresistant bacterial pathogens from Croatia: a multicentric study

PubMed Central

Bielen, Luka; Likić, Robert; Erdeljić, Viktorija; Mareković, Ivana; Firis, Nataša; Grgić-Medić, Marijana; Godan, Ana; Tomić, Ivan; Hunjak, Blaženka; Markotić, Alemka; Bejuk, Danijela; Tičić, Vladimira; Balzar, Silvana; Bedenić, Branka

2018-01-01

Aim To determine in vitro susceptibility of multiresistant bacterial isolates to fosfomycin. Methods In this prospective in vitro study (local non-random sample, level of evidence 3), 288 consecutively collected multiresistant bacterial isolates from seven medical centers in Croatia were tested from February 2014 until October 2016 for susceptibility to fosfomycin and other antibiotics according to Clinical and Laboratory Standards Institute methodology. Susceptibility to fosfomycin was determined by agar dilution method, while disc diffusion were performed for in vitro testing of other antibiotics. Polymerase chain reaction and sequencing was performed for the majority of extended spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae (K. pneumoniae) and carbapenem-resistant isolates. Results The majority of 288 multiresistant bacterial isolates (82.6%) were susceptible to fosfomycin. The 236 multiresistant Gram-negative isolates showed excellent susceptibility to fosfomycin. Susceptibility rates were as follows: Escherichia coli ESBL 97%, K. pneumoniae ESBL 80%, Enterobacter species 85.7%, Citrobacter freundii 100%, Proteus mirabilis 93%, and Pseudomonas aeruginosa 60%. Of the 52 multiresistant Gram-positive isolates, methicillin-resistant Staphylococcus aureus showed excellent susceptibility to fosfomycin (94.4%) and vancomycin-resistant enterococcus showed low susceptibility to fosfomycin (31%). Polymerase chain reaction analysis of 36/50 ESBL-producing K. pneumoniae isolates showed that majority of isolates had CTX-M-15 beta lactamase (27/36) preceded by ISEcp insertion sequence. All carbapenem-resistant Enterobacter and Citrobacter isolates had blaVIM-1 metallo-beta-lactamase gene. Conclusion With the best in vitro activity among the tested antibiotics, fosfomycin could be an effective treatment option for infections caused by multiresistant Gram-negative and Gram-positive bacterial strains in the hospital setting. PMID:29740989

Development of molecular markers for zebrafish (Danio rerio) ovarian follicle growth assessment following in-vitro culture in cryopreservation studies.

PubMed

Anil, Siji; Rawson, David; Zhang, Tiantian

2018-05-29

Development of in vitro culture protocol for early stage ovarian follicles of zebrafish is important since cryopreserved early stage ovarian follicles would need to be matured in vitro following cryopreservation before they can be fertilised. Development of molecular markers for zebrafish (Danio rerio) ovarian follicle growth assessment following in vitro culture of early stage zebrafish ovarian follicles in ovarian tissue fragments is reported here for the first time although some work has been reported for in vitro culture of isolated early stage zebrafish ovarian follicles. The main aim of the present study was to develop molecular markers in an optimised in vitro culture protocol for stage I and stage II zebrafish ovarian follicles in ovarian tissue fragments. The effect of concentration of the hormones human chorionic gonadotropin and follicle stimulating hormones, and additives such as Foetal Bovine Serum and Bovine Serum Albumin were studied. The results showed that early stage zebrafish ovarian fragments containing stage I and stage II follicles which are cultured in vitro for 24 h in 20% FBS and 100mIU/ml FSH in 90% L-15 medium at 28 °C can grow to the size of stage II and stage III ovarian follicles respectively. More importantly the follicle growth from stage I to stage II and from stage II to stage III were confirmed using molecular markers such as cyp19a1a (also known as P450aromA) and vtg1 genes respectively. However, no follicle growth was observed following cryopreservation and in vitro culture. Copyright © 2018 Elsevier Inc. All rights reserved.

Quantifying statistical relationships between commonly used in vitro models for estimating lead bioaccessibility.

PubMed

Yan, Kaihong; Dong, Zhaomin; Liu, Yanju; Naidu, Ravi

2016-04-01

Bioaccessibility to assess potential risks resulting from exposure to Pb-contaminated soils is commonly estimated using various in vitro methods. However, existing in vitro methods yield different results depending on the composition of the extractant as well as the contaminated soils. For this reason, the relationships between the five commonly used in vitro methods, the Relative Bioavailability Leaching Procedure (RBALP), the unified BioAccessibility Research Group Europe (BARGE) method (UBM), the Solubility Bioaccessibility Research Consortium assay (SBRC), a Physiologically Based Extraction Test (PBET), and the in vitro Digestion Model (RIVM) were quantified statistically using 10 soils from long-term Pb-contaminated mining and smelter sites located in Western Australia and South Australia. For all 10 soils, the measured Pb bioaccessibility regarding all in vitro methods varied from 1.9 to 106% for gastric phase, which is higher than that for intestinal phase: 0.2 ∼ 78.6%. The variations in Pb bioaccessibility depend on the in vitro models being used, suggesting that the method chosen for bioaccessibility assessment must be validated against in vivo studies prior to use for predicting risk. Regression studies between RBALP and SRBC, RBALP and RIVM (0.06) (0.06 g of soil in each tube, S:L ratios for gastric phase and intestinal phase are 1:375 and 1:958, respectively) showed that Pb bioaccessibility based on the three methods were comparable. Meanwhile, the slopes between RBALP and UBM, RBALP and RIVM (0.6) (0.6 g soil in each tube, S:L ratios for gastric phase and intestinal phase are 1:37.5 and 1:96, respectively) were 1.21 and 1.02, respectively. The findings presented in this study could help standardize in vitro bioaccessibility measurements and provide a scientific basis for further relating Pb bioavailability and soil properties.

[The in vitro dissolution of total composition of the tablet of rhizomes of Ligusticum chuanxiong components and in vitro-in vivo correlation by the method of area under the absorbance-wavelength curve].

PubMed

Lai, Hong-qiang; Hu, Yue; Li, Xiao-dong

2015-06-01

To discuss the availability of evaluation on the dissolution studies of the multicomponents in traditional Chinese medicine, the in vitro dissolution of total composition of the tablet of rhizomes of Ligusticum chuanxiong components and its correlation with the in vivo were studied by the method of area under the absorbance-wavelength curve (AUAWC). Taken the tablet of rhizomes of Ligusticum chuanxiong components which is composed of sodium ferulate and ligustrazine hydrochloride as subject model, the dissolution tests were carried out with basket method. The plasma concentrations of tablets in different rats were determined by AUAWC at different interval times. The in vivo absorption percentage was calculated by Wagner-Nelson equation to evaluate the in vitro and in vivo correlation. According to the results, the cumulative dissolution in vitro of total composition of tablets of rhizomes of Ligusticum chuanxiong components at 60 min was 90.65% in water by AUAWC. The in vivo pharmacokinetics is fitted with an one-compartment model. The linear equation based on the cumulative dissolution rate (fr) and absorption percentage (fa) at 5, 10, 20, 30 and 60 min was fa = 0.819 7 fr+0.183 and the correlation coefficient was 0.959 5, which showed a good correlation between the in vitro dissolution and the in vivo absorption percentage. The method of AUAWC can be used accurately, feasibly and conveniently to evaluate the in vitro and in vivo correlation of total composition of tablets of rhizomes of Ligusticum chuanxiong components, which will provide better guidance to study the in vitro and in vivo correlation of sustained release preparation etc under complex system of traditional Chinese medicine in the future.

In vitro contraction of cytokinetic ring depends on myosin II but not on actin dynamics.

PubMed

Mishra, Mithilesh; Kashiwazaki, Jun; Takagi, Tomoko; Srinivasan, Ramanujam; Huang, Yinyi; Balasubramanian, Mohan K; Mabuchi, Issei

2013-07-01

Cytokinesis in many eukaryotes involves the contraction of an actomyosin-based contractile ring. However, the detailed mechanism of contractile ring contraction is not fully understood. Here, we establish an experimental system to study contraction of the ring to completion in vitro. We show that the contractile ring of permeabilized fission yeast cells undergoes rapid contraction in an ATP- and myosin-II-dependent manner in the absence of other cytoplasmic constituents. Surprisingly, neither actin polymerization nor its disassembly is required for contraction of the contractile ring, although addition of exogenous actin-crosslinking proteins blocks ring contraction. Using contractile rings generated from fission yeast cytokinesis mutants, we show that not all proteins required for assembly of the ring are required for its contraction in vitro. Our work provides the beginnings of the definition of a minimal contraction-competent cytokinetic ring apparatus.

An In Vitro and In Vivo Study of the α-Amylase Activity of Phaseolamin

PubMed Central

de Gouveia, Neire Moura; Alves, Fernanda Vieira; Furtado, Fabiana Barcelos; Scherer, Danielli Luana; Mundim, Antonio Vicente

2014-01-01

Abstract We evaluated the polypeptide profiles, inhibition of human salivary α-amylase activity, and hemagglutination properties of a commercial phaseolamin sample. We also performed an in vivo assay to investigate the effects of a commercial phaseolamin treatment (100, 500, or 1500 mg/kg) over 20 days on the glycemia, body weight, and serum biochemical parameters (total cholesterol, triglycerides, alanine aminotransferase, and aspartate aminotransferase) of nondiabetic and streptozotocin-induced diabetic rats. The in vitro evaluation showed defined protein profiles, low hemagglutination activity, and high α-amylase inhibition. None of the experimental groups treated with phaseolamin or acarbose showed decreases in body weight. Our data demonstrate that phaseolamin inhibits amylase activity in vitro, reduces blood glucose levels, decreases or attenuates some of the renal and hepatic effects of diabetes in streptozotocin-induced rats, and could therefore have therapeutic potential in the treatment or prevention of the complications of diabetes. PMID:24650210

In vitro and in vivo antioxidant activities of polysaccharide purified from aloe vera (Aloe barbadensis) gel.

PubMed

Kang, Min-Cheol; Kim, Seo Young; Kim, Yoon Taek; Kim, Eun-A; Lee, Seung-Hong; Ko, Seok-Chun; Wijesinghe, W A J P; Samarakoon, Kalpa W; Kim, Young-Sun; Cho, Jin Hun; Jang, Hyeang-Su; Jeon, You-Jin

2014-01-01

The in vitro and in vivo antioxidant potentials of a polysaccharide isolated from aloe vera gel were investigated. Enzymatic extracts were prepared from aloe vera gel by using ten digestive enzymes including five carbohydrases and five proteases. Among them, the highest yield was obtained with the Viscozyme extract and the same extract showed the best radical scavenging activity. An active polysaccharide was purified from the Viscozyme extract using ethanol-added separation and anion exchange chromatography. Purified aloe vera polysaccharide (APS) strongly scavenged radicals including DPPH, hydroxyl and alkyl radicals. In addition, APS showed a protective effect against AAPH-induced oxidative stress and cell death in Vero cells as well as in the in vivo zebrafish model. In this study, it is proved that both the in vitro and in vivo antioxidant potentials of APS could be further utilized in relevant industrial applications. Copyright © 2013 Elsevier Ltd. All rights reserved.

Oral exposure to polystyrene nanoparticles affects iron absorption

NASA Astrophysics Data System (ADS)

Mahler, Gretchen J.; Esch, Mandy B.; Tako, Elad; Southard, Teresa L.; Archer, Shivaun D.; Glahn, Raymond P.; Shuler, Michael L.

2012-04-01

The use of engineered nanoparticles in food and pharmaceuticals is expected to increase, but the impact of chronic oral exposure to nanoparticles on human health remains unknown. Here, we show that chronic and acute oral exposure to polystyrene nanoparticles can influence iron uptake and iron transport in an in vitro model of the intestinal epithelium and an in vivo chicken intestinal loop model. Intestinal cells that are exposed to high doses of nanoparticles showed increased iron transport due to nanoparticle disruption of the cell membrane. Chickens acutely exposed to carboxylated particles (50 nm in diameter) had a lower iron absorption than unexposed or chronically exposed birds. Chronic exposure caused remodelling of the intestinal villi, which increased the surface area available for iron absorption. The agreement between the in vitro and in vivo results suggests that our in vitro intestinal epithelium model is potentially useful for toxicology studies.

Paradoxical Growth of Candida dubliniensis Does Not Preclude In Vivo Response to Echinocandin Therapy ▿

PubMed Central

Mariné, Marçal; Pastor, F. Javier; Sahand, Ismail H.; Pontón, José; Quindós, Guillermo; Guarro, Josep

2009-01-01

Candida dubliniensis commonly shows paradoxical or trailing growth effects in vitro in the presence of echinocandins. We tested the in vitro activities of anidulafungin, caspofungin, and micafungin against clinical isolates of C. dubliniensis and evaluated the efficacy of these drugs in two murine models of systemic infection. The three echinocandins were similarly effective in the treatment of experimental disseminated infections with C. dubliniensis strains showing or not showing abnormal growth in vitro. PMID:19786599

Development of bupivacaine decorated reduced graphene oxide and its local anesthetic effect-In vivo study.

PubMed

Zhang, Zhi; Zhang, Xin; Li, Aixiang; Ma, Chuangen

2018-03-01

The present works aims to develop bupivacaine modified reduced graphene oxide (BPV/RGO), and comparative evaluation of their anesthetic effect with free bupivacaine (BPV). The prepared BPV/RGO was studied by using various spectroscopic and microscopic characterization studies. In vitro drug release from BPV/RGO was studied using HPLC analysis. The cytotoxicity of BPV/RGO was studied against fibroblast (3T3) cells. In vivo evaluation of anesthetic effects was performed on animal models. BPV/RGO showed a prolonged in vitro release and lower cytotoxicity when compared to free BPV. Also, BPV/RGO showed a significantly prolonged analgesic effect when compared to free BPV. Further, the prepared BPV/RGO drug delivery system demonstrated to function as gifted to overcome the drawbacks of free BPV and other available drug delivery systems by prolonging the anesthetic effect with poor cytotoxicity. Copyright © 2018. Published by Elsevier B.V.

Development of an Extended-Release Formulation for Apremilast and a Level A in Vitro-in Vivo Correlation Study in Beagle Dogs.

PubMed

Tang, Meiqiong; Hu, Ping; Huang, Shigui; Zheng, Qiang; Yu, Hao; He, Yun

2016-11-01

The primary objective of the present study was to develop extended-release matrix formulations of apremilast for the oral delivery and to study their in vitro and in vivo correlation. Five extended-release formulations containing hydroxypropylmethylcellulose (HPMC) as the retarding excipient with different release rate of apremilast were prepared. Dissolution tests of all the formulated tablets were performed in water, pH 4.0 and 6.8 buffer solutions. The in vitro release kinetics was studied and supported by Korsmeyer-Peppas's equation as it presented highest values of correlation coefficients (r 2 up to 0.966). Among all formulated tablets, F2 (HPMC 25%) and F4 (HPMC 35%) were selected to perform an in vivo study in beagle dogs to obtain various pharmacokinetic parameters, i.e., peak plasma concentration (C max ), time to peak plasma concentration (t max ), area under the plasma-concentration vs. time curve (AUC). Higher t max and t 1/2 , lower C max and elimination coefficient (K e ) were observed for both extended formulations compared to marketed immediate-release products (Otezla ® ). Level A in vitro-in vivo correlations were created with the help of Wagner-Nelson and numeric deconvolution methods. Both formulations showed good in vitro-in vivo correlations whose accuracies were further verified by an internal validation.

In vitro maturation and fertilization of oocytes from unstimulated ovaries in infertile women with polycystic ovary syndrome.

PubMed

Zhao, Jun-Zhao; Zhou, Wei; Zhang, Wei; Ge, Hong-Shan; Huang, Xue-Feng; Lin, Jin-Ju

2009-06-01

To evaluate the effects of in vitro maturation and fertilization of oocytes from unstimulated ovaries in infertile women with polycystic ovary syndrome (PCOS). Retrospective study. Reproductive Medicine Center, First Affiliated Hospital of Wenzhou Medical College, People's Republic of China. One hundred eighteen women with PCOS undergoing 152 cycles of in vitro maturation treatment. Oocyte retrieval was carried out by ultrasound-guided puncture on days 9-14 of the cycle. The oocytes were cultured in vitro using maturation culture medium, which consisted of M-199 + 20% fetal bovine serum (FBS) + 75 mIU/mL recombinant FSH +/- 0.5 IU/mL hCG. After the oocytes had matured in vitro, fertilization and embryo transfer were performed. Rates of clinical pregnancy, multiple pregnancies, and live birth. Relatively optimal laboratory results were obtained in this study. Embryo transfer was performed in 140 cycles, with a clinical pregnancy rate (PR) of 40.0% per transfer. Fifty-six babies have been born and there are 10 ongoing pregnancies. The overall multiple PR was 33.93%. Our results show that using in vitro matured oocytes from unstimulated ovaries could be offered as an alternative to conventional IVF in women with PCOS, and future work should address ways to decrease the incidence of multiple pregnancies.

Comparative study of lipophilic and hydrophilic antioxidants from in vivo and in vitro grown Coriandrum sativum.

PubMed

Dias, Maria Inês; Barros, Lillian; Sousa, Maria João; Ferreira, Isabel C F R

2011-06-01

Coriander is commonly used for medicinal purposes, food applications, cosmetics and perfumes. Herein, the production of antioxidants in vegetative parts (leaves and stems) of in vivo and in vitro grown samples was compared. In vitro samples were clone A- with notorious purple pigmentation in stems and leaves and clone B- green. Seeds were also studied as they are used to obtain in vivo and in vitro vegetative parts. Lipophilic (tocopherols, carotenoids and chlorophylls) and hydrophilic (sugars, ascorbic acid, phenolics, flavonols and anthocyanins) compounds were quantified. The antioxidant activity was evaluated by radical scavenging activity, reducing power and lipid peroxidation inhibition. The in vivo sample showed the highest antioxidant activity mainly due to its highest levels of hydrophilic compounds. Otherwise, in vitro samples, mainly clone A, gave the highest concentration in lipophilic compounds but a different profile when compared to the in vivo sample. Clones A and B revealed a lack of β-carotene, β- and δ-tocopherols, a decrease in α-tocopherol, and an increase in γ-tocopherol and clorophylls in comparison to the in vivo sample. In vitro culture might be useful to explore the plants potentialities for industrial applications, controlling environmental conditions to produce higher amounts of some bioactive products.

Can in vitro systems capture the characteristic differences between the flexion-extension kinematics of the healthy and TKA knee?

PubMed

Varadarajan, Kartik M; Harry, Rubash E; Johnson, Todd; Li, Guoan

2009-10-01

In vitro systems provide a powerful means to evaluate the efficacy of total knee arthroplasty (TKA) in restoring normal knee kinematics. The Oxford knee rig (OKR) and the robotic knee testing system (RKTS) represent two systems that have been extensively used to study TKA biomechanics. Nonetheless, a frequently asked question is whether in vitro simulations can capture the in vivo behavior of the knee. Here, we compared the flexion-extension kinematics of intact knees and knees after TKA tested on the OKR and RKTS, to results of representative in vivo studies. The goal was to determine if the in vitro systems could capture the key kinematic features of knees in healthy subjects and TKA patients. Results showed that the RKTS and the OKR can replicate the femoral rollback and 'screw home' tibial rotation between 0 degrees and 30 degrees flexion seen in healthy subjects, and the reduced femoral rollback and absence of 'screw home' motion in TKA patients. The RKTS also replicated the overall internally rotated position of the tibia beyond 30 degrees flexion. However, ability of the OKR to replicate the internally rotated position of the knee beyond 30 degrees flexion was inconsistent. These data could aid in validation of new in vitro systems and physiologic interpretations of in vitro results.

Stryphnodendron adstringens and purified tannin on Pythium insidiosum: in vitro and in vivo studies.

PubMed

Trolezi, Rodrigo; Azanha, Juliana Maziero; Paschoal, Natália Rodrigues; Chechi, Jéssica Luana; Dias Silva, Marcelo José; Fabris, Viciany Eric; Vilegas, Wagner; Kaneno, Ramon; Fernandes Junior, Ary; Bosco, Sandra de Moraes Gimenes

2017-02-23

Pythium insidiosum is the etiological agent of pythiosis, an emerging life-threatening infectious disease in tropical and subtropical regions. The pathogen is a fungus-like organism resistant to antifungal therapy, for this reason, most cases need extensive surgical debridments as treatment, but depending on the size and anatomical region of the lesion, such approach is unfeasible. We investigate the fungicidal effect and toxicity of crude bark extract of Stryphnodendron adstringens and commercially available tannin on Pythium insidiosum both in vitro and in vivo. Standardized fragments of mycelia of fifteen isolates of P. insidiosum were tested with different concentrations of bark extract (10 to 30% v/v) and tannin (0.5, 1.0 and 1.5 mg/mL). For in vivo study, fifteen rabbits were experimentally infected with zoospores of P. insidiosum and treated by oral and intralesional applications of bark extract and tannin. Acute toxicity tests with both substances were also performed in rats. In vitro studies showed fungicidal effect for both substances at different concentrations and the SEM showed alteration on the cell wall surface of the pathogen. All infected rabbits developed a firm nodular mass that reached around 90 mm 2 ninety days after inoculation, but neither the intralesional inoculation of tannin, nor the oral administration of crude extract and tannin were able to promote remission of the lesions. Lesions developed by rabbits presented an encapsulated abscess being quite different of naturally acquired pythiosis, which is characterized by ulcerated lesions. Since no toxicity was observed in rats or rabbits inoculated with these products, while in vitro experiments showed direct antifungal effect, therapeutic activity of S. adstringens and tannin should be clinically tested as an alternative for healing wounds in naturally acquired pythiosis.

Cross-study projections of genomic biomarkers: an evaluation in cancer genomics.

PubMed

Lucas, Joseph E; Carvalho, Carlos M; Chen, Julia Ling-Yu; Chi, Jen-Tsan; West, Mike

2009-01-01

Human disease studies using DNA microarrays in both clinical/observational and experimental/controlled studies are having increasing impact on our understanding of the complexity of human diseases. A fundamental concept is the use of gene expression as a "common currency" that links the results of in vitro controlled experiments to in vivo observational human studies. Many studies--in cancer and other diseases--have shown promise in using in vitro cell manipulations to improve understanding of in vivo biology, but experiments often simply fail to reflect the enormous phenotypic variation seen in human diseases. We address this with a framework and methods to dissect, enhance and extend the in vivo utility of in vitro derived gene expression signatures. From an experimentally defined gene expression signature we use statistical factor analysis to generate multiple quantitative factors in human cancer gene expression data. These factors retain their relationship to the original, one-dimensional in vitro signature but better describe the diversity of in vivo biology. In a breast cancer analysis, we show that factors can reflect fundamentally different biological processes linked to molecular and clinical features of human cancers, and that in combination they can improve prediction of clinical outcomes.

Transcranial Cavitation Detection in Primates during Blood-Brain Barrier Opening – A Performance Assessment Study

PubMed Central

Wu, Shih-Ying; Tung, Yao-Sheng; Marquet, Fabrice; Downs, Matthew Eric; Sanchez, Carlos Sierra; Chen, Cherry Chen; Ferrera, Vincent

2014-01-01

Focused ultrasound (FUS) has been shown promise in treating the brain locally and noninvasively. Transcranial passive cavitation detection (PCD) provides methodology of monitoring the treatment in real time, while the skull effects remain a major challenge for its translation to the clinic. In this study, we investigated the sensitivity, reliability, and limitations of PCD through primate (macaque and human) skulls in vitro. The results were further correlated with the in vivo macaque studies including the transcranial PCD calibration and real-time monitoring of BBB opening, with magnetic resonance imaging assessing the opening and safety. The stable cavitation doses using harmonics (SCDh) and ultraharmonics (SCDu), the inertial cavitation dose (ICD), and the cavitation signal-to-noise ratio (SNR) were quantified based on the PCD signals. Results showed that through the macaque skull the pressure threshold for detecting the SCDh remained the same as without the skull in place, while it increased for the SCDu and ICD; through the human skull, it increased for all cavitation doses. The transcranial PCD was found reliable both in vitro and in vivo when the transcranial cavitation SNR exceeded the 1-dB detection limit through the in vitro macaque (attenuation: 4.92 dB/mm) and human (attenuation: 7.33 dB/mm) skull. In addition, using long pulses enabled reliable PCD monitoring and facilitate BBB opening at low pressures. The in vivo results showed that the SCDh became detectable at pressures as low as 100 kPa; the ICD, at 250 kPa while it could occur at lower pressures; the SCDu, at 700 kPa and was less reliable at lower pressures. Real-time monitoring of PCD was further implemented during BBB opening, with successful and safe opening achieved at 250–600 kPa in both the thalamus and the putamen. In conclusion, this study shows that transcranial PCD in macaques in vitro and in vivo as well as humans in vitro is reliable by improving the cavitation SNR beyond the 1-dB detection limit. PMID:24859660

CHARACTERISTICS OF GROWTH OF SARCOMA AND CARCINOMA CULTIVATED IN VITRO

PubMed Central

Lambert, Robert A.; Hanes, Frederic M.

1911-01-01

1. The transplantable sarcomata of rats and mice grow very readily by the method of cultivating tissues in vitro. 2. Sarcomatous tissue grows in conformity to a type which may be regarded as characteristic for tissues of mesenchymal origin. 3. The growth of sarcoma cells in vitro consists in ameboid wandering into the surrounding plasma, karyokinetic proliferation. and evidences of active metabolism on the part of the cells. 4. Mouse carcinomata can be cultivated in vitro. The outgrowth of carcinoma cells assumes a sheet-like form, only one cell in thickness. They migrate into the plasma by ameboid movement, the advancing edge showing numerous prolongations of the cytoplasm into pseudopods. 5. Karyokinetic figures are frequently seen in growing carcinoma cells. The cells show evidences of active metabolism. 6. Both sarcoma and carcinoma cells cultivated in vitro show active phagocytosis; carmin particles placed in the plasma are taken up rapidly by the growing cells. PMID:19867430

In vitro antioxidant activity of extracts from the leaves of Abies pindrow Royle.

PubMed

Gupta, D; Bhardwaj, R; Gupta, R K

2011-01-01

Traditionally, the leaves of Abies pindrow Royle are employed as an ayurvedic remedy for fever, hypoglycaemic, respiratory and inflammatory conditions. In this study, dichloromethane, methanol and acetone extracts of A. pindrow leaves were analysed for their phytochemical content and in vitro antioxidant activities. The methanol extract exhibited highest antioxidant activity while acetone extract showed presence of relatively high total phenol and flavonoids contents. The present study provides evidence that extracts of Abies pindrow leaves are a potential source of natural antioxidants and could serve as a base for future drugs.

Controlled-release tablet formulation of isoniazid.

PubMed

Jain, N K; Kulkarni, K; Talwar, N

1992-04-01

Guar (GG) and Karaya gums (KG) alone and in combination with hydroxy-propylmethylcellulose (HPMC) were evaluated as release retarding materials to formulate a controlled-release tablet dosage form of isoniazid (1). In vitro release of 1 from tablets followed non-Fickian release profile with rapid initial release. Urinary excretion studies in normal subjects showed steady-state levels of 1 for 13 h. In vitro and in vivo data correlated (r = 0.9794). The studies suggested the potentiality of GG and KG as release retarding materials in formulating controlled-release tablet dosage forms of 1.

Comparative evaluation of single and bilayered lamotrigine floating tablets

PubMed Central

Lakshmi, PK; Sridhar, M; Shruthi, B

2013-01-01

Aim: The purpose of this study was to prepare lamotrigine (LM) bilayered and single layered floating tablets and to compare their release profiles. Materials and Methods: LM floating tablets were prepared by direct compression method. Drug, hydroxy propyl methyl cellulose K4M, lactose monohydrate and polyvinylpyrrolidone K30 constitute controlled release layer components and floating layer components includes polymers and sodium bicarbonate. The prepared tablets were evaluated for physicochemical parameters such as hardness, friability, weight variation, thickness, floating lag time (FLT), floating time, in vitro buoyancy study, in vitro release studies. The drug-polymer interaction was studied by fourier transform infrared and differential scanning calorimetry. Results and Discussion: The FLT of all the formulations were within the prescribed limits (<3 min). When ethyl cellulose was used as floating layer component, tablets showed good buoyancy effect but eroded within 6-8 h. Hence it was replaced with hydroxypropyl cellulose -M hydrophilic polymer, which showed good FLT and floating duration for 16 h. Formulation LFC4 was found to be optimized with dissolution profile of zero order kinetics showing fickian diffusion. A comparative study of bilayered and single layered tablets of LM showed a highest similarity factor of 83.03, difference factor of 2.74 and t-test (P < 0.05) indicates that there is no significant difference between them. Conclusion: Though bilayered tablet possess many advantages, single layered tablet would be economical, cost-effective and reproducible for large scale production in the industry. However, the results of present study demonstrated that the in vitro development of bilayered gastro retentive floating tablets with controlled drug release profile for LM is feasible. PMID:24167788

Cytologic Effects of Air Force Chemicals

DTIC Science & Technology

1980-11-01

Studies of DNA replication and repair in cell cultures have shown that hydrazine, although highly toxic to cells, does not damage DNA and thus...interfere directly with DNA replication in Chinese hamster ovary cells grown in vitro, nor does it affect DNA repair synthesis in CCL-185 human lung cells...vitro with chemicals and monitoring their effect on DNA replication and repair. This method has been used to show that the alkylating agents MMS and 4

In-line pressure-flow module for in vitro modelling of haemodynamics and biosensor validation

NASA Technical Reports Server (NTRS)

Koenig, S. C.; Schaub, J. D.; Ewert, D. L.; Swope, R. D.; Convertino, V. A. (Principal Investigator)

1997-01-01

An in-line pressure-flow module for in vitro modelling of haemodynamics and biosensor validation has been developed. Studies show that good accuracy can be achieved in the measurement of pressure and of flow, in steady and pulstile flow systems. The model can be used for development, testing and evaluation of cardiovascular-mechanical-electrical anlogue models, cardiovascular prosthetics (i.e. valves, vascular grafts) and pressure and flow biosensors.

In vitro study of LDL transport under pressurized (convective) conditions.

PubMed

Cancel, Limary M; Fitting, Andrew; Tarbell, John M

2007-07-01

It is difficult to assess the transport pathways that carry low-density lipoprotein (LDL) into the artery wall in vivo, and there has been no previous in vitro study that has examined transendothelial transport under physiologically relevant pressurized (convective) conditions. Therefore, we measured water, albumin, and LDL fluxes across bovine aortic endothelial cell (BAEC) monolayers in vitro and determined the relative contributions of vesicles, paracellular transport through "breaks" in the tight junction, and "leaky" junctions associated with dying or dividing cells. Our results show that leaky junctions are the dominant pathway for LDL transport (>90%) under convective conditions and that albumin also has a significant component of transport through leaky junctions (44%). Transcellular transport of LDL by receptor-mediated processes makes a minor contribution (<10%) to overall transport under convective conditions.

Bromelain enzyme from pineapple: in vitro activity study under different micropropagation conditions.

PubMed

Vilanova Neta, Jaci Lima; da Silva Lédo, Ana; Lima, Aloisio André Bonfim; Santana, José Carlos Curvelo; Leite, Nadjma Souza; Ruzene, Denise Santos; Silva, Daniel Pereira; de Souza, Roberto Rodrigues

2012-09-01

The aim of this work was to evaluate the activity of bromelain in pineapple plants (Ananas comosus var. Comosus), Pérola cultivar, produced in vitro in different culture conditions. This enzyme, besides its pharmacological effects, is also employed in food industries, such as breweries and meat processing. In this work, the enzymatic activity was evaluated in the tissues of leaves and stems of plants grown in culture medium without plant growth regulator. The most significant levels of bromelain were observed in leaf tissue after 4 months of culture in vitro in medium with a filter paper bridge, followed by medium gelled by the agar. The results of this study, regarding the different structures of the pineapple (leaves and stems) in vitro showed that the activity of bromelain varied depending on the culture conditions, the time and structure of which was quantified, ensuring a viable strategy in the production of seedlings with high levels of bromelain in subsequent phases of micropropagation.

Bioadhesive films containing benzocaine: correlation between in vitro permeation and in vivo local anesthetic effect.

PubMed

de Araujo, Daniele Ribeiro; Padula, Cristina; Cereda, Cíntia Maria Saia; Tófoli, Giovana Radomille; Brito, Rui Barbosa; de Paula, Eneida; Nicoli, Sara; Santi, Patrizia

2010-08-01

The aim of this work was to develop anesthetic bioadhesive films containing benzocaine and study their in vitro skin permeation and in vivo performance, in comparison with commercial formulations. Films containing 3% and 5% w/w of benzocaine were prepared and characterized by weight, drug content, thickness and morphology. In vitro permeation assays were performed in vertical diffusion cells using full-thickness pig ear skin as barrier. Intensity and duration of analgesia were evaluated in rats by tail-flick test, and skin histological analysis was carried out. Tail-flick test showed that the duration of benzocaine-induced analgesia was significantly prolonged with the films compared to commercial creams, in agreement with the higher in vitro permeation. Histological analysis of the rat tail skin did not reveal morphological tissue changes nor cell infiltration signs after application of the commercial creams or films. Results from our study indicate that the films developed in this work can be considered as innovative dermal/transdermal therapeutic systems for benzocaine local delivery.

Techniques to Evaluate Dental Erosion: A Systematic Review of Literature

PubMed Central

Joshi, Mahasweta; Joshi, Nikhil; Angadi, Prabhakar; Raikar, Sonal

2016-01-01

This article reviews different techniques for evaluating dental erosion, weighs the advantages and disadvantages of these techniques, and presents the latest trends in the study of dental erosion. In May 2014, an initial search was carried out in the PubMed/MEDLINE database of indexed journals from 1975 to 2013 using the following keywords: dental erosion; dental erosion In-vitro; and dental erosion in-vivo. Bibliographic citations from the papers found were then used to find other useful sources. The authors categorize the techniques into three classes: in-vitro, in-vivo and in-vitro/in-vivo. The article discusses the instrumentation required to use each of these techniques, as well as their rationale, merits and applications. The emergence of in-vitro/in-vivo techniques offers the potential to accurately quantify tooth wear in clinical situations. Cross-sectional as well as longitudinal studies show that these techniques will improve diagnosis, treatment planning and management of dental erosion. PMID:27891489

In vitro stabilization of a low-tin bone-imaging agent (99mTc-Sn-HEDP) by ascorbic acid.

PubMed

Tofe, A J; Francis, M D

1976-09-01

The presence of oxidants in the 99mTc-pertechnetate and of oxygen in diagnostic kits containing low concentrations of Sn(II) has a detrimental effect upon in vitro and in vivo stability. Maintaining a nitrogen atmosphere or increasing the Sn(II) concentration inhibits the formation of 99mTcO4-. However, the latter remedy is likely to cause uptake in the reticuloendothelial system and has been associated with false positive or negative brain scans. We used ascorbic acid (an antioxidant) to ensure the in vitro stability with the low-Sn(II) bone agent disodium etidronate. In vitro stability studies by instant thin-layer chromatography, using high-acitivity generators and "instant pertechnetate," yielded less than 2% free pertechnetate at 24 hr after preparation. Distribution studies in guinea pigs show neither altered distribution of the bone agent nor abnormal distribution of ascorbic acid, suggesting its sole function as a noncomplexing stabilizer.

In vitro co-cultures of Pinus pinaster with Bursaphelenchus xylophilus: a biotechnological approach to study pine wilt disease.

PubMed

Faria, Jorge M S; Sena, Inês; Vieira da Silva, Inês; Ribeiro, Bruno; Barbosa, Pedro; Ascensão, Lia; Bennett, Richard N; Mota, Manuel; Figueiredo, A Cristina

2015-06-01

Co-cultures of Pinus pinaster with Bursaphelenchus xylophilus were established as a biotechnological tool to evaluate the effect of nematotoxics addition in a host/parasite culture system. The pinewood nematode (PWN), Bursaphelenchus xylophilus, the causal agent of pine wilt disease (PWD), was detected for the first time in Europe in 1999 spreading throughout the pine forests in Portugal and recently in Spain. Plant in vitro cultures may be a useful experimental system to investigate the plant/nematode relationships in loco, thus avoiding the difficulties of field assays. In this study, Pinus pinaster in vitro cultures were established and compared to in vivo 1 year-old plantlets by analyzing shoot structure and volatiles production. In vitro co-cultures were established with the PWN and the effect of the phytoparasite on in vitro shoot structure, water content and volatiles production was evaluated. In vitro shoots showed similar structure and volatiles production to in vivo maritime pine plantlets. The first macroscopic symptoms of PWD were observed about 4 weeks after in vitro co-culture establishment. Nematode population in the culture medium increased and PWNs were detected in gaps of the callus tissue and in cavities developed from the degradation of cambial cells. In terms of volatiles main components, plantlets, P. pinaster cultures, and P. pinaster with B. xylophilus co-cultures were all β- and α-pinene rich. Co-cultures may be an easy-to-handle biotechnological approach to study this pathology, envisioning the understanding of and finding ways to restrain this highly devastating nematode.

Use of external metabolizing systems when testing for endocrine disruption in the T-screen assay

DOE Office of Scientific and Technical Information (OSTI.GOV)

Taxvig, Camilla, E-mail: camta@food.dtu.dk; Olesen, Pelle Thonning; Nellemann, Christine

2011-02-01

Although, it is well-established that information on the metabolism of a substance is important in the evaluation of its toxic potential, there is limited experience with incorporating metabolic aspects into in vitro tests for endocrine disrupters. The aim of the current study was a) to study different in vitro systems for biotransformation of ten known endocrine disrupting chemicals (EDs): five azole fungicides, three parabens and 2 phthalates, b) to determine possible changes in the ability of the EDs to bind and activate the thyroid receptor (TR) in the in vitro T-screen assay after biotransformation and c) to investigate the endogenousmore » metabolic capacity of the GH3 cells, the cell line used in the T-screen assay, which is a proliferation assay used for the in vitro detection of agonistic and antagonistic properties of compounds at the level of the TR. The two in vitro metabolizing systems tested the human liver S9 mix and the PCB-induced rat microsomes gave an almost complete metabolic transformation of the tested parabens and phthalates. No marked difference the effects in the T-screen assay was observed between the parent compounds and the effects of the tested metabolic extracts. The GH3 cells themselves significantly metabolized the two tested phthalates dimethyl phthalate (DMP) and diethyl phthalate (DEP). Overall the results and qualitative data from the current study show that an in vitro metabolizing system using liver S9 or microsomes could be a convenient method for the incorporation of metabolic and toxicokinetic aspects into in vitro testing for endocrine disrupting effects.« less

Liquid crystalline systems for transdermal delivery of celecoxib: in vitro drug release and skin permeation studies.

PubMed

Estracanholli, Eder André; Praça, Fabíola Silva Garcia; Cintra, Ana Beatriz; Pierre, Maria Bernadete Riemma; Lara, Marilisa Guimarães

2014-12-01

Liquid crystalline systems of monoolein/water could be a promising approach for the delivery of celecoxib (CXB) to the skin because these systems can sustain drug release, improve drug penetration into the skin layers and minimize side effects. This study evaluated the potential of these systems for the delivery of CXB into the skin based on in vitro drug release and skin permeation studies. The amount of CXB that permeated into and/or was retained in the skin was assayed using an HPLC method. Polarizing light microscopy studies showed that liquid crystalline systems of monoolein/water were formed in the presence of CXB, without any changes in the mesophases. The liquid crystalline systems decreased drug release when compared to control solution. Drug release was independent of the initial water content of the systems and CXB was released from cubic phase systems, irrespective of the initial water content. The systems released the CXB following zero-order release kinetics. In vitro drug permeation studies showed that cubic phase systems allowed drug permeation and retention in the skin layers. Cubic phase systems of monoolein/water may be promising vehicles for the delivery of CXB in/through the skin because it improved CXB skin permeation compared with the control solution.

[High non-specific binding of the beta(1) -selective radioligand 2-(125)I-ICI-H].

PubMed

Riemann, B; Law, M P; Kopka, K; Wagner, St; Luthra, S; Pike, V W; Neumann, J; Kirchhefer, U; Schmitz, W; Schober, O; Schäfers, M

2003-08-01

As results of cardiac biopsies suggest, myocardial beta(1) -adrenoceptor density is reduced in patients with chronic heart failure. However, changes in cardiac beta(2)-adrenoceptors vary. With suitable radiopharmaceuticals single photon emission computed tomography (SPECT) and positron emission tomography (PET) offer the opportunity to assess beta-adrenoceptors non-invasively. Among the novel racemic analogues of the established beta(1)-selective adrenoceptor antagonist ICI 89.406 the iodinated 2-I-ICI-H showed high affinity and selectivity to beta(1)-adrenoceptors in murine ventricular membranes. The aim of this study was its evaluation as a putative sub-type selective beta(1)-adrenergic radioligand in cardiac imaging. Competition studies in vitro and in vivo were used to investigate the kinetics of 2-I-ICI-H binding to cardiac beta-adrenoceptors in mice and rats. In addition, the radiosynthesis of 2-(125)I-ICI-H from the silylated precursor 2-SiMe(3)-ICI-H was established. The specific activity was 80 GBq/ micro mol, the radiochemical yield ranged from 70 to 80%. The unlabelled compound 2-I-ICI-H showed high beta(1)-selectivity and -affinity in the in vitro competition studies. In vivo biodistribution studies apparently showed low affinity to cardiac beta-adrenoceptors. The radiolabelled counterpart 2-(125)I-ICI-H showed a high degree of non-specific binding in vitro and no specific binding to cardiac beta(1)-adrenoceptors in vivo. Because of its high non-specific binding 2-(125)I-ICI-H is no suitable radiotracer for imaging in vivo.

Evaluation of high resolution ultrasound as a tool for assessing the 3D volume of blood clots during in vitro thrombolysis.

PubMed

Auboire, Laurent; Escoffre, Jean-Michel; Fouan, Damien; Jacquet, Jean-René; Ossant, Frédéric; Grégoire, Jean-Marc; Bouakaz, Ayache

2017-07-24

Thrombosis is a major cause of several diseases, i.e. myocardial infarction, cerebral stroke and pulmonary embolism. Thrombolytic therapies are required to induce fast and efficient recanalization of occluded vessels. To evaluate the in vitro efficacy of these thrombolytic strategies, measuring clot dissolution is essential. This study aimed to evaluate and validate high resolution ultrasound as a tool to assess the exact volume of clots in 3D and in real time during in vitro thrombolytic drug testing. This new method was validated by measuring the effects of concentration range of recombinant tissue type plasminogen activator on a blood clot during complete occlusion or 70% stenosis of a vessel. This study shows that high resolution ultrasound imaging allows for a real-time assessment of the 3D volume of a blood clot with negligible inter- and intra-operator variabilities. The conclusions drawn from this study demonstrate the promising potential of high resolution ultrasound imaging for the in vitro assessment of new thrombolytic drugs.

Influence of Bicarbonate/Low-GDP Peritoneal Dialysis Fluid (Bicavera) on In Vitro and Ex Vivo Epithelial-to-Mesenchymal Transition of Mesothelial Cells

PubMed Central

Fernández–Perpén, Antonio; Pérez–Lozano, María Luisa; Bajo, María–Auxiliadora; Albar–Vizcaino, Patricia; Correa, Pilar Sandoval; del Peso, Gloria; Castro, María–José; Aguilera, Abelardo; Ossorio, Marta; Peter, Mirjam E.; Passlick–Deetjen, Jutta; Aroeira, Luiz S.; Selgas, Rafael; López–Cabrera, Manuel; Sánchez–Tomero, J. Antonio

2012-01-01

♦ Background: Peritoneal membrane damage induced by peritoneal dialysis (PD) is largely associated with epithelial-to-mesenchymal transition (EMT) of mesothelial cells (MCs), which is believed to be a result mainly of the glucose degradation products (GDPs) present in PD solutions. ♦ Objectives: This study investigated the impact of bicarbonate-buffered, low-GDP PD solution (BicaVera: Fresenius Medical Care, Bad Homburg, Germany) on EMT of MCs in vitro and ex vivo. ♦ Methods: In vitro studies: Omentum-derived MCs were incubated with lactate-buffered standard PD fluid or BicaVera fluid diluted 1:1 with culture medium. Ex vivo studies: From 31 patients randomly distributed to either standard or BicaVera solution and followed for 24 months, effluents were collected every 6 months for determination of EMT markers in effluent MCs. ♦ Results: Culturing of MCs with standard fluid in vitro resulted in morphology change to a non-epithelioid shape, with downregulation of E-cadherin (indicative of EMT) and strong induction of vascular endothelial growth factor (VEGF) expression. By contrast, in vitro exposure of MCs to bicarbonate/low-GDP solution had less impact on both EMT parameters. Ex vivo studies partially confirmed the foregoing results. The BicaVera group, with a higher prevalence of the non-epithelioid MC phenotype at baseline (for unknown reasons), showed a clear and significant trend to gain and maintain an epithelioid phenotype at medium- and longer-term and to show fewer fibrogenic characteristics. By contrast, the standard solution group demonstrated a progressive and significantly higher presence of the non-epithelioid phenotype. Compared with effluent MCs having an epithelioid phenotype, MCs with non-epithelioid morphology showed significantly lower levels of E-cadherin and greater levels of fibronectin and VEGF. In comparing the BicaVera and standard solution groups, MCs from the standard solution group showed significantly higher secretion of interleukin 8 and lower secretion of collagen I, but no differences in the levels of other EMT-associated molecules, including fibronectin, VEGF, E-cadherin, and transforming growth factor β1. Peritonitis incidence was similar in both groups. Functionally, the use of BicaVera fluid was associated with higher transport of small molecules and lower ultrafiltration capacity. ♦ Conclusions: Effluent MCs grown ex vivo from patients treated with bicarbonate/low-GDP BicaVera fluid showed a trend to acquire an epithelial phenotype, with lower production of proinflammatory cytokines and chemokines (such as interleukin 8) than was seen with MCs from patients treated with a lactate-buffered standard PD solution. PMID:22215656

A comparison of the erosive potential of different beverages in primary and permanent teeth using an in vitro model.

PubMed

Lussi, A; Kohler, N; Zero, D; Schaffner, M; Megert, B

2000-04-01

The aim of this study was to compare the erosive potential of different beverages and foodstuffs in primary and permanent teeth. Sixty primary and 60 permanent human teeth were immersed for 3 min in the solution under study (5 teeth per treatment group). Surface microhardness was measured before and after exposure. Initial (baseline) surface microhardness was lower for primary teeth than for permanent teeth. In both primary and permanent teeth, Sprite showed the highest decrease in surface microhardness, whereas yogurt showed an increase in surface microhardness in the primary teeth. Overall decrease was 27.2 +/- 17.5 KHN (mean +/- SD) for primary and 25.9 +/- 15.6 KHN for permanent teeth. The comparison of the erosive susceptibility in this in vitro model showed that primary teeth were not more susceptible to erosion compared to permanent teeth.

Synthesis, spectroscopic, thermogravimetric and antimicrobial studies of mixed ligands complexes

NASA Astrophysics Data System (ADS)

Mahmoud, Walaa H.; Mahmoud, Nessma F.; Mohamed, Gehad G.; El-Sonbati, Adel Z.; El-Bindary, Ashraf A.

2015-09-01

An interesting series of mixed ligand complexes have been synthesized by the reaction of metal chloride with guaifenesin (GFS) in the presence of 2-aminoacetic acid (HGly) (1:1:1 molar ratio). The elemental analysis, magnetic moments, molar conductance, spectral (UV-Vis, IR, 1H NMR and ESR) and thermal studies were used to characterize the isolated complexes. The molecular structure of GFS is optimized theoretically and the quantum chemical parameters are calculated. The IR showed that the ligand (GFS) acts as monobasic tridentate through the hydroxyl, phenoxy etheric and methoxy oxygen atoms and co-ligand (HGly) as monobasic bidentate through the deprotonated carboxylate oxygen atom and nitrogen atom of amino group. The molar conductivities showed that all the complexes are non-electrolytes except Cr(III) complex is electrolyte. Electronic and magnetic data proposed the octahedral structure for all complexes under investigation. ESR spectrum for Cu(II) revealed data which confirm the proposed structure. Antibacterial screening of the compounds were carried out in vitro on gram positive (Bacillus subtilis and Staphylococcus aureus), gram negative (Escherichia coli and Neisseria gonorrhoeae) bacteria and for in vitro antifungal activity against Candida albicans organism. However, some complexes showed more chemotherapeutic efficiency than the parent GFS drug. The complexes were also screened for their in vitro anticancer activity against the breast cell line (MFC7) and the results obtained showed that they exhibit a considerable anticancer activity.

In-vitro and clinical evaluation of transurethral laser-induced prostatectomy (TULIP)

NASA Astrophysics Data System (ADS)

van Swol, Christiaan F. P.; Verdaasdonck, Rudolf M.; Mooibroek, Jaap; Boon, Tom A.

1993-05-01

Transurethral ultrasound-guided laser induced prostatectomy (TULIP) is a recent development in the treatment of benign prostatic hyperplasia. The system is based upon Nd:YAG laser irradiation delivered by a right angled fiber. The dosimetry used in a clinical situation is mostly based upon animal studies. In this study, the light and temperature distribution in the prostate during Nd:YAG laser irradiation were modeled using Monte Carlo and finite differences theory. The results of this model were compared with in vitro experiments. The influence of the different parameters involved, e.g., the scanning speed and the power of the laser beam, were evaluated. Initial results show the temperature distribution and thus the therapeutic effect of the TULIP procedure. Until now 36 patients have been treated successfully. The mean in-hospital time was somewhat shorter than for a TURP treatment while the results were comparable. These treatments, however, show the need for a better understanding of the mechanisms involved. Modeling and subsequent in vitro and in vivo measurements might improve the understanding and safe and successful application of prostate treatment using laser based systems.

Anti-protein aggregation is a potential target for preventing delayed neuronal death after transient ischemia.

PubMed

Ge, Pengfei; Luo, Yinan; Wang, Haifeng; Ling, Feng

2009-12-01

Brain ischemia has been an important risk factor for human being health, there is no effective medicine can be used to protect delayed neuronal injury or death secondary to blood reperfusion following ischemia. Recent discovery shows protein aggregation is an important factor resulting in ischemia-induced neuron death. Therefore, we propose the hypothesis that inhibiting protein aggregation may be an effective way to prevent delayed neuronal death after transient ischemia. At present, in vitro studies show some chemicals such as 4PBA (sodium 4-phenylbutyrate) and trehalose have the features of antagonizing protein aggregation in vitro. Moreover, polyQ-binding peptide (QBP1), geldanamycin, amino acids and amino acid derivatives have been also used in vitro to decrease aggregation and to increase protein stability. Although in vivo and systematical study should be performed to evaluate their effects of anti-protein aggregation, this enlightening us on using them to protect ischemic-induced neuronal death, and find new potential chemicals or methods which could be effective in keeping protein stable and prevent forming aggregates.

Onychopharmacokinetics of terbinafine hydrochloride penetration from a novel topical formulation into the human nail in vitro.

PubMed

Hui, Xiaoying; Lindahl, Åke; Lamel, Sonia; Maibach, Howard I

2013-09-01

This study determined the onychopharmacokinetics, nail absorption, distribution, and penetration of [¹⁴C]-terbinafine HCl in a new topical formulation into/through the human finger nail using the in vitro finite dose model. This study determined the penetration rate of terbinafine HCl from multiple doses of topical formulation applied daily for 14 days. Results showed that the total dose recovery (mass balance) was almost 100%. The concentration of terbinafine HCl in the deeper nail plate (ventral/intermediate layers) and the cotton-pad nail bed samples after the 14-day treatment were 613 ± 145 and (±S.D.) and 27 ± 1.2 µg/cm³ (or 1.9 ± 0.6 µg/cm³ daily) on average, respectively. In comparison with nail concentration data from the literature for other topical terbinatine formulations, our results show that higher amounts of terbinafine HCl reached the deep nail plate and/or the nail bed after a 14-day topical treatment with this topical formulation in vitro.

Effective inhibition of nasopharyngeal carcinoma in vitro and in vivo by targeting glycolysis with oxamate.

PubMed

Li, Xiaobing; Lu, Wenhua; Hu, Yumin; Wen, Shijun; Qian, Chaonan; Wu, Wenjing; Huang, Peng

2013-11-01

Elevated aerobic glycolysis in cancer cells (Warburg effect) has been observed in many tumor types including nasopharyngeal carcinoma (NPC), which can often be detected clinically using FDG-PET. However, the role of glycolysis in supporting the growth of NPC cells and its therapeutic implications still remain to be investigated. In the present study, we showed that the LDH inhibitor oxamate significantly suppressed NPC cell proliferation in vitro and tumor growth in vivo, yet exhibited minimum toxicity to normal nasopharyngeal epithelial cells in vitro and was well tolerated in mice. Moreover, oxamate exhibited cytotoxic effect in NPC cells under hypoxia. Mechanistic study showed that oxamate significantly inhibited LDH activity, leading to a substantial decrease in glucose uptake and lactate production. Combination of oxamate with a mitochondrial respiratory complex I inhibitor resulted in a significant depletion of cellular ATP and a synergistic killing of cancer cells. Our results suggest that inhibition of glycolysis by oxamate is an effective therapeutic strategy for treatment of NPC and that combination of this compound with mitochondrial-targeted agents may improve the therapeutic activity.

Anticoagulant effects of a Cannabis extract in an obese rat model.

PubMed

Coetzee, C; Levendal, R-A; van de Venter, M; Frost, C L

2007-05-01

Blood coagulation studies were conducted to determine the possible anti-/prothrombotic effect of an organic cannabis extract and the three major cannabinoids, THC, CBD and CBN. The in vitro effect of the cannabis extract on thrombin activity produced an IC50 value of 9.89 mg/ml, compared to THC at 1.79 mg/ml. It was also found that the extract, THC and CBN showed considerable inhibition of thrombin-induced clot formation in vitro with IC50 values of 600, 87 and 83 microg/ml for the extract, THC and CBN respectively. In an in vivo model used to determine clotting times of lean and obese rats treated with a cannabis extract, 50% clotting times were found to be 1.5 and 2 fold greater than their respective control groups, supporting the results obtained in the in vitro model. The study thus shows that Cannabis sativa and the cannabinoids, THC and CBN, display anticoagulant activity and may be useful in the treatment of diseases such as type 2 diabetes in which a hypercoagulable state exists.

Role of gastric antioxidant and anti-Helicobactor pylori activities in antiulcerogenic activity of plantain banana (Musa sapientum var. paradisiaca).

PubMed

Goel, R K; Sairam, K; Rao, C V

2001-07-01

Studies with plantain banana (Musa sapientum var. paradisiaca) have indicated its ulcer protective and healing activities through its predominant effect on various mucosal defensive factors [Sanyal et.al, Arch Int Pharmacodyn, 149 (1964) 393; 155 (1965) 244]. Oxidative stress and Helicobactorpylori colonization are considered to be important factors in the pathogenesis of gastric ulcers. In the present study methanolic extract of plantain banana pulp (BE) was evaluated for its (i) antiulcer and antioxidant activities in 2 hr cold restraint stress and (ii) anti-H.pylori activity in vitro. The extract (BE, 50 mg/kg, twice daily for 5 days) showed significant antiulcer effect and antioxidant activity in gastric mucosal homogenates, where it reversed the increase in ulcer index, lipid peroxidation and super oxide dismutase values induced by stress. However it did not produce any change in catalase values, which was significantly decreased by stress. Further, in the in vitro study. BE (0.32-1,000 microg/ml) did not show any anti-H.pylori activity. The results suggest absence of anti-H. pyloric activity of methanolic extract of banana in vitro and its antioxidant activity may be involved in its ulcerprotective activity.

Hollow fiber: a biophotonic implant for live cells

NASA Astrophysics Data System (ADS)

Silvestre, Oscar F.; Holton, Mark D.; Summers, Huw D.; Smith, Paul J.; Errington, Rachel J.

2009-02-01

The technical objective of this study has been to design, build and validate biocompatible hollow fiber implants based on fluorescence with integrated biophotonics components to enable in fiber kinetic cell based assays. A human osteosarcoma in vitro cell model fiber system has been established with validation studies to determine in fiber cell growth, cell cycle analysis and organization in normal and drug treated conditions. The rationale for implant development have focused on developing benchmark concepts in standard monolayer tissue culture followed by the development of in vitro hollow fiber designs; encompassing imaging with and without integrated biophotonics. Furthermore the effect of introducing targetable biosensors into the encapsulated tumor implant such as quantum dots for informing new detection readouts and possible implant designs have been evaluated. A preliminary micro/macro imaging approach has been undertaken, that could provide a mean to track distinct morphological changes in cells growing in a 3D matrix within the fiber which affect the light scattering properties of the implant. Parallel engineering studies have showed the influence of the optical properties of the fiber polymer wall in all imaging modes. Taken all together, we show the basic foundation and the opportunities for multi-modal imaging within an in vitro implant format.

New Coumarin Derivatives as Potent Selective COX-2 Inhibitors: Synthesis, Anti-Inflammatory, QSAR, and Molecular Modeling Studies.

PubMed

Dawood, Dina H; Batran, Rasha Z; Farghaly, Thoraya A; Khedr, Mohammed A; Abdulla, Mohamed M

2015-12-01

Two new series of coumarin derivatives incorporating thiazoline and thiazolidinone moieties were designed, synthesized, and investigated in vivo for their anti-inflammatory activities using the carrageenan-induced rat paw edema model and in vitro for their inhibitory activities against the human cyclooxygenase (COX)-1 and COX-2 isoforms. Most of the synthesized compounds demonstrated exceptionally high in vivo anti-inflammatory activity and displayed superior GI safety profiles (0-7% ulceration) as compared to indomethacin. All the bioactive compounds showed in vitro high affinity and selectivity toward the COX-2 isoenzyme, compared to the reference celecoxib with IC50 values ranging from 0.31 to 0.78 μM. The ethyl thiosemicarbazone 2b, thiazoline derivatives 3a, 3b, 5b, 6a, and 7f, and the thiazolidinone compounds 8b and 9a showed the highest in vivo and in vitro anti-inflammatory activities with remarkable COX-2 selectivity. Quantitative structure-activity relationship study (QSAR) was done and resulted in a highly predictive power R(2) (0.908). A molecular docking study revealed a relationship between the docking affinity and the biological results. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Assessing the Relevance of in vitro Studies in Nanotoxicology by Examining Correlations between in vitro and in vivo Data

PubMed Central

Han, Xianglu; Corson, Nancy; Wade-Mercer, Pamela; Gelein, Robert; Jiang, Jingkun; Sahu, Manoranjan; Biswas, Pratim; Finkelstein, Jacob N.; Elder, Alison; Oberdörster, Günter

2012-01-01

There is an urgent need for in vitro screening assays to evaluate nanoparticle (NP) toxicity. However, the relevance of in vitro assays is still disputable. We administered doses of TiO2 NPs of different sizes to alveolar epithelial cells in vitro and the same NPs by intratracheal instillation in rats in vivo to examine the correlation between in vitro and in vivo responses. The correlations were based on toxicity rankings of NPs after adopting NP surface area as dose metric, and response per unit surface area as response metric. Sizes of the anatase TiO2 NPs ranged from 3 to 100 nm. A cell-free assay for measuring reactive oxygen species (ROS) was used, and lactate dehydrogenase (LDH) release, and protein oxidation induction were the in vitro cellular assays using a rat lung Type I epithelial cell line (R3/1) following 24 hr incubation. The in vivo endpoint was number of PMNs in bronchoalveolar lavage fluid (BALF) after exposure of rats to the NPs via intratracheal instillation. Slope analyses of the dose response curves shows that the in vivo and in vitro responses were well correlated. We conclude that using the approach of steepest slope analysis offers a superior method to correlate in vitro with in vivo results of NP toxicity and for ranking their toxic potency. PMID:22487507

Raman spectroscopic approach to monitor the in vitro cyclization of creatine → creatinine

NASA Astrophysics Data System (ADS)

Gangopadhyay, Debraj; Sharma, Poornima; Singh, Sachin Kumar; Singh, Pushkar; Tarcea, Nicolae; Deckert, Volker; Popp, Jürgen; Singh, Ranjan K.

2015-01-01

The creatine → creatinine cyclization, an important metabolic phenomenon has been initiated in vitro at acidic pH and studied through Raman spectroscopic and DFT approach. The equilibrium composition of neutral, zwitterionic and protonated microspecies of creatine has been monitored with time as the reaction proceeds. Time series Raman spectra show clear signature of creatinine formation at pH 3 after ∼240 min at room temperature and reaction is faster at higher temperature. The spectra at pH 1 and pH 5 do not show such signature up to 270 min implying faster reaction rate at pH 3.

Evaluation of disintegrants functionality for orodispersible mini tablets.

PubMed

Soulairol, Ian; Chaheen, Mohammad; Tarlier, Nicolas; Aubert, Adrien; Bataille, Bernard; Sharkawi, Tahmer

2017-11-01

This work evaluates the functionalities of different superdisintegrants (SD) for manufacturing orodispersible mini tablets (ODMT) by direct compression. Twenty-three formulations varying in SD type, concentration, and lubricant were used to manufacture ODMT. The ODMT were then characterized for the following properties: friability, porosity, tensile strength, in vivo and in vitro disintegration time (DT). The results show that the presence, type, and concentration of SD did not influence friability, porosity, or tablet tensile strength. With regards to in vivo DT, only cross-linked poly (vinyl pyrrolidone) improved DT in all the tested formulations. Results also showed that when using microcrystalline cellulose (MCC) above 20% in the formulation, DT is longer. Cross-linked carboxymethyl cellulose accelerates DT when the MCC content is less than 20%. As for cross-linked carboxymethyl starch and calcium alginate showed no improvement on DT. Results for in vitro DT were all shorter than in vivo results and there was no correlation with the in vivo evaluation. This study shows that there is a need to develop better in vitro testing that precisely simulates in vivo conditions and that are adapted to ODMT. This standardization of the test methods for ODMTs must be accompanied by an improvement in the comprehension of SD mechanisms.

Effect of medium-chain triglycerides on the release behavior of Endostar encapsulated PLGA microspheres.

PubMed

Meng, Boyu; Li, Ling; Hua, Su; Wang, Qingsong; Liu, Chunhui; Xu, Xiangyang; Yin, Xiaojin

2010-09-15

The incomplete release of Endostar from PLGA microspheres was observed in our previous study. In the present study, we focused on the effect of medium-chain triglycerides (MCT) on the in vitro/in vivo release behavior of Endostar encapsulated PLGA microspheres, which were prepared by a water-in-oil-in-water (W/O/W) double-emulsion method with or without MCT. The in vitro accumulated release of Endostar from microspheres co-encapsulated with 30% MCT was found to be 79.04% after a 30-day incubation period in PBS (pH 7.4) at 37 degrees C. However, the accumulated release of Endostar from MCT-free microspheres was found to be only 32.22%. Pouches containing Endostar encapsulated PLGA microspheres were implanted subcutaneously in rats. The effect of MCT on the in vivo release showed a similar trend to the in vitro release. After 30 days, only 9.87% of the total encapsulated Endostar was retained in microspheres co-encapsulated with 30% MCT, while 42.25% of Endostar was retained in MCT-free microspheres. The co-encapsulation of MCT provided the microspheres with a porous surface, which significantly improved the in vitro/in vivo release of Endostar from PLGA microspheres. In addition, in vitro experiments showed that MCT co-encapsulated PLGA microspheres had more inter-connected pores, faster degradation of PLGA, and faster swelling of microspheres, which helped to explain the mechanism of the effect of MCT on improving the release of Endostar from PLGA microspheres. Copyright 2010 Elsevier B.V. All rights reserved.

In vitro kinetic interactions of DEET, pyridostigmine and organophosphorus pesticides with human cholinesterases.

PubMed

Wille, Timo; Thiermann, Horst; Worek, Franz

2011-04-25

The simultaneous use of the repellent DEET, pyridostigmine, and organophosphorus pesticides has been assumed as a potential cause for the Gulf War Illness and combinations have been tested in different animal models. However, human in vitro data on interactions of DEET with other compounds are scarce and provoked the present in vitro study scrutinizing the interactions of DEET, pyridostigmine and pesticides with human acetylcholinesterase (hAChE) and butyrylcholinesterase (hBChE). DEET showed to be a weak and reversible inhibitor of hAChE and hBChE. The IC(50) of DEET was calculated to be 21.7mM DEET for hAChE and 3.2mM DEET for hBChE. The determination of the inhibition kinetics of pyridostigmine, malaoxon and chlorpyrifos oxon with hAChE in the presence of 5mM DEET resulted in a moderate reduction of the inhibition rate constant k(i). The decarbamoylation velocity of pyridostigmine-inhibited hAChE was not affected by DEET. In conclusion, the in vitro investigation of interactions between human cholinesterases, DEET, pyridostigmine, malaoxon and chlorpyrifos oxon showed a weak inhibition of hAChE and hBChE by DEET. The inhibitory potency of the tested cholinesterase inhibitors was not enhanced by DEET and it did not affect the regeneration velocity of pyridostigmine-inhibited AChE. Hence, this in vitro study does not give any evidence of a synergistic effect of the tested compounds on human cholinesterases. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

A novel glycyrrhetinic acid-modified oxaliplatin liposome for liver-targeting and in vitro/vivo evaluation

PubMed Central

Chen, Jingde; Jiang, Hong; Wu, Yin; Li, Yandong; Gao, Yong

2015-01-01

In this study, oxaliplatin (OX) liposomes surface-modified with glycyrrhetinic acid (GA) were developed by the film-dispersion method. Their morphology, physical and chemical properties, and in vitro release performance were investigated. The transmission electron microscope (TEM) image showed that most liposomes were spherical particles with similar size and uniform dispersion. Both OX-liposomes and GA-OX-liposomes had an average size of 90 nm. They were negatively charged, with zeta potentials of −20.6 and −21.3 mV, respectively, and the entrapment efficiency values of both were higher than 94%. In vitro data showed that the application of liposomes could prolong the OX release. The relatively high correlation coefficient values obtained from analyzing the amount of drug released versus the square root of time depicted that release followed the Weibull model. No significant changes were observed after the addition of GA to the liposomes. In vivo, the relatively long time to reach the maximum plasma concentration of OX-liposomes suggested a sustained-release profile of liposomes, which was consistent with the results of the in vitro release study. The increased area under the curve and maximum plasma concentration of OX-liposomes and GA-OX-liposomes demonstrated an increased absorption. The drug concentration in tissues indicated that the GA-modified liposomes delivered OX mainly to liver after intravenous administration. In addition, no severe signs, such as appearance of epithelial necrosis or sloughing of epithelial cells, were detected in histology studies. PMID:25945038

In vitro nematicidal effect of Chenopodium ambrosioides and Castela tortuosa n-hexane extracts against Haemonchus contortus (Nematoda) and their anthelmintic effect in gerbils.

PubMed

Zamilpa, A; García-Alanís, C; López-Arellano, M E; Hernández-Velázquez, V M; Valladares-Cisneros, M G; Salinas-Sánchez, D O; Mendoza-de Gives, P

2018-05-06

The in vitro nematicidal effect of Chenopodium ambrosioides and Castela tortuosa n-hexane extracts (E-Cham and E-Cato, respectively) on Haemonchus contortus infective larvae (L3) and the anthelmintic effect of these extracts against the pre-adult stage of the parasite in gerbils were evaluated using both individual and combined extracts. The in vitro confrontation between larvae and extracts was performed in 24-well micro-titration plates. The results were considered 24 and 72 h post confrontation. The in vivo nematicidal effect was examined using gerbils as a study model. The extracts from the two assessed plants were obtained through maceration using n-hexane as an organic agent. Gerbils artificially infected with H. contortus L3 were treated intraperitoneally with the corresponding extract either individually or in combination. The results showed that the highest individual lethal in vitro effect (96.3%) was obtained with the E-Cham extract at 72 h post confrontation at 40 mg/ml, followed by E-Cato (78.9%) at 20 mg/ml after 72 h. The highest combined effect (98.7%) was obtained after 72 h at 40 mg/ml. The in vivo assay showed that the individual administration of the E-Cato and E-Cham extracts reduced the parasitic burden in gerbils by 27.1% and 45.8%, respectively. Furthermore, the anthelmintic efficacy increased to 57.3% when both extracts were administered in combination. The results of the present study show an important combined nematicidal effect of the two plant extracts assessed against L3 in gerbils.

Gastroprotective mechanism of Bauhinia thonningii Schum.

PubMed

Abdelwahab, Siddig Ibrahim; Taha, Manal Mohamed Elhassan; Abdulla, Mahmood Ameen; Nordin, Norazie; Hadi, A Hamid A; Mohan, Syam; Jayapalan, Jaime Jacqueline; Hashim, Onn Haji

2013-06-21

Bauhinia thonningii Schum. (Cesalpiniaceae) is locally known as Tambarib and used to treat various diseases including gastric ulcer. The current study aims to evaluate the gastroprotecive mechanism(s) of methanolic (MEBT) and chloroform (CEBT) extracts of Bauhinia thonningii leaves on ethanol-induced gastric ulceration. Gastric acidity, quantification and histochemistry of mucus, gross and microscopic examination, nitric oxide, lipid peroxidation, 2D gel electrophoresis, mass spectroscopy and biochemical tests were utilized to assess the mechanism(s) underlying the gastroprotective effects of MEBT and CEBT. Effect of these extracts into lipopolysaccharide/interferon-γ stimulated rodent cells were done in vitro. In vitro and in vivo toxicity studies were also conducted. Antioxidant activities of MEBT and CEBT were examined using DPPH, FRAP and ORAC assays. Phytochemical analyses of MEBT and CEBT were conducted using chemical and spectroscopic methods. Gross and histological features confirmed the anti-ulcerogenic properties of Bauhinia thonningii. Gastroprotective mechanism of MEBT was observed to be mediated through the modulation of PAS-reactive substances, MDA and proteomics biomarkers (creatine kinase, malate dehydrogenase, ATP synthase, actin and thioredoxin). MEBT and CEBT showed no significant in vitro and in vivo effects on nitric oxide. Methanolic extract (MEBT) showed superior gastroprotective effects, polyphenolic content and antioxidant activities compared to CEBT. The plant extracts showed no in vitro or in vivo toxicity. It could be concluded that MEBT possesses anti-ulcer activity, which could be attributed to the inhibition of ethanol-induced oxidative damage and the intervention in proteomic pathways but not the nitric oxide pathway. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Investigating the correlation between in vivo absorption and in vitro release of fenofibrate from lipid matrix particles in biorelevant medium.

PubMed

Borkar, Nrupa; Xia, Dengning; Holm, René; Gan, Yong; Müllertz, Anette; Yang, Mingshi; Mu, Huiling

2014-01-23

Lipid matrix particles (LMP) may be used as better carriers for poorly water-soluble drugs than liquid lipid carriers because of reduced drug mobilization in the formulations. However, the digestion process of solid lipid particles and their effect on the absorption of poorly water-soluble drugs are not fully understood. This study aimed at investigating the effect of particle size of LMP on drug release in vitro as well as absorption in vivo in order to get a better understanding on the effect of degradation of lipid particles on drug solubilisation and absorption. Fenofibrate, a model poorly water-soluble drug, was incorporated into LMP in this study using probe ultrasound sonication. The resultant LMP were characterised in terms of particle size, size distribution, zeta potential, entrapment efficiency, in vitro lipolysis and in vivo absorption in rat model. LMP of three different particle sizes i.e. approximately 100 nm, 400 nm, and 10 μm (microparticles) were produced with high entrapment efficiencies. The in vitro lipolysis study showed that the recovery of fenofibrate in the aqueous phase for 100 nm and 400 nm LMP was significantly higher (p<0.05) than that of microparticles after 30 min of lipolysis, suggesting that nano-sized LMP were digested to a larger extent due to greater specific surface area. The 100 nm LMP showed faster initial digestion followed by 400 nm LMP and microparticles. The area under the plasma concentration-time curve (AUC) following oral administration of 100 nm LMP was significantly higher (p<0.01) than that of microparticles and fenofibrate crystalline suspension (control). However, no significant difference was observed between the AUCs of 100 nm and 400 nm LMP. The same rank order on the in vivo absorption and the in vitro response was observed. The recovery (%) of fenofibrate partitioning into the aqueous phase during in vitro lipolysis and the AUC of plasma concentration-time curve of fenofibric acid was in the order of 100 nm LMP>microparticles>control. In summary, the present study demonstrated the particle size dependence of bioavailability of fenofibrate loaded LMP in rat model which correlates well with the in vitro drug release performed in the biorelevant medium. Copyright © 2013 Elsevier B.V. All rights reserved.

Transbuccal delivery of chlorpheniramine maleate from mucoadhesive buccal patches.

PubMed

Sekhar, K Chandra; Naidu, K V S; Vishnu, Y Vamshi; Gannu, Ramesh; Kishan, V; Rao, Y Madhusudan

2008-01-01

This article describes buccal permeation of chlorpheniramine maleate (CPM) and its transbuccal delivery using mucoadhesive buccal patches. Permeation of CPM was calculated in vitro using porcine buccal membrane and in vivo in healthy humans. Buccal formulations were developed with hydroxyethylcellulose (HEC) and evaluated for in vitro release, moisture absorption, mechanical properties, and bioadhesion, and optimized formulation was subjected for bioavailability studies in healthy human volunteers. In vitro flux of CPM was calculated to be 0.14 +/- 0.03 mg.h(-1).cm(-2) and buccal absorption also was demonstrated in vivo in human volunteers. In vitro drug release and moisture absorbed were governed by HEC content and formulations exhibited good tensile and mucoadhesive properties. Bioavailability from optimized buccal patch was 1.46 times higher than the oral dosage form and the results showed statistically significant difference.

Toxicity evaluation of chromium picolinate nanoparticles in vivo and in vitro in rat.

PubMed

Chen, Shih-Yi; Lien, Tu-Fa

2013-02-01

The toxicity of nanoCrpic is still not understood and needs further investigation. Thus, this study investigated the effect of chromium picolinate nanoparticles (nanoCrpic) on the toxicity in vivo and in vitro in rat. In the in vivo study, 36 rats (Wistar, 8-week-old) were randomly divided into the control group (fed basal diet), the low-dose (300 ppb, μg/kg), and high-dose (1,000 ppb) nanoCrpic groups. The trial was conducted for 2 months; at the final stage of the trial, the rats were sacrificed, liver and kidney were examined, and samples of tissues were taken for histological examination. Hepatocytes isolated from 10-week-old Wistar male rats were used for in vitro study to examine the degree of DNA damage following exposure to 0 and 0.294 mM of H(2)O(2) for 30 min. Incubation medium was supplemented with 0 (control), 100, and 300 ppb nanoCrpic. In vivo study indicated that no lesions of liver or kidney were detected in 300 and 1,000 ppb nanoCrpic fed rats. The in vitro study evaluated DNA damage according to the percentage and distance of the fragments migration and revealed that there was insignificant difference between the nanoCrpic and control groups (p > 0.05). This study indicated that nanoCrpic at 300-1,000 ppb in vivo and at 100-300 ppb in vitro showed no signs of toxicity to rats.

In Vitro Antimicrobial Susceptibility Patterns of Blastocystis

PubMed Central

Bush, Stephen; Ellis, John; Harkness, John; Stark, Damien

2015-01-01

Blastocystis is the most common human enteric protist with controversial clinical significance. Metronidazole is considered a first-line treatment for Blastocystis infection; however, there has been increasing evidence for the lack of efficacy of this treatment. Treatment failure has been reported in several clinical cases, and recent in vitro studies have suggested the occurrence of metronidazole-resistant strains. In this study, we tested 12 Blastocystis isolates from 4 common Blastocystis subtypes (ST1, ST3, ST4, and ST8) against 12 commonly used antimicrobials (metronidazole, paromomycin, ornidazole, albendazole, ivermectin, trimethoprim-sulfamethoxazole [TMP-SMX], furazolidone, nitazoxanide, secnidazole, fluconazole, nystatin, and itraconazole) at 10 different concentrations in vitro. It was found that each subtype showed little sensitivity to the commonly used metronidazole, paromomycin, and triple therapy (furazolidone, nitazoxanide, and secnidazole). This study highlights the efficacy of other potential drug treatments, including trimethoprim-sulfamethoxazole and ivermectin, and suggests that current treatment regimens be revised. PMID:25987633

Treatment of Recurrent Eczematous External Otitis with Honey Eardrops: A Proof-of-Concept Study.

PubMed

Henatsch, Darius; Nabuurs, Cindy H; van de Goor, Rens M; Wolffs, Petra F; Stokroos, Robert J

2017-10-01

Eczematous external otitis is a chronic inflammatory disease and often difficult to treat. Our objective was to investigate the clinical effect and in vitro antibacterial potential of medical honey eardrops as treatment of eczematous external otitis. In a prospective study, 15 patients diagnosed with recurrent eczematous external otitis were treated with medical honey eardrops for 2 weeks. The following clinical outcomes were evaluated: visual analog scale of ear complaints, score of eczema, and eradication of bacterial infection. Furthermore, the antibacterial effect of honey eardrops against different bacterial strains was tested in vitro. Treatment resulted in less discomfort and itching and decreased signs of eczema, with high patient satisfaction and without adverse reactions. Honey eardrops showed a strong in vitro inhibitory activity against all tested strains but did not eradicate Staphylococcus aureus infection in vivo. The results of this preliminary study indicate a possible role of honey eardrops in eczematous ear disease.

Cancer cells recovering from damage exhibit mitochondrial restructuring and increased aerobic glycolysis.

PubMed

Akakura, Shin; Ostrakhovitch, Elena; Sanokawa-Akakura, Reiko; Tabibzadeh, Siamak

2014-06-13

Instead of relying on mitochondrial oxidative phosphorylation, most cancer cells rely heavily on aerobic glycolysis, a phenomenon termed as "the Warburg effect". We considered that this effect is a direct consequence of damage which persists in cancer cells that recover from damage. To this end, we studied glycolysis and rate of cell proliferation in cancer cells that recovered from severe damage. We show that in vitro Damage-Recovered (DR) cells exhibit mitochondrial structural remodeling, display Warburg effect, and show increased in vitro and in vivo proliferation and tolerance to damage. To test whether cancer cells derived from tumor microenvironment can show similar properties, we isolated Damage-Recovered (T(DR)) cells from tumors. We demonstrate that T(DR) cells also show increased aerobic glycolysis and a high proliferation rate. These findings show that Warburg effect and its consequences are induced in cancer cells that survive severe damage. Copyright © 2014 Elsevier Inc. All rights reserved.

RNA-Seq transcriptome profiling of mouse oocytes after in vitro maturation and/or vitrification.

PubMed

Gao, Lei; Jia, Gongxue; Li, Ai; Ma, Haojia; Huang, Zhengyuan; Zhu, Shien; Hou, Yunpeng; Fu, Xiangwei

2017-10-16

In vitro maturation (IVM) and vitrification have been widely used to prepare oocytes before fertilization; however, potential effects of these procedures, such as expression profile changes, are poorly understood. In this study, mouse oocytes were divided into four groups and subjected to combinations of in vitro maturation and/or vitrification treatments. RNA-seq and in silico pathway analysis were used to identify differentially expressed genes (DEGs) that may be involved in oocyte viability after in vitro maturation and/or vitrification. Our results showed that 1) 69 genes were differentially expressed after IVM, 66 of which were up-regulated. Atp5e and Atp5o were enriched in the most significant gene ontology term "mitochondrial membrane part"; thus, these genes may be promising candidate biomarkers for oocyte viability after IVM. 2) The influence of vitrification on the transcriptome of oocytes was negligible, as no DEGs were found between vitrified and fresh oocytes. 3) The MII stage is more suitable for oocyte vitrification with respect to the transcriptome. This study provides a valuable new theoretical basis to further improve the efficiency of in vitro maturation and/or oocyte vitrification.

Impact of head and neck radiotherapy on the mechanical behavior of composite resins and adhesive systems: A systematic review.

PubMed

Madrid Troconis, Cristhian Camilo; Santos-Silva, Alan Roger; Brandão, Thaís Bianca; Lopes, Marcio Ajudarte; de Goes, Mario Fernando

2017-11-01

To analyze the evidence regarding the impact of head and neck radiotherapy (HNRT) on the mechanical behavior of composite resins and adhesive systems. Searches were conducted on PubMed, Embase, Scopus and ISI Web of Science databases using "Radiotherapy", "Composite resins" and "Adhesive systems" as keywords. Selected studies were written in English and assessed the mechanical behavior of composite resins and/or adhesive systems when bonding procedure was conducted before and/or after a maximum radiation dose ≥50Gy, applied under in vitro or in vivo conditions. In total, 115 studies were found but only 16 were included, from which five evaluated the effect of in vitro HNRT on microhardness, wear resistance, diametral tensile and flexural strength of composite resins, showing no significant negative effect in most of reports. Regarding bond strength of adhesive systems, 11 studies were included from which five reported no meaningful negative effect when bonding procedure was conducted before simulated HNRT. Conversely, five studies showed that bond strength diminished when adhesive procedure was done after in vitro radiation therapy. Only two studies about dental adhesion were conducted after in vivo radiotherapy but the results were not conclusive. The mechanical behavior of composite resins and adhesive systems seems not to be affected when in vitro HNRT is applied after bonding procedure. However, bond strength of adhesive systems tends to decrease when simulated radiotherapy is used immediately before bonding procedure. Studies assessing dentin bond strength after in-vivo HNRT were limited and controversial. Copyright © 2017 The Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

Novel Uses of In Vitro Data to Develop Quantitative Biological Activity Relationship Models for in Vivo Carcinogenicity Prediction.

PubMed

Pradeep, Prachi; Povinelli, Richard J; Merrill, Stephen J; Bozdag, Serdar; Sem, Daniel S

2015-04-01

The availability of large in vitro datasets enables better insight into the mode of action of chemicals and better identification of potential mechanism(s) of toxicity. Several studies have shown that not all in vitro assays can contribute as equal predictors of in vivo carcinogenicity for development of hybrid Quantitative Structure Activity Relationship (QSAR) models. We propose two novel approaches for the use of mechanistically relevant in vitro assay data in the identification of relevant biological descriptors and development of Quantitative Biological Activity Relationship (QBAR) models for carcinogenicity prediction. We demonstrate that in vitro assay data can be used to develop QBAR models for in vivo carcinogenicity prediction via two case studies corroborated with firm scientific rationale. The case studies demonstrate the similarities between QBAR and QSAR modeling in: (i) the selection of relevant descriptors to be used in the machine learning algorithm, and (ii) the development of a computational model that maps chemical or biological descriptors to a toxic endpoint. The results of both the case studies show: (i) improved accuracy and sensitivity which is especially desirable under regulatory requirements, and (ii) overall adherence with the OECD/REACH guidelines. Such mechanism based models can be used along with QSAR models for prediction of mechanistically complex toxic endpoints. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A review study on the effect of Iranian herbal medicines against in vitro replication of herpes simplex virus

PubMed Central

Moradi, Mohammad-Taghi; Rafieian-Kopaei, Mahmoud; Karimi, Ali

2016-01-01

Objective: There are a number of published data indicating in vitro anti-HSV activity of some of Iranian herbal extracts with no systematic review to discuss these results. Therefore, this article was aimed to review and discuss the methods carried out and the phytochemistry and bioactivity of the extracts used and also conclusions provided in these publications. Materials and Methods: Published articles both in English (from Medline, Science Direct, EMBASE, Scopus, Pro Quest, Google scholar, Cochrane Library) and in Persian (from SID, Iran Medex and Magiran) databases, from 1966 to October 2014 were incorporated in this review. The in vitro studies that lacked CC50, IC50, were excluded. Results: Only 42 published reports were found to examine Iranian herbs against HSV replication in vitro. Seventeen out of 42 studies in which 23 kinds of medicinal plants were subjected to crude extraction were included. The review of data showed that some of the herbal extracts including Hyssopus officinalis methanolic extract, Melissa officinalis aqueous extract, Quercus persica L. hydroalcoholic extract and Securigeras ecuridaca methanolic extract with selective index (SI) of 234, 877, >778 and 250, respectively were highly effective against HSV in vitro. Conclusion: More comprehensive studies using more advanced methods are needed to be done to achieve promising anti-HSV agents from the bioactive compounds isolated from these herbs. PMID:27761420

In vitro percutaneous absorption studies and in vivo evaluation of anti-inflammatory activity of essential fatty acids (EFA) from fish oil extracts.

PubMed

Puglia, Carmelo; Tropea, Salvatore; Rizza, Luisa; Santagati, Natale Alfredo; Bonina, Francesco

2005-08-11

The aim of the present study was to evaluate the in vitro percutaneous absorption and the in vivo anti-inflammatory activity of EPA and DHA fatty acids from three oily extracts, obtained by acetonic extractions from the entrails of different varieties of Mediterranean fishes such as mackerel (Scomber scombrus), sardine (Sardina pilchardus) and horse mackerel (Trachurus mediterraneus). In the first part of our research, we focused our attention on the characterization of the oily extracts to determine their omega-3 polyunsaturated fatty acid content, then, we evaluated the in vitro percutaneous absorption through excised human skin (stratum corneum/epidermis membranes; SCE) of EPA and DHA contained in the extracts. In the second part, the fish oil which guaranteed the best in vitro permeation profile of these omega-3 fatty acids was studied in order to evaluate its inhibiting ability towards the in vivo UVB-induced skin erythema. From the results obtained, all the fish oils tested in this study presented significant amounts of omega-3 fatty acids EPA and DHA, and particularly sardine oil extract showed higher concentrations of these substances compared to the other two fish oils. The in vitro experiments revealed interesting fluxes of these compounds from sardine extract through the stratum corneum/epidermis membranes and an appreciable anti-inflammatory activity against UVB-induced erythema in human volunteers was also observed.

In vitro molecular machine learning algorithm via symmetric internal loops of DNA.

PubMed

Lee, Ji-Hoon; Lee, Seung Hwan; Baek, Christina; Chun, Hyosun; Ryu, Je-Hwan; Kim, Jin-Woo; Deaton, Russell; Zhang, Byoung-Tak

2017-08-01

Programmable biomolecules, such as DNA strands, deoxyribozymes, and restriction enzymes, have been used to solve computational problems, construct large-scale logic circuits, and program simple molecular games. Although studies have shown the potential of molecular computing, the capability of computational learning with DNA molecules, i.e., molecular machine learning, has yet to be experimentally verified. Here, we present a novel molecular learning in vitro model in which symmetric internal loops of double-stranded DNA are exploited to measure the differences between training instances, thus enabling the molecules to learn from small errors. The model was evaluated on a data set of twenty dialogue sentences obtained from the television shows Friends and Prison Break. The wet DNA-computing experiments confirmed that the molecular learning machine was able to generalize the dialogue patterns of each show and successfully identify the show from which the sentences originated. The molecular machine learning model described here opens the way for solving machine learning problems in computer science and biology using in vitro molecular computing with the data encoded in DNA molecules. Copyright © 2017. Published by Elsevier B.V.

A new chemical approach to optimize the in vitro SPF method on the HD6 PMMA plate.

PubMed

Marguerie, S; Pissavini, M; Baud, A; Carayol, T; Doucet, O

2012-01-01

In a previous study, we demonstrated that control of the roughness of molded PMMA plates improves in vitro SPF reproducibility. However, in vitro/vivo deviations are still observed. Sunscreens show different behavior during spreading on the HD6 surface according to the formulation, resulting in a more or less homogenous distribution. The hydrophilic nature of HD6 appears to contribute significantly during spreading. Two different sunscreens offering a homogenous and non-homogenous distribution were investigated to check if the interfacial tension between product and substrate has a real influence on the spreading quality. Using microscopic observations, we attempted to correlate the in vitro SPF results with the product's spreading property. In order to reduce this interfacial tension, an HD6 pretreatment with an amphoteric surfactant, cocamidopropyl betain, was performed. In vitro SPF on "pretreated HD6" was examined using a cohort of 30 products. This pretreatment led to reliable results, demonstrating good association with the in vivo SPF.

In vitro degradation of neurotensin in human plasma.

PubMed

Lee, Y C; Uttenthal, L O; Smith, H A; Bloom, S R

1986-01-01

To study the degradation of neurotensin in plasma in vitro, fresh human plasma was incubated with neurotensin in the presence and absence of the peptidase inhibitors pepstatin A, EDTA, PMSF and aprotinin. The half-time of disappearance of neurotensin at 37 degrees C was calculated to be 226 min in vitro as opposed to 1.4 min in vivo when measured by radioimmunoassay with a C-terminally directed neurotensin antiserum. Both gel filtration and reversed phase high-pressure liquid chromatography (HPLC) showed that the main degradation product of neurotensin in human plasma in vitro was chromatographically and immunologically identical to neurotensin 1-8 and HPLC also demonstrated the formation of neurotensin 1-11. The loss of neurotensin incubated in human plasma in vitro was greatly reduced by EDTA but not by the other peptidase inhibitors tested. In this respect peptidase(s) responsible for the degradation of neurotensin in plasma differ from those present in brain homogenates. EDTA may be of importance in the preservation of neurotensin in plasma samples.

Bisphenol A release from orthodontic adhesives measured in vitro and in vivo with gas chromatography.

PubMed

Moreira, Marília Rodrigues; Matos, Leonardo Gontijo; de Souza, Israel Donizeti; Brigante, Tamires Amabile Valim; Queiroz, Maria Eugênia Costa; Romano, Fábio Lourenço; Nelson-Filho, Paulo; Matsumoto, Mírian Aiko Nakane

2017-03-01

The objectives of this study were to quantify in vitro the Bisphenol A (BPA) release from 5 orthodontic composites and to assess in vivo the BPA level in patients' saliva and urine after bracket bonding with an orthodontic adhesive system. For the in-vitro portion of this study, 5 orthodontic composites were evaluated: Eagle Spectrum (American Orthodontics, Sheboygan, Wis), Enlight (Ormco, Orange, Calif), Light Bond (Reliance Orthodontic Products, Itasca, Ill), Mono Lok II (Rocky Mountain Orthodontics, Denver, Colo), and Transbond XT (3M Unitek, Monrovia, Calif). Simulating intraoral conditions, the specimens were immersed in a water/ethanol solution, and the BPA (ng.g -1 ) liberation was measured after 30 minutes, 24 hours, 1 day, 1 week, and 1 month by the gas chromatography system coupled with mass spectrometry. Twenty patients indicated for fixed orthodontic treatment participated in the in-vivo study. Saliva samples were collected before bracket bonding and then 30 minutes, 24 hours, 1 day, 1 week, and 1 month after bonding the brackets. Urine samples were collected before bonding and then at 1 day, 1 week, and 1 month after bonding. The results were analyzed statistically using analysis of variance and Tukey posttest, with a significance level of 5%. All composites evaluated in vitro released small amounts of BPA. Enlight composite showed the greatest release, at 1 month. Regarding the in-vivo study, the mean BPA level in saliva increased significantly only at 30 minutes after bonding in comparison with measurements recorded before bonding. All orthodontic composites released BPA in vitro. Enlight and Light Bond had, respectively, the highest and lowest BPA releases in vitro. The in-vivo experiment showed that bracket bonding with the Transbond XT orthodontic adhesive system resulted in increased BPA levels in saliva and urine. The levels were significant but still lower than the reference dose for daily ingestion. Copyright © 2016 American Association of Orthodontists. Published by Elsevier Inc. All rights reserved.

Comparative evaluation of the antimicrobial activity of natural extracts of Morinda citrifolia, papain and aloe vera (all in gel formulation), 2% chlorhexidine gel and calcium hydroxide, against Enterococcus faecalis: An in vitro study

PubMed Central

Bhardwaj, Anuj; Ballal, Suma; Velmurugan, Natanasabapathy

2012-01-01

Aim: A comparative evaluation of the antimicrobial activity of natural extracts of Morinda citrifolia, papain, and aloe vera (all in gel formulations), 2% chlorhexidine gel and calcium hydroxide, against Enterococcus faecalis—an in vitro study. Materials and Methods: The antimicrobial efficacy was assessed in vitro using dentin shavings collected at 2 depths of 200 and 400 μm. The total colony forming units at the end of 1, 3, and 5 days were assessed. Results: The overall percentage inhibition of bacterial growth (200 and 400 μm depth) was 100% with chlorhexidine gel. This was followed by M. citrifolia gel (86.02%), which showed better antimicrobial efficacy as compared with aloe vera gel (78.9%), papain gel (67.3%), and calcium hydroxide (64.3%). There was no statistical difference between data at 200 and 400 μm depth. Conclusion: Chlorhexidine gel showed the maximum antimicrobial activity against E. faecalis, whereas calcium hydroxide showed the least. Among the natural intracanal medicaments, M. citrifolia gel consistently exhibited good inhibition up to the 5th day followed by aloe vera gel and papain gel. PMID:22876022

In Vivo and In Vitro Toxicity Evaluation of Hydroethanolic Extract of Kalanchoe brasiliensis (Crassulaceae) Leaves

PubMed Central

Lima, Adley Antoninni Neves; Soares, Luiz Alberto Lira

2018-01-01

The species Kalanchoe brasiliensis, known as “Saião,” has anti-inflammatory, antimicrobial, and antihistamine activities. It also has the quercetin and kaempferol flavonoids, which exert their therapeutic activities. With extensive popular use besides the defined therapeutical properties, the study of possible side effects is indispensable. The objective of this study is to evaluate the toxicity in vitro and in vivo from the hydroethanolic extract of the leaves of K. brasiliensis. The action of the extract (concentrations from 0.1 to 1000 uL/100 uL) in normal and tumor cells was evaluated using the MTT method. Acute toxicity and subchronic toxicity were evaluated in mice with doses of 250 to 1000 mg/kg orally, following recognized protocols. The in vitro results indicated cytotoxic activity for 3T3 cell line (normal) and 786-0 (kidney carcinoma), showing the activity to be concentration-dependent, reaching 92.23% cell inhibition. In vivo, the extract showed no significant toxicity; only liver changes related to acute toxicity and some signs of liver damage, combining biochemical and histological data. In general, the extract showed low or no toxicity, introducing itself as safe for use with promising therapeutic potential. PMID:29593788

Genotoxicity of tungsten carbide-cobalt (WC-Co) nanoparticles in vitro: mechanisms-of-action studies.

PubMed

Moche, Hélène; Chevalier, Dany; Vezin, Hervé; Claude, Nancy; Lorge, Elisabeth; Nesslany, Fabrice

2015-02-01

We showed previously that tungsten carbide-cobalt (WC-Co) nanoparticles (NP) can be used as a nanoparticulate positive control in some in vitro mammalian genotoxicity assays. Here, we investigate the mechanisms of action involved in WC-Co NP genotoxicity in L5178Y mouse lymphoma cells and primary human lymphocytes, in vitro. Data from the micronucleus assay coupled with centromere staining and from the chromosome-aberration assay show the involvement of both clastogenic and aneugenic events. Experiments with the formamidopyrimidine DNA glycosylase (FPG)-modified comet assay showed a slight (non-significant) increase in FPG-sensitive sites in the L5178Y mouse lymphoma cells but not in the human lymphocytes. Electron paramagnetic resonance spin-trapping results showed the presence of hydroxyl radicals (•OH) in WC-Co NP suspensions, with or without cells, but with time-dependent production in the presence of cells. However, a significant difference in •OH production was observed between human lymphocytes from two different donors. Using H2O2, we showed that WC-Co NP can participate in Fenton-like reactions. Thus, •OH might be produced either via intrinsic generation by WC-Co NP or through a Fenton-like reaction in the presence of cells. Copyright © 2015 Elsevier B.V. All rights reserved.

Effect of chosen lactic acid bacteria strains on Staphylococcus aureus in vitro as well as in meat and raw sausages.

PubMed

Gomółka-Pawlicka, M; Uradziński, J; Wiszniewska, A

2004-01-01

The present study was aimed at determining the influence of 15 strains of lactic acid bacteria on the growth of 2 Staphylococcus aureus strains in vitro as well as in meat and raw sausages. The investigations were performed within the framework of three alternate stages which differed in respect to the products studied, the number of Lactobacillus sp. strains and, partly, methodological approach. The study also considered water activity (a(w)) and pH of the products investigated. The results obtained are demonstrated in 5 diagrams. It was found that among 15 strains of Lactobacillus aureus investigated only one strain, Lactobacillus helveticus T 78, showed antagonistic effect on studied strains of Staphylococcus aureus both in vitro as well as in meat and raw sausages. Five other strains of Lactobacillus spp. displayed the antagonistic effect in vitro only. The temperature and incubation time of sausages, but also the type of sausage stuffing were found to have a distinct or slight influence, respectively, on the antagonistic interaction between the bacteria. However, this phenomenon was affected by neither a(w) nor pH.

Cell adhesion in zebrafish myogenesis: distribution of intermediate filaments, microfilaments, intracellular adhesion structures and extracellular matrix.

PubMed

Costa, Manoel L; Escaleira, Roberta C; Jazenko, Fernanda; Mermelstein, Claudia S

2008-10-01

To overcome the limitations of in vitro studies, we have been studying myogenesis in situ in zebrafish embryos, at a sub-cellular level. While in previous works we focused on myofibrillogenesis and some aspects of adhesion structures, here we describe in more detail cell adhesion structures and interactions among cytoskeletal components, membrane and extracellular matrix during zebrafish muscle development. We studied the intermediate filaments, and we describe the full range of desmin distribution in zebrafish development, from perinuclear to striated, until its deposition around the intersomite septa of older somites. This adhesion structure, positive for desmin and actin, has not been previously observed in myogenesis in vitro. We also show that actin is initially located in the intersomite septum region whereas it is confined to the myofibrils later on. While actin localization changes during development, the adhesion complex proteins vinculin, paxillin, talin, dystrophin, laminin and fibronectin always appear exclusively at the intersomite septa, and appear to be co-distributed, even though the extracellular proteins accumulates before the intracellular ones. Contrary to the adhesion proteins, that are continuously distributed, desmin and sarcomeric actin form triangular aggregates among the septa and the cytoskeleton. We studied the cytoskeletal linker plectin as well, and we show that it has a distribution similar to desmin and not to actin. We conclude that the in situ adhesion structures differ from their in vitro counterparts, and that the actual zebrafish embryo myogenesis is quite different than that which occurs in in vitro systems. Copyright 2008 Wiley-Liss, Inc.

Comparative study of the in vitro activity of various antifungal drugs against Scedosporium spp. in aerobic and hyperbaric atmosphere versus normal atmosphere.

PubMed

Farina, C; Marchesi, G; Passera, M; Diliberto, C; Russello, G

2012-06-01

Scedosporium spp. have been observed with increasing frequency over the last decade in immunocompromised patients and trauma patients. This mould is often multi-drug resistant and its mortality rate remains very high. The primary goal of this study was to obtain data concerning the in vitro susceptibility of 13 Scedosporium strains comparing the in vitro incubation in aerobic versus hyperbaric conditions. Chemosensitivity of thirteen Scedosporium strains was evaluated after a 72h-incubation in a normoxic (21% O2) normobaric (1 ATA) atmosphere versus a hyperoxic (100% O2) hyperbaric (2-3 ATA), and after a re-incubation at room temperature for an additional 72h. All S. apiospermum and S. prolificans strains showed no growth after incubation in hyperbaric hyperoxic atmosphere. However, when plates were then maintained at room temperature in aerobic conditions, growth was systematically observed from 36 to 96h, and Minimal inhibitory concentration (MIC) values were the same obtained after incubation in aerobic conditions. These results suggest impressive in vitro fungistatic activity of the hyperoxic hyperbaric atmosphere, even if its effect is strictly time-dependent. This preliminary in vitro study has potential clinical relevance because it focuses on examining in vitro combination therapy using hyperoxic hyperbaric conditions plus a single antifungal agent, rather than using combinations of different antifungal drugs, to potentially increase the antifungal activity. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

Comparative evaluation of antiplatelet effect of lycopene with aspirin and the effect of their combination on platelet aggregation: An in vitro study

PubMed Central

Sawardekar, Swapna B.; Patel, Tejal C.; Uchil, Dinesh

2016-01-01

Introduction: The objective was to compare antiplatelet effect of lycopene with aspirin and to study effect of combination of the two on platelet aggregation in vitro, using platelets from healthy volunteers. Materials and Methods: Platelets were harvested; platelet count of platelet-rich plasma adjusted to 2.5 Χ 105/μL. Aspirin (140 μmol/L) and lycopene (4, 6, 8, 10, and 12 μmol/L) were studied in vitro against adenosine-5’- diphosphate (ADP) (2.5 μM/L) and collagen Results: All the concentrations of lycopene (4–12 μmol/L) exhibited reduction in maximum platelet aggregation induced by aggregating agents ADP and collagen (P < 0.01 vs. vehicle) and were comparable with aspirin. Lycopene at concentration 10 μmol/L showed maximum platelet inhibition (47.05% ± 19.56%) against ADP, whereas lycopene at concentration 8 μmol/L showed maximum platelet inhibition (54.26% ± 30.71%) against collagen. Four μmol/L of lycopene combined with 140 μmol/L and 70 μmol/L aspirin showed greater inhibition of platelets as compared to aspirin 140 μmol/L alone, against both ADP and collagen. Conclusion: The study favorably compares lycopene and aspirin with respect to their antiplatelet activities against ADP and collagen. Lycopene can be considered as a potential target for modifying the thrombotic and pro-inflammatory events associated with platelet activation. PMID:26997718

Development, evaluation and pharmacokinetics of time-dependent ketorolac tromethamine tablets.

PubMed

Vemula, Sateesh Kumar; Veerareddy, Prabhakar Reddy

2013-01-01

The present study was intended to develop a time-dependent colon-targeted compression-coated tablets of ketorolac tromethamine (KTM) using hydroxypropyl methylcellulose (HPMC) that release the drug slowly but completely in the colonic region by retarding the drug releases in stomach and small intestine. KTM core tablets were prepared by direct compression method and were compression coated with HPMC. The formulation is optimized based on the in vitro drug release studies and further evaluated by X-ray imaging technique in healthy humans to ensure the colonic delivery. To prove these results, in vivo pharmacokinetic studies in human volunteers were designed to study the in vitro-in vivo correlation. From the in vitro dissolution study, optimized formulation F3 showed negligible drug release (6.75 ± 0.49%) in the initial lag period followed by slow release (97.47 ± 0.93%) for 24 h which clearly indicates that the drug is delivered to the colon. The X-ray imaging studies showed that the tablets reached the colon without disintegrating in upper gastrointestinal system. From the pharmacokinetic evaluation, the immediate-release tablets producing peak plasma concentration (C(max)) was 4482.74 ng/ml at 2 h T(max) and colon-targeted tablets showed C(max) = 3562.67 ng/ml at 10 h T(max). The area under the curve for the immediate-release and compression-coated tablets was 10595.14 and 18796.70 ng h/ml and the mean resident time was 3.82 and 10.75 h, respectively. Thus, the compression-coated tablets based on time-dependent approach were preferred for colon-targeted delivery of ketorolac.

In vitro study of antioxidant activity and phenolic content of Chrysanthemum balsamita varieties.

PubMed

Benedec, Daniela; Filip, Lorena; Vlase, Laurian; Bele, Constantin; Sevastre, Bogdan; Raita, Oana; Olah, Neli-Kinga; Hanganu, Daniela

2016-07-01

The purpose of our study was to identify the phenolic substances of two varieties of Chrysanthemum balsamita (balsamita and tanacetoides) and to measure the overall antioxidant activity. The phenolic compounds were determined by HPLC. The evaluation of the polyphenolic content was performed by colorimetric analysis. The antioxidant activity was measured by three in vitro assay models: the DPPH, the silver nanoparticles antioxidant capacity (SNPAC) and EPR radical detection. Using HPLC-MS analysis, phenolic acids, flavonoids and flavonoid aglycone were detected. The highest antioxidant activity was showed by Chrysanthemum balsamita var. balsamita, while the lowest for the Chrysanthemum balsamita var. tanacetoides extract, in accord with the polyphenolic content. The results show that Chrysanthemum balsamita var. balsamita might be a source of antioxidant flavonoids, especially rutin and isoquercitrin.

Nanocapsule-mediated cytosolic siRNA delivery for anti-inflammatory treatment.

PubMed

Jiang, Ying; Hardie, Joseph; Liu, Yuanchang; Ray, Moumita; Luo, Xiang; Das, Riddha; Landis, Ryan F; Farkas, Michelle E; Rotello, Vincent M

2018-06-05

The use of nanoparticle-stabilized nanocapsules for cytosolic siRNA delivery for immunomodulation in vitro and in vivo is reported. These NPSCs deliver siRNA directly to the cytosol of macrophages in vitro with concomitant knockdown of gene expression. In vivo studies showed directed delivery of NPSCs to the spleen, enabling gene silencing of macrophages, with preliminary studies showing 70% gene knockdown at a siRNA dose of 0.28 mg/kg. Significantly, the delivery of siRNA targeting tumor necrosis factor-α efficiently silenced TNF-α expression in LPS-challenged mice, demonstrating efficacy in modulating immune response in an organ-selective manner. This research highlights the potential of the NPSC platform for targeted immunotherapy and further manipulation of the immune system. Copyright © 2018 Elsevier B.V. All rights reserved.

Activities of Tannins--From In Vitro Studies to Clinical Trials.

PubMed

Sieniawska, Elwira

2015-11-01

Tannins are considered as valuable plant secondary metabolites providing many benefits for human health. In this review information was gathered about bioactivity in vitro and in vivo, as well as about conducted clinical trials. The literature research was based on ScienceDirect, Scopus, and Cochrane databases and presents a wide range of tested activities of tannins. The described clinical trials verify laboratory tests and show the effective health benefits taken from supplementation with tannins.

In vitro and in vivo effects of deferoxamine in neonatal acute leukemia.

PubMed

Estrov, Z; Tawa, A; Wang, X H; Dubé, I D; Sulh, H; Cohen, A; Gelfand, E W; Freedman, M H

1987-03-01

A six week old infant with acute leukemia failed to attain remission with chemotherapy. Because we previously demonstrated that the iron chelator deferoxamine (DFO) has antiproliferative properties and modulatory effects on cell differentiation, a protocol was designed for in vitro study and for clinical use in the patient. At diagnosis, blast cells were morphologically undifferentiated, had nondiagnostic cytochemistry, showed an abnormal karyotype (t[4;11]), expressed markers of B cell lineage, and demonstrated C mu gene rearrangement. Tissue culture of marrow or blood cells yielded colonies of leukemic blasts. Increasing concentrations of DFO produced a dose-dependent suppression of patient's blast colony growth in vitro, and blasts within colonies showed a marked change in surface antigen expression from lymphoid to myelomonocytic markers, became monocytic in appearance, and developed intense staining for nonspecific esterase. When DFO was given intravenously to the patient as a single agent for 48 hours, blasts no longer expressed lymphoid antigens and became strongly positive for myelomonocytic markers, identical to the in vitro findings. Intravenous DFO halted rising peripheral blood blast cell numbers and allowed a several-fold increase in normal hematopoietic progenitor colony growth. When combined with low-dose cytosine arabinoside in the treatment protocol, DFO caused striking leukemic cytoreduction. Our findings indicate that DFO has antileukemic properties by virtue of its effects on proliferation and differentiation, and they prompt further experimental and clinical studies with this agent.

An insight of in vitro transport of PEGylated non-ionic surfactant vesicles (NSVs) across the intestinal polarized enterocyte monolayers.

PubMed

Primavera, Rosita; Palumbo, Paola; Celia, Christian; Cinque, Benedetta; Carata, Elisabetta; Carafa, Maria; Paolino, Donatella; Cifone, Maria Grazia; Di Marzio, Luisa

2018-06-01

PEGylated non-ionic surfactant-based vesicles (NSVs) are promising drug delivery systems for the local, oral and systemic administrations of therapeutics. The aim of this study was to test the cellular biocompatibility and transport of Nile Red-loaded NSVs (NR-NSVs) across the Caco-2-cell monolayers, which represent an in vitro model of human intestinal epithelium. The NR-NSVs assumed a spherical shape with a mean size of 140 nm, and a narrow size distribution. The NR-NSVs did not modify Caco-2 cell viability, which remained unaltered in vitro up to a concentration of 1 mM. The transport studies demonstrated that the NR-NSVs moved across the Caco-2 monolayers without affecting the transepithelial electrical resistance. These results were supported by flow cytometry analysis, which demonstrated that NR-NSVs were internalized inside the Caco-2 cells. Nanoparticle tracking and Transmission Electron Microscopy (TEM) analysis showed the presence of NR-NSVs in the basolateral side of the Caco-2 monolayers. TEM images also showed that NSVs were transported intact across the Caco-2 monolayers, thus demonstrating a predominant transcytosis mechanism of transport through endocytosis. The NSVs did not affect the integrity of the membrane barrier in vitro, and can potentially be used in clinics to increase the oral bioavailability and delivery of therapeutics. Copyright © 2018 Elsevier B.V. All rights reserved.

Structural Characterization and In Vitro Antioxidant Activity of Kojic Dipalmitate Loaded W/O/W Multiple Emulsions Intended for Skin Disorders

PubMed Central

Marcussi, Diana Gleide; Calixto, Giovana Maria Fioramonti; Corrêa, Marcos Antonio

2015-01-01

Multiple emulsions (MEs) are intensively being studied for drug delivery due to their ability to load and increase the bioavailability of active lipophilic antioxidant, such as kojic dipalmitate (KDP). The aim of this study was to structurally characterize developed MEs by determining the average droplet size (Dnm) and zeta potential (ZP), performing macroscopic and microscopic analysis and analyzing their rheological behavior and in vitro bioadhesion. Furthermore, the in vitro safety profile and antioxidant activity of KDP-loaded MEs were evaluated. The developed MEs showed a Dnm of approximately 1 micrometer and a ZP of −13 mV, and no change was observed in Dnm or ZP of the system with the addition of KDP. KDP-unloaded MEs exhibited ‘‘shear thinning” flow behavior whereas KDP-loaded MEs exhibited Newtonian behavior, which are both characteristic of antithixotropic materials. MEs have bioadhesion properties that were not influenced by the incorporation of KDP. The results showed that the incorporation of KDP into MEs improved the safety profile of the drug. The in vitro antioxidant activity assay suggested that MEs presented a higher capacity for maintaining the antioxidant activity of KDP. ME-based systems may be a promising platform for the topical application of KDP in the treatment of skin disorders. PMID:25785265

Inhibitor of endocytosis impairs gene electrotransfer to mouse muscle in vivo.

PubMed

Markelc, Bostjan; Skvarca, Eva; Dolinsek, Tanja; Kloboves, Veronika Prevodnik; Coer, Andrej; Sersa, Gregor; Cemazar, Maja

2015-06-01

Application of electric pulses (electroporation/electropermeabilization) is an effective method for gene transfer (i.e. gene electrotransfer (GET)) in vitro and in vivo. Currently, the mechanisms by which the DNA enters the cell are not yet fully understood. Experimental evidence is building up that endocytosis is the main mechanism by which the DNA, which is later expressed, enters the cell. Therefore the aim of our study was to elucidate whether inhibitors of endocytosis, methyl-β-cyclodextrin (MβCD), Concanavalin A (ConA) and Dynasore, can impair the transfection efficacy of GET in vitro in B16F1 murine melanoma and in vivo in m. tibialis cranialis in mice. We show that MβCD--general inhibitor of endocytosis--can almost prevent GET of EGFP-N1 plasmid in vitro, that ConA--inhibitor of clathrin mediated endocytosis--also abrogates GET but to a lesser extent, and when using Dynasore--reversible inhibitor of dynamin--there is no effect on GET efficacy, if endocytosis is blocked for only 5 min after GET. Moreover, MβCD also reduced GET efficacy in vivo in m. tibialis cranialis and this effect was long lasting. The results of this study show that endocytosis is probably the main mechanism of entrance of DNA after GET in vitro and also in vivo. Copyright © 2014 Elsevier B.V. All rights reserved.

Preparation and In Vitro/Ex Vivo Evaluation of Moxifloxacin-Loaded PLGA Nanosuspensions for Ophthalmic Application

PubMed Central

Mudgil, Meetali; Pawar, Pravin K.

2013-01-01

The aim of the present investigation was to prepare a colloidal ophthalmic formulation to improve the residence time of moxifloxacin. Moxifloxacin-loaded poly(dl-lactide-co-glycolide) (PLGA) nanosuspensions were prepared by using the solvent evaporation technique. The nanosuspensions were characterised physically by using different techniques like particle size, zeta potential, FTIR, DSC, and XRD analysis. In vitro and ex vivo studies of nanosuspensions were carried out using a modified USP dissolution apparatus and all-glass Franz diffusion cells, respectively. The antibacterial activities of the nanosuspension and marketed formulations were performed against S. aureus and P. aeroginosa. The moxifloxacin-loaded PLGA nanosuspensions showed uniform particle size, ranging between 164–490 nm with negative zeta potential for all batches. The percentage entrapment efficiency of the drug-loaded nano-suspension was found to be between 84.09 to 92.05%. In vitro drug release studies suggest that all of the formulations showed extended drug release profiles and follow Korsemeyer-Peppas release kinetics. In vitro corneal permeability was found to be comparable with that of the marketed formulation across isolated goat cornea, indicating the suitability of the nanosuspension formulation in the ophthalmic delivery of moxifloxacin. The optimised nano-suspension was found to be more active against S. aureus and P. aeruginosa compared to the marketed eye drops. PMID:23833723

Preparation and in vitro characterization of thermosensitive and mucoadhesive hydrogels for nasal delivery of phenylephrine hydrochloride.

PubMed

Xu, Xiaofeng; Shen, Yan; Wang, Wei; Sun, Chunmeng; Li, Chang; Xiong, Yerong; Tu, Jiasheng

2014-11-01

The aim of the present work was to develop a nasal delivery system of phenylephrine hydrochloride (PE) in spray form to make prolonged remedy of nasal congestion. The formulations contain the thermosensitive hydrogel, i.e., Poloxamer 407 (P407) and Poloxamer 188 (P188) mixtures, and mucoadhesives, i.e., ε-polylysine (ε-PL) and low molecular weight sodium hyaluronate (MW 11,000Da). The in vitro characterizations of formulations including rheology studies, texture profiles and in vitro mucoadhesion potential were investigated after gelation temperatures measurements. The results showed that the concentration of P407 or P188 had significant influence on gelation temperature and texture profiles. The addition of mucoadhesives, though lowered the gel strength of formulations, increased interaction with mucin. After screening, two formulations (i.e., 1.0% PE/0.5% ε-PL/17% P407/0.5% P188 or Formulation A; and 1.0% PE/0.5% HA/17% P407/0.8% P188 or Formulation B) presenting suitable gelation temperatures (∼32°C) were used for further studies on in vitro release behaviors and mucosa ciliotoxicity. Both formulations showed sustained release of PE for up to 8h and similar toxicity to saline, the negative control. Thus, the thermosensitive and mucoadhesive PE-containing hydrogels are promising to achieve prolonged decongestion in nasal cavity. Copyright © 2014 Elsevier B.V. All rights reserved.

The Effect of Myricetin on Pharmacokinetics of Atomoxetine and its Metabolite 4-Hydroxyatomoxetine In Vivo and In Vitro.

PubMed

Lan, Tian; Hu, Xiao-Xia; Liang, Bing-Qing; Pan, Wen-He; Zhou, Quan; Yuan, Ling-Jing; Hu, Guo-Xin

2017-04-01

Atomoxetine is the first non-stimulant drug to be approved for the treatment of ADHD, while the effect of myricetin on the pharmacokinetic of atomoxetine in rats or human is still unknown. The present work was to study the impact of myricetin on the metabolism of atomoxetine both in vivo and in vitro. Twenty healthy male Sprague-Dawley rats were randomly divided into four groups: A (control group), B (100 mg/kg myricetin), C (50 mg/kg myricetin), and D (25 mg/kg myricetin). A single dose of atomoxetine (10 mg/kg) was administrated half an hour later. In addition, human and rat liver microsomes were performed to determine the effect of myricetin on the metabolism of atomoxetine in vitro. Group B, C, D all increased the C max and AUC of atomoxetine, but decreased the C max and AUC of 4-hydroxyatomoxetine. Moreover, myricetin showed inhibitory effect on human and rat microsomes, the IC 50 of myricetin was 8.651 and 35.45 µmol/L, respectively. Our study showed that myricetin could significantly inhibit the formation of atomoxetine metabolite both in vivo and in vitro. It is recommended that the effect of myricetin on the metabolism of atomoxetine should be noted and atomoxetine plasma concentration should be monitored.

Leptin accelerates the pathogenesis of heterotopic ossification in rat tendon tissues via mTORC1 signaling.

PubMed

Jiang, Huaji; Chen, Yuhui; Chen, Guorong; Tian, Xinggui; Tang, Jiajun; Luo, Lei; Huang, Minjun; Yan, Bin; Ao, Xiang; Zhou, Wen; Wang, Liping; Bai, Xiaochun; Zhang, Zhongmin; Wang, Liang; Xian, Cory J

2018-02-01

Leptin, an adipocyte-derived cytokine associated with bone metabolism, is believed to play a critical role in the pathogenesis of heterotopic ossification (HO). The effect and underlying action mechanism of leptin were investigated on osteogenic differentiation of tendon-derived stem cells (TDSCs) in vitro and the HO formation in rat tendons. Isolated rat TDSCs were treated with various concentrations of leptin in the presence or absence of mTORC1 signaling specific inhibitor rapamycin in vitro. A rat model with Achilles tenotomy was employed to evaluate the effect of leptin on HO formation together with or without rapamycin treatment. In vitro studies with TDSCs showed that leptin increased the expression of osteogenic biomarkers (alkaline phosphatase, runt-related transcription factor 2, osterix, osteocalcin) and enhanced mineralization of TDSCs via activating the mTORC1 signal pathway (as indicated by phosphorylation of p70 ribosomal S6 kinase 1 and p70 ribosomal S6). However, mTORC1 signaling blockade with rapamycin treatment suppressed leptin-induced osteogenic differentiation and mineralization. In vivo studies showed that leptin promoted HO formation in the Achilles tendon after tenotomy, and rapamycin treatment blocked leptin-induced HO formation. In conclusion, leptin can promote TDSC osteogenic differentiation and heterotopic bone formation via mTORC1 signaling in both vitro and vivo model, which provides a new potential therapeutic target for HO prevention. © 2017 Wiley Periodicals, Inc.

Association between in vitro platinum resistance in the EDR assay and clinical outcomes for ovarian cancer patients.

PubMed

Holloway, Robert W; Mehta, Rita S; Finkler, Neil J; Li, Kuo-Tung; McLaren, Christine E; Parker, Ricardo J; Fruehauf, John P

2002-10-01

The initial clinical response to platinum is a major determinant of outcome for patients with ovarian cancer. This retrospective study was undertaken to correlate the response and survival of newly diagnosed advanced ovarian cancer patients who received platinum-based therapy with in vitro drug response to cisplatin or carboplatin measured as percentage cell inhibition (PCI) in the in vitro Extreme Drug Resistance (EDR) assay. Outcomes for newly diagnosed ovarian cancer patients with tumor specimens submitted in a serial fashion for the EDR assay were studied. EDR assay results for cisplatin and carboplatin were correlated with clinical outcome for 79 evaluable chemotherapy nai;ve cases who presented with advanced (stages IIC, III, and IV) ovarian cancer. Stage IV and suboptimally debulked stage IIIc accounted for 16 cases, while 63 cases were optimally debulked Stage III/IIc. All patients were treated with platinum-based combination chemotherapy at a single institution. In vitro results for patient tumors were classified as low drug resistance (PCI > median), intermediate drug resistance [PCI between the median and 1 standard deviation (SD) below the median], or extreme drug resistance (PCI more than 1 SD below the median). For the purpose of this analysis, in vitro EDR to either cisplatin or carboplatin was considered to represent extreme resistance to platinum (EDRP), while the absence of EDR to either cisplatin or carboplatin was considered to represent low resistance to platinum (LDRP). Patients demonstrating relative in vitro resistance to paclitaxel and non-cross-resistance to cyclophosphamide and/or doxorubicin received cyclophosphamide plus platinum (CP); cyclophosphamide, doxorubicin, and platinum (CAP); or platinum alone in place of paclitaxel plus platinum (TP). Progression-free survival (PFS) and overall survival (OS) were correlated with EDR assay results. Median PFS was 6 months for the 17 cases exhibiting EDRP, compared to 24 months for the 62 cases exhibiting LDRP in vitro [relative risk (RR) 3.78, confidence intervals (CI) 1.82-7.83], adjusted for stage, debulking status, in vitro response to 3-OH-cyclophosphamide, and histological grade. Estimated overall 5-year survival was 19% for patients with tumors showing EDRP, compared to 68% for patients with tumors showing LDRP (RR 2.32, CI 1.06-5.07). Patients treated with CP (n = 20) showed no significant difference in OS compared to patients treated with TP (n = 54), CAP (n = 4), or cisplatin (n = 1) alone. In vitro platinum response remained an independent predictor of PFS and OS in multivariate analyses adjusted for CP versus TP, CAP, or platinum administration, and adjusted for debulking status. Median PFS for all 79 patients was 22 months, with an estimated 5-year survival of 57%. Patients with tumors demonstrating in vitro EDR to platinum were at significantly increased risk for progression and death when treated with standard platinum-based regimens. Such patients may therefore benefit from entry onto trials with novel agents or combinations.

Formulation and in vitro/in vivo evaluation of levodopa transdermal delivery systems.

PubMed

Lee, Kyung Eun; Choi, Yun Jung; Oh, Byu Ree; Chun, In Koo; Gwak, Hye Sun

2013-11-18

This study aims to investigate the feasibility of Levodopa transdermal delivery systems (TDSs). Levodopa TDSs were formulated using various vehicles and permeation enhancers, and in vitro permeation and in vivo pharmacokinetic studies were carried out. In the in vitro study, ester-type vehicles showed relatively high enhancing effects; propylene glycol monocaprylate and propylene glycol monolaurate showed the highest permeation fluxes from both solution and pressure sensitive adhesive (PSA) TDS formulations. Lag time was dramatically shortened with PSA TDS formulations as compared with solution formulations. In the in vivo study, the addition of fatty acids increased blood drug concentrations regardless of the kind or concentration of fatty acid; the AUCinf increased up to 8.7 times as compared with propylene glycol (PG) alone. PSA TDS containing 10% linoleic acid exhibited prolonged Tmax as compared with oral form. Total clearance of L-dopa from PSA TDSs was significantly lower than from oral form (up to 86.8 times). Especially, PSA TDS containing 10% linoleic acid (LOA) revealed 76.2 fold higher AUCinf than oral administration. Based on our results, the L-dopa PSA TDS containing PG with 10% LOA could be used as a good adjuvant therapy for Parkinson's disease patients who experience symptom fluctuation by L-dopa oral administration. Copyright © 2013 Elsevier B.V. All rights reserved.

Polydimethylsiloxane pressure sensors for force analysis in tension band wiring of the olecranon.

PubMed

Zens, Martin; Goldschmidtboeing, Frank; Wagner, Ferdinand; Reising, Kilian; Südkamp, Norbert P; Woias, Peter

2016-11-14

Several different surgical techniques are used in the treatment of olecranon fractures. Tension band wiring is one of the most preferred options by surgeons worldwide. The concept of this technique is to transform a tensile force into a compression force that adjoins two surfaces of a fractured bone. Currently, little is known about the resulting compression force within a fracture. Sensor devices are needed that directly transduce the compression force into a measurement quality. This allows the comparison of different surgical techniques. Ideally the sensor devices ought to be placed in the gap between the fractured segments. The design, development and characterization of miniaturized pressure sensors fabricated entirely from polydimethylsiloxane (PDMS) for a placement within a fracture is presented. The pressure sensors presented in this work are tested, calibrated and used in an experimental in vitro study. The pressure sensors are highly sensitive with an accuracy of approximately 3 kPa. A flexible fabrication process for various possible applications is described. The first in vitro study shows that using a single-twist or double-twist technique in tension band wiring of the olecranon has no significant effect on the resulting compression forces. The in vitro study shows the feasibility of the proposed measurement technique and the results of a first exemplary study.

Cinnamon: Potential Role in the Prevention of Insulin Resistance, Metabolic Syndrome, and Type 2 Diabetes

PubMed Central

Qin, Bolin; Panickar, Kiran S.; Anderson, Richard A.

2010-01-01

Metabolic syndrome is associated with insulin resistance, elevated glucose and lipids, inflammation, decreased antioxidant activity, increased weight gain, and increased glycation of proteins. Cinnamon has been shown to improve all of these variables in in vitro, animal, and/or human studies. In addition, cinnamon has been shown to alleviate factors associated with Alzheimer's disease by blocking and reversing tau formation in vitro and in ischemic stroke by blocking cell swelling. In vitro studies also show that components of cinnamon control angiogenesis associated with the proliferation of cancer cells. Human studies involving control subjects and subjects with metabolic syndrome, type 2 diabetes mellitus, and polycystic ovary syndrome all show beneficial effects of whole cinnamon and/or aqueous extracts of cinnamon on glucose, insulin, insulin sensitivity, lipids, antioxidant status, blood pressure, lean body mass, and gastric emptying. However, not all studies have shown positive effects of cinnamon, and type and amount of cinnamon, as well as the type of subjects and drugs subjects are taking, are likely to affect the response to cinnamon. In summary, components of cinnamon may be important in the alleviation and prevention of the signs and symptoms of metabolic syndrome, type 2 diabetes, and cardiovascular and related diseases. PMID:20513336

Potential in vitro antioxidant and protective effects of Gymnema montanum H. on alloxan-induced oxidative damage in pancreatic beta-cells, HIT-T15.

PubMed

Ramkumar, Kunga Mohan; Manjula, Chinnasamy; Sankar, Lakshmanan; Suriyanarayanan, Sarvajayakesavalu; Rajaguru, Palanisamy

2009-09-01

The present study describes the antioxidant activities of ethanol extract from Gymnema montanum (GLEt) which is an endemic plant of India. Antioxidant activity of the GLEt was studied in vitro based on scavenging of hydroxyl radicals, superoxide anions, nitric oxide, hydrogen peroxide, peroxynitrite, reducing power and inhibition of lipid peroxidation estimated in terms of thiobarbituric acid reactive substances (TBARS). Further, we examined its protective effect against alloxan-induced oxidative stress in pancreatic beta-cells, HIT-T15 by measuring the free radical generation, malonaldehyde formation and antioxidant levels such as CAT, GPx and GSH. Results showed that G. montanum leaves exhibited significant antioxidant activities measured by various in vitro model systems. The HIT-T15 cell line studies showed the tendency of GLEt to increase antioxidant levels meanwhile decrease the free radical formation and inhibit the lipid peroxidation. The antioxidant activity was found to be well correlated with the phenolic phytochemicals present in the extract. GC-MS analyses revealed the presence of few phenolic compounds in the extract. As this plant has already been demonstrated for a variety of medicinal properties from our laboratory, results of this study suggest that G. montanum is an interesting source for antioxidant compounds and useful for various therapeutic applications.

In vitro comparison of antimicrobial activity of aqueous decoction of Coriandrum sativum, and Dentol Drop with chlorhexidine on Streptococcus mutans.

PubMed

Moradian, Hamid; Bazargani, Abdollah; Rafiee, Azade; Nazarialam, Ali

2013-09-01

Dental caries is still remained as a major health problem. This problem has created a new interest to search for new antimicrobial agents from various sources including medicinal plants. Since limited data is available so far regarding the antibacterial effect of Coriandrum sativum seed and Dentol Drop against Streptococcus mutans, this study aims to assess this activity. This experimental study was conducted in Shiraz University of Medical Sciences. In vitro comparison of antimicrobial activity of aqueous decoction of Coriandrum sativum seed and Dentol drop with chlorhexidine against Streptococcus mutans was evaluated using disk diffusion and broth microdilution assays. Positive and negative controls were considered. The data was statistically analyzed by applying Kruskal-Wallis and Tukey post-hoc test to compare the groups using SPSS software (version 17). Dentol drop showed a remarkable antibacterial activity, in comparison with chlorhexidine, against S. mutans in the disk diffusion (p value = 0.005), and broth microdilution assays (p value = 0.0001). Based on the results of this study, Coriandrum sativum seed did not posses any antibacterial property. Coriandrum sativum seed showed no anti-Streptococcus mutans activity. Dentol drop exhibited a remarkable antibacterial activity against S. mutans when tested in vitro. Dentol drop can be further studied as a preventive measure for dental caries.

In vitro and in vivo antimicrobial activities of seeds of Caesalpinia bonduc (Lin.) Roxb.

PubMed

Arif, Tasleem; Mandal, T K; Kumar, Naresh; Bhosale, J D; Hole, Archana; Sharma, G L; Padhi, M M; Lavekar, G S; Dabur, Rajesh

2009-05-04

Caesalpinia bonduc (Lin.) Roxb. is a known drug in Ayurveda to treat various diseases specifically tumors, cysts and cystic fibrosis (CF). The aim of this study was to assess in vitro as well as in vivo antimicrobial activity of Caesalpinia bonduc seeds. The in vitro antimicrobial activities of seed coat and seed kernel extracts were investigated by microbroth dilution assay. In vivo activities of hydro-alcoholic extracts were investigated in rat models of chronic Pseudomonas aeruginosa pneumonia mimicking that in patients with cystic fibrosis. Various extracts of plant seeds exhibited in vitro antimicrobial activities in a range of 22-350 microg/ml. The extracts also showed activity against methicillin resistant (MR) Staphylococcus aureus and ampicillin resistant (AR) Pseudomonas aeruginosa as in the sensitive strains. In rat model of chronic Pseudomonas aeruginosa pneumonia, hydro-alcoholic extracts of Caesalpinia bonduc seed kernel (CBSK) and Caesalpinia bonduc seed coat (CBSC) were injected subcutaneously in the test groups of animals. The control groups were treated with cortisone and saline. Two weeks after challenge with Pseudomonas aeruginosa, the CBSK treated animals showed a significant bacterial clearance from the lungs (P<0.04) and less severe incidence of lung abscess (P<0.05). Results showed that Caesalpinia bonduc may have the potential to be promising natural medicine, with other forms of treatments, for CF patients with chronic Pseudomonas aeruginosa lung infections.

Study on antibacterial effect of medlar and hawthorn compound extract in vitro.

PubMed

Niu, Yang; Nan, Yi; Yuan, Ling; Wang, Rong

2013-01-01

This paper evaluated the antibacterial effect of medlar and hawthorn compound extract in vitro. Water extract method and ethanol extraction method was adopted to prepare the compound extracts, and disc diffusion method and improved test tube doubling dilution method were used to conduct the antibacterial test on the two common pathogenic bacteria, Staphylococcus aureus and Klebsiella pneumonia, in vitro. The results showed that medlar and hawthorn compound extract was moderately sensitive to Staphylococcus aureus, while its inhibiting effect on Klebsiella pneumoniae was particularly significant, moreover, the antibacterial effect of ethanol extract was better than water extract. Medlar and hawthorn compounds had good antibacterial effect on the two pathogenic bacteria.

Evaluation of in-vitro antibacterial activity and anti-inflammatory activity for different extracts of Rauvolfia tetraphylla L. root bark.

PubMed

Ganga Rao, B; Umamaheswara Rao, P; Sambasiva Rao, E; Mallikarjuna Rao, T; Praneeth D, V S

2012-10-01

To assess the in-vitro antibacterial activity and anti-inflammatory activity of orally administered different extracts (Hydro-alcoholic, methanolic, ethyl acetate and hexane) of Rauvolfia tetraphylla (R. tetraphylla) root bark in Carrageenan induced acute inflammation in rats. In-vitro antibacterial activity was evaluated for extracts against four Gram positive and four Gram negative bacteria by using cylinder plate assay. Hydro-alcoholic extract (70% v/v ethanol) at 200, 400 and 800 mg/kg doses and methanolic, ethyl acetate and hexane extracts at doses 100, 200 and 400 mg/kg were tested for anti-inflammatory activity in Carrageenan induced rat paw oedema model and paw thickness was measured every one hour up to 6 hrs. All extracts of R. tetraphylla root bark showed good zone of inhibition against tested bacterial strains. In Carrageenan induced inflammation model, hydro-alcoholic and methanolic extract of R. tetraphylla root bark at three different doses produced significant (P<0.001) reduction when compared to vehicle treated control group and hexane, ethyl acetate extracts. In the present study extracts of R. tetraphylla root bark shows good in-vitro antibacterial activity and in-vivo anti-inflammatory activity in rats.

Specialization of the DNA-Cleaving Activity of a Group I Ribozyme Through In Vitro Evolution

NASA Technical Reports Server (NTRS)

Tsang, Joyce; Joyce, Gerald F.

1996-01-01

In an earlier study, an in vitro evolution procedure was applied to a large population of variants of the Tetrahymena group 1 ribozyme to obtain individuals with a 10(exp 5)-fold improved ability to cleave a target single-stranded DNA substrate under simulated physiological conditions. The evolved ribozymes also showed a twofold improvement, compared to the wild-type, in their ability to cleave a single-stranded RNA substrate. Here, we report continuation of the in vitro evolution process using a new selection strategy to achieve both enhanced DNA and diminished RNA-cleavage activity. Our strategy combines a positive selection for DNA cleavage with a negative selection against RNA binding. After 36 "generations" of in vitro evolution, the evolved population showed an approx. 100-fold increase in the ratio of DNA to RNA-cleavage activity. Site-directed mutagenesis experiment confirmed the selective advantage of two covarying mutations within the catalytic core of ribozyme that are largely responsible for this modified behavior. The population of ribozymes has now undergone a total of 63 successive generations of evolution, resulting in an average 28 mutations relative to the wild-type that are responsible for the altered phenotype.

Potential uses of terahertz pulse imaging in dentistry: caries and erosion detection

NASA Astrophysics Data System (ADS)

Longbottom, Christopher; Crawley, David A.; Cole, Bryan E.; Arnone, Donald D.; Wallace, Vincent P.; Pepper, Michael

2002-06-01

TeraHertz Pulse Imaging (TPI) is a relatively new imaging modality for medical and dental imaging. The aim of the present study was to make a preliminary assessment of the potential uses of TPI in clinical dentistry, particularly in relation to caries detection and the detection and monitoring of erosion. Images were obtained in vitro using a new TPI system developed by TeraView Ltd. We present data showing that TPI in vitro images of approximal surfaces of whole teeth demonstrate a distinctive shadowing in the presence of natural carious lesions in enamel. The thickness of this enamel shadowing appears to be related to lesion depth. The use of non-ionizing radiation to image such lesions non-destructively in vitro represents a significant step towards such measurements in vivo. In addition, data is presented which indicates that TPI may have a potential role in the detection and monitoring of enamel erosion. In vitro experiments on whole incisor teeth show that TPI is capable of detecting relatively small artificially induced changes in the buccal or palatal surface of the enamel of these teeth. Imaging of enamel thickness at such a resolution without ionizing radiation would represent a significant breakthrough if applicable in vivo.

Effect of the lectins wheat germ agglutinin (WGA) and Ulex europaeus agglutinin (UEA-I) on the alpha-amylase secretion of rat pancreas in vitro and in vivo.

PubMed

Mikkat, U; Damm, I; Schröder, G; Schmidt, K; Wirth, C; Weber, H; Jonas, L

1998-05-01

Lectins are able to bind to cholecystokinin (CCK) receptors and other glycosylated membrane proteins. The lectins wheat germ agglutinin (WGA) and Ulex europaeus agglutinin (UEA-I) are used for affinity chromatography to isolate the highly glycosylated CCK-A receptor of pancreatic acinar cells. According to the working hypothesis that lectin binding to the CCK receptor should alter the ligand-receptor interaction, the effect of WGA and UEA-I on CCK-8-induced enzyme secretion was studied on isolated rat pancreatic acini in vitro. In vitro both lectins showed a dosage-dependent inhibition of CCK-8-induced alpha-amylase secretion of acini over 60 min. WGA showed a strong inhibitory effect on amylase secretion, approximately 40%, in vitro. UEA-I caused a smaller, but significant decrease, approximately 20%, in enzyme secretion of isolated acini. Additionally, both lectins inhibited cerulein/secretin- or cerulein-induced pancreatic secretion of rats in vivo, but not after secretin alone. The results are discussed with respect to a possible influence of both lectins on the interaction of CCK or cerulein with the CCK-A receptor.

Intracellular Doppler Signatures of Platinum Sensitivity Captured by Biodynamic Profiling in Ovarian Xenografts

NASA Astrophysics Data System (ADS)

Merrill, Daniel; An, Ran; Sun, Hao; Yakubov, Bakhtiyor; Matei, Daniela; Turek, John; Nolte, David

2016-01-01

Three-dimensional (3D) tissue cultures are replacing conventional two-dimensional (2D) cultures for applications in cancer drug development. However, direct comparisons of in vitro 3D models relative to in vivo models derived from the same cell lines have not been reported because of the lack of sensitive optical probes that can extract high-content information from deep inside living tissue. Here we report the use of biodynamic imaging (BDI) to measure response to platinum in 3D living tissue. BDI combines low-coherence digital holography with intracellular Doppler spectroscopy to study tumor drug response. Human ovarian cancer cell lines were grown either in vitro as 3D multicellular monoculture spheroids or as xenografts in nude mice. Fragments of xenografts grown in vivo in nude mice from a platinum-sensitive human ovarian cell line showed rapid and dramatic signatures of induced cell death when exposed to platinum ex vivo, while the corresponding 3D multicellular spheroids grown in vitro showed negligible response. The differences in drug response between in vivo and in vitro growth have important implications for predicting chemotherapeutic response using tumor biopsies from patients or patient-derived xenografts.

In vitro and in vivo comparison of a tailored magnetic particle imaging blood pool tracer with Resovist.

PubMed

Kaul, Michael Gerhard; Mummert, Tobias; Jung, Caroline; Salamon, Johannes; Khandhar, Amit P; Ferguson, R Matthew; Kemp, Scott J; Ittrich, Harald; Krishnan, Kannan M; Adam, Gerhard; Knopp, Tobias

2017-05-07

Optimizing tracers for individual imaging techniques is an active field of research. The purpose of this study was to perform in vitro and in vivo magnetic particle imaging (MPI) measurements using a new monodisperse and size-optimized tracer, LS-008, and to compare it with the performance of Resovist, the standard MPI tracer. Magnetic particle spectroscopy (MPS) and in vitro MPI measurements were performed in concerns of concentration and amount of tracer in a phantom. In vivo studies were carried out in healthy FVB mice. The first group (n  =  3) received 60 µl LS-008 (87 mM) and the second (n  =  3) diluted Resovist of the same concentration and volume. Tracer injections were performed with a syringe pump during a dynamic MPI scan. For anatomic referencing MRI was applied beforehand of the MPI measurements. Summing up MPS examinations and in vitro MPI experiments, LS-008 showed better sensitivity and spatial resolution than Resovist. In vivo both tracers can visualize the propagation of the bolus through the inferior vena cava. MPI with LS-008 did show less temporal fluctuation artifacts and the pulsation of blood due to respiratory and cardiac cycle was detectable. With LS-008 the aorta was distinguishable from the caval vein while with Resovist this failed. A liver vessel and a vessel structure leading cranially could only be observed with LS-008 and not with Resovist. Beside these structural advantages both tracers showed very different blood half-life. For LS-008 we found 88 min. Resovist did show a fast liver accumulation and a half-life of 13 min. Only with LS-008 the perfusion fraction in liver and kidney was measureable. MPI for angiography can be significantly improved by applying more effective tracers. LS-008 shows a clear improvement concerning the delineation while resolving a larger number of vessels in comparison to Resovist. Therefore, in aspects of quality and quantity LS-008 is clearly favorable for angiographic and perfusion studies.

In vitro and in vivo comparison of a tailored magnetic particle imaging blood pool tracer with Resovist

NASA Astrophysics Data System (ADS)

Kaul, Michael Gerhard; Mummert, Tobias; Jung, Caroline; Salamon, Johannes; Khandhar, Amit P.; Ferguson, R. Matthew; Kemp, Scott J.; Ittrich, Harald; Krishnan, Kannan M.; Adam, Gerhard; Knopp, Tobias

2017-05-01

Optimizing tracers for individual imaging techniques is an active field of research. The purpose of this study was to perform in vitro and in vivo magnetic particle imaging (MPI) measurements using a new monodisperse and size-optimized tracer, LS-008, and to compare it with the performance of Resovist, the standard MPI tracer. Magnetic particle spectroscopy (MPS) and in vitro MPI measurements were performed in concerns of concentration and amount of tracer in a phantom. In vivo studies were carried out in healthy FVB mice. The first group (n  =  3) received 60 µl LS-008 (87 mM) and the second (n  =  3) diluted Resovist of the same concentration and volume. Tracer injections were performed with a syringe pump during a dynamic MPI scan. For anatomic referencing MRI was applied beforehand of the MPI measurements. Summing up MPS examinations and in vitro MPI experiments, LS-008 showed better sensitivity and spatial resolution than Resovist. In vivo both tracers can visualize the propagation of the bolus through the inferior vena cava. MPI with LS-008 did show less temporal fluctuation artifacts and the pulsation of blood due to respiratory and cardiac cycle was detectable. With LS-008 the aorta was distinguishable from the caval vein while with Resovist this failed. A liver vessel and a vessel structure leading cranially could only be observed with LS-008 and not with Resovist. Beside these structural advantages both tracers showed very different blood half-life. For LS-008 we found 88 min. Resovist did show a fast liver accumulation and a half-life of 13 min. Only with LS-008 the perfusion fraction in liver and kidney was measureable. MPI for angiography can be significantly improved by applying more effective tracers. LS-008 shows a clear improvement concerning the delineation while resolving a larger number of vessels in comparison to Resovist. Therefore, in aspects of quality and quantity LS-008 is clearly favorable for angiographic and perfusion studies.

Vitamin K2 Ameliorates Damage of Blood Vessels by Glucocorticoid: a Potential Mechanism for Its Protective Effects in Glucocorticoid-induced Osteonecrosis of the Femoral Head in a Rat Model

PubMed Central

Zhang, Yuelei; Yin, Junhui; Ding, Hao; Zhang, Changqing; Gao, You-Shui

2016-01-01

Glucocorticoid has been reported to decrease blood vessel number and harm the blood supply in the femoral head, which is recognized to be an important mechanism of glucocorticoid-induced osteonecrosis of the femoral head (ONFH). To prevent glucocorticoid-induced ONFH, medication that promotes both bone formation and angiogenesis would be ideal. Vitamin K2 has been revealed to play an important role in bone metabolism; however, few studies have focused on the effect of Vitamin K2 on new vascular formation. Thus, this study aimed to investigate whether Vitamin K2 promoted new blood vessel formation in the presence of glucocorticoids, both in vitro and in vivo. The effect of Vitamin K2 on viability, migration, in vitro tube formation, and VEGF, vWF, CD31, KDR, Flt and PDGFB in EAhy926 incubated with or without dexamethasone were elucidated. VEGF, TGF-β and BMP-2, angiogenesis-related proteins secreted by osteoblasts, were also detected in the osteoblast-like cell line of MG63. In addition, blood vessels of the femoral head in rats administered with or without methylprednisolone and Vitamin K2 were evaluated using angiography and CD31 staining. In vitro studies showed that Vitamin K2 significantly protected endothelial cells from dexamethasone-induced apoptosis, promoted endothelial cell migration and in vitro tube formation. Angiogenesis-related proteins both in EAhy926 and MG63 were also upregulated by Vitamin K2 when cotreated with dexamethasone. In vivo studies showed enhanced blood vessel volume and CD31-positive staining cells in rats cotreated with VK2 and methylprednisolone compared to rats treated with methylprednisolone only. Collectively, Vitamin K2 has the ability to promote angiogenesis in vitro and to ameliorate vessels of the femoral head in glucocorticoid-treated rats in vivo, indicating that Vitamin K2 is a promising drug that may be used to prevent steroid-induced ONFH. PMID:27313492

Vitamin K2 Ameliorates Damage of Blood Vessels by Glucocorticoid: a Potential Mechanism for Its Protective Effects in Glucocorticoid-induced Osteonecrosis of the Femoral Head in a Rat Model.

PubMed

Zhang, Yuelei; Yin, Junhui; Ding, Hao; Zhang, Changqing; Gao, You-Shui

2016-01-01

Glucocorticoid has been reported to decrease blood vessel number and harm the blood supply in the femoral head, which is recognized to be an important mechanism of glucocorticoid-induced osteonecrosis of the femoral head (ONFH). To prevent glucocorticoid-induced ONFH, medication that promotes both bone formation and angiogenesis would be ideal. Vitamin K2 has been revealed to play an important role in bone metabolism; however, few studies have focused on the effect of Vitamin K2 on new vascular formation. Thus, this study aimed to investigate whether Vitamin K2 promoted new blood vessel formation in the presence of glucocorticoids, both in vitro and in vivo. The effect of Vitamin K2 on viability, migration, in vitro tube formation, and VEGF, vWF, CD31, KDR, Flt and PDGFB in EAhy926 incubated with or without dexamethasone were elucidated. VEGF, TGF-β and BMP-2, angiogenesis-related proteins secreted by osteoblasts, were also detected in the osteoblast-like cell line of MG63. In addition, blood vessels of the femoral head in rats administered with or without methylprednisolone and Vitamin K2 were evaluated using angiography and CD31 staining. In vitro studies showed that Vitamin K2 significantly protected endothelial cells from dexamethasone-induced apoptosis, promoted endothelial cell migration and in vitro tube formation. Angiogenesis-related proteins both in EAhy926 and MG63 were also upregulated by Vitamin K2 when cotreated with dexamethasone. In vivo studies showed enhanced blood vessel volume and CD31-positive staining cells in rats cotreated with VK2 and methylprednisolone compared to rats treated with methylprednisolone only. Collectively, Vitamin K2 has the ability to promote angiogenesis in vitro and to ameliorate vessels of the femoral head in glucocorticoid-treated rats in vivo, indicating that Vitamin K2 is a promising drug that may be used to prevent steroid-induced ONFH.

Development of a Test Method for the Evaluation of DNA Damage in Mouse Spermatogonial Stem Cells

PubMed Central

Jeon, Hye Lyun; Yi, Jung-Sun; Kim, Tae Sung; Oh, Youkyung; Lee, Hye Jeong; Lee, Minseong; Bang, Jin Seok; Ko, Kinarm; Ahn, Il Young; Ko, Kyungyuk; Kim, Joohwan; Park, Hye-Kyung; Lee, Jong Kwon; Sohn, Soo Jung

2017-01-01

Although alternative test methods based on the 3Rs (Replacement, Reduction, Refinement) are being developed to replace animal testing in reproductive and developmental toxicology, they are still in an early stage. Consequently, we aimed to develop alternative test methods in male animals using mouse spermatogonial stem cells (mSSCs). Here, we modified the OECD TG 489 and optimized the in vitro comet assay in our previous study. This study aimed to verify the validity of in vitro tests involving mSSCs by comparing their results with those of in vivo tests using C57BL/6 mice by gavage. We selected hydroxyurea (HU), which is known to chemically induce male reproductive toxicity. The 50% inhibitory concentration (IC50) value of HU was 0.9 mM, as determined by the MTT assay. In the in vitro comet assay, % tail DNA and Olive tail moment (OTM) after HU administration increased significantly, compared to the control. Annexin V, PI staining and TUNEL assays showed that HU caused apoptosis in mSSCs. In order to compare in vitro tests with in vivo tests, the same substances were administered to male C57BL/6 mice. Reproductive toxicity was observed at 25, 50, 100, and 200 mg/kg/day as measured by clinical measures of reduction in sperm motility and testicular weight. The comet assay, DCFH-DA assay, H&E staining, and TUNEL assay were also performed. The results of the test with C57BL/6 mice were similar to those with mSSCs for HU treatment. Finally, linear regression analysis showed a strong positive correlation between results of in vitro tests and those of in vivo. In conclusion, the present study is the first to demonstrate the effect of HU-induced DNA damage, ROS formation, and apoptosis in mSSCs. Further, the results of the current study suggest that mSSCs could be a useful model to predict male reproductive toxicity. PMID:28443181

A hybrid design to optimize preparation of lopinavir loaded solid lipid nanoparticles and comparative pharmacokinetic evaluation with marketed lopinavir/ritonavir coformulation.

PubMed

Ravi, Punna Rao; Vats, Rahul; Dalal, Vikas; Murthy, Aditya Narasimha

2014-07-01

To prepare stearic acid-based lopinavir (LPV) loaded solid lipid nanoparticles (SLNs) using a hybrid design and compare in-vivo performance of optimized formulation with marketed LPV/ritonavir (RTV) coformulation. LPV SLNs were prepared by hot melt emulsion technique and optimized using Plackett-Burman design and Box-Behnken design. Physical characterization studies were conducted for the optimized SLNs. Comparative oral pharmacokinetic studies and tissue distribution studies of optimized SLNs and LPV/RTV coformulation were done in Wistar rats. In-vitro metabolic stability and intestinal permeability studies for LPV SLNs were undertaken to elucidate the mechanism involved in the pharmacokinetic improvement of LPV. Optimized SLNs exhibited nanometeric size (223 nm) with high entrapment efficiency (83%). In-vitro drug release study of SLNs showed biphasic sustained release behaviour. Significant increase in oral bioavailability of LPV from LPV SLNs (5 folds) and LPV/RTV coformulation (3.7 folds) was observed as compared with free LPV. LPV SLNs showed better tissue distribution of LPV in HIV reservoirs than LPV/RTV coformulation. In-vitro studies demonstrated that SLNs provided metabolic protection of LPV and were endocytosized during absorption. SLNs enhanced oral bioavailability and improved distribution profile of LPV to HIV reservoirs and hence could be better alternative to LPV/RTV coformulation. © 2014 Royal Pharmaceutical Society.

An in vitro study of cryopreserved and fresh human arteries: a comparison with ePTFE prostheses and human arteries studied non-invasively in vivo.

PubMed

Armentano, Ricardo L; Santana, Daniel Bia; Cabrera Fischer, Edmundo I; Graf, Sebastián; Cámpos, Héctor Pérez; Germán, Yanina Zócalo; Carmen Saldías, Maria Del; Alvarez, Inés

2006-02-01

The surgical options in arterial reconstruction are: the use of autologous arteries; autologous veins; or expanded polytetrafluoroethylene (ePTFE) grafts. However, the development of intimal hyperplasia when using veins or ePTFE grafts has been associated with graft failure. Since autologous arteries are not always available, the use of cryopreserved arteries has to be considered. The aims of this study were: (a) to compare the viscoelastic properties of stored cryopreserved arteries and fresh arteries by in vitro analysis; and (b) to compare the viscoelastic properties of arteries measured non-invasively in normotensive patients, with fresh arteries, cryopreserved arteries, and ePTFE segments. The viscoelastic studies were performed in normotensive patients using stress-strain analysis with non-invasive measurement of pressure and diameter in the common carotid artery, and in vitro measurements of pressure and diameter in arteries and prostheses. The in vitro studies showed that the elastic modulus (E), viscous modulus (eta), Stiffness Index (SI), Peterson modulus (Ep), and the pulse wave velocity (PWV) values for human cryopreserved carotid arteries were similar to the values obtained non-invasively in normotensive subjects (P>0.05) and to human fresh vessels (P>0.05). In vitro, the SI, Ep, PWV, and E values of ePTFE were significantly higher than the observed values in subjects and with fresh and cryopreserved arteries (P<0.05); on the other hand the ePTFE eta values were the lowest (P<0.05). We concluded that cryopreserved arteries have similar viscoelastic properties to those obtained in vivo in the arteries of normotensive subjects and in vitro in fresh arteries. Consequently, we conclude that the cryopreservation procedure does not modify the mechanical properties of the arterial wall.

The effect of orthodontic bonding materials on dental plaque accumulation and composition in vitro.

PubMed

Badawi, H; Evans, R D; Wilson, M; Ready, D; Noar, J H; Pratten, J

2003-08-01

The aim of this study was to investigate the accumulation and composition of microcosm dental plaque on different orthodontic bonding materials using an in vitro model. Microcosm plaques were grown on discs of a range of bonding materials in a constant depth film fermentor. The biofilms were derived from human saliva and supplied with artificial saliva as a source of nutrients. The number of viable bacteria in the biofilms was determined and the streptococci present were identified to species level. The results showed that there was no significant difference in bacterial accumulation between different bonding materials, however, biofilms grown on materials which were fluoride releasing, did not contain Streptococcus mutans. This in vitro study has shown that the use of fluoride-releasing bonding materials may support the growth of supragingival plaque, which does not contain S. mutans.

Comparative study of the corrosion behavior of peripheral stents in an accelerated corrosion model: experimental in vitro study of 28 metallic vascular endoprostheses

PubMed Central

Paprottka, Karolin J.; Paprottka, Philipp M.; Reiser, Maximilian F.; Waggershauser, Tobias

2015-01-01

PURPOSE Clinical cases of stent-fractures show that corrosion behavior might play a role in these fractures. Implanted in vivo, especially in combination with other implanted foreign materials, these metallic products are exposed to special conditions, which can cause a process of corrosion. Here, we aimed to test the corrosion potential of stents made of different materials in an in vitro setting. METHODS A total of 28 peripheral stents of different materials (nitinol, cobalt-chromium-nickel, tantalum, V4A) and surface treatments (electropolish, mechanical polish, no polish) were tested in vitro. Corrosion was accelerated by applying a constant voltage of 3.5 V and amperage of 1.16 mA in 0.9% NaCl. RESULTS Nitinol stents showed the lowest susceptibility to corrosion and the longest period without damage. The Memotherm II® (BARD Angiomed®) was the only stent that showed neither macroscopic nor microscopic damages. The worst performing material was cobalt-chromium-nickel, which showed corrosion damages about ten times earlier compared to nitinol. Considering the reasons for termination of the test, nitinol stents primarily showed length deficits, while V4A and tantalum stents showed fractures. Cobalt-chromium-nickel stents had multiple fractures or a complete lysis in equal proportions. When placed in direct contact, nitinol stents showed best corrosion resistance, regardless of what material they were combined with. In terms of polishing treatments, electropolished stents performed the best, mechanical-polished stents and those without polishing treatment followed. CONCLUSION The analysis of corrosion behavior may be useful to select the right stent fulfilling the individual needs of the patient within a large number of different stents. PMID:26268301

Influence of pH on in vitro disintegration of phosphate binders.

PubMed

Stamatakis, M K; Alderman, J M; Meyer-Stout, P J

1998-11-01

Hyperphosphatemia, a common complication in patients with end-stage renal disease, is treated with oral phosphate-binding medications that restrict phosphorus absorption from the gastrointestinal (GI) tract. Impaired product performance, such as failure to disintegrate and/or dissolve in the GI tract, could limit the efficacy of the phosphate binder. Disintegration may be as important as dissolution for predicting in vitro product performance for medications that act locally on the GI tract, such as phosphate binders. Furthermore, patients with end-stage renal disease have a wide range in GI pH, and pH can influence a product's performance. The purpose of this study was to determine the effect of pH on in vitro disintegration of phosphate binders. Fifteen different commercially available phosphate binders (seven calcium carbonate tablet formulations, two calcium acetate tablet formulations, three aluminum hydroxide capsule formulations, and three aluminum hydroxide tablet formulations) were studied using the United States Pharmacopeia (USP) standard disintegration apparatus. Phosphate binders were tested in simulated gastric fluid (pH 1.5), distilled water (pH 5.1), and simulated intestinal fluid (pH 7.5). Product failure was defined as two or more individual tablets or capsules failing to disintegrate completely within 30 minutes. Results indicate that 9 of the 15 phosphate binders tested showed statistically significant differences in disintegration time (DT) based on pH. The percentage of binders that passed the disintegration study test in distilled water, gastric fluid, and intestinal fluid were 80%, 80%, and 73%, respectively. The findings of this study show that the disintegration of commercially available phosphate binders is highly variable. The pH significantly affected in vitro disintegration in the majority of phosphate binders tested; how significantly this affects in vivo performance has yet to be studied.

Anticancer Effects of Extracts from the Fruit of Morinda Citrifolia (Noni) in Breast Cancer Cell Lines.

PubMed

Sharma, K; Pachauri, S D; Khandelwal, K; Ahmad, H; Arya, A; Biala, P; Agrawal, S; Pandey, R R; Srivastava, A; Srivastav, A; Saxena, J K; Dwivedi, A K

2016-03-01

Morinda citrifolia L. (NONI) fruits have been used for thousands of years for the treatment of many health problems including cancer, cold, diabetes, flu, hypertension, and pain. Plant extracts have reported several therapeutic benefits, but extraction of individual compound from the extract often exhibits limited clinical utility as the synergistic effect of various natural ingredients gets lost. They generally constitute polyphenols and flavonoids. Studies have suggested that these phytochemicals, especially polyphenols, display high antioxidant properties, which help to reduce the risk of degenerative diseases, such as cancer and cardiovascular diseases. Several in-vitro and in-vivo studies have shown that Noni fruits have antioxidant, anti-inflammatory, anti-dementia, liver-protective, anticancer, analgesic, and immunomodulatory effects. Till date about 7 in vitro cancer studies have been done, but a detailed in vitro study including cell cycle and caspase activation assay on breast cancer cell line has not been done. In the present study different Noni fruit fractions have tested on cancer cell lines MCF-7, MDA-MB-231 (breast adenocarcinoma) and one non-cancer cell line HEK-293 (Human embryonic kidney). Out of which ethylacetate extract showed a higher order of in vitro anticancer activity profile. The ethylacetate extract strongly inhibited the proliferation of MCF-7, MDA-MB-231 and HEK-293 cell lines with IC50 values of 25, 35, 60 µg/ml respectively. The extract showed increase in apoptotic cells in MCF-7 and MDA-MB-231 cells and arrested the cell cycle in the G1/S phase in MCF-7 and G0/G1 phase in MDA-MB-231 cells. Noni extract also decreases the intracellular ROS generation and mitochondrial membrane potential. © Georg Thieme Verlag KG Stuttgart · New York.

Preclinical and clinical in vitro in vivo correlation of an hGH dextran microsphere formulation.

PubMed

Vlugt-Wensink, K D F; de Vrueh, R; Gresnigt, M G; Hoogerbrugge, C M; van Buul-Offers, S C; de Leede, L G J; Sterkman, L G W; Crommelin, D J A; Hennink, W E; Verrijk, R

2007-12-01

To investigate the in vitro in vivo correlation of a sustained release formulation for human growth hormone (hGH) based on hydroxyethyl methacrylated dextran (dex-HEMA) microspheres in Pit-1 deficient Snell dwarf mice and in healthy human volunteers. A hGH-loaded microsphere formulation was developed and tested in Snell dwarf mice (pharmacodynamic study) and in healthy human volunteers (pharmacokinetic study). Single subcutaneous administration of the microspheres in mice resulted in a good correlation between hGH released in vitro and in vivo effects for the hGH-loaded microsphere formulation similar to daily injected hGH indicating a retained bioactivity. Testing the microspheres in healthy volunteers showed an increase (over 7-8 days) in hGH serum concentrations (peak concentrations: 1-2.5 ng/ml). A good in vitro in vivo correlation was obtained between the measured and calculated (from in vitro release data) hGH serum concentrations. Moreover, an increased serum concentration of biomarkers (insulin-like growth factor-I (IGF-I), IGF binding protein-3 (IGFBP-3) was found again indicating that bioactive hGH was released from the microspheres. Good in vitro in vivo correlations were obtained for hGH-loaded dex-HEMA microspheres, which is an important advantage in predicting the effect of the controlled drug delivery product in a clinical situations.

Preclinical and Clinical In Vitro In Vivo Correlation of an hGH Dextran Microsphere Formulation

PubMed Central

de Vrueh, R.; Gresnigt, M. G.; Hoogerbrugge, C. M.; van Buul-Offers, S. C.; de Leede, L. G. J.; Sterkman, L. G. W.; Crommelin, D. J. A.; Hennink, W. E.; Verrijk, R.

2007-01-01

Purpose To investigate the in vitro in vivo correlation of a sustained release formulation for human growth hormone (hGH) based on hydroxyethyl methacrylated dextran (dex-HEMA) microspheres in Pit-1 deficient Snell dwarf mice and in healthy human volunteers. Materials and Methods A hGH-loaded microsphere formulation was developed and tested in Snell dwarf mice (pharmacodynamic study) and in healthy human volunteers (pharmacokinetic study). Results Single subcutaneous administration of the microspheres in mice resulted in a good correlation between hGH released in vitro and in vivo effects for the hGH-loaded microsphere formulation similar to daily injected hGH indicating a retained bioactivity. Testing the microspheres in healthy volunteers showed an increase (over 7–8 days) in hGH serum concentrations (peak concentrations: 1–2.5 ng/ml). A good in vitro in vivo correlation was obtained between the measured and calculated (from in vitro release data) hGH serum concentrations. Moreover, an increased serum concentration of biomarkers (insulin-like growth factor-I (IGF-I), IGF binding protein-3 (IGFBP-3) was found again indicating that bioactive hGH was released from the microspheres. Conclusions Good in vitro in vivo correlations were obtained for hGH-loaded dex-HEMA microspheres, which is an important advantage in predicting the effect of the controlled drug delivery product in a clinical situations. PMID:17929148

Antimicrobial effect of chlorhexidine digluconate in dentin: In vitro and in situ study.

PubMed

Borges, Fátima Maria Cavalcante; de Melo, Mary Anne Sampaio; Lima, Juliana Paiva Marques; Zanin, Iriana Carla Junqueria; Rodrigues, Lidiany Karla Azevedo

2012-01-01

The aim of this study was to evaluate a very short-term in vitro and in situ effect of 2% chlorhexidine-digluconate-based (CHX) cavity cleanser on the disinfection of dentin demineralized by cariogenic bacteria. Human dentin slabs were randomly allocated and used in 2 distinct phases, in vitro and in situ, for obtaining demineralized dentin. In vitro, the slabs (n=15) were immersed for 5 days in BHI broth inoculated with Streptococcus mutans CTT 3440. In situ, a double-blind design was conducted in one phase of 14 days, during which 20 volunteers wore palatal devices containing two human dental dentin slabs. On 5(th) day in vitro and 14(th) day in situ, the slabs were allocated to the two groups: Control group (5 μl of 0.9% NaCl solution) and CHX group (5 μl of 2% chlorhexidine digluconate solution, Cavity Cleanser™ BISCO, Schaumburg, IL, EUA), for 5 minutes. The microbiological analyses were performed immediately before and after the treatments. The log reductions means found for CHX treatment on tested micro organisms were higher when compared to Control group either in vitro or in situ conditions. Our results showed that CHX was effective in reducing the cultivable microbiota in contaminated dentin. Furthermore, although the use of chlorhexidine-digluconate-based cavity disinfectant did not completely eliminate the viable microorganisms, it served as a suitable agent to disinfect tooth preparations.

Use of external metabolizing systems when testing for endocrine disruption in the T-screen assay.

PubMed

Taxvig, Camilla; Olesen, Pelle Thonning; Nellemann, Christine

2011-02-01

Although, it is well-established that information on the metabolism of a substance is important in the evaluation of its toxic potential, there is limited experience with incorporating metabolic aspects into in vitro tests for endocrine disrupters. The aim of the current study was a) to study different in vitro systems for biotransformation of ten known endocrine disrupting chemicals (EDs): five azole fungicides, three parabens and 2 phthalates, b) to determine possible changes in the ability of the EDs to bind and activate the thyroid receptor (TR) in the in vitro T-screen assay after biotransformation and c) to investigate the endogenous metabolic capacity of the GH3 cells, the cell line used in the T-screen assay, which is a proliferation assay used for the in vitro detection of agonistic and antagonistic properties of compounds at the level of the TR. The two in vitro metabolizing systems tested the human liver S9 mix and the PCB-induced rat microsomes gave an almost complete metabolic transformation of the tested parabens and phthalates. No marked difference the effects in the T-screen assay was observed between the parent compounds and the effects of the tested metabolic extracts. The GH3 cells themselves significantly metabolized the two tested phthalates dimethyl phthalate (DMP) and diethyl phthalate (DEP). Overall the results and qualitative data from the current study show that an in vitro metabolizing system using liver S9 or microsomes could be a convenient method for the incorporation of metabolic and toxicokinetic aspects into in vitro testing for endocrine disrupting effects. Copyright © 2010 Elsevier Inc. All rights reserved.

Drug repurposing: In-vitro anti-glycation properties of 18 common drugs

PubMed Central

Rasheed, Saima; Sánchez, Sara S.; Yousuf, Sammer; Honoré, Stella M.; Choudhary, M. Iqbal

2018-01-01

Drug repositioning or repurposing, i.e. identifying new indications for existing drugs, has gained increasing attention in the recent years. This approach enables the scientists to discover “new targets” for known drugs in a cost and time efficient manner. Glycation, the non-enzymatic reaction of sugars with proteins or nucleic acids to form early glycation (Amadori or fructosamine) products, is a key molecular basis of diabetic complications. Inhibiting the process of non-enzymatic protein glycation is one of the key strategies to prevent glycation-mediated diabetic complications. The present study focuses on the anti-glycation activity of 18 drugs, commonly used for the treatment of gastrointestinal, central nervous system, inflammatory diseases, bacterial infections, and gout. This study was carried out by using two in-vitro protein anti-glycation assay models. Results revealed that nimesulide (3), a non-steroidal anti-inflammatory drug, possesses a good anti-glycation activity in in-vitro BSA-MG and BSA-glucose glycation models with IC50 values of 330.56 ± 2.90, and 145.46 ± 16.35 μM, respectively. Phloroglucinol dihydrate (11), a drug used for the treatment of gastrointestinal diseases, showed a weak activity in BSA-MG glycation model (IC50 = 654.89 ± 2.50 μM), while it showed a good activity in BSA-glucose assay (IC50 = 148.23 ± 0.15 μM). Trimethylphloroglucinol (9), a drug used for the treatment of pain related to functional disorders of the digestive and biliary tracts, also showed a good antiglycation activity in BSA-MG model (IC50 = 321.15 ± 1.26 μM), while it was found to be inactive in in-vitro BSA-glucose assay (IC50 = 12.95% inhibition). These activities of drugs were compared with the anti-glycation activity of the standard, rutin (IC50 = 294.5 ± 1.50 μM in BSA-MG glycation model, and IC50 = 86.94 ± 0.24 μM in BSA- glucose model). Rest of the drugs exhibited a relatively weak antiglycation activity. This study identifies nimesulide (3), and phloroglucinol dihydrate (11) as new inhibitors of in-vitro protein glycation for further investigations as potential anti-diabetic agents. PMID:29300762

Bone morphogenetic protein 7 (BMP-7) influences tendon-bone integration in vitro.

PubMed

Schwarting, Tim; Lechler, Philipp; Struewer, Johannes; Ambrock, Marius; Frangen, Thomas Manfred; Ruchholtz, Steffen; Ziring, Ewgeni; Frink, Michael

2015-01-01

Successful graft ingrowth following reconstruction of the anterior cruciate ligament is governed by complex biological processes at the tendon-bone interface. The aim of this study was to investigate in an in vitro study the effects of bone morphogenetic protein 7 (BMP-7) on tendon-bone integration. To study the biological effects of BMP-7 on the process of tendon-bone-integration, two independent in vitro models were used. The first model involved the mono- and coculture of bovine tendon specimens and primary bovine osteoblasts with and without BMP-7 exposure. The second model comprised the mono- and coculture of primary bovine osteoblasts and fibroblasts. Alkaline phosphatase (ALP), lactate dehydrogenase (LDH), lactate and osteocalcin (OCN) were analyzed by ELISA. Histological analysis and electron microscopy of the tendon specimens were performed. In both models, positive effects of BMP-7 on ALP enzyme activity were observed (p<0.001). Additionally, similar results were noted for LDH activity and lactate concentration. BMP-7 stimulation led to a significant increase in OCN expression. Whereas the effects of BMP-7 on tendon monoculture peaked during an early phase of the experiment (p<0.001), the cocultures showed a maximal increase during the later stages (p<0.001). The histological analysis showed a stimulating effect of BMP-7 on extracellular matrix formation. Organized ossification zones and calcium carbonate-like structures were only observed in the BMP-stimulated cell cultures. This study showed the positive effects of BMP-7 on the biological process of tendon-bone integration in vitro. Histological signs of improved mineralization were paralleled by increased rates of osteoblast-specific protein levels in primary bovine osteoblasts and fibroblasts. Our findings indicated a role for BMP-7 as an adjuvant therapeutic agent in the treatment of ligamentous injuries, and they emphasized the importance of the transdifferentiation process of tendinous fibroblasts at the tendon-bone interface.

Evaluation of anthelmintic potential of the Ethiopian medicinal plant Embelia schimperi Vatke in vivo and in vitro against some intestinal parasites.

PubMed

Debebe, Yared; Tefera, Mesfin; Mekonnen, Walelign; Abebe, Dawit; Woldekidan, Samuel; Abebe, Abiy; Belete, Yehualashet; Menberu, Temesgen; Belayneh, Bethelhem; Tesfaye, Berhanu; Nasir, Ibrahim; Yirsaw, Kidist; Basha, Hirut; Dawit, Asrat; Debella, Asfaw

2015-06-18

Embelia schimperi has been used for the treatment of intestinal parasites especially tapeworm infestations for centuries in Ethiopia. However, there is lack of scientific based evidences regarding the efficacy, safety and phytochemical analysis of this plant despite its frequent use as an anthelmintic. This study has therefore evaluated the efficacy and acute toxicity of E. schimperi thereby generating relevant preclinical information. The anthelmintic activities of the crude hydroalcoholic extract of E. schimperi and the isolated compound, embelin, were conducted using in vivo and in vitro models against the dwarf tapeworm, Hymenolepis nana, and the hookworm, Necator americanus, respectively. LD50 of the crude hydroalcoholic extract was determined using Swiss albino mice following the OECD guidelines. Chemical characterization of the isolated embelin was conducted using UV-spectroscopy, HPLC and NMR. In the acute toxicity study no prominent signs of toxicity and mortality were recorded among the experimental animals at the highest administered dose. Hence the LD50 of the plant was found to be higher than 5000 mg/kg. In vivo cestocidal activity of the crude hydroalcoholic extract of E. schimperi showed 100% parasite clearance at 1000 mg/kg, while the diammonium salt of embelin showed 85.3% parasite clearance at 750 mg/kg. The in vitro anthelminthic activity study revealed that the LC50 value of the crude extract and albendazole were 228.7 and 51.33 μg/mL, respectively. The results clearly indicated that the hydroalcoholic extract of E. schimperi and the diammonium salt of the isolated compound embelin had anthelmintic activity against hookworm larva in vitro and H. nana in vivo. Hence the findings of this study showed Embelia schimperi appears to possess some anthelmintic activity that may support the usage of these plants by local traditional healers to treat helminthic infestations.

Preparation and in vitro characterization of pluronic-attached polyamidoamine dendrimers for drug delivery.

PubMed

Gu, Zhuojun; Wang, Meng; Fang, Qiongyan; Zheng, Huaiyu; Wu, Feiyue; Lin, Dai; Xu, Ying; Jin, Yi

2015-05-01

Polyamidoamine (PAMAM) dendrimers have attracted lots of interest as drug carriers. And little study about whether pluronic-attached PAMAM dendrimers could be potential drug delivery systems has been carried on. Pluronic F127 (PF127) attached PAMAM dendrimers were designed as novel drug carriers. Two conjugation ratios of PF127-attached PAMAM dendrimers were synthesized. (1)H nuclear magnetic resonance ((1)H-NMR), Fourier transform infrared spectrum (FTIR), element analysis and ninhydrin assay were used to characterize the conjugates. Size, zeta potential and critical micelle concentrations (CMC) were also detected. And DOX was incorporated into the hydrophobic interior of the conjugates. Studies on their drug loading and drug release were carried on. Furthermore, hemolysis and cytotoxicity assay were used to evaluate the toxicity of the conjugates. PF127 was successfully conjugated to the fifth generation PAMAM dendrimer at two molar ratios of 19% and 57% (PF127 to surface amine per PAMAM dendrimer molecular). The conjugates showed an increased size and a reduced zeta potential. And higher CMC values were obtained than pure PF127. Compared with unconjugated PAMAM dendrimer, PF127 conjugation significantly reduced the hemolytic toxicity and cytotoxicity of PAMAM dendrimer in vitro. The encapsulation results showed that the ability to encapsulate DOX by the conjugate of 19% conjugation ratio was better than that of 57% conjugation ratio. And the maximum is ∼12.87 DOX molecules per conjugate molecule. Moreover, the complexes showed a sustained release behavior compared to pure DOX. Findings from the in vitro study show that the PF127-attached PAMAM dendrimers may be potential carriers for drug delivery.

Activity of antibacterial, antiviral, anti-inflammatory in compounds andrographolide salt.

PubMed

Wen, Li; Xia, Nan; Chen, Xianghong; Li, Yingxiu; Hong, Yi; Liu, YaJie; Wang, ZiZhen; Liu, YaJie

2014-10-05

Andrographolide sulfonic acid sodium salt (ASS) was synthesized to increase the the solubility of Andrographolide in aqueous solution. We have studied its pharmacological effect of antibiosis, anti-inflammatory and immunoregulation. Cylinder-plate method was used to study ASS׳s in vitro antibacterial activity, and its protection for mice infected by Staphylococcus aureus and Shigella dysenteriae. Various inflammation models, including the auricular edema induced by xylene in mice, CMC-Na induced air pounch model and the paw edema induced by albumen in rats were used to explore the characteristic of ASS׳s anti-inflammation effect. We built up the immune model by injecting chicken red cells in enter celiac of mice and study the effect of ASS on immunoregulation, taking andrographolide as the positive control. bacteriostasis in vivo and in vitro experiments show that ASS has a weak antibacterial effect and no bactericidal effect, but can reduce the mice mortality of Staphylococcus aureus infected. Anti-inflammatory experiments show that ASS can reduce the mouse ear swelling induced by xylene and rat paw swelling induced by egg albumin, and lessen leukocytes in air bag caused by CMCNa, and lower IL1 not ably in rat serum. Immune tests indicate that ASS can get spleen and thymus gain weight and increase rate of abdominal macrophage phagocytosis of mice. The result of bacteriostasis shows that ASS has weak in vitro antibacterial effect. ASS shows significant effects of anti-inflammation and improving immunity, thus enables the mice against bacteria better. Copyright © 2014 Elsevier B.V. All rights reserved.

An in vitro and in vivo evaluation of new potential trans-sialidase inhibitors of Trypanosoma cruzi predicted by a computational drug repositioning method.

PubMed

Lara-Ramirez, Edgar E; López-Cedillo, Julio Cesar; Nogueda-Torres, Benjamin; Kashif, Muhammad; Garcia-Perez, Carlos; Bocanegra-Garcia, Virgilio; Agusti, Rosalía; Uhrig, María Laura; Rivera, Gildardo

2017-05-26

Chagas disease is one of the most important neglected parasitic diseases afflicting developed and undeveloped countries. There are currently limited options for inexpensive and secure pharmacological treatment. In this study, we employed a structure-based virtual screening protocol for 3180 FDA-approved drugs for repositioning of them as potential trans-sialidase inhibitors. In vitro and in vivo evaluations were performed for the selected drugs against trypomastigotes from the INC-5 and NINOA strains of T. cruzi. Also, inhibition of sialylation by the trans-sialidase enzyme reaction was evaluated using high-performance anion-exchange chromatography with pulse amperometric detection to confirm the mechanism of action. Results from the computational study showed 38 top drugs with the best binding-energies. Four compounds with antihistaminic, anti-hypertensive, and antibiotic properties showed better trypanocidal effects (LC 50 range = 4.5-25.8 μg/mL) than the reference drugs, nifurtimox and benznidazole (LC 50 range = 36.1-46.8 μg/mL) in both strains in the in vitro model. The anti-inflammatory, sulfasalazine showed moderate inhibition (37.6%) of sialylation in a trans-sialidase enzyme inhibition reaction. Sulfasalazine also showed the best trypanocidal effects in short-term in vivo experiments on infected mice. This study suggests for the first time that the anti-inflammatory sulfasalazine could be used as a lead compound to develop new trans-sialidase inhibitors. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Proflavine derivatives as fluorescent imaging agents of amyloid deposits.

PubMed

Garin, Dominique; Oukhatar, Fatima; Mahon, Andrew B; Try, Andrew C; Dubois-Dauphin, Michel; Laferla, Frank M; Demeunynck, Martine; Sallanon, Marcelle Moulin; Chierici, Sabine

2011-04-15

A series of proflavine derivatives for use to further image Aβ amyloid deposits were synthesized and characterized. Aged 3xTg-AD (23 months old) mice hippocampus sections incubated with these derivatives revealed preferential labeling of amyloid plaques. Furthermore an in vitro binding study showed an inhibitory effect, although moderate, of these compounds on Aβ(40) fibril formation. This study highlights the potential of proflavine as a molecular scaffold for designing new Aβ imaging agents, its native fluorescence allowing in vitro neuropathological staining in AD damaged brain sections. Copyright © 2011 Elsevier Ltd. All rights reserved.

Fluorescent composite scaffolds made of nanodiamonds/polycaprolactone

NASA Astrophysics Data System (ADS)

Cao, Li; Hou, Yanwen; Lafdi, Khalid; Urmey, Kirk

2015-11-01

Polycaprolactone (PCL) has been widely studied for biological applications. Biodegradable PCL fibrous scaffold can work as an appropriate substrate for tissue regeneration. In this letter, fluorescent nanodiamonds (FNDs) were prepared after surface passivation with octadecylamine. The FNDs were then mixed with PCL polymer and subsequently electrospun into FNDs/PCL fibrous scaffolds. The obtained scaffolds not only exhibited photoluminescence, but also showed reinforced mechanical strength. Toxicity study indicated FNDs/PCL scaffolds were nontoxic. This biocompatible fluorescent composite fibrous scaffold can support in vitro cell growth and also has the potential to act as an optical probe for tissue engineering application in vitro and in vivo.

Fucoxanthin bioavailability from fucoxanthin-fortified milk: In vivo and in vitro study.

PubMed

Mok, Il-Kyoon; Lee, Jae Kwon; Kim, Jeong Hwa; Pan, Cheol-Ho; Kim, Sang Min

2018-08-30

Our previous study reported the improved stability of fucoxanthin (FX) fortified in whole milk (WM) and skimmed milk (SM). In this study, in vivo and in vitro FX bioavailability were investigated using FX-fortified milk (FX-SM and FX-WM) and microalga Phaeodactylum tricornutum biomass (Pt-powder). Organ tissue accumulation of FX and its metabolites (FXOH: fucoxanthinol, AXA: amarouciaxanthin A) after repeated oral administration was in the following order: FX-SM > FX-WM > Pt-powder. In vivo pharmacokinetic study with a single oral administration also demonstrated that the absorption of FXOH and AXA was the highest for FX-SM. To reinforce the in vivo results, in vitro-simulated digestion and Caco-2 cell uptake assays were performed, which revealed that FX-SM showed the highest FX bioaccessibility (release from food matrices) and cellular uptake efficiency of FX and FXOH. In conclusion, skimmed milk was validated as an excellent food matrix for FX application in terms of stability and bioavailability. Copyright © 2018 Elsevier Ltd. All rights reserved.

Factors Affecting the Longevity and Strength in an In Vitro Model of the Bone–Ligament Interface

PubMed Central

Paxton, Jennifer Z.; Donnelly, Kenneth; Keatch, Robert P.; Grover, Liam M.

2010-01-01

The interfaces between musculoskeletal tissues with contrasting moduli are morphologically and biochemically adapted to allow the transmission of force with minimal injury. Current methods of tissue engineering ligaments and tendons do not include the interface and this may limit the future clinical success of engineered musculoskeletal tissues. This study aimed to use solid brushite cement anchors to engineer intact ligaments from bone-to-bone, creating a functional musculoskeletal interface in vitro. We show here that modifying anchor shape and cement composition can alter both the longevity and the strength of an in vitro model of the bone–ligament interface: with values reaching 23 days and 21.6 kPa, respectively. These results validate the use of brushite bone cement to engineer the bone–ligament interface in vitro and raise the potential for future use in ligament replacement surgery. PMID:20431953

Evaluation of anti-inflammatory potential of leaf extracts of Skimmia anquetilia

PubMed Central

Kumar, Vijender; Bhat, Zulfiqar Ali; Kumar, Dinesh; Khan, NA; Chashoo, IA

2012-01-01

Objective To evaluate anti-inflammatory potential of leaf extract of Skimmia anquetilia by in-vitro and in-vivo anti-inflammatory models. Methods Acute toxicity study was carried out to determine the toxicity level of different extract using acute toxic class method as described in Organization of Economic Co-operation and Development Guidelines No.423. Carrageenan (1% w/w) was administered and inflammation was induced in rat paw. The leaf extracts of Skimmia anquetilia were evaluated for anti-inflammatory activity by in-vitro human red blood cell (HRBC) membrane stabilization method and in-vivo carrangeenan-induced rat paw edema method. Results The in-vitro membrane stabilizing test showed petroleum ether (PE), chloroform (CE), ethyl acetate (EE), methanol (ME) and aqueous extracts (AE) showed 49.44%, 59.39%, 60.15%, 68.40% and 52.18 % protection, respectively as compared to control groups. The in-vivo results of CE, EE and ME showed 58.20%, 60.17% and 67.53% inhibition of inflammation after 6h administration of test drugs in albino rats. The potency of the leaf extracts of Skimmia anquetilia were compared with standard diclofenac (10 mg/kg) which showed 74.18% protection in in-vitro HRBC membrane stabilization test and 71.64% inhibition in in-vivo carrangeenan-induced rat paw edema model. The ME showed a dose dependent significant (P< 0.01) anti-inflammatory activity in human red blood cell membrane stabilization test and reduction of edema in carrageenan induced rat paw edema. Conclusions The present investigation has confirmed the anti-inflammatory activity of Skimmia anquetilia due to presence of bioactive phytoconstitutes for the first time and provide the pharmacological evidence in favor of traditional claim of Skimmia anquetilia as an anti- inflammatory agent. PMID:23569983

In vitro transcription of two human rotaviruses.

PubMed Central

Flores, J; Myslinski, J; Kalica, A R; Greenberg, H B; Wyatt, R G; Kapikian, A Z; Chanock, R M

1982-01-01

The RNA polymerase activities of a cultivatable (Wa) and a noncultivatable (DS-1) strain of human rotavirus were studied. Under optimal conditions, transcription of all of their RNA segments occurred, as evidenced by the hybridization of labeled transcripts to genomic RNA. Cross-hybridization between the two viruses showed that none of their 11 genes were completely homologous. The transcription products could be translated in vitro, yielding proteins with an electrophoretic pattern resembling that obtained with proteins labeled in vivo during infection with the Wa virus. Images PMID:6292446

Studies on strain sensitivity of Trichomonas vaginalis to metronidazole.

PubMed Central

Meingassner, J G; Havelec, L; Mieth, H

1978-01-01

Ninety-four strains of Trichomonas vaginalis isolated from unselected patients in Vienna during the period from November 1975 to November 1975 were subjected to sensitivity tests against metronidazole in vitro and in vivo. Nearly 90% of the isolates examined showed in vitro growth inhibition at concentrations of 0.4 to 1.6 microgram metronidazole/ml and only about 10% at concentrations of less than 0.4 microgram/ml. All isolates showed some inhibition of growth after incubation for 48 hours with 3.2 microgram metroidazole/ml. Tests on treatment carried out on experimentally infected mice revealed that all strains of T. vaginalis examined were sensitive to metronidazole (MDE: less than 3 x 35 mg/kg peroral). PMID:305807

Assessing the risk of pH-dependent absorption for new molecular entities: a novel in vitro dissolution test, physicochemical analysis, and risk assessment strategy.

PubMed

Mathias, Neil R; Xu, Yan; Patel, Dhaval; Grass, Michael; Caldwell, Brett; Jager, Casey; Mullin, Jim; Hansen, Luke; Crison, John; Saari, Amy; Gesenberg, Christoph; Morrison, John; Vig, Balvinder; Raghavan, Krishnaswamy

2013-11-04

Weak base therapeutic agents can show reduced absorption or large pharmacokinetic variability when coadministered with pH-modifying agents, or in achlorhydria disease states, due to reduced dissolution rate and/or solubility at high gastric pH. This is often referred to as pH-effect. The goal of this study was to understand why some drugs exhibit a stronger pH-effect than others. To study this, an API-sparing, two-stage, in vitro microdissolution test was developed to generate drug dissolution, supersaturation, and precipitation kinetic data under conditions that mimic the dynamic pH changes in the gastrointestinal tract. In vitro dissolution was assessed for a chemically diverse set of compounds under high pH and low pH, analogous to elevated and normal gastric pH conditions observed in pH-modifier cotreated and untreated subjects, respectively. Represented as a ratio between the conditions, the in vitro pH-effect correlated linearly with clinical pH-effect based on the Cmax ratio and in a non-linear relationship based on AUC ratio. Additionally, several in silico approaches that use the in vitro dissolution data were found to be reasonably predictive of the clinical pH-effect. To explore the hypothesis that physicochemical properties are predictors of clinical pH-effect, statistical correlation analyses were conducted using linear sequential feature selection and partial least-squares regression. Physicochemical parameters did not show statistically significant linear correlations to clinical pH-effect for this data set, which highlights the complexity and poorly understood nature of the interplay between parameters. Finally, a strategy is proposed for implementation early in clinical development, to systematically assess the risk of clinical pH-effect for new molecular entities that integrates physicochemical analysis and in vitro, in vivo and in silico methods.

In vitro and in vivo studies of biodegradable fine grained AZ31 magnesium alloy produced by equal channel angular pressing.

PubMed

Ratna Sunil, B; Sampath Kumar, T S; Chakkingal, Uday; Nandakumar, V; Doble, Mukesh; Devi Prasad, V; Raghunath, M

2016-02-01

The objective of the present work is to investigate the role of different grain sizes produced by equal channel angular pressing (ECAP) on the degradation behavior of magnesium alloy using in vitro and in vivo studies. Commercially available AZ31 magnesium alloy was selected and processed by ECAP at 300°C for up to four passes using route Bc. Grain refinement from a starting size of 46μm to a grain size distribution of 1-5μm was successfully achieved after the 4th pass. Wettability of ECAPed samples assessed by contact angle measurements was found to increase due to the fine grain structure. In vitro degradation and bioactivity of the samples studied by immersing in super saturated simulated body fluid (SBF 5×) showed rapid mineralization within 24h due to the increased wettability in fine grained AZ31 Mg alloy. Corrosion behavior of the samples assessed by weight loss and electrochemical tests conducted in SBF 5× clearly showed the prominent role of enhanced mineral deposition on ECAPed AZ31 Mg in controlling the abnormal degradation. Cytotoxicity studies by MTT colorimetric assay showed that all the samples are viable. Additionally, cell adhesion was excellent for ECAPed samples particularly for the 3rd and 4th pass samples. In vivo experiments conducted using New Zealand White rabbits clearly showed lower degradation rate for ECAPed sample compared with annealed AZ31 Mg alloy and all the samples showed biocompatibility and no health abnormalities were noticed in the animals after 60days of in vivo studies. These results suggest that the grain size plays an important role in degradation management of magnesium alloys and ECAP technique can be adopted to achieve fine grain structures for developing degradable magnesium alloys for biomedical applications. Copyright © 2015 Elsevier B.V. All rights reserved.

In vitro production of M. × piperita not containing pulegone and menthofuran.

PubMed

Bertoli, Alessandra; Leonardi, Michele; Krzyzanowska, Justyna; Oleszek, Wieslaw; Pistelli, Luisa

2012-01-01

The essential oils (EOs) and static headspaces (HSs) of in vitro plantlets and callus of Mentha x piperita were characterized by GC-MS analysis. Leaves were used as explants to induce in vitro plant material. The EO yields of the in vitro biomass were much lower (0.1% v/w) than those of the parent plants (2% v/w). Many typical mint volatiles were emitted by the in vitro production, but the callus and in vitro plantelet EOs were characterized by the lack of both pulegone and menthofuran. This was an important difference between in vitro and in vivo plant material as huge amounts of pulegone and menthofuran may jeopardise the safety of mint essential oil. Regarding the other characteristic volatiles, menthone was present in reduced amounts (2%) in the in vitro plantlets and was not detected in the callus, even if it represented the main constituent of the stem and leaf EOs obtained from the cultivated mint (26% leaves; 33% stems). The M. piperita callus was characterized by menthol (9%) and menthone (2%), while the in vitro plantlet EO showed lower amounts of both these compounds in favour of piperitenone oxide (45%). Therefore, the established callus and in vitro plantlets showed peculiar aromatic profiles characterized by the lack of pulegone and menthofuran which have to be monitored in the mint oil for their toxicity.

[Differentiation of bone marrow derived from mesenchymal stem cells into cardiomyocyte-like cells induced by co-culture with rat myocardial cells].

PubMed

Zhang, Rong-Li; Jiang, Er-Lie; Wang, Mei; Zhou, Zheng; Zhai, Wen-Jing; Zhai, Wei-Hua; Wang, Hua; Wang, Zhi-Yong; Bao, Yu-Shi; DU, Hong; Han, Ming-Zhe

2008-10-01

The study was purposed to investigate the differentiation ability of mesenchymal stem cells (MSCs) into myocardial cells in vitro. Rat bone marrow-derived MSCs were labeled and co-cultured with neonatal rat cardiomyocytes (CM) for 5 - 7 days. The expression of cell surface antigens was detected by flow cytometry, and the expression of muscle-specific marker myosin and troponin T in labeled cells was detected by immunofluorescence. The results showed that in vitro cultured MSCs expressed CD90, CD44, CD105, CD54, not expressed CD34, CD45, CD31. After co-cultured with neonatal rat CM, labeled MSCs differentiated into cardiomyocyte-like cells expressing myosin and troponin T. It is concluded that MSCs can differentiate into cardiomyocyte-like cells when co-cultured with neonatal myocardial cells in vitro. In co-culture of two kind of cells in ratio of four to one showed obvious efficacy differentiating MSCs into CMs.

Effects of Chain Length and Degree of Unsaturation of Fatty Acids on Structure and in Vitro Digestibility of Starch-Protein-Fatty Acid Complexes.

PubMed

Zheng, Mengge; Chao, Chen; Yu, Jinglin; Copeland, Les; Wang, Shuo; Wang, Shujun

2018-02-28

The effects of chain length and degree of unsaturation of fatty acids (FAs) on structure and in vitro digestibility of starch-protein-FA complexes were investigated in model systems. Studies with the rapid visco analyzer (RVA) showed that the formation of ternary complex resulted in higher viscosities than those of binary complex during the cooling and holding stages. The results of differential scanning calorimetry (DSC), Raman, and X-ray diffraction (XRD) showed that the structural differences for ternary complexes were much less than those for binary complexes. Starch-protein-FA complexes presented lower in vitro enzymatic digestibility compared with starch-FAs complexes. We conclude that shorter chain and lower unsaturation FAs favor the formation of ternary complexes but decrease the thermal stability of these complexes. FAs had a smaller effect on the ordered structures of ternary complexes than on those of binary complexes and little effect on enzymatic digestibility of both binary and ternary complexes.

In-vitro Antioxidant Activities of the Ethanolic Extracts of Some Contained-Allantoin Plants

PubMed Central

Selamoglu, Zeliha; Dusgun, Cihan; Akgul, Hasan; Gulhan, Mehmet Fuat

2017-01-01

It has been investigated the in-vitro antioxidant properties of ethanol extracts of the contained-allantoin plants in this study. Contained-allantoined plant samples Plantago lanceolata, Plantago major, Robinia pseudoacacia, Platanus orientalis and Aesculus hippocastanum were tested at different concentrations. The antioxidant activities of plant samples were analysed by 1,1- diphenyl-2-picrylhydrazyl (DPPH) radical scavenging method, cupric reducing antioxidant capacity (CUPRAC), reducing power assay and β-carotene bleaching method. Plantago major plant showed the highest antioxidant capacity compared to other plant extracts in results of the in-vitro assays including 1,1- diphenyl-2-picrylhydrazyl (DPPH) radical scavenging method with 90.25 %, cupric reducing antioxidant capacity (CUPRAC) with 1.789 %, reducing power assay (FRAP) with 1.321 % and β-carotene bleaching method with 78.01 % in 1 mg/mL. The lowest antioxidant activity was determined in Robinia pseudoacacia plant. In conclusion, allantoin shows antioxidant properties and it has the positive effect on total antioxidant capacity.

In vitro and in vivo antileishmanial and trypanocidal studies of new N-benzene- and N-naphthalenesulfonamide derivatives.

PubMed

Galiana-Roselló, Cristina; Bilbao-Ramos, Pablo; Dea-Ayuela, M Auxiliadora; Rolón, Miriam; Vega, Celeste; Bolás-Fernández, Francisco; García-España, Enrique; Alfonso, Jorge; Coronel, Cathia; González-Rosende, M Eugenia

2013-11-27

We report in vivo and in vitro antileishmanial and trypanocidal activities of a new series of N-substituted benzene and naphthalenesulfonamides 1-15. Compounds 1-15 were screened in vitro against Leishmania infantum , Leishmania braziliensis , Leishmania guyanensis , Leishmania amazonensis , and Trypanosoma cruzi . Sulfonamides 6e, 10b, and 10d displayed remarkable activity and selectivity toward T. cruzi epimastigotes and amastigotes. 6e showed significant trypanocidal activity on parasitemia in a murine model of acute Chagas disease. Moreover, 6e, 8c, 9c, 12c, and 14d displayed interesting IC50 values against Leishmania spp promastigotes as well as L. amazonensis and L. infantum amastigotes. 9c showed excellent in vivo activity (up to 97% inhibition of the parasite growth) in a short-term treatment murine model for acute infection by L. infantum. In addition, the effect of compounds 9c and 14d on tubulin as potential target was assessed by confocal microscopy analysis applied to L. infantum promastigotes.

Semi-purification procedures of prions from a prion-infected brain using sucrose has no influence on the nonenzymatic glycation of the disease-associated prion isoform.

PubMed

Choi, Yeong-Gon; Kim, Jae-Il; Choi, Eun-Kyoung; Carp, Richard I; Kim, Yong-Sun

2016-01-01

Previous studies have shown that the Nε-carboxymethyl group is linked to not only one or more N-terminal Lys residues but also to one or more Lys residues of the protease-resistant core region of the pathogenic prion isoform (PrPSc) in prion-infected brains. Using an anti-advanced glycation end product (AGE) antibody, we detected nonenzymatically glycated PrPSc (AGE-PrPSc) in prion-infected brains following concentration by a series of ultracentrifugation steps with a sucrose cushion. In the present study, the levels of in vitro nonenzymatic glycation of PrPSc using sucrose were investigated to determine whether sucrose cushion can artificially and nonenzymatically induce in vitro glycation during ultracentrifugation. The first insoluble pellet fraction following the first ultracentrifugation (PU1st) collected from 263K scrapie-infected brains was incubated with sucrose, glucose or colloidal silica coated with polyvinylpyrrolidone (percoll). None of the compounds in vitro resulted in AGE-PrPSc. Nonetheless, glucose and percoll produced AGEs in vitro from other proteins within PU1st of the infected brains. This reaction could lead to the AGE-modified polymer(s) of nonenzymatic glycation-prone protein(s). This study showed that PrPSc is not nonenzymatically glycated in vitro with sucrose, glucose or percoll and that AGE-modified PrPSc can be isolated and enriched from prion-infected brains.

Lack of in vitro-in vivo correlation for a UC781-releasing vaginal ring in macaques.

PubMed

McConville, Christopher; Smith, James M; McCoy, Clare F; Srinivasan, Priya; Mitchell, James; Holder, Angela; Otten, Ron A; Butera, Salvatore; Doncel, Gustavo F; Friend, David R; Malcolm, R Karl

2015-02-01

This study describes the preclinical development of a matrix-type silicone elastomer vaginal ring device designed to provide controlled release of UC781, a non-nucleoside reverse transcriptase inhibitor. Testing of both human- and macaque-sized rings in a sink condition in vitro release model demonstrated continuous UC781 release in quantities considered sufficient to maintain vaginal fluid concentrations at levels 82-860-fold higher than the in vitro IC50 (2.0 to 10.4 nM) and therefore potentially protect against mucosal transmission of HIV. The 100-mg UC781 rings were well tolerated in pig-tailed macaques, did not induce local inflammation as determined by cytokine analysis and maintained median concentrations in vaginal fluids of UC781 in the range of 0.27 to 5.18 mM during the course of the 28-day study. Analysis of residual UC781 content in rings after completion of both the in vitro release and macaque pharmacokinetic studies revealed that 57 and 5 mg of UC781 was released, respectively. The pharmacokinetic analysis of a 100-mg UC781 vaginal ring in pig-tailed macaques showed poor in vivo-in vitro correlation, attributed to the very poor solubility of UC781 in vaginal fluid and resulting in a dissolution-controlled drug release mechanism rather than the expected diffusion-controlled mechanism.

IFITM3 restricts the morbidity and mortality associated with influenza.

PubMed

Everitt, Aaron R; Clare, Simon; Pertel, Thomas; John, Sinu P; Wash, Rachael S; Smith, Sarah E; Chin, Christopher R; Feeley, Eric M; Sims, Jennifer S; Adams, David J; Wise, Helen M; Kane, Leanne; Goulding, David; Digard, Paul; Anttila, Verneri; Baillie, J Kenneth; Walsh, Tim S; Hume, David A; Palotie, Aarno; Xue, Yali; Colonna, Vincenza; Tyler-Smith, Chris; Dunning, Jake; Gordon, Stephen B; Smyth, Rosalind L; Openshaw, Peter J; Dougan, Gordon; Brass, Abraham L; Kellam, Paul

2012-03-25

The 2009 H1N1 influenza pandemic showed the speed with which a novel respiratory virus can spread and the ability of a generally mild infection to induce severe morbidity and mortality in a subset of the population. Recent in vitro studies show that the interferon-inducible transmembrane (IFITM) protein family members potently restrict the replication of multiple pathogenic viruses. Both the magnitude and breadth of the IFITM proteins' in vitro effects suggest that they are critical for intrinsic resistance to such viruses, including influenza viruses. Using a knockout mouse model, we now test this hypothesis directly and find that IFITM3 is essential for defending the host against influenza A virus in vivo. Mice lacking Ifitm3 display fulminant viral pneumonia when challenged with a normally low-pathogenicity influenza virus, mirroring the destruction inflicted by the highly pathogenic 1918 'Spanish' influenza. Similar increased viral replication is seen in vitro, with protection rescued by the re-introduction of Ifitm3. To test the role of IFITM3 in human influenza virus infection, we assessed the IFITM3 alleles of individuals hospitalized with seasonal or pandemic influenza H1N1/09 viruses. We find that a statistically significant number of hospitalized subjects show enrichment for a minor IFITM3 allele (SNP rs12252-C) that alters a splice acceptor site, and functional assays show the minor CC genotype IFITM3 has reduced influenza virus restriction in vitro. Together these data reveal that the action of a single intrinsic immune effector, IFITM3, profoundly alters the course of influenza virus infection in mouse and humans.

Nectin-like 4 Complexes with Choline Transporter-like Protein-1 and Regulates Schwann Cell Choline Homeostasis and Lipid Biogenesis in Vitro*

PubMed Central

Heffernan, Corey; Jain, Mohit R.; Liu, Tong; Kim, Hyosung; Barretto, Kevin; Li, Hong; Maurel, Patrice

2017-01-01

Nectin-like 4 (NECL4, CADM4) is a Schwann cell-specific cell adhesion molecule that promotes axo-glial interactions. In vitro and in vivo studies have shown that NECL4 is necessary for proper peripheral nerve myelination. However, the molecular mechanisms that are regulated by NECL4 and affect peripheral myelination currently remain unclear. We used an in vitro approach to begin identifying some of the mechanisms that could explain NECL4 function. Using mass spectrometry and Western blotting techniques, we have identified choline transporter-like 1 (CTL1) as a putative complexing partner with NECL4. We show that intracellular choline levels are significantly elevated in NECL4-deficient Schwann cells. The analysis of extracellular d9-choline uptake revealed a deficit in the amount of d9-choline found inside NECL4-deficient Schwann cells, suggestive of either reduced transport capabilities or increased metabolization of transported choline. An extensive lipidomic screen of choline derivatives showed that total phosphatidylcholine and phosphatidylinositol (but not diacylglycerol or sphingomyelin) are significantly elevated in NECL4-deficient Schwann cells, particularly specific subspecies of phosphatidylcholine carrying very long polyunsaturated fatty acid chains. Finally, CTL1-deficient Schwann cells are significantly impaired in their ability to myelinate neurites in vitro. To our knowledge, this is the first demonstration of a bona fide cell adhesion molecule, NECL4, regulating choline homeostasis and lipid biogenesis. Phosphatidylcholines are major myelin phospholipids, and several phosphorylated phosphatidylinositol species are known to regulate key aspects of peripheral myelination. Furthermore, the biophysical properties imparted to plasma membranes are regulated by fatty acid chain profiles. Therefore, it will be important to translate these in vitro observations to in vivo studies of NECL4 and CTL1-deficient mice. PMID:28119456

Establishment of 3D Co-Culture Models from Different Stages of Human Tongue Tumorigenesis: Utility in Understanding Neoplastic Progression.

PubMed

Sawant, Sharada; Dongre, Harsh; Singh, Archana Kumari; Joshi, Shriya; Costea, Daniela Elena; Mahadik, Snehal; Ahire, Chetan; Makani, Vidhi; Dange, Prerana; Sharma, Shilpi; Chaukar, Devendra; Vaidya, Milind

2016-01-01

To study multistep tumorigenesis process, there is a need of in-vitro 3D model simulating in-vivo tissue. Present study aimed to reconstitute in-vitro tissue models comprising various stages of neoplastic progression of tongue tumorigenesis and to evaluate the utility of these models to investigate the role of stromal fibroblasts in maintenance of desmosomal anchoring junctions using transmission electron microscopy. We reconstituted in-vitro models representing normal, dysplastic, and malignant tissues by seeding primary keratinocytes on either fibroblast embedded in collagen matrix or plain collagen matrix in growth factor-free medium. The findings of histomorphometry, immunohistochemistry, and electron microscopy analyses of the three types of 3D cultures showed that the stratified growth, cell proliferation, and differentiation were comparable between co-cultures and their respective native tissues; however, they largely differed in cultures grown without fibroblasts. The immunostaining intensity of proteins, viz., desmoplakin, desmoglein, and plakoglobin, was reduced as the disease stage increased in all co-cultures as observed in respective native tissues. Desmosome-like structures were identified using immunogold labeling in these cultures. Moreover, electron microscopic observations revealed that the desmosome number and their length were significantly reduced and intercellular spaces were increased in cultures grown without fibroblasts when compared with their co-culture counterparts. Our results showed that the major steps of tongue tumorigenesis can be reproduced in-vitro. Stromal fibroblasts play a role in regulation of epithelial thickness, cell proliferation, differentiation, and maintenance of desmosomalanchoring junctions in in-vitro grown tissues. The reconstituted co-culture models could help to answer various biological questions especially related to tongue tumorigenesis.

Prediction of pharmacokinetic and toxicological parameters of a 4-phenylcoumarin isolated from geopropolis: In silico and in vitro approaches.

PubMed

da Cunha, Marcos Guilherme; Franco, Gilson César Nobre; Franchin, Marcelo; Beutler, John A; de Alencar, Severino Matias; Ikegaki, Masaharu; Rosalen, Pedro Luiz

2016-11-30

In silico and in vitro methodologies have been used as important tools in the drug discovery process, including from natural sources. The aim of this study was to predict pharmacokinetic and toxicity (ADME/Tox) properties of a coumarin isolated from geopropolis using in silico and in vitro approaches. Cinnamoyloxy-mammeisin (CNM) isolated from Brazilian M. scutellaris geopropolis was evaluated for its pharmacokinetic parameters by in silico models (ACD/Percepta™ and MetaDrug™ software). Genotoxicity was assessed by in vitro DNA damage signaling PCR array. CNM did not pass all parameters of Lipinski's rule of five, with a predicted low oral bioavailability and high plasma protein binding, but with good predicted blood brain barrier penetration. CNM was predicted to show low affinity to cytochrome P450 family members. Furthermore, the predicted Ames test indicated potential mutagenicity of CNM. Also, the probability of toxicity for organs and tissues was classified as moderate and high for liver and kidney, and moderate and low for skin and eye irritation, respectively. The PCR array analysis showed that CNM significantly upregulated about 7% of all DNA damage-related genes. By exploring the biological function of these genes, it was found that the predicted CNM genotoxicity is likely to be mediated by apoptosis. The predicted ADME/Tox profile suggests that external use of CNM may be preferable to systemic exposure, while its genotoxicity was characterized by the upregulation of apoptosis-related genes after treatment. The combined use of in silico and in vitro approaches to evaluate these parameters generated useful hypotheses to guide further preclinical studies. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Establishment of 3D Co-Culture Models from Different Stages of Human Tongue Tumorigenesis: Utility in Understanding Neoplastic Progression

PubMed Central

Sawant, Sharada; Dongre, Harsh; Singh, Archana Kumari; Joshi, Shriya; Costea, Daniela Elena; Mahadik, Snehal; Ahire, Chetan; Makani, Vidhi; Dange, Prerana; Sharma, Shilpi; Chaukar, Devendra; Vaidya, Milind

2016-01-01

To study multistep tumorigenesis process, there is a need of in-vitro 3D model simulating in-vivo tissue. Present study aimed to reconstitute in-vitro tissue models comprising various stages of neoplastic progression of tongue tumorigenesis and to evaluate the utility of these models to investigate the role of stromal fibroblasts in maintenance of desmosomal anchoring junctions using transmission electron microscopy. We reconstituted in-vitro models representing normal, dysplastic, and malignant tissues by seeding primary keratinocytes on either fibroblast embedded in collagen matrix or plain collagen matrix in growth factor-free medium. The findings of histomorphometry, immunohistochemistry, and electron microscopy analyses of the three types of 3D cultures showed that the stratified growth, cell proliferation, and differentiation were comparable between co-cultures and their respective native tissues; however, they largely differed in cultures grown without fibroblasts. The immunostaining intensity of proteins, viz., desmoplakin, desmoglein, and plakoglobin, was reduced as the disease stage increased in all co-cultures as observed in respective native tissues. Desmosome-like structures were identified using immunogold labeling in these cultures. Moreover, electron microscopic observations revealed that the desmosome number and their length were significantly reduced and intercellular spaces were increased in cultures grown without fibroblasts when compared with their co-culture counterparts. Our results showed that the major steps of tongue tumorigenesis can be reproduced in-vitro. Stromal fibroblasts play a role in regulation of epithelial thickness, cell proliferation, differentiation, and maintenance of desmosomalanchoring junctions in in-vitro grown tissues. The reconstituted co-culture models could help to answer various biological questions especially related to tongue tumorigenesis. PMID:27501241

Simian virus 40 major late promoter: an upstream DNA sequence required for efficient in vitro transcription.

PubMed Central

Brady, J; Radonovich, M; Thoren, M; Das, G; Salzman, N P

1984-01-01

We have previously identified an 11-base DNA sequence, 5'-G-G-T-A-C-C-T-A-A-C-C-3' (simian virus 40 [SV40] map position 294 to 304), which is important in the control of SV40 late RNA expression in vitro and in vivo (Brady et al., Cell 31:625-633, 1982). We report here the identification of another domain of the SV40 late promoter. A series of mutants with deletions extending from SV40 map position 0 to 300 was prepared by nuclease BAL 31 treatment. The cloned templates were then analyzed for efficiency and accuracy of late SV40 RNA expression in the Manley in vitro transcription system. Our studies showed that, in addition to the promoter domain near map position 300, there are essential DNA sequences between nucleotide positions 74 and 95 that are required for efficient expression of late SV40 RNA. Included in this SV40 DNA sequence were two of the six GGGCGG SV40 repeat sequences and an 11-nucleotide segment which showed strong homology with the upstream sequences required for the efficient in vitro and in vivo expression of the histone H2A gene. This upstream promoter sequence supported transcription with the same efficiency even when it was moved 72 nucleotides closer to the major late cap site. In vitro promoter competition analysis demonstrated that the upstream promoter sequence, independent of the 294 to 304 promoter element, is capable of binding polymerase-transcription factors required for SV40 late gene transcription. Finally, we show that DNA sequences which control the specificity of RNA initiation at nucleotide 325 lie downstream of map position 294. Images PMID:6321950

Genotoxicity evaluation of carbon monoxide and 1, 3-butadiene using a new joint technology - the in vitro γH2AX HCS assay combined with air-liquid interface system.

PubMed

Zhang, Sen; Chen, Huan; Wang, An; Liu, Yong; Hou, Hongwei; Hu, Qingyuan

2018-05-21

To investigate the genotoxicity of gaseous toxicants CO and 1,3-butadiene in vitro, a novel combination technology-the in vitro γH2AX high content screening assay combined with air-liquid interface system was established. The results showed that this new technology was available and effective. Based on the joint technology, genotoxicity of CO and 1,3-butadiene was evaluated further in this study. The results showed that treatment concentrations (0, 20%,40%, 80% and 100%, v/v) and exposure time (15, 30, 45, 60 and 90 min) of CO both had no statistically significant effects on the induction of γH2AX (p > 0.05). However, 1,3-butadiene can induce significant γH2AX (p < 0.01) in A549 cells in a dose/time-dependent manner both in the absence and presence of rat liver S9. When the concentrations of 1,3-butadiene were more than 80%, a higher γH2AX level could be induced than the 1.5-fold of vehicle controls after 1 h of treatment. Overall, this new technology can be used a complementary tool to evaluate the genotoxicity of airborne toxicants in vitro based on the in vitro γH2AX high content screening assay combined with air-liquid interface system. Based on the joint technology, CO was not genotoxic in A549 cells, while 1,3-butadiene showed significant genotoxicity in the dose/time-dependency on the induction of γH2AX.

Evaluation of Anthelmintic Activity and Composition of Pumpkin (Cucurbita pepo L.) Seed Extracts—In Vitro and in Vivo Studies

PubMed Central

Grzybek, Maciej; Kukula-Koch, Wirginia; Strachecka, Aneta; Jaworska, Aleksandra; Phiri, Andrew M.; Paleolog, Jerzy; Tomczuk, Krzysztof

2016-01-01

A significant number of studies report growing resistance in nematodes thriving in both humans and livestock. This study was conducted to evaluate the in vitro and in vivo anthelmintic efficiency of Curcubita pepo (C. pepo) L. hot water extract (HWE), cold water extract (CWE) or ethanol extract (ETE) on two model nematodes: Caenorhabditis elegans (C. elegans) and Heligmosoides bakeri (H. bakeri). Methods: Raman, IR and LC-MS spectroscopy analyses were performed on the studied plant material to deliver qualitative and quantitative data on the composition of the obtained extracts: ETE, HWE and CWE. The in vitro activity evaluation showed an impact of C. pepo extracts on C. elegans and different developmental stages of H. bakeri. The following in vivo experiments on mice infected with H. bakeri confirmed inhibitory properties of the most active pumpkin extract selected by the in vitro study. All of the extracts were found to contain cucurbitine, aminoacids, fatty acids, and-for the first time-berberine and palmatine were identified. All C. pepo seed extracts exhibited a nematidicidal potential in vitro, affecting the survival of L1 and L2 H. bakeri larvae. The ETE was the strongest and demonstrated a positive effect on H. bakeri eggs hatching and marked inhibitory properties against worm motility, compared to a PBS control. No significant effects of pumpkin seed extracts on C. elegans integrity or motility were found. The EtOH extract in the in vivo studies showed anthelmintic properties against both H. bakeri fecal egg counts and adult worm burdens. The highest egg counts reduction was observed for the 8 g/kg dose (IC50 against H. bakeri = 2.43; 95% Cl = 2.01–2.94). A decrease in faecal egg counts (FEC) was accompanied by a significant reduction in worm burden of the treated mice compared to the control group. Conclusions: Pumpkin seed extracts may be used to control of Gastrointestinal (G.I.) nematode infections. This relatively inexpensive alternative to the currently available chemotherapeutic should be considered as a novel drug candidate in the nearest future. PMID:27598135

Evaluation of Anthelmintic Activity and Composition of Pumpkin (Cucurbita pepo L.) Seed Extracts-In Vitro and in Vivo Studies.

PubMed

Grzybek, Maciej; Kukula-Koch, Wirginia; Strachecka, Aneta; Jaworska, Aleksandra; Phiri, Andrew M; Paleolog, Jerzy; Tomczuk, Krzysztof

2016-09-01

A significant number of studies report growing resistance in nematodes thriving in both humans and livestock. This study was conducted to evaluate the in vitro and in vivo anthelmintic efficiency of Curcubita pepo (C. pepo) L. hot water extract (HWE), cold water extract (CWE) or ethanol extract (ETE) on two model nematodes: Caenorhabditis elegans (C. elegans) and Heligmosoides bakeri (H. bakeri). Raman, IR and LC-MS spectroscopy analyses were performed on the studied plant material to deliver qualitative and quantitative data on the composition of the obtained extracts: ETE, HWE and CWE. The in vitro activity evaluation showed an impact of C. pepo extracts on C. elegans and different developmental stages of H. bakeri. The following in vivo experiments on mice infected with H. bakeri confirmed inhibitory properties of the most active pumpkin extract selected by the in vitro study. All of the extracts were found to contain cucurbitine, aminoacids, fatty acids, and-for the first time-berberine and palmatine were identified. All C. pepo seed extracts exhibited a nematidicidal potential in vitro, affecting the survival of L1 and L2 H. bakeri larvae. The ETE was the strongest and demonstrated a positive effect on H. bakeri eggs hatching and marked inhibitory properties against worm motility, compared to a PBS control. No significant effects of pumpkin seed extracts on C. elegans integrity or motility were found. The EtOH extract in the in vivo studies showed anthelmintic properties against both H. bakeri fecal egg counts and adult worm burdens. The highest egg counts reduction was observed for the 8 g/kg dose (IC50 against H. bakeri = 2.43; 95% Cl = 2.01-2.94). A decrease in faecal egg counts (FEC) was accompanied by a significant reduction in worm burden of the treated mice compared to the control group. Pumpkin seed extracts may be used to control of Gastrointestinal (G.I.) nematode infections. This relatively inexpensive alternative to the currently available chemotherapeutic should be considered as a novel drug candidate in the nearest future.

In vitro effectiveness of anidulafungin against Candida sp. biofilms.

PubMed

Rosato, Antonio; Piarulli, Monica; Schiavone, Brigida Pia Immacolata; Catalano, Alessia; Carocci, Alessia; Carrieri, Antonio; Carone, Addolorata; Caggiano, Giuseppina; Franchini, Carlo; Corbo, Filomena; Montagna, Maria Teresa

2013-12-01

This study furnishes deeper insights to previous works on anidulafungin, demonstrating the potent activity against Candida strains planktonic cells and biofilms. Candida sp., associated with many biomaterial-related infections, give rise to infective pathologies typically associated with biofilm formation. We recently determined the in vitro antifungal activities of echinocandin anidulafungin in association with some antifungal drugs against some Candida strains in their planktonic states. A total of 11 Candida strains biofilms were tested in this study: six Candida albicans, three C. parapsilosis and two C. tropicalis. All yeast isolates and ATCC strains were stored at -20°C in glycerol stocks and were subcultured on antimicrobial agent-free Sabouraud dextrose agar plates. MIC endpoints were determined colorimetrically by using the indicator 2,3-bis(2-methoxy-4-nitro-5-sulphophenyl)-5-[(phenylamino)carbonyl]-2H-tetrazolium hydroxide (XTT) with menadione as electron-coupling agent. The activity of anidulafungin was assessed using in vitro microbiological model relevant for clinical practice. Anidulafungin showed a strong activity in vitro against both planktonic and biofilms cells, and our study confirms that high anidulafungin concentrations might establish paradoxical growth effect in C. albicans and C. tropicalis biofilms.

Development, characterization, and in vitro and in vivo evaluation of benzocaine- and lidocaine-loaded nanostructrured lipid carriers.

PubMed

Puglia, Carmelo; Sarpietro, Maria Grazia; Bonina, Francesco; Castelli, Francesco; Zammataro, Magda; Chiechio, Santina

2011-05-01

The present study concerns the in vitro and in vivo evaluation of benzocaine (BENZO) and lidocaine (LIDO) topical delivery from nanostructured lipid carriers (NLCs). Morphology and dimensional distribution of NLCs have been, respectively, characterized by differential scanning calorimetry (DSC) and photon correlation spectroscopy. The release pattern of BENZO and LIDO from NLCs was evaluated in vitro determining drug percutaneous absorption through excised human skin. Radiant heat tail-flick test was carried out in mice to determine the antinociceptive effect of BENZO and LIDO from NLC. DSC studies revealed that the inner oil phase of NLC plays a significant role in stabilizing the particle architecture and increasing the drug solubility. In vitro evidences show that BENZO and LIDO, when incorporated in viscosized NLC dispersions, exhibited a lower flux with respect to formulations containing the free drugs in the aqueous phase. In vivo study enabled to demonstrate that BENZO and LIDO can be released in a prolonged fashion when incorporated into lipid carriers. The results obtained pointed out NLC capability to act as an effective drug reservoir, thus prolonging the anesthetic effect of BENZO and LIDO. Copyright © 2010 Wiley-Liss, Inc.

Inflammatory response study of gellan gum impregnated duck's feet derived collagen sponges.

PubMed

Song, Jeong Eun; Lee, Seon Eui; Cha, Se Rom; Jang, Na Keum; Tripathy, Nirmalya; Reis, Rui L; Khang, Gilson

2016-10-01

Tissue engineered biomaterials have biodegradable and biocompatible properties. In this study, we have fabricated sponges using duck's feet derived collagen (DC) and gellan gum (GG), and further studied its inflammatory responses. The as-prepared duck's feet DC/GG sponges showed the possibility of application as a tissue engineering material through in vitro and in vivo experiments. The physical and chemical properties of sponges were characterized by compression strength, porosity, and scanning electron microscopy, etc. In vitro cell viability were investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assay. An inflammatory response was studied after seeding RAW264.7 cells on as-fabricated sponges using reverse transcriptase-polymerase chain reaction. In vivo studies were carried out by implanting in subcutaneous nude mouse followed by extraction, histological staining. Collectively, superior results were showed by DC/GG sponges than GG sponge in terms of physical property and cell proliferation and thus can be considered as a potential candidate for future tissue engineering applications.

Effects of P-MAPA Immunomodulator on Toll-Like Receptors and p53: Potential Therapeutic Strategies for Infectious Diseases and Cancer

PubMed Central

2012-01-01

Background Compounds that can act as agonists for toll-like receptors (TLRs) may be promising candidates for the development of drugs against infectious diseases and cancer. The present study aimed to characterize the immunomodulatory effects of P-MAPA on TLRs in vitro and in vivo, as well as to investigate its potential as adjuvant therapy in infectious diseases and cancer. Methods For these purposes, the activity of P-MAPA on TLRs was assayed in vitro through NF-κB activation in HEK293 cells expressing a given TLR, and using an in vivo animal model for bladder cancer (BC). The antimicrobial activity of P-MAPA was tested against Mycobacterium tuberculosis (TB) in vitro in an MIC assay, and in vivo using an aerosol infection model of murine tuberculosis. Antitumor effects of P-MAPA were tested in an animal model with experimentally induced BC. Moxifloxacin (MXF) and Bacillus Calmette-Guerin (BCG) were used as positive controls in the animal models. Results The results showed that P-MAPA, administered alone or in combination with MXF, induced significant responses in vivo against TB. In contrast, the compound did not show antimicrobial activity in vitro. P-MAPA showed a significant stimulatory effect on human TLR2 and TLR4 in vitro. In BC, TLR2, TLR4 and p53 protein levels were significantly higher in the P-MAPA group than in the BCG group. The most common histopathological changes in each group were papillary carcinoma in BC group, low-grade intraepithelial neoplasia in BCG group and simple hyperplasia in P-MAPA group. Concerning the toxicological analysis performed during BC treatment, P-MAPA did not show evidence for hepatotoxicity and nephrotoxicity. Conclusions In conclusion, P-MAPA acted as TLR ligand in vitro and improved the immunological status in BC, increasing TLR2 and TLR4 protein levels. P-MAPA immunotherapy was more effective in restoring p53 and TLRs reactivities and showed significantly greater antitumor activity than BCG. The activation of TLRs and p53 may provide a hypothetical mechanism for the therapeutic effects in both cancer and infectious diseases. Taken together data obtained will encourage the further investigation of P-MAPA as a potential candidate for the treatment of cancer and infectious diseases. PMID:22709446

Effects of P-MAPA Immunomodulator on Toll-Like Receptors and p53: Potential Therapeutic Strategies for Infectious Diseases and Cancer.

PubMed

Fávaro, Wagner J; Nunes, Odilon S; Seiva, Fabio Rf; Nunes, Iseu S; Woolhiser, Lisa K; Durán, Nelson; Lenaerts, Anne J

2012-06-18

Compounds that can act as agonists for toll-like receptors (TLRs) may be promising candidates for the development of drugs against infectious diseases and cancer. The present study aimed to characterize the immunomodulatory effects of P-MAPA on TLRs in vitro and in vivo, as well as to investigate its potential as adjuvant therapy in infectious diseases and cancer. For these purposes, the activity of P-MAPA on TLRs was assayed in vitro through NF-κB activation in HEK293 cells expressing a given TLR, and using an in vivo animal model for bladder cancer (BC). The antimicrobial activity of P-MAPA was tested against Mycobacterium tuberculosis (TB) in vitro in an MIC assay, and in vivo using an aerosol infection model of murine tuberculosis. Antitumor effects of P-MAPA were tested in an animal model with experimentally induced BC. Moxifloxacin (MXF) and Bacillus Calmette-Guerin (BCG) were used as positive controls in the animal models. The results showed that P-MAPA, administered alone or in combination with MXF, induced significant responses in vivo against TB. In contrast, the compound did not show antimicrobial activity in vitro. P-MAPA showed a significant stimulatory effect on human TLR2 and TLR4 in vitro. In BC, TLR2, TLR4 and p53 protein levels were significantly higher in the P-MAPA group than in the BCG group. The most common histopathological changes in each group were papillary carcinoma in BC group, low-grade intraepithelial neoplasia in BCG group and simple hyperplasia in P-MAPA group. Concerning the toxicological analysis performed during BC treatment, P-MAPA did not show evidence for hepatotoxicity and nephrotoxicity. In conclusion, P-MAPA acted as TLR ligand in vitro and improved the immunological status in BC, increasing TLR2 and TLR4 protein levels. P-MAPA immunotherapy was more effective in restoring p53 and TLRs reactivities and showed significantly greater antitumor activity than BCG. The activation of TLRs and p53 may provide a hypothetical mechanism for the therapeutic effects in both cancer and infectious diseases. Taken together data obtained will encourage the further investigation of P-MAPA as a potential candidate for the treatment of cancer and infectious diseases.

In vitro antiplasmodial and cytotoxic properties of some medicinal plants from western Burkina Faso.

PubMed

Sanon, Souleymane; Gansane, Adama; Ouattara, Lamoussa P; Traore, Abdoulaye; Ouedraogo, Issa N; Tiono, Alfred; Taramelli, Donatella; Basilico, Nicoletta; Sirima, Sodiomon B

2013-01-01

Resistance of malaria parasites to existing drugs complicates treatment, but an antimalarial vaccine that could protect against this disease is not yet available. It is therefore necessary to find new effective and affordable medicines. Medicinal plants could be a potential source of antimalarial agents. Some medicinal plants from Burkina Faso were evaluated for their antiplasmodial and cytotoxic properties in vitro . Crude dichloromethane, methanol, water-methanol, aqueous and alkaloids extracts were prepared for 12 parts of 10 plants. Chloroquine-resistant malaria strain K1 was used for the in vitro sensibility assay. The Plasmodium lactacte dehydrogenase technique was used to determine the 50% inhibitory concentration of parasites activity (IC 50 ). The cytotoxic effects were determined with HepG2 cells, using the tetrazolium-based colorimetric technique, and the selectivity index (SI) was calculated. Sixty crude extracts were prepared. Seven extracts from Terminalia avicenoides showed IC 50 < 5 µg/mL. The IC 50 of dichloromethane, methanol, aqueous and alkaloids extracts ranged between 1.6 µg/mL and 4.5 µg/mL. Three crude extracts from Combretum collinum and three from Ficus capraefolia had an IC 50 ranging between 0.2 µg/mL and 2.5 µg/mL. Crude extracts from these three plants had no cytotoxic effect, with SI > 1. The other plants have mostly moderate or no antimalarial effects. Some extracts from Cordia myxa , Ficus capraefolia and Opilia celtidifolia showed cytotoxicity, with an SI ranging between 0.4 and 0.9. Our study showed a good antiplasmodial in vitro activity of Terminalia avicenoides, Combretum collinum and Ficus capraefolia . These three plants may contain antiplasmodial molecules that could be isolated by bio-guided phytochemical studies.

In vitro antiplasmodial and cytotoxic properties of some medicinal plants from western Burkina Faso

PubMed Central

Gansane, Adama; Ouattara, Lamoussa P.; Traore, Abdoulaye; Ouedraogo, Issa N.; Tiono, Alfred; Taramelli, Donatella; Basilico, Nicoletta; Sirima, Sodiomon B.

2013-01-01

Background Resistance of malaria parasites to existing drugs complicates treatment, but an antimalarial vaccine that could protect against this disease is not yet available. It is therefore necessary to find new effective and affordable medicines. Medicinal plants could be a potential source of antimalarial agents. Some medicinal plants from Burkina Faso were evaluated for their antiplasmodial and cytotoxic properties in vitro. Methods Crude dichloromethane, methanol, water-methanol, aqueous and alkaloids extracts were prepared for 12 parts of 10 plants. Chloroquine-resistant malaria strain K1 was used for the in vitro sensibility assay. The Plasmodium lactacte dehydrogenase technique was used to determine the 50% inhibitory concentration of parasites activity (IC50). The cytotoxic effects were determined with HepG2 cells, using the tetrazolium-based colorimetric technique, and the selectivity index (SI) was calculated. Results Sixty crude extracts were prepared. Seven extracts from Terminalia avicenoides showed IC50 < 5 µg/mL. The IC50 of dichloromethane, methanol, aqueous and alkaloids extracts ranged between 1.6 µg/mL and 4.5 µg/mL. Three crude extracts from Combretum collinum and three from Ficus capraefolia had an IC50 ranging between 0.2 µg/mL and 2.5 µg/mL. Crude extracts from these three plants had no cytotoxic effect, with SI > 1. The other plants have mostly moderate or no antimalarial effects. Some extracts from Cordia myxa, Ficus capraefolia and Opilia celtidifolia showed cytotoxicity, with an SI ranging between 0.4 and 0.9. Conclusion Our study showed a good antiplasmodial in vitro activity of Terminalia avicenoides, Combretum collinum and Ficus capraefolia. These three plants may contain antiplasmodial molecules that could be isolated by bio-guided phytochemical studies. PMID:29043169

Microfluidic neurite guidance to study structure-function relationships in topologically-complex population-based neural networks.

PubMed

Honegger, Thibault; Thielen, Moritz I; Feizi, Soheil; Sanjana, Neville E; Voldman, Joel

2016-06-22

The central nervous system is a dense, layered, 3D interconnected network of populations of neurons, and thus recapitulating that complexity for in vitro CNS models requires methods that can create defined topologically-complex neuronal networks. Several three-dimensional patterning approaches have been developed but none have demonstrated the ability to control the connections between populations of neurons. Here we report a method using AC electrokinetic forces that can guide, accelerate, slow down and push up neurites in un-modified collagen scaffolds. We present a means to create in vitro neural networks of arbitrary complexity by using such forces to create 3D intersections of primary neuronal populations that are plated in a 2D plane. We report for the first time in vitro basic brain motifs that have been previously observed in vivo and show that their functional network is highly decorrelated to their structure. This platform can provide building blocks to reproduce in vitro the complexity of neural circuits and provide a minimalistic environment to study the structure-function relationship of the brain circuitry.

Effect of the Combination Hot Water - Calcium Chloride on the In Vitro Growth of Colletotrichum gloeosporioides and the Postharvest Quality of Infected Papaya

PubMed Central

Ayón-Reyna, Lidia Elena; López-Valenzuela, José Ángel; Delgado-Vargas, Francisco; López-López, Martha Edith; Molina-Corral, Francisco Javier; Carrillo-López, Armando; Vega-García, Misael Odín

2017-01-01

Anthracnose of papaya fruit caused by the fungus Colletotrichum gloeosporioides is one of the most economically important postharvest diseases. Hot water immersion (HW) and calcium chloride (Ca) treatments have been used to control papaya postharvest diseases; however, the effect of the combination HW-Ca on the pathogen growth and the development of the disease in infected papaya fruit has been scarcely studied. The aim of this study was to evaluate the effect of the HW-Ca treatment on the in vitro growth of C. gloesporioides conidia and the quality of infected papaya. In vitro, the HW-Ca treated conidia showed reduced mycelial growth and germination. In vivo, the HW-Ca treatment of infected papaya delayed for 5 days the onset of the anthracnose symptoms and improved the papaya postharvest quality. The combined treatment HW-Ca was better than any of the individual treatments to inhibit the in vitro development of C. gloeosporioides and to reduce the negative effects of papaya anthracnose. PMID:29238280

Exposure to cadmium during in vitro maturation at environmental nanomolar levels impairs oocyte fertilization through oxidative damage: A large animal model study.

PubMed

Martino, N A; Marzano, G; Mangiacotti, M; Miedico, O; Sardanelli, A M; Gnoni, A; Lacalandra, G M; Chiaravalle, A E; Ciani, E; Bogliolo, L; Minervini, F; Pizzi, F; Dell'Aquila, M E

2017-04-01

Cadmium is a highly toxic heavy metal with negative effects on oocyte fertilization. The aim of this study was to analyse whether cadmium-induced impairment of fertilization is caused by mitochondria dysfunction and oxidative stress in the cumulus-oocyte complex (COC). Preliminarily, 19 trace element levels were measured in ovaries from juvenile and adult ewes and age-related cadmium ovarian bioaccumulation at nanomolar concentrations was found. COCs from juvenile and adult ewes, exposed during in vitro maturation to 1nM or 100nM CdCl 2 , and subjected to in vitro fertilization showed significantly lower fertilization rates in exposed COCs compared with controls. In vitro matured exposed and control COCs underwent confocal microscopy analysis of mitochondria activity and reactive oxygen species (ROS) levels and lipid peroxidation (LPO) assay at cumulus cell and oocyte level. In both age groups, cadmium at nanomolar concentrations induced cumulus-oocyte mitochondria over-activity and oxidative damage which were related to impaired oocyte fertilization. Copyright © 2017 Elsevier Inc. All rights reserved.

Microfluidic neurite guidance to study structure-function relationships in topologically-complex population-based neural networks

NASA Astrophysics Data System (ADS)

Honegger, Thibault; Thielen, Moritz I.; Feizi, Soheil; Sanjana, Neville E.; Voldman, Joel

2016-06-01

The central nervous system is a dense, layered, 3D interconnected network of populations of neurons, and thus recapitulating that complexity for in vitro CNS models requires methods that can create defined topologically-complex neuronal networks. Several three-dimensional patterning approaches have been developed but none have demonstrated the ability to control the connections between populations of neurons. Here we report a method using AC electrokinetic forces that can guide, accelerate, slow down and push up neurites in un-modified collagen scaffolds. We present a means to create in vitro neural networks of arbitrary complexity by using such forces to create 3D intersections of primary neuronal populations that are plated in a 2D plane. We report for the first time in vitro basic brain motifs that have been previously observed in vivo and show that their functional network is highly decorrelated to their structure. This platform can provide building blocks to reproduce in vitro the complexity of neural circuits and provide a minimalistic environment to study the structure-function relationship of the brain circuitry.

In Vitro Activity of Rifampicin Combined with Daptomycin or Tigecycline on Staphylococcus haemolyticus Biofilms.

PubMed

Szczuka, Ewa; Grabska, Katarzyna; Kaznowski, Adam

2015-08-01

Staphylococcus haemolyticus is of increasing concern as a cause of several biofilm-associated infections, and today, it represents the second most common organism among clinical isolates of coagulase-negative staphylococci. However, little is known regarding the treatment of infections caused by these bacteria. In this study, we characterize the biofilm formed by S. haemolyticus strains isolated from bloodstream infections and assess in vitro the activity of rifampicin combined with daptomycin or tigecycline against bacteria growing in a biofilm. The results of our studies indicated that the majority (78 %) of methicillin-resistant Staphylococcus haemolyticus strains have the ability to form a biofilm in vitro. None of these strains carried icaADBC genes indicating that they form biofilm via ica-independent mechanisms. The molecular characterization of the biofilm showed that proteins are the predominant matrix component and play a major role in biofilm structure. Extracellular DNA and polysaccharides, other than polysaccharide intercellular adhesin, are also present in the biofilm matrix, but they play a minor role. The images obtained by confocal laser scanning microscopy showed that most S. haemolyticus strains formed a dense biofilm with a low number of dead cells. In vitro study demonstrated excellent activity of tigecycline in combination with rifampicin against cell growth in the proteinous biofilm. The BIC (biofilm inhibitory concentration) value for tigecycline/rifampicin ranged from 0.062 to 1 µg/ml, whereas for daptomycin/rifampicin from 0.125 to 2 µg/ml. These results indicated that the tigecycline/rifampicin combination was more effective against ica-independent biofilm, formed by S. haemolyticus strains, than the daptomycin/rifampicin combination.

Microarray analysis in rat liver slices correctly predicts in vivo hepatotoxicity.

PubMed

Elferink, M G L; Olinga, P; Draaisma, A L; Merema, M T; Bauerschmidt, S; Polman, J; Schoonen, W G; Groothuis, G M M

2008-06-15

The microarray technology, developed for the simultaneous analysis of a large number of genes, may be useful for the detection of toxicity in an early stage of the development of new drugs. The effect of different hepatotoxins was analyzed at the gene expression level in the rat liver both in vivo and in vitro. As in vitro model system the precision-cut liver slice model was used, in which all liver cell types are present in their natural architecture. This is important since drug-induced toxicity often is a multi-cellular process involving not only hepatocytes but also other cell types such as Kupffer and stellate cells. As model toxic compounds lipopolysaccharide (LPS, inducing inflammation), paracetamol (necrosis), carbon tetrachloride (CCl(4), fibrosis and necrosis) and gliotoxin (apoptosis) were used. The aim of this study was to validate the rat liver slice system as in vitro model system for drug-induced toxicity studies. The results of the microarray studies show that the in vitro profiles of gene expression cluster per compound and incubation time, and when analyzed in a commercial gene expression database, can predict the toxicity and pathology observed in vivo. Each toxic compound induces a specific pattern of gene expression changes. In addition, some common genes were up- or down-regulated with all toxic compounds. These data show that the rat liver slice system can be an appropriate tool for the prediction of multi-cellular liver toxicity. The same experiments and analyses are currently performed for the prediction of human specific toxicity using human liver slices.

Tissue compatibility and pharmacokinetics of three potential subcutaneous injectables for low-pH drug solutions.

PubMed

Wu, Zimei; Tucker, Ian G; Razzak, Majid; McSporran, Keith; Medlicott, Natalie J

2010-07-01

The aim of the study was to investigate the tissue tolerance and bioavailability of four formulations containing 5% ricobendazole solubilised at low pH, following subcutaneous injection in sheep. Formulations were: a water-in-oil emulsion, a microemulsion, a hydroxypropyl-beta-cyclodextrin (HP-beta-CD, 20%) drug solution, and a low-pH drug solution (reference). In-vitro cytotoxicity of the formulations was investigated in L929 fibroblasts using MTS viability and lactate dehydrogenase leakage assays. Each formulation and respective vehicle was injected into either side of the back of a sheep to investigate the tissue tolerance and pharmacokinetics. In-vitro studies suggested that both the emulsion and the microemulsion are unlikely to give a burst release of the low-pH drug solution in aqueous media. The microemulsion showed the greatest in-vitro cytotoxic effect but no significant difference was observed between the other formulations. In sheep, the three new formulations and vehicles caused little or no injection-site reactions compared with a marked response to the reference formulation. Bioavailabilities of HP-beta-CD formulation, emulsion and microemulsion formulations, relative to the reference formulation, were 194, 155 and 115%, respectively. The three new subcutaneous injectables showed promise for reducing irritation of low-pH solubilised ricobendazole. HP-beta-CD significantly enhanced the drug absorption. Controlling the burst release of the low-pH drug solution may improve tissue tolerance and minimise post-injection precipitation, and hence increase drug bioavailability. The in-vitro cytotoxicity studies did not predict the in-vivo irritation effects.

In vitro analysis of platelet function in acute aneurysmal subarachnoid haemorrhage.

PubMed

von der Brelie, Christian; Subai, Alexander; Limperger, Verena; Rohde, Veit; Dempfle, Astrid; Boström, Azize

2018-04-01

Platelet function might play an essential role in the pathogenesis of delayed cerebral ischemia (DCI) after aneurysmal subarachnoid haemorrhage (SAH). Thus, impaired platelet function and disturbed primary haemostasis induced by intake of acetylsalicylic acid (ASA) might influence the rate of DCI. Primary haemostasis and platelet function can be measured with in vitro diagnosis (platelet function analyser test, PFA 100). The aim of this study is to evaluate the rate of DCI, haemorrhagic complications and the neurological outcome. Two groups were compared (patients with regular platelet function versus patients with impaired platelet function). This is a retrospective observational study. An initial cohort of 787 patients with SAH has been treated from January 2005 to September 2012. Seventy-nine patients (10%) with aneurysmal SAH, a history of ASA medication and PFA testing within the first 24 h after aneurysm rupture have been included. The overall rate of DCI in the present study was 43%. In vitro platelet function testing showed pathological primary haemostasis in 69.6%. The DCI rate was higher in patients with regular tested primary haemostasis (p = 0.02, OR = 3.16, 95%CI = [1.19; 8.83]). However, outcome assessment by mGOS did not show a significant difference between the groups. Patients with impaired primary haemostasis did not display a higher rate of haemorrhagic complications. Impairment of primary haemostasis resulting from an impairment of platelet function at an early stage after SAH might lead to a lower rate of DCI. In vitro testing of platelet function might be useful to predict the occurrence of DCI in the course.

Gelatin nanoparticle-mediated intranasal delivery of substance P protects against 6-hydroxydopamine-induced apoptosis: an in vitro and in vivo study.

PubMed

Lu, Cui-Tao; Jin, Rong-Rong; Jiang, Yi-Na; Lin, Qian; Yu, Wen-Ze; Mao, Kai-Li; Tian, Fu-Rong; Zhao, Ya-Ping; Zhao, Ying-Zheng

2015-01-01

The aim of this study was to investigate the protective role of intranasally administered substance P-loaded gelatin nanoparticles (SP-GNPs) against 6-hydroxydopamine (6-OHDA)-induced apoptosis in vitro and in vivo, and to provide a new strategy for treating brain pathology, such as Parkinson's disease. SP-GNPs were prepared by a water-in-water emulsion method, and their stability, encapsulating efficiency, and loading capacity were evaluated. PC-12 cells were used to examine the enhancement of growth and inhibition of apoptosis by SP-GNPs in vitro using MTT assays. In the in vivo study, hemiparkinsonian rats were created by intracerebroventricular injection of 6-OHDA. The rats then received intranasal SP-GNPs daily for 2 weeks. Functional improvement was assessed by quantifying rotational behavior, and the degree of apoptosis was assessed by immunohistochemical staining for caspase-3 in the substantia nigra region. PC-12 cells with 6-OHDA-induced disease treated with SP-GNPs showed higher cell viability than their untreated counterparts, and cell viability increased as the concentration of substance P (SP) increased, indicating that SP could enhance cell growth and inhibit the cell apoptosis induced by 6-OHDA. Rats with 6-OHDA-induced hemiparkinsonism treated with SP-GNPs made fewer rotations and showed less staining for caspase-3 than their counterparts not treated with SP, indicating that SP protects rats with 6-OHDA-induced hemiparkinsonism from apoptosis and therefore demonstrates their functional improvement. Intranasal delivery of SP-GNPs protects against 6-OHDA-induced apoptosis both in vitro and in vivo.

Microarray analysis in rat liver slices correctly predicts in vivo hepatotoxicity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Elferink, M.G.L.; Olinga, P.; Draaisma, A.L.

2008-06-15

The microarray technology, developed for the simultaneous analysis of a large number of genes, may be useful for the detection of toxicity in an early stage of the development of new drugs. The effect of different hepatotoxins was analyzed at the gene expression level in the rat liver both in vivo and in vitro. As in vitro model system the precision-cut liver slice model was used, in which all liver cell types are present in their natural architecture. This is important since drug-induced toxicity often is a multi-cellular process involving not only hepatocytes but also other cell types such asmore » Kupffer and stellate cells. As model toxic compounds lipopolysaccharide (LPS, inducing inflammation), paracetamol (necrosis), carbon tetrachloride (CCl{sub 4}, fibrosis and necrosis) and gliotoxin (apoptosis) were used. The aim of this study was to validate the rat liver slice system as in vitro model system for drug-induced toxicity studies. The results of the microarray studies show that the in vitro profiles of gene expression cluster per compound and incubation time, and when analyzed in a commercial gene expression database, can predict the toxicity and pathology observed in vivo. Each toxic compound induces a specific pattern of gene expression changes. In addition, some common genes were up- or down-regulated with all toxic compounds. These data show that the rat liver slice system can be an appropriate tool for the prediction of multi-cellular liver toxicity. The same experiments and analyses are currently performed for the prediction of human specific toxicity using human liver slices.« less

Torsemide Fast Dissolving Tablets: Development, Optimization Using Box-Bhenken Design and Response Surface Methodology, In Vitro Characterization, and Pharmacokinetic Assessment.

PubMed

El-Shenawy, Ahmed A; Ahmed, Mahmoud M; Mansour, Heba F; Abd El Rasoul, Saleh

2017-08-01

The present study planed to develop new fast dissolving tablets (FDTs) of torsemide. Solid dispersions (SDs) of torsemide and sorbitol (3:1) or polyvinylpyrrolidone (PVP) k25 were prepared. The prepared SDs were evaluated for in-vitro dissolution. Fourier transform infrared spectroscopy and differential scanning calorimetry for SDs revealed no drug/excipient interactions and transformation of torsemide to the amorphous form. Torsemide/sorbitol SD was selected for formulation of torsemide FDTs by direct compression method. Box-Bhenken factorial design was employed to design 15 formulations using croscarmellose sodium and crospovidone at different concentrations. The response surface methodology was used to analyze the effect of changing these concentrations (independent variables) on disintegration time (Y 1 ), percentage friability (Y 2 ), and amount torsemide released at 10 min. The physical mixtures of torsemide and the used excipients were evaluated for angle of repose, Hausner's ratio, and Carr's index. The prepared FDTs tablets were evaluated for wetting and disintegration time, weight variation, drug content, percentage friability, thickness, hardness, and in vitro release. Based on the in-vitro results and factorial design characterization, F10 and F7 were selected for bioavailability studies following administration to Albino New Zealand rabbits. They showed significantly higher C max and (AUC 0-12 ) and shorter T max than those obtained after administration of the corresponding ordinary commercial Torseretic ® tablets. Stability study was conducted for F10 that showed good stability upon storage at 30°C/75% RH and 40°C/75% RH for 3 months.

Preparation & in vitro evaluation of 90Y-DOTA-rituximab

PubMed Central

Kameswaran, Mythili; Pandey, Usha; Dash, Ashutosh; Samuel, Grace; Venkatesh, Meera

2016-01-01

Background & objectives: Radioimmunotherapy is extensively being used for the treatment of non-Hodgkin's lymphoma (NHL). Use of rituximab, a chimeric anti-CD20 antibody directed against the CD20 antigen in combination with suitable beta emitters is expected to result in good treatment response by its cross-fire and bystander effects. The present work involves the conjugation of p-isothiocyanatobenzyl DOTA (p-SCN-Bn-DOTA) to rituximab, its radiolabelling with 90Y and in vitro and in vivo evaluation to determine its potential as a radioimmunotherapeutic agent. Methods: Rituximab was conjugated with p-SCN-Bn-DOTA at 1:1 antibody: DOTA molar ratio. The number of DOTA molecules linked to one molecule of rituximab was determined by radioassay and spectroscopic assay. Radiolabelling of rituximab with 90Y was carried out and its in vitro stability was evaluated. In vitro cell binding studies were carried out in Raji cells expressing CD20 antigen. Biodistribution studies were carried out in normal Swiss mice. Results: Using both radioassay and spectroscopic method, it was determined that about five molecules of DOTA were linked to rituximab. Radiolabelling of the rituximab conjugate with 90Y and subsequent purification on PD-10 column gave a product with radiochemical purity (RCP) > 98 per cent which was retained at > 90 per cent up to 72 h when stored at 37°C. In vitro cell binding experiments of 90Y-DOTA-rituximab with Raji cells exhibited specific binding of 20.7 ± 0.1 per cent with 90Y-DOTA-rituximab which reduced to 15.5 ± 0.2 per cent when incubated with cold rituximab. The equilibrium constant Kd for 90Y-DOTA-Rituximab was determined to be 3.38 nM. Radiolabelled antibody showed clearance via hepatobiliary and renal routes and activity in tibia was found to be quite low indicating in vivo stability of 90Y-DOTA-rituximab. Interpretation & conclusions: p-SCN-Bn-DOTA was conjugated with rituximab and radiolabelling with 90Y was carried out. In vitro studies carried out in Raji cells showed the specificity of the radiolabelled conjugate suggesting the potential uitability of the formulation as a radiopharmaceutical for therapy of NHL. PMID:26997015

Preparation & in vitro evaluation of ⁹⁰Y-DOTA-rituximab.

PubMed

Kameswaran, Mythili; Pandey, Usha; Dash, Ashutosh; Samuel, Grace; Venkatesh, Meera

2016-01-01

Radioimmunotherapy is extensively being used for the treatment of non-Hodgkin's lymphoma (NHL). Use of rituximab, a chimeric anti-CD20 antibody directed against the CD20 antigen in combination with suitable beta emitters is expected to result in good treatment response by its cross-fire and bystander effects. The present work involves the conjugation of p-isothiocyanatobenzyl DOTA (p-SCN-Bn-DOTA) to rituximab, its radiolabelling with [90] Y and in vitro and in vivo evaluation to determine its potential as a radioimmunotherapeutic agent. Rituximab was conjugated with p-SCN-Bn-DOTA at 1:1 antibody: DOTA molar ratio. The number of DOTA molecules linked to one molecule of rituximab was determined by radioassay and spectroscopic assay. Radiolabelling of rituximab with 90 Y was carried out and its in vitro stability was evaluated. In vitro cell binding studies were carried out in Raji cells expressing CD20 antigen. Biodistribution studies were carried out in normal Swiss mice. Using both radioassay and spectroscopic method, it was determined that about five molecules of DOTA were linked to rituximab. Radiolabelling of the rituximab conjugate with [90] Y and subsequent purification on PD-10 column gave a product with radiochemical purity (RCP) > 98 per cent which was retained at > 90 per cent up to 72 h when stored at 37°C. In vitro cell binding experiments of 90 Y-DOTA-rituximab with Raji cells exhibited specific binding of 20.7 ± 0.1 per cent with [90] Y-DOTA-rituximab which reduced to 15.5 ± 0.2 per cent when incubated with cold rituximab. The equilibrium constant K d for 90 Y-DOTA-Rituximab was determined to be 3.38 nM. Radiolabelled antibody showed clearance via hepatobiliary and renal routes and activity in tibia was found to be quite low indicating in vivo stability of [90] Y-DOTA-rituximab. p-SCN-Bn-DOTA was conjugated with rituximab and radiolabelling with 90 Y was carried out. In vitro studies carried out in Raji cells showed the specificity of the radiolabelled conjugate suggesting the potential uitability of the formulation as a radiopharmaceutical for therapy of NHL.

Torque Loss After Miniscrew Placement: An In-Vitro Study Followed by a Clinical Trial.

PubMed

Migliorati, Marco; Drago, Sara; Barberis, Fabrizio; Schiavetti, Irene; Dalessandri, Domenico; Benedicenti, Stefano; Biavati, Armando Silvestrini

2016-01-01

To evaluate torque loss a week after insertion, both in an in vivo and an in vitro experimental setup were designed. In the in vivo setup a total of 29 miniscrews were placed in 20 patients who underwent orthodontic treatment. Maximum insertion torque (MIT) was evaluated at insertion time (T1). A week later, insertion torque was measured again by applying a quarter turn (T2); no load was applied on the screw during the first week. In the in vitro setup a total of 20 miniscrews were placed in pig rib bone samples. MIT was evaluated at insertion time (T1). Bone samples were kept in saline solution and controlled environment for a week during which the solution was refreshed every day. Afterwards, torque was measured again by applying a quarter turn (T2). The comparison of MIT over time was done calculating the percentage difference of the torque values between pre- and post-treatment and using the parametric two independent samples t-test or the non-parametric Mann-Whitney test. After a week unloaded miniscrews showed a mean loss of rotational torque of 36.3% and 40.9% in in vitro and in in vivo conditions, respectively. No statistical differences were found between the two different setups. Torque loss was observed after the first week in both study models; in vitro experimental setup provided a reliable study model for studying torque variation during the first week after insertion.

Torque Loss After Miniscrew Placement: An In-Vitro Study Followed by a Clinical Trial

PubMed Central

Migliorati, Marco; Drago, Sara; Barberis, Fabrizio; Schiavetti, Irene; Dalessandri, Domenico; Benedicenti, Stefano; Biavati, Armando Silvestrini

2016-01-01

To evaluate torque loss a week after insertion, both in an in vivo and an in vitro experimental setup were designed. In the in vivo setup a total of 29 miniscrews were placed in 20 patients who underwent orthodontic treatment. Maximum insertion torque (MIT) was evaluated at insertion time (T1). A week later, insertion torque was measured again by applying a quarter turn (T2); no load was applied on the screw during the first week. In the in vitro setup a total of 20 miniscrews were placed in pig rib bone samples. MIT was evaluated at insertion time (T1). Bone samples were kept in saline solution and controlled environment for a week during which the solution was refreshed every day. Afterwards, torque was measured again by applying a quarter turn (T2). The comparison of MIT over time was done calculating the percentage difference of the torque values between pre- and post-treatment and using the parametric two independent samples t-test or the non-parametric Mann–Whitney test. After a week unloaded miniscrews showed a mean loss of rotational torque of 36.3% and 40.9% in in vitro and in in vivo conditions, respectively. No statistical differences were found between the two different setups. Torque loss was observed after the first week in both study models; in vitro experimental setup provided a reliable study model for studying torque variation during the first week after insertion. PMID:27386011

In vitro and in vivo evaluations of three computer-aided shade matching instruments.

PubMed

Yuan, Kun; Sun, Xiang; Wang, Fu; Wang, Hui; Chen, Ji-hua

2012-01-01

This study evaluated the accuracy and reliability of three computer-aided shade matching instruments (Shadepilot, VITA Easyshade, and ShadeEye NCC) using both in vitro and in vivo models. The in vitro model included the measurement of five VITA Classical shade guides. The in vivo model utilized three instruments to measure the central region of the labial surface of maxillary right central incisors of 85 people. The accuracy and reliability of the three instruments in these two evaluating models were calculated. Significant differences were observed in the accuracy of instruments both in vitro and in vivo. No significant differences were found in the reliability of instruments between and within the in vitro and the in vivo groups. VITA Easyshade was significantly different in accuracy between in vitro and in vivo models, while no significant difference was found for the other two instruments. Shadepilot was the only instrument tested in the present study that showed high accuracy and reliability both in vitro and in vivo. Significant differences were observed in the L*a*b* values of the 85 natural teeth measured using three instruments in the in vivo assessment. The pair-agreement rates of shade matching among the three instruments ranged from 37.7% to 48.2%, and the incidence of identical shade results shared by all three instruments was 25.9%. As different L*a*b* values and shade matching results were reported for the same tooth, a combination of the evaluated shade matching instruments and visual shade confirmation is recommended for clinical use.

Oral cancer radiotherapy affects enamel microhardness and associated indentation pattern morphology

PubMed Central

Seyedmahmoud, R.; Thiagarajan, G.; Gorski, J. P.; Reed Edwards, R.; McGuire, J. D.

2017-01-01

Objectives The aim of this study is to determine the effects of in vitro and in vivo high-dose radiotherapy on microhardness and associated indentation pattern morphology of enamel. Materials and methods The inner, middle, and outer microhardness of enamel was evaluated using three experimental groups: control (non-radiated); in vitro irradiated; in vivo irradiated. In vitro specimens were exposed to simulated radiotherapy, and in vivo specimens were extracted teeth from oral cancer patients previously treated with radiotherapy. Indentations were measured via SEM images to calculate microhardness values and to assess the mechanomorphological properties of enamel before and after radiotherapy. Results Middle and outer regions of enamel demonstrated a significant decrease in microhardness after in vitro and in vivo irradiation compared to the control group (p < 0.05). Two indentation patterns were observed: pattern A—presence of microcracks around indent periphery, which represents local dissipation of deformation energy; pattern B—clean, sharp indents. The percentage of clean microindentation patterns, compared to controls, was significantly higher following in vitro and in vivo irradiation in all enamel regions. The highest percentage of clean microindentations (65%) was observed in the in vivo irradiated group in the inner region of enamel near the dentin-enamel junction. Conclusions For the first time, this study shows that in vitro and in vivo irradiation alters enamel microhardness. Likewise, the indentation pattern differences suggest that enamel may become more brittle following in vitro and in vivo irradiation. Clinical relevance The mechanomorphological property changes of enamel following radiation may be a contributory component of pathologic enamel delamination following oral cancer radiotherapy. PMID:29151196

miRNA studies in in vitro and in vivo activated hepatic stellate cells

PubMed Central

Maubach, Gunter; Lim, Michelle Chin Chia; Chen, Jinmiao; Yang, Henry; Zhuo, Lang

2011-01-01

AIM: To understand which and how different miRNAs are implicated in the process of hepatic stellate cell (HSC) activation. METHODS: We used microarrays to examine the differential expression of miRNAs during in vitro activation of primary HSCs (pHSCs). The transcriptome changes upon stable transfection of rno-miR-146a into an HSC cell line were studied using cDNA microarrays. Selected differentially regulated miRNAs were investigated by quantitative real-time polymerase chain reaction during in vivo HSC activation. The effect of miRNA mimics and inhibitor on the in vitro activation of pHSCs was also evaluated. RESULTS: We found that 16 miRNAs were upregulated and 26 were downregulated significantly in 10-d in vitro activated pHSCs in comparison to quiescent pHSCs. Overexpression of rno-miR-146a was characterized by marked upregulation of tissue inhibitor of metalloproteinase-3, which is implicated in the regulation of tumor necrosis factor-α activity. Differences in the regulation of selected miRNAs were observed comparing in vitro and in vivo HSC activation. Treatment with miR-26a and 29a mimics, and miR-214 inhibitor during in vitro activation of pHSCs induced significant downregulation of collagen type I transcription. CONCLUSION: Our results emphasize the different regulation of miRNAs in in vitro and in vivo activated pHSCs. We also showed that miR-26a, 29a and 214 are involved in the regulation of collagen type I mRNA. PMID:21734783

Comparison of MeHg-induced toxicogenomic responses across in vivo and in vitro models used in developmental toxicology.

PubMed

Robinson, Joshua F; Theunissen, Peter T; van Dartel, Dorien A M; Pennings, Jeroen L; Faustman, Elaine M; Piersma, Aldert H

2011-09-01

Toxicogenomic evaluations may improve toxicity prediction of in vitro-based developmental models, such as whole embryo culture (WEC) and embryonic stem cells (ESC), by providing a robust mechanistic marker which can be linked with responses associated with developmental toxicity in vivo. While promising in theory, toxicogenomic comparisons between in vivo and in vitro models are complex due to inherent differences in model characteristics and experimental design. Determining factors which influence these global comparisons are critical in the identification of reliable mechanistic-based markers of developmental toxicity. In this study, we compared available toxicogenomic data assessing the impact of the known teratogen, methylmercury (MeHg) across a diverse set of in vitro and in vivo models to investigate the impact of experimental variables (i.e. model, dose, time) on our comparative assessments. We evaluated common and unique aspects at both the functional (Gene Ontology) and gene level of MeHg-induced response. At the functional level, we observed stronger similarity in MeHg-response between mouse embryos exposed in utero (2 studies), ESC, and WEC as compared to liver, brain and mouse embryonic fibroblast MeHg studies. These findings were strongly correlated to the presence of a MeHg-induced developmentally related gene signature. In addition, we identified specific MeHg-induced gene expression alterations associated with developmental signaling and heart development across WEC, ESC and in vivo systems. However, the significance of overlap between studies was highly dependent on traditional experimental variables (i.e. dose, time). In summary, we identify promising examples of unique gene expression responses which show in vitro-in vivo similarities supporting the relevance of in vitro developmental models for predicting in vivo developmental toxicity. Copyright © 2011 Elsevier Inc. All rights reserved.

Synthesis, structure, theoretical and experimental in vitro antioxidant/pharmacological properties of α-aryl, N-alkyl nitrones, as potential agents for the treatment of cerebral ischemia.

PubMed

Samadi, Abdelouahid; Soriano, Elena; Revuelta, Julia; Valderas, Carolina; Chioua, Mourad; Garrido, Ignacio; Bartolomé, Begoña; Tomassolli, Isabelle; Ismaili, Lhassane; González-Lafuente, Laura; Villarroya, Mercedes; García, Antonio G; Oset-Gasque, María J; Marco-Contelles, José

2011-01-15

The synthesis, structure, theoretical and experimental in vitro antioxidant properties using the DPPH, ORAC, and benzoic acid, as well as preliminary in vitro pharmacological activities of (Z)-α-aryl and heteroaryl N-alkyl-nitrones 6-15, 18, 19, 21, and 23, is reported. In the in vitro antioxidant activity, for the DPPH radical test, only nitrones bearing free phenol groups gave the best RSA (%) values, nitrones 13 and 14 showing the highest values in this assay. In the ORAC analysis, the most potent radical scavenger was nitrone indole 21, followed by the N-benzyl benzene-type nitrones 10 and 15. Interestingly enough, the archetypal nitrone 7 (PBN) gave a low RSA value (1.4%) in the DPPH test, or was inactive in the ORAC assay. Concerning the ability to scavenge the hydroxyl radical, all the nitrones studied proved active in this experiment, showing high values in the 94-97% range, the most potent being nitrone 14. The theoretical calculations for the prediction of the antioxidant power, and the potential of ionization confirm that nitrones 9 and 10 are among the best compounds in electron transfer processes, a result that is also in good agreement with the experimental values in the DPPH assay. The calculated energy values for the reaction of ROS (hydroxyl, peroxyl) with the nitrones predict that the most favourable adduct-spin will take place between nitrones 9, 10, and 21, a fact that would be in agreement with their experimentally observed scavenger ability. The in vitro pharmacological analysis showed that the neuroprotective profile of the target molecules was in general low, with values ranging from 0% to 18.7%, in human neuroblastoma cells stressed with a mixture of rotenone/oligomycin-A, being nitrones 18, and 6-8 the most potent, as they show values in the range 24-18.4%. Crown Copyright © 2010. Published by Elsevier Ltd. All rights reserved.

Sustained Release of Lidocaine from Solvent-Free Biodegradable Poly[(d,l)-Lactide-co-Glycolide] (PLGA): In Vitro and In Vivo Study.

PubMed

Kau, Yi-Chuan; Liao, Chia-Chih; Chen, Ying-Chi; Liu, Shih-Jung

2014-09-16

Local anesthetics are commonly used for pain relief by regional nerve blocking. In this study, we fabricated solvent-free biodegradable pellets to extend the duration of lidocaine release without any significant local or systemic toxicity levels. To manufacture the pellets, poly[(d,l)-lactide-co-glycolide] (PLGA) was first pre-mixed with lidocaine powder into different ratios. The powder mixture was then compressed with a mold (diameter of 1, 5, 8 or 10 mm) and sintered at 65 °C to form pellets. The in vitro release study showed that the lidocaine/PLGA pellets exhibited a tri-phase release behavior (a burst, a diffusion-controlled release and a degradation-dominated release) and reached completion around day 28. Scanning electron microscope (SEM) photos show that small channels could be found on the surfaces of the pellets on day 2. Furthermore, the polymer matrix swelled and fell apart on day 7, while the pellets became viscous after 10 days of in vitro elution. Perineural administration of the lidocaine/PLGA pellets produced anti-hypersensitivity effects lasting for at least 24 h in rats, significant when compared to the control group (a pure PLGA was pellet administered). In addition, no inflammation was detected within the nerve and in the neighboring muscle by histopathology.

Significant blockade of multiple receptor tyrosine kinases by MGCD516 (Sitravatinib), a novel small molecule inhibitor, shows potent anti-tumor activity in preclinical models of sarcoma.

PubMed

Patwardhan, Parag P; Ivy, Kathryn S; Musi, Elgilda; de Stanchina, Elisa; Schwartz, Gary K

2016-01-26

Sarcomas are rare but highly aggressive mesenchymal tumors with a median survival of 10-18 months for metastatic disease. Mutation and/or overexpression of many receptor tyrosine kinases (RTKs) including c-Met, PDGFR, c-Kit and IGF1-R drive defective signaling pathways in sarcomas. MGCD516 (Sitravatinib) is a novel small molecule inhibitor targeting multiple RTKs involved in driving sarcoma cell growth. In the present study, we evaluated the efficacy of MGCD516 both in vitro and in mouse xenograft models in vivo. MGCD516 treatment resulted in significant blockade of phosphorylation of potential driver RTKs and induced potent anti-proliferative effects in vitro. Furthermore, MGCD516 treatment of tumor xenografts in vivo resulted in significant suppression of tumor growth. Efficacy of MGCD516 was superior to imatinib and crizotinib, two other well-studied multi-kinase inhibitors with overlapping target specificities, both in vitro and in vivo. This is the first report describing MGCD516 as a potent multi-kinase inhibitor in different models of sarcoma, superior to imatinib and crizotinib. Results from this study showing blockade of multiple driver signaling pathways provides a rationale for further clinical development of MGCD516 for the treatment of patients with soft-tissue sarcoma.

Significant blockade of multiple receptor tyrosine kinases by MGCD516 (Sitravatinib), a novel small molecule inhibitor, shows potent anti-tumor activity in preclinical models of sarcoma

PubMed Central

Musi, Elgilda; de Stanchina, Elisa; Schwartz, Gary K.

2016-01-01

Sarcomas are rare but highly aggressive mesenchymal tumors with a median survival of 10–18 months for metastatic disease. Mutation and/or overexpression of many receptor tyrosine kinases (RTKs) including c-Met, PDGFR, c-Kit and IGF1-R drive defective signaling pathways in sarcomas. MGCD516 (Sitravatinib) is a novel small molecule inhibitor targeting multiple RTKs involved in driving sarcoma cell growth. In the present study, we evaluated the efficacy of MGCD516 both in vitro and in mouse xenograft models in vivo. MGCD516 treatment resulted in significant blockade of phosphorylation of potential driver RTKs and induced potent anti-proliferative effects in vitro. Furthermore, MGCD516 treatment of tumor xenografts in vivo resulted in significant suppression of tumor growth. Efficacy of MGCD516 was superior to imatinib and crizotinib, two other well-studied multi-kinase inhibitors with overlapping target specificities, both in vitro and in vivo. This is the first report describing MGCD516 as a potent multi-kinase inhibitor in different models of sarcoma, superior to imatinib and crizotinib. Results from this study showing blockade of multiple driver signaling pathways provides a rationale for further clinical development of MGCD516 for the treatment of patients with soft-tissue sarcoma. PMID:26675259

Design and in vivo evaluation of carvedilol buccal mucoadhesive patches.

PubMed

Thimmasetty, J; Pandey, G S; Babu, P R Sathesh

2008-07-01

The buccal region offers an attractive route of administration for systemic drug delivery. Carvedilol (dose, 3.125-25 mg) is beta-adrenergic antagonist. Its oral bioavailability is 25-35% because of first pass metabolism. Buccal absorption studies of a carvedilol solution in human volunteers showed 32.86% drug absorption. FTIR and UV spectroscopic methods revealed that there was no interaction between carvedilol and polymers. Carvedilol patches were prepared using HPMC, carbopol 934, eudragit RS 100, and ethylcellulose. The patches were evaluated for their thickness uniformity, folding endurance, weight uniformity, content uniformity, swelling behaviour, tensile strength, and surface pH. In vitro release studies were conducted for carvedilol-loaded patches in phosphate buffer (pH, 6.6) solution. Patches exhibited drug release in the range of 86.26 to 98.32% in 90 min. Data of in vitro release from patches were fit to different equations and kinetic models to explain release profiles. Kinetic models used were zero and first-order equations, Hixon-Crowell, Higuchi, and Korsmeyer-Peppas models. In vivo drug release studies in rabbits showed 90.85% of drug release from HPMC-carbopol patch while it was 74.63 to 88.02% within 90 min in human volunteers. Good correlation among in vitro release and in vivo release of carvedilol was observed.

Respiration in vitro: I. Spontaneous activity.

PubMed

Hamada, O; Garcia-Rill, E; Skinner, R D

1992-01-01

The present report describes respiratory-like activity recorded from intercostal muscles in the neonatal rat in vitro brain stem-spinal cord, rib-attached preparation. In this preparation from 1- to 4-day-old rats, spontaneous rhythmic and synchronized upward movements of the rib cage coincided with the recorded muscle activity. Spontaneous respiratory-like activity showed a frequency in the range of 0.05-0.2 Hz, with single-, double-, and mixed-burst patterns. Spontaneous activity declined over time, but increased in frequency as temperature increased. Multilevel recordings showed a cephalocaudal order of bursting of intercostal muscles. Brain stem transections at the prepontine level did not affect spontaneous frequency, whereas premedullary transections resulted in an increase in spontaneous respiratory frequency. High spinal transections eliminated spontaneous respiratory-like activity. These results suggest that there is a well-organized pontomedullary pattern generator for respiratory-like activity in this preparation, which can be modulated by temperature. The characteristics of these electromyographic (EMG) recordings allow comparison with previous in vitro studies of respiratory-like activity using nerve activity and in vivo studies using EMG activity. These results provide basic information on the spontaneous activity of this preparation as a prelude to the study of the effects of electrical stimulation of the spinal cord to induce respiratory-like activity, as described in the companion article.

Ameliorative effect of alkaloid extract of Cyclea peltata (Poir.) Hook. f. & Thoms. roots (ACP) on APAP/CCl4 induced liver toxicity in Wistar rats and in vitro free radical scavenging property.

PubMed

Shine, Varghese Jancy; Latha, Panikamparambil Gopalakrishnan; Suja, Somasekharan Nair Rajam; Anuja, Gangadharan Indira; Raj, Gopan; Rajasekharan, Sreedharan Nair

2014-02-01

To evaluate the hepatoprotective and antioxidant properties of alkaloid extract of Cyclea peltata (C. peltata) against paracetamol/carbon tetra chloride induced liver damage in Wistar rats. In vivo paracetamol/carbon tetrachloride induced liver damage in Wistar rats, in vitro free radical scavenging studies, HPTLC estimation of tetrandrine and direct analysis in real time- mass spectrometry of alkaloid extract of C. peltata were used for the validation. The results showed that pretreatment with alkaloid extract of C. peltata caused significant reduction of serum glutamate pyruvate transaminase, serum glutamate oxaloacetate transaminase, serum alkaline phosphatase, serum cholesterol, liver malondialdehyde levels. The reduced glutathione, catalase, superoxide dismutase levels in liver were increased with alkaloid extract of C. peltata treatment. These results were almost comparable to silymarin and normal control. Histopathological studies also substantiated the biochemical findings. The in vitro hydroxyl, superoxide and DPPH scavenging study of alkaloid extract of C. peltata showed significant free radical scavenging property. The hepatoprotective property of alkaloid extract of C. peltata against paracetamol/carbon tetrachloride may be due the synergistic action of alkaloids especially tetrandrine, fangchinoline through free radical scavenging and thus preventing oxidative stress.

Cytotoxic evaluation of hydroxyapatite-filled and silica/hydroxyapatite-filled acrylate-based restorative composite resins: An in vitro study.

PubMed

Chadda, Harshita; Naveen, Sangeetha Vasudevaraj; Mohan, Saktiswaren; Satapathy, Bhabani K; Ray, Alok R; Kamarul, Tunku

2016-07-01

Although the physical and mechanical properties of hydroxyapatite-filled dental restorative composite resins have been examined, the biocompatibility of these materials has not been studied in detail. The purpose of this in vitro study was to analyze the toxicity of acrylate-based restorative composite resins filled with hydroxyapatite and a silica/hydroxyapatite combination. Five different restorative materials based on bisphenol A-glycidyl methacrylate (bis-GMA) and tri-ethylene glycol dimethacrylate (TEGDMA) were developed: unfilled (H0), hydroxyapatite-filled (H30, H50), and silica/hydroxyapatite-filled (SH30, SH50) composite resins. These were tested for in vitro cytotoxicity by using human bone marrow mesenchymal stromal cells. Surface morphology, elemental composition, and functional groups were determined by scanning electron microscopy (SEM), energy-dispersive x-ray spectroscopy (EDX), and Fourier-transformed infrared spectroscopy (FTIR). The spectra normalization, baseline corrections, and peak integration were carried out by OPUS v4.0 software. Both in vitro cytotoxicity results and SEM analysis indicated that the composite resins developed were nontoxic and supported cell adherence. Elemental analysis with EDX revealed the presence of carbon, oxygen, calcium, silicon, and gold, while the presence of methacrylate, hydroxyl, and methylene functional groups was confirmed through FTIR analysis. The characterization and compatibility studies showed that these hydroxyapatite-filled and silica/hydroxyapatite-filled bis-GMA/TEGDMA-based restorative composite resins are nontoxic to human bone marrow mesenchymal stromal cells and show a favorable biologic response, making them potential biomaterials. Copyright © 2016 Editorial Council for the Journal of Prosthetic Dentistry. Published by Elsevier Inc. All rights reserved.

Pre-clinical Evaluation of a Cyanine-Based SPECT Probe for Multimodal Tumor Necrosis Imaging.

PubMed

Stammes, Marieke A; Knol-Blankevoort, Vicky T; Cruz, Luis J; Feitsma, Hans R I J; Mezzanotte, Laura; Cordfunke, Robert A; Sinisi, Riccardo; Dubikovskaya, Elena A; Maeda, Azusa; DaCosta, Ralph S; Bierau, Katja; Chan, Alan; Kaijzel, Eric L; Snoeks, Thomas J A; van Beek, Ermond R; Löwik, Clemens W G M

2016-12-01

Recently we showed that a number of carboxylated near-infrared fluorescent (NIRF) cyanine dyes possess strong necrosis avid properties in vitro as well as in different mouse models of spontaneous and therapy-induced tumor necrosis, indicating their potential use for cancer diagnostic- and prognostic purposes. In the previous study, the detection of the cyanines was achieved by whole body optical imaging, a technique that, due to the limited penetration of near-infrared light, is not suitable for investigations deeper than 1 cm within the human body. Therefore, in order to facilitate clinical translation, the purpose of the present study was to generate a necrosis avid cyanine-based NIRF probe that could also be used for single photon emission computed tomography (SPECT). For this, the necrosis avid NIRF cyanine HQ4 was radiolabeled with 111 indium, via the chelate diethylene triamine pentaacetic acid (DTPA). The necrosis avid properties of the radiotracer [ 111 In]DTPA-HQ4 were examined in vitro and in vivo in different breast tumor models in mice using SPECT and optical imaging. Moreover, biodistribution studies were performed to examine the pharmacokinetics of the probe in vivo. Using optical imaging and radioactivity measurements, in vitro, we showed selective accumulation of [ 111 In]DTPA-HQ4 in dead cells. Using SPECT and in biodistribution studies, the necrosis avidity of the radiotracer was confirmed in a 4T1 mouse breast cancer model of spontaneous tumor necrosis and in a MCF-7 human breast cancer model of chemotherapy-induced tumor necrosis. The radiotracer [ 111 In]DTPA-HQ4 possessed strong and selective necrosis avidity in vitro and in various mouse models of tumor necrosis in vivo, indicating its potential to be clinically applied for diagnostic purposes and to monitor anti-cancer treatment efficacy.

Astrocytes from adult Wistar rats aged in vitro show changes in glial functions.

PubMed

Souza, Débora Guerini; Bellaver, Bruna; Raupp, Gustavo Santos; Souza, Diogo Onofre; Quincozes-Santos, André

2015-11-01

Astrocytes, the most versatile cells of the central nervous system, play an important role in the regulation of neurotransmitter homeostasis, energy metabolism, antioxidant defenses and the anti-inflammatory response. Recently, our group characterized cortical astrocyte cultures from adult Wistar rats. In line with that work, we studied glial function using an experimental in vitro model of aging astrocytes (30 days in vitro after reaching confluence) from newborn (NB), adult (AD) and aged (AG) Wistar rats. We evaluated metabolic parameters, such as the glucose uptake, glutamine synthetase (GS) activity, and glutathione (GSH) content, as well as the GFAP, GLUT-1 and xCT expression. AD and AG astrocytes take up less glucose than NB astrocytes and had decreased GLUT1 expression levels. Furthermore, AD and AG astrocytes exhibited decreased GS activity compared to NB cells. Simultaneously, AD and AG astrocytes showed an increase in GSH levels, along with an increase in xCT expression. NB, AD and AG astrocytes presented similar morphology; however, differences in GFAP levels were observed. Taken together, these results improve the knowledge of cerebral senescence and represent an innovative tool for brain studies of aging. Copyright © 2015 Elsevier Ltd. All rights reserved.

Antioxidant properties and inhibitory effects of Satureja khozestanica essential oil on LDL oxidation induced-CuSO4 in vitro

PubMed Central

Bagheri, Shahrokh; Ahmadvand, Hassan; Khosrowbeygi, Ali; Ghazanfari, Farshid; Jafari, Narges; Nazem, Habibolah; Hosseini, Reza Haji

2013-01-01

Objective To assess various antioxidative activities of Satureja khozestanica essential oil (SKE) and its effect on oxidation of low density lipoprotein (LDL) induced by CuSO4 in vitro by monitoring the formation of conjugated dienes and malondialdehyde (MDA). Methods The formation of conjugated dienes, lag time and MDA were measured. Inhibition of this Cu-induced oxidation was studied in the presence of several concentrations of SKE. Also total antioxidant activity and free radical scavenging of SKE were determinated. Results It was demonstrated that SKE was able to inhibit LDL oxidation and decrease the resistance of LDL against oxidation. The inhibitory effects of SKE on LDL oxidation were dose-dependent at concentrations ranging from 50 to 200 µg/mL. Total antioxidant capacity of SKE was (3.20±0.40) nmol of ascorbic acid equivalents/g SKE. The SKE showed remarkable scavenging activity on 2, 2-diphenyl-picrylhydrazyl, IC50 (5.30±0.11) ng/mL. Conclusions This study shows that SKE is a source of potent antioxidants and prevents the oxidation of LDL in vitro and it may be suitable for use in food and pharmaceutical applications. PMID:23570012

Spermicidal activity of Indian seaweeds: an in vitro study.

PubMed

Prakash, S; Ravikumar, S; Reddy, K V R; Kannapiran, E

2014-05-01

Contraceptive properties of seaweeds are still stands as lacuna; in this context, the screening of in vitro male contraceptive properties of crude ethanolic extract of Indian seaweeds against normal human sperm is carried out. In total, twelve seaweeds were screened for in vitro spermicidal activity. Among these twelve seaweeds, Halimeda gracilis showed 100% inhibition of human spermatozoa at 10 mg ml(-1) concentration in 20 s and its EC50 value was 2.05 mg ml(-1) in 20 s. Further, dose- and time-dependent spermicidal assay revealed that the sperm was completely immobilised for 20 s. Plasma membrane of sperm was damaged due to the exposure of H. gracilis extract. MTT assay with H. gracilis extract showed 88.5% of cytotoxic incidence. H. gracilis extract tested for cytotoxicity against Artemia salina recorded LC50 value of 34.8 μg ml(-1) . Phytochemical analysis of H. gracilis extract evidenced the presence of alkaloids, flavonoids, proteins and sugars. Results of this study clearly inferred that the synergistic effect of active principles reside within the H. gracilis extract had shown better contraceptive activity. © 2013 Blackwell Verlag GmbH.

Effect of light polarization on the efficiency of photodynamic therapy of basal cell carcinomas: an in vitro cellular study.

PubMed

JalalKamali, M; Nematollahi-Mahani, S N; Shojaei, M; Shamsoddini, A; Arabpour, N

2018-02-01

In an in vitro study, the effect of light polarization on the efficiency of 5-aminolaevulinic acid (ALA) photodynamic therapy (PDT) of basal cell carcinoma (BCC) was investigated. Three states of light polarization (non-polarized, linearly polarized, and circularly polarized) were considered. Cells were exposed to green (532 pm 20 nm) irradiation from light emitting diodes. Cell survival was measured by the colorimetric assay (WST-1) and Trypan blue staining. The colorimetric assay showed a pronounced decrease in the cell viability (up to 30%) using polarized light compared to the non-polarized one in the wavelength region used. Similar results were obtained by the cell counting method (20-30% increase in cell death). The observed effect was dependent on the concentration of photosensitizer. The effect is more expressed in the case of linearly polarized light compared to the circularly polarized one. Results show that the use of polarized light increases the efficiency of in vitro ALA-PDT of BCC. Utilizing polarized light, it is possible to obtain the same effect from PDT by lower concentrations of photosensitizer. Additionally, the concentration dependency of PDT response and photo-bleaching is also reduced.

Successful transplantation of in vitro expanded human cadaver corneal endothelial precursor cells on to a cadaver bovine's eye using a nanocomposite gel sheet.

PubMed

Parikumar, Periyasamy; Haraguchi, Kazutoshi; Ohbayashi, Akira; Senthilkumar, Rajappa; Abraham, Samuel J K

2014-05-01

In vitro expansion of human corneal endothelial precursor (HCEP) cells has been reported via production of cell aggregated spheres. However, to translate this procedure in human patients warrants maintaining the position of the eyeballs facing down for 36 h, which is not feasible. In this study, we report a method using a nanocomposite (NC) gel sheet to accomplish the integration of HCEP cells to the endothelium of cadaver bovine's eyes. HCEP cells were isolated from the corneal endothelium of a cadaver human eye and then expanded using a thermoreversible gelation polymer (TGP) as reported earlier. For the study, three cadaver bovine eyes were used. The NC gel sheets were inserted into the bovine eyes', aligned and suture-fixed in position under the host endothelium. HCEP cells previously expanded in the TGP were harvested and injected using a 26-gauge syringe between the endothelium and the NC gel sheet. The eyes were left undisturbed for three hours following which the NC gel sheets were gently removed. The corneas were harvested and subjected to histopathological studies. Histopathological studies showed that all the three corneas used for NC gel sheet implantation showed the presence of engrafted HCEP cells, seen as multi-layered cells over the native endothelium of the bovine cornea. Examination of the NC gel sheets used for implantation showed that only very few corneal endothelial cells remained on the sheets amounting to what could be considered negligible. The use of the NC gel sheet makes HCEP cell transplantation feasible for human patients. Further in vitro basic studies followed by translational studies are necessary to bring this method for clinical application in appropriate indications.

Effect of phototherapy on shear bond strength of resin cements to zirconia ceramics: a systematic review and meta-analysis of in-vitro studies.

PubMed

Al-Aali, Khulud Abdulrahman

2018-05-11

The present study systematically reviewed the literature to investigate the effect of phototherapy on the shear bond strength (SBS) of resin cement to zirconia ceramic. electronic databases including MEDLINE (PubMed), ISI Web of Science, Scopus, ScIELO, LILACS and EMBASE until April 2018. The addressed focused question was: Does phototherapy increase the SBS of resin cement to zirconia ceramics?" A total of 8 in-vitro studies were included in the qualitative and quantitative analysis. The mean SBS for phototherapy ranged from 4.1 to 18.95 MPa while mean SBS for sandblasted zirconia-composite specimens ranged from 3.98 to 23.35 MPa in the included studies. Qualitative analysis showed 3 studies favoured application of phototherapy in significantly increasing SBS, while 4 studies indicated sandblasting showed significantly greater SBS of resin cement to zirconia ceramics. Considering the effects of phototherapy, significant heterogeneity for SBS (Q value = 136.37, p<0.0001, I 2  = 94.87%) was noticed among both the groups. The overall mean difference for SBS (SMD = -0.59, 95% CI = -1.99 to -0.80, p = 0.402) was not significant between phototherapy and sandblast (control) groups. Whether the effect of phototherapy on increasing the SBS of resin cement to zirconia ceramic is debatable. Further in-vitro studies should be performed in order to obtain strong conclusions. Copyright © 2018. Published by Elsevier B.V.

Proteopolymersomes: in vitro production of a membrane protein in polymersome membranes.

PubMed

Nallani, Madhavan; Andreasson-Ochsner, Mirjam; Tan, Cherng-Wen Darren; Sinner, Eva-Kathrin; Wisantoso, Yudi; Geifman-Shochat, Susana; Hunziker, Walter

2011-12-01

Polymersomes are stable self-assembled architectures which mimic cell membranes. For characterization, membrane proteins can be incorporated into such bio-mimetic membranes by reconstitution methods, leading to so-called proteopolymersomes. In this work, we demonstrate the direct incorporation of a membrane protein into polymersome membranes by a cell-free expression system. Firstly, we demonstrate pore formation in the preformed polymersome membrane using α-hemolysin. Secondly, we use claudin-2, a protein involved in cell-cell interactions, to demonstrate the in vitro expression of a membrane protein into these polymersomes. Surface plasmon resonance (Biacore) binding studies with the claudin-2 proteopolymersomes and claudin-2 specific antibodies are performed to show the presence of the in vitro expressed protein in polymersome membranes.

In vitro antifungal activity of terpinen-4-ol, eugenol, carvone, 1,8-cineole (eucalyptol) and thymol against mycotoxigenic plant pathogens.

PubMed

Morcia, C; Malnati, M; Terzi, V

2012-01-01

The aim of this study was to examine the effect of five naturally occurring compounds from essential oils on 10 different species of mycotoxigenic fungi involved in several plant diseases. The antifungal activities of terpinen-4-ol, eugenol, carvone, 1,8-cineole (eucalyptol) and thymol were observed in vitro on Fusarium subglutinans, Fusarium cerealis, Fusarium verticillioides, Fusarium proliferatum, Fusarium oxysporum, Fusarium sporotrichioides, Aspergillus tubingensis, Aspergillus carbonarius, Alternaria alternata and Penicillium sp. The naturally occurring compounds tested showed toxic effects on in vitro mycelium growth of all fungal species but with different level of potency. The results are encouraging for further investigations of in planta antifungal activities of these essential oils components.

Inhibition of p53 Mutant Peptide Aggregation In Vitro by Cationic Osmolyte Acetylcholine Chloride.

PubMed

Chen, Zhaolin; Kanapathipillai, Mathumai

2017-01-01

Mutations of tumor suppressor protein p53 are present in almost about 50% of all cancers. It has been reported that the p53 mutations cause aggregation and subsequent loss of p53 function, leading to cancer progression. Here in this study we focus on the inhibitory effects of cationic osmolyte molecules acetylcholine chloride, and choline on an aggregation prone 10 amino acid p53 mutant peptide WRPILTIITL, and the corresponding wildtype peptide RRPILTIITL in vitro. The characterization tools used for this study include Thioflavin- T (ThT) induced fluorescence, transmission electron microscopy (TEM), congo red binding, turbidity, dynamic light scattering (DLS), and cell viability assays. The results show that acetylcholine chloride in micromolar concentrations significantly inhibit p53 mutant peptide aggregation in vitro, and could be promising candidate for p53 mutant/ misfolded protein aggregation inhibition. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Quinazoline clubbed 1,3,5-triazine derivatives as VEGFR2 kinase inhibitors: design, synthesis, docking, in vitro cytotoxicity and in ovo antiangiogenic activity.

PubMed

Pathak, Prateek; Shukla, Parjanya Kumar; Kumar, Vikas; Kumar, Ankit; Verma, Amita

2018-04-16

A series of quinazoline clubbed 1,3,5-triazine derivatives (QCT) were synthesized and evaluated for their in vitro anticancer activity against HeLa (human cervical cancer), MCF-7 (human breast cancer cell), HL-60 (human promyelocytic leukemia cell), HepG2 (human Hepatocellular carcinoma cell), and one normal cell line HFF (human foreskin fibroblasts). In vitro assay result encouraged to further move towards in ovo anticancer evaluation using chick embryo. The series of QCT derivatives showed higher anticancer and antiangiogenic activity against HeLa and MCF-7 cell lines. In the series, synthetic molecule 8d, 8l, and 8m displayed significant activity. Further, these results substantiated by docking study on VGFR2. SAR study concluded that the potency of drugs depends on the nature of aliphatic substitution and the heterocyclic ring system.

Effect of Injection Molding Melt Temperatures on PLGA Craniofacial Plate Properties during In Vitro Degradation.

PubMed

de Melo, Liliane Pimenta; Salmoria, Gean Vitor; Fancello, Eduardo Alberto; Roesler, Carlos Rodrigo de Mello

2017-01-01

The purpose of this article is to present mechanical and physicochemical properties during in vitro degradation of PLGA material as craniofacial plates based on different values of injection molded temperatures. Injection molded plates were submitted to in vitro degradation in a thermostat bath at 37 ± 1°C by 16 weeks. The material was removed after 15, 30, 60, and 120 days; then bending stiffness, crystallinity, molecular weights, and viscoelasticity were studied. A significant decrease of molecular weight and mechanical properties over time and a difference in FT-IR after 60 days showed faster degradation of the material in the geometry studied. DSC analysis confirmed that the crystallization occurred, especially in higher melt temperature condition. DMA analysis suggests a greater contribution of the viscous component of higher temperature than lower temperature in thermomechanical behavior. The results suggest that physical-mechanical properties of PLGA plates among degradation differ per injection molding temperatures.

In vitro and in vivo studies on the transport of PEGylated silica nanoparticles across the blood-brain barrier.

PubMed

Liu, Dan; Lin, Bingqian; Shao, Wei; Zhu, Zhi; Ji, Tianhai; Yang, Chaoyong

2014-02-12

Transport of PEGylated silica nanoparticles (PSiNPs) with diameters of 100, 50, and 25 nm across the blood-brain barrier (BBB) was evaluated using an in vitro BBB model based on mouse cerebral endothelial cells (bEnd.3) cultured on transwell inserts within a chamber. In vivo animal experiments were further performed by noninvasive in vivo imaging and ex vivo optical imaging after injection via carotid artery. Confocal fluorescence studies were carried out to evaluate the uptake of PSiNPs by brain endothelial cells. The results showed that PSiNPs can traverse the BBB in vitro and in vivo. The transport efficiency of PSiNPs across BBB was found to be size-dependent, with increased particle size resulting in decreased efficiency. This work points to the potential application of small sized silica nanoparticles in brain imaging or drug delivery.

Experimental research on the in vitro antitumor effects of Crataegus sanguinea.

PubMed

Sun, Jianling; Gao, Guolan; Gao, YuLian; Xiong, LiJuan; Li, Xiaoying; Guo, Jihong; Zhang, Yueming

2013-09-01

Crataegus sanguinea is a wild plant, which has been widely grown in the north and south of the Tianshan mountains in Xinjiang. In order to explore their anti-cancer properties, edible wild plants from Xinjiang have been tested for their antitumor properties. We used Ames tests, mouse bone marrow polychromatic erythrocytes micronucleus tests, and tumor cells cultured in vitro to study the anti-mutagenic and anti-tumor effects of C. sanguinea extract. We have shown that C. sanguinea has anti-mutagenic effect, but no mutagenicity. Cell culture in vitro experiments show that there is no inhibition of growth or increase in cell death on normal mouse fibroblasts, but a stronger inhibition of cell growth and an increase in cell death of Hep-2 and MGC-803 tumor cells. The results of this study illustrate that C. sanguinea extract has both anti-mutagenic and anti-tumor effects.

Efficient ibuprofen delivery from anhydrous semisolid formulation based on a novel cross-linked silicone polymer network: an in vitro and in vivo study.

PubMed

Aliyar, Hyder; Huber, Robert; Loubert, Gary; Schalau, Gerald

2014-07-01

The use of silicone as a primary polymer in topical semisolid pharmaceutical formulations is infrequent. Recent development of novel silicone materials provides an opportunity to investigate their drug delivery efficiencies. In this study, an anhydrous semisolid formulation was prepared using a novel cross-linked silicone polymer network swollen in isododecane. Similar formulations were prepared using petrolatum, an acrylic, or a cellulose polymer. All formulations contained 5% ibuprofen (IBP). In vitro permeability was evaluated for all formulations and a commercial product using human cadaver epidermis. The silicone formulation delivered IBP more efficiently than all other formulations in terms of flux, cumulative amount, and percent drug release. The silicone formulation showed the maximum flux of 85.9 μg . cm(-2) . h(-1) and a cumulative IBP release of 261.6 μg in 8 h, whereas the benchmark showed 20.1 μg . cm(-2) . h(-1) and 30.9 μg, respectively. An in vivo study conducted on rats showed calculated blood AUCs of 59.2 and 17.6 μg . h/g (p < 0.003) for the silicone formulation and the benchmark, respectively. The IBP in excised rat skin was 264 ± 59 μg/g for the silicone formulation and 102 ± 5 μg/g for the benchmark. The results obtained from the in vitro and in vivo studies demonstrate efficient topical IBP delivery by the silicone formulation. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association.

Nanocarriers as phototherapeutic drug delivery system: Appraisal of three different nanosystems in an in vivo and in vitro exploratory study.

PubMed

Ricci-Junior, Eduardo; de Oliveira de Siqueira, Luciana Betzler; Rodrigues, Raphaela Aparecida Schuenck; Sancenón, Félix; Martínez-Máñez, Ramón; de Moraes, João Alfredo; Santos-Oliveira, Ralph

2018-03-01

The use of nanosystems as diagnosing and therapy systems is increasing each year. There are several nanosystems available and the most prominent ones are: mesoporous silica, nanoemulsion and polymeric nanoparticles. With characteristics like low toxicology, and easy-producing process they have advantages when compared with the traditional system used, as they show specific targeting, controlled release, and higher penetration. In this study we tested three different nanocarriers (polymeric nanoparticles, nanoemulsion and mesoporous silica) containing phthalocyanineas possible PDT drugs (nanodrugs). They were tested in vitro and in vivo: cells and healthy mice, respectively, in order to understand the biological behavior and reach the initial conclusions. The results in cells showed that a dose response was observed with different concentrations of the three nanocarriers. The results in animal showed that all nanosystems have potential for application in PDT, since they were able to produce a visible effect in healthy animals. Copyright © 2017 Elsevier B.V. All rights reserved.

A miniaturized glucose biosensor for in vitro and in vivo studies.

PubMed

Yang, Yang-Li; Huang, Jian-Feng; Tseng, Ta-Feng; Lin, Chia-Ching; Lou, Shyh-Liang

2008-01-01

A miniaturized wireless glucose biosensor has been developed to perform in vitro and in vivo studies. It consists of an external control subsystem and an implant sensing subsystem. The implant subsystem consists of a micro-processor, which coordinates circuitries of radio frequency, power regulator, command demodulator, glucose sensing trigger and signal read-out. Except for a set of sensing electrodes, the micro-processor, the circuitries and a receiving coil were hermetically sealed with polydimethylsiloxane. The electrode set is a substrate of silicon oxide coated with platinum, which includes a working electrode and a reference electrode. Glucose oxidase was immobilized on the surface of the working electrode. The implant subsystem bi-directionally communicates with the external subsystem via radio frequency technologies. The external subsystem wirelessly supplies electricity to power the implant, issues commands to the implant to perform tasks, receives the glucose responses detected by the electrode, and relays the response signals to a computer through a RS-232 connection. Studies of in vitro and in vivo were performed to evaluate the biosensor. The linear response of the biosensor is up to 15 mM of glucose in vitro. The results of in vivo study show significant glucose variations measured from the interstitial tissue fluid of a diabetes rat in fasting and non-fasting periods.

Surface Chemistry Interactions of Cationorm with Films by Human Meibum and Tear Film Compounds

PubMed Central

Georgiev, Georgi As.; Yokoi, Norihiko; Nencheva, Yana; Peev, Nikola; Daull, Philippe

2017-01-01

Cationorm® (CN) cationic nanoemulsion was demonstrated to enhance tear film (TF) stability in vivo possibly via effects on tear film lipid layer (TFLL). Therefore the interactions of CN with human meibum (MGS) and TFLL in vitro and in vivo deserve special study. MGS and CN were spread at the air/water interface of a Langmuir surface balance to ensure a range of MGS/CN oil phase ratios: 20/1, 10/1, 5/1, 3/1, 2/1 and 1/1. The films capability to reorganize during dynamic area changes was evaluated via the surface pressure-area compression isotherms and step/relaxation dilatational rheology studies. Films structure was monitored with Brewster angle microscopy. CN/TFLL interactions at the ocular surface were monitored with non-contact specular microscopy. The in vitro studies of MGS/CN layers showed that (i) CN inclusion (at fixed MGS content) increased film elasticity and thickness and that (ii) CN can compensate for moderate meibum deficiency in MGS/CN films. In vivo CN mixed with TFLL in a manner similar to CN/MGS interactions in vitro, and resulted in enhanced thickness of TFLL. In vitro and in vivo data complement each other and facilitated the study of the composition-structure-function relationship that determines the impact of cationic nanoemulsions on TF. PMID:28718823

Fertilisation in the horse and paracrine signalling in the oviduct.

PubMed

Goudet, Ghylène

2011-01-01

The mammalian oviduct plays a crucial role in the preparation of gametes for fertilisation (transport and final maturation) and fertilisation itself. An increasing number of studies offers a comprehensive overview of the functions of the oviduct and its secretions, but this topic has had limited investigation in the horse. Limited data are available on the final oocyte maturation in the equine oviduct. However, in vitro and in vivo systems have been established to analyse the influence of equine oviduct epithelial cells (OEC) during maturation on the potential of oocytes for fertilisation and development. Most studies focus on the role of the oviduct in equine sperm function, such as spermatozoa transport, attachment to oviduct epithelium, viability, motility and capacitation. Moreover, some possible candidate molecules for sperm-oviducal interactions have been identified in the horse. Finally, the low efficiency of conventional in vitro fertilisation and the in vivo fertilisation of equine oocytes transferred into the oviduct of an inseminated mare predicted an influence of oviduct in equine fertilisation. Actually, in vivo and in vitro experiments demonstrated a role of the oviduct in equine fertilisation. Moreover, recent studies showed a beneficial effect of homologous and heterologous OEC on equine in vitro fertilisation, and some candidate molecules have been studied.

Impact of an Anticaries Mouthrinse on In Vitro Remineralization and Microbial Control

PubMed Central

Sun, Frank C.; Engelman, E. Eric; McGuire, James A.; Kosmoski, Gabrielle; Carratello, Lauren; Ricci-Nittel, Danette; Zhang, Jane Z.; Schemehorn, Bruce R.; Gambogi, Robert J.

2014-01-01

Objective. The objective of this research was to evaluate the caries control potential of a new fluoride mouthrinse that also contained antimicrobial agents and a biofilm disrupting agent using different in vitro models. Methods. Four in vitro studies were conducted to assess the performance of this three pronged approach to caries control: (1) traditional enamel fluoride uptake, (2) surface microhardness study using pH cycling model and subsequent fluoride uptake, (3) a salivary biofilm flow-through study to determine the anti-microbial activity, and (4) a single species biofilm model measuring effect on biofilm matrix disruption. Results. The data showed that a LISTERINE rinse with fluoride, essential oils and xylitol was superior in promoting enamel fluoride uptake and in enhancing antimicrobial activity over traditional commercially available fluoridated products. An increase of the surface microhardness was observed when the LISTERINE rinse was used in combination with fluoridated toothpaste versus the fluoridated toothpaste alone. Finally, it was demonstrated that xylitol solutions disrupted and reduced the biovolume of biofilm matrix of mature Streptococcus mutans. Conclusion. These in vitro studies demonstrated that a fluoride mouthrinse with antimicrobial agent and biofilm matrix disrupting agent provided multifaceted and enhanced anti-caries efficacy by promoting remineralization, reducing acidogenic bacteria and disrupting biofilm matrix. PMID:24648842

New investigations into the genotoxicity of cobalt compounds and their impact on overall assessment of genotoxic risk.

PubMed

Kirkland, David; Brock, Tom; Haddouk, Hasnaà; Hargeaves, Victoria; Lloyd, Melvyn; Mc Garry, Sarah; Proudlock, Raymond; Sarlang, Séverine; Sewald, Katherina; Sire, Guillaume; Sokolowski, Andrea; Ziemann, Christina

2015-10-01

The genotoxicity of cobalt metal and cobalt compounds has been widely studied. Several publications show induction of chromosomal aberrations, micronuclei or DNA damage in mammalian cells in vitro in the absence of S9. Mixed results were seen in gene mutation studies in bacteria and mammalian cells in vitro, and in chromosomal aberration or micronucleus assays in vivo. To resolve these inconsistencies, new studies were performed with soluble and poorly soluble cobalt compounds according to OECD-recommended protocols. Induction of chromosomal damage was confirmed in vitro, but data suggest this may be due to oxidative stress. No biologically significant mutagenic responses were obtained in bacteria, Tk(+/-) or Hprt mutation tests. Negative results were also obtained for chromosomal aberrations (in bone marrow and spermatogonia) and micronuclei at maximum tolerated doses in vivo. Poorly soluble cobalt compounds do not appear to be genotoxic. Soluble compounds do induce some DNA and chromosomal damage in vitro, probably due to reactive oxygen. The absence of chromosome damage in robust GLP studies in vivo suggests that effective protective processes are sufficient to prevent oxidative DNA damage in whole mammals. Overall, there is no evidence of genetic toxicity with relevance for humans of cobalt substances and cobalt metal. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Curcumin-docetaxel co-loaded nanosuspension for enhanced anti-breast cancer activity.

PubMed

Sahu, Bhanu P; Hazarika, Hemanga; Bharadwaj, Rituraj; Loying, Pojul; Baishya, Rinku; Dash, Suvakanta; Das, Malay K

2016-08-01

A curcumin-docetaxel co-loaded nanosuspension with increased anti-breast cancer activity was developed. Curcumin is a potential anticancer agent with p-glycoprotein (p-gp) inhibiting activity may be co-administered with docetaxel as a nanosuspension to enhance its anticancer effect by increasing the oral bioavailability and decreasing drug efflux. Nanosuspensions of curcumin and docetaxel were prepared by precipitation-homozenisation technique and evaluated for particle size, polydispersity, zeta potential and drug release. The in vitro MTT assay was conducted using MCF-7 for anti-breast cancer activity. The in vivo biodistribution by radiolabeling and tumor inhibition study was conducted in mice. Homogenous nanosuspensions of 80 ± 20 nm were obtained with increased solubility. The drugs as nanosuspensions showed higher cytotoxicity on MCF-7 cell line compared to their suspensions due to the increased in vitro cellular uptake. Due to this increased solubility, sensitization of tumor cells and inhibition of p-gp the in-vivo results showed greater tumor inhibition rate of up to 70% in MCF-7 treated mice. Histopathological results showed higher apoptotic activity and reduced level of angiogenesis. The in vitro and in vivo study of the nanosuspensions has shown that Co-administration of Curcumin as a p-gp inhibitor with docetaxel may have the potential to increase the anti-breast cancer efficacy of both drugs.

Enhancing the efficiency of bortezomib conjugated to pegylated gold nanoparticles: an in vitro study on human pancreatic cancer cells and adenocarcinoma human lung alveolar basal epithelial cells.

PubMed

Coelho, Sílvia Castro; Almeida, Gabriela M; Santos-Silva, Filipe; Pereira, Maria Carmo; Coelho, Manuel A N

2016-08-01

Gold nanoparticles have become promising vectors for cancer diagnosis and treatment. The present study investigates the effect of bortezomib (BTZ), a proteasome inhibitor, conjugated with pegylated gold nanoparticles (PEGAuNPs) in pancreatic and lung cancer cells. Synthesized gold nanoparticles (PEGAuNPs) were conjugated with bortezomib antitumor drug. We investigated the cytotoxicity induced by BTZ conjugated with functionalized gold nanoparticles in vitro, in the human pancreatic (S2-013) and lung (A549) cancer cell lines. We found an efficient of conjugation of BTZ with PEGAuNPs. In vitro assays showed that after 72 h' incubation with PEGAuNPs-BTZ cancer cells revealed alterations in morphology; also for S2-013 and A549 cancer cells, the IC50 value of free BTZ is respectively 1.5 and 4.3 times higher than the IC50 value of PEGAuNPs-BTZ. Furthermore, for TERT-HPNE, the IC50 value is around 63 times lower for free BTZ than the conjugated nanovehicle. Cell growth inhibition results showed a remarkable enhancement in the effect of BTZ when conjugated with AuNPs. Our findings showed that conjugation with PEGAuNPs enhance the BTZ growth-inhibition effect on human cancer cells (S2-013 and A549) and decreases its toxicity against normal cells (TERT-HPNE).

Short communication: Effect of casein haplotype on angiotensin-converting enzyme inhibitory and antioxidant capacities of milk casein from Italian Holstein cows before and following in vitro digestion with gastrointestinal enzymes.

PubMed

Perna, Annamaria; Simonetti, Amalia; Gambacorta, Emilio

2016-09-01

The aim of this work was to investigate the effect of casein haplotype (αS1, β, and κ) on antioxidative and angiotensin-converting enzyme (ACE) inhibitory capacities of milk casein from Italian Holstein cows before and following in vitro digestion with gastrointestinal enzymes. The antioxidant capacity was measured using 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid and ferric-reducing antioxidant power assays, whereas ACE inhibition was determined by ACE-inhibitory assay. The ACE-inhibitory and antioxidant capacities of milk casein increased during in vitro gastrointestinal digestion. Casein haplotype significantly influenced the antioxidative and ACE-inhibitory capacities of digested casein. In particular, BB-A(2)A(1)-AA casein and BB-A(1)A(1)-AA casein showed the highest ACE-inhibitory capacity, BB-A(2)A(2)-AA casein showed the highest antioxidant capacity, whereas BB-A(2)A(2)-BB casein showed the lowest biological capacity. To date, few studies have been done on the effect of casein haplotype on biological capacity of milk casein, thus the present study sets the basis for a new knowledge that could lead to the production of milk with better nutraceutical properties. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Probiotic Microorganisms Inhibit Epithelial Cell Internalization of Botulinum Neurotoxin Serotype A

PubMed Central

Lam, Tina I.; Tam, Christina C.; Stanker, Larry H.; Cheng, Luisa W.

2016-01-01

Botulinum neurotoxins (BoNTs) are some of the most poisonous natural toxins known to man and are threats to public health and safety. Previous work from our laboratory showed that both BoNT serotype A complex and holotoxin can bind and transit through the intestinal epithelia to disseminate in the blood. The timing of BoNT/A toxin internalization was shown to be comparable in both the Caco-2 in vitro cell culture and in the oral mouse intoxication models. Probiotic microorganisms have been extensively studied for their beneficial effects in not only maintaining the normal gut mucosa but also protection from allergens, pathogens, and toxins. In this study, we evaluate whether probiotic microorganisms will block BoNT/A uptake in the in vitro cell culture system using Caco-2 cells. Several probiotics tested (Saccharomyces boulardii, Lactobacillus acidophilus, Lactobacillus rhamnosus LGG, and Lactobacillus reuteri) blocked BoNT/A uptake in a dose-dependent manner whereas a non-probiotic strain of Escherichia coli did not. We also showed that inhibition of BoNT/A uptake was not due to the degradation of BoNT/A nor by sequestration of toxin via binding to probiotics. These results show for the first time that probiotic treatment can inhibit BoNT/A binding and internalization in vitro and may lead to the development of new therapies. PMID:27999281

Probiotic Microorganisms Inhibit Epithelial Cell Internalization of Botulinum Neurotoxin Serotype A.

PubMed

Lam, Tina I; Tam, Christina C; Stanker, Larry H; Cheng, Luisa W

2016-12-16

Botulinum neurotoxins (BoNTs) are some of the most poisonous natural toxins known to man and are threats to public health and safety. Previous work from our laboratory showed that both BoNT serotype A complex and holotoxin can bind and transit through the intestinal epithelia to disseminate in the blood. The timing of BoNT/A toxin internalization was shown to be comparable in both the Caco-2 in vitro cell culture and in the oral mouse intoxication models. Probiotic microorganisms have been extensively studied for their beneficial effects in not only maintaining the normal gut mucosa but also protection from allergens, pathogens, and toxins. In this study, we evaluate whether probiotic microorganisms will block BoNT/A uptake in the in vitro cell culture system using Caco-2 cells. Several probiotics tested ( Saccharomyces boulardii , Lactobacillus acidophilus , Lactobacillus rhamnosus LGG, and Lactobacillus reuteri ) blocked BoNT/A uptake in a dose-dependent manner whereas a non-probiotic strain of Escherichia coli did not. We also showed that inhibition of BoNT/A uptake was not due to the degradation of BoNT/A nor by sequestration of toxin via binding to probiotics. These results show for the first time that probiotic treatment can inhibit BoNT/A binding and internalization in vitro and may lead to the development of new therapies.

Mechanism of age-dependent involution in embryonic chick notochords.

PubMed

Ghanem, E; Cornelissen, M; Thierens, H; De Ridder, L

1996-07-15

To study the possible mechanism of the age-dependent involution of the notochord, isolated mesenchyme-free notochords of chick embryos were cultured in vitro and compared with their counterparts in vivo. Two different aspects were evaluated: (1) DNA synthesis measured by [3H]thymidine incorporation and visualized by autoradiography and (2) cell death quantified by counting the number of pyknotic nuclei. The results demonstrate that [3H]thymidine uptake by notochords shows an age-dependent decrease in vitro as well as in vivo. The number of [3H]thymidine-labelled notochord cells, however, is higher in vitro than in vivo. At the same time, there is an age-dependent increase in pyknosis in the notochord in vivo and in vitro. So, during the aging process, the number of both pyknotic nuclei and of [3H]thymidine-labelled nuclei suggest a high turnover of notochord cells in vitro. From these results, we can conclude that the process of involution in aging notochord seems to be controlled by a programmed intrinsic process, which might be influenced partially by the microenvironment in vivo.

In vitro generation of viral-antigen dependent cytotoxic T-cells from ginbuna crucian carp, Carassius auratus langsdorfii

DOE Office of Scientific and Technical Information (OSTI.GOV)

Somamoto, Tomonori, E-mail: somamoto@agr.kyushu-u.ac.j; Okamoto, Nobuaki; Nakanishi, Teruyuki

2009-06-20

Little is known about antigen-specific T-cell responses to viruses in teleosts due to a lack of a suitable experimental system using inbred or clonal animals. In the present study we have successfully induced an in vitro generation of virus-specific cytotoxic T-cells (CTLs) from isogeneic ginbuna crucian carp. Responder cells (primarily lymphocytes) from crucian carp haematopoietic necrosis virus (CHNV)-infected fish were capable of proliferating after stimulation in vitro with CHNV-infected syngeneic stimulator cells (primarily lymphocytes and macrophages). The effector cells collected 8 and 12 days after the in vitro stimulation efficiently lysed CHNV-infected syngeneic cells, but not CHNV-infected allogeneic cells ormore » different virus (EVA)-infected syngeneic cells. Furthermore, in situ hybridization analysis showed that some effector cells binding to a CHNV-infected target were TCRbeta or CD8alpha positive. These results provide evidence that the teleost effector cells generated in vitro correspond to virus-specific CTL and they recognize virus-infected target cells in a similar manner of mammalian counterparts.« less

Pattern Learning, Damage and Repair within Biological Neural Networks

NASA Astrophysics Data System (ADS)

Siu, Theodore; Fitzgerald O'Neill, Kate; Shinbrot, Troy

2015-03-01

Traumatic brain injury (TBI) causes damage to neural networks, potentially leading to disability or even death. Nearly one in ten of these patients die, and most of the remainder suffer from symptoms ranging from headaches and nausea to convulsions and paralysis. In vitro studies to develop treatments for TBI have limited in vivo applicability, and in vitro therapies have even proven to worsen the outcome of TBI patients. We propose that this disconnect between in vitro and in vivo outcomes may be associated with the fact that in vitro tests assess indirect measures of neuronal health, but do not investigate the actual function of neuronal networks. Therefore in this talk, we examine both in vitro and in silico neuronal networks that actually perform a function: pattern identification. We allow the networks to execute genetic, Hebbian, learning, and additionally, we examine the effects of damage and subsequent repair within our networks. We show that the length of repaired connections affects the overall pattern learning performance of the network and we propose therapies that may improve function following TBI in clinical settings.

Biodegradation performance of a chitosan coated magnesium-zinc-tricalcium phosphate composite as an implant.

PubMed

Zhao, Jun; Chen, Liangjian; Yu, Kun; Chen, Chang; Dai, Yilong; Qiao, Xueyan; Yan, Yang

2014-09-01

A Mg-Zn-tricalcium phosphate composite with a chitosan coating was prepared in this investigation to study its biodegradation performance both in vitro and in vivo conditions. The in vitro test results show that the immersion corrosion rate, the pH values of the simulated body fluids and the released metal ion concentration of the chitosan coated composite are all lower than those of the uncoated composite. The in vitro cytotoxicity test shows that the chitosan coated specimens is safe for cellular applications. When the chitosan coated composite is tested in vivo, the concentration of metal ions from the composite observed in the venous blood of Zelanian rabbits is less than the uncoated composite specimens. The chitosan coating slows down the in vivo degradation of the composite after surgery. In vivo testing also indicates that the chitosan coated composite is harmless to important visceral organs, including the heart, kidneys, and liver of the rabbits. The new bone formation surrounding the chitosan coated composite implant shows that the composite improves the concrescence of the bone tissues. The chitosan coating is an effective corrosion resistant layer that reduces the hydrogen release of the implant composite, thereby decreasing the subcutaneous gas bubbles formed.

A homeopathic remedy from arnica, marigold, St. John's wort and comfrey accelerates in vitro wound scratch closure of NIH 3T3 fibroblasts.

PubMed

Hostanska, Katarina; Rostock, Matthias; Melzer, Joerg; Baumgartner, Stephan; Saller, Reinhard

2012-07-18

Drugs of plant origin such as Arnica montana, Calendula officinalis or Hypericum perforatum have been frequently used to promote wound healing. While their effect on wound healing using preparations at pharmacological concentrations was supported by several in vitro and clinical studies, investigations of herbal homeopathic remedies on wound healing process are rare. The objective of this study was to investigate the effect of a commercial low potency homeopathic remedy Similasan® Arnica plus Spray on wound closure in a controlled, blind trial in vitro. We investigated the effect of an ethanolic preparation composed of equal parts of Arnica montana 4x, Calendula officinalis 4x, Hypericum perforatum 4x and Symphytum officinale 6x (0712-2), its succussed hydroalcoholic solvent (0712-1) and unsuccussed solvent (0712-3) on NIH 3T3 fibroblasts. Cell viability was determined by WST-1 assay, cell growth using BrdU uptake, cell migration by chemotaxis assay and wound closure by CytoSelect ™Wound Healing Assay Kit which generated a defined "wound field". All assays were performed in three independent controlled experiments. None of the three substances affected cell viability and none showed a stimulating effect on cell proliferation. Preparation (0712-2) exerted a stimulating effect on fibroblast migration (31.9%) vs 14.7% with succussed solvent (0712-1) at 1:100 dilutions (p < 0.001). Unsuccussed solvent (0712-3) had no influence on cell migration (6.3%; p > 0.05). Preparation (0712-2) at a dilution of 1:100 promoted in vitro wound closure by 59.5% and differed significantly (p < 0.001) from succussed solvent (0712-1), which caused 22.1% wound closure. Results of this study showed that the low potency homeopathic remedy (0712-2) exerted in vitro wound closure potential in NIH 3T3 fibroblasts. This effect resulted from stimulation of fibroblasts motility rather than of their mitosis.

A combined approach to investigate the toxicity of an industrial landfill's leachate: Chemical analyses, risk assessment and in vitro assays

DOE Office of Scientific and Technical Information (OSTI.GOV)

Baderna, D., E-mail: diego.baderna@marionegri.it; Maggioni, S.; Boriani, E.

2011-05-15

Solid wastes constitute an important and emerging problem. Landfills are still one of the most common ways to manage waste disposal. The risk assessment of pollutants from landfills is becoming a major environmental issue in Europe, due to the large number of sites and to the importance of groundwater protection. Furthermore, there is lack of knowledge for the environmental, ecotoxicological and toxicological characteristics of most contaminants contained into landfill leacheates. Understanding leachate composition and creating an integrated strategy for risk assessment are currently needed to correctly face the landfill issues and to make projections on the long-term impacts of amore » landfill, with particular attention to the estimation of possible adverse effects on human health and ecosystem. In the present study, we propose an integrated strategy to evaluate the toxicity of the leachate using chemical analyses, risk assessment guidelines and in vitro assays using the hepatoma HepG2 cells as a model. The approach was applied on a real case study: an industrial waste landfill in northern Italy for which data on the presence of leachate contaminants are available from the last 11 years. Results from our ecological risk models suggest important toxic effects on freshwater fish and small rodents, mainly due to ammonia and inorganic constituents. Our results from in vitro data show an inhibition of cell proliferation by leachate at low doses and cytotoxic effect at high doses after 48 h of exposure. - Research highlights: {yields} We study the toxicity of leachate from a non-hazardous industrial waste landfill. {yields} We perform chemical analyses, risk assessments and in vitro assays on HepG2 cells. {yields} Risk models suggest toxic effects due to ammonia and inorganic constituents. {yields} In vitro assays show that leachate inhibits cell proliferation at low doses. {yields} Leachate can induce cytotoxic effects on HepG2 cells at high doses.« less

In vivo maternal and in vitro BPA exposure effects on hypothalamic neurogenesis and appetite regulators.

PubMed

Desai, Mina; Ferrini, Monica G; Han, Guang; Jellyman, Juanita K; Ross, Michael G

2018-07-01

In utero exposure to the ubiquitous plasticizer, bisphenol A (BPA) is associated with offspring obesity. As food intake/appetite is one of the critical elements contributing to obesity, we determined the effects of in vivo maternal BPA and in vitro BPA exposure on newborn hypothalamic stem cells which form the arcuate nucleus appetite center. For in vivo studies, female rats received BPA prior to and during pregnancy via drinking water, and newborn offspring primary hypothalamic neuroprogenitor (NPCs) were obtained and cultured. For in vitro BPA exposure, primary hypothalamic NPCs from healthy newborns were utilized. In both cases, we studied the effects of BPA on NPC proliferation and differentiation, including putative signal and appetite factors. Maternal BPA increased hypothalamic NPC proliferation and differentiation in newborns, in conjunction with increased neuroproliferative (Hes1) and proneurogenic (Ngn3) protein expression. With NPC differentiation, BPA exposure increased appetite peptide and reduced satiety peptide expression. In vitro BPA-treated control NPCs showed results that were consistent with in vivo data (increase appetite vs satiety peptide expression) and further showed a shift towards neuronal versus glial fate as well as an increase in the epigenetic regulator lysine-specific histone demethylase1 (LSD1). These findings emphasize the vulnerability of stem-cell populations that are involved in life-long regulation of metabolic homeostasis to epigenetically-mediated endocrine disruption by BPA during early life. Copyright © 2018. Published by Elsevier Inc.

Clinical disintegration time of orally disintegrating tablets clinically available in Japan in healthy volunteers.

PubMed

Yoshita, Tomohiro; Uchida, Shinya; Namiki, Noriyuki

2013-01-01

Disintegration time is an important characteristic of orally disintegrating tablets (ODTs), and evaluation of disintegration time is a key step in formulation development, manufacturing, and clinical practice. In this study, we aimed to clarify the clinical disintegration time of ODTs that are currently used clinically, and to evaluate its correlation with the in vitro disintegration time of ODTs which was measured using Tricorptester, a newly developed disintegration testing apparatus. The clinical disintegration time of 17 ODT products was measured in healthy volunteers (n=9-10; age range, 21-28 years). A randomized single-blind trial was performed; each tablet was placed on the tongues of the participants, and it disintegrated in their oral cavities. No significant difference was observed in the clinical disintegration time of each ODT among the 3 groups to which the subjects were randomly assigned. The clinical disintegration time of the 17 ODT products was between 17.6 s and 33.8 s. The in vitro disintegration time of 26 clinically used ODT products measured using Tricorptester ranged between 4.40 s and 30.4 s. A significant positive correlation was observed between in vitro and clinical disintegration times (r=0.79; p<0.001). This study shows that all the tested products, which are clinically available in Japan, showed good disintegration and that the disintegration time varied according to the product. In addition, the in vitro disintegration time of ODTs measured using Tricorptester is a good reflection of the disintegration time in the oral cavity.

In vitro mechanical integrity of hydroxyapatite coated magnesium alloy.

PubMed

Kannan, M Bobby; Orr, Lynnley

2011-08-01

The mechanical integrity of resorbable implants during service, especially in load bearing orthopaedic applications, is critical. The high degradation rate of resorbable magnesium and magnesium-based implants in body fluid may potentially cause premature in-service failure. In this study, a magnesium alloy (AZ91) was potentiostatically coated with hydroxyapatite at different cathodic voltages in an attempt to enhance the mechanical integrity. The mechanical integrity of the uncoated and hydroxyapatite coated alloys was evaluated after in vitro testing of the coated samples in simulated body fluid (SBF). The uncoated alloy showed 40% loss in the mechanical strength after five days exposure to SBF. However, the hydroxyapatite coated alloy exposed to SBF showed 20% improvement in the mechanical strength as compared to that of the uncoated alloy. The alloy coated potentiostatically at -2 V performed better than the -3 V coated alloy. The cross-sectional analysis of the coatings revealed relatively uniform coating thickness for the -2 V coated alloy, whereas the -3 V coated alloy exhibited areas of uneven coating. This can be attributed to the increase in hydrogen evolution on the alloy during -3 V coating as compared to -2 V coating. The scanning electron micrographs of the in vitro tested alloy revealed that hydroxyapatite coating significantly reduced the localized corrosion of the alloy, which is critical for better in-service mechanical integrity. Thus, the study suggests that the in vitro mechanical integrity of resorbable magnesium-based alloy can be improved by potentiostatic hydroxyapatite coating. © 2011 IOP Publishing Ltd

The effect of formulation additives on in vitro dissolution-absorption profile and in vivo bioavailability of telmisartan from brand and generic formulations.

PubMed

Borbás, Enikő; Nagy, Zsombor K; Nagy, Brigitta; Balogh, Attila; Farkas, Balázs; Tsinman, Oksana; Tsinman, Konstantin; Sinkó, Bálint

2018-03-01

In this study, brand and four generic formulations of telmisartan, an antihypertensive drug, were used in in vitro simultaneous dissolution-absorption, investigating the effect of different formulation additives on dissolution and on absorption through an artificial membrane. The in vitro test was found to be sensitive enough to show even small differences between brand and generic formulations caused by the use of different excipients. By only changing the type of filler from sorbitol to mannitol in the formulation, the flux through the membrane was reduced by approximately 10%. Changing the salt forming agent as well resulted in approximately 20% of flux reduction compared to the brand formulation. This significant difference was clearly shown in the published in vivo results as well. The use of additional lactose monohydrate in the formulation also leads to approximately 10% reduction in flux. The results show that by changing excipients, the dissolution of telmisartan was not altered significantly, but the flux through the membrane was found to be significantly changed. These results pointed out the limitations of traditional USP dissolution tests and emphasized the importance of simultaneously measuring dissolution and absorption, which allows the complex effect of formulation excipients on both processes to be measured. Moreover, the in vivo predictive power of the simultaneous dissolution-absorption test was demonstrated by comparing the in vitro fluxes to in vivo bioequivalence study results. Copyright © 2018 Elsevier B.V. All rights reserved.

TAT improves in vitro transportation of fortilin through midgut and into hemocytes of white shrimp Litopenaeus vannamei

NASA Astrophysics Data System (ADS)

Zhou, Yi; Zhang, Wenbing; Mai, Kangsen; Xu, Wei; Zhang, Yanjiao; Ai, Qinghui; Wang, Xiaojie

2012-06-01

Fortilin is a multifunctional protein implicated in many important cellular processes. Since injection of Pm-fortilin reduces shrimp mortality caused by white spot syndrome virus (WSSV), there is potential application of fortilin in shrimp culture. In the present study, in order to improve trans-membrane transportation efficiency, the protein transduction domain of the transactivator of transcription (TAT) peptide was fused to fortilin. The Pichia pastoris yeast expression system, which is widely accepted in animal feeds, was used for production of recombinant fusion protein. Green fluorescence protein (GFP) was selected as a reporter because of its intrinsic visible fluorescence. The fortilin, TAT and GFP fusion protein were constructed. Their trans-membrane transportation efficiency and effects on immune response of shrimp were analyzed in vitro. Results showed that TAT peptide improved in vitro uptake of fortilin into the hemocytes and midgut of Litopenaeus vannamei. The phenoloxidase (PO) activity of hemocytes incubated with GFP-Fortilin or GFP-Fortilin-TAT was significantly increased compared with that in the control without expressed fortilin. The PO activity of hemocytes incubated with 200 μg mL-1 GFP-Fortilin-TAT was significantly higher than that in the group with the same concentration of GFP-Fortilin. Hemocytes incubated with GFP-Fortilin-TAT at all concentrations showed significantly higher nitric oxide synthase (NOS) activity than those in the control or in the GFP-Fortilin treatment. The present in vitro study indicated that TAT fusion protein improved the immune effect of fortilin.

In vitro digestion and absorption of BDE-28, -47, -99 and -153 in indoor dust and its implication in risk assessment.

PubMed

Kang, Yuan; Pan, Weijian; Liang, Siyun; Zeng, Lixuan; Zhang, Qiuyun; Luo, Jiwen; Guo, Xinmei

2016-12-01

The bioaccessibility of polybrominated diphenyl ethers (PBDEs) in indoor dust was estimated by a series of in vitro digestion methods. However, the absorption of PBDEs by intestinal cells after in vitro digestion was seldom studied. In the present study, the bioaccessibility of BDE-28, 47, 99 and 153 in indoor dust was firstly investigated by using the in vitro digestion method. Bioaccessibility in intestinal phase (BDE-28: 24.5-30.1%; BDE-47: 6.99-13.0; BDE-99: 1.61-14.2%; and BDE-153 5.97-24.4%.) was higher than that in gastric phase (BDE-28: 38.3-58.0; BDE-47: 9.62-30.9%; BDE-99: 9.71-24.3%; and BDE-153: 13.8-57.4%). The organic matter contents in indoor dust showed variable influence on the bioaccessibility of PBDEs. For the Caco-2 uptake assay, the BDE-28 showed greatest transport rate from medium to cell (K mc : 0.525h -1 ), followed by -47, -99 and -153. The K mc of PBDEs was significantly negative correlated with its corresponding K OW value. Similar pattern was found for the maximum uptake flux (J u, max ) and the transport rate from cell to medium (K cm ). The combination of bioacessibility and the absorption factor by Caco-2 cells could be used to estimate human intake of PBDEs via indoor dust would avoid overestimate the health risk. Copyright © 2016 Elsevier Inc. All rights reserved.

Pharmacokinetics and pharmacodynamics of CD4-anchoring bi-functional fusion inhibitor in monkeys.

PubMed

Liu, Xingrong; Ou, Ying C; Zhang, Jun; Ahene, Ago; Clark, Douglas; Hsieh, Su-Chun; Cooper, Matthew; Ji, Changhua

2014-03-01

This study was to characterize the pharmacokinetics (PK) and pharmacodynamics (PD) of a chimeric protein, CD4-anchoring bi-functional fusion inhibitor (CD4-BFFI), in monkeys and assess the feasibility for HIV-1 treatment in humans. The serum concentrations of CD4-BFFI and CD4 receptors were determined and modeled using a target-mediated drug disposition (TMDD) model following intravenous administration of 1 or 10 mg/kg in monkeys. In vitro CD4 internalization was examined in human peripheral blood mononuclear cells. Noncompartmental analysis showed a decrease in clearance (1.35 to 0.563 mL/h/kg) and an increase in half-lives (35 to 50 h) with increasing doses. Dose-dependent CD4 occupancy was observed. The TMDD model reasonably captured the PK/PD profiles and suggested greater degradation rate constant for the free CD4 than the bound CD4. In vitro assay showed CD4-BFFI did not reduce the internalization of cell surface CD4. The simulated serum concentrations of CD4-BFFI were 20-fold above its in vitro IC50 for HIV-1 at 3 mg/kg weekly or biweekly following subcutaneous administration in humans. The TMDD modeling and in vitro CD4 internalization study indicate that CD4-BFFI does not induce CD4 internalization and CD4-BFFI short half-life is likely due to normal CD4 internalization. The simulated human PK supports CD4-BFFI as a promising anti-HIV-1 agent.

Bioaccessibility of cadmium in fresh and cooked Agaricus blazei Murill assessed by in vitro biomimetic digestion system.

PubMed

Sun, Liping; Liu, Gaoxiang; Yang, Meizhizi; Zhuang, Yongliang

2012-05-01

Bioaccessibility of cadmium (Cd) in fresh and cooked Agaricus blazei Murill (AbM) was studied by an in vitro biomimetic digestion system in this paper. The results showed that the Cd content in fresh AbM was 10.27 mg kg(-1) DM. The cooking treatments of boiling and microwaving with water significantly decreased Cd contents in fresh AbM by 36.4% and 30.2% (P<0.05), respectively. Cd in fresh AbM showed the highest bioaccessibility of 77.8% during the biomimetic digestion in stomach, followed by that of 69.4% from the gastrointestinal digestion. Cooking treatments also significantly lowered the bioaccessibility of Cd (P<0.05). Cd in boiled AbM showed 50.7% and 46.1% bioaccessibility during the gastric and gastrointestinal procedures. While, Cd in microwaved AbM showed 58.2% and 50.4% bioaccessibility. This study confirmed that the health risk assessment of AbM depending on the total Cd levels in fresh AbM was inaccurate, especially for the products domestically cooked. Copyright © 2012 Elsevier Ltd. All rights reserved.

In vitro physical, chemical, and biological evaluation of commercially available metal orthodontic brackets.

PubMed

Kim, Joo Hyoung; Cha, Jung Yul; Hwang, Chung Ju

2012-12-01

This in vitro study was undertaken to evaluate the physical, chemical, and biological properties of commercially available metal orthodontic brackets in South Korea, because national standards for these products are lacking. FOUR BRACKET BRANDS WERE TESTED FOR DIMENSIONAL ACCURACY, (MANUFACTURING ERRORS IN ANGULATION AND TORQUE), CYTOTOXICITY, COMPOSITION, ELUTION, AND CORROSION: Archist (Daeseung Medical), Victory (3M Unitek), Kosaka (Tomy), and Confidence (Shinye Odontology Materials). The tested rackets showed no significant differences in manufacturing errors in angulation, but Confidence brackets showed a significant difference in manufacturing errors in torque. None of the brackets were cytotoxic to mouse fibroblasts. The metal ion components did not show a regular increasing or decreasing trend of elution over time, but the volume of the total eluted metal ions increased: Archist brackets had the maximal Cr elution and Confidence brackets appeared to have the largest volume of total eluted metal ions because of excessive Ni elution. Confidence brackets showed the lowest corrosion resistance during potentiodynamic polarization. The results of this study could potentially be applied in establishing national standards for metal orthodontic brackets and in evaluating commercially available products.

Muscle mitochondrial metabolism and calcium signaling impairment in patients treated with statins

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sirvent, P., E-mail: pascal.sirvent@univ-bpclermont.fr; CHRU Montpellier, 34295 Montpellier; Clermont Université, Université Blaise Pascal, EA 3533, Laboratoire des Adaptations Métaboliques à l'Exercice en conditions Physiologiques et Pathologiques

2012-03-01

The most common and problematic side effect of statins is myopathy. To date, the patho-physiological mechanisms of statin myotoxicity are still not clearly understood. In previous studies, we showed that acute application in vitro of simvastatin caused impairment of mitochondrial function and dysfunction of calcium homeostasis in human and rat healthy muscle samples. We thus evaluated in the present study, mitochondrial function and calcium signaling in muscles of patients treated with statins, who present or not muscle symptoms, by oxygraphy and recording of calcium sparks, respectively. Patients treated with statins showed impairment of mitochondrial respiration that involved mainly the complexmore » I of the respiratory chain and altered frequency and amplitude of calcium sparks. The muscle problems observed in statin-treated patients appear thus to be related to impairment of mitochondrial function and muscle calcium homeostasis, confirming the results we previously reported in vitro. -- Highlights: ► The most common and problematic side effect of statins is myopathy. ► Patients treated with statins showed impairment of mitochondrial respiration. ► Statins-treated patients showed altered frequency and amplitude of calcium sparks.« less

Medicinal activities of the leaves of Musa sapientum var. sylvesteris in vitro

PubMed Central

Sahaa, Repon Kumer; Acharyaa, Srijan; Shovon, Syed Sohidul Haque; Royb, Priyanka

2013-01-01

Objective This study is to investigate the medicinal value of methanolic extract of the leaves of Musa sapientum var. sylvesteris in Bangladesh. Methods Several biochemical assays, thin layer chormatogarphy and ultra-violet spectroscopy were used to detect the presence of various types of compounds in this extract. Antioxidant effects were measured by DPPH scavenging assay, total reducing assay and hydrogen peroxide scavenging assay. Receptor binding activities and hydrogen peroxide induced hemolysis assay were performed by hemagglutination assay and hemolysis assay using erythrocytes. Disk diffusion assay was performed to show the antibacterial effect of the extract. Results Methanolic extract of the leaves showed antioxidant and antibacterial activity in vitro. The extract showed hemaglutination inhibition activities and hydrogen peroxide induced hemolysis inhibition activity of human red blood cells. Conclusion Musa sapientum var. sylvesteris can be an useful medicinal plant. PMID:23730561

A study of the uptake and biodistribution of nano-titanium dioxide using in vitro and in vivo models of oral intake

NASA Astrophysics Data System (ADS)

MacNicoll, Alan; Kelly, Mick; Aksoy, Hatice; Kramer, Evelien; Bouwmeester, Hans; Chaudhry, Qasim

2015-02-01

Certain food additives may contain a sizeable fraction of particles in the nanoscale. However, little is known about the fate, behaviour and toxicological effects of orally-ingested nanoparticles. This study investigated the uptake and biodistribution of nano- and larger-sized titanium dioxide (TiO2) using an in vitro model of gut epithelium and in vivo in rat. The results of the in vivo study showed that oral administration of 5 mg/kg body weight of TiO2 nano- or larger particles did not lead to any significant translocation of TiO2 (measured as titanium) either to blood, urine or to various organs in rat at any of the time intervals studied over a 96 h post-administration period. Different methods used for dispersing particles did not affect the uptake, and orally administered TiO2 was found excreted in the faeces over a period of time. The in vitro study provided further evidence for the lack of translocation of TiO2 across the gut epithelium model. The overall evidence from both in vivo and in vitro studies did not support that oral ingestion of nano- or larger particles of TiO2 via food would result in any significant internal exposure of the consumer to the nanoparticles. The dietary TiO2 nanoparticles are likely to be excreted in the faeces.

Gelatin- and hydroxyapatite-based cryogels for bone tissue engineering: synthesis, characterization, in vitro and in vivo biocompatibility.

PubMed

Kemençe, Nevsal; Bölgen, Nimet

2017-01-01

The aim of this study was the synthesis and characterization of gelatin- and hydroxyapatite (osteoconductive component of bone)-based cryogels for tissue-engineering applications. Preliminary in vitro and in vivo biocompatibility tests were conducted. Gelatin- and hydroxyapatite-based cryogels of varying concentrations were synthesized using glutaraldehyde as the crosslinking agent. Chemical structure, pore morphology, pore size distribution, mechanical properties, swelling characteristics and degradation profiles of the synthesized cryogels were demonstrated by Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), mercury porosimetry, a mechanical test device, swelling ratio tests and weight loss measurements, respectively. In vitro cell viability and in vivo biocompatility tests were performed in order to show the performance of the cryogels in the biological environment. Changing the concentrations of gelatin, hydroxyapatite and crosslinker changed the chemical structure, pore size and pore size distribution of the cryogels, which in turn resulted in the ultimate behaviour (mechanical properties, swelling ratio, degradation profile). In vitro cell culture tests showed the viability of the cells. The cryogels did not show any cytotoxic effects on the cells. Clinical outcomes and the gross pathological results demonstrated that there was no necrosis noted in the abdominal and thoracic regions at the end of implantation and the implanted cryogel was found to be non-irritant and non-toxic at 12 weeks of implantation. Copyright © 2013 John Wiley & Sons, Ltd. Copyright © 2013 John Wiley & Sons, Ltd.

Mono-epoxy-tocotrienol-α enhances wound healing in diabetic mice and stimulates in vitro angiogenesis and cell migration.

PubMed

Xu, Cheng; Bentinger, Magnus; Savu, Octavian; Moshfegh, Ali; Sunkari, Vivekananda; Dallner, Gustav; Swiezewska, Ewa; Catrina, Sergiu-Bogdan; Brismar, Kerstin; Tekle, Michael

2017-01-01

Diabetes mellitus is characterized by hyperglycemia and capillary hypoxia that causes excessive production of free radicals and impaired antioxidant defense, resulting in oxidative stress and diabetes complications such as impaired wound healing. We have previously shown that modified forms of tocotrienols possess beneficial effects on the biosynthesis of the mevalonate pathway lipids including increase in mitochondrial CoQ. The aim of this study is to investigate the effects of mono-epoxy-tocotrienol-α on in vitro and in vivo wound healing models as well as its effects on mitochondrial function. Gene profiling analysis and gene expression studies on HepG2 cells and human dermal fibroblasts were performed by microarray and qPCR, respectively. In vitro wound healing using human fibroblasts was studied by scratch assay and in vitro angiogenesis using human dermal microvascular endothelial cells was studied by the tube formation assay. In vivo wound healing was performed in the diabetic db/db mouse model. For the study of mitochondrial functions and oxygen consumption rate Seahorse XF-24 was employed. In vitro, significant increase in wound closure and cell migration (p<0.05) both in normal and high glucose and in endothelial tube formation (angiogenesis) (p<0.005) were observed. Microarray profiling analysis showed a 20-fold increase of KIF26A gene expression and 11-fold decrease of lanosterol synthase expression. Expression analysis by qPCR showed significant increase of the growth factors VEGFA and PDGFB. The epoxidated compound induced a significantly higher basal and reserve mitochondrial capacity in both HDF and HepG2 cells. Additionally, in vivo wound healing in db/db mice, demonstrated a small but significant enhancement on wound healing upon local application of the compound compared to treatment with vehicle alone. Mono-epoxy-tocotrienol-α seems to possess beneficial effects on wound healing by increasing the expression of genes involved in cell growth, motility and angiogenes as well as on mitochondrial function. Copyright © 2017 Elsevier Inc. All rights reserved.

Preparation and characterization of oxybenzone-loaded gelatin microspheres for enhancement of sunscreening efficacy.

PubMed

Patel, M; Jain, Sunil K; Yadav, Awesh K; Gogna, D; Agrawal, G P

2006-01-01

The objective of our present study was to prepare and evaluate gelatin microspheres of oxybenzone to enhance its sunscreening efficacy. The gelatin microspheres of oxybenzone were prepared by emulsion method. Process parameters were analyzed to optimize the formulation. The in vitro drug release study was performed in pH 7.4 using cellulose acetate membrane. Microspheres prepared using oxybenzone:gelatin ratio of 1:6 showed slowest drug release and those prepared with oxybenzone:gelatin ratio of 1:2 showed fastest drug release. The gelatin microspheres of oxybenzone were incorporated in aloe vera gel. Sun exposure method using sodium nitroprusside solution was used for in vitro sunscreen efficacy testing. The formulation C5 containing oxybenzone-bearing gelatin microspheres in aloe vera gel showed best sunscreen efficacy. The formulations were evaluated for skin irritation test in human volunteers, sun protection factor, and minimum erythema dose in albino rats. These studies revealed that the incorporation of sunscreening agent-loaded microspheres into aloe vera gel greatly increased the efficacy of sunscreen formulation more than four times.

In vitro anthelmintic activity of active compounds of the fringed rue Ruta chalepensis against dairy ewe gastrointestinal nematodes.

PubMed

Ortu, E; Sanna, G; Scala, A; Pulina, G; Caboni, P; Battacone, G

2017-07-01

Infections by gastrointestinal nematodes negatively affect small ruminant health and at the same time cause substantial economic losses worldwide. Because resistance to conventional anthelmintic compounds is growing, target studies evaluating the effectiveness of alternative ingredients of botanical origin on gastrointestinal nematodes are needed. In this study, we evaluated the in vitro anthelmintic activity of Ruta chalepensis L. extracts on the third-stage larvae of sheep gastrointestinal nematodes. A methanol extract showed the highest anthelmintic activity, with an EC50 = 0.10 ± 0.06 mg/ml after 96 h, while the essential oil had an EC50 = 1.45 ± 1.22 mg/ml after 48 h. Moreover, three secondary metabolites of the essential oil, i.e. 2-decanone, 2-nonanone and 2-undecanone, showed EC50 values of 0.07 ± 0.06, 0.25 ± 0.29 and 0.88 ± 0.73 mg/ml at 24 h, respectively. The present study indicated that the R. chalepensis methanol extract, the essential oil and its metabolites 2-decanone, 2-nonanone and 2-undecanone showed promising anthelmintic activity on gastrointestinal nematodes.

Polymeric nanoparticles containing diazepam: preparation, optimization, characterization, in-vitro drug release and release kinetic study

NASA Astrophysics Data System (ADS)

Bohrey, Sarvesh; Chourasiya, Vibha; Pandey, Archna

2016-03-01

Nanoparticles formulated from biodegradable polymers like poly(lactic-co-glycolic acid) (PLGA) are being extensively investigated as drug delivery systems due to their two important properties such as biocompatibility and controlled drug release characteristics. The aim of this work to formulated diazepam loaded PLGA nanoparticles by using emulsion solvent evaporation technique. Polyvinyl alcohol (PVA) is used as stabilizing agent. Diazepam is a benzodiazepine derivative drug, and widely used as an anticonvulsant in the treatment of various types of epilepsy, insomnia and anxiety. This work investigates the effects of some preparation variables on the size and shape of nanoparticles prepared by emulsion solvent evaporation method. These nanoparticles were characterized by photon correlation spectroscopy (PCS), transmission electron microscopy (TEM). Zeta potential study was also performed to understand the surface charge of nanoparticles. The drug release from drug loaded nanoparticles was studied by dialysis bag method and the in vitro drug release data was also studied by various kinetic models. The results show that sonication time, polymer content, surfactant concentration, ratio of organic to aqueous phase volume, and the amount of drug have an important effect on the size of nanoparticles. Hopefully we produced spherical shape Diazepam loaded PLGA nanoparticles with a size range under 250 nm with zeta potential -23.3 mV. The in vitro drug release analysis shows sustained release of drug from nanoparticles and follow Korsmeyer-Peppas model.

In vitro comparison of antimicrobial activity of aqueous decoction of Coriandrum sativum, and Dentol Drop with chlorhexidine on Streptococcus mutans

PubMed Central

Moradian, Hamid; Bazargani, Abdollah; Rafiee, Azade; Nazarialam, Ali

2013-01-01

Background and objectives Dental caries is still remained as a major health problem. This problem has created a new interest to search for new antimicrobial agents from various sources including medicinal plants. Since limited data is available so far regarding the antibacterial effect of Coriandrum sativum seed and Dentol Drop against Streptococcus mutans, this study aims to assess this activity. Materials and Methods This experimental study was conducted in Shiraz University of Medical Sciences. In vitro comparison of antimicrobial activity of aqueous decoction of Coriandrum sativum seed and Dentol drop with chlorhexidine against Streptococcus mutans was evaluated using disk diffusion and broth microdilution assays. Positive and negative controls were considered. The data was statistically analyzed by applying Kruskal-Wallis and Tukey post-hoc test to compare the groups using SPSS software (version 17). Results Dentol drop showed a remarkable antibacterial activity, in comparison with chlorhexidine, against S. mutans in the disk diffusion (p value = 0.005), and broth microdilution assays (p value = 0.0001). Based on the results of this study, Coriandrum sativum seed did not posses any antibacterial property. Conclusion Coriandrum sativum seed showed no anti-Streptococcus mutans activity. Dentol drop exhibited a remarkable antibacterial activity against S. mutans when tested in vitro. Dentol drop can be further studied as a preventive measure for dental caries. PMID:24475330

Endoscopic diode-laser applications in airway surgery

NASA Astrophysics Data System (ADS)

Pankratov, Michail M.; Wang, Zhi; Rebeiz, Elie E.; Perrault, Donald F., Jr.; Shapshay, Stanley M.; Gleich, Lyon L.

1994-09-01

A technique was developed to secure small mucosal grafts onto the airway wound with fibrin/albumin tissue adhesive mixed with ICG dye and irradiated with a 810 nm diode laser. An in vitro study of the tensile strength produced strong mucosal soldering which was adequate to fix grafts in place. In vivo studies showed that wounds with mucosal grafts were completely covered by regenerated squamous cells in 1 week and by ciliated epithelium in 2 weeks. Excellent healing was observed at 6 and 14 days postoperatively and the histology at 28 days found normal epithelium over the vocal cord lesion. This soldering technique is a less traumatic treatment for patients with extensive lesions of the larynx of various origin. Diode laser soldering with ICG-doped fibrin tissue adhesive was evaluated in tracheal anastomosis as a substitute for absorbable sutures. In vitro studies demonstrated strong anastomoses with minimal tissue damage. In vivo animal study showed that these anastomoses had less fibrosis and tissue damage than control animals repaired with sutures only.

(68)Ga-DOTATATE PET in juvenile angiofibroma.

PubMed

Gronkiewicz, Zuzanna; Kukwa, Wojciech; Krolicki, Leszek; Cyran-Chlebicka, Agata; Pawlak, Dariusz; Stankiewicz, Czeslaw; Krzeski, Antoni; Górnicka, Barbara; Wolosz, Dominika; Kunikowska, Jolanta

2016-06-01

As somatostatin receptors (SSTRs) may be overexpressed in rapidly growing vessels, the aim of this study was the analysis of in vivo and in vitro SSTR2A expression in juvenile angiofibroma (JA). A group of six male adolescents with a diagnosis of primary, recurrent/residual JA was enrolled in the study. All patients underwent (68)Ga-DOTATATE PET/computed tomography (CT) followed by immunohistochemical staining for SSTR expression. (68)Ga-DOTATATE PET/CT showed accumulation in areas matching the pathologic tissue in the nasopharynx of all patients studied with SUVmax of 5.1 ± 0.9 (ranging from 3.6 to 6.4). In all cases, the immunohistochemical examination showed a presence of SSTR2A with a high staining index. In vitro SSTR2A cytoplasm expression was found to be high in all tumor specimens. However, the uptake of (68)Ga-DOTATATE was weak in the PET/CT studies. We postulate that the intracellular localization of the SSTR2A in JA may cause this discrepancy.

Modulation of ovarian steroidogenesis by adiponectin during delayed embryonic development of Cynopterus sphinx.

PubMed

Anuradha; Krishna, Amitabh

2014-09-01

The aim of present study was to evaluate role of adiponectin in ovarian steroidogenesis during delayed embryonic development of Cynopterus sphinx. This study showed significantly low circulating adiponectin level and a decline in expression of adiponectin receptor 1 (AdipoR1) in the ovary during the period of delayed embryonic development as compared with the normal development. The adiponectin treatment in vivo during the period of delayed development caused significantly increased in circulating progesterone and estradiol levels together with increased expression of AdipoR1 in the ovary. The in vitro study confirmed the stimulatory effect of adiponectin on progesterone synthesis. Both in vivo and in vitro studies showed that the effects of adiponectin on ovarian steroidogenesis were mediated through increased expression of luteinizing hormone-receptor, steroidogenic acute regulatory protein and 3β-hydroxyl steroid dehydrogenase enzyme. The adiponectin treatment may also promote progesterone synthesis by modulating ovarian angiogenesis, cell survival and rate of apoptosis. Copyright © 2014 Elsevier Ltd. All rights reserved.

In vitro activity of novel anti-MRSA cephalosporins and comparator antimicrobial agents against staphylococci involved in prosthetic joint infections.

PubMed

Isnard, Christophe; Dhalluin, Anne; Malandain, Damasie; Bruey, Quentin; Auzou, Michel; Michon, Jocelyn; Giard, Jean-Christophe; Guérin, François; Cattoir, Vincent

2018-02-05

Ceftaroline and ceftobiprole are new parenteral cephalosporins with potent activity against methicillin-resistant (MR) staphylococci, which are the leading cause of prosthetic joint infections (PJIs). The aim of this study was to determine and compare the in vitro activities of both molecules against staphylococcal isolates recovered from clinically documented PJIs. A collection of 200 non-duplicate clinical isolates [100 Staphylococcus aureus and 100 coagulase-negative staphylococci (CoNS), including 19 and 27 MR isolates, respectively] was studied. Minimum inhibitory concentrations (MICs) of oxacillin, ceftaroline, ceftobiprole, vancomycin, teicoplanin, clindamycin, levofloxacin, linezolid and daptomycin were determined by the broth microdilution method. Bactericidal activity (at 4× MIC) of ceftaroline, ceftobiprole, vancomycin, teicoplanin, linezolid and daptomycin was assessed by time-kill assay. Among the S. aureus isolates, 100% were susceptible to ceftaroline (MIC 50/90 , 0.25/0.5μg/mL) and 98% were susceptible to ceftobiprole (MIC 50/90 , 0.5/1μg/mL), regardless of their methicillin resistance. The two ceftobiprole-non-susceptible strains (including one MRSA) showed MICs at 4mg/L. Against CoNS isolates, ceftaroline and ceftobiprole exhibited in vitro potency with MIC 50/90 values at 0.06/0.25μg/mL and 0.25/1μg/mL, respectively. At 4× MIC, ceftaroline and ceftobiprole showed rapid and marked bactericidal activity against both S. aureus and CoNS (after 24/12h and 12/6h of incubation, respectively), whilst none of the other molecules tested had a bactericidal effect by 24h. This study showed that ceftaroline and ceftobiprole have excellent in vitro activity against clinical isolates of staphylococci involved in PJIs. These molecules may therefore represent promising alternatives for the treatment of such infections. Copyright © 2018 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.

Functional cardiotoxicity assessment of cosmetic compounds using human-induced pluripotent stem cell-derived cardiomyocytes.

PubMed

Chaudhari, Umesh; Nemade, Harshal; Sureshkumar, Poornima; Vinken, Mathieu; Ates, Gamze; Rogiers, Vera; Hescheler, Jürgen; Hengstler, Jan Georg; Sachinidis, Agapios

2018-01-01

There is a large demand of a human relevant in vitro test system suitable for assessing the cardiotoxic potential of cosmetic ingredients and other chemicals. Using human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), we have already established an in vitro cardiotoxicity assay and identified genomic biomarkers of anthracycline-induced cardiotoxicity in our previous work. Here, five cosmetic ingredients were studied by the new hiPSC-CMs test; kojic acid (KJA), triclosan (TS), triclocarban (TCC), 2,7-naphthalenediol (NPT), and basic red 51 (BR51) based on cytotoxicity as well as ATP assays, beating rate, and genomic biomarkers to determine the lowest observed effect concentration (LOEC) and no observed effect concentration (NOEC). The LOEC for beating rate were 400, 10, 3, >400, and 3 µM for KJA, TS, TCC, NPT, and BR51, respectively. The corresponding concentrations for cytotoxicity or ATP depletion were similar, with the exception of TS and TCC, where the cardiomyocyte-beating assay showed positive results at non-cytotoxic concentrations. Functional analysis also showed that the individual compounds caused different effects on hiPSC-CMs. While exposure to KJA, TS, TCC, and BR51 induced significant arrhythmic beating, NPT slightly decreased cell viability, but did not influence beating. Gene expression studies showed that TS and NPT caused down-regulation of cytoskeletal and cardiac ion homeostasis genes. Moreover, TS and NPT deregulated genomic biomarkers known to be affected also by anthracyclines. The present study demonstrates that hiPSC-CMs can be used to determine LOECs and NOECs in vitro, which can be compared to human blood concentrations to determine margins of exposure. Our in vitro assay, which so far has been tested with several anthracyclines and cosmetics, still requires validation by larger numbers of positive and negative controls, before it can be recommended for routine analysis.

In vitro and in vivo evaluation of curcumin loaded lauroyl sulphated chitosan for enhancing oral bioavailability.

PubMed

Shelma, R; Sharma, Chandra P

2013-06-05

Curcumin has been demonstrated as a potent anticancer agent but its clinical application has been limited by its poor aqueous solubility and bioavailability. Here we describe encapsulation of curcumin in the lauroyl sulphated chitosan with a view to improve its bioavailability. In vitro antioxidant activity of extract of curcumin loaded matrix was investigated and exhibited dose dependent radical scavenging and reducing activity. Cytotoxicity studies carried out with curcumin loaded carrier on C6 cell line and were found to be toxic. Its in vitro effects on proliferation using the C6 cell lines also studied and observed antiproliferation of C6 cell line. Plasma concentration of curcumin-time profiles from pharmacokinetic studies in rats after oral administration showed a 11.5-fold increased pharmacological availability of curcumin with encapsulated curcumin compared with native curcumin. Overall we demonstrate that the curcumin loaded matrix has shown a superior pharmacological availability in vivo over curcumin. Copyright © 2013 Elsevier Ltd. All rights reserved.

In Vitro Antimicrobial Susceptibility Patterns of Blastocystis.

PubMed

Roberts, Tamalee; Bush, Stephen; Ellis, John; Harkness, John; Stark, Damien

2015-08-01

Blastocystis is the most common human enteric protist with controversial clinical significance. Metronidazole is considered a first-line treatment for Blastocystis infection; however, there has been increasing evidence for the lack of efficacy of this treatment. Treatment failure has been reported in several clinical cases, and recent in vitro studies have suggested the occurrence of metronidazole-resistant strains. In this study, we tested 12 Blastocystis isolates from 4 common Blastocystis subtypes (ST1, ST3, ST4, and ST8) against 12 commonly used antimicrobials (metronidazole, paromomycin, ornidazole, albendazole, ivermectin, trimethoprim-sulfamethoxazole [TMP-SMX], furazolidone, nitazoxanide, secnidazole, fluconazole, nystatin, and itraconazole) at 10 different concentrations in vitro. It was found that each subtype showed little sensitivity to the commonly used metronidazole, paromomycin, and triple therapy (furazolidone, nitazoxanide, and secnidazole). This study highlights the efficacy of other potential drug treatments, including trimethoprim-sulfamethoxazole and ivermectin, and suggests that current treatment regimens be revised. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Transferrin-appended PEGylated nanoparticles for temozolomide delivery to brain: in vitro characterisation.

PubMed

Jain, Aviral; Chasoo, Gousia; Singh, Shashank K; Saxena, Ajit K; Jain, Sanjay K

2011-01-01

Polymer-based nanotechnologies are proposed to be an alternative for drug administration, delivery and targeting to those of conventional formulations. The blood brain barrier is frequently a rate-limiting factor in determining permeation of a drug into brain. In this study, the surface-engineered long-circulating PLGA nanoparticles (NPs) were assessed for brain-specific delivery. Long circulating NPs of PLGA- and PEG-synthesised copolymer were prepared by emulsification solvent evaporation method. Further, the surface of PEGylated NPs was modified by anchoring transferrin (Tf) ligand for receptor-mediated targeting to brain. NPs were characterised for shape and size, zeta potential, entrapment efficiency and in vitro drug release. In vitro cytotoxicity studies were performed on human cancer cell lines. Confocal Laser Scanning Microscopy studies show the enhanced uptake of Tf-appended PEGylated NPs and their localisation in the brain tissues. Hence, the specific role of Tf ligand on PEGylated NPs for brain delivery was confirmed.

Assessment of Aprotinin Loaded Microemulsion Formulations for Parenteral Drug Delivery: Preparation, Characterization, in vitro Release and Cytotoxicity Studies.

PubMed

Okur, Neslihan Üstündağ; Özdemir, Derya İlem; Kahyaoğlu, Şennur Görgülü; Şenyiğit, Zeynep Ay; Aşıkoğlu, Makbule; Genç, Lütfi; Karasulu, H Yeşim

2015-01-01

The object of the current study was to prepare novel microemulsion formulations of aprotinin for parenteral delivery and to compare in vitro characteristics and release behaviour of different Technetium-99m ((99m)Tc)-Aprotinin loaded microemulsion formulations. In addition, cytotoxicity of microemulsion formulation was evaluated with cell culture studies on human immortalized pancreatic duct epithelial-like cells. For this aim, firstly, pseudo-ternary phase diagrams were plotted to detect the formulation region and optimal microemulsions were characterized for their thermodynamic stability, conductivity, particle size, zeta potential, viscosity, pH and in vitro release properties. For in vitro release studies aprotinin was labelled with (99m)Tc and labelling efficiency, radiochemical purity and stability of the radiolabeled complex were determined by several chromatography techniques. Radiolabeling efficiency of (99m)Tc-Aprotinin was found over than 90% without any significant changes up to 6 hours after labelling at room temperature. After that, in vitro release studies of (99m)Tc-Aprotinin loaded microemulsions were performed with two different methods; dissolution from diffusion cells and dialysis bags. Both methods showed that release rate of (99m)Tc- Aprotinin from microemulsion could be controlled by microemulsion formulations. Drug release from the optimized microemulsion formulations was found lower compared to drug solution at the end of six hours. According to stability studies, the optimized formulation was found to be stable over a period of 12 months. Also, human immortalized pancreatic duct epithelial-like cells were used to evaluate the cytotoxicity of optimum formulation. Developed microemulsion did not reveal cytotoxicity. In conclusion the present study indicated that the M1-APT microemulsion is appropriate for intravenous application of aprotinin.

Survival of human pre-antral follicles after cryopreservation of ovarian tissue, follicular isolation and in vitro culture in a calcium alginate matrix.

PubMed

Amorim, Christiani A; Van Langendonckt, Anne; David, Anu; Dolmans, Marie-Madeleine; Donnez, Jacques

2009-01-01

Ovarian tissue cryopreservation is a promising technique to safeguard fertility in cancer patients. However, in some types of cancer, there is a risk of transmitting malignant cells present in the cryopreserved tissue. To avoid such a risk, pre-antral follicles could be isolated from ovarian tissue and grown in vitro. On the basis of this assumption, the aim of our study was to investigate in vitro survival and growth of pre-antral follicles after cryopreservation of ovarian tissue and follicular isolation, followed by encapsulation in alginate beads. Ovarian biopsies from four patients were frozen and thawed. Pre-antral follicles were then isolated and embedded in an alginate matrix before in vitro culture for 7 days. Small pre-antral follicles (42.98 +/- 9.06 microm) from frozen-thawed tissue can survive and develop after enzymatic isolation and in vitro culture. A total of 159 follicles were incubated in a three-dimensional system (alginate hydrogel) and, after 7 days, all of them showed an increase in size (final size 56.73 +/- 13.10 microm). The survival rate of the follicles was 90% (oocyte and all granulosa cells viable). Our preliminary results indicate that alginate hydrogels may be a suitable system for in vitro culture of isolated human pre-antral follicles. However, more studies are required to establish whether follicular morphology and functionality can be maintained using this matrix.

In vitro-in vivo activity relationship of substituted benzimidazole cell division inhibitors with activity against Mycobacteria tuberculosis

PubMed Central

Knudson, Susan E.; Kumar, Kunal; Awasthi, Divya; Ojima, Iwao; Slayden, Richard A.

2014-01-01

Structure based drug design was used to develop a compound library of novel 2,5,6- and 2,5,7-trisubstituted benzimidazoles. Three structural analogs, SB-P1G10, SB-P8B2 and SB-P3G2 were selected from this library based on previous studies for advanced study. In vitro studies revealed that SB-P8B2 and SB-P3G2 had sigmoidal kill-curves while in contrast SB-P1G10 showed a narrow zonal susceptibility. The in vitro studies also demonstrated that exposure to SB-P8B2 or SB-P3G2 was bactericidal, while SB-P1G10 treatment never resulted in complete killing. The dose curves for the three compounds against clinical isolates were comparable to their respective dose curves in the laboratory strain of M. tuberculosis. SB-P8B2 and SB-P3G2 exhibited antibacterial activity against non-replicating bacilli under low oxygen conditions. SB-P3G2 and SB-P1G10 were assessed in acute short-term animal models of tuberculosis, which showed that SB-P3G2 treatment demonstrated activity against M. tuberculosis. Together, these studies reveal an in vitro- in vivo relationship of the 2,5,6-trisubstituted benzimidazoles that serves as a criterion for advancing this class of cell division inhibitors into more resource intensive in vivo efficacy models such as the long-term murine model of tuberculosis and Pre-IND PK/PD studies. Specifically, these studies are the first demonstration of efficacy and an in vitro–in vivo activity relationship for 2,5,6-trisubstituted benzimidazoles. The in vivo activity presented in this manuscript substantiates this class of cell division inhibitors as having potency and efficacy against M. tuberculosis. PMID:24746463

The effect of pH on cell viability, cell migration, cell proliferation, wound closure, and wound reepithelialization: In vitro and in vivo study.

PubMed

Kruse, Carla R; Singh, Mansher; Targosinski, Stefan; Sinha, Indranil; Sørensen, Jens A; Eriksson, Elof; Nuutila, Kristo

2017-04-01

Wound microenvironment plays a major role in the process of wound healing. It contains various external and internal factors that participate in wound pathophysiology. The pH is an important factor that influences wound healing by changing throughout the healing process. Several previous studies have investigated the role of pH in relation to pathogens but studies concentrating on the effects of pH on wound healing itself are inconclusive. The purpose of this study was to comprehensively and in a controlled fashion investigate the effect of pH on wound healing by studying its effect on human primary keratinocyte and fibroblast function in vitro and on wound healing in vivo. In vitro, primary human keratinocytes and fibroblasts were cultured in different levels of pH (5.5-12.5) and the effect on cell viability, proliferation, and migration was studied. A rat full-thickness wound model was used to investigate the effect of pH (5.5-9.5) on wound healing in vivo. The effect of pH on inflammation was monitored by measuring IL-1 α concentrations from wounds and cell cultures exposed to different pH environments. Our results showed that both skin cell types tolerated wide range of pH very well. They further demonstrated that both acidic and alkaline environments decelerated cell migration in comparison to neutral environments and interestingly alkaline conditions significantly enhanced cell proliferation. Results from the in vivo experiments indicated that a prolonged, strongly acidic wound environment prevents both wound closure and reepithelialization while a prolonged alkaline environment did not have any negative impact on wound closure or reepithelialization. Separately, both in vitro and in vivo studies showed that prolonged acidic conditions significantly increased the expression of IL-1 α in fibroblast cultures and in wound fluid, whereas prolonged alkaline conditions did not result in elevated amounts of IL-1 α. © 2017 by the Wound Healing Society.

Effect of ambient light exposure of media and embryos on development and quality of porcine parthenogenetically activated embryos.

PubMed

Li, Rong; Liu, Ying; Pedersen, Hanne Skovsgaard; Callesen, Henrik

2015-06-01

Light exposure is a common stress factor during in vitro handling of oocytes and embryos that originates from both microscope and ambient light. In the current study, the effect of two types of ambient light (daylight and laboratory light) on porcine parthenogenetically activated (PA) embryos was tested in two experiments: (1) ambient light on medium subsequently used for embryo in vitro development; and (2) ambient light exposure on activated oocytes before in vitro development. The results from Experiment 1 showed that exposure of culture medium to both types of ambient light decreased the percentage of blastocysts that showed good morphology, only after 24 h exposure. The results from Experiment 2 revealed a reduction in both blastocyst formation and quality when activated oocytes were exposed to both types of ambient light. This effect was seen after only 1 h exposure and increased with time. In conclusion, exposure to ambient light can be harmful to embryo development, both when medium is exposed for a long period of time and, to a greater extent, when the embryo itself is exposed for >1 h. In practice, it is therefore recommended to protect both culture medium and porcine embryos against ambient light during in vitro handling in the laboratory.

Control-release microcapsule of famotidine loaded biomimetic synthesized mesoporous silica nanoparticles: Controlled release effect and enhanced stomach adhesion in vitro.

PubMed

Li, Jing; Wang, Hongyu; Yang, Baixue; Xu, Lu; Zheng, Nan; Chen, Hongtao; Li, Sanming

2016-01-01

In the present work, control-release microcapsule of famotidine (FMT) loaded biomimetic synthesized mesoporous silica nanoparticles (B-MSNs) was developed, and controlled release effect and stomach adhesion of this formulation in vitro were mainly investigated. B-MSN was previously synthesized and it was amorphous mesoporous nanoparticles with helical channels. Cytotoxicity of B-MSN was studied using human breast cancer cells (MCF-7) and the result indicated that cytotoxicity of B-MSN can be neglected. After loading FMT into B-MSN, specific surface area, pore volume and pore diameter of B-MSN were obviously reduced. In vitro dissolution test showed that B-MSN had the ability to slow down FMT release for 15 min. In order to prolong controlled release effect and remained the advantage of B-MSN (improve drug stability due to its rigid silica framework), the combined application of control-release microcapsule (using cellulose and hydroxypropyl methylcellulose K15M as excipients) with B-MSN was designed. It was obvious that newly designed formulation significantly controlled FMT release with Fickian diffusion mechanism and showed enhanced stomach adhesion in vitro, which has significant value in widening the application of B-MSN in formulation design. Copyright © 2015 Elsevier B.V. All rights reserved.

Essential oils to control Botrytis cinerea in vitro and in vivo on plum fruits.

PubMed

Aminifard, Mohammad Hossein; Mohammadi, Samane

2013-01-01

The consequence of misusing chemical biocides in controlling pests and diseases has drawn the attention of policy makers to the development of methods potentially available in nature for this purpose. In the present study the inhibitory effects of black caraway, fennel and peppermint essential oils against Botrytis cinerea were tested at various concentrations in vitro and in vivo. The in vitro results showed that the growth of B. cinerea was completely inhibited by the application of black caraway and fennel oils at concentrations of 400 and 600 µL L⁻¹ respectively. The in vivo results indicated that black caraway, fennel and peppermint oils at all applied concentrations inhibited B. cinerea growth on plum fruits compared with the control. In addition, all three oils at higher concentrations showed positive effects on fruit quality characteristics such as titrable acidity, total soluble solids, carbohydrate content, pH and weight loss percentage. Thus the oils inhibited the infection of plum fruits by B. cinerea and increased their storage life. This research confirms the antifungal effects of black caraway, fennel and peppermint essential oils both in vitro and in vivo on plum fruits postharvest. Therefore these essential oils could be an alternative to chemicals to control postharvest phytopathogenic fungi on plum fruits. Copyright © 2012 Society of Chemical Industry.

Evaluation of in-vitro antibacterial activity and anti-inflammatory activity for different extracts of Rauvolfia tetraphylla L. root bark

PubMed Central

Ganga Rao, B.; Umamaheswara Rao, P.; Sambasiva Rao, E.; Mallikarjuna Rao, T.; Praneeth. D, V. S.

2012-01-01

Objective To assess the in-vitro antibacterial activity and anti-inflammatory activity of orally administered different extracts (Hydro-alcoholic, methanolic, ethyl acetate and hexane) of Rauvolfia tetraphylla (R. tetraphylla) root bark in Carrageenan induced acute inflammation in rats. Methods In-vitro antibacterial activity was evaluated for extracts against four Gram positive and four Gram negative bacteria by using cylinder plate assay. Hydro-alcoholic extract (70% v/v ethanol) at 200, 400 and 800 mg/kg doses and methanolic, ethyl acetate and hexane extracts at doses 100, 200 and 400 mg/kg were tested for anti-inflammatory activity in Carrageenan induced rat paw oedema model and paw thickness was measured every one hour up to 6 hrs. Results All extracts of R. tetraphylla root bark showed good zone of inhibition against tested bacterial strains. In Carrageenan induced inflammation model, hydro-alcoholic and methanolic extract of R. tetraphylla root bark at three different doses produced significant (P<0.001) reduction when compared to vehicle treated control group and hexane, ethyl acetate extracts. Conclusions In the present study extracts of R. tetraphylla root bark shows good in-vitro antibacterial activity and in-vivo anti-inflammatory activity in rats. PMID:23569853

Osteogenic Treatment Initiating a Tissue-Engineered Cartilage Template Hypertrophic Transition.

PubMed

Fu, J Y; Lim, S Y; He, P F; Fan, C J; Wang, D A

2016-10-01

Hypertrophic chondrocytes play a critical role in endochondral bone formation as well as the progress of osteoarthritis (OA). An in vitro cartilage hypertrophy model can be used as a platform to study complex molecular mechanisms involved in these processes and screen new drugs for OA. To develop an in vitro cartilage hypertrophy model, we treated a tissue-engineered cartilage template, living hyaline cartilaginous graft (LhCG), with osteogenic medium for hypertrophic induction. In addition, endothelial progenitor cells (EPCs) were seeded onto LhCG constructs to mimic vascular invasion. The results showed that osteogenic treatment significantly inhibited the synthesis of endostatin in LhCG constructs and enhanced expression of hypertrophic marker-collagen type X (Col X) and osteogenic markers, as well as calcium deposition in vitro. Upon subcutaneous implantation, osteogenic medium-treated LhCG constructs all stained positive for Col X and showed significant calcium deposition and blood vessel invasion. Col X staining and calcium deposition were most obvious in osteogenic medium-treated only group, while there was no difference between EPC-seeded and non-seeded group. These results demonstrated that osteogenic treatment was of the primary factor to induce hypertrophic transition of LhCG constructs and this model may contribute to the establishment of an in vitro cartilage hypertrophy model.

The antibacterial activity of syringopicroside, its metabolites and natural analogues from Syringae Folium.

PubMed

Zhou, Zhengyuan; Han, Na; Liu, Zhihui; Song, Zehai; Wu, Peng; Shao, Jingxuan; Zhang, Jia Ming; Yin, Jun

2016-04-01

In the present study, the in vitro antibacterial activity of an effective fraction (ESF) from Syringae Folium (SF) on Methicillin-resistant Staphylococcus aureus (MRSA) was evaluated and then its in vivo activity was evaluated by using the MRSA-infected mouse peritonitis model. The ESF showed a significant in vitro and in vivo activity on decreasing the Minimum Inhibitory Concentrations (MICs) and increasing the survival rate of mouse from 42.8% to 100%. Six iridoid glucosides (IGs) of ESF were characterized by UPLC-TOF-MS method and also isolated by column chromatography. Most of them showed in vitro anti MRSA activity. Syringopicroside (Sy), the major compound of IGs, was found to increase the survival rate from 42.8% to 92.8% of the MRSA-infected mouse, which revealed Sy is also the main active components of ESF. In order to know why the effect of oral administration of SF is better than its injections in clinic and the metabolites of Sy, seven metabolites of Sy were isolated from rat urine and identified on the basis of NMR and MS spectra. Most of metabolites possessed stronger in vitro anti-MRSA activity than that of Sy, which furtherly proved the clinical result. Copyright © 2016 Elsevier B.V. All rights reserved.

Resveratrol-Loaded Lipid Nanocarriers: Correlation between In Vitro Occlusion Factor and In Vivo Skin Hydrating Effect

PubMed Central

Parenti, Carmela; Turnaturi, Rita

2017-01-01

Lipid nanocarriers show occlusive properties that may be related to their ability to improve skin hydration. The aim of this work was to evaluate the relationship between in vitro occlusion factor and in vivo skin hydration for three types of lipid nanocarriers: nanoemulsions (NEs), solid lipid nanoparticles (SLNs) and nanostructured lipid carriers (NLCs). These lipid nanocarriers were loaded with trans-resveratrol (RSV) and incorporated in gel vehicles. In vitro occlusion factor was in the order SLNs > NLCs > NEs. Gels containing unloaded or RSV loaded lipid nanocarriers were applied on the back of a hand of 12 healthy volunteers twice a day for one week, recording skin hydration changes using the instrument Soft Plus. An increase of skin hydration was observed for all lipid nanocarriers (SLNs > NLCs > NEs). RSV loading into these nanocarriers did not affect in vitro and in vivo lipid nanocarriers effects. A linear relationship (r2 = 0.969) was observed between occlusion factor and in vivo increase of skin hydration. Therefore, the results of this study showed the feasibility of using the occlusion factor to predict in vivo skin hydration resulting from topical application of different lipid nanocarriers loading an active ingredient with no inherent hydrating activity. PMID:29232856

Influence of Aloe vera on water absorption and enzymatic in vitro degradation of alginate hydrogel films.

PubMed

Pereira, Rúben F; Carvalho, Anabela; Gil, M H; Mendes, Ausenda; Bártolo, Paulo J

2013-10-15

This study investigates the influence of Aloe vera on water absorption and the in vitro degradation rate of Aloe vera-Ca-alginate hydrogel films, for wound healing and drug delivery applications. The influence of A. vera content (5%, 15% and 25%, v/v) on water absorption was evaluated by the incubation of the films into a 0.1 M HCl solution (pH 1.0), acetate buffer (pH 5.5) and simulated body fluid solution (pH 7.4) during 24h. Results show that the water absorption is significantly higher for films containing high A. vera contents (15% and 25%), while no significant differences are observed between the alginate neat film and the film with 5% of A. vera. The in vitro enzymatic degradation tests indicate that an increase in the A. vera content significantly enhances the degradation rate of the films. Control films, incubated in a simulated body fluid solution without enzymes, are resistant to the hydrolytic degradation, exhibiting reduced weight loss and maintaining its structural integrity. Results also show that the water absorption and the in vitro degradation rate of the films can be tailored by changing the A. vera content. Copyright © 2013 Elsevier Ltd. All rights reserved.

Effects of humic acids in vitro.

PubMed

Vašková, Janka; Veliká, Beáta; Pilátová, Martina; Kron, Ivan; Vaško, Ladislav

2011-06-01

Humic acids are known for their overall positive health and productivity effects in animal feeding trials and, controversially, as an aetiological factor of cancer. We tried to assess the in vitro effect of humic acids from a selected source in Slovakia when used at recommended prophylactic dosage. We investigated antioxidant properties, enzymatic and non-enzymatic antioxidant defence system in liver mitochondria and cultured cancer cell lines in vitro. We observed a significant decrease in superoxide dismutase activity after humic acids treatment irrespective of dissolving in dimethyl sulphoxide or direct addition to mitochondria suspension in a respiration medium. Activities of other antioxidant enzymes measured, such as glutathione peroxidase and glutathione reductase, showed no significant differences from the control as well as the reduced glutathione content. Percentage of inhibition by humic acids of superoxide radical indicated lower efficacy compared with that of hydroxyl radical. Survival of six different cancer cells lines indicated that only the acute T lymphoblastic leukaemia cell line was sensitive to the tested humic acids. Despite relatively low solubility in aqueous solutions, humic acids from the selected source participated in redox regulation. By recapturing the radicals, humic acids reloaded the antioxidant defensive mechanism. Results from in vitro study conducted with humic acids from the natural source showed potential of these substances as promising immunity enhancing agents.

In vitro/in vivo characterization of nanoemulsion formulation of metronidazole with improved skin targeting and anti-rosacea properties.

PubMed

Yu, Meng; Ma, Huixian; Lei, Mingzhu; Li, Nan; Tan, Fengping

2014-09-01

Topical skin treatment was limited due to the lack of suitable delivery system with significant cutaneous localization and systemic safety. The aim of this study was to develop and optimize a nanoemulsion (NE) to enhance targeting localization of metronidazole (MTZ) in skin layers. In vitro studies were used to optimize NE formulations, and a series of experiments were carried in vitro and in vivo to validate the therapeutic efficacy of MTZ-loaded optimal NE. NE type selection and D-optimal design study were applied to optimize NE formulation with maximum skin retention and minimum skin penetration. Three formulation variables: Oil X1 (Labrafil), Smix X2 (a mixture of Cremophor EL/Tetraethylene glycol, 2:1 w/w) and water X3 were included in D-design. The system was assessed for skin retention Y1, cumulative MTZ amount after 24 h Y2 and droplet size Y3. Following optimization, the values of formulation components (X1, X2 and X3) were 4.13%, 16.42% and 79.45%, respectively. The optimized NE was assessed for viscosity, droplet size, morphological study and in vitro permeation in pig skin. Distributions of MTZ were validated by confocal laser scanning microscopy (CLSM). Active agent of NE transferred into deeper skin and localized in epidermal/dermal layers after 24 h, which showed significant advantages of the optimal NE over Gel. The skin targeting localization and minimal systemic escape of optimal NE was further proved by in vivo study on rat skin. Current in vitro-in vivo correlation (IVIVC) enabled the prediction of pharmacokinetic profile of MTZ from in vitro permeation results. Further, the in vivo anti-rosacea efficacy of optimal formulation was investigated by pharmacodynamics study on mice ear. Copyright © 2014 Elsevier B.V. All rights reserved.

Studies on penetration of antibiotic in bacterial cells in space conditions (7-IML-1)

NASA Technical Reports Server (NTRS)

Tixador, R.

1992-01-01

The Cytos 2 experiment was performed aboard Salyut 7 in order to test the antibiotic sensitivity of bacteria cultivated in vitro in space. An increase of the Minimal Inhibitory Concentration (MIC) in the inflight cultures (i.e., an increase of the antibiotic resistance) was observed. Complementary studies of the ultrastructure showed a thickening of the cell envelope. In order to confirm the results of the Cytos 2 experiment, we performed the ANTIBIO experiment during the D1 mission to try to differentiate, by means of the 1 g centrifuge in the Biorack, between the biological effects of cosmic rays and those caused by microgravity conditions. The originality of this experiment was in the fact that it was designed to test the antibiotic sensitivity of bacteria cultivated in vitro during the orbital phase of the flight. The results show an increase in resistance to Colistin in in-flight bacteria. The MIC is practically double in the in-flight cultures. A cell count of living bacteria in the cultures containing the different Colistin concentrations showed a significant difference between the cultures developed during space flight and the ground based cultures. The comparison between the 1 g and 0 g in-flight cultures show similar behavior for the two sets. Nevertheless, a small difference between the two sets of ground based control cultures was noted. The cultures developed on the ground centrifuge (1.4 g) present a slight decrease in comparison with the cultures developed in the static rack (1 g). In order to approach the mechanisms of the increase of antibiotic resistance on bacteria cultivated in vitro in space, we have proposed the study on penetration of antibiotics in bacterial cells in space conditions. This experiment was selected for the International Microgravity Laboratory 1 (IML-1) mission.

Evaluation of anti-diabetic and anti-tumoral activities of bioactive compounds from Phoenix dactylifera L's leaf: In vitro and in vivo approach.

PubMed

Chakroun, Mouna; Khemakhem, Bassem; Mabrouk, Hazem Ben; El Abed, Hanen; Makni, Mohamed; Bouaziz, Mohamed; Drira, Noureddine; Marrakchi, Naziha; Mejdoub, Hafedh

2016-12-01

Among various chronic disorders, cancer and diabetes mellitus are the most common disorders. This study was designed to evaluate the effectiveness of hydroalcoholic extract of Phoenix dactylifera L. leaves (HEPdL) in animal models of type II diabetes in vitro/in vivo and in a human melanoma-derived cell line (IGR-39). A liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis was also performed to determine the amount of phenolic and flavonoid compounds in this plant. The physicochemical results by LC-MS/MS analysis of HEPdL showed the presence of 10 phenolic compounds. The in vitro study showed that the extract exhibited a more specific and potent inhibitor of α-glucosidase than α-amylase with an IC 50 value of 20±1μg/mL and 30±0.8μg/mL, respectively. More importantly, the in vivo study of the postprandial hyperglycemia activity with (20mg/kg) of HEPdL showed a decrease in plasma glucose levels after 60min in resemblance to the glucor (acarbose) (50mg/kg) effect. The oral administration of HEPdL (20mg/kg) in alloxan-induced diabetic mices for 28days showed a more significant anti-diabetic activity than that of the drug (50mg/kg). Moreover, cytotoxicity effects of HEPdL in IGR-39 cancer cell lines were tested by MTT assay. This extract was effective in inhibiting cancer cells growth (IGR-39) at dose 35 and 75μg/mL. These results confirm ethnopharmacological significance of the plant and could be taken further for the development of an effective pharmaceutical drug against diabetes and cancer. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Genotoxicity of two heavy metal compounds: lead nitrate and cobalt chloride in Polychaete Perinereis cultrifera.

PubMed

Singh, Nisha; Bhagat, Jacky; Ingole, Baban S

2017-07-01

The present study explores the in vivo and in vitro genotoxic effects of lead nitrate, [Pb(NO 3 ) 2 ] a recognized environmental pollutant and cobalt chloride (CoCl 2 ), an emerging environmental pollutant in polychaete Perinereis cultrifera using comet assay. Despite widespread occurrence and extensive industrial applications, no previous published reports on genotoxicity of these compounds are available in polychaete as detected by comet assay. Polychaetes were exposed in vivo to Pb(NO 3 ) 2 (0, 100, 500, and 1000 μg/l) and CoCl 2 (0, 100, 300, and 500 μg/l) for 5 days. At 100 μg/l Pb(NO 3 ) 2 concentration, tail DNA (TDNA) values in coelomocytes were increase by 1.16, 1.43, and 1.55-fold after day 1, day 3, and day 5, whereas, OTM showed 1.12, 2.33, and 2.10-fold increase in in vivo. Pb(NO 3 ) 2 showed a concentration and time-dependent genotoxicity whereas CoCl 2 showed a concentration-dependent genotoxicity in in vivo. A concentration-dependent increase in DNA damage was observed in in vitro studies for Pb(NO 3 ) 2 and CoCl 2 . DNA damage at 500 μg/L showed almost threefold increase in TDNA and approximately fourfold increase in OTM as compared to control in in vitro. Our studies suggest that Pb(NO 3 ) 2 and CoCl 2 have potential to cause genotoxic damage, with Pb(NO 3 ) 2 being more genotoxic in polychaete and should be used more carefully in industrial and other activities. Graphical abstract.

Triamcinolone acetonide activates an anti-inflammatory and folate receptor-positive macrophage that prevents osteophytosis in vivo.

PubMed

Siebelt, Michiel; Korthagen, Nicoline; Wei, Wu; Groen, Harald; Bastiaansen-Jenniskens, Yvonne; Müller, Christina; Waarsing, Jan Hendrik; de Jong, Marion; Weinans, Harrie

2015-12-05

Triamcinolone acetonide (TA) is used for osteoarthritis management to reduce pain, and pre-clinical studies have shown that TA limits osteophyte formation. Osteophyte formation is known to be facilitated by synovial macrophage activation. TA injections might influence macrophage activation and subsequently reduce osteophytosis. Although widely applied in clinical care, the mechanism through which TA exerts this effect remains unknown. In this animal study, we investigated the in vivo effects of TA injections on macrophage activation, osteophyte development and joint degeneration. Furthermore, in vitro macrophage differentiation experiments were conducted to further explain working mechanisms of TA effects found in vivo. Osteoarthritis was induced in rat knees using papain injections and a running protocol. Untreated and TA-treated animals were longitudinally monitored for 12 weeks with in vivo micro-computed tomography (μCT) to measure subchondral bone changes. Synovial macrophage activation was measured in vivo using folate receptor β (FRβ)-targeted single-photon emission computed tomography/computed tomography. Articular cartilage was analyzed at 6 and 12 weeks with ex vivo contrast-enhanced μCT and histology. To further explain the outcomes of our in vivo study, TA on macrophages was also studied in vitro. These cultured macrophages were either M1- or M2-activated, and they were analyzed using fluorescence-activated cell sorting for CD163 and FRβ expression as well as for messenger RNA (mRNA) expression of interleukin (IL)-10. Our in vivo study showed that intra-articular injections with TA strongly enhanced FRβ(+) macrophage activation. Despite stimulated macrophage activation, osteophyte formation was fully prevented. There was no beneficial effect of TA against cartilage degradation or subchondral bone sclerosis. In vitro macrophage cultures showed that TA strongly induced monocyte differentiation towards CD163(+) and FRβ(+) macrophages. Furthermore, TA-stimulated M2 macrophages showed enhanced IL-10 expression at the mRNA level. TA injections potently induce a CD163(+)- and FRβ(+)-activated macrophage with anti-inflammatory characteristics such as reduced IL-10 production in vitro and lack of osteophytosis in vivo.

Enhancing the Therapeutic Efficacy of Cancer Treatment With Cannabinoids

PubMed Central

Yasmin-Karim, Sayeda; Moreau, Michele; Mueller, Romy; Sinha, Neeharika; Dabney, Raymond; Herman, Allen; Ngwa, Wilfred

2018-01-01

Over the years, many in vitro and in vivo studies have shown the antineoplastic effects of cannabinoids (CBDs), with reports advocating for investigations of combination therapy approaches that could better leverage these effects in clinical translation. This study explores the potential of combination approaches employing CBDs with radiotherapy (RT) or smart biomaterials toward enhancing therapeutic efficacy during treatment of pancreatic and lung cancers. In in vitro studies, clonogenic assay results showed greater effective tumor cell killing, when combining CBDs and RT. Meanwhile, in vivo study results revealed major increase in survival when employing smart biomaterials for sustained delivery of CBDs to tumor cells. The significance of these findings, considerations for further research, and viable roadmap to clinical translation are discussed. PMID:29740535

Electrochemical properties of titanium nitride nerve stimulation electrodes: an in vitro and in vivo study

PubMed Central

Meijs, Suzan; Fjorback, Morten; Jensen, Carina; Sørensen, Søren; Rechendorff, Kristian; Rijkhoff, Nico J. M.

2015-01-01

The in vivo electrochemical behavior of titanium nitride (TiN) nerve stimulation electrodes was compared to their in vitro behavior for a period of 90 days. Ten electrodes were implanted in two Göttingen minipigs. Four of these were used for electrical stimulation and electrochemical measurements. Five electrodes were kept in Ringer's solution at 37.5°C, of which four were used for electrical stimulation and electrochemical measurements. The voltage transients measured in vivo were 13 times greater than in vitro at implantation and they continued to increase with time. The electrochemical properties in vivo and the tissue resistance (Rtissue) followed a similar trend with time. There was no consistent significant difference between the electrochemical properties of the in vivo and in vitro electrodes after the implanted period. The differences between the in vivo and in vitro electrodes during the implanted period show that the evaluation of electrochemical performance of implantable stimulation electrodes cannot be substituted with in vitro measurements. After the implanted period, however, the performance of the in vivo and in vitro electrodes in saline was similar. In addition, the changes observed over time during the post-implantation period regarding the electrochemical properties of the in vivo electrodes and Rtissue were similar, which indicates that these changes are due to the foreign body response to implantation. PMID:26300717

Antiplasmodial activity of novel keto-enamine chalcone-chloroquine based hybrid pharmacophores.

PubMed

Sashidhara, Koneni V; Kumar, Manoj; Modukuri, Ram K; Srivastava, Rajeev Kumar; Soni, Awakash; Srivastava, Kumkum; Singh, Shiv Vardan; Saxena, J K; Gauniyal, Harsh M; Puri, Sunil K

2012-05-01

A series of novel keto-enamine chalcone-chloroquine based hybrids were synthesized following new methodology developed in our laboratory. The synthesized compounds were screened against chloroquine sensitive strain (3D7) of Plasmodium falciparum in an in vitro model. Some of the compounds were showing comparable antimalarial activity at par with chloroquine. Compounds with significant in vitro antimalarial activity were then evaluated for their in vivo efficacy in Swiss mice against Plasmodium yoelii (chloroquine resistant N-67 strain), wherein compounds 25 and 27 each showed an in vivo suppression of 99.9% parasitaemia on day 4. Biochemical studies reveal that inhibition of hemozoin formation is the primary mechanism of action of these analogues. Copyright © 2012 Elsevier Ltd. All rights reserved.

[Application research of DEI technique based on synchrotron X-ray source in imaging rabbit eyeball in vitro].

PubMed

Yin, Hong-xia; Huang, Zhi-feng; Wang, Zhen-chang; Liu, Zhao-hui; Li, Yong; Zhu, Pei-ping

2010-03-23

To study the application of DEI technique in imaging the small structures of rabbit eyeball. DEI technique was used to image the eyeball of New Zealand white rabbit in vitro. The experiments were performed using beamline 4W1A at the topography station of Beijing Synchrotron Radiation Facility (BSRF). DEI image showed clearly the fine structures of the rabbit eyeball, such as the transparent cornea, the sclera, the ciliaris, and the ciliary body. DEI is a new X-ray imaging modality which achieves high contrast and spatial resolution. It also showed obvious effect of edge enhancement. DEI has good potential in observing the micro-structures of eyeballs and other small organs.

Light-driven changes in energy metabolism directly entrain the cyanobacterial circadian oscillator

PubMed Central

Rust, Michael J.; Golden, Susan S.; O'Shea, Erin K.

2012-01-01

Circadian clocks are self-sustained biological oscillators that can be entrained by environmental cues. Though this phenomenon has been studied in many organisms, the molecular mechanisms of entrainment remain unclear. Three cyanobacterial proteins and ATP are sufficient to generate oscillations in phosphorylation in vitro. We show that changes in illumination that induce a phase shift in cultured cyanobacteria also cause changes in the ATP/ADP ratio. When these nucleotide changes are simulated in the in vitro oscillator, they cause phase shifts similar to those observed in vivo. Physiological concentrations of ADP inhibit kinase activity in the oscillator and a mathematical model constrained by data shows that this effect is sufficient to quantitatively explain entrainment of the cyanobacterial circadian clock. PMID:21233390

Effect of different in vitro culture extracts of black pepper (Piper nigrum L.) on toxic metabolites-producing strains.

PubMed

Ahmad, Nisar; Abbasi, Bilal Haider; Fazal, Hina

2016-03-01

In the present study, the effect of different in vitro cultures (callus, in vitro shoots) and commercially available peppercorn extract was investigated for its activity against toxic metabolite-producing strains (Escherichia coli, Pseudomonas aeroginosa, Salmonella typhi, Bacillus subtilis, Bacillus cereus, Staphylococcus aureus, and Candida albicans). These in vitro cultures were extracted with ethanol, hexane, and chloroform, and the antipathogenic activity was determined by well-diffusion method. Hexane extract of callus showed 22 mm zone of inhibition against B. cereus, 23 mm against S. aureus, while regenerated shoots and seeds have shown 24.3 and 26 mm zones of inhibition. The ethanolic extracts of regenerated Piper shoots have shown 25 mm activity against S. aureus, 21 mm against B. cereus, and 16 mm in the case of C. albicans in comparison with standard antibiotics. Peppercorn extracts in chloroform and ethanol had shown activities against B. cereus (23.6 mm) and B. subtilis (23.5 mm). During in vitro organogenesis and morphogenesis, cells and tissues produced a comparable phytochemicals profile like mother plant. Morphogenesis is critically controlled by the application of exogenous plant-growth regulators. Such addition alters the hormonal transduction pathways, and cells under in vitro conditions regenerate tissues, which are dependant on the physiological state of cells, and finally enhance the production of secondary metabolites. To the best of our knowledge, this is the first report to compare the antimicrobial potential of in vitro regenerated tissues and peppercorn with standard antibiotics. In conclusion, most of the extracts showed pronounced activities against all the pathogenic microbes. This is a preliminary work, and the minimum inhibitory concentration values needs to be further explored. Regenerated tissues of P. nigrum are a good source of biologically active metabolites for antimicrobial activities, and callus culture presented itself as a good candidate for such activities. © The Author(s) 2013.

Mumps outbreaks in a highly vaccinated population: Investigation of a neutralization titre against the current circulating wildtype genotype G5 mumps virus.

PubMed

Kenny, Lena; O'Kelly, Edwin; Connell, Jeff; De Gascun, Cillian; Hassan, Jaythoon

2016-01-01

Mumps outbreaks continue to occur globally, despite high levels of uptake of the mumps vaccine. In order to address immunity to the current circulating wildtype virus, we sought to determine a mumps G5 specific IgG quantitative value which correlates with genotype G5 specific neutralization ability in vitro. Sera from 199 individuals including controls and acute mumps cases were assessed for mumps specific IgG titres using five different enzyme immunoassays coated with antigen from different mumps virus strains. A subset of 66 sera was also assessed for in vitro neutralizing antibody against a contemporary circulating genotype G5 mumps virus. For all the different antigenic targets, mumps specific IgG titres were higher in patients following acute mumps infection compared to controls. In acute mumps infected patients, females showed significantly higher serum titres of anti-G5 IgG compared to males (p<0.05). Furthermore, control males did not show any change in G5 specific IgG with increasing age whereas females show a progressive rise in titre. Linear regression analysis revealed a significant association between the mumps G5 specific IgG levels in the EIA and the in vitro neutralization titres (r(2)=0.59). Specific IgG to the current circulating genotype G5 mumps strain showed significantly lower titres in males which supports our previous observation that there is a male gender bias in cases of acute mumps infection. Furthermore, in this preliminary study, the data indicate that genotype G5 specific IgG levels of >40 RU/ml are required for neutralization capability to be observed in vitro. Copyright © 2015 Elsevier B.V. All rights reserved.

[Preparation and in vitro dissolution of magnolol solid dispersion].

PubMed

Tang, Lan; Qiu, Shuai-Bo; Wu, Lan; Lv, Long-Fei; Lv, Hui-Xia; Shan, Wei-Guang

2016-02-01

In this study, solid dispersion system of magnolol in croscarmellose sodium was prepared by using the solvent evaporation method, in order to increase the drug dissolution. And its dissolution behavior, stability and physical characteristics were studied. The solid dispersion was prepared with magnolol and croscarmellose sodium, with the proportion of 1∶5, the in vitro dissolution of magnolol solid dispersion was up to 80.66% at 120 min, which was 6.9 times of magnolol. The results of DSC (differential scanning calorimetry), IR (infra-red) spectrum and SEM (scanning electron microscopy) showed that magnolol existed in solid dispersion in an amorphous form. After an accelerated stability test for six months, the drug dissolution and content in magnolol solid dispersion showed no significant change. So the solid dispersion prepared with croscarmellose sodium as the carrier can remarkably improve the stability and dissolution of magnolol. Copyright© by the Chinese Pharmaceutical Association.

Radioprotective activity of Gentiana lutea extract and mangiferin.

PubMed

Menkovic, Nebojsa; Juranic, Zorica; Stanojkovic, Tatjana; Raonic-Stevanovic, Tatjana; Savikin, Katarina; Zdunić, Gordana; Borojevic, Nenad

2010-11-01

Radioprotective/sensitizing actions of Gentiana lutea aqueous-ethanol extract and mangiferin on radiation-induced effects on different types of cells were investigated. The study focused on the decreasing survival of normal human immunocompetent cells, the survival of the malignant cells in vitro, and the survival of ex vivo irradiated cells before and after consumption of the extract by healthy volunteers. The in vitro experiments showed that mangiferin could inhibit cytotoxic action of ionizing irradiation (doses of 6 and 8 Gy) only on normal resting human PBMC, not stimulated for proliferation. Orally consumed G. lutea extract showed the potential to reduce the cytotoxic effect of x-ray irradiation on normal human immunocompetent cells PBMC of some healthy people, without changing the susceptibility of malignant cells to be destroyed by irradiation. Since the radioprotective effect was individually dependent, further clinical studies are needed. Copyright © 2010 John Wiley & Sons, Ltd.

Enhancing Skin Permeation of Biphenylacetic Acid (BPA) Using Salt Formation with Organic and Alkali Metal Bases

PubMed Central

Pawar, Vijay; Naik, Prashant; Giridhar, Rajani; Yadav, Mange Ram

2015-01-01

In the present study, a series of organic and alkali metal salts of biphenylacetic acid (BPA) have been prepared and evaluated in vitro for percutaneous drug delivery. The physicochemical properties of BPA salts were determined using solubility measurements, DSC, and IR. The DSC thermogram and FTIR spectra confirmed the salt formation with organic and alkali metal bases. Among the series, salts with organic amines (ethanolamine, diethanolamine, triethanolamine, and diethylamine) had lowered melting points while the alkali metal salt (sodium) had a higher melting point than BPA. The in vitro study showed that salt formation improves the physicochemical properties of BPA, leading to improved permeability through the skin. Amongst all the prepared salts, ethanolamine salt (1b) showed 7.2- and 5.4-fold higher skin permeation than the parent drug at pH 7.4 and 5.0, respectively, using rat skin. PMID:26839810

Cholangiocytes derived from human induced pluripotent stem cells for disease modeling and drug validation.

PubMed

Sampaziotis, Fotios; de Brito, Miguel Cardoso; Madrigal, Pedro; Bertero, Alessandro; Saeb-Parsy, Kourosh; Soares, Filipa A C; Schrumpf, Elisabeth; Melum, Espen; Karlsen, Tom H; Bradley, J Andrew; Gelson, William Th; Davies, Susan; Baker, Alastair; Kaser, Arthur; Alexander, Graeme J; Hannan, Nicholas R F; Vallier, Ludovic

2015-08-01

The study of biliary disease has been constrained by a lack of primary human cholangiocytes. Here we present an efficient, serum-free protocol for directed differentiation of human induced pluripotent stem cells into cholangiocyte-like cells (CLCs). CLCs show functional characteristics of cholangiocytes, including bile acids transfer, alkaline phosphatase activity, γ-glutamyl-transpeptidase activity and physiological responses to secretin, somatostatin and vascular endothelial growth factor. We use CLCs to model in vitro key features of Alagille syndrome, polycystic liver disease and cystic fibrosis (CF)-associated cholangiopathy. Furthermore, we use CLCs generated from healthy individuals and patients with polycystic liver disease to reproduce the effects of the drugs verapamil and octreotide, and we show that the experimental CF drug VX809 rescues the disease phenotype of CF cholangiopathy in vitro. Our differentiation protocol will facilitate the study of biological mechanisms controlling biliary development, as well as disease modeling and drug screening.

Immunomodulator 'mushroom beta glucan' induces Wnt/β catenin signalling and improves wound recovery in tilapia and rat skin: a histopathological study.

PubMed

Hsiao, Chien-Mei; Wu, Yu-Sheng; Nan, Fan-Hua; Huang, Shih-Ling; Chen, Lynette; Chen, Shiu-Nan

2016-12-01

The present study aims to investigate the effects of mushroom beta glucan (MBG) on wound recovery in partial hepatectomy (PH) in Nile tilapia (Oreochromis niloticus) and in rat skin wound healing examination. Following PH, we focussed on the effects on liver repair ability using in vitro and in vivo tests. In vitro, we examined whether the MBG has an impact on liver cell proliferation, mainly through 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) assays and bromodeoxyuridine (BrdU) cell proliferation assay detection method. Results showed that MBG treatment was remarkable in enhancing cell proliferation of hepatocytes and in maintaining the cellular viability. Immunohistochemical staining to analyse Wnt/β-catenin signalling also showed that MBG has the effect of promoting cell proliferation of liver tissues after PH surgery. © 2015 Medicalhelplines.com Inc and John Wiley & Sons Ltd.

The immunomodulating V and W proteins of Nipah virus determine disease course.

PubMed

Satterfield, Benjamin A; Cross, Robert W; Fenton, Karla A; Agans, Krystle N; Basler, Christopher F; Geisbert, Thomas W; Mire, Chad E

2015-06-24

The viral determinants that contribute to Nipah virus (NiV)-mediated disease are poorly understood compared with other paramyxoviruses. Here we use recombinant NiVs (rNiVs) to examine the contributions of the NiV V and W proteins to NiV pathogenesis in a ferret model. We show that a V-deficient rNiV is susceptible to the innate immune response in vitro and behaves as a replicating non-lethal virus in vivo. Remarkably, rNiV lacking W expression results in a delayed and altered disease course with decreased respiratory disease and increased terminal neurological disease associated with altered in vitro inflammatory cytokine production. This study confirms the V protein as the major determinant of pathogenesis, also being the first in vivo study to show that the W protein modulates the inflammatory host immune response in a manner that determines the disease course.

False HDAC Inhibition by Aurone Compound.

PubMed

Itoh, Yukihiro; Suzuki, Miki; Matsui, Taiji; Ota, Yosuke; Hui, Zi; Tsubaki, Kazunori; Suzuki, Takayoshi

2016-01-01

Fluorescence assays are useful tools for estimating enzymatic activity. Their simplicity and manageability make them suitable for screening enzyme inhibitors in drug discovery studies. However, researchers need to pay attention to compounds that show auto-fluorescence and quench fluorescence, because such compounds lower the accuracy of the fluorescence assay systems by producing false-positive or negative results. In this study, we found that aurone compound 7, which has been reported as a histone deacetylase (HDAC) inhibitor, gave false-positive results. Although compound 7 was identified by an in vitro HDAC fluorescence assay, it did not show HDAC inhibitory activity in a cell-based assay, leading us to suspect its in vitro HDAC inhibitory activity. As a result of verification experiments, we found that compound 7 interferes with the HDAC fluorescence assay by quenching the HDAC fluorescence signal. Our findings underscore the faults of fluorescence assays and call attention to careless interpretation.

Leishmanicidal activity of nine novel flavonoids from Delphinium staphisagria.

PubMed

Ramírez-Macías, Inmaculada; Marín, Clotilde; Díaz, Jesús G; Rosales, María José; Gutiérrez-Sánchez, Ramón; Sánchez-Moreno, Manuel

2012-01-01

To evaluate the in vitro leishmanicidal activity of nine flavonoid derivatives from Delphinium staphisagria against L. infantum and L. braziliensis. The in vitro activity of compounds 1-9 was assayed on extracellular promastigote and axenic amastigote forms and on intracellular amastigote forms of the parasites. Infectivity and cytotoxicity tests were carried on J774.2 macrophage cells using Glucantime as the reference drug. The mechanisms of action were analysed performing metabolite excretion and transmission electronic microscope ultrastructural alteration studies. Nine flavonoids showed leishmanicidal activity against promastigote as well as amastigote forms of Leishmania infantum and L. braziliensis. These compounds were nontoxic to mammalian cells and were effective at similar concentrations up to or lower than that of the reference drug (Glucantime). The results showed that 2(″)-acetylpetiolaroside (compound 8) was clearly the most active. This study has demonstrated that flavonoid derivatives are active against L. infantum and L. braziliensis.

A novel single compartment in vitro model for electrophysiological research using the perfluorocarbon FC-770.

PubMed

Choudhary, M; Clavica, F; van Mastrigt, R; van Asselt, E

2016-06-20

Electrophysiological studies of whole organ systems in vitro often require measurement of nerve activity and/or stimulation of the organ via the associated nerves. Currently two-compartment setups are used for such studies. These setups are complicated and require two fluids in two separate compartments and stretching the nerve across one chamber to the other, which may damage the nerves. We aimed at developing a simple single compartment setup by testing the electrophysiological properties of FC-770 (a perfluorocarbon) for in vitro recording of bladder afferent nerve activity and electrical stimulation of the bladder. Perflurocarbons are especially suitable for such a setup because of their high oxygen carrying capacity and insulating properties. In male Wistar rats, afferent nerve activity was recorded from postganglionic branches of the pelvic nerve in vitro, in situ and in vivo. The bladder was stimulated electrically via the efferent nerves. Organ viability was monitored by recording spontaneous contractions of the bladder. Additionally, histological examinations were done to test the effect of FC-770 on the bladder tissue. Afferent nerve activity was successfully recorded in a total of 11 rats. The bladders were stimulated electrically and high amplitude contractions were evoked. Histological examinations and monitoring of spontaneous contractions showed that FC-770 maintained organ viability and did not cause damage to the tissue. We have shown that FC-770 enables a simple, one compartment in vitro alternative for the generally used two compartment setups for whole organ electrophysiological studies.

Effects of Stirring and Fluid Perfusion on the In Vitro Degradation of Calcium Phosphate Cement/PLGA Composites.

PubMed

An, Jie; Leeuwenburgh, Sander C G; Wolke, Joop G C; Jansen, John A

2015-11-01

In vitro degradation rates of calcium phosphate bioceramics are investigated using a large variation of soaking protocols that do not all match the dynamic conditions of the perfused physiological environment. Therefore, we studied the effect of stirring and fluid perfusion on the in vitro degradation rate of apatitic calcium phosphate cements (CPC) containing poly(lactic-co-glycolic acid) (PLGA) microspheres. The composites were soaked in phosphate-buffered saline up to 6 weeks under unstirred, stirred, or perfused conditions followed by analysis of mass loss, compression strength, porosity, crystal phase composition, and morphology of the cement composites. The results showed that fluid perfusion reduced the decrease in pH and corresponding degradation rates, while nonperfused soaking conditions (i.e., stirred and unstirred conditions) resulted into more extensive acidification, the rate of which increased with stirring. After 2 weeks, the formation of a secondary brushite phase was observed for cement composites soaked under nonperfused (i.e., stirred and unstirred) conditions, whereas this phase was not detected in cements soaked under perfused conditions. The degradation rate of cement composites decreased in the order unstirred>stirred>perfused, as evidenced by quantification of mass loss, compression strength, and pore morphology. To summarize, we have demonstrated that soaking conditions strongly affected the in vitro degradation process of CPCs. As a consequence, it can be concluded that the experimental design of current in vitro degradation studies does not allow for correlation to (pre-)clinical studies.

Sustained Release of Green Tea Polyphenols from Liposomal Nanoparticles; Release Kinetics and Mathematical Modelling.

PubMed

Prakash Upputuri, Ravi Theaj; Azad Mandal, Abul Kalam

2017-01-01

Background: Green tea polyphenols (GTP) are known to have several health benefits. In spite of these benefits, its application as a therapeutic agent is limited due to some of its limitations such as stability, bioavailability, and biotransformation. To overcome these limitations, liposomal nanoparticles have been used as a carrier of the GTP. Objective: Encapsulation of GTP to the liposomal nanoparticles in order to achieve a sustained release of the GTP and to determine the drug release kinetics and the mechanism of the release. Materials and Methods: GTP encapsulated liposomal nanoparticles were prepared using phosphatidyl choline and cholesterol. The synthesized particles were characterized for their particle size and morphology. In vitro release studies were carried out, followed by drug release kinetics, and determining the mechanism of release. In vitro , antioxidant assay was determined following 2,2-diphenyl-1-picrylhydrazyl (DPPH) method. Results: Atomic force microscope (AFM) and high resolution scanning electron microscope (HR SEM) images showed spherical particles of the size of 64.5 and 252 nm. An encapsulation efficiency as high as 77.7% was observed with GTP concentration of 5 mg.mL -1 . In vitro release studies showed that the loading concentrations of GTP were independent to the cumulative percentage of the drug release. GTP release by varying the pH and temperature showed a direct correlation between the release parameter and the percentage of drug release. The higher the pH and temperature, the higher was the percentage of the drug release. The release data showed a good correlation with Zero order kinetics and the mechanism of the release being anomalous mode. Radical scavenging activity of the released GTP showed a potent scavenging activity. Conclusion: GTP encapsulated liposomal nanoparticles could be used as a delivery vehicle for achieving a sustained release.

Matrix-Embedded Cytokines to Simulate Osteoarthritis-Like Cartilage Microenvironments

PubMed Central

Murab, Sumit; Chameettachal, Shibu; Bhattacharjee, Maumita; Das, Sanskrita; Kaplan, David L.

2013-01-01

In vivo, cytokines noncovalently bind to the extracellular matrix (ECM), to facilitate intimate interactions with cellular receptors and potentiate biological activity. Development of a biomaterial that simulates this type of physiological binding and function is an exciting proposition for designing controlled advanced delivery systems for simulating in vivo conditions in vitro. We have decorated silk protein with sulfonated moieties through diazonium coupling reactions to noncovalently immobilize pro-inflammatory cytokines interleukin-1 beta (IL-1β) and tumor necrosis factor alpha (TNF-α) in such a biomimetic manner. After adsorption of the cytokines to the diazonium-modified silk matrix, constant release of cytokines up to at least 3 days was demonstrated, as an initial step to simulate an osteoarthritic (OA) microenvironment in vitro. Matrix-embedded cytokines induced the formation of multiple elongated processes in chondrocytes in vitro, akin to what is seen in OA cartilage in vivo. Gene expression profiles with this in vitro tissue model of OA showed significant similarities to profiles from explanted OA cartilage tissues collected from patients who underwent total knee replacement surgery. The common markers of OA, including COL, MMP, TIMP, ADAMTS, and metallothioneins, were upregulated at least 35-fold in the in vitro model when compared to the control—non-OA in vitro generated tissue-engineered cartilage. The microarray data were validated by reverse transcriptase–polymerase chain reaction. Mechanistically, protein interaction studies indicated that TNF-α and IL-1β synergistically controlled the equilibrium between MMPs and their inhibitors, TIMPs, resulting in ECM degradation through the MAPK pathway. This study offers a promising initial step toward establishing a relevant in vitro OA disease model, which can be further modified to assess signaling mechanisms, responses to cell or drug treatments and patient-specific features. PMID:23470228

A feasibility study of prepubertal and over mature aged local goat in relation to results of In Vitro growth culture to obtain additional M-II oocyte resources

NASA Astrophysics Data System (ADS)

Ciptadi, Gatot; Ihsan, M. Nur; Rahayu, Sri; Widjaja, D. H. K.; Mudawamah, Mudawamah

2017-11-01

The aims of this research are to study the potential source of mature (M-II) oocytes of domestic animals using follicles isolated from prepubertal and over mature aged Indonesian local goats, resulting from an in vitro growth (IVG) method. This method of IVG could provide a new source of M-II oocytes for embryo production. In Indonesia, a very limited number of a good quality oocytes are available for research purposes, as there is a limited number of reproductive females slaughtered, which is dominated by prepubertal and old mature aged animals. IVG culture systems could be improved as an alternative method to provide a new source of a good quality oocytes for in vitro maturation of M-II oocytes. From a number of prepubertal and mature aged goats slaughtered in a local abattoir, the small oocytes in the preantral follicles were cultured in vitro to normal oocyte growth. The methods used in this research are experimental. Follicles were isolated, cultured in vitro for 14 days individually using a sticky medium containing 4% (w/v) polyvinylpyrrolidone in TCM 199 10% Fetal Bovine Serum supplemented with Follicle Stimulating Hormone, which was then evaluated for their follicle development and oocyte quality. The research results showed that a minimum follicle size and oocyte diameter is needed (>100 um) for early evaluation of maturation to be achieved, meanwhile oocytes recovered from IVG after being cultured in vitro for maturation resulted in a very low rate of maturation. However, in the future, IVG of the preantral follicles of Indonesian local goat could be considered as an alternative source of oocytes for both research purposes and embryo production in vitro.

Sunscreen tests: correspondence between in vitro data and values reported by the manufacturers.

PubMed

Garoli, Denis; Pelizzo, Maria Guglielmina; Bernardini, Bianca; Nicolosi, Piergiorgio; Alaibac, Mauro

2008-12-01

In vitro sunscreen tests are diffusively used to test both the sun protection factor (SPF) and the photo-stability of filters. Spectrophotometric measurements of the absorbance of ultraviolet radiations through a sunscreen applied on a suitable substrate allow a rapid evaluation of its protection factor both at short and long wavelength ultraviolet radiation (UVB and UVA). The objective of this study has been to demonstrate if Teflon can be adopted as substrate both for SPF evaluation and photo-stability tests. Moreover, we have investigated if there is a correspondence between in vitro SPF measurements and values reported by manufacturers on sunscreens. Teflon has been used to perform several photo-stability tests by irradiating the filters with different wavebands and analyzing the combined effect of UV and infrared (IR) light. Similar analyses have been carried out using PMMA Plates, which is the standard substrate for UVA in vitro test. We have demonstrated that it is possible to establish a good correspondence between in vitro SPF and values reported by manufacturers on sunscreens. We have also verified that the in vitro/label SPF correlation curve depends on the quantity of product applied while this does not seem to be true for other parameters like Critical Wavelength and UVA ratio. With regard to photo-stability studies, our results indicate for the first time that IR irradiation may have a role on photo-degradation. The results show that there is a good correlation between the in vitro SPF determined by the present method and the SPF reported by the manufacturer. The compatibility of the results obtained using Teflon and PMMA Plates demonstrates that Teflon can be utilized for both SPF determination and photo-stability tests.

[In vitro metabolism of fenbendazole prodrug].

PubMed

Wen, Ai-Dan; Duan, Li-Ping; Liu, Cong-Shan; Tao, Yi; Xue, Jian; Wu, Ning-Bo; Jiang, Bin; Zhang, Hao-Bing

2013-02-01

Synthesized fenbendazole prodrug N-methoxycarbonyl-N'-(2-nitro-4-phenylthiophenyl) thiourea (MPT) was analyzed in vitro in artificial gastric juice, intestinal juice and mouse liver homogenate model by using HPLC method, and metabolic curve was then generated. MPT was tested against Echinococcus granulosus protoscolices in vitro. The result showed that MPT could be metabolized in the three biological media, and to the active compound fenbendazole in liver homogenate, with a metabolic rate of 7.92%. Besides, the prodrug showed a weak activity against E. granulosus protoscolices with a mortality of 45.9%.

In vitro activity of the polyether ionophorous antibiotic monensin against the cyst form of Toxoplasma gondii.

PubMed

Couzinet, S; Dubremetz, J F; Buzoni-Gatel, D; Jeminet, G; Prensier, G

2000-10-01

Toxoplasma gondii. The experiments were conducted in vitro using 2 methods; cysts produced either in mice or in cell culture were exposed to monensin in vitro, and the infectivity of the parasites was then assessed in vivo or in vitro. The data obtained from these 2 systems of evaluation showed that monensin inhibits the infectivity and the viability of the bradyzoites. Its activity was time and concentration dependent. The first effects were observed at very low drug concentrations (i.e. 0.0001 microg/ml). Immunofluorescence and electron microscopy analysis showed significant cytological alterations of the monensin-treated bradyzoites: they were swollen, had a large number of vacuoles in their cytoplasm and were found lysed at higher concentrations in ionophore.

Characterizing parameters of Jatropha curcas cell cultures for microgravity studies

NASA Astrophysics Data System (ADS)

Vendrame, Wagner A.; Pinares, Ania

2013-06-01

Jatropha (Jatropha curcas) is a tropical perennial species identified as a potential biofuel crop. The oil is of excellent quality and it has been successfully tested as biodiesel and in jet fuel mixes. However, studies on breeding and genetic improvement of jatropha are limited. Space offers a unique environment for experiments aiming at the assessment of mutations and differential gene expression of crops and in vitro cultures of plants are convenient for studies of genetic variation as affected by microgravity. However, before microgravity studies can be successfully performed, pre-flight experiments are necessary to characterize plant material and validate flight hardware environmental conditions. Such preliminary studies set the ground for subsequent spaceflight experiments. The objectives of this study were to compare the in vitro growth of cultures from three explant sources (cotyledon, leaf, and stem sections) of three jatropha accessions (Brazil, India, and Tanzania) outside and inside the petriGAP, a modified group activation pack (GAP) flight hardware to fit petri dishes. In vitro jatropha cell cultures were established in petri dishes containing a modified MS medium and maintained in a plant growth chamber at 25 ± 2 °C in the dark. Parameters evaluated were surface area of the explant tissue (A), fresh weight (FW), and dry weight (DW) for a period of 12 weeks. Growth was observed for cultures from all accessions at week 12, including subsequent plantlet regeneration. For all accessions differences in A, FW and DW were observed for inside vs. outside the PetriGAPs. Growth parameters were affected by accession (genotype), explant type, and environment. The type of explant influenced the type of cell growth and subsequent plantlet regeneration capacity. However, overall cell growth showed no abnormalities. The present study demonstrated that jatropha in vitro cell cultures are suitable for growth inside PetriGAPs for a period of 12 weeks. The parameters evaluated in this study provide the basic ground work and pre-flight assessment needed to justify a model for microgravity studies with jatropha in vitro cell cultures. Future studies should focus on results of experiments performed with jatropha in vitro cultures in microgravity.

High Modulus Biodegradable Polyurethanes for Vascular Stents: Evaluation of Accelerated in vitro Degradation and Cell Viability of Degradation Products

PubMed Central

Sgarioto, Melissa; Adhikari, Raju; Gunatillake, Pathiraja A.; Moore, Tim; Patterson, John; Nagel, Marie-Danielle; Malherbe, François

2015-01-01

We have recently reported the mechanical properties and hydrolytic degradation behavior of a series of NovoSorb™ biodegradable polyurethanes (PUs) prepared by varying the hard segment (HS) weight percentage from 60 to 100. In this study, the in vitro degradation behavior of these PUs with and without extracellular matrix (ECM) coating was investigated under accelerated hydrolytic degradation (phosphate buffer saline; PBS/70°C) conditions. The mass loss at different time intervals and the effect of aqueous degradation products on the viability and growth of human umbilical vein endothelial cells (HUVEC) were examined. The results showed that PUs with HS 80% and below completely disintegrated leaving no visual polymer residue at 18 weeks and the degradation medium turned acidic due to the accumulation of products from the soft segment (SS) degradation. As expected the PU with the lowest HS was the fastest to degrade. The accumulated degradation products, when tested undiluted, showed viability of about 40% for HUVEC cells. However, the viability was over 80% when the solution was diluted to 50% and below. The growth of HUVEC cells is similar to but not identical to that observed with tissue culture polystyrene standard (TCPS). The results from this in vitro study suggested that the PUs in the series degraded primarily due to the SS degradation and the cell viability of the accumulated acidic degradation products showed poor viability to HUVEC cells when tested undiluted, however particles released to the degradation medium showed cell viability over 80%. PMID:26000274

[The study on the proliferation and the apoptosis factors in vitro of Kölliker organ supporting cells in the cochlea of newborn rat].

PubMed

He, Yuanyuan; Yang, Jun

2015-01-01

To study the apoptosis/proliferation of Kölliker organ supporting cells and to understand the prompting apoptosis factors in vivo in the supporting cells in the Kölliker organ by changing the environment of the cultured supporting cells in the Kliker organ in vitro, via the separation, culture and purification of the supporting cells in the K6lliker organ. A combinatorial approach of enzymatic digestion and mechanical separation was employed to isolate and culture in vitro pure Kölliker organ supporting cells. The purity was tested by flow cytometry assay. And K6lliker organ supporting cells were harvested to detect the rate and cycle of apoptosis by flow cytometry after Annexin V/PI staining, to test the cell growth curve by MTT assay, and to observe the differential expressions of the Bcl-2, Caspase-3, Caspase-8 and Caspase-9 through the Realtime PCR and Western blot. The calcium, potassium and glutamate concentrations in the culture medium of these cells in vitro were changed to detect the survival rate of cells by MTT assay. The purity of K6lliker organ supporting cells by flow cytometry assay was 96. 56%. And these cells showed no significant difference in apoptosis, but an evident linear growth. The results of Realtime PCR and Western blot showed that the expression of Bcl-2, Caspase-3, Caspase-8 and Caspase-9 mRNA and protein in all different time points kept stable. Furthermore, the elevation of extracellular Ca2+ might contribute to decrease the cell viability of supporting cells. And K+ participated regulation of cell viability in a concentration-depending way. However, glutamate appeared to be a protective factor in high concentration. There is no significant apoptosis in vitro of the supporting cells in the Kölliker organ of rats, showing a linear growth. The Ca2+ in high concentration might contribute to the apoptosis factor of these cells. However, the K+ and glutamate appear to be protective factors in high concentration.