Search results for: in vitro fertilization

Authors: Rajinikanth Siddalingam, Poonguzhali Subramanian

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The present study aims to prepare and evaluate the self-nano emulsifying drug delivery (SNEDDS) system to enhance the dissolution rate of a poorly soluble drug dutasteride. The formulation was prepared using capryol PGMC, Cremophor EL, and polyethylene glycol (PEG) 400 as oil, surfactant and co-surfactant, respectively. The pseudo-ternary phase diagrams with presence and absence of drug were plotted to find out the nano emulsification range and also to evaluate the effect of dutasteride on the emulsification behavior of the phases. Prepared SNEDDS formulations were evaluated for its particle size distribution, nano emulsifying properties, robustness to dilution, self-emulsification time, turbidity measurement, drug content and in-vitro dissolution. The optimized formulations are further evaluated for heating cooling cycle, centrifugation studies, freeze-thaw cycling, particle size distribution and zeta potential were carried out to confirm the stability of the formed SNEDDS formulations. The particle size, zeta potential and polydispersity index of the optimized formulation found to be 35.45 nm, -15.45 and 0.19, respectively. The in vitro results are revealed that the prepared formulation enhanced the dissolution rate of dutasteride significantly as compared with pure drug. The in vivo studies in was conducted using rats and the results are revealed that SNEDDS formulation has enhanced the bioavailability of dutasteride drug significantly as compared with raw drug. Based the results, it was concluded that the dutasteride-loaded SNEDDS shows potential to enhance the dissolution of dutasteride, thus improving the bioavailability and therapeutic effects.

Keywords: self-emulsifying drug delivery system, dutasteride, enhancement of bioavailability, dissolution enhancement

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Authors: Irina Mikajlo, Jaroslav Záhora, Helena Dvořáčková, Jaroslav Hynšt, Jakub Elbl

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Strong anthropogenic impact has uncontrolled consequences on the nature of the soil. Hence, up-to-date sustainable methods of soil state improvement are essential. Investigators provide the evidence that biochar can positively effects physical, chemical and biological soil properties and the abundance of mycorrhizal fungi which are in the focus of this study. The main aim of the present investigation is to demonstrate the effect of two types of plant growth promoting bacteria (PGPB) inoculums along with the beech wood biochar and mineral N additives on mycorrhizal colonization. Experiment has been set up in laboratory conditions with containers filled with arable soil from the protection zone of the main water source ‘Brezova nad Svitavou’. Lactuca sativa (lettuce) has been selected as a model plant. Based on the obtained data, it can be concluded that mycorrhizal colonization increased as the result of combined influence of biochar and PGPB inoculums amendment. In addition, correlation analyses showed that the numbers of main groups of cultivated bacteria were dependent on the degree of mycorrhizal colonization.

Keywords: Arbuscular mycorrhiza, biochar, PGPB inoculum, soil microorganisms

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Authors: Jae Bok Heo, Yun Mi Lee, Hee Rang Yun

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Several GTPases are required for ribosome biogenesis and assembly. We recently characterized rice (Oryza sativa) nuclear/nucleolar GTPase 2 (OsNug2), belonging to the YlqF/YawG family of GTPases, as playing a role in pre-60S ribosomal subunit maturation. To investigate the potential factors involved in regulating the function of OsNug2, yeast two-hybrid screens were carried out using OsNug2 as bait. Rice serine/threonine kinase 1 (OsSTK1) was identified as a potential interacting protein candidate. In vitro pull down and bimolecular fluorescence complementation assays confirmed the interaction between OsNug2 and OsSTK1, and like green fluorescent protein-tagged OsNug2, green fluorescent protein-tagged OsSTK1 was targeted to the nucleus of Arabidopsis protoplasts. OsSTK1 was not found to affect the GTP-binding activity of OsNug2; however, when recombinant OsSTK1 was included in OsNug2 assay reaction mixtures, OsSTK1 increased the GTPase activity of OsNug2. To test whether OsSTK1 phosphorylates OsNug2 in vitro, a kinase assay was performed. OsSTK1 was found to have weak autophosphorylation activity and strongly phosphorylated serine 209 of OsNug2. Yeast complementation testing resulted in a GAL::OsNug2(S209N) mutant-harboring yeast strain exhibiting a growth-defective phenotype on galactose medium at 39°C, divergent from that of a yeast strain harboring GAL::OsNug2. The intrinsic GTPase activity of mutant OsNug2(S209N) was found to be similar to that of OsNug2, was not fully enhanced upon weak binding of OsSTK1. Our findings reported here indicate that OsSTK1 functions as a positive regulator protein of OsNug2 by enhancing the GTPase activity of OsNug2, and that the phosphorylation of serine 209 of OsNug2 is essential for the complete function of OsNug2 in ribosome biogenesis.

Keywords: OsSTK1, OsNug2, GTPase activity, GTP binding activity, phosphorylation

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Authors: Hajji Lobna, Chattaoui Mayssa, Regaieg Hajer, M'Hamdi-Boughalleb Naima, Rhouma Ali, Horrigue-Raouani Najet

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In this study, three local isolates of Trichoderma (Tr1: T. viride, Tr2: T. harzianum and Tr3: T. asperellum) were isolated and evaluated for their biocontrol effectiveness under in vitro conditions and in greenhouse. In vitro bioassay revealed a biopotential control against Fusarium oxysporum f. sp. radicis lycopersici and Meloidogyne javanica (RKN) separately. All species of Trichoderma exhibited biocontrol performance and (Tr1) Trichoderma viride was the most efficient. In fact, growth rate inhibition of Fusarium oxysporum f. sp. radicis lycopersici (FORL) was reached 75.5% with Tr1. Parasitism rate of root-knot nematode was 60% for juveniles and 75% for eggs with the same one. Pots experiment results showed that Tr1 and Tr2, compared to chemical treatment, enhanced the plant growth and exhibited better antagonism against root-knot nematode and root-rot fungi separated or combined. All Trichoderma isolates revealed a bioprotection potential against Fusarium oxysporum f. sp. radicis lycopersici. When pathogen fungi inoculated alone, Fusarium wilt index and browning vascular rate were reduced significantly with Tr1 (0.91, 2.38%) and Tr2 (1.5, 5.5%), respectively. In the case of combined infection with Fusarium and nematode, the same isolate of Trichoderma Tr1 and Tr2 decreased Fusarium wilt index at 1.1 and 0.83 and reduced the browning vascular rate at 6.5% and 6%, respectively. Similarly, the isolate Tr1 and Tr2 caused maximum inhibition of nematode multiplication. Multiplication rate was declined at 4% with both isolates either tomato infected by nematode separately or concomitantly with Fusarium. The chemical treatment was moderate in activity against Meloidogyne javanica and Fusarium oxysporum f. sp. radicis lycopersici alone and combined.

Keywords: trichoderma spp., meloidogyne javanica, Fusarium oxysporum f.sp.radicis lycopersici, biocontrol

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Authors: Esmaeil Kouhgardi, Saeedeh Dashti, Hakimeh Fekrandish

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This research has been conducted to study the method of culture and comparing growth and biomass of Gracilariacorticata cultured on rope and net for 50 days through two treatments (first treatment: culture of alga on net and the second treatment: culture of alga on rope and each treatment was repeated by four cases). During culture period, the water of aquariums was replaced once every two days for 40-50%. Also, 0.3-0.5 grams of Urea fertilizer was added to the culture environment for fertilization. Moreover, some of the environmental factors such as pH, salinity and temperature of the environment were measured on a daily basis. During the culture period, extent of longitudinal growth of the species of both treatments was equal. The said length was reached from 8-10 cm to 10.5-13 cm accordingly. The resulted weight in repetitions of the first treatment was higher than that of the second treatment in such a way as in the first treatment, its weight reached from 10 grams to 21.119 grams and in the second treatment, its weight reached from 10 grams to 17.663 grams. On a whole, it may be stated that that kind of alga being studied has a considerable growth with respect to its volume. The results have revealed that the percentage of daily growth and wet weight at the end of the first treatment was higher than that of the second treatment and it was registered as 0.934, 6.072 and 811.432 in the first treatment and 0.797, 4.990 and 758.071 in the second treatment respectively. This difference is significant (P < 0.05). Growth and biomass of G. corticata through culture on net was more emphasizing on numerous branches due to wider bed. Moreover, higher level of the species in this method was exposed to sunlight and this increased biosynthesis and eventually increases of growth and biomass.

Keywords: red alga, growth, biomass, culture, net, rope

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Authors: Mostafa Heidari

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Nitrogen fertilization has played a significant role in increasing crop yield, and solving problems of hunger and malnutrition worldwide. However, excessive of heavy metals such as arsenic can interfere on growth and reduced grain yield. In order to investigate the effects of different concentrations of arsenic and nitrogen fertilizer on photosynthetic pigments and antioxidant enzyme activities in safflower (cv. Goldasht), a factorial plot experiment as randomized complete block design with three replication was conducted in university of Zabol. Arsenic treatment included: A1= control or 0, A2=30, A3=60 and A4=90 mg. kg-1 soil from the Na2HASO4 source and three nitrogen levels including W1=75, W2=150 and W3=225 kg.ha-1 from urea source. Results showed that, arsenic had a significant effect on the activity of antioxidant enzymes. By increasing arsenic levels from A1 to A4, the activity of ascorbate peroxidase (APX) and gayacol peroxidase (GPX) increased and catalase (CAT) was decreased. In this study, arsenic had no significant on chlorophyll a, b and cartoneid content. Nitrogen and interaction between arsenic and nitrogen treatment, except APX, had significant effect on CAT and GPX. The highest GPX activity was obtained at A4N3 treatment. Nitrogen increased the content of chlorophyll a, b and cartoneid.

Keywords: arsenic, physiological parameters, oxidative enzymes, nitrogen

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Authors: Ahmed Altalb

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Fertilizers are natural materials, or industrial contain nutrients, which help to improve soil fertility and is considered (nitrogen, phosphorus, and potassium) is important elements for the growth of crops properly. Fertilization is necessary in order to improve the quality of agricultural products and the recovery in agricultural activities. The use of organic fertilizers and chemical lead to reduce the loss of nutrients in agricultural soils, and this leads to an increase in the production of agricultural crops. Fertilizers are one of the key factors in the increase of agricultural production as well as other factors such as irrigation and improved seeds and Prevention and others; the fertilizers will continue to be a cornerstone of the agriculture in order to produce the food to feed of world population. The use of fertilizers has become commonplace today, especially the chemical fertilizers for the development of agricultural production, due to the provision of nutrients for plants and in high concentrations and easily dissolves in water and ease of use. The choose the right type of fertilizer depends on the soil type and the type of crop. In this subject, find the relationship between the agricultural extension and the optimal use of fertilizers. The extension plays the important role in the advise and educate of farmers in how they optimal use the fertilizers in a scientific way. This article aims to identify the concept the fertilizers. Identify the role of fertilizers in increasing the agricultural production, identify the role of agricultural extension in the optimal use of fertilizers and rural development.

Keywords: agricultural, extension, fertilizers, production

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Authors: Mayuva Youngsabanant-Areekijseree, Chanikarn Srinark, S. Sengsai, Mayuree Pumipaiboon

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Our project was aimed at morphology of oocytes, follicle cells and follicular fluid proteins study of Large White pig (at local slaughter house in Nakhon Pathom Province). The porcine oocytes and follicular fluid of healthy small follicles (1-2 mm), medium follicles (3-6 mm in diameters) and large follicles (7-8 mm and 10 mm in diameter) were aspirated and collected from the ovary by sterile technique. Then, the oocytes and the follicle cells were separated from the fluid. The oocytes were round shape and surrounded by zona pellucida with numerous layers of cumulus cells. Based on the number of cumulus cell layers surrounding oocytes, the oocytes were classified into 5 types, which were intact-, multi-, partial-cumulus layer oocyte, completely denuded oocyte and degenerative oocyte. The collected oocytes showed high percentages of intact- and multi- cumulus cell layers in the small follicles (53.48%) medium follicles (56.94%) and large follicles (56.52%) which have high potential to develop into mature oocytes in vitro. Proteins from follicular fluid of 3 size follicles were separated by SDS-PAGE and LC/MS/MS. The molecular weight of follicular fluid proteins from the small follicles were 24, 60-65, 79, 110, 140, 160, and > 220 kDa. Meanwhile, the follicular fluid protein from medium and large follicle contained 52, 65, 79, 90, 110, 120, 160, 190 and > 220 kDa. Almost all proteins played important roles in promoting and regulating growth and development of oocytes and ovulation. This finding was an initial tool for in vitro testing and applied biotechnology research. Acknowledgements: The project was funded by a grant from Silpakorn University Research & Development Institute (SURDI) and Faculty of Science, Silpakorn University, Thailand.

Keywords: follicular fluid protein, LC/MS/MS, porcine oocyte, SDS-PAGE, reproductive biology

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Authors: C. E. Chinma, S. O. Azeez, J. C. Anuonye, O. B. Ocheme, C. M. Yakubu, S. James, E. U. Ohuoba, I. A. Baba

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There is growing interest in the use of rice bran protein in food formulation due to its hypoallergenic protein, high nutritional value and health promoting potentials. For the first time, the amino acid profile, protein digestibility, antioxidant, and functional properties of protein concentrate from some local varieties of rice bran from Nigeria were studied for possible food applications. Protein concentrates were prepared from rice bran and analysed using standard methods. Results showed that protein content of Kwandala, Yardass, Jeep, and Jamila were 69.24%, 69.97%, 68.73%, and 71.62%, respectively while total essential amino acid were 52.71, 53.03, 51.86, and 55.75g/100g protein, respectively. In vitro protein digestibility of protein concentrate from Kwandala, Yardass, Jeep and Jamila were 90.70%, 91.39%, 90.57% and 91.63% respectively. DPPH radical inhibition of protein from Kwandala, Yardass, Jeep, and Jamila were 48.15%, 48.90%, 47.56%, and 53.29%, respectively while ferric reducing ability power were 0.52, 0.55, 0.47 and 0.67mmol TE per gram, respectively. Protein concentrate from Jamila had higher onset (92.57oC) and denaturation temperature (102.13oC), and enthalpy (0.72J/g) than Jeep (91.46oC, 101.76oC, and 0.68J/g, respectively), Kwandala (90.32oC, 100.54oC and 0.57J/g, respectively), and Yardass (88.94oC, 99.45oC, and 0.51J/g, respectively). In vitro digestibility of protein from Kwandala, Yardas, Jeep, and Jamila were 90.70%, 91.39%, 90.57% and 91.63% respectively. Oil absorption capacity of Kwandala, Yardass, Jeep, and Jamila were 3.61, 3.73, 3.40, and 4.23g oil/g sample respectively, while water absorption capacity were 4.19, 4.32, 3.55 and 4.48g water/g sample, respectively. Protein concentrates had low bulk density (0.37-0.43g/ml). Protein concentrate from Jamila rice bran had the highest foam capacity (37.25%), followed by Yardass (34.20%), Kwandala (30.14%) and Jeep (28.90%). Protein concentrates showed low emulsifying and gelling capacities. In conclusion, protein concentrate prepared from these local rice bran varieties could serve as functional ingredients in food formulations and for enriching low protein foods.

Keywords: rice bran protein, amino acid profile, protein digestibility, antioxidant and functional properties

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Authors: Richa Sharma, Uma Nahar, Sadhna Sharma, Indu Verma

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Counteracting the deadly pathogen Mycobacterium tuberculosis (M. tb) effectively is still a global challenge. Scrutinizing alternative weapons like antimicrobial peptides to strengthen existing tuberculosis artillery is urgently required. Considering the antimycobacterial potential of Human Beta Defensin 1 (HBD-1) along with isoniazid, the present study was designed to explore the ability of HBD-1 to act against active and dormant M. tb. HBD-1 was screened in silico using antimicrobial peptide prediction servers to identify its short antimicrobial motif. The activity of both HBD-1 and its selected motif (Pep B) was determined at different concentrations against actively growing M. tb in vitro and ex vivo in monocyte derived macrophages (MDMs). Log phase M. tb was grown along with HBD-1 and Pep B for 7 days. M. tb infected MDMs were treated with HBD-1 and Pep B for 72 hours. Thereafter, colony forming unit (CFU) enumeration was performed to determine activity of both peptides against actively growing in vitro and intracellular M. tb. The dormant M. tb models were prepared by following two approaches and treated with different concentrations of HBD-1 and Pep B. Firstly, 20-22 days old M. tbH37Rv was grown in potassium deficient Sauton media for 35 days. The presence of dormant bacilli was confirmed by Nile red staining. Dormant bacilli were further treated with rifampicin, isoniazid, HBD-1 and its motif for 7 days. The effect of both peptides on latent bacilli was assessed by colony forming units (CFU) and most probable number (MPN) enumeration. Secondly, human PBMC granuloma model was prepared by infecting PBMCs seeded on collagen matrix with M. tb(MOI 0.1) for 10 days. Histopathology was done to confirm granuloma formation. The granuloma thus formed was incubated for 72 hours with rifampicin, HBD-1 and Pep B individually. Difference in bacillary load was determined by CFU enumeration. The minimum inhibitory concentrations of HBD-1 and Pep B restricting growth of mycobacteria in vitro were 2μg/ml and 20μg/ml respectively. The intracellular mycobacterial load was reduced significantly by HBD-1 and Pep B at 1μg/ml and 5μg/ml respectively. Nile red positive bacterial population, high MPN/ low CFU count and tolerance to isoniazid, confirmed the formation of potassium deficienybaseddormancy model. HBD-1 (8μg/ml) showed 96% and 99% killing and Pep B (40μg/ml) lowered dormant bacillary load by 68.89% and 92.49% based on CFU and MPN enumeration respectively. Further, H&E stained aggregates of macrophages and lymphocytes, acid fast bacilli surrounded by cellular aggregates and rifampicin resistance, indicated the formation of human granuloma dormancy model. HBD-1 (8μg/ml) led to 81.3% reduction in CFU whereas its motif Pep B (40μg/ml) showed only 54.66% decrease in bacterial load inside granuloma. Thus, the present study indicated that HBD-1 and its motif are effective antimicrobial players against both actively growing and dormant M. tb. They should be further explored to tap their potential to design a powerful weapon for combating tuberculosis.

Keywords: antimicrobial peptides, dormant, human beta defensin 1, tuberculosis

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Authors: Nandini Nalika, Suhel Parvez

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Nanotechnology has emerged to play a vital role in developing all through the industrial world with an immense production of nanomaterials including nanoparticles (NPs). Many toxicological studies have confirmed that due to unique small size and physico-chemical properties of NPs (1-100nm), they can be potentially hazardous. Metallic NPs of small size have been shown to induce higher levels of cellular oxidative stress and can easily pass through the Blood Brain Barrier (BBB) and significantly accumulate in brain. With the wide applications of titanium dioxide nanoparticles (TNPs) in day-to-day life in form of cosmetics, paints, sterilisation and so on, there is growing concern regarding the deleterious effects of TNPs on central nervous system and mitochondria appear to be important cellular organelles targeted to the pro-oxidative effects of NPs and an important source that contribute significantly for the production of reactive oxygen species after some toxicity or an injury. The aim of our study was to elucidate the effect of TNPs in anatase form with different concentrations (5-50 µg/ml) following with various oxidative stress markers in isolated brain mitochondria as an in vitro model. Oxidative stress was determined by measuring the different oxidative stress markers like lipid peroxidation as well as the protein carbonyl content which was found to be significantly increased. Reduced glutathione content and major glutathione metabolizing enzymes were also modulated signifying the role of glutathione redox cycle in the pathophysiology of TNPs. The study also includes the mitochondrial enzymes (Complex 1, Complex II, complex IV, Complex V ) and the enzymes showed toxicity in a relatively short time due to the effect of TNPs. The study provide a range of concentration that were toxic to the neuronal cells and data pointing to a general toxicity in brain mitochondria by TNPs, therefore, it is in need to consider the proper utilization of NPs in the environment.

Keywords: mitochondria, nanoparticles, brain, in vitro

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Authors: Twin Aji Kusumagiani, Eleonora Agustine, Dini Fitriani

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The value changes of soil physical properties in the agricultural area are giving impacts on soil fertility. This can be caused by excessive usage of inorganic fertilizers and imbalances on organic fertilization. Soil physical parameters that can be measured include soil electrical conductivity, water content volume, soil porosity, dielectric permittivity, etc. This study used the electrical conductivity and volume water content as the measured physical parameters. The study was conducted on volcanic soil obtained from agricultural land conditioned with NPK fertilizer and salt in a certain amount. The dimension of the conditioned soil being used is 1 x 1 x 0.5 meters. By using this method, we can delineate the soil electrical conductivity value of land due to changes in the provision of inorganic NPK fertilizer and the salinity in the soil. Zone with the additional 1 kg of salt has the dimension of 60 cm in width, 20 cm in depth and 1 cm in thickness while zone with the additional of 10 kg NPK fertilizer has the dimensions of 70 cm in width, 20 cm in depth and 3 cm in thickness. This salt addition resulted in EC values changes from the original condition. Changes of the EC value tend to occur at a depth of 20 to 40 cm on the line 1B at 9:45 dS/cm and line 1C of 9.35 dS/cm and tend to have the direction to the Northeast.

Keywords: EC, electrical conductivity, VWC, volume water content, NPK fertilizer, salt, volcanic soil

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Authors: Rashi Rajput, Manisha Singh

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Escitalopram oxalate (ETP), an FDA approved antidepressant drug from the category of SSRI (selective serotonin reuptake inhibitor) and is used in treatment of general anxiety disorder (GAD), major depressive disorder (MDD).When taken orally, it is metabolized to S-demethylcitalopram (S-DCT) and S-didemethylcitalopram (S-DDCT) in the liver with the help of enzymes CYP2C19, CYP3A4 and CYP2D6. Hence, causing side effects such as dizziness, fast or irregular heartbeat, headache, nausea etc. Therefore, targeted and sustained drug delivery will be a helpful tool for increasing its efficacy and reducing side effects. The present study is designed for formulating mucoadhesive nanoparticle formulation for the same Escitalopram loaded polymeric nanoparticles were prepared by ionic gelation method and characterization of the optimised formulation was done by zeta average particle size (93.63nm), zeta potential (-1.89mV), TEM (range of 60nm to 115nm) analysis also confirms nanometric size range of the drug loaded nanoparticles along with polydispersibility index of 0.117. In this research, we have studied the in vitro drug release profile for ETP nanoparticles, through a semi permeable dialysis membrane. The three important characteristics affecting the drug release behaviour were – particle size, ionic strength and morphology of the optimised nanoparticles. The data showed that on increasing the particle size of the drug loaded nanoparticles, the initial burst was reduced which was comparatively higher in drug. Whereas, the formulation with 1mg/ml chitosan in 1.5mg/ml tripolyphosphate solution showed steady release over the entire period of drug release. Then this data was further validated through mathematical modelling to establish the mechanism of drug release kinetics, which showed a typical linear diffusion profile in optimised ETP loaded nanoparticles.

Keywords: ionic gelation, mucoadhesive nanoparticle, semi-permeable dialysis membrane, zeta potential

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Authors: Rajendra Jangde, Deependra Singh

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Present work was based with objective to release of the antimicrobial and debriding agent in sustained manner at the wound surface. In order to provide a long-lasting antimicrobial action and moist environment on wound space, Biocompatible moist system was developed for complete healing. In the present study, a biocompatible moist system of PVA-gelatin hydrogel was developed capable of carrying multiple drugs- Quercetin and Cabopol in controlled manner for effective and complete wound healing. Carbopol and Quercetin were prepared by thin film hydration techniques and optimized system was incorporated in PVA-Gelatin slurry. PVA-Gelatin hydrogels were prepared by freeze thaw method. The prepared dispersion was casted into films to prepare multiphase hydrogel system and characterized by in vitro and in vivo studies. Results revealed the uniform dispersion of microspheres in a three-dimensional matrix of the PVA-Gelatin hydrogel observed at different magnifications. The in vitro release data showed typical biphasic release pattern, i.e., a burst release followed by a slower sustained release for 5 days. Prepared system was found to be stable under both normal and accelerated conditions. Histopathological study showed significant (p<0.05) increase in fibroblast cells, collagen fibres and blood vessels formation. All parameters such as wound contraction, tensile strength, histopathological and biochemical parameters- hydroxyproline content, protein level, etc. were observed significant (p<0.05) in comparison to control group. Present results suggest an accelerated re-epithelialization under moist wound environment with delivery of multiple drugs effective at different stages of wound healing cascade with minimum disturbance of wound bed.

Keywords: multiphase hydrogel, optimization quercetin, wound healing

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Authors: Sineenat Kempubpha, Phornpa-Ngan Muadmuang, Putthamas Phetmuangprab, Surin Promphet, Sopita Bandit

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Introduction: Discarded pericarp of mangosteen (Garcinia mangostana) is a benefit to be developed as veterinary phytopharmacal products since it made up of abundance pharmacological active compounds. The active compounds of mangosteen pericarp not only act as an antihistamine, an anti-inflammatory, heart disease and HIV therapeutic substances but also act as antibacterial and antifungal agents. Aim: This study was an in vitro procedural attempt to determine the antibacterial effects of mangosteen pericarp 95% ethanol extract on the main causative pathogen of canine superficial pyoderma, Staphylococcus pseudintermedius. Methods: S. pseudintermedius were collected from various sites of the skin of canine superficial pyoderma dogs and were revived and lawn cultured. The S. pseudintermedius growth inhibition study was determined by disc diffusion technique, the mangosteen pericarp crude extracted was dissolved in 3 types of solvents (95% ethanol, 2% DMSO and distilled water, respectively). The micro broth dilution technique was used for determining both minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values. Statistical analysis was done by calculating the mean of the zones of inhibition of tested microorganisms. Results: S. pseudintermedius growth inhibition study showed that the inhibition efficacy of 95% ethanol was greater than the inhibition efficacy of 2% DMSO and distilled water (9.10±0.18 mm, 6.95±0.60 mm and 6.80±0.18 mm, respectively). The MIC value was 125 µg/ml and the MBC value was 1 mg/ml. Conclusion: Mangosteen pericarp extract dissolved with 95% ethanol showed the highest zone of inhibition against the tested microorganisms. The MIC value was 125 µg/ml and the MBC value was 1 mg/ml which suggests its potent antibacterial action against S. pseudintermedius. However, further analytical studies are needed to isolate the key molecules of mangosteen pericarp for higher effect on canine superficial pyoderma microorganism therapeutic products.

Keywords: mangosteen, Garcinia mangostana, Staphylococcus pseudintermedius, canine superficial pyoderma, in vitro study

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Authors: Khayra Zerrouki, Noureddine Djebli, Esra Eroglu, Afife Mat, Ozhan Gul

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Neurodegenerative diseases of the human brain comprise a variety of disorders that affect an increasing percentage of the population. Alzheimer’s disease (AD) is a complex, multifactorial, heterogeneous mental illness, which is characterized by an age-dependent loss of memory and an impairment of multiple cognitive functions, but this 10 last years it concerns the population most and most young. Hypericum perforatum has traditionally been used as an external anti-inflammatory and healing remedy for the treatment of swellings, wounds and burns, diseases of the alimentary tract and psychological disorders. It is currently of great interest due to new and important therapeutic applications. In this study, the chemical composition of methanolic extract of Hypericum perforatum (HPM) was analysed by using high performance liquid chromatography – diode array detector (HPLC-DAD). The in vitro antioxidant activity of HPM was evaluated by using several antioxidant tests. HSM exhibits inhibitory capacity against posphatidylcholine liposome peroxidation, induced with iron and ascorbic acid, scavenge DPPH and superoxide radicals and act as reductants. The cytotoxic activity of HSM was also determined by using MTT cell viability assay on HeLa and NRK-52E cell lines. The in vivo activity studies in Swiss mice were determined by using behavioral, memory tests and histological study. According to tests results HPM that may be relevant to the treatment of cognitive disorders. The results of chemical analysis showed a hight level of hyperforin and quercitin that had an important antioxidant activity proved in vitro with the DPPH, anti LPO and SOD; this antioxidant activity was confirmed in vivo after the non-toxic results by means of improvement in behavioral and memory than the reducing shrunken in pyramidal cells of mice brains.

Keywords: AlCl3, alzheimer, mice, neuroprotective, neurotoxicity, phytotherapy

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Authors: Tooba N. Shamsi, Romana Perveen, Sadaf Fatima

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Protease Inhibitors (PIs) are widespread in nature, produced by animals, plants and microorganisms. They play vital role in various biological activities by keeping a check on activity of proteases. Present study aims to investigate antioxidant and anti-inflammatory properties of PPI from Cajanus cajan (CCTI) and Phaseolus limensis (LBTI). PPI was purified from C. cajan (PUSA-992) by ammonium sulfate precipitation followed by ion exchange chromatography. The anti-oxidant activity was analyzed by two most common radical scavenging assays of FRAP (ferric reducing antioxidant power) and DPPH (1,1- diphenyl-2-picrylhydrazyl). Also, in-vitro anti-inflammatory activity was evaluated using albumin denaturation assay and membrane stabilization assay at different concentrations. Ascorbic acid and aspirin were used as a standards for antioxidant and anti-inflammatory assays respectively. The PPIs were also checked for antimicrobial activity against a number of bacterial strains. The CCTI and LBTI showed DPPH radical scavenging activity in a concentration–dependent manner with IC50 values 544 µg/ml and 506 µg/ml respectively comparative to ascorbic acid which was 258 µg/ml. Following FRAP assay, it was evaluated that LBTI had 87.5% and CCTI showed 84.4% antioxidant activity, taking value of standard ascorbic acid to be 100%. The PPIs also showed in-vitro anti‐inflammatory activity by inhibiting the heat induced albumin denaturation with IC50 values of 686 µg/ml and 615 µg/ml for CCTI and LBTI respectively compared to the standard (aspirin) which was 70.8 µg/ml. Red blood cells membrane stabilization with IC50 values of 641 µg/ml and 587 µg/ml for CCTI and LBTI respectively against aspirin which showed IC50 value of 70.4 µg/ml. PPIs showed antibacterial activity against 7 known strains while there was apparently no action against fungi.

Keywords: Cajanus cajan, Phaseolus limensis, Lima beans, protein protease inhibitor, antioxidant, anti-inflammatory, antimicrobial activity

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Authors: Wiam El Kheir, Anais Dumais, Maude Beaudoin, Bernard Marcos, Nick Virgilio, Benoit Paquette, Nathalie Faucheux, Marc-Antoine Lauzon

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The high infiltrative pattern of glioblastoma multiforme cells (GBM) is the main cause responsible for the actual standard treatments failure. The tumor high heterogeneity, the interstitial fluid flow (IFF) and chemokines guides GBM cells migration in the brain parenchyma resulting in tumor recurrence. Drug delivery systems emerged as an alternative approach to develop effective treatments for the disease. Some recent studies have proposed to harness the effect CXC-lchemokine-ligand-12 to direct and control the cancer cell migration through delivery system. However, the dynamics of the brain environment on the delivery system remains poorly understood. Nanoparticles (NPs) and hydrogels are known as good carriers for the encapsulation of different agents and control their release. We studied the release of CXCL12 (free or loaded into NPs) from an alginate-based hydrogel under static and indirect perfusion (IP) conditions. Under static conditions, the main phenomena driving CXCL12 release from the hydrogel was diffusion with the presence of strong interactions between the positively charged CXCL12 and the negatively charge alginate. CXCL12 release profiles were independent from the initial mass loadings. Afterwards, we demonstrated that the release could tuned by loading CXCL12 into Alginate/Chitosan-Nanoparticles (Alg/Chit-NPs) and embedded them into alginate-hydrogel. The initial burst release was substantially attenuated and the overall cumulative release percentages of 21%, 16% and 7% were observed for initial mass loadings of 0.07, 0.13 and 0.26 µg, respectively, suggesting stronger electrostatic interactions. Results were mathematically modeled based on Fick’s second law of diffusion framework developed previously to estimate the effective diffusion coefficient (Deff) and the mass transfer coefficient. Embedding the CXCL12 into NPs decreased the Deff an order of magnitude, which was coherent with experimental data. Thereafter, we developed an in-vitro 3D model that takes into consideration the convective contribution of the brain IFF to study CXCL12 release in an in-vitro microenvironment that mimics as faithfully as possible the human brain. From is unique design, the model also allowed us to understand the effect of IP on CXCL12 release in respect to time and space. Four flow rates (0.5, 3, 6.5 and 10 µL/min) which may increase CXCL12 release in-vivo depending on the tumor location were assessed. Under IP, cumulative percentages varying between 4.5-7.3%, 23-58.5%, 77.8-92.5% and 89.2-95.9% were released for the three initial mass loadings of 0.08, 0.16 and 0.33 µg, respectively. As the flow rate increase, IP culture conditions resulted in a higher release of CXCL12 compared to static conditions as the convection contribution became the main driving mass transport phenomena. Further, depending on the flow rate, IP had a direct impact on CXCL12 distribution within the simulated brain tissue, which illustrates the importance of developing such 3D in-vitro models to assess the efficiency of a delivery system targeting the brain. In future work, using this very model, we aim to understand the impact of the different phenomenon occurring on GBM cell behaviors in response to the resulting chemokine gradient subjected to various flow while allowing them to express their invasive characteristics in an in-vitro microenvironment that mimics the in-vivo brain parenchyma.

Keywords: 3D culture system, chemokines gradient, glioblastoma multiforme, kinetic release, mathematical modeling

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Authors: Martin Hewing, Katharina Hölzle

Abstract:

Organizations increasingly use environmental stimuli and ideas from users within participatory innovation processes in order to tap new sources of knowledge. The research presented in this article focuses on users who shape the distant edges of markets and currently are not using products and services from a domain– so called potential users. Those users at the peripheries are perceived to contribute more novel information, by which they better reflect shifts in needs and behavior than current users in the core market. Their contributions in collaborative and creative problem-solving processes and how they generate ideas for discontinuous innovations are of particular interest. With an experimental design, we compare ideas from potential and current users and analyze the effects of cognitive distance in collaboration and the utilization of explicit and tacit knowledge. We find potential users to generate more original ideas, particularly when they collaborate with someone experienced within the domain. Their ideas are most obviously characterized by an increased level of surprise and unusualness compared to dominant designs, which is rooted in contexts and does not require technological leaps. Collaboration with potential users can therefore result in new ways to leverage technological competences. Furthermore, the cross-fertilization arising from cognitive distance between a potential and a current user is asymmetric due to differences in the nature of their utilized knowledge and personal objectives. This paper discusses implications for innovation research and the management of early innovation processes.

Keywords: user collaboration, co-creation, discontinuous innovation, innovation research

Procedia PDF Downloads 482

Authors: M. H. Rajikin, S. M. M. Syairah, A. R. Sharaniza

Abstract:

Effects of nicotine on pre-partum body weight and preimplantation embryonic development has been reported previously. Present study was conducted to determine the effects of annatto (Bixa orellana)-derived delta-tocotrienol (TCT) (with presence of 10% gamma-TCT isomer) on the nicotine-induced reduction in body weight and 8-cell embryonic growth in mice. Twenty four 6-8 weeks old (23-25g) female balb/c mice were randomly divided into four groups (G1-G4; n=6). Those groups were subjected to the following treatments for 7 consecutive days: G1 (control) were gavaged with 0.1 ml tocopherol stripped corn oil, G2 was subcutaneously (s.c.) injected with 3 mg/kg/day of nicotine, G3 received concurrent treatment of nicotine (3 mg/kg/day) and 60 mg/kg/day of δ-TCT mixture (contains 90% delta & 10% gamma isomers) and G4 was given 60 mg/kg/day of δ-TCT mixture alone. Body weights were recorded daily during the treatment. On Day 8, females were superovulated with 5 IU Pregnant Mare’s Serum Gonadotropin (PMSG) for 48 hours followed with 5 IU human Chorionic Gonadotropin (hCG) before mated with males at the ratio of 1:1. Females were sacrificed by cervical dislocation for embryo collection 48 hours post-coitum. Collected embryos were cultured in vitro. Results showed that throughout Day 1 to Day 7, the body weight of nicotine treated group (G2) was significantly lower (p<0.05) than that of G1, G3 and G4. Intervention with δ-TCT mixture (G3) managed to increase the body weight close to the control group. This is also observed in the group treated with δ-TCT mixture alone (G4). The development of 8-cell embryos following in vitro culture (IVC) was totally inhibited in G2. Intervention with δ-TCT mixture (G3) resulted in the production of 8-cell embryos, although it was not up to that of the control group. Treatment with δ-TCT mixture alone (G4) caused significant increase in the average number of produced 8-cell embryo compared to G1. Present data indicated that δ-TCT mixture was able to reverse the body weight loss in nicotine treated mice and the development of 8-cell embryos was also improved.

Keywords: δ-tocotrienol, body weight, nicotine, preimplantation embryonic development

Procedia PDF Downloads 333

Authors: K. K. Balavenkatraman, B. Bertschi, K. Bigot, A. Grevot, A. Doelemeyer, S. D. Chibout, A. Wolf, F. Pognan, N. Manevski, O. Kretz, P. Swart, K. Litherland, J. Ashton-Chess, B. Ling, R. Wettstein, D. J. Schaefer

Abstract:

Skin toxicity is poorly detected during preclinical studies, and drug-induced side effects in humans such as rashes, hyperplasia or more serious events like bullous pemphigus or toxic epidermal necrolysis represent an important hurdle for clinical development. In vitro keratinocyte-based epidermal skin models are suitable for the detection of chemical-induced irritancy, but do not recapitulate the biological complexity of full skin and fail to detect potential serious side-effects. Normal healthy skin explants may represent a valuable complementary tool, having the advantage of retaining the full skin architecture and the resident immune cell diversity. This study investigated several conditions for the maintenance of good morphological structure after several days of culture and the retention of phase II metabolism for 24 hours in skin explants in vitro. Human skin samples were collected with informed consent from patients undergoing plastic surgery and immediately transferred and processed in our laboratory by removing the underlying dermal fat. Punch biopsies of 4 mm diameter were cultured in an air-liquid interface using transwell filters. Different cultural conditions such as the effect of calcium, temperature and cultivation media were tested for a period of 14 days and explants were histologically examined after Hematoxylin and Eosin staining. Our results demonstrated that the use of Williams E Medium at 32°C maintained the physiological integrity of the skin for approximately one week. Upon prolonged incubation, the upper layers of the epidermis become thickened and some dead cells are present. Interestingly, these effects were prevented by addition of EGFR inhibitors such as Afatinib or Erlotinib. Phase II metabolism of the skin such as glucuronidation (4-methyl umbeliferone), sulfation (minoxidil), N-acetyltransferase (p-toluidene), catechol methylation (2,3-dehydroxy naphthalene), and glutathione conjugation (chlorodinitro benzene) were analyzed by using LCMS. Our results demonstrated that the human skin explants possess metabolic activity for a period of at least 24 hours for all the substrates tested. A time course for glucuronidation with 4-methyl umbeliferone was performed and a linear correlation was obtained over a period of 24 hours. Longer-term culture studies will indicate the possible evolution of such metabolic activities. In summary, these results demonstrate that human skin explants maintain a normal structure for several days in vitro and are metabolically active for at least the first 24 hours. Hence, with further characterisation, this model may be suitable for the study of drug-induced toxicity.

Keywords: human skin explant, phase II metabolism, epidermal growth factor receptor, toxicity

Procedia PDF Downloads 264

Authors: Marisa Rio, Sharanya Bola, Richard H. W. Funk, Gerald Gerlach

Abstract:

Cell migration, wound healing and regeneration are some of the physiological phenomena in which electric fields (EFs) have proven to have an important function. Physiologically, cells experience electrical signals in the form of transmembrane potentials, ion fluxes through protein channels as well as electric fields at their surface. As soon as a wound is created, the disruption of the epithelial layers generates an electric field of ca. 40-200 mV/mm, directing cell migration towards the wound site, starting the healing process. In vitro electrotaxis, experiments have shown cells respond to DC EFs polarizing and migrating towards one of the poles (cathode or anode). A standard electrotaxis experiment consists of an electrotaxis chamber where cells are cultured, a DC power source and agar salt bridges that help delaying toxic products from the electrodes to attain the cell surface. The electric field strengths used in such an experiment are uniform and homogeneous. In contrast, the endogenous electric field strength around a wound tend to be multi-field and non-homogeneous. In this study, we present a custom device that enables electrotaxis experiments in non-homogeneous DC electric fields. Its main feature involves the replacement of conventional metallic electrodes, separated from the electrotaxis channel by agarose gel bridges, through electrolyte-filled microchannels. The connection to the DC source is made by Ag/AgCl electrodes, incased in agarose gel and placed at the end of each microfluidic channel. An SU-8 membrane closes the fluidic channels and simultaneously serves as the single connection from each of them to the central electrotaxis chamber. The electric field distribution and current density were numerically simulated with the steady-state electric conduction module from ANSYS 16.0. Simulation data confirms the application of nonhomogeneous EF of physiological strength. To validate the biocompatibility of the device cellular viability of the photoreceptor-derived 661W cell line was accessed. The cells have not shown any signs of apoptosis, damage or detachment during stimulation. Furthermore, immunofluorescence staining, namely by vinculin and actin labelling, allowed the assessment of adhesion efficiency and orientation of the cytoskeleton, respectively. Cellular motility in the presence and absence of applied DC EFs was verified. The movement of individual cells was tracked for the duration of the experiments, confirming the EF-induced, cathodal-directed motility of the studied cell line. The in vitro monolayer wound assay, or “scratch assay” is a standard protocol to quantitatively access cell migration in vitro. It encompasses the growth of a confluent cell monolayer followed by the mechanic creation of a scratch, representing a wound. Hence, wound dynamics was monitored over time and compared for control and applied the electric field to quantify cellular population motility.

Keywords: DC non-homogeneous electric fields, electrotaxis, microfluidic biochip, wound healing

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Authors: Kuan Yin Hsiao, Shyh Ming Kuo, Yi Jhen Wu, Chin Wen Chuang, Chuen-Fu Lin, Wei-qing Yang, Han Hsiang Huang

Abstract:

Anti-tumor effects of natural products, like compounds from marine sponges and soft corals, have been investigated for decades. Polymeric nanoparticles prepared from biodegradable and biocompatible molecules, such as Hyaluronan (HA), Chitosan (CHI) and gelatin have been widely studied. Encapsulation of anti-cancer therapies by the biopolymeric nanoparticles in drug delivery system is potentially capable of improving the therapeutic effects and attenuating their toxicity. In the current study, the anti-bladder cancer effects of heteronemin extracted from the sponge Hippospongia sp. with or without HA and CHI nanoparticle encapsulation were assessed and evaluated in vitro. Results showed that IC50 (half maximal inhibitory concentration) of heteronemin toward T24 human bladder cancer cell viability is approximately 0.18 µg/mL. Both plain and HA nanoparticles-encapsulated heteronemin at 0.2 and 0.4 µg/mL significantly reduced T24 cell viability (P<0.001) while HA nanoparticles-encapsulated heteronemin showed weaker viability-inhibitory effects on L929 fibroblasts compared with plain heteronemin at the identical concentrations. HA and CHI nanoparticles-encapsulated heteronemin exhibited significantly stronger inhibitory effects against migration of T24 human bladder cancer cell than those exerted by plain heteronemin at the same concentrations (P<0.001). The flow cytometric results showed that 0.2 µg/mL HA and CHI nanoparticles-encapsulated heteronemin induced higher early apoptosis rate than that induced by plain heteronemin at the same concentration. These results show that HA and CHI nanoparticle encapsulation is able to elevate anti-migratory and apoptosis-inducing effects exerted by heteronemin against bladder cancer cells in vitro. The in vivo anti-bladder cancer effects of the compound with or without HA/CHI nanoparticle encapsulation will be further investigated and examined using murine tumor models. The data obtained from this study will extensively evaluate of the anti-bladder cancer effects of heteronemin as well as HA/CHI-encapsulated heteronemin and pave a way to develop potential bladder cancer treatment.

Keywords: heteronemin, nanoparticles, hyaluronan, chitosan, bladder cancer

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Authors: Brian H. Marsh

Abstract:

Nitrogen fertility is an important component for optimum potato yield and quality. Best management practices are necessary in regards to N applications to achieve these goals without applying excess N with may contribute to ground water contamination. Eight potato fields in the Southern San Joaquin Valley were sampled for nitrogen inputs and uptake, tuber and vine dry matter and residual soil nitrate-N. The fields had substantial soil nitrate-N prior to the potato crop. Nitrogen fertilizer was applied prior to planting and in irrigation water as needed based on in-season petiole sampling in accordance with published recommendations. Average total nitrogen uptake was 237 kg ha^-1 on 63.5 Mg ha^-1 tuber yield and nitrogen use efficiency was very good at 81 percent. Sixty-nine percent of the plant nitrogen was removed in tubers. Soil nitrate-N increased 14 percent from pre-plant to post-harvest averaged across all fields and was generally situated in the upper soil profile. Irrigation timing and amount applied did not move water into the lower profile except for a single location where nitrate also moved into the lower soil profile. Pre-plant soil analysis is important information to be used. Rotation crops having deeper rooting growth would be able to utilize nitrogen that remained in the soil profile.

Keywords: potato, nitrogen fertilization, irrigation management, leaching potential

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Authors: M. Das, B. P. Sarma, G. Ahmed

Abstract:

Human diet can be a major source of antioxidants. Poly¬phenols, which are organic compounds present in the regular human diet, have good antioxidant property. Most of the diseases are detected too late and that cause irre¬versible damage to the body. Therefore food that forms the natural source of antioxidants can prevent free radi¬cals from damaging our body tissues. Dillenia indica and Garcinia penducalata are two major fruits, easily available in Assam, North eastern Indian state. In the present study, the in vitro antioxi¬dant properties of the fruits of these plants are compared as the decoction of these fruits form a major part of Assamese cuisine. DPPH free radical scavenging activity of the methanol, petroleum ether and water extracts of G. penducalata and D. indica fruits were carried out by the methods of Cotelle A et al. (1996). Different concentrations ranging from 10–110 ug/ml of the extracts were added to 100 uM of DPPH (2,2, Diphenyl-2-picryl hydrazyl) and the absor¬bance was read at 517 nm after incubation. Ascorbic acid was used as the standard. Different concentrations of the methanol, petroleum ether and water extracts of G. penducalata and D. indica fruits were mixed with sodium nitroprusside and incubated. Griess reagent was added to the mixtures and their optical density was read at 546 nm following the method of Marcocci et al. (1994). Ascorbic acid was used as the standard. In order to find the scavenging activity of the extracts against hydroxyl radicals, the method of Kunchandy & Ohkawa (1990) was followed.The superoxide scavenging activity of the methanol, petroleum ether and water extracts of the fruits was deter¬mined by the method of Robak & Gryglewski (1998).Six replicates were maintained in each of the experiments and their SEM was evaluated based on which, non linear regres¬sion (curve fit), exponential growth were derived to calculate the IC50 values of the SAWE and standard compounds. All the statistical analyses were done by using paired t test. The hydroxyl radical scavenging activity of the various extracts of D. indica exhibited IC50 values < 110 ug/ml concentration, the scavenging activity of the extracts of G. penducalata was surprisingly>110 ug/ml.Similarly the oxygen free radical scavenging activity of the different extracts of D. indica exhibited an IC50 value of <110 ug/ml but the methanolic extract of the same exhib¬ited a better free radical scavenging activity compared to that of vitamin C. The methanolic extract of D. indica exhibited an IC50 value better than that of vitamin C. The DPPH scavenging activities of the various extracts of D. indica and G. penducalata were <110 ug/ml but the methanolic extract of D. indica exhibited an IC50 value bet¬ter than that of vitaminc C.The higher amounts of phenolic content in the methanolic extract of D. indica might be one of the major causes for its enhanced in vitro antioxidant activity.The present study concludes that Dillenia indica and Garcinia penducalata both possesses anti oxidant activi¬ties. The anti oxidant activity of Dillenia indica is superior to that of Garcinia penducalata due to its higher phenolic content

Keywords: antioxidants, free radicals, phenolic, scavenging

Procedia PDF Downloads 572

Authors: Q. Ni, J. A. Serna, D. Holland, X. Wang

Abstract:

The objective of this study is to develop Nuclear Magnetic Resonance (NMR) techniques to enhance bone related research applied on normal and disuse (Biglycan knockout) mice bone in vitro by using both low-field and high-field NMR simultaneously. It is known that the total amplitude of T₂ relaxation envelopes, measured by the Carr-Purcell-Meiboom-Gill NMR spin echo train (CPMG), is a representation of the liquid phase inside the pores. Therefore, the NMR CPMG magnetization amplitude can be transferred to the volume of water after calibration with the NMR signal amplitude of the known volume of the selected water. In this study, the distribution of mobile water, porosity that can be determined by using low-field (20 MHz) CPMG relaxation technique, and the pore size distributions can be determined by a computational inversion relaxation method. It is also known that the total proton intensity of magnetization from the NMR free induction decay (FID) signal is due to the water present inside the pores (mobile water), the water that has undergone hydration with the bone (bound water), and the protons in the collagen and mineral matter (solid-like protons). Therefore, the components of total mobile and bound water within bone that can be determined by low-field NMR free induction decay technique. Furthermore, the bound water in solid phase (mineral and organic constituents), especially, the dominated component of calcium hydroxyapatite (Ca₁₀(OH)₂(PO₄)₆) can be determined by using high-field (400 MHz) magic angle spinning (MAS) NMR. With MAS technique reducing NMR spectral linewidth inhomogeneous broadening and susceptibility broadening of liquid-solid mix, in particular, we can conduct further research into the ¹H and ³¹P elements and environments of bone materials to identify the locations of bound water such as OH- group within minerals and bone architecture. We hypothesize that with low-field and high-field magic angle spinning NMR can provide a more complete interpretation of water distribution, particularly, in bound water, and these data are important to access bone quality and predict the mechanical behavior of bone.

Keywords: bone, mice bone, NMR, water in bone

Procedia PDF Downloads 151

Authors: Thomas Louis, Irina Primac, Florent Morfoisse, Tania Durre, Silvia Blacher, Agnes Noel

Abstract:

In vitro and in vivo lymphangiogenesis assays are essential for the identification of potential lymphangiogenic agents and the screening of pharmacological inhibitors. In the present study, we analyse three biological models: in vitro lymphatic endothelial cell spheroids, in vivo ear sponge assay, and in vivo lymph node colonisation by tumour cells. These assays provide suitable 3D models to test pro- and anti-lymphangiogenic factors or drugs. 3D images were acquired by confocal laser scanning and light sheet fluorescence microscopy. Virtual scan microscopy followed by 3D reconstruction by image aligning methods was also used to obtain 3D images of whole large sponge and ganglion samples. 3D reconstruction, image segmentation, skeletonisation, and other image processing algorithms are described. Fixed and time-lapse imaging techniques are used to analyse lymphatic endothelial cell spheroids behaviour. The study of cell spatial distribution in spheroid models enables to detect interactions between cells and to identify invasion hierarchy and guidance patterns. Global measurements such as volume, length, and density of lymphatic vessels are measured in both in vivo models. Branching density and tortuosity evaluation are also proposed to determine structure complexity. Those properties combined with vessel spatial distribution are evaluated in order to determine lymphangiogenesis extent. Lymphatic endothelial cell invasion and lymphangiogenesis were evaluated under various experimental conditions. The comparison of these conditions enables to identify lymphangiogenic agents and to better comprehend their roles in the lymphangiogenesis process. The proposed methodology is validated by its application on the three presented models.

Keywords: 3D image segmentation, 3D image skeletonisation, cell invasion, confocal microscopy, ear sponges, light sheet microscopy, lymph nodes, lymphangiogenesis, spheroids

Procedia PDF Downloads 351

Authors: Amanina Iymia Jeffree, Reena Thriumani, Mohammad Iqbal Omar, Ammar Zakaria, Yumi Zuhanis Has-Yun Hashim, Ali Yeon Md Shakaff

Abstract:

Metabolite profiling is a strategy to be approached in the pattern recognition method focused on three types of cancer cell line that driving the most to death specifically lung, breast, and colon cancer. The purpose of this study was to discriminate the VOCs pattern among cancerous and control group based on metabolite profiling. The sampling was executed utilizing the cell culture technique. All culture flasks were incubated till 72 hours and data collection started after 24 hours. Every running sample took 24 minutes to be completed accordingly. The comparative metabolite patterns were identified by the implementation of headspace-solid phase micro-extraction (HS-SPME) sampling coupled with gas chromatography-mass spectrometry (GCMS). The optimizations of the main experimental variables such as oven temperature and time were evaluated by response surface methodology (RSM) to get the optimal condition. Volatiles were acknowledged through the National Institute of Standards and Technology (NIST) mass spectral database and retention time libraries. To improve the reliability of significance, it is of crucial importance to eliminate background noise which data from 3rd minutes to 17th minutes were selected for statistical analysis. Targeted metabolites, of which were annotated as known compounds with the peak area greater than 0.5 percent were highlighted and subsequently treated statistically. Volatiles produced contain hundreds to thousands of compounds; therefore, it will be optimized by chemometric analysis, such as principal component analysis (PCA) as a preliminary analysis before subjected to a pattern classifier for identification of VOC samples. The volatile organic compound profiling has shown to be significantly distinguished among cancerous and control group based on metabolite profiling.

Keywords: in vitro cancer cell line, metabolite profiling, pattern recognition, volatile organic compounds

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Authors: Maryam Farzaneh, Seyyedeh Nafiseh Hassani, Hossein Baharvand

Abstract:

Objective: Chicken gonadal tissue has a two population such primordial germ cells (PGCs) and stromal cells (somatic cells). PGCs and embryonic germ cells (EGCs) that is a pluripotent type of PGCs in long-term culture are suitable sources for the production of chicken pluripotent stem cell lines, transgenic birds, vaccine and recombinant protein production. In general, the effect of growth factors such bFGF and mouse LIF on derivation of PGCs in vitro are important and in this study we could see the unique effect of small molecules such PD032 and SB43 as a chemical, in comparison to growth factors. Materials and Methods: After incubation of fertilized chicken egg up to 6 days and isolation of primary gonadal tissues and culture of mixed cells like PGCs and stromal cells. PGCs proliferate in the present of fetal calf serum (FCS) and small molecules and in another group bFGF, that these factors are important for PGCs culture and derivation. Somatic cells produce a multilayer feeder under the PGCs in primary culture and PGCs make a small cluster under these cells. Results: In present of small molecules and high volume of FCS (15%), the present of EGCs as a pluripotent stem cells were clear four weeks, that they had a positive immune-staining and periodic acid-Schiff staining (PAS), but in present of growth factors like bFGF without any chemicals, the present of PGCs were clear but after 7 until 10 days, there were disappear. Conclusion: Until now we have seen many researches about derivation and maintenance of chicken PGCs, in the hope of understanding the mechanisms that occur during germline development and production of a therapeutic product by transgenic birds. There are still many unknowns in this area and this project will try to have efficient conditions for identification of suitable culture medium for long-term culture of PGCs in vitro without serum and feeder cells.

Keywords: chicken gonadal primordial germ cells, pluripotent stem cells, growth factors, small molecules, transgenic birds

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Authors: Jeena Gupta

Abstract:

In the today’s world of pharmaceutical products, one should not forget the healing properties of inexpensive food materials especially spices. They are known to possess hidden pharmaceutical ingredients, imparting them the qualities of being anti-microbial, anti-oxidant, anti-inflammatory and anti-carcinogenic. Further aberrant epigenetic regulatory mechanisms like DNA methylation, histone modifications or altered microRNA expression patterns, which regulates gene expression without changing DNA sequence, contribute significantly in the development of various diseases. Changing lifestyles and diets exert their effect by influencing these epigenetic mechanisms which are thus the target of dietary phytochemicals. Bioactive components of plants have been in use since ages but their potential to reverse epigenetic alterations and prevention against diseases is yet to be explored. Spices being rich repositories of many bioactive constituents are responsible for providing them unique aroma and taste. Some spices like curcuma and garlic have been well evaluated for their epigenetic regulatory potential, but for others, it is largely unknown. We have evaluated the biological activity of phyto-active components of Fennel, Cardamom and Fenugreek by in silico molecular modeling, in vitro and in vivo studies. Ligand-based similarity studies were conducted to identify structurally similar compounds to understand their biological phenomenon. The database searching has been done by using Fenchone from fennel, Sabinene from cardamom and protodioscin from fenugreek as a query molecule in the different small molecule databases. Moreover, the results of the database searching exhibited that these compounds are having potential binding with the different targets found in the Protein Data Bank. Further in addition to being epigenetic modifiers, in vitro study had demonstrated the antimicrobial, antifungal, antioxidant and cytotoxicity protective effects of Fenchone, Sabinene and Protodioscin. To best of our knowledge, such type of studies facilitate the target fishing as well as making the roadmap in drug design and discovery process for identification of novel therapeutics.

Keywords: epigenetics, spices, phytochemicals, fenchone

Procedia PDF Downloads 130