Search results for: gene module

Authors: Yassir Adam Shuaib, Stefan Niemann, Eltahir Awad Khalil, Ulrich Schaible, Lothar Heinz Wieler, Mohammed Ahmed Bakhiet, Abbashar Osman Mohammed, Mohamed Abdelsalam Abdalla, Elvira Richter

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Tuberculosis (TB) is a chronic bacterial disease of humans and animals and it is characterized by the progressive development of specific granulomatous tubercle lesions in affected tissues. In a six-month study, from June to November 2014, a total of 2,304 carcasses of cattle, camel, sheep, and goats slaughtered at East and West Gaash slaughterhouses, Kassala, were investigated during postmortem, in parallel, 101 sputum samples from TB suspected patients at Kassala and El-Gadarif Teaching Hospitals were collected in order to investigate tuberculosis in animals and humans. Only 0.1% carcasses were found with suspected TB lesions in the liver and lung and peritoneal cavity of two sheep and no tuberculous lesions were found in the carcasses of cattle, goats or camels. All samples, tissue lesions and sputum, were decontaminated by the NALC-NaOH method and cultured for mycobacterial growth at the NRZ for Mycobacteria, Research Center Borstel, Germany. Genotyping and molecular characterization of the grown strains were done by line probe assay (GenoType CM and MTBC) and 16S rDNA, rpoB gene, and ITS sequencing, spoligotyping, MIRU-VNTR typing and next generation sequencing (NGS). Culture of the specimens revealed growth of organisms from 81.6% of all samples. Mycobacterium tuberculosis (76.2%), M. intracellulare (14.2%), mixed infection with M. tuberculosis and M. intracellulare (6.0%) and mixed infection with M. tuberculosis and M. fortuitum and with M. intracellulare and unknown species (1.2%) were detected in the sputum samples and unknown species (1.2%) were detected in the samples of one of the animals tissues. From the 69 M. tuberculosis strains, 25 (36.2%) were showing either mono-drug-resistant or multi-drug-resistant or poly-drug-resistant but none was extensively drug-resistant. In conclusion, the prevalence of TB in animals was very low while in humans M. tuberculosis-Delhi/CAS lineage was responsible for most cases and there was an evidence of MDR transmission and acquisition.

Keywords: animal, human, slaughterhouse, Sudan, tuberculosis

Procedia PDF Downloads 342

Authors: Suchithra V., Shreedhanya, Kavya Menon, Vidya Niranjan

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Skin metagenomics has a wide range of applications with direct relevance to the health of the organism. It gives us insight to the diverse community of microorganisms (the microbiome) harbored on the skin. In the recent years, it has become increasingly apparent that the interaction between skin microbiome and the human body plays a prominent role in immune system development, cancer development, disease pathology, and many other biological implications. Next Generation Sequencing has led to faster and better understanding of environmental organisms and their mutual interactions. This project is studying the human skin microbiome of different individuals having varied skin conditions. Bacterial 16S rRNA data of skin microbiome is downloaded from SRA toolkit provided by NCBI to perform metagenomics analysis. Twelve samples are selected with two controls, and 3 different categories, i.e., sex (male/female), skin type (moist/intermittently moist/sebaceous) and occlusion (occluded/intermittently occluded/exposed). Quality of the data is increased using Cutadapt, and its analysis is done using FastQC. USearch, a tool used to analyze an NGS data, provides a suitable platform to obtain taxonomy classification and abundance of bacteria from the metagenome data. The statistical tool used for analyzing the USearch result is METAGENassist. The results revealed that the top three abundant organisms found were: Prevotella, Corynebacterium, and Anaerococcus. Prevotella is known to be an infectious bacterium found on wound, tooth cavity, etc. Corynebacterium and Anaerococcus are opportunist bacteria responsible for skin odor. This result infers that Prevotella thrives easily in sebaceous skin conditions. Therefore it is better to undergo intermittently occluded treatment such as applying ointments, creams, etc. to treat wound for sebaceous skin type. Exposing the wound should be avoided as it leads to an increase in Prevotella abundance. Moist skin type individuals can opt for occluded or intermittently occluded treatment as they have shown to decrease the abundance of bacteria during treatment.

Keywords: bacterial 16S rRNA , next generation sequencing, skin metagenomics, skin microbiome, taxonomy

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Authors: Mukolwe D. Lubembe, Odongo O. David, Githaka Naftali, Kanduma Esther, Marinda Oosthuizen, Kgomotso P. Sibeko

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Theileriosis is a tick-borne disease of cattle caused by an apicomplexan protozoan parasite of the genus Theileria. In eastern and southern Africa, Theileria infections in cattle are caused by the species Theileria parva whose natural reservoir is the African buffalo (Syncerus caffer). Currently, East Coast Fever (ECF) caused by the cattle-derived Theileria parva is still a major problem in eastern Africa and some parts of southern Africa but not in South Africa following its eradication in the 1950s. However, Corridor disease (CD) caused by the buffalo-derived Theileria parva still remains a concern in South Africa. The diversity of Theileria parva in South Africa in comparison to other affected countries is poorly defined yet its known to be the survival strategy of this parasite. We assessed the antigenic diversity of Theileria parva isolates from Buffalo and cattle at the wildlife-livestock interface comparing samples from South Africa, Zimbabwe, Kenya, Tanzania, and Uganda. Antigenic epitopes of eight schizont antigen genes (Tp1, Tp3, Tp4, Tp5, Tp6, Tp7, Tp8 and Tp10) were amplified by PCR from genomic DNA extracted from blood samples collected from cattle and buffalo at the wildlife-livestock interface. Amplicons were purified and then sequenced on NGS platform. Full length open reading frames (ORFs) of two schizont antigen genes (Tp2 and Tp9) and one sporozoite antigen gene, p67 were also amplified from genomic DNA. Amplicons were then purified and cloned for sequencing. Analysis was based on sequence differences in the genes. Preliminary results show an extensively diverse population of Theileria parva circulating in buffalo and cattle populations at the wildlife-livestock interface. Diversity of the antigen genes contributes to the evasion of the immune system of the host by Theileria parva. This possess a concern in that, some of the Theileria parva populations may re-assort and become adapted to cattle to cause a form of theileriosis that is as fatal as ECF in areas where ECF was eradicated or is absent

Keywords: Theileria parva, east coast fever, corridor diseases, antigen genes, diversity

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Authors: África Vicario, Fernando J. Álvarez, Felipe Parralejo, Fernando Aranda

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The irrigation systems of traditional agriculture, such as gravity-fed irrigation, produce a great waste of water because, generally, there is no control over the amount of water supplied in relation to the water needed. The AIRON Project tries to solve this problem by implementing an IoT-based system to sensor the irrigation plots so that the state of the crops and the amount of water used for irrigation can be known remotely. The IoT system consists of a sensor network that measures the humidity of the soil, the weather conditions (temperature, relative humidity, wind and solar radiation) and the irrigation water flow. The communication between this network and a central gateway is conducted by means of long-range wireless communication that depends on the characteristics of the irrigation plot. The main objective of the AIRON project is to deploy an IoT sensor network in two different plots of the irrigation community of Aranjuez in the Spanish region of Madrid. The first plot is 2 km away from the central gateway, so LoRa has been used as the base communication technology. The problem with this plot is the absence of mains electric power, so devices with energy-saving modes have had to be used to maximize the external batteries' use time. An ESP32 SOC board with a LoRa module is employed in this case to gather data from the sensor network and send them to a gateway consisting of a Raspberry Pi with a LoRa hat. The second plot is located 18 km away from the gateway, a range that hampers the use of LoRa technology. In order to establish reliable communication in this case, the long-term evolution (LTE) standard is used, which makes it possible to reach much greater distances by using the cellular network. As mains electric power is available in this plot, a Raspberry Pi has been used instead of the ESP32 board to collect sensor data. All data received from the two plots are stored on a proprietary server located at the irrigation management company's headquarters. The analysis of these data by means of machine learning algorithms that are currently under development should allow a short-term prediction of the irrigation water demand that would significantly reduce the waste of this increasingly valuable natural resource. The major finding of this work is the real possibility of deploying a remote sensing system for irrigated plots by using Commercial-Off-The-Shelf (COTS) devices, easily scalable and adaptable to design requirements such as the distance to the control center or the availability of mains electrical power at the site.

Keywords: internet of things, irrigation water control, LoRa, LTE, smart farming

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Authors: Shih-Ping Liu, Cheng-Hsuan Chang, Yu-Chuen Huang, Shih-Yin Chen, Woei-Cherng Shyu

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Embryonic stem cells (ES) and induced pluripotnet stem cells (iPS) are both pluripotent stem cells. For mouse stem cells culture technology, leukemia inhibitory factor (LIF) was used to maintain the pluripotency of stem cells in vitro. However, LIF is an expensive reagent. The goal of this study was to find out a pure compound extracted from Chinese herbal medicine that could maintain stem cells pluripotency to replace LIF and improve the iPS generation efficiency. From 20 candidates traditional Chinese medicine we found that Cordycepsmilitaris triggered the up-regulation of stem cells activating genes (Oct4 and Sox2) expression levels in MEF cells. Cordycepin, a major active component of Cordycepsmilitaris, also could up-regulate Oct4 and Sox2 gene expression. Furthermore, we used ES and iPS cells and treated them with different concentrations of Cordycepin (replaced LIF in the culture medium) to test whether it was useful to maintain the pluripotency. The results showed higher expression levels of several stem cells markers in 10 μM Cordycepin-treated ES and iPS cells compared to controls that did not contain LIF, including alkaline phosphatase, SSEA1, and Nanog. Embryonic body formation and differentiation confirmed that 10 μM Cordycepin-containing medium was capable to maintain stem cells pluripotency after four times passages. For mechanism analysis, microarray analysis indicated extracellular matrix and Jak/Stat signaling pathway as the top two deregulated pathways. In ECM pathway, we determined that the integrin αVβ5 expression levels and phosphorylated Src levels increased after Cordycepin treatment. In addition, the phosphorylated Jak2 and phosphorylated Sat3 protein levels were increased after Cordycepin treatment and suppressed with the Jak2 inhibitor, AG490. The expression of cytokines associated with Jak2/Stat3 signaling pathway were also up-regulated by Q-PCR and ELISA assay. Lastly, we used Oct4-GFP MEF cells to test iPS generation efficiency following Cordycepin treatment. We observed that 10 Μm Cordycepin significantly increased the iPS generation efficiency in day 21. In conclusion, we demonstrated Cordycepin could maintain the pluripotency of stem cells through both of ECM and Jak2/Stat3 signaling pathway and improved iPS generation efficiency.

Keywords: cordycepin, iPS cells, Jak2/Stat3 signaling pathway, molecular biology

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Authors: Feixiong Chen, Naoufel Haddour, Marie Frenea-Robin, Yves MéRieux, Yann Chevolot, Virginie Monnier

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Neonatal thrombopenia occurs when the mother generates antibodies against her baby’s platelet antigens. It is particularly critical for newborns because it can cause coagulation troubles leading to intracranial hemorrhage. In this case, diagnosis must be done quickly to make platelets transfusion immediately after birth. Before transfusion, platelet antigens must be tested carefully to avoid rejection. The majority of thrombopenia (95 %) are caused by antibodies directed against Human Platelet Antigen 1a (HPA-1a) or 5b (HPA-5b). The common method for antigen platelets detection is polymerase chain reaction allowing for identification of gene sequence. However, it is expensive, time-consuming and requires significant blood volume which is not suitable for newborns. We propose to develop a point-of-care device based on double functionalized magnetic colloids with 1) antibodies specific to antigen platelets and 2) highly sensitive electroactive molecules in order to be detected by an electrochemical microsensor. These magnetic colloids will be used first to isolate platelets from other blood components, then to capture specifically platelets bearing HPA-1a and HPA-5b antigens and finally to attract them close to sensor working electrode for improved electrochemical signal. The expected advantages are an assay time lower than 20 min starting from blood volume smaller than 100 µL. Our functionalization procedure based on amine dendrimers and NHS-ester modification of initial carboxyl colloids will be presented. Functionalization efficiency was evaluated by colorimetric titration of surface chemical groups, zeta potential measurements, infrared spectroscopy, fluorescence scanning and cyclic voltammetry. Our results showed that electroactive molecules and antibodies can be immobilized successfully onto magnetic colloids. Application of a magnetic field onto working electrode increased the detected electrochemical signal. Magnetic colloids were able to capture specific purified antigens extracted from platelets.

Keywords: Magnetic Nanoparticles , Electroactive Molecules, Antibody, Platelet

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Authors: Esam Mohammed Al-shaebi

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One of the most crucial approaches for treating human diseases, particularly parasite infections, is nanomedicine. One of the most significant protozoan diseases that impact farm and domestic animals is coccidiosis. While, amprolium is one of the traditional anticoccidial medication, the advent of drug-resistant strains of Eimeria necessitates the development of novel treatments. The goal of the current investigation was to determine whether biosynthesized selenium nanoparticles (Bio-SeNPs) using Azadirachta indica leaves extract might treat mice with Eimeria papillata infection in the jejunal tissue. Five groups of seven mice each were used, as follows: Group 1: Non-infected-non-treated (negative control). Group 2: Non-infected treated group with Bio-SeNPs (0.5 mg/kg of body weight). Groups 3-5 were orally inoculated with 1×103 sporulated oocysts of E. papillata. Group 3: Infected-non-treated (positive control). Group 4: Infected and treated group with Bio-SeNPs (0.5 mg/kg). Group 5: Infected and treated group with the Amprolium. Groups 4 and 5 daily received oral administration (for 5 days) of Bio-SeNPs and anticoccidial medication, respectively, after infection. Bio-SeNPs caused a considerable reduction in oocyst output in mice feces (97.21%). This was also accompanied by a significant reduction in the number of developmental parasitic stages in the jejunal tissues. Glutathione reduced (GSH), glutathione peroxidase (GPx), and superoxide dismutase (SOD) levels were dramatically reduced by the Eimeria parasite, whereas, nitric oxide (NO) and malonaldehyde (MDA) levels were markedly elevated. The amount of goblet cells and MUC2 gene expression were used as apoptotic indicators, and both were considerably downregulated by infection. However, infection markedly increased the expression of inflammatory cytokines (IL-6 and TNF-α) and the apoptotic genes (Caspase-3 and BCL2). Bio-SeNPs were administrated to mice to drastically lower body weight, oxidative stress, and inflammatory and apoptotic indicators in the jejunal tissue. Our research thus showed the involvement of Bio-SeNPs in protecting mice with E. papillata infections against jejunal damage.

Keywords: coccidiosis, nanoparticles, azadirachta indica, oxidative stress

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Authors: Jerome Joshi

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The paper presents the modelling of financial markets using the Stochastic Pi Calculus model. The Stochastic Pi Calculus model is mainly used for biological applications; however, the feature of this model promotes its use in financial markets, more prominently in high frequency trading. The trading system can be broadly classified into exchange, market makers or intermediary traders and fundamental traders. The exchange is where the action of the trade is executed, and the two types of traders act as market participants in the exchange. High frequency trading, with its complex networks and numerous market participants (intermediary and fundamental traders) poses a difficulty while modelling. It involves the participants to seek the advantage of complex trading algorithms and high execution speeds to carry out large volumes of trades. To earn profits from each trade, the trader must be at the top of the order book quite frequently by executing or processing multiple trades simultaneously. This would require highly automated systems as well as the right sentiment to outperform other traders. However, always being at the top of the book is also not best for the trader, since it was the reason for the outbreak of the ‘Hot – Potato Effect,’ which in turn demands for a better and more efficient model. The characteristics of the model should be such that it should be flexible and have diverse applications. Therefore, a model which has its application in a similar field characterized by such difficulty should be chosen. It should also be flexible in its simulation so that it can be further extended and adapted for future research as well as be equipped with certain tools so that it can be perfectly used in the field of finance. In this case, the Stochastic Pi Calculus model seems to be an ideal fit for financial applications, owing to its expertise in the field of biology. It is an extension of the original Pi Calculus model and acts as a solution and an alternative to the previously flawed algorithm, provided the application of this model is further extended. This model would focus on solving the problem which led to the ‘Flash Crash’ which is the ‘Hot –Potato Effect.’ The model consists of small sub-systems, which can be integrated to form a large system. It is designed in way such that the behavior of ‘noise traders’ is considered as a random process or noise in the system. While modelling, to get a better understanding of the problem, a broader picture is taken into consideration with the trader, the system, and the market participants. The paper goes on to explain trading in exchanges, types of traders, high frequency trading, ‘Flash Crash,’ ‘Hot-Potato Effect,’ evaluation of orders and time delay in further detail. For the future, there is a need to focus on the calibration of the module so that they would interact perfectly with other modules. This model, with its application extended, would provide a basis for researchers for further research in the field of finance and computing.

Keywords: concurrent computing, high frequency trading, financial markets, stochastic pi calculus

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Authors: Anyimc, C. J. Aneke, J. O. Orji, O. Nworie, U. C. C. Egbule

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The microbial examination and antimicrobial susceptibility profile of microorganism isolated from the salt mining site in Ebonyi state were evaluated in the present study using a standard microbiological technique. A total of 300 samples were randomly collected in three sample groups (A, B, and C) of 100 each. Isolation, Identification and characterization of organization present on the soil samples were determined by culturing, gram-staining and biochemical technique. The result showed the following organisms were isolated with their frequency as follow: Bacillus species (37.3%) and Staphylococcus species(23.5%) had the highest frequency in the whole Sample group A and B while Klebsiella specie (15.7%), Pseudomonas species(13.7%), and Erwinia species (9.8%) had the least. Rhizopus species (42.0%) and Aspergillus species (26.0%) were the highest fungi isolated, followed by Penicillum species (20.0%) while Mucor species (4.0%), and Fusarium species (8.0%) recorded the least. Sample group C showed high microbial population of all the microbial isolates when compared to sample group A and B. Disc diffusion method was used to determine the susceptibility of isolated bacteria to various antibiotics (oxfloxacin, pefloxacin, ciprorex, augumentin, gentamycin, ciproflox, septrin, ampicillin), while agar well diffusion method was used to determine the susceptibility of isolated fungi to some antifungal drugs (metronidazole, ketoconazole, itraconazole fluconazole). The antibacterial activity of the antibiotics used showed that ciproflux has the best inhibitory effect on all the test bacteria. Ketoconazole showed the highest inhibitory effect on the fungal isolates, followed by itraconazole, while metronidazole and fluconazole showed the least inhibitory effect on the entire test fungal isolates. Hence, the multiple drug resistance of most isolates to appropriate drugs of choice are of great public health concern and cells for periodic monitoring of antibiograms to detect possible changing patterns. Microbes isolated in the salt mining site can also be used as a source of gene(s) that can increase salt tolerance in different crop species through genetic engineering.

Keywords: microorganisms, antibacterial, antifungal, resistance, salt mining site, Ebonyi State

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Authors: Patarasuda Chaisupa, R. Clay Wright

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The selection of appropriate biomolecular targets is a crucial aspect of biopharmaceutical development. The Auxin-Inducible Degron Degradation (AID) technology has demonstrated remarkable potential in efficiently and rapidly degrading target proteins, thereby enabling the identification and acquisition of drug targets. The AID system also offers a viable method to deplete specific proteins, particularly in cases where the degradation pathway has not been exploited or when the adaptation of proteins, including the cell environment, occurs to compensate for the mutation or gene knockout. In this study, we have engineered an improved AID system tailored to deplete proteins of interest. This AID construct combines the auxin-responsive E3 ubiquitin ligase binding domain, AFB2, and the substrate degron, IAA17, fused to the target genes. Essential genes of fungi with the lowest percent amino acid similarity to human and plant orthologs, according to the Basic Local Alignment Search Tool (BLAST), were cloned into the AID construct in S. cerevisiae (AID-tagged strains) using a modular yeast cloning toolkit for multipart assembly and direct genetic modification. Each E3 ubiquitin ligase and IAA17 degron was fused to a fluorescence protein, allowing for real-time monitoring of protein levels in response to different auxin doses via cytometry. Our AID system exhibited high sensitivity, with an EC50 value of 0.040 µM (SE = 0.016) for AFB2, enabling the specific promotion of IAA17::target protein degradation. Furthermore, we demonstrate how this improved AID system enhances quantitative functional studies of various proteins in fungi. The advancements made in auxin-inducible protein degradation in this study offer a powerful approach to investigating critical target protein viability in fungi, screening protein targets for drugs, and regulating intracellular protein abundance, thus revolutionizing the study of protein function underlying a diverse range of biological processes.

Keywords: synthetic biology, bioengineering, molecular biology, biotechnology

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Authors: Liaqat Ali, Hubert E. Blum, Türkân Sakinc

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Enterococcus faecium is an emerging multidrug-resistant nosocomial pathogen increased dramatically worldwide and causing bacteremia, endocarditis, urinary tract and surgical site infections in immunocomprised patients. The capsular polysaccharides that contribute to pathogenesis through evasion of the host innate immune system are also involved in hindering leukocyte killing of enterococci. The gene cluster (enterococcal polysaccharide antigen) of E. faecalis encoding homologues of many genes involved in polysaccharide biosynthesis. We identified two putative loci with 22 kb and 19 kb which contained 11 genes encoding for glycosyltransferases (GTFs); this was confirmed by using genome comparison of already sequenced strains that has no homology to known capsule genes and the epa-locus. The polysaccharide-conjugate vaccines have rapidly emerged as a suitable strategy to combat different pathogenic bacteria, therefore, we investigated a polysaccharide biosynthesis CapD protein in E. faecium contains 336 amino acids and had putative function for N-linked glycosylation. The deletion/knock-out capD mutant was constructed and complemented by homologues recombination method and confirmed by using PCR and sequencing. For further characterization and functional analysis, in-vitro cell culture and in-vivo a mouse infection models were used. Our ΔcapD mutant shows a strong hydrophobicity and all strains exhibited biofilm production. Subsequently, the opsonic activity was tested in an opsonophagocytic assay which shows increased in mutant compared complemented and wild type strains but more than two fold decreased in colonization and adherence was seen on surface of uroepithelial cells. However, a significant higher bacterial colonialization was observed in capD mutant during animal bacteremia infection. Unlike other polysaccharides biosynthesis proteins, CapD does not seems to be a major virulence factor in enterococci but further experiments and attention is needed to clarify its function, exact mechanism and involvement in pathogenesis of enteroccocal nosocomial infections eventually to develop a vaccine/ or targeted therapy.

Keywords: E. faecium, pathogenesis, polysaccharides, biofilm formation

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Authors: Maha Z. Rizk, Sami A. Fattah, Heba M. Darwish, Sanaa A. Ali, Mai O. Kadry

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Although it is known that nano-TiO2 and other nanoparticles can induce liver toxicity, the mechanisms and the molecular pathogenesis are still unclear. The present study investigated some biochemical indices of nano-sized Titanium dioxide (TiO2 NPS) toxicity in mice liver and the ameliorative efficacy of individual and combined doses of idebenone, carnosine and vitamin E. Nano-anatase TiO2 (21 nm) was administered as a total oral dose of 2.2 gm/Kg daily for 2 weeks followed by the afore-mentioned antioxidants daily either individually or in combination for 1month. TiO2-NPS induced a significant elevation in serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and hepatic oxidative stress biomarkers [lipid peroxides (LP), and nitric oxide levels (NOX), while it significantly reduced glutathione reductase (GR), reduced glutathione (GSH) and glutathione peroxidase(GPX) levels. Moreover the quantitative RT-PCR analysis showed that nano-anatase TiO2 can significantly alter the mRNA and protein expressions of the fibrotic factors TGF-B1, VEGFand Smad-2. Histopathological examination of hepatic tissue reinforced the previous biochemical results. Our results also implied that inflammatory responses and liver injury may be involved in nano-anatase TiO2-induced liver toxicity Tumor necrosis factor-α (TNF-α) and Interleukin -6 (IL-6) and increased the percent of DNA damage which was assessed by COMET assay in addition to the apoptotic marker Caspase-3. Moreover mRNA gene expression observed by RT-PCR showed a significant overexpression in nuclear factor relation -2 (Nrf2), nuclear factor kappa beta (NF-Kβ) and the apoptotic factor (bax), and a significant down regulation in the antiapoptotic factor (bcl2) level. In conclusion idebenone, carnosine and vitamin E ameliorated the deviated previously mentioned parameters with variable degrees with the most pronounced role in alleviating the hazardous effect of TiO2 NPS toxicity following the combination regimen.

Keywords: Nano-anatase TiO2, TGF-B1, SMAD-2

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Authors: Karol Rodriguez-Peña, Martha L. Macias-Rubalcava, Leticia Rocha-Zavaleta, Sergio Sanchez

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A bioassay-guided study for anti-cancer compounds from endophytes of the Mexican medicinal plant Amphipteryygium adstringens resulted in the isolation of a streptomycete capable of producing a group of compounds with high cytotoxic activity. Microorganisms from surface sterilized samples of various sections of the plant were isolated and all the actinomycetes found were evaluated for their potential to produce compounds with cytotoxic activity against cancer cell lines MCF7 (breast cancer) and HeLa (cervical cancer) as well as the non-tumoural cell line HaCaT (keratinocyte). The most active microorganism was picked for further evaluation. The identification of the microorganism was carried out by 16S rDNA gene sequencing, finding the closest proximity to Streptomyces scabrisporus, but with the additional characteristic that the strain isolated in this study was capable of producing colorful compounds never described for this species. Crude extracts of dichloromethane and ethyl acetate showed IC50 values of 0.29 and 0.96 μg/mL for MCF7, 0.51 and 1.98 μg/mL for HeLa and 0.96 and 2.7 μg/mL for HaCaT. Scaling the fermentation to 10 L in a bioreactor generated 1 g of total crude extract, which was fractionated by silica gel open column to yield 14 fractions. Nine of the fractions showed cytotoxic activity. Fraction 4 was chosen for subsequent purification because of its high activity against cancerous cell lines, lower activity against keratinocytes. HPLC-UV-MS/ESI was used for the evaluation of this fraction, finding at least 10 different compounds with high values of m/z (≈588). Purification of the compounds was carried out by preparative thin-layer chromatography. The prevalent compound was Steffimycin B, a molecule known for its antibiotic and cytotoxic activities and also for its low solubility in aqueous solutions. Along with steffimycin B, another five compounds belonging to the steffimycin family were isolated and at this moment their structures are being elucidated, some of which display better solubility in water: an attractive property for the pharmaceutical industry. As a conclusion to this study, the isolation of endophytes resulted in the discovery of a strain capable of producing compounds with high cytotoxic activity that need to be studied for their possible utilization.

Keywords: amphipterygium adstringens, cytotoxicity, streptomyces scabrisporus, steffimycin

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Authors: U. Pruthviraj, Praveen Kumar R. A. K. Athul, K. V. Gangadharan, S. Rao Shrikantha

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An open source based autonomous unmanned marine surface vehicle (UMSV) is developed for some of the marine applications such as pollution control, environmental monitoring and thermal imaging. A double rotomoulded hull boat is deployed which is rugged, tough, quick to deploy and moves faster. It is suitable for environmental monitoring, and it is designed for easy maintenance. A 2HP electric outboard marine motor is used which is powered by a lithium-ion battery and can also be charged from a solar charger. All connections are completely waterproof to IP67 ratings. In full throttle speed, the marine motor is capable of up to 7 kmph. The motor is integrated with an open source based controller using cortex M4F for adjusting the direction of the motor. This UMSV can be operated by three modes: semi-autonomous, manual and fully automated. One of the channels of a 2.4GHz radio link 8 channel transmitter is used for toggling between different modes of the USMV. In this electric outboard marine motor an on board GPS system has been fitted to find the range and GPS positioning. The entire system can be assembled in the field in less than 10 minutes. A Flir Lepton thermal camera core, is integrated with a 64-bit quad-core Linux based open source processor, facilitating real-time capturing of thermal images and the results are stored in a micro SD card which is a data storage device for the system. The thermal camera is interfaced to an open source processor through SPI protocol. These thermal images are used for finding oil spills and to look for people who are drowning at low visibility during the night time. A Real Time clock (RTC) module is attached with the battery to provide the date and time of thermal images captured. For the live video feed, a 900MHz long range video transmitter and receiver is setup by which from a higher power output a longer range of 40miles has been achieved. A Multi-parameter probe is used to measure the following parameters: conductivity, salinity, resistivity, density, dissolved oxygen content, ORP (Oxidation-Reduction Potential), pH level, temperature, water level and pressure (absolute).The maximum pressure it can withstand 160 psi, up to 100m. This work represents a field demonstration of an open source based autonomous navigation system for a marine surface vehicle.

Keywords: open source, autonomous navigation, environmental monitoring, UMSV, outboard motor, multi-parameter probe

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Authors: Vijaya Madhuri Devraj, Swarnalatha Guditi, Kiran Kumar Bokara, Gangadhar Taduri

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Cell-based strategies may open therapeutic approaches that promote tolerance through manipulation of macrophages to increase long-term transplant survival rates and minimize side effects of the current immune suppressive regimens. The aim of the present study was, therefore, to test and compare the therapeutic potential of MSC and Tregs on macrophage polarization to develop an alternate cell-based treatment option in kidney transplantation. In the current protocol, macrophages from kidney transplant recipients with graft dysfunction were co-cultured with MSCs and Treg cells with and without cell-cell contact on transwell plates, further to quantitatively assess macrophage polarization in response to MSC and Treg treatment over time, M1 and M2 cell surface markers were used. Additionally, multiple soluble analytes were analyzed in cell supernatant by using bead-based immunoassays. Furthermore, to confirm our findings, gene expression analysis was done. MSCs induced the formation of M2 macrophages more than Tregs when macrophages M0 were cultured in transwell without cell contact. From this, we deduced the mechanism that soluble factors present in the MSCs condition media are involved in skewing of macrophages towards type 2 macrophages; similarly, in co-culture with cell-cell contact, MSCs resulted in more M2 type macrophages than Tregs. And an important finding of this study is the combination of both MSC-Treg showed significantly effective and consistent results in both with and without cell contact setups. Hence, it is suggestive to prefer MSCs over Tregs for secretome-based therapy and a combination of both for either therapy for effective transplantation outcomes. Our findings underline a key role of Tregs and MSCs in promoting macrophage polarization towards anti-inflammatory type. The study has great importance in prolongation of allograft and patient survival without any rejection by cell-based therapy, which induce self-tolerance and controlling infection.

Keywords: graft rejection, graft tolerance, macrophage polarization, mesenchymal stem cells, regulatory T cells, transplant immunology

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Authors: Ali Bahkali, Rayan Yousef Booq, Mohammad Khiyami

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Soil samples were collected from five different regions in the Kingdom of Saudi Arabia. Microbiological methods included dilution methods and pour plates to isolate and purify bacteria soil. The ability of isolates to develop biopolymer was investigated on petri dishes containing elements and substance concentrations stimulating developing biopolymer. Fluorescent stains, Nile red and Nile blue were used to stain the bacterial cells developing biopolymers. In addition, Sudan black was used to detect biopolymers in bacterial cells. The isolates which developed biopolymers were identified based on their gene sequence of 1 6sRNA and their ability to grow and synthesize PHAs on mineral medium supplemented with 1% dates molasses as the only carbon source under nitrogen limitation. During the study 293 bacterial isolates were isolated and detected. Through the initial survey on the petri dishes, 84 isolates showed the ability to develop biopolymers. These bacterial colonies developed a pink color due to accumulation of the biopolymers in the cells. Twenty-three isolates were able to grow on dates molasses, three strains of which showed the ability to accumulate biopolymers. These strains included Bacillus sp., Ralstonia sp. and Microbacterium sp. They were detected by Nile blue A stain with fluorescence microscopy (OLYMPUS IX 51). Among the isolated strains Ralstonia sp. was selected after its ability to grow on molasses dates in the presence of a limited nitrogen source was detected. The optimum conditions for formation of biopolymers by isolated strains were investigated. Conditions studied included, best incubation duration (2 days), temperature (30°C) and pH (7-8). The maximum PHB production was raised by 1% (v1v) when using concentrations of dates molasses 1, 2, 3, 4 and 5% in MSM. The best inoculated with 1% old inoculum (1= OD). The ideal extraction method of PHA and PHB proved to be 0.4% sodium hypochlorite solution, producing a quantity of polymer 98.79% of the cell's dry weight. The maximum PHB production was 1.79 g/L recorded by Ralstonia sp. after 48 h, while it was 1.40 g/L produced by R.eutropha ATCC 17697 after 48 h.

Keywords: bacteria forming polyhydroxyalkanoate, detection, molecular, Saudi Arabia

Procedia PDF Downloads 323

Authors: Mir Mohammad Badrul Hasan, Anwar Abdkader, Chokri Cherif

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With the increasing demand and use of carbon fibre reinforced composites (CFRC), disposal of the carbon fibres (CF) and end of life composite parts is gaining tremendous importance on the issue especially of sustainability. Furthermore, a number of processes (e. g. pyrolysis, solvolysis, etc.) are available currently to obtain recycled CF (rCF) from end-of-life CFRC. Since the CF waste or rCF are neither allowed to be thermally degraded nor landfilled (EU Directive 1999/31/EC), profitable recycling and re-use concepts are urgently necessary. Currently, the market for materials based on rCF mainly consists of random mats (nonwoven) made from short fibres. The strengths of composites that can be achieved from injection-molded components and from nonwovens are between 200-404 MPa and are characterized by low performance and suitable for non-structural applications such as in aircraft and vehicle interiors. On the contrary, spinning rCF to yarn constructions offers good potential for higher CFRC material properties due to high fibre orientation and compaction of rCF. However, no investigation is reported till yet on the direct comparison of the mechanical properties of thermoplastic CFRC manufactured from virgin CF filament yarn and spun yarns from staple rCF. There is a lack of understanding on the level of performance of the composites that can be achieved from hybrid yarns consisting of rCF and PA6 fibres. In this drop back, extensive research works are being carried out at the Textile Machinery and High-Performance Material Technology (ITM) on the development of new thermoplastic CFRC from hybrid yarns consisting of rCF. For this purpose, a process chain is developed at the ITM starting from fibre preparation to hybrid yarns manufacturing consisting of staple rCF by mixing with thermoplastic fibres. The objective is to apply such hybrid yarns for the manufacturing of load bearing textile reinforced thermoplastic CFRCs. In this paper, the development of innovative multi-component core-sheath hybrid yarn structures consisting of staple rCF and polyamide 6 (PA 6) on a DREF-3000 friction spinning machine is reported. Furthermore, Unidirectional (UD) CFRCs are manufactured from the developed hybrid yarns, and the mechanical properties of the composites such as tensile and flexural properties are analyzed. The results show that the UD composite manufactured from the developed hybrid yarns consisting of staple rCF possesses approximately 80% of the tensile strength and E-module to those produced from virgin CF filament yarn. The results show a huge potential of the DREF-3000 friction spinning process to develop composites from rCF for high-performance applications.

Keywords: recycled carbon fibres, hybrid yarn, friction spinning, thermoplastic composite

Procedia PDF Downloads 230

Authors: Willem J. A. Louw

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The wave of new advanced computing technology that has been developed during the recent past has significantly changed the way we communicate, collaborate and collect information. It has created a new technology environment and paradigm in which our children and students grow-up and this impacts on their learning. Research confirmed that Generation Y students have a preference for learning in the new technology environment. The challenge or question is: How do we adjust our teaching and learning to make the most of these changes. The complexity of effective and efficient teaching and learning must not be underestimated and changes must be preceded by proper objective research to prevent any haphazard developments that could do more harm than benefit. A blended learning approach has been used in the Forestry department for a few numbers of years including the use of electronic-peer assisted learning (e-pal) in a fixed-computer set-up within a learning management system environment. It was decided to extend the investigation and do some exploratory research by using a range of different Tablet devices. For this purpose, learning activities or assignments were designed to cover aspects of communication, collaboration and collection of information. The Moodle learning management system was used to present normal module information, to communicate with students and for feedback and data collection. Student feedback was collected by using an online questionnaire and informal discussions. The research project was implemented in 2013, 2014 and 2015 amongst first and third-year students doing a forestry three-year technical tertiary qualification in commercial plantation management. In general, more than 80% of the students alluded to that the device was very useful in their learning environment while the rest indicated that the devices were not very useful. More than ninety percent of the students acknowledged that they would like to continue using the devices for all of their modules whilst the rest alluded to functioning efficiently without the devices. Results indicated that information collection (access to resources) was rated the highest advantageous factor followed by communication and collaboration. The main general advantages of using Tablets were listed by the students as being mobility (portability), 24/7 access to learning material and information of any kind on a user friendly device in a Wi-Fi environment, fast computing process speeds, saving time, effort and airtime through skyping and e-mail, and use of various applications. Ownership of the device is a critical factor while the risk was identified as a major potential constraint. Significant differences were reported between the different types and quality of Tablets. The preferred types are those with a bigger screen and the ones with overall better functionality and quality features. Tablets significantly increase the collaboration, communication and information collection needs of the students. It does, however, not replace the need of a computer/laptop because of limited storage and computation capacity, small screen size and inefficient typing.

Keywords: tablets, teaching, blended learning, tablet quality

Procedia PDF Downloads 228

Authors: Michael P. Mannino, Gerald W. Hart

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The majority of glucose metabolism proceeds through glycolytic pathways such as glycolysis or pentose phosphate pathway, however, about 5% is shunted through the hexosamine biosynthetic pathway, producing uridine diphosphate N-acetyl glucosamine (UDP-GlcNAc). This precursor can then be incorporated into complex oligosaccharides decorating the cell surface or remain as an intracellular post-translational-modification (PTM) of serine/threonine residues (O-GlcNAcylation, OGN), which has been identified on over 4,000 cytosolic or nuclear proteins. Intracellular OGN has major implications on cellularprocesses, typically by modulating protein localization, protein-protein interactions, protein degradation, and gene expression. Additionally, OGN is known to have an extensive cross-talk with phosphorylation, be in a competitive or cooperative manner. Unlike other PTMs there are only two cycling enzymes that are capable of adding or removing the GlcNAc moiety, O-linked N-aceytl glucosamine Transferase (OGT) and O-linked N-acetyl glucoamidase (OGA), respectively. The activity of OGT has been shown to be sensitive to cellular UDP-GlcNAc levels, even changing substrate affinity. Owing to this and that the concentration of UDP-GlcNAc is related to the metabolisms of glucose, amino acid, fatty acid, and nucleotides, O-GlcNAc is often referred to as a nutrient sensing rheostat. Indeed OGN is known to regulate several signaling pathways as a result of nutrient levels, such as insulin signaling. Dysregulation of OGN is associated with several disease states such as cancer, diabetes, and neurodegeneration. Improvements in glycomics over the past 10-15 years has significantly increased the OGT substrate pool, suggesting O-GlcNAc’s involvement in a wide variety of signaling pathways. However, O-GlcNAc’s role at the receptor level has only been identified in a case-by-case basis of known pathways. Examining the OGN of the plasma membrane (PM) may better focus our understanding of O-GlcNAc-effected signaling pathways. In this current study, PM fractions were isolated from several cell types via ultracentrifugation, followed by purification and MS/MS analysis in several cell lines. This process was repeated with or without OGT/OGA inhibitors or with increased/decreased glucose levels in media to ascertain the importance of OGN. Various pathways are followed up on in more detailed studies employing methods to localize OGN at the PM specifically.

Keywords: GlcNAc, nutrient sensitive, post-translational-modification, receptor

Procedia PDF Downloads 81

Authors: Holm Zerbe, Laura Macias, Hans-Joachim Schuberth, Wolfram Petzl

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Mastitis is among the most important production diseases in cows. It accounts for large parts of antimicrobial drug use in the dairy industry worldwide. Due to the imminent normative to reduce the use of antimicrobial drugs in livestock, new ways for therapy and prophylaxis of mastitis are needed. Recently epigenetic regulation of inflammation by chromatin modifications has increasingly drawn attention. Currently, some epigenetic modifiers have already been approved for the use in humans, however little is known about their actions in the bovine system. The aim of our study was to investigate whether three selected epigenetic modifiers (Vitamin D3, SAHA and S2101) influence the initial immune response towards mastitis pathogens in bovine udder tissue in vitro. Tissue explants of the teat cistern and udder parenchyma were collected from 21 cows and were incubated for 36 hours in the absence and presence of epigenetic modifiers. Additionally, the tissue was stimulated with heat-inactivated particles of Escherichia coli and Staphylococcus aureus, which are regarded as two of the most important mastitis pathogens. After incubation, the explants were tested by RT-qPCR for transcript abundances of immune-related candidate genes. Gene expression was validated in culture supernatants by an AlphaLISA assay. Furthermore, the culture supernatants were analyzed for their chemotactic capacity through a chemotaxis assay. Statistical analysis of data was performed with the program ‘R’ version 3.2.3. Vitamin D3 had no effect on the immune response of udder tissue in vitro after stimulation with mastitis pathogens. The epigenetic modifiers SAHA and S2101 however significantly blocked the pathogen-induced upregulation of CXCL8, TNFα, S100A9 and LAP (P < 0.05). The regulation of IL10 was not affected by treatment with SAHA and S2101. Transcript abundances for CXCL8 were reflected by IL8 contents and chemotactic activity in culture supernatants. In conclusion, these data show the potential of epigenetic modifiers (SAHA and S2101) to block overshooting inflammation in the udder. Thus epigenetic modifiers may serve in future as immune modulators for the treatment and/or prophylaxis of clinical mastitis. (Funded by Deutsche Forschungsgemeinschaft PE 1495/2-1).

Keywords: mastitis, cattle, epigenetics, immunomodulation

Procedia PDF Downloads 204

Authors: Guanyu Jiang, Donghai Xu, Huanteng Liu

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After the Fukushima-Daiichi accident, the development of accident tolerant fuel cladding materials to improve reactor safety has become a hot topic in the field of nuclear industry. ZrO₂ has a satisfactory neutron economy and can guarantee the fission chain reaction process, which enables it to be a promising coating for zirconium alloy cladding. Maintaining a good corrosion resistance in primary coolant loop during normal operations of Pressurized Water Reactors is a prerequisite for ZrO₂ as a protective coating on zirconium alloy cladding. Research on the corrosion performance of ZrO₂ coating in nuclear water chemistry is relatively scarce, and existing reports failed to provide an in-depth explanation for the failure causes of ZrO₂ coating. Herein, a detailed corrosion process of ZrO₂ coating in lithiated water at 360 °C and 18.5 MPa was proposed based on experimental research and molecular dynamics simulation. Lithiated water with different LiOH solutions in the present work was deaerated and had a dissolved oxygen concentration of < 10 ppb. The concentration of Li (as LiOH) was determined to be 2.3 ppm, 70 ppm, and 500 ppm, respectively. Corrosion tests were conducted in a static autoclave. Modeling and corresponding calculations were operated on Materials Studio software. The calculation of adsorption energy and dynamics parameters were undertaken by the Energy task and Dynamics task of the Forcite module, respectively. The protective effect and failure mechanism of ZrO₂ coating on Zry-4 under varied LiOH concentrations was further revealed by comparison with the coating corrosion performance in pure water (namely 0 ppm Li). ZrO₂ coating provided a favorable corrosion protection with the occurrence of localized corrosion at low LiOH concentrations. Factors influencing corrosion resistance mainly include pitting corrosion extension, enhanced Li+ permeation, short-circuit diffusion of O²⁻ and ZrO₂ phase transformation. In highly-concentrated LiOH solutions, intergranular corrosion, internal oxidation, and perforation resulted in coating failure. Zr ions were released to coating surface to form flocculent ZrO₂ and ZrO₂ clusters due to the strong diffusion and dissolution tendency of α-Zr in the Zry-4 substrate. Considering that primary water of Pressurized Water Reactors usually includes 2.3 ppm Li, the stability of ZrO₂ make itself a candidate fuel cladding coating material. Under unfavorable conditions with high Li concentrations, more boric acid should be added to alleviate caustic corrosion of ZrO₂ coating once it is used. This work can provide some references to understand the service behavior of nuclear coatings under variable water chemistry conditions and promote the in-pile application of ZrO₂ coating.

Keywords: ZrO₂ coating, Zry-4, corrosion behavior, failure mechanism, LiOH concentration

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Authors: Lorena Blasco-Arcas, Francesca Pucciarelli

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Business contexts have become globalised and digitalised, which requires that managers develop a strong sense of cross-cultural intelligence while working in geographically distant teams by means of digital technologies. How to better equip future managers on these kinds of skills has been put forward as a critical issue in Business Schools. In pursuing these goals, higher education is shifting from a passive lecture approach, to more active and experiential learning approaches that are more suitable to learn skills. For example, through the use of case studies, proposing plausible business problem to be solved by students (or teams of students), these institutions have focused for long in fostering learning by doing. Though, case studies are no longer enough as a tool to promote active teamwork and experiential learning. Moreover, digital advancements applied to educational settings have enabled augmented classrooms, expanding the learning experience beyond the class, which increase students’ engagement and experiential learning. Different authors have highlighted the benefits of digital engagement in order to achieve a deeper and longer-lasting learning and comprehension of core marketing concepts. Clickers, computer-based simulations and business games have become fairly popular between instructors, but still are limited by the fact that are fictional experiences. Further exploration of real digital platforms to implement real, live projects in the classroom seem relevant for marketing and business education. Building on this, this paper describes the development of an experiential learning activity in class, in which students developed a communication campaign in teams using the BuzzFeed platform, and subsequently implementing the campaign by using other social media platforms (e.g. Facebook, Instagram, Twitter…). The article details the procedure of using the project for a marketing module in a Bachelor program with students located in France, Italy and Spain campuses working on multi-campus groups. Further, this paper describes the project outcomes in terms of students’ engagement and analytics (i.e. visits achieved). the project included a survey in order to analyze and identify main aspects related to how the learning experience is influenced by the cultural competence developed through working in geographically distant and culturally diverse teamwork. Finally, some recommendations to use project-based social media tools while working with virtual teamwork in the classroom are provided.

Keywords: cultural competences, experiential learning, social media, teamwork, virtual group work

Procedia PDF Downloads 156

Authors: Junichi Hosoi

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Epidermal Langerhans cells reside in upper layer of epidermis and play a role in immune surveillance. The finding of the close association of nerve endings to Langerhans cells triggered the research on systemic regulation of Langerhans cells. They disappear from epidermis after exposure to environmental and internal stimuli and reappear about a week later. Myeloid progenitor cells are assumed to be one of the sources of Langerhans cells. We examined the effects of cortisol on the reappearance of Langerhans cells in vitro. Cord-blood derived CD34-positive cells were cultured in the medium supplemented with stem cell factor/Flt3 ligand/granulocyte macrophage-colony stimulating factor/tumor necrosis factor alpha/bone morphologic protein 7/transforming growth factor beta in the presence or absence of cortisol. Cells were analyzed by flow cytometry for CD1a (cluster differentiation 1a), a marker of Langerhans cells and dermal dendritic cells, and CD39 (cluster differentiation factor 39), extracellular adenosine triphosphatase. Both CD1a-positive cells and CD39-positive cells were decreased by treatment with cortisol (suppression by 35% and 22% compared to no stress hormone, respectively). Differentiated Langerhans cells are attracted to epidermis by chemokines that are secreted from keratinocytes. Epidermal keratinocytes were cultured in the presence or absence of cortisol and analyzed for the expression of CCL2 (C-C motif chemokine ligand 2) and CCL20 (C-C motif chemokine ligand 20), which are typical attractants of Langerhans cells, by quantitative reverse transcriptase polymerase chain reaction. The expression of both chemokines, CCL2 and CCL20, were suppressed by treatment with cortisol (suppression by 38% and 48% compared to no stress hormone, respectively). We examined the possible regulation of the suppression by cortisol with plant extracts. The extracts of Ganoderma lucidum and Iris protected the suppression of the differentiation to CD39-positive cells and also the suppression of the gene expression of LC-chemoattractants. These results suggest that cortisol, which is either systemic or locally produced, blocks the supply of epidermal Langerhans cells at 2 steps, differentiation from the precursor and attraction to epidermis. The suppression is possibly blocked by some plant extracts.

Keywords: Langerhans cell, stress, CD39, chemokine

Procedia PDF Downloads 159

Authors: Pedro G. Morouço

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One utmost challenge in Tissue Engineering is the production of 3D constructs capable of mimicking the functional hierarchy of native tissues. This is well stated for osteochondral tissue due to the complex mechanical functional unit based on the junction of articular cartilage and bone. Thus, the aim of the present study was to develop a new additive manufacturing system coupling micro-extrusion with hydrogels printing. An integrated system was developed with 2 main features: (i) the printing of up to three distinct hydrogels; (ii) in coordination with the printing of a thermoplastic structural support. The hydrogel printing module was projected with a ‘revolver-like’ system, where the hydrogel selection was made by a rotating mechanism. The hydrogel deposition was then controlled by pressured air input. The use of specific components approved for medical use was incorporated in the material dispensing system (Nordson EDF Optimum® fluid dispensing system). The thermoplastic extrusion modulus enabled the control of required extrusion temperature through electric resistances in the polymer reservoir and the extrusion system. After testing and upgrades, a hydrogel modulus with 3 syringes (3cm3 capacity each), with a pressure range of 0-2.5bar, a rotational speed of 0-5rpm, and working with needles from 200-800µm was obtained. This modulus was successfully coupled to the extrusion system that presented a temperature up to 300˚C, a pressure range of 0-12bar, and working with nozzles from 200-500µm. The applied motor could provide a velocity range 0-2000mm/min. Although, there are distinct printing requirements for hydrogels and polymers, the novel system could develop hybrid scaffolds, combining the 2 moduli. The morphological analysis showed high reliability (n=5) between the theoretical and obtained filament and pore size (350µm and 300µm vs. 342±4µm and 302±3µm, p>0.05, respectively) of the polymer; and multi-material 3D constructs were successfully obtained. Human tissues present very distinct and complex structures regarding their mechanical properties, organization, composition and dimensions. For osteochondral regenerative medicine, a multiphasic scaffold is required as subchondral bone and overlying cartilage must regenerate at the same time. Thus, a scaffold with 3 layers (bone, intermediate and cartilage parts) can be a promising approach. The developed system may give a suitable solution to construct those hybrid scaffolds with enhanced properties. The present novel system is a step-forward regarding osteochondral tissue engineering due to its ability to generate layered mechanically stable implants through the double-printing of hydrogels with thermoplastics.

Keywords: 3D bioprinting, bone regeneration, cartilage regeneration, regenerative medicine, tissue engineering

Procedia PDF Downloads 137

Authors: Miceska Simona, Škof Erik, Frković Grazio Snježana, Jeričević Anja, Smrkolj Špela, Cvjetićanin Branko, Novaković Srdjan, Grčar Kuzmanov Biljana, Kloboves-Prevodnik Veronika

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Objectives: High-grade serous ovarian cancer (HGSOC) is characterized by the dissemination of the tumor cells (TC) in the peritoneal cavity forming malignant ascites at the time of diagnosis or recurrence. Still, cytology itself has been underutilized as a modality for the diagnosis of HGSOC from ascites, and histological examination from the tumor tissue is yet the only validated method used. The objective of this study was to evaluate the reliability of cytology in the diagnosis of HGSOC in relation to the histopathological examination. Methods: The study included 42 patients with histologically confirmed HGSOC, accompanied by malignant ascites. To confirm the malignancy of the TC in the ascites and to define their immunophenotype, immunohistochemical reaction (IHC) of the following antigens: Calretinin, MOC, WT1, PAX8, p53, p16 & Ki-67 was evaluated on ascites cytospins and tissue blocks. For complete cytological determination of HGSOC, BRCA 1/2 gene mutation was determined from ascites, tissue block, and blood. BRCA1/2 mutation from blood was performed to define the type of mutation, somatic vs germline. Results: Among 42 patients, the immunophenotype of HGSOC from ascites was confirmed in 36 cases (86%). For more profound analysis, the patients were divided in 3 groups regarding the number of TC present in the ascites: patients with less than 10% TC, 10% TC, and more than 10% TC. From all included patients, in the group with less than 10% TC, there were 10 cases, and only 5 of them(50%) showed HGSOC phenotype; 12 cases had equally 10% of TC, and 11 cases (92%) showed HGSOC phenotype; 20 cases had more than 10% TC and all of them (100%) confirmed the HGSOC immunophenotype from ascites. Only 33 patients were eligible for further BRCA1/2 analysis. Eleven BRCA1/2 mutations were detected from thetissue block: 6 germline and 5 somatic. In 2 cases with less than 10% TC, BRCA1/2 mutation was not detected; 4 cases had 10% TC, and 2 of them (50%) confirmed the mutation; 4 cases had more than 10% TC, and all showed 100% reliability with the tumor tissue. Conclusions: Cytology is a highly reliable method for determining the immunophenotype of HGSOC and BRCA1/2 mutation if more than 10% of tumor cells are present in the ascites. This may present an additional non-invasive clinical approach for fast and effective diagnose in the future, especially in inoperable conditions or relapses.

Keywords: cytology, ascites, high-grade serous ovarian cancer, immunophenotype, BRCA1/2

Procedia PDF Downloads 167

Authors: G. Krishnamoorthy, S. Anandhakumar

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The effect of D-Lysine (D-Lys) on collagen with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide(EDC)/N-hydroxysuccinimide(NHS) initiated cross linking using experimental and modelling tools are evaluated. The results of the Coll-D-Lys-EDC/NHS scaffold also indicate an increase in the tensile strength (TS), percentage of elongation (% E), denaturation temperature (Td), and decrease the decomposition rate compared to L-Lys-EDC/NHS. Scanning electron microscopic (SEM) and atomic force microscopic (AFM) analyses revealed a well ordered with properly oriented and well-aligned structure of scaffold. The D-Lys stabilizes the scaffold against degradation by collagenase than L-Lys. The cell assay showed more than 98% fibroblast viability (NIH3T3) and improved cell adhesions, protein adsorption after 72h of culture when compared with native scaffold. Cell attachment after 74h was robust, with cytoskeletal analysis showing that the attached cells were aligned along the fibers assuming a spindle-shape appearance, despite, gene expression analyses revealed no apparent alterations in mRNA levels, although cell proliferation was not adversely affected. D-Lysine (D-Lys) plays a pivotal role in the self-assembly and conformation of collagen fibrils. The D-Lys assisted EDC/NHS initiated cross-linking induces the formation of an carboxamide by the activation of the side chain -COOH group, followed by aminolysis of the O-iso acylurea intermediates by the -NH2 groups are directly joined via an isopeptides bond. This leads to the formation of intra- and inter-helical cross links. Modeling studies indicated that D-Lys bind with collagen-like peptide (CLP) through multiple H-bonding and hydrophobic interactions. Orientational changes in collagenase on CLP-D-Lys are observed which may decrease its accessibility to degradation and stabilize CLP against the action of the former. D-Lys has lowest binding energy and improved fibrillar-assembly and staggered alignment without the undesired structural stiffness and aggregations. The proteolytic machinery is not well equipped to deal with Coll-D-Lys than Coll-L-Lys scaffold. The information derived from the present study could help in designing collagenolytically stable heterochiral collagen based scaffold for biomedical applications.

Keywords: collagen, collagenase, collagen like peptide, D-lysine, heterochiral collagen scaffold

Procedia PDF Downloads 369

Authors: Thor Alexis Sazon, Alexander Guzman-Urbina, Yasuhiro Fukushima

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This study outlines the method of how to develop a surrogate life cycle model based on fuzzy logic using three fuzzy inference methods: (1) the conventional Fuzzy Inference System (FIS), (2) the hybrid system of Data Analytics and Fuzzy Inference (DAFIS), which uses data clustering for defining the membership functions, and (3) the Adaptive-Neuro Fuzzy Inference System (ANFIS), a combination of fuzzy inference and artificial neural network. These methods were demonstrated with a case study where the Global Warming Potential (GWP) and the Levelized Cost of Energy (LCOE) of solar photovoltaic (PV) were estimated using Solar Irradiation, Module Efficiency, and Performance Ratio as inputs. The effects of using different fuzzy inference types, either Sugeno- or Mamdani-type, and of changing the number of input membership functions to the error between the calibration data and the model-generated outputs were also illustrated. The solution spaces of the three methods were consequently examined with a sensitivity analysis. ANFIS exhibited the lowest error while DAFIS gave slightly lower errors compared to FIS. Increasing the number of input membership functions helped with error reduction in some cases but, at times, resulted in the opposite. Sugeno-type models gave errors that are slightly lower than those of the Mamdani-type. While ANFIS is superior in terms of error minimization, it could generate solutions that are questionable, i.e. the negative GWP values of the Solar PV system when the inputs were all at the upper end of their range. This shows that the applicability of the ANFIS models highly depends on the range of cases at which it was calibrated. FIS and DAFIS generated more intuitive trends in the sensitivity runs. DAFIS demonstrated an optimal design point wherein increasing the input values does not improve the GWP and LCOE anymore. In the absence of data that could be used for calibration, conventional FIS presents a knowledge-based model that could be used for prediction. In the PV case study, conventional FIS generated errors that are just slightly higher than those of DAFIS. The inherent complexity of a Life Cycle study often hinders its widespread use in the industry and policy-making sectors. While the methodology does not guarantee a more accurate result compared to those generated by the Life Cycle Methodology, it does provide a relatively simpler way of generating knowledge- and data-based estimates that could be used during the initial design of a system.

Keywords: solar photovoltaic, fuzzy logic, inference system, artificial neural networks

Procedia PDF Downloads 131

Authors: Eryati Darwin, Eka Fithra Elfi, Indria Hafizah

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Background: Physical exercise induces a pattern of hormonal and immunological responses that prevent endothelial dysfunction by maintaining the availability of nitric oxide (NO). Regular and moderate exercise stimulates NO release, that can be considered as protective factor of cardiovascular diseases, while strenuous exercise induces increased levels in a number of pro-inflammatory and anti-inflammatory cytokines. Pro-inflammatory cytokines tumor necrosis factor-α (TNF-α) triggers endothelial activation which results in an increased vascular permeability. Nuclear gene factor kappa B (NF-κB) activates biological effect of TNF-α. Aim of Study: To determine the effect of physical exercise on the endothelial and skeletal muscle, we measured the level of NF-κB on rats’ serum, macrophages, and myocytes after strenuous physical exercise. Methods: 30 male Rattus norvegicus in the age of eight weeks were randomly divided into five groups (each containing six), and there were treated groups (T) and control group (C). The treated groups obtain strenuous physical exercise by ran on treadmill at 32 m/minutes for 1 hour or until exhaustion. Blood samples, myocytes of gastrocnemius muscle, and intraperitoneal macrophages were collected sequentially. There were investigated immediately, 2 hours, 6 hours, and 24 hours (T1, T2, T3, and T4) after sacrifice. The levels of NF-κB were measured by ELISA methods. Results: From our study, we found that the levels of NF-κB on myocytes in treated group from which its specimen was taken immediately (T1), 2 hours after treadmill (T2), and 6 hours after treadmill (T3) were significantly higher than control group (p<0.05), while the group from which its specimen was taken 24 hours after treadmill, was no significantly different (p>0.05). Also on macrophages, NF-κB in treated groups T1, T2, and T3 was significantly higher than control group (p<0.05), but there was no difference between T4 and control group (p>0.05). The level of serum NF-κB was not significantly different between treatment group as well as compared to control group (p>0.05). Serum NF-κB was significantly higher than the level on macrophages and myocytes (p<0.05). Conclusion: This study demonstrated that strenuous physical exercise stimulates the activation of NF-κB that plays a role in vascular inflammation and muscular damage, and may be recovered after resting period.

Keywords: endothelial function, inflammation, NFkB, physical exercise

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Authors: Mohammed Abdullah Alshahrani

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The research focuses on investigating the use of partial least squares (PLS) methodology for addressing challenges associated with high-dimensional correlated data. Recent technological advancements have led to experiments producing data characterized by a large number of variables compared to observations, with substantial inter-variable correlations. Such data patterns are common in chemometrics, where near-infrared (NIR) spectrometer calibrations record chemical absorbance levels across hundreds of wavelengths, and in genomics, where thousands of genomic regions' copy number alterations (CNA) are recorded from cancer patients. PLS serves as a widely used method for analyzing high-dimensional data, functioning as a regression tool in chemometrics and a classification method in genomics. It handles data complexity by creating latent variables (components) from original variables. However, applying PLS can present challenges. The study investigates key areas to address these challenges, including unifying interpretations across three main PLS algorithms and exploring unusual negative shrinkage factors encountered during model fitting. The research presents an alternative approach to addressing the interpretation challenge of predictor weights associated with PLS. Sparse estimation of predictor weights is employed using a penalty function combining a lasso penalty for sparsity and a Cauchy distribution-based penalty to account for variable dependencies. The results demonstrate sparse and grouped weight estimates, aiding interpretation and prediction tasks in genomic data analysis. High-dimensional data scenarios, where predictors outnumber observations, are common in regression analysis applications. Ordinary least squares regression (OLS), the standard method, performs inadequately with high-dimensional and highly correlated data. Copy number alterations (CNA) in key genes have been linked to disease phenotypes, highlighting the importance of accurate classification of gene expression data in bioinformatics and biology using regularized methods like PLS for regression and classification.

Keywords: partial least square regression, genetics data, negative filter factors, high dimensional data, high correlated data

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Authors: Binaya Tamang, Buddhhi Raj Pokhrel, Narayan Gautam

Abstract:

Background: There may be a connection between thyroid function and vitamin D status since both bind to similar nuclear hormone receptors and have similar response regions on gene promoters. The purpose of the current study was to investigate the relationship between thyroid hormones and vitamin D levels in females who were attending a tertiary care center in western Nepal and were either hypothyroid or euthyroid. Methods: This hospital-based cross-sectional study was carried out between March 2020 and March 2021 by the Biochemistry department of the Universal College of Medical Sciences (UCMS), Bhairahawa, Province No. 5, Nepal, in cooperation with Internal medicine. Prior to the study, institutional review committee approval (UCMS/IRC/008/20) was acquired from UCMS. Women who visited the Internal Medicine OPD of UCMS and were advised to get a thyroid function test (TFT) were included in the study population. Only those who were willing to participate in the study were enrolled after the goals and advantages of the study had been explained to them. Participants who had recently used vitamin D supplements and medications that affected thyroid hormones were excluded. The participants gave their consent verbally and in writing. After getting the consent, a convenient sample technique was applied. Serum was isolated after drawing 3 ml of blood in a plain vial. Chemiluminescence assay was used to analyze vitamin D and thyroid hormones (MAGLUMI 2000). SPSS version 16.0 for Windows was used to conduct the statistical analysis. Statistical significance was defined as a P-value < 0.05. Results: Majority of the study population (n=214, 71%) had insufficient serum vitamin D levels. Among the thyroid groups, the median Vitamin D levels were significantly lower in hypothyroid (16.88 ng/ml) as compared to the euthyroid groups (25.01 ng/ml) (P<0.001). Similarly, serum Vitamin D levels were considerably lower in the obese population (16.86 ng/ml) as compared to the normal BMI group (24.90 ng/ml) (P<0.001) as well as in the vegetarian (15.43 ng.ml) than mixed diet consumer (24.89 ng/ml) (P<0.01). Even after the adjustment for these variables, the Vitamin D levels were significantly lower in the hypothyroid population than in the euthyroid group (P<0.001). Conclusion: Comparing the hypothyroid population to the euthyroid, the median serum vitamin D levels were considerably lower. We were alarmed to see that the majority of euthyroid participants also had low levels of vitamin D. Therefore if left untreated, low vitamin D levels in hypothyroid patients could worsen their health further.

Keywords: vitamin D, thyroid hormones, euthyroid, hypothyroid, Nepal

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