Search results for: oral squamous cell carcinoma

Authors: Lethukuthula Ngobese, Abin Gupthar, Patrick Govender

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The ability of yeast cell wall-derived mannoproteins (glycoproteins) to positively contribute to oenological properties has been a key factor that stimulates research initiatives into these industrially important glycoproteins. In addition, and from a fundamental research perspective, yeast cell wall glycoproteins are involved in a wide range of biological interactions. To date, and to the best of our knowledge, our understanding of the fine molecular structure of these mannoproteins is fairly limited. Generally, the amino acid sequences of their protein moieties have been established from structural and functional analysis of the genomic sequence of these yeasts whilst far less information is available on the glycosyl moieties of these mannoproteins. A novel strategy was devised in this study that entails the genetic engineering of yeast strains that over-express and release cell wall-associated glycoproteins into the liquid growth medium. To this end, the Flo1p mannoprotein was overexpressed in Saccharomyces cerevisiae laboratory strains bearing a specific deletion in KNR4 and GPI7 genes involved in cell wall biosynthesis that have been previously shown to extracellularly hyper-secrete cell wall-associated glycoproteins. A polymerase chain reaction (PCR) -based cloning strategy was employed to generate transgenic yeast strains in which the native cell wall FLO1 glycoprotein-encoding gene is brought under transcriptional control of the constitutive PGK1 promoter. The modified Helm’s flocculation assay was employed to assess flocculation intensities of a Flo1p over-expressing wild type and deletion mutant as an indirect measure of their abilities to release the desired mannoprotein. The flocculation intensities of the transformed strains were assessed and all the strains showed similar intensities (>98% flocculation). To assess if mannoproteins were released into the growth medium, the supernatant of each strain was subjected to the BCA protein assay and the transformed Δknr4 strain showed a considerable increase in protein levels. This study has the potential to produce mannoproteins in sufficient quantities that may be employed in future investigations to understand their molecular structures and mechanisms of interaction to the benefit of both fundamental and industrial applications.

Keywords: glycoproteins, genetic engineering, flocculation, over-expression

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Authors: Virginia Dickson-Swift, Amanda Kenny, Jane Farmer, Sarah Larkins, Karen Carlisle, Helen Hickson

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In this presentation, we will focus on the methods of Remote Services Futures (RSF), an evidence-based method of community participation that was developed in Scotland. Using oral health as the focus, we will discuss the ways that RSF can be used to achieve sustainable engagement with stakeholders from various parts of the community. We will describe our findings of using RSF methods to engage with rural communities, including the steps involved and what happened when we asked people about the oral health services that they thought were needed in their community. We found that most community members started by thinking that a public dental clinic was required in every community, which is not a sustainable health service delivery option. Through a series of facilitated workshops, communities were able to discuss and prioritise their needs and develop a costed plan for their community which will ensure sustainable service delivery into the future. Our study highlights the complexities of decision making in rural communities. It is important to ensure that when communities participate in health care planning that the outcomes are practical, feasible and sustainable.

Keywords: community participation, sustainable health planning, Remote Services Futures, rural communities

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Authors: William Kelly, Sorelle Veigne, Xianhua Li, Zuyi Huang, Shyamsundar Subramanian, Eugene Schaefer

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The ambr15™ bioreactor is a single-use microbioreactor for cell line development and process optimization. The ambr system offers fully automatic liquid handling with the possibility of fed-batch operation and automatic control of pH and oxygen delivery. With operating conditions for large scale biopharmaceutical production properly scaled down, micro bioreactors such as the ambr15™ can potentially be used to predict the effect of process changes such as modified media or different cell lines. In this study, gassing rates and dilution rates were varied for a semi-continuous cell culture system in the ambr15™ bioreactor. The corresponding changes to metabolite production and consumption, as well as cell growth rate and therapeutic protein production were measured. Conditions were identified in the ambr15™ bioreactor that produced metabolic shifts and specific metabolic and protein production rates also seen in the corresponding larger (5 liter) scale perfusion process. A Dynamic Flux Balance model was employed to understand and predict the metabolic changes observed. The DFB model-predicted trends observed experimentally, including lower specific glucose consumption when CO₂ was maintained at higher levels (i.e. 100 mm Hg) in the broth. A Computational Fluid Dynamic (CFD) model of the ambr15™ was also developed, to understand transfer of O₂ and CO₂ to the liquid. This CFD model predicted gas-liquid flow in the bioreactor using the ANSYS software. The two-phase flow equations were solved via an Eulerian method, with population balance equations tracking the size of the gas bubbles resulting from breakage and coalescence. Reasonable results were obtained in that the Carbon Dioxide mass transfer coefficient (kLa) and the air hold up increased with higher gas flow rate. Volume-averaged kLa values at 500 RPM increased as the gas flow rate was doubled and matched experimentally determined values. These results form a solid basis for optimizing the ambr15™, using both CFD and FBA modelling approaches together, for use in microscale simulations of larger scale cell culture processes.

Keywords: cell culture, computational fluid dynamics, dynamic flux balance analysis, microbioreactor

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Authors: Murali Aarthy, Sanjeev Kumar Singh

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High risk human papillomaviruses are highly associated with the carcinoma of the cervix and the other genital tumors. Cervical cancer develops through the multistep process in which increasingly severe premalignant dysplastic lesions called cervical intraepithelial neoplastic progress to invasive cancer. The oncoprotein E7 of human papillomavirus expressed in the lower epithelial layers drives the cells into S-phase creating an environment conducive for viral genome replication and cell proliferation. The replication of the virus occurs in the terminally differentiating epithelium and requires the activation of cellular DNA replication proteins. To date, no suitable drug molecule is available to treat HPV infection whereas identification of potential drug targets and development of novel anti-HPV chemotherapies with unique mode of actions are expected. Hence, our present study aimed to identify the potential inhibitors analogous to EGCG, a green tea molecule which is considered to be safe to use for mammalian systems. A 3D similarity search on the natural small molecule library from natural product database using EGCG identified 11 potential hits based on their similarity score. The structure based docking strategies were implemented in the potential hits and the key interacting residues of protein with compounds were identified through simulation studies and binding free energy calculations. The conformational changes between the apoprotein and the complex were analyzed with the simulation and the results demonstrated that the dynamical and structural effects observed in the protein were induced by the compounds and indicated the dominance to the oncoprotein. Overall, our study provides the basis for the structural insights of the identified potential hits and EGCG and hence, the analogous compounds identified can be potent inhibitors against the HPV 16 E7 oncoprotein.

Keywords: EGCG, oncoprotein, molecular dynamics simulation, analogues

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Authors: Khaoula Bensaida, Osama Eljamal

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The electrical energy generation through Microbial Fuel Cells (MFCs) using microorganisms is a renewable and sustainable approach. It creates truly an efficient technology for power production and wastewater treatment. MFC is an electrochemical device which turns wastewater into electricity. The most important part of MFC is microbes. Nano zero-valent Iron NZVI technique was successfully applied in degrading the chemical pollutants and cleaning wastewater. However, the use of NZVI for enhancing the current production is still not confirmed yet. This study aims to confirm the effect of these particles on the current generation by using MFC. A constructed microbial fuel cell, which utilizes domestic wastewater, has been considered for wastewater treatment and bio-electricity generation. The two electrodes were connected to an external resistor (200 ohms). Experiments were conducted in two steps. First, the MFC was constructed without adding NZVI particles (Control) while at a second step, nanoparticles were added with a concentration of 50mg/L. After 20 hours, the measured voltage increased to 5 and 8mV, respectively. To conclude, the use of zero-valent iron in an MFC system can increase electricity generation.

Keywords: bacterial growth, electricity generation, microbial fuel cell MFC, nano zero-valent iron NZVI.

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Authors: Leo Nnamdi Ozurumba-Dwight

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Messenger ribooxynucleic acid (mRNA) molecules are compositional, protein-based. These proteins, encoding mRNA molecules (which collectively connote the transcriptome), when analyzed by RNA sequencing (RNAseq), unveils the nature of gene expression in the RNA. The obtained gene expression provides clues of cellular traits and their dynamics in presentations. These can be studied in relation to function and responses. RNAseq is a practical concept in Genomics as it enables detection and quantitative analysis of mRNA molecules. Single cell and spatial transcriptomics both present varying avenues for expositions in genomic characteristics of single cells and pooled cells in disease conditions such as cancer, auto-immune diseases, hematopoietic based diseases, among others, from investigated biological tissue samples. Single cell transcriptomics helps conduct a direct assessment of each building unit of tissues (the cell) during diagnosis and molecular gene expressional studies. A typical technique to achieve this is through the use of a single-cell RNA sequencer (scRNAseq), which helps in conducting high throughput genomic expressional studies. However, this technique generates expressional gene data for several cells which lack presentations on the cells’ positional coordinates within the tissue. As science is developmental, the use of complimentary pre-established tissue reference maps using molecular and bioinformatics techniques has innovatively sprung-forth and is now used to resolve this set back to produce both levels of data in one shot of scRNAseq analysis. This is an emerging conceptual approach in methodology for integrative and progressively dependable transcriptomics analysis. This can support in-situ fashioned analysis for better understanding of tissue functional organization, unveil new biomarkers for early-stage detection of diseases, biomarkers for therapeutic targets in drug development, and exposit nature of cell-to-cell interactions. Also, these are vital genomic signatures and characterizations of clinical applications. Over the past decades, RNAseq has generated a wide array of information that is igniting bespoke breakthroughs and innovations in Biomedicine. On the other side, spatial transcriptomics is tissue level based and utilized to study biological specimens having heterogeneous features. It exposits the gross identity of investigated mammalian tissues, which can then be used to study cell differentiation, track cell line trajectory patterns and behavior, and regulatory homeostasis in disease states. Also, it requires referenced positional analysis to make up of genomic signatures that will be sassed from the single cells in the tissue sample. Given these two presented approaches to RNA transcriptomics study in varying quantities of cell lines, with avenues for appropriate resolutions, both approaches have made the study of gene expression from mRNA molecules interesting, progressive, developmental, and helping to tackle health challenges head-on.

Keywords: transcriptomics, RNA sequencing, single cell, spatial, gene expression.

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Authors: Guillaume Soubeyran, Fabrice Micoud, Benoit Morin, Jean-Philippe Poirot-Crouvezier, Magali Reytier

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Proton Exchange Membrane Fuel Cell (PEMFC) technology is considered as a solution for the reduction of CO2 emissions. However, this technology still meets several challenges for high-scale industrialization. In this context, the increase of durability remains a critical aspect for competitiveness of this technology. Fortunately, performance degradations in nominal operating conditions is partially reversible, meaning that if specific conditions are applied, a partial recovery of fuel cell performance can be achieved, while irreversible degradations can only be mitigated. Thus, it is worth studying the optimal conditions to rejuvenate these reversible degradations and assessing the long-term impact of such procedures on the performance of the cell. Reversible degradations consist mainly of anode Pt active sites poisoning by carbon monoxide at the anode, heterogeneities in water management during use, and oxidation/deactivation of Pt active sites at the cathode. The latter is identified as a major source of reversible performance loss caused by the presence oxygen, high temperature and high cathode potential that favor platinum oxidation, especially in high efficiency operating points. Hence, we studied here a recovery procedure aiming at reducing the platinum oxides by decreasing cathode potential during operation. Indeed, the application of short air starvation phase leads to a drop of cathode potential. Cell performances are temporarily increased afterwards. Nevertheless, local temperature and current heterogeneities within the cells are favored and shall be minimized. The consumption of fuel during the recovery phase shall also be considered to evaluate the global efficiency. Consequently, the purpose of this work is to find an optimal compromise between the recovery of reversible degradations by air starvation, the increase of global cell efficiency and the mitigation of irreversible degradations effects. Different operating parameters have first been studied such as cell voltage, temperature and humidity in single cell set-up. Considering the global PEMFC system efficiency, tests showed that reducing duration of recovery phase and reducing cell voltage was the key to ensure an efficient recovery. Recovery phase frequency was a major factor as well. A specific method was established to find the optimal frequency depending on the duration and voltage of the recovery phase. Then, long-term degradations have also been studied by applying FC-DLC cycles based on NEDC cycles on a 4-cell short stack by alternating test sequences with and without recovery phases. Depending on recovery phase timing, cell efficiency during the cycle was increased up to 2% thanks to a mean voltage increase of 10 mV during test sequences with recovery phases. However, cyclic voltammetry tests results suggest that the implementation of recovery phases causes an acceleration of the decrease of platinum active areas that could be due to the high potential variations applied to the cathode electrode during operation.

Keywords: durability, PEMFC, recovery procedure, reversible degradation

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Authors: H. Oudira, A. Saifi

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The therapeutic utility of certain Auger emitters such as iodine-125 depends on their position within the cell nucleus . Or diagnostically, and to maintain as low as possible cell damage, it is preferable to have radionuclide localized outside the cell or at least the core. One solution to this problem is to consider markers capable of conveying anticancer drugs to the tumor site regardless of their location within the human body. The objective of this study is to simulate the impact of a complex such as bleomycin on single and double strand breaks in the DNA molecule. Indeed, this simulation consists of the following transactions: - Construction of BLM -Fe- DNA complex. - Simulation of the electron’s transport from the metastable state excitation of Fe 57 by the Monte Carlo method. - Treatment of chemical reactions in the considered environment by the diffusion equation. For physical, physico-chemical and finally chemical steps, the geometry of the complex is considered as a sphere of 50 nm centered on the binding site , and the mathematical method used is called step by step based on Monte Carlo codes.

Keywords: concentration, yield, radical species, bleomycin, excitation, DNA

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Authors: Pyi Phyo Maung

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In the study, the calculations of the permeability coefficient, values of the volume and porosity of a unit cell of a woven fabric before and after deformation based on the geometrical parameters are presented. Two types of carbon woven fabric structures were investigated: standard type, which integrated the filament, has a cross sectional shape of a cylinder and spread tow type, which has a rectangular cross sectional shape. The space antennas reflector, which distinctive feature is the presence of the surface of double curvature, is considered as the object of the research. Modeling of the kinetics of the process of impregnation of the reflector for the two types of carbon fabric’s unit cell structures was performed using software RAM-RTM. This work also investigated the influence of the grid angle between warp and welt of the unit cell on the duration of impregnation process. The results showed that decreasing the angle between warp and welt of the unit cell, the decreasing of the permeability values were occurred. Based on the results of calculation samples of the reflectors, their quality was determined. The comparisons of the theoretical and experimental results have been carried out. Comparison of the two textile structures (standard and spread tow) showed that the standard textiles with circular cross section were impregnated faster than spread tows, which have a rectangular cross section.

Keywords: vacuum assistant resin infusion, impregnation time, shear angle, reflector and modeling

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Authors: Sara Pereira-Nogueira, Tim Worbs, Marc Permanyer-Bosser, Reinhold Förster

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While the entry mechanism of lymphocytes into the lymph node via the blood are well described, it is still largely unknown how cells enter lymph nodes that arrive via afferent lymphatics. In order to address this, our group has established a micro-injection technique in mice through which cells are delivered directly into the lymphatic vessel immediately afferent to the popliteal lymph node. Injected cells can then be tracked via multi-colour fluorescence or 2-photon microscopy, and their localization can be analysed within the popliteal or downstream lymph nodes by immunohistology. Since naïve B cells express the chemokine receptor CXCR5 we intra-lymphatically co-injected B cells derived from wildtype and Cxcr5-deficient mice. While CXCR5 does not play a role in guiding B cells out of the subcapsular sinus, it affects their positioning within the lymph node parenchyma, since CXCR5-deficient B cells are impaired in migrating into the B cell follicle. The knowledge obtained by studying B-cell migration may prove beneficial in clinical settings regarding tumor metastasis or autoimmune diseases.

Keywords: afferent lymphatics, B cell migration, chemokine, intra-lymphatic injection

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Authors: Fatemeh Abbasi, Arne Hofemeier, Timo Betz

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Muscle growth and regeneration critically depend on satellite cells (SCs) which are muscle stem cells located between the basal lamina and myofibres. Upon damage, SCs become activated, enter the cell cycle, and give rise to myoblasts that form new myofibres, while a sub-population self-renew and re-populate the muscle stem cell niche. In aged muscle as well as in certain muscle diseases such as muscular dystrophy, some of the SCs lose their regenerative ability. Although it is demonstrated that the chemical composition of SCs quiescent niche is different from the activated niche, the mechanism initially activated in the SCs remains unknown. While extensive research efforts focused on potential chemical activation, no such factor has been identified to the author’s best knowledge. However, it is substantiated that niche mechanics affects SCs behaviors, such as stemness and engraftment. We hypothesize that mechanical stress in the healthy niche (homeostasis) is different from the regenerative niche and that this difference could serve as an early signal activating SCs upon fiber damage. To investigate this hypothesis, we develop a biomimetic system to reconstitute both, the mechanical and the chemical environment of the SC niche. Cells will be confined between two elastic polyacrylamide (PAA) hydrogels with controlled elastic moduli and functionalized surface chemistry. By controlling the distance between the PAA hydrogel surfaces, we vary the compression forces exerted by the substrates on the cells, while the lateral displacement of the upper hydrogel will create controlled shear forces. To establish such a system, a simplified system is presented. We engineered a sandwich-like configuration of two elastic PAA layer with stiffnesses between 1 and 10 kPa and confined a precursor myoblast cell line (C2C12) in between these layers. Our initial observations in this sandwich model indicate that C2C12 cells show different behaviors under mechanical compression if compared to a control one-layer gel without compression. Interestingly, this behavior is stiffness-dependent. While the shape of C2C12 cells in the sandwich consisting of two stiff (10 kPa) layers was much more elongated, showing almost a neuronal phenotype, the cell shape in a sandwich situation consisting of one stiff and one soft (1 kPa) layer was more spherical. Surprisingly, even in proliferation medium and at very low cell density, the sandwich situation stimulated cell differentiation with increased striation and myofibre formation. Such behavior is commonly found for confluent cells in differentiation medium. These results suggest that mechanical changes in stiffness and applied pressure might be a relevant stimulation for changes in muscle cell behavior.

Keywords: C2C12 cells, compression, force, satellite cells, skeletal muscle

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Authors: Rima Harche, Abdelkader Mouheb

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The Algiers refinery as all the other refineries always suffers from the problem of stopping of the tubes of heat exchanger. For that a study experimental of this phenomenon was undertaken in site on the cell of heat exchangers E101 (E101 CBA and E101 EDF) intended for the heating of the crude before its fractionation, which are exposed to the problem of the fouling on the side tubes exchangers. It is of tube-calenders type with head floating. Each cell is made up of three heat exchangers, laid out in series.

Keywords: fouling, fluid temperatue , oil, tubular heat exchanger, fouling resistance, modeling, heat transfer coefficient

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Authors: Leo Nnamdi Ozurumba-Dwight

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Single-cell genomic analytical systems have proved to be a platform to isolate bulk cells into selected single cells for genomic, proteomic, and related metabolomic studies. This is enabling systematic investigations of the level of heterogeneity in a diverse and wide pool of cell populations. Single cell technologies, embracing techniques such as high parameter flow cytometry, single-cell sequencing, and high-resolution images are playing vital roles in these investigations on messenger ribonucleic acid (mRNA) molecules and related gene expressions in tracking the nature and course of disease conditions. This entails targeted molecular investigations on unit cells that help us understand cell behavoiur and expressions, which can be examined for their health implications on the health state of patients. One of the vital good sides of single-cell RNA sequencing (scRNA seq) is its probing capacity to detect deranged or abnormal cell populations present within homogenously perceived pooled cells, which would have evaded cursory screening on the pooled cell populations of biological samples obtained as part of diagnostic procedures. Despite conduction of just single-cell transcriptome analysis, scRNAseq now permits comparison of the transcriptome of the individual cells, which can be evaluated for gene expressional patterns that depict areas of heterogeneity with pharmaceutical drug discovery and clinical treatment applications. It is vital to strictly work through the tools of investigations from wet lab to bioinformatics and computational tooled analyses. In the precise steps for scRNAseq, it is critical to do thorough and effective isolation of viable single cells from the tissues of interest using dependable techniques (such as FACS) before proceeding to lysis, as this enhances the appropriate picking of quality mRNA molecules for subsequent sequencing (such as by the use of Polymerase Chain Reaction machine). Interestingly, scRNAseq can be deployed to analyze various types of biological samples such as embryos, nervous systems, tumour cells, stem cells, lymphocytes, and haematopoietic cells. In haematopoietic cells, it can be used to stratify acute myeloid leukemia patterns in patients, sorting them out into cohorts that enable re-modeling of treatment regimens based on stratified presentations. In immunotherapy, it can furnish specialist clinician-immunologist with tools to re-model treatment for each patient, an attribute of precision medicine. Finally, the good predictive attribute of scRNAseq can help reduce the cost of treatment for patients, thus attracting more patients who would have otherwise been discouraged from seeking quality clinical consultation help due to perceived high cost. This is a positive paradigm shift for patients’ attitudes primed towards seeking treatment.

Keywords: immunotherapy, transcriptome, re-modeling, mRNA, scRNA-seq

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Authors: Anthony S. Machi, Joseph Minardi

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We present a case report of left atrial myxoma diagnosed by bedside transesophageal (TEE) ultrasound. Left atrial myxoma is the most common benign cardiac tumor and can obstruct blood flow and cause valvular insufficiency. Common symptoms consist of dyspnea, pulmonary edema and other features of left heart failure in addition to thrombus release in the form of tumor fragments. The availability of bedside ultrasound equipment is essential for the quick diagnosis and treatment of various emergency conditions including cardiac neoplasms. A 48-year-old Caucasian female with a four-year history of an untreated renal mass and anemia presented to the ED with two months of sharp, intermittent, bilateral flank pain radiating into the abdomen. She also reported intermittent vomiting and constipation along with generalized body aches, night sweats, and 100-pound weight loss over last year. She had a CT in 2013 showing a 3 cm left renal mass and a second CT in April 2016 showing a 3.8 cm left renal mass along with a past medical history of diverticulosis, chronic bronchitis, dyspnea on exertion, uncontrolled hypertension, and hyperlipidemia. Her maternal family history is positive for breast cancer, hypertension, and Type II Diabetes. Her paternal family history is positive for stroke. She was a current everyday smoker with an 11 pack/year history. Alcohol and drug use were denied. Physical exam was notable for a Grade II/IV systolic murmur at the right upper sternal border, dyspnea on exertion without angina, and a tender left lower quadrant. Her vitals and labs were notable for a blood pressure of 144/96, heart rate of 96 beats per minute, pulse oximetry of 96%, hemoglobin of 7.6 g/dL, hypokalemia, hypochloremia, and multiple other abnormalities. Physicians ordered a CT to evaluate her flank pain which revealed a 7.2 x 8.9 x 10.5 cm mixed cystic/solid mass in the lower pole of the left kidney and a filling defect in the left atrium. Bedside TEE was ordered to follow up on the filling defect. TEE reported an ejection fraction of 60-65% and visualized a mobile 6 x 3 cm mass in the left atrium attached to the interatrial septum extending into the mitral valve. Cardiothoracic Surgery and Urology were consulted and confirmed a diagnosis of left atrial myxoma and clear cell renal cell carcinoma. The patient returned a week later due to worsening nausea and vomiting and underwent emergent nephrectomy, lymph node dissection, and colostomy due to a necrotic colon. Her condition declined over the next four months due to lung and brain metastases, infections, and other complications until she passed away.

Keywords: bedside ultrasound, echocardiography, emergency medicine, left atrial myxoma

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Authors: Ya L. V., Benjamin Ong Wei Lin, Wanli Niu, Benjamin Seah Chin Tat

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This article introduces a methodology that improves reliability and cyclability of 2nd-life Li-ion battery system repurposed as an energy storage system (ESS). Most of the 2nd-life retired battery systems in the market have module/pack-level state-of-health (SOH) indicator, which is utilized for guiding appropriate depth-of-discharge (DOD) in the application of ESS. Due to the lack of cell-level SOH indication, the different degrading behaviors among various cells cannot be identified upon reaching retired status; in the end, considering end-of-life (EOL) loss and pack-level DOD, the repurposed ESS has to be oversized by > 1.5 times to complement the application requirement of reliability and cyclability. This proposed battery grading algorithm, using non-invasive methodology, is able to detect outlier cells based on historical voltage data and calculate cell-level historical maximum temperature data using semi-analytic methodology. In this way, the individual battery cell in the 2nd-life battery system can be graded in terms of SOH on basis of the historical voltage fluctuation and estimated historical maximum temperature variation. These grades will have corresponding DOD grades in the application of the repurposed ESS to enhance system reliability and cyclability. In all, this introduced battery grading algorithm is non-invasive, compatible with all kinds of retired Li-ion battery systems which lack of cell-level SOH indication, as well as potentially being embedded into battery management software for preventive maintenance and real-time cyclability optimization.

Keywords: battery grading algorithm, 2nd-life repurposing battery system, semi-analytic methodology, reliability and cyclability

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Authors: David A. Ullisch, Yantree D. Sankar-Thomas, Stefan Wilke, Thomas Selge, Matthias Pump, Thomas Leibold, Kai Schütte, Gilbert Gorr

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Plants have been considered as a source of natural substances for ages. Secondary metabolites from plants are utilized especially in medical applications but are more and more interesting as cosmetical ingredients and in the field of nutraceuticals. However, supply of compounds from natural harvest can be limited by numerous factors i.e. endangered species, low product content, climate impacts and cost intensive extraction. Especially in the pharmaceutical industry the ability to provide sufficient amounts of product and high quality are additional requirements which in some cases are difficult to fulfill by plant harvest. Whereas in many cases the complexity of secondary metabolites precludes chemical synthesis on a reasonable commercial basis, plant cells contain the biosynthetic pathway – a natural chemical factory – for a given compound. A promising approach for the sustainable production of natural products can be plant cell fermentation (PCF®). A thoroughly accomplished development process comprises the identification of a high producing cell line, optimization of growth and production conditions, the development of a robust and reliable production process and its scale-up. In order to address persistent, long lasting production, development of cryopreservation protocols and generation of working cell banks is another important requirement to be considered. So far the most prominent example using a PCF® process is the production of the anticancer compound paclitaxel. To demonstrate the power of plant suspension cultures here we present three case studies: 1) For more than 17 years Phyton produces paclitaxel at industrial scale i.e. up to 75,000 L in scale. With 60 g/kg dw this fully controlled process which is applied according to GMP results in outstanding high yields. 2) Thapsigargin is another anticancer compound which is currently isolated from seeds of Thapsia garganica. Thapsigargin is a powerful cytotoxin – a SERCA inhibitor – and the precursor for the derivative ADT, the key ingredient of the investigational prodrug Mipsagargin (G-202) which is in several clinical trials. Phyton successfully generated plant cell lines capable to express this compound. Here we present data about the screening for high producing cell lines. 3) The third case study covers ingenol-3-mebutate. This compound is found in the milky sap of the intact plants of the Euphorbiacae family at very low concentrations. Ingenol-3-mebutate is used in Picato® which is approved against actinic keratosis. Generation of cell lines expressing significant amounts of ingenol-3-mebutate is another example underlining the strength of plant cell culture. The authors gratefully acknowledge Inspyr Therapeutics for funding.

Keywords: Ingenol-3-mebutate, plant cell culture, sustainability, thapsigargin

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Authors: Jienny Lee, In-Soo Cho, Sang-Ho Cha

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Adult mesenchymal stem cells (MSCs) have been investigated using preclinical approaches for tissue regeneration. Porcine MSCs (pMSCs) are capable of growing and attaching to plastic with a fibroblast-like morphology and then differentiating into bone, adipose, and cartilage tissues in vitro. This study was conducted to investigate the proliferating abilities, differentiation potentials, and multipotency of miniature pig adipose tissue-derived MSCs (mpAD-MSCs) with or without long-term cryopreservation, considering that cryostorage has the potential for use in clinical applications. After confirming the characteristics of the mpAD-MSCs, we examined the effect of long-term cryopreservation (> 2 years) on expression of cell surface markers (CD34, CD90 and CD105), proliferating abilities (cumulative population doubling level, doubling time, colony-forming unit, and MTT assay) and differentiation potentials into mesodermal cell lineages. As a result, the expression of cell surface markers is similar between thawed and fresh mpAD-MSCs. However, long-term cryopreservation significantly lowered the differentiation potentials (adipogenic, chondrogenic, and osteogenic) of mpAD-MSCs. When compared with fresh mpAD-MSCs, thawed mpAD-MSCs exhibited lower expression of mesodermal cell lineage-related genes such as peroxisome proliferator-activated receptor-g2, lipoprotein lipase, collagen Type II alpha 1, osteonectin, and osteocalcin. Interestingly, long-term cryostoraged mpAD-MSCs exhibited significantly higher cell viability than the fresh mpAD-MSCs. Long-term cryopreservation induced a 30% increase in the cell viability of mpAD-MSCs when compared with the fresh mpAD-MSCs at 5 days after thawing. However, long-term cryopreservation significantly lowered expression of stemness markers such as Oct3/4, Sox2, and Nanog. Furthermore, long-term cryopreservation negatively affected expression of senescence-associated genes such as telomerase reverse transcriptase and heat shock protein 90 of mpAD-MSCs when compared with the fresh mpAD-MSCs. The results from this study might be important for the successful application of MSCs in clinical trials after long-term cryopreservation.

Keywords: mesenchymal stem cells, cryopreservation, stemness, senescence

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Authors: Vineeth Siripuram, Abhineet Nigam

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A bioreactor is an engineered system that supports a biologically active environment. Along the years, the advancements in bioreactors have been widely accepted all over the world for varied applications ranging from sewage treatment to tissue cloning. Driven by tissue and organ shortage, tissue engineering has emerged as an alternative to transplantation for the reconstruction of lost or damaged organs. In this study, Computational fluid dynamics (CFD) has been used to model porous medium flow in scaffolds (taken from the literature) with different flow patterns. A detailed analysis of different scaffold geometries and their influence on cell deposition in the perfusion system is been carried out using Computational fluid dynamics (CFD). Considering the fact that, the scaffold should mimic the organs or tissues structures in a three-dimensional manner, certain assumptions were made accordingly. The research on scaffolds has been extensively carried out in different bioreactors. However, there has been less focus on the morphology of the scaffolds and the flow patterns in which the perfusion system is laid upon. The objective of this paper is to employ a computational approach using CFD simulation to determine the optimal morphology and the anisotropic measurements of the various samples of scaffolds. Using predictive computational modelling approach, variables which exert dominant effects on the cell deposition within the scaffold were prioritised and corresponding changes in morphology of scaffold and flow patterns in the perfusion systems are made. A Eulerian approach was carried on in multiple CFD simulations, and it is observed that the morphological and topological changes in the scaffold perfusion system are of great importance in the commercial applications of scaffolds.

Keywords: cell seeding, CFD, flow patterns, modelling, perfusion systems, scaffold

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Authors: Vongsathorn Ngampuak, Yutachai Chookaew, Wipawee Dejtisakdi

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Dunaliella salina has great potential as a system for generating commercially valuable products, including beta-carotene, pharmaceuticals, and biofuels. Our goal is to improve this potential by enhancing growth rate and other properties of D. salina under optimal growth conditions. We used ethyl methane sulfonate (EMS) to generate random mutants in D. salina KU11, a strain classified in Thailand. In a preliminary experiment, we first treated D. salina cells with 0%, 0.8%, 1.0%, 1.2%, 1.44% and 1.66% EMS to generate a killing curve. After that, we randomly picked 30 candidates from approximately 300 isolated survivor colonies from the 1.44% EMS treatment (which permitted 30% survival) as an initial test of the mutant screen. Among the 30 survivor lines, we found that 2 strains (mutant #17 and #24) had significantly improved growth rates and cell number accumulation at stationary phase approximately up to 1.8 and 1.45 fold, respectively, 2 strains (mutant #6 and #23) had significantly decreased growth rates and cell number accumulation at stationary phase approximately down to 1.4 and 1.35 fold, respectively, while 26 of 30 lines had similar growth rates compared with the wild type control. We also analyzed cell size for each strain and found there was no significant difference comparing all mutants with the wild type. In addition, mutant #24 had shown an increase of biomass accumulation approximately 1.65 fold compared with the wild type strain on day 5 that was entering early stationary phase. From these preliminary results, it could be feasible to identify D. salina mutants with significant improved growth rate, cell accumulation and biomass production compared to the wild type for the further study; this makes it possible to improve this microorganism as a platform for biotechnology application.

Keywords: Dunaliella salina, ethyl methyl sulfonate, growth rate, biomass

Procedia PDF Downloads 241

Authors: Iulianna Taritsa, Kuldeep Neote, Eric Fossel

Abstract:

Lysosome-induced immunogenic cell death (LIICD) is a powerful mechanism of targeting cancer cells that kills circulating malignant cells and primes the host’s immune cells against future remission. Current immunotherapies for cancer are limited in preventing recurrence – a gap that can be bridged by training the immune system to recognize cancer neoantigens. Lysosomal leakage can be induced therapeutically to traffic antigens from dying cells to dendritic cells, which can later present those tumorigenic antigens to T cells. Previous research has shown that oxidative agents administered in the tumor microenvironment can initiate LIICD. We generated a fusion protein between an oxidative agent known as xanthine oxidase (XO) and a mini-antibody specific for EGFR/HER2-sensitive breast tumor cells. The anti-EGFR single domain antibody fragment is uniquely sourced from llama, which is functional without the presence of a light chain. These llama micro-antibodies have been shown to be better able to penetrate tissues and have improved physicochemical stability as compared to traditional monoclonal antibodies. We demonstrate that the fusion protein created is stable and can induce early markers of immunogenic cell death in an in vitro human breast cancer cell line (SkBr3). Specifically, we measured overall cell death, as well as surface-expressed calreticulin, extracellular ATP release, and HMGB1 production. These markers are consensus indicators of ICD. Flow cytometry, luminescence assays, and ELISA were used respectively to quantify biomarker levels between treated versus untreated cells. We also included a positive control group of SkBr3 cells dosed with doxorubicin (a known inducer of LIICD) and a negative control dosed with cisplatin (a known inducer of cell death, but not of the immunogenic variety). We looked at each marker at various time points after cancer cells were treated with the XO/antibody fusion protein, doxorubicin, and cisplatin. Upregulated biomarkers after treatment with the fusion protein indicate an immunogenic response. We thus show the potential for this fusion protein to induce an anticancer effect paired with an adaptive immune response against EGFR/HER2+ cells. Our research in human cell lines here provides evidence for the success of the same therapeutic method for patients and serves as the gateway to developing a new treatment approach against breast cancer.

Keywords: apoptosis, breast cancer, immunogenic cell death, lysosome

Procedia PDF Downloads 199

Authors: Olfat A. Fadali, Mohamed S. Mahmoud, Omnia H. Abdelraheem, Shimaa G. Mohammed

Abstract:

The effect of electromagnetic field (EMF) on the removal of edible oil from oil-in-water emulsion by means of electrocoagulation was investigated in rectangular batch electrochemical cell with DC current. Iron (Fe) plate anodes and stainless steel cathodes were employed as electrodes. The effect of different magnetic field intensities (1.9, 3.9 and 5.2 tesla), three different positions of EMF (below, perpendicular and parallel to the electrocoagulation cell), as well as operating time; had been investigated. The application of electromagnetic field (5.2 tesla) raises percentage of oil removal from 72.4% for traditional electrocoagulation to 90.8% after 20 min.

Keywords: electrocoagulation, electromagnetic field, Oil-water emulsion, edible oil

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Authors: Navarro C., Jahier B., Gard C.

Abstract:

Diarrhoea is a common disorder in both cats and dogs. Recent guidelines highlight the importance of gut microbiota and the use of strategies such as prebiotics, probiotics, postbiotics, and fecal transplants for modulating the microbiota. The objective of this study was to evaluate the palatability of a flavoured oral paste containing prebiotics (brewer’s yeast products, fructo-oligosaccharide), probiotics (Enterococcus faecium), and postbiotics (lactic ferment products), in dogs and cats. Material and methods: Healthy adult animals (cats and dogs) from various breeds received the tested product (Ultradiar® Biotic, MP Labo, France) at the recommended dosage over a small quantity of kibbles: animals less than 2 kg bodyweight received 2 ml per day, animals between2 and 6 kg received 4 ml per day, animals between 6 and 12 kg received 5 ml per day, animals between 12 and 30 kg received 8 ml per day, and animals weighing more than 30 kg received 10 ml per day. For each animal, the investigator noted the intake (immediate in less than 2 seconds, delayed after 2 seconds, no intake), the consumption of the product (no consumption, partial consumption ≤ 5%, < 50%, 50% ≤ x < 95%, ≥ 95%, total consumption). Acceptability was defined as the percentage of dogs having consumed more than 95% of the product. Results: Thirty-seven dogs were included: 19 small size, 11 medium size, and 7 large size dogs. Thirty-six dogs (97%) took the product, with 65% showing immediate intake. Only one small-sized dog did not take the product. Among the 36 dogs who took the product, 19 (53%) had a complete consumption, 13 (36%) consumed more than 95% of the product, 3 dogs consumed more than 50% (and less than 95%), and one dog consumed less than 50%. The acceptability rate was 86%. Thirty-seven cats were included. Twenty-eight cats (76%) took the product, with 8% showing immediate intake. Among those 28 cats, 7 (25%) consumed more than 95% of the product, 13 (47%) consumed more than 50% (and less than 95%), 6 consumed less than 50% and 2 cats consumed less than 5%. Conclusion: The flavoured oral paste, Ultradiar® Biotic, was well-accepted by both dogs and cats, with higher acceptability observed in dogs compared to cats. These results suggest that the product is palatable and can be usefully administered to support gastrointestinal health in companion animals. Further studies should explore the clinical benefits of this formulation in managing gastrointestinal disorders.

Keywords: cat, dog, palatability, prebiotic, probiotic

Procedia PDF Downloads 39

Authors: Jimmy Jiun-Ming Su, Yuan-Min Lin

Abstract:

Bioprinting has been applied to produce 3D cellular constructs for tissue engineering. Microextrusion printing is the most common used method. However, printing low viscosity bioink is a challenge for this method. Herein, we developed a new 3D printing system to fabricate cell-laden hydrogels via a DLP-based projector. The bioprinter is assembled from affordable equipment including a stepper motor, screw, LED-based DLP projector, open source computer hardware and software. The system can use low viscosity and photo-polymerized bioink to fabricate 3D tissue mimics in a layer-by-layer manner. In this study, we used gelatin methylacrylate (GelMA) as bioink for stem cell encapsulation. In order to reinforce the printed construct, surface modified hydroxyapatite has been added in the bioink. We demonstrated the silanization of hydroxyapatite could improve the crosslinking between the interface of hydroxyapatite and GelMA. The results showed that the incorporation of silanized hydroxyapatite into the bioink had an enhancing effect on the mechanical properties of printed hydrogel, in addition, the hydrogel had low cytotoxicity and promoted the differentiation of embedded human bone marrow stem cells (hBMSCs) and retinal pigment epithelium (RPE) cells. Moreover, this bioprinting system has the ability to generate microchannels inside the engineered tissues to facilitate diffusion of nutrients. We believe this 3D bioprinting system has potential to fabricate various tissues for clinical applications and regenerative medicine in the future.

Keywords: bioprinting, cell encapsulation, digital light processing, GelMA hydrogel

Procedia PDF Downloads 181

Authors: Mengying Wang, Dongjing Liu, Holger Schwender, Ping Wang, Hongping Zhu, Tao Wu, Terri H Beaty

Abstract:

Background: Maternal smoking is a recognized risk factor for nonsyndromic cleft lip with or without cleft palate (NSCL/P). It has been reported that the effect of maternal smoking on oral clefts is mediated through genes that influence nicotine dependence. The polymorphisms of cholinergic receptor nicotinic alpha (CHRNA) and beta (CHRNB) subunits genes have previously shown strong associations with nicotine dependence. Here, we attempted to investigate whether the above genes are associated with clefting risk through testing for potential gene-gene (G×G) and gene-environment (G×E) interaction. Methods: We selected 120 markers in 14 genes associated with nicotine dependence to conduct transmission disequilibrium tests among 806 Chinese NSCL/P case-parent trios ascertained in an international consortium which conducted a genome-wide association study (GWAS) of oral clefts. We applied Cordell’s method using “TRIO” package in R to explore G×G as well as G×E interaction involving environmental tobacco smoke (ETS) based on conditional logistic regression model. Results: while no SNP showed significant association with NSCL/P after Bonferroni correction, we found signals for G×G interaction between 10 pairs of SNPs in CHRNA3, CHRNA5, and CHRNB4 (p<10-8), among which the most significant interaction was found between RS3743077 (CHRNA3) and RS11636753 (CHRNB4, p<8.2×10-12). Linkage disequilibrium (LD) analysis revealed only low level of LD between these markers. However, there were no significant results for G×ETS interaction. Conclusion: This study fails to detect association between nicotine dependence genes and NSCL/P, but illustrates the importance of taking into account potential G×G interaction for genetic association analysis in NSCL/P. This study also suggests nicotine dependence genes should be considered as important candidate genes for NSCL/P in future studies.

Keywords: Gene-Gene Interaction, Maternal Smoking, Nicotine Dependence, Non-Syndromic Cleft Lip with or without Cleft Palate

Procedia PDF Downloads 337

Authors: Md. M. Hossain, Regan Roat, Jenica Christopherson, Colette Free, Zhiguang Guo

Abstract:

Long noncoding RNA (lncRNA) mediated post-transcriptional gene regulation, and their epigenetic landscapes have been shown to be involved in many human diseases. However, their regulation in diabetes through governing islet’s β-cell function and survival needs to be elucidated. Due to the technical and ethical constraints, it is difficult to study their role in β-cell function and survival in human under in vivo condition. In this study, humanized mice have been developed through transplanting human pancreatic islet under the kidney capsule of NOD.SCID mice and induced β-cell death leading to diabetes condition to study lncRNA mediated regulation. For this, human islets from 3 donors (3000 IEQ, purity > 80%) were transplanted under the kidney capsule of STZ induced diabetic NOD.scid mice. After at least 2 weeks of normoglycecemia, lymphocytes from diabetic NOD mice were adoptively transferred and islet grafts were collected once blood glucose reached > 200 mg/dl. RNA from human donor islets, islet grafts from humanized mice with either adoptive lymphocyte transfer (ALT) or PBS control (CTL) were ribodepleted; barcoded fragment libraries were constructed and sequenced on the Ion Proton sequencer. lncRNA expression in isolated human islets, islet grafts from humanized mice with and without induced β-cell death and their regulation in human islets function in vitro under glucose challenge, cytokine mediated inflammation and induced apoptotic condition were investigated. Out of 3155 detected lncRNAs, 299 that highly expressed in islets were found to be significantly downregulated and 224 upregulated in ALT compared to CTL. Most of these are found to be collocated within 5 kb upstream and 1 kb downstream of 788 up- and 624 down-regulated mRNAs. Genomic Regions Enrichment of Annotations Analysis revealed deregulated and collocated genes are related to pancreas endocrine development; insulin synthesis, processing, and secretion; pancreatitis and diabetes. Many of them, that found to be located within enhancer domains for islet specific gene activity, are associated to the deregulation of known islet/βcell specific transcription factors and genes that are important for β-cell differentiation, identity, and function. RNA sequencing analysis revealed aberrant lncRNA expression which is associated to the deregulated mRNAs in β-cell function as well as in molecular pathways related to diabetes. A distinct set of candidate lncRNA isoforms were identified as highly enriched and specific to human islets, which are deregulated in human islets from donors with different BMIs and with type 2 diabetes. These RNAs show an interesting regulation in cultured human islets under glucose stimulation and with induced β-cell death by cytokines. Aberrant expression of these lncRNAs was detected in the exosomes from the media of islets cultured with cytokines. Results of this study suggest that the islet specific lncRNAs are deregulated in human islet with β-cell death, hence important in diabetes. These lncRNAs might be important for human β-cell function and survival thus could be used as biomarkers and novel therapeutic targets for diabetes.

Keywords: β-cell, humanized mouse, pancreatic islet, LncRNAs

Procedia PDF Downloads 164

Authors: Myoung-Jin Kim, Yeong-Seop Ahn, Hyun-Jee Yang, Hyoung-Kyu Song

Abstract:

This paper proposes the new cooperative communication scheme for the wireless communication system. When the receiver is located in the edge of coverage, the signal from the transmitter is distorted by the inter-cell interference (ICI) and power reduction by distance. In order to improve communication performance, the proposed scheme adds the relay. By using the relay, the receiver receives the signal from the transmitter and relay at the same time. Therefore, the new cooperative communication scheme obtains diversity gain and is improved by the relay.

Keywords: cooperative communication, diversity gain, OFDM, MIMO

Procedia PDF Downloads 609

Authors: Masho Mebrahtom Gebrehiwot, Gidey Yirga

Abstract:

Tigray highlands in northern Ethiopia, is characterized by a wide range of ecological conditions and climate. The siege of Tigray is believed to cause the deaths of nearly 600,000 civilians mainly due to starvation and lack of medicine. In this study, the most important edible and medicinal plants used during the siege of Tigray were investigated. Semi-structured interviews, observation and guided field walks were used in 500 informants (300 males and 200 females) selected randomly from two districts. A total of 25 species of medicinal plants were collected and identified for treating 30 human ailments. Furthermore, a total of 21 edible plants were also collected and identified. Nearly 68.75% of these species were wild and harvested mainly for their leaves and the remedies were administered through dermal, nasal and oral routes. Oral and dermal applications were the highest and most used route of application. Famen foods significantly saved thousands of human lives during the siege of Tigray both in urban and rural communities. We suggest domestication of some of the wild medicinal plants for long term conservation of the species. Documentation of farmers’ knowledge, attitude and practices of ethnobotany would be very important before the indigenous knowledge is lost forever.

Keywords: ethnobotany, tigray, siege, application

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Authors: Neil Erick Q. Madariaga, Noel B. Linsangan

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Body Mass Index (BMI) is one of the different ways to monitor the health of a person. It is based on the height and weight of the person. This study aims to compute for the BMI using an Android tablet by obtaining the height of the person by using a camera and measuring the weight of the person by using a weighing scale or load cell. The height of the person was estimated by applying background subtraction to the image captured and applying different processes such as getting the vanishing point and applying Artificial Neural Network. The weight was measured by using Wheatstone bridge load cell configuration and sending the value to the computer by using Gizduino microcontroller and Bluetooth technology after the amplification using AD620 instrumentation amplifier. The application will process the images and read the measured values and show the BMI of the person. The study met all the objectives needed and further studies will be needed to improve the design project.

Keywords: body mass index, artificial neural network, vanishing point, bluetooth, wheatstone bridge load cell

Procedia PDF Downloads 324

Authors: Tarek Elnimr, Reda Morsy, Assem El Fert, Aziza Ismail

Abstract:

Hepatitis is an inflammation of the liver. The condition can be self-limiting or can progress to fibrosis, cirrhosis or liver cancer. Disease caused by the hepatitis virus, the virus can cause hepatitis infection, ranging in severity from a mild illness lasting a few weeks to a serious, lifelong illness. A growing body of evidence indicates that many trace elements play important roles in a number of carcinogenic processes that proceed with various mechanisms. To examine the status of trace elements during the development of hepatic carcinoma, we determined the iron, copper, zinc and selenium levels in some biological samples of patients at different stages of viral hepatic disease. We observed significant changes in the iron, copper, zinc and selenium levels in the biological samples of patients hepatocellular carcinoma, relative to those of healthy controls. The mean hair, nail, RBC, serum and whole blood copper levels in patients with hepatitis virus were significantly higher than that of the control group. In contrast the mean iron, zinc, and selenium levels in patients having hepatitis virus were significantly lower than those of the control group. On the basis of this study, we identified the impact of natural supplements to improve the treatment of viral liver damage, using the level of some trace elements such as, iron, copper, zinc and selenium, which might serve as biomarkers for increases survival and reduces disease progression. Most of the elements revealed diverse and random distribution in the samples of the donor groups. The correlation study pointed out significant disparities in the mutual relationships among the trace elements in the patients and controls. Principal component analysis and cluster analysis of the element data manifested diverse apportionment of the selected elements in the scalp hair, nail and blood components of the patients compared with the healthy counterparts.

Keywords: hepatitis, hair, nail, blood components, trace element, nutrition therapy, multivariate analysis, correlation, ICP-MS

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Authors: Neda Sinaei, Davood Zare, Mehrdad Azin

Abstract:

Polyhydroxyalkanoates (PHAs) are biocompatible and biodegradable polymers produced by a wide range of bacterial strains. These biopolymers are significantly studied for drug delivery and tissue engineering applications because of their fascinating physicochemical properties. Polyhydroxybutyrate (PHB) scaffold that has been extracted from a novel bacteria using oil wastewater was selected to study. Some physical parameters affecting scaffold properties such as PHB concentration, solvent evaporation speed, and ultrasonic time were investigated. Scanning electron microscopy was used to evaluate the porosity. Afterward, the biocompatibility of PHB scaffold was assessed. Initial results showed the highly porous PHB scaffold structure with a variety of pore sizes. Subsequent results indicated that more unique pore sizes can be obtained by optimizing physical factors. It would be noticed that the morphology of the pore structure was accordingly affected by ultrasonic time. Hence, In vitro cell viability tests on the PHB scaffold using human foreskin fibroblasts revealed strong cell attachment and proliferation supports. Therefore, it can be concluded that the cost-effective PHB scaffold has the potential using as a biomaterial cell adhesion substrate in therapeutic applications.

Keywords: Polyhydroxybutyrate, biocompatible, scaffold, porous, tissue engineering

Procedia PDF Downloads 232