Search results for: single cell

Authors: Samineh Barmaki, Ville Jokinen, Esko Kankuri

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We report a microfluidic chip that can be used to modify the gaseous microenvironment of a cell-culture in ambient atmospheric conditions. The aim of the study is to show the cellular response to nitric oxide (NO) under hypoxic (oxygen < 5%) condition. Simultaneously targeting to hypoxic and nitric oxide will provide an opportunity for NO‑based therapeutics. Studies on cellular responses to lowered oxygen concentration or to gaseous mediators are usually carried out under a specific macro environment, such as hypoxia chambers, or with specific NO donor molecules that may have additional toxic effects. In our study, the chip consists of a microfluidic layer and a cell culture well, separated by a thin gas permeable polydimethylsiloxane (PDMS) membrane. The main design goal is to separate the gas oxygen scavenger and NO donor solutions, which are often toxic, from the cell media. Two different types of gas exchangers, titled 'pool' and 'meander' were tested. We find that the pool design allows us to reach a higher level of oxygen depletion than meander (24.32 ± 19.82 %vs -3.21 ± 8.81). Our microchip design can make the cells culture more simple and makes it easy to adapt existing cell culture protocols. Our first application is utilizing the chip to create hypoxic conditions on targeted areas of cell culture. In this study, oxygen scavenger sodium sulfite generates hypoxia and its effect on human embryonic kidney cells (HEK-293). The PDMS membrane was coated with fibronectin before initiating cell cultures, and the cells were grown for 48h on the chips before initiating the gas control experiments. The hypoxia experiments were performed by pumping of O₂-depleted H₂O into the microfluidic channel with a flow-rate of 0.5 ml/h. Image-iT® reagent as an oxygen level responser was mixed with HEK-293 cells. The fluorescent signal appears on cells stained with Image-iT® hypoxia reagent (after 6h of pumping oxygen-depleted H₂O through the microfluidic channel in pool area). The exposure to different levels of O₂ can be controlled by varying the thickness of the PDMS membrane. Recently, we improved the design of the microfluidic chip, which can control the microenvironment of two different gases at the same time. The hypoxic response was also improved from the new design of microchip. The cells were grown on the thin PDMS membrane for 30 hours, and with a flowrate of 0.1 ml/h; the oxygen scavenger was pumped into the microfluidic channel. We also show that by pumping sodium nitroprusside (SNP) as a nitric oxide donor activated under light and can generate nitric oxide on top of PDMS membrane. We are aiming to show cellular microenvironment response of HEK-293 cells to both nitric oxide (by pumping SNP) and hypoxia (by pumping oxygen scavenger solution) in separated channels in one microfluidic chip.

Keywords: hypoxia, nitric oxide, microenvironment, microfluidic chip, sodium nitroprusside, SNP

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Authors: Denis Mustafov, Emmanouil Karteris, Maria Braoudaki

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Glioblastoma multiform (GBM) is a heterogenous primary brain tumour that kills most affected patients. To the authors best knowledge, despite all research efforts there is no early diagnostic biomarker for GBM. MicroRNAs (miRNAs) are short non-coding RNA molecules which are deregulated in many cancers. The aim of this research was to determine miRNAs with a diagnostic impact and to potentially identify promising therapeutic targets for glioblastoma multiform. In silico analysis was performed to identify deregulated miRNAs with diagnostic relevance for glioblastoma. The expression profiles of the chosen miRNAs were then validated in vitro in the human glioblastoma cell lines A172 and U-87MG. Briefly, RNA extraction was carried out using the Trizol method, whilst miRNA extraction was performed using the mirVANA miRNA isolation kit. Quantitative Real-Time Polymerase Chain Reaction was performed to verify their expression. The presence of five target proteins within the A172 cell line was evaluated by Western blotting. The expression of the CORO1C protein within 32 GBM cases was examined via immunohistochemistry. The miRNAs identified in silico included miR-21-5p, miR-34a and miR-128a. These miRNAs were shown to target deregulated GBM genes, such as CDK6, E2F3, BMI1, JAG1, and CORO1C. miR-34a and miR-128a showed low expression profiles in comparison to a control miR-RNU-44 in both GBM cell lines suggesting tumour suppressor properties. Opposing, miR-21-5p demonstrated greater expression indicating that it could potentially function as an oncomiR. Western blotting revealed expression of all five proteins within the A172 cell line. In silico analysis also suggested that CORO1C is a target of miR-128a and miR-34a. Immunohistochemistry demonstrated that 75% of the GBM cases showed moderate to high expression of CORO1C protein. Greater understanding of the deregulated expression of miR-128a and the upregulation of CORO1C in GBM could potentially lead to the identification of a promising diagnostic biomarker signature for glioblastomas.

Keywords: non-coding RNAs, gene expression, brain tumours, immunohistochemistry

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Authors: Hala M. El-hanbuli, Mohammed F. Darweesh

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The incidence rates of any tumor vary hugely with geographical location. Basal Cell Carcinoma (BCC) is one of the most common skin cancer that has many histopathologic subtypes. Objective: The aim was to study the histopathological features of BCC cases that were received in the Pathology Department, Kasr El-Aini hospital, Cairo University, Egypt during the period from Jan 2004 to Dec 2013 and to evaluate the clinical characters through the patient data available in the request sheets. Methods: Slides and data of BCC cases were collected from the archives of the pathology department, Kasr El-Aini hospital. Revision of all available slides and histological classification of BCC according to WHO (2006) was done. Results: A total number of 310 cases of BCC representing about 65% from the total number of malignant skin tumors examined during the 10-years duration in the department. The age ranged from 8 to 84 years, the mean age was (55.7 ± 15.5). Most of the patients (85%) were above the age of 40 years. There was a slight male predominance (55%). Ulcerated BCC was the most common gross picture (60%), followed by nodular lesion (30%) and finally the ulcerated nodule (10%). Most of the lesions situated in the high-risk sites (77%) where the nose was the most common site (35%) followed by the periocular area (22%), then periauricular (15%) and finally perioral (5%). No lesion was reported outside the head. The tumor size was less than 2 centimeters in 65% of cases, and from 2-5 centimeters in the lesions' greatest dimension in the rest of cases. Histopathological reclassification revealed that the nodular BCC was the most common (68%) followed by the pigmented nodular (18.75%). The histologic high-risk groups represented (7.5%) about half of them (3.75%) being basosquamous carcinoma. The total incidence for multiple BCC and 2nd primary was 12%. Recurrent BCC represented 8%. All of the recurrent lesions of BCC belonged to the histologic high-risk group. Conclusion: Basal Cell Carcinoma is the most common skin cancer in the 10-year survey. Histopathological diagnosis and classification of BCC cases are essential for the determination of the tumor type and its biological behavior.

Keywords: basal cell carcinoma, high risk, histopathological features, statistical analysis

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Authors: Halima Albalushi, Mohadese Boroojerdi, Murtadha Alkhabori

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Introduction: Maintenance of stem cell properties during culture necessitates the recreation of the natural cell niche. Studies reported the promising outcome of mesenchymal stem cells (MSC) properties maintenance after using extracellular matrix such as CELLstart™, which is the recommended coating material for stem cells cultured in serum-free and xeno-free conditions. Laminin-521 is known as a crucial adhesion protein, which is found in natural stem cell niche, and plays an important role in facilitating the maintenance of self-renewal, pluripotency, standard morphology, and karyotype of human pluripotent stem cells (PSCs). The aim of this study is to investigate the effects of Laminin-521 on human umbilical cord-derived mesenchymal stem cells (UC-MSC) characteristics as a step toward clinical application. Methods: Human MSC were isolated from the umbilical cord via the explant method. Umbilical cord-derived-MSC were cultured in serum-free and xeno-free conditions in the presence of Laminin-521 for six passages. Cultured cells were evaluated by morphology and expansion index for each passage. Phenotypic characterization of UC-MSCs cultured on Laminin-521 was evaluated by assessment of cell surface markers. Results: Umbilical cord derived-MSCs formed small colonies and expanded as a homogeneous monolayer when cultured on Laminin-521. Umbilical cord derived-MSCs reached confluence after 4 days in culture. No statistically significant difference was detected in all passages when comparing the expansion index of UC-MSCs cultured on LN-521 and CELLstart™. Phenotypic characterization of UC-MSCs cultured on LN-521 using flow cytometry revealed positive expression of CD73, CD90, CD105 and negative expression of CD34, CD45, CD19, CD14 and HLA-DR.Conclusion: Laminin-521 is comparable to CELLstart™ in supporting UC-MSCs expansion and maintaining their characteristics during culture in xeno-free and serum-free culture conditions.

Keywords: mesenchymal stem cells, culture, laminin-521, xeno-free serum-free

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Authors: Sanjay Kumar Behera, Kanhu Charan Patra

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A GIS-based method has been applied for the determination of soil erosion and sediment yield in a small watershed in Ong River basin, Odisha, India. The method involves spatial disintegration of the catchment into homogenous grid cells to capture the catchment heterogeneity. The gross soil erosion in each cell was calculated using Universal Soil Loss Equation (USLE) by carefully determining its various parameters. The concept of sediment delivery ratio is used to route surface erosion from each of the discretized cells to the catchment outlet. The process of sediment delivery from grid cells to the catchment outlet is represented by the topographical characteristics of the cells. The effect of DEM resolution on sediment yield is analyzed using two different resolutions of DEM. The spatial discretization of the catchment and derivation of the physical parameters related to erosion in the cell are performed through GIS techniques.

Keywords: DEM, GIS, sediment delivery ratio, sediment yield, soil erosion

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Authors: Shima Fasahat

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This research is an experimental research which was done about microbial fuel cells in order to study them for electricity generating and wastewater treatment. These days, it is very important to find new, clean and sustainable ways for energy supplying. Because of this reason there are many researchers around the world who are studying about new and sustainable energies. There are different ways to produce these kind of energies like: solar cells, wind turbines, geothermal energy, fuel cells and many other ways. Fuel cells have different types one of these types is microbial fuel cell. In this research, an MFC was built in order to study how it can be used for electricity generating and wastewater treatment. The microbial fuel cell which was used in this research is a reactor that has two tanks with a catalyst solution. The chemical reaction in microbial fuel cells is a redox reaction. The microbial fuel cell in this research is a two chamber MFC. Anode chamber is an anaerobic one (ABR reactor) and the other chamber is a cathode chamber. Anode chamber consists of stabilized sludge which is the source of microorganisms that do redox reaction. The main microorganisms here are: Propionibacterium and Clostridium. The electrodes of anode chamber are graphite pages. Cathode chamber consists of graphite page electrodes and catalysts like: O₂, KMnO₄ and C₆N₆FeK₄. The membrane which separates the chambers is Nafion117. The reason of choosing this membrane is explained in the complete paper. The main goal of this research is to generate electricity and treating wastewater. It was found that when you use electron receptor compounds like: O_2, MnO₄, C₆N₆FeK₄ the velocity of electron receiving speeds up and in a less time more current will be achieved. It was found that the best compounds for this purpose are compounds which have iron in their chemical formula. It is also important to pay attention to the amount of nutrients which enters to bacteria chamber. By adding extra nutrients in some cases the result will be reverse.  By using ABR the amount of chemical oxidation demand reduces per day till it arrives to a stable amount.

Keywords: anaerobic baffled reactor, bioenergy, electrode, energy efficient, microbial fuel cell, renewable chemicals, sustainable

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Authors: Patricia P. M. C. Lourenco, Sang Yong Kim, Anaisa D. A. De Sena

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K-Pop fandom is a paradoxical dichotomy of two conceptual contexts: the Korean single fandom and the international fandom; both strongly influence K-Pop brand attitude. Collectivist, South Korea’s fans showcase their undivided support to one artist comeback towards earning a triple-crown in domestic music charts. In contrast, individualist international fans collectively ship a plethora of artists and collaborate amongst themselves to the continuous expansion of K-Pop into a mainstream cultural glocalization in international music charts. The distinct idiosyncrasies between the two groups creates a heterogeneous K-Pop brand attitude that is challenging to tackle marketing wise for lack of homogeneity in the sub-cultural K-Pop fandom.

Keywords: K-Pop fandom, single-fandom, multi-fandom, individualism, collectivism, brand attitude, sub-culture

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Authors: Aditi Mittal, Nishit Gupta, Tina Dadu, Anil Handoo

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Introduction: Hematopoietic stem cell transplantation (HSCT) is a curative treatment done for hematologic malignancies and other clinical conditions. Its main objective is to reconstitute the hematopoietic system of the recipient by administering an infusion of donor hematopoietic stem cells. Transplant engraftment is the first sign of bone marrow recovery. The main objective of this study is to assess immature platelet fraction (IPF) and immature reticulocyte fraction (IRF) as early indicators of post-hematopoietic stem cell transplant engraftment. Methods: Patients of all age groups and both genders undergoing both autologous and allogeneic transplants were included in the study. All the CBC samples were run on Mindray CAL-8000 (BC-6800 plus; Shenzhen, China) analyser and assessed for IPF and IRF. Neutrophil engraftment was defined as the first of three consecutive days with an ANC >0.5 x 109/L and platelet engraftment with a count >20 x 109/L. The cut-off values for IRF were calculated as 13.5% with a CV of 5% and for IPF was 19% with a CV of 12%. Results: The study sample comprised 200 patients, of whom 116 had undergone autologous HSCT, and 84 had undergone allogeneic HSCT. We observed that IRF anticipated the neutrophil recovery by an average of 5 days prior to IPF. Though there was no significant variation in IPF and IRF for the prediction of platelet recovery, IRF was preceded by 1 or 2 days to IPF in 25% of cases. Conclusions: Both IPF and IRF can be used as reliable parameters as predictors for post-transplant engraftment; however, IRF seems to be more reliable than IPF as a simple, inexpensive, and widely available tool for predicting marrow recovery several days before engraftment.

Keywords: transplantation, stem cells, reticulocyte, engraftment

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Authors: Sahar Nasr, Lin Li, Edwin Wang

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Bladder cancer (BLCA) is known as the 13th cause of death among cancer patients worldwide, and ~575,000 new BLCA cases are diagnosed each year. Urothelial carcinoma (UC) is the most prevalent subtype among BLCA patients, which can be categorized into muscle-invasive bladder cancer (MIBC) and non-muscle-invasive bladder cancer (NMIBC). Currently, various therapeutic options are available for UC patients, including (1) transurethral resection followed by intravesical instillation of chemotherapeutics or Bacillus Calmette-Guérin for NMIBC patients, (2) neoadjuvant platinum-based chemotherapy (NAC) plus radical cystectomy is the standard of care for localized MIBC patients, and (3) systematic chemotherapy for metastatic UC. However, conventional treatments may lead to several challenges for treating patients. As an illustration, some patients may suffer from recurrence of the disease after the first line of treatment. Recently, immune checkpoint therapy (ICT) has been introduced as an alternative treatment strategy for the first or second line of treatment in advanced or metastatic BLCA patients. Although ICT showed lucrative results for a fraction of BLCA patients, ~80% of patients were not responsive to it. Therefore, novel treatment methods are required to augment the ICI response rate within BLCA patients. It has been shown that the infiltration of T-cells into the tumor microenvironment (TME) is positively correlated with the response to ICT within cancerous patients. Therefore, the goal of this study is to enhance the infiltration of cytotoxic T-cells into TME through the identification of target genes within the tumor that are responsible for the non-T-cell inflamed TME and their inhibition. BLCA bulk RNA-sequencing data from The Cancer Genome Atlas (TCGA) and immune score for TCGA samples were used to determine the Pearson correlation score between the expression of different genes and immune score for each sample. The genes with strong negative correlations were selected (r < -0.2). Thereafter, the correlation between the expression of each gene and survival in BLCA patients was calculated using the TCGA data and Cox regression method. The genes that are common in both selected gene lists were chosen for further analysis. Afterward, BLCA bulk and single-cell RNA-sequencing data were ranked based on the expression of each selected gene and the top and bottom 25% samples were used for pathway enrichment analysis. If the pathways related to the T-cell infiltration (e.g., antigen presentation, interferon, or chemokine pathways) were enriched within the low-expression group, the gene was included for downstream analysis. Finally, the selected genes will be used to calculate the correlation between their expression and the infiltration rate of the activated CD+8 T-cells, natural killer cells and the activated dendric cells. A list of potential target genes has been identified and ranked based on the above-mentioned analysis and criteria. SUN-1 got the highest score within the gene list and other identified genes in the literature as benchmarks. In conclusion, inhibition of SUN1 may increase the tumor-infiltrating lymphocytes and the efficacy of ICI in BLCA patients. BLCA tumor cells with and without SUN-1 CRISPR/Cas9 knockout will be injected into the syngeneic mouse model to validate the predicted SUN-1 effect on increasing tumor-infiltrating lymphocytes.

Keywords: data analysis, gene expression analysis, gene identification, immunoinformatic, functional genomics, transcriptomics

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Authors: Catrin Morgan, Atholl Johnston

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Creatine supplementation should theoretically increase total muscle creatine and so enhance the generation of intramuscular phosphocreatine and subsequent ATP formation. The use of creatine as a potential ergogenic aid in sport has been an area of significant scientific research for a number of years. However the effect of creatine supplementation and swimming performance is a relatively new area of research and is the subject of this review. In swimming creatine supplementation could help maintain maximal power output, aid recovery and increase lean body mass. After investigating the underlying theory and science behind creatine supplementation, a literature review was conducted to identify the best evidence looking at the effect of creatine supplementation on swimming performance. The search identified 27 potential studies, and of these 17 were selected for review. The studies were then categorised into single sprint performance, which involves swimming a short distance race, or repeated interval performance, which involves swimming a series of sprints with intervals of rest between them. None of the studies on the effect of creatine controlled for the multiple confounding factors associated with measurement of swimming performance. The sample size in the studies was limited and this reduced the reliability of the studies and introduced the possibility of bias. The studies reviewed provided insufficient evidence to determine if creatine supplementation is beneficial to swimming performance. However, what data there was supported the use of creatine supplementation in repeated interval swimming rather than in single sprint swimming. From a review of the studies, it was calculated on average, there was a 1.37% increase in swimming performance with the use of creatine for repeated intervals and a 0.86% increase in performance for single sprint. While this may seem minor, it should be remembered that swimming races are often won by much smaller margins. In the 2012 London Olympics the Men’s 100 metres freestyle race was won by a margin of only 0.01 of a second. Therefore any potential benefit could make a dramatic difference to the final outcome of the race. Overall more research is warranted before the benefits of creatine supplementation in swimming performance can be further clarified.

Keywords: creatine supplementation, repeated interval, single sprint, swimming performance

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Authors: Vishnu Varthan Vaithiyalingam Jagannathan, Lakshmi Karunanidhi Santhanalakshmi, Srividya Ammayappan Rajam

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Formononetin is the O-Methoxy Flavonol that has many pharmacological activities, which belongs to the flavonoid family. In the current study, activity of this molecule was evaluated in lung cancer cell lines. In general, flavonoids possess certain anticancer mechanism. Being a flavonoid subfamily, this molecule was subjected to evaluate cytotoxicity assay by MTT ((3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide)) stain, mode of cell death assay stained by acridine orange and ethidium bromide and Evaluation of Apoptosis pathway (extrinsic or intrinsic) by Caspase 3/7 stain and Rhodamine-123 stain. From the results, we could able to confirm that the investigatory molecule formononetin has anticancer activity and in future, the study will propose to evaluate the formononetin action against genetic changes occurs during lung cancer progression.

Keywords: Caspase 3/7, formononetin, lung cancer, Rhodamine-123

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Authors: Kang-Hyuk Lee, Wei Yan, Sang-Im Yoo

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Recently, Ca₁₋ₓLaₓFe₁₂O₁₉ (Ca-La M-type) hexaferrites have been reported to possess higher crystalline anisotropy compared with SrFe₁₂O₁₉ (Sr M-type) hexaferrite without reducing its saturation magnetization (Ms), resulting in higher coercivity (Hc). While iron deficiency is known to be helpful for the growth and the formation of NiZn spinel ferrites, the effect of iron deficiency in Ca-La M-type hexaferrites has never been reported yet. In this study, therefore, we tried to investigate the effect of iron deficiency on the magnetic properties of Ca₀.₅La₀.₅Fe₁₂₋yO₁₉₋δ hexaferrites prepared by solid state reaction. As-calcined powder was pressed into pellets and sintered at 1275~1325℃ for 4 h in air. Samples were characterized by powder X-ray diffraction (XRD), vibrating sample magnetometer (VSM), and scanning electron microscope (SEM). Powder XRD analyses revealed that Ca₀.₅La₀.₅Fe₁₂₋yO₁₉₋δ (0.75 ≦ y ≦ 2.15) ferrites calcined at 1250-1300℃ for 12 h in air were composed of single phase without the second phases. With increasing the iron deficiency, y, the lattice parameters a, c and unite cell volumes were decreased first up to y=10.25 and then increased again. The highest Ms value of 77.5 emu/g was obtainable from the sample of Ca₀.₅La₀.₅Fe₁₂₋yO₁₉₋δ sintered at 1300℃ for 4 h in air. Detailed microstructures and magnetic properties of Ca-La M-type hexagonal ferrites will be presented for a discussion

Keywords: Ca-La M-type hexaferrite, magnetic properties, iron deficiency, hexaferrite

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Authors: Abdul Matin, Muhammad Ajmal, Uzma Yunus, Noaman-ul Haq, Hafiz M. Shohaib, Ambreen G. Muazzam

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Carbon nanoparticles (CNPs) have unique properties that are useful for the diagnosis and treatment of cancer due to their precise properties like small size (ideal for delivery within the body) stability in solvent and tunable surface chemistry for targeted delivery. Here, highly fluorescent, monodispersed and water-soluble CNPs were synthesized directly from a suitable carbohydrate source (glucose and sucrose) by one-step acid assisted ultrasonic treatment at 35 KHz for 4 hours. This method is green, simple, rapid and economical and can be used for large scale production and applications. The average particle sizes of CNPs are less than 10nm and they emit bright and colorful green-blue fluorescence under the irradiation of UV-light at 365nm. The CNPs were characterized by scanning electron microscopy, fluorescent spectrophotometry, Fourier transform infrared spectrophotometry, ultraviolet-visible spectrophotometry and TGA analysis. Fluorescent CNPs were used as fluorescent probe and nano-carriers for anticancer drug. Functionalized CNPs (with ethylene diamine) were attached with anticancer drug-Methotrexate. In vitro bioactivity and biocompatibility of CNPs-drug conjugates was evaluated by LDH assay and Sulforhodamine B assay using human lung carcinoma cell lines (H157). Our results reveled that CNPs showed biocompatibility and CNPs-anticancer drug conjugates have shown potent cytotoxic effects and high antitumor activities in lung cancer cell lines. CNPs are proved to be excellent substitute for conventional drug delivery cargo systems and anticancer therapeutics in vitro. Our future studies will be more focused on using the same nanoparticles in vivo model system.

Keywords: carbon nanoparticles, carbon nanoparticles-methotrexate conjugates, human lung carcinoma cell lines, lactate dehydrogenase, methotrexate

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Authors: Rayan B. M. Ameen, Ian P. Jones, Yu Lung Chiu

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Single alpha-beta colony micro-pillars have been manufactured from a polycrystalline commercial Ti-6Al-4V sample using Focused Ion Beam (FIB). Each pillar contained two alpha lamellae separated by a thin fillet of beta phase. A nano-indenter was then used to conduct uniaxial micro-compression tests on Ti alloy single crystals, using a diamond flat tip as a compression platen. By controlling the crystal orientation along the micro-pillar using Electron back scattering diffraction (EBSD) different slip systems have been selectively activated. The advantage of the micro-compression method over conventional mechanical testing techniques is the ability to localize a single crystal volume which is characterizable after deformation. By matching the stress-strain relations resulting from micro-compression experiments to TEM (Transmission Electron Microscopy) studies of slip transmission mechanisms through the α-β interfaces, some proper constitutive material parameters such as the role of these interfaces in determining yield, strain-hardening behaviour, initial dislocation density and the critical resolved shear stress are suggested.

Keywords: α/β-Ti alloy, focused ion beam, micro-mechanical test, nano-indentation, transmission electron diffraction, plastic flow

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Authors: Chevonne Reynolds, Res Altwegg, Andrew Balmford, Claire N. Spottiswoode

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Modern agriculture has revolutionised the planet’s capacity to support humans, yet has simultaneously had a greater negative impact on biodiversity than any other human activity. Balancing the demand for food with the conservation of biodiversity is one of the most pressing issues of our time. Biodiversity-friendly farming (‘land sharing’), or alternatively, separation of conservation and production activities (‘land sparing’), are proposed as two strategies for mediating the trade-off between agriculture and biodiversity. However, there is much debate regarding the efficacy of each strategy, as this trade-off has typically been addressed by short term studies at fine spatial scales. These studies ignore processes that are relevant to biodiversity at larger scales, such as meta-population dynamics and landscape connectivity. Therefore, to better understand species response to agricultural land-use and provide evidence to underpin the planning of better production landscapes, we need to determine the merits of each strategy at larger scales. In South Africa, a remarkable citizen science project - the South African Bird Atlas Project 2 (SABAP2) – collates an extensive dataset describing the occurrence of birds at a 5-min by 5-min grid cell resolution. We use these data, along with fine-resolution data on agricultural land-use, to determine which strategy optimises the agriculture-biodiversity trade-off in a southern African context, and at a spatial scale never considered before. To empirically test this trade-off, we model bird species population density, derived for each 5-min grid cell by Royle-Nicols single-species occupancy modelling, against both the amount and configuration of different types of agricultural production in the same 5-min grid cell. In using both production amount and configuration, we can show not only how species population densities react to changes in yield, but also describe the production landscape patterns most conducive to conservation. Furthermore, the extent of both the SABAP2 and land-cover datasets allows us to test this trade-off across multiple regions to determine if bird populations respond in a consistent way and whether results can be extrapolated to other landscapes. We tested the land sparing/sharing trade-off for 281 bird species across three different biomes in South Africa. Overall, a higher proportion of species are classified as losers, and would benefit from land sparing. However, this proportion of loser-sparers is not consistent and varies across biomes and the different types of agricultural production. This is most likely because of differences in the intensity of agricultural land-use and the interactions between the differing types of natural vegetation and agriculture. Interestingly, we observe a higher number of species that benefit from agriculture than anticipated, suggesting that agriculture is a legitimate resource for certain bird species. Our results support those seen at smaller scales and across vastly different agricultural systems, that land sparing benefits the most species. However, our analysis suggests that land sparing needs to be implemented at spatial scales much larger than previously considered. Species persistence in agricultural landscapes will require the conservation of large tracts of land, and is an important consideration in developing countries, which are undergoing rapid agricultural development.

Keywords: agriculture, birds, land sharing, land sparing

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Authors: Peter Antwi Buah, Yingbin Zhang, Jianxian He, Chenlin Xiang, Delali Atsu Y. Bakah

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Near-fault ground motions with velocity pulses are considered to cause significant damage to structures or slopes compared to ordinary ground motions without velocity pulses. The double pulsed pulse-like ground motion is as well known to be stronger than the single pulse. This study has numerically justified this perspective by studying the dynamic response of a homogeneous rock slope subjected to four pulse-like and two non-pulse-like ground motions using the Fast Lagrangian Analysis of Continua in 3 Dimensions (FLAC3D) software. Two of the pulse-like ground motions just have a single pulse. The results show that near-fault ground motions with velocity pulses can cause a higher dynamic response than regular ground motions. The amplification of the peak ground acceleration (PGA) in horizontal direction increases with the increase of the slope elevation. The seismic response of the slope under double pulse ground motion is stronger than that of the single pulse ground motion. The PGV amplification factor under the effect of the non-pulse-like records is also smaller than those under the pulse-like records. The velocity pulse strengthens the earthquake damage to the slope, which results in producing a more strong dynamic response.

Keywords: velocity pulses, dynamic response, PGV magnification effect, elevation effect, double pulse

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Authors: Rumeysa Gunduz, Dirk Koester, Thomas Schack

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Previous behavioral studies have shown that working memory (WM) and manual actions share limited capacity cognitive resources, which in turn results in execution costs of manual actions in WM. However, to the best of our knowledge, there is no study investigating the neurophysiology of execution costs. The current study aims to fill this research gap investigating the neurophysiology of execution costs of grasping in WM phases (encoding, maintenance, retrieval) considering verbal and visuospatial domains of WM. A WM-grasping dual task paradigm was implemented to examine execution costs. Baseline single task required performing verbal or visuospatial version of a WM task. Dual task required performing the WM task embedded in a high precision grasp to place task. 30 participants were tested in a 2 (single vs. dual task) x 2 (visuo-spatial vs. verbal WM) within subject design. Event related potentials (ERPs) were extracted for each WM phase separately in the single and dual tasks. Memory performance for visuospatial WM, but not for verbal WM, was significantly lower in the dual task compared to the single task. Encoding related ERPs in the single task revealed different ERPs of verbal WM and visuospatial WM at bilateral anterior sites and right posterior site. In the dual task, bilateral anterior difference disappeared due to bilaterally increased anterior negativities for visuospatial WM. Maintenance related ERPs in the dual task revealed different ERPs of verbal WM and visuospatial WM at bilateral posterior sites. There was also anterior negativity for visuospatial WM. Retrieval related ERPs in the single task revealed different ERPs of verbal WM and visuospatial WM at bilateral posterior sites. In the dual task, there was no difference between verbal WM and visuospatial WM. Behavioral and ERP findings suggest that execution of grasping shares cognitive resources only with visuospatial WM, which in turn results in domain specific execution costs. Moreover, ERP findings suggest unique patterns of costs in each WM phase, which supports the idea that each WM phase reflects a separate cognitive process. This study not only contributes to the understanding of cognitive principles of manual action control, but also contributes to the understanding of WM as an entity consisting of separate modalities and cognitive processes.

Keywords: dual task, grasping execution, neurophysiology, working memory domains, working memory phases

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Authors: Vinaya Kambappa, Chinnadurai Mani, Komaraiah Palle

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Non-small cell-lung cancer (NSCLC) cells have increased expression of EGFR, which makes them a potential target for cancer therapy. Based on molecular docking and previous reports, we designed and synthesized quinazoline derivatives as potent EGFR inhibitors. Among the derivatives, three compounds showed good antiproliferative activity against A-549 and H-1299 cells. Furthermore, these compounds inhibited EGFR signaling exhibiting diminishing p-EGFR and its downstream proteins like p-Akt, p-Erk1/2, and p-mTOR; however, it did not alter the levels of EGFR, Akt, Erk1/2 and mTOR proteins. Flow cytometric analysis indicated the accumulation of cells at G1 phase suggesting induction of apoptosis, which was further confirmed by annexin V/propidium iodide staining. Our study suggested that quinazoline scaffold can be developed as novel EGFR kinase inhibitors for cancer therapy.

Keywords: apoptosis, non-small cell-lung cancer cells, EGFR, quinazoline

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Authors: Arnab Sarkar, Michael Huang, Chuang Ren, Jun Li

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The increasing importance of computing in modern society has brought substantial growth in the demand for more computational power. In some problem domains such as scientific simulations, available computational power still sets a limit on what can be practically explored in computation. For many types of code, there is non-uniformity in the utility of computation. That is not every piece of computation contributes equally to the quality of the result. If this non-uniformity is understood well and exploited effectively, we can much more effectively utilize available computing power. In this paper, we discuss a case study of exploring such non-uniformity in a particle-in-cell simulation platform. We find both the existence of significant non-uniformity and that it is generally straightforward to exploit it. We show the potential of order-of-magnitude effective performance gain while keeping the comparable quality of output. We also discuss some challenges in both the practical application of the idea and evaluation of its impact.

Keywords: approximate computing, landau damping, non uniform utility computing, particle-in-cell

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Authors: Natalie Riley, Anita Quigley, Robert M. I. Kapsa, George W. Greene

Abstract:

As the fields of medicine and bionics grow rapidly in technological advancement, the future and success of it depends on the ability to effectively interface between the artificial and the biological worlds. The biggest obstacle when it comes to implantable, electronic medical devices, is maintaining a ‘clean’, low noise electrical connection that allows for efficient sharing of electrical information between the artificial and biological systems. Implant fouling occurs with the adhesion and accumulation of proteins and various cell types as a result of the immune response to protect itself from the foreign object, essentially forming an electrical insulation barrier that often leads to implant failure over time. Lubricin (LUB) functions as a major boundary lubricant in articular joints, a unique glycoprotein with impressive anti-adhesive properties that self-assembles to virtually any substrate to form a highly ordered, ‘telechelic’ polymer brush. LUB does not passivate electroactive surfaces which makes it ideal, along with its innate biocompatibility, as a coating for implantable bionic electrodes. It is the aim of the study to investigate LUB’s anti-fouling properties and its potential as a safe, bioinspired material for coating applications to enhance the performance and longevity of implantable medical devices as well as reducing the frequency of implant replacement surgeries. Native, bovine-derived LUB (N-LUB) and recombinant LUB (R-LUB) were applied to gold-coated mylar surfaces. Fibroblast, chondrocyte and neural cell types were cultured and grown on the coatings under both passive and electrically stimulated conditions to test the stability and anti-adhesive property of the LUB coating in the presence of an electric field. Lactate dehydrogenase (LDH) assays were conducted as a directly proportional cell population count on each surface along with immunofluorescent microscopy to visualize cells. One-way analysis of variance (ANOVA) with post-hoc Tukey’s test was used to test for statistical significance. Under both passive and electrically stimulated conditions, LUB significantly reduced cell attachment compared to bare gold. Comparing the two coating types, R-LUB reduced cell attachment significantly compared to its native counterpart. Immunofluorescent micrographs visually confirmed LUB’s antiadhesive property, R-LUB consistently demonstrating significantly less attached cells for both fibroblasts and chondrocytes. Preliminary results investigating neural cells have so far demonstrated that R-LUB has little effect on reducing neural cell attachment; the study is ongoing. Recombinant LUB coatings demonstrated impressive anti-adhesive properties, reducing cell attachment in fibroblasts and chondrocytes. These findings and the availability of recombinant LUB brings into question the results of previous experiments conducted using native-derived LUB, its potential not adequately represented nor realized due to unknown factors and impurities that warrant further study. R-LUB is stable and maintains its anti-fouling property under electrical stimulation, making it suitable for electroactive surfaces.

Keywords: anti-fouling, bioinspired, cell attachment, lubricin

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Authors: Aberdam Edith, Sangari Linda, Petit Isabelle, Aberdam Daniel

Abstract:

Background and Rationale: Transparent avascularised cornea is essential for normal vision and depends on limbal stem cells (LSC) that reside between the cornea and the conjunctiva. Ocular burns or injuries may destroy the limbus, causing limbal stem cell deficiency (LSCD). The cornea becomes vascularised by invaded conjunctival cells, the stroma is scarring, resulting in corneal opacity and loss of vision. Grafted autologous limbus or cultivated autologous LCS can restore the vision, unless the two eyes are affected. Alternative cellular sources have been tested in the last decades, including oral mucosa or hair follicle epithelial cells. However, only partial success has been achieved by the use of these cells since they were not able to uniformly commit into corneal epithelial cells. Human pluripotent stem cells (iPSC) display both unlimited growth capacity and ability to differentiate into any cell type. Our goal was to design a standardized and reproducible protocol to produce transplantable autologous LSC from patients through cell reprogramming technology. Methodology: First, keratinocyte primary culture was established from a small number of plucked hair follicles of healthy donors. The resulting epithelial cells were reprogrammed into induced pluripotent stem cells (iPSCs) and further differentiate into corneal epithelial cells (CEC), according to a robust protocol that recapitulates the main step of corneal embryonic development. qRT-PCR analysis and immunofluorescent staining during the course of differentiation confirm the expression of stage specific markers of corneal embryonic lineage. First appear ectodermal progenitor-specific cytokeratins K8/K18, followed at day 7 by limbal-specific PAX6, TP63 and cytokeratins K5/K14. At day 15, K3/K12+-corneal cells are present. To amplify the iPSC-derived LSC (named COiPSC), intact small epithelial colonies were detached and cultivated in limbal cell-specific medium. In that culture conditions, the COiPSC can be frozen and thaw at any passage, while retaining their corneal characteristics for at least eight passages. To evaluate the potential of COiPSC as an alternative ocular toxicity model, COiPSC were treated at passage P0 to P4 with increasing amounts of SDS and Benzalkonium. Cell proliferation and apoptosis of treated cells was compared to LSC and the SV40-immortalized human corneal epithelial cell line (HCE) routinely used by cosmetological industrials. Of note, HCE are more resistant to toxicity than LSC. At P0, COiPSC were systematically more resistant to chemical toxicity than LSC and even to HCE. Remarkably, this behavior changed with passage since COiPSC at P2 became identical to LSC and thus closer to physiology than HCE. Comparative transcriptome analysis confirmed that COiPSC from P2 are similar to a mixture of LSC and CEC. Finally, by organotypic reconstitution assay, we demonstrated the ability of COiPSC to produce a 3D corneal epithelium on a stromal equivalent made of keratocytes. Conclusion: COiPSC could become valuable for two main applications: (1) an alternative robust tool to perform, in a reproducible and physiological manner, toxicity assays for cosmetic products and pharmacological tests of drugs. (2). COiPSC could become an alternative autologous source for cornea transplantation for LSCD.

Keywords: Limbal stem cell deficiency, iPSC, cornea, limbal stem cells

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Authors: F. Amirarfaei, K. Khorasani

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In this paper, reconfigurable consensus achievement of a team of agents with marginally stable linear dynamics and single input channel has been considered. The control algorithm is based on a first order linear protocol. After occurrence of a LOE fault in one of the actuators, using the imperfect information of the effectiveness of the actuators from fault detection and identification module, the control gain is redesigned in a way to still reach consensus. The idea is based on the modeling of change in effectiveness as change of Laplacian matrix. Then as special cases of this class of systems, a team of single integrators as well as double integrators are considered and their behavior subject to a LOE fault is considered. The well-known relative measurements consensus protocol is applied to a leaderless team of single integrator as well as double integrator systems, and Gersgorin disk theorem is employed to determine whether fault occurrence has an effect on system stability and team consensus achievement or not. The analyses show that loss of effectiveness fault in actuator(s) of integrator systems affects neither system stability nor consensus achievement.

Keywords: multi-agent system, actuator fault, stability analysis, consensus achievement

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Authors: Doreen Duri, Danai Zhou, Babil Stray-Pedersen, Collet Dandara

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Given the high prevalence of HIV/AIDS in sub-Saharan Africa, and the elusive search for a cure, understanding the pharmacogenetics of currently used drugs is critical in populations from the most affected regions. Compared to Asian and Caucasian populations, African population groups are more genetically diverse, making it difficult to extrapolate findings from one ethnic group to another. This study aimed to investigate the role of genetic variation in CYP2B6 (c.516G>T and c.983T>C) single nucleotide polymorphisms on plasma nevirapine levels among HIV-infected adult Zimbabwean patients. Using a cross-sectional study, patients on nevirapine-containing HAART, having reached steady state (more than six weeks on treatment) were recruited to participate. Blood samples were collected after patients provided consent and samples were used to extract DNA for genetic analysis or to measure plasma nevirapine levels. Genetic analysis was carried out using PCR and RFLP or Snapshot for the two single nucleotide polymorphisms; CYP2B6 c.516G>T and c.983T>C, while LC-MS/MS was used in analyzing nevirapine concentration. CYP2B6 c.516G>T and c.983T>C significantly predicted plasma nevirapine concentration with the c.516T and c.983T being associated with elevated plasma nevirapine concentrations. Comparisons of the variant allele frequencies observed in this group to those reported in some African, Caucasian and Asian populations showed significant differences. We conclude that pharmacogenetics of nevirapine can be creatively used to determine patients who are likely to develop nevirapine-associated side effects as well as too low plasma concentrations for viral suppression.

Keywords: allele frequencies, genetically diverse, nevirapine, single nucleotide polymorphism

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Authors: Roxane A. Legaie, Kjiana E. Schwab, Caroline E. Gargett

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Studies looking at the changes in gene expression from RNAseq data often make use of linear models. It is also common practice to focus on a subset of data for a comparison of interest, leaving aside the samples not involved in this particular comparison. This work shows the importance of including all observations in the modeling process to better estimate variance parameters, even when the samples included are not directly used in the comparison under test. The human endometrium is a dynamic tissue, which undergoes cycles of growth and regression with each menstrual cycle. The mesenchymal stem cells (MSCs) present in the endometrium are likely responsible for this remarkable regenerative capacity. However recent studies suggest that MSCs also plays a role in the pathogenesis of endometriosis, one of the most common medical conditions affecting the lower abdomen in women in which the endometrial tissue grows outside the womb. In this study we compared gene expression profiles between MSCs and non-stem cell counterparts (‘non-MSC’) obtained from women with (‘E’) or without (‘noE’) endometriosis from RNAseq. Raw read counts were used for differential expression analysis using a linear model with the limma-voom R package, including either all samples in the study or only the samples belonging to the subset of interest (e.g. for the comparison ‘E vs noE in MSC cells’, including only MSC samples from E and noE patients but not the non-MSC ones). Using the full dataset we identified about 100 differentially expressed (DE) genes between E and noE samples in MSC samples (adj.p-val < 0.05 and |logFC|>1) while only 9 DE genes were identified when using only the subset of data (MSC samples only). Important genes known to be involved in endometriosis such as KLF9 and RND3 were missed in the latter case. When looking at the MSC vs non-MSC cells comparison, the linear model including all samples identified 260 genes for noE samples (including the stem cell marker SUSD2) while the subset analysis did not identify any DE genes. When looking at E samples, 12 genes were identified with the first approach and only 1 with the subset approach. Although the stem cell marker RGS5 was found in both cases, the subset test missed important genes involved in stem cell differentiation such as NOTCH3 and other potentially related genes to be used for further investigation and pathway analysis.

Keywords: differential expression, endometriosis, linear model, RNAseq

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Authors: Osamede Asowata, Christo Pienaar, Johan Bekker

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Photovoltaic (PV) modules may become a trend for future PV systems because of their greater flexibility in distributed system expansion, easier installation due to their nature, and higher system-level energy harnessing capabilities under shaded or PV manufacturing mismatch conditions. This is as compared to the single or multi-string inverters. Novel residential scale PV arrays are commonly connected to the grid by a single DC–AC inverter connected to a series, parallel or series-parallel string of PV panels, or many small DC–AC inverters which connect one or two panels directly to the AC grid. With an increasing worldwide interest in sustainable energy production and use, there is renewed focus on the power electronic converter interface for DC energy sources. Three specific examples of such DC energy sources that will have a role in distributed generation and sustainable energy systems are the photovoltaic (PV) panel, the fuel cell stack, and batteries of various chemistries. A high-efficiency inverter using Metal Oxide Semiconductor Field-Effect Transistors (MOSFETs) for all active switches is presented for a non-isolated photovoltaic and AC-module applications. The proposed configuration features a high efficiency over a wide load range, low ground leakage current and low-output AC-current distortion with no need for split capacitors. The detailed power stage operating principles, pulse width modulation scheme, multilevel bootstrap power supply, and integrated gate drivers for the proposed inverter is described. Experimental results of a hardware prototype, show that not only are MOSFET efficient in the system, it also shows that the ground leakage current issues are alleviated in the proposed inverter and also a 98 % maximum associated driver circuit is achieved. This, in turn, provides the need for a possible photovoltaic panel switching technique. This will help to reduce the effect of cloud movements as well as improve the overall efficiency of the system.

Keywords: grid connected photovoltaic (PV), Matlab efficiency simulation, maximum power point tracking (MPPT), module integrated converters (MICs), multilevel converter, series connected converter

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Authors: Rupali Mane

Abstract:

Cadmium sulfide (Cds) thin films were chemically deposited at room temperature, from aqueous ammonia solution using CdCl₂ (Cadmium chloride) as a Cd²⁺ and CS(NH₂)₂ (Thiourea) as S² ion sources. ‘as-deposited’ films were uniform, well adherent to the glass substrate, secularly reflective and yellowish in color. The ‘as-deposited ’Cds layers grew with nano-crystalline in nature and exhibit cubic structure, with blue-shift in optical band gap. The films were annealed in air atmosphere for two hours at different temperatures and further characterized for compositional, structural, morphological and optical properties. The XRD and SEM studies clearly revealed the systematic changes in morphological and structural form of Cds films with an improvement in the crystal quality. The annealed films showed ‘red-shift’ in the optical spectra after thermal treatment. The Thiourea modified CdS film could be good to provide solar cell application.

Keywords: cadmium sulfide, thin films, nano-crystalline, XRD

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

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

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

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Authors: Rachida Houssa, Hassan Rhinane, Fadoumo Ali Malouw, Amina Oulmaalem

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Since the early 70s, the Moroccan Atlantic sea was subjected to the pressure of the bottom trawling, one of the most destructive techniques seabed that cause havoc on fishing catch, nonselective, and responsible for more than half of all releases of fish around the world. The present paper aims to map and assess the impact of the activity of the bottom trawling of the Moroccan Atlantic coast. For this purpose, a dataset of thirty years, between 1962 and 1999, from foreign fishing vessels using bottom trawling, has been used and integrated in a GIS. To estimate the extent and the importance of the geographical distribution of the trawling effort, the Moroccan Atlantic area was divided into a grid of cells of 25 km2 (5x5 km). This grid was joined to the effort trawling data, creating a new entity with a table containing spatial overlay grid with the polygon of swept surfaces. This mapping model allowed to quantify the used fishing effort versus time and to generate the trace indicative of trawling efforts on the seabed. Indeed, for a given year, a grid cell may have a swept area equal to 0 (never been touched by the trawl) or 25 km2 (the trawled area is similar to the cell size) or may be 100 km2 indicating that for this year, the scanned surface is four times the cell area. The results show that the total cumulative sum of trawled area is approximately 28,738,326 km2, scattered throughout the Atlantic coast. 95% of the overall trawling effort is located in the southern zone, between 29°N and 20°30'N. Nearly 5% of the trawling effort is located in the northern coastal region, north of 33°N. The center area between 33°N and 29°N is the least swept by Russian commercial vessels because in this region the majority of the area is rocky, and non trawlable.

Keywords: GIS, Moroccan Atlantic Ocean, seabed, trawling

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Authors: Mahya Naghipoor

Abstract:

Purpose: Non-small cell lung cancer (NSCLC) is the most common lung cancer type. Epidermal growth factor receptor (EGFR) mutation is the main reason which causes NSCLC. Computed tomography (CT) is used for diagnosis and prognosis of lung cancers because of low price and little invasion. Semantic analyses of qualitative CT features are based on visual evaluation by radiologist. However, the naked eye ability may not assess all image features. On the other hand, radiomics provides the opportunity of quantitative analyses for CT images features. The aim of this review study was comparing accuracy of semantic and radiomics features in prognosis of EGFR mutation in NSCLC. Methods: For this purpose, the keywords including: non-small cell lung cancer, epidermal growth factor receptor mutation, semantic, radiomics, feature, receiver operating characteristics curve (ROC) and area under curve (AUC) were searched in PubMed and Google Scholar. Totally 29 papers were reviewed and the AUC of ROC analyses for semantic and radiomics features were compared. Results: The results showed that the reported AUC amounts for semantic features (ground glass opacity, shape, margins, lesion density and presence or absence of air bronchogram, emphysema and pleural effusion) were %41-%79. For radiomics features (kurtosis, skewness, entropy, texture, standard deviation (SD) and wavelet) the AUC values were found %50-%86. Conclusions: In conclusion, the accuracy of radiomics analysis is a little higher than semantic in prognosis of EGFR mutation in NSCLC.

Keywords: lung cancer, radiomics, computer tomography, mutation

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Authors: Saskya E. Carrera P., Ben Hankamer, Melanie Oey

Abstract:

The green alga C. reinhardtii provides a platform for the cheap, scalable, and safe production of complex proteins. Despite gene expression in photosynthetic organisms being tightly regulated by light, most expression studies have analysed chloroplast recombinant protein production under constant light. Here the influence of illumination time and intensity on GFP and a GFP-PlyGBS (bacterial-lysin) fusion protein expression was investigated. The expression of both proteins was strongly influenced by the light regime (6-24 hr illumination per day), the light intensity (0-450 E m⁻²s⁻¹) and growth condition (photoautotrophic, mixotrophic and heterotrophic). Heterotrophic conditions resulted in relatively low recombinant protein yields per unit volume, despite high protein yields per cell, due to low growth rates. Mixotrophic conditions exhibited the highest yields at 6 hrs illumination at 200µE m⁻²s⁻¹ and under continuous low light illumination (13-16 mg L⁻¹ GFP and 1.2-1.6 mg L⁻¹ GFP-PlyGBS), as these conditions supported good cell growth and cellular protein yields. A ~23-fold increase in protein accumulation per cell and ~9-fold increase L⁻¹ culture was observed compared to standard constant 24 hr illumination for GFP-PlyGBS. The highest yields under photoautotrophic conditions were obtained under 9 hrs illumination (6 mg L⁻¹ GFP and 2.1 mg L⁻¹ GFP-PlyGBS). This represents a ~4-fold increase in cellular protein accumulation for GFP-PlyGBS. On a volumetric basis the highest yield was at 15 hrs illumination (~2-fold increase L⁻¹ over the constant light for GFP-PlyGBS). Optimising illumination conditions to balance growth and protein expression can thus significantly enhance overall recombinant protein production in C. reinhardtii cultures.

Keywords: chlamydomonas reinhardtii, light, mixotrophic, recombinant protein

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