Search results for: breast cancer stem cells (bCSCs)

Authors: Bing–Jing Zhao, Chang-Kui Liu, Hong Wang, Min Hu

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Electron beam rapid manufacturing (EBRM) or Selective laser melting is an additive manufacturing process that uses 3D CAD data as a digital information source and energy in the form of a high-power laser beam or electron beam to create three-dimensional metal parts by fusing fine metallic powders together.Object:The present study was conducted to evaluate the mechanical properties ,the phase transformation,the corrosivity and the biocompatibility of Ti-6Al-4V by EBRM,SLM and forging technique.Method: Ti-6Al-4V alloy standard test pieces were manufactured by EBRM, SLM and forging technique according to AMS4999,GB/T228 and ISO 10993.The mechanical properties were analyzed by universal test machine. The phase transformation was analyzed by X-ray diffraction and scanning electron microscopy. The corrosivity was analyzed by electrochemical method. The biocompatibility was analyzed by co-culturing with mesenchymal stem cell and analyzed by scanning electron microscopy (SEM) and alkaline phosphatase assay (ALP) to evaluate cell adhesion and differentiation, respectively. Results: The mechanical properties, the phase transformation, the corrosivity and the biocompatibility of Ti-6Al-4V by EBRM、SLM were similar to forging and meet the mechanical property requirements of AMS4999 standard. a­phase microstructure for the EBM production contrast to the a’­phase microstructure of the SLM product. Mesenchymal stem cell adhesion and differentiation were well. Conclusion: The property of the Ti-6Al-4V alloy manufactured by EBRM and SLM technique can meet the medical standard from this study. But some further study should be proceeded in order to applying well in clinical practice.

Keywords: 3D printing, Electron Beam Rapid Manufacturing (EBRM), Selective Laser Melting (SLM), Computer Aided Design (CAD)

Procedia PDF Downloads 444

Authors: Samira Khalaji, , Fenneke Klein Jan, Kay-E. Gottschalk, Eugenia Makrantonaki, Karin Scharffetter-Kochanek

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Biological aging is a multi-dimensional process that takes place over a whole range of scales from the nanoscopic alterations within individual cells, over transformations in tissues and organs and to changes of the whole organism. On the single cell level, aging involves mutation of genes, differences in gene expression levels as well as altered posttranslational modifications of proteins. A variety of proteins is affected, including proteins of the cell cytoskeleton and migration machinery. Previous work quantified the expression of cytoskeleton proteins on the gene and protein levels in senescent and young fibroblasts. Their results show that senescent skin fibroblasts have an upregulated expression of the intermediate filament (IF) protein vimentin in contrast to actin and tubulin, which are downregulated. IFs play an important role in providing mechanical stability of cells. However, the mechanical properties of IFs depending on cellular senescence or age of the donor has not been studied so far. Hence, we employed passive microrheology on primary human dermal fibroblasts from female donors with age of 28 years (young) and 86 years (old) as model of in vivo aging and human normal dermal fibroblast from 11-year old male with CPD 17-35 (young) and CPD 58-59 (senescence) as a model of in vitro replicative senescence. In contrast to the expectations, our primary results show no significant differences in the viscoelastic properties of fibroblasts depending on age of the donor or cellular replicative senescence.

Keywords: aging, cytoskeleton, fibroblast, mechanical properties

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Authors: Ahmed Banda, Alaa Maghrabi, Aiman Fakieh

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Terahertz (THz) range lies in the area between 0.1 to 10 THz. The process of generating and detecting THz can be done through different techniques. One of the most familiar techniques is done through a photoconductive antenna (PCA). The process of generating THz radiation at PCA includes applying a laser pump in femtosecond and DC voltage difference. However, photocurrent is generated at PCA, which its value is affected by different parameters (e.g., dielectric properties, DC voltage difference and incident power of laser pump). THz radiation is used for biomedical applications. However, different biomedical fields need new technologies to meet patients’ needs (e.g. blood-related conditions). In this work, a novel method to check the red blood cells (RBCs) concentration of a blood sample using PCA is presented. RBCs constitute 44% of total blood volume. RBCs contain Hemoglobin that transfers oxygen from lungs to body organs. Then it returns to the lungs carrying carbon dioxide, which the body then gets rid of in the process of exhalation. The configuration has been simulated and optimized using COMSOL Multiphysics. The differentiation of RBCs concentration affects its dielectric properties (e.g., the relative permittivity of RBCs in the blood sample). However, the effects of four blood samples (with different concentrations of RBCs) on photocurrent value have been tested. Photocurrent peak value and RBCs concentration are inversely proportional to each other due to the change of dielectric properties of RBCs. It was noticed that photocurrent peak value has dropped from 162.99 nA to 108.66 nA when RBCs concentration has risen from 0% to 100% of a blood sample. The optimization of this method helps to launch new products for diagnosing blood-related conditions (e.g., anemia and leukemia). The resultant electric field from DC components can not be used to count the RBCs of the blood sample.

Keywords: biomedical applications, photoconductive antenna, photocurrent, red blood cells, THz radiation

Procedia PDF Downloads 192

Authors: Jinhui Bak, Bumjin Kim

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The purpose of this study is to analyze how the scientific and mathematical fields (STEM) and liberal arts (A) work together in the STEAM program. In the future STEAM programs that have been designed and developed, the humanities will act not just as a 'tool' for science technology and mathematics, but as a 'core' content to have an equivalent status. STEAM was first introduced to the Republic of Korea in 2011 when the Ministry of Education emphasized fostering creative convergence talent. Many programs have since been developed under the name STEAM, but with the majority of programs focusing on technology education, arts and humanities are considered secondary. As a result, arts is most likely to be accepted as an option that can be excluded from the teachers who run the STEAM program. If what we ultimately pursue through STEAM education is in fostering STEAM literacy, we should no longer turn arts into a tooling area for STEM. Based on this consciousness, this study analyzed over 160 STEAM programs in middle and high schools, which were produced and distributed by the Ministry of Education and the Korea Science and Technology Foundation from 2012 to 2017. The framework of analyses referenced two criteria presented in the related prior studies: normative convergence and technological convergence. In addition, we divide Arts into fine arts and liberal arts and focused on Korean Language Course which is in liberal arts and analyzed what kind of curriculum standards were selected, and what kind of process the Korean language department participated in teaching and learning. In this study, to ensure the reliability of the analysis results, we have chosen to cross-check the individual analysis results of the two researchers and only if they are consistent. We also conducted a reliability check on the analysis results of three middle and high school teachers involved in the STEAM education program. Analyzing 10 programs selected randomly from the analyzed programs, Cronbach's α .853 showed a reliable level. The results of this study are summarized as follows. First, the convergence ratio of the liberal arts was lowest in the department of moral at 14.58%. Second, the normative convergence is 28.19%, which is lower than that of the technological convergence. Third, the language and achievement criteria selected for the program were limited to functional areas such as listening, talking, reading and writing. This means that the convergence of Korean language departments is made only by the necessary tools to communicate opinions or promote scientific products. In this study, we intend to compare these results with the STEAM programs in the United States and abroad to explore what elements or key concepts are required for the achievement criteria for Korean language and curriculum. This is meaningful in that the humanities field (A), including Korean, provides basic data that can be fused into 'equivalent qualifications' with science (S), technical engineering (TE) and mathematics (M).

Keywords: Korean STEAM Programme, liberal arts, STEAM curriculum, STEAM Literacy, STEM

Procedia PDF Downloads 141

Authors: Danielle C. B. Honorato, Rafael H. Carvalho, Adriana L. Soares, Ana Paula F. R. L. Bracarense, Paulo D. Guarnieri, Massami Shimokomaki, Elza I. Ida

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Over the past decade, the Brazilian production of turkey meat increased by more than 50%, indicating that the turkey meat is considered a great potential for the Brazilian economy contributing to the growth of agribusiness at the marketing international scenario. However, significant color changes may occur during its processing leading to the pale, soft and exudative (PSE) appearance on the surface of breast meat due to the low water holding capacity (WHC). Changes in PSE meat functional properties occur due to the myofibrils proteins denaturation caused by a rapid postmortem glycolysis resulting in a rapid pH decline while the carcass temperature is still warm. The aim of this study was to analyze the physical, chemical and histological characteristics of PSE turkey meat obtained from a Brazilian commercial processing plant. The turkey breasts samples were collected (n=64) at the processing line and classified as PSE at L* ≥ 53 value. The pH was also analyzed after L* measurement. In sequence, PSE meat samples were evaluated for WHC, cooking loss (CL), shear force (SF), myofibril fragmentation index (MFI), protein denaturation (PD) and histological evaluation. The abnormal color samples presented lower pH values, 16% lower fiber diameter, 11% lower SF and 2% lower WHC than those classified as normal. The CL, PD and MFI were, respectively, 9%, 18% and 4% higher in PSE samples. The Pearson correlation between the L* values and CL, PD and MFI was positive, while that SF and pH values presented negative correlation. Under light microscopy, a shrinking of PSE muscle cell diameter was approximately 16% shorter in relation to normal samples and an extracellular enlargement of endomysium and perimysium sheaths as the consequence of higher water contents lost as observed previously by lower WHC values. Thus, the results showed that PSE turkey breast meat presented significant changes in their physical, chemical and histological characteristics that may impair its functional properties.

Keywords: functional properties, histological evaluation, meat quality, PSE

Procedia PDF Downloads 442

Authors: Chih-Chang Yeh, Min-Fang Yang, Hsin-I Chang

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Chitosan is a cationic polysaccharide derived from the partial deacetylation of chitin. Hyaluronic acid (HA), chondroitin sulfate (CS) and heparin (HP) are anionic glycosaminoglycans (GCGs) which can regulate osteogenic activity. In this study, chitosan membranes were prepared by glutaraldehyde crosslinking reaction and then complexed with three different types of GCGs. 7F2 osteoblasts-like cells and macrophages Raw264.7 were used as models to study the influence of chitosan membranes on osteometabolism. Although chitosan membranes are highly hydrophilic, the membranes associated with GCG-chitosan complexes showed about 60-70% cell attachment. Furthermore, the membranes associated with HP-chitosan complexes could increase ALP activity in comparison with chitosan films only. Three types of the membranes associated with GCG-chitosan complexes could significantly inhibit LPS induced-nitric oxide expression. In addition, chitosan membranes associated with HP and HA can down-regulate tartrate-resistant acid phosphatase (TRAP) activity but not CS-chitosan complexes. Based on these results, we conclude that chitosan membranes associated with HP can increase ALP activity in osteoblasts and chitosan membranes associated with HP and HA reduce TRAP activity in osteoclasts.

Keywords: osteoblast, osteoclast, chitosan, glycosaminoglycan

Procedia PDF Downloads 510

Authors: Ali Mashayekh, Mahdiye Khorasani, Thomas weyh

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The demand for battery electric vehicles (BEV) is steadily increasing, and it can be assumed that electric mobility will dominate the market for individual transportation in the future. Regarding BEVs, the focus of state-of-the-art research and development is on vehicle batteries since their properties primarily determine vehicles' characteristic parameters, such as price, driving range, charging time, and lifetime. State-of-the-art battery packs consist of invariable configurations of battery cells, connected in series and parallel. A promising alternative is battery systems based on multilevel inverters, which can alter the configuration of the battery cells during operation via semiconductor switches. The main benefit of such topologies is that a three-phase AC voltage can be directly generated from the battery pack, and no separate power inverters are required. Therefore, modular battery systems based on different multilevel inverter topologies and reconfigurable battery systems are currently under investigation. Another advantage of the multilevel concept is that the possibility to reconfigure the battery pack allows battery cells with different states of charge (SoC) to be connected in parallel, and thus low-loss balancing can take place between such cells. In contrast, in conventional battery systems, parallel connected (hard-wired) battery cells are discharged via bleeder resistors to keep the individual SoCs of the parallel battery strands balanced, ultimately reducing the vehicle range. Different multilevel inverter topologies and reconfigurable batteries have been described in the available literature that makes the before-mentioned advantages possible. However, what has not yet been described is how an intelligent operating algorithm needs to look like to keep the SoCs of the individual battery strands of a modular battery system with integrated power electronics balanced. Therefore, this paper suggests an SoC balancing approach for Battery Modular Multilevel Management (BM3) converter systems, which can be similarly used for reconfigurable battery systems or other multilevel inverter topologies with parallel connectivity. The here suggested approach attempts to simultaneously utilize all converter modules (bypassing individual modules should be avoided) because the parallel connection of adjacent modules reduces the phase-strand's battery impedance. Furthermore, the presented approach tries to reduce the number of switching events when changing the switching state combination. Thereby, the ohmic battery losses and switching losses are kept as low as possible. Since no power is dissipated in any designated bleeder resistors and no designated active balancing circuitry is required, the suggested approach can be categorized as a proactive balancing approach. To verify the algorithm's validity, simulations are used.

Keywords: battery management system, BEV, battery modular multilevel management (BM3), SoC balancing

Procedia PDF Downloads 112

Authors: I. C. Orabueze, A. A. Amudalat, S. A. Adesegun, A. A. Usman

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Dental and associated oral diseases are increasingly affecting a considerable portion of the population and are considered some of the major causes of tooth loss, discomfort, mouth odor and loss of confidence. This study focused on the ethnobotanical survey of medicinal plants used in oral therapy and evaluation of the antimicrobial activities of methanolic extracts of two selected plants from the survey for their efficacy against dental microorganisms. The ethnobotanical survey was carried out in six herbal markets in Lagos State, Nigeria by oral interviewing and information obtained from an old family manually complied herbal medication book. Methanolic extracts of Olax subscorpioidea (stem bark) and Bridelia ferruginea (stem bark) were assayed for their antimicrobial activities against clinical oral isolates (Aspergillus fumigatus, Candida albicans, Streptococcus spp, Staphylococcus aureus, Lactobacillus acidophilus and Pseudomonas aeruginosa). In vitro microbial technique (agar well diffusion method and minimum inhibitory concentration (MIC) assay) were employed for the assay. Chlorhexidine gluconate was used as the reference drug for comparison with the extract results. And the preliminary phytochemical screening of the constituents of the plants were done. The ethnobotanical survey produced plants (28) of diverse family. Different parts of plants (seed, fruit, leaf, root, bark) were mentioned but 60% mentioned were either the stem or the bark. O. subscorpioidea showed considerable antifungal activity with zone of inhibition ranging from 2.650 – 2.000 cm against Aspergillus fumigatus but no such encouraging inhibitory activity was observed in the other assayed organisms. B. ferruginea showed antibacterial sensitivity against Streptococcus spp, Staphylococcus aureus, Lactobacillus acidophilus and Pseudomonas aeruginosa with zone of inhibitions ranging from 3.400 - 2.500, 2.250 - 1.600, 2.700 - 1.950, 2.225 – 1.525 cm respectively. The minimum inhibitory concentration of O. subscorpioidea against Aspergillus fumigatus was 51.2 mg ml-1 while that of B. ferruginea against Streptococcus spp was 0.1mg ml-1 and for Staphylococcus aureus, Lactobacillus acidophilus and Pseudomonas aeruginosa were 25.6 mg ml-1. A phytochemical analysis reveals the presence of alkaloids, saponins, cardiac glycoside, tannins, phenols and terpenoids in both plants, with steroids only in B. ferruginea. No toxicity was observed among mice given the two methanolic extracts (1000 mg Kg-1) after 21 days. The barks of both plants exhibited antimicrobial properties against periodontal diseases causing organisms assayed, thus up-holding their folkloric use in oral disorder management. Further research could be done viewing these extracts as combination therapy, checking for possible synergistic value in toothpaste and oral rinse formulations for reducing oral bacterial flora and fungi load.

Keywords: antimicrobial activities, Bridelia ferruginea, dental disinfection, methanolic extract, Olax subscorpioidea, ethnobotanical survey

Procedia PDF Downloads 229

Authors: Mahesh Pal, Tripti Mishra, Chandana Rao, Dalip Upreti

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Cancer has been considered to be a very dreadful disease. Holmskioldia sanguinea is a large climbing shrub found in the Himalayas at an altitude of 5,000 ft and preliminary investigation showed the excellent yield of andrographolide and subjected for the anticancer activity. Protective effect of Holmskioldia sanguinea leaf ethanolic extract has been investigated against Ehrlich ascites carcinoma (EAC) and Daltons ascites lymphoma (DAL) in Swiss albino mice to evaluate the possible mechanism of action. The enzymatic antioxidant status was studied on tumor bearing mice, which shows the potential of the compound to possess significant free radical scavenging property and revealed significant tumor regression and prolonged survival time. The isolated bioactive molecule andrographolide from Holmskioldia sanguinea yields (2.5%) in subject to HPTLC/HPLC analysis. The cellular defense system constituting the superoxide dismutase, catalyses was enhanced whereby the lipid peroxidation content was restricted to a larger extent. The Holmskioldia sanguinea is a new source of andrographolide and demonstrated the potency in treatment of cancer.

Keywords: Holmskioldia sanguinea, tumor, mice, andrographolide

Procedia PDF Downloads 241

Authors: Samar Omar A. Rabah, Sahar Ragab El Hadad, Fatmah Albani

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Since the discovery of Agarwood (Incense tree), many studies reported its characteristic effects and variable benefits, as either to produce Arabian Incense or as a traditional medicine against many diseases. Laboratory experiments were carried out on the effect of different concentrations of Incense smoke inhalation on the lung weight and tissue in female mice. This research derives its importance from the fact that Incense is heavily used in Saudi Arabia in the absence of thorough studies of its effects on health. Eighty animals are used in this study, and they are divided into four groups, each is 20 animals. Three groups are exposed to different concentrations (2, 4 and 6 gm) of Incense smoke daily for three months, and the fourth group is the control. At the end of each month, five animals from each group were dissected. Obtained data showed an increase but not significant in animal body and lung weight, this results return to natural increase as a result of normal growth of animals. Light microscope reveals some changes in the lung tissue, such as focal emphysema, rupture in the alveolar walls, hemorrhage, congestion, edema and few peri-bronchial lymphoid cells. After continuous exposure to Incense smoke focal necrosis and degradation are observed in some cells of epithelial bronchioles. Also, fibrosis of peri-bronchial, thickening in alveolar walls and aggregation of lymphoid cells are demonstrated in some lungs sections. according to the above manifestations it could be concluded that exposure to Incense smoke causes pulmonary harmful effects. Therefore, we can recommend that Incense smoke will be used only in open places to reduce its harms.

Keywords: incense smoke, lungs, histological changes of lungs, agarwood

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Authors: Gyati Shilakari Asthana, Abhay Asthana

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Purpose: The therapeutic potential of oligonucleotide (ODN) is primarily dependent upon its safe and efficient delivery to specific cells overcoming degradation and maximizing cellular uptake in vivo. The present study is focused to design low molecular weight chitosan nanoconstructs to meet the requirements of safe and effectual delivery of ODNs. LMW-chitosan is a biodegradable, water soluble, biocompatible polymer and is useful as a non-viral vector for gene delivery due to its better stability in water. Methods: LMW chitosan ODN nanoparticles (CHODN NPs) were formulated by self assembled method using various N/P ratios (moles ratio of amine groups of CH to phosphate moieties of ODNs; 0.5:1, 1:1, 3:1, 5:1 and 7:1) of CH to ODN. The developed CHODN NPs were evaluated with respect to gel retardation assay, particle size, zeta potential and cytotoxicity and transfection efficiency. Results: Complete complexation of CH/ODN was achieved at the charge ratio of 0.5:1 or above and CHODN NPs displayed resistance against DNase I. On increasing the N/P ratio of CH/ODN, particle size of the NPs decreased whereas zeta potential (ZV) value increased. No significant toxicity was observed at all CH concentrations. The transfection efficiency was increased on increasing N/P ratio from 1:1 to 3:1, whereas it was decreased with further increment in N/P ratio upto 7:1. Maximum transfection of CHODN NPs with both the cell lines (Raw 267.4 cells and Hela cells) was achieved at N/P ratio of 3:1. The results suggest that transfection efficiency of CHODN NPs is dependent on N/P ratio. Conclusion: Thus the present study states that LMW chitosan nanoparticulate carriers would be acceptable choice to improve transfection efficiency in vitro as well as in vivo delivery of oligonucleotide.

Keywords: LMW-chitosan, chitosan nanoparticles, biocompatibility, cytotoxicity study, transfection efficiency, oligonucleotide

Procedia PDF Downloads 483

Authors: Arunporn Itharat, Kamonmas Srikwan, Srisopa Ruangnoo, Pakakrong Thongdeeying

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Background: The rhizomes of Smilax glabra (SG) has long been used in Traditional Chinese and Thai herbal medicine to treat a variety of infectious diseases and immunological disorders. Objective: To investigate the in vitro anti-allergic activities of crude extracts and pure isolated flavonoid compounds from SG by determination of inhibitory effects on antigen-induced release of β-hexosaminidase from RBL-2H3 cells. Methods: The in vitro inhibitory effects of crude aqueous and organic extracts on beta-hexosaminidase release in RBL-2H3 cells were evaluated as an in vitro indication of possible anti-allergic activity in vivo. Bioassay-guided fractionation of extracts was used to isolate flavonoid compounds from the ethanolic extracts. Results: The 95% and 50% ethanolic extracts of SG showed remarkably high anti-allergic activity, with IC50 values of 5.74 ± 2.44 and 23.54 ± 4.75 μg/ml, much higher activity than that for Ketotifen (IC50 58.90 μM). The water extract had negligible activity (IC50 > 100 μg/ml). The two isolated flavonols, Engeletin and Astilbin, showed weak anti-allergic activity, IC50 values 97.46 ± 2.04 and > 100 μg/ml, respectively. Conclusions: The 95% and 50% ethanolic extracts of SG showed strong anti-allergic activity but two flavonol constituents did not show any significant anti-allergic activity. These findings suggest that a combination of effects of various phytochemicals in crude extracts used in traditional medicine are responsible for the purported anti-allergic activity of SG herbal preparations. The plethora of constituents in crude extracts, as yet unidentified, are likely to be acting synergistically to account for the strong observed anti-allergic in vitro activity.

Keywords: Smilax glabra, anti-allergic activity, RBL-2H3 cells, flavonoid compounds

Procedia PDF Downloads 651

Authors: Mohamed Moussaoui

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A series of thirty-one potential inhibitors targeting the epidermal growth factor receptor kinase (EGFR), derived from quinazoline, underwent 3D-QSAR analysis using CoMFA and CoMSIA methodologies. The training and test sets of quinazoline derivatives were utilized to construct and validate the QSAR models, respectively, with dataset alignment performed using the lowest energy conformer of the most active compound. The best-performing CoMFA and CoMSIA models demonstrated impressive determination coefficients, with R² values of 0.981 and 0.978, respectively, and Leave One Out cross-validation determination coefficients, Q², of 0.645 and 0.729, respectively. Furthermore, external validation using a test set of five compounds yielded predicted determination coefficients, R² test, of 0.929 and 0.909 for CoMFA and CoMSIA, respectively. Building upon these promising results, eighteen new compounds were designed and assessed for drug likeness and ADMET properties through in silico methods. Additionally, molecular docking studies were conducted to elucidate the binding interactions between the selected compounds and the enzyme. Detailed molecular dynamics simulations were performed to analyze the stability, conformational changes, and binding interactions of the quinazoline derivatives with the EGFR kinase. These simulations provided deeper insights into the dynamic behavior of the compounds within the active site. This comprehensive analysis enhances the understanding of quinazoline derivatives as potential anti-cancer agents and provides valuable insights for lead optimization in the early stages of drug discovery, particularly for developing highly potent anticancer therapeutics

Keywords: 3D-QSAR, CoMFA, CoMSIA, ADMET, molecular docking, quinazoline, molecular dynamic, egfr inhibitors, lung cancer, anticancer

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Authors: Aishik Deb, Rituparna Sinha

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We have developed an algorithm to detect the abnormal segment/"structural variation in the genome across a number of samples. We have worked on simulated as well as real data from the BAM Files and have designed a segmentation algorithm where abnormal segments are detected. This algorithm aims to improve the accuracy and performance of the existing CNV-CH algorithm. The next-generation sequencing (NGS) approach is very fast and can generate large sequences in a reasonable time. So the huge volume of sequence information gives rise to the need for Big Data and parallel approaches of segmentation. Therefore, we have designed a map-reduce approach for the existing CNV-CH algorithm where a large amount of sequence data can be segmented and structural variations in the human genome can be detected. We have compared the efficiency of the traditional and map-reduce algorithms with respect to precision, sensitivity, and F-Score. The advantages of using our algorithm are that it is fast and has better accuracy. This algorithm can be applied to detect structural variations within a genome, which in turn can be used to detect various genetic disorders such as cancer, etc. The defects may be caused by new mutations or changes to the DNA and generally result in abnormally high or low base coverage and quantification values.

Keywords: cancer detection, convex hull segmentation, map reduce, next generation sequencing

Procedia PDF Downloads 118

Authors: Donghee Kang

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Background: Cancer pain is very difficult to control as the mechanism of pain is varied, and the patient has several co-morbidities. The use of Intravenous Patient-Controlled Analgesia (IV-PCA) can effectively control underlying pain and breakthrough pain. Ketamine is used in many pain patients due to its unique analgesic effect. In this study, it was checked whether there was a difference in the amount of analgesic usage, pain control degree, and side effects between patients who controlled pain with fentanyl-based IV-PCA and those who added Ketamine for pain control. Methods: Among the patients referred to this department for cancer pain, IV-PCA was applied to patients who were taking sufficient oral analgesics but could not control them or had blood clotting disorders that made the procedure difficult, and this patient group was targeted. In IV-PCA, 3000 mcg of Fentanyl, 160 mg of Nefopam, and 0.3 mg of Ramosetrone were mixed with normal saline to make a total volume of 100 ml. Group F used this IV-PCA as it is, and group K mixed 250 mg of Ketamine with normal saline to make a total volume of 100 ml. For IV-PCA, the basal rate was 0.5ml/h, the bolus was set to 1ml when pressed once, and the lockout time was set to 15 minutes. If pain was not controlled after IV-PCA application, 500 mcg of Fentanyl was added, and if excessive sedation or breathing difficulties occurred, the use was stopped for one hour. After that, the degree of daily pain control, analgesic usage, and side effects were investigated for seven days using this IV-PCA. Results: There was no difference between the two groups in the demographic data. Both groups had adequate pain control. Initial morphine milligram equivalents did not differ between the two groups, but the total amount of Fentanyl used for seven days was significantly different between the two groups [p=0.014], and group F used more Fentanyl through IV-PCA. In addition, the amount of sleeping pills used during the seven days was higher in Group F [p<0.01]. Overall, there was no difference in the frequency of side effects between the two groups, but the nausea was more frequent in Group F [p=0.031]. Discussion: When the two groups were compared, pain control was good in both groups. This seems to be because Fentanyl-based IV-PCA showed an adequate pain control effect. However, there was a significant difference in the total amount of opioid (Fentanyl) used, which is thought to be the opioid-sparing effect of Ketamine. Also, among the side effects, nausea was significantly less, which is thought to be possible because the amount of opioids used in the Ketamine group was small. The frequency of requesting sleeping pills was significantly less in the group using Ketamine, and it seems that Ketamine also helped improve sleep quality. In conclusion, using Ketamine with an opioid to control pain seems to have advantages. IV-PCA, which can be used effectively when other procedures are difficult, is more effective and safer when used together with Ketamine than opioids alone.

Keywords: cancer pain, intravenous patient-controlled analgesia, Ketamine, opioid

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Authors: Arpit Gite

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Purpose: We have evaluated the role of Total Neoadjuvant Therapy in patients with Locally Advanced Rectal cancer by giving Chemoradiotherapy followed by consolidation chemotherapy (CRT-CNCT) and, after that, the strategy of wait and watch. Methods: In this prospective case series, we evaluated the results of three locally advanced Rectal cancers, two cases Stage II (cT3N0) and one case Stage III ( cT4aN2). All three patients' growth was 4-6 cm from the anal verge. We have treated with Chemoradiotherapy to dose of 45Gy/25 Fractions to elective nodal regions (Inguinal node in anal canal Involvement)and Primary and mesorectum (Phase I) followed by 14.4Gy/8 Fractions to Primary and Mesorectum(Phase II) to a total dose of 59.4Gy/33 Fractions with concurrent chemotherapy Tab Capecitabine 825mg/m2 PO BD with Radiation therapy. After 6 weeks of completion of Chemoradiotherapy, advised six cycles of consolidative chemotherapy, CAPEOX regimen, Oxaliplatin 130mg/m2 on day 1 and Capecitabine 1000mg/m2 PO BD on days 1-14 repeated on a 21-day cycle for a total of six cycles. The primary endpoint is Disease-free survival (DFS); the secondary endpoint is adverse events related to chemoradiotherapy. Radiation toxicity is assessed by RTOG criteria, and chemotherapy toxicity is assessed by Common Terminology Criteria for Adverse Events (CTCAE) Version 5.0. Results: After 6 weeks of completion of Chemoradiotherapy, we did PET-CT of all three patients; all three patients had a clinically complete response and we advised 6 cycles of consolidative chemotherapy. After completion of consolidative chemotherapy, again PET-CT and sigmoidoscopy, all three patients had complete response on PET-CT and no lesions on sigmoidoscopy and kept all three patients on wait and watch.2 patients had Grade 2 skin toxicities,1 patient had Grade 1 skin toxicity, .2 patients had Grade 2 lower GI toxicities, and 1 patient had Grade lower GI toxicity, both according to RTOG criteria. 3 patients had Grade 2 diarrhea due to capecitabine, and 1 patient had Grade 1 thrombocytopenia due to oxaliplatin assessed by Common Terminology Criteria for Adverse Events (CTCAE) Version 5.0. Conclusion: Sphincter Preservation is possible with this regimen in those who don’t want to opt for surgery or in case of low-lying rectal cancer.

Keywords: locally advanced rectal cancer, sphincter preservation, chemoradiotherapy, consolidative chemotherapy

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Authors: Yousuf M. Al Suleimani, Robin Hiley

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The endogenous phospholipid lysophosphatidylinositol (LPI) was recently identified as a novel ligand for the G protein-coupled receptor 55 (GPR55) and an inducer of intracellular Ca2+ [Ca2+]i release. This study attempts to characterize Ca2+ signals provoked by LPI in HEK293 cells engineered to stably express human GPR55 and to test cannabinoid ligand activity at GPR55. The study shows that treatment with LPI stimulates a sustained, oscillatory Ca2+ release. The response is characterized by an initial rapid rise, which is mediated by the Gαq-PLC-IP3 pathway, and this is followed by prolonged oscillations that require RhoA activation. Ca2+ oscillations are initiated by intracellular mechanisms and extracellular Ca2+ is only required to replenish Ca2+ lost from the cytoplasm. Analysis of cannabinoid ligand activity at GPR55 revealed no clear effect of the endocannabinoid anandamide, however, rimonabant and the CB1 receptor antagonist AM251 evoked GPR55-mediated [Ca2+]i. Thus, LPI is likely to be a key plasma membrane mediator of signaling events and changes in gene expression through GPR55 activation.

Keywords: lysophosphatidylinositol, calcium, GPR55, cannabinoid

Procedia PDF Downloads 341

Authors: Benchouala Amira, Bojic Clement, Poupin Pascal, Cossu Leguille-carole

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The objective of this study is to evaluate in vitro the cytotoxic and androgenic potential of several antifungal molecules (amphotericin B, econazole, ketoconazole and miconazole) on MDA-Kb2 cell lines. This biological model is an effective tool for the detection of endocrine disruptors because it responds well to the main agonist of the androgen receptor (testosterone) and also to an antagonist: flutamide. The cytotoxicity of each chemical compound tested was measured using an MTT assay (tetrazolium salt, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) which measures the activity of the reductase function of mitochondrial succinate dehydrogenase enzymes of cultured cells. This complementary cytotoxicity test is essential to ensure that the effects of reduction in luminescence intensity observed during androgenic tests are only attributable to the anti-androgenic action of the compounds tested and not to their possible cytotoxic properties. Tests of the androgenic activity of antifungals show that these compounds do not have the capacity to induce transcription of the luciferase gene. These compounds do not exert an androgenic effect on MDA-Kb2 cells in culture for the environmental concentrations tested. The addition of flutamide for the same tested concentrations of antifungal molecules reduces the luminescence induced by amphotericin B, econazole and miconazole, which is explained by a strong interaction of these molecules with flutamide which may have a greater toxic effect than when tested alone. The cytotoxicity test shows that econazole and ketoconazole can cause cell death at certain concentrations tested. This cell mortality is perhaps induced by a direct or indirect action on deoxyribonucleic acid (DNA), ribonucleic acid (RNA) or proteins necessary for cell division.

Keywords: cytotoxicity, androgenic potential, antifungals, MDA-Kb2

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Authors: Jiangang Shen

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Olfactory dysfunction is a common symptom companied by anxiety- and depressive-like behaviors in depressive patients. Chronic stress triggers hormone responses and inhibits the proliferation and differentiation of neural stem cells (NSCs) in the hippocampus and subventricular zone (SVZ)-olfactory bulb (OB), contributing to depressive behaviors and olfactory dysfunction. However, the cellular signaling molecules to regulate chronic stress mediated olfactory dysfunction are largely unclear. Adaptor proteins containing the pleckstrin homology domain, phosphotyrosine binding domain, and leucine zipper motif (APPLs) are multifunctional adaptor proteins. Herein, we tested the hypothesis that APPL2 could inhibit hippocampal neurogenesis by affecting glucocorticoid receptor (GR) signaling, subsequently contributing to depressive and anxiety behaviors as well as olfactory dysfunctions. The major discoveries are included: (1) APPL2 Tg mice had enhanced GR phosphorylation under basic conditions but had no different plasma corticosterone (CORT) level and GR phosphorylation under stress stimulation. (2) APPL2 Tg mice had impaired hippocampal neurogenesis and revealed depressive and anxiety behaviors. (3) GR antagonist RU486 reversed the impaired hippocampal neurogenesis in the APPL2 Tg mice. (4) APPL2 Tg mice displayed higher GR activity and less capacity for neurogenesis at the olfactory system with lesser olfactory sensitivity than WT mice. (5) APPL2 negatively regulates olfactory functions by switching fate commitments of NSCs in adult olfactory bulbs via interaction with Notch1 signaling. Furthermore, baicalin, a natural medicinal compound, was found to be a promising agent targeting APPL2/GR signaling and promoting adult neurogenesis in APPL2 Tg mice and chronic corticosterone-induced depression mouse models. Behavioral tests revealed that baicalin had antidepressant and olfactory-improving effects. Taken together, APPL2 is a critical therapeutic target for antidepressant treatment.

Keywords: APPL2, hippocampal neurogenesis, depressive behaviors and olfactory dysfunction, stress

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Authors: Bagus Indrawan Hardi

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Cellular Automata are widely used in land use modeling, it has been proven powerful to simulate land use change for small scale in many large cities in the world. In this paper, we try to implement CA for land use modeling in unique city in Indonesia, Karawang. Instead the complex numerical implementation, CA are simple, and it is accurate and also highly dependable on the on the rules (rule based). The most important to do in CA is how we form and calculate the neighborhood effect. The neighborhood effect represents the environment and relationship situation between the occupied cell and others. We adopted 196 cells of circular neighborhood with 8 cells of radius. For the results, CA works well in this study, we exhibit several analyzed and proceed of zoomed part in Karawang region. The rule set can handle the complexity in land use modeling. However, we cannot strictly believe of the result, many non-technical parameters, such as politics, natural disaster activities, etc. may change the results dramatically.

Keywords: cellular automata (CA), land use change, spatial dynamics, urban sprawl

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Authors: Chaoran Liu

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Bioactive polysaccharides and polysaccharide-protein complex derived from mushrooms and fungi have a wide range of immunomodulatory activity with low side-effects and have therefore the potential to be developed as an adjuvant in cancer therapies. Mushrooms sclerotium is rich in polysaccharides and the polysaccharides isolated from the sclerotium of Polyporus rhinocerus have shown potent in vivo and in vitro immunomodulatory effects. Macrophages are considered to be an important component of the innate immune response against bacterial infection and cancer. To better understanding the immunomodulatory effects and its underlying mechanisms of sclerotial water-soluble polysaccharides extracted from P. rhinocerus on macrophages, the objectives of this study are to purify the water-soluble novel sclerotial polysaccharides and to characterize the structure and properties as well as to study the detailed molecular mechanisms of the in vitro immunomodulating effects in murine macrophages. The hot water-soluble fraction PRW from the sclerotium of P. rhinocerus was obtained using solvent extraction. PRW was further fractionated by membrane ultrafiltration to a give a fraction (PRW1) with molecular mass less than 50 kDa. PRW1 was characterized to be a polysaccharide-protein complex composed of 45.7% polysaccharide and 44.2% protein. The chemical structure of the carbohydrate moiety of PRW1 was elucidated by GC and FTIR to be mainly beta-D-glucan with trace amount of galactose and mannose. The immunomodulatory effects of PRW1 on murine RAW 264.7 macrophages were demonstrated in terms of the increase in nitric oxide production and cytokine production. Mechanistically, PRW1 initiates ERK phosphorylation to activate macrophages within 15 min and significantly improves the expression level of inducible NOS (iNOS) from 6 h after treatment. In summary, this study indicates that PRW1 is a potent immunomodulatory agent for macrophages and suggests that mushroom sclerotia from Polyporus rhinocerus requires for further investigation in cancer research.

Keywords: Polyporus rhinocerus, mushroom sclerotia, Polysaccharide-Protein Complex, macrophage activation

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Authors: A. Saurabh Singh, B. Raghvendra, C. Prabhakar Singh

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Solid Oxide Fuel Cells (SOFCs) are one of the most attractive electrochemical energy conversion systems, as these devices present a clean energy production, thus promising high efficiencies and low environmental impact. The electrodes are the main components that decisively control the performance of a SOFC. Conventional, anode materials (like Ni-YSZ) are operates at very high temperature. Therefore, cost-effective materials which operate at relatively lower temperatures are still required. In present study, we have synthesized La doped Strontium Titanate via solid state reaction route. The structural, microstructural and density of the pellet have been investigated employing XRD, SEM and Archimedes Principle, respectively. The electrical conductivity of the systems has been determined by impedance spectroscopy techniques. The electrical conductivity of the Lanthanum Strontium Titanate (LST) has been found to be higher than the composite Ni-YSZ system at 700 °C.

Keywords: IT-SOFC, LST, Lanthanum Strontium Titanate, electrical conductivity

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Authors: Farah Diab, Mohamad Khalil, Francesca Storace, Francesca Baldini, Piero Portincasaa, Giulio Lupidi, Laura Vergani

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Thymbra spicata is a thyme-like plant belonging to the Lamiaceae family that shows a global distribution, especially in the eastern Mediterranean region. Leaves of T. spicata contain large amounts of phenols such as phenolic acids (rosmarinic acid), phenolic monoterpenes (carvacrol), and flavonoids. In Lebanon, T. spicata is currently used as a culinary herb in salad and infusion, as well as for traditional medicinal purposes. Carvacrol (5-isopropyl-2-methyl phenol), the most abundant polyphenol in the organic extract and essential oils, has a great array of pharmacological properties. In fact, carvacrol is largely employed as a food additive and neutraceutical agent. Our aim is to investigate the beneficial effects of T. spicata ethanolic extract (TE) and its main component, carvacrol, using in vitro models of hepatic steatosis and endothelial dysfunction. As a further point, we focused on investigating if and how the binding of carvacrol to albumin, the physiological transporter for drugs in the blood, might be altered by the presence of high levels of fatty acids (FAs), thus impairing the carvacrol bio-distribution in vivo. For that reason, hepatic FaO cells treated with exogenous FAs such as oleate and palmitate mimic hepatosteatosis; endothelial HECV cells exposed to hydrogen peroxide are a model of endothelial dysfunction. In these models, we measured lipid accumulation, free radical production, lipoperoxidation, and nitric oxide release before and after treatment with carvacrol. The carvacrol binding to albumin with/without high levels of long-chain FAs was assessed by absorption and emission spectroscopies. Our findings show that both TE and carvacrol (i) counteracted lipid accumulation in hepatocytes by decreasing the intracellular and extracellular lipid contents in steatotic FaO cells; (ii) decreased oxidative stress in endothelial cells by significantly reducing lipoperoxidation and free radical production, as well as, attenuating the nitric oxide release; (ii) high levels of circulating FAs reduced the binding of carvacrol to albumin. The beneficial effects of TE and carvacrol on both hepatic and endothelial cells point to a nutraceutical potential. However, high levels of circulating FAs, such as those occurring in metabolic disorders, might hinder the carvacrol transport, bio-distribution, and pharmacodynamics.

Keywords: carvacrol, endothelial dysfunction, fatty acids, non-alcoholic fatty liver diseases, serum albumin

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Authors: Rui Chen, Jinfeng Zhang, Chun-Sing Lee

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In the past couple of decades, nanoscale metal-organic frameworks (NMOFs) have been highlighted as promising delivery platforms for biomedical applications, which combine many potent features such as high loading capacity, progressive biodegradability and low cytotoxicity. While NMOF has been extensively used as carriers for drugs of different modalities, so far there is no report on exploiting the advantages of NMOF for combination therapy. Herein, we prepared core-shell nanoparticles, where each nanoparticle contains a single graphitic-phase carbon nitride (g-C3N4) nanosheet encapsulated by a zeolitic-imidazolate frameworks-8 (ZIF-8) shell. The g-C3N4 nanosheets are effective visible-light photosensitizer for photodynamic therapy (PDT). When hosting DOX (doxorubicin), the as-synthesized core-shell nanoparticles could realize combinational photo-chemo therapy and provide dual-color fluorescence imaging. Therefore, we expect NMOFs-based core-shell nanoparticles could provide a new way to achieve much-enhanced cancer therapy.

Keywords: carbon nitride, combination therapy, drug delivery, nanoscale metal-organic frameworks

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Authors: Elin Julianti, Marlia Singgih, Masayoshi Arai, Jianyu Lin, Masteria Yunovilsa Putra, Muhammad Azhari, Agnia S. Muharam

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The search for a source of new medicinal compounds with anticancer activity from natural products has become important to resolve the ineffectiveness problem of pancreatic cancer therapy. Fungal marine microorganisms are prolific sources of bioactive natural products. In this present study, the ethyl acetate extract of cultured broth of Trichoderma longibrachiatum marine sponge-derived fungi exhibited selective cytotoxicity against human pancreatic carcinoma PANC-1 cells cultured under glucose-deficient conditions (IC50 = 98,4 µg/mL). The T. longibrachiatum was fermented by the static method at room temperature for 60 days. The culture broth was extracted using ethyl acetate by liquid-liquid extraction method. The liquid-liquid extraction was conducted toward the ethyl extract by using 90% MeOH-H₂O and n-|Hexane as a solvent. The extract of 90% MeOH-H₂O was fractionated by liquid extraction using by C₁₈ reversed-phase vacuum flash chromatography using mixtures of MeOH-H₂O, from 50:50 to 100:0, and 1% TFA MeOH as the eluents to yield six fractions. The fraction 2 (MeOH-H2O, 70:30) and fraction 3 (MeOH-H2O, 80:20) showed moderate cytotoxicity with IC50 value of 119.3 and 274.7 µg/mL, respectively. Fraction 4 (MeOH-H₂O, 90:10) showed the highest cytotoxicity activity with IC₅₀value of < 10 µg/mL. The chemical compounds of the fractions that are responsible for cytotoxic activity are potent for further investigation.

Keywords: cytotoxic activity, trichoderma longibrachiatum, marine-derived fungi, PANC-1 cell line

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Authors: Mohammad Y. Alfaifi

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Anything that poses a challenge or a threat to our well-being is a stress. Understanding the genetic material and cellular response of rats threatened with Repeated swimming stress provides insights that can influence human health. The aim of the present study was to assess the genetical damage and cytological changes caused by exposure of the test organism (Rattus rattus) to forced swimming stress. For this purpose, animals have been submerged in water path 15 minutes daily for 2 weeks. Following that, we performed a micronuclei (MN) test using MNNCE (Micronucleated normocromatic erythrocytes) and MNPCE (Micronucleated polychromatic erythrocytes), NDI (Nuclear division index) and cytological parameters using NDCI (nuclear division cytotoxicity index), necrotic and apoptotic cells in rat's bone marrow samples. Results showed that there was a slightly but not significant increase in the frequency of micronucleated as well as in cytological parameters in bone marrow cells.

Keywords: submergence stress, micronucleus, NDI, NDCI, toxicity, chromosomal aberrations

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Authors: Amenah Dhannoon, Ciaran Martin Hurley, Laura Wrafter, Podraic J. Regan

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Introduction: The COVID-19 pandemic continues to present unprecedented challenges for healthcare systems. This has resulted in the pragmatic shift in the practice of plastic surgery units worldwide. During this period, many units reported a significant fall in urgent melanoma referrals, leading to patients presenting with advanced disease requiring more extensive surgery and inferior outcomes. Our objective was to evaluate our unit's experience with both non-invasive and invasive melanoma during the COVID-19 pandemic and characterize our experience and contrast it to that experienced by our neighbors in the UK, mainland Europe and North America. Methods: a retrospective chart review was performed on all patients diagnosed with invasive and non-invasive cutaneous melanoma between March to December of 2019 (control) compared to 2020 (COVID-19 pandemic) in a single plastic surgery unit in Ireland. Patient demographics, referral source, surgical procedures, tumour characteristics, radiological findings, oncological therapies and follow-up were recorded. All data were anonymized and stored in Microsoft Excel. Results: A total of 589 patients were included in the study. Of these, 314 (53%) with invasive melanoma, compared to 275 (47%) with the non-invasive disease. Overall, more patients were diagnosed with both invasive and non-invasive melanoma in 2020 than in 2019 (p<0.05). However, significantly longer waiting times in 2020 (64 days) compared to 2019 (28 days) (p<0.05), with the majority of the referral being from GP in 2019 (83%) compared to 61% in 2020. Positive sentinel lymph node were higher in 2019 at 56% (n=28) compared to 24% (n=22) in 2020. There was no statistically significant difference in the tutor characteristics or metastasis status. Discussion: While other countries have noticed a fall in the melanoma diagnosis. Our units experienced a higher number of disease diagnoses. This can be due to multiple reasons. In Ireland, the government reached an early agreement with the private sector to continue elective surgery on an urgent basis in private hospitals. This allowed access to local anesthetic procedures and local skin cancer cases were triaged to non-COVID-19 provider centers. Our unit also adapted a fast, effective and minimal patient contact strategy for triaging skin cancer based on telemedicine. Thirdly, a skin cancer nurse specialist maintained patient follow-ups and triaging a dedicated email service. Finally, our plastic surgery service continued to maintain a virtual complex skin cancer multidisciplinary team meeting during the pandemic, ensuring local clinical governance has adhered to each clinical case. Conclusion: Our study highlights that with the prompt efficient restructuring of services, we could reserve successful management of skin cancer even in the most devastating times. It is important to reflect on the success during the pandemic and emphasize the importance of preparation for a potentially difficult future

Keywords: malignant melanoma, skin cancer, COVID-19, triage

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Authors: Francesca Lombardi, Francesca Rosaria Augello, Benedetta Cinque, Maria Grazia Cifone, Paola Palumbo

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Glioblastoma multiforme (GBM) is a high-grade primary brain tumor refractory to current forms of treatment. The survival benefits of patients with GBM remain unsatisfactory due to the intrinsic or acquired resistance to temozolomide (TMZ), an alkylating agent, used as the first-line chemotherapeutic drug to treat GBM patients. Its cytotoxic effect is visualized by the induction of O6-methylguanine (O6MeG) within DNA. Cyclooxygenase-2 (COX-2), an inflammation-associated enzyme, has been implicated in tumorigenesis and progression of GBM, its inhibition shows anticancer activities. In the present study, it was verified if the combination of a COX-2 selective inhibitor, NS398, with TMZ could counteract the TMZ resistance. In particular, the effect of NS398 mixed with TMZ was investigated in the GBM TMZ-resistant cell line, T98G. Cells were pretreated with NS398 (100µM, 24 hours) and then exposed to TMZ alone (200µM), NS398 alone, or both for 72 hours, after which cell growth rate and cycle phases, as well as apoptosis level, were evaluated. Coadministration of NS398 and TMZ caused a significant decrease in cell growth and a progressive increase of dead cells detected by trypan blue staining. Moreover, a significant level of apoptotic cell percentage and alteration of cell cycle phases were observed in T98G treated with TMZ-NS398 combination when compared to untreated cells or TMZ-treated cells. TMZ-resistant tumors, as GBM, express elevated levels of DNA repair enzyme O6-methylguanine-DNA methyltransferase (MGMT). The mixture drastically reduced MGMT expression in the TMZ-resistant cell line T98G, known to express high levels of MGMT basically. Moreover, while TMZ alone did not influence the COX-2 protein expression, the combination successfully reduced it. In conclusion, these results demonstrated that NS398 enhanced the efficacy of TMZ through cell number reduction, apoptosis induction, and decreased MGMT levels, suggesting the ability of drug combination to reduce the chemoresistance. This drug combination deserves attention and could be considered as a promising therapeutic strategy for GBM patients.

Keywords: COX-2, COX-2 inhibitor, glioblastoma, NS398, T98G, temozolomide

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Authors: John G. Skelly, Peter K. Dearden, Thomas W. R. Harrop, Sarah N. Inwood, Joseph Guhlin

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The Argentine stem weevil (ASW; L. bonariensis) is an economically important pasture pest in New Zealand, which causes about $200 million of damage per annum. Microctonus hyperodae (Mh), a parasite of the ASW in its natural range in South America, was introduced into New Zealand to curb the pasture damage caused by the ASW. Mh is an endoparasitic wasp that lays its eggs in the ASW halting its reproduction. Mh was initially successful at preventing ASW proliferation and reducing pasture damage. The effectiveness of Mh has since declined due to decreased parasitism rates and has resulted in increased pasture damage. Although the mechanism through which ASW has developed resistance to Mh has not been discovered, it has been proposed to be due to the different reproductive modes used by Mh and the ASW in New Zealand. The ASW reproduces sexually, whereas Mh reproduces asexually, which has been hypothesised to have allowed the ASW to ‘out evolve’ Mh. Other species within the Microctonus genus reproduce both sexually and asexually. Strains of Microctonus aethiopoides (Ma), a species closely related to Mh, reproduce either by sexual or asexual reproduction. Comparing the genomes of sexual and asexual Microctonus may allow for the identification of the mechanism of asexual reproduction and other characteristics that may improve Mh as a biocontrol agent. The genomes of Mh and three strains of Ma, two of which reproduce sexually and one reproduces asexually, have been sequenced and annotated. The French (MaFR) and Moroccan (MaMO) reproduce sexually, whereas the Irish strain (MaIR) reproduces asexually. Like Mh, The Ma strains are also used as biocontrol agents, but for different weevil species. The genomes of Mh and MaIR were subsequently upgraded using Hi-C, resulting in a set of high quality, highly contiguous genomes. A subset of the genes involved in mitosis and meiosis, which have been identified though the use of Hidden Markov Models generated from genes involved in these processes in other Hymenoptera, have been catalogued in Mh and the strains of Ma. Meiosis and mitosis genes were broadly conserved in both sexual and asexual Microctonus species. This implies that either the asexual species have retained a subset of the molecular components required for sexual reproduction or that the molecular mechanisms of mitosis and meiosis are different or differently regulated in Microctonus to other insect species in which these mechanisms are more broadly characterised. Bioinformatic analysis of the chemoreceptor compliment in Microctonus has revealed some variation in the number of olfactory receptors, which may be related to host preference. Phylogenetic analysis of olfactory receptors highlights variation, which may be able to explain different host range preferences in the Microctonus. Hi-C clustering implies that Mh has 12 chromosomes, and MaIR has 8. Hence there may be variation in gene regulation between species. Genome alignment of Mh and MaIR implies that there may be large scale genome structural variation. Greater insight into the genetics of these agriculturally important group of parasitic wasps may be beneficial in restoring or maintaining their biocontrol efficacy.

Keywords: argentine stem weevil, asexual, genomics, Microctonus hyperodae

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Authors: Thauane Silva, Gabrielle do Vale, Andre Ferreira, Marilda Siqueira, Thiago Moreno L. Souza, Milene D. Miranda

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The exposure of A(H1N1)pdm09-infected epithelial cells (HeLa) to HIV-1 viral particles, or its gp120, enhanced interferon-induced transmembrane protein (IFITM3) content, a viral restriction factor (RF), resulting in a decrease in influenza replication. The gp120 binds to CCR5 (R5) or CXCR4 (X4) cell receptors during HIV-1 infection. Then, it is possible that the endogenous ligands of these receptors also modulate the expression of IFITM3 and other cellular factors that restrict influenza virus replication. Thus, the aim of this study is to analyze the role of cellular receptors R5 and X4 in modulating RFs in order to inhibit the replication of the influenza virus. A549 cells were treated with 2x effective dose (ED50) of endogenous R5 or X4 receptor agonists, CCL3 (20 ng/ml), CCL4 (10 ng/ml), CCL5 (10 ng/ml) and CXCL12 (100 ng/mL) or exogenous agonists, gp120 Bal-R5, gp120 IIIB-X4 and its mutants (5 µg/mL). The interferon α (10 ng/mL) and oseltamivir (60 nM) were used as a control. After 24 h post agonists exposure, the cells were infected with virus influenza A(H3N2) at 2 MOI (multiplicity of infection) for 1 h. Then, 24 h post infection, the supernatant was harvested and, the viral titre was evaluated by qRT-PCR. To evaluate IFITM3 and SAM and HD domain containing deoxynucleoside triphosphate triphosphohydrolase 1 (SAMHD1) protein levels, A549 were exposed to agonists for 24 h, and the monolayer was lysed with Laemmli buffer for western blot (WB) assay or fixed for indirect immunofluorescence (IFI) assay. In addition to this, we analyzed other RFs modulation in A549, after 24 h post agonists exposure by customized RT² Profiler Polymerase Chain Reaction Array. We also performed a functional assay in which SAMHD1-knocked-down, by single-stranded RNA (siRNA), A549 cells were infected with A(H3N2). In addition, the cells were treated with guanosine to assess the regulatory role of dNTPs by SAMHD1. We found that R5 and X4 agonists inhibited influenza replication in 54 ± 9%. We observed a four-fold increase in SAMHD1 transcripts by RFs mRNA quantification panel. After 24 h post agonists exposure, we did not observe an increase in IFITM3 protein levels through WB or IFI assays, but we observed an upregulation up to three-fold in the protein content of SAMHD1, in A549 exposed to agonists. Besides this, influenza replication enhanced in 20% in cell cultures that SAMDH1 was knockdown. Guanosine treatment in cells exposed to R5 ligands further inhibited influenza virus replication, suggesting that the inhibitory mechanism may involve the activation of the SAMHD1 deoxynucleotide triphosphohydrolase activity. Thus, our data show for the first time a direct relationship of SAMHD1 and inhibition of influenza replication, and provides perspectives for new studies on the signaling modulation, through cellular receptors, to induce proteins of great importance in the control of relevant infections for public health.

Keywords: chemokine receptors, gp120, influenza, virus restriction factors

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