Search results for: live cells imaging
Commenced in January 2007
Frequency: Monthly
Edition: International
Paper Count: 5497

Search results for: live cells imaging

5197 Effect of Moringa Oleifera on Liveweight Reproductive Tract Dimention of Giant African Land Snail (Archachatina marginata)

Authors: J. A. Abiona, O. O. Fabinu, O. O. Ehimiyein, A. O. Ladokun, M. O. Abioja, J. O. Daramola, O. E. Oke, O. A. Osinowo, O. M. Onagbesan

Abstract:

A study was conducted on the effect of Moringa oleifera on liveweight and reproductive tract dimension of Giant African Land Snail (Archachatina marginata). Thirty two snails (32) with weight range of 100 – 150 g were used for this study. Eight snails (8) were subjected to each of the four treatments which were: Concentrate only, concentrate + 100g of Moringa oleifera, concentrate + 200g of Moringa oleifera and concentrate + 300g of Moringa oleifera. Parameters monitored were: Shell length, shell width, shell circumference and weekly live weight. Reproductive tract dimension taken include: Organ weight (ORGWT), reproductive tract weight (REPTWT), reproductive tract length (REPTLNT), ovo-tesis weight (OVOWT), edible part weight (EDPTWT), albumen weight (ALBWT) and albumen length (ALBLNT). Shell dimensions and the live weight were measured and recorded on a weekly basis with a tape rule and a sensitive weighing scale. After nine weeks, six snails were randomly selected from each treatment and dissected. Their reproductive tracts were removed and dimensions were taken. The result showed that ORGWT, OVOWT, ALBWT, ALBLNT, REPTLNT and REPTWT were not significantly affected (P>0.05) by different levels of Moringa oleifera inclusions with concentrate. However, Moringa oleifera inclusion with concentrate at different levels had significant effect (P<0.001) on Live weight, shell length and shell diameters of the animal. Snails given 300 g of Moringa oleifera per kilogramme of concentrate gave the highest live weight and shell length together with shell diameter. It was however recommended from this study that inclusion of Moringa oleifera leave meal into snail feed at 300 g per kg of concentrate would enhance live weight and shell parameters (length and width).

Keywords: reproductive tract, giant African land snails, Moringa oleifera, live weight, shell dimension

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5196 Prevalence of Down Syndrome: A Single-Center Study in Bandung, Indonesia

Authors: Bremmy Laksono, Riksa Parikrama, Nur A. Rosyada, Willyanti Soewondo, Dadang S. H. Effendi, Eriska Rianti, Arlette S. Setiawan, Ine Sasmita, Risti S. Primanti, Erna Kurnikasari, Yunia Sribudiani

Abstract:

Down syndrome (DS) is a chromosomal abnormality characterised by complete 21 chromosome trisomy (classical or non-disjunction), or partial 21 chromosome trisomy (mosaicism), or chromosome rearrangement involving chromosome 21 (translocation). This study was carried out to describe the frequency of DS patients in a research institution in the city of Bandung, Indonesia. This descriptive study also provides a picture of the residential location and surrounding area of their dwellings. This study involved people with DS in various age whose chromosome were evaluated by conventional karyotyping method and FISH. Data were collected from 60 patients with DS from a total 150 patients during the period of September 2015 to August 2016 who were referred to Cell Culture and Cytogenetics Laboratory, Faculty of Medicine Universitas Padjadjaran, Indonesia. Results showed that the most common type of DS was non-disjunction (93%), followed by mosaicism (5%), no patient with translocation DS (0%), and a very rare type of tetrasomy 21 (2%). There were 39 males (65%) and 21 females (35%) of DS patient. Most of them live in suburban area beyond Bandung city (55%) while the rest live inside urban area of Bandung city (45%). They live mostly in dense area of greater Bandung area (65%) and only a few live in mid-density area (25%) and the least live in sparse populated area (10%). Their houses are mostly located in residential estate area (55%), nearby industrial area (37%), and around agricultural area (8%). Based on the study, it could be concluded that non-disjunction DS is the most common type. DS patients referred to the laboratory mostly came from dense residential zone in suburban area outside Bandung city. The low number of DS patients referred to the laboratory for chromosome analysis was the highlight to improve health service for people with genetic disorder. This study offered several information regarding area of DS patients’ residence and the condition of neighbourhood in Bandung city where they live as well.

Keywords: chromosome, descriptive, Down syndrome, prevalence

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5195 The Role of Immunologic Diamonds in Dealing with Mycobacterium Tuberculosis; Responses of Immune Cells in Affliction to the Respiratory Tuberculosis

Authors: Seyyed Mohammad Amin Mousavi Sagharchi, Elham Javanroudi

Abstract:

Introduction: Tuberculosis (TB) is a known disease with hidden features caused by Mycobacterium tuberculosis (MTB). This disease, which is one of the 10 deadliest in the world, has caused millions of deaths in recent decades. Furthermore, TB is responsible for infecting about 30% population of world. Like any infection, TB can activate the immune system by locating and colonization in the human body, especially in the alveoli. TB is granulomatosis, so MTB can absorb the host’s immune cells and other cells to form granuloma. Method: Different databases (e.g., PubMed) were recruited to prepare this paper and fulfill our goals to search and find effective papers and investigations. Results: Immune response to MTB is related to T cell killers and contains CD1, CD4, and CD8 T lymphocytes. CD1 lymphocytes can recognize glycolipids, which highly exist in the Mycobacterial fatty cell wall. CD4 lymphocytes and macrophages form granuloma, and it is the main line of immune response to Mycobacteria. On the other hand, CD8 cells have cytolytic function for directly killing MTB by secretion of granulysin. Other functions and secretion to the deal are interleukin-12 (IL-12) by induction of expression interferon-γ (INF-γ) for macrophages activation and creating a granuloma, and tumor necrosis factor (TNF) by promoting macrophage phagolysosomal fusion. Conclusion: Immune cells in battle with MTB are macrophages, dendritic cells (DCs), neutrophils, and natural killer (NK) cells. These immune cells can recognize the Mycobacterium by various receptors, including Toll-like receptors (TLRs), Nod-like receptors (NLRs), and C-type lectin receptors (CLRs) located in the cell surface. In human alveoli exist about 50 dendritic macrophages, which have close communication with other immune cells in the circulating system and epithelial cells to deal with Mycobacteria. Against immune cells, MTB handles some factors (e.g., cordfactor, O-Ag, lipoarabinomannan, sulfatides, and adenylate cyclase) and practical functions (e.g., inhibition of macrophages).

Keywords: mycobacterium tuberculosis, immune responses, immunological mechanisms, respiratory tuberculosis

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5194 A Prospective Study on the Efficacy of Mesenchymal Stem Cells in Intervertebral Disc Regeneration

Authors: Prabhu Thangaraju, Manoj Deepak, A. Sivakumar

Abstract:

Removal of inter vertebral disc along with spinal fusion has many disadvantages such as causing stress fractures. If it is possible regenerate the spine it would be possible avoid the complications of the surgery and achieve better results. Our study involves the use of mesenchymal stem cells in regenerating the discs. Our study involved 10 patients who presented with degenerative disc disease between 2008-2011 in our hospital. After adequate pre-operative check prepared mesenchymal stem cells were injected into the disc spaces. These patients were subjected to conservative therapy for a minimum of six weeks before they were accepted into the study. They were followed up regularly for a minimum of 2years with serial radiographs and MRI. 8 out of the 10 patients had completed reduction in the pain. The T2 weighted MRI images in 9 out of the 10 patients showed a bright signal compared the previous Images which indicated that there was improvement in the hydration levels. From the case study of 10 patients who were subjected to mesenchymal cell therapy in our hospital, we can conclude that the use of mesenchymal cells in treatment of intervertebral disc degeneration in a safe and effective option.

Keywords: mesenchymal stem cells, intervertebral disc, the spine, disc degeneration

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5193 Exploring the Success of Live Streaming Commerce in China: A Literature Analysis

Authors: Ming Gao, Matthew Tingchi Liu, Hoi Ngan Loi

Abstract:

Live streaming refers to the video contents generated by broadcasters and shared with viewers in real-time by uploading them to short-video platforms. In recent years, individual KOL broadcasters have successfully made use of live streams to sell a large amount of goods to the consumers. For example, Wei Ya, the Number 1 broadcaster in Taobao Live, sold products worth RMB 2.7 billion (USD 0.38 billion) in 2018. Regarding the success of live streaming commerce (LSC) in China, this study explores the elements of the booming LSC industry and attempts to explain the reasons behind its prosperity. A systematic review of industry reports and academic papers was conducted to summarize the latest findings in this field. And the results of this investigation showed that a live streaming eco-system has been established by the LSC players, namely, the platform, the broadcaster, the product supplier, and the viewer. In this eco-system, all players have complementary advantages and needs, and their close cooperation leads to a win-win situation. For instance, platforms and broadcasters have abundant internet traffic, which needs to be monetized, while product suppliers have mature supply chains and the need of promoting the products. In addition, viewers are attached to the LSC platforms to get product information, bargains, and entertainment. This study highlights the importance of the mass-personal hybrid communication nature of live streaming because its interpersonal communication feature increases consumers’ positive experiences, while its mass media broadcasting feature facilitates product promotion. Another innovative point of this study lies in its inclusion of the special characteristic of Chinese Internet culture - entertainment. The entertaining genres of the live streams created by broadcasters serve as down-to-earth approaches to reach their audiences easily. Further, the nature of video, i.e., the dynamic and salient stimulus, is emphasized in this study. Since video is more engaging, it can attract viewers in a quick and easy way. Meanwhile, the abundant, interesting, high-quality, and free short videos have added “stickiness” to platforms by retaining users and prolonging their staying time on the platforms. In addition, broadcasters’ important characters, such as physical attractiveness, humor, sex appeal, kindness, communication skills, and interactivity, are also identified as important factors that influence consumers’ engagement and purchase intention. In conclusion, all players have their own proper places in this live streaming eco-system, in which they work seamlessly to give full play to their respective advantages, with each player taking what it needs and offering what it has. This has contributed to the success of live streaming commerce in China.

Keywords: broadcasters, communication, entertainment, live streaming commerce, viewers

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5192 Advantages of Multispectral Imaging for Accurate Gas Temperature Profile Retrieval from Fire Combustion Reactions

Authors: Jean-Philippe Gagnon, Benjamin Saute, Stéphane Boubanga-Tombet

Abstract:

Infrared thermal imaging is used for a wide range of applications, especially in the combustion domain. However, it is well known that most combustion gases such as carbon dioxide (CO₂), water vapor (H₂O), and carbon monoxide (CO) selectively absorb/emit infrared radiation at discrete energies, i.e., over a very narrow spectral range. Therefore, temperature profiles of most combustion processes derived from conventional broadband imaging are inaccurate without prior knowledge or assumptions about the spectral emissivity properties of the combustion gases. Using spectral filters allows estimating these critical emissivity parameters in addition to providing selectivity regarding the chemical nature of the combustion gases. However, due to the turbulent nature of most flames, it is crucial that such information be obtained without sacrificing temporal resolution. For this reason, Telops has developed a time-resolved multispectral imaging system which combines a high-performance broadband camera synchronized with a rotating spectral filter wheel. In order to illustrate the benefits of using this system to characterize combustion experiments, measurements were carried out using a Telops MS-IR MW on a very simple combustion system: a wood fire. The temperature profiles calculated using the spectral information from the different channels were compared with corresponding temperature profiles obtained with conventional broadband imaging. The results illustrate the benefits of the Telops MS-IR cameras for the characterization of laminar and turbulent combustion systems at a high temporal resolution.

Keywords: infrared, multispectral, fire, broadband, gas temperature, IR camera

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5191 Biodegradation of Chlorpyrifos in Real Wastewater by Acromobacter xylosoxidans SRK5 Immobilized in Calcium Alginate

Authors: Saira Khalid, Imran Hashmi

Abstract:

Agrochemical industries produce huge amount of wastewater containing pesticides and other harmful residues. Environmental regulations make it compulsory to bring pesticides to a minimum level before releasing wastewater from industrial units.The present study was designed with the objective to investigate biodegradation of CP in real wastewater using bacterial cells immobilized in calcium alginate. Bacterial strain identified as Acromobacter xylosoxidans SRK5 (KT013092) using 16S rRNA nucleotide sequence analysis was used. SRK5 was immobilized in calcium alginate to make calcium alginate microspheres (CAMs). Real wastewater from industry having 50 mg L⁻¹ of CP was inoculated with free cells or CAMs and incubated for 96 h at 37˚C. CP removal efficiency with CAMs was 98% after 72 h of incubation, and no lag phase was observed. With free cells, 12h of lag phase was observed. After 96 h of incubation 87% of CP removal was observed when inoculated with free cells. No adsorption was observed on vacant CAMs. Phytotoxicity assay demonstrated considerable loss in toxicity. Almost complete COD removal was achieved at 96 h with CAMs. Study suggests the use of immobilized cells of SRK5 for bioaugmentation of industrial wastewater for CP degradation instead of free cells.

Keywords: biodegradation, chlorpyrifos, immobilization, wastewater

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5190 Comparison of the Classification of Cystic Renal Lesions Using the Bosniak Classification System with Contrast Enhanced Ultrasound and Magnetic Resonance Imaging to Computed Tomography: A Prospective Study

Authors: Dechen Tshering Vogel, Johannes T. Heverhagen, Bernard Kiss, Spyridon Arampatzis

Abstract:

In addition to computed tomography (CT), contrast enhanced ultrasound (CEUS), and magnetic resonance imaging (MRI) are being increasingly used for imaging of renal lesions. The aim of this prospective study was to compare the classification of complex cystic renal lesions using the Bosniak classification with CEUS and MRI to CT. Forty-eight patients with 65 cystic renal lesions were included in this study. All participants signed written informed consent. The agreement between the Bosniak classifications of complex renal lesions ( ≥ BII-F) on CEUS and MRI were compared to that of CT and were tested using Cohen’s Kappa. Sensitivity, specificity, positive and negative predictive values (PPV/NPV) and the accuracy of CEUS and MRI compared to CT in the detection of complex renal lesions were calculated. Twenty-nine (45%) out of 65 cystic renal lesions were classified as complex using CT. The agreement between CEUS and CT in the classification of complex cysts was fair (agreement 50.8%, Kappa 0.31), and was excellent between MRI and CT (agreement 93.9%, Kappa 0.88). Compared to CT, MRI had a sensitivity of 96.6%, specificity of 91.7%, a PPV of 54.7%, and an NPV of 54.7% with an accuracy of 63.1%. The corresponding values for CEUS were sensitivity 100.0%, specificity 33.3%, PPV 90.3%, and NPV 97.1% with an accuracy 93.8%. The classification of complex renal cysts based on MRI and CT scans correlated well, and MRI can be used instead of CT for this purpose. CEUS can exclude complex lesions, but due to higher sensitivity, cystic lesions tend to be upgraded. However, it is useful for initial imaging, for follow up of lesions and in those patients with contraindications to CT and MRI.

Keywords: Bosniak classification, computed tomography, contrast enhanced ultrasound, cystic renal lesions, magnetic resonance imaging

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5189 Regression for Doubly Inflated Multivariate Poisson Distributions

Authors: Ishapathik Das, Sumen Sen, N. Rao Chaganty, Pooja Sengupta

Abstract:

Dependent multivariate count data occur in several research studies. These data can be modeled by a multivariate Poisson or Negative binomial distribution constructed using copulas. However, when some of the counts are inflated, that is, the number of observations in some cells are much larger than other cells, then the copula based multivariate Poisson (or Negative binomial) distribution may not fit well and it is not an appropriate statistical model for the data. There is a need to modify or adjust the multivariate distribution to account for the inflated frequencies. In this article, we consider the situation where the frequencies of two cells are higher compared to the other cells, and develop a doubly inflated multivariate Poisson distribution function using multivariate Gaussian copula. We also discuss procedures for regression on covariates for the doubly inflated multivariate count data. For illustrating the proposed methodologies, we present a real data containing bivariate count observations with inflations in two cells. Several models and linear predictors with log link functions are considered, and we discuss maximum likelihood estimation to estimate unknown parameters of the models.

Keywords: copula, Gaussian copula, multivariate distributions, inflated distributios

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5188 Indoleamine 2,3 Dioxygenase and Regulatory T Cells in Acute Myeloid Leukemia

Authors: Iman M. Mansour, Rania A. Zayed, Fadwa S. Abdel-Azim, Lamyaa H. Abdel-Latif

Abstract:

Background and Objectives: The microenvironment of acute myeloid leukemia (AML) is suppressive for immune cells. Regulatory T cells (Tregs) have been recognized to play a role in helping leukemic cells to evade immunesurveillance. The mesenchymal stem cells (MSCs) are essential contributors in immunomodulation of the microenvironment as they can promote differentiation of Tregs via the indoleamine 2,3-dioxygenase (IDO) pathway. The aim of the present work was to evaluate the expression of IDO in bone marrow derived MSCs and to study its correlation to percentage of Tregs. Methods: 37 adult bone marrow samples were cultured in appropriate culture medium to isolate MSCs. Successful harvest of MSCs was determined by plastic adherence, morphology and positive expression of CD271 and CD105; negative expression of CD34 and CD45 using flowcytometry. MSCs were examined for IDO expression by immunocytochemistry using anti-IDO monoclonal antibody. CD4+ CD25+ cells (Tregs) were measured in bone marrow samples by flowcytometry. Results: MSCs were successfully isolated from 20 of the 37 bone marrow samples cultured. MSCs showed higher expression of IDO and Tregs percentage was higher in AML patients compared to control subjects (p=0.002 and p<0.001 respectively). A positive correlation was found between IDO expression and Tregs percentage (p value=0.012, r=0.5). Conclusion: In this study, we revealed an association between high IDO expression in MSCs and elevated levels of Tregs which has an important role in the pathogenesis of AML, providing immunosuppressive microenvironment.

Keywords: acute myeloid leukemia, indoleamine 2, 3-dioxygenase, mesenchymal stem cells, T regulatory cells

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5187 BSYJ Promoting Homing and Differentiation of Mesenchymal Stem Cells at the Retina of Age-Related Macular Degeneration Model Mice Induced by Sodium Iodate

Authors: Lina Liang, Kai Xu, Jing Zhang

Abstract:

Purpose: Age-related macular degeneration (AMD) is a major leading cause of visual impairment and blindness with no cure currently established. Cell replacement is discussed as a potential therapy for AMD. Besides intravitreal injection and subretinal injection, intravenous administration has been explored as an alternative route. This study is to observe the effect of BSYJ, a traditional Chinese medicine on the homing and differentiation of mesenchymal stem cells transplanted via tail vein injection in an age-related macular degeneration mouse model. Methods: Four-week-old C57BL/6J mice were injected with 40 mg/kg NaIO₃ to induce age-related macular degeneration model. At the second day after NaIO₃ injection, 1×10⁷ GFP labeled bone marrow-derived mesenchymal stem cells (GFP-MSCs) were transplanted via tali vein injection into the experimental mice. Then the mice were randomly divided into two groups, gavaged with either BSYJ solution (BSYJ group, n=12) or distilled water (DW group, n=12). 12 age-matched healthy C57BL/6J mice were fed regularly as normal control. At day 7, day 14, and day 28 after treatment, retina flat mounting was used to detect the homing of mesenchymal stem cells at the retina. Double-labeling immunofluorescence was used to determine the differentiation of mesenchymal stem cells. Results: At 7, 14, 28 days after treatment, the numbers of GFP-MSCs detected by retina flatmount were 10.2 ± 2.5, 14.5 ± 3.4 and 18.7 ± 5.8, respectively in the distilled water group, while 15.7 ± 3.8, 32.3 ± 3.5 and 77.3 ± 6.4 in BSYJ group, the differences between the two groups were significant (p < 0.05). At 28 days after treatment, it was shown by double staining immunofluorescence that there were more GFP positive cells in the retina of BSYJ group than that of the DW group, but none of the cells expressed RPE specific genes such as RPE65 and CRALBP, or photoreceptor genes such as recoverin and rhodopsin either in BSYJ group or DW group. However, GFAP positive cells were found among the cells labeled with GFP, and the double labeling cells were much more in the BSYJ group than the distilled water group. Conclusion: BSYJ could promote homing of mesenchymal stem cells at the retina of age-related macular degeneration model mice induced by NaIO₃, and the differentiation towards to glial cells. Acknowledgement: National Natural Foundation of China (No: 81473736, 81674033,81973912).

Keywords: BSYJ, differentiation, homing, mesenchymal stem cells

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5186 Digital Phase Shifting Holography in a Non-Linear Interferometer using Undetected Photons

Authors: Sebastian Töpfer, Marta Gilaberte Basset, Jorge Fuenzalida, Fabian Steinlechner, Juan P. Torres, Markus Gräfe

Abstract:

This work introduces a combination of digital phase-shifting holography with a non-linear interferometer using undetected photons. Non-linear interferometers can be used in combination with a measurement scheme called quantum imaging with undetected photons, which allows for the separation of the wavelengths used for sampling an object and detecting it in the imaging sensor. This method recently faced increasing attention, as it allows to use of exotic wavelengths (e.g., mid-infrared, ultraviolet) for object interaction while at the same time keeping the detection in spectral areas with highly developed, comparable low-cost imaging sensors. The object information, including its transmission and phase influence, is recorded in the form of an interferometric pattern. To collect these, this work combines the method of quantum imaging with undetected photons with digital phase-shifting holography with a minimal sampling of the interference. With this, the quantum imaging scheme gets extended in its measurement capabilities and brings it one step closer to application. Quantum imaging with undetected photons uses correlated photons generated by spontaneous parametric down-conversion in a non-linear interferometer to create indistinguishable photon pairs, which leads to an effect called induced coherence without induced emission. Placing an object inside changes the interferometric pattern depending on the object’s properties. Digital phase-shifting holography records multiple images of the interference with determined phase shifts to reconstruct the complete interference shape, which can afterward be used to analyze the changes introduced by the object and conclude its properties. An extensive characterization of this method was done using a proof-of-principle setup. The measured spatial resolution, phase accuracy, and transmission accuracy are compared for different combinations of camera exposure times and the number of interference sampling steps. The current limits of this method are shown to allow further improvements. To summarize, this work presents an alternative holographic measurement method using non-linear interferometers in combination with quantum imaging to enable new ways of measuring and motivating continuing research.

Keywords: digital holography, quantum imaging, quantum holography, quantum metrology

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5185 Synergistic Anti-Proliferation Effect of PLK-1 Inhibitor and Livistona Chinensis Fruit Extracts on Lung Adenocarcinoma A549 Cells

Authors: Min-Chien Su, Tzu-Hsuan Hsu, Guan-Xuan Wu, Shyh-Ming Kuo

Abstract:

Lung cancer is one of the clinically challenging malignant diseases worldwide. For efficient therapeutics in cancer, combination therapy has developed to acquire a better outcome. PLK-1 was one of the major factors affecting cell mitosis in cancer cells, its inhibitor Bi6727 was proven effective in treating several different cancers namely oral cancer, colon cancer and lung cancer. Despite its low toxicity toward normal cells compared to traditional chemotherapy, it is still yet to be evaluated in detail. Livistona Chinensis (LC) is a Chinese herb that used as a traditional prescription to treat lung cancer. Due to the uncertainty of the efficacy of LC, we utilized a water extraction method to extract the Livistona Chinensis and then lyophilized into powder for further study. In this study we investigated the antiproliferation activities of Bi6727 and LC extracts (LCE) on A549 non-small lung cancer cells. The IC50 of Bi6727 and LCE on A549 are 60 nM and 0.8 mg/mL, respectively. The fluorescent staining images shown nucleolus damage in cells treated with Bi6727 and mitochondrial damage after treated with LCE. A549 cells treated with Bi6727 and LCE showed increased expression of Bax, Caspase-3 and Caspase-9 proteins from Western blot assay. LCE also inhibited A549 cells growth keeping cells at G2-M phase from cell cycle assay. Apoptosis assay results showed that LCE induced late apoptosis of A549 cells. JC-1 assay showed that the mitochondria damaged at the LCE concentration of 0.4 mg/mL. In our preliminary anti-proliferation test of combined LCE and Bi-6727 on A549 cells, we found a dramatically decrease in proliferation after treated with LCE first for 24-h and then Bi-6727 for extra 24-h. This was an important finding regarding synergistic anti-proliferation effect of these drugs, However, the usage, the application sequence of LCE and Bi-6727 on A549 cells and their related mechanisms still need to be evaluated. In summary, the drugs exerted anti-proliferation effect on A549 cells independently. We hopefully combine the usage of these two drugs will bring a different and potential outcome in treating lung cancer.

Keywords: anti-proliferation, A549, Livistona Chinensis fruit extracts, PLK-1 inhibitor

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5184 Intensive Crosstalk between Autophagy and Intracellular Signaling Regulates Osteosarcoma Cell Survival Response under Cisplatin Stress

Authors: Jyothi Nagraj, Sudeshna Mukherjee, Rajdeep Chowdhury

Abstract:

Autophagy has recently been linked with cancer cell survival post drug insult contributing to acquisition of resistance. However, the molecular signaling governing autophagic survival response is poorly explored. In our study, in osteosarcoma (OS) cells cisplatin shock was found to activate both MAPK and autophagy signaling. An activation of JNK and autophagy acted as pro-survival strategy, while ERK1/2 triggered apoptotic signals upon cisplatin stress. An increased sensitivity of the cells to cisplatin was obtained with simultaneous inhibition of both autophagy and JNK pathway. Furthermore, we observed that the autophagic stimulation upon drug stress regulates other developmentally active signaling pathways like the Hippo pathway in OS cells. Cisplatin resistant cells were thereafter developed by repetitive drug exposure followed by clonal selection. Basal levels of autophagy were found to be high in resistant cells to. However, the signaling mechanism leading to autophagic up-regulation and its regulatory effect differed in OS cells upon attaining drug resistance. Our results provide valuable clues to regulatory dynamics of autophagy that can be considered for development of improved therapeutic strategy against resistant type cancers.

Keywords: JNK, autophagy, drug resistance, cancer

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5183 Neighbourhood Design for Independent Living of Adults with Intellectual Disability

Authors: Cate MacMillan, Nicholas J. Stevens, Johanna Rosier, Steven Boyd

Abstract:

Choosing where to live is an important decision for anybody, however, this decision is more complex if you are an adult with intellectual disability. Our research asked adults with intellectual disability, parents and carers and disability, housing and built environment decision makers what they considered important in deciding where to live. If medical advances continue to improve the longevity of adults with intellectual disability, many of these adults will outlive their parents. With appropriate community support, and in appropriately designed neighbourhoods, many will be able to live independently. Our research suggests that the key to achieving independent living as an adult with intellectual disability is not so much about the house but the type of neighbourhood and its design. This paper presents the results of interviews and details a practical approach which will better inform urban development decision-makers in establishing safe, inclusive and accessible neighbourhood design.

Keywords: inclusion, independent living, intellectual disability, neighbourhoods, systems thinking, urban design and planning

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5182 Fluorescence Gold Nanoparticles: Sensing Properties and Cytotoxicity Studies in MCF-7 Human Breast Cancer Cells

Authors: Cristina Núñez, Rufina Bastida, Elena Labisbal, Alejandro Macías, María T. Pereira, José M. Vila

Abstract:

A highly selective quinoline-based fluorescent sensor L was designed in order to functionalize gold nanoparticles (GNPs@L). The cytotoxicity of compound L and GNPs@L on the MCF-7 breast cancer cells was explored and it was observed that L and GNPs@L compounds induced apoptosis in MCF-7 cancer cells. The cellular uptake of the hybrid system GNPs@L was studied using confocal laser scanning microscopy (CLSM).

Keywords: cytotoxicity, fluorescent probes, nanoparticles, quinoline

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5181 A Comparative Study for Various Techniques Using WEKA for Red Blood Cells Classification

Authors: Jameela Ali, Hamid A. Jalab, Loay E. George, Abdul Rahim Ahmad, Azizah Suliman, Karim Al-Jashamy

Abstract:

Red blood cells (RBC) are the most common types of blood cells and are the most intensively studied in cell biology. The lack of RBCs is a condition in which the amount of hemoglobin level is lower than normal and is referred to as “anemia”. Abnormalities in RBCs will affect the exchange of oxygen. This paper presents a comparative study for various techniques for classifyig the red blood cells as normal, or abnormal (anemic) using WEKA. WEKA is an open source consists of different machine learning algorithms for data mining applications. The algorithm tested are Radial Basis Function neural network, Support vector machine, and K-Nearest Neighbors algorithm. Two sets of combined features were utilized for classification of blood cells images. The first set, exclusively consist of geometrical features, was used to identify whether the tested blood cell has a spherical shape or non-spherical cells. While the second set, consist mainly of textural features was used to recognize the types of the spherical cells. We have provided an evaluation based on applying these classification methods to our RBCs image dataset which were obtained from Serdang Hospital-Malaysia, and measuring the accuracy of test results. The best achieved classification rates are 97%, 98%, and 79% for Support vector machines, Radial Basis Function neural network, and K-Nearest Neighbors algorithm respectively

Keywords: red blood cells, classification, radial basis function neural networks, suport vector machine, k-nearest neighbors algorithm

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5180 Wound Healing Potential and Comparison of Mummy Substance Effect on Adipose and Wharton’s Jelly-Derived Mesenchymal Stem Cells Co-Cultured with Human Fibroblast

Authors: Sepideh Hassanpour Khodaei

Abstract:

Background/Objectives: The purpose of this study is to evaluate the effect of mummy substances on two issues of proliferation and production of matrix protein synthesis in wound healing. Methods: The methodology used for this aim involves isolating mesenchymal stem cells and human fibroblasts procured at Pastor Institute, Iran. The cells were treated with mummy substances separately and co-cultured between ASCs and WJSCs, and fibroblasts. Proliferation was assessed by Ki67 method in monolayer conditions. Synthesis of components of extracellular matrix (ECM) such as collagen type I, type III, and fibronectin 1 (FN1) was determined by qPCR. Results: The effects of adipocyte stem cells (ASCs), Wharton Jelly Stem Cells (WJSCs), and Mummy material on fibroblast proliferation and migration were evaluated. The present finding underlined the importance of Mummy material, ASCs, and WJSCs in the proliferation and migration of fibroblast cells. Furthermore, the expression of collagen I, III, and FN1 was increased in the presence of the above material and cells. Conclusion: This study presented an effective in vitro method for the healing process. Hence, the prospect of utilizing Mummy material and stem cell-based therapies in wound healing as a therapeutic approach is promising.

Keywords: mummy material, wound healing, adipose tissue, Wharton’s jelly

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5179 Normal Hematopoietic Stem Cell and the Toxic Effect of Parthenolide

Authors: Alsulami H., Alghamdi N., Alasker A., Almohen N., Shome D.

Abstract:

Most conventional chemotherapeutic agents which are used for the treatment of cancers not only eradicate cancer cells but also affect normal hematopoietic Stem cells (HSCs) that leads to severe pancytopenia during treatment. Therefore, a need exists for novel approaches to treat cancer without or with minimum effect on normal HSCs. Parthenolide (PTL), a herbal product occurring naturally in the plant Feverfew, is a potential new chemotherapeutic agent for the treatment of many cancers such as acute myeloid leukemia (AML) and chronic lymphocytic leukemia (CLL). In this study we investigated the effect of different PTL concentrations on the viability of normal HSCs and also on the ability of these cells to form colonies after they have been treated with PTL in vitro. Methods: In this study, 24 samples of bone marrow and cord blood were collected with consent, and mononuclear cells were separated using density gradient separation. These cells were then exposed to various concentrations of PTL for 24 hours. Cell viability after culture was determined using 7ADD in a flow cytometry test. Additionally, the impact of PTL on hematopoietic stem cells (HSCs) was evaluated using a colony forming unit assay (CFU). Furthermore, the levels of NFҝB expression were assessed by using a PE-labelled anti-pNFκBP65 antibody. Results: this study showed that there was no statistically significant difference in the percentage of cell death between untreated and PTL treated cells with 5 μM PTL (p = 0.7), 10 μM PTL (p = 0.4) and 25 μM (p = 0.09) respectively. However, at higher doses, PTL caused significant increase in the percentage of cell death. These results were significant when compared to untreated control (p < 0.001). The response of cord blood cells (n=4) on the other hand was slightly different from that for bone marrow cells in that the percentage of cell death was significant at 100 μM PTL. Therefore, cord blood cells seemed more resistant than bone marrow cells. Discussion &Conclusion: At concentrations ≤25 μM PTL has a minimum or no effect on HSCs in vitro. Cord blood HSCs are more resistant to PTL compared to bone marrow HSCs. This could be due to the higher percentage of T-lymphocytes, which are resistant to PTL, in CB samples (85% in CB vs. 56% in BM. Additionally, CB samples contained a higher proportion of CD34+ cells, with 14.5% of brightly CD34+ cells compared to only 1% in normal BM. These bright CD34+ cells in CB were mostly negative for early-stage stem cell maturation antigens, making them young and resilient to oxidative stress and high concentrations of PTL.

Keywords: stem cell, parthenolide, NFKB, CLL

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5178 Conformal Coating Technology Applicable to Cell Therapeutics Using Click-Reactive Biocompatible Polymers

Authors: Venkat Garigapati

Abstract:

Cell-based therapies are limited due to underlying host immune system activity. Microencapsulation of living cells to overcome this issue has some serious drawbacks, such as limitations of nutrient and oxygen diffusion, which pose a threat to the function and longevity of cells. The conformal coating could overcome the issues which are generally involved in traditional microencapsulation. Some of the theoretical advantages of conformal coating include superior nutrient and oxygen supply to cells, prolonged lifespan, improved drug-secreting cell functionality and an opportunity to load high cell doses in small volumes. Despite several advantages to the conformal coating, there are no suitable methods available to apply to living cells. The ultra-thin conformal coating was achieved utilizing click-reactive methacryloyloxyethyl phosphorylcholine (MPC) polymers, which are capable of specifically reacting one polymer to another at neutral pH in the aqueous isotonic system at the desired temperature suitable for living cells without the need of deleterious initiators. ARPE-19 (Adult Retinal Pigment Epithelial cell line-19) cell-spheroids and rat pancreatic islets were used in the formulation studies. The in vitro studies of coated ARPE-19 cell-spheroids and rat islets indicate that the coat was intact; cells were viable and functioning. The in vitro study results revealed that the conformal coating technology seems promising and in vivo studies are being planned.

Keywords: cells, hydrogel, conformal coating, microencapsulation, insulin

Procedia PDF Downloads 91
5177 Analysis of Selected Hematological Variables during Three Different Menstrual Phases between Sedentary and Sports Women

Authors: G. Vasanthi

Abstract:

The purpose of the study was to analyse the red blood cells and white blood cells during three different menstrual phases between sedentary and sports women. To achieve this purpose, fifteen female sedentary post graduate students (M.A., M.Sc.) and fifteen students of Master of Physical Education and Sports (M.P.Ed.) women who regularly involved in vigouous sports training and participated in sports competition on different games were selected by adopting random sampling method. All the students were hostelers and their age group was between 20 to 22 years. The blood sample were collected during the mid-period of the three different phases to calculate the red blood cells and white blood cells. The data collected were treated statistically by using analysis of variance. The results reveal that the RBC and WBC is found to be significant between sedentary and sports women during the three different menstrual phases.

Keywords: RBC, WBC, menstrual, proliferative, secretary, sedentary women, sports women

Procedia PDF Downloads 504
5176 Design and Implementation of a Lab Bench for Synthetic Aperture Radar Imaging System

Authors: Karthiyayini Nagarajan, P. V. RamaKrishna

Abstract:

Radar Imaging techniques provides extensive applications in the field of remote sensing, majorly Synthetic Aperture Radar(SAR) that provide high resolution target images. This paper work puts forward the effective and realizable signal generation and processing for SAR images. The major units in the system include camera, signal generation unit, signal processing unit and display screen. The real radio channel is replaced by its mathematical model based on optical image to calculate a reflected signal model in real time. Signal generation realizes the algorithm and forms the radar reflection model. Signal processing unit provides range and azimuth resolution through matched filtering and spectrum analysis procedure to form radar image on the display screen. The restored image has the same quality as that of the optical image. This SAR imaging system has been designed and implemented using MATLAB and Quartus II tools on Stratix III device as a System(lab bench) that works in real time to study/investigate on radar imaging rudiments and signal processing scheme for educational and research purposes.

Keywords: synthetic aperture radar, radio reflection model, lab bench

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5175 Enhancing Sensitization of Cervical Cancer Cells to γ-Radiation Ellagic Acid

Authors: Vidhula Ahire, Amit Kumar, K. P. Mishra, Gauri Kulkarni

Abstract:

Herbal polyphenols have gained significance because of their increasing promise in prevention and treatment of cancer. Therefore, development of a dietary compound as an effective radiosensitizer and a radioprotector is highly warranted for cervical cancer patients undergoing therapy. This study describes the cytotoxic effects of the flavonoid, ellagic acid (EA) when administered either alone or in combination with gamma radiation on cervical cancer HeLa cells in vitro. Apoptotic index and proliferation were measured by using trypan blue assay. Reproductive cell death was analyzed by clonogenic assay. Propidium iodide staining for flowcytometry was performed to analyze cell cycle modulation. Nuclear and mitochondrial changes were studied with specific dyes. DNA repair kinetics was analyzed by immunofluorescence assay. Evaluation and comparison of EA effects were performed with other clinically used breast cancer drugs. When tumor cells were exposed to 2 and 4 Gy of irradiation in presence of EA (10 μM), it yielded a synergistic cytotoxic effect on cervical cancer cells whereas in NIH3T3 cells it reversed the injury caused by irradiation and abetted in the regaining of normal healthy cells. At 24h ~25foci/cell was observed and 2.6 fold decrease in the mitochondrial membrane potential. Up to 40% cell were arrested in the G1 phase and 20-36% cells exhibited apoptosis. Our results demonstrate the role of increased apoptosis and cell cycle modulation in the mechanism of EA mediated radiosensitization of cervical cancer cells and thus advocating EA as an adjuvant for preclinical trials in cancer chemo- radiotherapy.

Keywords: cervical cancer, ellagic acid, sensitization, radiation therapy

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5174 Stem Cell Differentiation Toward Secretory Progenitors after Intestinal Ischemia-Reperfusion in a Rat is Accompanied by Inhibited Notch Signaling Cascade

Authors: Igor Sukhotnik

Abstract:

Objectives: Notch signaling is thought to act to drive cell versification in the lining of the small intestine. When Notch signaling is blocked, proliferation ceases, and epithelial cells become secretory. The purpose of the present study was to evaluate the role of Notch signaling pathway in stem cell differentiation in a rat model of intestinal ischemia-reperfusion (IR). Methods: Male Sprague-Dawley rats were randomly divided into four experimental groups: Sham-24 and Sham-48 rats underwent laparotomy and were killed 24 or 48 h later, respectively; IR-24 and IR-48 rats underwent occlusion of SMA and portal vein for 30 min followed by 24 or 48 h of reperfusion, respectively. Notch-related gene and protein expression were determined using Real Time PCR, Western blotting and immunohistochemistry. Wax histology and immunohistochemistry was used to determine cell differentiation toward absorptive (enterocytes) or secretory progenitors (goblet cells, enteroendocrine cells or Paneth cells). Results: IR-48 rats exhibited a significant decrease in Notch-1 protein expression (Western blot) that was coincided with a significant decrease in the number of Notch-1 positive cells (immunohistochemistry) in jejunum and ileum as well as Hes-1 positive cells in jejunum and ileum compared to Sham-48 rats. A significant down-regulation of Notch signaling related genes and proteins in IR animals was accompanied by a significant increase in the number of goblet and Paneth cells and decreased number of absorptive cells compared to control rats. Conclusions: Forty-eight hours following intestinal IR in rats, inhibited Notch signaling pathway was accompanied by intestinal stem cells differentiation toward secretory progenitors.

Keywords: Intestine, notch, ischemia-reperfusion, cell differentiation, secretory

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5173 Enhancing Power Conversion Efficiency of P3HT/PCBM Polymer Solar Cells

Authors: Nidal H. Abu-Zahra, Mahmoud Algazzar

Abstract:

In this research, n-dodecylthiol was added to P3HT/PC70BM polymer solar cells to improve the crystallinity of P3HT and enhance the phase separation of P3HT/PC70BM. The improved crystallinity of P3HT/PC70BM doped with 0-5% by volume of n-dodecylthiol resulted in improving the power conversion efficiency of polymer solar cells by 33%. In addition, thermal annealing of the P3HT/PC70MB/n-dodecylthiolcompound showed further improvement in crystallinity with n-dodecylthiol concentration up to 2%. The highest power conversion efficiency of 3.21% was achieved with polymer crystallites size L of 11.2nm, after annealing at 150°C for 30 minutes under a vacuum atmosphere. The smaller crystallite size suggests a shorter path of the charge carriers between P3HT backbones, which could be beneficial to getting a higher short circuit current in the devices made with the additive.

Keywords: n-dodecylthiol, congugated PSC, P3HT/PCBM, polymer solar cells

Procedia PDF Downloads 283
5172 Annona muricata Leaves Induced Mitochondrial-Mediated Apoptosis in A549 Cells

Authors: Soheil Zorofchian Moghadamtousi, Habsah Abdul Kadir, Mohammadjavad Paydar, Elham Rouhollahi, Hamed Karimian

Abstract:

The present study was designed to evaluate the molecular mechanisms of Annona muricata leaves ethyl acetate extract (AMEAE) against lung cancer A549 cells. Cell viability analysis revealed the selective cytotoxic effect of AMEAE towards A549 cells. Treatment of A549 cells with AMEAE significantly elevated the reactive oxygen species formation, followed by attenuation of mitochondrial membrane potential via upregulation of Bax and downregulation of Bcl-2, accompanied by cytochrome c release to the cytosol. The released cytochrome c triggered the activation of caspase-9 followed by caspase-3. In addition, AMEAE-induced apoptosis was accompanied by cell cycle arrest at G1 phase. Our data showed for the first time that AMEAE inhibited the proliferation of A549 cells, leading to cell cycle arrest and programmed cell death through activation of the mitochondrial-mediated signaling pathway.

Keywords: Annona muricata, lung cancer, apoptosis, mitochondria

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5171 Viscoelastic Cell Concentration in a High Aspect Ratio Microchannel Using a Non-Powered Air Compressor

Authors: Jeonghun Nam, Seonggil Kim, Hyunjoo Choi, Chae Seung Lim

Abstract:

Quantification and analysis of rare cells are challenging in clinical applications and cell biology due to its extremely small number in blood. In this work, we propose a viscoelastic microfluidic device for continuous cell concentration without sheath flows. Due to the viscoelastic effect on suspending cells, cells with the blockage ratio higher than 0.1 could be tightly focused at the center of the microchannel. The blockage ratio was defined as the particle diameter divided by the channel width. Finally, cells were concentrated through the center outlet and the additional suspending medium was removed to the side outlets. Since viscoelastic focusing is insensitive to the flow rate higher than 10 μl/min, the non-powered hand pump sprayer could be used with no accurate control of the flow rate, which is suitable for clinical settings in resource-limited developing countries. Using multiple concentration processes, high-throughput concentration of white blood cells in lysed blood sample was achieved by ~ 300-fold.

Keywords: cell concentration, high-throughput, non-powered, viscoelastic fluid

Procedia PDF Downloads 288
5170 Determination of Circulating Tumor Cells in Breast Cancer Patients by Electrochemical Biosensor

Authors: Gökçe Erdemir, İlhan Yaylım, Serap Erdem-Kuruca, Musa Mutlu Can

Abstract:

It has been determined that the main reason for the death of cancer disease is caused by metastases rather than the primary tumor. The cells that leave the primary tumor and enter the circulation and cause metastasis in the secondary organs are called "circulating tumor cells" (CTCs). The presence and number of circulating tumor cells has been associated with poor prognosis in many major types of cancer, including breast, prostate, and colorectal cancer. It is thought that knowledge of circulating tumor cells, which are seen as the main cause of cancer-related deaths due to metastasis, plays a key role in the diagnosis and treatment of cancer. The fact that tissue biopsies used in cancer diagnosis and follow-up are an invasive method and are insufficient in understanding the risk of metastasis and the progression of the disease have led to new searches. Liquid biopsy tests performed with a small amount of blood sample taken from the patient for the detection of CTCs are easy and reliable, as well as allowing more than one sample to be taken over time to follow the prognosis. However, since these cells are found in very small amounts in the blood, it is very difficult to capture them and specially designed analytical techniques and devices are required. Methods based on the biological and physical properties of the cells are used to capture these cells in the blood. Early diagnosis is very important in following the prognosis of tumors of epithelial origin such as breast, lung, colon and prostate. Molecules such as EpCAM, vimentin, and cytokeratins are expressed on the surface of cells that pass into the circulation from very few primary tumors and reach secondary organs from the circulation, and are used in the diagnosis of cancer in the early stage. For example, increased EpCAM expression in breast and prostate cancer has been associated with prognosis. These molecules can be determined in some blood or body fluids to be taken from patients. However, more sensitive methods are required to be able to determine when they are at a low level according to the course of the disease. The aim is to detect these molecules found in very few cancer cells with the help of sensitive, fast-sensing biosensors, first in breast cancer cells reproduced in vitro and then in blood samples taken from breast cancer patients. In this way, cancer cells can be diagnosed early and easily and effectively treated.

Keywords: electrochemical biosensors, breast cancer, circulating tumor cells, EpCAM, Vimentin, Cytokeratins

Procedia PDF Downloads 261
5169 Imaging of Underground Targets with an Improved Back-Projection Algorithm

Authors: Alireza Akbari, Gelareh Babaee Khou

Abstract:

Ground Penetrating Radar (GPR) is an important nondestructive remote sensing tool that has been used in both military and civilian fields. Recently, GPR imaging has attracted lots of attention in detection of subsurface shallow small targets such as landmines and unexploded ordnance and also imaging behind the wall for security applications. For the monostatic arrangement in the space-time GPR image, a single point target appears as a hyperbolic curve because of the different trip times of the EM wave when the radar moves along a synthetic aperture and collects reflectivity of the subsurface targets. With this hyperbolic curve, the resolution along the synthetic aperture direction shows undesired low resolution features owing to the tails of hyperbola. However, highly accurate information about the size, electromagnetic (EM) reflectivity, and depth of the buried objects is essential in most GPR applications. Therefore hyperbolic curve behavior in the space-time GPR image is often willing to be transformed to a focused pattern showing the object's true location and size together with its EM scattering. The common goal in a typical GPR image is to display the information of the spatial location and the reflectivity of an underground object. Therefore, the main challenge of GPR imaging technique is to devise an image reconstruction algorithm that provides high resolution and good suppression of strong artifacts and noise. In this paper, at first, the standard back-projection (BP) algorithm that was adapted to GPR imaging applications used for the image reconstruction. The standard BP algorithm was limited with against strong noise and a lot of artifacts, which have adverse effects on the following work like detection targets. Thus, an improved BP is based on cross-correlation between the receiving signals proposed for decreasing noises and suppression artifacts. To improve the quality of the results of proposed BP imaging algorithm, a weight factor was designed for each point in region imaging. Compared to a standard BP algorithm scheme, the improved algorithm produces images of higher quality and resolution. This proposed improved BP algorithm was applied on the simulation and the real GPR data and the results showed that the proposed improved BP imaging algorithm has a superior suppression artifacts and produces images with high quality and resolution. In order to quantitatively describe the imaging results on the effect of artifact suppression, focusing parameter was evaluated.

Keywords: algorithm, back-projection, GPR, remote sensing

Procedia PDF Downloads 453
5168 Synergistic Cytotoxicity of Cisplatin and Taxol in Overcoming Taxol Resistance through the Inhibition of LDHA in Oral Squamous Cell Carcinoma

Authors: Lin Feng, Ling-Ling E., Hong-Chen Liu

Abstract:

The development of chemoresistance in patients represents a major challenge in cancer treatment. Lactate dehydrogenase‑A (LDHA) is one of the principle isoforms of LDH that is expressed in breast tissue, controlling the conversion of pyruvate to lactate and also playing a significant role in the metabolism of glucose. The aim of this study was to identify whether LDHA was involved in oral cancer cell resistance to Taxol and whether the downregulation of LDHA, as a result of cisplatin treatment, may overcome Taxol resistance in human oral squamous cells. The OECM‑1 oral epidermal carcinoma cell line was used, which has been widely used as a model of oral cancer in previous studies. The role of LDHA in Taxol and cisplatin resistance was investigated and the synergistic cytotoxicity of cisplatin and/or Taxol in oral squamous cells was analyzed. Cell viability was analyzed by MTT assay, LDHA expression was analyzed by western blot analysis and siRNA transfection was performed to knock down LDHA expression. The present study results showed that decreased levels of LDHA were responsible for the resistance of oral cancer cells to cisplatin (CDDP). CDDP treatments downregulated LDHA expression and lower levels of LDHA were detected in the CDDP‑resistant oral cancer cells compared with the CDDP‑sensitive cells. By contrast, the Taxol‑resistant cancer cells showed elevated LDHA expression levels. In addition, small interfering RNA‑knockdown of LDHA sensitized the cells to Taxol but desensitized them to CDDP treatment while exogenous expression of LDHA sensitized the cells to CDDP, but desensitized them to Taxol. The present study also revealed the synergistic cytotoxicity of CDDP and Taxol for killing oral cancer cells through the inhibition of LDHA. This study highlights LDHA as a novel therapeutic target for overcoming Taxol resistance in oral cancer patients using the combined treatments of Taxol and CDDP.

Keywords: cisplatin, Taxol, carcinoma, oral squamous cells

Procedia PDF Downloads 419