Search results for: hepatocellular carcinoma cells

Authors: Susan Hall, Gary D. Grant, Catherine McDermott, Devinder Arora

Abstract:

Introduction /Aim: The kynurenine pathway is thought to play an important role in the pathophysiology of numerous neurodegenerative diseases including depression, Alzheimer’s disease, and Parkinson’s disease. Numerous neuroactive compounds, including the neurotoxic 3-hydroxyanthranilic acid, 3-hydroxykynurenine and quinolinic acid and the neuroprotective kynurenic acid and picolinic acid, are produced through the metabolism of kynurenine and are thought to be the causative agents responsible for neurodegeneration. The toxicity of 3-hydroxykynurenine, 3-hydroxyanthranilic acid and quinolinic acid has been widely evaluated and demonstrated in primary cell cultures but to date only 3-hydroxykynurenine and 3-hydroxyanthranilic acid have been shown to cause toxicity in immortal tumour cells. The aim of this study was to evaluate the toxicity of kynurenine metabolites, both individually and in combination, on SH-SY5Y neuroblastoma cells after 24 and 72 h exposure in order to explore a cost-effective model to study their neurotoxic effects and potential protective agents. Methods: SH-SY5Y neuroblastoma cells were exposed to various concentrations of the neuroactive kynurenine metabolites, both individually and in combination, for 24 and 72 h, and viability was subsequently evaluated using the Resazurin (Alamar blue) proliferation assay. Furthermore, the effects of these compounds, alone and in combination, on specific death pathways including apoptosis, necrosis and free radical production was evaluated using various assays. Results: Consistent with literature, toxicity was shown with short-term 24-hour treatments at 1000 μM concentrations for both 3-hydroxykynurenine and 3-hydroxyanthranilic acid. Combinations of kynurenine metabolites showed modest toxicity towards SH-SY5Y neuroblastoma cells in a concentration-dependent manner. Specific cell death pathways, including apoptosis, necrosis and free radical production were shown to be increased after both 24 and 72 h exposure of SH-SY5Y neuroblastoma cells to 3-hydroxykynurenine and 3-hydroxyanthranilic acid and various combinations of neurotoxic kynurenine metabolites. Conclusion: It is well documented that neurotoxic kynurenine metabolites show toxicity towards primary human neurons in the nanomolar to low micromolar concentration range. Results show that the concentrations required to show significant cell death are in the range of 1000 µM for 3-hydroxykynurenine and 3-hydroxyanthranilic acid and toxicity of quinolinic acid towards SH-SY5Y was unable to be shown. This differs significantly from toxicities observed in primary human neurons. Combinations of the neurotoxic metabolites were shown to have modest toxicity towards these cells with increased toxicity and activation of cell death pathways observed after 72 h exposure. This study suggests that the 24 h model is unsuitable for use in neurotoxicity studies, however, the 72 h model better represents the observations of the studies using primary human neurons and may provide some benefit in providing a cost-effective model to assess possible protective agents against kynurenine metabolite toxicities.

Keywords: kynurenine metabolites, neurotoxicity, quinolinic acid, SH-SY5Y neuroblastoma

Procedia PDF Downloads 419

Authors: M. Borgie, Z. Dagher, F. Ledoux, A. Verdin, F. Cazier, H. Greige, P. Shirali, D. Courcot

Abstract:

In October 2013, the International Agency for Research on Cancer (IARC) classified outdoor air pollution and fine particulate matter (PM2.5) as carcinogenic to humans. Despite the clearly relationship established by epidemiological studies between PM exposure and the onset of respiratory and cardiovascular diseases, uncertainties remain about the physiopathological mechanisms responsible for these diseases. The aim of this work was to evaluate the toxicological effects of two samples of atmospheric PM2.5 collected at urban and rural sites on human bronchial epithelial cells, BEAS-2B, especially to investigate the metabolic activation of organic compounds, the alteration of epigenetic mechanisms (i.e. microRNAs genes expression), the phosphorylation of H2AX and the telomerase activity. Our results showed a significant increase in CYP1A1, CYP1B1, and AhRR genes expression, miR-21 gene expression, H2AX phosphorylation and telomerase activity in BEAS-2B cells after their exposure to PM2.5, both in a dose and site-dependent manner. These results showed that PM2.5, especially urban PM, are able to induce the expression of metabolizing enzymes which can provide metabolic biotransformation of organic compounds into more toxic and carcinogenic metabolites, and to induce the expression of the oncomiR miR-21 which promotes cell growth and enhances tumor invasion and metastasis in lung cancer. In addition, our results have highlighted the role of PM2.5 in the activation of telomerase, which can maintain the telomeres length and subsequently preventing cell death, and have also demonstrated the ability of PM2.5 to induce DNA breaks and thus to increase the risk of mutations or chromosomal translocations that lead to genomic instability. All these factors may contribute to cell abnormalities, and thus the development of cancer.

Keywords: BEAS-2B cells, carcinogenesis, epigenetic alterations and genotoxicity, PM2.5

Procedia PDF Downloads 382

Authors: Kirti Hira, A. Sajeli Begum, S. Mahibalan, Poorna Chandra Rao

Abstract:

Introduction: Cancer is one of the leading causes of death worldwide. Although existing therapy effectively kills cancer cells, they do affect normal growing cells leading to many undesirable side effects. Hence there is need to develop effective as well as safe drug molecules to combat cancer, which is possible through phyto-research. The currently available plant-derived blockbuster drugs are the example for this. In view of this, an investigation was done to identify cytotoxic lead molecules from Hedyotis umbellata (Family Rubiaceae), a widely distributed weed in India. Materials and Methods: The methanolic extract of the whole plant of H. umbellata (MHU), prepared through Soxhlet extraction method was further fractionated with diethyl ether and n-butanol, successively. MHU, ether fraction (EMHU) and butanol fraction (BMHU) were lyophilized and were tested for the cytotoxic effect using 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay against non-small cell lung cancer (NSCLC) A549 cell lines. The potentially active EMHU was subjected to chromatographic purification using normal-phase silica columns, in order to isolate the responsible bioactive compounds. The isolated pure compounds were tested for their cytotoxic effect by MTT assay against A549 cells. Compound-3, which was found to be most active, was characterized using IR, 1H- and 13C-NMR and MS analysis. The study was further extended to decipher the mechanism of action of cytotoxicity of compound-3 against A549 cells through various in vitro cellular models. Cell cycle analysis was done using flow cytometry following PI (Propidium Iodide) staining. Protein analysis was done using Western blot technique. Results: Among MHU, EMHU, and BMHU, the non-polar fraction EMHU demonstrated a significant dose-dependent cytotoxic effect with IC50 of 67.7μg/ml. Chromatography of EMHU yielded seven compounds. MTT assay of isolated compounds explored compound-3 as potentially active one, which inhibited the growth of A549 cells with IC50value of 14.2μM. Further, compound-3 was identified as cedrelopsin, a coumarin derivative having molecular weight of 260. Results of in vitro mechanistic studies explained that cedrelopsin induced cell cycle arrest at G2/M phase and down-regulated the expression of G2/M regulatory proteins such as cyclin B1, cdc2, and cdc25C, dose dependently. This is the first report that explores the cytotoxic mechanism of cedrelopsin. Conclusion: Thus a potential small lead molecule, cedrelopsin isolated from H. umbellata, showing antiproliferative effect mediated by G2/M arrest in A549 cells was discovered. The effect of cedrelopsin against other cancer cell lines followed by in vivo studies can be performed in future to develop a new drug candidate.

Keywords: A549, cedrelopsin, G2/M phase, Hedyotis umbellata

Procedia PDF Downloads 176

Authors: Almudena Espin Perez, Tyler Risom, Carl Pelz, Isabel English, Robert M. Angelo, Rosalie Sears, Andrew J. Gentles

Abstract:

Pancreatic ductal adenocarcinoma (PDAC) is one of the most deadly malignancies. The role of the tumor microenvironment (TME) is gaining significant attention in cancer research. Despite ongoing efforts, the nature of the interactions between tumors, immune cells, and stromal cells remains poorly understood. The cell-intrinsic properties that govern cell lineage plasticity in PDAC and extrinsic influences of immune populations require technically challenging approaches due to the inherently heterogeneous nature of PDAC. Understanding the cell lineage plasticity of PDAC will improve the development of novel strategies that could be translated to the clinic. Members of the team have demonstrated that the acquisition of ductal to neuroendocrine lineage plasticity in PDAC confers therapeutic resistance and is a biomarker of poor outcomes in patients. Our approach combines computational methods for deconvolving bulk transcriptomic cancer data using CIBERSORTx and high-throughput single-cell imaging using Multiplexed Ion Beam Imaging (MIBI) to study lineage plasticity in PDAC and its relationship to the infiltrating immune system. The CIBERSORTx algorithm uses signature matrices from immune cells and stroma from sorted and single-cell data in order to 1) infer the fractions of different immune cell types and stromal cells in bulked gene expression data and 2) impute a representative transcriptome profile for each cell type. We studied a unique set of 300 genomically well-characterized primary PDAC samples with rich clinical annotation. We deconvolved the PDAC transcriptome profiles using CIBERSORTx, leveraging publicly available single-cell RNA-seq data from normal pancreatic tissue and PDAC to estimate cell type proportions in PDAC, and digitally reconstruct cell-specific transcriptional profiles from our study dataset. We built signature matrices and optimized by simulations and comparison to ground truth data. We identified cell-type-specific transcriptional programs that contribute to cancer cell lineage plasticity, especially in the ductal compartment. We also studied cell differentiation hierarchies using CytoTRACE and predict cell lineage trajectories for acinar and ductal cells that we believe are pinpointing relevant information on PDAC progression. Collaborators (Angelo lab, Stanford University) has led the development of the Multiplexed Ion Beam Imaging (MIBI) platform for spatial proteomics. We will use in the very near future MIBI from tissue microarray of 40 PDAC samples to understand the spatial relationship between cancer cell lineage plasticity and stromal cells focused on infiltrating immune cells, using the relevant markers of PDAC plasticity identified from the RNA-seq analysis.

Keywords: deconvolution, imaging, microenvironment, PDAC

Procedia PDF Downloads 128

Authors: Malgorzata J. Rybak-Smith, Benedicte Thiebaut, Simon Johnson, Peter Bishop, Helen E. Townley

Abstract:

Gadolinium doped titania (TiO2:Gd) nanoparticles (NPs) can be activated by X-ray radiation to generate Reactive Oxygen Species (ROS), which can be effective in killing cancer cells. As such, treatment with these NPs can be used to enhance the efficacy of conventional radiotherapy. Incorporation of the NPs in to tumour tissue will permit the extension of radiotherapy to currently untreatable tumours deep within the body, and also reduce damage to neighbouring healthy cells. In an attempt to find a fast and scalable method for the synthesis of the TiO2:Gd NPs, the use of Flame Spray Pyrolysis (FSP) was investigated. A series of TiO2 NPs were generated with 1, 2, 5 and 7 mol% gadolinium dopant. Post-synthesis, the TiO2:Gd NPs were silica-coated to improve their biocompatibility. Physico-chemical characterisation was used to determine the size and stability in aqueous suspensions of the NPs. All analysed TiO2:Gd NPs were shown to have relatively high photocatalytic activity. Furthermore, the FSP synthesized silica-coated TiO2:Gd NPs generated enhanced ROS in chemico. Studies on rhabdomyosarcoma (RMS) cell lines (RD & RH30) demonstrated that in the absence of irradiation all TiO2:Gd NPs were inert. However, application of TiO2:Gd NPs to RMS cells, followed by irradiation, showed a significant decrease in cell proliferation. Consequently, our studies showed that the X-ray-activatable TiO2:Gd NPs can be prepared by a high-throughput scalable technique to provide a novel and affordable anticancer therapy.

Keywords: cancer, gadolinium, ROS, titania nanoparticles, X-ray

Procedia PDF Downloads 431

Authors: Viviane A. O. Silva, Angela M. Costa, Glaucia N. M. Hajj, Ana Preto, Aline Tansini, Martin Roffé, Peter Jordan, Rui M. Reis

Abstract:

Autophagy is a recycling and degradative system suggested to be a major cell death pathway in cancer cells. Autophagy pathway is interconnected with the endocytosis pathways sharing the same ultimate lysosomal destination. Lysosomes are crucial regulators of cell homeostasis, responsible to downregulate receptor signalling and turnover. It seems highly likely that derailed endocytosis can make major contributions to several hallmarks of cancer. WNK2, a member of the WNK (with-no-lysine [K]) subfamily of protein kinases, had been found downregulated by its promoter hypermethylation, and has been proposed to act as a specific tumour-suppressor gene in brain tumors. Although some contradictory studies indicated WNK2 as an autophagy modulator, its role in cancer cell death is largely unknown. There is also growing evidence for additional roles of WNK kinases in vesicular traffic. Aim: To evaluate the role of WNK2 in autophagy and endocytosis on glioma context. Methods: Wild-type (wt) A172 cells (WNK2 promoter-methylated), and A172 transfected either with an empty vector (Ev) or with a WNK2 expression vector, were used to assess the cellular basal capacities to promote autophagy, through western blot and flow-cytometry analysis. Additionally, we evaluated the effect of WNK2 on general endocytosis trafficking routes by immunofluorescence. Results: The re-expression of ectopic WNK2 did not interfere with autophagy-related protein light chain 3 (LC3-II) expression levels as well as did not promote mTOR signaling pathway alteration when compared with Ev or wt A172 cells. However, the restoration of WNK2 resulted in a marked increase (8 to 92,4%) of Acidic Vesicular Organelles formation (AVOs). Moreover, our results also suggest that WNK2 cells promotes delay in uptake and internalization rate of cholera toxin B and transferrin ligands. Conclusions: The restoration of WNK2 interferes in vesicular traffic during endocytosis pathway and increase AVOs formation. This results also suggest the role of WNK2 in growth factor receptor turnover related to cell growth and homeostasis and associates one more time, WNK2 silencing contribution in genesis of gliomas.

Keywords: autophagy, endocytosis, glioma, WNK2

Procedia PDF Downloads 370

Authors: Jihoon Yang, Jeong II Choi

Abstract:

Mincle is expressed in macrophages and is members of immunoreceptors induced after exposure to various stimuli and stresses. Mincle receptor activation promotes the production of these substances by increasing the transcription of inflammatory cytokines and chemokines. Cytokines, which play an important role in the initiation and maintenance of such inflammatory pain diseases, have a significant effect on sensory neurons in addition to their enhancement and inhibitory effects on immune and inflammatory cells as mediators of cell interaction. Glial cells in the central nervous system play a critical role in development and maintenance of chronic pain states. Microglia are tissue-resident macrophages in the central nervous system, and belong to a group of mononuclear phagocytes. In the central nervous system, mincle receptor is present in neurons and glial cells of the brain.This study was performed to identify the Mincle receptor in the spinal cord and to investigate the effect of Mincle receptor activation on nociception and the changes of microglia. Materials and Methods: C-type lectins(Mincle) was identified in spinal cord of Male Sprague–Dawley rats. Then, mincle receptor ligand (TDB), via an intrathecal catheter. Mechanical allodynia was measured using von Frey test to evaluate the effect of intrathecal injection of TDB. Result: The present investigation shows that the intrathecal administration of TDB in the rat produces a reliable and quantifiable mechanical hyperalgesia. In addition, The mechanical hyperalgesia after TDB injection gradually developed over time and remained until 10 days. Mincle receptor is identified in the spinal cord, mainly expressed in neuronal cells, but not in microglia or astrocyte. These results suggest that activation of mincle receptor pathway in neurons plays an important role in inducing activation of microglia and inducing mechanical allodynia.

Keywords: mincle, spinal cord, pain, microglia

Procedia PDF Downloads 161

Authors: Jesus Alfredo Ortega Granados, Pandiyan Thangarasu

Abstract:

Antibiotic tetracycline presents as a micro-contaminant in fresh water, wastewater and soils, causing environmental and health problems. In this work, tetracycline (TC) has been employed as chemo-sensor for the recognition of Al³⁺ without interring other ions, and the results show that it enhances the fluorescence intensity for Al³⁺ and there is no interference from other coexisting cation ions (Cd²⁺, Ni²⁺, Co²⁺, Sr²⁺, Mg²⁺, Fe³⁺, K⁺, Sm³⁺, Ag⁺, Na⁺, Ba²⁺, Zn²⁺, and Mn²⁺). For the addition of Cu²⁺ to [TET-Al³⁺], it appears that the intensity of fluorescence has been quenched. Other combinations of metal ions in addition to TC do not change the fluorescence behavior. The stoichiometry determined by Job´s plot for the interaction of TC with Al³⁺ was found to be 1:1. Importantly, the detection of Al³⁺⁺ successfully employed in the real samples like living cells, and it was found that TC efficiently performs as a fluorescent probe for Al³⁺ ion in living systems, especially in Saccharomyces cerevisiae; this is confirmed by confocal laser scanning microscopy.

Keywords: chemo-sensor, recognition of Al³⁺ ion, Saccharomyces cerevisiae, tetracycline,

Procedia PDF Downloads 191

Authors: Chahrazed Bendenia, Souhila Bendenia, Samia Moulebhar, Hanaa Merad-Dib, Sarra Merabet, Sid Ahmed Khantar, Baghdad Hadri

Abstract:

In recent years, organic solar cells (OSCs) have become the fundamental concern of researchers thanks to their advantages in terms of flexibility, manufacturing processes and low cost. The performance of these devices is influenced by various factors, such as the layers introduced in the stacking of the solar cell realized. In our work, the modeling of a reverse OSC under AM1.5G illumination will be determined. The photo-active polymer/fullerene layer will be analyzed from the polymer variation of this layer using the SCAPS simulator to extract the J-V characteristics: open circuit voltage (Voc), short circuit current (Jsc), filling factor (FF) and power conversion efficiency (η). The results obtained indicated that the materials used have a significant impact on improving the photovoltaic parameters of the devices studied.

Keywords: solar, polymer, simulator, characteristics

Procedia PDF Downloads 78

Authors: Afzaal Bashir, Sunaina Afzaal

Abstract:

Background: Facial contour deformities associated with pigmentary changes are of major concern for plastic surgeons, both being important and difficult in treating such issues. No definite ideal treatment option is available to simultaneously address both the contour defect as well as related pigmentation. Objectives: The aim of the current study is to compare the long-term effects of conventional adipose tissue grafting and ex-vivo expanded Mesenchymal Stem Cells enriched adipose tissue grafting for the treatment of contour deformities with pigmentary changes on the face. Material and Methods: In this study, eighty (80) patients with contour deformities of the face with hyperpigmentation were recruited after informed consent. Two techniques i.e., conventional fat grafting (C-FG) and fat grafts enriched with expanded adipose stem cells (FG-ASCs), were used to address the pigmentation. Both techniques were explained to patients, and enrolled patients were divided into two groups i.e., C-FG and FG-ASCS, per patients’ choice and satisfaction. Patients of the FG-ASCs group were treated with fat grafts enriched with expanded adipose stem cells, while patients of the C-FGs group were treated with conventional fat grafting. Patients were followed for 12 months, and improvement in face pigmentation was assessed clinically as well as measured objectively. Patient satisfaction was also documented as highly satisfied, satisfied, and unsatisfied. Results: Mean age of patients was 24.42(±4.49), and 66 patients were females. The forehead was involved in 61.20% of cases, the cheek in 21.20% of cases, the chin in 11.20% of cases, and the nose in 6.20% of cases. In the GF-ASCs group, the integrated color density (ICD) was decreased (1.08×10⁶ ±4.64×10⁵) as compared to the C-FG group (2.80×10⁵±1.69×10⁵). Patients treated with fat grafts enriched with expanded adipose stem cells were significantly more satisfied as compared to patients treated with conventional fat grafting only. Conclusion: Mesenchymal stem cell-enriched autologous fat grafting is a preferred option for improving the contour deformities related to increased pigmentation of face skin.

Keywords: hyperpigmentation, color density, enriched adipose tissue graft, fat grafting, contour deformities, Image J

Procedia PDF Downloads 113

Authors: Lanlan Xiao, Yang Liu, Shuo Chen, Bingmei Fu

Abstract:

Most cancer-related deaths are due to metastasis. Metastasis is a complex, multistep processes including the detachment of cancer cells from the primary tumor and the migration to distant targeted organs through blood and/or lymphatic circulations. During hematogenous metastasis, the emigration of tumor cells from the blood stream through the vascular wall into the tissue involves arrest in the microvasculature, adhesion to the endothelial cells forming the microvessel wall and transmigration to the tissue through the endothelial barrier termed as extravasation. The narrow slit between endothelial cells that line the microvessel wall is the principal pathway for tumor cell extravasation to the surrounding tissue. To understand this crucial step for tumor hematogenous metastasis, we used Dissipative Particle Dynamics method to investigate an individual cell passing through a narrow slit numerically. The cell membrane was simulated by a spring-based network model which can separate the internal cytoplasm and surrounding fluid. The effects of the cell elasticity, cell shape and cell surface area increase, and slit size on the cell transmigration through the slit were investigated. Under a fixed driven force, the cell with higher elasticity can be elongated more and pass faster through the slit. When the slit width decreases to 2/3 of the cell diameter, the spherical cell becomes jammed despite reducing its elasticity modulus by 10 times. However, transforming the cell from a spherical to ellipsoidal shape and increasing the cell surface area only by 3% can enable the cell to pass the narrow slit. Therefore the cell shape and surface area increase play a more important role than the cell elasticity in cell passing through the narrow slit. In addition, the simulation results indicate that the cell migration velocity decreases during entry but increases during exit of the slit, which is qualitatively in agreement with the experimental observation.

Keywords: dissipative particle dynamics, deformability, surface area increase, cell migration

Procedia PDF Downloads 336

Authors: Zellagui Rahima, Chaumont Denis, Boughelout Abderrahman, Adnane Mohamed

Abstract:

Thin films of CdxZn1-xS were deposed by chemical bath deposition on glass substrates for photovoltaic applications. The thin films CdZnS were synthesized by chemical bath (CBD) with different deposition protocols for optimized the parameter of deposition as the temperature, time of deposition, concentrations of ion and pH. Surface morphology, optical and chemical composition properties of thin film CdZnS were investigated by SEM, EDAX, spectrophotometer. The transmittance is 80% in visible region 300 nm – 1000 nm; it has been observed in that films the grain size is between 50nm and 100nm measured by SEM image and we also note that the shape of particle is changing with the change in concentration. This result favors of application these films in solar cells; the chemical analysis with EDAX gives information about the presence of Cd, Zn and S elements and investigates the stoichiometry.

Keywords: thin film, solar cells, transmition, cdzns

Procedia PDF Downloads 262

Authors: Diego Luján Villarreal

Abstract:

Retinal prosthetic devices aim to repair some vision in visual impairment patients by stimulating electrically neural cells in the visual system. In this study, the 3D linear electrode carrier is presented. A simulation framework was developed by placing the 3D carrier 1 mm away from the fovea center at the highest-density cell. Cell stimulation is verified in COMSOL Multiphysics by developing a 3D computational model which includes the relevant retinal interface elements and dynamics of the voltage-gated ionic channels. Current distribution resulting from low threshold amplitudes produces a small volume equivalent to the volume confined by individual cells at the highest-density cell using small-sized electrodes. Delicate retinal tissue is protected by excessive charge density

Keywords: retinal prosthetic devices, visual devices, retinal implants., visual prosthetic devices

Procedia PDF Downloads 114

Authors: Dona Pamoda W. Jayatunga, Veer Bala Gupta, Eugene Hone, Ralph N. Martins

Abstract:

Being a neurodegenerative disorder, Alzheimer’s disease (AD) affects a majority of the elderly demented worldwide. The key risk factors for AD are age, metabolic syndrome, allele status of APOE gene, head injuries and lifestyle. The progressive nature of AD is characterized by symptoms of multiple cognitive deficits exacerbated over time, leading to death within a decade from clinical diagnosis. However, it is revealed that AD originates via a prodromal phase that spans from one to few decades before symptoms first manifest. The key pathological hallmarks of AD brains are deposition of amyloid beta (Aβ) plaques and neurofibrillary tangles (NFT). However, the yet unknown etiology of the disease fails to distinguish mitochondrial dysfunction between a cause or an outcome. The absence of early diagnosis tools and definite therapies for AD have permitted recruits of nutraceutical-based approaches aimed at reducing the risk of AD by modulating lifestyle or be used as preventive tools during AD prodromal state before widespread neurodegeneration begins. The objective of the present study was to investigate beneficial effects of luteolin, a plant-based flavone compound, against AD. The neuroprotective effects of luteolin on amyloid beta (Aβ) (1-42)-induced neurotoxicity was measured using cultured human neuroblastoma BE(2)-M17 cells. After exposure to 20μM Aβ (1-42) for 48 h, the neuroblastoma cells exhibited marked apoptotic death. Co-treatment of 20μM Aβ (1-42) with luteolin (0.5-5μM) significantly protected the cells against Aβ (1-42)-induced toxicity, as assessed by the MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2(4sulfophenyl)-2H-tetrazolium, inner salt; MTS] reduction assay and the lactate dehydrogenase (LDH) cell death assay. The results suggest that luteolin prevents Aβ (1-42)-induced apoptotic neuronal death. However, further studies are underway to determine its protective mechanisms in AD including the activity against tau hyperphosphorylation and mitochondrial dysfunction.

Keywords: Aβ (1-42)-induced toxicity, Alzheimer’s disease, luteolin, neuroblastoma cells

Procedia PDF Downloads 150

Authors: Kakali Bhadra

Abstract:

Harmalol administration caused remarkable reduction in proliferation of HepG₂ cells with GI₅₀ of 14.2 mM, without showing much cytotoxicity in embryonic liver cell line, WRL-68. Data from circular dichroism and differential scanning calorimetric analysis of harmalol-CT DNA complex shows conformational changes with prominent CD perturbation and stabilization of CT DNA by 8 ^oC. Binding constant and stoichiometry was also calculated using the above biophysical techniques. Further, dose dependent apoptotic induction ability of harmalol was studied in HepG₂ cells using different biochemical assays. Generation of ROS, DNA damage, changes in cellular external and ultramorphology, alteration of membrane, formation of comet tail, decreased mitochondrial membrane potential and a significant increase in Sub G_o/G₁ population made the cancer cell, HepG₂, prone to apoptosis. Up regulation of p53 and caspase 3 further indicated the apoptotic role of harmalol.

Keywords: apoptosis, beta carboline alkaloid, comet assay, cytotoxicity, ROS

Procedia PDF Downloads 209

Authors: Muslim Idan Mohsin, Mohammed Jasim Al-Shamarti, Rusul Idan Mohsin, Ali A. Majeed

Abstract:

One of the causative agents of the lower respiratory tract (LRT) is Pseudomonas aeruginosa, which can lead to severe infection associated with a lung infection. There are many cytokines that are secreted in response to bacterial infection, in particular interleukin IL-36 cytokine in response to P. aeruginosa infection. The involvement of IL-36 in the P. aeruginosa infection could be a clue to find a specific way for treatments of different inflammatory and degenerative lung diseases. IL36 promotes primary immune response via binding to the IL-36 receptor (IL-36R). Indeed, an overactivity of IL-36 might be an initiating factor for many immunopathologic sceneries in pneumonia. Here we demonstrate if the IL-36 cytokine increases in response P. aeruginosa infection that is isolated from lower respiratory tract infection (LRT). We demonstrated that IL-36 expression significantly unregulated in human lung epithelial (A549) cells after infected by P. aeruginosa at mRNA level.

Keywords: IL36, Pseudomonas aeruginosa, LRT infection, A549 cells

Procedia PDF Downloads 234

Authors: Taleb Lotfi, Benarbia Maheidine, Allam Hamza, Boutira Fatima, Boukerche Abdelbaki

Abstract:

Introduction and purpose of the study: This is a retrospective single-center study with an analytical aim to determine the prognostic factors for relapse in patients treated with radiotherapy after total laryngectomy with lymph node dissection for laryngeal cancer at the Emir Abdelkader cancer center in Oran (Algeria). Material and methods: During the study period from January 2014 to December 2018, eighty-nine patients (n=89) with squamous cell carcinoma of the larynx were treated with postoperative radiotherapy. Relapse-free survival was studied in the univariate analysis according to pre-treatment criteria using Kaplan-Meier survival curves. We performed a univariate analysis to identify relapse factors. Statistically significant factors have been studied in the multifactorial analysis according to the Cox model. Results and statistical analysis: The average age was 62.7 years (40-86 years). It was a squamous cell carcinoma in all cases. Postoperatively, the tumor was classified as pT3 and pT4 in 93.3% of patients. Histological lymph node involvement was found in 36 cases (40.4%), with capsule rupture in 39% of cases, while the limits of surgical excision were microscopically infiltrated in 11 patients (12.3%). Chemotherapy concomitant with radiotherapy was used in 67.4% of patients. With a median follow-up of 57 months (23 to 104 months), the probabilities of relapse-free survival and five-year overall survival are 71.2% and 72.4%, respectively. The factors correlated with a high risk of relapse were locally advanced tumor stage pT4 (p=0.001), tumor site in case of subglottic extension (p=0.0003), infiltrated surgical limits R1 (p=0.001), l lymph node involvement (p=0.002), particularly in the event of lymph node capsular rupture (p=0.0003) as well as the time between surgery and adjuvant radiotherapy (p=0.001). However, in the subgroup analysis, the major prognostic factors for disease-free survival were subglottic tumor extension (p=0.001) and time from surgery to adjuvant radiotherapy (p=0.005). Conclusion: Combined surgery and postoperative radiation therapy are effective treatment modalities in the management of laryngeal cancer. Close cooperation of the entire cervicofacial oncology team is essential, expressed during a multidisciplinary consultation meeting, with the need to respect the time between surgery and radiotherapy.

Keywords: laryngeal cancer, laryngectomy, postoperative radiotherapy, survival

Procedia PDF Downloads 106

Authors: Sina Mahdavi

Abstract:

Multiple sclerosis (MS) is a progressive inflammatory autoimmune disease of the CNS that affects the myelination process in the central nervous system (CNS). Complex interactions of various "environmental or infectious" factors may act as triggers in autoimmunity and disease progression. The association between viral infections, especially human immunodeficiency virus (HIV) and MS is one potential cause that is not well understood. This study aims to summarize the available data on human HIV infection in MS disease progression. In this study, the keywords "Multiple sclerosis", "Human immunodeficiency virus ", and "Central nervous system" in the databases PubMed, and Google Scholar between 2017 and 2022 were searched and 15 articles were chosen, studied, and analyzed. Revealed histologic signs of "MS-like illness" in the setting of HIV, which comprised widespread demyelination with reactive astrocytes, foamy macrophages, and perivascular infiltration with inflammatory cells, all of which are compatible with MS lesions. Human immunodeficiency virus causes dysfunction of the immune system, especially characterized by hypergammaglobulinemia and chronic activation of B cells. Activation of B cells leads to increased synthesis of immunoglobulin and finally to an excess of free light chains. Free light chains may be involved in autoimmune responses against neurons. There is a high expression of HIV during the course of MS, which indicates the relationship between HIV and MS, that this virus can play a role in the development of MS by creating an inflammatory state. Therefore, measures to modulate the expression of HIV may be effective in reducing inflammatory processes in demyelinated areas of MS patients.

Keywords: multiple sclerosis, human immunodeficiency virus, central nervous system, autoimmunity

Procedia PDF Downloads 84

Authors: Navkiran Kaur, Apoorva Mathur, Abhishree Agarwal, Sakshi Gupta, Tuhin Rashmi

Abstract:

Aim: Breast cancer is the most common cancer in women worldwide, and resistance to the current therapeutics, often concurrently, is an increasing clinical challenge. Glycosylation of proteins is one of the most important post-translational modifications. It is widely known that aberrant glycosylation has been implicated in many different diseases due to changes associated with biological function and protein folding. Alterations in cell surface glycosylation, can promote invasive behavior of tumor cells that ultimately lead to the progression of cancer. In breast cancer, there is an increasing evidence pertaining to the role of glycosylation in tumor formation and metastasis. In the present study, an attempt has been made to study the disease associated sialoglycoproteins in breast cancer by using bioinformatics tools. The sequence will be retrieved from UniProt database. A database in the form of a word document was made by a collection of FASTA sequences of breast cancer gene sequence. Glycosylation was studied using yinOyang tool on ExPASy and Differential genes expression and protein analysis was done in context of breast cancer metastasis. The number of residues predicted O-glc NAc threshold containing 50 aberrant glycosylation sites or more was detected and recorded for individual sequence. We found that the there is a significant change in the expression profiling of glycosylation patterns of various proteins associated with breast cancer. Differential aberrant glycosylated proteins in breast cancer cells with respect to non-neoplastic cells are an important factor for the overall progression and development of cancer.

Keywords: breast cancer, bioinformatics, cancer, metastasis, glycosylation

Procedia PDF Downloads 294

Authors: Maryam Kiani

Abstract:

This study focuses on the development of composite nanomaterials based on SiO2 and carbon black for oxygen reduction reaction (ORR) electrocatalysts in batteries and fuel cells. The aim was to explore the potential of these composite materials as efficient catalysts for ORR, which is a critical process in energy conversion devices. The SiO2/carbon black composite nanomaterials were synthesized using a facile and scalable method. The morphology, structure, and electrochemical properties of the materials were characterized using various techniques, including scanning electron microscopy (SEM), X-ray diffraction (XRD), and electrochemical measurements. The results demonstrated that the incorporation of SiO2 into the carbon black matrix enhanced the ORR performance of the composite material. The composite nanomaterials exhibited improved electrocatalytic activity, enhanced stability, and increased durability compared to pure carbon black. The presence of SiO2 facilitated the formation of active sites, improved electron transfer, and increased the surface area available for ORR. This study contributes to the advancement of battery and fuel cell technology by offering a promising approach for the development of high-performance ORR electrocatalysts. The SiO2/carbon black composite nanomaterials show great potential for improving the efficiency and durability of energy conversion devices, leading to more sustainable and efficient energy solutions.

Keywords: oxygen reduction reaction, batteries, fuel cells, electrrocatalyst

Procedia PDF Downloads 119

Authors: Narine Ghazaryan, Joana Catarina Macedo, Sara Vaz, Naira Ayvazyan, Elsa Logarinho

Abstract:

Macrovipera lebetina obtusa (MLO) is a poisonous snake in Armenia. Obtustatin represents the shortest known monomeric disintegrin, isolated from the snake venom of MLO, and is known to specifically inhibit α1β1 integrin. Its oncostatic effect is due to the inhibition of angiogenesis, which likely arises from α1β1 integrin inhibition in the endothelial cells. To explore the therapeutic potential of the MLO snake venom and obtustatin, we studied activity of obtustatin and MLO venom in vitro, by testing their efficacy in human dermal microvascular endothelial cells (HMVEC-D) and in vivo, using chick embryo chorioallantoic membrane assay (CAM assay). Our in vitro results showed that obtustatin in comparison with MLO venom did not exhibit cytotoxic activity in HMVEC-D cells in comparison to MLO venom. But in vivo results have shown that 4µg /embryo (90 µM) of obtustatin inhibited angiogenesis induced by FGF2 by 17% while MLO snake venom induced 22% reduction of the angiogenic index. The concentration of obtustatin in the crude MLO venom was 0.3 nM, which is 300.000 times less than the concentration of the obtustatin itself. Given this enormous difference in concentration, it is likely that some components of the crude venom contribute to the observed anti-angiogenic effect. Hypotheses will be ascertained to justify this action: components in the MLO venom may increase obtustatin efficacy or have independent but synergic anti-angiogenic activities.

Keywords: angiogenesis, alpa1 beta 1 integrin, Macrovipera lebetina obtusa, obtustatin

Procedia PDF Downloads 197

Authors: Shuo Li, Yannick Coffinier, Chann Lagadec, Fabrizio Cleri, Katsuhiko Nishiguchi, Akira Fujiwara, Soo Hyeon Kim, Nicolas Clément

Abstract:

The need for single cell detection and analysis techniques has increased in the past decades because of the heterogeneity of individual living cells, which increases the complexity of the pathogenesis of malignant tumors. In the search for early cancer detection, high-precision medicine and therapy, the technologies most used today for sensitive detection of target analytes and monitoring the variation of these species are mainly including two types. One is based on the identification of molecular differences at the single-cell level, such as flow cytometry, fluorescence-activated cell sorting, next generation proteomics, lipidomic studies, another is based on capturing or detecting single tumor cells from fresh or fixed primary tumors and metastatic tissues, and rare circulating tumors cells (CTCs) from blood or bone marrow, for example, dielectrophoresis technique, microfluidic based microposts chip, electrochemical (EC) approach. Compared to other methods, EC sensors have the merits of easy operation, high sensitivity, and portability. However, despite various demonstrations of low limits of detection (LOD), including aptamer sensors, arrayed EC sensors for detecting single-cell have not been demonstrated. In this work, a new technique based on 20-nm-thick nanopillars array to support cells and keep them at ideal recognition distance for redox-labeled aptamers grafted on the surface. The key advantages of this technology are not only to suppress the false positive signal arising from the pressure exerted by all (including non-target) cells pushing on the aptamers by downward force but also to stabilize the aptamer at the ideal hairpin configuration thanks to a confinement effect. With the first implementation of this technique, a LOD of 13 cells (with5.4 μL of cell suspension) was estimated. In further, the nanosupported cell technology using redox-labeled aptasensors has been pushed forward and fully integrated into a single-cell electrochemical aptasensor array. To reach this goal, the LOD has been reduced by more than one order of magnitude by suppressing parasitic capacitive electrochemical signals by minimizing the sensor area and localizing the cells. Statistical analysis at the single-cell level is demonstrated for the recognition of cancer cells. The future of this technology is discussed, and the potential for scaling over millions of electrodes, thus pushing further integration at sub-cellular level, is highlighted. Despite several demonstrations of electrochemical devices with LOD of 1 cell/mL, the implementation of single-cell bioelectrochemical sensor arrays has remained elusive due to their challenging implementation at a large scale. Here, the introduced nanopillar array technology combined with redox-labeled aptamers targeting epithelial cell adhesion molecule (EpCAM) is perfectly suited for such implementation. Combining nanopillar arrays with microwells determined for single cell trapping directly on the sensor surface, single target cells are successfully detected and analyzed. This first implementation of a single-cell electrochemical aptasensor array based on Brownian-fluctuating redox species opens new opportunities for large-scale implementation and statistical analysis of early cancer diagnosis and cancer therapy in clinical settings.

Keywords: bioelectrochemistry, aptasensors, single-cell, nanopillars

Procedia PDF Downloads 118

Authors: Daphne Meza, Louie Abejar, David A. Rubenstein, Wei Yin

Abstract:

Endothelial cell (ECs) morphology and function is highly impacted by the mechanical stresses these cells experience in vivo. Any change in the mechanical environment can trigger pathological EC responses. A detailed understanding of EC morphological response and function upon subjection to individual and simultaneous mechanical stimuli is needed for advancement in mechanobiology and preventive medicine. To investigate this, a programmable device capable of simultaneously applying physiological fluid shear stress (FSS) and cyclic strain (CS) has been developed, characterized and validated. Its validation was performed both experimentally, through tracer tracking, and theoretically, through the use of a computational fluid dynamics model. The effectiveness of the device was evaluated through EC morphology changes under mechanical loading conditions. Changes in cell morphology were evaluated through: cell and nucleus elongation, cell alignment and junctional actin production. The results demonstrated that the combined FSS-CS stimulation induced visible changes in EC morphology. Upon simultaneous fluid shear stress and biaxial tensile strain stimulation, cells were elongated and generally aligned with the flow direction, with stress fibers highlighted along the cell junctions. The concurrent stimulation from shear stress and biaxial cyclic stretch led to a significant increase in cell elongation compared to untreated cells. This, however, was significantly lower than that induced by shear stress alone, indicating that the biaxial tensile strain may counteract the elongating effect of shear stress to maintain the shape of ECs. A similar trend was seen in alignment, where the alignment induced by the concurrent application of shear stress and cyclic stretch fell in between that induced by shear stress and tensile stretch alone, indicating the opposite role shear stress and tensile strain may play in cell alignment. Junctional actin accumulation was increased upon shear stress alone or simultaneously with tensile stretch. Tensile stretch alone did not change junctional actin accumulation, indicating the dominant role of shear stress in damaging EC junctions. These results demonstrate that the shearing-stretching device is capable of applying well characterized dynamic shear stress and tensile strain to cultured ECs. Using this device, EC response to altered mechanical environment in vivo can be characterized in vitro.

Keywords: cyclic stretch, endothelial cells, fluid shear stress, vascular biology

Procedia PDF Downloads 378

Authors: Kevin Kellner, Nga Lao, Orla Coleman, Paula Meleady, Niall Barron

Abstract:

Chinese Hamster Ovary (CHO) cells are the prominent cell line used in biopharmaceutical production. To improve yields and find beneficial bioprocess phenotypes genetic engineering plays an essential role in recent research. The miR-23 cluster, specifically miR-24 and miR-27, was first identified as differentially expressed during hypothermic conditions suggesting a role in proliferation and productivity in CHO cells. In this study, we used sponge decoy technology to stably deplete the miRNA expression of the cluster. Furthermore, we implemented the CRISPR/Cas9 system to knockdown miRNA expression. Sponge constructs were designed for an imperfect binding of the miRNA target, protecting from RISC mediated cleavage. GuideRNAs for the CRISPR/Cas9 system were designed to target the seed region of the miRNA. The expression of mature miRNA and precursor were confirmed using RT-qPCR. For both approaches stable expressing mixed populations were generated and characterised in batch cultures. It was shown, that CRISPR/Cas9 can be implemented in CHO cells with achieving high knockdown efficacy of every single member of the cluster. Targeting of one miRNA member showed that its genomic paralog is successfully targeted as well. The stable depletion of miR-24 using CRISPR/Cas9 showed increased growth and specific productivity in a CHO-K1 mAb expressing cell line. This phenotype was further characterized using quantitative label-free LC-MS/MS showing 186 proteins differently expressed with 19 involved in proliferation and 26 involved in protein folding/translation. Targeting miR-27 in the same cell line showed increased viability in late stages of the culture compared to the control. To evaluate the phenotype in an industry relevant cell line; the miR-23 cluster, miR-24 and miR-27 were stably depleted in a Fc fusion CHO-S cell line which showed increased batch titers up to 1.5-fold. In this work, we highlighted that the stable depletion of the miR-23 cluster and its members can improve the bioprocess phenotype concerning growth and productivity in two different cell lines. Furthermore, we showed that using CRISPR/Cas9 is comparable to the traditional sponge decoy technology.

Keywords: Chinese Hamster ovary cells, CRISPR/Cas9, microRNAs, sponge decoy technology

Procedia PDF Downloads 200

Authors: Rudra Vaghela, P. K. Kulkarni

Abstract:

The aim of the present research was to develop oral Amphotericin B (AmB) nanocrystals (Nc) grafted with suitable ligand in order to enhance drug transport across the intestinal epithelial barrier and subsequently, active uptake by macrophages. AmB Nc were prepared by liquid anti-solvent precipitation technique (LAS). Poloxamer 188 was used to stabilize the prepared AmB Nc and grafted with mannose for actively targeting M cells in Peyer’s patches. To prevent shedding of the stabilizer and ligand, N,N’-Dicyclohexylcarbodiimide (DCC) was used as a cross-linker. The prepared AmB Nc were characterized for particle size, PDI, zeta potential, X-ray diffraction (XRD) and surface morphology using scanning electron microscope (SEM) and evaluated for drug content, in vitro drug release and cell uptake studies using caco-2 cells. The particle size of stabilized AmB Nc grafted with WGA was in the range of 287-417 nm with negative zeta potential between -18 to -25 mV. XRD studies revealed crystalline nature of AmB Nc. SEM studies revealed that ungrafted AmB Nc were irregular in shape with rough surface whereas, grafted AmB Nc were found to be rod-shaped with smooth surface. In vitro drug release of AmB Nc was found to be 86% at the end of one hour. Cellular studies revealed higher invasion and uptake of AmB Nc towards caco-2 cell membrane when compared to ungrafted AmB Nc. Our findings emphasize scope on developing oral delivery system for passively targeting M cells in Peyer’s patches.

Keywords: leishmaniasis, amphotericin b nanocrystals, macrophage targeting, LAS technique

Procedia PDF Downloads 304

Authors: Gabriel Hunduma

Abstract:

Asymptomatic intra-thoracic neurogenic tumours are rare. Tumours arising from the intercostal nerves of the chest wall are exceedingly rare. This paper reports an incidental discovery of a neurogenic intercostal tumour while being investigated for Coronavirus Disease 2019 (COVID-19). A 54-year-old female underwent a thoracotomy and resection for an intercostal tumour. Pre-operative images showed an intrathoracic mass, and the biopsy revealed a schwannoma. The most common presenting symptom recorded in literature is chest pain; however, our case remained asymptomatic despite the size of the mass and thoracic area it occupied. After an extensive search of the literature, COVID-19 was found to have an influence on the development of certain cells in breast cancer. Hence there is a possibility that COVID-19 played a role in progressing the development of the schwannoma cells.

Keywords: thoracic surgery, intercostal schwannoma, chest wall oncology, COVID-19

Procedia PDF Downloads 214

Authors: Omar Nouri, Wafa Mnejja, Nejla Fourati, Fatma Dhouib, Wicem Siala, Ilhem Charfeddine, Afef Khanfir, Jamel Daoud

Abstract:

Background and objective: Induction chemotherapy (IC) followed by concomitant chemo radiotherapy with intensity modulated radiation (IMRT) technique is actually the recommended treatment modality for locally advanced nasopharyngeal carcinomas (NPC). The aim of this study was to evaluate the prognostic factors predicting loco regional relapse with this new treatment protocol. Patients and methods: A retrospective study of 52 patients with NPC treated between June 2016 and July 2019. All patients received IC according to the protocol of the Head and Neck Radiotherapy Oncology Group (Gortec) NPC 2006 (3 TPF courses) followed by concomitant chemo radiotherapy with weekly cisplatin (40 mg / m2). Patients received IMRT with integrated simultaneous boost (SIB) of 33 daily fractions at a dose of 69.96 Gy for high-risk volume, 60 Gy for intermediate risk volume and 54 Gy for low-risk volume. Median age was 49 years (19-69) with a sex ratio of 3.3. Forty five tumors (86.5%) were classified as stages III - IV according to the 2017 UICC TNM classification. Loco regional relapse (LRR) was defined as a local and/or regional progression that occurs at least 6 months after the end of treatment. Survival analysis was performed according to Kaplan-Meier method and Log-rank test was used to compare anatomy clinical and therapeutic factors that may influence loco regional free survival (LRFS). Results: After a median follow up of 42 months, 6 patients (11.5%) experienced LRR. A metastatic relapse was also noted for 3 of these patients (50%). Target volumes coverage was optimal for all patient with LRR. Four relapses (66.6%) were in high-risk target volume and two (33.3%) were borderline. Three years LRFS was 85,9%. Four factors predicted loco regional relapses: histologic type other than undifferentiated (UCNT) (p=0.027), a macroscopic pre chemotherapy tumor volume exceeding 100 cm³ (p=0.005), a reduction in IC doses exceeding 20% (p=0.016) and a total cumulative cisplatin dose less than 380 mg/m² (p=0.0.34). TNM classification and response to IC did not impact loco regional relapses. Conclusion: For nasopharyngeal carcinoma, tumors with initial high volume and/or histologic type other than UCNT, have a higher risk of loco regional relapse. Therefore, they require a more aggressive therapeutic approaches and a suitable monitoring protocol.

Keywords: loco regional relapse, modulation intensity radiotherapy, nasopharyngeal carcinoma, prognostic factors

Procedia PDF Downloads 128

Authors: H. Zouaoui, D. Abdi, B. Nessark, F. Habelhames, A. Bahloul

Abstract:

Among the conjugated organic polymers, the polythiophenes constitute a particularly important class of conjugated polymers, which has been extensively studied for the relation between the geometrical structure and the optic and electronic properties, while the polythiophene is an intractable material. They are, furthermore, chemically and thermally stable materials, and are very attractive for exploitation of their physical properties. The polythiophenes are extensively studied due to the possibility of synthesizing low band gap materials by using substituted thiophenes as precursors. Low band gap polymers may convert visible light into electricity and some photoelectrochemical cells based on these materials have been prepared. Polythiophenes (PThs) are good candidates for polymer optoelectronic devices such as polymer solar cells (PSCs) polymer light-emitting diodes (PLEDs) field-effect transistors (FETs) electrochromics and biosensors. In this work, MnO2 has been synthesized by hydrothermal method and analyzed by infrared spectroscopy. The polybithiophene+MnO2 composite films were electrochemically prepared by cyclic voltammetry technic on a conductor glass substrate ITO (indium–tin-oxide). The composite films are characterized by cyclic voltammetry, impedance spectroscopy and photoelectrochemical analyses. The results confirmed the presence of manganese dioxide nanoparticles in the polymer layer. An application has been made by using these deposits as an electrode in a photoelectrochemical cell for measuring photocurrent tests. The composite films show a significant photocurrent intensity 80 μA.cm-2.

Keywords: polybithiophene, MnO2, photoelectrochemical cells, composite films

Procedia PDF Downloads 353

Authors: Cóndor C. José, Rodrigues E. Camila, Noronha L. Irene, Dos Santos Fernando, Irigoyen M. Claudia, Andrade Lúcia

Abstract:

Sepsis induces alterations in hemodynamics and autonomic nervous system (ASN). The autonomic activity can be calculated by measuring heart rate variability (HRV) that represents the complex interplay between ASN and cardiac pacemaker cells. Wharton’s jelly mesenchymal stem cells (WJ-MSCs) are known to express genes and secreted factors involved in neuroprotective and immunological effects, also to improve the survival in experimental septic animals. We hypothesized, that WJ-MSCs present an important role in the autonomic activity and in the hemodynamic effects in a cecal ligation and puncture (CLP) model of sepsis. Methods: We used flow cytometry to evaluate WJ-MSCs phenotypes. We divided Wistar rats into groups: sham (shamoperated); CLP; and CLP+MSC (106 WJ-MSCs, i.p., 6 h after CLP). At 24 h post-CLP, we recorded the systolic arterial pressure (SAP) and heart rate (HR) over 20 min. The spectral analysis of HR and SAP; also the spontaneous baroreflex sensitivity (measure by bradycardic and tachycardic responses) were evaluated after recording. The one-way ANOVA and the post hoc Student– Newman– Keuls tests (P< 0.05) were used to data comparison Results: WJ-MSCs were negative for CD3, CD34, CD45 and HLA-DR, whereas they were positive for CD73, CD90 and CD105. The CLP group showed a reduction in variance of overall variability and in high-frequency power of HR (heart parasympathetic activity); furthermore, there is a low-frequency reduction of SAP (blood vessels sympathetic activity). The treatment with WJ-MSCs improved the autonomic activity by increasing the high and lowfrequency power; and restore the baroreflex sensitive. Conclusions: WJ-MSCs attenuate the impairment of autonomic control of the heart and vessels and might therefore play a protective role in sepsis. (Supported by FAPESP).

Keywords: baroreflex response, heart rate variability, sepsis, wharton’s jelly-derived mesenchymal stem cells

Procedia PDF Downloads 303

Authors: Magdalena Rusady Goey, Gordon Strathdee, Neil Perkins

Abstract:

Background: Acute lymphoblastic leukaemia (ALL) is the most frequent type of childhood malignancy, accounting for 25% of all cases. TWIST2, a basic helix-loop-helix transcription factor, has been implicated in ALL development. Prior studies found that TWIST2 undergoes epigenetic silencing in more than 50% cases of ALL through promoter hypermethylation and suggested that re-expression of TWIST2 may inhibit cell growth/survival of leukaemia cell lines. TWIST2 has also been implicated as a regulator of NF-kappaB activity, which is constitutively active in leukaemia. Here, we use a lentiviral transductions system to confirm the importance of TWIST2 in controlling leukaemia cell growth and to investigate whether this is achieved through altered regulation of NF-kappaB activity. Method: Re-expression of TWIST2 in leukaemia cell lines was achieved using lentiviral-based transduction. The lentiviral vector also expresses enhanced green fluorescent protein (eGFP), allowing transduced cells to be tracked using flow cytometry. Analysis of apoptosis and cell proliferation were done using annexinV and VPD450 staining, respectively. Result and Discussion: TWIST2-expressing cells were rapidly depleted from a mixed population in ALL cell lines (NALM6 and Reh), indicating that TWIST2 inhibited cell growth/survival of ALL cells. In contrast, myeloid cell lines (HL60 and K562) were comparatively insensitive to TWIST2 re-expression. Analysis of apoptosis and cell proliferation found no significant induction of apoptosis, but did find a rapid induction of proliferation arrest in TWIST2-expressing Reh and NALM6 cells. Initial experiment with NF-kappaB inhibitor demonstrated that inhibition of NF-kappaB has similar impact on cell proliferation in the ALL cell lines, suggesting that TWITST2 may induce cell proliferation arrest through inhibition of NF-kappaB. Conclusion: The results of this study suggest that epigenetic inactivation of TWIST2 in primary ALL leads to increased proliferation, potentially by altering the regulation of NF-kappaB.

Keywords: leukaemia, acute lymphoblastic leukaemia, NF-kappaB, TWIST2, lentivirus

Procedia PDF Downloads 346