Search results for: in vitro cell line study

Authors: Cheng-Cao Sun, Shu-Jun Li, Cuili Yang, Yongyong Xi, Liang Wang, Feng Zhang, De-Jia Li

Abstract:

Hsa-miRNA-326 (miR-326) has recently been discovered having anticancer efficacy in different organs. However, the role of miR-326 on non-small cell lung cancer (NSCLC) is still ambiguous. In this study, we investigated the role of miR-326 on the development of NSCLC. The results indicated that miR-326 was significantly down-regulated in primary tumor tissues and very low levels were found in NSCLC cell lines. Ectopic expression of miR-326 in NSCLC cell lines significantly suppressed cell growth as evidenced by cell viability assay, colony formation assay and BrdU staining, through inhibition of cyclin D1, cyclin D2, CDK4, and up-regulation of p57(Kip2) and p21(Waf1/Cip1). In addition, miR-326 induced apoptosis, as indicated by concomitantly with up-regulation of key apoptosis protein cleaved caspase-3, and down-regulation of anti-apoptosis protein Bcl2. Moreover, miR-326 inhibited cellular migration and invasiveness through inhibition of matrix metalloproteinases (MMP)-7 and MMP-9. Further, oncogene CCND1 was revealed to be a putative target of miR-326, which was inversely correlated with miR-326 expression in NSCLC. Taken together, our results demonstrated that miR-326 played a pivotal role on NSCLC through inhibiting cell proliferation, migration, invasion, and promoting apoptosis by targeting oncogenic CCND1.

Keywords: hsa-miRNA-326 (miR-326), cyclin D1, non-small cell lung cancer (NSCLC), proliferation, apoptosis

Procedia PDF Downloads 276

Authors: F. Bosca, F. Foglietta, F. Turci, E. Calcio Gaudino, S. Mana, F. Dosio, R. Canaparo, L. Serpe, A. Barge

Abstract:

In recent years, medical science has improved chemotherapy, radiation therapy and adjuvant therapy and has developed newer targeted therapies as well as refining surgical techniques for removing cancer. However, the chances of surviving the disease depend greatly on the type and location of the cancer and the extent of the disease at the start of treatment. Moreover, mainstream forms of cancer treatment have side effects which range from the unpleasant to the fatal. Therefore, the continuation of progress in anti-cancer therapy may depend on placing emphasis on other existing but less thoroughly investigated therapeutic approaches such as Sonodynamic Therapy (SDT). SDT is based on the local activation of a so called 'sonosensitizer', a molecule able to be excited by ultrasound, the radical production as a consequence of its relaxation processes and cell death due to different mechanisms induced by radical production. The present work deals with synthesis, characterization and preliminary in vitro test of Single Walled Carbon Nanotubes (SWCNT) decorated with porphyrins and biological vectors. The SWCNT’s surface was modified exploiting 1, 3-dipolar cycloaddition or Dies Alder reactions. For this purpose, different porphyrins scaffolds were ad-hoc synthesized using also non-conventional techniques. To increase cellular specificity of porphyrin-conjugated SWCNTs and to improve their ability to be suspended in aqueous solution, the modified nano-tubes were grafted with suitable glutamine or hyaluronic acid derivatives. These nano-sized sonosensitizers were characterized by several methodologies and tested in vitro on different cancer cell lines.

Keywords: sonodynamic therapy, porphyrins synthesis and modification, SWNCT grafting, hyaluronic acid, anti-cancer treatment

Procedia PDF Downloads 368

Authors: Martyna Chotomska, Aleksandra Studzinska, Marta Lisowska, Justyna Szubert, Aleksandra Tabis, Jacek Bania, Arkadiusz Miazek

Abstract:

Objectives: Assessing antigen-specific T cell responses is of utmost importance for the pre-clinical testing of prototype vaccines against intracellular pathogens and tumor antigens. Mainly two types of in vitro assays are used for this purpose 1) enzyme-linked immunospot (ELISpot) and 2) intracellular cytokine staining (ICS). Both are time-consuming, relatively expensive, and require manual dexterity. Here, we assess if a straightforward detection of luciferase activity in blood samples of transgenic reporter mice expressing a secreted Lucia luciferase under the transcriptional control of IFN-γ promoter parallels the sensitivity of IFNγ ELISpot assay. Methods: IFN-γ-LUCIA mouse strain carrying multiple copies of Lucia luciferase transgene under the transcriptional control of IFNγ minimal promoter were generated by pronuclear injection of linear DNA. The specificity of transgene expression and mobilization was assessed in vitro using transgenic splenocytes exposed to various mitogens. The IFN-γ-LUCIA mice were immunized with 50mg of ovalbumin (OVA) emulsified in incomplete Freund’s adjuvant three times every two weeks by subcutaneous injections. Blood samples were collected before and five days after each immunization. Luciferase activity was assessed in blood serum. Peripheral blood mononuclear cells were separated and assessed for frequencies of OVA-specific IFNγ-secreting T cells. Results: We show that in vitro cultured splenocytes of IFN-γ-LUCIA mice respond by 2 and 3 fold increase in secreted luciferase activity to T cell mitogens concanavalin A and phorbol myristate acetate, respectively but fail to respond to B cell-stimulating E.coli lipopolysaccharide. Immunization of IFN-γ-LUCIA mice with OVA leads to over 4 fold increase in luciferase activity in blood serum five days post-immunization with a barely detectable increase in OVA-specific, IFNγ-secreting T cells by ELISpot. Second and third immunizations, further increase the luciferase activity and coincidently also increase the frequencies of OVA-specific T cells by ELISpot. Conclusions: We conclude that minimally invasive monitoring of luciferase secretions in blood serum of IFN-γ-LUCIA mice constitutes a sensitive method for evaluating primary and memory Th1 responses to protein antigens. As such, this method may complement existing methods for rapid immunogenicity assessment of prototype vaccines.

Keywords: ELISpot, immunogenicity, interferon-gamma, reporter mice, vaccines

Procedia PDF Downloads 143

Authors: Yogesh D. Bansod, Jiri Bursa

Abstract:

The quantitative study of cell mechanics is of paramount interest since it regulates the behavior of the living cells in response to the myriad of extracellular and intracellular mechanical stimuli. The novel experimental techniques together with robust computational approaches have given rise to new theories and models, which describe cell mechanics as a combination of biomechanical and biochemical processes. This review paper encapsulates the existing continuum-based computational approaches that have been developed for interpreting the mechanical responses of living cells under different loading and boundary conditions. The salient features and drawbacks of each model are discussed from both structural and biological points of view. This discussion can contribute to the development of even more precise and realistic computational models of cell mechanics based on continuum approaches or on their combination with microstructural approaches, which in turn may provide a better understanding of mechanotransduction in living cells.

Keywords: cell mechanics, computational models, continuum approach, mechanical models

Procedia PDF Downloads 336

Authors: Hicham Gourgue, Ahmed Aharoune, Ahmed Ihlal

Abstract:

We present in this work our model of road traffic emissions (line sources) and dispersion of these emissions, named DISPOLSPEM (Dispersion of Poly Sources and Pollutants Emission Model). In its emission part, this model was designed to keep the consistent bottom-up and top-down approaches. It also allows to generate emission inventories from reduced input parameters being adapted to existing conditions in Morocco and in the other developing countries. While several simplifications are made, all the performance of the model results are kept. A further important advantage of the model is that it allows the uncertainty calculation and emission rate uncertainty according to each of the input parameters. In the dispersion part of the model, an improved line source model has been developed, implemented and tested against a reference solution. It provides improvement in accuracy over previous formulas of line source Gaussian plume model, without being too demanding in terms of computational resources. In the case study presented here, the biggest errors were associated with the ends of line source sections; these errors will be canceled by adjacent sections of line sources during the simulation of a road network. In cases where the wind is parallel to the source line, the use of the combination discretized source and analytical line source formulas minimizes remarkably the error. Because this combination is applied only for a small number of wind directions, it should not excessively increase the calculation time.

Keywords: air pollution, dispersion, emissions, line sources, road traffic, urban transport

Procedia PDF Downloads 416

Authors: Minhee Son, Kyung Nam Kim

Abstract:

This study explores the plan of the Korean and Japanese governments to transition into the hydrogen economy. Two motor companies, Hyundai Motor Company from Korea and Toyota from Japan, released the Hydrogen Fuel Cell Vehicle to monopolize the green energy automobile market. Although, they are the main countries which emit greenhouse gas, hydrogen energy can bring from a certain industry places, such as chemical plants and steel mills. Recent, the two countries have been focusing on the hydrogen industry including a fuel cell vehicle, a hydrogen station, a fuel cell plant, a residential fuel cell. The purpose of this paper is to find out the differences of the policies in the two countries to be hydrogen societies. We analyze the behavior of the public and private sectors in Korea and Japan about hydrogen energy and fuel cells for the transition of the hydrogen economy. Finally we show the similarities and differences of both countries in hydrogen fuel cells. And some cities have feature such as Hydrogen cities. Hydrogen energy can make impact environmental sustainability.

Keywords: fuel cell, hydrogen city, hydrogen fuel cell vehicle, hydrogen station, hydrogen energy

Procedia PDF Downloads 453

Authors: S. Bouchami, M.N. Lakhoua

Abstract:

The automotive market is a dynamic market that continues to grow. That’s why several companies belonging to this sector adopt a quality improvement approach. Wanting to be competitive and successful in the environment in which they operate, these companies are dedicated to establishing a system of quality management to ensure the achievement of the objective quality, improving the products and process as well as the satisfaction of the customers. In this paper, the management of the quality and the improvement of a production line in an industrial company is presented. In fact, the project is divided into two essential parts: the creation of the technical line documentation and the quality assurance documentation and the resolution of defects at the line, as well as those claimed by the customer. The creation of the documents has required a deep understanding of the manufacturing process. The analysis and problem solving were done through the implementation of PDCA (Plan Do Check Act) and FTA (Fault Tree Analysis). As perspective, in order to better optimize production and improve the efficiency of the production line, a study on the problems associated with the supply of raw materials should be made to solve the problems of stock-outs which cause delays penalizing for the industrial company.

Keywords: quality management, documentary system, Plan Do Check Act (PDCA), fault tree analysis (FTA) method

Procedia PDF Downloads 118

Authors: M. Rahimnejad, B. Vahidi, B. Ebrahimi Hoseinzadeh, F. Yazdian, P. Motamed Fath, R. Jamjah

Abstract:

In this study, we developed and simulated nano-drug delivery systems efficacy in compare to free drug prescription. Computational models can be utilized to accelerate experimental steps and control the experiments high cost. Molecular dynamics simulation (MDS), in particular NAMD was utilized to better understand the anti-cancer drug interaction with cell membrane model. Paclitaxel (PTX) and dipalmitoylphosphatidylcholine (DPPC) were selected for the drug molecule and as a natural phospholipid nanocarrier, respectively. This work focused on two important interaction parameters between molecules in terms of center of mass (COM) and van der Waals interaction energy. Furthermore, we compared the simulation results of the PTX interaction with the cell membrane and the interaction of DPPC as a nanocarrier loaded by the drug with the cell membrane. The molecular dynamic analysis resulted in low energy between the nanocarrier and the cell membrane as well as significant decrease of COM amount in the nanocarrier and the cell membrane system during the interaction. Thus, the drug vehicle showed notably better interaction with the cell membrane in compared to free drug interaction with the cell membrane.

Keywords: anti-cancer drug, center of mass, interaction energy, molecular dynamics simulation, nanocarrier

Procedia PDF Downloads 310

Authors: H. Rezzouk, M. Hatti, H. Rahmani, S. Atoui

Abstract:

This paper discusses the design and analysis of a hybrid PV-Fuel cell energy system destined to power a DC load. The system is composed of a photovoltaic array, a fuel cell, an electrolyzer and a hydrogen tank. HOMER software is used in this study to calculate the optimum capacities of the power system components that their combination allows an efficient use of solar resource to cover the hourly load needs. The optimal system sizing allows establishing the right balance between the daily electrical energy produced by the power system and the daily electrical energy consumed by the DC load using a 28 KW PV array, a 7.5 KW fuel cell, a 40KW electrolyzer and a 270 Kg hydrogen tank. The variation of powers involved into the DC bus of the hybrid PV-fuel cell system has been computed and analyzed for each hour over one year: the output powers of the PV array and the fuel cell, the input power of the elctrolyzer system and the DC primary load. Equally, the annual variation of stored hydrogen produced by the electrolyzer has been assessed. The PV array contributes in the power system with 82% whereas the fuel cell produces 18%. 38% of the total energy consumption belongs to the DC primary load while the rest goes to the electrolyzer.

Keywords: electrolyzer, hydrogen, hydrogen fueled cell, photovoltaic

Procedia PDF Downloads 459

Authors: Yingjeng James Li, Yun Jyun Ou, Chih Chi Hsu, Chiao-Chih Hu

Abstract:

A fuel cell is a device that produces electric power by reacting fuel and oxidant electrochemically. There is no pollution produced from a fuel cell if hydrogen is employed as the fuel. Therefore, a fuel cell is considered as a zero emission device and is a source of green power. A membrane electrode assembly (MEA) is the key component of a fuel cell. It is, therefore, beneficial to develop MEAs with high performance. In this study, an MEA for proton exchange membrane fuel cell (PEMFC) was prepared from a 15-micron thick reinforced PEM. The active area of such MEA is 25 cm2. Carbon supported platinum (Pt/C) was employed as the catalyst for both anode and cathode. The platinum loading is 0.6 mg/cm2 based on the sum of anode and cathode. Commercially available carbon papers coated with a micro porous layer (MPL) serve as gas diffusion layers (GDLs). The original thickness of the GDL is 250 μm. It was compressed down to 163 μm when assembled into the single cell test fixture. Polarization curves were taken by using eight different test conditions. At our standard test condition (cell: 70 °C; anode: pure hydrogen, 100%RH, 1.2 stoic, ambient pressure; cathode: air, 100%RH, 3.0 stoic, ambient pressure), the cell current density is 1250 mA/cm2 at 0.6 V, and 2400 mA/cm2 at 0.4 V. At self-humidified condition and cell temperature of 55 °C, the cell current density is 1050 mA/cm2 at 0.6 V, and 2250 mA/cm2 at 0.4 V. Hydrogen crossover rate of the MEA is 0.0108 mL/min*cm2 according to linear sweep voltammetry experiments. According to the MEA’s Pt loading and the cyclic voltammetry experiments, the Pt electrochemical surface area is 60 m2/g. The ohmic part of the impedance spectroscopy results shows that the membrane resistance is about 60 mΩ*cm2 when the MEA is operated at 0.6 V.

Keywords: fuel cell, membrane electrode assembly, proton exchange membrane, reinforced

Procedia PDF Downloads 270

Authors: Rohana Abdullah, Md Nizam Abd Rahman, Seri Rahayu Kamat

Abstract:

Operator efficiency aspect is gaining importance in ensuring optimized usage of resources especially in the semi-automated manufacturing environment. This paper addresses a case study done to solve operator efficiency and line balancing issue at a semiconductor assembly and test manufacturing. A Man-to-Machine (M2M) work study technique is used to study operator current utilization and determine the optimum allocation of the operators to the machines. Critical factors such as operator activity, activity frequency and operator competency level are considered to gain insight on the parameters that affects the operator utilization. Equipment standard time and overall equipment efficiency (OEE) information are also gathered and analyzed to achieve a balanced and optimized production.

Keywords: operator efficiency, optimized production, line balancing, industrial and manufacturing engineering

Procedia PDF Downloads 703

Authors: Leo Nnamdi Ozurumba-Dwight

Abstract:

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

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

Procedia PDF Downloads 99

Authors: Anna-Mari Kok, Carel B. Oosthuizen, Namrita Lall

Abstract:

Tuberculosis (TB) is a disease caused by the bacterial pathogen Mycobacterium tuberculosis. It is associated with high mortality rates in both developing and developed countries. Many higher plants are found that are medicinally associated with tuberculosis infection. Plants belonging to thirteen families were selected, based on their traditional usage for tuberculosis and its associated symptoms. Eight plants showed the best antimycobacterial activities (MIC-value ≤ 500.0 µg/ml) against M. tuberculosis H37Rv. LS was found to have a minimum inhibitory concentration (MIC) of 125 µg/ml whereas, Tulbaghia violacea, Heteromorpha arborescens, Sutherlandia frutescens, Eucalyptus deglupta, and Plectranthus neochilus were found to have a MIC value of 250 µg/ml against M. tuberculosis H37Rv. Cytotoxicity values on U937 and HepG2 cells were obtained and the IC50 values ranged between 40 ±4.30 and > 400 µg/ml for the U937 cell line and 72.4 ±1.50 and > 400 µg/ml for the HepG2 cell line. Heteromorpha arborescens had the lowest IC50 value in both cell lines and therefore showed moderate levels of toxicity. Of the 19 samples that underwent the 2, 2- diphenyl- 1- picrylhydrazyl (DPPH) antioxidant assay, Eucalyptus deglupta and Melianthus major showed significant free radical scavenging activities with concentrations of 1.33 and 1.32 µg/ml respectively for the inhibition of DPPH. Hepatotoxicity induced by acetaminophen identified Searsia lancea with hepatoprotective activity of 59.37% at a ¼ IC50 concentration. Out of the 7 samples that were investigated for their immunomodulatory capabilities, Eucalyptus deglupta produced the most IL-12 with Sutherlandia frutescens also showing positive results for IL-12 production. In the present study, Eucalyptus deglupta showed the most promising results with good activity against M. tuberculosis with an MIC-value of 250 µg/ml. It also has potent antioxidant activity with an IC50 value of 1.33 µg/ml. This sample also stimulated high production of the cytokine, IL-12. Searsia lancea showed moderate antimycobacterial acticvity with an MIC-value of 500 µg/ml. The antioxidant potential also showed promising results with an IC50 value of 4.50 µg/ml. The hepatoprotective capability of Searsia lancea was 59.34% at a ¼ IC50 concentration. Another sample Sutherlandia frutescens showed effective antimycobacterial activity with an MIC-value of 250 µg/ml. It also stimulated production of IL-12 with 13.43 pg/ml produced. These three samples can be considered for further studies for the consideration as adjuvants for current tuberculosis treatment.

Keywords: adjuvant, hepatoprotection, immunomodulation, tuberculosis

Procedia PDF Downloads 279

Authors: Cheng-Cao Sun, Shu-Jun Li, Cuili Yang, Yongyong Xi, Liang Wang, Feng Zhang, De-Jia Li

Abstract:

MicroRNAs (miRNAs) act as key regulators of multiple cancers. Hsa-miR-329 (miR-329) functions as a tumor suppressor in some malignancies. However, its role on lung cancer remains poorly understood. In this study, we investigated the role of miR-329 on the development of lung cancer. The results indicated that miR-329 was decreased in primary lung cancer tissues compared with matched adjacent normal lung tissues and very low levels were found in a non-small cell lung cancer (NSCLC) cell lines. Ectopic expression of miR-329 in lung cancer cell lines substantially repressed cell growth as evidenced by cell viability assay, colony formation assay and BrdU staining, through inhibiting cyclin D1, cyclin D2, and up-regulatiing p57(Kip2) and p21(WAF1/CIP1). In addition, miR-329 promoted NSCLC cell apoptosis, as indicated by up-regulation of key apoptosis gene cleaved caspase-3, and down-regulation of anti-apoptosis gene Bcl2. Moreover, miR-329 inhibited cellular migration and invasiveness through inhibiting matrix metalloproteinases (MMP)-7 and MMP-9. Further, oncogene MET was revealed to be a putative target of miR-329, which was inversely correlated with miR-329 expression. Furthermore, down-regulation of MET by siRNA performed similar effects to over-expression of miR-329. Collectively, our results demonstrated that miR-329 played a pivotal role in lung cancer through inhibiting cell proliferation, migration, invasion, and promoting apoptosis by targeting oncogenic MET.

Keywords: hsa-miRNA-329(miR-329), MET, non-small cell lung cancer (NSCLC), proliferation, apoptosis

Procedia PDF Downloads 378

Authors: C. Kridtayopas, W. Danvilai, P. Sopannarath, A. Kayan, W. Loongyai

Abstract:

Both Betong chicken (KU Line) and Thai Native chickens were the high quality of the meat and low carcass fat compared to broiler chickens. The objective of this study was to determine the growth performance, carcass characteristic, meat quality and association of polymorphism in the ApoVLDL-II gene with fat accumulation in the female broiler, Thai Native and Betong (KU line) chickens at 4-14 weeks. The chickens were used and reared under the same environment and management (100 chicks per breed). The results showed that body weight (BW) of broiler chickens was significantly higher than Thai Native and Betong (KU line) chickens (P < 0.01) through all the experiment. At 4-8 weeks of age, feed conversion ratio (FCR) of broiler chickens was significantly better than Thai Native and Betong (KU line) chickens (P < 0.01), then increased at week 8-14. The percentage of breast, abdominal fat and subcutaneous fat of broiler chickens was significantly greater than Thai Native and Betong (KU line) chickens (P < 0.01). However, Thai Native chickens showed the highest percentage of liver (P < 0.01) when compared to other breeds. In addition, the percentage of wing of Thai Native and Betong (KU line) chickens were significantly (P < 0.01) higher than broiler chickens. Meat quality was also determined and found that, pH of breast meat left from slaughter 45 minutes (pH45) and 24 hours (pH24) of broiler was significantly higher than Thai Native and Betong (KU line) (P < 0.01) whereas the percentage of drip loss, thawing loss, cooking loss and shear force was not significantly different between breeds. The polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique was used to genotype the polymorphism in the ApoVLDL-II gene in the broiler, Thai Native and Betong (KU line) chickens. The results found that, the polymorphism in the ApoVLDL-II gene at VLDL6 loci was not associated with fat accumulation in those studied population.

Keywords: ApoVLDL-II gene, Betong (KU line) chickens, broiler chickens, carcass characteristic, growth performance, meat quality, Thai native chickens

Procedia PDF Downloads 178

Authors: Harika Atmaca, Emir Bozkurt, Latife Merve Oktay, Selim Uzunoglu, Ruchan Uslu, Burçak Karaca

Abstract:

Microarrays have been developed for highly parallel enzyme-linked immunosorbent assay (ELISA) applications. The most common protein arrays are produced by using multiple monoclonal antibodies, since they are robust molecules which can be easily handled and immobilized by standard procedures without loss of activity. Protein expression profiling with protein array technology allows simultaneous analysis of the protein expression pattern of a large number of proteins. Trabectedin, a tetrahydroisoquinoline alkaloid derived from a Caribbean tunicate, Ecteinascidia turbinata, has been shown to have antitumor effects. Here, we used a novel proteomic approach to explore the mechanism of action of trabectedin in prostate cancer cell line PC-3 by apoptosis antibody microarray. XTT cell proliferation kit and Cell Death Detection Elisa Plus Kit (Roche) was used for measuring cytotoxicity and apoptosis. Human Apoptosis Protein Array (R&D Systems) which consists of 35 apoptosis related proteins was used to assess the omic protein expression pattern. Trabectedin induced cytotoxicity and apoptosis in prostate cancer cells in a time and concentration-dependent manner. The expression levels of the death receptor pathway molecules, TRAIL-R1/DR4, TRAIL R2/DR5, TNF R1/TNFRSF1A, FADD were significantly increased by 4.0-, 21.0-, 4.20- and 11.5-fold by trabectedin treatment in PC-3 cells. Moreover, mitochondrial pathway related pro-apoptotic proteins Bax, Bad, Cytochrome c, and Cleaved Caspase-3 expressions were induced by 2.68-, 2.07-, 2.8-, and 4.5-fold and the expression levels of anti-apoptotic proteins Bcl-2 and Bcl-XL were reduced by 3.5- and 5.2-fold in PC-3 cells. Proteomic (antibody microarray) analysis suggests that the mechanism of action of trabectedin may be exerted via the induction of both intrinsic and extrinsic apoptotic pathways. The antibody microarray platform can be utilised to explore the molecular mechanism of action of novel anticancer agents.

Keywords: trabectedin, prostate cancer, omic protein expression profile, apoptosis

Procedia PDF Downloads 419

Authors: Celso Afonso Klein Jr., Henrique Cantarelli, Fernando Portella, Keiichi Hosaka, Eduardo Reston, Fabricio Collares, Roberto Zimmer

Abstract:

TTo evaluate the influence of preheating self-adhesive cement at 39ºC on cell migration, cytotoxicity and degree of conversion. RelyX U200, Set PP and MaxCem Elite were subjected to a degree of conversion analysis (FTIR-ATR). For the cytotoxicity analysis, extracts (24 h and 7 days) were placed in contact with NIH/3T3 cells. For cell migration, images were captured of each sample until the possible closure of the cleft occurred. In the results of the degree of conversion, preheating did not improve the conversion of cement. For the MTT, preheating did not improve the results within 24 hours. However, it generated positive results within 7 days for the Set PP resin cement. For cell migration, high rates of cell death were found in all groups. It is concluded that preheating at 39ºC caused a positive effect only in increasing the cell viability of the Set PP resin cement and that both materials analyzed are highly cytotoxic.

Keywords: dental cements, resin cements, degree of conversion, cytotoxicity, cell migration assays

Procedia PDF Downloads 38

Authors: Pinar Ercan, Sedef N. El

Abstract:

The goals of this study were to determine the total anthocyanin and procyanidin contents and their in vitro bioaccessibilities of apple, red grape and cinnamon by a static in vitro digestion method reported by the COST FA1005 Action INFOGEST, as well as in vitro inhibitory effects of these food samples on starch and lipid digestive enzymes. While the highest total anthocyanin content was found in red grape (164.76 ± 2.51 mg/100 g), the highest procyanidin content was found in cinnamon (6432.54±177.31 mg/100 g) among the selected food samples (p<0.05). The anthocyanin bioaccessibilities were found as 10.23±1 %, 8.23±0.64 %, and 8.73±0.70 % in apple, red grape, and cinnamon, respectively. The procyanidin bioaccessibilities of apple, red grape, and cinnamon were found as 17.57±0.71 %, 14.08±0.74 % and 18.75±1.49 %, respectively. The analyzed apple, red grape and cinnamon showed the inhibitory activity against α-glucosidase (IC50 544.27±21.94, 445.63±15.67, 1592±17.58 μg/mL, respectively), α-amylase (IC50 38.41±7.26, 56.12±3.60, 3.54±0.86 μg/mL, respectively), and lipase (IC50 52.65±2.05, 581.70±54.14, 49.63±2.72 μg/mL, respectively). Red grape sample showed the highest inhibitory activity against α-glucosidase, cinnamon showed the highest inhibitory activity against α-amylase and lipase according to IC50 (concentration of inhibitor required to produce a 50% inhibition of the initial rate of reaction) and Catechin equivalent inhibition capacity (CEIC50) values. This study reported that apple, grape and cinnamon samples can inhibit the activity of digestive enzymes in vitro. The consumption of these samples would be used in conjunction with a low-calorie diet for body weight management.

Keywords: anthocyanin, α-amylase, α-glucosidase, lipase, procyanidin

Procedia PDF Downloads 151

Authors: M. Marjaneh, M. Y. Narazah, H. Shahrul

Abstract:

Array-based gene expression analysis is a powerful tool to profile expression of genes and to generate information on therapeutic effects of new anti-cancer compounds. Anti-apoptotic effect of thymoquinone was studied in MCF7 breast cancer cell line using gene expression profiling with cDNA micro array. The purity and yield of RNA samples were determined using RNeasyPlus Mini kit. The Agilent RNA 6000 Nano LabChip kit evaluated the quantity of the RNA samples. AffinityScript RT oligo-dT promoter primer was used to generate cDNA strands. T7 RNA polymerase was used to convert cDNA to cRNA. The cRNA samples and human universal reference RNA were labelled with Cy-3-CTP and Cy-5-CTP, respectively. Feature Extraction and GeneSpring software analysed the data. The single experiment analysis revealed involvement of 64 pathways with up-regulated genes and 78 pathways with down-regulated genes. The MAPK and p38-MAPK pathways were inhibited due to the up-regulation of PTPRR gene. The inhibition of p38-MAPK suggested up-regulation of TGF-ß pathway. Inhibition of p38 - MAPK caused up-regulation of TP53 and down-regulation of Bcl2 genes indicating involvement of intrinsic apoptotic pathway. Down-regulation of CARD16 gene as an adaptor molecule regulated CASP1 and suggested necrosis-like programmed cell death and involvement of caspase in apoptosis. Furthermore, down-regulation of GPCR, EGF-EGFR signalling pathways suggested reduction of ER. Involvement of AhR pathway which control cytochrome P450 and glucuronidation pathways showed metabolism of Thymoquinone. The findings showed differential expression of several genes in apoptosis pathways with thymoquinone treatment in estrogen receptor-positive breast cancer cells.

Keywords: cDNA microarray, thymoquinone, CARD16, PTPRR, CASP10

Procedia PDF Downloads 324

Authors: Yu-Chen Hu

Abstract:

Gene editing by CRISPR and gene regulation by microRNA or CRISPR activation have dramatically changed the way to manipulate cellular gene expression and cell fate. In recent years, various gene editing and gene manipulation technologies have been applied to control stem cell differentiation to enhance tissue regeneration. This research will focus on how to develop CRISPR, CRISPR activation (CRISPRa), CRISPR inhibition (CRISPRi), as well as bi-directional CRISPR-AI gene regulation technologies to control cell differentiation and bone regeneration. Moreover, in this study, CRISPR/Cas13d-mediated RNA editng for miRNA editing and bone regeneration will be discussed.

Keywords: gene therapy, bone regeneration, stem cell, CRISPR, gene regulation

Procedia PDF Downloads 58

Authors: Taraneh Estedlal

Abstract:

The objective of this study was to review in-vivo and in-vitro studies to compare the effectiveness of bulk-fill and conventional resin composites with regard to marginal adaptation, polymerization shrinkage, and other mechanical properties.PubMed and Scopus databases was investigated for in-vitro studies and randomized clinical trials comparing incidence of fractures, color stability, marginal adaptation, pain and discomfort, recurrent caries, occlusion, pulpal reaction, and proper proximal contacts of restorations made with conventional and bulk resins. The failure rate of conventional and flowable bulk-fill resin composites was not significantly different to sculptable bulk-fill resin composites. The objective of this study was to review in-vivo and in-vitro studies to compare the effectiveness of bulk-fill and conventional resin composites with regard to marginal adaptation, polymerization shrinkage, and other mechanical properties. PubMed and Scopus databases was investigated for in-vitro studies and randomized clinical trials comparing one of the pearlier mentioned properties between bulk-fill and control composites. Despite differences in physical and in-vitro properties, failure rate of conventional and flowable bulk-fill resin composites was not significantly different to sculptable bulk-fill resin composites.

Keywords: polymerization shrinkage, color stability, marginal adaptation, recurrent caries, occlusion, pulpal reaction

Procedia PDF Downloads 128

Authors: Hadriana Bansi, Elizabeth Wina, Toto Toharmat

Abstract:

Acacia villosa is thornless shrub legume which contents high crude protein. However, the utilization of A. villosa as ruminant feed is limited by its secondary compounds. The aim of this article is to find out the maximum of substitution A. villosa in sheep ration. The nutritional evaluation consisted of in vitro two stages, in vivo, and in vitro gas production trials. The secondary compounds of A. villosa also were analyzed. Evaluating digestibility of increasing level of substitution A. villosa replacing Pennisetum purpureum was using in vitro two stages. The substitution of 30% A. villosa was compared to 100% P. purpureum by in vitro gas production technique and in vivo digestibility. The results of two stages in vitro showed that total phenol, condensed tannin, and non-protein amino acid (NPAA) were high. Substitution 15% A. villosa reached the highest digestibility for both dry matter (DM) and crude protein (CP) which were 67% and 86% respectively, but it was shown that DM and CP digestibility of substitution 30% of A. villosa was still high which were 61.82% and 75-67% respectively. The pattern of gas production showed that first 8 hours total gas production substitution of 30% A. villosa was higher than 100% P. purpureum and declined after 10 hours incubation. In vivo trials showed that substitution of 30% A. villosa significantly increased CP intake, CP digestibility, and nitrogen retention. It can be concluded that substitution A. villosa until 30% still gave the good impact even though it has high secondary compounds.

Keywords: Acacia villosa, digestibility, gas production, secondary compounds

Procedia PDF Downloads 131

Authors: Dalita G. S. M. Cavalcante, Elton A. P. dos Reis, Andressa S. Gomes, Caroline S. Danna, Leandra Ernest Kerche-Silva, Eidi Yoshihara, Aldo E. Job

Abstract:

In order to minimize environmental impacts, a composite was developed from mixture of leather shavings (LE) with natural rubber (NR), which patent is already deposited. The new material created can be used in applications such as floors e heels for shoes. Besides these applications, the aim is to use this new material for the production of products for the textile industry, such as boots, gloves and bags. But the question arises, as to biocompatibility of this new material. This is justified because the structure of the leather shavings has chrome. The trivalent chromium is usually not toxic, but the hexavalent chromium can be highly toxic and genotoxic for living beings, causing damage to the DNA molecule and contributing to the formation of cancer. Based on this, the objective of this study is evaluate the possible genotoxic effects of the new composite, using as system - test two cell lines (MRC-5 and CHO-K1) by comet assay. For this, the production of the composite was performed in three proportions: for every 100 grams of NR was added 40 (E40), 50 (E50) or 60 (E60) grams of LE. The latex was collected from the rubber tree (Hevea brasiliensis). For vulcanization of the NR, activators and accelerators were used. The two cell lines were exposed to the new composite in its three proportions using elution method, that is, cells exposed to liquid extracts obtained from the composite for 24 hours. For obtaining the liquid extract, each sample of the composite was crushed into pieces and mixed with an extraction solution. The quantification of total chromium and hexavalent chromium in the extracts were performed by Optical Emission Spectrometry by Inductively Coupled Plasma (ICP-OES). The levels of DNA damage in cells exposed to both extracts were monitored by alkaline version of the comet assay. The results of the quantification of metals in ICP-OES indicated the presence of total chromium in different extracts, but were not detected presence of hexavalent chromium in any extract. Through the comet assay were not found DNA damage of the CHO-K1 cells exposed to both extracts. As for MRC-5, was found a significant increase in DNA damage in cells exposed to E50 and E60. Based on the above data, it can be asserted that the extracts obtained from the composite were highly genotoxic for MRC-5 cells. These biological responses do not appear to be related to chromium metal, since there was a predominance of trivalent chromium in the extracts, indicating that during the production process of the new composite, there was no formation of hexavalent chromium. In conclusion it can infer that the leather shavings containing chromium can be reused, thereby reducing the environmental impacts of this waste. Already on the composite indicates to its incorporation in applications that do not aim at direct contact with the human skin, and it is suggested the chain of composite production be studied, in an attempt to make it biocompatible so that it may be safely used by the textile industry.

Keywords: cell line, chrome, genotoxicity, leather, natural rubber

Procedia PDF Downloads 180

Authors: Yunhui Chen, Damon Kent, Matthew Dargusch

Abstract:

Synthetic scaffolds are a highly promising new approach to replace both autografts and allografts to repair and remodel damaged bone tissue. Biocompatible porous titanium scaffold was manufactured through a powder metallurgy approach. Magnesium powder was used as space holder material which was compacted with titanium powder and removed during sintering. Evaluation of the porosity and mechanical properties showed a high level of compatibility with human bone. Interconnectivity between pores is higher than 95% for porosity as low as 30%. The elastic moduli are 39 GPa, 16 GPa and 9 GPa for 30%, 40% and 50% porosity samples which match well to that of natural bone (4-30 GPa). The yield strengths for 30% and 40% porosity samples of 315 MPa and 175 MPa are superior to that of human bone (130-180 MPa). In-vitro cell culture tests on the scaffold samples using Human Mesenchymal Stem Cells (hMSCs) demonstrated their biocompatibility and indicated osseointegration potential. The scaffolds allowed cells to adhere and spread both on the surface and inside the pore structures. With increasing levels of porosity/interconnectivity, improved cell proliferation is obtained within the pores. It is concluded that samples with 30% porosity exhibit the best biocompatibility. The results suggest that porous titanium scaffolds generated using this manufacturing route have excellent potential for hard tissue engineering applications.

Keywords: scaffolds, MG-63 cell culture, titanium, space holder

Procedia PDF Downloads 210

Authors: Ainoa Guinart, Maria Korpidou, Daniel Doellerer, Cornelia Palivan, Ben L. Feringa

Abstract:

Stimuli responsive systems arise from the need to meet unsolved needs of current molecular drugs. Our study presents the design of a delivery system with high spatiotemporal control and tuneable release profiles. We study the incorporation of a hydrophobic synthetic molecular motor into PDMS-b-PMOXA block copolymer vesicles to create a self-assembled system. We prove their successful incorporation and selective activation by low powered visible light (λ 430 nm, 6.9 mW). We trigger the release of a fluorescent dye with high release efficiencies over sequential cycles (up to 75%) with the ability to turn on and off the release behaviour on demand by light irradiation. Low concentrations of photo-responsive units are proven to trigger release down to 1 mol% of molecular motor. Finally, we test our system in relevant physiological conditions using a lung cancer cell line and the encapsulation of an approved drug. Similar levels of cell viability are observed compared to the free-given drugshowing the potential of our platform to deliver functional drugs on demand with the same efficiency and lower toxicity.

Keywords: molecular motor, polymer, drug delivery, light-responsive, cancer, selfassembly

Procedia PDF Downloads 97

Authors: Jinfeng Zhang, Chun-Sing Lee

Abstract:

The use of different nanocarriers for delivering hydrophobic pharmaceutical agents to tumor sites has garnered major attention. Despite the merits of these nanocarriers, further studies are needed for improving their drug loading capacities (typically less than 10%) and reducing their potential systemic toxicity. So development of alternative self-carried nanodrug delivery strategies without using any inert carriers is highly desirable. In this study, we developed a self-carried theranostic curcumin (Cur) nanodrug for highly effective cancer therapy in vitro and in vivo with real-time monitoring of drug release. With a biocompatible C18PMH-PEG functionalization, the Cur nanoparticles (NPs) showed excellent dispersibility and outstanding stability in physiological environment, with drug loading capacity higher than 78 wt.%. Both confocal microscopy and flow cytometry confirmed the cellular fluorescent “OFF-ON” activation and real-time monitoring of Cur molecule release, showing its potential for cancer diagnosis. In vitro and in vivo experiments clearly show that therapeutic efficacy of the PEGylated Cur NPs is much better than that of free Cur. This self-carried theranostic strategy with real-time monitoring of drug release may open a new way for simultaneous cancer therapy and diagnosis.

Keywords: drug delivery, in vitro and in vivo cancer therapy, real-time monitoring, self-carried

Procedia PDF Downloads 374

Authors: Elvin P. Chizenga, Heidi Abrahamse

Abstract:

Cancer cell resistance to therapy is the main cause of treatment failures and the poor prognosis of cancer convalescence. The progression of cervical cancer to other parts of the genitourinary system and the reported recurrence rates are overwhelming. Current treatments, including surgery, chemo and radiation have been inefficient in eradicating the tumor cells. These treatments are also associated with poor prognosis and reduced quality of life, including fertility loss. This has inspired the need for the development of new treatment modalities to eradicate cervical cancer successfully. Photodynamic Therapy (PDT) is a modern treatment modality that induces cell death by photochemical interactions of light and a photosensitizer, which in the presence of molecular oxygen, yields a set of chemical reactions that generate Reactive Oxygen Species (ROS) and other free radical species causing cell damage. Enhancing PDT using modified drug delivery can increase the concentration of the photosensitizer in the tumor cells, and this has the potential to maximize its therapeutic efficacy. In cervical cancer, all infected cells constitutively express genes of the E6 and E7 HPV viral oncoproteins, resulting in high concentrations of E6 and E7 in the cytoplasm. This provides an opportunity for active targeting of cervical cancer cells using immune-mediated drug delivery to maximize therapeutic efficacy. The use of nanoparticles in PDT has also proven effective in enhancing therapeutic efficacy. Gold nanoparticles (AuNps) in particular, are explored for their use in biomedicine due to their biocompatibility, low toxicity, and enhancement of drug uptake by tumor cells. In this present study, a biomolecule comprising of AuNPs, anti-E6 monoclonal antibodies, and Aluminium Phthalocyanine photosensitizer was synthesized for use in targeted PDT of cervical cancer. The AuNp-Anti-E6-Sulfonated Aluminium Phthalocyanine mix (AlPcSmix) photosensitizing biomolecule was synthesized by coupling AuNps and anti-E6 monoclonal antibodies to the AlPcSmix via Polyethylene Glycol (PEG) chemical links. The final product was characterized using Transmission Electron Microscope (TEM), Zeta Potential, Uv-Vis Spectrophotometry, Fourier Transform Infrared Spectroscopy (FTIR), and X-ray diffraction (XRD), to confirm its chemical structure and functionality. To observe its therapeutic role in treating cervical cancer, cervical cancer cells, HeLa cells were seeded in 3.4 cm² diameter culture dishes at a concentration of 5x10⁵ cells/ml, in vitro. The cells were treated with varying concentrations of the photosensitizing biomolecule and irradiated using a 673.2 nm wavelength of laser light. Post irradiation cellular responses were performed to observe changes in morphology, viability, proliferation, cytotoxicity, and cell death pathways induced. Dose-Dependent response of the cells to treatment was demonstrated as significant morphologic changes, increased cytotoxicity, and decreased cell viability and proliferation This study presented a synthetic biomolecule for targeted PDT of cervical cancer. The study suggested that PDT using this AuNp- Anti-E6- AlPcSmix photosensitizing biomolecule is a very effective treatment method for the eradication of cervical cancer cells, in vitro. Further studies in vivo need to be conducted to support the use of this biomolecule in treating cervical cancer in clinical settings.

Keywords: anti-E6 monoclonal antibody, cervical cancer, gold nanoparticles, photodynamic therapy

Procedia PDF Downloads 101

Authors: S. Vinotha, I. Thabrew, S. Sri Ranjani

Abstract:

Hot aqueous and methanol extracts of the two selected herbal medicines such are Vellarugu Chooranam (V.C) and Amukkirai Chooranam (A.C) were examined for total phenolic and flavonoid contents and in-vitro antioxidant activity using four different methods. The total phenolic and flavonoid contents in methanol extract of V.C were found to be higher (44.41±1.26 mg GAE⁄g; 174.44±9.32 mg QE⁄g) than in the methanol extract of A.C (20.56±0.67 mg GAE⁄g;7.21±0.85 mg QE⁄g). Hot methanol and aqueous extracts of both medicines showed low antioxidant activity in DPPH, ABTS, and FRAP methods and Iron chelating activity not found at highest possible concentration. V.C contains higher concentrations of total phenolic and flavonoid contents than A.C and can also exert greater antioxidant activity than A.C, although the activities demonstrated were lower than the positive control Trolox. The in-vitro antioxidant activity was not related with the total phenolic and flavonoid contents of the methanol and aqueous extracts of both herbal medicines (A.C and V.C).

Keywords: activity, different extracts, herbal medicines, in-vitro antioxidant

Procedia PDF Downloads 379

Authors: Hawraa Zbeeb, Hala Khalifeh, Mohamad Khalil, Francesca Storace, Francesca Baldini, Giulio Lupidi, Laura Vergani

Abstract:

Sarcopoterium spinosum, a widely distributed spiny shrub belonging to the Rosaceae family, is rich in essential and beneficial constituents. In fact, S. spinosum fruits and roots are traditionally used as herbal medicine in the eastern Mediterranean landscape, and this shrub is mentioned as a medicinal plant in a large number of ethnobotanical surveys. Aqueous root extracts from S. spinosum are used by traditional medicinal practitioners for weight loss treatment of diabetes and pain. Moreover, the anti-diabetic activity of S. spinosum root extract has been reported in different studies, but the beneficial effects of aerial parts, especially fruits, have not been elucidated yet. The aim of the present study was to investigate the in vitro antioxidant and lipid-lowering properties of an ethanolic extract from S. spinosum fruits using both hepatic (FaO) and endothelial (HECV) cells in an attempt to evaluate its possible employment as a nutraceutical supplement. First of all, in vitro spectrophotometric assays were employed to characterize the extract. The total phenol content (TPC) was evaluated by Folin–Ciocalteu spectrophotometric method and the radical scavenging activity was tested by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2, 2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assays. After that, the beneficial effects of the extract were tested on cells. FaO cells treated for 3 hours with 0.75 mM oleate/palmitate mix (1:2 molar ratio) mimic in vitro a moderate hepato-steatosis. HECV cells exposed for 1 hour to 100 µM H₂O₂ mimic an oxidative insult leading to oxidative stress conditions. After the metabolic and oxidative insult, both cell lines were treated with increasing concentrations of the S. spinosum extract (1, 10, 25 µg/mL) for 24 hours. The results showed the S. spinosum ethanolic extract is rather rich in phenols (TPC of 18.6 mgGAE/g dry extracts). Moreover, the extract showed a good scavenging ability in vitro (IC₅₀ 15.9 µg/ml and 10.9 µg/ml measured by DPPH and ABTS assays, respectively). When the extract was tested on cells, the results showed that it could ameliorate some markers of cell dysfunction. The three concentrations of the extract led to a significant decrease in the intracellular triglyceride (TG) content in steatotic FaO cells measured by spectrophotometric assay. On the other hand, HECV cells treated with increasing concentrations of the extract did not result in a significant decrease in both lipid peroxidation measured by the Thiobarbituric Acid Reactive Substances (TBARS) assay, and in reactive oxygen species (ROS) production measured by fluorometric analysis after DCF staining. Interestingly, the ethanolic extract was able to accelerate the wound repair of confluent HECV cells with respect to H₂O₂-insulted cells as measured by T-scratch assay. Taken together, these results seem to indicate that the ethanol extract from S. spinosum fruits is rich in phenol compounds and plays considerable lipid-lowering activity in vitro on steatotic hepatocytes and accelerates wound healing repair on endothelial cells. In light of that, the ethanolic extract from S. spinosum fruits could be a potential candidate for nutraceutical applications.

Keywords: antioxidant activity, ethanolic extract, lipid-lowering activity, phenolic compounds, Sarcopoterium spinosum fruits

Procedia PDF Downloads 148

Authors: Rasha M. Hussein, Reem M. Hashem, Laila A. Rashed

Abstract:

Alzheimer’s disease is the most common dementia disease in the elderly. It is characterized by the accumulation of extracellular amyloid β (Aβ) peptides and intracellular hyper-phosphorylated tau protein. In addition, recent evidence indicates that accumulation of intracellular amyloid β peptides may play a role in Alzheimer’s disease pathogenesis. This suggests that intracellular Heat Shock Proteins (HSP) that maintain the protein quality control in the cell might be potential candidates for disease amelioration. DNAJB6, a member of DNAJ family of HSP, effectively prevented the aggregation of poly glutamines stretches associated with Huntington’s disease both in vitro and in cells. In addition, DNAJB6 was found recently to delay the aggregation of Aβ42 peptides in vitro. In the present study, we investigated the ability of DNAJB6 to prevent the aggregation of both intracellular and extracellular Aβ peptides using transfection of HEK293 cells with Aβ-GFP and recombinant Aβ42 peptides respectively. We performed western blotting and immunofluorescence techniques. We found that DNAJB6 can prevent Aβ-GFP aggregation, but not the seeded aggregation initiated by extracellular Aβ peptides. Moreover, DNAJB6 required interaction with HSP70 to prevent the aggregation of Aβ-GFP protein and its J-domain was essential for this anti-aggregation activity. Interestingly, overexpression of other DNAJ proteins as well as HSPB1 suppressed Aβ-GFP aggregation efficiently. Our findings suggest that DNAJB6 is a promising candidate for the inhibition of Aβ-GFP mediated aggregation through a canonical HSP70 dependent mechanism.

Keywords: Aβ, Alzheimer’s disease, chaperone, DNAJB6, aggregation

Procedia PDF Downloads 488