Search results for: microplate antimicrobial assay

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

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

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

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Authors: Jade Edey, Andrew Bennett, Gareth Hathway

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Nuclear receptor-related 1 protein (also known as Nurr1 or NR4A2) is an orphan nuclear receptor which plays a vital role in the development, survival and maintenance of dopaminergic (DA) neurons particularly in the substantia nigra (SN). Increasing research has investigated Nurr1’s additional role within microglia and astrocytes where it has been suggested to act as a negative regulator of inflammation; potentially offering neuroprotection. Considering both DA neurodegeneration and neuroinflammation are commonly accepted constituents of Parkinson’s Disease (PD), understanding the mechanisms by which Nurr1 regulates inflammatory processes could provide an attractive therapeutic target. Nurr1 regulates inflammation via a transrepressive mechanism possibly dependent upon SUMOylation. In addition, Nurr1 can transactivate numerous genes involved in DA synthesis, such as Tyrosine Hydroxylase (TH). A C-terminal splice variant of Nurr1, Nurr-1a, has been reported in both neuronal and glial cells. However, research into its transcriptional activity is minimal. We employed in vitro methods such as SUMO-Pulldown experiments alongside Luciferase reporter assays to investigate the SUMOylation status and transactivation capabilities of Nurr1 and Nurr-1a respectively. The SUMO-Pulldown assay demonstrated Nurr-1a undergoes significantly more SUMO modification than its full-length variant. Consequently, despite having less transcriptional activation than Nurr1, Nurr1a may play a more prominent role in repression of microglial inflammation. Contrary to published literature we also identified that SUMOylation enhances transcriptional activation by Nurr1 and Nurr1a. SUMOylation-dependent increases in Nurr1 and Nurr1a transcriptional activation were only evident in neuronal SHSY5Y cells but not in HEK293 cells. This research provides novel insight into the regulation of Nurr-1a and indicates differential effects of SUMOylation dependent regulation in neuronal and inflammatory cells.

Keywords: nuclear receptors, Parkinson’s disease, inflammation, transcriptional regulation

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Authors: Aysegul Alyamac, Sukru Gulec

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Ankaferd Blood Stopper (ABS) is a plant extract used to stop bleeding caused by injuries and surgical interventions. ABS also involved in wound healing of intestinal mucosal damage due to oxidative stress and inflammation. Inflammatory Bowel Disease (IBD) is a common chronic disorder of the gastrointestinal tract that causes abdominal pain, diarrhea, and gastrointestinal bleeding, and increases the risk of colon cancer. Inflammation is an essential factor in the development of IBD. The various studies have been performed about the physiological effects of ABS; however, ABS dependent mechanism on colonic inflammation has not been elucidated. Thus, the protective effect of ABS on colonic inflammation was investigated in this study. The Caco-2 and RAW 264.7 murine macrophage cells were used as a model of in vitro colonic inflammation. RAW 264.7 cells were treated with lipopolysaccharide (LPS) for 12 hours to induce the inflammation, and a conditional medium was obtained. Caco-2 cells were treated with 15 µl/ml ABS for 4 hours, then incubated with conditional medium and the cells also were incubated with 15 µl/ml ABS and conditional medium together for 4 hours. Tumor necrosis factor alpha (TNF-α) protein levels were targeted in testing inflammatory condition and its level was significantly increased (25 fold, p<0.001) compared to the control group by using Enzyme-Linked Immunosorbent Assay (ELISA) method. The COX-2 mRNA level was used as a marker gene to show the possible anti-inflammatory effect of ABS in Caco-2 cells. RAW cells-derived conditional medium significantly (3.3 fold, p<0.001) induced cyclooxygenase-2 (COX-2) mRNA levels in Caco-2 cells. The pretreatment of Caco-2 cells caused a significant decrease (3.3 fold, p<0.001) in COX-2 mRNA levels relative to conditional medium given group. Furthermore, COX-2 mRNA level was significantly reduced (4,7 fold, p<0.001) in ABS and conditional medium treated group. These results suggest that ABS might have an anti-inflammatory effect in vitro.

Keywords: Ankaferd blood stopper, CaCo-2, colonic inflammation, RAW 264.7

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Authors: Alejandra Betancur Sánchez, Carmen Elena Zapata Sánchez, Juan Bautista López Ortiz

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Adverse effects and increased air pollution has raised concerns about regulatory policies and has fostered the development of new air quality standards; this is due to the complexity of the composition and the poorly understood reactions in the atmospheric environment. Toxic compounds act as environmental agents having various effects, from irritation to death of cells and tissues. A toxic agent is defined an adverse response in a biological system. There is a particular class that produces some kind of alteration in the genetic material or associated components, so they are recognized as genotoxic agents. Within cells, they interact directly or indirectly with DNA, causing mutations or interfere with some enzymatic repair processes or in the genesis or polymerization of proteinaceous material involved in chromosome segregation. An air pollutant may cause or contribute to increased mortality or serious illness and even pose a potential danger to human health. The aim of this study was to evaluate the effect on the viability and the genotoxic potential on the cell lines CHO-K1 and Jurkat and peripheral blood of particulate matter PM T lymphocytes 2.5 obtained from filters collected three monitoring stations network air quality Aburrá Valley. Tests, reduction of MTT, trypan blue, NRU, comet assay, sister chromatid exchange (SCE) and chromosomal aberrations allowed evidence reduction in cell viability in cell lines CHO-K1 and Jurkat and damage to the DNA from cell line CHOK1, however, no significant effects were observed in the number of SCEs and chromosomal aberrations. The results suggest that PM2.5 material has genotoxic potential and can induce cancer development, as has been suggested in other studies.

Keywords: PM2.5, cell line Jurkat, cell line CHO-K1, cytotoxicity, genotoxicity

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Authors: Mahboubeh Kabiri, Saba Aslani

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Among various approaches used for the treatment of cardiovascular diseases, the occlusion of the small-diameter vascular graft (SDVG) is still an unresolved problem which seeks further research to address them. Though autografts are now the gold standards to be replaced for blocked coronary arteries, they suffer from inadequate quality and quantity. On the other hand, the major problems of the tissue engineered grafts are thrombosis and intimal hyperplasia. Provision of a suitable spatiotemporal release pattern of anticoagulant agents such as heparin and aspirin can be a step forward to overcome such issues . Herein, we fabricated electrospun scaffolds from FDA (Food and Drug Administration) approved poly-L-lactic acid (PLLA) with aspirin loaded into the nanofibers. Also, we surface coated the scaffolds with Amniotic Membrane lysate as a source for natural elastic polymers and a mimic of endothelial basement membrane. The scaffolds were characterized thoroughly structurally and mechanically for their morphology, fiber orientation, tensile strength, hydrophilicity, cytotoxicity, aspirin release and cell attachment support. According to the scanning electron microscopy (SEM) images, the size of fibers ranged from 250 to 500 nm. The scaffolds showed appropriate tensile strength expected for vascular grafts. Cellular attachment, growth, and infiltration were proved using SEM and MTT (3-(4,5-Dimethylthiazol-2-Yl)-2,5-Diphenyltetrazolium Bromide) assay. Drug-loaded scaffolds showed a sustained release profile of aspirin in 7 days. An enhanced cytocompatibility was observed in AM-coated electrospun PLLA fibers compared to uncoated scaffolds. Our results together indicated that AM lysate coated ASA releasing scaffolds have promising potentials for development of a biocompatible SDVG.

Keywords: vascular tissue engineering, vascular grafts, anticoagulant agent, aspirin, amniotic membrane

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Authors: Ibrahim M. Labouta, Gina N. Tageldin, Salwa M. Fahmy, Hayam M. Ashour, Mounir A. Khalil, Tamer M. Ibrahim, Nefertiti A. El-Nikhely

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Bruton’s tyrosine kinase (BTK) is a critical effector molecule in B cell antigen receptor (BCR) signaling transduction. It regulates B cell proliferation, development and survival. Since BTK is widely expressed in many B cell leukaemias and lymphomas, targeting BTK by small molecules inhibitors became an attractive idea as new treatment modalities for B cell mediated hematologic malignancies. Ibrutinib is the 1st generation BTK inhibitor, approved by FDA for treatment of relapsed mantle cell lymphoma (MCL) and chronic lymphocytic leukemia (CLL). It binds irreversibly to the unique cysteine (Cys481) within the ATP-binding pocket of BTK. Besides ibrutinib, many irreversible covalent BTK inhibitors comprising pyrimidine nucleus such as spebrutinib (phase IIb) showed high selectivity and potency when compared to it. In this study, the designed compounds were based on 5-cyano-2-methylsulfanyl pyrimidine core and decorated with electrophilic warheads which are essential for the optimal activity for targeted covalent inhibition (TCI). However, modifications at pyrimidine C4 or C6 were made by introduction of substituted amines which are provided to behave differently. The synthesized derivatives were evaluated for their anticancer activity in leukemia cell lines (e.g. THP-1). Results showed that, some derivatives exhibited antiproliferative activity with IC50 ranged from 5-50 μM, The in vitro enzymatic inhibitory assay for these compounds against BTK is still under investigation. Nevertheless, we could conclude from the initial biological screening that, the synthesized 4 or 6-subsitituted aminopyrimidines represent promising and novel antileukemic agents. Meanwhile, further studies are still needed to attribute this activity through targeting BTK enzyme and inhibition of BCR signaling pathway.

Keywords: BTK inhibitors, hematologic malignancies, structure based drug design (SBDD), targeted covalent inhibitors (TCI)

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Authors: Anna Curtin, Matthew Thibodeau, Heather Buckley

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Water scarcity is characterized by a lack of access to clean and affordable drinking water, as well as water for hygienic and economic needs. The amount of people effected by water scarcity is expected to increase in the coming years due to climate change, population growth, and pollution, amongst other things. In response, scientists are pursuing cost effective drinking water treatment methods, often with a focus on alternative water sources. Desalination of seawater via reverse osmosis is one promising alternative method. Desalination of seawater via reverse osmosis, however, is limited significantly by biofouling of the filtration membrane. Biofouling is the buildup of microorganisms in a biofilm at the water-membrane interface. It clogs the membrane, decreasing the efficiency of filtration, consequently increasing operational and maintenance costs. Although effective, existing chemical treatment methods can damage the membrane, decreasing the lifespan of the membrane; create antibiotic resistance; and cause harm to humans and the environment if they pass through the membrane into the permeate. The current project focuses on applying safer preservatives used in home and personal care products to RO membranes to investigate the biofouling treatment efficacy. Currently, many of these safer preservatives have only been tested on cells in planktonic phase in suspension cultures, not on cells in biofilms. The results of suspension culture tests are not applicable to biofouling scenarios because organisms in planktonic phase in suspension cultures exhibit different morphological, chemical, and metabolic characteristics than those in a biofilm. Testing antifoulant efficacy of safer preservatives on biofilms will provide more applicable results to biofouling on RO membranes. To do this, biofilms will be grown on 96-well-plates and minimum inhibitory concentrations (MIC90) and log-reductions will be calculated for various safer preservatives. Results from these tests will be used to guide doses for tests of safer preservatives in a bench-scale RO system.

Keywords: reverse osmosis, biofouling, preservatives, antimicrobial, safer alternative, green chemistry

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Authors: Sneha Kumari, Ravi Krishnan Elangovan

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This abstract describes the development of a high-speed tracking method by modification in motor components for nanoparticle attachment. Myosin motors are nano-sized protein machines powering movement that defines life. These miniature molecular devices serve as engines utilizing chemical energy stored in ATP to produce useful mechanical energy in the form of a few nanometre displacement events leading to force generation that is required for cargo transport, cell division, cell locomotion, translated to macroscopic movements like running etc. With the advent of in vitro motility assay (IVMA), detailed functional studies of the actomyosin system could be performed. The major challenge with the currently available IVMA for tracking actin filaments is a resolution limitation of ± 50nm. To overcome this, we are trying to develop Single Molecule IVMA in which nanoparticle (GNP/QD) will be attached along or on the barbed end of actin filaments using CapZ protein and visualization by a compact TIRF module called ‘cTIRF’. The waveguide-based illumination by cTIRF offers a unique separation of excitation and collection optics, enabling imaging by scattering without emission filters. So, this technology is well equipped to perform tracking with high precision in temporal resolution of 2ms with significantly improved SNR by 100-fold as compared to conventional TIRF. Also, the nanoparticles (QD/GNP) attached to actin filament act as a point source of light coffering ease in filament tracking compared to conventional manual tracking. Moreover, the attachment of cargo (QD/GNP) to the thin filament paves the way for various nano-technological applications through their transportation to different predetermined locations on the chip

Keywords: actin, cargo, IVMA, myosin motors and single-molecule system

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Authors: Jeena Gupta

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In the today’s world of pharmaceutical products, one should not forget the healing properties of inexpensive food materials especially spices. They are known to possess hidden pharmaceutical ingredients, imparting them the qualities of being anti-microbial, anti-oxidant, anti-inflammatory and anti-carcinogenic. Further aberrant epigenetic regulatory mechanisms like DNA methylation, histone modifications or altered microRNA expression patterns, which regulates gene expression without changing DNA sequence, contribute significantly in the development of various diseases. Changing lifestyles and diets exert their effect by influencing these epigenetic mechanisms which are thus the target of dietary phytochemicals. Bioactive components of plants have been in use since ages but their potential to reverse epigenetic alterations and prevention against diseases is yet to be explored. Spices being rich repositories of many bioactive constituents are responsible for providing them unique aroma and taste. Some spices like curcuma and garlic have been well evaluated for their epigenetic regulatory potential, but for others, it is largely unknown. We have evaluated the biological activity of phyto-active components of Fennel, Cardamom and Fenugreek by in silico molecular modeling, in vitro and in vivo studies. Ligand-based similarity studies were conducted to identify structurally similar compounds to understand their biological phenomenon. The database searching has been done by using Fenchone from fennel, Sabinene from cardamom and protodioscin from fenugreek as a query molecule in the different small molecule databases. Moreover, the results of the database searching exhibited that these compounds are having potential binding with the different targets found in the Protein Data Bank. Further in addition to being epigenetic modifiers, in vitro study had demonstrated the antimicrobial, antifungal, antioxidant and cytotoxicity protective effects of Fenchone, Sabinene and Protodioscin. To best of our knowledge, such type of studies facilitate the target fishing as well as making the roadmap in drug design and discovery process for identification of novel therapeutics.

Keywords: epigenetics, spices, phytochemicals, fenchone

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Authors: Esther N. Maina, Anna Skowronska, Sridhar Chaganti, Malcolm A. Taylor, Paul G. Murray, Tatjana Stankovic

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Epstein-Barr virus (EBV) has been implicated in the pathogenesis of human tumours of B-cell origin. The demonstration that a proportion of Hodgkin lymphomas and all Burkitt’s lymphomas harbour EBV suggests that the virus contributes to the development of these malignancies. However, the mechanisms of lymphomagenesis remain largely unknown. To determine whether EBV causes DNA damage and alters DNA damage response in cells of EBV-driven lymphoma origin, Germinal Centre (GC) B cells were infected with EBV and DNA damage responses to gamma ionising radiation (IR) assessed at early time points (12hr – 72hr) after infection and prior to establishment of lymphoblastoid (LCL) cell lines. In the presence of EBV, we observed induction of spontaneous DNA DSBs and downregulation of ATM-dependent phosphorylation in response to IR. This downregulation coincided with reduced ability of infected cells to repair IR-induced DNA double-strand breaks, as measured by the kinetics of gamma H2AX, a marker of double-strand breaks, and by the tail moment of the comet assay. Furthermore, we found that alteration of DNA damage responses coincided with the expression of LMP-1 protein. The presence of the EBV virus did not affect the localization of the ATM-dependent DNA repair proteins to sites of damage but instead lead to an increased expression of PP5, a phosphatase that regulates ATM function. The impact of the virus on DNA repair was most prominent 24h after infection, suggesting that this time point is crucial for the viral establishment in B cells. Our results suggest that during an early infection EBV virus dampens crucial cellular responses to DNA double-strand breaks which facilitate successful viral infection, but at the same time might provide the mechanism for tumor development.

Keywords: EBV, ATM, DNA damage, germinal center cells

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Authors: Dmitry Nosik, Nickolay Nosik, Elli Kaplina, Olga Lobach, Marina Chataeva, Lev Rasnetsov

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Background and aims: Human Immunodeficiency Virus (HIV) infection is very often associated with Herpes Simplex Virus (HSV) infection but HIV patients are treated with a cocktail of antiretroviral drugs which are toxic. The use of an antiviral drug which will be active against both viruses like ferrovir found in our previous studies is rather actual. Earlier we had shown that Fullerene poly-amino capronic acid (FPACA) was active in case of monoinfection of HIV-1 or HSV-1. The aim of the study was to analyze the efficiency of FPACA against mixed infection of HIV and HSV. Methods: The peripheral blood lymphocytes, CEM, MT-4 cells were simultaneously infected with HIV-1 and HSV-1. FPACA was added 1 hour before infection. Cells viability was detected by MTT assay, virus antigens detected by ELISA, syncytium formation detected by microscopy. The different multiplicity of HIV-1/HSV-1 ratio was used. Results: The double viral HIV-1/HSV-1 infection was more cytopathic comparing with monoinfections. In mixed infection by the HIV-1/HSV-1 concentration of HIV-1 antigens and syncytium formations increased by 1,7 to 2,3 times in different cells in comparison with the culture infected with HIV-1 alone. The concentration of HSV-1 increased by 1,5-1,7 times, respectively. Administration of FPACA (1 microg/ml) protected cells: HIV-1/HSV-1 (1:1) – 80,1%; HIV-1/HSV-1 (1:4) – 57,2%; HIV-1/HSV-1 (1:8) – 46,3 %; HIV-1/HSV-1 (1:16) – 17,0%. Virus’s antigen levels were also reduced. Syncytium formation was totally inhibited in all cases of mixed infection. Conclusion: FPACA showed antiviral activity in case of mixed viral infection induced by Human Immunodeficiency Virus and Herpes Simplex Virus. The effect of viral inhibition increased with the multiplicity of HIV-1 in the inoculum. The mechanism of FPACA action is connected with the blocking of the virus particles adsorption to the cells and it could be suggested that it can have an antiviral activity against some other viruses too. Now FPACA could be considered as a potential drug for treatment of HIV disease complicated with opportunistic herpes viral infection.

Keywords: antiviral drug, human immunodeficiency virus (hiv), herpes simplex virus (hsv), mixed viral infection

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Authors: David Guillermo Piedrahita Marquez, Hector Suarez Mahecha, Jairo Humberto Lopez

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The propolis are substances obtained from the beehive have an action against pathogens, prooxidant substances and free radicals because of its polyphenols content, this has motivated the use of these compounds in the food and pharmaceutical industries. However, due to their organoleptic properties and their ability to react with other compounds, their application has been limited; therefore, the objective of this research was to propose a mechanism to protect propolis and mitigate side effects granted by its components. To achieve the stated purpose ethanolic extracts of propolis (EEP) from three samples from Santander were obtained and their antioxidant and antimicrobial activity were evaluated in order to choose the extract with the biggest potential. Subsequently mixtures of the extract with maltodextrin were prepared by spray drying varying concentration and temperature, finally the yield, the physicochemical, and antioxidant properties of the products were measured. It was concluded that Socorro propolis was the best for the production of microencapsulated due to their activity against pathogenic strains, for its large percentage of DPPH radical inactivation and for its high phenolic content. In spray drying, the concentration of bioactive had a greater impact than temperature and the conditions set allowed a good performance and the production of particles with high antioxidant potential and little chance of proliferation of microorganisms. Also, it was concluded that the best conditions that allowed us to obtain the best particles were obtained after drying a mixture 1:2 ( EEP: Maltodextrin), besides the concentration is the most important variable in the spray drying process, at the end we obtained particles of different sizes and shape and the uniformity of the surface depend on the temperature. After watching the previously mentioned microparticles by scanning electron microscopy (SEM) it was concluded that most of the particles produced during the spray dry process had a spherical shape and presented agglomerations due to the moisture content of the ethanolic extracts of propolis (EEP), the morphology of the microparticles contributed to the stability of the final product and reduce the loss of total phenolic content.

Keywords: spray drying, propolis, maltodextrin, encapsulation, scanning electron microscopy

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Authors: N. Zarghami, M. Mohammadinejad, A. Akbarzadeh, Y. Pilehvar-Soltanahmadi, F. Zarghami

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Breast cancer is known to be the most common cancer in women. Cyclin D1 is a proto-oncogene and over expression of cyclin D1 is directly associated with tumorgenesis. Cyclin D1 is overexpressed in more than 50% of breast cancer cases. Curcumin is derived from turmeric (curcuma longa) and chrysin is a component that could be extracted from many plants and honey. These two plants derived compounds are believed to assist in inhibition of the cancer cells growth and reducing cyclin D1 expression. In this work, the hypothesis is to combine curcumin and chrysin in order to analyze the potential synergistic effect in inhibition of cell proliferation and down regulation of cyclin D1. In addition, use of PLGA-PEG to improve bioavailability of pure curcumin and chrysin, while reinforcing the potential effect of this combination. PLGA-PEG nanoparticles were synthesized and characterized with FT-IR and 1HNMR methods. Although morphological features were analyzed by SEM. Afterward curcumin and chrysin were encapsulated with synthesized PLGA-PEG and MTT-assay was performed to measure cytotoxicity effect of these plant constitutes. T-47D cells were treated with proper concentration of these constituents and Real-time PCR was carried out to evaluate cyclin D1 expression levels. Curcumin, chrysin and combination of curcumin –chrysin in intact and nano-capsulated form affected T-47D cells in time and dose dependent manner and the combination of these compounds had synergistic effects. Real-time PCR results, revealed that curcumin, chrysin and combination of curcumin-chrysin in pure and encapsulated form inhibited cyclin D1 expression. Compared to pure components, different concentrations of nano-curcumin, nano chrysin and nano-combination caused further decline in cyclin D12 expression by 5-11%, 8-22% and 6-18% respectively. Our results demonstrated that, combination of chrysin-curcumin had synergistic effect and nano capsulated form of this component had grater inhibition on cyclin D1 expression.

Keywords: breast cancer, cyclin D1, curcumin, chrysin, nanoparticles

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Authors: Pedro Cabrales, Scott Caroen, Tony R. Reid, Bryan Oronsky

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Introduction and Purpose RRx-001 is an NLRP3 inhibitor and Nrf2 agonist in Phase 3 trials for the treatment of cancer. The purpose of this study was to examine whether treatment with RRx-001, given itsanti-inflammatory and antioxidant properties, improvedexercise and skeletal muscle oxidative capacity in mice on the generalpremiss that better health outcomes correlatewith more activity. Material and Methods Male and female adult mice (n=6 per group) were subjected to an endurance exercise capacity (EEC)test until exhaustion on a motorized treadmill after 3 once weekly doses of either RRx-001 5 mg/kg, RRx-001 2 mg/kg, or vehicle. The EEC protocol consisted of a treadmill velocity of 30meters per min at an uphill inclination (slope of 10%) until the mice reached fatigue, which was defined as the inability of the mice to maintain the appropriate pace despitecontinuous hand stimulation for 1 min. The concentration of malondialdehyde (MDA), an indicator of lipid peroxidation, and creatine kinase (CK), an indicator of muscle damage, in the blood samples collected immediately after the acute exercise was determined with a commercial ELISA assay kit. ResultsThe exhaustive exercise times of the RRx-001 groups were significantly longer than that of the vehicle group (p<0.05) by weeks 2 and 3. In addition, MDA levels in the gastrocnemius, soleus, and extensor digitorum longus muscles were significantly lower than those of the vehicle group were (p<0.05), as were the serum CK levels(p<0.05). ConclusionsIn conclusion, this study found that RRx-001 has anti-fatigue properties, as evidenced by an increase in exercise capacity with RRx-001 treatment, and protects against strenuous exercise-induced muscle damage and lipid peroxidation. This data potentially supports the use of RRx-001 in the clinic to improve exercise performance and reduce physical fatigue.

Keywords: RRx-001, anti-fatigue, muscle protection, increased exercise tolerance, lipid peroxidation

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Authors: Ziyad S. Haidar

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Background: Saliva plays a major role in maintaining oral and dental health (consequently, general health and well-being). Where it normally bathes the oral cavity and acts as a clearing agent. This becomes more apparent when the amount and quality of salivare significantly reduced due to medications, salivary gland neoplasms, disorders such as Sjögren’s syndrome, and especially ionizing radiation therapy for tumors of the head and neck, the fifth most common malignancy worldwide, during which the salivary glands are included within the radiation field or zone. Clinically, patients affected by salivary gland dysfunction often opt to terminate their radiotherapy course prematurely because they become malnourished and experience a significant decrease in their quality of life. Accordingly, the development of an alternative treatment to restore or regenerate damaged salivary gland tissue is eagerly awaited. Likewise, the formulation of a radioprotection modality and early damage prevention strategy is also highly desirable. Objectives: To assess the pre-clinical radio-protective effect as well as the reparative/regenerative potential of layer-by-layer self-assembled lipid-polymer-based core-shell nanocapsules designed and fine-tuned in this experimental work for the sequential (ordered) release of dual cytokines, following a single local administration (direct injection) into a murine sub-mandibular salivary gland model of irradiation. Methods: The formulated core-shell nanocapsules were characterized by physical-chemical-mechanically pre-/post-loading with the drugs (in solution and powder formats), followed by optimizing the pharmaco-kinetic profile. Then, nanosuspensions were administered directly into the salivary glands, 24hrs pre-irradiation (PBS, un-loaded nanocapsules, and individual and combined vehicle-free cytokines were injected into the control glands for an in-depth comparative analysis). External irradiation at an elevated dose of 18Gy (revised from our previous 15Gy model) was exposed to the head-and-neck region of C57BL/6 mice. Salivary flow rate (un-stimulated) and salivary protein content/excretion were regularly assessed using an enzyme-linked immunosorbent assay (3-month period). Histological and histomorphometric evaluation and apoptosis/proliferation analysis followed by local versus systemic bio-distribution and immuno-histochemical assays were then performed on all harvested major organs (at the distinct experimental end-points). Results: Monodisperse, stable, and cytocompatible nanocapsules capable of maintaining the bioactivity of the encapsulant within the different compartments with the core and shell and with controlled/customizable pharmaco-kinetics, resulted, as is illustrated in the graphical abstract (Figure) below. The experimental animals demonstrated a significant increase in salivary flow rates when compared to the controls. Herein, salivary protein content was comparable to the pre-irradiation (baseline) level. Histomorphometry further confirmed the biocompatibility and localization of the nanocapsules, in vivo, into the site of injection. Acinar cells showed fewer vacuoles and nuclear aberration in the experimental group, while the amount of mucin was higher in controls. Overall, fewer apoptotic activities were detected by a Terminal deoxynucleotidyl Transferase (TdT) dUTP Nick-End Labeling (TUNEL) assay and proliferative rates were similar to the controls, suggesting an interesting reparative and regenerative potential of irradiation-damaged/-dysfunctional salivary glands. The Figure below exemplifies some of these findings. Conclusions: Biocompatible, reproducible, and customizable self-assembling layer-by-layer core-shell delivery system is formulated and presented. Our findings suggest that localized sequential bioactive delivery of dual cytokines (in specific dose and order) can prevent irradiation-induced damage via reducing apoptosis and also has the potential to promote in situ proliferation of salivary gland cells; maxSALIVA is scalable (Good Manufacturing Practice or GMP production for human clinical trials) and patent-pending.

Keywords: saliva, head and neck cancer, nanotechnology, controlled drug delivery, xerostomia, mucositis, biopolymers, innovation

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Authors: Dada. O.Akinola, Uche. I. Ebele, Bamiro .A.Rafatu, Akinbami A. Akinsegun, Dada O. Adeyemi, Adeyemi. O. Ibukun, Okunowo O.Bolanle, Abdulateef O. Kareem, Ibrahim.N. Ismaila, Dosu Rihanat

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Background: Hyperglycaemia alters qualitatively and quantitatively all the full blood count parameters. The alterations among other factors are responsible for the macrovascular and microvascular complications associated with diabetes mellitus (DM). This study is aimed at correlating haematological parameters in DM subjects with their fasting blood glucose (FBG) and anthropometric parameters. Materials and Methods: This was a cross-sectional study of participants attending DM clinic of Lagos State University Teaching Hospital (LASUTH), Ikeja. The study recruited one hundred and two (102) DM subjects and one hundred (100) non-DM controls. Venous blood samples were collected for full blood count (FBC) assay while FBG was done, structured questionnaires were administered, and anthropometric measurements of all participants were done. Data were analyzed with Statistical Package for Social Science (SPSS) version 23. P was set at ≤0.05. Results: The mean age of DM patients was 64.32± 11.31 years. Using a haemoglobin concentration cut-off of 11g/dl, 39.2%, and 13% DM and control participants respectively had values lower than 11g/dl. A total of 22.5% and 3% of DM and controls respectively gave a history of previous blood transfusion.White blood cells count and platelet count means were (6.12±1.60 and 5.30±7.52,p=0.59) and (213.31±73.58 and 228.91±73.21,p = 0.26) *109/L in DM subjects and controls respectively. FBG and all the anthropometric data in DM subjects were significantly higher than in controls. Conclusions: The prevalence of anaemia in DM subjects was three times higher than in controls. The white blood cell count was higher but not statistically significant in DM compared with controls. But platelet count was higher but not statistically significant in controls compared with DM subjects.

Keywords: haematological profile, diabetes mellitus, anthropometric data, fasting blood glucose

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Authors: Rania Hanafi Mahmoud Said, Rasha Mohamed Taha

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Background: A recent study has explored the potential of an approach to treating infected root canals using a combination of Mineral Trioxide Aggregate (MTA) and Tetracycline-loaded Nanochitosan. Material and methods: Forty dogs were included in the study, with infected periapical areas induced by leaving access openings in their teeth for four months. Bacteriological samples from the infected root canals were collected and managed anaerobically to identify and count the different microorganisms present. The most common microorganisms detected were Prevotella oris, Fusobacterium nucleatum, Streptococcus viridans, Enterococcus faecalis, Clostridium subterminale, Porphyromonas gingivalis, and Peptostreptococcus anaerobius. The dogs were divided into four groups based on the sealant used to treat the infected periapical areas: Group I: Negative control (no treatment) Group II: Positive control (MTA only) Group III: MTA + tetracycline Group IV: MTA + tetracycline loaded on nanochitosan Results: Periapical areas in Group IV showed significantly more bone healing than those in Groups I, II, and III. The newly formed bone was evaluated radiographically, histologically, and immunohistochemically using Osteopontin (OSP) antibodies. Data collected was statistically analysed using SPSS software at a 0.05 significance level. Conclusion: The study concluded that the combined use of Tetracycline-loaded Nanochitosan and MTA presents a promising approach for the treatment of infected root canals. The potent antimicrobial activity of Tetracycline-loaded Nanochitosan, along with the biocompatibility and desirable properties of MTA, may synergistically contribute to improved clinical outcomes in endodontic therapy. This study has important implications for the clinical management of infected root canals. The combination of Tetracycline-loaded Nanochitosan and MTA could provide a more effective and efficient means of treating these challenging cases. Further research is needed to confirm these findings in humans and to optimize the treatment protocol.

Keywords: mineral trioxide aggregate, tetracycline-loaded nanochitosan, periapical infection, osteopontine

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Authors: Vikash Yadav, Ashish Arora

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Microorganisms have self-defense mechanisms to protect themselves from toxic environments. Phenolic acid decarboxylase(pad) is responsible for the defense against toxicity caused by phenolic acids, converting them into less toxic vinyl derivatives. The transcription of the pad gene is regulated by a negative transcription factor, phenolic acid decarboxylase regulators (PadR), in a substrate-inducible manner. The PadR family members share the conserved DNA-binding features and interact with the operator DNA using a winged helix-turn-helix (wHTH) motif, which contains a three-helix motif and a β-stranded wing. The members of this family function as transcriptional regulators that are involved in various cellular survival processes, such as toxin production, detoxification, multidrug resistance, antibiotic biosynthesis, and carbon catabolism. Rv1176 of Mycobacterium tuberculosis H37Rv has been assigned to the PadR family protein that remains to be structurally and functionally uncharacterized. To reveal the structural mechanism by which Rv1176 could regulates effector-responsive transcription, several experiments were performed, including Electrophoretic Mobility Shift Assay (EMSA) for DNA protein interaction, differential scanning calorimetry (DSC) and Differential Scanning Fluorimetry (DSF) for temperature and ligand-dependent protein stability, Circular Dichroism (CD) spectroscopy for secondary structure analysis. Further, to evaluate the functional role of Rv1176, the intracellular survival of recombinant M. smegmatis was examined in murine macrophage cell line J774A.1 and different stressed conditions like oxidative, pH, and nutritive stress. All these studies demonstrated that Rv1176 could behave as a transcription regulator and its expression in recombinant M. smegmatis increases intracellular survival.

Keywords: EMSA, Mycobacterium tuberculosis, PadR family protein, transcriptional regulator

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Authors: Azar Heidarizadi, Mahdieh Salimi, Hossein Mozdarani

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Introduction: As a complex disease, breast cancer results from several genetic and epigenetic changes. Lactoferrin, a member of the transferrin family, is reported to have a number of biological functions, including DNA synthesis, immune responses, iron transport, etc., any of which could play a role in tumor progression. The aim of this study was to investigate the bioinformatics data and experimental assay to find the pattern of promoter methylation and gene expression of LTF in breast cancer in order to study its potential role in cancer management. Material and Methods: In order to evaluate the methylation status of the LTF promoter, we studied the MS-PCR and Real-Time PCR on samples from patients with breast cancer and normal cases. 67 patient samples were conducted for this study, including tumoral, plasma, and normal tissue adjacent samples, as well as 30 plasma from normal cases and 10 tissue breast reduction cases. Subsequently, bioinformatics analyses such as cBioPortal databases, string, and genomatix were conducted to disclose the prognostic value of LTF in breast cancer progression. Results: The analysis of LTF expression showed an inverse relationship between the expression level of LTF and the stages of tissues of breast cancer patients (p<0.01). In fact, stages 1 and 2 had a high expression in LTF, while, in stages 3 and 4, a significant reduction was observable (p < 0.0001). LTF expression frequently alters with a decrease in the expression in ER⁺, PR⁺, and HER2⁺ patients (P < 0.01) and an increase in the expression in the TNBC, LN¯, ER¯, and PR- patients (P < 0.001). Also, LTF expression is significantly associated with metastasis and lymph node involvement factors (P < 0.0001). The sensitivity and specificity of LTF were detected, respectively. A negative correlation was detected between the results of level expression and methylation of the LTF promoter. Conclusions: The altered expression of LTF observed in breast cancer patients could be considered as a promotion in cell proliferation and metastasis even in the early stages of cancer.

Keywords: LTF, expression, methylation, breast cancer

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Authors: Abul Hashem

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The experiment was conducted to find out the effect of different levels of Moringa oleifiera leaf extract and synthetic antioxidant (Beta Hydroxyl Anisole) on fresh and preserved beef meatballs. For this purpose, ground beef samples were divided into five treatment groups. They are treated as control, synthetic antioxidant, 0.1%, 0.2% and 0.3% Moringa oleifera leaf extract as T1, T2, T3, T4 and T5, respectively. Five kinds of meatballs were made and biscuit crushed and egg albumin was mixed with beef meatballs and cooking was practiced properly. Proximate analysis, sensory tests (color, flavor, tenderness, juiciness, overall acceptability), cooking loss, pH value, free fatty acids (FFA), thiobarbituric acid values (TBARS), peroxide value(POV) and microbiological examination were determined in order to evaluate the effect of Moringa oleifiera leaf extract as natural antioxidant & antimicrobial activities in comparing to BHA (Beta Hydroxyl Anisole) at first day before freezing and for maintaining meatballs qualities on the shelf life of beef meat balls stored for 60 days under frozen condition. Freezing temperature was -20˚C. Days of intervals of experiment were on 0, 15th, 30th, and 60th days. Dry matter content of all the treatment groups differ significantly (p<0.05). On the contrary, DM content increased significantly (p<0.05) with the advancement of different days of intervals. CP content of all the treatments were increased significantly (p<0.05) among the different treatment groups. EE content at different treatment levels differ significantly (p<0.05). Ash content at different treatment levels was also differ significantly (p<0.05). FFA values, TBARS, POV were decreased significantly (p<0.05) at different treatment levels. Color, odor, tenderness, juiciness, overall acceptability, raw PH, cooked pH were increased at different treatment levels significantly (p<0.05). The cooking loss (%) at different treatment levels were differ significantly (p<0.05). TVC (logCFU/g), TCC (logCFU/g) and TYMC (logCFU/g) was decreased significantly (p<0.05) at different treatment levels comparison to control. Considering CP, tenderness, juiciness, overall acceptability, cooking loss, FFA, POV, TBARS and microbial parameters it can be concluded that Moringa oleifera leaf extract at 0.1%, 0.2% and 0.3% can be used instead of 0.1% synthetic antioxidant BHA in beef meatballs.

Keywords: antioxidant, beef meatball, BHA, moringa leaf extract, quality

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Authors: Arfan Ali, Muhammad Shahzad Iqbal, Idrees Ahmad Nasir

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Potato (Solanum tuberosum L.) is ranked among the top leading staple foods in the world. Salinity adversely affects potato crop yield and quality. Therefore, increased level of salt tolerance is a key factor to ensure high yield. The present study focused on the Agrobacterium-mediated transformation of Atriplex canescens betaine aldehyde dehydrogenase (BADH) gene, using single, double and triple CAMV35s promoter to improve salt tolerance in potato. Detection of seven potato lines harboring BADH gene, followed by identification of T-DNA insertions, determination of transgenes copies no through Southern Hybridization and quantification of BADH protein through Enzyme Linked Immunosorbent Assay were considered in this study. The results clearly depict that the salt tolerance of potato was found to be promoter-dependent, as the potato transgenic lines with triple promoter showed 4.4 times more glycine betaine production which consequently leads towards high resistance to salt stress as compared to transgenic potato lines with single and double promoters having least production of glycine betaine. Moreover, triple promoter transgenic potato lines have also shown lower levels of H2O2, malondialdehyde (MDA), relative electrical conductivity, high proline and chlorophyll content as compared other two lines having a single and double promoter. Insilco analysis also confirmed that Atriplex canescens BADH has the tendency to interact with sodium ions and water molecules. Taken together these facts it can be concluded that over-expression of BADH under triple CAMV35s promoter with more glycine betaine, chlorophyll & MDA contents, high relative quantities of other metabolites results in an enhanced level of salt tolerance in potato.

Keywords: Atriplex canescens, BADH, CAMV35s promotor, potato, Solanum tubersum

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Authors: Mohamad Mohsen Homayouni, Niloofar Taghipour, Ahmad Reza Memar, Niloofar Khalaji, Hamed Kiani, Seyyed Javad Seyyed Tabaei

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Background and Objective: Cryptosporidium is a coccidian protozoan parasite; its oocysts in surface water are a global health problem. Due to the low number of parasites in the water resources and the lack of laboratory culture, rapid and sensitive method for detection of the organism in the water resources is necessarily required. We applied modified acid-fast staining and PCR for the detection of the Cryptosporidium spp. and analysed the genotypes in 55 samples collected from surface water. Methods: Over a period of nine months, 55 surface water samples were collected from the five rivers in Tehran, Iran. The samples were filtered by using cellulose acetate membrane filters. By acid fast method, initial identification of Cryptosporidium oocyst were carried out on surface water samples. Then, nested PCR assay was designed for the specific amplification and analysed the genotypes. Results: Modified Ziehl-Neelsen method revealed 5–20 Cryptosporidium oocysts detected per 10 Liter. Five out of the 55 (9.09%) surface water samples were found positive for Cryptosporidium spp. by Ziehl-Neelsen test and seven (12.7%) were found positive by nested PCR. The staining results were consistent with PCR. Seven Cryptosporidium PCR products were successfully sequenced and five gp60 subtypes were detected. Our finding of gp60 gene revealed that all of the positive isolates were Cryptosporidium parvum and belonged to subtype families IIa and IId. Conclusion: Our investigations were showed that collection of water samples were contaminated by Cryptosporidium, with potential hazards for the significant health problem. This study provides the first report on detection and genotyping of Cryptosporidium species from surface water samples in Iran, and its result confirmed the low clinical incidence of this parasite on the community.

Keywords: Cryptosporidium spp., membrane filtration, subtype, surface water, Iran

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Authors: Cristian Moisa, Andreea Lupitu, Adriana Csakvari, Dana G. Radu, Leonard Marian Olariu, Georgeta Pop, Dorina Chambre, Lucian Copolovici, Dana Copolovici

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Green synthesis of nanoparticles (NPs) represents a promising, accessible, eco-friendly, and safe process with significant applications in biotechnology, pharmaceutical sciences, and farming. The aim of our study was to obtain silver nanoparticles, using plant wastes extracts resulted in the essential oils extraction process: Thymus vulgaris L., Origanum vulgare L., Lavandula angustifolia L., and in hemp processing for seed and fibre, Cannabis sativa. Firstly, we obtained aqueous extracts of thyme, oregano, lavender, and hemp (two monoicous and one dioicous varieties), all harvested in western part of Romania. Then, we determined the chemical composition of the extracts by liquid-chromatography coupled with diode array and mass spectrometer detectors. The compounds identified in the extracts were in agreement with earlier published data, and the determination of the antioxidant activity of the obtained extracts by DPPH (2,2-diphenyl-1-picrylhydrazyl) and ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)) assays confirmed their antioxidant activity due to their total polyphenolic content evaluated by Folin-Ciocalteu assay. Then, the silver nanoparticles (AgNPs) were successfully biosynthesised, as was demonstrated by UV-VIS, FT-IR spectroscopies, and SEM, by reacting AgNO₃ solution and plant extracts. AgNPs were spherical in shape, with less than 30 nm in diameter, and had a good bactericidal activity against Gram-positive (Staphylococcus aureus) and Gram-negative bacteria (Escherichia coli, Klebsiella pneumoniae, Pseudomonas fluorescens).

Keywords: plant wastes extracts, chemical composition, high performance liquid chromatography mass spectrometer, HPLC-MS, scanning electron microscopy, SEM, silver nanoparticles

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Authors: Mohammadjavad Mehrabanpour, Maryam Ranjbar Bushehri, Dorsa Mehrabanpour

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Respiratory diseases are the most important problems in the country’s poultry industry, particularly when it comes to broiler flocks. Ornithobacterium rhinotracheale (ORT) is a species that causes poor performance in growth rate, egg production, and mortality. This pathogen causes a respiratory infection including pulmonary alveolar inflammation, and pneumonia of birds throughout the world. Newcastle disease (ND) is a highly contagious disease in poultry, and also, it causes considerable losses to the poultry industry. The aim of this study was to evaluate the simultaneous occurrence of ORT and ND and NDV isolation by inoculation in embryonated eggs and confirmed by RT-PCR in broiler chicken flocks in Fars province. In this study, 318 blood and 85 tissue samples (brain, trachea, liver, and cecal tonsils) were collected from 15 broiler chicken farms. Survey serum antibody titers against ORT by using a commercial enzyme-linked immunosorbent assay (ELISA) kit performed. Evaluation of antibody titer against ND virus is performed by hemagglutination inhibition test. Virus isolation with chick embryo eggs 9-11 and RT-PCR method were carried out. A total of 318 serum samples, 135 samples (42.5%) were positive for antibodies to ORT and titer of HI antibodies against NDV in 122 serum samples (38/4%) were 7-10 (log2) and 61 serum samples (19/2%) had occurrence antibody titer against Newcastle virus and ORT. Results of the present study indicated that 20 tissue samples were positive in embryonated egg and in rapid hemagglutination (HA) test. HI test with specific ND positive serum confirmed that 6 of 20 samples. PCR confirmed that all six samples were positive and PCR products of samples indicated 535-base pair fragments in electrophrosis. Due to the great economic importance of these two diseases in the poultry industry, it is necessary to design and implement a comprehensive plan for prevention and control of these diseases.

Keywords: ELISA, Ornithobacterium rhinotracheale, newcastle disease, seroprevalence

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Authors: Stelvin Sebastian, Andriya Annie Tom, Joyalanna Babu, Merin Joshy

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Introduction: The worldwide increase in the use of antibiotics in poultry and livestock industry to treat and prevent bacterial diseases and as growth promoters in feeds has led to the problem of development of antibiotic resistance both in animals and human population. Aim: To study the pattern of antibiotic use and prevalence of multidrug-resistant Escherichia coli in poultry chickens in selected farms in Muvattupuzha and to compare the spread of multidrug-resistant bacteria from poultry environment to UTI patients. Methodology: Two farms from each of 6 localities in Muvattupuzha were selected. A questionnaire on the pattern of antibiotic use and various farming practices were surveyed from farms. From each farm, 60samples of fresh fecal matter, litter from inside, litter from the outside shed, agricultural soil and control soil were collected, and antimicrobial susceptibility testing of E. coli was done. Antibiogram of UTI patients was collected from the secondary care hospital included in the study, and those were compared with resistance patterns of poultry samples. Results: From survey response antibiotics such as ofloxacin, enrofloxacin, levofloxacin, ciprofloxacin, colistin, ceftriaxone, neomycin, cephalexin, and oxytetracycline were used for treatment and prevention of infections in poultry. 31of 48 samples (51.66%) showed E. coli growth. 7 of 15 antibiotics (46.6%) showed resistance. Ampicillin, amoxicillin, meropenem, tetracycline showed 100% resistance to all samples. Statistical analysis confirmed similar resistance pattern in the poultry environment and UTI patients for antibiotics such as ampicillin, amoxicillin, amikacin, and ofloxacin. Conclusion: E. coli were resistant not only to extended-spectrum beta-lactams but also to carbapenems, which may be disseminated to the environment where litter was used as manure. This may due to irrational use of antibiotics in chicken or from their use in poultry feed as growth promoters. The study concludes the presence of multidrug-resistant E.coli in poultry and its spread to environment and humans, which may cause potentially serious implications for human health.

Keywords: multidrug resistance, escherichia coli, urinary tract infection, poultry

Procedia PDF Downloads 130

Authors: Vivek B. Ravindran, Aravind Surapaneni, Rebecca Traub, Sarvesh K. Soni, Andrew S. Ball

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Parasitic diseases have a devastating, long-term impact on human health and welfare. More than two billion people are infected with soil-transmitted helminths (STHs), including the roundworms (Ascaris), hookworms (Necator and Ancylostoma) and whipworm (Trichuris) with majority occurring in the tropical and subtropical regions of the world. Despite its low prevalence in developed countries, the removal of STHs from wastewater remains crucial to allow the safe use of sludge or recycled water in agriculture. Conventional methods such as incubation and optical microscopy are cumbersome; consequently, the results drastically vary from person-to-person observing the ova (eggs) under microscope. Although PCR-based methods are an alternative to conventional techniques, it lacks the ability to distinguish between viable and non-viable helminth ova. As a result, wastewater treatment industries are in major need for radically new and innovative tools to detect and quantify STHs eggs with precision, accuracy and being cost-effective. In our study, we focus on the following novel and innovative techniques: -Recombinase polymerase amplification and Surface enhanced Raman spectroscopy (RPA-SERS) based detection of helminth ova. -Use of metal nanoparticles and their relative nanozyme activity. -Colorimetric detection, differentiation and enumeration of genera of helminth ova using hydrolytic enzymes (chitinase and lipase). -Propidium monoazide (PMA)-qPCR to detect viable helminth ova. -Modified assay to recover and enumerate helminth eggs from fresh raw sewage. -Transcriptome analysis of ascaris ova in fresh raw sewage. The aforementioned techniques have the potential to replace current conventional and molecular methods thereby producing a standard protocol for the determination and enumeration of helminth ova in sewage sludge.

Keywords: colorimetry, helminth, PMA-QPCR, nanoparticles, RPA, viable

Procedia PDF Downloads 282

Authors: Lukman Ahmad Jamil, Heather M. Wallace

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Introduction: Numerous epidemiological studies have shown a positive as well as negative association and no association in some cases between chronic use of calcium channel blockers and the increased risk of developing cancer. However, these associations were enmeshed with controversies in the absence of laboratory based studies to back up those claims. Aim: The aim of this study was to determine in mechanistic terms the association between the long-term administration of nifedipine and diltiazem and increased risk of developing cancer using the human embryonic kidney (HEK293) cell line. Methods: Cell counting using the Trypan blue dye exclusion and 3-4, 5-Dimethylthiazol-2-yl-2, 5-diphenyl-tetrazolium bromide (MTT) assays were used to investigate the effect of nifedipine and diltiazem on the growth pattern of HEK293 cells. Protein assay using modified Lowry method and analysis of intracellular polyamines concentration using Liquid Chromatography – Tandem Mass Spectrometry (LC-MS) were performed to ascertain the mechanism through which chronic use of nifedipine increases the risk of developing cancer. Results: Both nifedipine and diltiazem significantly increased the proliferation of HEK293 cells dose and time dependently. This proliferative effect after 24, 48 and 72-hour incubation period was observed at 0.78, 1.56 and 25 µM for nifedipine and 0.39, 1.56 and 25 µM for diltiazem, respectively. The increased proliferation of the cells was found to be statistically significantly (p<0.05). Furthermore, the increased proliferation of the cells induced by nifedipine was associated with the increase in the protein content and elevated intracellular polyamines concentration level. Conclusion: The chronic use of nifedipine is associated with increased proliferation of cells with concomitant elevation of polyamines concentration and elevated polyamine levels have been implicated in many malignant transformations and hence, these provide a possible explanation on the link between long term use of nifedipine and development of some human cancers. Further studies are needed to evaluate the cause of this association.

Keywords: cancer, nifedipine, polyamine, proliferation

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Authors: Aliro Villacorta, Laura Rubio, Mohamed Alaraby, Montserrat López Mesas, Victor Fuentes-Cebrian, Oscar H. Moriones, Ricard Marcos, Alba Hernández.

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Micro and nano plastics (MNPLs) are emergent environmental pollutants requiring urgent information on their potential risks to human health. One of the problems associated with the evaluation of their undesirable effects is the lack of real samples matching those resulting from the environmental degradation of plastic wastes. To such end, we propose an easy method to obtain polyethylene terephthalate nano plastics from water plastic bottles (PET-NPLs) but, in principle, applicable to any other plastic goods sources. An extensive characterization indicates that the proposed process produces uniform samples of PET-NPLs of around 100 nm, as determined by using a multi-angle and dynamic light scattering methodology. An important point to be highlighted is that to avoid the metal contamination resulting from methods using metal blades/burrs for milling, trituration, or sanding, we propose to use diamond burrs to produce metal-free samples. To visualize the toxicological profile of the produced PET-NPLs, we have evaluated their ability to be internalized by cells, their cytotoxicity, and their ability to induce oxidative stress and induce DNA damage. In this preliminary approach, we have detected their cellular uptake, but without the induction of significant biological effects. Thus, no relevant increases in toxicity, reactive oxygen species (ROS) induction, or DNA damage -as detected with the comet assay- have been observed. The use of real samples, as produced in this study, will generate relevant data in the discussion about the potential health risks associated with MNPLs exposures.

Keywords: nanoplastics, polyethylene terephthalate, physicochemical characterization, cell uptake, cytotoxicity

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Authors: Younus Mohammad, Anita B. Fallah, Ben J. Boyd, Shakila B. Rizwan

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N-acylethanolamines (NAEs) are endogenous lipids, which have neuromodulatory properties. NAEs have shown neuroprotective properties in various neurodegenerative diseases including Alzheimer's disease, Parkinson's disease and ischemic stroke. However, NAEs are eliminated rapidly in vivo by enzymatic hydrolysis. We propose to encapsulate NAEs in liquid crystalline nanoparticles (cubosomes) to increase their biological half-life and explore their therapeutic potential. Recently, we have reported the co-formulation and nanostructural characterization of cubosomes containing the NAE, oleoylethanolamide and a synthetic cubosome forming lipid phytantriol. Here, we report on the formulation of cubosomes with the NAE, linoleoylethanolamide (LEA) as the core cubosome forming lipid. LEA-cubosomes were formulated in the presence of three different steric stabilisers: two brain targeting ligands, Tween 80 and Pluronic P188 and a control, Pluronic F127. Size, morphology and internal structure of formulations were characterized by dynamic light scattering (DLS), cryogenic transmission electron microscopy (Cryo–TEM) and small angle X–ray scattering (SAXS), respectively. Chemical stability of LEA in formulations was investigated using high-performance liquid chromatography (HPLC). Cytotoxicity of formulations towards human cerebral microvascular endothelial cell line (hCMEC/D3) was also investigated using an MTT (3-[4, 5- dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide) assay. All cubosome formulations had mean particle size of less than 250 nm and were uniformly distributed with polydispersity indices less than 0.2. Cubosomes produced had a bicontinuous cubic internal structure with an Im3m space group but different lattice parameters, indicating the different modes of interaction between the stabilisers and LEA. LEA in formulations was found to be chemically stable. At concentrations of up to 20 µg/mL LEA in the presence of all the stabilisers, greater than 80% cell viability was observed.

Keywords: blood-brain barrier, cubosomes, linoleoyl ethanolamide, N-acylethanolamines (NAEs)

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

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

Keywords: cisplatin, Taxol, carcinoma, oral squamous cells

Procedia PDF Downloads 390