Search results for: cellular inflammation

Authors: Dugeree Otgongerel, Kyong Jin Cho, Yu-Han Kim, Sangmee Ahn Jo

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Excessive activation of glutamate receptor is thought to be involved in retinal ganglion cell (RGC) death after ischemia- reperfusion damage. Glutamate carboxypeptidase II (GCPII) is an enzyme responsible for the synthesis of glutamate. Several studies showed that inhibition of GCPII prevents or reduces cellular damage in brain diseases. Thus, in this study, we examined the expression of GCPII in rat retina and the role of GCPII in acute high IOP ischemia-reperfusion damage of eye by using a GCPII inhibitor, 2-(phosphonomethyl) pentanedioic acid (2-PMPA). Animal model of ischemia-reperfusion was induced by raising the intraocular pressure for 60 min and followed by reperfusion for 3 days. Rats were randomly divided into four groups: either intra-vitreous injection of 2-PMPA (11 or 110 ng per eye) or PBS after ischemia-reperfusion, 2-PMPA treatment without ischemia-reperfusion and sham-operated normal control. GCPII immunoreactivity in normal rat retina was detected weakly in retinal nerve fiber layer (RNFL) and retinal ganglionic cell layer (RGL) and also inner plexiform layer (IPL) and outer plexiform layer (OPL) strongly where are co-stained with an anti-GFAP antibody, suggesting that GCPII is expressed mostly in Muller and astrocytes. Immunostaining with anti-BRN antibody showed that ischemia- reperfusion caused RGC death (31.5 %) and decreased retinal thickness in all layers of damaged retina, but the treatment of 2-PMPA twice at 0 and 48 hour after reperfusion blocked these retinal damages. GCPII level in RNFL layer was enhanced after ischemia-reperfusion but was blocked by PMPA treatment. This result was confirmed by western blot analysis showing that the level of GCPII protein after ischemia- reperfusion increased by 2.2- fold compared to control, but this increase was blocked almost completely by 110 ng PMPA treatment. Interestingly, GFAP immunoreactivity in the retina after ischemia- reperfusion followed by treatment with PMPA showed similar pattern to GCPII, increase after ischemia-reperfusion but reduction to the normal level by PMPA treatment. Our data demonstrate that increase of GCPII protein level after ischemia-reperfusion injury is likely to cause glial activation and/or retinal cell death which are mediated by glutamate, and GCPII inhibitors may be useful in treatment of retinal disorders in which glutamate excitotoxicity is pathogenic.

Keywords: glutamate carboxypepptidase II, glutamate excitotoxicity, ischemia-reperfusion, retinal ganglion cell

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Authors: Abdulkarem Qasem Dammag, Basema Qasim Dammag, Jian Dai

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Change in Land use and Land cover (LU/LC) has a profound impact on the area's natural, economic, and ecological development, and the search for drivers of land cover change is one of the fundamental issues of LU/LC change. The study aimed to assess the temporal and Spatio-temporal dynamics of LU/LC in the past and to predict the future using Landsat images by exploring the characteristics of different LU/LC types. Spatio-temporal patterns of LU/LC change in Ibb Governorate, Yemen, were analyzed based on RS and GIS from 1990, 2005, and 2020. A socioeconomic survey and key informant interviews were used to assess potential drivers of LU/LC. The results showed that from 1990 to 2020, the total area of vegetation land decreased by 5.3%, while the area of barren land, grassland, built-up area, and waterbody increased by 2.7%, 1.6%, 1.04%, and 0.06%, respectively. Based on socio-economic surveys and key informant interviews, natural factors had a significant and long-term impact on land change. In contrast, site construction and socio-economic factors were the main driving forces affecting land change in a short time scale. The analysis results have been linked to the CA-Markov Land Use simulation and forecasting model for the years 2035 and 2050. The simulation results revealed from the period 2020 to 2050, the trend of dynamic changes in land use, where the total area of barren land decreased by 7.0% and grassland by 0.2%, while the vegetation land, built-up area, and waterbody increased by 4.6%, 2.6%, and 0.1 %, respectively. Overall, these findings provide LULC's past and future trends and identify drivers, which can play an important role in sustainable land use planning and management by balancing and coordinating urban growth and land use and can also be used at the regional level in different levels to provide as a reference. In addition, the results provide scientific guidance to government departments and local decision-makers in future land-use planning through dynamic monitoring of LU/LC change.

Keywords: LU/LC change, CA-Markov model, driving forces, change detection, LU/LC change simulation

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Authors: Y. Landkocz, C. Lepers, P.J. Martin, B. Fougère, F. Roy Saint-Georges. A. Verdin, F. Cazier, F. Ledoux, D. Courcot, F. Sichel, P. Gosset, P. Shirali, S. Billet

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In 2013, the International Agency for Research on Cancer (IARC) classified air pollution and fine particles as carcinogenic to humans. Causal relationships exist between elevated ambient levels of airborne particles and increase of mortality and morbidity including pulmonary diseases, like lung cancer. However, due to a double complexity of both physicochemical Particulate Matter (PM) properties and tumor mechanistic processes, mechanisms of action remain not fully elucidated. Furthermore, because of several common properties between air pollution PM and tobacco smoke, like the same route of exposure and chemical composition, potential mechanisms of synergy could exist. Therefore, smoking could be an aggravating factor of the particles toxicity. In order to identify some mechanisms of action of particles according to their size, two samples of PM were collected: PM0.03 2.5 and PM0.33 2.5 in the urban-industrial area of Dunkerque. The overall cytotoxicity of the fine particles was determined on human bronchial cells (BEAS-2B). Toxicological study focused then on the metabolic activation of the organic compounds coated onto PM and some genetic and epigenetic changes induced on a co-culture model of BEAS-2B and alveolar macrophages isolated from bronchoalveolar lavages performed in smokers and non-smokers. The results showed (i) the contribution of the ultrafine fraction of atmospheric particles to genotoxic (eg. DNA double-strand breaks) and epigenetic mechanisms (eg. promoter methylation) involved in tumor processes, and (ii) the influence of smoking on the cellular response. Three main conclusions can be discussed. First, our results showed the ability of the particles to induce deleterious effects potentially involved in the stages of initiation and promotion of carcinogenesis. The second conclusion is that smoking affects the nature of the induced genotoxic effects. Finally, the in vitro developed cell model, using bronchial epithelial cells and alveolar macrophages can take into account quite realistically, some of the existing cell interactions existing in the lung.

Keywords: air pollution, fine and ultrafine particles, genotoxic and epigenetic alterations, smoking

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Authors: Ludmila A. Gavriliuc, Jerry McLarty, Heather E. Kleiner, J. Michael Mathis

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Abstract -- Background: The citrus coumarine Auraptene (Aur) is an effective chemopreventive agent, as manifested in many models of diseases and cancer. Nuclear factor erythroid 2-related factor (Nrf2) is an important regulator of genes induced by oxidative stress, such as glutathione S-transferases, heme oxygenase-1, and peroxiredoxin 1, by activating the antioxidant response element (ARE). Genetic and biochemical evidence has demonstrated that glutathione (GSH) and glutathione-dependent enzymes, glutathione reductase (GR), glutathione peroxidases (GPs), glutathione S-transferases (GSTs) are responsible for the control of intracellular reduction-oxidation status and participate in cellular adaptation to oxidative stress. The effect of Aur on the activity of GR, GPs (Se-GP and Se-iGP), and content of GSH in the liver, kidney, and spleen is insufficiently explored. Aim: Our goal was the examination of the Aur influence on the redox-system of GSH in Nrf2 wild type and Nrf2 knockout mice via activation of Nrf2 and ARE. Methods: Twenty female mice, 10 Nrf2 wild-type (WT) and 10 Nrf2 (-/-) knockout (KO), were bred and genotyped for our study. The activity of GR, Se-GP, Se-iGP, GST, G6PD, CytP450 reductase, catalase (Cat), and content of GSH were analyzed in the liver, kidney, and spleen using Spectrophotometry methods. The results of the specific activity of enzymes and the amount of GSH were analyzed with ANOVA and Spearman statistical methods. Results: Aur (200 mg/kg) treatment induced hepatic GST, GR, Se-GP activity and inhibited their activity in the spleen of mice, most likely via activation of the ARE through Nrf2. Activation in kidney Se-GP and G6PD by Aur is also controlled, apparently through Nrf2. Results of the non-parametric Spearman correlation analysis indicated the strong positive correlation between GR and G6PD only in the liver in WT control mice (r=+0.972; p < 0.005) and in the kidney KO control mice (r=+0.958; p < 0.005). The observed low content of GSH in the liver of KO mice indicated an increase in its participation in the neutralization of toxic substances with the absence of induction of GSH-dependent enzymes, such as GST, GR, Se-GP, and Se-iGP. Activation of CytP450 in kidney and spleen and Cat in the liver in KO mice probably revealed another regulatory mechanism for these enzymes. Conclusion: Thereby, obtained results testify that Aur can modulate the activity of genes and antioxidant enzymatic redox-system of GSH, responsible for the control of intracellular reduction-oxidation status.

Keywords: auraptene, glutathione, GST, Nrf2

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Authors: Xiaojie Cheng, Ulrike Frank, Feng Zhao, Karin Pritsch

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Ragweed (Ambrosia artemisiifolia) is an invasive weed that has become an increasing global problem. In addition to affecting land use and crop yields, ragweed has a strong impact on human health as it produces highly allergenic pollen. Global warming will result in an earlier and longer pollen season enhanced pollen production and an increase in pollen allergenicity with a negative effect on atopic patients. The aims of this study were to investigate the effects of increasing temperature, the future climate scenario in the Munich area, southern Germany, predicted on the basis of RCP8.5 until the end of 2050s, or/and NO₂, a major air pollutant, 1) on the vegetative and reproductive characteristics of ragweed plants, 2) on the total allergenicity of ragweed pollen, 3) on the total pollen proteomic patterns. Ragweed plants were cultivated for the whole plant vegetation period under controlled conditions either under ambient climate conditions or 4°C higher temperatures with or without additional NO₂. Higher temperature resulted in bigger plant sizes, longer male inflorescences, and longer pollen seasons. The total allergenic potential of the pollen was accessed by dot blot using serum from ragweed pollen sensitized patients. The comparative immunoblot analysis revealed that the in vivo fumigation of ragweed plants with elevated NO₂-concentrations significantly increased the allergenic potential of the pollen, and in combination with increased temperature, the allergenic potential was even higher. On the other hand, label-free protein quantification by liquid chromatography-tandem mass spectrometry (LC-MS/MS) was performed. The results showed that more proteins were significantly up- and down-regulated under higher temperatures with/without elevated NO₂ conditions. Most of the highly expressed proteins were participating intensively in the metabolic process, the cellular process, and the stress defense process. These findings suggest that rising temperature and elevated NO₂ are important environmental factors for higher abiotic stress activities, catalytic activities, and thus higher allergenic potential observed in pollen proteins.

Keywords: climate change, NO₂, pollen proteome, ragweed, temperature

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Authors: Kamila Dus-Szachniewicz, Artur Bednarkiewicz, Katarzyna Gdesz-Birula, Slawomir Drobczynski

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In modern oncology hyperthermia (HT) is defined as a controlled tumor heating. HT treatment temperatures range between 40–48 °C and can selectively damage heat-sensitive cancer cells or limit their further growth, usually with minimal injury to healthy tissues. Despite many advantages, conventional whole-body and regional hyperthermia have clinically relevant side effects, including cardiac and vascular disorders. Additionally, the lack of accessibility of deep-seated tumor sites and impaired targeting micrometastases renders HT less effective. It is believed that above disadvantages can significantly overcome by the application of biofunctionalized microparticles, which can specifically target tumor sites and become activated by an external stimulus to provide a sufficient cellular response. In our research, the unique optical tweezers system have enabled capturing the silica microparticles, primary cells and tumor spheroids in highly controllable and reproducible environment to study the impact of localized heat stimulation on normal and pathological cell and within multicellular tumor spheroid. High throughput spheroid model was introduced to better mimic the response to HT treatment on tumors in vivo. Additionally, application of local heating of tumor spheroids was performed in strictly controlled conditions resembling tumor microenvironment (temperature, pH, hypoxia, etc.), in response to localized and nonhomogeneous hyperthermia in the extracellular matrix, which promotes tumor progression and metastatic spread. The lack of precise control over these well- defined parameters in basic research leads to discrepancies in the response of tumor cells to the new treatment strategy in preclinical animal testing. The developed approach enables also sorting out subclasses of cells, which exhibit partial or total resistance to therapy, in order to understand fundamental aspects of the resistance shown by given tumor cells in response to given therapy mode and conditions. This work was funded by the National Science Centre (NCN, Poland) under grant no. UMO-2017/27/B/ST7/01255.

Keywords: cancer spheroids, hyperthermia, microparticles, optical tweezers

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Authors: Mauricio A. Sarabia-Vallejos, Carmen M. Gonzalez-Henriquez, Adolfo Del Campo-Garcia, Aitzibier L. Cortajarena, Juan Rodriguez-Hernandez

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In this study, it was explored the formation and the morphological differences between wrinkled hydrogel patterns obtained via generation of surface instabilities. The slight variations in the polymerization conditions produce important changes in the material composition and pattern structuration. The compounds were synthesized using three main components, i.e. an amphiphilic monomer, hydroxyethyl methacrylate (HEMA), a hydrophobic monomer, trifluoroethyl methacrylate (TFMA), and a hydrophilic crosslinking agent, poly(ethylene glycol) diacrylate (PEGDA). The first part of this study was related to the formation of wrinkled surfaces using only HEMA and PEGDA and varying the amount of water added in the reaction. The second part of this study involves the gradual insertion of TFMA into the hydrophilic reaction mixture. Interestingly, the manipulation of the chemical composition of this hydrogel affects both surface morphology and physicochemical characteristics of the patterns, inducing transitions from one particular type of structure (wrinkles or ripples) to different ones (creases, folds, and crumples). Contact angle measurements show that the insertion of TFMA produces a slight decrease in surface wettability of the samples, remaining however highly hydrophilic (contact angle below 45°). More interestingly, by using confocal Raman spectroscopy, important information about the wrinkle formation mechanism is obtained. The procedure involving two consecutive thermal and photopolymerization steps lead to a “pseudo” two-layer system. Thus, upon photopolymerization, the surface is crosslinked to a higher extent than the bulk and water evaporation drives the formation of wrinkled surfaces. Finally, cellular, and bacterial proliferation studies were performed to the samples, showing that the amount of TFMA included in each sample slightly affects the proliferation of both (bacteria and cells), but in the case of bacteria, the morphology of the sample also plays an important role, importantly reducing the bacterial proliferation.

Keywords: antibiofouling properties, hydrophobic/hydrophilic balance, morphologic characterization, wrinkled hydrogel patterns

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Authors: Mustafa M. Donma, Orkide Donma

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The impact of metabolic syndrome (MetS) on cognition and functions of the brain is being investigated. Iron deficiency and deficiencies of B9 (folate) as well as B12 (cobalamin) vitamins are best-known nutritional anemias. They are associated with cognitive disorders and learning difficulties. The antidepressant effects of vitamin D are known and the deficiency state affects mental functions negatively. The aim of this study is to investigate possible correlations of MetS with serum brain-derived neurotrophic factor (BDNF), iron, folate, cobalamin and vitamin D in pediatric patients. 30 children, whose age- and sex-dependent body mass index (BMI) percentiles vary between 85 and 15, 60 morbid obese children with above 99^th percentiles constituted the study population. Anthropometric measurements were taken. BMI values were calculated. Age- and sex-dependent BMI percentile values were obtained using the appropriate tables prepared by the World Health Organization (WHO). Obesity classification was performed according to WHO criteria. Those with MetS were evaluated according to MetS criteria. Serum BDNF was determined by enzyme-linked immunosorbent assay. Serum folate was analyzed by an immunoassay analyzer. Serum cobalamin concentrations were measured using electrochemiluminescence immunoassay. Vitamin D status was determined by the measurement of 25-hydroxycholecalciferol [25-hydroxy vitamin D3, 25(OH)D] using high performance liquid chromatography. Statistical evaluations were performed using SPSS for Windows, version 16. The p values less than 0.05 were accepted as statistically significant. Although statistically insignificant, lower folate and cobalamin values were found in MO children compared to those observed for children with normal BMI. For iron and BDNF values, no alterations were detected among the groups. Significantly decreased vitamin D concentrations were noted in MO children with MetS in comparison with those in children with normal BMI (p ≤ 0.05). The positive correlation observed between iron and BDNF in normal-BMI group was not found in two MO groups. In THE MetS group, the partial correlation among iron, BDNF, folate, cobalamin, vitamin D controlling for waist circumference and BMI was r = -0.501; p ≤ 0.05. None was calculated in MO and normal BMI groups. In conclusion, vitamin D should also be considered during the assessment of pediatric MetS. Waist circumference and BMI should collectively be evaluated during the evaluation of MetS in children. Within this context, BDNF appears to be a key biochemical parameter during the examination of obesity degree in terms of mental functions, cognition and learning capacity. The association observed between iron and BDNF in children with normal BMI was not detected in MO groups possibly due to development of inflammation and other obesity-related pathologies. It was suggested that this finding may contribute to mental function impairments commonly observed among obese children.

Keywords: brain-derived neurotrophic factor, iron, vitamin B9, vitamin B12, vitamin D

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Authors: Kumari Rekha, Mathur K Dhananjay, Maheshwari Deepanshu, Nautiyal Nidhi, Shubham Smriti, Laal Deepika, Sinha Swati, Kumar Anupam, Biswas Subhrajit, Shiv Kumar Sarin

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Background: Mesenchymal stem cells are multipotent stem cells derived from mesoderm and are used for therapeutic purposes because of their self-renewal, homing capacity, Immunomodulatory capability, low immunogenicity and mitochondrial transfer signaling. MSCs have the ability to regulate the mechanism of both innate as well as adaptive immune responses through the modulation of cellular response and the secretion of inflammatory mediators. Different sources of MSC are UC MSC, BM MSC, Dental Pulp, and Adipose MSC. The most frequent source used is umbilical cord tissue due to its being easily available and free of limitations of collection procedures from respective hospitals. The immunosuppressive role of MSCs is particularly interesting for clinical use since it confers resistance to rejection by the host immune response. Methodology: In this study, T helper cells (TH4), Cytotoxic T cells (CD-8), immunoregulatory cells (CD25 +FOXP3+) are compared from isolated MSC from three different methods, UC Dissociation Kit (Miltenyi), Explant Culture and Collagenase Type-IV. To check the immunomodulatory property, these MSCs were seeded with PBMC(Coculture) in CD3 coated 24 well plates. Cd28 antibody was added in coculture for six days. The coculture was analyzed in FACS Verse flow cytometry. Results: From flow cytometry analysis of coculture, it found that All over T helper cells (CD4+) number p<0.0264 increases in (All Enzymes) MSC rather than explant MSC(p>0.0895) as compared to Collagenase(p>0.7889) in a coculture of Activated T cell and Mesenchymal Stem Cell. Similar T reg cells (CD25+, FOXP3+) expression p<0.0234increases in All Enzymes), decreases in Explant and Collagenase. Experiments have shown that MSCs can also directly prevent the cytotoxic activity of CD8 lymphocytes mainly by blocking their proliferation rather than by inhibiting the cytotoxic effect. And promoting the t-reg cells, which helps in the mediation of immune response in various diseases. Conclusion: MSC suppress Cytotoxic CD8 T cell and Enhance immunoregulatory T reg (CD4+, CD25+, FOXP3+) Cell expression. Thus, MSC maintains a proper balance(ratio) between CD4 T cells and Cytotoxic CD8 T cells.

Keywords: MSC, disease, T cell, T regulatory

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Authors: Parimelazhagan Thangaraj

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Natural products serves human kind as a source of all drugs and higher plants provide most of these therapeutic agents. These products are widely recognized in the pharmaceutical industry for their broad structural diversity as well as their wide range of pharmacological activities. Euphorbiaceae is one of the important families with significant pharmacological activities, of which many species has been used traditionally for the treatment of various ailments. Breynia retusa belongs to the family Euphorbiaceae is used to cure ailments like body pain, skin inflammation, hyperglycaemia, diarrhoea, dysentery and toothache. Flowers and young leaves of B. retusa are cooked and eaten, roots are used for meningitis. The juice of the stem is used in conjunctivtis and leaves as poultice to hasten suppuration. Based on the strong evidences of traditional uses of Breynia retusa, the present study was focused on neutraceuticals evaluation of the species with special reference to oxidative stress and diabetes. Both leaves and stem of B. retusa were extracted with different solvents and analyzed for radical scavenging ability wherein ABTS.+ (8396.95±1529.01 µM TEAC/g extract), phosphomolybdenum (17.34±0.08 g AAE/100 g extract) and FRAP (6075.66±414.28 µM Fe (II) E/mg extract) assays showed good radical scavenging activity in stem. Furthermore, leaf extracts showed good radical inhibition in DPPH (2.4 µg/mL), metal ion (27.44±0.09 mg EDTAE/g extract) scavenging methods. The α-amylase and α-glucosidase inhibitors are currently used for diabetic treatment as oral hypoglycemic agents. The inhibitory effects of the B. retusa leaf and stem ethyl acetate extracts showed good inhibition on α-amylase (96.25% and 95.69 respectively) and α-glucosidase (54.50% and 50.87% respectively) enzymes compared to standard acarbose. The proximate composition analysis of B. retusa leaves contains higher amount of total carbohydrates (14.08 g Glucose equivalents/100 g sample), ash (19.04 %) and crude fibre (0.52 %). The examination of mineral profile explored that the leaves was rich in calcium (1891 ppm), sulphur (1406 ppm), copper (2600 ppm) and magnesium (778 ppm). Leaves sample revealed very minimal amount of anti-nutrient contents like trypsin (14.08±0.03 TIU/mg protein) and tannin (0.011±0.001 mg TAE/g sample). The low anti nutritional factors may not pose any serious nutritional problems when these leaves are consumed. In conclusion, it is very clear that dietary compounds from B. retusa are suitable and promising for the development of safe food products and natural additives. Based on the studies, it may be concluded that nutritional composition, antioxidant and anti-diabetic activities this species can be used as future therapeutic medicine.

Keywords: Breynia retusa, nutraceuticals, antioxidant, anti diabetic

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Authors: Epameinondas Xanthakis, Erik Kaunisto, Alain Le-Bail, Lilia Ahrné

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Fruits and vegetables are characterized as perishable food matrices due to their short shelf life as several deterioration mechanisms are being involved. Prior to the common preservation methods like freezing or canning, fruits and vegetables are being blanched in order to inactivate deteriorative enzymes. Both conventional blanching pretreatments and conventional freezing methods hide drawbacks behind their beneficial impacts on the preservation of those matrices. Conventional blanching methods may require longer processing times, leaching of minerals and nutrients due to the contact with the warm water which in turn leads to effluent production with large BOD. An important issue of freezing technologies is the size of the formed ice crystals which is also critical for the final quality of the frozen food as it can cause irreversible damage to the cellular structure and subsequently to degrade the texture and the colour of the product. Herein, the developed microwave blanching methodology and the results regarding quality aspects and enzyme inactivation will be presented. Moreover, heat transfer phenomena, mass balance, temperature distribution, and enzyme inactivation (such as Pectin Methyl Esterase and Ascorbic Acid Oxidase) of our microwave blanching approach will be evaluated based on measurements and computer modelling. The present work is part of the COLDμWAVE project which aims to the development of an innovative environmentally sustainable process for blanching and freezing of fruits and vegetables with improved textural and nutritional quality. In this context, COLDµWAVE will develop tailored equipment for MW blanching of vegetables that has very high energy efficiency and no water consumption. Furthermore, the next steps of this project regarding the development of innovative pathways in MW assisted freezing to improve the quality of frozen vegetables, by exploring in depth previous results acquired by the authors, will be presented. The application of MW assisted freezing process on fruits and vegetables it is expected to lead to improved quality characteristics compared to the conventional freezing. Acknowledgments: COLDμWAVE has received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grand agreement No 660067.

Keywords: blanching, freezing, fruits, microwave blanching, microwave

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Authors: Andleeb Zahra, Itrat Rubab, Sumaira Malik, Amina Khan, Muhammad Jawad Khan, M. Qaiser Fatmi

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Oral squamous cell carcinoma (OSCC) is one of the most common cancers worldwide. Recent studies have highlighted the role of miRNA in disease pathology, indicating its potential use in an early diagnostic tool. miRNAs are small, double stranded, non-coding RNAs that regulate gene expression by deregulating mRNAs. miRNAs play important roles in modifying various cellular processes such as cell growth, differentiation, apoptosis, and immune response. Dis-regulated expression of miRNAs is known to affect the cell growth, and this may function as tumor suppressors or oncogenes in various cancers. Objectives: The main objectives of this study were to characterize the extracellular miRNAs involved in oral cancer (OC) to assist early detection of cancer as well as to propose a list of genes that can potentially be used as biomarkers of OC. We used gene expression data by microarrays already available in literature. Materials and Methods: In the first step, a total of 318 miRNAs involved in oral carcinoma were shortlisted followed by the prediction of their target genes. Simultaneously, the differentially expressed genes (DEGs) of oral carcinoma from all experiments were identified. The common genes between lists of DEGs of OC based on experimentally proven data and target genes of each miRNA were identified. These common genes are the targets of specific miRNA, which is involved in OC. Finally, a list of genes was generated which may be used as biomarker of OC. Results and Conclusion: In results, we included some of pathways in cancer to show the change in gene expression under the control of specific miRNA. Ingenuity pathway analysis (IPA) provided a list of major biomarkers like CDH2, CDK7 and functional enrichment analysis identified the role of miRNA in major pathways like cell adhesion molecules pathway affected by cancer. We observed that at least 25 genes are regulated by maximum number of miRNAs, and thereby, they can be used as biomarkers of OC. To better understand the role of miRNA with respect to their target genes further experiments are required, and our study provides a platform to better understand the miRNA-OC relationship at genomics level.

Keywords: biomarkers, gene expression, miRNA, oral carcinoma

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Authors: Johanna Perdomo, Riki Lamont, Edmund Pickering, Naomi C. Paxton, Maria A. Woodruff

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Background: Tissue Engineering (TE) has combined advanced materials, such as biomaterials, to create affordable scaffolds and dynamic systems to generate stimulation of seeded cells on these scaffolds, improving and maintaining the cellular growth process in a cell culture. However, Few TE skin products have been clinically translated, and more research is required to produce highly biomimetic skin substitutes that mimic the native elasticity of skin in a controlled manner. Therefore, this work will be focused on the fabrication of a novel mechanical system to enhance the TE treatment approaches for the reparation of damaged tissue skin. Aims: To archive this, a soft robotic device will be created to emulate different deformation of skin stress. The design of this soft robot will allow the attachment of scaffolds, which will then be mechanically actuated. This will provide a novel and highly adaptable platform for dynamic cell culture. Methods: Novel, low-cost soft robot is fabricated via 3D printed moulds and silicone. A low cost, electro-mechanical device was constructed to actuate the soft robot through the controlled combination of positive and negative air pressure to control the different state of movements. Mechanical tests were conducted to assess the performance and calibration of each electronic component. Similarly, pressure-displacement test was performed on scaffolds, which were attached to the soft robot, applying various mechanical loading regimes. Lastly, digital image correlation test was performed to obtain strain distributions over the soft robot’s surface. Results: The control system can control and stabilise positive pressure changes for long hours. Similarly, pressure-displacement test demonstrated that scaffolds with 5µm of diameter and wavy geometry can displace at 100%, applying a maximum pressure of 1.5 PSI. Lastly, during the inflation state, the displacement of silicone was measured using DIC method, and this showed a parameter of 4.78 mm and strain of 0.0652. Discussion And Conclusion: The developed soft robot system provides a novel and low-cost platform for the dynamic actuation of tissue scaffolds with a target towards dynamic cell culture.

Keywords: soft robot, tissue engineering, mechanical stimulation, dynamic cell culture, bioreactor

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Authors: Ji Jiazheng, Wang Jingrao, Jin Xin, Zhang Hong

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Diabetes mellitus is a metabolic disease characterized by high blood sugar. A long-standing hyperglycemic state can lead to various tissue damage. Diabetic retinopathy is the most common and widely studied ocular complication and has become the leading cause of blindness in my country. At the same time, diabetes has profound clinically relevant effects on the cornea, leading to keratopathy and vision-threatening. The cornea is an avascular tissue and is sensitive to hyperglycemia, Keratopathy caused by diabetes is usually chronic, they are called diabetic keratopathy or diabetic neurotrophic keratopathy, leading to several diabetic corneal complications including delayed epithelial wound healing, recurrent erosions, neuropathy, loss of sensitivity. Corneal stem cell dysfunction in diabetic patients as an important influencing factor of diabetic keratopathy. The consequences of this condition are often underestimated. The limbus is located between the cornea and the sclera tissue. The limbal stroma consists of a series of radial elevations with fibrovascular centers known as palisades of Vogt (POV). Previous studies have shown that palisades of Vogt (POV), as the main site of limbal stem cells, plays an important role in the homeostasis of the corneal epithelium. Therefore, POV plays a vital role in the healing of corneal epithelial surgery and postoperative evaluation. IVCM can observe the condition of the corneal epithelium at the cellular level. It has profound significance and guidance for the evaluation of limbal and limbal stem cells. We have previously observed structural changes in POV in HSK and HZO patients on IVCM. At present, there have been reports involving limbal stem cell dysfunction in diabetic patients, but the specific pathogenesis is still unclear. However, there are no studies on POV morphological changes in patients with DM. Therefore, we performed statistics and compared the correlation between POV morphological changes and corneal epithelial basal cell density, corneal nerves, and length of disease in DM patients and normal humans using IVCM studies. At the same time, fundoscopy was used to observe the correlation between the thickness of RNFL and the thickness of GCC and POV in diabetic patients. And to observe the correlation between SVD, DVD and POV for research.

Keywords: confocal microscopy, fundus, limbal stem cells, diabetes

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Authors: Jannis Kountouras, Nikolaos Kapetanakis, Stergios A. Polyzos, Apostolis Papaeftymiou, Panagiotis Katsinelos, Ioannis Venizelos, Christina Nikolaidou, Christos Zavos, Iordanis Romiopoulos, Elena Tsiaousi, Evangelos Kazakos, Michael Doulberis

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Background & Aims: Helicobacter pylori infection (Hp-I) has been recognized as a substantial risk agent involved in gastrointestinal (GI) tract oncogenesis by stimulating cancer stem cells (CSCs), oncogenes, immune surveillance processes, and triggering GI microbiota dysbiosis. We aimed to investigate the possible involvement of active Hp-I in the sequence: chronic inflammation–adenoma–colorectal cancer (CRC) development. Methods: Four pillars were investigated: (i) endoscopic and conventional histological examinations of patients with CRC, colorectal adenomas (CRA) versus controls to detect the presence of active Hp-I; (ii) immunohistochemical determination of the presence of Hp; expression of CD44, an indicator of CSCs and/or bone marrow-derived stem cells (BMDSCs); expressions of oncogene Ki67 and anti-apoptotic Bcl-2 protein; (iii) expression of CD45, indicator of immune surveillance locally (assessing mainly T and B lymphocytes locally); and (iv) correlation of the studied parameters with the presence or absence of Hp-I. Results: Among 50 patients with CRC, 25 with CRA, and 10 controls, a significantly higher presence of Hp-I in the CRA (68%) and CRC group (84%) were found compared with controls (30%). The presence of Hp-I with accompanying immunohistochemical expression of CD44 in biopsy specimens was revealed in a high proportion of patients with CRA associated with moderate/severe dysplasia (88%) and CRC patients with moderate/severe degree of malignancy (91%). Comparable results were also obtained for Ki67, Bcl-2, and CD45 immunohistochemical expressions. Concluding Remarks: Hp-I seems to be involved in the sequence: CRA – dysplasia – CRC, similarly to the upper GI tract oncogenesis, by several pathways such as the following: Beyond Hp-I associated insulin resistance, the major underlying mechanism responsible for the metabolic syndrome (MetS) that increase the risk of colorectal neoplasms, as implied by other Hp-I related MetS pathologies, such as non-alcoholic fatty liver disease and upper GI cancer, the disturbance of the normal GI microbiota (i.e., dysbiosis) and the formation of an irritative biofilm could contribute to a perpetual inflammatory upper GIT and colon mucosal damage, stimulating CSCs or recruiting BMDSCs and affecting oncogenes and immune surveillance processes. Further large-scale relative studies with a pathophysiological perspective are necessary to demonstrate in-depth this relationship.

Keywords: Helicobacter pylori, colorectal cancer, colorectal adenomas, gastrointestinal oncogenesis

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Authors: Ruth Green, Samantha Milton, Rinal Desai

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Background/Aims: Eye pain/swelling/redness is a common presentation to Barnet General Hospital (a district general hospital), pediatric emergency department, and is managed by both the pediatric and emergency teams. The management of each child differs dramatically depending on the healthcare professional who reviews them. There also appears to be confusion in diagnosis between periorbital cellulitis, pre-septal cellulitis, and orbital cellulitis. Pre septal cellulitis refers to an inflammation of the eyelids and soft tissue anterior to the orbital septum. In contrast, orbital cellulitis is a serious, rapidly progressive infection of soft tissues located posterior to the orbital septum. Pre-septal cellulitis is more prevalent and less serious than orbital cellulitis, although it may be part of a continuous spectrum if untreated. Pre-septal cellulitis should there be diagnosed and treated urgently to prevent spread to the septum. For the purpose of the audit, the term periorbital cellulitis has been used as an umbrella term for all spectrums of this infection. The audit aimed to look at, how as a whole, the department is diagnosing and managing orbital and pre-septal cellulitis. Gold Standard: Patients of the same age and diagnosis should be treated with the same medication, advice, and follow-up. Method: Data was collected retrospectively from pediatric patients ( < 18years) who attended the emergency department from June 2019 to February 2020 who had been coded as pre-septal cellulitis, periorbital cellulitis, orbital cellulitis, or eye pain/swelling/redness. Demographics, signs and symptoms, management, and follow-up were recorded for all patients with any of the diagnoses of pre-septal, periorbital, or orbital cellulitis. A Microsoft Excel spreadsheet was used to record the anonymised data. Results: There were vast discrepancies in the diagnosis, management, and follow-up of patients with periorbital cellulitis. Conclusion/Discussion: The audit concluded there is no uniform approach to managing periorbital cellulitis in Barnet General Hospital Paediatric Emergency Department. Healthcare professionals misdiagnosed conjunctivitis as periorbital cellulitis, and adequate steps did not appear to be documented on excluding red flag signs and symptoms of patients presenting. There was no consistency in follow-up, with some patients having timely phone reviews or clinical reviews for mild symptoms. Advice given by the staff was appropriate, and patients did return when symptoms got worse and were treated accordingly. Plan: Given the inconsistency, a gold standard care pathway or local easily accessible clinical guideline can be developed to help with the diagnosis and management of periorbital cellulitis. Along with this, a teaching session can be carried out for the staff of the pediatric team and emergency department to disseminate the teaching. Following the introduction of a guideline and teaching sessions, patients notes can be re-reviewed to check improvement in patient care.

Keywords: periorbital cellulitis, preseptal cellulitis, orbital cellulitis, erythematous eyelid

Procedia PDF Downloads 108

Authors: Reshu Saxena, R. K. Tripathi

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HIV-1 transmission and spread involves significant host-virus interaction. Potential targets for prevention of HIV-1 lies at the site of mucosal barriers. Thus a better understanding of how HIV-1 infects target cells at such sites and lead their invasion is required, with prime focus on the host determinants regulating HIV-1 spread. HIV-1 Nef is important for viral infectivity and pathogenicity. It promotes HIV-1 replication, facilitating immune evasion by interacting with various host factors and altering cellular pathways via multiple protein-protein interactions. In this study nef was sequenced from HIV-1 patients, and showed specific mutations revealing sequence variability in nef. To explore the difference in Nef functionality based on sequence variability we have studied the effects of HIV-1 Nef in human SupT1 T cell line and (THP-1) monocyte-macrophage cell lines through proteomics approach. 2D-Gel Electrophoresis in control and Nef-transfected SupT1 cells demonstrated several differentially expressed proteins with significant modulation of alpha-enolase. Through further studies, effects of Nef on alpha-enolase regulation were found to be cell lineage-specific, being stimulatory in macrophages/monocytes, inhibitory in T cells and without effect in HEK-293 cells. Cell migration and invasion studies were employed to determine biological function affected by Nef mediated regulation of alpha-enolase. Cell invasion was enhanced in THP-1 cells but was inhibited in SupT1 cells by wildtype nef. In addition, the modulation of enolase and cell invasion remained unaffected by a unique nef variant. These results indicated that regulation of alpha-enolase expression and invasive property of host cells by Nef is sequence specific, suggesting involvement of a particular motif of Nef. To precisely determine this site, we designed a heptapeptide including the suggested alpha-enolase regulating sequence of nef and a nef mutant with deletion of this site. Macrophages/monocytes being the major cells affected by HIV-1 at mucosal barriers, were particularly investigated by the nef mutant and peptide. Both the nef mutant and heptapeptide led to inhibition of enhanced enolase expression and increased invasiveness in THP-1 cells. Together, these findings suggest a possible mechanism of host invasion by HIV-1 through Nef mediated regulation of alpha-enolase and identifies a potential therapeutic target for HIV-1 entry at mucosal barriers.

Keywords: HIV-1 Nef, nef variants, host-virus interaction, tissue invasion

Procedia PDF Downloads 386

Authors: Hamid Abbasi, Neda Jourabchi, Ranasadat Abedi, Kiarash Tajernarenj, Mehdi Farhoudi, Sarvin Sanaie

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Background: Alpha lipoic acid (ALA), fat- and water-soluble, coenzyme with sulfuret content, has received considerable attention for its potential therapeutic role in diabetes, cardiovascular diseases, cancers, and central nervous disease. This investigation aims to evaluate the probable protective effects of ALA in stroke patients. Methods: Based on Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines, This meta-analysis was performed. The PICO criteria for this meta-analysis were as follows: Population/Patients (P: stroke patients); Intervention (I: ALA); Comparison (C: control); Outcome (O: blood glucose, lipid profile, oxidative stress, inflammatory factors).In addition, Studies that were excluded from the analysis consisted of in vitro, in vivo, and ex vivo studies, case reports, quasi-experimental studies. Scopus, PubMed, Web of Science, EMBASE databases were searched until August 2023. Results: Of 496 records that were screened in the title/abstract stage, 9 studies were included in this meta-analysis. The sample sizes in the included studies vary between 28 and 90. The result of risk of bias was performed via risk of bias (RoB) in randomized-controlled trials (RCTs) based on the second version of the Cochrane RoB assessment tool. 8 studies had a definitely high risk of bias. Discussion: To the best of our knowledge, The present meta-analysis is the first study addressing the effectiveness of ALA supplementation in enhancing post-stroke metabolic markers, including lipid profile, oxidative stress, and inflammatory indices. It is imperative to acknowledge certain potential limitations inherent in this study. First of all, type of treatment (oral or intravenous infusion) could alter the bioavailability of ALA. Our study had restricted evidence regarding the impact of ALA supplementation on included outcomes. Therefore, further research is warranted to develop into the effects of ALA specifically on inflammation and oxidative stress. Funding: The research protocol was approved and supported by the Student Research Committee, Tabriz University of Medical Sciences (grant number: 72825). Registration: This study was registered in the International prospective register of systematic reviews (PROSPERO ID: CR42023461612).

Keywords: alpha-lipoic acid, lipid profile, blood glucose, inflammatory factors, oxidative stress, meta-analysis, post-stroke

Procedia PDF Downloads 36

Authors: Sudhir Kumar, V. R. Sinha

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Background: The topical and systemic toxicity associated with present nonmelanoma skin cancer (NMSC) treatment therapy using 5-Fluorouracil (5-FU) make it necessary to develop a novel delivery system having lesser toxicity and better control over drug release. Solid lipid nanoparticles offer many advantages like: controlled and localized release of entrapped actives, nontoxicity, and better tolerance. Aim:-To investigate safety and efficacy of 5-FU loaded solid lipid nanoparticles as a topical delivery system for the treatment of nonmelanoma skin cancer. Method: Topical solid lipid nanoparticles of 5-FU were prepared using Compritol 888 ATO (Glyceryl behenate) as lipid component and pluronic F68 (Poloxamer 188), Tween 80 (Polysorbate 80), Tyloxapol (4-(1,1,3,3-Tetramethylbutyl) phenol polymer with formaldehyde and oxirane) as surfactants. The SLNs were prepared with emulsification method. Different formulation parameters viz. type and ratio of surfactant, ratio of lipid and ratio of surfactant:lipid were investigated on particle size and drug entrapment efficiency. Results: Characterization of SLNs like–Transmission Electron Microscopy (TEM), Differential Scannig calorimetry (DSC), Fourier transform infrared spectroscopy (FTIR), Particle size determination, Polydispersity index, Entrapment efficiency, Drug loading, ex vivo skin permeation and skin retention studies, skin irritation and histopathology studies were performed. TEM results showed that shape of SLNs was spherical with size range 200-500nm. Higher encapsulation efficiency was obtained for batches having higher concentration of surfactant and lipid. It was found maximum 64.3% for SLN-6 batch with size of 400.1±9.22 nm and PDI 0.221±0.031. Optimized SLN batches and marketed 5-FU cream were compared for flux across rat skin and skin drug retention. The lesser flux and higher skin retention was obtained for SLN formulation in comparison to topical 5-FU cream, which ensures less systemic toxicity and better control of drug release across skin. Chronic skin irritation studies lacks serious erythema or inflammation and histopathology studies showed no significant change in physiology of epidermal layers of rat skin. So, these studies suggest that the optimized SLN formulation is efficient then marketed cream and safer for long term NMSC treatment regimens. Conclusion: Topical and systemic toxicity associated with long-term use of 5-FU, in the treatment of NMSC, can be minimized with its controlled release with significant drug retention with minimal flux across skin. The study may provide a better alternate for effective NMSC treatment.

Keywords: 5-FU, topical formulation, solid lipid nanoparticles, non melanoma skin cancer

Procedia PDF Downloads 488

Authors: Emma R. Arakelova, Stepan G. Grigoryan, Flora G. Arsenyan, Nelli S. Babayan, Ruzanna M. Grigoryan, Natalia K. Sarkisyan

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Novel nanosize zinc oxide composites of doxorubicin obtained by deposition of 180 nm thick zinc oxide film on the drug surface using DC-magnetron sputtering of a zinc target in the form of gels (PEO+Dox+ZnO and Starch+NaCMC+Dox+ZnO) were studied for drug delivery applications. The cancer specificity was revealed both in in vitro and in vivo models. The cytotoxicity of the test compounds was analyzed against human cancer (HeLa) and normal (MRC5) cell lines using MTT colorimetric cell viability assay. IC50 values were determined and compared to reveal the cancer specificity of the test samples. The mechanistic study of the most active compound was investigated using Flow cytometry analyzing of the DNA content after PI (propidium iodide) staining. Data were analyzed with Tree Star FlowJo software using cell cycle analysis Dean-Jett-Fox module. The in vivo anticancer activity estimation experiments were carried out on mice with inoculated ascitic Ehrlich’s carcinoma at intraperitoneal introduction of doxorubicin and its zinc oxide compositions. It was shown that the nanosize zinc oxide film deposition on the drug surface leads to the selective anticancer activity of composites at the cellular level with the range of selectivity index (SI) from 4 (Starch+NaCMC+Dox+ZnO) to 200 (PEO(gel)+Dox+ZnO) which is higher than that of free Dox (SI = 56). The significant increase in vivo antitumor activity (by a factor of 2-2.5) and decrease of general toxicity of zinc oxide compositions of doxorubicin in the form of the above mentioned gels compared to free doxorubicin were shown on the model of inoculated Ehrlich's ascitic carcinoma. Mechanistic studies of anticancer activity revealed the cytostatic effect based on the high level of DNA biosynthesis inhibition at considerable low concentrations of zinc oxide compositions of doxorubicin. The results of studies in vitro and in vivo behavior of PEO+Dox+ZnO and Starch+NaCMC+Dox+ZnO composites confirm the high potential of the nanosize zinc oxide composites as a vector delivery system for future application in cancer chemotherapy.

Keywords: anticancer activity, cancer specificity, doxorubicin, zinc oxide

Procedia PDF Downloads 384

Authors: Elif Tugce Aksun Tumerkan

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Fluorescent proteins (FPs) have become very popular since green fluorescent protein discovered from crystal jellyfish. It is known that Anthozoa species have a wide range of chromophore organisms, and the initial crystal structure for non-fluorescent chromophores obtained from the reef-building coral has been determined. There are also differently coloured pigments in non-bioluminescent Anthozoa zooxanthellate and azooxanthellate which are frequently members of the GFP-like protein family. The development of fluorescent proteins (FPs) and their applications is an outstanding example of basic science leading to practical biotechnological and medical applications. Fluorescent proteins have several applications in science and are used as important indicators in molecular biology and cell-based research. With rising interest in cell biology, FPs have used as biosensor indicators and probes in pharmacology and cell biology. Using fluorescent proteins in genetically encoded metabolite sensors has many advantages than chemical probes for metabolites such as easily introduced into any cell or organism in any sub-cellular localization and giving chance to fixing to fluoresce of different colours or characteristics. There are different factors effects to signalling mechanism when they used as a biosensor. While there are wide ranges of research have been done on the significance and applications of fluorescent proteins, the cell signalling response of FPs and target cell are less well understood. In this study, it was aimed to clarify the response of adaptive mechanisms of coral species such as pH, temperature and symbiotic relationship and target cells properties on the signalling capacity. Corals are a rich natural source of fluorescent proteins that change with environmental conditions such as light, heat stress and injury. Adaptation mechanism of coral species to these types of environmental variations is important factor due to FPs properties have affected by this mechanism. Since fluorescent proteins obtained from nature, their own ecological property like the symbiotic relationship is observed very commonly in coral species and living conditions have the impact on FPs efficiency. Target cell properties also have an effect on signalling and visualization. The dynamicity of detector that used for reading fluorescence and the level of background fluorescence are key parameters for the quality of the fluorescent signal. Among the factors, it can be concluded that coral species adaptive characteristics have the strongest effect on FPs signalling capacity.

Keywords: biosensor, cell biology, environmental conditions, fluorescent protein, sea anemone

Procedia PDF Downloads 148

Authors: Zeina Bou Diab, Arij Daou

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The neural mechanisms of sequential behaviors are intensively studied, with songbirds a focus for learned vocal production. We are studying the premotor nucleus HVC at a nexus of multiple pathways contributing to song learning and production. The HVC consists of multiple classes of neuronal populations, each has its own cellular, electrophysiological and functional properties. During singing, a large subset of motor cortex analog-projecting HVCRA neurons emit a single 6-10 ms burst of spikes at the same time during each rendition of song, a large subset of basal ganglia-projecting HVCX neurons fire 1 to 4 bursts that are similarly time locked to vocalizations, while HVCINT neurons fire tonically at average high frequency throughout song with prominent modulations whose timing in relation to song remains unresolved. This opens the opportunity to define models relating explicit HVC circuitry to how these neurons work cooperatively to control learning and singing. We developed conductance-based Hodgkin-Huxley models for the three classes of HVC neurons (based on the ion channels previously identified from in vitro recordings) and connected them in several physiologically realistic networks (based on the known synaptic connectivity and specific glutaminergic and gabaergic pharmacology) via different architecture patterning scenarios with the aim to replicate the in vivo firing patterning behaviors. We are able, through these networks, to reproduce the in vivo behavior of each class of HVC neurons, as shown by the experimental recordings. The different network architectures developed highlight different mechanisms that might be contributing to the propagation of sequential neural activity (continuous or punctate) in the HVC and to the distinctive firing patterns that each class exhibits during singing. Examples of such possible mechanisms include: 1) post-inhibitory rebound in HVCX and their population patterns during singing, 2) different subclasses of HVCINT interacting via inhibitory-inhibitory loops, 3) mono-synaptic HVCX to HVCRA excitatory connectivity, and 4) structured many-to-one inhibitory synapses from interneurons to projection neurons, and others. Replication is only a preliminary step that must be followed by model prediction and testing.

Keywords: computational modeling, neural networks, temporal neural sequences, ionic currents, songbird

Procedia PDF Downloads 43

Authors: Stephanie PB Caligiuri, Harold M Aukema, Delfin Rodriguez-Leyva, Amir Ravandi, Randy Guzman, Grant N. Pierce

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Background: Hypertension leads to cardiac and cerebral events and therefore is the leading risk factor attributed to death in the world. Oxylipins may be mediators in these events as they can regulate vascular tone and inflammation. Oxylipins are derived from fatty acids. Dietary flaxseed is rich in the n3 fatty acid, alpha-linolenic acid, and, therefore, may have the ability to change the substrate profile of oxylipins. As a result, this could alter blood pressure. Methods: A randomized, double-blinded, controlled clinical trial, the Flax-PAD trial, was used to assess the impact of dietary flaxseed on blood pressure (BP), and to also assess the relationship of plasma oxylipins to BP in 81 patients with peripheral arterial disease (PAD). Patients with PAD were chosen for the clinical trial as they are at an increased risk for hypertension and cardiac and cerebral events. Thirty grams of ground flaxseed were added to food products to consume on a daily basis for 6 months. The control food products contained wheat germ, wheat bran, and mixed dietary oils instead of flaxseed. Central BP, which is more significantly associated to organ damage, cardiac, and cerebral events versus brachial BP, was measured by pulse wave analysis at baseline and 6 months. A plasma profile of 43 oxylipins was generated using solid phase extraction, HPLC-MS/MS, and stable isotope dilution quantitation. Results: At baseline, the central BP (systolic/diastolic) in the placebo and flaxseed group were, 131/73 ± 2.5/1.4 mmHg and 128/71 ± 2.6/1.4 mmHg, respectively. After 6 months of intervention, the flaxseed group exhibited a decrease in blood pressure of 4.0/1.0 mmHg. The 6 month central BP in the placebo and flaxseed groups were, 132/74 ± 2.9/1.8 mmHg and 124/70 ± 2.6/1.6 mmHg (P<0.05). Correlation and logistic regression analyses between central blood pressure and oxylipins were performed. Significant associations were observed between central blood pressure and 17 oxylipins, primarily produced from arachidonic acid. Every 1 nM increase in 16-hydroxyeicosatetraenoic acid (HETE) increased the odds of having high central systolic BP by 15-fold, of having high central diastolic BP by 6-fold and of having high central mean arterial pressure by 15-fold. In addition, every 1 nM increase in 5,6-dihydroxyeicosatrienoic acid (DHET) and 11,12-DHET increased the odds of having high central mean arterial pressure by 45- and 18-fold, respectively. Flaxseed induced a significant decrease in these as well as 4 other vasoconstrictive oxylipins. Conclusion: Dietary flaxseed significantly lowered blood pressure in patients with PAD and hypertension. Plasma oxylipins were strongly associated with central blood pressure and may have mediated the flaxseed-induced decrease in blood pressure.

Keywords: hypertension, flaxseed, oxylipins, peripheral arterial disease

Procedia PDF Downloads 437

Authors: S. D. N. K. Bathige, G. I. Godahewa, Navaneethaiyer Umasuthan, Jehee Lee

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Kunitz-type serine protease inhibitors (KTIs) are identified in various organisms including animals, plants and microbes. These proteins shared single or multiple Kunitz inhibitory domains link together or associated with other types of domains. Characteristic Kunitz type domain composed of around 60 amino acid residues with six conserved cysteine residues to stabilize by three disulfide bridges. KTIs are involved in various physiological processes, such as ion channel blocking, blood coagulation, fibrinolysis and inflammation. In this study, two Kunitz-type domain containing protein was identified from rock bream database and designated as RbKunitz. The coding sequence of RbKunitz encoded for 507 amino acids with 56.2 kDa theoretical molecular mass and 5.7 isoelectric point (pI). There are several functional domains including MANEC superfamily domain, PKD superfamily domain, and LDLa domain were predicted in addition to the two characteristic Kunitz domain. Moreover, trypsin interaction sites were also identified in Kunitz domain. Homology analysis revealed that RbKunitz shared highest identity (77.6%) with Takifugu rubripes. Completely conserved 28 cysteine residues were recognized, when comparison of RbKunitz with other orthologs from different taxonomical groups. These structural evidences indicate the rigidity of RbKunitz folding structure to achieve the proper function. The phylogenetic tree was constructed using neighbor-joining method and exhibited that the KTIs from fish and non-fish has been evolved in separately. Rock bream was clustered with Takifugu rubripes. The SYBR Green qPCR was performed to quantify the RbKunitz transcripts in different tissues and challenged tissues. The mRNA transcripts of RbKunitz were detected in all tissues (muscle, spleen, head kidney, blood, heart, skin, liver, intestine, kidney and gills) analyzed and highest transcripts level was detected in gill tissues. Temporal transcription profile of RbKunitz in rock bream blood tissues was analyzed upon LPS (lipopolysaccharide), Poly I:C (Polyinosinic:polycytidylic acid) and Edwardsiella tarda challenge to understand the immune responses of this gene. Compare to the unchallenged control RbKunitz exhibited strong up-regulation at 24 h post injection (p.i.) after LPS and E. tarda injection. Comparatively robust expression of RbKunits was observed at 3 h p.i. upon Poly I:C challenge. Taken together all these data indicate that RbKunitz may involve into to immune responses upon pathogenic stress, in order to protect the rock bream.

Keywords: Kunitz-type, rock bream, immune response, serine protease inhibitor

Procedia PDF Downloads 349

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

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

Keywords: autophagy, endocytosis, glioma, WNK2

Procedia PDF Downloads 346

Authors: Tayyaba Bari, Muhammad Hamza Anjum, Samra Kanwal, Fakhera Ikram

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Osteoarthritis, a degenerative condition, affects more than 213 million individuals globally. Since articular cartilage has no or limited vessels, therefore, after deteriorating, it is unable to rejuvenate. Traditional approaches for cartilage repair, like autologous chondrocyte implantation, microfracture and cartilage transplantation are often associated with postoperative complications and lead to further degradation. Decellularized human umbilical cord has gained interest as a viable treatment for cartilage repair. Decellularization removes all cellular contents as well as debris, leaving a biologically active 3D network known as extracellular matrix (ECM). This matrix is biodegradable, non-immunogenic and provides a microenvironment for homeostasis, growth and repair. UC derived bioink function as 3D scaffolding material, not only mediates cell-matrix interactions but also adherence, proliferation and propagation of cells for 3D organoids. This study comprises different physical, chemical and biological approaches to optimize the decellularization of human umbilical cord (UC) tissues followed by the solubilization of these tissues to bioink formation. The decellularization process consisted of two cycles of freeze thaw where the umbilical cord at -20˚C was thawed at room temperature followed by dissection in small sections from 0.5 to 1cm. Similarly decellularization with ionic and non-ionic detergents Sodium dodecyl sulfate (SDS) and Triton-X 100 revealed that both concentrations of SDS i.e 0.1% and 1% were effective in complete removal of cells from the small UC tissues. The results of decellularization was further confirmed by running them on 1% agarose gel. Histological analysis revealed the efficacy of decellularization, which involves paraffin embedded samples of 4μm processed for Hematoxylin-eosin-safran and 4,6-diamidino-2-phenylindole (DAPI). ECM preservation was confirmed by Alcian Blue, and Masson’s trichrome staining on consecutive sections and images were obtained. Sulfated GAG’s content were determined by 1,9-dimethyl-methylene blue (DMMB) assay, similarly collagen quantification was done by hydroxy proline assay. This 3D bioengineered scaffold will provide a typical atmosphere as in the extracellular matrix of the tissue, which would be seeded with the mesenchymal cells to generate the desired 3D ink for in vitro and in vivo cartilage regeneration applications.

Keywords: umbilical cord, 3d printing, bioink, tissue engineering, cartilage regeneration

Procedia PDF Downloads 61

Authors: Jasmeet Kaur, Anup Singh, Shashikant Iyengar, Arun Kumar, Ira Sahay

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Latent autoimmune diabetes in adults (LADA) is an irreversible autoimmune disease that affects insulin production. LADA is characterized by the production of Glutamic acid decarboxylase (GAD) antibodies, which is similar to type 1 diabetes. Individuals with LADA may eventually develop overt diabetes and require insulin. In this condition, the pancreas produces little or no insulin, which is a hormone used by the body to allow glucose to enter cells and produce energy. While type 1 diabetes was traditionally associated with children and teenagers, its prevalence has increased in adults as well. LADA is frequently misdiagnosed as type 2 diabetes, especially in adulthood when type 2 diabetes is more common. LADA develops in adulthood, usually after age 30. Managing LADA involves metabolic control with exogenous insulin and prolonging the life of surviving beta cells, thereby slowing the disease's progression. This case study examines the impact of approximately 3 months of low-carbohydrate dietary intervention in a 42-year-old woman with LADA who was initially misdiagnosed as having type 2 diabetes. Her c-peptide was 0.13 and her HbA1c was 9.3% when this trial began. Low-carbohydrate interventions have been shown to improve blood sugar levels, including fasting, post-meal, and random blood sugar levels, as well as haemoglobin levels, blood pressure, energy levels, sleep quality, and satiety levels. The use of low-carbohydrate dietary intervention significantly reduces both hypo- and hyperglycaemia events. During the 3 months of the study, there were 2 to 3 hyperglycaemic events owing to physical stress and a single hypoglycaemic event. Low-carbohydrate dietary therapies lessen insulin dose inaccuracy, which explains why there were fewer hyperglycaemic and hypoglycaemic events. In three months, the glycated haemoglobin (HbA1c) level was reduced from 9.3% to 6.3%. These improvements occur without the need for caloric restriction or physical activity. Stress management was crucial aspect of the treatment plan as stress-induced neuroendocrine hormones can cause immunological dysregulation. Additionally, supplements that support immune system and reduce inflammation were used as part of the treatment during the trial. Long-term studies are needed to track disease development and corroborate the claim that such dietary treatments can prolong the honeymoon phase in LADA. Various factors can contribute to additional autoimmune attacks, so measuring c-peptide is crucial on a regular basis to determine whether insulin levels need to be adjusted.

Keywords: autoimmune, diabetes, LADA, low_carb, nutrition

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Authors: Avijit Dey, Antony Gomes, Subir Chandra Dasgupta

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Tea (Camellia sinensis) is the most popular beverages in the world and is ranked second after the water. Tea has been considered as a health promoting beverage since ancient times due to its health-promoting activity. Recently, immunomodulatory, anti-arthritic, antioxidant, anticancer and cardioprotective activity of tea has been established. Very few studies have demonstrated the effect of high dose of black tea on health. The aim of the present study was to evaluate the role of low & high dose of Black Tea Extract (BTE) on the different physiological parameters of mother and pups during prenatal and postnatal developmental period in the experimental rodent. BTE was orally administered in LD (50mg BTE/kg/day) and HD (100mg BTE/kg/day) except control groups of rats (n=6/group) throughout the prenatal (day 0-21) and postnatal (day 21-42) periods. During prenatal period (0, 7th, 14th, 20th days) body weight, urinary calcium, magnesium, urea and creatinine was measured. In postnatal period physical (0, 10th, 21th days) parameters of pups like body weight, cranial length, cranial diameter, neck width, tail length, craniosacral length of pups were analyzed. Liver and lungs from pups and kidney spleen, etc. from mothers were collected on day 42 for histopathological studies. The comparative urine strip and morphology of RBC was also analyzed by SEM from mothers of different groups on day 42. The level of cytokines like IL-1alpha, IL-1beta, IL-6, IL-10, TNF-alpha were analysed by enzyme-linked immunosorbent assay (ELISA) on day 0, day 20 and day 42. The body weight of LD and HD mothers were also significantly (P<0.05) less than control mothers at 20th day of pregnancy and there was also significant changes in urinary calcium, urea, creatinine. The bio morphometric analysis of pups showed significant alteration (P<0.05) in HD groups relative to control. Some histological alterations were also observed in pups and mothers. Comparative urine strip analysis and morphology of RBC showed significant changes in treated groups. LD and HD treated mothers showed an increase in proinflammatory cytokines like IL-1beta, TNF-alpha and decrease in anti-inflammatory cytokine-like IL-10 on day 20 compared to PC mothers. This study clearly indicated that high dose of BTE possesses detrimental effect on pregnant mother and the pup. Further studies are in progress to elucidate the molecular mechanism of actions. This project work has been sponsored by National Tea Research Foundation vide Project Sanction No.: 17 (305)/2013/4423 dated 11th March, 2014. All experimental protocols described in the study were approved by animal ethics committee.

Keywords: black tea extract, pregnancy, prenatal and postnatal development, inflammation

Procedia PDF Downloads 252

Authors: E. M. Condori-Peñaloza, S. S. Costa

Abstract:

Eleusine indica L, known as goose-grass, is considered a troublesome weed that can cause important economic losses in the agriculture worldwide. However, this grass is used as a medicinal plant in some regions of Brazil to treat influenza and pneumonia. In Africa and Asia, it is used to treat malaria and as diuretic, anti-helminthic, among other uses. Despite its therapeutic potential, little is known about the chemical composition and bioactive compounds of E. indica. Hitherto, two major flavonoids, schaftoside and vitexin, were isolated from aerial part of the species and showed inhibitory activity on lung neutrophil influxes in mice, suggesting a beneficial effect on airway inflammation. Therefore, the aim of this study was to analyze the chemical profile of aqueous extracts from aerial parts of Eleusine indica specimens using high performance liquid chromatography (HPLC-DAD) and 1H nuclear magnetic resonance spectroscopy (NMR), with emphasis on phenolic compounds. Specimens of E. indica were collected in Minas Gerais state, Brazil. Aerial parts of fresh plants were extracted by decoction (10% p/v). After spontaneous precipitation of the aqueous extract at 10-12°C for 24 hours, the supernatant obtained was frozen and lyophilized. After that, 1 g of the extract was dissolved into 25 mL of water and fractionated on a reverse phase chromatography column (RP-2), eluted with a gradient of H2O/EtOH. Five fractions were obtained. The extract and fractions had their chemical profile analyzed by using HPLC-DAD (C-18 column: 20 μL, 256 -365 nm; gradient water 0.01% phosphoric acid/ acetonitrile. The extract was also analyzed by NMR (1H, 500 MHz, D2O) in order to access its global chemical composition. HPLC-DAD analyses of crude extract allowed the identification of ten phenolic compounds. Fraction 1, eluted with 100% water, was poor in phenolic compounds and no major peak was detected. In fraction 2, eluted with 100% water, it was possible to observe one major peak at retention time (RT) of 23.75 minutes compatible with flavonoid; fraction 3, also eluted with 100% water, showed four peaks at RT= 21.47, 23.52, 24.33 and 25.84 minutes, all of them compatible with flavonoid. In fraction 4, eluted with 50%/ethanol/50% water, it was possible to observe 3 peaks compatible with flavonoids at RT=24.65, 26.81, 27.49 minutes, and one peak (28.83 min) compatible with a phenolic acid derivative. Finally, in fraction 5, eluted with 100% ethanol, no phenolic substance was detected. The UV spectra of all flavonoids detected were compatible with the flavone subclass (λ= 320-345 nm). The 1H NMR spectra of aerial parts extract showed signals in three regions: δ 0.8-3.0 ppm (aliphatic compounds), δ 3.0-5.5 ppm corresponding to carbohydrates (signals most abundant and overlapped), and δ 6.0-8.5 ppm (aromatic compounds). Signals compatible with flavonoids (rings A and B) could also be detected in the crude extract spectra. These results suggest the presence of several flavonoids in E. indica, which reinforces their therapeutic potential. The pharmacological activities of Eleusine indica extracts and fractions will be further evaluated.

Keywords: flavonoids, HPLC, NMR, phenolic compounds

Procedia PDF Downloads 288

Authors: Tiffanie-Marie Borg, Yasmin Zeinolabediny, Nima Heidari, Ali Noorani, Mark Slevin, Angel Cullen, Stefano Olgiati, Alberto Zerbi, Alessandro Danovi, Adrian Wilson

Abstract:

Knee Osteoarthritis (OA) is a critical cause of disability globally. In recent years, there has been growing interest in non-invasive treatments, such as intra-articular injection of micro-fragmented fat (MFAT), showing great potential in treating OA. Mesenchymal stem cells (MSCs), originating from pericytes of micro-vessels in MFAT, can differentiate into mesenchymal lineage cells such as cartilage, osteocytes, adipocytes, and osteoblasts. Secretion of growth factor and cytokines from MSCs have the capability to inhibit T cell growth, reduced pain and inflammation, and create a micro-environment that through paracrine signaling, can promote joint repair and cartilage regeneration. Here we have shown, for the first time, data supporting the hypothesis that women respond better in terms of improvements in pain and function to MFAT injection compared to men. Historically, women have been underrepresented in studies, and studies with both sexes regularly fail to analyse the results by sex. To mitigate this bias and quantify it, we describe a technique using reproducible statistical analysis and replicable results with Open Access statistical software R to calculate the magnitude of this difference. Genetic, hormonal, environmental, and age factors play a role in our observed difference between the sexes. This observational, intention-to-treat study included the complete sample of 456 patients who agreed to be scored for pain (visual analogue scale (VAS)) and function (Oxford knee score (OKS)) at baseline regardless of subsequent changes to adherence or status during follow-up. We report that a significantly larger number of women responded to treatment than men: [90% vs. 60% change in VAS scores with 87% vs. 65% change in OKS scores, respectively]. Women overall had a stronger positive response to treatment with reduced pain and improved mobility and function. Pre-injection, our cohort of women were in more pain with worse joint function which is quite common to see in orthopaedics. However, during the 2-year follow-up, they consistently maintained a lower incidence of discomfort with superior joint function. This data clearly identifies a clear need for further studies to identify the cell and molecular biological and other basis for these differences and be able to utilize this information for stratification in order to improve outcome for both women and men.

Keywords: gender differences, micro-fragmented adipose tissue, knee osteoarthritis, stem cells

Procedia PDF Downloads 168