Search results for: protein complementation assay

Authors: Latife Merve Oktay, Berrin Tugrul

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Hydroxytyrosol (HT) is a phenolic phytochemical molecule derived from the hydrolysis of oleuropein, which originates during the maturation of the olives. It has recently received particular attention because of its antioxidant, anti-proliferative, pro-apoptotic and anti-inflammatory activities. In this study, we investigated the cytotoxic and apoptotic effects of hydroxytyrosol and its effects on phosphatidylinositol 3-kinase/Akt (PI3K/Akt) and extracellular signal-regulated kinase 1/2 (ERK 1/2) signaling pathways in human ovarian cancer cell lines OVCAR-3 and MDAH-2774. XTT cell proliferation kit, Cell Death Detection Elisa Plus Kit (Roche) and Human Apoptosis Array (R&D Systems) were used to determine the cytotoxic and apoptotic effects of HT in OVCAR-3 and MDAH-2774 cell lines at 24, 48, 72, and 96 h. Effect of HT on PI3K/Akt and ERK 1/2 signaling pathways were investigated by using specific inhibitors of these pathways. IC50 values of HT were found to be 102.3 µM in MDAH-2774 cells at 72 h and 51.5 µM in OVCAR-3 cells at 96 h. Apoptotic effect of HT in MDAH-2774 cells was the highest at 50 µM at 72 h, and kept decreasing at 100 and 150 µM concentrations and was not seen at 200 µM and higher concentrations. Highest apoptotic effect was seen at 100 µM concentration in OVCAR-3 cells at 96 h, however apoptotic effect was decreased over 100 µM concentrations. According to antibody microarray results, HT increased the levels of pro-apoptotic molecules Bad, Bax, active caspase-3, Htra2/Omi by 2.0-, 1.4-, 1.2-, 4.2-fold, respectively and also increased the levels of pro-apoptotic death receptors TRAIL R1/DR4, TRAIL R2/DR5, FAS/TNFRSF6 by 2.1-, 1.7-, 1.6-fold, respectively, however, it decreased the level of Survivin by 1.6-fold which is one of the inhibitor of apoptosis protein (IAP) family in MDAH-2774 cells. In OVCAR-3 cells, HT decreased the levels of anti-apoptotic proteins Bcl-2, pro-caspase 3 by 3.1-, 8.2-fold, respectively and IAP family proteins CIAP-1, CIAP-2, XIAP, Livin, Survivin by 6.5-, 6.0-, 3.2-, 2.2-, 2.7-fold, respectively and increased the level of cytochrome-c by 1.2-fold. We have shown that HT shows its cytotoxic and apoptotic effect through inhibiting ERK 1/2 signaling pathway in both OVCAR-3 and MDAH-2774 cells. Further studies are needed to investigate molecular mechanisms and modulatory effects of hydroxytyrosol.

Keywords: apoptosis, cytotoxicity, hydroxytyrosol, ovarian cancer

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Authors: Mustafa M. Donma, Orkide Donma

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Spexin, expressed in central nervous system, has attracted much interest in feeding behavior, obesity, diabetes, energy metabolism and cardiovascular functions. Fetuin A is known as negative acute phase reactant synthesized in the liver. So far, it has become a major concern of many studies in numerous clinical states. The relationship between the concentrations of spexin as well as fetuin A and the risk for cardiovascular diseases (CVDs) were also investigated. Eosinophils, suggested to be associated with the development of CVDs, are introduced as early indicators of cardiometabolic complications. Patients with elevated platelet count, associated with hypercoagulable state in the body, are also more liable to CVDs. In this study, the aim is to examine the profiles of spexin and fetuin A concomitant with the course of variations detected in eosinophil as well as platelet counts in morbid obese children. Thirty-four children with normal-body mass index (N-BMI) and fifty-one morbid obese (MO) children participated in the study. Written-informed consent forms were obtained prior to the study. Institutional ethics committee approved the study protocol. Age- and sex-adjusted BMI percentile tables prepared by World Health Organization were used to classify healthy and obese children. Mean age ± SEM of the children were 9.3 ± 0.6 years and 10.7 ± 0.5 years in N-BMI and MO groups, respectively. Anthropometric measurements of the children were taken. Body mass index values were calculated from weight and height values. Blood samples were obtained after an overnight fasting. Routine hematologic and biochemical tests were performed. Within this context, fasting blood glucose (FBG), insulin (INS), triglycerides (TRG), high density lipoprotein-cholesterol (HDL-C) concentrations were measured. Homeostatic model assessment for insulin resistance (HOMA-IR) values were calculated. Spexin and fetuin A levels were determined by enzyme-linked immunosorbent assay. Data were evaluated from the statistical point of view. Statistically significant differences were found between groups in terms of BMI, fat mass index, INS, HOMA-IR and HDL-C. In MO group, all parameters increased as HDL-C decreased. Elevated concentrations in MO group were detected in eosinophils (p<0.05) and platelets (p>0.05). Fetuin A levels decreased in MO group (p>0.05). However, decrease was statistically significant in spexin levels for this group (p<0.05). In conclusion, these results have suggested that increases in eosinophils and platelets exhibit behavior as cardiovascular risk factors. Decreased fetuin A behaved as a risk factor suitable to increased risk for cardiovascular problems associated with the severity of obesity. Along with increased eosinophils, increased platelets and decreased fetuin A, decreased spexin was the parameter, which reflects best its possible participation in the early development of CVD risk in MO children.

Keywords: cardiovascular diseases , eosinophils , fetuin A , pediatric morbid obesity , platelets , spexin

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Authors: Belinda Vega, Claudio Alvarez, Héctor Flores, Marcia Oliva, Katherine Alveal, Teresa Toro, María José Tapia, Fanny Guzmán

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With the increase in global temperatures and the decrease of oxygen (O2) concentration in the oceans, fish cultures are exposed to frequent fluctuations in dissolved O2 (DO) concentration that can cause chronic stress in the animals, altering the normal functioning of their immune system and making them vulnerable to infections, consequently increasing morbidity and mortality in the farms with economic losses. The mucosal organs (skin -and mucus-, gills, gut, and nasal mucosa) are the first line of defense of the fish against pathogens. Therefore, the objective of this study is to evaluate the effect of hypoxia on the antimicrobial activity of epidermal mucus from corvina drum (Cilus Gilberti), a native marine species with the potential for the diversification of aquaculture in Chile. To achieve this, the epidermal mucus of juveniles (~220g) kept under normoxia (7 mg/L DO) and hypoxia (2 mg/L DO) environmental conditions was collected after 6 weeks, as well as after 6 days of intraperitoneal inoculation with lipopolysaccharide from Vibrio anguillarum to induce an immune response in the fish. Total protein extracts of the mucus were used for bactericidal activity and lysozyme and peroxidase activity assays. Although the mucus from both experimental groups showed inhibitory effects on the bacterial growth of Vibrio anguillarum and Vibrio ordalli, this effect was more long-lasting in the normoxia group. We also observed a notable reduction in the presence of lysozyme in the mucus from fish exposed to hypoxia, with no differences in peroxidase content. Future proteomic studies of corvina mucus associated with the environmental conditions studied in this work will allow the isolation and identification of peptides with antimicrobial activity, those responsible for the results obtained. This will help establish strategies aimed at minimizing the impacts of hypoxia on the defense responses of corvina drum against potential pathogens. Funding: FONDECYT 3200440 and FONDECYT 1210056

Keywords: Cilus gilberti, mucus, antimicrobial activity, HYPOXIA

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Authors: Christen Kutz

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Objective: The aim of this study is to review a case series of 4 high-level Olympic and Division 1 wrestlers who experienced debilitating posterior circulation ischemic strokes during or after a competitive wrestling event and to identify risk factors, etiology and outcomes of stroke in young, healthy elite wrestlers. Background: Stroke occurs in one in 10,000 people under age 64. In young adults, the most common causes of stroke are cardiac embolism, hypercoagulable state, and vasculopathy. One-third of these strokes occur in young, fit individuals. There is little published literature about ischemic strokes that occur in wrestlers. Based on the nature of wrestling, the risk of injury or dissection to neurovascular structures may be a possible theory, but very few case reports exist. Methodology: 4 wrestlers under the age of 44 with a known history of ischemic stroke participated in individual interviews either in person or virtually. Each of the wrestlers provided their demographic information, wrestling background, clinical presentation at the time of stroke, imaging results, identification of potential risk factors, acute treatment and recovery. Results: 3 white male Division 1 wrestlers (2 Lehigh University, 1 Lock Haven University) and 1 black male 2008 Olympian experienced posterior circulation strokes. Case #1 felt a “pop” while wrestling (lateral medullary infarct, possible vertebral artery dissection); Case #2 awoke with severe vertigo, sweating, and vomiting after wrestling the previous day (left cerebellar infarct, (+) protein S deficiency); Case #3 severe vertigo, ataxia, and sensation of impending doom after wrestling earlier that week (left cerebellar infarct, hypoplastic left vertebral artery (+) anti-cardiolipin antibodies). Case #4 severe dizziness, confusion (left cerebellar stroke, vertebral artery dissection, small PFO). Conclusion: 3 wrestlers were started on anti-platelet therapy, risk factors were modified, and returned to their sport. 1 wrestler was placed on anti-coagulation and retired from competition.

Keywords: stroke, wrestling, Olympic, posterior circulation

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Authors: Maryam Farzaneh, Seyyedeh Nafiseh Hassani, Hossein Baharvand

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Objective: Chicken gonadal tissue has a two population such primordial germ cells (PGCs) and stromal cells (somatic cells). PGCs and embryonic germ cells (EGCs) that is a pluripotent type of PGCs in long-term culture are suitable sources for the production of chicken pluripotent stem cell lines, transgenic birds, vaccine and recombinant protein production. In general, the effect of growth factors such bFGF and mouse LIF on derivation of PGCs in vitro are important and in this study we could see the unique effect of small molecules such PD032 and SB43 as a chemical, in comparison to growth factors. Materials and Methods: After incubation of fertilized chicken egg up to 6 days and isolation of primary gonadal tissues and culture of mixed cells like PGCs and stromal cells. PGCs proliferate in the present of fetal calf serum (FCS) and small molecules and in another group bFGF, that these factors are important for PGCs culture and derivation. Somatic cells produce a multilayer feeder under the PGCs in primary culture and PGCs make a small cluster under these cells. Results: In present of small molecules and high volume of FCS (15%), the present of EGCs as a pluripotent stem cells were clear four weeks, that they had a positive immune-staining and periodic acid-Schiff staining (PAS), but in present of growth factors like bFGF without any chemicals, the present of PGCs were clear but after 7 until 10 days, there were disappear. Conclusion: Until now we have seen many researches about derivation and maintenance of chicken PGCs, in the hope of understanding the mechanisms that occur during germline development and production of a therapeutic product by transgenic birds. There are still many unknowns in this area and this project will try to have efficient conditions for identification of suitable culture medium for long-term culture of PGCs in vitro without serum and feeder cells.

Keywords: chicken gonadal primordial germ cells, pluripotent stem cells, growth factors, small molecules, transgenic birds

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Authors: Yang Yang, Luciana Magalhães Rebelo Alencar, Martha Sahylí Ortega Pijeira, Beatriz da Silva Batista, Alefe Roger Silva França, Erick Rafael Dias Rates, Ruana Cardoso Lima, Sara Gemini-Piperni, Ralph Santos-Oliveira

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Radium-²²³ dichloride ([²²³Rₐ]RₐCl₂) is an alpha particle-emitting radiopharmaceutical currently approved for the treatment of patients with castration-resistant prostate cancer, symptomatic bone metastases, and no known visceral metastatic disease. [²²³Rₐ]RₐCl₂ is bone-seeking calcium mimetic that bonds into the newly formed bone stroma, especially osteoblastic or sclerotic metastases, killing the tumor cells by inducing DNA breaks in a potent and localized manner. Nonetheless, the successful therapy of osteosarcoma as primary bone tumors is still a challenge. Nanomicelles are colloidal nanosystems widely used in drug development to improve blood circulation time, bioavailability, and specificity of therapeutic agents, among other applications. In addition, the enhanced permeability and retention effect of the nanosystems, and the renal excretion of the nanomicelles reported in most cases so far, are very attractive to achieve selective and increased accumulation in tumor site as well as to increase the safety of [²²³Rₐ]RₐCl₂ in the clinical routine. In the present work, [²²³Rₐ]RₐCl₂ nanomicelles were produced, characterized, in vitro evaluated, and compared with pure [²²³Rₐ]RₐCl2 solution using SAOS2 osteosarcoma cells. The [²²³Rₐ]RₐCl₂ nanomicelles were prepared using the amphiphilic copolymer Pluronic F127. The dynamic light scattering analysis of freshly produced [²²³Rₐ]RₐCl₂ nanomicelles demonstrated a mean size of 129.4 nm with a polydispersity index (PDI) of 0.303. After one week stored in the refrigerator, the mean size of the [²²³Rₐ]RₐCl₂ nanomicelles increased to 169.4 with a PDI of 0.381. Atomic force microscopy analysis of [223Rₐ]RₐCl₂ nanomicelles exhibited spherical structures whose heights reach 1 µm, suggesting the filling of 127-Pluronic nanomicelles with [²²³Rₐ]RₐCl₂. The viability assay with [²²³Rₐ]RₐCl₂ nanomicelles displayed a dose-dependent response as it was observed using pure [²²³Rₐ]RₐCl2. However, at the same dose, [²²³Rₐ]RₐCl₂ nanomicelles were 20% higher efficient in killing SAOS2 cells when compared with pure [²²³Rₐ]RₐCl₂. These findings demonstrated the effectiveness of the nanosystem validating the application of nanotechnology in targeted alpha therapy with [²²³Ra]RₐCl₂. In addition, the [²²³Rₐ]RaCl₂nanomicelles may be decorated and incorporated with a great variety of agents and compounds (e.g., monoclonal antibodies, aptamers, peptides) to overcome the limited use of [²²³Ra]RₐCl₂.

Keywords: nanomicelles, osteosarcoma, radium dichloride, targeted alpha therapy

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Authors: G. S. Sumanasekara, A. Balasuriya

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Introduction: In Sri Lanka studies reveal higher trend in prevalence of diabetes. The office employees have sedentary life style and their eating patterns changed due to nutritional transition. Further overall, urban and rural pre diabetes is also increasing. Objectives - Study the general food pattern of office employees and its relation to overweight/obesity and prevalence of diabetes among them. Method: The data was collected from office employees between 30-60 years (n-400).Data analyzed using SPSS 16 version.The Study design was a descriptive cross sectional study. The study setting was Ministry of Economic Development. Anthropometric measurements and blood glucose assessed by trained nurses. Dietary pattern was studied through a food frequency questionairre thereby calculated daily nutrient intakes. Results: Mean age of office employees were 38.98 SD (7.033) CI=95%) and 245 females (61.2%) 155 males (38.8 %) ,Nationality includes Sinhala (67.5%), Tamil(20%), and Muslims (12.5%).Owerweight(7,1.8%), obese male(36,9%), obese female(66,16%)/ diabetes/obese(18,4.5%) out of 127(31.8%) who were above the normal BMI whereas 273(68.2) were within the normal. Mean BMI was 24.1593.Mean Blood sugar level was 104.646,SD(16.018).12% consume tobacco products,17.8 consumed alcohol.15.8% had nutrition training. Two main dietary patterns identified who were vegetarians and non vegetarians .Mean energy intake 1727.1, (SD 4.97), Mean protein consumption(11.33, SD 1.811), Mean fat consumption(24.07, SD 4.131),Mean CHO consumption (64.56, SD 4.54), Mean Fibre (30.05, SD 17.9), Mean cholesterol(16.85, SD 17.22), Energy intake was higher in non vegetarians and larger propotion of energy derived from proteins , and fat. Their carbohydrate and cholesterol intake was also higher. Tamils were mostly vegetarians. Mainly BMI were within normal range(18.5-23.5) whereas Muslims who had higher energy intakes showed BMI above the normal. Conclusion: Two distinct dietary patterns identified. Different ethnic groups consume different diets with different nutrient composition. Dietary pattern has a relation to overweight. Overweight related to high blood glucose levels but some overweight subjects do not show any relation.

Keywords: obesity, overweight, diabetes, dietary pattern, nutrition, BMI, non communicable disease

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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: Kanokwan Sansuwan, Orapint Jintasataporn, Srinoy Chumkam

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Asian seabass is one of the economically important fish of Thailand and other countries in the Southeast Asia. Zinc is an essential trace metal to fish and vital to various biological processes and function. It is required for normal growth and indispensable in the diet. Therefore, the artificial diets offered to intensively cultivated fish must possess the zinc content required by the animal metabolism for health maintenance and high weight gain rates. However, essential elements must also be in an available form to be utilized by the organism. Thus, this study was designed to evaluate the application of different zinc forms, including organic Zinc (zinc amino acid complex) and inorganic Zinc (zinc sulfate), as feed additives in diets for Asian seabass. Three groups with five replicates of fish (mean weight 22.54 ± 0.80 g) were given a basal diet either unsupplemented (control) or supplemented with 50 mg Zn kg−¹ sulfate (ZnSO₄) or Zinc Amino Acid Complex (ZnAA) respectively. Feeding regimen was initially set at 3% of body weight per day, and then the feed amount was adjusted weekly according to the actual feeding performance. The experiment was conducted for 10 weeks. Fish supplemented with ZnAA had the highest values in all studied growth indicators (weight gain, average daily growth and specific growth rate), followed by fish fed the diets with the ZnSO₄, and lowest in fish fed the diets with the control. Lysozyme and superoxide dismutase (SOD) activity of fish supplemented with ZnAA were significantly higher compared with all other groups (P < 0.05). Fish supplemented with ZnSO₄ exhibited significant increase in digestive enzyme activities (protease, pepsin and trypsin) compared with ZnAA and the control (P < 0.05). However, no significant differences were observed for RNA and protein in muscle (P > 0.05). The results of the present work suggest that ZnAA are a better source of trace elements for Asian seabass, based on growth performance and immunity indices examined in this study.

Keywords: Asian seabass, growth performance, zinc amino acid complex (ZnAA), zinc sulfate (ZnSO₄)

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Authors: Marwa Elkalla, E. Ali Abdelfadil, Ali. Mohamed. M. Sami, Ali M. Abdel-Monem

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To assess the impact of the blood collection technique on clinical pathology markers and to establish reference intervals, a study was performed using normal, healthy C57BL/6 mice. Both sexes were employed, and they were randomly assigned to different groups depending on the phlebotomy technique used. The blood was drawn in one of four ways: intracardiac (IC), caudal vena cava (VC), caudal vena cava (VC) plus a peritoneal collection of any extravasated blood, or retroorbital phlebotomy (RO). Several serum biochemistries, such as a liver function test, a complete blood count with differentials, and a platelet count, were analysed from the blood and serum samples analysed. Red blood cell count, haemoglobin (p >0.002), hematocrit, alkaline phosphatase, albumin, total protein, and creatinine were all significantly greater in female mice. Platelet counts, specific white blood cell numbers (total, neutrophil, lymphocyte, and eosinophil counts), globulin, amylase, and the BUN/creatinine ratio were all greater in males. The VC approach seemed marginally superior to the IC approach for the characteristics under consideration and was linked to the least variation among both sexes. Transaminase levels showed the greatest variation between study groups. The aspartate aminotransferase (AST) values were linked with decreased fluctuation for the VC approach, but the alanine aminotransferase (ALT) values were similar between the IC and VC groups. There was a lot of diversity and range in transaminase levels between the MC and RO groups. We found that the RO approach, the only one tested that allowed for repeated sample collection, yielded acceptable ALT readings. The findings show that the test results are significantly affected by the phlebotomy technique and that the VC or IC techniques provide the most reliable data. When organising a study and comparing data to reference ranges, the ranges supplied here by collection method and sex can be utilised to determine the best approach to data collection. The authors suggest establishing norms based on the procedures used by each individual researcher in his or her own lab.

Keywords: clinical, pathology, blood, effect

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Authors: Muhammad Dildar Gogi, Muhammad Jalal Arif, Muhammad Junaid Nisar, Mubashir Iqbal, Waleed Afzal Naveed, Muhammad Ahsan Khan, Ahmad Nawaz, Muhammad Sufian, Muhammad Arshad, Amna Jalal

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Fruit flies of Bactrocera genus cause heavy losses in fruits and vegetables globally and insecticide-application for their control creates issues of ecological backlash, environmental pollution, and food safety. There is need to explore alternatives and food-baits application is considered safe for the environment and effective for fruit fly management. Present experiment was carried out to assess the attractancy of five phagostimulant-Mixtures (PHS-Mix) prepared by mixing banana-squash, mulberry, protein-hydrolysate and molasses with some phagostimulant-lure sources including beef extract, fish extract, yeast, starch, rose oil, casein and cedar oil in five different ratios i.e., PHS-Mix-1 (1 part of all ingredients), PHS-Mix-2 (1 part of banana with 0.75 parts of all other ingredients), PHS-Mix-3 (1 part of banana with 0.5 parts of all other ingredients), PHS-Mix-4 (1 part of banana with 0.25 parts of all other ingredients) and PHS-Mix-5 (1 part of banana with 0.125 parts of all other ingredients). These were evaluated in comparison with a standard (GF-120). PHS-Mix-4 demonstrated 40.5±1.3-46.2±1.6% AI for satiated flies (class-II i.e., moderately attractive) and 59.5±2.0-68.6±3.0% AI for starved flies (class-III i.e., highly attractive) for both B. dorsalis and B. zonata in olfactometric study while the same exhibited 51.2±0.53% AI (class-III i.e., highly attractive) for B. zonata and 45.4±0.89% AI (class-II i.e., moderately attractive) for B. dorsalis in field study. PHS-Mix-1 proved non-attractive (class-I) and moderately attractive (class-II) phagostimulant in olfactometer and field studies, respectively. PHS-Mix-2 exhibited moderate attractiveness for starved lots in olfactometer and field-lot in field studies. PHS-Mix-5 proved non-attractive to starved and satiated lots of B. zonata and B. dorsalis females in olfactometer and field studies. Overall PHS-Mix-4 proved better phagostimulant-mixture followed by PHS-Mix-3 which was categorized as class-II (moderately attractive) phagostimulant for starved and satiated lots of female flies of both species in olfactometer and field studies; hence these can be exploited for fruit fly management.

Keywords: attractive index, field conditions, olfactometer, Tephritid flies

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Authors: Vishvas N. Patel, Mehul Chorawala

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Currently, according to the authors best knowledge there are less effective therapeutic agents to limit intestinal mucosa damage associated with inflammatory bowel disease (IBD). Clinical studies have shown beneficial effects of several probiotics in patients of IBD. Probiotics are live organisms; confer a health benefit to the host by modulating immunoinflammation and oxidative stress. Although probiotics in murine and human improve disease severity, very little is known about the specific contribution of cell wall contents of probiotics in IBD. Herein, we investigated the ameliorative potential of cell wall contents of Lactobacillus casei (LC) in lipopolysaccharide (LPS)-induced murine colitis. Methods: Colitis was induced in LPS-sensitized rats by intracolonic instillation of LPS (50 µg/rat) for consecutive 14 days. Concurrently, cell wall contents isolated from 103, 106 and 109 CFU of LC was given subcutaneously to each rat for 21 days, considering sulfasalazine (100 mg/kg, p.o.) as standard. The severity of colitis was assessed by body weight loss, food intake, stool consistency, rectal bleeding, colon weight/length, spleen weight and histological analysis. Colonic inflammatory markers (myeloperoxidase (MPO) activity, C-reactive protein and proinflammatory cytokines) and oxidative stress markers (malondialdehyde, reduced glutathione and nitric oxide) were also assayed. Results: Cell wall contents of isolated from 106 and 109 CFU of LC significantly improved the severity of colitis by reducing body weight loss and diarrhea & bleeding incidence, improving food intake, colon weight/length, spleen weight and microscopic damage to the colonic mucosa. The treatment also reduced levels of inflammatory and oxidative stress markers and boosted antioxidant molecule. However, cell wall contents of isolated from 103 were ineffective. Conclusion: In conclusion, cell wall contents of LC attenuate LPS-induced colitis by modulating immuno-inflammation and oxidative stress.

Keywords: probiotics, Lactobacillus casei, immuno-inflammation, oxidative stress, lipopolysaccharide, colitis

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Authors: Delphine Morales, Florian Lombart, Agathe Truchot, Pauline Maire, Pascale Vigneron, Antoine Galmiche, Catherine Lok, Muriel Vayssade

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Targeted therapy molecules are used as a first-line treatment for metastatic melanoma with B-Raf mutation. Nevertheless, these molecules can cause side effects to patients and are efficient on 50 to 60 % of them. Indeed, melanoma cell sensitivity to targeted therapy molecules is dependent on tumor microenvironment (cell-cell and cell-extracellular matrix interactions). To better unravel factors modulating cell sensitivity to B-Raf inhibitor, we have developed and compared several melanoma models: from metastatic melanoma cells cultured as monolayer (2D) to a co-culture in a 3D dermal equivalent. Cell response was studied in different melanoma cell lines such as SK-MEL-28 (mutant B-Raf (V600E), sensitive to Vemurafenib), SK-MEL-3 (mutant B-Raf (V600E), resistant to Vemurafenib) and a primary culture of dermal human fibroblasts (HDFn). Assays have initially been performed in a monolayer cell culture (2D), then a second time on a 3D dermal equivalent (dermal human fibroblasts embedded in a collagen gel). All cell lines were treated with Vemurafenib (a B-Raf inhibitor) for 48 hours at various concentrations. Cell sensitivity to treatment was assessed under various aspects: Cell proliferation (cell counting, EdU incorporation, MTS assay), MAPK signaling pathway analysis (Western-Blotting), Apoptosis (TUNEL), Cytokine release (IL-6, IL-1α, HGF, TGF-β, TNF-α) upon Vemurafenib treatment (ELISA) and histology for 3D models. In 2D configuration, the inhibitory effect of Vemurafenib on cell proliferation was confirmed on SK-MEL-28 cells (IC50=0.5 µM), and not on the SK-MEL-3 cell line. No apoptotic signal was detected in SK-MEL-28-treated cells, suggesting a cytostatic effect of the Vemurafenib rather than a cytotoxic one. The inhibition of SK-MEL-28 cell proliferation upon treatment was correlated with a strong expression decrease of phosphorylated proteins involved in the MAPK pathway (ERK, MEK, and AKT/PKB). Vemurafenib (from 5 µM to 10 µM) also slowed down HDFn proliferation, whatever cell culture configuration (monolayer or 3D dermal equivalent). SK-MEL-28 cells cultured in the dermal equivalent were still sensitive to high Vemurafenib concentrations. To better characterize all cell population impacts (melanoma cells, dermal fibroblasts) on Vemurafenib efficacy, cytokine release is being studied in 2D and 3D models. We have successfully developed and validated a relevant 3D model, mimicking cutaneous metastatic melanoma and tumor microenvironment. This 3D melanoma model will become more complex by adding a third cell population, keratinocytes, allowing us to characterize the epidermis influence on the melanoma cell sensitivity to Vemurafenib. In the long run, the establishment of more relevant 3D melanoma models with patients’ cells might be useful for personalized therapy development. The authors would like to thank the Picardie region and the European Regional Development Fund (ERDF) 2014/2020 for the funding of this work and Oise committee of "La ligue contre le cancer".

Keywords: 3D human skin model, melanoma, tissue engineering, vemurafenib efficiency

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Authors: Thomais Tsoulia, Arvind Y. M. Sundaram, Stine Braaen, Øyvind Haugland, Espen Rimstad, Øystein Wessel, Maria K. Dahle

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Fish red blood cells (RBC) are nucleated, and in addition to their function in gas exchange, they have been characterized as mediators of immune responses. Salmonid RBC are the major target cells of Piscineorthoreovirus (PRV), a virus associated with heart and skeletal muscle inflammation (HSMI) in farmed Atlantic salmon. The activation of antiviral response genesin RBChas previously been described in ex vivo and in vivo PRV-infection models, but not explored in the initial virus encounter phase. In the present study, mRNA transcriptome responses were explored in erythrocytes from individual fish, kept ex vivo, and exposed to purified PRV for 24 hours. The responses were compared to responses in macrophage-like salmon head kidney (SHK-1) and endothelial-like Atlantic salmon kidney (ASK) cells, none of which support PRV replication. The comparative analysis showed that the antiviral response to PRV was strongest in the SHK-1 cells, with a set of 80 significantly induced genes (≥ 2-fold upregulation). In RBC, 46 genes were significantly upregulated, while ASK cells were not significantly responsive. In particular, the transcriptome analysis of RBC revealed that PRV significantly induced interferon regulatory factor 1 (IRF1) and interferon-induced protein with tetratricopeptide repeats 5-like (IFIT9). However, several interferon-regulated antiviral genes which have previously been reported upregulated in PRV infected RBC in vivo (myxovirus resistance (Mx), interferon-stimulated gene 15 (ISG15), toll-like receptor 3 (TLR3)), were not significantly induced after 24h of virus stimulation. In contrast to RBC, these antiviral response genes were significantly upregulated in SHK-1. These results confirm that RBC are involved in the innate immune response to viruses, but with a delayed antiviral response compared to SHK-1. A notable difference is that interferon regulatory factor 1 (IRF-1) is the most strongly induced gene in RBC, but not among the significantly induced genes in SHK-1. Putative differences in the binding, recognition, and response to PRV, and any link to effects on the ability of PRV to replicate remains to be explored.

Keywords: antiviral responses, atlantic salmon, piscine orthoreovirus-1, red blood cells, RNA-seq

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Authors: Cherif Y., Ghariani R., Derbal S., Farhati S., Ben Dahmen F., Abdallah M.

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Introduction: Tuberculosis (TBC) is a frequent infection and is still a major public health problem in Tunisia. The aim of this work is to focus on diagnostic and therapeutic characteristics of TBC in patients referred to our internal medicine department. Patients and Methods: The study was retrospective and descriptive of a cohort of consecutive cases treated from January 2016 to December 2019, collecting patients with latent or patent TBC. Twenty-eight medical records of adults diagnosed with TBC were reviewed. Results: Twenty-eight patients, including 18 women and 10 men, were diagnosed with TBC. Their mean age is 48 years (range: 22-78 years). Five patients have a medical history of diabetes mellitus, 1 patient was followed for systemic lupus erythematosus treated with corticosteroids and immunosuppressant drugs, and another was treated with corticosteroids for Mac Duffy syndrome. The TBC is latent in 12 cases and patent in 16 cases. The most common symptoms were fever and weight loss and were found in 10 cases, a cough in 2 cases, sputum in 3 cases, lymph nodes in 4 cases, erythema nodosum in 2 cases, and neurological signs in 3 cases. Lymphopenia is noticed in 3 cases and a biological inflammatory syndrome in 18 of the cases. The purified protein derivate reaction was positive in 17 cases, anergic in 3 cases, negative in 5 cases, and not done in 3 cases. The acid-fast bacilli stain culture was strongly positive in one patient. The histopathological study was conclusive in 11 patients and showed granulomatosis with caseous necrosis. TBC was pulmonary in 7 patients, lymph node in 7 cases, peritoneal in 7 cases, digestive in 1 case, neuromeningeal in 3 cases, and thyroïd in 1 case. Seven patients had multifocal TBC. All the patients received anti-tuberculosis treatment with a mean duration of 8 months with no failure or relapse with an average follow-up time of 10.58 months. Conclusion: Diagnosis and management of TBC remain essential to avoid serious complications. The survey is necessary to ensure timely detection and treatment of infected adults to decrease its incidence. The best treatment remains preventive through vaccination and improving social and economic conditions.

Keywords: tuberculosis, infection, autoimmune disease, granulomatosis

Procedia PDF Downloads 165

Authors: Ipsita Biswas, Arnab Sarkar, Ashikur Rahaman, Gopeswar Mukherjee, Subhrangsu Chatterjee, Shamee Bhattacharjee, Deba Prasad Mandal

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One of the major reasons for the high mortality rate of lung cancer is the substantial delays in disease detection at late metastatic stages. It is of utmost importance to understand the detailed molecular signaling and detect the molecular markers that can be used for the early diagnosis of cancer. Several studies explored the emerging roles of long noncoding RNAs (lncRNAs) in various cancers as well as lung cancer. A long non-coding RNA LINC00273 was recently discovered to promote cancer cell migration and invasion, and its positive correlation with the pathological stages of metastasis may prove it to be a potential target for inhibiting cancer cell metastasis. Comparing real-time expression of LINC00273 in various human clinical cancer tissue samples with normal tissue samples revealed significantly higher expression in cancer tissues. This long intergenic noncoding RNA was found to be highly expressed in human liver tumor-initiating cells, human gastric adenocarcinoma AGS cell line, as well as human non-small cell lung cancer A549 cell line. SiRNA and shRNA-induced knockdown of LINC00273 in both in vitro and in vivo nude mice significantly subsided AGS and A549 cancer cell migration and invasion. LINC00273 knockdown also reduced TGF-β induced SNAIL, SLUG, VIMENTIN, ZEB1 expression, and metastasis in A549 cells. Plenty of reports have suggested the role of microRNAs of the miR200 family in reversing epithelial to mesenchymal transition (EMT) by inhibiting ZEB transcription factors. In this study, hsa-miR-200a-3p was predicted via IntaRNA-Freiburg RNA tools to be a potential target of LINC00273 with a negative free binding energy of −8.793 kcal/mol, and this interaction was verified as a confirmed target of LINC00273 by RNA pulldown, real-time PCR and luciferase assay. Mechanistically, LINC00273 accelerated TGF-β induced EMT by sponging hsa-miR-200a-3p which in turn liberated ZEB1 and promoted prometastatic functions in A549 cells in vitro as verified by real-time PCR and western blotting. The similar expression patterns of these EMT regulatory pathway molecules, viz. LINC00273, hsa-miR-200a-3p, ZEB1 and TGF-β, were also detected in various clinical samples like breast cancer tissues, oral cancer tissues, lung cancer tissues, etc. Overall, this LINC00273 mediated EMT regulatory signaling can serve as a potential therapeutic target for the prevention of lung cancer metastasis.

Keywords: epithelial to mesenchymal transition, long noncoding RNA, microRNA, non-small-cell lung carcinoma

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Authors: Wenpeng You, Maciej Henneberg

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Background: Sugar consumption has been overwhelmingly advocated as a major dietary offender to obesity prevalence. Meat intake has been hypothesized as an obesity contributor in previous publications, but a moderate amount of meat to be included in our daily diet still has been suggested in many dietary guidelines. Comparable sugar and meat exposure data were obtained to assess the difference in relationships between the two major food groups and obesity prevalence at population level. Methods: Population level estimates of obesity and overweight rates, per capita per day exposure of major food groups (meat, sugar, starch crops, fibers, fats and fruits) and total calories, per capita per year GDP, urbanization and physical inactivity prevalence rate were extracted and matched for statistical analysis. Correlation coefficient (Pearson and partial) comparisons with Fisher’s r-to-z transformation and β range (β ± 2 SE) and overlapping in multiple linear regression (Enter and Stepwise) were used to examine potential differences in the relationships between obesity prevalence and sugar exposure and meat exposure respectively. Results: Pearson and partial correlations (controlled for total calories, physical inactivity prevalence, GDP and urbanization) analyses revealed that sugar and meat exposures correlated to obesity and overweight prevalence significantly. Fisher's r-to-z transformation did not show statistically significant difference in Pearson correlation coefficients (z=-0.53, p=0.5961) or partial correlation coefficients (z=-0.04, p=0.9681) between obesity prevalence and both sugar exposure and meat exposure. Both Enter and Stepwise models in multiple linear regression analysis showed that sugar and meat exposure were most significant predictors of obesity prevalence. Great β range overlapping in the Enter (0.289-0.573) and Stepwise (0.294-0.582) models indicated statistically sugar and meat exposure correlated to obesity without significant difference. Conclusion: Worldwide sugar and meat exposure correlated to obesity prevalence at the same extent. Like sugar, minimal meat exposure should also be suggested in the dietary guidelines.

Keywords: meat, sugar, obesity, energy surplus, meat protein, fats, insulin resistance

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Authors: Onuoha Ogbonnaya Gideon, Ishaq Hafsah Yusuf

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Introduction: Meat is a perishable food of good nutrition. Meat ranks among the most significant, nutritious, and favored food items available to most locals. It is a good source of protein (17-19%), depending on sources, and contains appreciable amounts of fat and moisture. However, it has a very short shelf life due mainly to its high moisture, fat, and other nutrient contents. Meat spoilage can result from microbial proliferation as well as inherent enzymes in the meat tissues. Bacteria contamination and permeability to both oxygen and water vapor are major concerns associated with spoilage of meat and its storage. Packaging is fundamental in the preservation and presentation of food. Red clay is a very common substance; hydrous aluminum phyllosilicate, sometimes with varying amounts of iron, magnesium, alkali metals, alkaline earth, and cation formed from sedimentary rocks. Furthermore, red clay is an extremely absorbent material and develops plasticity when wet due to the molecular film of water surrounding the clay particles but can become hard, impervious, brittle, and non-brittle and non-plastic when dry. In developing countries, the high cost of refrigeration technologies and most other methods of preserving meat are exorbitant and thus can be substituted with the less expensive and readily available red clay for the preservation of meat. Methodology: 1000g of lean meat was diced into cubes of 10g each. The sample was then divided into four groups labelled raw meat (RMC); raw in 10% brine solution (RMB), boiled meat (BMC), and fried meat (FMC). It was then encapsulated with 2mm thick red clay and then heated in a muffle furnace at a temperature of 600OC for 30min. The samples were kept on a bench top for 30 days, and a storage study was carried out. Results: Our findings showed a decrease in value during storage for the physiochemical properties of all the sample; pH values decreased [RMC (7.05-7.6), RMB (8.46-7.0), BMC (6.0-5.0), FMC (4.08-3.9)]; free fatty acid content decreased with storage time [RMC (32.6%-31%), RMB (30.2%-28.6%), BMC (30.5%-27.4%), FMC (25.6%-23.8%)]; total soluble solid value decreased [RMC16.20-15.07, RMB (17.22-16.04), BMC (17.05-15.54), FMC (15.3-14.9)]. Conclusion: This result shows that encapsulation with red clay reduced all the values analyzed and thus has the potential to extend the shelf life of stored meat.

Keywords: red clay, encapsulating, shelf life, physicochemical properties, lean meat

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Authors: Safee Chaudhary, Mahnoor Naseer Gondal, Hira Anees Awan, Abdul Rehman, Ammar Arif, Risham Hussain, Huma Khawar, Zainab Arshad, Muhammad Faizyab Ali Chaudhary, Waleed Ahmed, Muhammad Umer Sultan, Bibi Amina, Salaar Khan, Muhammad Moaz Ahmad, Osama Shiraz Shah, Hadia Hameed, Muhammad Farooq Ahmad Butt, Muhammad Ahmad, Sameer Ahmed, Fayyaz Ahmed, Omer Ishaq, Waqar Nabi, Wim Vanderbauwhede, Bilal Wajid, Huma Shehwana, Muhammad Tariq, Amir Faisal

Abstract:

Cancer is a manifestation of multifactorial deregulations in biomolecular pathways. These deregulations arise from the complex multi-scale interplay between cellular and extracellular factors. Such multifactorial aberrations at gene, protein, and extracellular scales need to be investigated systematically towards decoding the underlying mechanisms and orchestrating therapeutic interventions for patient treatment. In this work, we propose ‘TISON’, a next-generation web-based multiscale modeling platform for clinical systems oncology. TISON’s unique modeling abstraction allows a seamless coupling of information from biomolecular networks, cell decision circuits, extra-cellular environments, and tissue geometries. The platform can undertake multiscale sensitivity analysis towards in silico biomarker identification and drug evaluation on cellular phenotypes in user-defined tissue geometries. Furthermore, integration of cancer expression databases such as The Cancer Genome Atlas (TCGA) and Human Proteome Atlas (HPA) facilitates in the development of personalized therapeutics. TISON is the next-evolution of multiscale cancer modeling and simulation platforms and provides a ‘zero-code’ model development, simulation, and analysis environment for application in clinical settings.

Keywords: systems oncology, cancer systems biology, cancer therapeutics, personalized therapeutics, cancer modelling

Procedia PDF Downloads 179

Authors: Chakrapani Pullagummi, Arun Jyothi Bheemagani, B. Chandra Sekhar Singh, Prem Kumar, A. Roja Rani

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Diabetes mellitus describes a metabolic disorder of multiple aetiology characterized by chronic hyperglycaemia with disturbances of carbohydrate, fat and protein metabolism resulting from defects in insulin secretion and its action. The objective of present study was alloxan induced diabetes in S.D (Sprague Dawley) rats, treated with leaf extract of Andrographis paniculata and bark extract of Erythrina indica. Plant extract treated rats were analyzed biochemically and molecularly. on normal and diabetic rats. The changes in MDA (lipid peroxidation) and glucose (by GOD method) levels in blood of both normal and diabetic rat were analyzed. Diabetes induced rats were treated with methanolic extracts of Andrographis paniculata leaf and Erythrina indica bark which are of medicinal importance. Later after inducing diabetes the rats were treated with medicinal plant extracts, Andrographis paniculata leaf and Erythrina indica bark which are well known for their anti diabetic and antioxidative property in order to control the glucose and MDA levels. The blood plasma of diabetic and normal rats was analyzed for the levels of MDA (lipid peroxidation) and glucose levels. Results of this study suggested that the Andrographis paniculata leaf and Erythrina indica can be used as a potential natural antidiabetic agent for treating and postponing the appearance of complications that arise due to Diabetes. Molecular study deals with the analysis of binding mechanism of 2 selected natural compounds from Andrographis and Erythrina extracts against the novel target for type T2D namely PPAR-γ compared with Rosiglitazone (standard compound). The results revealed that most of the selected herbal lead compounds were effective targets against the receptors. These compounds showed favorable interactions with the amino acid residues thereby substantiating their proven efficacy as anti-diabetic compounds.

Keywords: andrographis paniculata, erythrina indica, alloxan, lipid peroxidation, blood glucose level, PPAR-γ

Procedia PDF Downloads 450

Authors: Surbhi Surbhi, Andrea Erni, Gunter Meister, Harold Cremer, Christophe Beclin

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MicroRNAs (miRNAs) are short 20-23 nucleotide long non-coding RNAs; there are 2605 miRNA in humans and 1936 miRNA in mouse in total (miRBase). The nervous system expresses the most abundant miRNA and most diverse. MiRNAs play a role in many steps during neurogenesis, like cell proliferation, differentiation, neural patterning, axon pathfinding, etc. Moreover, in vitro studies suggested a role in the regulation of local translation at the synapse, thus controlling neuronal plasticity. However, due to the specific structure of miRNA molecules, an in-vivo confirmation of the general role of miRNAs in the control of neuronal plasticity is still pending. For example, their small size and their high level of sequence homology make difficult the analysis of their cellular and sub-cellular localization in-vivo by in-situ hybridization. Moreover, it was found that only 40% of the expressed miRNA molecules in a cell are included in RNA-Induced Silencing Complexes (RISC) and, therefore, involved in inhibitory interactions while the rest is silent. Definitively, the development of new tools is needed to have a better understanding of the cellular function of miRNAs, in particular their role in neuronal plasticity. Here we describe a new technique called in-vivo AGO-APP designed to investigate miRNA expression and function in-vivo. This technique is based on the expression of a small peptide derived from the human RISC-complex protein TNRC6B, called T6B, which binds all known Argonaute (Ago) proteins with high affinity allowing the efficient immunoprecipitation of AGO-bound miRNAs. We have generated two transgenic mouse lines conditionally expressing T6B either ubiquitously in the cell or targeted at the synapse. A comparison of the repertoire of miRNAs immuno-precipitated from mature neurons of both mouse lines will provide us with a list of miRNAs showing a specific activity at the synapse. The physiological role of these miRNAs will be subsequently addressed through gain and loss of function experiments.

Keywords: RNA-induced silencing complexes, TNRC6B, miRNA, argonaute, synapse, neuronal plasticity, neurogenesis

Procedia PDF Downloads 92

Authors: Obadire F. O., Obadire, S. O., Adeoti R. F., Pirgozliev V.

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An experiment was conducted to determine the efficacy of utilizing African black plum seed nut (ABPNBD) meal on hematological and biochemical indices of broiler chicken ration formulated to substitute wheat offal. A total of 150- 1day-old, male Agrited birds were reared for 28 days of the experiment. The birds were assigned to five dietary treatments, with ten birds per treatment replicated 3 times. Experimental diets were formulated by supplementing the milled African black plum nut at (0, 5, 10, 12.5, and 15%) inclusion levels in the starter broiler’s ratio designated as T1 (control diet containing no ABPBD), Treatments (T2, 3,4 and 5) contained ABPNBD at 5, 10, 12.5, and 15%, respectively, in a completely randomized design. The hematological and biochemical indices of the birds were determined. The result revealed that all hematological parameters measured were significant (P <0.05) except for WBC. Increasing inclusion levels of ABPNBD decreased the PCV, HB, and RBC of the birds across the treatment groups. Birds fed 12.5 and 15% ABPNBD diets recorded the least of the parameters. The result of the serum biochemical indices showed significant (P < 0.05) influence for all parameters measured except for alanine transaminase (ALT), (AST), and creatinine. The total protein (TP), albumin, globulin, and glucose values were reduced across the treatment group as ABPNBD inclusion increased. Birds fed above 10% ABPNBD recorded the lowest value of TP, albumin, globulin, and glucose when compared with birds on a control diet and other treatments. The uric acid ranged from 3.85 to 2 .13 mmol/L, while creatinine ranged from 62.00 to 53.50 mmol/l. AST ranged between 8.50 u/l (5%) to 7.90 u/l (10%). ALT ranged between 7.50 u/l (12.5%) to 5.50 u/l (5 and 10%). In conclusion, dietary inclusion of African black plum up to 10% has no detrimental effect on the health of the starter chickens. Meanwhile, inclusion above 10% revealed a negative effect on some blood parameters measured. Therefore, African black plum should be supplemented with probable probiotics or subjected to different processing methods if to be used at a 15% inclusion level for optimal results.

Keywords: African black plum seed, starter broiler chickens, hematological and serum biochemical indices, (Vitex doniana)

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Authors: Farhana Nosheen, Moeez Ahmad

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Throughout the food chain, the food waste from initial agricultural production to final household consumption has become a serious concern for global sustainability because of its adverse impacts on food security, natural resources, the environment, and human health. About a third of tomatoes (Lycopersicon esculentum L.) delivered to processing plants end as processing waste. The amount of such waste material is estimated to have increased with the emergence of mechanical harvesting. Experiments were made to determine the nutritional profile and antioxidant activity of tomato processing waste and to explore the bioactive compound in tomato waste, i.e., Lycopene. Tomato Variety of ‘SAHARA F1’ was used to make tomato waste. The tomatoes were properly cleaned, and then unwanted impurities were removed properly. The tomatoes were blanched at 90 ℃ for 5 minutes. After which, the skin of the tomatoes was removed, and the remaining part passed through the electric pulper. The pulp and seeds were collected separately. The seeds and skin of tomatoes were mixed and saved in a sterilized jar. The samples of tomato waste were found to contain 89.11±0.006 g/100g moisture, 10.13±0.115 g/100g protein, 2.066±0.57 g/100g fat, 4.81±0.10 g/100g crude fiber, and 4.06±0.057 g/100g ash and NFE 78.92±0.066 g/100g. The results confirmed that tomato waste contains a considerable amount of Lycopene 51.0667±0.00577 mg/100g and exhibited good antioxidant properties. Total phenolics showed average contents of 122.9600±0.01000 mg GAE/100g, of which flavonoids accounted for 41.5367±0.00577 mg QE/100g. Antioxidant activity of tomato processing waste was found 0.6833±0.00577 mmol Trolox/100g. Unsaturated fatty acids represent the major portion of total fatty acids, Linoleic acid being the major one. The mineral content of tomato waste showed a good amount of potassium 3030.1767 mg/100g and calcium 131.80 mg/100g, respectively were present in it. These findings suggest that tomato processing waste is rich in nutrients, antioxidants, fatty acids, and minerals. I recommend that this waste should be sun-dried to be used in the combination of feed of the animals. It can also be used in making some other products like lycopene tea or several other health-beneficial products.

Keywords: food waste, tomato, bioactive compound, sustainable management

Procedia PDF Downloads 73

Authors: Lamzira Ebralidze, Aleksandre Tsertsvadze, Dali Berashvili, Aliosha Bakuridze

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Nowadays, there is an increasing demand for biologically active substances from vegetable, animal, and mineral resources. In terms of the use of natural compounds, pharmaceutical, cosmetic, and nutrition industry has big interest. The biggest drawback of conventional extraction methods is the need to use a large volume of organic extragents. The removal of the organic solvent is a multi-stage process. And their absolute removal cannot be achieved, and they still appear in the final product as impurities. A large amount of waste containing organic solvent damages not only human health but also has the harmful effects of the environment. Accordingly, researchers are focused on improving the extraction methods, which aims to minimize the use of organic solvents and energy sources, using alternate solvents and renewable raw materials. In this context, green extraction principles were formed. Green Extraction is a need of today’s environment. Green Extraction is the concept, and it totally corresponds to the challenges of the 21st century. The extraction of biologically active compounds based on green extraction principles is vital from the view of preservation and maintaining biodiversity. Novel technologies of green extraction are known, such as "cold methods" because during the extraction process, the temperature is relatively lower, and it doesn’t have a negative impact on the stability of plant compounds. Novel technologies provide great opportunities to reduce or replace the use of organic toxic solvents, the efficiency of the process, enhance excretion yield, and improve the quality of the final product. The objective of the research is the development of green technologies of flavonoids containing preparations. Methodology: At the first stage of the research, flavonoids containing preparations (Tincture Herba Leonuri, flamine, rutine) were prepared based on conventional extraction methods: maceration, bismaceration, percolation, repercolation. At the same time, the same preparations were prepared based on green technologies, microwave-assisted, UV extraction methods. Product quality characteristics were evaluated by pharmacopeia methods. At the next stage of the research technological - economic characteristics and cost efficiency of products prepared based on conventional and novel technologies were determined. For the extraction of flavonoids, water is used as extragent. Surface-active substances are used as co-solvent in order to reduce surface tension, which significantly increases the solubility of polyphenols in water. Different concentrations of water-glycerol mixture, cyclodextrin, ionic solvent were used for the extraction process. In vitro antioxidant activity will be studied by the spectrophotometric method, using DPPH (2,2-diphenyl-1- picrylhydrazyl) as an antioxidant assay. The advantage of green extraction methods is also the possibility of obtaining higher yield in case of low temperature, limitation extraction process of undesirable compounds. That is especially important for the extraction of thermosensitive compounds and maintaining their stability.

Keywords: extraction, green technologies, natural resources, flavonoids

Procedia PDF Downloads 101

Authors: Kan Yu, Khui Hung Lee, Eben Afrifa-Yamoah, Jing Guo, Katrina Harrison, Jack Goldblatt, Nicholas Pachter, Jitian Xiao, Guicheng Brad Zhang

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Background and Significance of the Study: Congenital Heart Defects (CHDs) are the most common malformation at birth and one of the leading causes of infant death. Although the exact etiology remains a significant challenge, epigenetic modifications, such as DNA methylation, are thought to contribute to the pathogenesis of congenital heart defects. At present, no existing DNA methylation biomarkers are used for early detection of CHDs. The existing CHD diagnostic techniques are time-consuming and costly and can only be used to diagnose CHDs after an infant was born. The present study employed a machine learning technique to analyse genome-wide methylation data in children with and without CHDs with the aim to find methylation biomarkers for CHDs. Methods: The Illumina Human Methylation EPIC BeadChip was used to screen the genome‐wide DNA methylation profiles of 24 infants diagnosed with congenital heart defects and 24 healthy infants without congenital heart defects. Primary pre-processing was conducted by using RnBeads and limma packages. The methylation levels of top 600 genes with the lowest p-value were selected and further investigated by using a random forest approach. ROC curves were used to analyse the sensitivity and specificity of each biomarker in both training and test sample sets. The functionalities of selected genes with high sensitivity and specificity were then assessed in molecular processes. Major Findings of the Study: Three genes (MIR663, FGF3, and FAM64A) were identified from both training and validating data by random forests with an average sensitivity and specificity of 85% and 95%. GO analyses for the top 600 genes showed that these putative differentially methylated genes were primarily associated with regulation of lipid metabolic process, protein-containing complex localization, and Notch signalling pathway. The present findings highlight that aberrant DNA methylation may play a significant role in the pathogenesis of congenital heart defects.

Keywords: biomarker, congenital heart defects, DNA methylation, random forest

Procedia PDF Downloads 128

Authors: Chang-Hui Shen, Paulina Konarzewska

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Cell homeostasis is regulated by vacuolar activity and it has been shown that lipid composition of the vacuole plays an important role in vacuolar function. The major phosphoinositide species present in the vacuolar membrane include phosphatidylinositol 3,5-biphosphate (PI(3,5)P₂) which is generated from PI(3)P controlled by Fab1p. Deletion of FAB1 gene reduce the synthesis of PI(3,5)P₂ and thus result in enlarged or fragmented vacuoles, with neutral vacuolar pH due to reduced vacuolar H⁺-ATPase activity. These mutants also exhibited poor growth at high extracellular pH and in the presence of CaCl₂. Conversely, VPS34 regulates the synthesis of PI(3)P from phosphatidylinositol (PI), and the lack of Vps34p results in the reduction of vacuolar activity. Although the cellular observations are clear, it is still unknown about the molecular mechanism between the phospholipid biosynthesis pathway and vacuolar activity. Since both VPS34 and FAB1 are important in vacuolar activity, we hypothesize that the molecular mechanism of vacuolar function might be regulated by the transcriptional regulators of phospholipid biosynthesis. In this study, we study the role of the major phospholipid biosynthesis transcription factor, INO2, in the regulation of vacuolar activity. We first performed qRT-PCR to examine the effect of Ino2p on the expression of VPS34 and FAB1. Our results showed that VPS34 was upregulated in the presence of inositol for both WT and ino2Δ cells. However, FAB1 was only upregulated significantly in ino2Δ cells. This indicated that Ino2p might be the negative regulator for FAB1 expression. Next, growth sensitivity experiment showed that WT, vma3Δ, and ino2Δ grew well in growth medium buffered to pH 5.5 containing 10 mM CaCl₂. As cells were switched to growth medium buffered to pH 7 containing CaCl₂ WT, ino2Δ and opi1Δ showed growth reduction, whereas vma3Δ was completely nonviable. As the concentration of CaCl₂ was increased to 60 mM, ino2Δ cells showed moderate growth reduction compared to WT. This result suggests that ino2Δ cells have better vacuolar activity. Microscopic analysis and vacuolar acidification were employed to further elucidate the importance of INO2 in vacuolar homeostasis. Analysis of vacuolar morphology indicated that WT and vma3Δ cells displayed vacuoles that occupied a small area of the cell when grown in media buffered to pH 5.5. Whereas, ino2Δ displayed fragmented vacuoles. On the other hand, all strains grown in media buffered to pH 7, exhibited enlarged vacuoles that occupied most of the cell’s surface. This indicated that the presence of INO2 may play negative effect in vacuolar morphology when cells are grown in media buffered to pH 5.5. Furthermore, vacuolar acidification assay showed that only vma3Δ cells displayed notably less acidic vacuoles as cells were grown in media buffered to pH 5.5 and pH 7. Whereas, ino2Δ cells displayed more acidic pH compared to WT at pH7. Taken together, our results demonstrated the molecular mechanism of the vacuolar activity regulated by the phospholipid biosynthesis transcription factors Ino2p. Ino2p negatively regulates vacuolar activity through the expression of FAB1.

Keywords: vacuole, phospholipid, homeostasis, Ino2p, FAB1

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Authors: Tomasz Borowski, Dawid Sołoducha, Agata Markowska-Szczupak, Aneta Wesołowska, Marian Kordas, Rafał Rakoczy

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Essential oils (EOs) are fragrant volatile oils obtained from plants. These are used for cooking (for flavor and aroma), cleaning, beauty (e.g., rosemary essential oil is used to promote hair growth), health (e.g. thyme essential oil cures arthritis, normalizes blood pressure, reduces stress on the heart, cures chest infection and cough) and in the food industry as preservatives and antioxidants. Rosemary and thyme essential oils are considered the most eminent herbs based on their history and medicinal properties. They possess a wide range of activity against different types of bacteria and fungi compared with the other oils in both in vitro and in vivo studies. However, traditional uses of EOs are limited due to rosemary and thyme oils in high concentrations can be toxic. In light of the accessible data, the following hypothesis was put forward: Low frequency rotating magnetic field (RMF) increases the antimicrobial potential of EOs. The aim of this work was to investigate the antimicrobial activity of commercial Salvia Rosmarinus L. and Thymus vulgaris L. essential oil from Polish company Avicenna-Oil under Rotating Magnetic Field (RMF) at f = 25 Hz. The self-constructed reactor (MAP) was applied for this study. The chemical composition of oils was determined by gas chromatography coupled with mass spectrometry (GC-MS). Model bacteria Escherichia coli K12 (ATCC 25922) was used. Minimum inhibitory concentrations (MIC) against E. coli were determined for the essential oils. Tested oils in very small concentrations were prepared (from 1 to 3 drops of essential oils per 3 mL working suspensions). From the results of disc diffusion assay and MIC tests, it can be concluded that thyme oil had the highest antibacterial activity against E. coli. Moreover, the study indicates the exposition to the RMF, as compared to the unexposed controls causing an increase in the efficacy of antibacterial properties of tested oils. The extended radiation exposure to RMF at the frequency f= 25 Hz beyond 160 minutes resulted in a significant increase in antibacterial potential against E. coli. Bacteria were killed within 40 minutes in thyme oil in lower tested concentration (1 drop of essential oils per 3 mL working suspension). Rapid decrease (>3 log) of bacteria number was observed with rosemary oil within 100 minutes (in concentration 3 drops of essential oils per 3 mL working suspension). Thus, a method for improving the antimicrobial performance of essential oil in low concentrations was developed. However, it still remains to be investigated how bacteria get killed by the EOs treated by an electromagnetic field. The possible mechanisms relies on alteration in the permeability of ionic channels in ionic channels in the bacterial cell walls that transport in the cells was proposed. For further studies, it is proposed to examine other types of essential oils and other antibiotic-resistant bacteria (ARB), which are causing a serious concern throughout the world.

Keywords: rotating magnetic field, rosemary, thyme, essential oils, Escherichia coli

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Authors: Grace McCambridge, Alanna Keady, Madhur Agrawal, Dequina Nicholas Alvarado, Barbara Nikolajczyk, Leena Panneerseelan-Bharath

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Age is a non-modifiable risk factor for the inflammation that underlies pathologies such as type 2 diabetes mellitus (T2DM). Inflammation, as indicated by circulating cytokines, rises in aging, but mechanisms that promote this ‘inflammaging’ remain poorly defined. Furthermore, downstream consequences of inflammaging, including the development of an inflammatory profile that predicts comorbidities like T2DM, remain speculative. We tested the possibility that natural aging-associated changes in autophagy, a process that is compromised in both aging and T2DM, regulates inflammatory profiles in older subjects. Our data showed that circulating CD4⁺ T cells from older compared to younger subjects have (i) defects in autophagy; (ii) higher mitochondria accumulation; (iii) a failure to metabolically shift from oxidative phosphorylation to anaerobic glycolysis upon αCD3/CD28 activation; (iv) more reactive oxygen species (ROS) accumulation; and (v) a cytokine profile that recapitulates the Th17 profile that predicts T2DM. ROS scavenging in cells from older subjects restored mitochondrial mass and membrane potential (indicators of improved autophagy) and reduced Th17 cytokines to amounts made by T cells from younger subjects. Knock-down of the autophagy protein Atg3 in T cells from younger subjects increased mitochondrial accumulation and Th17 cytokines. To begin translating these findings to clinical practice, we showed that physiological concentrations of the diabetes drug metformin (100 µM) added in vitro enhanced autophagy, prevented mitochondria and ROS accumulation, increased anaerobic glycolysis, and decreased Th17 cytokines in activated CD4⁺ T cells from older subjects. Metformin therefore improves autophagy and multiple downstream pro-inflammatory mechanisms CD4⁺ T cells from older subjects. We conclude that autophagy improvement ameliorates the development of a T2DM-predictive Th17 profile in aging, and thus holds promise for delay or prevention of aging-associated metabolic decline.

Keywords: autophagy, mitochondrial turnover, ROS, glycolysis

Procedia PDF Downloads 127

Authors: Ugwuowo Leonard Chidi, Oparaji Chetachukwu Jecinta., Ogidi Chibuzor Agafenachukwu, Onuoha Rebecca Obianuju

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This experiment was conducted to investigate the growth performance, haematology and serum biochemistry indices of broiler birds fed diets containing Neem leaf meal. A total of 96 unsexed day-old broiler birds were allocated to four treatments of T1, T2, T3 and T4 and replicated three times with eight birds per replicate in a Completely Randomized Design. The treatments were diets containing 2.0, 4.0, 6.0 and 8.0% Neem leaf meal respectively. Growth performances, packed cell volume, red blood cell count, haemoglobin, white blood cell count, lymphocytes, mean corpuscular volume, mean corpuscular haemoglobin concentration, platelet count, aspartate amino transaminase, alanine amino transaminase, alkaline phosphate, cholesterol, albumin, globulin, urea, glucose, total protein and creatinine were evaluated. Results showed that there were no significant differences (P>0.05) in all the growth performance parameters among the treatments. The results of the experiment showed that there were significant differences (P<0.05) in all the heamatological and serum biochemistry parameters at finisher phases. Mean corpuscular volume, white blood cell count, lymphocytes, red blood cell count, haemoglobin, platelet count, creatinine and triglyceride increased and were highest in treatment two while treatment four had the least values in mean corpuscular volume, urea, white blood cell, haemoglobin and triglyceride. This implies that the levels of inclusion of Neem leaf meal in this experiment did not affect the growth performance of the broiler chicks but the haematological and serum biochemistry indices were affected. Treatment two with a 4% inclusion level of Neem leaf meal has shown the capacity to replace vaccines and antibiotics in broilers due to the positive effects it had on both the haematological and serum biochemistry.

Keywords: leaf meal, broiler, Aziradicta indica, serum biochemistry, haematology

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Authors: Javed Sidiqi, Stephen Baezinger, Stephen Wegulo

Abstract:

Wheat (Triticum aestivum L.) production continues to be challenged by foliar fungal diseases although significant improvement has been made to manage the diseases through developing resistant varieties and the fungicide use to ensure sufficient wheat is produced to meet the growing population’s need. Significant crop losses have been recorded in the history of grain production and yield losses due to fungal diseases, and the trend continues to threat food security in the world and particularly in the less developed countries. The impact of individual fungal diseases on grain yield has been studied extensively to determine crop losses. However, there is limited research available to find out the combined effects of fungal diseases on grain yield and the ways to effectively manage the diseases. Therefore, the objectives of this research were to study the effect of fungal pathogens on grain yield of pre-released winter wheat genotypes in fungicide treated and untreated plots, and to determine whether S7b gene was present in ‘Gage’ wheat as previously hypothesized. Sixty winter wheat genotypes in fungicide treated and untreated plots were studied across four environments. There was a significant effect of fungicide on grain yield consistently across four environments in three years. Fungicide treated wheat lines demonstrated (4,496 kg/ ha-1) grain yield compared to (3,147 kg/ ha-1) grain yield in untreated wheat lines indicating 43% increased grain yield due to severity of foliar fungal diseases. Furthermore, fungicide application also caused an increase in protein concentration from 153 (g kg-1) to 164 (g kg-1) in treated plots in along with test weight from 73 to 77 (kg hL-1) respectively. Gage wheat variety and ISr7b-Ra were crossed to determine presence of Sr7b in Gage. The F2 and F2:3 segregating families were screened and evaluated for stem rust resistance. The segregation of families fell within 15:1 ratio for two separate resistance genes suggesting that Sr7b segregates independently from an unknown resistance gene in Gage that needs to be characterized for its use in the future wheat breeding program to develop resistant wheat varieties.

Keywords: funicide, genetics, foliar diseases, grain

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