Search results for: tumor protein (P53)

Authors: A. G. Miah, U. Salma, K. Schellander

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Relaxin was first described in 1926 by Frederick Hisaw. Previously it was considered as only the hormone of pregnant mammals due to its important roles in pregnancy and parturition. From the last decade, the physiological role of relaxin in male reproduction has been given experimental attention, and the results have made it clear that relaxin can no longer be considered strictly as only the hormone of female reproduction. The accessory glands (specially, the prostate glands) of the male reproductive system are the source of seminal relaxin, which is secreted into the seminal plasma and saturated with spermatozoa just after ejaculation. Several studies have reported that relaxin has important roles in improving motility in human sperm. Thereafter, the growing interest on relaxin has intensified efforts to investigate the role of relaxin in other sperm physiological phenomena like, capacitation, acrosome reaction, and their mediating factors associated with successful fertilization. Therefore, this review aims to provide up-to-date information about the physiological roles of relaxin in sperm motility, capacitation, acrosome reaction, and their mediating factors, such as, utilization of glucose, cholesterol efflux, Ca2+-influx, intracellular cAMP and protein tyrosine phosphorylation. Some studies have shown relaxin to increase the percentage of progressive motility and induce capacitation and acrosome reaction through increasing the utilization of glucose and mediating the cholesterol efflux, Ca2+-influx, intracellular cAMP and protein tyrosine phosphorylation. Thus, the review suggests that the supplementation of relaxin into the capacitating medium may contribute the possible beneficial roles in fresh and cryopreserved spermatozoal prefertilization events.

Keywords: relaxin, physiological roles, prefertilizing activities, spermatozoa

Procedia PDF Downloads 572

Authors: Gladys Abiemwense Ibhaze, Anthony Henry Ekeocha, Adebowale Noah Fajemisin, Tope Oke, Caroline Tosin Alade,

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The study was conducted to evaluate the silage quality and acceptability of ensiled avocado pear skin (APS) with cassava peel (CSP) and brewers’ grain (BG) using eighteen (18) West African Dwarf goats with an average weight of 7.0±1.5 kg. The experimental diets; 1) 50% cassava peel+ 50% brewers’ grain, 2) 50% brewers’ grain+ 50% avocado pear skin, 3) 50% cassava peel +25% brewers’ grain+ 25% avocado pear skin were ensiled for 21 days. The experimental design was a completely randomized design (CRD). The chemical composition of the diets was investigated. The acceptability of the diets was evaluated for twelve (12) days. Results obtained showed that the crude protein content ranged from 12.18 – 12.47%, crude fiber (15.99-22.67%). Results obtained showed that diet 1 had the least pH value (4.0), followed by diet 3 (4.5) and diet 2 (5.2). All diets were firm in texture and maintained their initial color. The temperature ranged from 27-29 ⁰C with diet 2 having the highest temperature of 29 ⁰C. Acceptability of experimental diets varied (p < 0.05) significantly. Dry matter intake ranged from (426.22-686.73g/day) with animals on a diet one recording the highest dry matter intake. The coefficient of preference and percentage preference, also differed (p <0.05) significantly among the diets. Diet 1 had a coefficient of preference greater than unity. However, this was not significantly (p>0.05) different from diet two but differed from diet 3. Conclusively, APS could be included in goats’ diets in the absence of CSP during feed scarcity provided a rich source of protein is available.

Keywords: avocado pear skin, Brewers' grain, Cassava peel, preference

Procedia PDF Downloads 209

Authors: I. Karagjozova, B. Dejanova, J. Pluncevic, S. Petrovska, V. Antevska, L. Todorovska

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Introduction: Medical checking assessment is important in sports medicine. To follow the health condition in subjects who perform sports, body composition parameters, such as intracellular water, extracellular water, protein and mineral content, muscle and fat mass might be useful. The aim of the study was to show available parameters and to compare them to manual assessment. Material and methods: A number of 20 subjects (14 male and 6 female) at age of 20±2 years were determined in the study, 5 performed recreational sports, while others were professional ones. The mean height was 175±7 cm, the mean weight was 72±9 cm, and the body mass index (BMI) was 23±2 kg/m2. The measured compartments were as following: intracellular water (IW), extracellular water (EW), protein component (PC), mineral component (MC), skeletal muscle mass (SMM) and body fat mass (BFM). Lean balance were examined for right and left arm (LA), trunk (T), right leg (RL) and left leg (LL). The comparison was made between the calculation derived by manual made measurements, using Matejka formula and parameters obtained by body composition analyzer (BCA) - Inbody 720 BCA Biospace. Used parameters for the comparison were muscle mass (SMM), body fat mass (BFM). Results: BCA obtained values were for: IW - 22.6±5L, EW - 13.5±2 L, PC - 9.8±0.9 kg, MC - 3.5±0.3, SMM - 27±3 kg, BFM - 13.8±4 kg. Lean balance showed following values for: RA - 2.45±0.2 kg, LA - 2.37±0.4, T - 20.9±5 kg, RL - 7.43±1 kg, and LL - 7.49 ±1.5 kg. SMM showed statistical difference between manual obtained value, 51±01% to BCA parameter 45.5±3% (p<0.001). Manual obtained values for BFM was lower (17±2%) than BCA obtained one, 19.5±5.9% (p<0.02). Discussion: The obtained results showed appropriate values for the examined age, regarding to all examined parameters which contribute to overview the body compartments, important for sport performing. Due to comparison between the manual and BCA assessment, we may conclude that manual measurements may differ from the certain ones, which is confirmed by statistical significance.

Keywords: athletes, body composition, bio electrical impedance, sports medicine

Procedia PDF Downloads 479

Authors: Azza A. Ali, Doaa M. Abd El-Latif, Amany M. Gad, Yasser M. A. Elnahas, Karema Abu-Elfotuh

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Background: Exposure to Aluminium (Al) has been increased recently. It is found in food products, food additives, drinking water, cosmetics and medicines. Chronic consumption of Al causes oxidative stress and has been implicated in several chronic disorders. Liver is considered as the major site for detoxification while kidney is involved in the elimination of toxic substances and is a target organ of metal toxicity. Social isolation (SI) or protein malnutrition (PM) also causes oxidative stress and has negative impact on Al-induced nephrotoxicity as well as hepatotoxicity. Coenzyme Q10 (CoQ10) is a powerful intracellular antioxidant with mitochondrial membrane stabilizing ability while wheat grass is a natural product with antioxidant, anti-inflammatory and different protective activities, cocoa is also potent antioxidants and can protect against many diseases. They provide different degrees of protection from the impact of oxidative stress. Objective: To study the impact of social isolation together with Protein malnutrition on nephro- and hepato-toxicity induced by chronic Al exposure in rats as well as to investigate the postulated protection using a combination of Co Q10, wheat grass and cocoa. Methods: Eight groups of rats were used; four served as protected groups and four as un-protected. Each of them received daily for five weeks AlCl3 (70 mg/kg, IP) for Al-toxicity model groups except one group served as control. Al-toxicity model groups were divided to Al-toxicity alone, SI- associated PM (10% casein diet) and Al- associated SI&PM groups. Protection was induced by oral co-administration of CoQ10 (200mg/kg), wheat grass (100mg/kg) and cocoa powder (24mg/kg) combination together with Al. Biochemical changes in total bilirubin, lipids, cholesterol, triglycerides, glucose, proteins, creatinine and urea as well as alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), lactate deshydrogenase (LDH) were measured in serum of all groups. Specimens of kidney and liver were used for assessment of oxidative parameters (MDA, SOD, TAC, NO), inflammatory mediators (TNF-α, IL-6β, nuclear factor kappa B (NF-κB), Caspase-3) and DNA fragmentation in addition to evaluation of histopathological changes. Results: SI together with PM severely enhanced nephro- and hepato-toxicity induced by chronic Al exposure. Co Q10, wheat grass and cocoa combination showed clear protection against hazards of Al exposure either alone or when associated with SI&PM. Their protection were indicated by the significant decrease in Al-induced elevations in total bilirubin, lipids, cholesterol, triglycerides, glucose, creatinine and urea levels as well as ALT, AST, ALP, LDH. Liver and kidney of the treated groups also showed significant decrease in MDA, NO, TNF-α, IL-6β, NF-κB, caspase-3 and DNA fragmentation, together with significant increase in total proteins, SOD and TAC. Biochemical results were confirmed by the histopathological examinations. Conclusion: SI together with PM represents a risk factor in enhancing nephro- and hepato-toxicity induced by Al in rats. CoQ10, wheat grass and cocoa combination provide clear protection against nephro- and hepatotoxicity as well as the consequent degenerations induced by chronic Al-exposure even when associated with the risk of SI together with PM.

Keywords: aluminum, nephrotoxicity, hepatotoxicity, isolation and protein malnutrition, coenzyme Q10, wheatgrass, cocoa, nutrients combinations

Procedia PDF Downloads 252

Authors: Kuladip Jana, Bhaswati Banerjee, Parimal C. Sen

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Benzo(a)pyrene [B(a)P] is an environmental toxicant present mostly in cigarette smoke and car exhaust, is an aryl hydrocarbon receptor (AhR) ligand that exerts its toxic effects on both male and female reproductive systems along with carcinogenesis in skin, prostate, ovary, lung and mammary glands. Our study was focused on elucidating the molecular mechanism of B(a)P induced male reproductive toxicity and its prevention with phytochemical like resveratrol. In this study, the effect of B(a)P at different doses (0.1, 0.25, 0.5, 1 and 5 mg /kg body weight) was studied on male reproductive system of Wistar rat. A significant decrease in cauda epididymal sperm count and motility along with the presence of sperm head abnormalities and altered epididymal and testicular histology were documented following B(a)P treatment. B(a)P treatment resulted apoptotic sperm cells as observed by TUNEL and Annexin V-PI assay with increased Reactive Oxygen Species (ROS), altered sperm mitochondrial membrane potential (ΔΨm) with a simultaneous decrease in the activity of antioxidant enzymes and GSH status. TUNEL positive apoptotic cells also observed in testis as well as isolated germ and Leydig cells following B(a)P exposure. Western Blot analysis revealed the activation of p38 mitogen activated protein kinase (p38MAPK), cytosolic translocation of cytochrome-c, upregulation of Bax and inducible nitric oxide synthase (iNOS) with cleavage of poly ADP ribose polymerase (PARP) and down regulation of BCl2 in testis upon B(a)P treatment. The protein and mRNA levels of testicular key steroidogenesis regulatory proteins like steroidogenic acute regulatory protein (StAR), cytochrome P450 IIA1 (CYPIIA1), 3β hydroxy steroid dehydrogenase (3β HSD), 17β hydroxy steroid dehydrogenase (17β HSD) showed a significant decrease in a dose dependent manner while an increase in the expression of cytochrome P450 1A1 (CYP1A1), Aryl hydrocarbon Receptor (AhR), active caspase- 9 and caspase- 3 following B(a)P exposure. We conclude that exposure of benzo(a)pyrene caused testicular gamatogenic and steroidogenic disorders by induction of oxidative stress, inhibition of StAR and other steroidogenic enzymes along with activation of p38MAPK and initiated caspase-3 mediated germ and Leydig cell apoptosis. Next we investigated the role of resveratrol on B(a)P induced male reproductive toxicity. Our study highlighted that resveratrol co-treatment with B(a)P maintained testicular redox potential, increased serum testosterone level and prevented steroidogenic dysfunction with enhanced expression of major testicular steroidogenic proteins (CYPIIA1, StAR, 3β HSD,17β HSD) relative to treatment with B(a)P only. Resveratrol suppressed B(a)P-induced testicular activation of p38 MAPK, ATF2, iNOS and ROS production; cytosolic translocation of Cytochome c and Caspase 3 activation thereby prevented oxidative stress of testis and inhibited apoptosis. Resveratrol co-treatment also decreased B(a)P-induced AhR protein level, its nuclear translocation and subsequent CYP1A1 promoter activation, thereby decreased protein and mRNA levels of testicular cytochrome P4501A1 (CYP1A1) and prevented BPDE-DNA adduct formation. Our findings cumulatively suggest that resveratrol prevents activation of B(a)P by modulating the transcriptional regulation of CYP1A1 and acting as an antioxidant thus prevents B(a)P-induced oxidative stress and testicular apoptosis.

Keywords: benzo(a)pyrene, resveratrol, testis, apoptosis, cytochrome P450 1A1 (CYP1A1), aryl hydrocarbon receptor (AhR), p38 MAPK/ATF2/iNOS

Procedia PDF Downloads 238

Authors: Marin Kanarev, Delyan Stoyanov, Ivanna Popova, Nadezhda Petrova

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Thanks to increased survival rates in patients bearing oncological malignancies due to recent developments in anti-cancer therapies and diagnostic techniques, observation of clinical cases of metachronous cancers is more common and can provide more in-depth knowledge of their development and, as a result, help clinicians apply suitable therapy. This unusual case of three metachronous tumors presented the opportunity to follow their occurrence, progression, and treatment thoroughly. A 77-year-old male presented with carcinoma ventriculi of the pylorus region, which was surgically removed via upper subtotal stomach resection, a lateral antecolical gastro-enteroanastomosis, and a subsequent Braun anastomosis. An EOX chemotherapy regimen followed. A CT scan four months later showed no indication of recurrence or dissemination. The same scan, performed as a part of the follow-up plan two years later, showed an indication of neoplastic growth in the urinary bladder. After the patient had been directed to a urologist, the suspicion was confirmed, and the growth was histologically diagnosed as a carcinoma of the urinary bladder. An immunohistochemistry test showed an expression of PDL1 of less than 5%, which resulted in treatment with GemCis chemotherapy regimen that led to full remission. Two years and seven months after the first surgery, a CT scan showed again that the two carcinomas were gone. However, four months later, elevated tumor markers prompted a PET/CT scan, which showed data indicative of recurring neoplastic growth in the region of the stomach cardia. It was diagnosed as an adenocarcinoma infiltrating the esophagus. Preoperative chemotherapy with the ECF regimen was completed in four courses, and a CT scan showed no progression of the disease. In less than a month after therapy, the patient underwent laparotomy, debridement, gastrectomy, and a subsequent mechanical terminal-lateral esophago-jejunoanasthomosis. It was verified that the tumor originated from metastasis from the carcinoma ventriculi, which was located in the pylorus. In conclusion, this case report highlights the importance of patient follow-up and studying recurring neoplastic growth. Despite the absence of symptoms, clinicians should maintain a high level of suspicion when evaluating the patient data and choosing the most suitable therapy.

Keywords: carcinoma, follow-up, metachronous, neoplastic growth, recurrence

Procedia PDF Downloads 92

Authors: Danna Jia, Bin Li

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Background: Atopic dermatitis (AD) is a chronic and refractory inflammatory skin disease characterized by relapsing eczematous and pruritic skin lesions. The global prevalence of AD ranges from 1~ 20%, and its incidence rates are increasing. It affects individuals from infancy to adulthood, significantly impacting their daily lives and social activities. Despite its major health burden, the precise mechanisms underlying AD remain unknown. Understanding the genetic differences associated with AD is crucial for advancing diagnosis and targeted treatment development. This study aims to identify candidate genes of AD by using bioinformatics analysis. Methods: We conducted a comprehensive analysis of four pooled transcriptomic datasets (GSE16161, GSE32924, GSE130588, and GSE120721) obtained from the Gene Expression Omnibus (GEO) database. Differential gene expression analysis was performed using the R statistical language. The differentially expressed genes (DEGs) between AD patients and normal individuals were functionally analyzed using Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment. Furthermore, a protein-protein interaction (PPI) network was constructed to identify candidate genes. Results: Among the patient-level gene expression datasets, we identified 114 shared DEGs, consisting of 53 upregulated genes and 61 downregulated genes. Functional analysis using GO and KEGG revealed that the DEGs were mainly associated with the negative regulation of transcription from RNA polymerase II promoter, membrane-related functions, protein binding, and the Human papillomavirus infection pathway. Through the PPI network analysis, we identified eight core genes: CD44, STAT1, HMMR, AURKA, MKI67, and SMARCA4. Conclusion: This study elucidates key genes associated with AD, providing potential targets for diagnosis and treatment. The identified genes have the potential to contribute to the understanding and management of AD. The bioinformatics analysis conducted in this study offers new insights and directions for further research on AD. Future studies can focus on validating the functional roles of these genes and exploring their therapeutic potential in AD. While these findings will require further verification as achieved with experiments involving in vivo and in vitro models, these results provided some initial insights into dysfunctional inflammatory and immune responses associated with AD. Such information offers the potential to develop novel therapeutic targets for use in preventing and treating AD.

Keywords: atopic dermatitis, bioinformatics, biomarkers, genes

Procedia PDF Downloads 87

Authors: Huimin Zhao, Xiaoquan Liu, Haochen Liu

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The major challenge facing Alzheimer's Disease (AD) drug development is how to effectively improve cognitive function in clinical practice. Growing evidence indicates that stimulating hippocampal neurogenesis is a strategy for restoring cognition in animal models of AD. The mitogen-activated protein kinase (MAPK) pathway is a crucial factor in neurogenesis, which is negatively regulated by Dual-specificity phosphatase 16 (DUSP16). Transcriptome analysis of post-mortem brain tissue revealed up-regulation of DUSP16 expression in AD patients. Additionally, DUSP16 was involved in regulating the proliferation and neural differentiation of neural progenitor cells (NPCs). Nevertheless, whether the effect of DUSP16 on ameliorating cognitive disorders by influencing NPCs differentiation in AD mice remains unclear. Our study demonstrates an association between DUSP16 SNPs and clinical progression in individuals with mild cognitive impairment (MCI). Besides, we found that increased DUSP16 expression in both 3×Tg and SAMP8 models of AD led to NPC differentiation impairments. By silencing DUSP16, cognitive benefits, the induction of AHN and synaptic plasticity, were observed in AD mice. Furthermore, we found that DUSP16 is involved in the process of NPC differentiation by regulating c-Jun N-terminal kinase (JNK) phosphorylation. Moreover, the increased DUSP16 may be regulated by the ETS transcription factor (ELK1), which binds to the promoter region of DUSP16. Loss of ELK1 resulted in decreased DUSP16 mRNA and protein levels. Our data uncover a potential regulatory role for DUSP16 in adult hippocampal neurogenesis and provide a possibility to find the target of AD intervention.

Keywords: alzheimer's disease, cognitive function, DUSP16, hippocampal neurogenesis

Procedia PDF Downloads 75

Authors: B. Cilek Tatar, G. Sumnu, M. Oztop, E. Ayaz

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Encapsulation protects sensitive food ingredients against heat, oxygen, moisture and pH until they are released to the system. It can mask the unwanted taste of nutrients that are added to the foods for fortification purposes. Cherry laurels (Prunus laurocerasus) contain phenolic compounds which decrease the proneness to several chronic diseases such as types of cancer and cardiovascular diseases. The objective of this research was to study the effects of centrifugation, different coating materials and homogenization methods on microencapsulation of powders obtained from cherry laurels. In this study, maltodextrin and mixture of maltodextrin:whey protein with a ratio of 1:3 (w/w) were chosen as coating materials. Total solid content of coating materials was kept constant as 10% (w/w). Capsules were obtained from powders of freeze-dried cherry laurels through encapsulation process by silent crusher homogenizer or microfluidization. Freeze-dried cherry laurels were core materials and core to coating ratio was chosen as 1:10 by weight. To homogenize the mixture, high speed homogenizer was used at 4000 rpm for 5 min. Then, silent crusher or microfluidizer was used to complete encapsulation process. The mixtures were treated either by silent crusher for 1 min at 75000 rpm or microfluidizer at 50 MPa for 3 passes. Freeze drying for 48 hours was applied to emulsions to obtain capsules in powder form. After these steps, dry capsules were grounded manually into a fine powder. The microcapsules were analyzed for total antioxidant activity with DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging method. Prior to high speed homogenization, the samples were centrifuged (4000 rpm, 1 min). Centrifugation was found to have positive effect on total antioxidant activity of capsules. Microcapsules treated by microfluidizer were found to have higher total antioxidant activities than those treated by silent crusher. It was found that increasing whey protein concentration in coating material (using maltodextrin:whey protein 1:3 mixture) had positive effect on total antioxidant activity for both silent crusher and microfluidization methods. Therefore, capsules prepared by microfluidization of centrifuged mixtures can be selected as the best conditions for encapsulation of cherry laurel powder by considering their total antioxidant activity. In this study, it was shown that capsules prepared by these methods can be recommended to be incorporated into foods in order to enhance their functionality by increasing antioxidant activity.

Keywords: antioxidant activity, cherry laurel, microencapsulation, microfluidization

Procedia PDF Downloads 297

Authors: A. Pornpitakdumrong

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The aim of this research study was to develop recipe of Karanda ice cream as healthy promoting ice cream by high protein, low fat and naturally raw material, which found in local area. The results were found that appropriate condition for Karanda ice cream including incubation period, temperature and frozen time, which were 8-12 hours, -20 to -25 °C and 2-4 hours, respectively. Small fruit variety Karanda should selected only ripe fruits for Karanda ice cream made. Because of unripe fruits were contained resin and need to be air dried for reducing level of resin. Therefore, large fruit variety Karanda can be use both ripe and unripe fruits for Karanda ice cream made by without any astringent and bitter taste. However, small fruit variety Karanda was proper to made ice cream for trade, because occurring of industry to select the ripe fruits and commercially frozen, which be providing for the whole year compared with large variety fruits were rarely, low harvesting amount and short shelf life. Karanda ice cream produced from flesh part was attractive but was not accepted by consumers. It may due to resin contained with Karanda pulp, which led to be rough texture of ice cream. We were choose only Karanda juice, which was more appropriated and used Karanda juice with water by 1:1 ratio, because undiluted juice was sour taste. Most acceptance recipe of karanda ice cream product was sixth recipe by 91% of consumers, which was contained soy protein to made ice cream was delicate and swell, milk powder (little amount) to made ice cream was greasy, corn powder as stabilizer and undiluted coconut milk (little amount) to improve ice cream odor and similar to apricot odor.

Keywords: karonda fruits, Carissa carandas Linn, ice cream, healthy ice cream

Procedia PDF Downloads 412

Authors: Hazrat Salman Sidique, Muhammad Tahir Khan, Haq Aman Ullah, Muhammad Mobashar, Muhammad Ishtiaq Sohail Mehmood

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The poor quality conventional feed for the livestock production in Pakistan are wheat straw, tops of sugar cane and tree leaves. To enhance the nutritive value of feed, this study focused on investigating the effects of fibrolytic enzyme (Fibrozyme®, Alltech Inc. Company, USA) at different levels i.e. 0, 5, 10, and 15g/kg of total mix ration on feed intake, digestibility, milk yield and composition, and economics of the ration in Holstein Friesians cows. Twelve Holstein Friesians cows of almost the same age, and lactation stage were randomly allocated into 4 equal groups i.e. A, B, C, and D. Four experimental rations supplemented with Fibrozyme® 0g, 5g, 10g, and 15g/Kg of total mix ration were assigned to these sets correspondingly. The dry matter intake was linearly and significantly (P<0.05) improved. A significant effect of Fibrozyme® was observed for organic matter digestibility, ether extract digestibility, crude fiber digestibility, nitrogen free extract digestibility, and acid detergent fiber digestibility while the results were statistically non-significant for crude protein digestibility, neutral detergent fiber digestibility, and ash digestibility. Milk yield and composition except fat were significantly (P<0.05) increased in all Fibrozyme® treated groups. This study concludes that supplementation of Fibrozyme® at the rate of 15g/Kg total mix ration improved the dry matter intake, nutrients digestibility, and milk production and constituents like protein, lactose, and solid not fat. Therefore, treatment of total mix ration with Fibrozyme® was desirably reasonable and profitable.

Keywords: digestibility, fibrozyme, TMR, digestibility, lactating cow

Procedia PDF Downloads 114

Authors: Lalita Subedi, Jae Kyoung Chae, Yong Un Park, Cho Kyo Hee, Lee Jae Hyuk, Kang Min Cheol, Sun Yeou Kim

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Neuroinflammation may mediate the relationship between low levels of estrogens and neurodegenerative disease. Estrogens are neuroprotective and anti-inflammatory in neurodegenerative disease models. Due to the long term side effects of estrogens, researches have been focused on finding an effective phytoestrogens for biological activities. Daidzein present in soybeans and its active metabolite equol (7-hydroxy-3-(4'-hydroxyphenyl)-chroman) bears strong antioxidant and anticancer showed more potent anti-inflammatory and neuroprotective role in neuroinflammatory model confirmed its in vitro activity with molecular mechanism through NF-κB pathway. Three major CNS cells Microglia (BV-2), Astrocyte (C6), Neuron (N2a) were used to find the effect of equol in inducible nitric oxide synthase (iNOS), cyclooxygenase (COX-2), MAPKs signaling proteins, apoptosis related proteins by western blot analysis. Nitric oxide (NO) and prostaglandin E2 (PGE2) was measured by the Gries method and ELISA, respectively. Cytokines like tumor necrosis factor-α (TNF-α) and IL-6 were also measured in the conditioned medium of LPS activated cells with or without equol. Equol inhibited the NO production, PGE-2 production and expression of COX-2 and iNOS in LPS-stimulated microglial cells at a dose dependent without any cellular toxicity. At the same time Equol also showed promising effect in modulation of MAPK’s and nuclear factor kappa B (NF-κB) expression with significant inhibition of the production of proinflammatory cytokine like interleukin -6 (IL-6), and tumor necrosis factor -α (TNF-α). Additionally, it inhibited the LPS activated microglia-induced neuronal cell death by downregulating the apoptotic phenomenon in neuronal cells. Furthermore, equol increases the production of neurotrophins like NGF and increase the neurite outgrowth as well. In conclusion the natural daidzein metabolite equol are more active than daidzein, which showed a promising effectiveness as an anti-neuroinflammatory and neuroprotective agent via downregulating the LPS stimulated microglial activation and neuronal apoptosis. This work was supported by Brain Korea 21 Plus project and High Value-added Food Technology Development Program 114006-4, Ministry of Agriculture, Food and Rural Affairs.

Keywords: apoptosis, equol, neuroinflammation, phytoestrogen

Procedia PDF Downloads 363

Authors: Yamina Boukari, Nashiru Billa, Andrew Morris, Stephen Doughty, Kevin Shakesheff

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Poly (lactic-co-glycolic) acid (PLGA) is a synthetic polymer that can be used in bone tissue engineering with the aim of creating a scaffold in order to support the growth of cells. The formation of microspheres from this polymer is an attractive strategy that would allow for the development of an injectable system, hence avoiding invasive surgical procedures. The aim of this study was to develop a microsphere delivery system for use as an injectable scaffold in bone tissue engineering and evaluate various formulation parameters on its properties. Porous and lysozyme-containing PLGA microspheres were prepared using the double emulsion solvent evaporation method from various molecular weights (MW). Scaffolds were formed by sintering to contain 1 -3mg of lysozyme per gram of scaffold. The mechanical and physical properties of the scaffolds were assessed along with the release of lysozyme, which was used as a model protein. The MW of PLGA was found to have an influence on microsphere size during fabrication, with increased MW leading to an increased microsphere diameter. An inversely proportional relationship was displayed between PLGA MW and mechanical strength of formed scaffolds across loadings for low, intermediate and high MW respectively. Lysozyme release from both microspheres and formed scaffolds showed an initial burst release phase, with both microspheres and scaffolds fabricated using high MW PLGA showing the lowest protein release. Following the initial burst phase, the profiles for each MW followed a similar slow release over 30 days. Overall, the results of this study demonstrate that lysozyme can be successfully incorporated into porous PLGA scaffolds and released over 30 days in vitro, and that varying the MW of the PLGA can be used as a method of altering the physical properties of the resulting scaffolds.

Keywords: bone, microspheres, PLGA, tissue engineering

Procedia PDF Downloads 427

Authors: Faheema Khan

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To investigate the ability of sensitive and tolerant genotype of Glycine max to adapt to a saline environment in a field, we examined the growth performance, water relation and activities of antioxidant enzymes in relation to photosynthetic rate, chlorophyll a fluorescence, photosynthetic pigment concentration, protein and proline in plants exposed to salt stress. Ten soybean genotypes (Pusa-20, Pusa-40, Pusa-37, Pusa-16, Pusa-24, Pusa-22, BRAGG, PK-416, PK-1042, and DS-9712) were selected and grown hydroponically. After 3 days of proper germination, the seedlings were transferred to Hoagland’s solution (Hoagland and Arnon 1950). The growth chamber was maintained at a photosynthetic photon flux density of 430 μmol m−2 s−1, 14 h of light, 10 h of dark and a relative humidity of 60%. The nutrient solution was bubbled with sterile air and changed on alternate days. Ten-day-old seedlings were given seven levels of salt in the form of NaCl viz., T1 = 0 mM NaCl, T2=25 mM NaCl, T3=50 mM NaCl, T4=75 mM NaCl, T5=100 mM NaCl, T6=125 mM NaCl, T7=150 mM NaCl. The investigation showed that genotype Pusa-24, PK-416 and Pusa-20 appeared to be the most salt-sensitive. genotypes as inferred from their significantly reduced length, fresh weight and dry weight in response to the NaCl exposure. Pusa-37 appeared to be the most tolerant genotype since no significant effect of NaCl treatment on growth was found. We observed a greater decline in the photosynthetic variables like photosynthetic rate, chlorophyll fluorescence and chlorophyll content, in salt-sensitive (Pusa-24) genotype than in salt-tolerant Pusa-37 under high salinity. Numerous primers were verified on ten soybean genotypes obtained from Operon technologies among which 30 RAPD primers shown high polymorphism and genetic variation. The Jaccard’s similarity coefficient values for each pairwise comparison between cultivars were calculated and similarity coefficient matrix was constructed. The closer varieties in the cluster behaved similar in their response to salinity tolerance. Intra-clustering within the two clusters precisely grouped the 10 genotypes in sub-cluster as expected from their physiological findings.Salt tolerant genotype Pusa-37, was further analysed by 2-Dimensional gel electrophoresis to analyse the differential expression of proteins at high salt stress. In the Present study, 173 protein spots were identified. Of these, 40 proteins responsive to salinity were either up- or down-regulated in Pusa-37. Proteomic analysis in salt-tolerant genotype (Pusa-37) led to the detection of proteins involved in a variety of biological processes, such as protein synthesis (12 %), redox regulation (19 %), primary and secondary metabolism (25 %), or disease- and defence-related processes (32 %). In conclusion, the soybean plants in our study responded to salt stress by changing their protein expression pattern. The photosynthetic, biochemical and molecular study showed that there is variability in salt tolerance behaviour in soybean genotypes. Pusa-24 is the salt-sensitive and Pusa-37 is the salt-tolerant genotype. Moreover this study gives new insights into the salt-stress response in soybean and demonstrates the power of genomic and proteomic approach in plant biology studies which finally could help us in identifying the possible regulatory switches (gene/s) controlling the salt tolerant genotype of the crop plants and their possible role in defence mechanism.

Keywords: glycine max, salt stress, RAPD, genomic and proteomic variability

Procedia PDF Downloads 426

Authors: Van-Ngoc-Cuong Le, Ngoc-Quy Le

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Approaches to the diagnosis of ectopic solid organs in the abdominopelvic cavity include Accessory liver lobe, Accessory spleens (ectopic splenic tissue), Wandering spleen, Ectopic pancreatic tissue, Ectopic kidney (Pancake kidney), Cryptorchidism (undescended testis, ectopic testis), Ectopic endometriosis. The application of diagnostic imaging techniques, of which magnetic resonance imaging is the most important, includes a clinical case study and reports. Ectopic organs and tumors are easy to confuse. This is a concern, as well as practical challenges encountered and solutions adopted in the fields of Image Analysis.

Keywords: ectopic, accessory, wandering, tumor

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Authors: Mina Rabipour, Elena Pallaske, Floyd Hassenrück, Rocio Rebollido-Rios

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Protein tyrosine kinases, including Lyn kinase of the Src family kinases (SFK), regulate cell proliferation, survival, and differentiation. Lyn kinase has been implicated in various cancers, positioning it as a promising therapeutic target. However, the conserved ATP-binding pocket across SFKs makes developing selective inhibitors challenging. This study aims to address this limitation by exploring the potential for allosteric modulation of Lyn kinase, focusing on how its structural dynamics and specific oncogenic mutations impact its conformation and function. To achieve this, we combined homology modeling, molecular dynamics simulations, and data science techniques to conduct microsecond-length simulations. Our approach allowed a detailed investigation into the interplay between Lyn’s catalytic and regulatory domains, identifying key conformational states involved in allosteric regulation. Additionally, we evaluated the structural effects of Dasatinib, a competitive inhibitor, and ATP binding on Lyn active conformation. Notably, our simulations show that cancer-related mutations, specifically I364L/N and E290D/K, shift Lyn toward an inactive conformation, contrasting with the active state of the wild-type protein. This may suggest how these mutations contribute to aberrant signaling in cancer cells. We conducted a dynamical network analysis to assess residue-residue interactions and the impact of mutations on the Lyn intramolecular network. This revealed significant disruptions due to mutations, especially in regions distant from the ATP-binding site. These disruptions suggest potential allosteric sites as therapeutic targets, offering an alternative strategy for Lyn inhibition with higher specificity and fewer off-target effects compared to ATP-competitive inhibitors. Our findings provide insights into Lyn kinase regulation and highlight allosteric sites as avenues for selective drug development. Targeting these sites may modulate Lyn activity in cancer cells, reducing toxicity and improving outcomes. Furthermore, our computational strategy offers a scalable approach for analyzing other SFK members or kinases with similar properties, facilitating the discovery of selective allosteric modulators and contributing to precise cancer therapies.

Keywords: lyn tyrosine kinase, mutation analysis, conformational changes, dynamic network analysis, allosteric modulation, targeted inhibition

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Authors: Bhawna Chandravanshi, Ramesh Bhonde

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In the present study, we focused on the improvisation of islet survival in hypoxia. The Islet-like cell aggregates (ICAs) derived from Wharton's jelly mesenchymal stem cells (WJ-MSC) were cultured with and without WJ-MSC for 48h in hypoxia and normoxia and tested for their direct trophic effect on β cell survival. The WJ MSCs themselves secreted insulin upon glucose challenge and expressed the pancreatic markers at both transcription and translational level (C-peptide, Insulin, Glucagon and Glut 2). Direct contact of MSCs with ICAs facilitate the highest viability under hypoxia as evidenced by fluorescein diacetate/propidium iodide and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The cytokine analysis of the co-cultured ICAs revealed amplification of anti-inflammatory cytokine-like TGFβ and TNFα accompanied by depletion of pro-inflammatory cytokines. The increment in VEGF and PDGFa was also seen showing their ability to vascularize upon transplantation. This was further accompanied by reduction in total reactive oxygen species, nitric oxide, and super oxide ions and down-regulation of Caspase3, Caspase8, p53 and up regulation of Bcl2 confirming prevention of apoptosis in ICAs. There was a significant reduction in the expression of p38 protein in the presence of MSCs making the ICAs responsive to glucose. Taken together our data demonstrate for the first time that the WJ-MSC expressed pancreatic markers and their supplementation protected engineered islets against hypoxia, oxidative stress, and inflammatory cytokines by inhibiting p38 MAPK protein.

Keywords: hypoxia, islet-like cell aggregates, inflammatory cytokines, oxidative stress

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Authors: M. Cabral, A. Verdin, G. Garçon, A. Touré, C. Diop, M. Fall, S. Bouhsina, D. Dewaele, F.Cazier, A. Tall Dia, P. Shirali, A. Diouf

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This case-control study dealt with the health adverse effects within the population neighboring the Mbeubeuss waste dump, which is located near the district of Malika (Diamalaye II) in Dakar (Senegal). All the household and industrial waste arising from Dakar are stored in this open landfill without being covered and are therefore possible sources of Pb and Cd contaminated air emissions and lixiviates. The objective of this study is part of improving the health of the population neighboring Mbeubeuss by determining Pb and Cd concentrations both in environment and humans, and studying possible renal function alterations within the adults. Soil and air samples were collected in the control site (Darou Salam) and the waste dump neighboring site (Diamalaye II). Control and exposed adults were recruited as living in Darou Salam (n = 52) and in Diamalaye II (n = 77). Pb and Cd concentrations in soil, air and biological samples were determined. Moreover, we were interested in analyzing some impregnation (zinc protoporphyrin, d-aminolevulinic acid dehydratase) and oxidative stress biomarkers (malonedialdehyde, gluthatione status), in addition to several nephrotoxicity parameters (creatinuria, proteinuria, lactate dehydrogenase, CC16 protein, glutathione S-transferase-alpha and retinol binding protein) in blood and/or urine. The results showed the significant Pb and Cd contamination of the soil and air samples derived from the landfill, and therefore of the neighboring population of adults. This critical exposure to environmental Pb and Cd had some harmful consequences for their health, as shown by the reported oxidative stress and nephrotoxicity signs.

Keywords: Pb and Cd environmental exposure, impregnation markers, landfill, nephrotoxicity markers

Procedia PDF Downloads 447

Authors: Yew Mun Yip, Dawei Zhang

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Proteins are the biological machinery that executes specific vital functions in every cell of the human body by folding into their 3D structures. When a protein misfolds from its native structure, the machinery will malfunction and lead to misfolding diseases. Although in vitro experiments are able to conclude that the mutations of the amino acid sequence lead to incorrectly folded protein structures, these experiments are unable to decipher the folding process. Therefore, molecular dynamic (MD) simulations are employed to simulate the folding process so that our improved understanding of the folding process will enable us to contemplate better treatments for misfolding diseases. MD simulations make use of force fields to simulate the folding process of peptides. Secondary structures are formed via the hydrogen bonds formed between the backbone atoms (C, O, N, H). It is important that the hydrogen bond energy computed during the MD simulation is accurate in order to direct the folding process to the native structure. Since the atoms involved in a hydrogen bond possess very dissimilar electronegativities, the more electronegative atom will attract greater electron density from the less electronegative atom towards itself. This is known as the polarization effect. Since the polarization effect changes the electron density of the two atoms in close proximity, the atomic charges of the two atoms should also vary based on the strength of the polarization effect. However, the fixed atomic charge scheme in force fields does not account for the polarization effect. In this study, we introduce the polarized structure-specific backbone charge (PSBC) model. The PSBC model accounts for the polarization effect in MD simulation by updating the atomic charges of the backbone hydrogen bond atoms according to equations derived between the amount of charge transferred to the atom and the length of the hydrogen bond, which are calculated from quantum-mechanical calculations. Compared to other polarizable models, the PSBC model does not require quantum-mechanical calculations of the peptide simulated at every time-step of the simulation and maintains the dynamic update of atomic charges, thereby reducing the computational cost and time while accounting for the polarization effect dynamically at the same time. The PSBC model is applied to two different β-peptides, namely the Beta3s/GS peptide, a de novo designed three-stranded β-sheet whose structure is folded in vitro and studied by NMR, and the trpzip peptides, a double-stranded β-sheet where a correlation is found between the type of amino acids that constitute the β-turn and the β-propensity.

Keywords: hydrogen bond, polarization effect, protein folding, PSBC

Procedia PDF Downloads 271

Authors: Liudmila Garzanova, Lidia Ananyeva, Olga Koneva, Olga Ovsyannikova, Oxana Desinova, Mayya Starovoytova, Rushana Shayahmetova, Anna Khelkovskaya-Sergeeva

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Objectives. C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) are associated with severe course, increased morbidity and mortality in systemic sclerosis (SSc). The aim of our study was to assess changes in CRP and ESR in SSc patients during long-term RTX therapy. Methods. This study included 113 patients with SSc. Mean age was 48.1±13 years, female-85%. The mean disease duration was 6±5 years. The diffuse cutaneous subset of the disease had 55% of patients. All pts had interstitial lung disease (ILD). All patients received prednisolone at a mean dose of 11.6±4.8 mg/day, and 53 of them - were immunosuppressants at inclusion. Patients received RTX due to the ineffectiveness of previous therapy for ILD. The parameters were evaluated over the periods: at baseline (point 0), 13±2.3 month (point 1, n=113), 42±14 month (point 2, n=80) and 79±6.5 month (point 3, n=25) after initiation of RTX therapy. Cumulative mean dose of RTX at point 1 = 1.7±0.6g, at point 2 = 3±1.5g, and at point 3 = 3.8±2.4g. The results are presented in the form of mean values, delta(Δ)-difference between the baseline parameter and follow-up point. Results. There was an improvement in studied parameters on RTX therapy. There was a significant decrease of ESR, CRP and activity index (EScSG-AI) at all observation points (p=0.001). In point 1: ΔCRP was 6.7 mg/l, ΔESR = 7.4 mm/h, ΔActivity index (EScSG-AI) = 1.7. In point 2: ΔCRP was 8.7 mg/l, ΔESR = 7.5 mm/h, ΔActivity index (EScSG-AI) = 1.9. In point 3: ΔCRP was 16.1 mg/l, ΔESR = 11 mm/h, ΔActivity index (EScSG-AI) = 2.1. Conclusion. There was a significant decrease in CRP and ESR during long-term RTX therapy, which correlated with a decrease in the disease activity index. RTX is an effective treatment option for SSc with an elevation of acute-phase reactants.

Keywords: C-reactive protein, interstitial lung disease, systemic sclerosis, rituximab

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Authors: Sandra Smieszek, Jonathan Haines

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Introduction: Numerous research efforts have focused on searching for ‘longevity genes’. However, attempting to decipher the genetic component of the longevous phenotype have resulted in limited success and the mechanisms governing longevity remain to be explained. We conducted a genome-wide homozygosity analysis (GWHA) of the founder population of the Amish community in central Ohio. While genome-wide association studies using unrelated individuals have revealed many interesting longevity associated variants, these variants are typically of small effect and cannot explain the observed patterns of heritability for this complex trait. The Amish provide a large cohort of extended kinships allowing for in depth analysis via family-based approach excellent population due to its. Heritability of longevity increases with age with significant genetic contribution being seen in individuals living beyond 60 years of age. In our present analysis we show that the heritability of longevity is estimated to be increasing with age particularly on the paternal side. Methods: The present analysis integrated both phenotypic and genotypic data and led to the discovery of a series of variants, distinct for stratified populations across ages and distinct for paternal and maternal cohorts. Specifically 5437 subjects were analyzed and a subset of 893 successfully genotyped individuals was used to assess CHIP heritability. We have conducted the homozygosity analysis to examine if homozygosity is associated with increased risk of living beyond 90. We analyzed AMISH cohort genotyped for 614,957 SNPs. Results: We delineated 10 significant regions of homozygosity (ROH) specific for the age group of interest (>90). Of particular interest was ROH on chromosome 13, P < 0.0001. The lead SNPs rs7318486 and rs9645914 point to COL4A2 and our lead SNP. COL25A1 encodes one of the six subunits of type IV collagen, the C-terminal portion of the protein, known as canstatin, is an inhibitor of angiogenesis and tumor growth. COL4A2 mutations have been reported with a broader spectrum of cerebrovascular, renal, ophthalmological, cardiac, and muscular abnormalities. The second region of interest points to IRS2. Furthermore we built a classifier using the obtained SNPs from the significant ROH region with 0.945 AUC giving ability to discriminate between those living beyond to 90 years of age and beyond. Conclusion: In conclusion our results suggest that a history of longevity does indeed contribute to increasing the odds of individual longevity. Preliminary results are consistent with conjecture that heritability of longevity is substantial when we start looking at oldest fifth and smaller percentiles of survival specifically in males. We will validate all the candidate variants in independent cohorts of centenarians, to test whether they are robustly associated with human longevity. The identified regions of interest via ROH analysis could be of profound importance for the understanding of genetic underpinnings of longevity.

Keywords: regions of homozygosity, longevity, SNP, Amish

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Authors: Mahzad Yousefi, Azin Tavakoli

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Periodontal disease is an inflammatory reaction; therefore, it is predicted that changes may occur in some inflammatory parameters that can be detected in routine blood tests. The objective of this study was to evaluate the hematological and biochemistry changes that occur in dogs affected with advanced stages of periodontal disease. 87 dogs were diagnosed with periodontal disease (PD group), and 76 healthy dogs entered the study. The PD dogs had been affected with periodontitis stage 3 or 4 and were candidates for any dental extractions. The healthy dogs were either referred for annual checkups or for issuing health travel certificates that their owners asked for complete lab tests. Neither the diseased nor healthy subjects had a history of infectious, or other general health problems or surgery in the past 3 months. Age, as well as all hematologic including PCV, WBC and RBC count, Hb, MCV, MCH, MCHC, PLT, CBC, NLR, and biochemistry data, including total protein, albumin, glucose, BUN, Creatinine, ALT, AST, and ALP, were recorded and analyzed statistically. Results confirmed that aging has a significant direct relationship with the advanced stages of periodontal disease. Mild leukocytosis occurred in the diseased group; however, it was not significant. Also, the mean total protein of the PD group was lower than that of the healthy dogs, and serum levels of albumin were found to be lower significantly in the diseased group (P<0.05). Mean ±SD amount of Platelet, MCH, and ALT were significantly higher in the diseased group in comparison to the healthy dogs (P<0.05). No significant differences were reported in other evaluated parameters. It is concluded that CBC indices of PD dogs are not systemic inflammatory; however, only a decrease in albumin showed inflammatory responses. Some indices in routine laboratory tests can be changed significantly during advanced stages of the periodontal disease dogs.

Keywords: periodontal disease, dogs, hematological factors, advanced stages, blood tests

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Authors: M. Y. Abubakar, M. Yunisa, A. N. Muhammad

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Constraining the production Clarias gariepinus and Oreochromis niloticus culture is the prohibitive cost of feed. We assess the performance of the species fingerlings on diets substituted with composite. Four dietary treatments (0%, 25%, 45%, and 75%) for C. gariepinus and five (0%, 25%, 50%, 75%, and whole food composite) for O. niloticus were formulated and each fed to 15 fingerlings for C. gariepinus and 10 fingerlings for O. niloticus stocked in 75ltrs plastic bowls, replicated trice in a completely randomized design. The experiment lasted 56 days. Percent survival rate was significantly (p < 0.05) higher (57.78 ± 9.69) in C. gariepinus fed diet III. The growth and nutrient utilization indices were least in the fish fed diet IV, which was significantly (p < 0.05) lower than in other treatments. Fish fed dietary treatment III, recorded the best in growth and nutrient utilization indices and was significantly higher (p < 0.05) than those fed dietary treatments I & II which were non-significant (p > 0.05) and higher than those fed 75% substitution. Better profit index was in the fish fed diet with 50% substitution level. For O. niloticus, the survival (172.62 ± 39.03) was significantly higher (p < 0.05) in those fed 25% substituted diet. For growth indices, the least performed were those fed whole composite while other treatments were non-significant (p > 0.05) different from each other. In terms of nutrient utilization, fish fed diet substituted at 0%, 25%, 50% and 75% food composite had similar food conversion ratio and protein efficiency ratio. However, there was no significant difference in the profit index among the whole treatment. It can be concluded that food composite from Sokoto house-holds can optimally replace groundnut cake up to 50% level as a protein source in the diets of Clarias gariepinus and O. niloticus fingerlings without adverse effects on survival, growth, and nutrient utilization.

Keywords: food composite, nutrient utilization, C. gariepinus, O. niloticus household, substitution levels

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Authors: F. Yılmaz, Y. Saylan, A. Derazshamshir, S. Atay, A. Denizli

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Quartz crystal microbalance (QCM), high-resolution mass-sensing technique, measures changes in mass on oscillating quartz crystal surface by measuring changes in oscillation frequency of crystal in real time. Protein adsorption techniques via hydrophobic interaction between protein and solid support, called hydrophobic interaction chromatography (HIC), can be favorable in many cases. Some nanoparticles can be effectively applied for HIC. HIC takes advantage of the hydrophobicity of proteins by promoting its separation on the basis of hydrophobic interactions between immobilized hydrophobic ligands and nonpolar regions on the surface of the proteins. Lysozyme is found in a variety of vertebrate cells and secretions, such as spleen, milk, tears, and egg white. Its common applications are as a cell-disrupting agent for extraction of bacterial intracellular products, as an antibacterial agent in ophthalmologic preparations, as a food additive in milk products and as a drug for treatment of ulcers and infections. Lysozyme has also been used in cancer chemotherapy. The aim of this study is the synthesis of hydrophobic nanoparticles for Lysozyme detection. For this purpose, methacryoyl-L-phenylalanine was chosen as a hydrophobic matrix. The hydrophobic nanoparticles were synthesized by micro-emulsion polymerization method. Then, hydrophobic QCM nanosensor was characterized by Attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy, atomic force microscopy (AFM) and zeta size analysis. Hydrophobic QCM nanosensor was tested for real-time detection of Lysozyme from aqueous solution. The kinetic and affinity studies were determined by using Lysozyme solutions with different concentrations. The responses related to a mass (Δm) and frequency (Δf) shifts were used to evaluate adsorption properties.

Keywords: nanosensor, HIC, lysozyme, QCM

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Authors: Ved Vrat Verma, Rani Gupta, Manisha Goel

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γ-glutamyltranspeptidase (GGT: EC 2.3.2.2) is an N-terminal nucleophile hydrolase conserved in all three domains of life. GGT plays a key role in glutathione metabolism where it catalyzes the breakage of the γ-glutamyl bonds and transfer of γ-glutamyl group to water (hydrolytic activity) or amino acids or short peptides (transpeptidase activity). GGTs from bacteria, archaea, and eukaryotes (human, rat and mouse) are homologous proteins sharing >50% sequence similarity and conserved four layered αββα sandwich like three dimensional structural fold. These proteins though similar in their structure to each other, are quite diverse in their enzyme activity: some GGTs are better at hydrolysis reactions but poor in transpeptidase activity, whereas many others may show opposite behaviour. GGT is known to be involved in various diseases like asthma, parkinson, arthritis, and gastric cancer. Its inhibition prior to chemotherapy treatments has been shown to sensitize tumours to the treatment. Microbial GGT is known to be a virulence factor too, important for the colonization of bacteria in host. However, all known inhibitors (mimics of its native substrate, glutamate) are highly toxic because they interfere with other enzyme pathways. However, a few successful efforts have been reported previously in designing species specific inhibitors. We aim to leverage the diversity seen in GGT family (pathogen vs. eukaryotes) for designing specific inhibitors. Thus, in the present study, we have used DIVERGE software to identify sites in GGT proteins, which are crucial for the functional and structural divergence of these proteins. Since, type II divergence sites vary in clade specific manner, so type II divergent sites were our focus of interest throughout the study. Type II divergent sites were identified for pathogen vs. eukaryotes clusters and sites were marked on clade specific representative structures HpGGT (2QM6) and HmGGT (4ZCG) of pathogen and eukaryotes clade respectively. The crucial divergent sites within 15 A radii of the binding cavity were highlighted, and in-silico mutations were performed on these sites to delineate the role of these sites on the mechanism of catalysis and protein folding. Further, the amino acid network (AAN) analysis was also performed by Cytoscape to delineate assortative mixing for cavity divergent sites which could strengthen our hypothesis. Additionally, molecular dynamics simulations were performed for wild complexes and mutant complexes close to physiological conditions (pH 7.0, 0.1 M ionic strength and 1 atm pressure) and the role of putative divergence sites and structural integrities of the homologous proteins have been analysed. The dynamics data were scrutinized in terms of RMSD, RMSF, non-native H-bonds and salt bridges. The RMSD, RMSF fluctuations of proteins complexes are compared, and the changes at protein ligand binding sites were highlighted. The outcomes of our study highlighted some crucial divergent sites which could be used for novel inhibitors designing in a species-specific manner. Since, for drug development, it is challenging to design novel drug by targeting similar protein which exists in eukaryotes, so this study could set up an initial platform to overcome this challenge and help to deduce the more effective targets for novel drug discovery.

Keywords: γ-glutamyltranspeptidase, divergence, species-specific, drug design

Procedia PDF Downloads 273

Authors: Niharika Bala, Alaa Habib, Marianne Kozuch, Nancy Denslow, Sarah Glover, Abdel Alli

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The Coronavirus disease 2019 (COVID-19) is a viral infection caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and associated with systemic inflammation. Inflammation is an important process that follows infection and facilitates the repair of damaged tissue. Polyunsaturated fatty acids play an important role in the inflammatory process. These lipids can target transcription factors to modulate gene expression and protein function. Here, we evaluated whether differences in basal levels of different types of biomarkers can be detected in freshly frozen plasma samples from patients with and without COVID-19. Fatty acid methyl ester (FAME) analysis showed a decrease in arachidic acid and myristic acid but an increase in caprylic acid, palmitic acid, and eicosenoic acid in the plasma of COVID-19 patients compared to non-COVID19 patients. Multiple chemokines, including IP-10, MCP-1, and MIP-1 beta, were increased in the COVID-19 group compared to the non-COVID-19 group. Similarly, cytokines, including IL-1 alpha and IL-8, and cell adhesion and inflammatory response markers, including ICAM-1 and E-selectin, were greater in the plasma of COVID-19 patients compared to non-COVID-19 patients. A baseline signature of specific polyunsaturated fatty acids, cytokines, and chemokines present in the plasma after COVID-19 viral infection may serve as biomarkers that can be useful in various applications, including determination of severity of infection, indication of disease prognosis, and consideration for therapeutic options.

Keywords: SARS-CoV-2, COVID-19, fatty acids, inflammation, cytokines, chemokines

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Authors: Shreya Ghosh, Akansha Garg, Chayanika Kala, Ashwani Kumar Thakur

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Background: Granuloma formation is one of the characteristic features of tuberculosis. Besides, chronic inflammation underlying tuberculosis is often indicated by an increase in the concentration of serum amyloid A (SAA) protein. The connection between tuberculosis and SAA-driven secondary amyloidosis is well documented. However, SAA-derived amyloid deposition start sites are not well understood in tuberculosis and other chronic inflammatory conditions. It was hypothesized that granuloma could be a potential site for an amyloid deposition because both SAA protein and proteases that cleave SAA into aggregation-prone fragments are reported to be present in the granuloma. Here the authors have shown the presence of SAA-derived amyloid deposits in the granuloma of tuberculosis patients. Methodology: Over a period of two years, tuberculosis patients were screened, and biopsies were collected from the affected organs of the patients. The gold standard, Congo red dye staining, was used to identify amyloid deposits in the tissue sections of tuberculosis patients containing granulomatous structure. Results: 11 out of 150 FFPE biopsy specimens of tuberculosis patients showed eosinophilic hyaline-rich deposits surrounding granuloma. Upon Congo red staining, these deposits exhibited characteristic apple-green birefringence under polarized light, confirming amyloid deposits. Further, upon immunohistochemical staining with anti-SAA, the amyloid enriched areas showed positive immunoreactivity. Conclusion: In this pilot study, we have shown that granuloma can be a potential site for serum amyloid A-derived amyloid formation in tuberculosis patients. Moreover, the presence of amyloid gave significant cues that granuloma might be a probable amyloid deposition start in tuberculosis patients. This study will set a stage to expand the clinical and fundamental research in the understanding of amyloid formation in granuloma underlying tuberculosis and chronic inflammatory conditions.

Keywords: amyloid, granuloma, periphery, serum amyloid A, tuberculosis

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Authors: D. F. Carvalho, A. O. Uscamayta, J. C. Guerrero, H. F. Oliveira, P. M. Azevedo-Marques

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The creation of vector contours slices (ROIs) on body silhouettes in oncologic patients is an important step during the radiotherapy planning in clinic and hospitals to ensure the accuracy of oncologic treatment. The radiotherapy planning of patients is performed by complex softwares focused on analysis of tumor regions, protection of organs at risk (OARs) and calculation of radiation doses for anomalies (tumors). These softwares are supplied for a few manufacturers and run over sophisticated workstations with vector processing presenting a cost of approximately twenty thousand dollars. The Brazilian project SIPRAD (Radiotherapy Planning System) presents a proposal adapted to the emerging countries reality that generally does not have the monetary conditions to acquire some radiotherapy planning workstations, resulting in waiting queues for new patients treatment. The SIPRAD project is composed by a set of integrated and interoperabilities softwares that are able to execute all stages of radiotherapy planning on simple personal computers (PCs) in replace to the workstations. The goal of this work is to present an image processing technique, computationally feasible, that is able to perform an automatic contour delineation in patient body silhouettes (SIPRAD-Body). The SIPRAD-Body technique is performed in tomography slices under grayscale images, extending their use with a greedy algorithm in three dimensions. SIPRAD-Body creates an irregular polyhedron with the Canny Edge adapted algorithm without the use of preprocessing filters, as contrast and brightness. In addition, comparing the technique SIPRAD-Body with existing current solutions is reached a contours similarity at least 78%. For this comparison is used four criteria: contour area, contour length, difference between the mass centers and Jaccard index technique. SIPRAD-Body was tested in a set of oncologic exams provided by the Clinical Hospital of the University of Sao Paulo (HCRP-USP). The exams were applied in patients with different conditions of ethnology, ages, tumor severities and body regions. Even in case of services that have already workstations, it is possible to have SIPRAD working together PCs because of the interoperability of communication between both systems through the DICOM protocol that provides an increase of workflow. Therefore, the conclusion is that SIPRAD-Body technique is feasible because of its degree of similarity in both new radiotherapy planning services and existing services.

Keywords: radiotherapy, image processing, DICOM RT, Treatment Planning System (TPS)

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Authors: Caroline Mendes, Mary McNamara, Orla Howe

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For chemotherapy, a drug delivery system should be able to specifically target cancer cells and deliver the therapeutic dose without affecting normal cells. Folate receptors (FR) can be considered key targets since they are commonly over-expressed in cancer cells and they are the molecular marker used in this study. Here, cyclodextrin (CD) has being studied as a vehicle for delivering the chemotherapeutic drug, methotrexate (MTX). CDs have the ability to form inclusion complexes, in which molecules of suitable dimensions are included within the CD cavity. In this study, β-CD has been modified using folic acid so as to specifically target the FR molecular marker. Thus, the system studied here for drug delivery consists of β-CD, folic acid and MTX (CDEnFA:MTX). Cellular uptake of folic acid is mediated with high affinity by folate receptors while the cellular uptake of antifolates, such as MTX, is mediated with high affinity by the reduced folate carriers (RFCs). This study addresses the gene (mRNA) and protein expression levels of FRs and RFCs in the cancer cell lines CaCo-2, SKOV-3, HeLa, MCF-7, A549 and the normal cell line BEAS-2B, quantified by real-time polymerase chain reaction (real-time PCR) and flow cytometry, respectively. From that, four cell lines with different levels of FRs, were chosen for cytotoxicity assays of MTX and CDEnFA:MTX using the MTT assay. Real-time PCR and flow cytometry data demonstrated that all cell lines ubiquitously express moderate levels of RFC. These experiments have also shown that levels of FR protein in CaCo-2 cells are high, while levels in SKOV-3, HeLa and MCF-7 cells are moderate. A549 and BEAS-2B cells express low levels of FR protein. FRs are highly expressed in all the cancer cell lines analysed when compared to the normal cell line BEAS-2B. The cell lines CaCo-2, MCF-7, A549 and BEAS-2B were used in the cell viability assays. 48 hours treatment with the free drug and the complex resulted in IC50 values of 93.9 µM ± 9.2 and 56.0 µM ± 4.0 for CaCo-2 for free MTX and CDEnFA:MTX respectively, 118.2 µM ± 10.8 and 97.8 µM ± 12.3 for MCF-7, 36.4 µM ± 6.9 and 75.0 µM ± 8.5 for A549 and 132.6 µM ± 12.1 and 288.1 µM ± 16.3 for BEAS-2B. These results demonstrate that MTX is more toxic towards cell lines expressing low levels of FR, such as the BEAS-2B. More importantly, these results demonstrate that the inclusion complex CDEnFA:MTX showed greater cytotoxicity than the free drug towards the high FR expressing CaCo-2 cells, indicating that it has potential to target this receptor, enhancing the specificity and the efficiency of the drug.

Keywords: cyclodextrins, cancer treatment, drug delivery, folate receptors, reduced folate carriers

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Authors: Uchegbu Nneka Nkechi, Okoye Ogochukwu Peace

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The evaluation of the visco-elastic properties and microbial quality of a formulated oat-based dietetic food were investigated. Oat flour, pumpkin seed flour, carrot flour and skimmed milk powder were blended in varying proportions to formulate a product with codes OCF, which contains 70% oat flour, 10 % carrot flour, 10 % pumpkin seed flour and 10% skimmed milk powder, OCF which contains 65 % oat flour, 10 % carrot flour, 10 % pumpkin seed flour and 15 % skimmed milk powder, OCF which contains 60 % oat flour, 10 % carrot flour, 10 % pumpkin seed flour and 20 % skimmed milk powder, OCF which contains 55 % oat flour, 10 % carrot flour, 10 % pumpkin seed flour and 25 % skimmed milk powder and OF with 95 % oat as the commercial control. All the samples were assessed for their proximate composition, microbial quality and visco-elastic properties. The moisture content was highest at sample OF (10.73%) and lowest at OCF (7.10%) (P<0.05). Crude protein ranged from 13.38%-22.86%, with OCF having the highest (P<0.05) protein content and OF having the lowest. Crude fat was 3.74% for OCF and 6.31% for OF. Crude fiber ranged from 3.58% - 17.39% with OF having the lowest (P<0.05) fiber content and OCF having the highest. Ash content was 1.30% for OCF and 2.75% for OCF. There was no mold growth in the samples. The total viable ml/wl count ranged from 1.5×10³ cfu/g - 2.6×10³ cfu/g, with OCF having the lowest and OF having the highest (P<0.05) total viable count. The peak viscosity of the sample ranged from 75.00 cP-2895.00 cP, with OCF having the lowest and OF having the highest value. The final viscosity was 130.50 cP in OCF and 3572.50 cP in OF. The setback viscosity was 58.00 cP in OCF and 1680.50 cP in OF. The peak time was 6.93 mins in OCF to 5.57 mins in OF. There was no pasting temperature for all samples except the OF, which had 86.43. Sample OF was the highest in terms of overall acceptability. This study showed that the oat-based composite flour produced had a nutritional profile that would be acceptable for the aged population.

Keywords: dietetic, pumpkin, visco-elastic, microbial

Procedia PDF Downloads 201