Search results for: extracellular proteins

Authors: Masoud Sakhinia, Sajjad Ahmad

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Introduction: Though knowledge of the compositional makeup and structure of the limbal niche has progressed exponentially during the past decade, much is yet to be understood. Identifying the precise profile and role of the stromal makeup which spans the ocular surface may inform researchers of the most optimum conditions needed to effectively expand LESCs in vitro, whilst preserving their differentiation status and phenotype. Limbal fibroblasts, as opposed to corneal fibroblasts are thought to form an important component of the microenvironment where LESCs reside. Methods: The corneal stroma was tissue engineered in vitro using both limbal and corneal fibroblasts embedded within a tissue engineered 3D collagen matrix. The effect of these two different fibroblasts on LESCs and hTCEpi corneal epithelial cell line were then subsequently determined using phase contrast microscopy, histolological analysis and PCR for specific stem cell markers. The study aimed to develop an in vitro model which could be used to determine whether limbal, as opposed to corneal fibroblasts, maintained the stem cell phenotype of LESCs and hTCEpi cell line. Results: Tissue culture analysis was inconclusive and required further quantitative analysis for remarks on cell proliferation within the varying stroma. Histological analysis of the tissue-engineered cornea showed a comparable structure to that of the human cornea, though with limited epithelial stratification. PCR results for epithelial cell markers of cells cultured on limbal fibroblasts showed reduced expression of CK3, a negative marker for LESC’s, whilst also exhibiting a relatively low expression level of P63, a marker for undifferentiated LESCs. Conclusion: We have shown the potential for the construction of a tissue engineered human cornea using a 3D collagen matrix and described some preliminary results in the analysis of the effects of varying stroma consisting of limbal and corneal fibroblasts, respectively, on the proliferation of stem cell phenotype of primary LESCs and hTCEpi corneal epithelial cells. Although no definitive marker exists to conclusively illustrate the presence of LESCs, the combination of positive and negative stem cell markers in our study were inconclusive. Though it is less traslational to the human corneal model, the use of conditioned medium from that of limbal and corneal fibroblasts may provide a more simple avenue. Moreover, combinations of extracellular matrices could be used as a surrogate in these culture models.

Keywords: cornea, Limbal Stem Cells, tissue engineering, PCR

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Authors: Rohit Singh Dangi, Ravi Kant Pal, Monica Sundd

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Acyl-CoA binding protein (ACBP) is a housekeeping protein which functions as an intracellular carrier of acyl-CoA esters. Given the fact that the amastigote stage (blood stage) of Leishmania depends largely on fatty acids as the energy source, of which a large part is derived from its host, these proteins might have an important role in its survival. In Leishmania major, genome sequencing suggests the presence of six ACBPs, whose function remains largely unknown. For functional and structural characterization, one of the ACBP genes was cloned, and the protein was expressed and purified heterologously. Acyl-CoA ester binding and stoichiometry were analyzed by isothermal titration calorimetry and Dynamic light scattering. Our results shed light on high affinity of ACBP towards longer acyl-CoA esters, such as myristoyl-CoA to arachidonoyl-CoA with single binding site. To understand the binding mechanism & dynamics, Nuclear magnetic resonance assignments of this protein are being done. The protein's crystal structure was determined at 1.5Å resolution and revealed a classical topology for ACBP, containing four alpha-helical bundles. In the binding pocket, the loop between the first and the second helix (16 – 26AA) is four residues longer from other extensively studied ACBPs (PfACBP) and it curls upwards towards the pantothenate moiety of CoA to provide a large tunnel space for long acyl chain insertion.

Keywords: acyl-coa binding protein (ACBP), acyl-coa esters, crystal structure, isothermal titration, calorimetry, Leishmania

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Authors: Kkochorong Park, Keun Cheon Kim, Hyoban Lee, Eun Ju Lee, Bongsoo Kim

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Introduction of biomaterials such as DNA, RNA, proteins is important for many research areas. There are many methods to introduce biomaterials into living organisms like tissue and cells. To introduce biomaterials, several indirect methods including virus‐mediated delivery, chemical reagent (i.e., lipofectamine), electrophoresis have been used. Such methods are passive delivery using an endocytosis process of cell, reducing an efficiency of delivery. Unlike the indirect delivery method, it has been reported that a direct delivery of exogenous biomolecules into nucleus have been more efficient to expression or integration of biomolecules. Nano-sized material is beneficial for detect signal from cell or deliver stimuli/materials into the cell at cellular and molecular levels, due to its similar physical scale. Especially, because 1 dimensional (1D) nanomaterials such as nanotube, nanorod and nanowire with high‐aspect ratio have nanoscale geometry and excellent mechanical, electrical, and chemical properties, they could play an important role in molecular and cellular biology. In this study, by using single crystalline 1D noble metal nanowire, we fabricated nano-sized 1D injector which can successfully interface with living cells and directly deliver biomolecules into several types of cell line (i.e., stem cell, mammalian embryo) without inducing detrimental damages on living cell. This nano-bio technology could be a promising and robust tool for introducing exogenous biomaterials into living organism.

Keywords: DNA, gene delivery, nanoinjector, nanowire

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Authors: Zelin Zhou, Timothy Langrish

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Spray drying is the final stage of milk powder production. Traditionally, the quality of spray-dried milk powders has mainly been assessed using their physical properties, such as their moisture contents, while chemical changes occurring during the spray drying process have often been ignored. With growing concerns about food quality, it is necessary to establish a better understanding of heat-induced degradation due to the spray-drying process of skim milk. In this study, the extent of thermal degradation for skim milk in a pilot-scale spray dryer has been investigated using different inlet gas temperatures. The extent of heat-induced damage has been measured by the formation of advanced Maillard reaction products and the loss of soluble proteins at pH 4.6 as assessed by a fluorometric method. A significant increase in the extent of thermal degradation has been found when the inlet gas temperature increased from 170°C to 190°C, suggesting protein unfolding may play an important role in the kinetics of heat-induced degradation for milk in spray dryers. Colour changes of the spray-dried skim milk powders have also been analysed using a standard lighting box. Colourimetric analysis results were expressed in CIELAB colour space with the use of the E index (E) and the Chroma (C) for measuring the difference between colours and the intensity of the colours. A strong linear correlation between the colour intensity of the spray-dried skim milk powders and the formation of advanced Maillard reaction products has been observed.

Keywords: colour formation, Maillard reactions, spray drying, skim milk powder

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Authors: Ceyda Dadali, Yeşim Elmaci

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Coffee was introduced as an economic crop during the fifteenth century; nowadays it is the most important food commodity ranking second after crude oil. Desirable sensory properties make coffee one of the most often consumed and most popular beverages in the world. The coffee preparation method has a significant effect on flavor and composition of coffee brews. Three different extraction methodologies namely decoction, infusion and pressure methods have been used for coffee brew preparation. Each of these methods is related to specific granulation (coffee grind) of coffee powder, water-coffee ratio temperature and brewing time. Coffee is a mixture of 1500 chemical compounds. Chemical composition of coffee highly depends on brewing methods, coffee bean species and roasting time-temperature. Coffee contains a wide number of very important bioactive compounds, such as diterpenes: cafestol and kahweol, alkaloids: caffeine, theobromine and trigonelline, melanoidins, phenolic compounds. The phenolic compounds of coffee include chlorogenic acids (quinyl esters of hidroxycinnamic acids), caffeic, ferulic, p-coumaric acid. In coffee caffeoylquinic acids, feruloylquinic acids and di-caffeoylquinic acids are three main groups of chlorogenic acids constitues 6% -10% of dry weight of coffee. The bioavailability of chlorogenic acids in coffee depends on the absorption and metabolization to biomarkers in individuals. Also, the interaction of coffee polyphenols with other compounds such as dietary proteins affects the biomarkers. Since bioactive composition of coffee depends on brewing methods effect of coffee brewing method on bioactive compounds of coffee will be discussed in this study.

Keywords: bioactive compounds of coffee, biomarkers, coffee brew, effect of brewing

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Authors: Shirin jalili, Hadi Shirzad, Samaneh Nabavi, Somayeh Khanjani

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Biosensors in forensic analysis are ideal biological tools that can be used for rapid and sensitive initial screening and testing to detect of suspicious components like biological and chemical agent in crime scenes. The wide use of different biomolecules such as proteins, nucleic acids, microorganisms, antibodies and enzymes makes it possible. These biosensors have great advantages such as rapidity, little sample manipulation and high sensitivity, also Because of their stability, specificity and low cost they have become a very important tool to Forensic analysis and detection of crime. In crime scenes different substances such as rape samples, Semen, saliva fingerprints and blood samples, act as a detecting elements for biosensors. On the other hand, successful fluid recovery via biosensor has the propensity to yield a highly valuable source of genetic material, which is important in finding the suspect. Although current biological fluid testing techniques are impaired for identification of body fluids. But these methods have disadvantages. For example if they are to be used simultaneously, Often give false positive result. These limitations can negatively result the output of a case through missed or misinterpreted evidence. The use of biosensor enable criminal researchers the highly sensitive and non-destructive detection of biological fluid through interaction with several fluid-endogenous and other biological and chemical contamination at the crime scene. For this reason, using of the biosensors for detecting the biological fluid found at the crime scenes which play an important role in identifying the suspect and solving the criminal.

Keywords: biosensors, forensic analysis, biological fluid, crime detection

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Authors: Lucia Salvioni, Pietro Giorgio Lovaglio, Valerio Leoni, Miriam Colombo, Luisa Fiandra

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The involvement of plasmatic surfactant protein-D (SP-D) in pulmonary diseases has been long investigated, and over the last two years, more interest has been directed to determine its role as a marker of COVID-19. In this direction, several studies aimed to correlate pulmonary surfactant proteins with the clinical manifestations of the virus indicated SP-D as a prognostic biomarker of COVID-19 pneumonia severity. The present work has performed a retrospective study on a relatively large cohort of patients of Hospital Pio XI of Desio (Lombardia, Italy) with the aim to assess differences in the hematic SP-D concentrations among COVID-19 patients and healthy donors and the role of SP-D as a prognostic marker of severity and/or of mortality risk. The obtained results showed a significant difference in the mean of log SP-D levels between COVID-19 patients and healthy donors, so as between dead and survived patients. SP-D values were significantly higher for both hospitalized COVID-19 and dead patients, with threshold values of 150 and 250 ng/mL, respectively. SP-D levels at admission and increasing differences among follow-up and admission values resulted in the strongest significant risk factors of mortality. Therefore, this study demonstrated the role of SP-D as a predictive marker of SARS-CoV-2 infection and its outcome. A significant correlation of SP-D with patient mortality indicated that it is also a prognostic factor in terms of mortality, and its early detection should be considered to design adequate preventive treatments for COVID-19 patients.

Keywords: SARS-CoV-2 infection, COVID-19, surfactant protein-D (SP-D), mortality, biomarker

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Authors: Mohammad B. HabibiNajafi, Saeideh Sadat Fatemizadeh, Maryam Tavakoli

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In recent years, the consumption of functional foods, including foods containing probiotic bacteria, has come to notice. Milk proteins have been identified as a source of angiotensin-I-converting enzyme )ACE( inhibitory peptides and are currently the best-known class of bioactive peptides. In this study, the effects of adding prebiotic ingredients (inulin and wheat fiber) and fat percentage (0%, 2% and 3.5%) in yogurt containing probiotic Lactobacillus casei on physicochemical properties, degree of proteolysis, antioxidant and ACE-inhibitory activity within 21 days of storage at 5 ± 1 °C were evaluated. The results of statistical analysis showed that the application of prebiotic compounds led to a significant increase in water holding capacity, proteolysis and ACE-inhibitory of samples. The degree of proteolysis in yogurt increases as storage time elapses (P < 0.05) but when proteolysis exceeds a certain threshold, this trend begins to decline. Also, during storage time, water holding capacity reduced initially but increased thereafter. Moreover, based on our findings, the survival of Lactobacillus casei in samples treated with inulin and wheat fiber increased significantly in comparison to the control sample (P < 0.05) whereas the effect of fat percentage on the survival of probiotic bacteria was not significant (P = 0.095). Furthermore, the effect of prebiotic ingredients and the presence of probiotic cultures on the antioxidant activity of samples was significant (P < 0.05).

Keywords: probiotic yogurt, proteolysis, ACE-inhibitory, antioxidant activity

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Authors: Daria Grechishnikova, Maria Poptsova

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Transposable elements play an important role in the evolution of various species from bacteria to human. Long Interspersed Nuclear Elements (LINEs) and Short Interspersed Nuclear Elements (SINEs) are two major classes of retrotransposons that occupy a considerable part of any genome and their copy numbers can range form several hundreds to a million. Both LINEs and SINEs multiply through a copy-and-paste mechanism. LINEs encode proteins, which make them capable of self-propagation while SINEs are parasitic and require the machinery of LINEs to multiply. The mechanisms how LINE and SINE RNA is recognized by the LINE-encoded reverse transcriptase (RT) remain unclear. For some SINE-LINE pairs, it was shown that they share a common 3’-end with a stem-loop structure. Majority of the SINE-LINE pairs do not have a common 3’-end. Recently we have shown that in the human genome Alu-L1 pairs have structurally similar stem-loop structure at the 3’-end. Here we extended our analysis to a wide range of species and analyzed LINEs from 161 different species from Repbase and 217 SINE sequences from SINEBase. It appeared that all of the analyzed sequences contained stem-loop structures at the 3’-end. Here we conclude that it is very likely that a common evolutionary mechanism of transposon RNA recognition requires the presence of stem-loop structures at their 3’-end.

Keywords: LINE, SINE, mechanisms of retrotransposition, retrotransposons, stem-loop, stem-loop structures, transposons

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Authors: Ayodele A. Otaiku

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Purpose: Nigeria has become the largest importer of rice in Africa and second in the world, 2015. Investigate interactions of organic rice farming on soil quality and health from bio-waste converted to biofertilizer and its environmental impact on rice crop. Methodology: Bio-wastes, poultry waste, organic agriculture wastes, wood ash mixed with microbial inoculant organisms called OBD-Plus microbes (broad spectrum) composted in anaerobic digester to OBD-biofertilizer (2010 - 2012) uses microbes to build humus and other stable carbons. Two field experiments were carried out at Badeggi, Niger state in 2011 and 2012 to evaluate the response of lowland rice production using biofertilizer. The experimental field was laid out in a strip-plot design with five treatments and three replications and at twenty-one day old seedlings of FARO 44 and FARO 52 rice varieties were transplanted. Plots without fertiliser application served as control. Findings: The highest rice grain yield increase of 4.4 t/ha over the control in 2012 against the Nigeria average of lowland rice grain yields of 1.5 t/ha. The utilization of OBD-Biofertilizer can decrease the use of chemical nitrogen fertilizer, prevent the depletion of soil organic matter and reduce environmental pollution. Increasing the floodwater productivity and optimizing the recycling of nutrients cum grazer populations and disease by biocontrols microbes present in the OBD-Biofertilizer. Organic matter in the soil improves by 58% and C/N 15 (2011) and 13.35 (2012). Implications: OBD- Biofertilizer produce plant growth hormones such as indole acetic acid (IAA), glomalin related soil protein and extracellular enzymes as phosphatases that promote soil health and quality. Conclusion: Microorganisms can enhance nutrients use efficiency by increasing root surface area e.g., mycorrhizal, fungi, promoting other beneficial symbioses of the host plant and microbial interactions resulting to increase in soil organic matter. By 2030, climate change is projected to depress cereal production in Africa by 2 to 3 percent. Improved seeds and increased fertilizer use should more than compensate, but this factor will still weigh heavily on efforts to make progress.

Keywords: OBD-plus microbial consortia, OBD-biofertilizer, rice production, soil quality, sustainable agriculture

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Authors: Saqib Ali, Man-Qun Wang

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The Japanese pine sawyer, Monochamus alternatus Hope (Coleoptera: Cerambycidae), is a major pest of pines and it is also the key vector of the exotic pinewood nematode in China. In the present study, we cloned, expressed, and purified a chemosensory protein (CSP) in M. alternatus. We surveyed its expression in various developmental stages of male and female adult tissues and determined its binding affinities for different pine volatiles using a competitive binding fluorescence assay. A CSP known as CSP5 in M. alternatus was obtained from an antennal cDNA library and expressed in Escherichia coli. Quantitative reverse transcription polymerase chain reaction results indicated that the CSP5 gene was mainly expressed in male and female antennae. Competitive binding assays were performed to test the binding affinity of recombinant CSP5 to 13 odour molecules of pine volatiles. The results showed that CSP5 showed very strong binding abilities to myrcene, (+)-β-pinene, and (−)-isolongifolene, whereas the volatiles 2-methoxy-4-vinylphenol, p-cymene, and (+)-limonene oxide have relatively weak binding affinity at pH 5.0. Three volatiles myrcene, (+)-β-pinene, and (−)-isolongifolene may play crucial roles in CSP5 binding with ligands, but this needs further study for confirmation. The sensitivity of insect to host plant volatiles can effectively be used to control and monitor the population through mass trapping as part of integrated pest management programs.

Keywords: olfactory-specific protein, volatiles, competitive binding assay, expression characteristics, qPCR

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Authors: Elena K. Schneider, Johnny X. Huang, Vincenzo Carbone, Mark Baker, Mohammad A. K. Azad, Matthew A. Cooper, Jian Li, Tony Velkov

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Ivacaftor is a novel CF trans-membrane conductance regulator (CFTR) potentiator that improves the pulmonary function for cystic fibrosis patients bearing a G551D CFTR-protein mutation. Because ivacaftor is highly bound (>97%) to plasma proteins, there is the strong possibility that co-administered CF drugs that compete for the same plasma protein binding sites and impact the free drug concentration. This in turn could lead to drastic changes in the in vivo efficacy of ivacaftor and therapeutic outcomes. This study compares the binding affinity of ivacaftor and co-administered CF drugs for human serum albumin (HSA) and α1-acid glycoprotein (AGP) using surface plasmon resonance and fluorimetric binding assays that measure the displacement of site selective probes. Due to their high plasma protein binding affinities, drug-drug interactions between ivacaftor are to be expected with ducosate, montelukast, ibuprofen, dicloxacillin, omeprazole and loratadine. The significance of these drug-drug interactions is interpreted in terms of the pharmacodynamic/pharmacokinetic parameters and molecular docking simulations. The translational outcomes of the data are presented as recommendations for a staggered treatment regimen for future clinical trials which aims to maximize the effective free drug concentration and clinical efficacy of ivacaftor.

Keywords: human α-1-acid glycoprotein, binding affinity, human serum albumin, ivacaftor, cystic fibrosis

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Authors: M. M. Essa, S. Subash, M. Akbar, S. Al-Adawi, A. Al-Asmi, G. J. Guillemein

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Transgenic (tg) mice which contain an amyloid precursor protein (APP) gene mutation, develop extracellular amyloid beta (Aβ) deposition in the brain, and severe memory and behavioral deficits with age. These mice serve as an important animal model for testing the efficacy of novel drug candidates for the treatment and management of symptoms of Alzheimer's disease (AD). Several reports have suggested that oxidative stress is the underlying cause of Aβ neurotoxicity in AD. Date palm fruits contain very high levels of antioxidants and several medicinal properties that may be useful for improving the quality of life in AD patients. In this study, we investigated the effect of dietary supplementation of Omani date palm fruits on the memory, anxiety and learning skills along with inflammation in an AD mouse model containing the double Swedish APP mutation (APPsw/Tg2576). The experimental groups of APP-transgenic mice from the age of 4 months were fed custom-mix diets (pellets) containing 2% and 4% Date palm fruits. We assessed spatial memory and learning ability, psychomotor coordination, and anxiety-related behavior in Tg and wild-type mice at the age of 4-5 months and 18-19 months using the Morris water maze test, rota rod test, elevated plus maze test, and open field test. Further, inflammatory parameters also analyzed. APPsw/Tg2576 mice that were fed a standard chow diet without dates showed significant memory deficits, increased anxiety-related behavior, and severe impairment in spatial learning ability, position discrimination learning ability and motor coordination along with increased inflammation compared to the wild type mice on the same diet, at the age of 18-19 months In contrast, PPsw/Tg2576 mice that were fed a diet containing 2% and 4% dates showed a significant improvements in memory, learning, locomotor function, and anxiety with reduced inflammatory markers compared to APPsw/Tg2576 mice fed the standard chow diet. Our results suggest that dietary supplementation with dates may slow the progression of cognitive and behavioral impairments in AD. The exact mechanism is still unclear and further extensive research needed.

Keywords: Alzheimer's disease, date palm fruits, Oman, cognitive decline, memory loss, anxiety, inflammation

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Authors: Lu Yang, Keng Po Lai

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Metformin, a well-known clinical drug against diabetes, is found with potential anti-diabetic and anti-obese benefits, as reported in increasing evidences. However, the current clinical and experimental investigations are not to reveal the detailed mechanisms of metformin-anti-obesity/hypertension. We have used the bioinformatics strategy, including network pharmacology and molecular docking methodology, to uncover the key targets and pathways of bioactive compounds against clinical disorders, such as cancers, coronavirus disease. Thus, in this report, the in-silico approach was utilized to identify the hug targets, pharmacological function, and mechanism of metformin against obesity and hypertension. The networking analysis identified 154 differentially expressed genes of obesity and hypertension, 21 interaction genes, and 6 hug genes of metformin treating hypertensive obesity. As a result, the molecular docking findings indicated the potent binding capability of metformin with the key proteins, including interleukin 6 (IL-6) and chemokine (C-C motif) Ligand 2 (CCL2), in hypertensive obesity. The metformin-exerted anti-hypertensive obesity action involved in metabolic regulation, inflammatory reaction. And the anti-hypertensive obesity mechanisms of metformin were revealed, including regulation of inflammatory and immunological signaling pathways for metabolic homeostasis in tissue and microenvironmental melioration in blood pressure. In conclusion, our identified findings with bioinformatics analysis have demonstrated the detailed hug and pharmacological targets, biological functions, and signaling pathways of metformin treating hypertensive obesity.

Keywords: metformin, obesity, hypertension, bioinformatics findings

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Authors: Hulya Ilyasoglu Buyukkestelli, Sedef Nehir El

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Lutein is a member of xanthophyll group of carotenoids found in fruits and vegetables. Lutein accumulates in the macula region of the retina and known as macular pigment which absorbs damaging light in the blue wavelengths. The presence of lutein in retina has been related to decreased risk of two common eye diseases, age-related macular degeneration, and cataract. Being a strong antioxidant, it may also have effects on prevention some types of cancer, cardiovascular disease, cognitive dysfunction. Humans are not capable of synthesizing lutein de novo; therefore it must be provided naturally by the diet, fortified foods, and beverages or nutritional supplement. However, poor bioavailability and physicochemical stability limit its usage in the food industry. Poor solubility in digestive fluids and sensitivity to heat, light, and oxygen are both affect the stability and bioavailability of lutein. In this context, new technologies, delivery systems and formulations have been applied to improve stability and solubility of lutein. Nanotechnology, including nanoemulsion, nanocrystal, nanoencapsulation technology and microencapsulation by complex coacervation, spray drying are promising ways of increasing solubilization of lutein and stability of it in different conditions. Bioavailability of lutein is also dependent on formulations used, starch formulations and milk proteins, especially sodium caseinate are found effective in improving the bioavailability of lutein. Designing foods with highly bioavailable and stabile lutein needs knowledge about current technologies, formulations, and further needs. This review provides an overview of the new technologies and formulations used to improve bioavailability of lutein and also gives a future outlook to food researches.

Keywords: bioavailability, formulation, lutein, nanotechnology

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Authors: Rabaya Basori, Arup K. Raychaudhuri

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In recent years, a variety of semiconductor nanowires (NWs) has been synthesized and used as basic building blocks for the development of electronic and optoelectronic nanodevices. Dielectrophoresis (DEP) has been widely investigated as a scalable technique to trap and manipulate polarizable objects. This includes biological cells, nanoparticles, DNA molecules, organic or inorganic NWs and proteins using electric field gradients. In this article, we have used DEP force to localize nanowire growth by physical vapor deposition (PVD) method as well as control of NW diameter on field assisted growth of the NWs of CuTCNQ (Cu-tetracyanoquinodimethane); a metal-organic charge transfer complex material which is well known of resistive switching. We report a versatile analysis platform, based on a set of nanogap electrodes, for the controlled growth of nanowire. Non-uniform electric field and dielectrophoretic force is created in between two metal electrodes, patterned by electron beam lithography process. Suspended CuTCNQ nanowires have been grown laterally between two electrodes in the vicinity of electric field and dielectric force by applying external bias. Growth and diameter dependence of the nanowires on external bias has been investigated in the framework of these two forces by COMSOL Multiphysics simulation. This report will help successful in-situ nanodevice fabrication with constrained number of NW and diameter without any post treatment.

Keywords: nanowire, dielectrophoretic force, confined growth, controlled diameter, comsol multiphysics simulation

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Authors: Kanika Gupta, Ashok Kumar

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Biofilms are dense, highly hydrated cell clusters that are irreversibly attached to a substratum, to an interface or to each other, and are embedded in a self-produced gelatinous matrix composed of extracellular polymeric substances. Research in biofilm field has become very significant, as biofilm has shown high mechanical resilience and resistance to antibiotic treatment and constituted as a significant problem in both healthcare and other industry related to microorganisms. The massive information both stated and hidden in the biofilm literature are growing exponentially therefore it is not possible for researchers and practitioners to automatically extract and relate information from different written resources. So, the current work proposes and discusses the use of text mining techniques for the extraction of information from biofilm literature corpora containing 34306 documents. It is very difficult and expensive to obtain annotated material for biomedical literature as the literature is unstructured i.e. free-text. Therefore, we considered unsupervised approach, where no annotated training is necessary and using this approach we developed a system that will classify the text on the basis of growth and development, drug effects, radiation effects, classification and physiology of biofilms. For this, a two-step structure was used where the first step is to extract keywords from the biofilm literature using a metathesaurus and standard natural language processing tools like Rapid Miner_v5.3 and the second step is to discover relations between the genes extracted from the whole set of biofilm literature using pubmed.mineR_v1.0.11. We used unsupervised approach, which is the machine learning task of inferring a function to describe hidden structure from 'unlabeled' data, in the above-extracted datasets to develop classifiers using WinPython-64 bit_v3.5.4.0Qt5 and R studio_v0.99.467 packages which will automatically classify the text by using the mentioned sets. The developed classifiers were tested on a large data set of biofilm literature which showed that the unsupervised approach proposed is promising as well as suited for a semi-automatic labeling of the extracted relations. The entire information was stored in the relational database which was hosted locally on the server. The generated biofilm vocabulary and genes relations will be significant for researchers dealing with biofilm research, making their search easy and efficient as the keywords and genes could be directly mapped with the documents used for database development.

Keywords: biofilms literature, classifiers development, text mining, unsupervised learning approach, unstructured data, relational database

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Authors: Priya Kulkarni, Soumya Koppikar, Narendrakumar Wagh, Dhanshri Ingle, Onkar Lande, Abhay Harsulkar

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Cartilage is an extracellular matrix composed of aggrecan, which imparts it with a great tensile strength, stiffness and resilience. Disruption in cartilage metabolism leading to progressive degeneration is a characteristic feature of Osteoarthritis (OA). The process involves enzymatic depolymerisation of cartilage specific proteoglycan, releasing free glycosaminoglycan (GAG). This released GAG in synovial fluid (SF) of knee joint serves as a direct measure of cartilage loss, however, limited due to its invasive nature. Western Ontario and McMaster Universities Arthritis Index (WOMAC) is widely used for assessing pain, stiffness and physical-functions in OA patients. The scale is comprised of three subscales namely, pain, stiffness and physical-function, intends to measure patient’s perspective of disease severity as well as efficacy of prescribed treatment. Twenty SF samples obtained from OA patients were analysed for their GAG values in SF using DMMB based assay. LK 1.0 vernacular version was used to attain WOMAC scale. The results were evaluated using SAS University software (Edition 1.0) for statistical significance. All OA patients revealed higher GAG values compared to the control value of 78.4±30.1µg/ml (obtained from our non-OA patients). Average WOMAC calculated was 51.3 while pain, stiffness and function estimated were 9.7, 3.9 and 37.7, respectively. Interestingly, a strong statistical correlation was established between WOMAC function subscale and GAG (p = 0.0102). This subscale is based on day-to-day activities like stair-use, bending, walking, getting in/out of car, rising from bed. However, pain and stiffness subscale did not show correlation with any of the studied markers and endorsed the atypical inflammation in OA pathology. On one side, where knee pain showed poor correlation with GAG, it is often noted that radiography is insensitive to cartilage degenerative changes; thus OA remains undiagnosed for long. Moreover, active cartilage degradation phase remains elusive to both, patient and clinician. Through analysis of large number of OA patients we have established a close association of Kellgren-Lawrence grades and increased cartilage loss. A direct attempt to correlate WOMAC and radiographic progression of OA with various biomarkers has not been attempted so far. We found a good correlation in GAG levels in SF and the function subscale.

Keywords: cartilage, Glycosaminoglycan, synovial fluid, western ontario and McMaster Universities Arthritis Index

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Authors: M. Ekiert, T. Uhl, A. Mlyniec

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Tendons are the biological soft tissue structures composed of collagen, proteoglycan, glycoproteins, water and cells of extracellular matrix (ECM). Tendons, which primary function is to transfer force generated by the muscles to the bones causing joints movement, are exposed to many micro and macro damages. In fact, tendons and ligaments trauma are one of the most numerous injuries of human musculoskeletal system, causing for many people (particularly for athletes and physically active people), recurring disorders, chronic pain or even inability of movement. The number of tendons reconstruction and transplantation procedures is increasing every year. Therefore, studies on soft tissues storage conditions (influencing i.e. tissue aging) seem to be an extremely important issue. In this study, an atomic-scale investigation on the kinetics of decomposition of two selected tendon components – collagen type I (which forms a 60-85% of a tendon dry mass) and elastin protein (which combine with ECM creates elastic fibers of connective tissues) is presented. A molecular model of collagen and elastin was developed based on crystal structure of triple-helical collagen-like 1QSU peptide and P15502 human elastin protein, respectively. Each model employed 4 linear strands collagen/elastin strands per unit cell, distributed in 2x2 matrix arrangement, placed in simulation box filled with water molecules. A decomposition phenomena was simulated with molecular dynamics (MD) method using ReaxFF force field and periodic boundary conditions. A set of NVT-MD runs was performed for 1000K temperature range in order to obtained temperature-depended rate of production of decomposition by-products. Based on calculated reaction rates activation energies and pre-exponential factors, required to formulate Arrhenius equations describing kinetics of decomposition of tested soft tissue components, were calculated. Moreover, by adjusting a model developed for collagen, system scalability and correct implementation of the periodic boundary conditions were evaluated. An obtained results provide a deeper insight into decomposition of selected tendon components. A developed methodology may also be easily transferred to other connective tissue elements and therefore might be used for further studies on soft tissues aging.

Keywords: decomposition, molecular dynamics, soft tissue, tendons

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Authors: Anuradha Gupta, Neelam Bansal, Varinder Uppal

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The present investigation was made on 35 Indian buffalo fetuses ranging from 0.9 cm to 104 cm curved crown rump length (CVRL). The gross anatomical study revealed that all structures were developed at 13 cm CVRL (87 days) in group I. At 0.9 cm CVRL (32 days) the heart was unseptated and tubular and was clearly divided into common atrial chamber dorsally and primitive ventricle in 1.2 cm CVRL fetus (34 days). Septum primum appeared at 1.9 cm CVRL (37 days), truncal ridges at 2.5 cm CVRL (39 days) and foramen ovale in 3.0 cm CVRL (42 days) buffalo foetuses. At 7.6 cm CVRL (62 days) endocardial cushions fused to form left and right atrioventricular openings and four chambered heart was formed in 8.7 cm CVRL (66 days). Endocardium and epicardium was thicker in atria as compared to ventricles in all the age groups. Myocardium of atria was thin as compared to ventricles in all the age groups and was loosely arranged. Immature hyaline cartilage was first appeared at base of aorta in 62 cm CVRL (213 days) fetuses. Intercalated discs were seen in group III and aorta, pulmonary artery, coronary artery were well appreciated in 3.2 cm CVRL (43 days). Neutral and acid mucopolysaccharides were comparatively more in atria than ventricles. Basic proteins showed strong reaction in atrium and ventricle, and intense in conduction system. Lipids and phospholipids were more in myocardium and conduction system than endocardium and epicardium. All the histochemical moieties were comparatively more in tunica intima than media and adventitia of all the great vessels of heart.

Keywords: buffalo, fetal development, histochemistry, heart

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Authors: Renata Shitakubo, Hanyu Yangcheng, Jay-lin Jane, Fernanda Peroni Okita, Beatriz Cordenunsi

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In the degradation transitory starch model, the enzymatic activity of glucan/water dikinase (GWD) and phosphoglucan/water dikinase (PWD) are essential for the granule degradation. GWD and PWD phosphorylate glucose molecules in the positions C6 and C3, respectively, in the amylopectin chains. This action is essential to allow that β-amylase degrade starch granules without previous action of α-amylase. During banana starch degradation, as part of banana ripening, both α- and β-amylases activities and proteins were already detected and, it is also known that there is a GWD and PWD protein bounded to the starch granule. Therefore, the aim of this study was to quantify both Gluc-6P and Gluc-3P in order to estimate the importance of the GWD-PWD-β-amylase pathway in banana starch degradation. Starch granules were isolated as described by Peroni-Okita et al (Carbohydrate Polymers, 81:291-299, 2010), from banana fruit at different stages of ripening, green (20.7%), intermediate (18.2%) and ripe (6.2%). Total phosphorus content was determinate following the Smith and Caruso method (1964). Gluc-6P and Gluc-3P quantifications were performed as described by Lim et al (Cereal Chemistry, 71(5):488-493, 1994). Total phosphorous content in green banana starch is found as 0.009%, intermediary banana starch 0.006% and ripe banana starch 0.004%, both by the colorimetric method and 31P-NMR. The NMR analysis showed the phosphorus content in C6 and C3. The results by NMR indicate that the amylopectin is phosphorylate by GWD and PWD before the bananas become ripen. Since both the total content of phosphorus and phosphorylated glucose molecules at positions C3 and C6 decrease with the starch degradation, it can be concluded that this phosphorylation occurs only in the surface of the starch granule and before the fruit be harvested.

Keywords: starch, GWD, PWD, 31P-NMR

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Authors: Jarel Elgin Tolentino, Arby Denise Nera, Mary Rose Roco, Angela Vianca Aspa, Nikko Beltran, Else Dapat

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Drug resistance by cells has been the problem in the medical field for decades now. The use of medicinal plants as a source of creating powerful drugs has been nowadays recognized worldwide to treat such resistant diseases. In the present study, the potential for Diospyros philippinensis A. DC. to inhibit growth of both bacteria and cancer cell line was conducted. The leaf crude extracts were screened for the presence of phytochemicals and examined for potential bioactivities by employing several assays like Kirby-Bauer disc diffusion method, DPPH (2,2-diphenyl-1-picryl-hydrazyl-hydrate) assay and MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) tetrazolium assay for the antibacterial, antioxidant and cytotoxic activities of the extract, respectively. Phytochemical test results of the extracts revealed the presence of alkaloids, flavonoids, saponins, phenols, quinones, cardiac glycosides, phlobatannins, carbohydrate, cardenolides and proteins. The leaf extracts were found to exhibit antibacterial activity against gram-positive bacteria, high antioxidant activity (99.22% ± 0.005) but did not show any sign of cytotoxicity towards HCT116 (ATCC CCL-247). The study therefore concludes that D. philippinensis A. DC. leaf extract can be a source of antibacterial and chemopreventive agents. This claim may be used as basis for future investigation.

Keywords: bioassay, medicinal plants, plant crude extracts, phytochemical screening

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Authors: Jiun-Nan Hou, Tien-Huang Chen, Wei-June Chen

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Dengue viruses (DENVs) are naturally transmitted between humans by mosquito vectors. Mosquito cells usually survive DENV infection, allowing infected mosquitoes to retain an active status for virus transmission. In this study, we found that DENV2 virus infection in mosquito cells causes the unfolded protein response (UPR) that activates the protein kinase RNA-like endoplasmic reticulum kinase (PERK) signal pathway, leading to shutdown of global protein translation in infected cells which was apparently regulated by the PERK signal pathway. According to observation in this study, the PERK signal pathway in DENV2-infected C6/36 cells alleviates ER stress, and reduces initiator and effector caspases, as well as the apoptosis rate via shutdown of cellular proteins. In fact, phosphorylation of eukaryotic initiation factor 2ɑ (eIF2ɑ) by the PERK signal pathway may impair recruitment of ribosomes that bind to the mRNA 5’-cap structure, resulting in an inhibitory effect on canonical cap-dependent cellular protein translation. The resultant pro-survival “byproduct” of infected mosquito cells is undoubtedly advantageous for viral replication. This finding provides insights into elucidating the PERK-mediated modulating web that is actively involved in dynamic protein synthesis, cell survival, and viral replication in mosquito cells.

Keywords: cap-dependent protein translation, dengue virus, endoplasmic reticulum stress, mosquito cells, PERK signal pathway

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Authors: M. Acin-Albiac, P. Filannino, R. Coda, Carlo G. Rizzello, M. Gobbetti, R. Di Cagno

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Demand for smart management of large amounts of agro-food by-products has become an area of major environmental and economic importance worldwide. Brewers' spent grain (BSG), the most abundant by-product generated in the beer-brewing process, represents an example of valuable raw material and source of health-promoting compounds. To the date, the valorization of BSG as a food ingredient has been limited due to poor technological and sensory properties. Tailored bioprocessing through lactic acid bacteria (LAB) fermentation is a versatile and sustainable means for the exploitation of food industry by-products. Indigestible carbohydrates (e.g., hemicelluloses and celluloses), high phenolic content, and mostly lignin make of BSG a hostile environment for microbial survival. Hence, the selection of tailored starters is required for successful fermentation. Our study investigated the metabolic strategies of Leuconostoc pseudomesenteroides and Lactobacillus plantarum strains to exploit BSG as a food ingredient. Two distinctive BSG samples from different breweries (Italian IT- and Finish FL-BSG) were microbially and chemically characterized. Growth kinetics, organic acid profiles, and the evolution of phenolic profiles during the fermentation in two BSG model media were determined. The results were further complemented with gene expression targeting genes involved in the degradation cellulose, hemicelluloses building blocks, and the metabolism of anti-nutritional factors. Overall, the results were LAB genus dependent showing distinctive metabolic capabilities. Leuc. pseudomesenteroides DSM 20193 may degrade BSG xylans while sucrose metabolism could be furtherly exploited for extracellular polymeric substances (EPS) production to enhance BSG pro-technological properties. Although L. plantarum strains may follow the same metabolic strategies during BSG fermentation, the mode of action to pursue such strategies was strain-dependent. L. plantarum PU1 showed a great preference for β-galactans compared to strain WCFS1, while the preference for arabinose occurred at different metabolic phases. Phenolic compounds profiling highlighted a novel metabolic route for lignin metabolism. These findings will allow an improvement of understanding of how lactic acid bacteria transform BSG into economically valuable food ingredients.

Keywords: brewery by-product valorization, metabolism of plant phenolics, metabolism of lactic acid bacteria, gene expression

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Authors: A. Aslihan Gokaltun, Ali Tuncel

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The unique properties of capillary electrochromatography (CEC) such as high performance, high selectivity, low consumption of both reagents and analytes ensure this technique an attractive one for the separation of biomolecules including nucleosides and nucleotides, peptides, proteins, carbohydrates. Monoliths have become a well-established separation media for CEC in the format that can be compared to a single large 'particle' that does not include interparticular voids. Convective flow through the pores of monolith significantly accelerates the rate of mass transfer and enables a substantial increase in the speed of the separation. In this work, we propose a new approach for the preparation of cationic monolithic stationary phase for capillary electrochromatography. Instead of utilizing a charge bearing monomer during polymerization, the desired charge-bearing group is generated on the capillary monolith after polymerization by using the reactive moiety of the monolithic support via one-pot, simple reaction. Optimized monolithic column compensates the disadvantages of frequently used reversed phases, which are difficult for separation of polar solutes. Rapid separation and high column efficiencies are achieved for the separation of neutral analytes, nucleic acid bases and nucleosides in reversed phase mode. Capillary monolith showed satisfactory hydrodynamic permeability and mechanical stability with relative standard deviation (RSD) values below 2 %. A new promising, reactive support that has a 'ligand selection flexibility' due to its reactive functionality represent a new family of separation media for CEC.

Keywords: biomolecules, capillary electrochromatography, cationic monolith, neutral analytes

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Authors: Sarita Puri, Tapan K. Chaudhuri

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Chaperonin GroEL is a tetradecameric Escherichia coli protein having identical subunits of 57 kDa. The elucidation of thermodynamic parameters related to stability for the native GroEL is not feasible as it undergoes irreversible unfolding because of its large size (800kDa) and multimeric nature. Nevertheless, it is important to determine the thermodynamic stability parameters for the highly stable GroEL protein as it helps in folding and holding of many substrate proteins during many cellular stresses. Properly folded monomers work as building-block for the formation of native tetradecameric GroEL. Spontaneous refolding behavior of monomeric GroEL makes it suitable for protein-denaturant interactions and thermodynamic stability based studies. The urea mediated unfolding is a three state process which means there is the formation of one intermediate state along with native and unfolded states. The heat mediated denaturation is a two-state process. The unfolding process is reversible as observed by the spontaneous refolding of denatured protein in both urea and head mediated refolding processes. Analysis of folding/unfolding data provides a measure of various thermodynamic stability parameters for the monomeric GroEL. The proposed mechanism of unfolding of monomeric GroEL is a three state process which involves formation of one stable intermediate having folded apical domain and unfolded equatorial, intermediate domains. Research in progress is to demonstrate the importance of specific residues in stability and oligomerization of GroEL protein. Several mutant versions of GroEL are under investigation to resolve the above mentioned issue.

Keywords: equilibrium unfolding, monomeric GroEl, spontaneous refolding, thermodynamic stability

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Authors: Sirikwan Ponprateep, Anchalee Tassanakajon, Vichien Rimphanitchayakit

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A Kazal-type serine proteinase inhibitor, SPIPm2, was abundantly expressed in the hemocytes and secreted into shrimp plasma has anti-viral property against white spot syndrome virus (WSSV). To discover the molecular mechanism of antiviral activity, the binding assay showed that SPIPm2 bind to the components of viral particle and shrimp hemocyte. From our previous report, viral target protein of SPIPm2 was identified, namely WSV477 using yeast two-hybrid screening. WSV477 is an early gene product of WSSV and involved in viral propagation. In this study, the co-immunoprecipitation technique and Tandem Mass Spectrometry (LC-MS/MS) was used to identify the target protein of SPIPm2 from shrimp hemocyte. The target protein of SPIPm2 was cyclophilin A. In vertebrate, cyclophilin A or peptidylprolyl isomerase A was reported to be the immune suppressor interacted with cyclosporin A involved in immune defense response. The recombinant cyclophilin A from Penaeus monodon (rPmCypA) was produced in E.coli system and purified using Ni-NTA column to confirm the protein-protein interaction. In vitro pull-down assay showed the interaction between rSPIPm2 and rPmCypA. To study the biological function of these proteins, the expression analysis of immune gene in shrimp defense pathways will be investigated after rPmCypA administration.

Keywords: cyclophilin A, protein-protein interaction, Kazal-type serine proteinase inhibitor, Penaeus monodon

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Authors: V. M. Varagyan, G. A. Gevorgyan, K. V. Grigoryan, A. L. Varagyan

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Alaverdi region is situated in the northern part of the Republic of Armenia. Previous studies (1989) in Alaverdi region showed that due to soil irrigation with the highly polluted waters of the Debed and Shnogh rivers, the content of heavy metals in the brown forest steppe soils was significantly higher than the maximum permissible concentration as a result of which the fermentative activity in all the layers of the soils was stressed. Compared to the non-polluted soils, the activity of ferments in the plough layers of the highly polluted soils decreased by 44 - 68% (invertase – 60%, phosphatase – 44%, urease – 66%, catalase – 68%). In case of the soil irrigation with the polluted waters, a decrease in the intensity of fermentative reactions was conditioned by the high content of heavy metals in the soils and changes in chemical composition, physical and physicochemical properties. 20-year changes in the fermentative activity in the brown forest steppe soils in Alaverdi region were investigated. The activity of extracellular ferments in the soils was determined by the unification methods. The study has confirmed that self-recovery process occurs in soils previously polluted with heavy metals which can be revealed by fermentative activity. The investigations revealed that during 1989 – 2009, the activity of ferments in the plough layers of the medium and highly polluted soils increased by 31.2 – 52.6% (invertase – 31.2%, urease – 52.6%, phosphatase – 33.3%, catalase – 41.8%) and 24.1 – 87.0% (invertase – 40.4%, urease – 76.9%, phosphatase – 24.1%, catalase – 87.0%) respectively which indicated that the dynamic properties of the soils, which had been broken due to heavy metal pollution, were improved. In 1989, the activity of the Alaverdi copper smelting plant was temporarily stopped due to financial problems caused by the economic crisis and the absence of market, and the factory again started operation in 1997 and isn’t currently running at full capacity. As a result, the Debed river water has obtained a new chemical composition and comparatively good irrigation properties. Due to irrigation with this water, the gradually recovery of the soil dynamic properties, which had been broken due to irrigation with the waters polluted with heavy metals, was occurred. This is also explained by the fact that in case of irrigation with the partially cleaned water, the soil protective function against pollutants rose due to a content increase in humus and silt fractions. It is supposed that in case of the soil irrigation with the partially cleaned water, the intensity of fermentative reactions wasn’t directly affected by heavy metals.

Keywords: alaverdi region, heavy metal pollution, self-recovery, soil fermentative activity

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Authors: Chaomeng Liu, Li Li, Xiao Wang, Li Ren, Qinge Zhang

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Late-life depression (LLD) is a prevalent mental disorder among the elderly, frequently accompanied by significant cognitive decline, and has emerged as a worldwide public health concern. Microglia, astrocytes, and peripheral immune cells play pivotal roles in regulating inflammatory responses within the central nervous system (CNS) across diverse cerebral disorders. This review commences with the clinical research findings and accentuates the recent advancements pertaining to microglia and astrocytes in the neuroinflammation process of LLD. The reciprocal communication network between the CNS and immune system is of paramount importance in the pathogenesis of depression and cognitive decline. Stress-induced downregulation of tight and gap junction proteins in the brain results in increased blood-brain barrier permeability and impaired astrocyte function. Concurrently, activated microglia release inflammatory mediators, initiating the kynurenine metabolic pathway and exacerbating the quinolinic acid/kynurenic acid imbalance. Moreover, the balance between Th17 and Treg cells is implicated in the preservation of immune homeostasis within the cerebral milieu of individuals suffering from LLD. The ultimate objective of this review is to present future strategies for the management and treatment of LLD, informed by the most recent advancements in research, with the aim of averting or postponing the onset of AD.

Keywords: neuroinflammation, late-life depression, microglia, astrocytes, central nervous system, blood-brain barrier, Kynurenine pathway

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Authors: Yichao Liang, Biye Chen, Xiang Li, Steven R. Dimler

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This study investigated the effects of adding calcium and magnesium chloride on heat and storage stability of milk protein concentrate-soy protein isolate (8:2 respectively) mixtures containing 10% w/w total protein subjected to the in-container sterilization (115 °C x 15 min). The particle size does not change when emulsions are heated at pH between 6.7 and 7.3 irrespective of the mixed protein ratio. Increasing concentration of divalent cation salts resulted in an increase in protein particle size, dry sediment formation and sediment height and a decrease in pH, heat stability and hydration in milk protein concentrate-soy protein isolate mixtures solutions on sterilization at 115°C. Fortification of divalent cation salts in milk protein concentrate-soy protein isolate mixture solutions resulted in an accelerated protein sedimentation and two unique sediment regions during accelerated storage stability testing. Moreover, the heat stability decreased upon sterilization at 115°C, with addition of MgCl₂ causing a greater increase in sedimentation velocity and compressibility than CaCl₂. Increasing pH value of protein milk concentrate-soy protein isolate mixtures solutions from 6.7 to 7.2 resulted in an increase in viscosity following the heat treatment. The study demonstrated that the type and concentration of divalent cation salts used strongly impact heat and storage stability of milk protein concentrate-soy protein isolate mixture nutritional beverages.

Keywords: divalent cation salts, heat stability, milk protein concentrate, soy protein isolate, storage stability

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