Search results for: in vivo biomarkers

Authors: Bhumika Wadhwa, Fayaz Malik

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The serine/threonine protein kinase B (PKB) also known as Akt, is one of the multifaceted kinase in human kinome, existing in three isoforms. Akt plays a vital role in phosphoinositide 3-kinase (PI3K) mediated oncogenesis in various malignancies and is one of the attractive targets for cancer drug discovery. The functional significance of an individual isoform of Akt is not redundant in cancer cell proliferation and metastasis instead Akt isoforms play distinct roles during metastasis; thereby regulating EMT. This study aims to determine isoform specific functions of Akt in cancer. The results obtained suggest that Akt1 restrict tumor invasion, whereas Akt2 promotes cell migration and invasion by various techniques like MTT, wound healing and invasion assay. Similarly, qRT-PCR also revealed that Akt3 has shown promising results in promoting cancer cell migration. Contrary to pro-oncogenic properties attributed to Akt, it is to be understood how various isoforms of Akt compensates each other in the regulation of common pathways during cancer progression and drug resistance. In conclusion, this study aims to target selective isoforms which is essential to inhibit cancer. However, the question now is whether, and how much, Akt inhibition will be tolerated in the clinic remains to be answered and the experiments will have to address the question of which combinations of newly devised Akt isoform specific inhibitors exert a favourable therapeutic effect in in vivo models of cancer to provide the therapeutic window with minimal toxicity.

Keywords: Akt isoforms, cancer, drug resistance, epithelial mesenchymal transition

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Authors: Mariane G. Tadros, Shahira M. Ezzat, Maha M. Salama, Mohamed A. Farag

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In this study, the in vitro anticholinesterase activity of the volatile oils of both O. basilicum and O. africanum was investigated and both samples showed significant activity. As a result, the major constituents of the two oils were isolated using several column chromatography. Linalool, 1,8-cineol and eugenol were isolated from the volatile oil of O. basilicum and camphor was isolated from the volatile oil of O. africanum. The anticholinesterase activity of the isolated compounds were also evaluated where 1,8-cineol showed the highest inhibitory activity followed by camphor. To confirm these activities, learning and memory enhancing effects were tested in mice. Memory impairment was induced by scopolamine, a cholinergic muscarinic receptor antagonist. Anti-amnesic effects of both volatile oils and their terpenoids were investigated by the passive avoidance task in mice. We also examined their effects on brain acetylcholinesterase activity. Results showed that scopolamine-induced cognitive dysfunction was significantly attenuated by administration of the volatile oils and their terpenoids, eugenol and camphor, in the passive avoidance task and inhibited brain acetylcholinesterase activity. These results suggest that O. basilicum and O. africanum volatile oils can be good candidates for further studies on Alzheimer’s disease via their acetylcholinesterase inhibitory actions.

Keywords: Ocimum baselicum, Ocimum africanum, GC/MS analysis, anticholinesterase

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Authors: Al-Nouri Doha Mostfa, Al-Khalifa Abdulrahman Salih

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The effects of long-term supplementation with different dietary omega-6/omega-3 (ω-6/ω-3) polyunsaturated fatty acid (PUFAs) ratios on the bone marrow fatty acids level, plasma biomarkers of bone metabolism, and minerals content in bone were evaluated in rabbits. Weanling male and female New Zealand white rabbits were randomly assigned to five groups and fed ad libitum for 100 days on diets containing 70 g/kg different dietary oils which providing the following ω-6/ω-3 ratios: soy bean oil (SBO control, 8.68), sesame oil (SO, 21.75), fish oil (FO, 0.39), DHA algae oil (DHA, 0.63), and DHA and ARA algae oils (DHA/ARA, 0.68). The bone marrow arachidonic (ARA), eicosapentaenoic (EPA), and docosahexaenoic (DHA) fatty acid levels were significantly influenced by and reflected the dietary ω-6/ω-3 ratios fed to rabbits. Rabbits fed on the FO diet maintained a lower ω-6/ω-3 ratio and a higher EPA and DHA levels, those fed on the DHA/ARA diet maintained a lower ω-6/ω-3 ratio and a higher ARA level, while those fed on the SO diet maintained a higher ω-6/ω-3 ratio and a lower ARA level. Plasma alkaline phosphatase (ALP) activity was significantly higher in male and female rabbits fed the DHA/ARA diet compared with those fed the control, SO, FO, or DHA diets. There was a significant main effect of dietary treatment on femur calcium (Ca), phosphorous (P), magnesium (Mg), and zinc (Zn) contents in both genders. This study confirmed that different dietary oil sources with varying ω-6/ω-3 ratios significantly altered the fatty acids level of bone marrow. In addition, the significant elevation in minerals content and the maintenance of optimal Ca/P ratio in bone of DHA/ARA and DHA fed groups beside the significant elevation in ALP activity in the DHA/ARA fed group proved that marine algae oils may be promising dietary sources for promoting bone mineralization and formation, thus improving bone mass during the growth stage.

Keywords: arachidonic (ARA), docosahexaenoic (DHA), eicosapentaenoic (EPA), growing rabbits

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Authors: Shangqing Song, Bin Xu, Yajun Cheng, Zhong Wang

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miRNAs derived from exosomes exist in a body fluid such as urine were regarded as potential biomarkers for various human cancers diagnosis and prognosis, as mature miRNAs can be steadily preserved by exosomes. However, its potential value in clear cell renal cell carcinoma (ccRCC) diagnosis and prognosis remains unclear. In the present study, differentially expressed miRNAs from urinal exosomes were identified by next-generation sequencing (NGS) technology. The 16 differentially expressed miRNAs were identified between ccRCC patients and healthy donors. To explore the specific diagnosis biomarker of ccRCC, we validated these urinary exosomes from 70 early-stage renal cancer patients, 30 healthy people and other urinary system cancers, including 30 early-stage prostate cancer patients and 30 early-stage bladder cancer patients by qRT-PCR. The results showed that urinary exosome miR-30c-5p could be stably amplified and meanwhile the expression of miR-30c-5p has no significant difference between other urinary system cancers and healthy control, however, expression level of miR-30c-5p in urinary exosomal of ccRCC patients was lower than healthy people and receiver operation characterization (ROC) curve showed that the area under the curve (AUC) values was 0.8192 (95% confidence interval was 0.7388-0.8996, P= 0.0000). In addition, up-regulating miR-30c-5p expression could inhibit renal cell carcinoma cells growth. Lastly, HSP5A was found as a direct target gene of miR-30c-5p. HSP5A depletion reversed the promoting effect of ccRCC growth casued by miR-30c-5p inhibitor, respectively. In conclusion, this study demonstrated that urinary exosomal miR-30c-5p is readily accessible as diagnosis biomarker of early-stage ccRCC, and miR-30c-5p might modulate the expression of HSPA5, which correlated with the progression of ccRCC.

Keywords: clear cell renal cell carcinoma, exosome, HSP5A, miR-30c-5p

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Authors: Tahani Hallek, Dhaou Akrout, Riadh Ahmadi, Mabrouk Montacer

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The present work is an organic geochemical study of the Fahdene Formation outcrops at the Mahjouba region belonging to the Eastern part of the Kalaat Senan structure in northwestern Tunisia (the Kef-Tedjerouine area). The analytical study of the organic content of the samples collected, allowed us to point out that the Formation in question is characterized by an average to good oil potential. This fossilized organic matter has a mixed origin (type II and III), as indicated by the relatively high values of hydrogen index. This origin is confirmed by the C29 Steranes abundance and also by tricyclic terpanes C19/(C19+C23) and tetracyclic terpanes C24/(C24+C23) ratios, that suggest a marine environment of deposit with high plants contribution. We have demonstrated that the heterogeneity of organic matter between the marine aspect, confirmed by the presence of foraminifera, and the continental contribution, is the result of an episodic anomaly in relation to the sequential stratigraphy. Given that the study area is defined as an outer platform forming a transition zone between a stable continental domain to the south and a deep basin to the north, we have explained the continental contribution by successive forced regressions, having blocked the albian transgression, allowing the installation of the lowstand system tracts. This aspect is represented by the incised valleys filling, in direct contact with the pelagic and deep sea facies. Consequently, the Fahdene Formation, in the Kef-Tedjerouine area, consists of transgressive system tracts (TST) brutally truncated by extras of continental progradation; resulting in a mixed influence deposition having retained a heterogeneous organic material.

Keywords: molecular geochemistry, biomarkers, forced regression, deposit environment, mixed origin, Northern Tunisia

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Authors: Desissa Yadeta Muleta

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Purely organic long-persistent luminescence materials (OLPLMs) have been developed as emerging organic materials due to their simple production process, low preparation cost and better biocompatibilities. Notably, OLPLMs with afterglow-time-tunable long-persistent luminescence (LPL) characteristics enable higher-level protection applications and have great prospects in biological applications. The realization of these advanced performances depends on our ability to gradually tune LPL duration under ambient conditions, however, the strategies to achieve this are few due to the lack of unambiguous mechanisms. Here, we propose a two-step strategy to gradually tune LPL duration of OLPLMs over a wide range of seconds in water and air, by using derivatives as the guest and introducing a third-party material into the host-immobilized host–guest doping system. Based on this strategy, we develop an analysis method for deoxyribonucleic acid (DNA) content detection without DNA separation in aqueous samples, which circumvents the influence of the chromophore, fluorophore and other interferents in vivo, enabling a certain degree of in situ detection that is difficult to achieve using today’s methods. This work will expedite the development of afterglow-time-tunable OLPLMs and expand new horizons for their applications in data protection, bio-detection, and bio-sensing

Keywords: deoxyribonucliec acid, long persistent luminescent materials, water, air

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Authors: Debasmita Dubey, Rajesh Kumar Meher, Smruti Pragya Samal, Pradeep Kumar Naik

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Background: To determine antioxidant properties and anticancer activity by ROS and mitochondrial transmembrane potential mediated apoptosis against MCF7, MDA-MB-231, cell line. Methods: Leaf sample was extracted using methanol by microwave digestion technique. The antioxidant properties of the methanolic extract were determined by a DPPH scavenging assay. In vitro anticancer activity, mitochondrial transmembrane potential, apoptosis activity and DNA fragmentation study, as well as intracellular ROS activity of most potential leaf extract, were also determined by using the MDA-MB-231cell line. In vivo animal toxicity study was carried out using mice model. Results: Methanolic leaf extract has shown the highest antioxidant, as well as anticancer activity, is based on the assay conducted. For the identification of active phytochemicals from methanolic extract, High-resolution mass spectroscopy-LCMS was used. In vitro cytotoxicity study against MCF-7 and MDA-MB-231 cell line and IC 50 value was found to be 37.5µg/ml. From histopathological studies, no toxicity in liver and kidney tissue was identified. Conclusion: This plant tuber can be used as a regular diet to reduce the chance of breast cancer. Further, more studies should be conducted to isolate and identify the responsible compound.

Keywords: human breast adenocarcinoma, ROS, mitochondrial transmembrane, apoptosis

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Authors: Luai Z. Hasoun, Steven W. Bailey, Kitti K. Outlaw, June E. Ayling

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Human skin color is thought to have evolved to balance sufficient photochemical synthesis of vitamin D versus the need to protect not only DNA but also folate from degradation by ultraviolet light (UV). Although the risk of DNA damage and subsequent skin cancer is related to light skin color, the effect of UV on skin folate of any species is unknown. Here we show that UVA irradiation at 13 mW/cm2 for a total exposure of 187 J/cm2 (similar to a maximal daily equatorial dose) induced a significant loss of total folate in epidermis of ex vivo white skin. No loss was observed in black skin samples, or in the dermis of either color. Interestingly, while the concentration of 5 methyltetrahydrofolate (5-MTHF) fell in white epidermis, a concomitant increase of tetrahydrofolic acid was found, though not enough to maintain the total pool. These results demonstrate that UVA indeed not only decreases folate in skin, but also rearranges the pool components. This could be due in part to the reported increase of NADPH oxidase activity upon UV irradiation, which in turn depletes the NADPH needed for 5-MTHF biosynthesis by 5,10-methylenetetrahydrofolate reductase. The increased tetrahydrofolic acid might further support production of the nucleotide bases needed for DNA repair. However, total folate was lost at a rate that could, with strong or continuous enough exposure to ultraviolet radiation, substantially deplete light colored skin locally, and also put pressure on total body stores for individuals with low intake of folate.

Keywords: depletion, folate, human skin, ultraviolet

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Authors: Hsin-Lien Lin, Ju-Hui Fu

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Dendritic cells (DCs) are the major professional antigen-presenting cells for the development of optimal T-cell immunity. DCs can be used as pharmacological targets to screen novel biological modifiers for the treatment of harmful immune responses, such as transplantation rejection. Dryopteris crassirhizoma Nakai (Aspiadaceae) is used for traditional herbal medicine in the region of East Asia. The root of this fern plant has been listed for treating inflammatory diseases. Dryocrassin is the tetrameric phlorophenone component derived from Dryopteris. Here, we tested the immunomodulatory potential of dryocrassin on lipopolysaccharide (LPS)-stimulated activation of mouse bone marrow-derived DCs in vitro and in skin allograft transplantation in vivo. Results demonstrated that dryocrassin reduced the secretion of tumor necrosis factor-α, interleukin-6, and interleukin-12p70 by LPS-stimulated DCs. The expression of LPS-induced major histocompatibility complex class II, CD40, and CD86 on DCs was also blocked by dryocrassin. Moreover, LPS-stimulated DC-elicited allogeneic T-cell proliferation was lessened by dryocrassin. In addition, dryocrassin inhibited LPS-induced activation of IϰB kinase, JNK/p38 mitogen-activated protein kinase, as well as the translocation of NF-ϰB. Treatment with dryocrassin obviously diminished 2,4-dinitro-1-fluorobenzene- induced delayed-type hypersensitivity and prolonged skin allograft survival. Dryocrassin may be one of the potent immunosuppressive agents for transplant rejection through the destruction of DC maturation and function.

Keywords: dryocrassin, dendritic cells, immunosuppression, skin allograft

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Authors: Mai M. Farid, Ximeng Yang, Tomoharu Kuboyama, Yuna Inada, Chihiro Tohda

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Intro: Trigonella foenum-graecum (Fenugreek), from Fabaceae family is a well-known plant traditionally used as food and medicine. Many pharmacological effects of Trigonella foenum- graecum seeds extract (TF extract) were evaluated such as anti-diabetic, anti-tumor and anti-dementia effects using in vivo models. Regarding the anti-dementia effects of TF extract, diabetic rats, aluminum chloride-induced amnesia rats and scopolamine-injected mice were used previously for evaluation, which are not well established as Alzheimer’s disease models. In addition, those previous studies, active constituents in TF extract for memory function were not identified. Method: This study aimed to clarify the effect of TF extract on Alzheimer’s disease model, 5XFAD mouse that overexpresses mutated APP and PS1 genes and determine the major active constituent in the brain after oral intake of TF extract. Results: Trigonelline was detected in the cerebral cortex of 5XFAD mice after 24 hours of oral administration of TF extract by LC-MS/MS. Oral administration of TF extract for 17 days improved object location memory in 5XFAD mice. Conclusion: These results suggest that TF extract and its active constituents could be an expected therapeutic candidate for Alzheimer’s disease.

Keywords: Alzheimer's disease, LC-MS/MS, memory recovery, Trigonella foenum-graecum Seeds, 5XFAD mice

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Authors: Kent Salomonsson, Xuefang Zhao, Sara Kallin

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Human soft tissue is loaded and deformed by any activity, an effect known as a stress-strain relationship, and is often described by a load and tissue elongation curve. Several advances have been made in the fields of biology and mechanics of soft human tissue. However, there is limited information available on in vivo tissue mechanical characteristics and behavior. Confident mechanical properties of human soft tissue cannot be extrapolated from e.g. animal testing. Thus, there is need for non invasive methods to analyze mechanical characteristics of soft human tissue. In the present study, the internal mechanical conditions of the lower limb, which is subject to an external load, is studied by use of the finite element method. A detailed finite element model of the lower limb is made possible by use of MRI scans. Skin, fat, bones, fascia and muscles are represented separately and the material properties for them are obtained from literature. Previous studies have been shown to address macroscopic deformation features, e.g. indentation depth, to a large extent. However, the detail in which the internal anatomical features have been modeled does not reveal the critical internal strains that may induce hypoxia and/or eventual tissue damage. The results of the present study reveals that lumped material models, i.e. averaging of the material properties for the different constituents, does not capture regions of critical strains in contrast to more detailed models.

Keywords: FEM, tissue, indentation, properties

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Authors: Khadijeh Abhari, Seyed Shahram Shekarforoush, Saeid Hosseinzadeh

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An in vivo trial was conducted to evaluate the effects of Bacillus coagulans, and inulin, either separately or in combination, on lipid profile using a rat model. Thirty-two male Wistar rats were randomly divided into four groups (n=8) and fed as follows: standard diet (control), standard diet with 5% w/w long chain inulin (prebiotic), standard diet with 109 spores/day spores of B. coagulans by orogastric gavage (probiotic), and standard diet with 5% w/w long chain inulin and 109 spores/day of B. coagulans (synbiotic). Rats were fed the treatments for 30 days. Serum samples were collected 10, 20 and 30 days following onset of treatment. Total cholesterol, HDL and LDL cholesterol and triglycerides concentrations were analyzed. Results of this study showed that inulin potentially affected the lipid profile. An obvious decrease in serum total cholesterol and LDL-cholestrol of rats fed with inulin in synbiotic and prebiotic groups was seen in all sampling days. Inulin fed rats also demonstrated higher levels of HDL-cholesterol concentration; however this value in probiotic and control fed rats remains without significant change. According to the results of this study, B. coagulans did not contribute to any lipid profile changes after 30 days. Thus, further in vitro investigations on the characteristic of these bacteria could be useful to gain insights into understanding the treatment of probiotics in order to achieve the maximum beneficial effect.

Keywords: bacillus coagulans, inulin, rat, lipid profile, synbiotic diet

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Authors: Nadia A. Mohamed, Mehrevan M. Abdel-Moniem

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Zingiber officinale (ginger) has been cultivated for medicinal purposes due to their various proprieties both in vitro and in vivo, so we designed to evaluate the ginger’s potential effect on nephrin m RNA expression in cisplatin-induced nephrotoxic rats. Method: Forty male albino rats were divided into group I was injected (IP) with one ml saline, group II(cisplatin) injected (IP) with a single dose of 12 mg/kg cisplatin, group III (ginger) received (PO) 310 mg/kg for 30 successive days, and group IV(cisplatin and ginger) rats received ginger extract (310 mg/kg) daily for 20 successive days (PO), and then on day 20 of ginger extract administration each rat was injected(IP) with a single dose of 12 mg/kg cisplatin. The blood was sampled to assess urea, creatinine (SC), while the levels of malondialdehyde (MDA), nitric oxide (NO) and paraoxonase (PON1) were measured in kidney tissue homogenate. Expression of urinary nephrin gene (nephrin mRNA) was detected using qRT-PCR. Results: Treatment with ginger significantly decreased the levels of kidney function parameters as well as MDA and NO elevated by cisplatin injection, while PON1 was significantly reduced in the cisplatin group. However, the protection of male rats with ginger significantly increased the levels of nephrin gene expression and PON1 compared with the cisplatin-treated group. Our results generated a proposal on the ameliorating effect of ginger on nephrin mRNA gene expression reduction in cisplatin-induced nephrotoxicity.

Keywords: nephrin mRNA, ginger, cisplatin, nephrotoxicity

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Authors: Thomas Geninatti, Bruno Giacomo, Alessandro Grattoni

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Nanofluidic devices have been investigated for over a decade as promising platforms for the controlled release of therapeutics. The nanochannel drug delivery system (nDS), a membrane fabricated with high precision silicon techniques, capable of zero-order release of drugs by exploiting diffusion transport at the nanoscale originated from the interactions between molecules with nanochannel surfaces, showed the flexibility of the sustained release in vitro and in vivo, over periods of time ranging from weeks to months. To improve the implantable bio nanotechnology, in order to create a system that possesses the key features for achieve the suitable release of therapeutics, the next generation of nDS has been created. Platinum electrodes are integrated by e-beam deposition onto both surfaces of the membrane allowing low voltage (<2 V) and active temporal control of drug release through modulation of electrostatic potentials at the inlet and outlet of the membrane’s fluidic channels. Hence, a tunable administration of drugs is ensured from the nanochannel drug delivery system. The membrane will be incorporated into a peek implantable capsule, which will include drug reservoir, control hardware and RF system to allow suitable therapeutic regimens in real-time. Therefore, this new nanotechnology offers tremendous potential solutions to manage chronic disease such as cancer, heart disease, circadian dysfunction, pain and stress.

Keywords: nanochannel membrane, drug delivery, tunable release, personalized administration, nanoscale transport, biomems

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Authors: Dahi Ghareab Abdelsalam Ibrahim, R. H. Bakr

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Ionizing radiation, such as gamma radiation and X-rays, has many applications in medical diagnoses and cancer treatment. In this paper, we used the windowed Fourier transform to extract the complex image of the deformed red blood cells. The real values of the complex image are used to extract the best fitting of the deformed cell boundary. Male albino rats are irradiated by γ-rays from ⁶⁰Co. The male albino rats are anesthetized with ether, and then blood samples are collected from the eye vein by heparinized capillary tubes for studying the radiation-damaging effect in-vivo by the proposed windowed Fourier transform. The peripheral blood films are prepared according to the Brown method. The peripheral blood film is photographed by using an Automatic Image Contour Analysis system (SAMICA) from ELBEK-Bildanalyse GmbH, Siegen, Germany. The SAMICA system is provided with an electronic camera connected to a computer through a built-in interface card, and the image can be magnified up to 1200 times and displayed by the computer. The images of the peripheral blood films are then analyzed by the windowed Fourier transform method to extract the precise deformation from the best fitting. Based on accurate deformation evaluation of the red blood cells, diseases can be diagnosed in their primary stages.

Keywords: windowed Fourier transform, red blood cells, phase wrapping, Image processing

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Authors: Fadilah Aleanizy

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Colicins are a family of bacterial toxins that kill Escherichia coli and other closely related species. The mode of action of colicins involves binding to an outer membrane receptor and translocation across the cell envelope, leading to cytotoxicity through specific targets. The mechanism of colicin cytotoxicity includes a non-specific endonuclease activity or depolarization of the cytoplasmic membrane by pore-forming activity. For Group A colicins, translocation requires an interaction between the N-terminal domain of the colicin and a series of membrane- bound and periplasmic proteins known as the Tol system (TolB, TolR, TolA, TolQ, and Pal and the active domain must be translocated through the outer membranes. Protein-protein interactions are intrinsic to virtually every cellular process. The transient protein-protein interactions of the colicin include the interaction with much more complicated assemblies during colicin translocation across the cellular membrane to its target. The potassium release assay detects variation in the K+ content of bacterial cells (K+in). This assays is used to measure the effect of pore-forming colicins such as ColA on an indicator organism by measuring the changes of the K+ concentration in the external medium (K+out ) that are caused by cell killing with a K+ selective electrode. One of the goals of this work is to employ a quantifiable in-vivo method to spot which Tol protein are more implicated in the interaction with colicin A as it is translocated to its target.

Keywords: K+ efflux, Colicin A, Tol-proteins, E. coli

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Authors: Venkat Garigapati

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Cell-based therapies are limited due to underlying host immune system activity. Microencapsulation of living cells to overcome this issue has some serious drawbacks, such as limitations of nutrient and oxygen diffusion, which pose a threat to the function and longevity of cells. The conformal coating could overcome the issues which are generally involved in traditional microencapsulation. Some of the theoretical advantages of conformal coating include superior nutrient and oxygen supply to cells, prolonged lifespan, improved drug-secreting cell functionality and an opportunity to load high cell doses in small volumes. Despite several advantages to the conformal coating, there are no suitable methods available to apply to living cells. The ultra-thin conformal coating was achieved utilizing click-reactive methacryloyloxyethyl phosphorylcholine (MPC) polymers, which are capable of specifically reacting one polymer to another at neutral pH in the aqueous isotonic system at the desired temperature suitable for living cells without the need of deleterious initiators. ARPE-19 (Adult Retinal Pigment Epithelial cell line-19) cell-spheroids and rat pancreatic islets were used in the formulation studies. The in vitro studies of coated ARPE-19 cell-spheroids and rat islets indicate that the coat was intact; cells were viable and functioning. The in vitro study results revealed that the conformal coating technology seems promising and in vivo studies are being planned.

Keywords: cells, hydrogel, conformal coating, microencapsulation, insulin

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Authors: Shringika Soni, Utkarsh Jain, Nidhi Chauhan

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Aptamers are recently discovered ~80-100 bp long artificial oligonucleotides that not only demonstrated their applications in therapeutics; it is tremendously used in diagnostic and sensing application to detect different biomarkers and drugs. Synthesizing aptamers for proteins or genomic template is comparatively feasible in laboratory, but drugs or other chemical target based aptamers require major specification and proper optimization and validation. One has to optimize all selection, amplification, and characterization steps of the end product, which is extremely time-consuming. Therefore, we performed asymmetric PCR (polymerase chain reaction) for random oligonucleotides pool synthesis, and further use them in Systematic evolution of ligands by exponential enrichment (SELEX) for anti-psychotic drugs based aptamers synthesis. Anti-psychotic drugs are major tranquilizers to control psychosis for proper cognitive functions. Though their low medical use, their misuse may lead to severe medical condition as addiction and can promote crime in social and economical impact. In this work, we have approached the in-vitro SELEX method for ssDNA synthesis for anti-psychotic drugs (in this case ‘target’) based aptamer synthesis. The study was performed in three stages, where first stage included synthesis of random oligonucleotides pool via asymmetric PCR where end product was analyzed with electrophoresis and purified for further stages. The purified oligonucleotide pool was incubated in SELEX buffer, and further partition was performed in the next stage to obtain target specific aptamers. The isolated oligonucleotides are characterized and quantified after each round of partition, and significant results were obtained. After the repetitive partition and amplification steps of target-specific oligonucleotides, final stage included sequencing of end product. We can confirm the specific sequence for anti-psychoactive drugs, which will be further used in diagnostic application in clinical and forensic set-up.

Keywords: anti-psychotic drugs, aptamer, biosensor, ssDNA, SELEX

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Authors: O. I. Papagianni, P. Potsaki, K. Almpounioti, D. Chatzicharalampous, A. Voutsa, O. Katira, A. Michalaki, H. C. Karantonis, A. E. Koutelidakis

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High intakes of carbohydrates and fats have been associated with an increased risk of chronic diseases due to the role of postprandial oxidative stress. This pilot nutritional intervention aimed to examine the acute effect of consuming two different types of breadsticks prepared from durum wheat flour mixtures differing in total phenolic content on postprandial inflammatory and oxidant responses in healthy volunteers. A cross-over, controlled, and single-blind clinical trial was designed, and two isocaloric high-fat and high-carbohydrate meals were tested. Serum total, HDL- and LDL-cholesterol, triglycerides, glucose, CRP, uric acid, plasma total antioxidant capacity, and antiplatelet activity were determined in fasting and 30, 60, and 120 min after consumption. The results showed a better postprandial HDL-cholesterol and total antioxidant activity response in the intervention group. The choice of durum wheat flours with higher phenolic content and antioxidant activity is presented as promising for human health, and clinical studies will expand to draw safer conclusions.

Keywords: breadsticks, durum wheat flours, postprandial inflammation, postprandial oxidative stress, ex vivo antiplatelet activity

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Authors: Ľudmila Ballová, Slavomír Kurhajec, Eva Petrovová, Jarmila Eftimová

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Angiogenesis, a formation of new blood vessels from a pre-existing vasculature, plays an important role in pathologic processes such as the growth and metastasis of tumours. Tumours cannot grow beyond a few millimetres without blood supply from the newly formed blood vessels from the host tissue, a process called tumour-induced angiogenesis. The successful research of antiangiogenic treatment of cancer has focused on nutraceuticals with angiogenesis-modulating properties. Berberine, as a major active component of the bark of Phellodendron amurense Rupr., has shown antitumour activity by intervening into different steps of carcinogenesis. The influence of ethanolic extract of Phellodendron amurese bark on the angiogenesis was tested in vivo on chick chorioallantoic membrane (CAM). The irritancy of the CAM after the application of the crude bark extract dissolved in normal saline (10 mg/mL) was investigated on embryonic day 7. No significant signs of the irritancy, such as vasoconstriction, hyperaemia, haemorrhage or coagulation were observed which indicates the harmless character of the extract. A significant reduction in vessel sprouting and higher percentage of avascular zone was observed in the case of CAM treated with the extract in comparison with non-treated CAM (control), which is a proof of the antiangiogenic potential of the extract. These results could contribute to the development of novel drugs for the treatment of cancer or other diseases, in which angiogenesis plays a significant role.

Keywords: angiogenesis, berberine, chorioallantoic membrane, irritancy, phellodendron amurense

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Authors: Priyanka Sharma, Tarkewshwar Dubey, Hemalatha Siva

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Hemorrhoids are one of the most common anorectal diseases around the world. Severalfactors are involved in causing hemorrhoids including irregularbowel function (constipation, diarrhea), exercise, gravity, low fiberdiet, pregnancy, obesity, high abdominal pressure, prolongedsitting, genetic factors, and aging. Pain, bleeding, itching,swelling and anal discharge are the symptoms of the disease. Due to limitedmodern pharmacotherapeutic options available for treatment, theherbal medicines remain the choice of therapy. Blumea lacera (Burm f.) DC. belonging to the Asteraceae family is a common plain land weed of Bangladesh. Traditionally it has been used for treatment of hemorrhoids.Considering the above fact, present study was aimed to validate the ethnomedicinal use of B. lacera leaves on experimental hemorrhoids in rats. The anti-hemorrhoid activity was performed by using croton oil induced rat models. The parameters studied were assessment of TNF-α and IL-6, Evans blue exudation, macroscopic severity score, recto-anal coefficient, histomorphological scores. Also, in vivo antioxidant parameters and histopathological studies were also performed. All paramaters exhibited significant anti-hemorrhoid activity. Moreover ethanolic extract of B. lacera (EBL) leaves 400mg/kg showed ameliorative effect oncroton oil induced hemorrhoids.In conclusion, EBL exhibitedbeneficial effect on croton oil- induced hemorrhoids and validates its ethnomedicinal use in treatment of piles.

Keywords: haemorrhoids, IL-6, piles, TNF-α

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Authors: Fatemeh Zeinali Sehrig

Abstract:

Background: miRNAs play an important role in the post-transcriptional regulation of genes, including genes involved in DNA methylation (DNMTs), and are also important regulators of oncogenic pathways. The study of microRNAs and DNMTs in breast cancer allows the development of targeted treatments and early detection of this cancer. Methods and Materials: Clinical Patients and Samples: Institutional guidelines, including ethical approval and informed consent, were followed by the Ethics Committee (Ethics code: IR.IAU.TABRIZ.REC.1401.063) of Tabriz Azad University, Tabriz, Iran. In this study, tissues of 100 patients with breast cancer and tissues of 100 healthy women were collected from Noor Nejat Hospital in Tabriz. The basic characteristics of the patients with breast cancer included: 1)Tumor grade(Grade 3 = 5%, Grade 2 = 87.5%, Grade 1 = 7.5%), 2)Lymph node(Yes = 87.5%, No = 12.5%), 3)Family cancer history(Yes = 47.5%, No = 41.3%, Unknown = 11.2%), 4) Abortion history(Yes = 36.2%).In silico methods (data gathering, process, and build networks): Gene Expression Omnibus (GEO), a high-throughput genomic database, was queried for miRNAs expression profiles in breast cancer. For Experimental protocol Tissue Processing, Total RNA isolation, complementary DNA(cDNA) synthesis, and quantitative real time PCR (QRT-PCR) analysis were performed. Results: In the present study, we found significant (p.value<0.05) changes in the expression level of miRNAs and DNMTs in patients with breast cancer. In bioinformatics studies, the GEO microarray data set, similar to qPCR results, showed a decreased expression of miRNAs and increased expression of DNMTs in breast cancer. Conclusion: According to the results of the present study, which showed a decrease in the expression of miRNAs and DNMTs in breast cancer, it can be said that these genes can be used as important diagnostic and therapeutic biomarkers in breast cancer.

Keywords: gene expression omnibus, microarray dataset, breast cancer, miRNA, DNMT (DNA methyltransferases)

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Authors: Brian M. Mehling, Louis Quartararo, Marine Manvelyan, Paul Wang, Dong-Cheng Wu

Abstract:

Numerous investigations suggest that Mesenchymal Stem Cells (MSCs) in general represent a valuable tool for therapy of symptoms related to chronic inflammatory diseases. Blue Horizon Stem Cell Therapy Program is a leading provider of adult and children’s stem cell therapies. Uniquely we have safely and efficiently treated more than 600 patients with documenting each procedure. The purpose of our study is primarily to monitor the immune response in order to validate the safety of intravenous infusion of human umbilical cord blood derived MSCs (UC-MSCs), and secondly, to evaluate effects on biomarkers associated with chronic inflammation. Nine patients were treated for conditions associated with chronic inflammation and for the purpose of anti-aging. They have been given one intravenous infusion of UC-MSCs. Our study of blood test markers of 9 patients with chronic inflammation before and within three months after MSCs treatment demonstrates that there is no significant changes and MSCs treatment was safe for the patients. Analysis of different indicators of chronic inflammation and aging included in initial, 24-hours, two weeks and three months protocols showed that stem cell treatment was safe for the patients; there were no adverse reactions. Moreover data from follow up protocols demonstrates significant improvement in energy level, hair, nails growth and skin conditions. Intravenously administered UC-MSCs were safe and effective in the improvement of symptoms related to chronic inflammation. Further close monitoring and inclusion of more patients are necessary to fully characterize the advantages of UC-MSCs application in treatment of symptoms related to chronic inflammation.

Keywords: chronic inflammatory diseases, intravenous infusion, stem cell therapy, umbilical cord blood derived mesenchymal stem cells (UC-MSCs)

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Authors: Mohd Adil, Rosina Khan, Danishuddin, Asad U. Khan

Abstract:

The aim of this study was to evaluate the influence of the crude and active solvent fraction of Trachyspermum ammi on S. mutans cariogenicity, effect on expression of genes involved in biofilm formation and caries development in rats. GC–MS was carried out to identify the major components present in the crude and the active fraction of T. ammi. The crude extract and the solvent fraction exhibiting least MIC were selected for further experiments. Scanning electron microscopy was carried out to observe the effect of the extracts on S. mutans biofilm. Comparative gene expression analysis was carried out for nine selected genes. 2-Isopropyl-5-methyl-phenol was found as major compound in crude and the active fraction. Binding site of this compound within the proteins involved in biofilm formation was mapped with the help of docking studies. Real-time RT-PCR analyses revealed significant suppression of the genes involved in biofilm formation. All the test groups showed reduction in caries (smooth surface as well as sulcal surface caries) in rats. Moreover, it also provides new insight to understand the mechanism influencing biofilm formation in S. mutans. Furthermore, the data suggest the putative cariostatic properties of T. Ammi and hence can be used as an alternative medicine to prevent caries infection.

Keywords: bio-film, Streptococcus mutans, dental caries, bio-informatic

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Authors: Rishav Shrestha, Yong Zhang

Abstract:

The Anti-Stoke upconversion process has been used extensively for bioimaging and is recently being used for photoactivated therapy in cancer utilizing upconversion nanoparticles (UCNs). The UCNs have an excitation band at 980nm; 980nm laser excitation used to produce UV/Visible emissions also produce a heating effect. Light-to-heat conversion has been observed in nanoparticles(NPs) doped with neodymium(Nd) or ytterbium(Yb)/erbium(Er) ions. Despite laser-induced heating in Rare-earth doped NPs being proven to be a relatively efficient process, only few attempts to use them as photothermal agents in biosystems have been made up to now. Gold nanoparticles and carbon nanotubes are the most researched and developed for photothermal applications. Both have large heating efficiency and outstanding biocompatibility. However, they show weak fluorescence which makes them harder to track in vivo. In that regard, UCNs are attractive due to their excellent optical features in addition to their light-to-heat conversion and excitation by NIR, for imaging and spatiotemporally releasing drugs. In this work, we have utilized a simple method to coat Nd doped UCNs with thermoresponsive polymer PNIPAM on which 4-Hydroxytamoxifen (4-OH-T) is loaded. Such UCNs demonstrate a high loading efficiency and low leakage of 4-OH-T. Encouragingly, the release of 4-OH-T can be modulated by varying the power and duration of the NIR. Such UCNs were then used to demonstrate imaging and controlled photothermal release of 4-OH-T in MCF-7 breast cancer cells.

Keywords: cancer therapy, controlled release, photothermal release, upconversion nanoparticles

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Authors: Iman A. A. Kassem, Ayman A. Farghaly, Zeinab M. Hassan, Farouk R. Melek, Neveen S. Ghaly

Abstract:

Lipidium sativum L, an annual herb that grows to 50 cm, is known as an important member of family Brassicaceae. Besides its nutritional value, the seeds were widely used in folk medicine for treatment of cough, asthma, and headache. It was also reported to possess hypocholesterolemic, anti-inflammatory, antidiarrheal, antimicrobial and anticancer activities. In this study, the genoprotective properties of L. sativum seed methanolic extract (LSME) were evaluated in vivo. Three groups of mice were given LSME for five consecutive days at the three dose levels 25, 50 and 100 mg/kg b.wt. The three groups were then injected intraperitoneally with cyclophosphamide at a dose of 20 mg/kg b.wt. to induce DNA damage. A group received only cyclophosphamide (20 mg/kg b.wt.) served as control. LSME significantly inhibited the DNA aberrations in mice caused by cyclophosphamide in a dose-dependent manner in the two groups that received LSME at 50 and 100 mg/kg b.wt. dose levels. The chromosomal aberrations' inhibitory indices were calculated as 18 and 31 in mice bone marrow cells and 27 and 48 in mice spermatocytes, respectively. Phytochemical examination carried out by us revealed that flavonoids were the main chemical constituents of LSME. The major flavonoids kaempferol, kaempferol-3-O-rhamnoside, kaempferol-3-O-glucoside, quercetin, and quercetin-3-O-glucoside were isolated and characterized. It was concluded that the genoprotective effect of LSME might be attributed to the presence of flavonoids which are well-known for their antioxidant properties.

Keywords: cyclophosphamide, flavonoids, genoprotective effect, Lepidium sativum

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Authors: Mariam Azam, Sajjad Ur Rahman, Mukarram Bashir, Muhammad Tahir, Seemal Javaid, Jawad, Aoun Muhammad, Muhammad Zohaib, Hannan Khan

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Products of microbial origin are of great importance as they have proved their value in healthcare and nutrition, use of these microbial metabolites acquired from partially fermented soya hulls and wheat bran along with Saccharomyces cerevisiae (DL-22 S/N) substantiates to be a great source for an increase in the total milk production and quality yield.1×109 CFU/ml cells of Saccharomyces cerevisiae (DL-22 S/N) were further grown under in-vivo conditions for the assessment of quality milk production. Two groups with twelve cows, each having the same physical characteristics (Group A and Group B), were under study, Group A was daily fed with 12gm of biological metabolites and 22% protein-pelleted feed. On the other hand, the animals of Group B were provided with no metabolites in their feed. In thirty days of trial, improvement in the overall health, body score, milk protein, milk fat, yield, incidence rate of mastitis, ash, and solid not fat (SNF) was observed. The collected data showed that the average quality milk production was elevated up to 0.45 liter/h/d. However, a reduction in the milk fats up to 0.45% and uplift in the SNF value up to 0.53% of cow milk was also observed. At the same time, the incidence rate of mastitis recorded for the animals under trial was reduced to half, and improved non specific immunity was reported.

Keywords: microbial metabolites, post-biotics, animal supplements, animal nutrition, proteins, animal production, fermentation

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Authors: Sarim Ahmad

Abstract:

The Single Cell Gel Electrophoresis Assay (SCGE, known as comet assay) is a potential, uncomplicated, sensitive and state-of-the-art technique for quantitating DNA damage at individual cell level and repair from in vivo and in vitro samples of eukaryotic cells and some prokaryotic cells, being popular in its widespread use in various areas including human biomonitoring, genotoxicology, ecological monitoring and as a tool for research into DNA damage or repair in different cell types in response to a range of DNA damaging agents, cancer risk and therapy. The method involves the encapsulation of cells in a low-melting-point agarose suspension, lysis of the cells in neutral or alkaline (pH > 13) conditions, and electrophoresis of the suspended lysed cells, resulting in structures resembling comets as observed by fluorescence microscopy; the intensity of the comet tail relative to the head reflects the number of DNA breaks. The likely basis for this is that loops containing a break lose their supercoiling and become free to extend towards the anode. This is followed by visual analysis with staining of DNA and calculating fluorescence to determine the extent of DNA damage. This can be performed by manual scoring or automatically by imaging software. The assay can, therefore, predict an individual’s tumor sensitivity to radiation and various chemotherapeutic drugs and further assess the oxidative stress within tumors and to detect the extent of DNA damage in various cancerous and precancerous lesions of oral cavity.

Keywords: comet assay, single cell gel electrophoresis, DNA damage, early detection test

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Authors: Han Xiao

Abstract:

The development of nanomedicines has recently achieved several breakthroughs in the field of cancer treatment; however, the biocompatibility and targeted burst release of these medications remain a limitation, which leads to serious side effects and significantly narrows the scope of their applications. The self-assembly of intermediate filament protein (IFP) peptides was triggered by a hydrophobic cation drug 7-amino actinomycin D (7-AAD) to synthesize pH-activatable nanoparticles (NPs) that could simultaneously locate tumors and produce antitumor effects. The designed IFP peptide included a target peptide (arginine–glycine–aspartate), a negatively charged region, and an α-helix sequence. It also possessed the ability to encapsulate 7-AAD molecules through the formation of hydrogen bonds and hydrophobic interactions by a one-step method. 7-AAD molecules with excellent near-infrared fluorescence properties could be target delivered into tumor cells by NPs and released immediately in the acidic environments of tumors and endosome/lysosomes, ultimately inducing cytotoxicity by arresting the tumor cell cycle with inserted DNA. It is noteworthy that the IFP/7-AAD NPs tail vein injection approach demonstrated not only high tumor-targeted imaging potential, but also strong antitumor therapeutic effects in vivo. The proposed strategy may be used in the delivery of cationic antitumor drugs for precise imaging and cancer therapy.

Keywords: 7-amino actinomycin D, intermediate filament protein, nanoparticle, tumor image

Procedia PDF Downloads 138

Authors: Hye Kyung Kim, Myung-Gyou Kim, Kang-Hyun Leem

Abstract:

The deer velvet antler is well known for its traditional medicinal value and is widely used in the clinic. It has been considered to possess bone-strengthening activity. The goal of this study was to investigate the anti-osteoporotic effect of deer antler velvet on ovariectomized rats (OVX), and their possible mechanism of the action. In the first step, the in vitro effects of DAE on bone loss were determined. The proliferation, collagen content and alkaline phosphatase (ALP) activity of human osteoblastic MG-63 cells and osteoclastogenesis from bone marrow-derived precursor cells were measured. The in vivo experiment confirmed the positive effect of DAE on bone tissue. 3-month old female Sparague-Dawley rats were either sham operated or OVX, and administered DAE (20 and 100 mg/kg) for 4 weeks. DAE increased MG-63 cell proliferation and ALP activity in a dose-dependent manner. Collagen content was also increased by DAE treatment. However, the effect of DAE on bone resorption was not observed. OVX rats supplemented with DAE showed osteoprotective effects as the bone ALP level was increased and c-terminal telopeptide level was decreased by 100 mg/kg DAE treatment compared with OVX controls. Moreover, the tartrate-resistant acid phosphatase-5b level was also decreased by DAE treatment. The present study suggests that DAE is effective in preventing bone loss in OVX rats, and may be potential therapeutic agents for the treatment of postmenopausal osteoporosis.

Keywords: bone ALP, c-terminal telopeptide, deer antler, osteoporosis, ovariectomy, tartrate-resistant acid phosphatase-5b

Procedia PDF Downloads 245