scholarly journals Effect of Melatonin on Tau aggregation and Tau-mediated cell surface morphology

2019 ◽  
Author(s):  
Rashmi Das ◽  
Abhishek Ankur Balmik ◽  
Subashchandrabose Chinnathambi
Keyword(s):  
Cd Spectroscopy ◽  
Sem Analysis ◽  
Membrane Topology ◽  
Microtubule Assembly ◽  
Neuronal Protein ◽  
Transmission Electron ◽  
Tht Fluorescence ◽  
Melatonin Administration ◽  
Tau Aggregation

ABSTRACTTau is the major neuronal protein involved in the stabilization of microtubule assembly. In Alzheimer’s disease, Tau self assembles to form intracellular protein aggregates, which are toxic to cells. Various methods have been tried and tested to restrain the aggregation of Tau. Most of the agents tested for this purpose have limitations in their effectiveness and availability to neuronal cells. We tested melatonin against in vitro Tau aggregation and observed its effect on membrane topology, tubulin network and Tau phosphorylation in neuro2a and N9 cell lines. The aggregation and conformation of Tau was determined by ThT fluorescence and CD spectroscopy respectively. The morphology of Tau aggregates in presence and absence of melatonin was studied by transmission electron microscopy. Melatonin was found to reduce the formation of higher order oligomeric structures without affecting the overall aggregation kinetics of Tau. Melatonin also modulates and helps to maintain membrane topology as evidenced by FE-SEM analysis. Overall, melatonin administration shows mild anti-aggregation and cytoprotective effects.

Photo-excited Toluidine Blue Inhibits Full-Length Tau Aggregation in Alzheimer’ Disease

2019 ◽  
Author(s):  
Tushar Dubey ◽  
Nalini Gorantla ◽  
Kagepura Chandrashekara ◽  
Subashchandrabose Chinnathambi
Keyword(s):  
Toluidine Blue ◽  
Cell Biology ◽  
Cd Spectroscopy ◽  
Drosophila Model ◽  
Sds Page ◽  
Biophysical Analysis ◽  
Stressed Cells ◽  
Singlet Oxygen Species ◽  
Tau Aggregation

The aggregates of microtubule-associated protein Tau are the major hallmark of Alzheimer’sdisease. Tau aggregates accumulate intracellularly thus leading to generation of neuronal toxicity.Numerous approaches have been targeted against Tau protein aggregation, which include application of synthetic and natural compounds. Toluidine blue is a basic dye of phenothiazine family, which irradiation with 630 nm light converted to photo-excited form leading to generation of singlet oxygen species. In present work we studied the potency of Toluidine blue and photo-excited Toluidine blue against Tau aggregation. Biochemical and biophysical analysis using ThSfluorescence, SDS-PAGE, CD spectroscopy and electron microscopy suggested that Toluidine blueinhibits the aggregation of Tau in-vitro. The Photo-excited toluidine blue potentially dissolved the matured Tau fibrils, which was indicating disaggregation property of Toluidine blue. The cell biology studies including cytotoxicity assays, ROS production assays suggested Toluidine blue to be a biocompatible dye as reduced ROS levels and cytotoxicity was observed after exposure of Toluidine blue on Tau stressed cells. The photo-excited Toluidine blue modulates the cytoskeleton network in cells, which was supported by immunofluorescence studies of neuronal cells. The studies in UAS Tau E14 transgenic Drosophila model suggested that photo-excited Toluidine blue was potent to restore the survival and memory deficit of Drosophila. The overall findings of our studies suggests that Toluidine blue to be a potent molecule in rescuing the Tau-mediated pathology by inhibiting its aggregation, reducing the cytotoxicity; modulating the tubulin level and behavioral characteristics of Drosophila. Thus Toluidine blue can be addressed as a potent molecule against Alzheimer’s disease.

Photo-excited Toluidine Blue Inhibits Full-Length Tau Aggregation in Alzheimer’ Disease

2019 ◽  
Author(s):  
Tushar Dubey ◽  
Nalini Gorantla ◽  
Kagepura Chandrashekara ◽  
Subashchandrabose Chinnathambi
Keyword(s):  
Toluidine Blue ◽  
Cell Biology ◽  
Cd Spectroscopy ◽  
Drosophila Model ◽  
Sds Page ◽  
Biophysical Analysis ◽  
Stressed Cells ◽  
Singlet Oxygen Species ◽  
Tau Aggregation

The aggregates of microtubule-associated protein Tau are the major hallmark of Alzheimer’sdisease. Tau aggregates accumulate intracellularly thus leading to generation of neuronal toxicity.Numerous approaches have been targeted against Tau protein aggregation, which include application of synthetic and natural compounds. Toluidine blue is a basic dye of phenothiazine family, which irradiation with 630 nm light converted to photo-excited form leading to generation of singlet oxygen species. In present work we studied the potency of Toluidine blue and photo-excited Toluidine blue against Tau aggregation. Biochemical and biophysical analysis using ThSfluorescence, SDS-PAGE, CD spectroscopy and electron microscopy suggested that Toluidine blueinhibits the aggregation of Tau in-vitro. The Photo-excited toluidine blue potentially dissolved the matured Tau fibrils, which was indicating disaggregation property of Toluidine blue. The cell biology studies including cytotoxicity assays, ROS production assays suggested Toluidine blue to be a biocompatible dye as reduced ROS levels and cytotoxicity was observed after exposure of Toluidine blue on Tau stressed cells. The photo-excited Toluidine blue modulates the cytoskeleton network in cells, which was supported by immunofluorescence studies of neuronal cells. The studies in UAS Tau E14 transgenic Drosophila model suggested that photo-excited Toluidine blue was potent to restore the survival and memory deficit of Drosophila. The overall findings of our studies suggests that Toluidine blue to be a potent molecule in rescuing the Tau-mediated pathology by inhibiting its aggregation, reducing the cytotoxicity; modulating the tubulin level and behavioral characteristics of Drosophila. Thus Toluidine blue can be addressed as a potent molecule against Alzheimer’s disease.

Exploring the potential of pyrazoline containing molecules as Aβ aggregation inhibitors in Alzheimer’s disease

2020 ◽  
Vol 0 (0) ◽  
Author(s):  
Mihir Pramod Khambete ◽  
Lalit Pramod Khare ◽  
Akshay Bhupendra Kapadia ◽  
Mariam Sohel Degani
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Cd Spectroscopy ◽  
Aromatic Rings ◽  
Transmission Electron ◽  
Aβ Aggregation ◽  
Aggregation Inhibitors ◽  
Aggregation Inhibition ◽  
Amyloid Aβ

AbstractObjectivesAlzheimer’s disease (AD) is a chronic and progressive neurodegenerative disease in which one of the most prominent pathological features is accumulation of amyloid (Aβ) plaques. This occurs due to the process of aggregation from monomeric to polymeric forms of Aβ peptide and thus represents one of the attractive targets to treat AD.MethodsAfter initial evaluation of a set of molecules containing N-acetylpyrazoline moiety flanked by aromatic rings on both sides as Aβ aggregation inhibitors, the most potent molecules were further investigated for mechanistic insights. These were carried out by employing techniques such as circular dichroism (CD) spectroscopy, transmission electron microscopy (TEM), in vitro PAMPA-BBB (Blood–Brain Barrier) assay and cytotoxicity evaluation.ResultsTwo molecules among the exploratory set displayed Aβ aggregation inhibition comparable to standard curcumin. Among the follow-up molecules, several molecules displayed more inhibition than curcumin. These molecules displayed good inhibitory activity even at lower concentrations. CD and TEM confirmed the mechanism of Aβ aggregation. These molecules were found to alleviate Aβ induced cytotoxicity. BBB penetration studies highlighted the potential of these molecules to reach central nervous system (CNS).ConclusionsThus, several promising Aβ-aggregation inhibitors were obtained as a result of this study.

scholarly journals Micro-/Nano-Structured Ceramic Scaffolds That Mimic Natural Cancellous Bone

Materials ◽  
2021 ◽  
Vol 14 (6) ◽  
pp. 1439
Author(s):  
Anabel Díaz-Arca ◽  
Patricia Ros-Tárraga ◽  
María J. Martínez Tomé ◽  
Antonio H. De Aza ◽  
Luis Meseguer-Olmo ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Cancellous Bone ◽  
Cell Biology ◽  
Human Mesenchymal Stem Cells ◽  
Sem Analysis ◽  
Nano Structure ◽  
Transmission Electron ◽  
Surface Nanostructures ◽  
Surface Nanostructure

Micro-/nano-structured scaffolds with a weight composition of 46.6% α-tricalcium phosphate (α-TCP)—53.4% silicocarnotite (SC) were synthesized by the polymer replica method. The scanning electron microscopy (SEM) analysis of the scaffolds and natural cancellous bone was performed for comparison purposes. Scaffolds were obtained at three cooling rates via the eutectoid temperature (50 °C/h, 16.5 °C/h, 5.5 °C/h), which allowed the surface nanostructure and mechanical strength to be controlled. Surface nanostructures were characterized by transmission electron microscopy (TEM) and Raman analysis. Both phases α-TCP and SC present in the scaffolds were well-identified, looked compact and dense, and had neither porosities nor cracks. The non-cytotoxic effect was evaluated in vitro by the proliferation ability of adult human mesenchymal stem cells (ah-MSCs) seeded on scaffold surfaces. There was no evidence for cytotoxicity and the number of cells increased with culture time. A dense cell-hydroxyapatite layer formed until 28 days. The SEM analysis suggested cell-mediated extracellular matrix formation. Finally, scaffolds were functionalized with the alkaline phosphatase enzyme (ALP) to achieve biological functionalization. The ALP was successfully grafted onto scaffolds, whose enzymatic activity was maintained. Scaffolds mimicked the micro-/nano-structure and chemical composition of natural cancellous bone by considering cell biology and biomolecule functionalization.

scholarly journals FRET-based Tau seeding assay does not represent prion-like templated assembly of Tau fibers

2020 ◽  
Author(s):  
Senthilvelrajan Kaniyappan ◽  
Katharina Tepper ◽  
Jacek Biernat ◽  
Ram Reddy Chandupatla ◽  
Sabrina Hübschmann ◽  
...  
Keyword(s):  
Recipient Cell ◽  
Pathological State ◽  
Amyloid Fibers ◽  
Force Microscopy ◽  
Atomic Force ◽  
Transmission Electron ◽  
Repeat Domain ◽  
Scanning Transmission ◽  
Tau Aggregation

AbstractTau aggregation into amyloid fibers based on the cross-beta structure is a hallmark of several Tauopathies, including Alzheimer Disease (AD). Trans-cellular propagation of Tau with pathological conformation has been suggested as a key disease mechanism. This is thought to cause the spreading of Tau pathology in AD by templated conversion of naive Tau in recipient cells into a pathological state, followed by assembly of pathological Tau fibers, similar to the mechanism proposed for prion pathogenesis. In cell cultures, the process is usually monitored by a FRET assay where the recipient cell expresses the Tau repeat domain (TauRD, with pro-aggregant mutation, e.g., ΔK280 or P301L, ∼13.5 kDa) fused to GFP-based FRET pairs (YFP or CFP, ∼28 kD). Since the diameter of the reporter GFP (∼3 nm) is ∼6.5 times larger than the β-strand distance (0.47nm), this points to a potential steric clash. Hence, we investigated the influence of GFP tagged (N- or C-terminally) TauRD and TauFL (full-length Tau) on their aggregation behavior in vitro. Using biophysical methods (light scattering, atomic force microscopy (AFM), and scanning-transmission electron microscopy (STEM)), we found that the assembly of TauRDΔK-GFP was severely inhibited, even in the presence of nucleation enhancers (heparin and/or pre-formed PHFs from TauRDΔK). Some rare fiber-like particles had a very different subunit packing from proper PHFs, as judged by STEM. The mass per length (MPL) values of TauRDΔK fibrils are equivalent to 4.45 molecules/nm, close to the expected value for a paired-helical fiber with 2 protofilaments and cross-β structure. By contrast, the elongated particles formed by TauRDΔK-GFP have MPL values around ∼2, less than half of the values expected for PHFs, indicating that the subunit packing is distinct. Thus, both kinetic and structural observations are incompatible with a model whereby external Tau can form a template for PHF assembly of Tau-GFP in recipient cells. As a consequence, the observed local increase of FRET in recipient cells must be caused by other signalling processes.

Rapid Biological Synthesis of Silver Nanoparticles and It's In Vitro Anti-Bacterial and Larvicidal Activities

2020 ◽  
Vol 12 (5) ◽  
pp. 593-602
Author(s):  
Gattu Sampath ◽  
Douglas JH Shyu ◽  
Neelamegam Rameshkumar ◽  
Muthukalingan Krishnan ◽  
Krishnan Raguvaran ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Structural Damage ◽  
Sem Analysis ◽  
Biological Synthesis ◽  
Infrared Spectrometry ◽  
Average Crystalline Size ◽  
Transmission Electron ◽  
Golden Yellow ◽  
Larvicidal Activities

The current investigation explains the Acacia nilotica (A. nilotica) leaf extract capped silver nanoparticle (AgNPs) synthesis. The instant formation of AgNPs was confirmed by altering the color golden yellow to dark brown and the characteristic SPR band determines by the UV-visible spectrometer. The structural specification was assessed using Fourier transform infrared spectrometry (FTIR), X-ray diffraction crystallography (XRD), High-resolution transmission electron microscopy, Atomic force microscopy (AFM), energy dispersive electron microscopy, zeta potential, dynamic light scattering. The particle size ranged 20–80 nm, the average crystalline size 35.26 nm were determined by HR-TEM and XRD. Further in vitro antimicrobial activity was tested against different bacteria by combining antibiotic with the AgNPs. Among all the tested bacteria, the Staphylococcus aureus (S. aureus) showed the highest (16.2 ± 0.6 mm) zone of inhibition. The Scanning electron microscope (SEM) analysis clearly showed the structural damage of S. aureus and Escherichia coli (E. coli) by increasing AgNPs concentration at 4 hr treatment. In addition, the larvicidal assay was performed against Culex quinquefasciatus (C. quinquefasciatus) and Aedes aegypti (A. aegypti). The higher efficiency of AgNPs was absorbed in A. aegypti (LC50 = 279 22 ppm) and C. quinque- fasciatus (LC50 = 168.98 ppm). The results proved that the An-AgNPs act as a multi-faceted effect on bacterial and controlling vectors.

Management of Ralstonia solanacearum in tomato using ZnO nanoparticles synthesized through Matricaria chamomilla

Plant Disease ◽  
2021 ◽  
Author(s):  
Raja Asad Ali Khan ◽  
Yuanyang Tang ◽  
Ishrat Naz ◽  
Syed Sartaj Alam ◽  
Wenzhao Wang ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Ralstonia Solanacearum ◽  
Sem Analysis ◽  
In Vivo Studies ◽  
Bacterial Cells ◽  
Matricaria Chamomilla ◽  
Bacterial Wilt Disease ◽  
Transmission Electron

Matricaria chamomilla flower extract was used as a biocompatible material for synthesis of zinc oxide nanoparticles (ZnONPs). The synthesized NPs were evaluated for their antibacterial potential in vitro and in vivo against Ralstonia solanacearum that causes devastating bacterial wilt disease in tomato and other crops. Synthesized ZnONPs were further analyzed by UV-Visible spectroscopy, fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), transmission electron microscopy (TEM) and scanning electron microscopy (SEM) with energy dispersive spectroscopy (EDS). The synthesized polydisperse ZnONPs were found to be in the size range of 8.9 to 32.6 nm, and at 18.0 µg ml-1 exhibited maximum in vitro growth inhibition of R. solanacearum. SEM analysis of affected bacterial cells showed morphological deformation such as disruption of cell membrane, cell wall and leakage of cell contents. Results of in vivo studies also showed that application of ZnONPs to the artificially inoculated tomato plants with R. solanacearum significantly enhanced the plant growth by reducing bacterial soil population and disease severity as compared to untreated control. Biosynthesized ZnONPs could be an effective approach to control R. solanacearum.

Maytansine-Induced Mitotic Arrest in Human Lymphoblasts

1977 ◽  
Vol 35 ◽  
pp. 460-461
Author(s):  
Tai-Te Chao ◽  
John Sullivan ◽  
Awtar Krishan
Keyword(s):  
Electron Microscopy ◽  
Cell Cycle ◽  
Transmission Electron Microscopy ◽  
Tubulin Polymerization ◽  
Vinca Alkaloids ◽  
Antimitotic Activity ◽  
Transmission Electron ◽  
Flow Microfluorometry ◽  
Brain Tubulin

Maytansine, a novel ansa macrolide (1), has potent anti-tumor and antimitotic activity (2, 3). It blocks cell cycle traverse in mitosis with resultant accumulation of metaphase cells (4). Inhibition of brain tubulin polymerization in vitro by maytansine has also been reported (3). The C-mitotic effect of this drug is similar to that of the well known Vinca- alkaloids, vinblastine and vincristine. This study was carried out to examine the effects of maytansine on the cell cycle traverse and the fine struc- I ture of human lymphoblasts.Log-phase cultures of CCRF-CEM human lymphoblasts were exposed to maytansine concentrations from 10-6 M to 10-10 M for 18 hrs. Aliquots of cells were removed for cell cycle analysis by flow microfluorometry (FMF) (5) and also processed for transmission electron microscopy (TEM). FMF analysis of cells treated with 10-8 M maytansine showed a reduction in the number of G1 cells and a corresponding build-up of cells with G2/M DNA content.

Correlation Between Cellular Functions and Morphological Changes of Human Trophoblast in Vitro

1975 ◽  
Vol 33 ◽  
pp. 600-601
Author(s):  
John C. Garancis ◽  
Robert O. Hussa ◽  
Michael T. Story ◽  
Donald Yorde ◽  
Roland A. Pattillo
Keyword(s):  
Electron Microscopy ◽  
Cellular Proliferation ◽  
Morphological Changes ◽  
Culture Fluid ◽  
Cellular Functions ◽  
Human Trophoblast ◽  
Transmission Electron ◽  
Marked Activity ◽  
Malignant Trophoblast

Human malignant trophoblast cells in continuous culture were incubated for 3 days in medium containing 1 mM N6-O2'-dibutyryl cyclic adenosine 3':5'-monophosphate (dibutyryl cyclic AMP) and 1 mM theophylline. The culture fluid was replenished daily. Stimulated cultures secreted many times more chorionic gonadotropin and estrogens than did control cultures in the absence of increased cellular proliferation. Scanning electron microscopy revealed remarkable surface changes of stimulated cells. Control cells (not stimulated) were smooth or provided with varying numbers of microvilli (Fig. 1). The latter, usually, were short and thin. The surface features of stimulated cells were considerably different. There was marked increase of microvilli which appeared elongated and thick. Many cells were covered with confluent polypoid projections (Fig. 2). Transmission electron microscopy demonstrated marked activity of cytoplasmic organelles. Mitochondria were increased in number and size; some giant forms with numerous cristae were observed.

Hemolysis of erythrocytes by the hemolytic system from the blood-feeding stable fly, Stomoxys calcitrans

1992 ◽  
Vol 50 (1) ◽  
pp. 690-691
Author(s):  
H. J. Kirch ◽  
G. Spates ◽  
R. Droleskey ◽  
W.J. Kloft ◽  
J.R. DeLoach
Keyword(s):  
Blood Feeding ◽  
Stomoxys Calcitrans ◽  
Stable Fly ◽  
Digestive Physiology ◽  
Transmission Electron ◽  
Erythrocyte Lysis ◽  
Mammalian Blood ◽  
Bovine Erythrocytes ◽  
Potential Tool

Blood feeding insects have to rely on the protein content of mammalian blood to insure reproduction. A substantial quantity of protein is provided by hemoglobin present in erythrocytes. Access to hemoglobin is accomplished only via erythrocyte lysis. It has been shown that midgut homogenates from the blood feeding stable fly, Stomoxys calcitrans, contain free fatty acids and it was proposed that these detergent-like compounds play a major role as hemolysins in the digestive physiology of this species. More recently sphingomyelinase activity was detected in midgut preparations of this fly, which would provide a potential tool for the enzymatic cleavage of the erythrocyte's membrane sphingomyelin. The action of specific hemolytic factors should affect the erythrocyte's morphology. The shape of bovine erythrocytes undergoing in vitro hemolysis by crude midgut homogenates from the stable fly was examined by scanning and transmission electron microscopy.

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