scholarly journals Potential utilization of glycerol as crosslinker in starch films for application in Regenerative Dentistry

2021 ◽  
Vol 10 (16) ◽  
pp. e148101623640
Author(s):  
Simone Rodrigues ◽  
Mariana Fornazier ◽  
Denildo Magalhães ◽  
Reinaldo Ruggiero
Keyword(s):  
Bone Substitutes ◽  
In Vitro Assays ◽  
Mechanical Resistance ◽  
Protein Diffusion ◽  
Regenerative Dentistry ◽  
Physical Barriers ◽  
Starch Films ◽  
And Diffusion

Periodontal disease results in damage to dental insertion apparatus. Regenerative procedures are proposed to replace lost structures in the context of guided tissue regeneration (GTR), guided bone regeneration (GBR) techniques and frequently associate bone substitutes with physical barriers aiming at greater longevity and improvement of aesthetic pattern. This study evaluates the possibility of using glycerol as a starch films modifying agent, acting as a cross-linking agent, without compromising its plasticizing effect. Biodegradable cassava starch films were prepared incorporating glycerol at concentrations of 0, 15, 20, 30 and 40% aiming application at dental regenerative procedures. The characterization of films by microscopy (SEM), thermal analysis (DSC), spectroscopic (UV / Vis., FTIR, XRD), mechanical (Traction), and analysis of protein swelling, degradation and diffusion and physiological temperature) showed that the incorporation of glycerol in up to 20% attributed to the films a plasticizer character and in higher concentrations, conferred a greater interaction of glycerol (crosslinking) with the starch chains and a degradation time that allows the physical barrier in RTG and ROG. The films presented mechanical resistance, malleability and permissiveness to protein diffusion in the in vitro assays, which meet the current attributes that guide the use of these resources in biomaterials.

scholarly journals Functional Characterization of the mazEF Toxin-Antitoxin System in the Pathogenic Bacterium Agrobacterium tumefaciens

2021 ◽  
Vol 9 (5) ◽  
pp. 1107
Author(s):  
Wonho Choi ◽  
Yoshihiro Yamaguchi ◽  
Ji-Young Park ◽  
Sang-Hyun Park ◽  
Hyeok-Won Lee ◽  
...  
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Physiological Function ◽  
Functional Characterization ◽  
Site Directed Mutagenesis ◽  
In Vitro Assays ◽  
Cell Growth Inhibition ◽  
The Right

Agrobacterium tumefaciens is a pathogen of various plants which transfers its own DNA (T-DNA) to the host plants. It is used for producing genetically modified plants with this ability. To control T-DNA transfer to the right place, toxin-antitoxin (TA) systems of A. tumefaciens were used to control the target site of transfer without any unintentional targeting. Here, we describe a toxin-antitoxin system, Atu0939 (mazE-at) and Atu0940 (mazF-at), in the chromosome of Agrobacterium tumefaciens. The toxin in the TA system has 33.3% identity and 45.5% similarity with MazF in Escherichia coli. The expression of MazF-at caused cell growth inhibition, while cells with MazF-at co-expressed with MazE-at grew normally. In vivo and in vitro assays revealed that MazF-at inhibited protein synthesis by decreasing the cellular mRNA stability. Moreover, the catalytic residue of MazF-at was determined to be the 24th glutamic acid using site-directed mutagenesis. From the results, we concluded that MazF-at is a type II toxin-antitoxin system and a ribosome-independent endoribonuclease. Here, we characterized a TA system in A. tumefaciens whose understanding might help to find its physiological function and to develop further applications.

Abstract 3228: Development and characterization of in vitro assays to detect and quantitate tubulysin B hydrazide in biological samples

2017 ◽  
Author(s):  
Nikki L. Parker ◽  
Jonathan M. Shillingford ◽  
Melissa Nelson ◽  
Joseph A. Reddy ◽  
Christopher P. Leamon
Keyword(s):  
Biological Samples ◽  
In Vitro Assays

scholarly journals Method combining BAC film and positive staining for the characterization of DNA intermediates by dark-field electron microscopy

2020 ◽  
Vol 5 (1) ◽  
Author(s):  
Yann Benureau ◽  
Eliana Moreira Tavares ◽  
Ali-Akbar Muhammad ◽  
Sonia Baconnais ◽  
Eric Le Cam ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Dark Field ◽  
In Vitro Assays ◽  
Positive Staining ◽  
Synthetic Dna ◽  
Dark Field Imaging ◽  
Alkyl Ammonium

Abstract DNA intermediate structures are formed in all major pathways of DNA metabolism. Transmission electron microscopy (TEM) is a tool of choice to study their choreography and has led to major advances in the understanding of these mechanisms, particularly those of homologous recombination (HR) and replication. In this article, we describe specific TEM procedures dedicated to the structural characterization of DNA intermediates formed during these processes. These particular DNA species contain single-stranded DNA regions and/or branched structures, which require controlling both the DNA molecules spreading and their staining for subsequent visualization using dark-field imaging mode. Combining BAC (benzyl dimethyl alkyl ammonium chloride) film hyperphase with positive staining and dark-field TEM allows characterizing synthetic DNA substrates, joint molecules formed during not only in vitro assays mimicking HR, but also in vivo DNA intermediates.

Determinants of platelet number and regulation of thrombopoiesis

Hematology ◽  
2009 ◽  
Vol 2009 (1) ◽  
pp. 147-152 ◽  
Author(s):  
Kenneth Kaushansky
Keyword(s):  
Stem Cell ◽  
Progenitor Cell ◽  
In Vitro Assays ◽  
Cellular Mechanisms ◽  
Hematopoietic Stem ◽  
Platelet Production ◽  
Progenitor Cell Proliferation ◽  
Basic Biology

Abstract Our understanding of thrombopoiesis has improved greatly in the last two decades with the availability of in vitro assays of megakaryocyte progenitor cell growth, with the cloning and characterization of stem cell factor (SCF) and thrombopoietin (Tpo), the latter the primary humoral regulator of this process, and with the generation of genetically altered murine models of thrombopoietic failure and excess. While SCF affects developmentally early aspects of megakaryocyte growth, Tpo affects nearly all aspects of platelet production, from hematopoietic stem cell (HSC) self-renewal and expansion, through stimulation of megakaryocyte progenitor cell proliferation, to supporting their maturation into platelet-producing cells. The molecular and cellular mechanisms through which the marrow microenvironment and humoral mediators affect platelet production provide new insights into the interplay between intrinsic and extrinsic influences on hematopoiesis, and highlight new opportunities to translate basic biology into clinical advances.

scholarly journals YPR139c/LOA1encodes a novel lysophosphatidic acid acyltransferase associated with lipid droplets and involved in TAG homeostasis

2012 ◽  
Vol 23 (2) ◽  
pp. 233-246 ◽  
Author(s):  
Sophie Ayciriex ◽  
Marina Le Guédard ◽  
Nadine Camougrand ◽  
Gisèle Velours ◽  
Mario Schoene ◽  
...  
Keyword(s):  
Lysophosphatidic Acid ◽  
Lipid Droplets ◽  
Neutral Lipids ◽  
In Vitro Assays ◽  
Metabolic Labeling ◽  
Lysophosphatidic Acid Acyltransferase ◽  
Escherichia Coli Cells ◽  
Type Strains

For many years, lipid droplets (LDs) were considered to be an inert store of lipids. However, recent data showed that LDs are dynamic organelles playing an important role in storage and mobilization of neutral lipids. In this paper, we report the characterization of LOA1 (alias VPS66, alias YPR139c), a yeast member of the glycerolipid acyltransferase family. LOA1 mutants show abnormalities in LD morphology. As previously reported, cells lacking LOA1 contain more LDs. Conversely, we showed that overexpression results in fewer LDs. We then compared the lipidome of loa1Δ mutant and wild-type strains. Steady-state metabolic labeling of loa1Δ revealed a significant reduction in triacylglycerol content, while phospholipid (PL) composition remained unchanged. Interestingly, lipidomic analysis indicates that both PLs and glycerolipids are qualitatively affected by the mutation, suggesting that Loa1p is a lysophosphatidic acid acyltransferase (LPA AT) with a preference for oleoyl-CoA. This hypothesis was tested by in vitro assays using both membranes of Escherichia coli cells expressing LOA1 and purified proteins as enzyme sources. Our results from purification of subcellular compartments and proteomic studies show that Loa1p is associated with LD and active in this compartment. Loa1p is therefore a novel LPA AT and plays a role in LD formation.

scholarly journals RBP-L, a transcription factor related to RBP-Jkappa.

1997 ◽  
Vol 17 (5) ◽  
pp. 2679-2687 ◽  
Author(s):  
S Minoguchi ◽  
Y Taniguchi ◽  
H Kato ◽  
T Okazaki ◽  
L J Strobl ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Notch Receptor ◽  
In Vitro Assays ◽  
Functional Interaction ◽  
Protein Protein Interaction ◽  
Isolation And Characterization ◽  
Signalling Molecule

RBP-Jkappa is a sequence-specific DNA binding protein which plays a central role in signalling downstream of the Notch receptor by physically interacting with its intracellular region. Although at least four Notch genes exist in mammals, it is unknown whether each Notch requires a specific downstream signalling molecule. Here we report isolation and characterization of a mouse RBP-Jkappa-related gene named RBP-L that is expressed almost exclusively in lung, in contrast to the ubiquitous expression of RBP-Jkappa. For simplicity, we propose to call RBP-Jkappa RBP-J. The RBP-L protein bound to a DNA sequence almost identical to that of RBP-J. Surprisingly, RBP-L did not interact with any of the known four mouse Notch proteins. Although we found that RBP-L and EBNA-2 cooperated in transcriptional activation, they did not show significantly strong protein-protein interaction that can be detected by several in vivo and in vitro assays. This is again in contrast to physical association of RBP-J with EBNA-2. Several models to explain functional interaction between RBP-L and EBNA-2 are discussed.

scholarly journals Screening Assays for Epigenetic Targets Using Native Histones as Substrates

2011 ◽  
Vol 17 (1) ◽  
pp. 18-26 ◽  
Author(s):  
Alexander-Thomas Hauser ◽  
Elisabeth-Maria Bissinger ◽  
Eric Metzger ◽  
Antje Repenning ◽  
Uta-Maria Bauer ◽  
...  
Keyword(s):  
In Vitro Assays ◽  
The Past ◽  
Chemistry Research ◽  
Core Histones ◽  
Homogeneous Assay ◽  
Screening Assays ◽  
Set Up ◽  
Identification And Characterization

In the past years, a lot of attention has been given to the identification and characterization of selective and potent inhibitors of chromatin-modifying enzymes to better understand their specific role in transcriptional regulation. As aberrant histone methylation is involved in different pathological processes, the search for methyltransferase and demethylase inhibitors has emerged as a crucial issue in current medicinal chemistry research. High-throughput in vitro assays are important tools for the identification of new methyltransferase or demethylase inhibitors. These usually use oligopeptide substrates derived from histone sequences, although in many cases, they are not good substrates for these enzymes. Here, the authors report about the setup and establishment of in vitro assays that use native core histones as substrates, enabling an assay environment that better resembles native conditions. They have applied these substrates for the known formaldehyde dehydrogenase assay for the histone demethylase LSD1 and have established two new antibody-based assays. For LSD1, a heterogeneous assay format was set up, and a homogeneous assay was used for the characterization of the arginine methyltransferase PRMT1. Validation of the system was achieved with reference inhibitors in each case.

scholarly journals Use of in vivo and in vitro assays for the characterization of mammalian excision repair and isolation of repair proteins

1990 ◽  
Vol 236 (2-3) ◽  
pp. 223-238 ◽  
Author(s):  
J.H.J. Hoeijmakers ◽  
A.P.M. Eker ◽  
R.D. Wood ◽  
P. Robins
Keyword(s):  
Excision Repair ◽  
In Vitro Assays

3D Microtomography Characterization of Dental Implantology Bone Substitutes Used In Vivo

2013 ◽  
Vol 541 ◽  
pp. 97-113 ◽  
Author(s):  
Rossella Bedini ◽  
Deborah Meleo ◽  
Raffaella Pecci
Keyword(s):  
Bone Substitute ◽  
Histological Analysis ◽  
Research Work ◽  
Bone Substitutes ◽  
Human Bone ◽  
Histological Techniques ◽  
In Vitro Characterization

After a short introduction to bone substitute biomaterials and X-ray microtomography, this article describes a research work carried out for in-vitro characterization of bone substitute biomaterials as well as for in-vivo investigation of human bone grafted with biomaterials. Three different bone substitute biomaterials have been analyzed in-vitro by means of 3D microtomographic technique, while human bone samples grafted with bone substitute biomaterials are investigated by 3D microtomography and histological techniques. 3D images of bone substitutes and human bone samples with biomaterials have been obtained, together with morphometric parameters, by microtomography . 2D histological images have also been obtained by traditional technique only for human bone samples with biomaterials. Compared to traditional histological analysis, 3D microtomography shows better results for investigating bone tissue and bone substitute biomaterial, and in a short time. Nevertheless, histological analysis remains the best technique for the observation of soft tissue and blood vessels.

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