Miracle-dz/UniFAR-FTUs huggingface dataset:
Chloroquine causes lysosomal dysfunction in neural retina and RPE: Implications for retinopathy

Chronic use of chloroquine has been shown to induce numerous pathophysiological defects in the retina. This drug has the ability to alter pH of intracellular compartments and lysosomal function of the retinal pigment epithelium (RPE) and retinal neurons may constitute the basis of chloroquine retinopathy. The aim of the current study was to investigate pathogenic alterations in retinal cells continuously exposed to chloroquine using appropriate in vivo and in vitro models. Male hooded Lister rats were implanted with osmotic mini pumps which released chloroquine continuously over a period of seven days. The eyes were processed for electron microscopy and ultrastructural abnormalities determined in the neural retina and quantified using stereology in the retinal pigment epithelium (RPE). RPE were also exposed to chloroquine in vitro and lysosomal pH changes were investigated using a pH sensitive probe. Degradative capacity was also analysed using FITC labeled rod outer segments (ROS). Chloroquine-treated animals displayed several ultra-structural abnormalities including numerous membranous cytoplasmic bodies (MCBs) in retinal neurons. Cone photoreceptors displayed numerous MCBs although rods did not. The RPE of the treated groups all showed significantly higher numbers of lysosomal associated organelles (LAO) than the control group (p < 0.001). The in vitro experiments demonstrated chloroquine-mediated rises in lysosomal pH and an increase in lysosome/phagosome accumulation of ROS in the chloroquine treated group (p < 0.01). The current study demonstrates that chloroquine disrupts lysosomal function in retinal neurons and RPE. The evidence presented provides a clear pathogenic basis for the functional defects experienced by patients with chloroquine retinopathy.
Monitoring of metabolite gradients in tissue-engineered constructs

At present, the assessment of developing tissue-engineered constructs is almost always carried out destructively using biochemical or histological methods to determine cell number, viability and tissue growth throughout the construct. Since many of these experiments are long, taking weeks or even months to complete, simple and readily applicable non-destructive methods of monitoring changes in cell metabolism, viability and tissue deposition within the construct would be invaluable; such methods could point out adverse responses during the early stages of culture. Here, we describe the use of microdialysis for detecting local changes in cellular metabolism within a tissue-engineered construct. Three-dimensional constructs consisting of bovine articular chondrocytes entrapped in an alginate gel were cultured in a bioreactor for two weeks. Glucose and lactate were monitored by microdialysis, as the major nutrient and metabolite, respectively. Concentration gradients within the construct were evident, with the highest lactate concentrations in the construct centre. The local lactate concentration was a measure of cellular metabolic activity, decreasing as cellular activity fell and increasing as cellular activity was stimulated. Nutrient starvation and cell death in the construct centre could be readily detected in constructs deliberately cultured under adverse conditions. The results show that probe measurements can give an early warning of inappropriate local metabolic changes. Such information during the growth of tissue-engineered constructs would allow either corrective action or else an early end to an unsuccessful test.
The Immunomodulatory Properties of Periodontal Ligament Stem Cells Isolated from Inflamed Periodontal Granulation

Periodontitis is currently the main cause of tooth loss and as yet there is no appropriate method for establishing a functional and predictable periodontal regeneration. Tissue engineering involving seed cells provides a new prospect for periodontal regeneration. While periodontal ligament stem cells (PDLSCs) are a good choice for seed cells, it is not always possible to obtain the patients' own PDLSCs. We and others have found a type of stromal cells from inflamed periodontal granulation. These cells displayed similar differentiation properties to PDLSCs. Inflammation has a profound influence on the immunomodulatory properties of mesenchymal stem cells, which may affect therapeutic outcome. In this study, we assessed the immunomodulatory characteristics of these inflamed human (ih)PDLSCs. Along with the similarity in cell surface marker expressions, they also displayed immunomodulatory properties comparable to those in healthy human (hh)PDLSCs. Both hhPDLSCs and ihPDLSCs can suppress the proliferation and secretion of IFN-γ in peripheral blood mononuclear cells by indirect soluble mediators and direct cell-cell contact. Albeit with some quantitative variances, the gene expressions of inducible nitric oxide synthases, indoleamine 2,3 dioxygenase, cyclooxygenase-2, TNF-α-induced protein 6 and IL-10 in ihPDLSCs displayed similar patterns as those in hhPDLSCs. Taken together, our results suggest that ihPDLSCs can provide a promising alternative to hhPDLSCs in terms of evident similarities in immunomodulatory properties as well as their easier accessibility and availability.
Dimeric arrangement and structure of the membrane‐bound acetylcholine receptor studied by electron microscopy.

The acetylcholine receptor protein (AChR) from the electric organ of Torpedo marmorata is studied in its membrane‐bound form by electron microscopy and single‐particle image averaging. About half the molecule protrudes from the membrane surface by approximately 5 nm. The low‐resolution 3‐D structure of this hydrated portion, including its handedness, can be deduced from averaged axial and lateral projections and from freeze‐etched membrane surfaces. In native membrane fragments, a dimeric form of the AChR is observed and the relative orientation of the AChR monomers within the dimer is established. The dimers disappear upon disulfide reduction of the membrane preparations, whereas the average axial projections of the AChR monomer remain unaffected. Since the existence of disulfide bonds linking AChR monomers between their respective delta‐subunits is well documented, the approximate position of the delta‐subunit within the low‐resolution structure of the AChR molecule can be deduced from the structure of the dimers.
The structure of the mite allergen Blo t 1 explains the limited antibody cross‐reactivity to Der p 1

The Blomia tropicalis (Blo t) mite species is considered a storage mite in temperate climate zones and an important source of indoor allergens causing allergic asthma and rhinitis in tropical and subtropical regions. Here, we report the crystal structure of one of the allergens from Blo t, recombinant proBlo t 1 (rproBlo t 1), determined at 2.1 Å resolution. Overall, the fold of rproBlo t 1 is characteristic for the pro‐form of cysteine proteases from the C1A class. Structural comparison of experimentally mapped Der f 1/Der p1 IgG epitopes to the same surface patch on Blo t 1, as well as of sequence identity of surface‐exposed residues, suggests limited cross‐reactivity between these allergens and Blo t 1. This is in agreement with ELISA inhibition results showing that, although cross‐reactive human IgE epitopes exist, there are unique IgE epitopes for both Blo t 1 and Der p 1.
The effect of selected “desirable difficulties” on the ability to recall anatomy information

“Desirable difficulties” is a theory from cognitive science used to promote learning in a variety of contexts. The basic premise is that creating a cognitively challenging environment at the learning acquisition phase, by actively engaging learners in the retrieval of to‐be‐learned materials, promotes long‐term retention. In this study, the degree of desirable difficulties was varied to identify how cognitively challenging the learning acquisition phase must be to benefit university‐level students' learning of anatomy concepts. This is important to investigate as applied studies of desirable difficulties are less frequent than laboratory‐based studies and the implementation of this principle may need to be tailored to the specific field of study, such as anatomy. As such, a read‐read‐read‐read (R‐R‐R‐R) condition was compared to read‐generate‐read‐generate (R‐G‐R‐G) and read‐test‐read‐test (R‐T‐R‐T) conditions. The three conditions varied in terms of how effortful the retrieval task was during the learning acquisition phase. R‐R‐R‐R required little effort because participants passively read the materials four times. R‐G‐R‐G required some effort to generate a response as participants completed a word fragment task during the learning acquisition phase. R‐T‐R‐T was thought to be most demanding as participants performed a free recall task twice during the learning phase. With regard to the absolute amount of anatomy information recalled, the R‐T‐R‐T condition was superior at both immediate and delayed (one week) assessment points. Thus, instructors and learners of anatomy would benefit from embedding more free recall components, or self‐testing, into university‐level course work or study practices. Anat Sci Educ 8: 395–403. © 2014 American Association of Anatomists.
Comparison of CART-based localization and SVMs-based localization in WSN

Localization of sensor nodes is essential for wireless sensor network when it is applied to the special applications. We formed two models to estimate the location of sensor nodes, CART-based localization and SVMs-based Localization. During the training process, the received signal strength of the reference nodes is selected as the input of two models and the location information is regarded as the output of two models. During the localization process, the decision trees of CART and support vector machines are used to estimate the location of blindfolded nodes. We demonstrate the practicality and feasibility of the two models through simulations in the 100m×100m area.
Power transformer modeling based on wide band impedance and admittance measurements

This paper evaluates the transient behavior of a single-phase, two-winding, 127/12 V, 12 VA signal transformer according to two different high-frequency models. The first model is based on the direct measurement of the transformer's impedance matrix, while the second model consists in a combination of the admittance matrix and voltage transfer measurements. Both are alternatives to obtain a model with more information on the transformer's behavior at frequencies ranging from 10 Hz to 2 MHz. The admittance matrix resulting from the inversion of the measured impedance matrix (in the first model) and the modified admittance matrix (in the second model) are approximated by rational functions via the vector fitting technique, which leads to two equivalent RLC models that are stable and ready to be used in EMTP-like programs. The obtained models are first compared in the frequency domain with measurements carried out at the transformer terminals. Then, in order to validate the developed models in the time domain, comparisons between the theoretical and measured results of the transferred voltages from the primary to the secondary windings of the transformer are made. These comparisons consider distinct load conditions and the application of impulse waves with different time characteristics.
The NY SAFE Act: County Clerks and County-State Relations

The NY SAFE Act is a controversial gun control law in New York State. Since its passage on January 15, 2013, it has been the source of a continuing controversy. The primary purpose of this study is to explore the reactions of New York State (elected) county clerks tasked with the law’s implementation. In this phenomenological study, semi-structured interviews were conducted with 11 county clerks across New York State. The researcher asked various questions about how the SAFE Act has affected their offices. The researcher used thematic analysis to organize themes, which reveal that a lack of or inadequate intergovernmental communication and the absence of any requests for input from the county clerks before the SAFE Act was passed has resulted in dissatisfaction among the clerks. Additionally, the SAFE Act has resulted in significant increase in administrative workload, and prompted a generally negative reaction from the county clerk’s constituencies. The researcher concludes with several administrative recommendations based on the study’s findings. THE NY SAFE ACT: COUNTY CLERKS AND COUNTY-STATE RELATIONS 3
Ganglion cell types of the turtle retina that project to the optic tectum: Intracellular HRP injections of retrogradely, rhodamine-marked cell bodies

Abstract The turtle retina has been shown to have a variety of different morphological ganglion cell types as well as distinct physiological ganglion cell types. The major projection of the retina to the brain in nonmammalian vertebrates is to the optic tectum. In this study, we address the question of which retinal ganglion cell types project to the optic tectum in the turtle. Fluorescent rhodamine-labeled microspheres were used to trace the retinal ganglion cell projection to the superficial layers of the optic tectum. The fluorescent ganglion cell somata, retrogradely marked by transport from the contralateral optic tectum, were impaled with micropipettes containing rhodamine-horseradish peroxidase solution and this dye was iontophoresed into the cells under visual control. Most of the morphological ganglion cell types described in Golgi studies (Kolb, 1982; Kolb et al., 1988) were stained. Thus, the small cell types G1, G2, G3, G5, G6, and G7; the medium-sized types G10, G11, G12, G13, and G14; and the large-sized types G15, G16, G19, G20, and G21 project to the optic tectum in the turtle. We have added a new type, G2a, which proves to have some differences from the original G2 in branching pattern. We were unable to stain the small type G4, the medium-sized types G8 and G9, and the large cell types G17 and G18; this suggests that they might not project to the superficial layers of the dorsolateral optic tectum, at least, in the turtle.
[Utilization of Maternal and Child Health handbook in Japan].

OBJECTIVES In Japan, Maternal and Child Health (MCH) handbook have been widely used for more than fifty years. However, there has been no evaluation research from the users' point of view. We therefore conducted a questionnaire survey of guardians to evaluate how they utilize MCH handbooks. METHODS A well-structured questionnaire survey was carried out in 231 municipalities, towns and villages of four prefectures (Gifu, Hiroshima, Niigata and Shizuoka) and one city (Yokohama) in November and December 1999. The targets were 13,271 guardians who visited health stations for 18-month examinations of their children and agreed to participated in our research. The questionnaire covered situation of usage and loss by guardians, utility from the users' perspective, and suggestions for improvement. RESULTS We obtained answers from 10,900 guardians. As for reading rate and writing rate, 98.3% of respondents had read and 97.8% of them had written down something in relation. Only 0.9% of respondents had lost this MCH handbook. Generally, 87.0% of respondents answered that MCH handbook was helpful for child bearing and 81.6% of them said the record for immunization was useful. However, 34.1% of respondents answered it was not simple to utilize MCH handbook and 60.6% of them requested more detail on child bearing. As for dental health, the completion rate for information was low and only 21.3% of respondents reported for the dental record was useful. CONCLUSIONS The research shows MCH handbook are highly utilized in Japan by almost all guardians. However, there are still problems to be solved; many guardians feel that they are not simple to use and the section on dental health is not highly particularly helpful. Improvements should be made in future in light of the viewpoints of guardians and parents.
Improvement of HIV-1 and Human T Cell Lymphotropic Virus Type 1 Replication-Dependent Vectors via Optimization of Reporter Gene Reconstitution and Modification with Intronic Short Hairpin RNA

ABSTRACT Cell-to-cell transmission is an efficient mechanism to disseminate human immunodeficiency virus type 1 (HIV-1) and human T cell lymphotropic virus type 1 (HTLV-1). However, it has been challenging to quantify the level of cell-to-cell transmission because the virus-producing cells cannot be easily distinguished from infected target cells. We have previously described replication-dependent vectors that can quantify infection events in cocultured cells. These vectors contain an antisense-oriented promoter and reporter gene interrupted by a sense-oriented intron from the human gamma-globin gene. This strategy prevents expression of the reporter gene in the transfected cells but permits its expression in target cells after infection. However, the gamma-globin intron is not efficiently removed by splicing in the aforementioned vectors, thereby reducing the level of reporter gene expression after transduction into target cells. Here, we used two approaches to improve the replication-dependent vectors. First, we improved the splicing events that remove the gamma-globin intron by optimizing the intron insertion site within the reporter gene. Second, we improved the packaging of the spliced RNA without the gamma-globin intron by targeting the intron-containing RNA via microRNA 30 (miR30)-based short hairpin RNAs. Using two optimized fluorescent reporter vectors and flow cytometry, we determined that multiply HIV-1-infected cells were generated at a higher frequency in coculture than in cell-free infection; furthermore, this increase was dependent upon viruses bearing HIV-1 Env. Compared with previously described vectors, these improved vectors can quantify the infection in lymphocytes and in primary cells with a higher sensitivity and allow the detection and quantitation of multiply infected cells, providing better tools to study retroviral cell-mediated infection. IMPORTANCE The human-pathogenic retroviruses HTLV-1 and HIV-1 can be transmitted more efficiently in vivo via direct contact of infected cells with healthy target cells than through cell-free virion-mediated infection. Despite its importance, cell-to-cell transmission has been difficult to quantify because the previously infected cells and the newly infected cells are mixed together in the same culture. In the current study, we generated vectors that are significantly improved over the previously described replication-dependent vectors. As a result, these improved vectors can efficiently detect and quantify cell-to-cell transmission or new infection events in cells in mixed culture. These luciferase- or fluorescence protein-based reporter vectors can be used to quantify and study HIV-1 or HTLV-1 cell-mediated infection in a simple one-step transfection/infection assay.
Glycyrrhizin and isoliquiritigenin suppress the LPS sensor Toll‐like receptor 4/MD‐2 complex signaling in a different manner

Recent evidences suggest that the extracts of plant products are able to modulate innate immune responses. A saponin GL and a chalcone ILG are representative components of Glycyrrhiza uralensis, which attenuate inflammatory responses mediated by TLRs. Here, we show that GL and ILG suppress different steps of the LPS sensor TLR4/MD‐2 complex signaling at the receptor level. Extract of G. uralensis suppressed IL‐6 and TNF‐α production induced by lipid A moiety of LPS in RAW264.7 cells. Among various G. uralensis‐related components of saponins and flavanones/chalcones, GL and ILG could suppress IL‐6 production induced by lipid A in dose‐dependent manners in RAW264.7 cells. Furthermore, elevation of plasma TNF‐α in LPS‐injected mice was attenuated by passive administration of GL or ILG. GL and ILG inhibited lipid A‐induced NF‐κB activation in Ba/F3 cells expressing TLR4/MD‐2 and CD14 and BMMs. These components also inhibited activation of MAPKs, including JNK, p38, and ERK in BMMs. In addition, GL and ILG inhibited NF‐κB activation and IL‐6 production induced by paclitaxel, a nonbacterial TLR4 ligand. Interestingly, GL attenuated the formation of the LPS‐TLR4/MD‐2 complexes, resulting in inhibition of homodimerization of TLR4. Although ILG did not affect LPS binding to TLR4/MD‐2, it could inhibit LPS‐induced TLR4 homodimerization. These results imply that GL and ILG modulate the TLR4/MD‐2 complex at the receptor level, leading to suppress LPS‐induced activation of signaling cascades and cytokine production, but their effects are exerted at different steps of TLR4/MD‐2 signaling.
Differentiation of Separated Mouse Mammary Luminal Epithelial and Myoepithelial Cells Cultured on EHS Matrix Analyzed by Indirect Immunofluorescence of Cytoskeletal Antigens

We have previously demonstrated that purified virgin mouse mammary luminal epithelial and myoepithelial cells promiscuously express cell type-specific cytokeratins when they are cloned in vitro. Changes in cytokeratin expression may be indicators of the loss or change of the differentiated identity of a cell. To investigate the factors that may be responsible for the maintenance of differentiated cellular identity, specifically cell-cell and cell-matrix interactions, we cloned flow-sorted mouse mammary epithelial cells on the extracellular matrix (ECM) derived from the Engelbreth-Holm-Swarm murine sarcoma (EHS matrix). Changes in cell differentiation on EHS, compared with culture on glass, were analyzed by comparing patterns of cytokeratin expression. The results indicate that ECM is responsible for maintenance of the differentiated identity of basal/myoepithelial cells and prevents the inappropriate expression of luminal antigens seen on glass or plastic. Luminal cell identity in the form of retention of luminal markers and absence of basal/myoepithelial antigens, on the contrary, appears to depend on homotypic cell-cell contacts and interactions. The results also show that luminal cells (or a subpopulation of them) can generate a cell layer that expresses only basal cytokeratin markers (and no luminal cytokeratin markers) and may form a pluripotent compartment.
Molecular Dynamics Simulation of GM1 in Phospholipid Bilayer

Abstract We report molecular dynamics simulation of fully hydrated lipid bilayer of dimyristoyl phosphatidyl choline (DMPC) at room temperature with ganglioside GM1 attached to it in the upper layer under periodic boundary conditions. The simulation results indicate that the presence of a single GM1 molecule has local effects on the bilayer. Three sugar residues (GalNAc-Gal-Glc) of the pentasaccharide head group of GM1 remain on the lipid surface where as the NeuNAc residue extends out in the aqueous layer. The radial distribution functions suggest ordering of water molecules near the glycerol and carboxyl group of the sialic acid in the upper layer. One of the ceramide chains of GM1, the sphingosine chain, folds up and is stacked under the sugar residues lying on the surface. The other ceramide chain is inserted into the lipid bilayer. The arrangement of the polar head group as well as the acyl chains of the lipids which are immediate neighbours of the GM1 are modified compared to the non-neighbour ones and others at the lower layer. The time average conformation of GM1-pentasaccharide is stabilized by a number of inter residue hydrogen bonds that were observed experimentally. The trajectory average conformation of GM1-pentasaccharide was docked on to the cholera toxin molecule and the minimized complex reveals alternative binding modes between the toxin and the GM1-pentasaccharide moiety. The results of these simulation studies might help to understand the structure and nature of the effects of GM1 on the membrane at atomic resolution.