Miracle-dz/UniFAR-FTUs huggingface dataset:
A Basic Cluster in the N Terminus of Yellow Fever Virus NS2A Contributes to Infectious Particle Production

ABSTRACT The flavivirus NS2A protein is involved in the assembly of infectious particles. To further understand its role in this process, a charged-to-alanine scanning analysis was performed on NS2A encoded by an infectious cDNA clone of yellow fever virus (YFV). Fifteen mutants containing single, double, or triple charged-to-alanine changes were tested. Five of them did not produce infectious particles, whereas efficient RNA replication was detectable for two of the five NS2A mutants (R22A-K23A-R24A and R99A-E100A-R101A mutants). Prolonged cultivation of transfected cells resulted in the recovery of pseudorevertants. Besides suppressor mutants in NS2A, a compensating second-site mutation in NS3 (D343G) arose for the NS2A R22A-K23A-R24A mutant. We found this NS3 mutation previously to be suppressive for the NS2Aα cleavage site Q189S mutant, also deficient in virion assembly. In this study, the subsequently suggested interaction between NS2A and NS3 was proven by coimmunoprecipitation analyses. Using selectively permeabilized cells, we could demonstrate that the regions encompassing R22A-K23A-R24A and Q189S in NS2A are localized to the cytoplasm, where NS3 is also known to reside. However, the defect in particle production observed for the NS2A R22A-K23A-R24A and Q189S mutants was not due to a defect in physical interaction between NS2A and NS3, as the NS2A mutations did not interrupt NS3 interaction. In fact, a region just upstream of R22-K23-R24 was mapped to be critical for NS2A-NS3 interaction. Taken together, these data support a complex interplay between YFV NS2A and NS3 in virion assembly and identify a basic cluster in the NS2A N terminus to be critical in this process. IMPORTANCE Despite an available vaccine, yellow fever remains endemic in tropical areas of South America and Africa. To control the disease, antiviral drugs are required, and an understanding of the determinants of virion assembly is central to their development. In this study, we identified a basic cluster of amino acids in the N terminus of YFV NS2A which inhibited virion assembly upon mutation. The defect was rescued by a spontaneously occurring mutation in NS3. Our study proves an interaction between NS2A and NS3, which, remarkably, was maintained for the NS2A mutant in the presence and absence of the NS3 mutation. This suggests a role for other viral and/or cellular proteins in virion assembly. Residues important for YFV virion production reported here only partially coincided with those reported for other flaviviruses, suggesting that the determinants for particle production are virus specific. Reconstruction of a YFV encoding tagged NS2A paves the way to identify further NS2A interaction partners.
Degradation of fibronectin and vitronectin in chronic wound fluid: analysis by cell blotting, immunoblotting, and cell adhesion assays.

We used a combination of cell blotting, immunoblotting, and cell adhesion assays to analyze fibronectin and vitronectin in wound fluid from acute and chronic wounds. Acute wound fluid (e.g., suction blister fluid, mastectomy fluid) contained intact fibronectin and vitronectin as major cell adhesion proteins. In marked contrast, chronic wound fluid samples from three of 11 patients with venous stasis ulcers showed complete degradation of vitronectin and degradation of fibronectin into small molecular mass polypeptides less than 125 kDa. Three of these polypeptides--54, 93, and 125 kDa--were biologically active in promoting cell attachment and were recognized by monoclonal antibodies that bind fibronectin near the arg-gly-asp (RGD) domain. In wound fluid samples from the other eight of 11 patients, only slight degradation of vitronectin and fibronectin occurred, which resulted in a mixture of mostly intact molecules along with large fragments. Intact fibronectin in chronic wound fluid samples contained the ED-A domain, which showed that fibronectin synthesis occurred locally in the wound bed. Wound fluid containing extensively degraded vitronectin and fibronectin reversibly inhibited cell adhesion, and excess fetal bovine serum, but not purified fibronectin, neutralized the inhibitory effect. We suggest that protease activity in some chronic wounds may cause degradation of adhesion proteins and prevent cell adhesion necessary for normal wound closure.
Comparative Proteomic Analysis of the Haemophilus ducreyi Porin-Deficient Mutant 35000HP::P2AB

ABSTRACT Haemophilus ducreyi is an obligate human pathogen and the causative agent of the sexually transmitted, genital ulcerative disease chancroid. The genome of strain 35000HP contains two known porin proteins, OmpP2A and OmpP2B. Loss of OmpP2A and OmpP2B expression in the mutant 35000HP::P2AB resulted in no obvious growth defect or phenotype. Comparison of outer membrane profiles indicated increased expression of the 58.5-kDa chaperone, GroEL, in the porin-deficient mutant. A proteomics-based comparison resulted in the identification of 231 proteins present in membrane-associated protein samples, of which a subset of 56 proteins was differentially expressed at a level of 1.5-fold or greater in the porin-deficient strain 35000HP::P2AB relative to that in 35000HP. Twenty of the differentially expressed proteins were selected for real-time PCR, resulting in the validation of 90% of the selected subgroup. Proteins identified in these studies suggested a decreased membrane stability phenotype, which was verified by disk diffusion assay. Loss of OmpP2A and OmpP2B resulted in global protein expression changes which appear to compensate for the absence of porin expression in 35000HP::P2AB.
Synergistic anticancer effects of ganciclovir/thymidine kinase and 5-fluorocytosine/cytosine deaminase gene therapies.

BACKGROUND A bacterial enzyme, Escherichia coli cytosine deaminase, which converts the prodrug 5-fluorocytosine into the toxic drug 5-fluorouracil, and a viral enzyme, herpes simplex virus thymidine kinase, which converts ganciclovir from an inactive prodrug to a cytotoxic agent by phosphorylation, are being actively investigated for use in gene therapy for cancer. The purpose of this study was to determine whether combining these prodrug-activating gene therapies might result in enhanced anticancer effects. METHODS Rat 9L gliosarcoma cells were transfected with plasmids containing the E. coli cytosine deaminase gene (9L/CD cells), with plasmids containing the herpes simplex virus thymidine kinase gene (9L/TK cells), or with both expression plasmids (9L/CD-TK cells). The drug sensitivities of the cell lines were evaluated; in addition, the sensitivities of 9L and 9L/CD-TK cells mixed in varied proportions were measured. The effects of prodrug treatment on 9L/CD-TK tumor growth (i.e., size and volume) in nude mice were monitored. The isobologram method of Loewe and the multiple drug-effect analysis method of Chou-Talalay were used to measure the interaction between the two prodrug-activating gene therapies. To elucidate the mechanism of interaction, the phosphorylation of ganciclovir in 9L/CD-TK cells after varying prodrug treatments was studied. RESULTS AND CONCLUSIONS The presence of transfected cytosine deaminase and thymidine kinase genes in 9L gliosarcoma cells reduced cell survival, both in vitro and in vivo, following treatment with the relevant prodrugs; the effects of the two components appeared to be synergistic and related mechanistically to the enhancement of ganciclovir phosphorylation by thymidine kinase following 5-fluorouracil treatment.
Reduction of Diagnostic Window by New Fourth-Generation Human Immunodeficiency Virus Screening Assays

ABSTRACT In order to reduce the diagnostic window between the time of human immunodeficiency virus (HIV) infection and laboratory diagnosis, new screening enzyme-linked immunosorbent assays (ELISAs) which permit the simultaneous detection of HIV antigen and antibody have been developed. Two fourth-generation assays, HIV DUO (Biomérieux) and HIV Combi (Boehringer Mannheim), for the combined detection of HIV antigen and antibody, were compared with a third-generation assay (HIV-1/HIV-2 3rd Generation Plus enzyme immunoassay [EIA]; Abbott) and a p24 antigen test (HIV-1 Ag monoclonal; Abbott). A total of 17 seroconversion panels, 15 cell culture supernatants infected with different HIV type 1 (HIV-1) subtypes, and 255 potentially cross-reactive serum samples were tested. Ten seroconversions were detected an average of 8.1 days earlier with HIV DUO and 7.5 days earlier with HIV Combi than with the third-generation ELISA. Overall, in the 17 seroconversion panels tested, HIV DUO detected HIV-1 infection an average of 4.8 days and HIV Combi detected infection an average of 4.4 days earlier than HIV-1/HIV-2 3rd Generation Plus EIA. HIV antigen was detected with HIV DUO and HIV Combi in all of the 15 cell culture supernatants infected with different HIV-1 subtypes, including subtype O. With fourth-generation assays, considerably fewer false-positive results (n = 4 to 6) were obtained, in comparison with the third-generation EIA (n = 18). Fourth-generation assays permit an earlier diagnosis of HIV infection than third-generation antibody screening assays through the detection of p24 antigen, which may be present in serum samples from individuals with recent HIV infection prior to seroconversion.
REINNERVATION OF TRANSPLANTED PANCREATIC ISLETS: A COMPARISON AMONG ISLETS IMPLANTED INTO THE KIDNEY, SPLEEN, AND LIVER

Islets transplanted beneath the kidney capsule become reinnervated during the first 3–4 months after implantation by both afferent and efferent nerve fibers. To evaluate the importance of the implantation organ for this process, the present study compared both the degree and the types of nerve fibers reinnervating islets transplanted into the liver, kidney, and spleen. For this purpose, 150 syngeneic islets were grafted under the kidney capsule of C57BL/6 mice. In addition, the same animals were injected with 150 islets into the spleen or liver. All animals were killed 14 weeks after transplantation, after which the graft-bearing organs were processed for indirect immunofluorescence for neuropeptides and tyrosine hydroxylase (TH), and with acetyl cholinesterase (AchE) staining to visualize nerve fibers. Both afferent (containing substance P and/or calcitonin gene-related peptide) and parasympathetic (containing vasoactive intestinal peptide or AchE) nerve fibers were absent from islets implanted into the spleen; an occasional CGRP fiber was seen in islets implanted into the liver; and all these fibers were regularly seen in islets implanted beneath the renal capsule. The islets implanted into the liver or spleen contained a dense network of sympathetic (containing neuropeptide Y and TH) nerve fibers that was often more dense than in the islet grafts under the kidney capsule. One-fifth of islets implanted into the liver were, however, completely devoid of demonstrable nerve fibers. In conclusion, there are marked differences with regard to the pattern of reinnervation of islets transplanted to different implantation sites.
Red cell vacuoles: Their size and distribution under normal conditions and after splenectomy

The frequency of occurrence of vacuoles in red blood cells was studied by transmission electron microscopy. Small vacuoles were found in about 13% of the cell sections, and they had a mean diameter of 130 ± 72 nm (mean ± SD). It can be estimated that there were about 20 small vacuoles per erythrocyte. The frequency of vacuoles was similar in density‐separated cell fractions. In splenectomized patients, the small vacuoles were 4 times more frequent; there was again no difference in vacuole density between top and bottom fractions of density‐separated red blood cells. The bottom fraction of red blood cells from splenectomized patients, however, had a high incidence of large vacuoles (> 300 nm in diameter) and clustering of small vacuoles. These large vacuoles were probably the result of aggregation and fusion of small vacuoles, and their size allowed detection by light microscopy. Hence, the well‐known “pocked” or “pitted” red blood cells of splenectomized individuals were more frequent in the bottom fraction. We conclude that small vacuoles occur normally in erythrocytes, that they tend to cluster and fuse during cell aging, and that the spleen is capable of removing these structures when they reach a certain size.
Axial transmission techniques for bone assessment: an in vitro comparative study

The objective of this paper was to compare three different approaches using ultrasonic axial transmission in vitro. Forty-one fresh human radii with soft tissue removed were investigated. Measurements of ultrasound velocity of the fastest signal contribution have been performed with a commercial device (Sunlight Omnisense/spl trade/, 1.25 MHz), a prototype based on 1 MHz bi-directional axial transmission, and a prototype low frequency (200 kHz) device. In addition, the low frequency probe allowed the velocity measurement of a slower wave guided by the bone. Site-matched pQCT measurements provided bone properties, such as cortical thickness, cross-sectional area, and bone mineral density (BMD). All measurements were taken at the same location which corresponds to the clinical site of measurement. Our results suggest that different axial transmission approaches may reflect different bone properties. Therefore, a multi-frequency technique might be valuable to separate respective contributions of different bone properties (e. g., cortical thickness and BMD) with measured values of velocity.
Molecular Survey of β-Lactamases Conferring Resistance to Newer β-Lactams in Enterobacteriaceae Isolates from Polish Hospitals

ABSTRACT The first national survey of resistance to newer β-lactams in nosocomial populations of Enterobacteriaceae in Poland was performed. The study covered all nonrepetitive enterobacterial isolates cultured from specimens from inpatients in 13 regional secondary-care hospitals from November 2003 to January 2004. Among 2,388 isolates, the predominant species was Escherichia coli (59.6%), followed by Proteus mirabilis (14.5%) and Klebsiella spp. (8.5%). The frequency of extended-spectrum β-lactamases (ESBLs) was very high, with ESBLs present in 11.1% of all isolates and 40.4% of Klebsiella pneumoniae isolates, the latter value greatly exceeding that for E. coli (2.5%). The contribution of outbreak isolates was significant, resulting, for example, in a particularly high rate of ESBL producers among Serratia marcescens isolates (70.8%). The pool of ESBL types was overwhelmingly dominated (81.7%) by CTX-M-like β-lactamases CTX-M-3 (80.6%) and CTX-M-15, with SHV types (17.5%; SHV-2, SHV-5, and SHV-12) and sporadic TEM-like enzymes (0.7%; TEM-19 and TEM-48) being the next most frequent. Acquired AmpC-type cephalosporinases were observed exclusively in P. mirabilis, in 20.5% of the isolates of this species (compared with the frequency of ESBL producers of 11.5% of P. mirabilis isolates). All these cephalosporinases (CMY-12, CMY-15, and a novel variant, CMY-38) originated from Citrobacter freundii. Four isolates of E. coli (two isolates), K. pneumoniae (one isolate), and P. mirabilis (one isolate) produced class A inhibitor-resistant β-lactamases (TEM-30, TEM-32, TEM-37, and SHV-49), being the first of such producers identified in Poland. The survey documented both specific and more global characteristics of the epidemiology of the β-lactamase-mediated resistance in enterobacteria from Polish hospitals and demonstrated that the ESBL frequency has reached an alarming level.
The Production Life Cycle

Tasks in production have different dynamics in comparison with individual products. We build a general equilibrium model where three forces — offshoring, learning, and wage — interact to determine how production fragmentation evolves over time. We show that the production process follows a life cycle: its fragmentation emerges, deepens, and converges. Our model is stylized and versatile, which can be used to tractably characterize the loci of global output, productivity, welfare, and value added distribution as production fragmentation progresses. This article is protected by copyright. All rights reserved
Governmental Rationales for Installing Electronic Information Systems: A Quest for Responsive Social Work

Various authors have argued that electronic information systems (EIS) do not necessarily improve the responsiveness of social work. One of the main reasons given for these failings is the flawed implementation of EIS by professionals. However, we argue that the ongoing debate fails to be explicit about governmental rationales for installing EIS. This article presents the findings of a content analysis that aimed to uncover the governmental rationales for installing EIS in the Flemish child and welfare protection (CWP) system. Our analysis revealed three clusters of rationales. The first cluster supports the use of EIS as an instrument to better match supply and demand. The second cluster shows that the gathered data also serve as an instrument for accountability. A third cluster focuses on the aim of creating more uniformity. Based on our analysis, we argue that it is not flawed implementation that impedes the development of responsive social work; rather, the governmental rationales themselves are flawed.
The role of zinc supplementation in the inhibition of tissue damage caused by exposure to electromagnetic field in rat lung and liver tissues.

BACKGROUND The objective of the present study was to examine the effects of zinc supplementation on the oxidant damage in lung and liver tissues in rats exposed to a 50-Hz frequency magnetic field for 5 minutes every other day over a period of 6 months. METHODS The study included 24 adult male Sprague-Dawley rats, which were divided into the three groups in equal numbers: Group 1, the control group (G1); Group 2, the group exposed to an electromagnetic field (G2); and Group 3, the group, which was exposed to an EMF and supplemented with zinc (G3). At the end of the 6-month procedures, the animals were decapitated to collect lung and liver tissue samples, in which MDA was analyzed using the "TBARS method (nmol/g/protein)", GSH by the "biuret method (mg/g/protein)" and zinc levels by atomic emission (µg/dl). RESULTS MDA levels in lung and liver tissues in G2 were higher than those in G1 and G3, and the levels in G3 were higher than those in G1 (p<0.01). As for GSH levels, G3 had GSH levels higher than G1 and G2, and G2 had GSH levels higher than G1 (p<0.01). Zinc values in the liver and lung tissues were the highest in the Group 3 and the lowest in the Group 2 (p<0.01). CONCLUSION Results of the study indicated that exposure to an electromagnetic field caused cellular damage in lung and liver tissues and zinc supplementation inhibited the inflicted cellular damage. Another important result of this study that needs emphasis was that exposure to an electromagnetic field led to a significant decrease in zinc levels in lung and liver tissues (Tab. 3, Ref. 23).
Reducing perinatal complications and preterm delivery for patients undergoing in utero closure of fetal myelomeningocele: further modifications to the multidisciplinary surgical technique.

UNLABELLED OBJECT. : As more pediatric neurosurgeons become involved with fetal myelomeningocele closure efforts, examining refined techniques in the overall surgical approach that could maximize beneficial outcomes becomes critical. The authors compared outcomes for patients who had undergone a modified technique with those for patients who had undergone fetal repair as part of the earlier Management of Myelomeningocele Study (MOMS). METHODS Demographic and outcomes data were collected for a series of 43 delivered patients who had undergone in utero myelomeningocele closure at the Fetal Center at Vanderbilt from March 2011 through January 2013 (the study cohort) and were compared with data for 78 patients who had undergone fetal repair as part of MOMS (the MOMS cohort). For the study cohort, no uterine trocar was used, and uterine entry, manipulation, and closure were modified to minimize separation of the amniotic membrane. Weekly ultrasound reports were obtained from primary maternal-fetal medicine providers and reviewed. A test for normality revealed that distribution for the study cohort was normal; therefore, parametric statistics were used for comparisons. RESULTS The incidence of premature rupture of membranes (22% vs 46%, p = 0.011) and chorioamnion separation (0% vs 26%, p < 0.001) were lower for the study cohort than for the MOMS cohort. Incidence of oligohydramnios did not differ between the cohorts. The mean (± SD) gestational age of 34.4 (± 6.6) weeks for the study cohort was similar to that for the MOMS cohort (34.1 ± 3.1 weeks). However, the proportion of infants born at term (37 weeks or greater) was significantly higher for the study cohort (16 of 41; 39%) than for the MOMS cohort (16 of 78; 21%) (p = 0.030). Compared with 10 (13%) of 78 patients in the MOMS cohort, only 2 (4%) of 41 infants in the study cohort were delivered earlier than 30 weeks of gestation (p = 0.084, approaching significance). For the study cohort, 2 fetal deaths were attributed to the intervention, and both were believed to be associated with placental disruption; one of these mothers had previously unidentified thrombophilia. Mortality rates did not statistically differ between the cohorts. CONCLUSIONS These early results suggest that careful attention to uterine entry, manipulation, and closure by the surgical team can result in a decreased rate of premature rupture of membranes and chorioamnion separation and can reduce early preterm delivery. Although these results are promising, their confirmation will require further study of a larger series of patients.
Impact of premature activation of the right ventricle with programmed stimulation in Brugada syndrome

In Brugada syndrome (BrS), it has been reported that delayed activation in the RV is related to the development of type‐1 ECG, which is more critical than type‐2. On the other hand, the coexistence of complete right bundle‐branch block (CRBBB), which also causes delayed activation in the RV, sometimes makes typical BrS ECG misleading. We hypothesized that premature stimulation of the RV can unmask the influence of delayed activation in the RV and convert the morphology of ECG in BrS patients.