1,195 research outputs found
Detection and Characterisation of Lactobacillus spp. in the Bovine Uterus and Their Influence on Bovine Endometrial Epithelial Cells In Vitro
Bacterial infections and inflammation of the uterus are common in dairy cattle
after parturition. In particular, pathogenic bacteria that cause endometritis
have been the focus of research in cattle reproduction in the last ten years.
The aim of the present study was to identify commensal lactobacilli in the
bovine uterus and to examine their influence on the synthesis of pro-
inflammatory factors in bovine endometrial epithelial cells in vitro.
Lactobacillus species were isolated from healthy bovine uteri and further
characterised. Bovine endometrial epithelial cells in the second passage (n =
5 animals) were co-cultured with the autochthonous isolates L. buchneri, L.
ruminis and L. amylovorus as well as with a commercially available L.
vaginalis in different multiplicities of infection (MOI = 1, 5 and 10,
respectively). Endometrial epithelial cells cultured without bacteria served
as controls. At distinct points in time (2, 4 and 6 h) total RNA was extracted
from co-cultured epithelial cells and subjected to reverse transcription
quantitative PCR of pro-inflammatory factors. Furthermore, the release of such
factors by co-cultured epithelial cells was measured by ELISA or EIA after 24
and 48 h. L. ruminis and L. amylovorus induced increased interleukin (IL)
IL1A, IL6, IL8 and prostaglandin-endoperoxide synthase 2 mRNA levels and the
release of IL8 and prostaglandin F2α in endometrial epithelial cells compared
with control cells. In contrast, L. buchneri did not significantly influence
the expression and release of these factors. Toll-like receptors 2 and 6
transcripts were found unchanged in co-cultured and untreated epithelial cells
in vitro. However, endometrial epithelial cells of each animal showed
individual differences in the response to bacterial load. These results
suggest that Lactobacillus species are present in the bovine uterus, revealing
immunomodulatory properties
Fusoselect: cell-cell fusion activity engineered by directed evolution of a retroviral glycoprotein
Membrane fusion plays a key role in many biological processes including vesicle trafficking, synaptic transmission, fertilization or cell entry of enveloped viruses. As a common feature the fusion process is mediated by distinct membrane proteins. We describe here ‘Fusoselect', a universal procedure allowing the identification and engineering of molecular determinants for cell-cell fusion-activity by directed evolution. The system couples cell-cell fusion with the release of retroviral particles, but can principally be applied to membrane proteins of non-viral origin as well. As a model system, we chose a γ-retroviral envelope protein, which naturally becomes fusion-active through proteolytic processing by the viral protease. The selection process evolved variants that, in contrast to the parental protein, mediated cell-cell fusion in absence of the viral protease. Detailed analysis of the variants revealed molecular determinants for fusion competence in the cytoplasmic tail (CT) of retroviral Env proteins and demonstrated the power of Fusoselec
Fusoselect: cell–cell fusion activity engineered by directed evolution of a retroviral glycoprotein
Membrane fusion plays a key role in many biological processes including vesicle trafficking, synaptic transmission, fertilization or cell entry of enveloped viruses. As a common feature the fusion process is mediated by distinct membrane proteins. We describe here ‘Fusoselect’, a universal procedure allowing the identification and engineering of molecular determinants for cell–cell fusion-activity by directed evolution. The system couples cell–cell fusion with the release of retroviral particles, but can principally be applied to membrane proteins of non-viral origin as well. As a model system, we chose a γ-retroviral envelope protein, which naturally becomes fusion-active through proteolytic processing by the viral protease. The selection process evolved variants that, in contrast to the parental protein, mediated cell–cell fusion in absence of the viral protease. Detailed analysis of the variants revealed molecular determinants for fusion competence in the cytoplasmic tail (CT) of retroviral Env proteins and demonstrated the power of Fusoselect
Comprehensive characterization of molecular interactions based on nanomechanics
Molecular interaction is a key concept in our understanding of the biological mechanisms of life. Two physical properties change when one molecular partner binds to another. Firstly, the masses combine and secondly, the structure of at least one binding partner is altered, mechanically transducing the binding into subsequent biological reactions. Here we present a nanomechanical micro-array technique for bio-medical research, which not only monitors the binding of effector molecules to their target but also the subsequent effect on a biological system in vitro. This label-free and real-time method directly and simultaneously tracks mass and nanomechanical changes at the sensor interface using micro-cantilever technology. To prove the concept we measured lipid vesicle (approximately 748*10(6) Da) adsorption on the sensor interface followed by subsequent binding of the bee venom peptide melittin (2840 Da) to the vesicles. The results show the high dynamic range of the instrument and that measuring the mass and structural changes simultaneously allow a comprehensive discussion of molecular interactions
Selection of prebiotic oligonucleotides by cyclic phase separation
The emergence of functional oligonucleotides on early Earth required a
molecular selection mechanism to screen for specific sequences with prebiotic
functions. Cyclic processes such as daily temperature oscillations were
ubiquitous in this environment and could trigger oligonucleotide phase
separation. Here, we propose sequence selection based on phase separation
cycles realized through sedimentation in a system subjected to the feeding of
oligonucleotides. Using theory and experiments with DNA, we show
sequence-specific enrichment in the sedimented dense phase, in particular of
short 22-mer DNA sequences. The underlying mechanism selects for
complementarity, as it enriches sequences that tightly interact in the
condensed phase through base-pairing. Our mechanism also enables initially
weakly biased pools to enhance their sequence bias or to replace the most
abundant sequences as the cycles progress. Our findings provide an example of a
selection mechanism that may have eased screening for the first auto-catalytic
self-replicating oligonucleotides
A Robust and Rapid Method of Producing Soluble, Stable, and Functional G-Protein Coupled Receptors
Membrane proteins, particularly G-protein coupled receptors (GPCRs), are notoriously difficult to express. Using commercial E.coli cell-free systems with the detergent Brij-35, we could rapidly produce milligram quantities of 13 unique GPCRs. Immunoaffinity purification yielded receptors at >90% purity. Secondary structure analysis using circular dichroism indicated that the purified receptors were properly folded. Microscale thermophoresis, a novel label-free and surface-free detection technique that uses thermal gradients, showed that these receptors bound their ligands. The secondary structure and ligand-binding results from cell-free produced proteins were comparable to those expressed and purified from HEK293 cells. Our study demonstrates that cell-free protein production using commercially available kits and optimal detergents is a robust technology that can be used to produce sufficient GPCRs for biochemical, structural, and functional analyses. This robust and simple method may further stimulate others to study the structure and function of membrane proteins.United States. Defense Advanced Research Projects Agency (DARPA-HR0011-09-C-0012)Massachusetts Institute of Technology. Undergraduate Research Opportunities Progra
Long-term trends in hepatitis C prevalence, treatment uptake and liver-related events in the Swiss HIV Cohort Study
BACKGROUND AND AIMS: Treatment for chronic hepatitis C virus (HCV) infections changed dramatically in the last decade. We assessed changes in the prevalence of replicating HCV infection, treatment uptake and liver-related morbidity and mortality in persons with HIV (PWH) and hepatitis C in the Swiss HIV cohort study.
METHODS: We included all cohort participants between 2002 and 2021. We assessed yearly prevalence of replicating HCV infection, overall and liver-related mortality, as well as the yearly incidence of liver-related events in persons with at least one documented positive HCV-RNA.
RESULTS: Of 14 652 participants under follow-up, 2294 had at least one positive HCV-RNA measurement. Of those, 1316 (57%) ever received an HCV treatment. Treatment uptake increased from 8.1% in 2002 to a maximum of 32.6% in 2016. Overall, prevalence of replicating HCV infection declined from 16.5% in 2004 to 1.3% in 2021. HCV prevalence declined from 63.2% to 7.1% in persons who inject drugs, and from 4.1% to 0.6% in men who have sex with men. Among the 2294 persons with replicating HCV infection, overall mortality declined from a maximum of 3.3 per 100 patient-years (PY) to 1.1 per 100 PY, and incidence of liver-related events decreased from 1.4/100 PY to 0.2/100 PY.
CONCLUSIONS: The introduction of DAA therapy was associated with a more than 10-fold reduction in prevalence of replicating HCV infection in PWH, approaching the estimates in the general population. Overall mortality and liver-related events declined substantially in persons living with HIV and hepatitis C
Long-term trends in hepatitis C prevalence, treatment uptake and liver-related events in the Swiss HIV Cohort Study.
BACKGROUND AND AIMS
Treatment for chronic hepatitis C virus (HCV) infections changed dramatically in the last decade. We assessed changes in the prevalence of replicating HCV infection, treatment uptake and liver-related morbidity and mortality in persons with HIV (PWH) and hepatitis C in the Swiss HIV cohort study.
METHODS
We included all cohort participants between 2002 and 2021. We assessed yearly prevalence of replicating HCV infection, overall and liver-related mortality, as well as the yearly incidence of liver-related events in persons with at least one documented positive HCV-RNA.
RESULTS
Of 14 652 participants under follow-up, 2294 had at least one positive HCV-RNA measurement. Of those, 1316 (57%) ever received an HCV treatment. Treatment uptake increased from 8.1% in 2002 to a maximum of 32.6% in 2016. Overall, prevalence of replicating HCV infection declined from 16.5% in 2004 to 1.3% in 2021. HCV prevalence declined from 63.2% to 7.1% in persons who inject drugs, and from 4.1% to 0.6% in men who have sex with men. Among the 2294 persons with replicating HCV infection, overall mortality declined from a maximum of 3.3 per 100 patient-years (PY) to 1.1 per 100 PY, and incidence of liver-related events decreased from 1.4/100 PY to 0.2/100 PY.
CONCLUSIONS
The introduction of DAA therapy was associated with a more than 10-fold reduction in prevalence of replicating HCV infection in PWH, approaching the estimates in the general population. Overall mortality and liver-related events declined substantially in persons living with HIV and hepatitis C
ЧЕРНІГІВСЬКЕ ВИДАННЯ НОВОГО ЗАВІТУ 1717 РОКУ З ПРИСВЯТОЮ ГЕТЬМАНУ ІВАНУ СКОРОПАДСЬКОМУ
В умовах втрати більшості українських стародруків XVII – початку XVIII ст., неабияке значення для дослідження мають поодинокі знахідки, котрі іноді трапляються серед приватних зібрань або державних сховищ. Збереження і введення до наукового обігу подібних раритетів входить до важливих напрямків досліджень української історії козацької доби
Gauge Fluxes in F-theory and Type IIB Orientifolds
We provide a detailed correspondence between G_4 gauge fluxes in F-theory
compactifications with SU(n) and SU(n)x(1) gauge symmetry and their Type IIB
orientifold limit. Based on the resolution of the relevant F-theory Tate models
we classify the factorisable G_4-fluxes and match them with the set of
universal D5-tadpole free U(1)-fluxes in Type IIB. Where available, the global
version of the universal spectral cover flux corresponds to Type IIB gauge flux
associated with a massive diagonal U(1). In U(1)-restricted Tate models extra
massless abelian fluxes exist which are associated with specific linear
combinations of Type IIB fluxes. Key to a quantitative match between F-theory
and Type IIB is a proper treatment of the conifold singularity encountered in
the Sen limit of generic F-theory models. We also shed further light on the
brane recombination process relating generic and U(1)-restricted Tate models.Comment: 53 pages, 3 figures; v2: Refs added; v3: minor corrections to match
version published in JHE
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