The activation of Toll-like receptors (TLRs) by lipopolysaccharide or other ligands evokes a proinflammatory immune response,
which is not only capable of clearing invading pathogens but can also ...inflict damage to host tissues. It is therefore important
to prevent an overshoot of the TLR-induced response where necessary, and here we show that extracellular ATP is capable of
doing this in human monocytes. Using reverse transcription-PCR, we showed that monocytes express P2Y 1 , P2Y 2 , P2Y 4 , P2Y 11 , and P2Y 13 receptors, as well as several P2X receptors. To elucidate the function of these receptors, we first studied Ca 2+ signaling in single cells. ATP or UTP induced a biphasic increase in cytosolic Ca 2+ , which corresponded to internal Ca 2+ release followed by activation of store-operated Ca 2+ entry. The evoked Ca 2+ signals stimulated Ca 2+ -activated K + channels, producing transient membrane hyperpolarization. In addition, ATP promoted cytoskeleton reorganization and cell
migration; however, unlike chemoattractants, the migration was non-directional and further analysis showed that ATP did not
activate Akt, essential for sensing gradients. When TLR2, TLR4, or TLR2/6 were stimulated with their respective ligands, ATPγS
profoundly inhibited secretion of proinflammatory cytokines (tumor necrosis factor-α and monocyte chemoattractant protein-1)
but increased the production of interleukin-10, an anti-inflammatory cytokine. In radioimmune assays, we found that ATP (or
ATPγS) strongly increased cAMP levels, and, moreover, the TLR-response was inhibited by forskolin, whereas UTP neither increased
cAMP nor inhibited the TLR-response. Thus, our data suggest that ATP promotes non-directional migration and, importantly,
acts as a âhost tissue damageâ signal via the G s protein-coupled P2Y 11 receptor and increased cAMP to negatively regulate TLR signaling.
Superantigens trigger polyclonal activation of T lymphocytes with cytokine release that eventually may lead to lethal cytokine syndrome (toxic shock). In contrast, bacterial components that are ...recognized by Toll-like receptors (e.g. LPS or CpG DNA) primarily target macrophages and dendritic cells. We have analyzed whether superantigens and TLR ligands interact with each other. We found that superantigens synergize with LPS in an IFN- gamma -dependent pathway. More important, we found compelling evidence that superantigens prime the innate immune cell system to a subsequent challenge with endotoxin. This sensitization was critically dependent on T-cell derived IFN- gamma . When we analyzed the underlying molecular mechanisms, we additionally found that TLR stimulation enhanced IFN- gamma -mediated cellular responses. Moreover, TLR ligands induced proteins of the SOCS family thus shutting off IFN- gamma -mediated cellular activation. Since IFN- gamma is synthesized by T cells after superantigen triggering, these results show that superantigen and TLR pathways are interconnected and regulate each other. They further show that the outcome of this interaction may include activation as well as down-regulation of the respective response pattern.
Dependent on the excretion pattern, wastewater monitoring of viruses can be a valuable approach to characterizing their circulation in the human population. Using polyethylene glycol precipitation ...and reverse transcription-quantitative PCR, the occurrence of RNA of SARS-CoV-2 and influenza viruses A/B in the raw wastewater of two treatment plants in Germany between January and May 2022 was investigated. Due to the relatively high incidence in both exposal areas (plant 1 and plant 2), SARS-CoV-2-specific RNA was determined in all 273 composite samples analyzed (concentration of E gene: 1.3 × 10⁴ to 3.2 × 10⁶ gc/L). Despite a nation-wide low number of confirmed infections, influenza virus A was demonstrated in 5.2% (concentration: 9.8 × 10² to 8.4 × 10⁴ gc/L; plant 1) and in 41.6% (3.6 × 10³ to 3.0 × 10⁵ gc/L; plant 2) of samples. Influenza virus B was detected in 36.0% (7.2 × 10² to 8.5 × 10⁶ gc/L; plant 1) and 57.7% (9.6 × 10³ to 2.1 × 10⁷ gc/L; plant 2) of wastewater samples. The results of the study demonstrate the frequent detection of two primary respiratory viruses in wastewater and offer the possibility to track the epidemiology of influenza by wastewater-based monitoring.
Filtration through membranes with nanopores is typically associated with high transmembrane pressures and high energy consumption. This problem can be addressed by reducing the respective membrane ...thickness. Here, a simple procedure is described to prepare ultrathin membranes based on protein nanopores, which exhibit excellent water permeance, two orders of magnitude superior to comparable, industrially applied membranes. Furthermore, incorporation of either closed or open protein nanopores allows tailoring the membrane's ion permeability. To form such membranes, the transmembrane protein ferric hydroxamate uptake protein component A (FhuA) or its open‐pore variant are assembled at the air–water interface of a Langmuir trough, compressed to a dense film, crosslinked by glutaraldehyde, and transferred to various support materials. This approach allows to prepare monolayer or multilayer membranes with a very high density of protein nanopores. Freestanding membranes covering holes up to 5 μm in diameter are visualized by atomic force microscopy (AFM), helium ion microscopy, and transmission electron microscopy. AFM PeakForce quantitative nanomechanical property mapping (PeakForce QNM) demonstrates remarkable mechanical stability and elastic properties of freestanding monolayer membranes with a thickness of only 5 nm. The new protein membrane can pave the way to energy‐efficient nanofiltration.
Introducing a facile method, protein nanopores are used to prepare an ultrathin yet stable nanofiltration membrane. Advanced microscopic techniques enable visualization of molecular monolayer membranes covering microporous supports. Membranes made from open protein pores show higher ion permeability over those made from almost closed pores. Furthermore, such protein membranes have outstanding water permeance higher than conventional nanofiltration membranes.
We present an in-depth investigation of the aging effects in silicone molds for vacuum casting processes. Their lifetime is limited to a few production cycles due to contamination with the ...diisocyanate component of polyurethane casting materials. Using thermogravimetric analysis measurements, Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, and helium-ion-microscopy the chemical and physical mechanisms of the aging process have been identified. It has been shown that a diffusion process of diisocyanate into the cavity surface leads to the formation of interpenetrating polymer networks of polyurea derivatives in silicone rubber. This has been proven by extracting and analyzing polyurea of low molecular weights from the silicone.
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