RESUMEN
Synapsed cells can communicate using exocytosed nucleotides like adenosine triphosphate (ATP). Ectonucleotidases localized to synaptic junctions degrade nucleotides into metabolites like adenosine monophosphate (AMP) or adenosine. Oftentimes nucleotide degradation occurs in a sequential manner, of which ATP degradation by CD39 and CD73 is a representative example. Here, CD39 first converts ATP and adenosine diphosphate (ADP) into AMP, after which AMP is dephosphorylated into adenosine by CD73. Hence, the concerted activity of CD39 and CD73 can help shape cellular responses to extracellular ATP. In a previous study, we demonstrated that coupled CD39 and CD73 activity within synapse-like junctions is strongly controlled by the enzymes' co-localization, their surface charge densities, and the electrostatic potential of the surrounding cell membranes. In this study, we demonstrate that crowders within synaptic junctions, which can include globular proteins like cytokines and membrane-bound proteins, impact coupled CD39 and CD73 ectonucleotidase activity and, in turn, the availability of intrasynapse ATP. Specifically, we developed a spatially explicit, reaction-diffusion model for the coupled conversion of ATP â AMP and AMP â adenosine in a model synaptic junction with crowders that is solved via the finite element method. Our modeling results suggest that the association rate for ATP to CD39 is strongly influenced by the density of intrasynaptic protein crowders, as increasing crowder density generally suppressed ATP association kinetics. Much of this suppression can be rationalized based on a loss of configurational entropy. The surface charges of crowders can further influence the association rate, with the surprising result that favorable crowder-nucleotide electrostatic interactions can yield CD39 association rates that are faster than crowder-free configurations. However, attractive crowder-nucleotide interactions decrease the rate and efficiency of adenosine production, which in turn increases the availability of ATP and AMP within the synapse relative to crowder-free configurations. These findings highlight how CD39 and CD73 ectonucleotidase activity, electrostatics, and crowding within synapses influence the availability of nucleotides for intercellular communication.
Asunto(s)
Adenosina , Apirasa , Adenosina/metabolismo , Adenosina Difosfato , Adenosina Monofosfato , Adenosina Trifosfato/metabolismo , Apirasa/metabolismo , Sinapsis/metabolismoRESUMEN
Nucleotides comprise small molecules that perform critical signaling roles in biological systems. Adenosine-based nucleotides, including adenosine tri-, di-, and mono-phosphate, are controlled through their rapid degradation by diphosphohydrolases and ecto-nucleotidases (NDAs). The interplay between nucleotide signaling and degradation is especially important in synapses formed between cells, which create signaling 'nanodomains'. Within these 'nanodomains', charged nucleotides interact with densely-packed membranes and biomolecules. While the contributions of electrostatic and steric interactions within such nanodomains are known to shape diffusion-limited reaction rates, less is understood about how these factors control the kinetics of nucleotidase activity. To quantify these factors, we utilized reaction-diffusion numerical simulations of 1) adenosine triphosphate (ATP) hydrolysis into adenosine monophosphate (AMP) and 2) AMP into adenosine (Ado) via two representative nucleotidases, CD39 and CD73. We evaluate these sequentially-coupled reactions in nanodomain geometries representative of extracellular synapses, within which we localize the nucleotidases. With this model, we find that 1) nucleotidase confinement reduces reaction rates relative to an open (bulk) system, 2) the rates of AMP and ADO formation are accelerated by restricting the diffusion of substrates away from the enzymes, and 3) nucleotidase co-localization and the presence of complementary (positive) charges to ATP enhance reaction rates, though the impact of these contributions on nucleotide pools depends on the degree to which the membrane competes for substrates. As a result, these contributions integratively control the relative concentrations and distributions of ATP and its metabolites within the junctional space. Altogether, our studies suggest that CD39 and CD73 nucleotidase activity within junctional spaces can exploit their confinement and favorable electrostatic interactions to finely control nucleotide signaling.
Asunto(s)
Adenosina Trifosfatasas/metabolismo , Adenosina/metabolismo , Nucleótidos/metabolismo , Cinética , Transducción de Señal , Propiedades de SuperficieRESUMEN
Sweet chestnut, an Asiatic tree introduced in many parts of Europe including the United Kingdom, is planted for nut production, timber, and amenity. Its major threat is the disease called blight, caused by the fungus Cryphonectria parasitica, which infects through wounds by airborne spores. Field trapping using sticky rods rotating traps was performed in an infected area in Devon (between May 2021 and April 2023). An improved dual hydrolysis Taqman probes real-time PCR was used. The number of spores was calculated by comparing the cycle threshold to the Ct of standards with known amounts of conidia or known target fragment copies cloned into a plasmid. Weekly spore counts were in the range of around 60 to approximately 8.5 × 103, with fluctuations of peaks (mainly in late summer-autumn 2021) and troughs. The effects of weather parameters were modelled, finding correlations between spore numbers and temperature, humidity, dewpoint, rainfall, wind speed, and wind duration. Additionally, an insect trapping was performed to confirm the presence/absence and quantity of C. parasitica conidia potentially phoretic on some insects by using the same molecular approach. None of the ten collected insect species harboured spores of this fungus.
RESUMEN
Silica-based materials including zeolites are commonly used for wide-ranging applications including separations and catalysis. Substrate transport rates in these materials can significantly influence the efficiency of such applications. Two factors that contribute to transport rates include (1) the porosity of the silicate matrix and (2) nonbonding interactions between the diffusing species and the silicate surface. These contributions generally emerge from disparate length scales, namely, "microscopic" (roughly nanometer-scale) and "macroscopic" (roughly micron-scale), respectively. Here, we develop a simulation framework to estimate the simultaneous impact of these factors on methane mass transport in silicate channels. Specifically, we develop a model of methane transport using homogenization theory to obtain transport parameters valid at length scales of hundreds to thousands of nanometers. These parameters implicitly reflect interactions taking place at fractions of a nanometer. The inputs to the homogenization analysis are data from extensive molecular dynamics simulations that incorporate atomistic-scale interactions, processed to yield local diffusion coefficients and mean force potentials. With this model, we demonstrate how nuances in silicate hydration and silica/methane interactions impact methane diffusion rates in silicate materials, including the effects of silicate surface chemistry such as the presence of silanol groups. The molecular dynamics simulations indicate that methane diffusivity at the silica surface is lower than the bulk-like rates observed at the center of channels of sufficient width. However, potentials of mean force generally evidence attractive methane/silica interactions that enhance diffusion overall. By simultaneously accounting for both of these effects, we show that the effective diffusion coefficient for the nanoporous silicate can be approximately double the value of estimates assuming fully bulk-like behavior in the channel. This study therefore demonstrates the importance of determining diffusion coefficients and potentials of mean force at an atomistic level when estimating transport properties in bulk materials. Importantly, we provide a simple homogenization framework to incorporate these molecular-scale attributes into bulk material transport estimates. This hybrid homogenization/molecular dynamics approach will be of general use for describing small-molecule transport in materials with detailed molecular interactions.
Asunto(s)
Metano , Simulación de Dinámica Molecular , Difusión , Silicatos , AguaRESUMEN
Background. Traditionally, osteolysis around total knee replacements (TKRs) is treated with complete revision. In certain subsets, polyethylene insert exchange and bone grafting may be applicable. This study reports the clinical outcomes for selective bone grafting in patients with osteolysis without complete revision of the TKR. Methods. This retrospective study analyzes 10 TKRs (9 patients, 66.5 ± 6.1 years old) presenting with osteolysis and revised after 8.7 ± 1.9 years of in vivo function. At index TKR, all patients were implanted with uncemented prosthesis and modular polyethylene insert with anteroposterior articular constraint (Ultracongruent, Natural Knee II, Sulzer Medica). The surgical technique for treating the osteolysis included removal of necrotic bone tissue using curettage, filling of the defect with bone graft materials, and polyethylene insert exchange. Results. Patients have not exhibited any further complications associated with osteolysis after 5.1 ± 2.4 years of followup. Routine radiographic exams show total incorporation of the graft material into the previously lytic regions in all patients. Conclusion. In some TKRs with osteolysis and firmly fixed components, the removal of lytic tissue and subsequent defect filling with bone graft materials can be a viable solution. This case series shows complete resolution of osteolysis in all patients with no complications.