Microbiome metabolite quantification methods enabling insights into human health and disease.

Many of the health-associated impacts of the microbiome are mediated by its chemical activity, producing and modifying small molecules (metabolites). Thus, microbiome metabolite quantification has a central role in efforts to elucidate and measure microbiome function. In this review, we cover general considerations when designing experiments to quantify microbiome metabolites, including sample preparation, data acquisition and data processing, since these are critical to downstream data quality. We then discuss data analysis and experimental steps to demonstrate that a given metabolite feature is of microbial origin. We further discuss techniques used to quantify common microbial metabolites, including short-chain fatty acids (SCFA), secondary bile acids (BAs), tryptophan derivatives, N-acyl amides and trimethylamine N-oxide (TMAO). Lastly, we conclude with challenges and future directions for the field.

[Clinical characteristics and genetic analysis of a child with Char syndrome caused by TFAP2B gene variant].

OBJECTIVE: To explore the clinical features and genetic etiology of a child with Char syndrome. METHODS: A child who was presented at the Department of Child Health, Henan Children's Hospital in February 2022 was selected as the study subject. Clinical data of the child was collected, and peripheral blood samples of the child and her parents were collected for the extraction of genomic DNA. Whole exome sequencing was carried out, and candidate variants were verified by Sanger sequencing and bioinformatic analysis. RESULTS: The child had mainly manifested facial dysmorphism, patent ductus arteriosus, growth retardation, curving of fifth fingers and middle toes. Whole exome sequencing revealed that she has harbored a heterozygous c.944A>C (p.Glu315Ala) variant of the TFAP2B gene, which was verified to be de novo by Sanger sequencing. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was rated to be likely pathogenic (PM1+PM2_Supporting+PM6+PP3). CONCLUSION: The heterozygous c.944A>C (p.Glu315Ala) variant of the TFAP2B gene probably underlay the Char syndrome in this child. Above finding has expanded the mutational and phenotypic spectra of the TFAP2B gene, which has facilitated early identification and diagnosis of Char syndrome.

Selective Methane Oxidation to Methanol on ZnO/Cu2O/Cu(111) Catalysts: Multiple Site-Dependent Behaviors.

Because of the abundance of natural gas in our planet, a major goal is to achieve a direct methane-to-methanol conversion at medium to low temperatures using mixtures of methane and oxygen. Here, we report an efficient catalyst, ZnO/Cu2O/Cu(111), for this process investigated using a combination of reactor testing, scanning tunneling microscopy, ambient-pressure X-ray photoemission spectroscopy, density functional calculations, and kinetic Monte Carlo simulations. The catalyst is capable of methane activation at room temperature and transforms mixtures of methane and oxygen to methanol at 450 K with a selectivity of ∼30%. This performance is not seen for other heterogeneous catalysts which usually require the addition of water to enable a significant conversion of methane to methanol. The unique coarse structure of the ZnO islands supported on a Cu2O/Cu(111) substrate provides a collection of multiple centers that display different catalytic activity during the reaction. ZnO-Cu2O step sites are active centers for methanol synthesis when exposed to CH4 and O2 due to an effective O-O bond dissociation, which enables a methane-to-methanol conversion with a reasonable selectivity. Upon addition of water, the defected O-rich ZnO sites, introduced by Zn vacancies, show superior behavior toward methane conversion and enhance the overall methanol selectivity to over 80%. Thus, in this case, the surface sites involved in a direct CH4 → CH3OH conversion are different from those engaged in methanol formation without water. The identification of the site-dependent behavior of ZnO/Cu2O/Cu(111) opens a design strategy for guiding efficient methane reformation with high methanol selectivity.

Structure of Copper-Cobalt Surface Alloys in Equilibrium with Carbon Monoxide Gas.

We studied the structure of the copper-cobalt (CuCo) surface alloy, formed by Co deposition on Cu(110), in dynamic equilibrium with CO. Using scanning tunneling microscopy (STM), we found that, in vacuum at room temperature and at low Co coverage, clusters of a few Co atoms substituting Cu atoms form at the surface. At CO pressures in the Torr range, we found that up to 2.5 CO molecules can bind on a single Co atom, in carbonyl-like configurations. Based on high-resolution STM images, together with density functional theory calculations, we determined the most stable CuCo cluster structures formed with bound CO. Such carbonyl-like formation manifests in shifts in the binding energy of the Co core-level peaks in X-ray photoelectron spectra, as well as shifts in the vibrational modes of adsorbed CO in infrared reflection absorption spectra. The multiple CO adsorption on a Co site weakens the Co-CO bond and thus reduces the C-O bond scission probability. Our results may explain the different product distribution, including higher selectivity toward alcohol formation, when bimetallic CuCo catalysts are used compared to pure Co.

Direct Conversion of Methane to Methanol on Ni-Ceria Surfaces: Metal-Support Interactions and Water-Enabled Catalytic Conversion by Site Blocking.

The transformation of methane into methanol or higher alcohols at moderate temperature and pressure conditions is of great environmental interest and remains a challenge despite many efforts. Extended surfaces of metallic nickel are inactive for a direct CH4 → CH3OH conversion. This experimental and computational study provides clear evidence that low Ni loadings on a CeO2(111) support can perform a direct catalytic cycle for the generation of methanol at low temperature using oxygen and water as reactants, with a higher selectivity than ever reported for ceria-based catalysts. On the basis of ambient pressure X-ray photoemission spectroscopy and density functional theory calculations, we demonstrate that water plays a crucial role in blocking catalyst sites where methyl species could fully decompose, an essential factor for diminishing the production of CO and CO2, and in generating sites on which methoxy species and ultimately methanol can form. In addition to water-site blocking, one needs the effects of metal-support interactions to bind and activate methane and water. These findings should be considered when designing metal/oxide catalysts for converting methane to value-added chemicals and fuels.

Ceria-based model catalysts: fundamental studies on the importance of the metal-ceria interface in CO oxidation, the water-gas shift, CO2 hydrogenation, and methane and alcohol reforming.

Model metal/ceria and ceria/metal catalysts have been shown to be excellent systems for studying fundamental phenomena linked to the operation of technical catalysts. In the last fifteen years, many combinations of well-defined systems involving different kinds of metals and ceria have been prepared and characterized using the modern techniques of surface science. So far most of the catalytic studies have been centered on a few reactions: CO oxidation, the hydrogenation of CO2, and the production of hydrogen through the water-gas shift reaction and the reforming of methane or alcohols. Using model catalysts it has been possible to examine in detail correlations between the structural, electronic and catalytic properties of ceria-metal interfaces. In situ techniques (X-ray photoelectron spectroscopy, X-ray absorption spectroscopy, infrared spectroscopy, scanning tunneling microscopy) have been combined to study the morphological changes under reaction conditions and investigate the evolution of active phases involved in the cleavage of C-O, C-H and C-C bonds. Several studies with model ceria catalysts have shown the importance of strong metal-support interactions. In general, a substantial body of knowledge has been acquired and concepts have been developed for a more rational approach to the design of novel technical catalysts containing ceria.

A Dynamic Biochemomechanical Model of Geometry-Confined Cell Spreading.

Cell spreading is involved in many physiological and pathological processes. The spreading behavior of a cell significantly depends on its microenvironment, but the biochemomechanical mechanisms of geometry-confined cell spreading remain unclear. A dynamic model is here established to investigate the spreading of cells confined in a finite region with different geometries, e.g., rectangle, ellipse, triangle, and L-shape. This model incorporates both biophysical and biochemical mechanisms, including actin polymerization, integrin-mediated binding, plasma viscoelasticity, and the elasticity of membranes and microtubules. We simulate the dynamic configurational evolution of a cell under different geometric microenvironments, including the angular distribution of microtubule forces and the deformation of the nucleus. The results indicate that the positioning of the cell-division plane is affected by its boundary confinement: a cell divides in a plane perpendicular to its minimal principal axis of inertia of area. In addition, the effects of such physical factors as the adhesive bond density, membrane tension, and microtubule number are examined on the cell spreading dynamics. The theoretical predictions show a good agreement with relevant experimental results. This work sheds light on the geometry-confined spreading dynamics of cells and holds potential applications in regulating cell division and designing cell-based sensors.

Numerical and experimental investigation of GaN-based flip-chip light-emitting diodes with highly reflective Ag/TiW and ITO/DBR Ohmic contacts.

We demonstrate two types of GaN-based flip-chip light-emitting diodes (FCLEDs) with highly reflective Ag/TiW and indium-tin oxide (ITO)/distributed Bragg reflector (DBR) p-type Ohmic contacts. We show that a direct Ohmic contact to p-GaN layer using pure Ag is obtained when annealed at 600°C in N2 ambient. A TiW diffusion barrier layer covered onto Ag is used to suppress the agglomeration of Ag and thus maintain high reflectance of Ag during high temperature annealing process. We develop a strip-shaped SiO2 current blocking layer beneath the ITO/DBR to alleviate current crowding occurring in FCLED with ITO/DBR. Owing to negligibly small spreading resistance of Ag, however, our combined numerical and experimental results show that the FCLED with Ag/TiW has a more favorable current spreading uniformity in comparison to the FCLED with ITO/DBR. As a result, the light output power of FCLED with Ag/TiW is 7.5% higher than that of FCLED with ITO/DBR at 350 mA. The maximum output power of the FCLED with Ag/TiW obtained at 305.6 A/cm2 is 29.3% larger than that of the FCLED with ITO/DBR obtained at 278.9 A/cm2. The improvement appears to be due to the enhanced current spreading and higher optical reflectance provided by the Ag/TiW.

Unravelling hydrogen bonding interactions of tryptamine-water dimer from neutral to cation.

The neutral and cationic forms of tryptamine-water dimer present a variety of noncovalent interactions. To characterize these interactions, a series of complementary methods, including the quantum theory of atoms in molecules, noncovalent interaction plots, natural bond orbital analysis, and energy decomposition analysis, were used. For the first time, the existence of the three intermolecular H-bonds in the conformer-locked tryptamine-water dimer A-H2O are identified, highlighting a single water's role as one proton donor and two proton acceptors as it binds to tryptamine. Furthermore, upon threshold ionization of the A-H2O dimer, the network of the three intermolecular H-bonds is indeed preserved while the individual H-bonds' binding strengths are subject to change; this is attributed to the existence of the optically accessible minimum energy isomer A+-H2O in the cation. In addition, it is found that the global minimum energy isomer H+-H2O contains a single intermolecular H-bond, but is more stable, by ca. 3 kcal mol-1, than the local minimum energy isomer A+-H2O; this is due to the stronger intramolecular interaction of H+-H2O as opposed to A+-H2O.

In†Situ Investigation of Methane Dry Reforming on Metal/Ceria(111) Surfaces: Metal-Support Interactions and C-H Bond Activation at Low Temperature.

Studies with a series of metal/ceria(111) (metal=Co, Ni, Cu; ceria=CeO2 ) surfaces indicate that metal-oxide interactions can play a very important role for the activation of methane and its reforming with CO2 at relatively low temperatures (600-700 K). Among the systems examined, Co/CeO2 (111) exhibits the best performance and Cu/CeO2 (111) has negligible activity. Experiments using ambient pressure X-ray photoelectron spectroscopy indicate that methane dissociates on Co/CeO2 (111) at temperatures as low as 300 K-generating CHx and COx species on the catalyst surface. The results of density functional calculations show a reduction in the methane activation barrier from 1.07 eV on Co(0001) to 0.87 eV on Co2+ /CeO2 (111), and to only 0.05 eV on Co0 /CeO2-x (111). At 700 K, under methane dry reforming conditions, CO2 dissociates on the oxide surface and a catalytic cycle is established without coke deposition. A significant part of the CHx formed on the Co0 /CeO2-x (111) catalyst recombines to yield ethane or ethylene.

Ambient pressure XPS and IRRAS investigation of ethanol steam reforming on Ni-CeO2(111) catalysts: an in situ study of C-C and O-H bond scission.

Ambient-Pressure X-ray Photoelectron Spectroscopy (AP-XPS) and Infrared Reflection Absorption Spectroscopy (AP-IRRAS) have been used to elucidate the active sites and mechanistic steps associated with the ethanol steam reforming reaction (ESR) over Ni-CeO2(111) model catalysts. Our results reveal that surface layers of the ceria substrate are both highly reduced and hydroxylated under reaction conditions while the small supported Ni nanoparticles are present as Ni(0)/NixC. A multifunctional, synergistic role is highlighted in which Ni, CeOx and the interface provide an ensemble effect in the active chemistry that leads to H2. Ni(0) is the active phase leading to both C-C and C-H bond cleavage in ethanol and it is also responsible for carbon accumulation. On the other hand, CeOx is important for the deprotonation of ethanol/water to ethoxy and OH intermediates. The active state of CeOx is a Ce(3+)(OH)x compound that results from extensive reduction by ethanol and the efficient dissociation of water. Additionally, we gain an important insight into the stability and selectivity of the catalyst by its effective water dissociation, where the accumulation of surface carbon can be mitigated by the increased presence of surface OH groups. The co-existence and cooperative interplay of Ni(0) and Ce(3+)(OH)x through a metal-support interaction facilitate oxygen transfer, activation of ethanol/water as well as the removal of coke.

Dry Reforming of Methane on a Highly-Active Ni-CeO2 Catalyst: Effects of Metal-Support Interactions on C-H Bond Breaking.

Ni-CeO2 is a highly efficient, stable and non-expensive catalyst for methane dry reforming at relative low temperatures (700 K). The active phase of the catalyst consists of small nanoparticles of nickel dispersed on partially reduced ceria. Experiments of ambient pressure XPS indicate that methane dissociates on Ni/CeO2 at temperatures as low as 300 K, generating CHx and COx species on the surface of the catalyst. Strong metal-support interactions activate Ni for the dissociation of methane. The results of density-functional calculations show a drop in the effective barrier for methane activation from 0.9 eV on Ni(111) to only 0.15 eV on Ni/CeO2-x (111). At 700 K, under methane dry reforming conditions, no signals for adsorbed CHx or C species are detected in the C 1s XPS region. The reforming of methane proceeds in a clean and efficient way.

In situ and theoretical studies for the dissociation of water on an active Ni/CeO2 catalyst: importance of strong metal-support interactions for the cleavage of O-H bonds.

Water dissociation is crucial in many catalytic reactions on oxide-supported transition-metal catalysts. Supported by experimental and density-functional theory results, the effect of the support on OH bond cleavage activity is elucidated for nickel/ceria systems. Ambient-pressure O 1s photoemission spectra at low Ni loadings on CeO2 (111) reveal a substantially larger amount of OH groups as compared to the bare support. Computed activation energy barriers for water dissociation show an enhanced reactivity of Ni adatoms on CeO2 (111) compared with pyramidal Ni4 particles with one Ni atom not in contact with the support, and extended Ni(111) surfaces. At the origin of this support effect is the ability of ceria to stabilize oxidized Ni(2+) species by accommodating electrons in localized f-states. The fast dissociation of water on Ni/CeO2 has a dramatic effect on the activity and stability of this system as a catalyst for the water-gas shift and ethanol steam reforming reactions.

Time-integrated BMP signaling determines fate in a stem cell model for early human development.

How paracrine signals are interpreted to yield multiple cell fate decisions in a dynamic context during human development in vivo and in vitro remains poorly understood. Here we report an automated tracking method to follow signaling histories linked to cell fate in large numbers of human pluripotent stem cells (hPSCs). Using an unbiased statistical approach, we discover that measured BMP signaling history correlates strongly with fate in individual cells. We find that BMP response in hPSCs varies more strongly in the duration of signaling than the level. However, both the level and duration of signaling activity control cell fate choices only by changing the time integral. Therefore, signaling duration and level are interchangeable in this context. In a stem cell model for patterning of the human embryo, we show that signaling histories predict the fate pattern and that the integral model correctly predicts changes in cell fate domains when signaling is perturbed. Our data suggest that mechanistically, BMP signaling is integrated by SOX2.

Endogenous FGFs drive ERK-dependent cell fate patterning in 2D human gastruloids.

The role of FGF is the least understood of the morphogens driving mammalian gastrulation. Here we investigated the function of FGF in a stem cell model for human gastrulation known as a 2D gastruloid. We found a ring of FGF-dependent ERK activity that closely follows the emergence of primitive streak (PS)-like cells but expands further inward. We showed that this ERK activity pattern is required for PS-like differentiation and that loss of PS-like cells upon FGF receptor inhibition can be rescued by directly activating ERK. We further demonstrated that the ERK-ring depends on localized activation of basally localized FGF receptors (FGFR) by endogenous FGF gradients. We confirm and extend previous studies in analyzing expression of FGF pathway components, showing the main receptor to be FGFR1 and the key ligands FGF2/4/17, similar to the human and monkey embryo but different from the mouse. In situ hybridization and scRNA-seq revealed that FGF4 and FGF17 expression colocalize with the PS marker TBXT but only FGF17 is maintained in nascent mesoderm and endoderm. FGF4 and FGF17 reduction both reduced ERK activity and differentiation to PS-like cells and their derivatives, indicating overlapping function. Thus, we have identified a previously unknown role for FGF-dependent ERK signaling in 2D gastruloids and possibly the human embryo, driven by a mechanism where FGF4 and FGF17 signal through basally localized FGFR1 to induce PS-like cells.

Small molecule biomarkers predictive of Chagas disease progression.

Chagas disease (CD) is a neglected tropical disease caused by the parasitic protozoan Trypanosoma cruzi. However, only 20% to 30% of infected individuals will progress to severe symptomatic cardiac manifestations. Current treatments are benznidazole and nifurtimox, which are poorly tolerated regimens. Developing a biomarker to determine the likelihood of patient progression would be helpful for doctors to optimize patient treatment strategies. Such a biomarker would also benefit drug discovery efforts and clinical trials. In this study, we combined untargeted and targeted metabolomics to compare serum samples from T. cruzi-infected individuals who progressed to severe cardiac disease, versus infected individuals who remained at the same disease stage (non-progressors). We identified four unannotated biomarker candidates, which were validated in an independent cohort using both untargeted and targeted analysis techniques. Overall, our findings demonstrate that serum small molecules can predict CD progression, offering potential for clinical monitoring.

Basal actomyosin pulses expand epithelium coordinating cell flattening and tissue elongation.

Actomyosin networks constrict cell area and junctions to alter cell and tissue shape. However, during cell expansion under mechanical stress, actomyosin networks are strengthened and polarized to relax stress. Thus, cells face a conflicting situation between the enhanced actomyosin contractile properties and the expansion behaviour of the cell or tissue. To address this paradoxical situation, we study late Drosophila oogenesis and reveal an unusual epithelial expansion wave behaviour. Mechanistically, Rac1 and Rho1 integrate basal pulsatile actomyosin networks with ruffles and focal adhesions to increase and then stabilize basal area of epithelial cells allowing their flattening and elongation. This epithelial expansion behaviour bridges cell changes to oocyte growth and extension, while oocyte growth in turn deforms the epithelium to drive cell spreading. Basal pulsatile actomyosin networks exhibit non-contractile mechanics, non-linear structures and F-actin/Myosin-II spatiotemporal signal separation, implicating unreported expanding properties. Biophysical modelling incorporating these expanding properties well simulates epithelial cell expansion waves. Our work thus highlights actomyosin expanding properties as a key mechanism driving tissue morphogenesis.

Localized cardiac small molecule trajectories and persistent chemical sequelae in experimental Chagas disease.

Post-infectious conditions present major health burdens but remain poorly understood. In Chagas disease (CD), caused by Trypanosoma cruzi parasites, antiparasitic agents that successfully clear T. cruzi do not always improve clinical outcomes. In this study, we reveal differential small molecule trajectories between cardiac regions during chronic T. cruzi infection, matching with characteristic CD apical aneurysm sites. Incomplete, region-specific, cardiac small molecule restoration is observed in animals treated with the antiparasitic benznidazole. In contrast, superior restoration of the cardiac small molecule profile is observed for a combination treatment of reduced-dose benznidazole plus an immunotherapy, even with less parasite burden reduction. Overall, these results reveal molecular mechanisms of CD treatment based on simultaneous effects on the pathogen and on host small molecule responses, and expand our understanding of clinical treatment failure in CD. This link between infection and subsequent persistent small molecule perturbation broadens our understanding of infectious disease sequelae.

Localized cardiac metabolic trajectories and post-infectious metabolic sequelae in experimental Chagas disease.

Post-infectious conditions, where clinical symptoms fail to resolve even after pathogen clearance, present major health burdens. However, the mechanisms involved remain poorly understood. In Chagas disease (CD), caused by the parasite Trypanosoma cruzi, antiparasitic agents can clear T. cruzi but late-stage treatment does not improve clinical cardiac outcomes. In this study, we revealed differential metabolic trajectories of cardiac regions during T. cruzi infection, matching sites of clinical symptoms. Incomplete, region-specific, cardiac metabolic restoration was observed in animals treated with the antiparasitic benznidazole, even though parasites were successfully cleared. In contrast, superior metabolic restoration was observed for a combination treatment of reduced-dose benznidazole plus an immunotherapy (Tc24-C4 T. cruzi flagellar protein and TLR4 agonist adjuvant), even though parasite burden reduction was lower. Overall, these results provide a mechanism to explain prior clinical treatment failures in CD and to test novel candidate treatment regimens. More broadly, our results demonstrate a link between persistent metabolic perturbation and post-infectious conditions, with broad implications for our understanding of post-infectious disease sequelae.