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We sought to identify and quantitatively analyze calcium oxalate (CaOx) kidney stones on the order of micrometers, with a focus on the quantitative identification of calcium oxalate monohydrate (COM) and dihydrate (COD). We performed Fourier transform infrared (FTIR) spectroscopy, powder X-ray diffraction (PXRD), and microfocus X-ray computed tomography measurements (microfocus X-ray CT) and compared their results. An extended analysis of the FTIR spectrum focusing on the 780 cm-1 peak made it possible to achieve a reliable analysis of the COM/COD ratio. We succeeded in the quantitative analysis of COM/COD in 50-µm2 areas by applying microscopic FTIR for thin sections of kidney stones, and by applying microfocus X-ray CT system for bulk samples. The analysis results based on the PXRD measurements with micro-sampling, the microscopic FTIR analysis of thin sections, and the microfocus X-ray CT system observation of a bulk kidney stone sample showed roughly consistent results, indicating that all three methods can be used complementarily. This quantitative analysis method evaluates the detailed CaOx composition on the preserved stone surface and provides information on the stone formation processes. This information clarifies where and which crystal phase nucleates, how the crystals grow, and how the transition from the metastable phase to the stable phase proceeds. The phase transition affects the growth rate and hardness of kidney stones and thus provides crucial clues to the kidney stone formation process.
Asunto(s)
Oxalato de Calcio , Cálculos Renales , Humanos , Oxalato de Calcio/química , Cálculos Renales/diagnóstico por imagen , Cálculos Renales/química , Espectroscopía Infrarroja por Transformada de Fourier , Tomografía Computarizada por Rayos X , Rayos XRESUMEN
The pathogenesis of kidney stone formation includes multi-step processes involving complex interactions between mineral components and protein matrix. Calcium-binding proteins in kidney stones have great influences on the stone formation. The spatial distributions of these proteins in kidney stones are essential for evaluating the in vivo effects of proteins on the stone formation, although the actual distribution of these proteins is still unclear. We reveal micro-scale distributions of three different proteins, namely osteopontin (OPN), renal prothrombin fragment 1 (RPTF-1), and calgranulin A (Cal-A), in human kidney stones retaining original mineral phases and textures: calcium oxalate monohydrate (COM) and calcium oxalate dihydrate (COD). OPN and RPTF-1 were distributed inside of both COM and COD crystals, whereas Cal-A was distributed outside of crystals. OPN and RPTF-1 showed homogeneous distributions in COM crystals with mosaic texture, and periodically distributions parallel to specific crystal faces in COD crystals. The unique distributions of these proteins enable us to interpret the different in vivo effects of each protein on CaOx crystal growth based on their physico-chemical properties and the complex physical environment changes of each protein. This method will further allow us to elucidate in vivo effects of different proteins on kidney stone formation.
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Proteínas de Unión al Calcio/metabolismo , Cálculos Renales/diagnóstico por imagen , Riñón/patología , Osteopontina/metabolismo , Fragmentos de Péptidos/metabolismo , Precursores de Proteínas/metabolismo , Protrombina/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Oxalato de Calcio/química , Oxalato de Calcio/metabolismo , Cristalización/métodos , Femenino , Humanos , Riñón/metabolismo , Masculino , Microscopía Electrónica de Rastreo/métodos , Persona de Mediana EdadRESUMEN
It is important to reveal the exact cause of poor diffractivity in protein crystals in order to determine the accurate structure of protein molecules. It is shown that there is a large amount of local strain in subgrains of glucose isomerase crystals even though the overall crystal quality is rather high, as shown by clear equal-thickness fringes in X-ray topography. Thus, a large stress is exerted on the subgrains of protein crystals, which could significantly lower the resistance of the crystals to radiation damage. It is also demonstrated that this local strain can be reduced through the introduction of dislocations in the crystal. This suggests that the introduction of dislocations in protein crystals can be effective in enhancing the crystal quality of subgrains of protein crystals. By exploiting this effect, the radiation damage in subgrains could be decreased, leading to the collection of X-ray diffraction data sets with high diffractivity.
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Isomerasas Aldosa-Cetosa/química , Difracción de Rayos X/métodos , Conformación ProteicaRESUMEN
The microscopic understanding of the crystal growth and dissolution processes have been greatly advanced by the direct imaging of nanoscale step flows by atomic force microscopy (AFM), optical interferometry, and X-ray microscopy. However, one of the most fundamental events that govern their kinetics, namely, atomistic events at the step edges, have not been well understood. In this study, we have developed high-speed frequency modulation AFM (FM-AFM) and enabled true atomic-resolution imaging in liquid at â¼1 s/frame, which is â¼50 times faster than the conventional FM-AFM. With the developed AFM, we have directly imaged subnanometer-scale surface structures around the moving step edges of calcite during its dissolution in water. The obtained images reveal that the transition region with typical width of a few nanometers is formed along the step edges. Building upon insight in previous studies, our simulations suggest that the transition region is most likely to be a Ca(OH)2 monolayer formed as an intermediate state in the dissolution process. On the basis of this finding, we improve our understanding of the atomistic dissolution model of calcite in water. These results open up a wide range of future applications of the high-speed FM-AFM to the studies on various dynamic processes at solid-liquid interfaces with true atomic resolution.
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Nucleation, the primary step in crystallization, dictates the number of crystals, the distribution of their sizes, the polymorph selection, and other crucial properties of the crystal population. We used time-resolved liquid-cell transmission electron microscopy (TEM) to perform an in situ examination of the nucleation of lysozyme crystals. Our TEM images revealed that mesoscopic clusters, which are similar to those previously assumed to consist of a dense liquid and serve as nucleation precursors, are actually amorphous solid particles (ASPs) and act only as heterogeneous nucleation sites. Crystalline phases never form inside them. We demonstrate that a crystal appears within a noncrystalline particle assembling lysozyme on an ASP or a container wall, highlighting the role of heterogeneous nucleation. These findings represent a significant departure from the existing formulation of the two-step nucleation mechanism while reaffirming the role of noncrystalline particles. The insights gained may have significant implications in areas that rely on the production of protein crystals, such as structural biology, pharmacy, and biophysics, and for the fundamental understanding of crystallization mechanisms.
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The normal growth rates of the {110} faces of tetragonal hen egg-white lysozyme crystals, R, were measured as a function of the supersaturation σ parameter using a reflection type interferometer under µG at the International Space Station (NanoStep Project). Since water slightly evaporated from in situ observation cells during a long-term space station experiment for several months, equilibrium temperature T(e) changed, and the actual σ, however, significantly increased mainly due to the increase in salt concentration C(s). To correct σ, the actual C(s) and protein concentration C(p), which correctly represent the measured T(e) value in space, were first calculated. Second, a new solubility curve with the corrected C(s) was plotted. Finally, the revised σ was obtained from the new solubility curve. This correction method successfully revealed that the 2.8% water was evaporated from the solution, leading to 2.8% increase in the C(s) and C(p) of the solution.
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Medio Ambiente Extraterrestre , Muramidasa/química , Nave Espacial , Temperatura , Agua/química , Cristalización , Solubilidad , VolatilizaciónRESUMEN
We demonstrate the oscillatory phenomenon for the twisting growth of a triclinic crystal through in situ observation of the concentration field around the growing tip of a needle by high-resolution phase-shift interferometry.
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The formation of crystals from solution requires the initial self-assembly of units of matter into stable periodic structures reaching a critical size. The early stages of this process , called nucleation, are very difficult to visualize. Here we describe a novel method that allows real time observation of the dynamics of nucleation and dissolution of sodium chlorate clusters in an ionic liquid solution using in situ transmission electron microscopy. Using ionic liquids as solvent circumvents the problem of evaporation and charging, while the nucleation frequency was reduced by using saturated solutions. We observe simultaneous formation and dissolution of prenucleation clusters, suggesting that high-density fluctuations leading to solid cluster formation exist even under equilibrium conditions. In situ electron diffraction patterns reveal the simultaneous formation of crystalline nuclei of two polymorphic structures, the stable cubic phase and the metastable monoclinic phase, during the earliest stages of nucleation. These results demonstrate that molecules in solution can form clusters of different polymorphic phases independently of their respective solubility.
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The growth rate vs. supersaturation of a lysozyme crystal was successfully measured in situ together with the crystal surface observation and the concentration measurements onboard the International Space Station. A Michelson-type interferometer and a Mach-Zehnder interferometer were, respectively, employed for real-time growth rate measurements and concentration field measurements. The hardware development, sample preparation, operation, and analysis methods are described.
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Cristalización/instrumentación , Rayos Láser , Muramidasa/química , Ingravidez , Animales , Interferometría , Cinética , TemperaturaRESUMEN
The majority of water has vanished from modern meteorites, yet there remain signatures of water on ancient asteroids. How and when water disappeared from the asteroids is important, because the final fluid-concentrated chemical species played critical roles in the early evolution of organics and in the final minerals in meteorites. Here we show evidence of vestigial traces of water based on a nanometre-scale palaeomagnetic method, applying electron holography to the framboids in the Tagish Lake meteorite. The framboids are colloidal crystals composed of three-dimensionally ordered magnetite nanoparticles and therefore are only able to form against the repulsive force induced by the surface charge of the magnetite as a water droplet parches in microgravity. We demonstrate that the magnetites have a flux closure vortex structure, a unique magnetic configuration in nature that permits the formation of colloidal crystals just before exhaustion of water from a local system within a hydrous asteroid.
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Magnesite growth in chloride and sulfate-rich solutions has been examined at 90 °C in situ using phase-shift interferometry (PSI) and ex situ using atomic force microscopy (AFM) to evaluate the feasibility of cosequestering SO2 and CO2 in Mg-rich rocks. Although sulfate may assist desolvation at the magnesite surface, evidence for enhanced growth was only found at specific surface sites. The overall growth rates fit with those observed for chloride experiments in similarly saturated solutions. Thus, the formation of Mg-SO4 ion pairs in solution, which lowers the supersaturation with respect to magnesite, will have the dominant effect during sequestration. Lowering the activity of Mg(2+) ions in solution also inhibited the nucleation of other hydrated Mg-carbonate phases. As no evidence was found for sulfate incorporation into the growing magnesite, the presence of sulfate in solution will be detrimental to CO2 sequestration and is not expected to be cosequestered. The PSI data also emphasize the variability of reactivity over the surface and how this changes as a function of solution saturation and composition.
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Cloruros/análisis , Magnesio/análisis , Nanotecnología , Sulfatos/análisis , Microscopía de Fuerza Atómica , SolucionesRESUMEN
To start systematically investigating the quality improvement of protein crystals, the elementary growth processes of protein crystals must be first clarified comprehensively. Atomic force microscopy (AFM) has made a tremendous contribution toward elucidating the elementary growth processes of protein crystals and has confirmed that protein crystals grow layer by layer utilizing kinks on steps, as in the case of inorganic and low-molecular-weight compound crystals. However, the scanning of the AFM cantilever greatly disturbs the concentration distribution and solution flow in the vicinity of growing protein crystals. AFM also cannot visualize the dynamic behavior of mobile solute and impurity molecules on protein crystal surfaces. To compensate for these disadvantages of AFM, in situ observation by two types of advanced optical microscopy has been recently performed. To observe the elementary steps of protein crystals noninvasively, laser confocal microscopy combined with differential interference contrast microscopy (LCM-DIM) was developed. To visualize individual mobile protein molecules, total internal reflection fluorescent (TIRF) microscopy, which is widely used in the field of biological physics, was applied to the visualization of protein crystal surfaces. In this review, recent progress in the noninvasive in situ observation of elementary steps and individual mobile protein molecules on protein crystal surfaces is outlined.
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Cristalización/métodos , Microscopía/métodos , Proteínas/químicaRESUMEN
Three-dimensional colloidal crystals made of ferromagnetic particles, such as magnetite (Fe(3)O(4)), cannot be synthesized in principle because of the strong attractive magnetic interaction. However, we discovered colloidal crystals composed of polyhedral magnetite nanocrystallites of uniform size in the range of a few hundred nanometers in the Tagish Lake meteorite. Those colloidal crystals were formed 4.6 billion years ago and thus are much older than natural colloidal crystals on earth, such as opals, which formed about 100 million years ago. We found that the size of each individual magnetite particle determines its morphology, which in turn plays an important role in deciding the packing structure of the colloidal crystals. We also hypothesize that each particle has a flux-closed magnetic domain structure, which reduces the interparticle magnetic force significantly.
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In this study, a real-time optical system was developed to observe crystallization in a small spherical melt droplet (few millimeters in diameter) by containerless processing. This system can be used to simultaneously observe the inside and the surface of a transparent melt droplet, as well as its ambient gas atmosphere at high temperatures. A silicate melt with a diameter of approximately 2 mm and a composition of MgO:SiO(2)=48:52 was levitated using a gas-jet levitation system, and its crystallization process was successfully observed from 2385 K in real time with good contrast using the developed optical setup.