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Morphological and elemental mapping of gallstones using synchrotron microtomography and synchrotron X‐ray fluorescence spectroscopy
摘要： Background and Aim: Regional differences in gallstone (GS) composition are well documented in the Indian subcontinent. The reasons for the same are unknown. Etiopathogenesis of GS remains elusive despite advances in instrumentation. This was an in-depth analysis of the chemical, structural, and elemental composition of GS with special reference to synchroton studies. Methods: We used high-end sensitive analytical complementary microscopic and spectroscopic methods techniques, such as X-ray diffraction, scanning electron microscopy, Fourier transform infrared, synchrotron X-ray fluorescence spectroscopy (SR-XRF), and 2D and 3D synchrotron microtomography (SR-μCT), to study the ultra structure and trace element composition of three major types of GS (cholesterol, mixed, and pigment). SR-XRF quantified the trace elements in GS. Results: The cholesterol GS (monohydrate and anhydrate) were crystalline, with high calcium content. The pigment GS were amorphous, featureless, black, and fragile, with high calcium bilirubinate and carbonate salts. They had the highest concentration of iron (average 31.50 ppm) and copper (average 92.73 ppm), with bacterial inclusion. The mixed stones had features of both cholesterol and pigment GS with intermediate levels of copper (average 20.8 ppm) and iron (average 17.78 ppm). Conclusion: SR-μCT has, for the first time, provided cross-sectional computed imaging delineating the framework of GS and mineral distribution. It provided excellent mapping of cholesterol GS. SR-XRF confirmed that pigment GS had high concentrations of copper and iron with bacterial inclusions, the latter possibly serving as a nidus to the formation of these stones.
关键词： pigment,microtomography,gallstones,cholesterol,elemental,synchroton
更新于2025-11-21 11:18:25
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Cholesterol Functionalization of Gold Nanoparticles Enhances Photo-Activation of Neural Activity
摘要： Gold nanoparticles (AuNPs) attached to the extracellular leaflet of the plasma membrane of neurons can enable the generation of action potentials (APs) in response to brief pulses of light. Recently described techniques to stably bind AuNP bioconjugates directly to membrane proteins (ion channels) in neurons enable robust AP generation mediated by the photoexcited conjugate. However, a strategy that binds the AuNP to the plasma membrane in a non-protein-specific manner could represent a simple, single-step means of establishing light-responsiveness in multiple types of excitable neurons contained in the same tissue. Based on the ability of cholesterol to insert into the plasma membrane, here we test whether AuNP functionalization with linear dihydrolipoic acid-poly(ethylene) glycol (DHLA-PEG) chains that are distally terminated with cholesterol (AuNP-PEG-Chol) can enable light-induced AP generation in neurons. Dorsal root ganglion (DRG) neurons of rat were labelled with 20 nm diameter spherical AuNP-PEG-Chol conjugates wherein ~30% of the surface ligands (DHLA-PEG-COOH) were conjugated to PEG-Chol. Voltage recordings under current-clamp conditions showed that DRG neurons labeled in this manner exhibited a capacity for AP generation in response to microsecond and millisecond pulses of 532 nm light, a property attributable to the close tethering of AuNP-PEG-Chol conjugates to the plasma membrane facilitated by the cholesterol moiety. Light-induced AP and subthreshold depolarizing responses of the DRG neurons were similar to those previously described for AuNP conjugates targeted to channel proteins using large, multicomponent immunoconjugates. This likely reflected the AuNP-PEG-Chol’s ability, upon plasmonic light absorption and resultant slight and rapid heating of the plasma membrane, to induce a concomitant transmembrane depolarizing capacitive current. Notably, AuNP-PEG-Chol delivered to DRG neurons by inclusion in the buffer contained in the recording pipette/electrode enabled similar light-responsiveness, consistent with the activity of AuNP-PEG-Chol bound to the inner (cytofacial) leaflet of the plasma membrane. Our results demonstrate the ability of AuNP-PEG-Chol conjugates to confer timely stable and direct responsiveness to light in neurons. Further, this strategy represents a general approach for establishing excitable cell photosensitivity that could be of substantial advantage for exploring a given tissue’s suitability for AuNP-mediated photo-control of neural activity.
关键词： nanoparticle functionalization,cholesterol,action potential,neural photo-activation,optocapacitance,gold nanoparticles,photosensitivity,dorsal root ganglion cell
更新于2025-09-23 15:23:52
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Novel lanthanidomesogens based on poly(ethylene glycol) cholesterol ether: Liquid crystal phase behavior and luminescence properties
摘要： Biocompatible systems with luminescent properties attract significant interest as they are broadly used in bioanalysis and medical diagnostics. This paper describes new lanthanide liquid crystals based on an oxyethylated amphiphile with a cholesterol fragment (CholEO10). The complex of physicochemical methods (polarizing optical microscopy (POM), X-ray powder diffraction, FTIR-spectroscopy and luminescent analysis) was used to study liquid crystalline, structural and optical properties of these new biomesogens. Their phase behavior was compared with the behavior a lanthanum-containing analogue without a cholesterol fragment in its structure. A rigid hydrophobic fragment was found to exert a strong influence on lyotropic mesomorphism in aqueous media. The phase transition from a lamellar mesophase to a hexagonal one was observed with decrease of the CholEO10/La3+ complex concentration and addition of water. The study of luminescent properties of CholEO10/Ln3+ complexes, where Ln3+ is Dy3+, Tb3+, or Eu3+, revealed that their characteristic emission occurs in the yellow, green and red spectral ranges. Optical characteristics of new mesogens were estimated. Mesogenic CholEO10/Tb3+ and CholEO10/Eu3+ complexes demonstrate longer lifetime of their excited state as compared with the C12EO10/Ln3+/H2O complex.
关键词： Cholesterol derivatives,Liquid crystals,Luminescence,Poly(ethylene glycol),Lanthanide
更新于2025-09-23 15:21:21
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Usefulness of magnetically-controlled MNPs-enzymes microreactors for the fluorimetric determination of total cholesterol in serum
摘要： A new dynamic method containing a magnetically retained enzyme reactor (MRER) located in the reaction/detection zone of a flow injection (FI) system, has been used for the determination of total cholesterol in serum samples. The MRER was formed by a mixture ratio of 2/1 of immobilized enzymes cholesterol esterase (ChE) and cholesterol oxidase (COx) on magnetic nanoparticles (MNPs). The analytical signal is based on the fluorescence decreasing of the fluorophore naphtofluorescein (NF) due to its oxidation by the H2O2 formed in the enzymatic reactions. The dynamic range of the calibration graph was 1.55–100 mmol L?1 expressed as total cholesterol concentration (r2 = 0.9995, n = 5, r = 3), and the detection limit was 0.65 mmol L?1. The precision expressed as relative standard deviation (RSD %) was in the range of 4.7 and 0.6%. The method showed a sampling frequency of 10 h?1 and this method was applied to the determination of cholesterol in serum samples. The results were compared with those obtained using a previous automated clinical analyzer (ILab 600 analyzer). Also, recovery values ranging between 88.5 and 101.5% were achieved.
关键词： Fluorimetric,Naphtofluorescein,Electromagnet retention,Cholesterol,Serum,Enzyme magnetic nanoparticles
更新于2025-09-23 15:19:57
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Investigation of the Internalization of Fluorescently Labeled Lipophilic siRNA into Cultured Tumor Cells
摘要： The attachment of lipophilic molecules of natural origin, which have natural means for cell internalization, to small interfering RNA (siRNA) is an effective way of delivering siRNA to cells for biomedical purposes in vitro and in vivo. Earlier, we showed that the attachment of cholesterol to the 5'-end of the sense strand of nuclease-resistant siRNA through the optimized linker allows it to penetrate the cells and suppress the expression of the target gene. However, the effectiveness of the conjugates is different for cells of different origin, and in hematopoietic cells, they are not active, despite effective accumulation. In this work, we investigated the accumulation of fluorescently labeled cholesterol conjugates of siRNA using endocytosis inhibitors and showed that fluorescently labeled 5'-cholesterol conjugate of siRNAs penetrate KB-3-1 and K562 cells in several ways whose contribution differs depending on cell type and the presence of serum. In a serum-free medium, it was found that macropinocytosis and clathrin-dependent endocytosis contribute to the accumulation of the conjugate in KB-3-1 cells, while clathrin-dependent endocytosis makes the main contribution in K562 cells, while inhibitors of different types of endocytosis do not reduce the biological activity of the conjugate without a fluorescent label.
关键词： cell delivery mechanism,inhibitors of endocytosis,siRNA,cholesterol conjugate,chemical modifications
更新于2025-09-19 17:13:59
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In-Situ imaging detection of cell membrane and intracellular cholesterol via cascade reactions
摘要： Herein, an e?ective membrane-to-intracellular cholesterol detection strategy was designed based on cascade reactions. A biochip array was ?rstly fabricated by consecutively immobilizing luminol modi?ed gold nanoparticles (Au@luminol), soybean peroxidase (SBP) and cholesterol oxidase (ChoX) on the cellulose acetate (CA) membrane functionalized home-made micropore array. When cholesterol existed, it was oxidized by ChoX generating H2O2, which further triggered the CL reaction under the SBP catalysis, the CL signals were collected by a charge-coupled device (CCD). The proposed strategy exhibited a wide linear range from 0.12 μM to 1000 μM and relatively low detection limit (LOD) of 0.08 μM. Furthermore，it could be used to in-situ detect membrane cholesterol and intracelluar esteri?ed cholesterol in HepG2 cells. After activated HepG2 cells were added to the modi?ed biochip, membrane cholesterol was detected directly. Intracelluar esteri?ed cholesterol was detected through the introduction of triton X-100 and cholesteryl esterase (ChoE). Additionally, the cholesterol content in cells was changed after stimulated by drugs, such as apolipoprotein A-I (ApoA-I), pitavastatin or probucol. The correlation of the CL signal with the amount of cholesterol con?rmed that our strategy was feasible to simultaneously detect membrane and intracellular cholesterol at di?erent cellular states. The proposed strategy exhibited excellent sensitivity, selectivity, stability, and reproducibility in a simple, cheap way, which opened a new door for studying clinic treatment of the cholesterol-related diseases.
关键词： CL image,In-situ,Intracelluar cholesterol,Membrane cholesterol
更新于2025-09-10 09:29:36
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Cytotoxic and Membrane Cholesterol Effects of Ultraviolet Irradiation and Zinc Oxide Nanoparticles on Chinese Hamster Ovary Cells
摘要： Zinc Oxide (ZnO) nanoparticles are suspected to produce toxic effects toward mammalian cells; however, discrepancies in the extent of this effect have been reported between different cell lines. Simultaneously, high levels of ultraviolet (UV-C) radiation can have carcinogenic effects. The mechanism of this effect is also not well understood. Due to similarities in phenotype morphology after cell exposure to ZnO nanoparticles and UV-C irradiation, we emit the hypothesis that the toxicity of both these factors is related to damage of cellular membranes and affect their sterol content. Wild-type Chinese Hamster Ovary (CHO-K1) cells were exposed to ZnO nanoparticles or UV-C radiation. The amount of absorbed ZnO was determined by UV-visible spectroscopy and the changes in sterol pro?les were evaluated by gas chromatography. Cell viability after both treatments was determined by microscopy. Comparing morphology results suggested similarities in toxicology events induced by ZnO nanoparticles and UV exposure. UV-C exposure for 360 min disrupts the sterol metabolic pathway by increasing the concentration of cholesterol by 21.6-fold. This increase in cholesterol production supports the hypothesis that UV irradiation has direct consequences in initiating sterol modi?cations in the cell membrane.
关键词： ZnO,cholesterol,toxicity,sterol content,UV-radiation
更新于2025-09-09 09:28:46