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1.
Nano Lett ; 24(6): 1882-1890, 2024 Feb 14.
Article in English | MEDLINE | ID: mdl-38198287

ABSTRACT

Understanding the spatial organization of membrane proteins is crucial for unraveling key principles in cell biology. The reaction-diffusion model is commonly used to understand biochemical patterning; however, applying reaction-diffusion models to subcellular phenomena is challenging because of the difficulty in measuring protein diffusivity and interaction kinetics in the living cell. In this work, we investigated the self-organization of the plasmalemma vesicle-associated protein (PLVAP), which creates regular arrangements of fenestrated ultrastructures, using single-molecule tracking. We demonstrated that the spatial organization of the ultrastructures is associated with a decrease in the association rate by actin destabilization. We also constructed a reaction-diffusion model that accurately generates a hexagonal array with the same 130 nm spacing as the actual scale and informs the stoichiometry of the ultrastructure, which can be discerned only through electron microscopy. Through this study, we integrated single-molecule experiments and reaction-diffusion modeling to surpass the limitations of static imaging tools and proposed emergent properties of the PLVAP ultrastructure.


Subject(s)
Carrier Proteins , Membrane Proteins , Membrane Proteins/metabolism , Diffusion , Models, Biological
2.
Nat Commun ; 14(1): 7619, 2023 Nov 22.
Article in English | MEDLINE | ID: mdl-37993434

ABSTRACT

The biological process of aging is thought to result in part from accumulation of senescent cells in organs. However, the present study identified a subset of fibroblasts and smooth muscle cells which are the major constituents of organ stroma neither proliferative nor senescent in tissues of the elderly, which we termed "mid-old status" cells. Upregulation of pro-inflammatory genes (IL1B and SAA1) and downregulation of anti-inflammatory genes (SLIT2 and CXCL12) were detected in mid-old cells. In the stroma, SAA1 promotes development of the inflammatory microenvironment via upregulation of MMP9, which decreases the stability of epithelial cells present on the basement membrane, decreasing epithelial cell function. Remarkably, the microenvironmental change and the functional decline of mid-old cells could be reversed by a young cell-originated protein, SLIT2. Our data identify functional reversion of mid-old cells as a potential method to prevent or ameliorate aspects of aging-related tissue dysfunction.


Subject(s)
Aging , Cellular Senescence , Humans , Aged , Cellular Senescence/genetics , Aging/genetics , Epithelial Cells/physiology , Fibroblasts , Myocytes, Smooth Muscle
3.
Anal Chem ; 95(43): 15924-15932, 2023 10 31.
Article in English | MEDLINE | ID: mdl-37774148

ABSTRACT

In live cells, the plasma membrane is composed of lipid domains separated by hundreds of nanometers in dynamic equilibrium. Lipid phase separation regulates the trafficking and spatiotemporal organization of membrane molecules that promote signal transduction. However, visualizing domains with adequate spatiotemporal accuracy remains challenging because of their subdiffraction limit size and highly dynamic properties. Here, we present a single lipid-molecular motion analysis pipeline (lipid-MAP) for analyzing the phase heterogeneity of lipid membranes by detecting the instantaneous velocity change of a single lipid molecule using the excellent optical properties of nanoparticles, high spatial localization accuracy of single-molecule localization microscopy, and separation capability of the diffusion state of the hidden Markov model algorithm. Using lipid-MAP, individual lipid molecules were found to be in dynamic equilibrium between two statistically distinguishable phases, leading to the formation of small (∼170 nm), viscous (2.5× more viscous than surrounding areas), and transient domains in live cells. Moreover, our findings provide an understanding of how membrane compositional changes, i.e., cholesterol and phospholipids, affect domain formation. This imaging method can contribute to an improved understanding of spatiotemporal-controlled membrane dynamics at the molecular level.


Subject(s)
Phospholipids , Signal Transduction , Cell Membrane/metabolism , Phospholipids/metabolism , Membranes , Diffusion , Lipid Bilayers/metabolism
4.
JACS Au ; 2(7): 1596-1603, 2022 Jul 25.
Article in English | MEDLINE | ID: mdl-35911456

ABSTRACT

Live video recording of intracellular material transport is a promising means of deciphering the fascinating underlying mechanisms driving life at the molecular level. Such technology holds the key to realizing real-time observation at appropriate resolutions in three-dimensional (3D) space within living cells. Here, we report an optical microscopic method for probing endosomal dynamics with proper spatiotemporal resolution within 3D space in live cells: plasmonic dark-field STORM (pdf-STORM). We first confirmed that pdf-STORM has a spatial resolution comparable to that of scanning electron microscopy. Additionally, by observing two optical probes within a single organelle, we were able to track rotational movements and demonstrate the feasibility of using pdf-STORM to observe the angular displacements of an endosome during a "tug-of-war" over an extended period. Finally, we show various biophysical parameters of the hitherto unelucidated dynamics of endosomes-angular displacement is discontinuous and y-axis movement predominates and follows a long-tail distribution.

5.
Nano Lett ; 21(16): 6998-7004, 2021 08 25.
Article in English | MEDLINE | ID: mdl-34339204

ABSTRACT

Solar-driven reactive oxygen species (ROS) generation is an attractive disinfection technique for cell death and water purification. However, most photocatalysts require high stability in the water environment and the production of ROS with a sufficient amount and diffusion length to damage pathogens. Here, a ROS generation system was developed consisting of tapered crystalline silicon microwires coated with anatase titanium dioxide for a conformal junction. The system effectively absorbed >95% of sunlight over 300-1100 nm, resulting in effective ROS generation. The system was designed to produce various ROS species, but a logistic regression analysis with cellular survival data revealed that the diffusion length of the ROS is ∼9 µm, implying that the most dominant species causing cell damage is H2O2. Surprisingly, a quantitative analysis showed that only 15 min of light irradiation on the system would catalyze a local bactericidal effect comparable to the conventional germicidal level of H2O2 (∼3 mM).


Subject(s)
Hydrogen Peroxide , Sunlight , Cell Death , Reactive Oxygen Species , Titanium
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