Cell-bound membrane vesicles contain antioxidative proteins and probably have an antioxidative function in cells or a therapeutic potential

Atomic force microscopy reveals involvement of the cell envelope in biomechanical properties of sickle erythrocytes

Abstract Background Intracellular hemoglobin polymerization has been supposed to be the major determinant for the elevated rigidity/stiffness of sickle erythrocytes from sickle cell anemia (SCA) patients. However, the contribution of the cell envelope remains unclear. Results In this study, using atomic force microscopy (AFM), we compared the normal and sickled erythrocyte surfaces for stiffness and topography. AFM detected that sickle cells had a rougher surface and were stiffer than normal erythrocytes and that sickle cell ghosts had a rougher surface (for both outer and inner surfaces) and were thicker than normal ghosts, the latter implying a higher membrane-associated hemoglobin content/layer in the sickle cell envelope. Compared to healthy subjects, the SCA patients had lower plasma lipoprotein levels. AFM further revealed that a mild concentration of methyl-β-cyclodextrin (MβCD, a putative cholesterol-depleting reagent) could induce an increase in roughness of erythrocytes/ghosts and a decrease in thickness of ghosts for both normal and sickle cells, implying that MβCD can alter the cell envelope from outside (cholesterol in the plasma membrane) to inside (membrane-associated hemoglobin). More importantly, MβCD also caused a more significant decrease in stiffness of sickle cells than that of normal erythrocytes. Conclusions The data reveal that besides the cytosolic hemoglobin fibers, the cell envelope containing the membrane-associated hemoglobin also is involved in the biomechanical properties (e.g., stiffness and shape maintenance) of sickle erythrocytes.

A GSH-activated AIE-based polymer photosensitizer for killing cancer cells

A GSH-Activated AIE-Based Polymer Photosensitizer for Killing Cancer Cells

In situ isolation of nuclei or nuclear proteins from adherent cells: a simple, effective method with less cytoplasmic contamination

Abstract Background Isolation of nuclei or nuclear proteins is a prerequisite for western blot, nuclear proteome profiling, and other evaluations of nuclear proteins. Here, we developed a simple method for in situ isolation of nuclei or nuclear proteins by in situ removing the extranuclear part of adherent cells via a classical nonionic detergent triton X-100. Results First, the feasibility of our method was confirmed by confocal microscopy, atomic force microscopy, scanning electron microscopy, dynamic light scattering, immunofluorescence imaging, and time-lapse dynamic observation. Next, the optimal concentration range (approximately 0.1–1% for ~ 10 min) of triton X-100 and the optimal treatment time (< 30 min) of 0.1–1% Triton X-100 for our method were determined via western blotting of eight extra-/intra-nuclear proteins. Subsequently, the effectiveness, sensitivity, and cytoplasmic contamination of our method were tested by investigating the levels of phosphorylated p65 (a NF-κB subunit) in the nuclei of endothelial or tumor cells treated with/without lipopolysaccharide (LPS) via western blotting and by comparing with a commercial nuclear protein extraction kit (a classical detergent-based method). The data show that compared with the commercial kit our method obtained a higher yield of total nuclear proteins, a higher pP65 level in both control and LPS groups, and much lower content of GAPDH (as a reference for cytoplasmic contamination) in nuclei. Conclusions The in situ isolation of nuclei or nuclear proteins from adherent cells in this study is a simple, effective method with less cytoplasmic contamination. This method/strategy has the potential of improving the quality of downstream evaluations including western blotting and proteomic profiling.

Enhanced antibacterial activity of levofloxacin/hydroxypropyl-β-cyclodextrin inclusion complex: In vitro and in vivo evaluation

A new poly(norbornene)-based sensor for fluorescent ratiometric sensing of adenosine 5′-triphosphate

An amphiphilic tetraphenylethylene-based photosensitizer for cancer cell ablation and bacteria killing

Ganglioside GM3-Functionalized Reconstituted High-Density Lipoprotein (GM3-rHDL) as a Novel Nanocarrier Enhances Antiatherosclerotic Efficacy of Statins in apoE−/− C57BL/6 Mice

Previously, we found that exogenous ganglioside GM3 had an antiatherosclerotic efficacy and that its antiatherosclerotic efficacy could be enhanced by reconstituted high-density lipoprotein (rHDL). In this study, we hypothesized that GM3-functionalized rHDL (i.e., GM3-rHDL) as a nanocarrier can promote the efficacy of traditional antiatherosclerotic drugs (e.g., statins). To test this hypothesis, lovastatin (LT) was used as a representative of statins, and LT-loaded GM3-rHDL nanoparticle (LT-GM3-rHDL or LT@GM3-rHDL; a mean size of ~142 nm) and multiple controls (e.g., GM3-rHDL without LT, LT-loaded rHDL or LT-rHDL, and other nanoparticles) were prepared. By using two different microsphere-based methods, the presences of apolipoprotein A-I (apoA-I) and/or GM3 in nanoparticles and the apoA-I-mediated macrophage-targeting ability of apoA-I/rHDL-containing nanoparticles were verified in vitro. Moreover, LT-GM3-rHDL nanoparticle had a slowly sustained LT release in vitro and the strongest inhibitory effect on the foam cell formation of macrophages (a key event of atherogenesis). After single administration of rHDL-based nanoparticles, a higher LT concentration was detected shortly in the atherosclerotic plaques of apoE−/− mice than non-rHDL-based nanoparticles, suggesting the in vivo plaque-targeting ability of apoA-I/rHDL-containing nanoparticles. Finally, among all nanoparticles LT-GM3-rHDL induced the largest decreases in the contents of blood lipids and in the areas of atherosclerotic plaques at various aortic locations in apoE−/− mice fed a high-fat diet for 12 weeks, supporting that LT-GM3-rHDL has the best in vivo antiatherosclerotic efficacy among the tested nanoparticles. Our data imply that GM3-functionalized rHDL (i.e., GM3-rHDL) can be utilized as a novel nanocarrier to enhance the efficacy of traditional antiatherosclerotic drugs (e.g., statins).

In situ AFM detection of the stiffness of the in situ exposed cell nucleus

A new imidazolium/sulfonamide linked ferrocene-dansyl dyad for dual-channel recognition of anion

Effect of liver total sphingomyelin synthase deficiency on plasma lipid metabolism

A GSH-responsive PET-based fluorescent probe for cancer cells imaging

Enhanced Anti-Atherosclerotic Efficacy of pH-Responsively Releasable Ganglioside GM3 Delivered by Reconstituted High-Density Lipoprotein

Recently, the atheroprotective role of endogenous GM3 and an atherogenesis-inhibiting effect of exogenous GM3 suggested a possibility of exogenous GM3 being recruited as an anti-atherosclerotic drug. This study seeks to endow exogenous GM3 with atherosclerotic targetability via reconstituted high-density lipoprotein (rHDL), an atherosclerotic targeting drug nanocarrier. Unloaded rHDL, rHDL loaded with exogenous GM3 at a low concentration (GM3L-rHDL), and rHDL carrying GM3 at a relatively high concentration (GM3H-rHDL) were prepared and characterized. The inhibitory effect of GM3-rHDL on lipid deposition in macrophages was confirmed, and GM3-rHDL did not affect the survival of red blood cells. In vivo experiments using ApoE−/− mice fed a high fat diet further confirmed the anti-atherosclerotic efficacy of exogenous GM3 and demonstrated that GM3 packed in HDL nanoparticles (GM3-rHDL) has an enhanced anti-atherosclerotic efficacy and a reduced effective dose of GM3. Then, the macrophage- and atherosclerotic plaque-targeting abilities of GM3-rHD, most likely via the interaction of ApoA-I on GM3-rHDL with its receptors (e.g., SR-B1) on cells, were certified via a microsphere-based method and an aortic fragment-based method, respectively. Moreover, we found that solution acidification enhanced GM3 release from GM3-rHDL nanoparticles, implying the pH-responsive GM3 release when GM3-rHDL enters the acidic atherosclerotic plaques from the neutral blood. The rHDL-mediated atherosclerotic targetability and pH-responsive GM3 release of GM3-rHDL enhanced the anti-atherosclerotic efficacy of exogenous GM3. The development of the GM3-rHDL nanoparticle may help with the application of exogenous GM3 as a clinical drug. Moreover, the data imply that the GM3-rHDL nanoparticle has the potential of being recruited as a drug nanocarrier with atherosclerotic targetability and enhanced anti-atherosclerotic efficacy.

Methyl-β-cyclodextrin suppresses the monocyte-endothelial adhesion triggered by lipopolysaccharide (LPS) or oxidized low-density lipoprotein (oxLDL)

AFM detects the effects of acidic condition on the size and biomechanical properties of native/oxidized low-density lipoprotein

A New Poly(Norbornene)-Based Sensor for Fluorescent Ratiometric Sensing of Adenosine 5′-Triphosphate

Single-vesicle tracking reveals the potential correlation of the movement of cell-bound membrane vesicles (CBMVs) with cell migration

Fluorescent norbornene for sequential detection of mercury and biothiols

Isolated cell-bound membrane vesicles (CBMVs) as a novel class of drug nanocarriers

Abstract Background Cell-bound membrane vesicles (CBMVs) are a type of membrane vesicles different from the well-known extracellular vesicles (EVs). In recent years, the applications of EVs as drug delivery systems have been studied widely. A question may arise whether isolated CBMVs also have the possibility of being recruited as a drug delivery system or nanocarrier? Methods To test the possibility, CBMVs were isolated/purified from the surfaces of cultured endothelial cells, loaded with a putative antitumor drug doxorubicin (Dox), and characterized. Subsequently, cellular experiments and animal experiments using mouse models were performed to determine the in vitro and in vivo antitumor effects of Dox-loaded CBMVs (Dox-CBMVs or Dox@CBMVs), respectively. Results Both Dox-free and Dox-loaded CBMVs were globular-shaped and nanometer-sized with an average diameter of ~ 300–400 nm. Dox-CBMVs could be internalized by cells and could kill multiple types of cancer cells. The in vivo antitumor ability of Dox-CBMVs also was confirmed. Moreover, Quantifications of blood cells (white blood cells and platelets) and specific enzymes (aspartate aminotransferase and creatine kinase isoenzymes) showed that Dox-CBMVs had lower side effects compared with free Dox. Conclusions The data show that the CBMV-entrapped Doxorubicin has the antitumor efficacy with lower side effects. This study provides evidence supporting the possibility of isolated cell-bound membrane vesicles as a novel drug nanocarrier.

Dynamic AFM detection of the oxidation-induced changes in size, stiffness, and stickiness of low-density lipoprotein

Abstract Background Low-density lipoprotein (LDL) is an important plasma lipoprotein transporting lipids to peripheral tissues/cells. The oxidation of LDL plays critical roles in atherogenesis and its oxidized form (oxLDL) is an important risk factor of atherosclerosis. The biomechanical properties of LDL/oxLDL are closely correlated with the disease. To date, however, the oxidation-induced changes in size and biomechanical properties (stiffness and stickiness) of LDL particles are less investigated. Methods In this study, copper-induced LDL oxidation was confirmed by detecting electrophoretic mobility, malondialdehyde production, and conjugated diene formation. Then, the topographical and biomechanical mappings of LDL particles before/after and during oxidation were performed by using atomic force microscopy (AFM) and the size and biomechanical forces of particles were measured and quantitatively analyzed. Results Oxidation induced a significant decrease in size and stiffness (Young’s modulus) but a significant increase in stickiness (adhesion force) of LDL particles. The smaller, softer, and stickier characteristics of oxidized LDL (oxLDL) partially explains its pro-atherosclerotic role. Conclusions The data implies that LDL oxidation probably aggravates atherogenesis by changing the size and biomechanical properties of LDL particles. The data may provide important information for a better understanding of LDL/oxLDL and atherosclerosis.

Nanomolar detection of adenosine triphosphate (ATP) using a nanostructured fluorescent chemosensing ensemble

A simple nanostructured chemosensing ensemble for fluorescent turn-on sensing of ATP in aqueous solutions and inside living cells using the indicator displacement assay (IDA) method results in a very low detection limit of 6.8 nM.

Highly efficient crystal red fluorescent 1,2-squaraine dyes with excellent biocompatibility and bioimaging

Dynamic single-vesicle tracking of cell-bound membrane vesicles on resting, activated, and cytoskeleton-disrupted cells

Circulating Complement C3-Alpha Chain Levels Predict Survival of Septic Shock Patients

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Exogenous GM3 ganglioside inhibits atherosclerosis via multiple steps: A potential atheroprotective drug

Image and data processing algorithms for identifying cell-bound membrane vesicle trajectories and movement information

An amphiphilic pyrene-based probe for multiple channel sensing of mercury ions

Comparative investigation on the sizes and scavenger receptor binding of human native and modified lipoprotein particles with atomic force microscopy

Enhanced Antiatherosclerotic Efficacy of Statin-Loaded Reconstituted High-Density Lipoprotein via Ganglioside GM1 Modification

Multifunctional Fluorescent Nanoprobe for Sequential Detections of Hg²⁺ Ions and Biothiols in Live Cells

In situ AFM imaging of apolipoprotein A-I directly derived from plasma HDL

Phosphatidylethanolamine Deficiency Impairs<i>Escherichia coli</i>Adhesion by Downregulating Lipopolysaccharide Synthesis, Which is Reversible by High Galactose/Lactose Cultivation

Effects of MβCD on Lipoxygenase-Induced LDL Oxidation

A self-assembled amphiphilic imidazolium-based ATP probe

An amphiphilic imidazolium-based self-assembled probe forms aggregates and selectively recognizes aqueous ATP among other bioactive anions with a significant enhancement in fluorescence emission.

Identification and characterization of cell-bound membrane vesicles

Double-stranded DNA-scaffolded fluorescent probes for fluorescence imaging of cell-surface molecules

Double-stranded DNA-scaffolded fluorescent probes were developed for fluorescence imaging of molecules on cell surfaces.

Methyl-β-Cyclodextrin Impairs the Monocyte-Adhering Ability of Endothelial Cells by Down-Regulating Adhesion Molecules and Caveolae and Reorganizing the Actin Cytoskeleton

Effects of cyclodextrins on the structure of LDL and its susceptibility to copper-induced oxidation

A Pyrene-functionalized Polynorbornene for Ratiometric Fluorescence Sensing of Pyrophosphate

Imaging and force measurement of LDL and HDL by AFM in air and liquid

AFM visualization of cortical filaments/network under cell-bound membrane vesicles

A novel polynorbornene-based chemosensor for the fluorescence sensing of Zn²⁺ and Cd²⁺ and subsequent detection of pyrophosphate in aqueous solutions

A novel 8-hydroxyquinoline anchored polynorbornene for the fluorescence sensing of Zn2+/Cd2+ and subsequent detection of pyrophosphate in aqueous solutions and its biological application is introduced.

Effects of membrane lipid composition and antibacterial drugs on the rigidity of<i>Escherichia coli</i>: Different contributions of various bacterial substructures

Multiple drug-loaded electrospun PLGA/gelatin composite nanofibers encapsulated with mesoporous ZnO nanospheres for potential postsurgical cancer treatment

The novel PLGA/GE fibers encapsulated with DOX@mZnO were successfully fabricated, which enabled the delivery of two drugs with distinct rates.

Fixation-induced cell blebbing on spread cells inversely correlates with phosphatidylinositol 4,5-bisphosphate level in the plasma membrane

An AFM-based pit-measuring method for indirect measurements of cell-surface membrane vesicles

AFM of the Ultrastructural and Mechanical Properties of Lipid-Raft-Disrupted and/or Cold-Treated Endothelial Cells

Novel controlled drug delivery system for multiple drugs based on electrospun nanofibers containing nanomicelles

Effects of long-term serial cell passaging on cell spreading, migration, and cell-surface ultrastructures of cultured vascular endothelial cells

Effects of exogenous ganglioside GM1 on different stages of cell spreading studied by directly quantifying spreading rate

Determination of the lowest concentrations of aldehyde fixatives for completely fixing various cellular structures by real-time imaging and quantification

BMP2 promotes migration and invasion of breast cancer cells via cytoskeletal reorganization and adhesion decrease: an AFM investigation

AFM Investigation on Ox-LDL-Induced Changes in Cell Spreading and Cell-Surface Adhesion Property of Endothelial Cells

Elucidation and Identification of Double-Tip Effects in Atomic Force Microscopy Studies of Biological Structures

Sonodynamic effects of hematoporphyrin monomethyl ether on CNE-2 cells detected by atomic force microscopy

Liposome impaired the adhesion and spreading of HEK293 cells: an AFM study

AFM force measurements of the gp120–sCD4 and gp120 or CD4 antigen–antibody interactions

Cell-surface ultrastructural changes during the in vitro neuron-like differentiation of rat bone marrow-derived mesenchymal stem cells

Detection of erythrocytes influenced by aging and type 2 diabetes using atomic force microscope

Photoinactivation effects of hematoporphyrin monomethyl ether on Gram-positive and -negative bacteria detected by atomic force microscopy

WGA-QD probe-based AFM detects WGA-binding sites on cell surface and WGA-induced rigidity alternation

Cold Induces Micro- and Nano-Scale Reorganization of Lipid Raft Markers at Mounds of T-Cell Membrane Fluctuations

QD as a bifunctional cell-surface marker for both fluorescence and atomic force microscopy

Time-dependent surface adhesive force and morphology of RBC measured by AFM

Fluorescence-topographic NSOM directly visualizes peak-valley polarities of GM1/GM3 rafts in cell membrane fluctuations

The analysis of morphological distortion during AFM study of cells

NSOM/QD-based nanoscale immunofluorescence imaging of antigen-specific T-cell receptor responses during an in vivo clonal Vγ2Vδ2 T-cell expansion

AbstractNanoscale imaging of an in vivo antigen-specific T-cell immune response has not been reported. Here, the combined near-field scanning optical microscopy– and fluorescent quantum dot–based nanotechnology was used to perform immunofluorescence imaging of antigen-specific T-cell receptor (TCR) response in an in vivo model of clonal T-cell expansion. The near-field scanning optical microscopy/quantum dot system provided a best-optical-resolution (<50 nm) nano-scale imaging of Vγ2Vδ2 TCR on the membrane of nonstimulated Vγ2Vδ2 T cells. Before Ag-induced clonal expansion, these nonstimulating Vγ2Vδ2 TCRs appeared to be distributed differently from their αβ TCR counterparts on the cell surface. Surprisingly, Vγ2Vδ2 TCR nanoclusters not only were formed but also sustained on the membrane during an in vivo clonal expansion of Vγ2Vδ2 T cells after phosphoantigen treatment or phosphoantigen plus mycobacterial infection. The TCR nanoclusters could array to form nanodomains or microdomains on the membrane of clonally expanded Vγ2Vδ2 T cells. Interestingly, expanded Vγ2Vδ2 T cells bearing TCR nanoclusters or nanodomains were able to rerecognize phosphoantigen and to exert better effector function. These studies provided nanoscale insight into the in vivo T-cell immune response.

Atomic force microscopic investigation on the potential early intermediate stages of fibrillogenesis of fibronectin within fibrils

Chromosome imaging by atomic force microscopy: influencing factors and comparative evaluation

Atomic Force Microscopic Examination of Chromosomes Treated with Trypsin or Ethidium Bromide

Atomic force microscope tracking observation of Chinese hamster ovary cell mitosis

Membrane deformation of unfixed erythrocytes in air with time lapse investigated by tapping mode atomic force microscopy

Research on double-probe, double- and triple-tip effects during atomic force microscopy scanning

Atomic force microscopy imaging and 3-D reconstructions of serial thin sections of a single cell and its interior structures

Atomic force bio-analytics of polymerization and aggregation of phycoerythrin-conjugated immunoglobulin G molecules

Effect ofBungarus multicinctus crude venom and its components on tumor cells

DISEASED RED BLOOD CELLS STUDIED BY ATOMIC FORCE MICROSCOPY

In recent years, many mammalian cells, especially erythrocytes because of simpleness of their membrane surfaces, were widely studied by atomic force microscopy. In our study, diseased erythrocytes were taken from patients of lung cancer, myelodisplastic syndrome (MDS), and so on. We obtained many clear topographical images of numerous erythrocytes, single erythrocyte, and ultramicrostructure of erythrocyte membrane surfaces from normal persons and patients. By studying the red cells of lung cancer patients, we found that many erythrocytes of lung cancer patient have changed into echinocytes. One erythrocyte has 10–20 short projections, most of which, with a mean width of 589.0 nm and a length of 646.7 nm, are on the edge of cell. The projections in the center of echinocytes are lodged and embedded, but in conventional model of echinocytes, the projections in the center stretch outside cell membrane, so a novel model of erythrocytes was designed in our paper. After observation of microstructure of MDS patient's erythrocyte membrane surface, we found that many apertures with different diameters of tens to hundreds nanometers appeared on the surface of cell membrane. It can be concluded that AFM may be widely applied in clinic pathological inspection.