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1.
Anal Chem ; 93(5): 2871-2878, 2021 02 09.
Article in English | MEDLINE | ID: mdl-33455155

ABSTRACT

Here, we report three-dimensional (3-D) visualization of dendrimer-encapsulated Pt nanoparticles (Pt DENs) by using 3-D electron tomography to reveal intricate structural characteristics of their whole organic-inorganic hybrid nanostructure. We reconstructed the 3-D spatial volume of Pt DENs by back-projecting a tilt series of two-dimensional (2-D) projections of Pt nanoparticles encapsulated inside dendrimers negatively stained with uranyl acetate. The direct 3-D visualization of Pt DENs elucidated their encapsulation characteristics with the spatial imaging of Pt nanoparticles embraced inside dendrimers in three dimensions. The encapsulation characteristics of Pt DENs were further verified with selective electrochemical poisoning experiments. In addition, quantitative 3-D structural characterization of Pt DENs provided more accurate and precise size distributions of nanoparticles than those obtained from conventional 2-D transmission electron microscopy analysis relying only on a 3-D structure projected on a 2-D plane.

2.
Langmuir ; 34(25): 7436-7444, 2018 06 26.
Article in English | MEDLINE | ID: mdl-29856918

ABSTRACT

In this study, we report the controllable synthesis of dendrimer-encapsulated Pt nanoparticles (Pt DENs) utilizing repetitively coupled chemical reduction and galvanic exchange reactions. The synthesis strategy allows the expansion of the applicable number of Pt atoms encapsulated inside dendrimers to more than 1000 without being limited by the fixed number of complexation sites for Pt2+ precursor ions in the dendrimers. The synthesis of Pt DENs is achieved in a short period of time (i.e., ∼10 min) simply by the coaddition of appropriate amounts of Cu2+ and Pt2+ precursors into aqueous dendrimer solution and subsequent addition of reducing agents such as BH4-, resulting in fast and selective complexation of Cu2+ with the dendrimers and subsequent chemical reduction of the complexed Cu2+ while uncomplexed Pt2+ precursors remain oxidized. Interestingly, the chemical reduction of Cu2+, leading to the formation of Cu nanoparticles encapsulated inside the dendrimers, is coupled with the galvanic exchange of the Cu nanoparticles with the nearby Pt2+. This coupling repetitively proceeds until all of the added Pt2+ ions form into Pt nanoparticles encapsulated inside the dendrimers. In contrast to the conventional method utilizing direct chemical reduction, this repetitively coupled chemical reduction and galvanic exchange enables a substantial increase in the applicable number of Pt atoms up to 1320 in Pt DENs while maintaining the unique features of DENs.

3.
Anal Bioanal Chem ; 408(25): 7165-72, 2016 Oct.
Article in English | MEDLINE | ID: mdl-27325466

ABSTRACT

Here, we report highly enhanced electrochemiluminescence (ECL) of luminol in the presence of H2O2 on indium tin oxides (ITOs) modified with both of dendrimer-encapsulated Pt nanoparticles (Pt DENs) and chemically converted graphenes (CCGs). The ITO electrodes were electrochemically modified with size-monodisperse Pt DENs via electrooxidative grafting of the terminal amines of the dendrimers encapsulating Pt nanoparticles. The Pt DEN-modified ITOs were then decorated with CCG sheets via electrostatic attachments of graphene oxides (GOs) and subsequent chemical reduction of the GOs to the CCGs. The resulting CCG-Pt DEN/ITO electrodes exhibited highly catalyzed electrochemical oxidation of luminol/H2O2, leading to significantly enhanced ECL of the luminol/H2O2 system, i.e., ∼15-fold enhancement, compared to ECL emission from bare ITOs even at lower applied potentials, which allowed sensitive ECL-based analysis of H2O2 using the CCG-Pt DEN/ITOs. Graphical abstract We report the highly enhanced electrochemiluminescence of the luminol/H2O2 system on the indium tin oxide electrodes modified with both of Pt nanoparticles and chemically converted graphenes using amine-terminated dendrimers.

4.
Am J Transl Res ; 5(1): 69-79, 2013.
Article in English | MEDLINE | ID: mdl-23390567

ABSTRACT

BACKGROUND: Cell-cell communication occurs via a variety of mechanisms, including long distances (hormonal), short distances (paracrine and synaptic) or direct coupling via gap junctions, antigen presentation, or ligand-receptor interactions. We evaluated the possibility of neuro-hormonal independent, non-diffusible, physically disconnected pathways for cell-cell communication using dorsal root ganglion (DRG) neurons. METHODS: We assessed intracellular calcium ([Ca(2+)]) in primary culture DRG neurons that express ATP-sensitive P2X3, capsaicinsensitive TRPV1 receptors modulated by estradiol. Physically disconnected (dish-in-dish system; inner chamber enclosed) mouse DRG were cultured for 12 hours near: a) media alone (control 1), b) mouse DRG (control 2), c) human neuroblastoma SHSY-5Y cells (cancer intervention), or d) mouse DRG treated with KCl (apoptosis intervention). RESULTS: Chemosensitive receptors [Ca(2+)](i) signaling did not differ between control 1 and 2. ATP (10 µM) and capsaicin (100nM) increased [Ca(2+)](i) transients to 425.86 + 49.5 nM, and 399.21 ± 44.5 nM, respectively. 17ß-estradiol (100 nM) exposure reduced ATP (171.17 ± 48.9 nM) and capsaicin (175.01±34.8 nM) [Ca(2+)](i) transients. The presence of cancer cells reduced ATP- and capsaicin-induced [Ca(2+)](i) by >50% (p<0.05) and abolished the 17ß-estradiol effect. By contrast, apoptotic DRG cells increased initial ATP-induced [Ca(2+)](i), flux four fold and abolished subsequent [Ca(2+)](i), responses to ATP stimulation (p<0.001). Capsaicin (100nM) induced [Ca(2+)](i) responses were totally abolished. CONCLUSION: The local presence of apoptotic DRG or human neuroblastoma cells induced differing abnormal ATP and capsaicin-mediated [Ca(2+)](i) fluxes in normal DRG. These findings support physically disconnected, non-diffusible cell-to-cell signaling. Further studies are needed to delineate the mechanism(s) of and model(s) of communication.

5.
Neuroreport ; 23(9): 530-4, 2012 Jun 20.
Article in English | MEDLINE | ID: mdl-22581043

ABSTRACT

In women, pain symptoms and nociceptive thresholds vary with the reproductive cycle, suggesting the role of estrogen receptors (ERs) in modulating nociception. Our previous data strongly suggest an interaction between ERs and ATP-induced purinergic (P2X3) as well as ERs and capsaicin-induced vanilloid (TRPV1) receptors at the level of dorsal root ganglion (DRG) neurons. In this study, we investigated the expression of P2X3 and TRPV1 receptors by western blotting and immunohistochemistry in lumbosacral DRGs from wild type, ERα, and ERß knockout mice. We found a significant decrease for both P2X3 and TRPV1 in ERαKO and ERßKO. This phenomenon was visualized in L1, L2, L4, and L6 levels for P2X3 receptors and in L1, L2, and S2 levels for TRPV1 receptors. This tan interaction between P2X3/TRPV1 and ERs expression in sensory neurons may represent a novel mechanism that can explain the sex differences in nociception observed in clinical practice. The DRG is an important site of visceral afferent convergence and cross-sensitization and a potential target for designing new anti-nociceptive therapies.


Subject(s)
Estrogen Receptor alpha/genetics , Estrogen Receptor beta/genetics , Ganglia, Spinal/metabolism , Nociception/physiology , Receptors, Purinergic P2X3/metabolism , Sensory Receptor Cells/metabolism , TRPV Cation Channels/metabolism , Animals , Female , Immunohistochemistry , Mice , Mice, Knockout , Visceral Afferents/metabolism
6.
Biotechnol Bioeng ; 101(2): 357-68, 2008 Oct 01.
Article in English | MEDLINE | ID: mdl-18435481

ABSTRACT

Development of methods to engineer gamma-retroviral vectors capable of transducing target cells in a cell-specific manner could impact the future of the clinical application of gene therapy as well as the understanding of the biology of transfer gene vectors. Two molecular events are critical for controlling the entry of gamma-retroviral vectors to target cells: binding to cell-surface receptors and the subsequent fusion of viral vector membrane and cellular membrane. In this report, we evaluated a method to incorporate a membrane-bound antibody and a fusogenic molecule to provide binding and fusion functions respectively, into gamma-retroviral vectors for targeted gene delivery. An anti-CD20 antibody and a fusogenic protein derived from Sindbis virus glycoprotein could be efficiently co-displayed on the surface of viral vectors. Vectors bearing anti-CD20 antibody conferred their binding specificity to cells expressing CD20. Enhanced in vitro transduction towards CD20-expressing cells was observed for gamma-retroviral vectors displaying both an antibody and a fusogen. We found that the biological activity of the fusogen played an important role on the efficiency of such a targeting strategy and were able to engineer several mutant forms of the fusogen exhibiting elevated fusion function to improve the overall efficiency of targeted transduction. We devised an animal model to show that subcutaneous injection of such engineered vectors to the areas xenografted with target cells could achieve targeted gene delivery in vivo. Taken together, we demonstrated as proof-of-principle a flexible and modular two-molecule strategy for engineering targeting gamma-retroviral vectors.


Subject(s)
Antibodies/genetics , Gammaretrovirus/genetics , Gene Targeting , Gene Transfer Techniques , Genetic Vectors , Recombinant Fusion Proteins/genetics , Animals , Antibodies/immunology , Antigens, CD20/genetics , Antigens, CD20/immunology , Cell Line , Female , Gammaretrovirus/immunology , Genetic Therapy , Humans , Mice , Recombinant Fusion Proteins/immunology , Sindbis Virus/genetics , Transduction, Genetic , Viral Envelope Proteins/genetics , Viral Envelope Proteins/immunology
7.
Nat Biotechnol ; 26(3): 326-34, 2008 Mar.
Article in English | MEDLINE | ID: mdl-18297056

ABSTRACT

We report a method of inducing antigen production in dendritic cells by in vivo targeting with lentiviral vectors that specifically bind to the dendritic cell-surface protein DC-SIGN. To target dendritic cells, we enveloped the lentivector with a viral glycoprotein from Sindbis virus engineered to be DC-SIGN-specific. In vitro, this lentivector specifically transduced dendritic cells and induced dendritic cell maturation. A high frequency (up to 12%) of ovalbumin (OVA)-specific CD8(+) T cells and a significant antibody response were observed 2 weeks after injection of a targeted lentiviral vector encoding an OVA transgene into naive mice. This approach also protected against the growth of OVA-expressing E.G7 tumors and induced regression of established tumors. Thus, lentiviral vectors targeting dendritic cells provide a simple method of producing effective immunity and may provide an alternative route for immunization with protein antigens.


Subject(s)
Cancer Vaccines/immunology , Dendritic Cells/immunology , Dendritic Cells/metabolism , Genetic Vectors/genetics , Lentivirus/genetics , Animals , B-Lymphocytes/cytology , B-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/immunology , Cell Adhesion Molecules/metabolism , Cell Line , Dendritic Cells/cytology , Female , Gene Expression , Humans , Lectins, C-Type/metabolism , Mice , Mice, Inbred C57BL , Mice, Transgenic , Neoplasms, Experimental/immunology , Neoplasms, Experimental/therapy , Ovalbumin/genetics , Ovalbumin/immunology , Receptors, Cell Surface/metabolism , Sensitivity and Specificity , Sindbis Virus/genetics , Sindbis Virus/metabolism , Transduction, Genetic
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