Interaction of Nanogold with VEGF and VEGFR Labeled by Quantum Dots in Near-field Scanning Optical Microscopy
|Keywords||Near-field optical microscope Quantum dots Gold Nanoparticles Vascular endothelial growth factor|
The purpose of using gold nanoparticles can prevent the heparin-binding sites of vascular endothelial growth factor (Vascular endothelial growth factor, VEGF) 165 to its receptor VEGFR2 combination of characteristics based on near-field optical microscope combined with quantum dot technology nano gold blocking VEGF165 their body the VEGFR2 combination of molecular mechanism of action. The method MTT assay whether the gold nanoparticles inhibit containing heparin binding sites of VEGF 165-induced vascular endothelial cell proliferation, without VEGF121 as a control for the heparin binding site; 2. Atomic force microscope (atomic force microscope, AFM) observation gold nanoparticles before and after the role of VEGF in human umbilical vein endothelial cells (Human YANBIAN UNIVERSITY. Protective effects of the compound Danshen vein endothelial cells, HUVECs), surface ultrastructural changes; 3. VEGFR2 and VEGF165 molecules on the endothelial cell surface markers using quantum dots (quantum dots, QDs), before joining the different concentrations of gold nanoparticles before and after the application of high-resolution confocal microscopy and near-field optical microscope probe intermolecular interactions; 4. serum-free cultured human umbilical vein endothelial cells (HUVECs), added of VEGF165, then adding different concentrations of nano gold, for 5 min, the cells within the vascular endothelial growth factor receptor -2 downstream signaling protein phosphorylation of PLC-yl level detected by Western blot. 1. Vascular endothelial cell proliferation inhibition experiments: gold nanoparticles significantly inhibited VEGF165-induced endothelial cell proliferation, while no significant inhibitory effect on the lack of a heparin-binding sites VEGF121 detected; 2.AFM of nanometer gold and of VEGF165, VEGF121 before and after vascular endothelial cell ultrastructural changes of VEGF165-induced endothelial cell surface the pore or particles of many about the size of a few hundred nanometers, the membrane edge spread extends. Addition of VEGF165, gold nanoparticles, the cell surface pores or particles is greatly reduced. VEGF121-induced endothelial cell surface ultrastructure gold nanoparticles before and after adding no significant change; 3. Confocal microscopy and near-field optical microscope to detect the gold nanoparticles blocking the combination of VEGF165 to its cell surface receptor VEGFR2. 4.VEGF165 constant concentration (10μg / L), with the increase in the concentration of gold nanoparticle solution (125,250,500 nmol / L), gold nanoparticles inhibit phosphorylation of PLC-γ1 is more and more obvious, and the lack of heparin binding point the VEGF121 no significant inhibition. Conclusions gold nanoparticles inhibit VEGF165 vascular endothelial cell proliferation: 2 gold nanoparticles can prevent VEGF165 to its receptor VEGFR2 combination of heparin-binding sites, preventing its downstream cell signaling, thereby inhibiting VEGF165 induced vascular endothelial cell proliferation: 3. combined with quantum dots labeling technique, high-resolution near-field optical microscope is a powerful tool to study the interaction between the biological single molecule.