Fluorescent Nanodiamonds for Detecting Free-Radical Generation in Real Time during Shear Stress in Human Umbilical Vein Endothelial Cells

Abstract

Free-radical generation is suspected to play a key role in cardiovascular diseases. Another crucial factor is shear stress. Human umbilical vein endothelial cells (HUVECS), which form the lining of blood vessels, require a physiological shear stress to activate many vasoactive factors. These are needed for maintaining vascular cell functions such as nonthrombogenicity, regulation of blood flow, and vascular tone. Additionally, blood clots form at regions of high shear stress within a blood vessel. Here, we use a new method called diamond magnetometry which allows us to measure the dynamics of free-radical generation in real time under shear stress. This quantum sensing technique allows free-radical detection with nanoscale resolution at the single-cell level. We investigate radical formation in HUVECs in a microfluidic environment under different flow conditions typically found in veins and arteries. Here, we looked into free-radical formation before, during, and after flow. We found that the free-radical production varied depending on the flow conditions. To confirm the magnetometry results and to differentiate between radicals, we performed conventional fluorescent reactive oxygen species (ROS) assays specific for superoxide, nitric oxide, and overall ROS.

Single‐Particle Tracking and Trajectory Analysis of Fluorescent Nanodiamonds in Cell‐Free Environment and Live Cells

“…[39,40] Based on previous work under controlled conditions in a chemical environment, we can ascertain that the concentrations we can estimate from T1 measurements are indeed in this expected concentration range. [6,9] This can stem either from the particle arriving at a more radical-rich subcellular location or the cell being stressed and undergoing apoptosisthe bright-field images show that the cell has shrunk during the experiment, and some of the neighboring cells have died as well. Notably, the MSD curves show a larger spread than in experiment 2 (Figure 8E), which might reflect changes in the viscosity of the environment of the FND.…”

“…Diamond magnetometry is a promising technique, applied in materials science, [1,2] spin electronics, [3] quantum sensing, and, lately, for detecting magnetic field fluctuations in chemical [4][5][6] and biological samples, [7] including individual living cells. [8,9] For intracellular measurements, fluorescent diamond nanoparticles (Fluorescent nanodiamonds [FNDs]) can be used. FNDs, internalized by a cell, offer extremely high spatial resolution (on the order of 10-20 nm).…”

Recent Developments of Nanodiamond Quantum Sensors for Biological Applications

“…[ 120 , 121 ] However, the detection of radicals with NDs was not only realized until 2020. [ 13 , 14 , 122 , 123 ] In the work from the group of P. Cigler, [ 13 ] as illustrated in Figure 6 a,b,c , the NDs were coated with a poly(glycerol) shell containing covalently bound stable TEMPO radicals coupled with NV⁻ centers. The TEMPO radicals in the nanosensor are gradually reduced by ascorbates until they are completely removed.…”

Insight into a Fenton-like Reaction Using Nanodiamond Based Relaxometry

“…So far, diamond magnetometry has been used for nanoscale magnetic resonance measurements and successfully applied to measure the magnetic field from nanoparticles [ 10 ], 2D materials, domain walls in magnetic structures [ 11 ], gadolinium ions in solution [ 12 ], spin-labeled molecules, proteins with metallic parts [ 13 ] or even slices of cells [ 14 ]. Recently, the first measurements in living cells have been demonstrated [ 15 , 16 , 17 , 18 , 19 ]. Here, we use a specific form of diamond magnetometry called T1 or relaxometry measurements [ 20 ].…”