Abstract
In vivo cell tracking of therapeutic, tumor, and endothelial cells is an emerging field and a promising technique for imaging cardiovascular disease and cancer development. Site-specific labeling of endothelial cells with the MRI contrast agent superparamagnetic iron oxide (SPIO) in the absence of toxic agents is challenging. Therefore, the aim of this in vitro study was to find optimal parameters for efficient and safe SPIO-labeling of endothelial cells using ultrasound-activated CD31-targeted microbubbles for future MRI tracking. Ultrasound at a frequency of 1 MHz (10,000 cycles, repetition rate of 20 Hz) was used for varying applied peak negative pressures (10–160 kPa, i.e. low mechanical index (MI) of 0.01–0.16), treatment durations (0–30 s), time of SPIO addition (-5 min– 15 min with respect to the start of the ultrasound), and incubation time after SPIO addition (5 min– 3 h). Iron specific Prussian Blue staining in combination with calcein-AM based cell viability assays were applied to define the most efficient and safe conditions for SPIO-labeling. Optimal SPIO labeling was observed when the ultrasound parameters were 40 kPa peak negative pressure (MI 0.04), applied for 30 s just before SPIO addition (0 min). Compared to the control, this resulted in an approximate 12 times increase of SPIO uptake in endothelial cells in vitro with 85% cell viability. Therefore, ultrasound-activated targeted ultrasound contrast agents show great potential for effective and safe labeling of endothelial cells with SPIO.
Highlights
In vivo cell tracking is a very promising technique to visualize cells of interest inside the body
The total ultrasound exposure time was a key factor for superparamagnetic iron oxide nanoparticles (SPIO) uptake efficacy
SPIO uptake by ultrasound-activated targeted microbubbles (tMB) in vivo is expected as we recently reported that tMB bound to αvβ3 can stimulate endothelial cell drug uptake in vivo [50]
Summary
In vivo cell tracking is a very promising technique to visualize cells of interest inside the body It allows tracking of motile therapeutic cells like immune cells, stem cells, and endothelial progenitor cells to sites of inflammation, cancer, or ischemia [1,2,3,4,5]. The T2 and T2Ã-shortening MRI contrast agent superparamagnetic iron oxide nanoparticles (SPIO) of 80–180 nm in size [17] are often used [18, 19] They are relatively safe compounds [19,20,21,22], but most of in vitro cell labeling techniques for SPIO are not applicable in vivo, because of the high toxicity and broad systemic effects of transfection agents [23]. The tMB have a ligand added in their coating by which the tMB can adhere to disease-specific cell membrane biomarkers [31, 32]
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