[18F]FLT - A new stem cell label for in vivo tracking with positron emission tomography

Objectives Because of their extended differentiation capacity, mesenchymal stem cells (MSCs) have gained great interest in the field of regenerative medicine. For the development of therapeutic strategies, more knowledge on their in vivo fate is required. So far, MSCs were mainly labeled for that purpose with iron particles for MR imaging or with [18F]FDG for PET imaging. In case of brain applications, however, [18F]FDG is critical as a MSC label. This is due to the risk of image contrast lost in case of instability of the labeling. 3’-deoxy-3’-18F-fluorothymidine ([18F]FLT) might be an alternative. Thus, this project was initiated to, prior to testing [18F]FLT-labeled MSCs in a large animal stroke model, investigating the cellular uptake, vitality and binding of [18F]FLT by MSCs in vitro.

Methods Adult ovine mononuclear MSCs were used to investigate [18F]FLT uptake kinetics and cell toxicity over time (0-120 min) in relation to tracer dose (0.1-100 MBq). 200,000 cells were seeded over time in 2 ml 37°C PBS per well in a 12-well plate. Cell viability and the amount of tracer uptake after labeling were evaluated using Trypan Blue reagent and Gammacounter, respectively.

Results [18F]FLT uptake by the MSCs increased up to 60min incubation time to 0.34%ID/10,000 cells, followed by a plateau of the labeling signal up to 120min incubation time. 60 min after [18F]FLT labeling, approximately 70% of the cells had good cellular vitality. Increasing the concentration of [18F]FLT decreased cell vitality from 85% at 0.1 MBq to 59% at 100 MBq, which was accompanied by higher tracer elution from the cells. After 60min labeling, ~85% of the initially incorporated [18F]FLT was retained inside the MSCs, while this was the case for only 47% after 3hrs.

Conclusions In conclusion, [18F]FLT labeling of sheep MSCs is feasible. 60min incubation time with the tracer does not perturb the biologic and functional properties of adult sheep MSCs in vitro. This labeling method and optimization strategy has led to pilot studies investigating the biodistribution of [18F]FLT-labeled MSCs in mice using small-animal PET/MRI. Future studies will also determine whether [18F]FLT-labeled MSCs can be tracked over time in a large animal model to assess the therapeutic potential of MSCs after stroke.