Noninvasive PET Imaging and Tracking of Engineered Human Muscle Precursor Cells for Skeletal Muscle Tissue Engineering

Abstract

Transplantation of human muscle precursor cells (hMPCs) is envisioned for the treatment of various muscle diseases. However, a feasible non-invasive tool to monitor cell survival, migration and integration into the host tissue is still missing. Methods In this study, we designed an adenoviral delivery system to genetically modify hMPCs to express a signaling-deficient form of a human dopamine D2 receptor (hD2R). The gene expression levels of the receptor were evaluated by Reverse Transcriptase Polymerase Chain Reaction (RTPCR) and infection efficiency was visualized by fluorescent microscopy. Viability, proliferation and differentiation capacity of the transduced cells were confirmed and their sustained myogenic phenotype was shown by flow cytometry analysis and fluorescent microscopy. 18F-Fallypride and 18F-FMISO, two well-established PET radioligands, were successfully synthesized and evaluated for their potential to image engineered hMPCs in a mouse model. Furthermore, biodistribution studies and autoradiography were also performed to determine the extent of signal specificity. Results To address the feasibility of the presented approach for tracking of hMPCs in an in vivo model, we first evaluated the safety of the adenoviral gene-delivery, which showed no detrimental effects on the primary human cells. Specific binding of 18F-Fallypride to hD2R_hMPCs was demonstrated in vitro, as well as in vivo, by performing autoradiography, biodistribution and PET experiments, respectively. Furthermore, 18F-FMISO uptake was evaluated at different time-points after cell inoculation in vivo, showing high signal only at the early stages. Finally, histological assessment of the harvested tissues confirmed the sustained survival of the transplanted cells at different time-points with formation of muscle tissue at the site of injection. Conclusion We here propose a signaling-deficient human D2R as a potent reporter for in vivo hMPCs PET tracking by 18F-Fallypride. This approach is a significant step forward towards a potential non-invasive tracking of hMPC_hD2R cells and bioengineered muscle tissues in the clinic.

Injected Human Muscle Precursor Cells Overexpressing PGC-1α Enhance Functional Muscle Regeneration after Trauma

“…Additionally, although imaging reporter genes are available for fluorescence, bioluminescence, and MRI, only radionuclide-based reporter genes are currently investigated for use in patients [ 22 – 27 ]. In our previous work, we developed a feasible method for in vivo tracking of subcutaneously injected hMPCs using [ 18 F]Fallypride, a well-established dopamine 2 receptor (hD2R) PET ligand [ 28 ]. After the successful generation of adenoviruses for the overexpression of a signalling-deficient hD2R in hMPCs, we were able to establish an ex situ model for imaging of bioengineered muscle tissue [ 28 ].…”

“…In our previous work, we developed a feasible method for in vivo tracking of subcutaneously injected hMPCs using [ 18 F]Fallypride, a well-established dopamine 2 receptor (hD2R) PET ligand [ 28 ]. After the successful generation of adenoviruses for the overexpression of a signalling-deficient hD2R in hMPCs, we were able to establish an ex situ model for imaging of bioengineered muscle tissue [ 28 ]. This encouraged us to concentrate our further investigations towards the applicability of these methods in an in situ muscle crush model studying skeletal muscle regeneration, thereby coming closer to a model for studying urinary sphincter muscle restoration.…”

Accomplishments and challenges in stem cell imaging in vivo

“…For example the expression of dopamine D 2 receptor (D2R) which is normally predominantly found in the striata nigra. D2R expression is imaged with PET following the administration of 18 F-labeled fallypride, a dopamine ligand [50]. Gene reporters enable SC imaging at any time and at multiple time points by administering the radiotracer to detect the expressed protein [14,16,51].…”

Cell Tracking in Cancer Immunotherapy

“…A similar approach was used to detect transplanted pancreatic islet cells that express glucagon-like peptide 1 receptor (GLP-1R) by PET imaging after the injection of 64 Cu-DO3A-VS-Cys40-Exendin-4, showing persistent and specific uptake in the mouse pancreas (36). The mutated version of the dopamine receptor, D2R80A, that internalize 18 F-Fallypride, has also been proposed for imaging mesenchymal stem cells (37,38).…”