Longitudinal monitoring of cells is required in order to understand the role of delivered stem cells in therapeutic neovascularization. However, there is not an imaging technique that is capable of quantitative, longitudinal assessment of stem cell behaviors with high spatial resolution and sufficient penetration depth. In this study, in vivo and in vitro experiments were performed to demonstrate the efficacy of ultrasound-guided photoacoustic (US/PA) imaging to monitor mesenchymal stem cells (MSCs) labeled with gold nanotracers (Au NTs). The Au NT labeled MSCs, injected intramuscularly in the lower limb of the Lewis rat, were detected and spatially resolved. Furthermore, our quantitative in vitro cell studies indicate that US/PA imaging is capable of high detection sensitivity (1×104 cells/mL) of the Au NT labeled MSCs. Finally, Au NT labeled MSCs captured in the PEGylated fibrin gel system were imaged in vivo, as well as in vitro, over a one week time period, suggesting that longitudinal cell tracking using US/PA imaging is possible. Overall, Au NT labeling of MSCs and US/PA imaging can be an alternative approach in stem cell imaging capable of noninvasive, sensitive, quantitative, longitudinal assessment of stem cell behaviors with high spatial and temporal resolutions at sufficient depths.

Stem cell biology has become an important field in regenerative medicine and tissue engineering therapy since the discovery and characterization of mesenchymal stem cells. Stem cell populations have also been isolated from human dental tissues, including dental pulp stem cells, stem cells from human exfoliated deciduous teeth, stem cells from apical papilla, dental follicle progenitor cells, and periodontal ligament stem cells. Dental stem cells are relatively easily obtainable and exhibit high plasticity and multipotential capabilities. The dental stem cells represent a gold standard for neural-crest-derived bone reconstruction in humans and can be used for the repair of body defects in low-risk autologous therapeutic strategies. The bioengineering technologies developed for tooth regeneration will make substantial contributions to understand the developmental process and will encourage future organ replacement by regenerative therapies in a wide variety of organs such as the liver, kidney, and heart. The concept of developing tooth banking and preservation of dental stem cells is promising. Further research in the area has the potential to herald a new dawn in effective treatment of notoriously difficult diseases which could prove highly beneficial to mankind in the long run.

Background: Stem cells can differentiate into multiple cell types, and therefore can be used for cellular therapies, including tissue repair. However, the participation of stem cells in tissue repair and neovascularization is not well understood. Therefore, implementing a noninvasive, long-term imaging technique to track stem cells in vivo is needed to obtain a better understanding of the wound healing response. Generally, we are interested in developing an imaging approach to track mesenchymal stem cells (MSCs) in vivo after delivery via a polyethylene glycol modified fibrin matrix (PEGylated fibrin matrix) using MSCs loaded with gold nanoparticles as nanotracers. The objective of the current study was to assess the effects of loading MSCs with gold nanoparticles on cellular function. Methods: In this study, we utilized various gold nanoparticle formulations by varying size and surface coatings and assessed the efficiency of cell labeling using darkfield microscopy. We hypothesized that loading cells with gold nanotracers would not significantly alter cell function due to the inert and biocompatible characteristics of gold. The effect of nanoparticle loading on cell viability and cytotoxicity was analyzed using a LIVE/DEAD stain and an MTT assay. The ability of MSCs to differentiate into adipocytes and osteocytes after nanoparticle loading was also examined. In addition, nanoparticle loading and retention over time was assessed using inductively coupled plasma mass spectrometry (ICP-MS). Conclusion: Our results demonstrate that loading MSCs with gold nanotracers does not alter cell function and, based on the ICP-MS results, long-term imaging and tracking of MSCs is feasible. These findings strengthen the possibility of imaging MSCs in vivo, such as with optical or photoacoustic imaging, to understand better the participation and role of MSCs in neovascularization.

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Spectroscopic photoacoustic imaging has the potential to become a powerful tool that can estimate distributions of optically absorbing chromophores in the body. We have developed an algorithm to select imaging wavelengths for spectroscopic photoacoustics given the spectra of expected chromophores. The algorithm uses the smallest singular value of a matrix constructed from the absorption spectra as a criterion to remove extraneous wavelengths. The method performed significantly better than an approach where evenly spaced wavelengths were used in the presence of noise and wavelength-dependent attenuation of light in tissue. Finally, the algorithm was applied to photoacoustic imaging of a phantom containing indocyanine green dye and silica-coated gold nanorods, demonstrating significant improvements in the ability to estimate relative contrast agent concentrations compared to the case where evenly spaced wavelengths were chosen. In summary, our work provides a versatile framework to select optical wavelengths and evaluate sets of absorbers for spectroscopic photoacoustic imaging.

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Photoacoustic imaging, using targeted plasmonic metallic nanoparticles, is a promising noninvasive molecular imaging method. Analysis of the photoacoustic signal generated by plasmonic metallic nanoparticles is complex because of the dependence upon physical properties of both the nanoparticle and the surrounding environment. We studied the effect of the aggregation of gold nanoparticles on the photoacoustic signal amplitude. We found that the photoacoustic signal from aggregated silica-coated gold nanoparticles is greatly enhanced in comparison to disperse silica-coated gold nanoparticles. Because cellular uptake and endocytosis of nanoparticles results in their aggregation, these results have important implications for the application of plasmonic metallic nanoparticles towards quantitative molecular imaging.