Molecular Imaging with Aquaporin-Based Reporter Genes: Quantitative Considerations from Monte Carlo Diffusion Simulations

Abstract

Aquaporins provide a unique approach for imaging genetic activity in deep tissues by increasing the rate of cellular water diffusion, which generates a magnetic resonance contrast. However, distinguishing aquaporin signals from the tissue background is challenging because water diffusion is influenced by structural factors, such as cell size and packing density. Here, we developed a Monte Carlo model to analyze how cell radius and intracellular volume fraction quantitatively affect aquaporin signals. We demonstrated that a differential imaging approach based on subtracting signals at two diffusion times can improve specificity by unambiguously isolating aquaporin signals from the tissue background. We further used Monte Carlo simulations to analyze the connection between diffusivity and the percentage of cells engineered to express aquaporin and established a mapping that accurately determined the volume fraction of aquaporin-expressing cells in mixed populations. The quantitative framework developed in this study will enable a broad range of applications in biomedical synthetic biology, requiring the use of aquaporins to noninvasively monitor the location and function of genetically engineered devices in live animals.

Exploring the potential of water channels for developing MRI reporters and sensors without the need for exogenous contrast agents

“…We transduced HT22 cell lines with the above constructs and used fluorescence-activated cell sorting (FACS) to select stable cell lines based on eGFP fluorescence. Using stimulated echo diffusion-weighted MRI, as previously described in our work [22], we measured the apparent diffusivity of the ensuing cells and quantified their functionality in terms of the fold-change in diffusivity relative to wild-type cells lacking heterologous hAqp1 expression ( Fig. 1a ).…”

“…Expression of hAqp1 facilitates the passive diffusion of water molecules across the plasma membrane. Therefore, by selecting a sufficiently long Δ (on the order of 100 ms [22]) to allow a substantial portion of water molecules to cross the plasma membrane, diffusion-weighted MRI can be used to visualize cells engineered to express hAqp1 based on their increased diffusivity compared to native cells ( Fig. 1a ).…”

A Dual‐Gene Reporter‐Amplifier Architecture for Enhancing the Sensitivity of Molecular MRI by Water Exchange

“…Whole‐cell MRI measurements were performed using cell pellets, as described in our previous studies [20,41,59,60] . Briefly, prior to undergoing MRI, cells were seeded in in 10 cm tissue culture plates such that they reached confluence within approximately 48 h from the time of seeding.…”

“…Whole-cell MRI measurements were performed using cell pellets, as described in our previous studies. [20,41,59,60] Briefly, prior to under-going MRI, cells were seeded in in 10 cm tissue culture plates such that they reached confluence within approximately 48 h from the time of seeding. For cells harboring an inducible Aqp1 construct, doxycycline hyclate (0.01-1 μg/mL) was added to the culture media approximately 24 h after seeding.…”

“…[23,28,[38][39][40] This results in shortening of the T 1 of water molecules within Oatp1b3-expressing cells, thereby enabling their visualization through T 1 -weighted MRI (Figure 1a). Aqp1, on the other hand, is a channel protein that facilitates the free and selective exchange of water molecules across the cell membrane and has been adapted as a reporter gene for visualizing cells using diffusion-weighted MRI [20,41] (Figure 1b).…”