Enhanced positive-contrast visualization of paramagnetic contrast agents using phase images

Paramagnetic contrast agents (CAs) are widely used for a multitude of emerging cellular-molecular applications (1). Commonly used superparamagnetic iron-oxide (SPIO) particles, with iron-oxide core diameters ranging between 5 nm and 1 μm, create microscopic static magnetic field perturbations that extend 10-100 times the particle's diameter, thereby reducing the effective T2 and T2* of nearby water protons. The resulting T2- and T2*-weighted images often exhibit regions of hypointensity in magnitude images where agent has accumulated. Clinical-grade SPIO agents such as Resovist®, Feridex®, and Combidex® are used to enhance various pathologies. SPIO particles have been used to detect molecular epitopes by conjugating their surfaces to a variety of target entities, such as peptides or antibodies (1,2). Intracellular labeling with SPIO, either ex vivo or in situ, is a rapidly evolving approach for preclinical cell trafficking and inflammation studies (3). Clinical translation of cell tracking has been demonstrated in pilot studies (4). In addition, paramagnetic CAs also encompass intracellular labeling approaches that image gene expression by utilizing genetically encoded metalloproteins (1,5-9). In these approaches, metalloproteins, particularly from the ferritin family, are selectively overexpressed in specific host tissues. These iron storage proteins become paramagnetic by sequestering and aggregating iron from the organism, thereby imparting exogenous MRI contrast to transduced tissues. Across all of these important iron-based paramagnetic CA applications, a common challenge is distinguishing the image intensity dropout generated by the agents from other sources of endogenous image hypointensity. For example, endogenous T2- and T2*-weighted hypointensity may originate from disparate tissue relaxation times, magnetic susceptibility interface gradients, blood vessels, and low proton density in tissues. To address this challenge, previous studies have generated positive contrast images highlighting the presence of SPIOs by changing how images are acquired. Exemplary studies utilize differential imaging before and after agent delivery, specialized weighted contrast methods (10,11), spectrally selective excitation (12,13), gradient dephasing (14), quantum coherence imaging (15), and ultrashort echo time image subtraction (16). These techniques often complement magnitude images but may also carry limitations. For example, several methods require foreknowledge of the magnitude of the field disturbances caused by the iron oxides, and this information is used to configure critical MRI scan parameters such as excitation (carrier) frequency, bandwidth, and selective radiofrequency pulse shape (12), rephasing gradient amplitude (13) or echo times, and slice thickness (14). Additionally, some of these techniques demand high-end hardware (16) or additional anatomic scans that result in longer acquisition times (12-14). An alternative approach for addressing this common challenge is to highlight SPIO by generating positive-contrast images from MRI phase images (17,18). Because phase image pixel values correlate with local magnetic field, applying high-pass filters to phase images has been shown to accentuate spatial magnetic field variations (i.e., paramagnetic CA deposits). Postprocessing of phase images does not require altering image acquisition procedures, as is required for many of the prior art methods, as discussed above. In one study, a smoothing high-pass filter was applied to phase images of an air-water phantom and in vivo brain tissue (18). The filter used in this study, however, results in images with an effective resolution roughly two times lower than the original image (18). In a second study, a high-pass filter that does not reduce effective resolution was applied to phase images of bovine muscle tissue (17). More recently, the linear component of phase spatial variation was visualized by performing quadratic fits to k-space phase shifts (19). This method uses a variable-size sliding-window approach that avoids potential phase-unwrapping errors but sacrifices image resolution as a result of partial-voluming. These studies (17-19) provide strong visual evidence for enhanced sensitivity to magnetic susceptibility disturbances among tissues when using filtered phase maps. However, these studies either do not quantify sensitivity gains or sacrifice spatial resolution during the image generation process. This study aims to simultaneously preserve image resolution while quantifying sensitivity improvements that result from high-pass filtering phase images. Our resulting phase slope magnitude (PSM) images emphasize spatial magnetic field variations. The PSM image contrast-to-noise ratio (CNR) was calculated in regions surrounding paramagnetic CA distributions and compared against CNR values from conventional magnitude images. Initially, using PSM we analyzed an agarose gel phantom doped with SPIO particles. PSM analysis was also performed on a fixed mouse brain that had been stereotaxically inoculated in one hemisegment of the striatum with a replication-defective adenovirus (AdV). The AdV contained transferrin receptor (TfR-1) transgenes; overexpression of TfR in transduced cells upregulates ferritin, a paramagnetic cellular iron storage protein that produces a weak contrasting effect in T2 and T2*-weighted MRI scans (5,20). Last, PSM analysis was performed using in vivo data acquired in a mouse brain that had been inoculated with AdV containing transgenes for ferritin subunits. These transgenes cause the formation of paramagnetic holoferritin as brain cells sequester endogenous iron from the organism (5). The paramagnetic disturbance from ferritin is weaker compared to SPIO, and PSM images may make the difference between seeing and not seeing a targeted CA. Overall, PSM images exhibit a significant 1- to 8-fold CNR improvement in regions where paramagnetic agent is present compared to conventional magnitude images. A method for thresholding PSM images to eliminate noise from phase measurements is also described. The PSM method does not require extra MRI scans because it relies on phase images that are a by-product of conventional MRI scans; it can be used retrospectively on previously acquired data. PSM image generation requires no prior knowledge about the magnitude of the field disturbance created by arbitrary CA distributions and can be used in conjunction with any T2*-weighted MRI pulse sequence.