Research into cell therapies for cardiac regeneration is stymied by an inability to track in vivo cell survival, proliferation, and differentiation. MRI tracking of iron oxide nanoparticle labeled cells following cardiac implantation has been explored, however concerns of tissue retention of particles after cell death and difficulty differentiating between labeled cells and tissue necrosis remain major limitations. Chemical exchange saturation transfer (CEST) - MRI has been used to image cells labeled with paraCEST contrast agents1 or cells expressing CEST active reporter genes2 in stationary organs and tumors but not the heart. We developed a cardiorespiratory gated CEST-encoded steady state cine pulse sequence (cardioCEST), and demonstrated the ability to selectively visualize CEST contrast from Eu-HPDO3A labeled cells following intra-myocardial implantation3. Here we examined whether cardioCEST imaging could differentiate cell survival/proliferation from rejection in a longitudinal mouse model of cardiac cell therapy. Pulse Sequence Design: CEST encoding used a 2s train of Gaussian saturation pulses (720°, B1average = 14μT, bandwidth = 200Hz, duration = 8.8ms, number of pulses = 196, offsets = ±15ppm) prior to a constant repetition time cine gradient echo readout that was cardiorespiratory gated (TR/TE= 7.1/3.1ms, flip angle = 10°). After each saturation 4 averages of one phase-encode step were acquired for each cardiac phase, and dummy pulses maintained steady state magnetization during respiratory motion. Additional parameters were FOV=2.56x2.56cm, Matrix = 256x128, and slice thickness = 1mm. All imaging was performed on a 7T Bruker Clinscan (Bruker, Germany) scanner using a cylindrical volume coil for excitation and a 4-channel phased-array surface coil for reception. Cell and Animal Experiments: C2C12 cells derived from C3H mice were labeled with Eu-HPDO3A via hypotonic swelling4 that yielded 22,355.67 ng(Eu)/mg cell mass as determined by ICPMS. Implantation of ~10⁶ cells into the left ventricular wall was performed using a surgical pop-out technique5. In order to model the survival/proliferation vs. the rejection of implanted cells, labeled C2C12 cells were implanted into syngeneic C3H mice (survival, n=6) or C57B6 mice (rejection, n=7). Separately, saline-labeled C2C12 cells implanted into different C3H (n = 6) or C57B6 (n = 9) mice served as controls. Pairs of cardioCEST images were acquired in 2-3 mid-ventricular slices per mouse at 1, 10 and 20 days following cell implantation. Afterwards hearts were excised and stained using hematoxylin and eosin staining. Registration of cardioCEST image pairs was performed to account for slight movement between acquisitions. Maps of CEST contrast were generated by matrix calculation of MTRasym = (S_-15ppm – S_15ppm)/S_-15ppm*100. Regions of interest in mice receiving Eu-HPDO3A labeled cells were defined in the area of cell implantation (cell graft), and a control area in the septum. In mice receiving saline labeled cells, one region of interest was defined in the area of cell injection (saline graft). MTR_asym values were similar at the sites of implanted Eu-HPDO3A labeled cells in C3H and C57B6 mice (Figures 1, 2) at 1 day after injection. In C3H mice (proliferation model, Figure 1), MTRasym remained higher for 20 days in areas of implanted labeled cells compared to septal regions and saline labeled controls (Figure 2), but decreased compared to day 1. Elevated MTR_asym in C3H mice at day 20 was localized to areas of C2C12 cell proliferation as identified by histology (Figure 1). In C57B6 mice (rejection model), MTR_asym in the area of implanted cells decreased significantly within 10 days of injection. By 20 days post injection, MTR_asym values were similar in areas of implanted cells, control regions in the septum, and in control mice receiving saline labeled cells (Figures 1, 2). Also, MTR_asym was significantly (p < .01) lower in the cell graft in C57B6 mice compared to C3H mice at 10 and 20 days after injection. Finally, graft size in C3H mice was similar when calculated from cardioCEST images as compared to histology (Figure 3). Longitudinal cardioCEST imaging enabled in vivo monitoring of cell survival/proliferation or rejection in a mouse model of cardiac cell therapy. Following rejection of labeled cells, Eu-HPDO3A is likely cleared from the myocardium similar to other lanthanide-HPDO3A complexes, leading to elimination of heightened MTR_asym values by 20 days post implantation. In proliferating cells the dilution of label with cell division led to a reduction in MTRasym values by 20 days post implantation that remained elevated compared to non-labeled and rejected cells. Further investigation is needed to understand the association between MTR_asym values and cell density. Imaging of cell fate decisions using cardioCEST MRI could support the design of novel methods to improve cell survival and differentiation in cardioregenerative therapies.