Photoacoustic Microscopy, Optical Coherence Tomography, and Fluorescence Imaging-Guided Stem Cell Retinal Therapy

Purpose: Stem cell therapy has the potential to treat currently incurable diseases, such as geographic atrophy in macular degeneration. However, tracking stem cells after transplantation is a major challenge. This study demonstrates an advanced, non-invasive, high-resolution, multimodality platform technology for the longitudinal visualization of damaged retinal pigment epithelium (RPE) using photoacoustic microscopy (PAM), optical coherence tomography (OCT), and fluorescence microscopy (FM) in living rabbits.

Methods: Millisecond laser photocoagulation was applied to 12 New Zealand rabbits to create RPE damage. On day 4 post-laser treatment, each eye received a subretinal injection of 30 µL (3.3×10⁶ cells/ µL) human-induced pluripotent stem cells differentiated to RPE (hiPSC-RPE) cells labeled with ultrapure chain-like gold nanoparticle (CGNP) clusters conjugated with indocyanine green. The CGNP clusters have a red-shifted optical absorption in the near-infrared window, and their diameter of 7-8 nm after disassembly enables renal excretion. hiPSC-RPE cells were followed up to 8 months after transplantation by color fundus photography, PAM, OCT, and FM.

Results: PAM images at 650 nm showed the distribution of the hiPSC-RPE cells, demonstrating that the cells rapidly localized to laser burns within 1 week and remained at the laser burn sites with signal for a duration of 8 months. PAM images at 578 nm visualized the microvasculature. Cells were observed using FM up to 28 days post-injection with a significant reduction in fluorescence signal by 1 month. The co-registration of PAM and OCT images validated the location of hiPSC-RPE cells to RPE injury sites. Histological and immunofluorescence images confirmed the imaging results. TEM and confocal images demonstrated CGNP clusters within hiPSC-RPE cells without affecting the cell’s morphology, pigmentation, and RPE differentiation or function.

Conclusions: This research presents an innovative platform technology for the longitudinal imaging of cell-based therapies in living animals for a duration of 8 months using PAM, OCT, and FM imaging.