Abstract
Purpose :
Retinal pigment epithelium (RPE) cell transplantation is a promising therapy for retinal degenerative spectrum diseases. However, the in vivo observation of transplanted RPE cells remains to be the obstacle for the postoperative evaluation of therapeutic efficacy. Here we established a novel in vivo tracking system to localize and trace transplanted RPE cells in subretinal space.
Methods :
The combination of near-infrared fluorescent nanoprobe with fundus fluorescent imaging was applied in tracking transplanted human fetal RPE (hfRPE) cells transfected with GFP lentivirus. The nanoprobe was incubated with pre-transplanted hfRPE cells. Fluorescence of GFP and nanoprobe channel were captured in fundus, respectively. To further demonstrate the labeling of nanoprobe, immunofluorescent staining of distinct cell type markers was performed. Single-cell RNA sequencing was also applied to analyze the composition of nanoprobe-labeling cells and the microenvironment of subretinal xenotransplantation.
Results :
Nanoprobe remained stable in hfRPE cells in vitro. After transplantation, fluorescent area of nanoprobe in fundus was consistent with that of GFP, as demonstrated in immunofluorescent staining. However, with the long-term observation, the fluorescence of nanoprobe was expanding beyond GFP signal, possibly resulting from the phagocytosis of transplanted cells by macrophage which was corelated with the infiltration of IBA1+ cells to the transplanted site. Single-cell RNA sequencing analysis of nanoprobe+ cells further revealed the incorporation of immune cells and fibroblasts in addition to hfRPE. It shows the additional role of nanoprobe fluorescent tracking in the in-vivo indication of immune rejection and inflammation.
Conclusions :
The near-infrared nanoprobe can be applied as a reliable method to label and trace hfRPE cells combined with in vivo fundus fluorescent imaging. Furthermore, the long-term observation of nanoprobe fluorescence can predict the extent of immune activity, which may serve as a clinical indicator for the modulation of immunosuppressant regimen.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.