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Cellular transport of subretinal material into choroidal and scleral blood vessels: an electron microscopic study
1Department of Vitreoretinal Surgery, University Eye Clinic, Cologne, Germany.
Background:
The fate of indigestible material injected into the subretinal space of rats was investigated.
Methods:
The non-toxic dye Monastral Blue (MB), which cannot be digested within the lysosomal compartment, was injected transsclerally into the subretinal space of Long Evans and Wistar rats. After 5 and 12 days respectively the eyes were enucleated and examined by light and electron microscopy. Cryo sections were made of eyes 5 days after MB injection for the application of immunohistochemical techniques using markers for epithelial cells (cytokeratin) and macrophages (ED 1).
Results:
Retina, choroid and sclera were not altered in their morphology in the circumference of the MB-containing bubble generated by subretinal injection. After both 5 and 12 days no injected material was found extracellularly in the subretinal space. Especially high amounts of MB were found, in particular 5 days after injection, in lysosomes and melanosomes of RPE cells as well as in cells between choroidal melanocytes. Cells containing MB were seen in contact with choroidal and scleral blood vessels. These MB-containing cells in the choroid and in the sclera were positive for macrophage antibodies.
Conclusion:
Subretinal injection was confirmed as a suitable method for placing fluids into the subretinal space without affecting the morphology of the retina. Subretinal injected material was shown to be incorporated into lysosomes and melanosomes of RPE cells. The injected material was subsequently transported through Bruch's membrane to be finally removed from the eye via choroidal and scleral veins, the process involving macrophages.
Insights
Indigestible material injected into the rat subretinal space is incorporated by retinal pigment epithelial cells. This material is then transported out of the eye via choroidal and scleral blood vessels, involving macrophages.
Area of Science:
- Ophthalmology
- Retinal Biology
- Cellular Biology
Background:
- Investigating the fate of indigestible material in the subretinal space is crucial for understanding ocular drug delivery and disease progression.
- The subretinal space is a potential site for therapeutic agent delivery, but its capacity for material clearance is not fully understood.
Purpose of the Study:
- To investigate the fate and transport of indigestible material injected into the subretinal space of rats.
- To assess the impact of subretinal injection on retinal morphology and identify the cellular mechanisms of material clearance.
Main Methods:
- Subretinal injection of Monastral Blue (MB), an indigestible dye, into Long Evans and Wistar rats.
- Examination of enucleated eyes using light and electron microscopy at 5 and 12 days post-injection.
- Immunohistochemical analysis using cytokeratin and macrophage markers (ED 1) on cryo-sections.
Main Results:
- Subretinal injection did not alter the morphology of the retina, choroid, or sclera.
- Injected Monastral Blue was not found extracellularly in the subretinal space after 5 or 12 days.
- Monastral Blue was primarily found within lysosomes and melanosomes of retinal pigment epithelial (RPE) cells and in macrophages in the choroid and sclera.
- MB-containing cells were observed in contact with choroidal and scleral blood vessels, indicating a route for systemic removal.
Conclusions:
- Subretinal injection is a safe method for introducing materials into the subretinal space without causing morphological damage.
- Injected indigestible material is internalized by RPE cells and subsequently transported through Bruch's membrane.
- The clearance of subretinal material involves macrophages and removal via choroidal and scleral vasculature.