The Essential Role of the Choriocapillaris in Vision: Novel Insights from Imaging and Molecular Biology

Abstract

The choriocapillaris, a dense capillary network located at the posterior pole of the eye, is essential for supporting normal vision, supplying nutrients, and removing waste products from photoreceptor cells and the retinal pigment epithelium. The anatomical location, heterogeneity, and homeostatic interactions with surrounding cell types make the choroid complex to study both in vivo and in vitro. Recent advances in single-cell RNA sequencing, in vivo imaging, and in vitro cell modeling are vastly improving our knowledge of the choroid and its role in normal health and in age-related macular degeneration (AMD). Histologically, loss of endothelial cells (ECs) of the choriocapillaris occurs early in AMD concomitant with elevated formation of the membrane attack complex of complement. Advanced imaging has allowed us to visualize early choroidal blood flow changes in AMD in living patients, supporting histological findings of loss of choroidal ECs. Single-cell RNA sequencing is being used to characterize choroidal cell types transcriptionally and discover their altered patterns of gene expression in aging and disease. Advances in induced pluripotent stem cell protocols and 3D cultures will allow us to closely mimic the in vivo microenvironment of the choroid in vitro to better understand the mechanism leading to choriocapillaris loss in AMD.

Keywords: age-related macular degeneration; choroid; endothelial cell; iPSC; induced pluripotent stem cell; single-cell RNA sequencing.

Figure 1.

Relationship of photoreceptor cells, RPE and choriocapillaris. Section of human retina labeled with antibodies directed against rhodopsin (magenta), pooled anti-cone opsins (green), UEA-I, which binds vascular endothelial cells of the retina and choroid (red) and DAPI, which labels DNA (blue). Note the relatively sparse vasculature in the inner retina, absence of vascular elements in the outer retina, and dense, large diameter vessels in the choriocapillaris. Scalebar = 25μm. INL, inner nuclear layer; ONL, outer nuclear layer; IS, inner segments; OS, outer segments; RPE, retinal pigment epithelium; CC, choriocapillaris.

Figure 2.

Confocal projection stereo pair of the human choroidal vasculature labeled with UEA-I lectin. Note the very dense honeycomb appearance of the choriocapillaris (superficial) and the complexity of the intermediate and large vessels. A potential A-V shunt is indicated (asterisk).

Figure 3.

Short posterior ciliary artery penetrating the sclera (SC, margin indicated by dotted line) to supply the choroidal arterioles and ultimately choriocapillaris. Red, CC, choriocapillaris, CH, Saller’s and Hatler’s layers of the choroid, RPE, retinal pigment epithelium. Red, anti-smooth muscle actin antibody; green, anti-vascular adhesion protein antibody, scalebar = 100μm.

Figure 4

In addition to endothelial cells (asterisks), cells of the human choroid include immune cells, both within the stroma (M, macrophage) and circulating (L); fibroblasts (F); melanocytes (Me); Schwann cells (S), smooth muscle cells (SM) and pericytes (P). Lu, vascular lumen. Scalebar = 2μm.

Figure 5:

Choroidal arteries, veins, and choriocapillaris endothelial cells identified with single-cell RNA sequencing. A. UMAP plot of 10,576 choroidal endothelial cells isolated from 6 human donors (Voigt et al., 2020c). Each point represents the multidimensional transcriptome of one cell. Clusters of cells (red, blue, purple) are classified into different endothelial cell types (arteries, veins, choriocapillaris). B. Violin plots demonstrate that all choroidal endothelial cells highly express the pan-endothelial cell gene VWF. Choroidal arteries specifically express SEMA3G, choroidal veins specifically express DARC, and the choriocapillaris specifically expressed CA4.

Figure 6:

Ghost choriocapillaris vessel (asterisk) beneath intact RPE monolayer. This degenerated capillary segment appears normal when the choroid is labeled with antibodies directed against type I collagen (green, top panel), but is unreactive with the endothelial cell binding lectin UEA-I (red, middle panel). Lower panel depicts merged channels with DAPI nuclear counterstain. RPE, retinal pigment epithelium; CC. choriocapillaris, CH outer choroid.

Figure 7.

Choroidal visualization on a normal fluorescein angiogram. A: Seven seconds after dye injection, the retinal arteries have begun to fill. The large choroidal vessels are visible (white arrow) and there is patchy choroidal flush (blue arrow) in the posterior pole. B: Thirty seconds later, the retinal circulation has filled, the choroidal flush has become homogenous, and the large choroidal vessels are no longer visible.

Figure 8.

Choroidal visualization on a normal indocyanine green angiogram. A: Sixteen seconds after dye injection, the retinal and choroidal circulations have begun to fill. There is patchy choroidal filling by the nerve (arrow). B: Thirty seconds later, the entire retinal and choroidal circulations have filled. There is no choroidal flush to obscure the choroidal vessels. Vortex veins are indicated by arrowheads.

Figure 9.

Choroidal visualization via swept source optical coherence tomography (SS-OCT) in the central 12×12mm of a normal retina. A: SS-OCT angiogram of the choriocapillaris. The speckled pattern is normal. B: En face SS-OCT of the large choroidal vessels. The choroidal vessels appear dark (arrow; see text). The overlying retinal vessels cause shadowing artefact. The blue lines in A and B indicate the position of the OCT line scans in C and D, respectively. The purple segmentation lines in C and D define the depth of tissue depicted in the en face images (A and B).

Figure 10.

SS-OCTA of choroidal neovascularization in age-related macular degeneration. A: This 64 year-old patient was referred with a diagnosis of subretinal fluid from central serous chorioretinopathy. There were multiple large, soft drusen bilaterally.B: A 12×12mm SS-OCTA segmented to include detectable flow at the level of the RPE and a few microns below it showed a CNV (magnified view, inset). C: The corresponding SS-OCT line scan had an RPE elevation corresponding to the CNV.

Figure 11.

En face SS-OCT imaging to create choroidal thickness and vascularity maps. This patient with proliferative diabetic retinopathy was imaged with 12×12mm SS-OCT to generate a choroidal thickness map (A), choroidal vascularity map (B), and choroidal vascularity index map (C). Areas with a thicker choroid or larger CVI measurements are depicted with warmer colors in the respective topographic heat maps (A, C). See Russell et al. (manuscript in revision) for more details.