Novel Characterization and Live Imaging of Schlemm's Canal Expressing Prox-1

Abstract

Schlemm's canal is an important structure of the conventional aqueous humor outflow pathway and is critically involved in regulating the intraocular pressure. In this study, we report a novel finding that prospero homeobox protein 1 (Prox-1), the master control gene for lymphatic development, is expressed in Schlemm's canal. Moreover, we provide a novel in vivo method of visualizing Schlemm's canal using a transgenic mouse model of Prox-1-green fluorescent protein (GFP). The anatomical location of Prox-1+ Schlemm's canal was further confirmed by in vivo gonioscopic examination and ex vivo immunohistochemical analysis. Additionally, we show that the Schlemm's canal is distinguishable from typical lymphatic vessels by lack of lymphatic vessel endothelial hyaluronan receptor (LYVE-1) expression and absence of apparent sprouting reaction when inflammatory lymphangiogenesis occurred in the cornea. Taken together, our findings offer new insights into Schlemm's canal and provide a new experimental model for live imaging of this critical structure to help further our understanding of the aqueous humor outflow. This may lead to new avenues toward the development of novel therapeutic intervention for relevant diseases, most notably glaucoma.

Impaired angiopoietin/Tie2 signaling compromises Schlemm’s canal integrity and induces glaucoma

“…We and others recently reported that Prox1 is expressed in SC ECs (7,10) and that its expression is changed by AHO and VEGFR3 signaling (8,9). To determine the role of Prox1 in SC development, we generated a Prox1 iΔEC mouse by crossing the VE-cadherin-Cre-ER T2 mouse (34) with the Prox1 fl/fl mouse (39), administered tamoxifen starting at P5, and analyzed them at P7 ( Figure 11A).…”

“…Moreover, connecting fibrils stretching from the SC ECs into the surrounding cribriform plexus play a role in preventing SC collapse (14), similar to that of the LVanchoring filaments (6,15). Recently, we and others found that the SC identity is a mixture between LV and blood vessel (BV), as indicated by the expression of the lymphatic markers Prox1, VEGFR3, and integrin α9 (no LYVE-1 and podoplanin) and blood markers such as Tie2 (7)(8)(9)(10). The SC originates from the choroidal vein, and postnatally, SC ECs acquire a lymphatic fate by the upregulation of Prox1 (8-10), a master transcriptional factor of lymphatic ECs (LECs) (16, Primary open-angle glaucoma (POAG) is often caused by elevated intraocular pressure (IOP), which arises due to increased resistance to aqueous humor outflow (AHO).…”

“…SC is an encircling channel consisting of continuous, nonfenestrated endothelial cell (EC) monolayers at the periphery of the cornea (4,6) and shares morphological, functional, and molecular similarities with lymphatic vessels (LVs) (7)(8)(9)(10). Similar to the thoracic duct, which drains lymph into the subclavian vein through the lymphovenous valve (11), the SC constitutes a blind-ended tube that transfers AH and antigen-presenting cells to the systemic circulation through the episcleral vein (12,13).…”

Schlemm’s canal: more than meets the eye, lymphatics in disguise

“…Immunohistochemical staining to characterize the ECs of SC revealed that they were positive for certain lymphatic and blood endothelial markers, including PROX1, integrin α9, CD31, lin, exhibited corneal edema, inflammation, and dilated corneoscleral lymphatic vessels (12). Earlier this year, Truong et al discovered high expression of the lymphatic transcription factor prospero-related homeobox 1 (PROX1) on SC endothelium in the eye (13), linking expression of the lymphatic fate determinant PROX1 to a structure within the eye. In this issue of JCI, two independent yet complementary studies by Park et al (14) and Aspelund et al (15) use genetic mouse models and sophisticated lymphatic imaging techniques to offer insight into the developmental origin of SC and the pathology associated with its improper regulation.…”

All TIEd up: mechanisms of Schlemm’s canal maintenance

“…However, some functional characteristics, such as the absence of pericyte coverage, a discontinuous basement membrane, an absence of blood filling, and the known role of SC in AH and immune cell draining from the eye to the blood circulation, led to suggestions that SC shares traits with the lymphatic vasculature (9). This mystery was solved definitively in 2014, when several research teams independently described the developmental origins and molecular mechanisms of SC formation (10)(11)(12)(13)(14). SC was shown to develop postnatally from the choroidal vein and to present a hybrid molecular Figure 1.…”