The cornea is the outermost layer of the eye serving as the first barrier from the outside environment which plays a crucial role in a clear vision. It is maintained by keeping transparency and avascularity under the normal state. This nature makes the cornea an ideal site to study lymphangiogenesis (LG), new lymphatic vessel growth, that can be induced by inflammation and assessed without any background vessels. The lymphatic vessels have long been neglected over blood vessels due to its invisibility, but its importance and involvement in inflammation, cancer metastasis, and transplant rejection are now well known since the discovery of several lymphatic-specific markers in recent years. Another important event that occurs during LG is valvulogenesis (VG). Previous studies from our lab have shown that lymphatic vessels develop luminal valves in the cornea during LG, and these valves express integrin alpha 9 (Itga-9) that plays a critical role in directing lymph flow.
My study consists of three parts: 1) to investigate the dynamic changes of LG and VG during corneal inflammation. This was revealed by our newly developed intravital imaging system; 2) to assess the role of VG itself or VG and LG together in corneal transplant rejection; and 3) to explore the cellular origin of corneal lymphatic endothelial cells that comprises lymphatic vessel walls.
First, using the intravital imaging technology, I demonstrated the multifaceted dynamics of LG and VG associated with corneal transplantation, from the initiation to regression phases, and reported several novel and critical phenomena and mechanisms that couldn’t be detected by conventional ex vivo immunohistochemistry.
Secondly, I investigated corneal transplant rejection reaction after blocking VG alone or in combination of LG. The results showed that anti–Itga-9 treatment alone suppressed corneal VG after transplantation. While this treatment did not affect LG, it promoted corneal graft survival in the low-risk setting. This study has provided the first evidence on the critical role of VG in mediating graft rejection. Furthermore, I have shown that combined blockade of VEGFR-3 (vascular endothelial growth factor receptor-3) and Itga-9 significantly suppressed both LG and VG after corneal transplantation, and this treatment led to a markedly promoted survival rate in the high-risk setting.
Lastly, I have provided the first in vivo evidence showing that transplanted bone marrow (BM) derived cells integrated into the newly formed lymphatic vessels in inflamed host corneas with the live imaging system.
In summary, my study reveals novel molecular and cellular mechanisms that contribute to LG and VG in the cornea. It is our hope that understanding these mechanisms would lead to novel therapies not only for corneal transplant rejection and inflammation, but also for lymphatic-related disease occurring outside the eye, such as cancer metastasis and lymphedema.