Lymphatic vessels identified in failed corneal transplants with neovascularisation

Abstract

BackgroundCorneal transplant failure with neovascularisation is a leading indication for full-thickness grafts in patients. Lymphangiogenesis is implicated in the pathology of graft failure, and here we systematically evaluate failed human corneal transplants with neovascularisation for the presence of lymphatic vessels.MethodsNine failed grafts with neovascularisation, based on H&E staining with subsequent immunoperoxidase staining for CD31, a blood vessel marker, were selected. Lymphatics were investigated by immunohistochemical and immunofluorescence approaches using podoplanin as a lymphatic marker. In two of nine cases, fluorescence in situ hybridisation (FISH) was used for detection of lymphatic mRNAs including podoplanin, VEGFR-3 and LYVE-1. All immunofluorescence and FISH samples were compared with positive and negative controls and visualised by confocal microscopy.ResultsCorneal neovascularisation was established in all cases by H&E and further confirmed by CD31 immunoreactive profiles. Immunohistochemistry for the podoplanin antibody was positive in all cases and showed morphologies ranging from distinct luminal structures to elongated profiles. Simultaneous immunofluorescence using CD31 and podoplanin showed lymphatic vessels distinct from blood vessels. Podoplanin immunofluorescence was noted in seven of nine cases and revealed clear lumina of varying sizes, in addition to lumen-like and elongated profiles. The presence of lymphatic mRNA was confirmed by FISH studies using a combination of at least two of podoplanin, VEGFR-3 and LYVE-1 mRNAs.ConclusionsThe consistent finding of lymphatic vessels in failed grafts with neovascularisation implicates them in the pathogenesis of corneal transplant failure, and points to the lymphatics as a potential new therapeutic target.

Variable Responses to Corneal Grafts: Insights from Immunology and Systems Biology

“…Simultaneous immunofluorescence using CD31 and podoplanin distinguishes lymphatic vessels from blood vessels. Moreover, confirming the presence of lymphatic mRNA by identifying at least two of the following mRNAs: podoplanin, VEGFR-3, and LYVE-1, may improve the accuracy of detection post-transplantation [109].…”

Application of microscale culture technologies for studying lymphatic vessel biology

“…While lymphangiogenesis is essential to coordinate immune responses, increased lymphatic vessel growth may have unfavorable outcomes in certain settings . One such example is increased lymphangiogenesis enhanced immune responses in kidney and corneal transplant rejection . In addition, aberrant levels of lymphangiogenesis are common in numerous primary human cancers including melanoma, breast, lung, and head and neck .…”

“…117 One such example is increased lymphangiogenesis enhanced immune responses in kidney and corneal transplant rejection. 118,119 In addition, aberrant levels of lymphangiogenesis are common in numerous primary human cancers including melanoma, 120 breast, 121 lung, 122 and head and neck. 123,124 Furthermore, increased lymphangiogenesis, particularly at the margins of primary tumors, is strongly implicated in metastasis through the lymphatic vasculature.…”

Corneal Lymphangiogenesis: Current Pathophysiological Understandings and Its Functional Role in Ocular Surface Disease

“…Diamond et al found that podoplanin hi LVs were distinct from BVs in failed human corneal grafts. Moreover, VEGFR-3 and LYVE-1 mRNA levels were found to be elevated [ 85 ]. Previously, our group also found LVs in the corneal button from pseudophakic bullous keratopathy and HSV-1 keratitis [ 86 ].…”