Indications of lymphatic endothelial differentiation and endothelial progenitor cell activation in the pathology of proliferative diabetic retinopathy

Purpose: Proliferative diabetic retinopathy (PDR) is characterized by ischaemiaand inflammation-induced neovascularization, but the pathological vascular differentiation in PDR remains poorly characterized. Here, endothelial progenitor and growth properties, as well as potential lymphatic differentiation, were investigated in the neovascular membrane specimens from vitrectomized patients with PDR. Methods: The expression of pan-endothelial CD31 (PECAM-1), ETS-related gene (ERG), alpha-smooth muscle actin (alpha-SMA), and stem/progenitor cell marker CD117 (c-kit) and cell proliferation marker Ki67 was investigated along with the markers of lymphatic endothelial differentiation (vascular endothelial growth factor receptor (VEGFR)-3; prospero-related homeobox gene-1 (Prox-1), lymphatic vessel endothelial receptor [LYVE)-1 and podoplanin (PDPN)] by immunohistochemistry. Lymphocyte antigen CD45 and pan-macrophage marker CD68 were likewise investigated. Results: All specimens displayed CD31, ERG and alpha-SMA immunoreactivity in irregular blood vessels. Unexpectedly, VEGFR3 and Prox-1 lymphatic marker positive vessels were also detected in several tissues. Prox-1 was co-expressed with CD117 in lumen-lining endothelial cells and adjacent cells, representing putative endothelial stem/progenitor cells and pro-angiogenic perivascular cells. Immunoreactivity of CD45 and CD68 was detectable in all investigated diabetic neovessel specimens. PDPN immunoreactivity was also detected in irregular lumen-forming structures, but these cells lacked CD31 and ERG that mark blood and lymphatic endothelium. Conclusions: Although the inner part of human eye is physiologically devoid of lymphatic vessels, lymphatic differentiation associated with endothelial stem/progenitor cell activation may be involved in the pathogenesis of human PDR. Further studies are warranted to elucidate whether targeting lymphatic factors could be beneficial in the treatment of patients with the sight-threatening forms of DR.