Myofibroblasts in proliferative diabetic retinopathy can originate from infiltrating fibrocytes and through endothelial-to-mesenchymal transition (EndoMT)

Abstract

Myofibroblasts expressing α-smooth muscle actin (α-SMA) are the key cellular mediator of fibrosis. Fibrovascular epiretinal membranes from patients with proliferative diabetic retinopathy (PDR) are characterized by the accumulation of a large number of myofibroblasts. We explored the hypothesis that proliferating endothelial cells via endothelial-to-mesenchymal transition (EndoMT) and/or bone marrow-derived circulating fibrocytes contribute to the myofibroblast population present in PDR epiretinal membranes. Epiretinal membranes from 14 patients with PDR were studied by immunohistochemistry. All membranes contained neovessels expressing the endothelial cell marker CD31. CD31(+) endothelial cells co-expressed the fibroblast/myofibroblast markers fibroblast-specific protein-1 (FSP-1) and α-SMA, indicative for the occurrence of endoMT. In the stroma, cells expressing FSP-1, α-SMA, the leukocyte common antigen CD45, and the myelomonocytic marker CD11b were detected. Double labeling showed co-localization of CD45 with FSP-1 and α-SMA and co-localization of CD11b with α-SMA and matrix metalloproteinase-9, demonstrating the presence of infiltrating fibrocytes. In addition, we investigated the phenotypic changes that take place in human retinal microvascular endothelial cells following exposure to transforming growth factor-β1 (TGF-β1), connective tissue growth factor (CTGF) and the proinflammatory cytokines interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α). Retinal microvascular endothelial cells changed morphology upon cytokine exposure, lost the expression of endothelial cell markers (endothelial nitric oxide synthase and vascular endothelial-cadherin) and started to express mesenchymal markers (calponin, snail, transgelin and FSP-1). These results suggest that endothelial cells as well as circulating fibrocytes may differentiate into myofibroblasts in the diabetic eye and contribute to pathologic fibrosis in PDR.

Keywords: Endothelial cells; Endothelial-to-mesenchymal transition (EndoMT); Fibrocytes; Myofibroblasts; Proliferative diabetic retinopathy; Retina.