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(Investigative Ophthalmology and Visual Science. 2007;48:3051-3055.)
© 2007 by The Association for Research in Vision and Ophthalmology, Inc.
DOI:  10.1167/iovs.06-1083
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Endogenous TNF{alpha} Suppression of Neovascularization in Corneal Stroma in Mice

Shuko Fujita,1 Shizuya Saika,1 Winston Whei-Yang Kao,2 Kyoko Fujita,1 Takeshi Miyamoto,1 Kazuo Ikeda,3 Yuji Nakajima,3 and Yoshitaka Ohnishi1

1From the Department of Ophthalmology, Wakayama Medical University, Wakayama, Japan; the 2Department of Ophthalmology, University of Cincinnati Medical Center, Cincinnati, Ohio; and the 3Department of Anatomy, Graduate School of Medicine, Osaka City University, Osaka, Japan.

PURPOSE. To examine the role of tumor necrosis factor {alpha} (TNF{alpha}) in stromal neovascularization in injured cornea in vivo and in cytokine-enhanced vessel-like endothelial cell tube formation in vitro.

METHODS. An in vitro model of angiogenesis was used to examine the roles of TNF{alpha} on tube formation by human umbilical vein endothelial cells (HUVECs) cocultured with fibroblasts on induction by transforming growth factor ß1 (TGFß1) and vascular endothelial growth factor (VEGF). Central cauterization was used to induce stromal neovascularization in corneas of wild-type (WT) and TNF{alpha}-null (Tnf{alpha}–/–) mice. At 7, 14, or 21 days of injury, experimental mice were killed, and the eyes were enucleated and subjected to histologic and immunohistochemical examination and real-time reverse transcription–polymerase chain reaction.

RESULTS. HUVECs formed a vessel-like tube structure on the fibroblast feeder layer. Adding TGFß1, VEGF, or both augmented vessel-like tube formation by HUVECs cocultured with fibroblasts. Adding TNF{alpha} (5 ng/mL) completely abolished the formation of tube-like structures despite the presence or absence of TGFß1 or VEGF in coculture. In vivo, cauterization of the central cornea induced the formation of CD31+ new vessels surrounding the limbus in WT mice. More prominent central stromal neovascularization accompanied by increased expression of TGFß1 and VEGF was found in Tnf{alpha}–/– mice compared with WT mice.

CONCLUSIONS. In addition to inhibiting TGFß1 and VEGF expression by fibroblasts, endogenous TNF{alpha} may counter the induction effects of TGFß1 and VEGF on vascular endothelial cells and may block neovascularization.




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