Esther J. Kuiper

The angio-fibrotic switch of VEGF and CTGF in proliferative diabetic retinopathy.

Background In proliferative diabetic retinopathy (PDR), vascular endothelial growth factor (VEGF) and connective tissue growth factor (CTGF) cause blindness by neovascularization and subsequent fibrosis, but their relative contribution to both processes is unknown. We hypothesize that the balance between levels of pro-angiogenic VEGF and pro-fibrotic CTGF regulates angiogenesis, the angio-fibrotic switch, and the resulting fibrosis and scarring. Methods/Principal Findings VEGF and CTGF were measured by ELISA in 68 vitreous samples of patients with proliferative DR (PDR, N = 32), macular hole (N = 13) or macular pucker (N = 23) and were related to clinical data, including degree of intra-ocular neovascularization and fibrosis. In addition, clinical cases of PDR (n = 4) were studied before and after pan-retinal photocoagulation and intra-vitreal injections with bevacizumab, an antibody against VEGF. Neovascularization and fibrosis in various degrees occurred almost exclusively in PDR patients. In PDR patients, vitreous CTGF levels were significantly associated with degree of fibrosis and with VEGF levels, but not with neovascularization, whereas VEGF levels were associated only with neovascularization. The ratio of CTGF and VEGF was the strongest predictor of degree of fibrosis. As predicted by these findings, patients with PDR demonstrated a temporary increase in intra-ocular fibrosis after anti-VEGF treatment or laser treatment. Conclusions/Significance CTGF is primarily a pro-fibrotic factor in the eye, and a shift in the balance between CTGF and VEGF is associated with the switch from angiogenesis to fibrosis in proliferative retinopathy.

Expression of vascular endothelial growth factor, stromal cell-derived factor-1, and CXCR4 in human limb muscle with acute and chronic ischemia

Objective—Vascular endothelial growth factor (VEGF)-induced stromal cell-derived factor-1 (SDF-1) has been implicated in angiogenesis in ischemic tissues by recruitment of CXCR4-positive bone marrow-derived circulating cells with paracrine functions in preclinical models. Here, evidence for this is provided in patients with peripheral artery disease. Methods and Results—Expression patterns of VEGF, SDF-1, and CXCR4 were studied in amputated limbs of 16 patients. VEGF-A was expressed in vascular structures and myofibers. SDF-1 was expressed in endothelial and subendothelial cells, whereas CXCR4 was expressed in proximity to capillaries. VEGF-A, SDF-1, and CXCR4 expressions were generally decreased in ischemic muscle as compared with nonischemic muscle in patients with chronic ischemia (0.41-fold, 0.97-fold, and 0.54-fold induction [medians], respectively), whereas substantially increased in 2 patients with acute-on-chronic ischemia (3.5- to 65.8-fold, 3.9- to 19.0-fold, and 4.1- to 30.6-fold induction, respectively). Furthermore, these gene expressions strongly correlated with capillary area. Only acute ischemic tissue displayed a high percentage of hypoxia-inducible factor-1&agr;–positive nuclei. Conclusions—These data suggest that VEGF and SDF-1 function as pro-angiogenic factors in patients with ischemic disease by perivascular retention of CXCR4-positive cells. Furthermore, these genes are downregulated in chronic ischemia as opposed to upregulated in more acute ischemia. The VEGF-SDF-1-CXCR4 pathway is a promising target to treat chronic ischemic disease.

The role of CTGF in diabetic retinopathy

Connective tissue growth factor (CTGF, CCN2) contributes to fibrotic responses in diabetic retinopathy, both before clinical manifestations occur in the pre-clinical stage of diabetic retinopathy (PCDR) and in proliferative diabetic retinopathy (PDR), the late clinical stage of the disease. CTGF is a secreted protein that modulates the actions of many growth factors and extracellular matrix (ECM) proteins, leading to tissue reorganization, such as ECM formation and remodeling, basal lamina (BL) thickening, pericyte apoptosis, angiogenesis, wound healing and fibrosis. In PCDR, CTGF contributes to thickening of the retinal capillary BL and is involved in loss of pericytes. In this stage, CTGF expression is induced by advanced glycation end products, and by growth factors such as vascular endothelial growth factor (VEGF) and transforming growth factor (TGF)-β. In PDR, the switch from neovascularization to a fibrotic phase - the angio-fibrotic switch - in PDR is driven by CTGF, in a critical balance with vascular endothelial growth factor (VEGF). We discuss here the roles of CTGF in the pathogenesis of DR in relation to ECM remodeling and wound healing mechanisms, and explore whether CTGF may be a potential novel therapeutic target in the clinical management of early as well as late stages of DR.

Angiogenesis Is Not Impaired in Connective Tissue Growth Factor (CTGF) Knock-out Mice

Connective tissue growth factor (CTGF) is a member of the CCN family of growth factors. CTGF is important in scarring, wound healing, and fibrosis. It has also been implicated to play a role in angiogenesis, in addition to vascular endothelial growth factor (VEGF). In the eye, angiogenesis and subsequent fibrosis are the main causes of blindness in conditions such as diabetic retinopathy. We have applied three different models of angiogenesis to homozygous CTGF−/− and heterozygous CTGF+/− mice to establish involvement of CTGF in neovascularization. CTGF−/− mice die around birth. Therefore, embryonic CTGF−/−, CTGF+/−, and CTGF+/+ bone explants were used to study in vitro angiogenesis, and neonatal and mature CTGF+/− and CTGF+/+ mice were used in models of oxygen-induced retinopathy and laser-induced choroidal neovascularization. Angiogenesis in vitro was independent of the CTGF genotype in both the presence and the absence of VEGF. Oxygen-induced vascular pathology in the retina, as determined semi-quantitatively, and laser-induced choroidal neovascularization, as determined quantitatively, were also not affected by the CTGF genotype. Our data show that downregulation of CTGF levels does not affect neovascularization, indicating distinct roles of VEGF and CTGF in angiogenesis and fibrosis in eye conditions.