Inhibition of Connective Tissue Growth Factor Ameliorates Pathological Changes in Diabetic Retinopathy
Abstract Number: 1003-P
Authors: JENNIFER L. WINKLER, MAMDOUH H. KEDEES, YELENA GUZ, GLADYS TEITELMAN
Institutions: Brooklyn, NY
Results: Vascular Endothelial Growth Factor (VEGF) is responsible for many of the vascular changes that occur in Diabetic Retinopathy (DR). VEGF induces the expression of Connective Tissue Growth Factor (CTGF), a profibrotic factor shown to induce extracellular matrix (ECM) production and angiogenesis. We sought to determine whether CTGF levels are elevated during DR and if inhibition of CTGF has an effect on the pathological changes associated with the disease.
We found that CTGF mRNA and protein levels are increased by 7 and 5-folds respectively in retina of rats after 12 weeks of streptozotocin-induced diabetes. Immunohistochemical analysis indicated that hyperglycemia lead to elevated CTGF expression in retinal Müller cells. Quantitative Real-Time PCR demonstrated an upregulation of the mRNA levels of Fibronectin (3-fold), Laminin ß1 (2-fold), Collagen IV (2-fold) and VEGF (2-fold). Hyperglycemia for 12 weeks also induced extensive vascular leakage in retinal blood vessels. Exogenous insulin therapy for 12 weeks significantly reduced mRNA levels of CTGF and of the genes examined and, in addition, it prevented retinal vascular leakage.
Since insulin therapy is associated with oscillations in blood glucose levels, we developed a supplementary gene therapy. First, we tested the efficiency of three CTGF siRNA's in vitro to decrease the levels of Transforming Growth Factor-β (TGF-β)-induced CTGF expression in Rat 2 fibroblasts. The up-regulation of CTGF by TGF-β was inhibited by 60% after treatment with siRNA.
Next, we examined whether the augmentation of CTGF levels during diabetes in rats could be attenuated by the injection of CTGF-siRNA. After 12 weeks of hyperglycemia, one eye was injected with the in-vitro tested CTGF-siRNA and the contralateral eye was injected with a scrambled siRNA and eyes were examined three days later. Treatment with siRNA resulted in a 40%, 50% and 60% decrease in CTGF, Laminin and Collagen IV mRNA respectively and a 50% decrease in CTGF protein. These findings suggest that CTGF is a possible target for gene therapy to ameliorate the transient deleterious effect of insulin treatment in diabetes.