Assessment of retinas of diabetic mice with mild elevation of plasma homocysteine.

Abstract

Abstract: : Purpose: Modest elevation of homocysteine (Hcy), a sulfur–containing amino acid that enhances the vulnerability of neuronal cells to excitotoxic injury, is a risk factor in hypertension and neurodegenerative diseases. We have shown that high levels of Hcy injected intravitreally in mice lead to significant ganglion cell (RGC) loss (Moore et al, 2001). Some clinical studies implicate elevation of plasma Hcy in the pathogenesis of diabetic retinopathy, which is characterized by RGC loss as well as vasculopathy. The present study addressed whether modest elevation of plasma Hcy in the presence of diabetes accelerates neuronal cell loss in early stages of diabetic retinopathy. Methods: Founder mice, heterozygous for a mutation of the cystathionine beta synthase (CBS) gene, were used to establish our colony. CBS +/– have twice the normal plasma Hcy levels. Diabetes (DB) was induced in 3 wk old mice using 3 injections of streptozotocin (75 mg/Kg). Four groups of mice were studied: DB CBS+/–; non–DB CBS+/–; DB CBS+/+; non–DB CBS+/+. At 5 and 10 wks post onset of diabetes, eyes were obtained, embedded in OCT and 10µm thick cryosections prepared. Sections were evaluated morphologically. Results: Significant differences in retinas of DB versus non–DB mice were obtained 10 wks post–onset of diabetes. DB CBS+/– and +/+ mice had significantly fewer RGCs than non–DB CBS+/– and +/+ mice (10.0 ± 0.5 & 10.3 ± 0.5 versus 14.9 ± 0.5 & 15.8 ± 0.6 cells/100 µm retina length, respectively.) There was noticeable disruption of the nerve fiber layer (NFL) in retinas of DB CBS+/–, but not in the other groups. Measurements of the total retinal thickness revealed a marked decrease in DB CBS+/– and +/+ mice compared to non–DB CBS +/– and +/+ mice (197.4 ± 4.8 & 204.2 ± 8.2 versus 242.6 ± 6.22 & 257.9 ± 7.7 µm, respectively.) The decrease in total retinal thickness was due to decreased thickness of the inner nuclear layer and the length of inner/outer segments. Conclusions: There is a significant loss of RGCs and a significant decrease in total retinal thickness in DB CBS +/– and +/+ versus non–DB mice. A modest increase of plasma Hcy does not appear to exaggerate neuronal cell loss in early stages of diabetes; it is however associated with marked disruption of the NFL. Future studies will determine whether longer duration of diabetes and modest elevation of Hcy result in more severe neuronal cell loss and/or vasculopathy in diabetic retinopathy.

Keywords: diabetic retinopathy • ganglion cells • excitatory neurotransmitters