Regulation of copper metabolism in the diabetic retina and effects of treatment with the Cu(II) chelator triethylenetetramine (TETA)

Abstract

Purpose : Dysregulation of copper (Cu) homeostasis has been implicated in the pathology of diabetic organ damage. Free Cu ions may interact with peroxidases and superoxides to catalyse reactive oxygen species production. Oral treatment with the Cu(II) chelator triethylenetetramine (TETA) has been shown to reverse structural and functional changes in the diabetic heart and kidney. We hypothesised that there is also impaired Cu homeostasis in the diabetic retina and this contributes to the oxidative stress that has been observed in this condition. Therefore, the regulation of Cu metabolism was investigated in the diabetic retina as well as the potential for therapeutic or toxic effects from TETA treatment.

Methods : Male Sprague Dawley rats were administered saline or 55mg/kg of streptozotocin (STZ). STZ-treated rats were split into untreated and TETA-treated groups and maintained for 16 weeks. 30mg/rat/day of TETA-disuccinate was administered via drinking water. Eyes were enucleated and retinas were dissected and processed for analysis. mRNA expression of copper transporters and oxidative stress mediators was analysed using rt-qPCR.

Results : We demonstrate that mRNA expression of ceruloplasmin, a copper-bound iron ferroxidase, is increased during hyperglycaemia compared to control (p < 0.0001) and this was partially normalised by TETA treatment (p < 0.001). Metallothionein 1 and 2 mRNA expression was increased during hyperglycaemia compared to controls (p < 0.0001) and Metallothionein 2 expression was further increased by TETA treatment compared to untreated diabetic (p < 0.05). ATP7A expression was significantly decreased in TETA-treated retinas compared to both untreated control and diabetic retinas (p < 0.05).

Conclusions : The observed changes to ceruloplasmin expression suggest that there is increased copper in the diabetic retina which was partially normalised by TETA-treatment. Changes to metallothioneins may indicate a protective effect, as these sequester and store reactive Cu ions. Studies are ongoing to confirm that these changes are present at a protein level as is quantification of trace metals in the retina using inductively coupled plasma mass spectrometry.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.