Abstract

DO.19.02

Basement Membrane Alterations in diabetic retina of STZ- treated mice

Elizabeth Abari1, Markus Huemmeke2, Irina Semkova1, Norbert Kociok1, Mats Paulsson2, Antonia M. Joussen1
1Labor für Exp. Ophthalmologie, Universitäts-Augenklinik Düsseldorf, 2Institut für Biochemie und molekulare Medizin, Köln

Objective
The basement membrane (BM) is made up of the basal lamina and the lamina fibroreticularis. The major basement membranes of the eye are Bruch’s membrane, the inner limiting membrane (ILM), and the capillary BMs forming a part of the blood-retina barrier. All BMs are formed by members of three protein families: laminins, nidogens, collagen IV, and by the proteoglycan perlecan. We aim to investigate the expression of individual basement membrane components in STZ induced diabetes in the eye.
Methods
To identify abnormalities in the distribution of specific constituents, we analyzed eyes obtained from C57BL/6J mice that were rendered diabetic by five consecutive daily intraperitoneal injections of STZ (65 mg/kg). After 4 and 6 months of diabetes (defined by blood glucose >300 mg/dl) eyes were enucleated and examined by histological and immunohistochemical techniques. We used a panel of antibodies against the BM protein families (laminin, collagen, nidogen and perlecan) to describe differences in the composition between normal and diabetic retinal BMs. Western blot was performed to quantify the changes in the BM protein expression in diabetic and non-diabetic retinae.
Results
Compared to the controls, there was no difference in the perlecan expression in diabetic retina both 4 and 6 months of diabetes. The expression of collagen-IV was also not changed. In contrast, immunoreactivity for laminin-1, laminin alpha5 and gamma1 chain was enhanced in the capillaries and Bruch’s membrane of the diabetic retinae. Furthermore, immunoreactivity for nidogen-1 (normally moderate expressed in the Bruch’s membrane) appeared to increase in the retinae after 4 and 6 months of diabetes in comparison to the controls.  Western blot analysis showed approximately 2-fold increase in the nidogen-1 expression after 4 months of diabetes. Nidogen-2 (normally high expressed) was not changed.
Conclusions
These data give evidence for the participation of laminins, nidogens, collagen IV in the BM composition of the capillaries in diabetic retinopathy. It appears that nidogen-1 was up-regulated while the levels of collagen IV and laminin are unchanged. These changes in the BMs composition may affect both the assembly and function and ultimately lead to pathological consequences such as macula edema and retinal fibrosis.

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