Abstract 20549: Advanced Glycation End Product Modification of Laminin Alters Kir4.1 Expression in Retinal Glia Cells
Diabetes alters neurotransmitter metabolism of the body increasing susceptibility to cardiovascular disorders. Retinal vascular dysfunction in diabetes is viewed as a change in the neurovascular unit which exhibits striking similarities with the neurovascular unit of the brain. Diabetic retinopathy (DR), a major cause of adult blindness is linked to a change in the retinal neurovascular unit, leading to dysfunction between the physical and biochemical relationship of neurons, glia, and retinal vasculature. Retinal glia, or Müller cells, span across the retina to maintain water homeostasis and potassium concentration via inwardly rectifying Kir4.1 channels. Diabetes is linked to an increase in Advance Glycation End products (AGEs) and a decrease in Kir4.1 expression, but no reports show how AGEs-linked to the basement membrane (BM) impact normal Kir4.1 expression. Laminin, a BM-component, is necessary for the expression of Kir4.1 in Müller cells. In this study, we hypothesized that AGE-modification of laminin is detrimental to Kir4.1 expression, thereby disrupting Müller cell function. The AGE-modified laminin-coated substrates were prepared by coating Petri-dishes in laminin (20 μg/mL) and methylglyoxal (MGO; 10, 100, 1000 μM) for seven days at 37°C. The rat Müller cells (rMC-1) were then propagated on AGE-modified laminin dishes for 24 hours. The expression of Kir4.1 and Kcnj10 (gene for Kir4.1) was determined using immunofluorescence and qRT-PCR respectively. The immunofluorescence studies revealed a profound decrease in the Kir4.1 expression on AGE-modified substrates as compared to untreated laminin. The mRNA expression of Kcnj10 exhibited a 3.4-fold decrease (p < 0.05) on AGE-modified laminin when compared to untreated cells. Potassium conductance studies using a PBFI assay revealed a 1.5-fold increase (p<0.05) in intracellular potassium concentration, suggesting a defect in potassium conductance on AGE-modified laminin. In conclusion, our studies suggest that AGE-modification of laminin is detrimental to normal Kir4.1 expression. In the future, prevention of AGE-modification will be a useful strategy to prevent dysfunction of the Müller cell and neurovascular unit to control the cardiovascular risks associated with diabetes.
Author Disclosures: K.M. Thompson: None. Q. Luo: None. A.D. Bhatwadekar: None.
- © 2016 by American Heart Association, Inc.