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Abstract #26690 Published in IGR 12-3
Decreased carbohydrate metabolism enzyme activities in the glaucomatous trabecular meshwork
Junk AK; Goel M; Mundorf T; Rockwood EJ; Bhattacharya SKMolecular Vision 2010; 16: 1286-1291
Purpose: To determine whether activity of carbohydrate metabolism enzymes (aldolase, pyruvate kinase, isocitrate dehydrogenase, and malate dehydrogenase) are altered in the glaucomatous trabecular meshwork (TM) compared to controls. Methods: Tissue specimens were obtained from trabeculectomy (n=45 open angle glaucoma; Caucasian, average age 61(plus or minus)8 years of age of both genders) and from cadaver eyes (n=15 control and n=5 glaucoma; Caucasian, average age 63(plus or minus)4 years of both genders). Protein extracts from TM tissue were prepared in a non-denaturing buffer containing 0.1% genapol. Aldolase activity was measured spectrophotometrically at 240 nm absorbance using reaction of 3-phosphoglycerate with hydrazine to form hydrazone. Pyruvate kinase activity was measured by coupling lactate dehydrogenase with NADPH and pyruvate absorbance was measured at 340 nm. Isocitrate dehydrogenase activity was measured using reduction of NADP to NADPH at the characteristic absorbance at 340 nm. Malate dehydrogenase catalyzes the interconversion of Lmalate and oxaloacetate using NADP as a coenzyme, quantified by its absorbance at 340 nm. Results: Aldolase, pyruvate kinase, isocitrate dehydrogenase, and malate dehyrogenase activities in the glaucomatous TM tissue were found to be reduced 70, 50, 25, and 69 percent, respectively. SDS-PAGE analysis suggests the presence of 4-hydorxynonenal (HNE) modified isocitrate dehydrogenase protein in the glaucomatous TM tissue compared to controls. Conclusions: Several Krebs cycle enzyme activities are considerably reduced in glaucomatous TM. HNE modified isocitrate dehydrogenase activity is consistent with reduced inactivated form of the protein. Lipid peroxidation product modification of aldolase, pyruvate kinase, and isocitrate dehydrogenase serves as a likely reason for the reduction of enzyme activity.
A. K. Junk. Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, 900 NW 17th Street, Miami, FL 33136, United States. ajunk@med.miami.edu
Classification:
3.7 Biochemistry (Part of: 3 Laboratory methods)
2.5.1 Trabecular meshwork (Part of: 2 Anatomical structures in glaucoma > 2.5 Meshwork)
