Identification of ocular matrix metalloproteinases present within the aqueous humor and iridocorneal drainage angle tissue of normal and glaucomatous canine eyes

Purpose The purpose of this study was to identify the most active matrix metalloproteinases (MMP) within the aqueous humor and iridocorneal angle tissue involved in the normal canine eye, and to compare these results to the MMP activity in dogs with glaucoma. Animal studied Aqueous humor samples from 32 normal eyes and 26 glaucomatous eyes were obtained through aqueous centesis and analyzed for MMP activity. Iridocorneal angle tissue from 16 enucleated normal canine eyes and 5 enucleated glaucomatous eyes were dissected and homogenized into solution. Procedure Bradford total protein assays were determined for aqueous humor samples and iridocorneal angle tissue samples. Substrate gelatin zymography was performed using 0.2 mu L volumes of an MMP-2/MMP-9 control, 2 mu L volumes of aqueous humor samples, and 10 mu g weights of total protein from iridocorneal angle tissue. The presence of MMP gel bands were identified visually and measured quantitatively by densitometry technique. A statistical analysis was performed on the data using Student's t-test, multiple logistic Wald's chi-squared regression, Pearson correlations, and repeated measures analysis. Results Within the aqueous humor of canine eyes, MMP-2 latent form was found to be the most relevant MMP. The quantity of latent MMP-2 within the aqueous humor of the glaucoma samples was significantly increased compared to the normal aqueous samples (P < 0.0001). Glaucoma occurrence was associated with elevated aqueous humor latent MMP-2 (P = 0.0002). Within the canine iridocorneal angle tissue, MMP-9 latent form and MMP-2 active form were found to be the most relevant MMPs. MMP-2 active form was found to be significantly increased in the glaucoma tissue samples when compared to the normal tissue samples (P = 0.0044). MMP-9 latent form was also found to be significantly increased in glaucomatous tissue when compared to the normal eyes (P = 0.0002). Tissue MMP-9 latent form was found to be associated with glaucoma status (P = 0.042). Conclusion Glaucoma aqueous humor samples expressed a statistically increased latent MMP-2 when compared to normal eyes. Iridocorneal angle tissue from glaucomatous eyes expressed a statistically significant increase in active MMP-2 and latent MMP-9 when compared to normal eyes. These data demonstrate that there is an association between elevated levels of intraocular MMP-2 and MMP-9 and the presence of glaucoma.